1 2005 Abstracts Please note: The following abstracts have not been edited for content. They appear as submitted by the authors. Sunday, October 30, 2005 Soy Isoflavones and Cognition: A Review of the Clinical Data. L. Dye and C. Hill, Human Appetite Research Unit, Institute of Psychological Sciences, University of Leeds, Leeds, LS2 9JT, UK. The aim of this presentation is to provide a detailed examination of the human experimental studies of the effects of soy derived isoflavones (soy IF) on cognitive function in females. In contrast to the vast number of studies of health effects of soy IF, there have been only a small number of intervention studies of soy IF with cognitive endpoints. There have been three intervention studies in young adults. One study (File et al., 2001) examined young men and women and found that a high soy diet (100mg/d) for 10 weeks improved verbal and non-verbal episodic memory and mental flexibility. Gender specific effects were noted for verbal fluency and planning ability, with improved performance in females on the high soy diet, but not males. Another found effects on spatial performance after consumption of 900g of soybeans within 1 week, although this study suffers from the lack of a control group (Celec et al., 2005). In the only double-blind randomised placebo-controlled trial in young adults, Hill et al. (2005) administered 68mg/d soy IF (aglycone weight, 116mg/d conjugated weight) in a soy food matrix across two consecutive menstrual cycles. An improvement in long-term verbal memory in the first and last week of the intervention was observed. There have been 4 published RCTs of soy IF in postmenopausal women varying in duration from 3-12months with a variety of intake forms and doses (ranging from 60 to 110mg/d). Three further unpublished data sets will be reported upon. Generally soy IF appear to exert stronger cognitive effects in postmenopausal women compared to premenopausal women and effects seem to be mainly confined to improvements in memory and certain aspects of frontal lobe function. Variables such as age, time of exposure, dose and administration of isoflavones may be important mediating factors in the effects observed. Whilst these studies have provided some evidence of significant effects on cognition which are more pronounced in postmenopausal than younger women, it is not possible to know whether the effects reported in these relatively short term interventions will be sustainable in the long-term. The intriguing preliminary data warrant the funding of long-term trials Absorption of Soy Bioactives Long-Term Dietary Habits Affect Soy Isoflavone Metabolism and Accumulation in Prostatic Fluid in Caucasian Men. T. Hedlund1, P. Maroni1, P. Ferucci1, R. Dayton1, S. Barnes2, K. Jones2, R. Moore2, L. Ogden1, K. Wahala3, H. Sackett1, and K. Gray1, 1University of Colorado Health Sciences Center, Aurora, CO, USA, 2University of Alabama at Birmingham, Birmingham, AL, USA, 3Univ. of Helsinki, Helsinki, Finland. The soy isoflavones daidzein and genistein are believed to reduce prostate cancer risk in soy consumers. However, daidzein can be metabolized by the intestinal flora to form a variety of compounds with different bioactivities. In the current study, we investigated the influence of long-term dietary habits on daidzein metabolism in healthy Caucasian men (19-65 y old). A secondary goal was to compare plasma and prostatic fluid concentrations of 5 isoflavonoids: genistein, daidzein, equol, dihydrodaidzein, and O-desmethylangolensin. Baseline plasma levels of isoflavonoids were quantitated in 45 men by HPLC/electrospray ionization/ mass spectrometry. Participants then consumed a soy beverage daily for 1 wk, and post-soy isoflavonoid levels were quantitated in plasma and prostatic fluid. Equol was the only metabolite that appeared to be influenced by routine dietary habits. Stratified analyses revealed that men who had consumed ≥30 mg/d soy isoflavones for at least 2 y had 5.3-times the probability of producing equol than men who had consumed ≥5 mg/d (P=0.014). Additionally, those men who consumed animal meat regularly had 4.7-times the probability of producing equol than men who did not consume meat (P=0.023). Equol production was not linked to age, BMI, or the consumption of yogurt, dairy, fruit or American style fast food. Daidzein and its metabolites (but not genistein) were typically present at higher levels in prostate fluid than plasma (median = 4-13 times that in plasma). In conclusion, our data suggest that the ability of Caucasian men to produce equol is favorably influenced by the long-term consumption of high amounts of soy, and the consumption of meat. Lastly, the high concentrations of isoflavonoids in prostatic fluid increases the potential for these compounds to have direct effects in the prostate. Bioavailability of the Cancer Preventive Soy Peptide Lunasin in Animals. B.O. de Lumen1, C. Lim1, I. Reyes1, P. Vichayavilas1, H. Chu1, J. Lee1, R. Hurwitz1, Y. Fang1 M. Fitch1 and H. Jeong2 , 1Division of Nutritional Sciences & Toxicology, University of California, Berkeley, CA, USA, 2Andong University, Andong, Korea. Oral bioavailability of the cancer preventive soy peptide lunasin was determined in mice and rats. 3H-lunasin was administered by gavage to male and female CD-1 mice with lunasin-enriched soy (LES) and organs were collected at 3,6,9,12, and 24 hrs post-administration. 3Hlunasin is absorbed and distributed in the various tissues collected including those that are targets for the most common cancers such as lungs, mammary gland, prostate and colon. At 3 hrs, approximately 30% of the total dose is accounted in all the tissues collected, extrapolation to 1 hr gives a value of approximately 38% absorption. Sprague-Dawley rats were fed LES for 4 weeks. Lunasin extracted from the blood and liver of these rats are intact and bioactive. Inflammatory Diseases Dietary Soy Protein During Pregnancy and Lactation Reduces Renal Inflammation and Disease Progression in Young Adult Rat Offspring with Genetically Determined Kidney Disease. H. Aukema1,2, L. Cahill1, C. Peng1, D. Sankaran1, N. Bankovic-Calic1, and M. Ogborn1,2, 1University of Manitoba, Winnipeg, Manitoba, Canada, 2Manitoba Institute of Child Health, Winnipeg, Manitoba, Canada. Dietary soy protein ameliorates disease progression in several ani- Oral Presentation Abstracts Oral Presentation Abstracts 6th International Symposium on the Oral Presentation Abstracts 2 Role of Soy in Preventing and Treating Chronic Disease mal models of kidney disease when soy exposure is initiated at weaning or later in life. We previously have demonstrated the beneficial effects of dietary soy protein exposure initiated at weaning in the Han:SPRD-cy rat, a genetic model of renal disease characterized by renal cyst growth and the common renal pathologies of inflammation and fibrosis. Therefore, the objective of the current study was to determine whether exposure to soy protein in the diet earlier in life would have further benefits. Han:SPRD-cy rats were given either soy protein or casein based diets from 2 wk prior to conception up to weaning and/or in the post-weaning period in a 2x2 design. During young adulthood (at 70 d of age), the progression of renal disease was examined by histology. As previously shown, soy protein compared to casein in the postweaning diet reduced disease progression, as evidenced by less inflammatory cell infiltration, renal cyst growth and epithelial cell proliferation. With respect to maternal effects, renal inflammatory cell infiltration was reduced by dietary soy protein exposure in the pregnancy/lactation period alone, and was even lower when maternal exposure was combined with soy protein feeding in the post-weaning period. For cell proliferation, maternal exposure to soy protein was just as effective as post-weaning exposure in lowering renal cell hyperproliferation in diseased kidneys. Maternal soy protein compared to casein also resulted in less proteinuria, indicating better renal function. In conclusion, these studies demonstrate that maternal soy protein exposure has long-term beneficial effects on renal inflammation and disease progression in young adult offspring in this model of genetically determined renal disease. • Supported by the Manitoba Instute of Child Health Isoflavonoid-Free Soy Shows Anti-Inflammatory Activity in an Experimental Model for Chronic Nonbacterial Prostatitis. M. Kauppila1, J. Bernoulli1, E. Yatkin*1, N. Saarinen2, and R. Santti1, 1Dept. of Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland, 2Functional Foods Forum, University of Turku, Turku, Finland. There is emerging evidence to support a role for non-bacterial chronic inflammation in prostate carcinogenesis. Because of the fragmentary and conflicting information available about anti-inflammatory properties of soy in prostate, we studied the effects of soy in an animal model reproducing the clinical picture of men with chronic non-bacterial prostatitis. Prostatic inflammation was induced in Noble rats with combined treatment with testosterone and estradiol known to induce prostate cancer. When kept on soy-free diet, rats developed gradually perivascular, stromal, and glandular inflammation in dorsolateral prostate. The inflammatory cells consisted mainly of T-lymphocytes (CD3+ cells). At six weeks, the inflammation proceeded to glandular form and subset of T-lymphocytes, cytotoxic (CD8+) cells were observed intraepithelially. Testosterone concentration and prostate size remained within the normal range while estrogen and prolactin concentrations were elevated. At 9 weeks, a gradual decline was measured in serum testosterone concentration and prostate weight. Dietary soy reduced inflammatory changes in each category (perivascular, stromal and glandular) while body weight gain, prostate size and hormone concentrations in serum remained unaltered. When rats were kept on isoflavonoidfree soy, the content of isoflavones in urine markedly decreased, yet an equivalent decrease in the number of inflammatory foci was observed at 3 and 9 weeks. Neither soy nor isoflavonoid-free soy showed any signs of anti-estrogenic potency. If inflammatory etiology is found important in prostate carcinogenesis, anti-inflammatory therapy based on regular consumption of soy or isoflavonoid-free soy could be enhanced. • Supported by Raisio Group Foundation, Raisio, Finland. Monday, October 31, 2005 Equol A Review of the Equol Hypothesis. M. Kurzer, University of Minnesota, Saint Paul, Minnesota, USA. It has been proposed that the isoflavone metabolite equol may be largely responsible for the biological effects of soy consumption. This is of particular interest, given that only 20-40% of the population produces this metabolite after soy consumption. Studies have shown that equol producers have hormone profiles consistent with lowered risk of breast cancer, and that in some cases equol producers may respond better than non-producers when it comes to soy benefits on indicators of cardiovascular disease, bone health, cancer risk, and menopausal symptoms. At the same time, other studies have shown no differences in effects of soy consumption between subjects who produce equol and those who do not. Thus, the relevance of equol production remains controversial, although the wide variability in production is not. Since equol is produced via bacterial metabolism, it has generally been assumed that this wide variation in equol production is due to differences in colonic microflora. If equol is critically important, the large variability in equol production could explain some of the inconsistencies in results among studies that have evaluated the biological effects of soy consumption. In addition, if it is true that equol production improves response to soy, it becomes even more important to understand the factors that influence isoflavone metabolism, and to determine whether it is possible to turn an equol non-producer into a producer. This talk will briefly summarize the factors influencing equol production, its mechanisms of action, and data related to the equol hypothesis. Equol: A Comparison of the Effects of the Racemic Compound with that of the Purified S-Enantiomer on the Growth, Invasion and DNA Integrity of Breast and Prostate Cells in vitro. P. Magee1, M. Raschke2, C. Steiner2, J. Duffin1, B. Pool-Zobel2, T. Jokela3, K. Wahala3, and I. Rowland*2, 1University of Ulster, Coleraine, UK, 2Friedrich Schiller University, Jena, Germany, 3University of Helsinki, Helsinki, Finland. The ability of soy isoflavones to protect against hormone-dependent cancers, such as those of the breast and prostate has been intensively studied to date. Equol, an isoflavan, is produced by approximately 3040% of the population via the bacterial conversion of the soy isoflavone daidzein in the gut. Due to the presence of a chiral centre, equol exists as two distinct optically active isomers, R- and S-equol, though the enantiomer produced by metabolic reduction from isoflavones is known to be S-(-)equol. It has been postulated that the R- and S- enantiomers have different biological properties, given their different binding affinities for the estrogen receptor. In this study we have compared the biological effects of the purified S-equol enantiomer to that of racemic (R and S) equol on breast and prostate cancer cells in vitro. Effects on cell proliferation, invasion through matrigel and DNA integrity using the Comet assay were studied. Both racemic and S-equol inhibited the growth of the breast cancer cell-line MDA-MB-231 (>10mM) and the prostate cancer cell-lines LNCaP (>5mM) and LAPC-4 (>2.5mM) with both compounds exerting similar effects as assessed by DAPI staining. The compounds also showed equipotent effects in inhibiting the invasion of MDA-MB-231 and PC-3 cancer cells through matrigel. S-equol (1, 10, 30mM) was, however, unable to prevent DNA damage in MCF-7 or MCF-10A breast cells follow- 3 2005 Abstracts Probiotic and Prebiotic Effects on Soy Isoflavone Metabolism, Equol, and Lipids. T. Larkin1,3, L. Astheimer1,3, and W. Price3, 1Department of Biomedical Science, University of Wollongong, Wollongong, NSW, Australia, 2Department of Chemistry, University of Wollongong, Wollongong, NSW, Australia, 3Smart Foods Key Centre, Wollongong, NSW, Australia. Due to the essential role of gut microflora in isoflavone metabolism, particularly that to equol, it was hypothesized that concurrent intake of soy with either a probiotic (yoghurt) or prebiotic (resistant starch) may affect isoflavone metabolism and lipids. Thirty-one hyperlipidemic men and postmenopausal women completed this randomised, crossover study of two 5-week dietary periods separated by a 4-week washout. Intake of soy alone was compared with either probiotic (soy + probiotic) or prebiotic (soy + prebiotic) treatments for effects on plasma and urinary isoflavones and plasma lipids following a test soy meal. Five weeks of soy intake significantly increased circulating plasma daidzein and genistein levels, but did not affect plasma or urinary isoflavones after the test soy meal. There was a trend for increased plasma daidzein and genistein levels with both probiotic and prebiotic treatment, but no overall significant effects. Neither treatment induced or increased equol production, though there was a trend with probiotic treatment for increased plasma equol in equol-positive subjects (n = 12). Despite a lack of significant effects on isoflavone bioavailability, total cholesterol was significantly decreased with both probiotic and prebiotic treatment (-4.7 ± 2.0%, p = 0.038 and -5.5 ± 1.6%, p = 0.003, respectively) and LDL cholesterol was most significantly decreased with prebiotic treatment (-7.3 ± 2.2%, p = 0.005). Independent of the potential for probiotic or prebiotic treatment to affect isoflavone bioavailability, the synergistic action between soy and probiotic or prebiotic intake for lipid-lowering effects suggests these combinations may be useful in lipid management. • Specialty Cereals Pty Ltd., Paul’s Dairy, So Natural Foods, Penford Australia, Dr. Barbara Meyer, Maria de los Angeles. Urinary Excretion of Equol and the Risk of Breast Cancer in Japanese Women. C. Nagata1, T. Ueno2, S. Uchiyama2, K. Urata2, Y. Nagao3, C. Shibuya3, Y. Kashiki3, and H. Shimizu1, 1Gifu University Graduate School of Medicine, Gifu, Japan, 2Saga Nutraceuticals Research Institute, Otsuka Pharmaceutical Co., Ltd., Saga, Japan, 3Gihoku General Hospital, Gifu, Japan. The association between urinary excretion of equol and the risk of breast cancer was examined in a case-control study among Japanese women. The case subjects were 160 women newly diagnosed and histologically confirmed with breast cancer between 2000 and 2002 at a general hospital in Japan. A total of 456 controls individually matched to cases on age, menopausal status, and the date of sample collection were selected from those who attended a breast cancer mass screening at this hospital. Diet before the date of diagnosis for the cases and the date of the screening visit for the controls was assessed by a validated semiquantitative food frequency questionnaire. Spot urine samples from each subject were analyzed for urinary excretion of isoflavonoids. Urine samples from breast cancer cases were collected before cancer therapy. Dietary intake of soy products or soy isofolavones did not differ significantly between cases and controls. The percentage of women with detectable levels of equol (>1.13nmol/ml) in urine was significantly greater in controls than that in cases (19.6 vs 8.2%). Urinary levels of daidzein and genistein were significantly higher in cases than in controls. The status of excretion of equol (+ or -) was moderately positively associated with urinary levels of daidzein and genistein in controls. Urinary equol or the ability to produce equol may be associated with the risk of breast cancer. Treatment of Postmenopausal Monkeys with Equol Did Not Improve the Plasma Lipid Profile. S. Appt, T. Clarkson*, and H. Chen, Wake Forest University School of Medicine, Winston-Salem, North Carolina, USA. Background: Monkeys fed soy protein, compared to casein, have markedly improved plasma lipid profiles (increased HDLC and decreased LDL+VLDLC). Soy consumption results in high plasma equol concentrations, which might account for the large lipid benefits in monkeys compared to much smaller benefits observed in women in (only about a third of which produce equol). Materials and Methods: Fifteen postmenopausal monkeys, all fed a casein-lactalbumin based diet containing some added cholesterol (0.20 mg/Cal), were randomized into a control group (n=7) or an equol group (n=8). Racemic equol was added to the diet of the equol group in an amount in which the s-anisomer would be equivalent to the amount of daidzein in our previous soy protein studies (52.4 mg/1,800 Cals). Plasma HDLC, LDL+VLDLC and triglycerides (TG) were measured at baseline, 3, 6 and 8 months. Repeated measures ANCOVA, adjusting for baseline, was used to analyze the data. Results: Postprandial total plasma equol concentrations were about 470 nmol/L, comparable to monkeys fed soy protein containing daidzein equivalent to the equol s-anisomer fed in this study. After 8 months of treatment, there were no significant effects of equol on HDLC, LDL+VLDLC or TG. Discussion: We found no evidence that equol per se is the mediator of the plasma lipid benefits of soy protein fed to monkeys. There are limitations to the study: synthetic equol may be handled differently by the gut, may act differently administered with soy protein or the effects may be due to dehydrodaidzein or tetrahydrodaidzein. Cancer Developing a Soy Food Based Intervention Among Healthy Men. G. Maskarinec1, S. Hebshi1, Y. Morimoto1, S. Sharma1, A.A. Franke1, and F.Z. Stanczyk2, 1Cancer Research Center of Hawaii, Honolulu, HI, USA, 2University of Southern California, Los Angeles, CA, USA. Increasing prostate cancer risk among men who migrated from Asia to the United States led to the hypothesis that soy foods may be protective through an effect on circulating sex steroids. This pilot study Oral Presentation Abstracts ing exposure to 2-hydroxy-4-nonenal, menadione or benzo(a)pyrene-7,8dihydrodiol-9,10-epoxide. In contrast, DNA damage in MCF-10A cells was significantly reduced following exposure to 2-hydroxy-4-nonenal or menadione when cells were pretreated with racemic equol (10, 30mM). These findings suggest that racemic equol has strong antigenotoxic activity in contrast to the purified S-equol enantiomer implicating the R-, rather than the S-enantiomer as being responsible for the antioxidant effects of equol, a finding that may have implications for the in vivo chemoprotective properties of equol. This study was carried out with financial support from the Commission of the European Communities specific RTD programme “Quality of Life and Management of Living Resources” (project no QLK2000-00266). It does not necessarily reflect its views and in no way anticipates the Commission’s future policy in this area. 6th International Symposium on the Oral Presentation Abstracts 4 Role of Soy in Preventing and Treating Chronic Disease examined the feasibility of recruiting healthy men for a soy food based intervention, developed a dietary protocol that could achieve compliance, obtained preliminary hormonal data, and assessed emotional status during the intervention. Invitations were sent to 1000 Kaiser Permanente Hawaii members, ages 40-69 years, who had a normal PSA test. We received replies from 90 (9%) interested men and identified 25 eligible subjects. After a run-in period, 24 men (92% of eligible) were randomized and 23 subjects completed the study. In a cross-over design, half of the subjects completed either a high soy diet or maintained their usual diet. In the high soy diet, dairy, meat, and snacks were replaced with approximately 75mg of isoflavones as 2 servings of soy milk, tofu, or soy nuts. After 3 months and a 1-month washout period, participants crossed over to the other diet. Compliance was assessed by 24-hour recalls, soy logs, and urinary isoflavone excretion measured by high pressure liquid chromatography. Each subject donated 3 blood samples and 5 overnight urine samples and completed the Profile Of Mood States questionnaire. Testosterone and PSA were quantified in serum by highly specific immunoassays. The intervention and all laboratory analyses were completed recently. The statistical analysis will apply t-tests and mixed models to examine differences by diet while incorporating the repeated measures. • This pilot stuy was supported by a developmental grant from the Cancer Research Center of Hawaii. Soy Intake, Use of Menopausal Hormones, Body Size, and Breast Cancer Risk in Asian-American Women. A. Wu1, M. Yu2, C. Tseng1, and M. Pike1, 1University of Southern California, USA, 2University of Minnesota, USA. We conducted a large population-based case-control study of breast cancer among Asian American women in Los Angeles County which included 1386 women with breast cancer (494 Chinese, 384 Japanese, 508 Filipino) and 1226 control women without breast cancer (515 Chinese, 331 Japanese, 380 Filipino). Participants provided detailed information on menstrual and reproductive factors, use of exogenous hormones, family history, body size, dietary history, and other lifestyle factors in a structured in-person interview. Established risk factors such as nulliparity, family history of breast cancer, and use of menopausal hormones are significant risk factors in Asian-American women as in other western populations. Risk of breast cancer is significantly inversely associated with soy intake during adolescence and adult life; the significant risk reductions were found in both premenopausal and postmenopausal women after adjustment for relevant dietary and non-dietary variables. However, further analyses in postmenopausal women showed that the benefits of soy intake are found only in certain subgroups; namely women with low body size and non-users of menopausal hormones. These results and our interpretation of the findings will be discussed. This work was supported by grants (1RB-0287, 3PB-0102, 5PB0018) from the California Breast Cancer Research Program. Incident breast cancer cases for this study were collected by the USC Cancer Surveillance Program (CSP), which is supported under subcontract by the California Department of Health. The CSP is also part of the National Cancer Institute’s Division of Cancer Prevention and Control Surveillance, Epidemiology, and End Results Program, under contract no. N01CN25403 Antiestrogen Effects of Soybean Glyceollins in Postmenopausal Monkeys. C. Wood1, S. Appt1, T. Clarkson1, A. Franke2, S. Boue3, M. Burow4, and J.M. Cline*1, 1Wake Forest University School of Medicine, Winston-Salem, NC, USA, 2Cancer Research Center of Hawaii, Honolulu, HI, USA, 3Southern Regional Research Center, United States Dept. of Agriculture, New Orleans, LA, USA, 4Tulane University School of Medicine, New Orleans, LA, USA. Estrogens have an important role in the development of breast and uterine cancer. Glyceollins are isoflavonoid pterocarpans, present in stressed soybeans, which have potential estrogen-antagonist effects. In this short-term study we used a primate model to evaluate the antiestrogen effects of glyceollin-enriched soy protein (GLY) on biomarkers for cancer risk. Thirty ovariectomized adult female cynomolgus macaques were randomized to one of three dietary treatments (expressed as equivalent doses for women): (1) estradiol (E2, 1 mg/day) + casein lactalbumin (control); (2) E2 + soy protein isolate (SPI) containing 180 mg/day isoflavones; and (3) E2 + GLY containing 189 mg isoflavones and 134 mg glyceollins (I-III). Experimental diets were given for 3 weeks. Breast and uterine measures were taken before and after each treatment. Serum glyceollin concentrations were 134.2 nM (40.0% of daidzein+genistein) 4 hours after feeding and <1 nM 24 hours after feeding. Compared to baseline, E2 treatment induced significantly greater breast proliferation in the control group (p=0.02 in lobules; p=0.002 in ducts) and SPI group (p=0.04 in lobules; p=0.02 in ducts) but not in the GLY group (p=0.09 in lobules; p=0.93 in ducts). E2 significantly increased uterine area in all groups (p<0.01 for all); however, the change in uterine area was significantly lower in the GLY group versus the control (p=0.01) and SPI (p=0.049) groups. These findings demonstrate that soybean glyceollins are absorbed and rapidly cleared following dietary intake. Preliminary data suggest that glyceollins may enhance the antiestrogenic properties of standard soy protein isolate in the breast and uterus. • This work was supported by the NIH/NCCAM R01-AT00639 (JMC). Phytoprevent: A European Project on the Prevention of Breast and Prostate Cancer by Phytoestrogens. I. Rowland, University of Ulster, Coleraine, UK. Phytoprevent is a 3 year multi-disciplinary project funded by the European Union with the aim of investigating the role of foods rich in phytoestrogens (isoflavones and lignans) on reduction of risk of breast and prostate cancer. In analytical studies, HPLC CoulArray methods have been validated for the analysis of food, plasma, urine and faecal and an ID-GC-MS-SIM method has been established for the measurement of plasma estrogen/phytoestrogen levels. In vitro methods were used to study effects of isoflavones on various stages of cancer. Preincubation of breast and prostate cell lines with low concentrations of isoflavones decreased hydrogen peroxide-induced DNA damage. Isoflavones were found to significantly enhance tight junction integrity in the breast cancer cell-lines MCF-7 and MDA-MB-231 suggesting a beneficial effect on tumor promotion. Isoflavones inhibited invasion of MDA-MB-231 breast cancer cells through Matrigel at concentrations as low as 2.5uM. The anti-angiogenic activity of 16 synthetic phytoestrogen metabolites has been determined. Two synthetic phytoestrogens were shown to possess anti-angiogenic activity using proliferation and differentiation assays. Studies in the Tg.NK transgenic mouse mammary cancer model showed that perinatal exposure to isoflavones had no effect on tumor development, while post weaning and life long exposure to isoflavones delayed tumor development and reduced tumor incidence. In a human intervention study (22 days) with foods containing soy, changes in urinary estrogen profile were seen in the soy consumers that were consistent with lower breast cancer risk. This study was carried out with financial support from the Commission of European Communities specific RTD programme 5 2005 Abstracts The Combination of Soy and Flaxseed or Their Phytoestrogens Can Better Reduce the Growth of Breast Tumors than Soy or Genistein Alone While Causing Little Effects on Bone Health in Ovariectomized Nude Mice. L. Thompson, K. Power, N. Saarinen, J. Chen, and W. Ward, Department of Nutritional Sciences, University of Toronto, Toronto, ON, Canada. Phytoestrogen (PE)-rich diets have been associated with a lower risk of breast cancer (BC) and osteoporosis. In previous studies in ovariectomized (OVX) mice, flaxseed (FS), rich in lignan PE, reduced the growth of human MCF-7 estrogen receptor positive (ER+) BC, while the soy isoflavone PE, genistein (GEN), stimulated tumor growth. In this study, we determined whether the combination of soy and FS or their respective PE, isoflavone genistein (GEN) or the lignans enterolactone (EL) or enterodiol (ED), can more effectively reduce the growth of ER+ BC in OVX mice than soy or GEN alone while maintaining protective effects on bone. OVX mice with established MCF-7 tumors were fed the basal diet, 20% soy protein isolate (SPI), 10% FS, or 20% SPI + 10% FS for 25 weeks. All mice exhibited tumor regression but SPI induced late stage tumor growth, which was negated by combining it with FS. SPI maintained femur bone mineral density (BMD) and strength but FS slightly attenuated this effect when combined with SPI. Using the same model, mice with MCF-7 tumors were given daily injections of GEN, EL, ED, EL+ED+GEN, or vehicle control for 22 weeks. All mice showed tumor regression but GEN caused the least effect. The effect of combined PE on bone did not differ from the control and GEN. Tumor cell proliferation and apoptosis data agreed with the observed changes in tumor size. In conclusion, combining PE or PE-rich foods resulted in a better control of tumor growth while causing little effects on bone health. • Funded by NSERC Tuesday, November 1, 2005 Cardiovascular Disease Hypocholesterolemic Effect of Soy Proteins—Only Due to Protein and Only in Hypercholesterolemics. Cesare R. Sirtori, Dept. of Pharmacological Sciences and University Center for Dyslipidemias, Niguarda Hospital, Milano, Italy. The concept that animal proteins may be atherogenic versus the case of vegetable proteins dates back to Ignatowsky (1908) and had a formal proof in 1977 in a “model” study. This was: a) carried out in hypercholesterolemic individuals (lowest starting cholesterolemia 270 mg/dl); b) on in-patients, followed in metabolic ward conditions; c) based on a direct comparison according to a cross-over protocol of an exclusively soy protein and an animal protein based diet (Sirtori et al, Lancet 1977; i: 275). In a second part of the study the addition of cholesterol was tested on similar in-patients on the soy diet, proving no change in the remarkable cholesterol lowering activity of the experimental regimen (ie around 23% LDL-C reduction). No other study ever fulfilled all these qualifications. In a larger outpatient study in Italy/Switzerland the same soy product (later found to be phytoestro- gen-free) was given to 130 patients all with cholesterolemia >300 mg/dl (Descovich et al 1980; ii: 709); both similar reductions of total and LDL-C levels as well as a clear correlation between these reductions and baseline cholesterolemia were reported. A large number of studies have been, unfortunately, carried out in normolipidemic individuals with, as expectable, negative findings; in addition, essentially all published studies have been on dietary supplements. The cholesterol lowering mechanism of soy protein is by way of stimulation of LDL-receptor activity, as shown in animal (Lovati et al, J Nutr 2000; 130:2543) and in clinical studies (Lovati et al, J Clin Invest 1987; 80: 1498). LDL-receptor activity is normal in normolipidemic individuals, unlikely to respond to this treatment. The mechanism thus differs from that of statins, working on cholesterol biosynthesis and active also at low levels of cholesterolemia, and is somewhat similar to that of berberine, also an LDL-receptor upregulator (Kong et al, Nature Med 2004 ; 10: 1344). A matter of confusion in the field was the belief that phytoestrogens might be in some way responsible for cholesterol lowering. This belief was based on primate studies and also on human findings using ethanol extracted soy products. This drastic chemical treatment most likely destroys active protein components, since a more appropriate column extraction procedure maintains the hypocholesterolemic activity in the absence of phytoestrogens (Fukui et al, J Agric Food Chem 2002; 50: 5717). Due to this ill fated belief, US distributed soy proteins show grossly altered structure, as assessed from a proteomic evaluation (Gianazza et al, J Nutr 2003; 133: 9); surprisingly they still cause some reduction of cholesterolemia. A reputable meta-analysis (Anderson et al, N Engl J Med 1995; 333: 276) clearly showed that soy proteins (mainly supplements) reduce cholesterolemia to a minimal extent in normolipidemic individuals (mean reduction -5.2 mg/dl; - 3.3% ) versus a high effect in patients with marked cholesterol elevations (up to -71.5 mg/dl; - 19.6%). This type of findings allowed the preparation of a “predictive nomogram” for assessing the individual benefit of treatment: the cholesterol reductions reported in recent American studies (Lichtenstein et al, ATVB. 2002; 22: 1852; Jenkins et al, JAMA 2003; 290: 502) fit well with the nomogram. It is thus unfortunate that a recent US Report (Evidence Report Technology Assessment, No.126, Effects of Soy on Health Outcomes), pooling together all sorts of studies and eliminating many (including our early ones) came to negative conclusions. Indeed the majority of the studies were in normolipidemics, frequently treated with chemically unsuitable proteins. Superficial comments such as “Evidence suggests that initial estimates on the potency of soy protein were too high” have no basis: there was never an estimate, since it was clear from the onset that the cholesterol lowering activity is based first on the patient characteristics, second on the soy product and third, most likely, on the daily amount. Soy proteins are active only in hyperlipidemics; this may reduce their appeal, ie considering the present indication of statins in normolipidemic coronary patients. It leaves, however, ample space for clinical use (children, renal patients, etc.). In addition, similar to the case of milk proteins providing ACE-inhibitory peptides for hypertension (FitzGerald et al, J Nutr 2004; 134: 980S), it should not be long before soy protein varieties from adequate cultivars or fractions thereof may turn out to provide effective “pill like” cholesterol reducers. 7S Soy Globulin Peptide and Cholesterol Homeostasis: in vitro and in vivo Data. M. Lovati, C. Manzoni, S. Castiglioni, and E. Disconzi, Dept. of Pharmacological Sciences, University of Milano, Milano, Italy. 7S soy globulin and its alfa’ subunit have been shown to positively modulate LDL receptor activity and to decrease triglyceride synthesis Oral Presentation Abstracts ‘Quality of Life and Management of Human Resources’ (Project No QLK1-2000-00266.It does not necessarily reflect its views and in no way anticipates the Commission’s future policy in this area 6th International Symposium on the Oral Presentation Abstracts 6 Role of Soy in Preventing and Treating Chronic Disease both in vitro (HepG2 cells) and in vivo (rats fed cholesterol-rich diet). The aim of the present investigation was to evaluate the effect of a synthetic peptide, corresponding to a sequence which differs between the alfa and alfa’ subunits from 7S soy globulin, on cell cholesterol homeostasis both in HepG2 cells and in rats fed casein-cholesterol diet. HepG2 cells were incubated for 24 hr in minimum essential medium (MEM) + 5% LPDS in the presence/absence of whole 7S globulin (0.5 mg/mL) or its alfa’ subunit (0.125 mg/mL) or the synthetic peptide (105 M). At the end of incubation, cell membranes and nuclei were separated by validated procedures. Aliquots of membrane and nuclear fractions were run on 7.5-15% SDS-PAGE gels, transferred onto nitrocellulose, and overlaid with a polyclonal antibody against SREBP-1. Cell preincubation with whole 7S soy globulin caused an increase in the mature forms of SREBP-1, both in membranes (+ 90%) and in the nuclei ( + 130%), vs that found in the control HepG2 cells (MEM + 5% LPDS). Moreover, a lower level of mature nuclear SREBP-1 was detected in HepG2 cells exposed to both the alfa’ subunit and the synthetic peptide, compared to that found after exposure to whole 7S globulin. The oral administration (14 days) of synthetic peptide (5 mg/Kg b.w.) to caseincholesterol fed rats resulted in lower plasma lipid levels (cholesterol, 25%; triglycerides, -28%) vs values recorded in rats fed casein-cholesterol diet alone. These results represent the first in vivo evidence of the plasma lipid-lowering properties induced by the synthetic peptide; moreover, these data suggest that alfa’ subunit from 7S soy globulin, and its synthetic peptide, are responsible for the direct up-regulation of LDL receptors. The mechanism of this effect is likely to be through the SREBP-2 transcriptional factor, the activity of which we are currently investigating. • Supported in part by a grant from MIUR of Italy. Effect of Two Types of Soy Milk and Dairy Milk on Plasma Lipids in Hypercholesterolemic Adults: A Randomized Trial. C.D. Gardner1, J. Morris1, M. Messina2, A. Kiazand1, A. Varady1, and A. Franke3, 1Stanford University, Stanford, California, USA, 2 Loma Linda University, Loma Linda, California, USA, 3Cancer Research Center of Hawai‘I, 1236 Lauhala St., Honolulu, HI 96813, USA. Background–The literature on the hypocholesterolemic effect of soy proteins is inconsistent. The effect of soy proteins may differ by dose and by the level of hypercholesterolemia. Objective–To contrast plasma lipid responses between two commercially available soy milks, one made using whole soy beans and the other using soy protein isolate, and low-fat dairy milk. Design–Randomized clinical trial, cross-over design. Patients–Adults (n=28), 30-65 years of age, with pre-study LDLcholesterol concentrations of 160-220 mg/dL, not on lipid lowering medications and with overall Framingham risk score of< 10%. Intervention–Participants were required to consume sufficient milk to provide 25 g protein/day from each milk source, preferably distributed over three times/day. The protocol included 3 four-week milk consumption phases, each separated from the next by wash-out periods of at least four weeks. Results–The average LDL-C concentration at the end of each phase (mean +/- SD) was 161 +/- 20, 161 +/- 26 and 170 +/- 24 mg/dL for the whole soy bean milk, the soy protein isolate milk, and the dairy milk, respectively (p=0.9 between soy milks, p=0.02 for each soy milk vs. cow’s milk,). No significant differences by type of milk were observed for HDL-cholesterol or triacylglycerols. The only pair-wise difference in total-C/HDL-C ratio was a significantly lower ratio for the soy protein isolate milk relative to the dairy milk (p=0.02). Conclusion–The results of this study support a modest, 5% LDLcholesterol lowering effect of either type of soy milk, relative to dairy milk, when provided in amounts yielding 25 g protein/day to adults with elevated LDL-cholesterol concentrations for four weeks. Soy Food Effects on Serum Lipoproteins in Humans: Updated Meta-Analysis. J.W. Anderson, VA Medical Center and University of Kentucky, Lexington, KY, USA. Soy foods significantly affect serum lipoproteins by decreasing LDL-cholesterol (LDL-c) and triglycerides (TG) while increasing HDL-cholesterol (HDL-c). Since our 1995 meta-analysis, most studies have reported less impressive alterations in serum lipoproteins than the 12.9% decrease in LDL-c then reported for soy foods. Since 1995 at least 55 randomized controlled trials with soy foods have reported serum lipoprotein changes. To examine possible causes of differences in lipoprotein responses, we have analyzed these studies with a focus on LDL-c. Weighted changes for different soy foods are presented as changes from baseline (to estimate maximum effects) and net changes (adjusted for control diet changes). In 55 trials, soy protein consumption averaged 44 g/d fed for an average of 29 days. Serum LDL-c decreased 8.5% with net change of 4.5% (95% CI, 2.8-6.3%). In 16 studies soy was baked or cooked with LDL-c changes of -0.9% and net changes of -1.4% (not significant). Using soy protein once daily and <25 g/d was less effective than use of > 25 g/d and > 2x/d. In 24 studies with 25-50 g/d soy protein given > 2x/d, LDL-c reduction was 9.3% with a net reduction of 5.4% (95% CI, 4.1-6.8%). These studies reported significant changes in HDL-c (+3.7%, 95% CI, 0.4-4.1%) and TG (-8.0%, 95% CI, -2.8 to -13.1%). Further subgroup analyses will be presented. Recent studies indicate that soy peptides have potent hypocholesterolemic effects and that soy proteins with more intact peptides have significantly greater hypocholesterolemia than more fragmented preparations. Soy protein extraction or baking may fragment the most active hypocholesterolemic peptides. Effect of Soy Isoflavone Protein and Soy Lecithin on Endothelial Function in Healthy Postmenopausal Women. Z. Faridi, M. Evans, V. Njike, M. Hoxley, and D. Katz, Yale Griffin Prevention Research Center, Derby, Connecticut, USA. Objective: To assess the effects of soy isoflavone protein concentrate and soy lecithin on endothelial function measured as flow mediated dilation (FMD) of the brachial artery in healthy postmenopausal women. Design: Randomized, double-blind, placebo controlled crossover trial. Subjects/Setting: 25 subjects (mean age 61 years; BMI 25.46 kg/m2) were recruited from the general population of Southwestern Connecticut. Intervention: Subjects underwent endothelial function testing at baseline and following 4 weeks of randomly assigned treatments with intervening 4-week washout periods. Treatment assignments included: Soy isoflavone protein (25gm/day) and soy lecithin (20gm/day); soy isoflavone protein (25gm/day) and placebo lecithin; placebo protein and soy lecithin (20gm/day); and double placebo. Outcome Measures: FMD of the brachial artery, and serum lipid levels Results: 22 women completed the trial; pre-treatment FMD was 8.60 ± 7.20. No statistically significant difference was seen in FMD between treatment assignments. However, FMD was highest after treatment with soy protein plus lecithin (7.50 ± 9.85), followed by soy protein plus placebo lecithin (5.51 ± 10.11), placebo protein plus soy lecithin (5.35 ± 6.13), and lowest after double placebo (4.53 ± 7.84). 7 2005 Abstracts Workshop Presentation Role of Soy Foods in the Management of Obesity and Related Chronic Diseases: Summary of a Symposium at the University of Illinois. John W. Erdman Jr., Keith Cadwallader and Barbara P. Klein, Department of Food Science and Human Nutrition, University of Illinois, Urbana, IL, USA. Obesity among American adults has doubled over the past two decades and has nearly tripled for children and adolescents. The consequences of obesity include increased risk of a host of chronic diseases including diabetes, cardiovascular disease, cancer and bone and joint degeneration. Enhanced obesity rates are not limited to westernized countries, as most societies world-wide are now reporting higher proportions of severely overweight citizens. Soy foods have the potential for assisting in obesity management and reduction of subsequent chronic disease development. To address these issues, a two-day forum was held on May 25 and 26, 2005 at the University of Illinois featuring leading researchers from universities and the food industry. They reviewed the current state of knowledge regarding the management of obesity and related diseases and the roles of soy and its components. Presentations focused upon the state of knowledge of obesity incidence, effectiveness of weight loss programs, and impacts of obesity on cardiovascular disease, diabetes and other co-morbidities. It is well accepted that consumption of soy can improve serum lipid profiles. There is emerging evidence suggesting that soy-containing isoflavones may also affect adipogenesis and may be beneficial for diabetics. An industry panel discussed their experiences in utilization of soy foods for weight loss. This presentation will review the scientific outcomes from the symposium. In addition, gaps in current knowledge for the roles of soy in the management of obesity and related diseases will be discussed. Immune Function Soy Isoflavones Modulate Immune Function in Healthy Postmenopausal Women. T. Ryan-Borchers, J. Park, B. Chew, M. McGuire, L. Fournier, and K. Beerman, Washington State University, Pullman, WA, USA. Background: After menopause, the immune system may be compromised due to effects of aging and diminishing concentrations of estrogen, an immune-modulating hormone. Isoflavones, plant-derived compounds that possess antioxidant, anti-estrogenic and pro-estrogenic actions, may offer immunological benefits to women in this stage of life. Objective: The objective of this study was to evaluate the effects of soy isoflavones, both soymilk and supplement form, on markers of immunity and oxidative stress in postmenopausal women. Design: Postmenopausal women aged 50-65 y (n = 52) enrolled in this 16 wk double-blind, placebo-controlled trial were randomized to one of three experimental groups: 1) Control - 706 mL/d cow’s milk plus a placebo supplement; 2) Soymilk - 71.6 mg isoflavones derived from 706 mL/d soymilk plus a placebo supplement; and 3) Supplement - 70 mg isoflavones in a supplement plus 706 mL/d cow’s milk. Plasma and 24-h urine samples were obtained at baseline and at 16 wk. Immune variables included lymphocyte subsets, cytokine production, as well as markers of inflammation and oxidative damage. Results: Baseline immune variables did not differ among groups. Both isoflavone interventions increased (p < 0.05) β cell populations and concentrations of plasma interferon-gamma (IFN-γ) and decreased (p < 0.05) concentrations of 8-hydroxy-2- deoxy-guanosine (8-OHdg), an oxidative marker of DNA damage. Isoflavone treatments did not influence concentrations of urinary 8-isoprostane (8-iso), plasma interleukin-2 (IL2), tumor necrosis factor-alpha (TNF-α) or C-reactive protein (CRP). Conclusions: Soymilk and supplemental isoflavones modulate β cell populations and IFN-γ concentrations, and appear protective against DNA damage in postmenopausal women. Supported by a grant from the Washington State Attorney General’s office; soymilk provided by White Wave, Inc. and isoflavone (Novasoy®) and placebo supplements provided by the Archer Daniels Midland Co.We thank Bridgett Mathison for lab assistance; Heather Gardner, Livia Wagner and April Broderick for delivery of study treatments and data entry; and all of the women who graciously participated in this study. Effects of ImmuSoy as a Food Supplement for Altering Peanut Allergic Reactions. T. Zhang1, W. Pan2, M. Takebe3, H. Sampson1, and X. Li*1, 1Pediatrics, Allergy & Immunology, Mount Sinai School of Medicine, New York, NY, USA, 2Surgery, Harvard Medical School, Beth Israel Deaconess Medical Center, Boston, MA, USA, 3Nichimo Co., Ltd., Tokyo, Japan. RATIONALE: Probiotic foods were only recently clinical trials conducted to evaluate probiotics in the treatment of atopic dermatitis and food allergy, Lactobacillus rhamnosus GG (LGG) had some effect on the prevention of early atopic disease in high-risk infants. Recent studies shown that soy as a kind of allergen-specific immunotherapy can be used to desensitize/downregulate peanut-specific response in peanutallergic mice. ImmuSoy is a koji fungus (Aspergillus oryzae) and lactobacteria soybean fermentation product, based on traditional Japanese fermentation technology. It is believed that this unique food supplement would be beneficial for the immune system when food allergy happened. We hypothesized that ImmuSoy may be effective for treating peanut allergy. METHODS: This study was undertaken in mice to investigate whether specific doses of ImmuSoy would has better effect than LGG for desensitization peanut allergic mice. Peanut allergic mice (C3H/HeJ, n=8-10/group) were fed ImmuSoy containing chow (0.5 % and 1 %), LGG 0.5% containing chow or regular chow (control) for 4 weeks beginning at week 10 following peanut sensitization, and then were challenged with peanut. Anaphylactic scores, plasma histamine, serum peanut specific-IgE levels and splenocyte cytokine production to peanut stimulation were determined. RESULTS: All control mice developed anaphylaxis (median score 3.3) following peanut challenge, in contrast, 50% and 25% of low and high doses of ImmuSoy treated-mice developed anaphylaxis (median scores 1.0 and 0.33 respectively) while LGG treated mice have 80% and mean score around 2. The low and high dose ImmuSoy treated-mice showed 50.4 % and 80.7% reduction respectively in plasma histamine(LGG 20%), and 47.1% and 73.9% reduction respectively in serum IgE levels as compared to untreated mice (p<0.05)(LGG 20% P>0.05). Oral Presentation Abstracts Soy isoflavone protein and soy lecithin significantly increased HDL/LDL relative to baseline value (soy isoflavone protein and soy lecithin, 0.64 ± 0.19; soy isoflavone protein and placebo lecithin, 0.58 ± 0.17; placebo protein and soy lecithin, 0.65 ± 0.18; baseline, 0.49 ± 0.15). Conclusion: In this sample of healthy postmenopausal women, soy isoflavone protein and soy lecithin significantly improved the lipid profile. Improvement in endothelial function could not be confirmed. The study was funded by a grant from Health & Nutrition Group of Eridania Béghin-Say America, Inc. 6th International Symposium on the Oral Presentation Abstracts 8 Role of Soy in Preventing and Treating Chronic Disease Furthermore, IL-4, IL-5 and IL-13 production by splenocytes of high dose ImmuSoy-treated mice were reduced by 74.1% and 77.0% (LGG 75%) whereas IFN-γ production was increased by 34.8%(LGG increased 10%) compared to control splenocytes. CONCLUSIONS: LGG of 0.5% containing chow shown some protective effects on peanut allergic mice, but when we increase the dose of LGG even to 5 times higher than ImmuSoy 1%, no dose dependent or no better effects shown (data not shown here). Compared with LGG, ImmuSoy used as a food supplement has dose dependent protective effects against peanut-induced anaphylaxis in mouse peanut allergic model, which is associated with down downregulation of Th2 responses. This approach might be a potential novel therapy for peanut allergy. Diabetes Habitual Soyfood Consumption Improves Glycemic Control Among Postmenopausal Chinese Women: A One-Year Follow-up Study. S. Ho and Y. Chen, The Chinese University of Hong Kong, Hong Kong SAR. Objectives: Previous studies show soy supplementation improves glycemic control in animals and in diabetic subjects. Little data are available on their effects among healthy adults. We examined the effect of habitual soyfood intake on changes in serum fasting glucose over a one-year followup period. Methods: This study included 173 community-based postmenopausal Chinese women aged 48-62 years who had been recruited into a one-year randomized controlled trial of three doses of isolated soy isoflavones (0, 40 and 80 mg/d) on bone changes. Habitual soyfood intake was assessed using a validated food frequency questionnaire. Fasting serum glucose (FG) was measured at baseline and at the end of one-year. Results: The mean intake of habitual soy protein among the study subjects was 5.3g/d (sd = 4.46). Soy protein intake, changes in body mass index, and baseline glucose were significantly correlated with changes of FG over one year. Stepwise regression analysis showed that habitual soy protein was significantly and inversely associated with the absolute and percent changes in FG in women with initial glucose over the median (92.1mg/dl) rather than in those below the median after adjusting for supplemental isoflavones and other potential confounders (including age, years since menopause, baseline FG, changes in body mass index, dietary intakes of total fat, fruits, vegetables, and physical activities). Soy protein accounted for 14.1% (p=0.002) and 12.8% (p=0.003) of the variations of absolute and percent changes of FG, respectively; and it was estimated that a daily increase of ten-gram soy protein was associated with 13.5 mg/dl (or 9.3%) decreases in fasting glucose (p<0.001) in women with higher initial FG. Conclusion: Habitual soyfood intake improves glycemic control in women with higher baseline FG. Beneficial Metabolic Effect of Soy-Rich Diets in Young Firefighter Trainees. Y. Yamori1,2, M. Shibata1, M. Mori3, N. Ishiwata4, M. Tokoro2, and Y. Yamamoto1, 1Hyogo Prefecture Health Promotion Association, Kobe, Japan, 2Mukogawa Women’s University, Nisinomiya, Japan, 3Research Institute for Production Development, Kyoto, Japan, 4Atomi Junior College, Tokyo, Japan. Purpose: To investigate the effect on the indices related to lifestylerelated disease risks, of continuous daily soy-rich diet intake in healthy young Japanese men. Method: A randomized, cross-over controlled dietary intervention study on metabolic effects of soy-rich diets was carried out in cooperation with 100 healthy firefighter trainees aged 18-25. After informed consent was obtained, they were randomized into soy rich (S) and ordinary control (C) diet groups during the first and second intervention periods (45 and 44 days, respectively) with an interval of the washout period of 25 days. S diets consisting of common Japanese soy products, such as tofu, soymilk, etc. contained more soy protein (29.4g in average) a day than regular Japanese C diets (soy protein 8.6g), and both diets were nearly the same in daily total energy (2800-3300 Kcal) and total protein (105-120g) intakes. Questionnaires about dietary custom, body mass index (BMI), body fat (F), blood pressure (BP), and blood and 24-hour urine were examined 4 times before and at the end of both intervention periods. Results: From among 95 male participants, 43 completed all tests including 4 times of 24-hour urine collection. S group (18) showed significant increases in urinary isoflavones (ISO: 19.2→87.4µmol) after the intervention and significant reduction in systolic and diastolic BP, fasting glucose (G), BMI and F. In contrast, C group (25), whose urinary ISO excretions were not changed, showed no significant changes in these parameters. Significant inter-group differences were noted in changes from the baseline levels of G and HOMA-IR (homeostasis model assessment insulin resistance index: G×immuno reactive insulin). Conclusion: Daily intake of soy rich diets containing about 30g of soy protein decreased G and improved HOMA-IR significantly in comparison with regular diets, suggestive of the beneficial effect of soy-rich diets on insulin resistance even in young healthy men. (This study was supported by Hyogo Prefecture Government) Menopausal Symptoms The Effects of 100mg Soy Isoflavone Supplements on Menopausal Symptoms and Quality of Life: A DoubleBlind, Randomised, Placebo-Controlled, Crossover Trial in British Postmenopausal Women. C. Hill1, A. Cassidy2, J. Powell3, D. Talbot3, and L. Dye1, 1Institute of Psychological Sciences, University of Leeds, Leeds, West Yorkshire, UK, 2School of Medicine, Health Policy and Practice, University of East Anglia, Norwich, East Anglia, UK, 3Unilever R&D, Sharnbrook, Bedfordshire, UK. Research into the efficacy of soy isoflavones (soy IF) to relieve menopausal symptoms has yet to produce a unanimous conclusion. Attention has been largely focused on hot flushes, with other psychological, vaso-somatic and general somatic symptoms typically assessed before and after an intervention. The conflicting findings on these endpoints could be attributed to the methodological issues surrounding retrospective reporting and the use of psychometrically weak scales. Therefore this study adopted a prospective assessment of the severity and frequency of menopausal symptoms. Retrospective measurements of quality of life were also taken, which tapped into a non-symptomatic consideration of well being. This study employed a double-blind, randomised, placebo-controlled crossover trial of 100mg soy IF supplement over 8 weeks in 21 British postmenopausal women. No significant difference was noted between soy IF and placebo for total symptom score or for the psychological, vaso-somatic or general somatic subscales. However, specific reductions were observed in the frequency of feeling easily stirred up and the severity of early morning awakenings and sleeplessness. Daily counts and severity measures of hot flushes were also significantly reduced across all weeks of soy IF intervention, with greater efficacy in women with > 5 hot flushes/day. There were no interactions with time on treatment, with a generally lower level of these 9 2005 Abstracts Isoflavone Supplements Predominantly Containing Genistin/Genistein Reduce Hot Flushes: A Critical Analysis of Published Studies. P. Williamson-Hughes1, B. Flickinger1, M. Messina2, and M. Empie1, 1Archer Daniels Midland Company, Decatur, IL, USA, 2Nutrition Matters, Inc., Townsend, WA, USA. Several reviews and meta-analyses have evaluated the clinical evidence relating isoflavone treatment to menopausal hot flush symptom relief. The majority of these reviews and meta-analyses have included a range of isoflavone sources; often without differentiating the chemical identity of individual isoflavone molecules contained in the study product or the compositional ratio of isoflavones. We conducted a re-evaluation of the published literature specific to studies utilizing well characterized isoflavone-containing supplements to determine if observed effects, or lack thereof, were attributable to specific isoflavones. Eleven studies were stratified in relation to their specific isoflavone composition. Six studies described isoflavone products which provided greater than 10 milligram genistein per treatment (calculated as aglycone equivalents), administered to a total of 188 subjects. These six studies consistently reported a statistically significant effect in reducing hot flushes in post-menopausal women. In contrast, the remaining five studies (total of 167 subjects) used supplements containing high daidzein or formononetin and low genistin/genistein content; only one of these (30 subjects) reported an effective reduction in hot flush. Hence, reports finding that the relationship between isoflavone consumption and hot flush reduction inconclusive suffer from over generalization by including any product with the term isoflavones. Lack of discrimination between individual molecular entities contained in heterogeneous isoflavone sources can be misleading. In light of our observations, we conclude that higher genistein ratio extract products with a minimum genistein dose level are effective in reducing hot flushes and that evaluation of isoflavone effects should be related to content of individual isoflavones within supplements. Comparison of Isoflavones and Tibolone Regarding Vaginal Estrogenicity in Peri and Postmenopausal Women. U.D. Rohr1 and A. Jungbauer2, 1AHS, 1090 Vienna, Austria, 2Department of Biotechnology, BOKU, 1090 Vienna, Austria. Isoflavones are promising agents in chemoprevention. Clinical effects in peri- and postmenopausal women of isoflavones and tibolone in postmenopausal women are compared to binding affinities to a yeast estrogen receptor model. Tibolone a compound used in postmenopausal hormone therapy, is metabolized into 3 further active compounds. 80mg isoflavone per day were applied in 11 perimenopausal women and 36 postmenopausal women for 3 month. Competitive radio-ligand binding assays of different dilutions of the Isoflavone preparations resulted in a binding to estrogen receptor (ER)-alpha and beta in a dose-dependent manner. 2.5 mg Tibolone was applied daily in 24 post-menopausal women for 6 months. Tibolone and its delta-isomer did predominantly bind and transactivate progesterone receptor and androgen receptor, whereas its 3alpha-hydroxy- and 3beta-hydroxy-tibolone predominantly bind and transactivate ER-alpha. Isoflavones did show a preference for binding to the ER-beta. The PAP-smear test, to control for inflammation and cancer risk of the vagina and portio of the uterus, was conducted after 3 month application of either tibolone or isoflavones. Isoflavone reduced vaginal PAP-test (superfiscial cells) for perimenopausal women from 37578 to 16758 counts and for postmenopausal women from 49417 to 22875 counts. Tibolone increased cell counts in postmenopausal women in the PAP-test. Hypertrophic effects in the vagina as well as endometrium are believed to be facilitated by the ER-alpha and suppressed by ER-beta: Our in vivo results correlate with our in vitro results: ER-beta affinity of isoflavones could explain the reduction of cells in the PAP smear test and ER-alpha affinity of tibolone may be correlated to the increase of cells in the PAP smear test by tibolone. Wednesday, November 2, 2005 Can Soy Isoflavones Cause Endometrial Cancer? J.M. Cline1, C. Wood1, A. Franke2, D. Bland1, D. Foth3; 1Wake Forest University School of Medicine, USA, 2University of Hawaii, USA, 3University of Cologne, Germany. Estrogens markedly increase the risk of endometrial cancer in women, and uterotrophic effects of estrogens are well-documented in animal models including nonhuman primates. As many investigators have shown, high-dose soy-derived isoflavonoids have estrogen-agonist activity on the uterus in rodent models, including the induction of estrogen-dependent hyperplastic and neoplastic changes. A recent 5-year trial of isolated soy isoflavones given in pill form to postmenopausal women found a slight but significant increase in the incidence of endometrial hyperplasia. This observation and its interpretation should be carefully considered. Human observational studies show no evidence of estrogen agonist effects of soy food consumption, and several short-term trials of soy isoflavonoids in women have not shown any proliferation-inducing effect on the uterus. Similarly, our nonhuman primate studies spanning up to 3 years (~10 years of equivalent human lifespan) have shown no evidence of isoflavonoid-induced endometrial hyperplasia at dietary doses of soy isoflavonoids. We have shown a lack of uterotrophic effect in short-term nonhuman primate studies using up to 10 times the dietary exposure of soy isoflavonoids or purified equol. We have also demonstrated estrogen-antagonist effects of soy isoflavone supplementation on estrogen-induced increases in uterine size, a finding which parallels the pattern of endometrial cancer risk in the Shanghai breast cancer study. Finally, in a 2-year trial of women given soy protein isolate supplementation at 58 mg/day, we found no association between serum isoflavonoid concentrations and endometrial proliferation. These findings give us some confidence that there is not a significant uterotrophic effect of dietary soy isoflavonoids in the human or nonhuman primate endometrium, and in fact that there may be a protective effect in the presence of higher endogenous or exogenous estrogens. However, the potential remains for differences in risk:benefit ratio based on isoflavone formulation or individual characteristics. ACKNOWLEDGEMENTS • The Solae Company; NIH Grants NIH HL45666; NIH AT00639 Oral Presentation Abstracts symptoms across all weeks of soy IF. Health, sexual, emotional and occupational quality of life was not significantly improved with soy IF, and there was no difference between treatments on the Cognitive Failures Questionnaire. However, frequency of sleep problems was significantly reduced after 4 weeks of soy IF, with no further improvements at week 8. Increases in circulating levels of oestrogen were significantly correlated with the reduction of hot flushes, with no clear relationship with other symptoms. Therefore, whilst this study was able to detect some effects on specific symptoms, generally this more methodologically rigorous approach validated findings from earlier methodologically weaker studies. 6th International Symposium on the Oral Presentation Abstracts 10 Role of Soy in Preventing and Treating Chronic Disease Osteoporosis Effects of the Phytoestrogen Genistein on Bone Loss, Cardiovascular Risk Prevention and Climacteric Symptoms: A Two Year Double Blind Placebo Controlled Study. Interim Evaluation at One Year. F. Squadrito, R. D’Anna, F. Corrado, A. Gaudio, M. Atteritano, A. Bitto, D. Altavilla, and N. Frisina, University of Messina, Messina, Sicily, Italy. To confirm our previous findings on the phytoestrogen genistein (1,2) 389 postmenopausal women with a BMD at the femoral neck of <0.795 g/cm2 were randomly assigned to receive either genistein (54 mg/day) (n=198; age = 55±4; BMI= 24±3 ) or placebo (n. 191; age =54±3; BMI = 25±4) in a two years double blind study. Bone loss: Genistein reduced at 12 months the urinary excretion of deoxypiridinoline (-35±13 %; p< 0.01) and increased the serum levels of the bone-specific ALP (18±6%; p<0.01) and bone G1a protein (16±5 %). Genistein increased BMD in the femur and lumbar spine. Placebo treatment did not affect any of those parameters. Cardiovascular risk factors: By comparison with placebo, genistein treatment at 12 months decreased fasting glucose (-14%; p<0.01), fasting insulin (-51%; p<0.01) and HOMA-IR (-43%; p< 0.01) Genistein administration reduced fibrinogen (-18%; p< 0.05), urinary F(2) isoprostanes (24%; p<0.05) and plasma soluble ICAM (-16%; p<0.01). Climacteric symptoms: By comparison with placebo, genistein caused a reduction from baseline in both the mean number (-34%; p<0.01 vs placebo) and the mean severity score (-40%; p<0.05 vs placebo) of hot flushes. Safety assessment: After a 12-months genistein treatment no significant difference in endometrial thickness was observed and vaginal cytology (using the karyopycnotic index or the maturation value score) showed no change. Our 1-year data confirm that genistein is an effective and safe alternative in postmenopausal women. 1) Morabito N. et al. J Bone Miner Res. 2002;17(10):1904-12. 2) Squadrito F. et al. Am J Med. 2003;114(6):470-6. This work was supported by grant from MIUR (Ministero dell’Istruzione dell’Università e della Ricerca). Effects of Long Term Soy Dietary Supplementation on Bone Mineral Density. F. Lovrien1, J. Williams2, B. Meyer1, and D. Erger1, 1Sioux Valley Hospital, Sioux Falls, South Dakota, USA, 2University of South Dakota, Vermillion, South Dakota, USA. A few studies in postmenopausal (PMP) women suggest that a diet that includes soy may reduce bone loss. Findings in some studies may be confounded by short study periods. We examined whether a diet with soy protein for 2 y reduces losses in bone mineral density (BMD) in PMP women. Subjects 1-5 years PMP were randomly assigned: soy/hormone replacement therapy (HRT; n=34), casein/HRT (n=33), soy/no-HRT (n=28), casein/no-HRT (n=20). For 2 y, subjects ate a normal diet supplemented with 40 g soy (90 mg aglycone isoflavone) or casein daily. Isoflavones (mg/g soy): genistein, 2.1; daidzein, 1.1; glycitein, 0.2. Both supplements provided 1400 mg calcium. DEXA scans of hip and spine were taken before and after 1 and 2 years. Comparisons (% change) were made in BMD between soy/HRT and casein/HRT and between soy/no-HRT and casein/no-HRT groups with an unpaired t-test. In the spine, after 1 year BMD decreased 1.26±0.72% (mean±SE) and 2.21±0.56% in soy/no-HRT subjects and casein/no-HRT subjects (NS) and 0.02±1.00% and 2.73±0.71% after 2 years (P<0.05). Changes in BMD were similar between soy/HRT and casein/HRT after 1 and 2 years. In the hip, no differences occurred after 1 or 2 y when comparisons were made between soy/HRT and casein/HRT and between soy/no-HRT and casein/no-HRT groups. Our findings indicate that long-term dietary supplementation with soy can reduce bone loss in PMP women, but this protection may be selective for certain bones. Supported by the South Dakota Health Research Foundation and the South Dakota Soybean Research and Promotion Council Effect of Soy Protein With or Without Isoflavones on Bone Mineral Density in Postmenopausal Women. J.C Gallagher, Prema Rapuri, Sue Longsdon, Jeff Detter; Creighton University Medical School, Omaha NE 68131, USA. Soy protein or its isoflavones have been suggested to have a positive effect on bone that may help to prevent the development of osteoporosis. The objective of the present study was to determine the effect of soy protein with or without isoflavones on bone mineral density (BMD) in postmenopausal women aged 55-72 years over 24 months. Two hundred and thirty women were enrolled into the study. They were randomly assigned to one of the three supplement groups, 25 g soy protein without isoflavones, 25 g soy protein containing 90 mg of isoflavones (genistein and daidzein) or 25 g casein. Twenty four women did not receive any supplemental protein and served as non protein control. Spine, femoral neck, trochanter, total femur and total body BMD measurements were performed by dual energy absorptiometry at baseline and at the end (24 months) of the study. The percent change in BMD over baseline was calculated at the end of the treatment period. Data was analyzed by ANOVA and Bonferroni correction was used for multiple comparisons. There were no significant differences between the groups with respect to their age and body mass index. The baseline BMD was not significantly different between the four groups. Bone loss occurred in al three groups; our preliminary analyses indicate that the unadjusted percent change in BMD was not significantly different between the treatment groups and none of the protein treatment groups prevented bone loss. ACKNOWLEDGEMENTS • This work was supported by funding from Solae Company. Cognitive Function Cognitive Effects of Soy Isoflavones in Older Adults: Influence of Gender and ApoE Genotype. C. Gleason1,2, S. Meade1,2, N. Lane1,2, T. Ohrt1,2, and S. Asthana1,2, 1University of Wisconsin, Dept. of Medicine, Sect. of Geriatrics, Madison, WI, USA, 2Madison VA GRECC, Madison, WI, USA. Evidence from basic science suggests that estrogen has neuroprotective and neuromodulatory effects; however, findings from the Women’s Health Initiative Memory Study (WHIMS) have raised concerns over the feasibility of traditional hormone therapies because of the increased risk of serious adverse effects, including cognitive declines. Soy isoflavones may serve as a critically needed alternative therapy to traditional hormone therapy. The few human research projects that have examined the cognitive effects of soy isoflavones offer intriguing, but as yet preliminary support for isoflavones’ beneficial actions. Careful and systematic study is needed to ascertain the merit of this potential therapeutic. This randomized, placebo-controlled, double-blind design clinical pilot study evaluated the effects of soy isoflavones on cognition in older men and postmenopausal women. It was hypothesized that 11 2005 Abstracts brains. Protein carbonyls in brain homogenates were derivatized with dinitrophenylhydrazine (DNPH), then analyzed by 2D gel proteomics and Western blots with anti-DNP antibody. While the overall patterns of gel spots were similar, the 2D Western blots of brain proteins from the GSE-fed animals were reduced (20-40% of controls). Gel spots reactive with the anti-DNP antibody were identified by matrix-assisted laser desorption ionization time of flight mass spectrometry. In view of the common proteins affected by GSE and SPI, we predict that proteins similar to those protected by GSE from oxidation will be protected by SPI and studies are ongoing to examine these effects. The DNP-reactive proteins reported here are the first protein oxidations identified, using proteomic technologies, to be affected by dietary polyphenols. GSE, SPI, and related polyphenols may have pleiotropic actions, including on gene expression, protein modifications, and anti-oxidant activity. These studies were supported in part by a subcontract of grant P50-AT00477 from Purdue University (Connie Weaver, PI), by a grant from the United Soybean Board, and by grant DAMD17-01-0469 from the Dept of Defense Medical Research and Materiel Command Breast Cancer Research Program. The grape seed extract preparation was generously provided by Kikkoman Corporation (Chiba, Japan), and the soy protein preparations provided by Protein Technologies, Inc. (now Solae, Ltd.). The operation of the UAB Comprehensive Cancer Center Proteomics/Mass Spectrometry Shared Facility was supported in part by a grant (P30 CA13148) from the National Cancer Institute to the UAB Comprehensive Cancer Center (A. LoBuglio, PI). Purchase of proteomic and mass spectrometry instrumentation in the Shared Facility was enabled by Shared Instrumentation Grants to HK and SB from the National Center for Research Resources. Weight Control 16-Week Randomized, Controlled Trial of Soy vs. Casein Meal Replacements for Weight Management of Obese Women. J.W. Anderson 1, J. Fuller1, E. Konz1, and A. Tabor2, 1University of Kentucky, Lexington, KY, USA, 2Revival Soy, Kernersville, NC, USA. NIA grant K23 AG24302 Research staff and study volunteers from the Wisconsin Comprehensive Memory Program The General Clinical Research Center: grant M01 RR03186 from the National Center for Research Resources, National Institutes of Health; Madison GRECC of the Department of Veterans Affairs; Archer Daniels Midland provided Novasoy® brand isoflavones tablets and matching placebo. Prediction of Direct Anti-Oxidant Activity for Soy Isoflavones in Mammalian Brain. S. Eliuk, J. Deshane, L. Wilson, M. Kirk, S. Barnes, and H. Kim*, University of Alabama at Birmingham, Birmingham, Alabama, USA. Soy isoflavones protected against ovariectomy-induced cognitive dysfunction in rats, however the molecular basis for this is unknown. Proteomics of brains from animals given grape seed extract (GSE) or soy protein isoflavones (SPI) indicated that there were common proteins among those affected in the brains of animals that received either polyphenol mixture. Since polyphenols have been shown to have antioxidant activity, we hypothesized that dietary GSE or SPI would attenuate brain protein oxidations. We studied a transgenic mouse model of dementia, the Tg2576 mouse, which overexpresses amyloid precursor protein in the brain, and the animals become cognitively impaired over time, concomitant with the deposition of amyloid plaques in their Meal replacements (MR) are very effective adjuncts to lifestyle changes for weight loss and weight maintenance. Soy protein has specific effects on fat, glucose and lipid metabolism and promotes weight loss in animals. This randomized, controlled trial compared effects of soy meal replacements vs. casein meal replacements on body weight for obese women participating in an intensive lifestyle intervention. Volunteers consumed three MR shakes, either soy or casein, one MR entrée, and five fruits or vegetables daily. Soy protein consumption was 62 g/d with 147 mg/d of isoflavones (aglycones); casein intake was ~68 g/d. 43 subjects enrolled and 35 (17 soy, 18 casein) completed the 16week study. Soy and casein shakes were well tolerated with good adherence. With soy, subjects lost more visceral adipose tissue and had greater reductions in LDL-cholesterol than with casein but differences were not statistically significant. Outcome changes at 16 weeks for soy and casein, respectively, with mean (SE) and significance from baseline* were: weight, -12.8% (1.4)*,-14.0% (1.2)*; DEXA total fat, -21.8% (2.4)*, -23.7% (2.0)*; visceral fat by mid-abdominal computerized tomography, -30.3% (4.2)*, -27.2% (3.7)*; LDL-cholesterol, -12.5% (4.2)*, -7.7% (4.4)*; and serum triglycerides, -12.5% (8.1), -6.7% (7.7). This intensive intervention was associated with significant weight loss averaging 17 pounds at 8 weeks and 28 pounds at 16 weeks with total fat losses averaging 23% and visceral fat losses averaging 29%. Including 3 MR shakes, either soy or casein, with one portion-control entrée and 5 fruits or vegetables is a very effective weight loss intervention. • Supported in part by Revival Soy. Oral Presentation Abstracts isoflavone supplements would ameliorate age-associated cognitive declines for healthy older adults (i.e., decrease rate of decline, stabilize or enhance cognitive abilities) as compared with subjects on placebo. Data were obtained from 30 cognitively healthy older adults (50% women). Subjects received either 100 mg/day of soy isoflavones or a placebo for 6 months. Cognitive evaluations were conducted at baseline and at 1, 3, and 6 months after initiation of study medications. Cognitive data, collected while on treatment with either isoflavones or placebo were used to calculate ‘change from baseline’ scores. Women on isoflavones outperformed women on placebo on a test of category fluency (χ2 = 5.4, p=0.02). Men on treatment did not differ significantly from those on placebo in category fluency performance. The accompanying figure depicts these data. In addition to sex-differences, a genetic risk factor for Alzheimer’s disease, the ApoE gene was also found to influence response to isoflavone treatment. Presence of one or two copies of the ApoE 4 allele increases a person’s risk for Alzheimer’s disease by three- and nine-fold, respectively. Only men without this genetic risk factor appeared to benefit from isoflavone treatment on a measure of executive function, the Trail Making Test (χ2 = 3.94, p =0.05). Although preliminary, these data suggest that isoflavone treatment is associated with improvements in language and executive function when compared with placebo. However, these beneficial effects are moderated by subject characteristics, such as sex and ApoE genotype. 6th International Symposium on the Poster Abstracts 12 Role of Soy in Preventing and Treating Chronic Disease Poster Presentation Abstracts Please note: The following abstracts have not been edited for content. They appear as submitted by the authors. Session I Sunday, October 30, 2005 ..............1:00–7:00 p.m. Monday, October 31, 2005 .....7:30 a.m.–12:30 p.m. Athletic Performance Post-Exercise Consumption of Soy Protein Promotes General Protein Synthesis and mRNA Translation Factor Activity in Skeletal Muscle. T. Anthony1 and M. McNurlan2, 1Indiana University School of Medicine, Evansville, IN, USA, 2State University of New York, Stony Brook, NY, USA. The objective of this study was to comparatively assess the stimulation of skeletal muscle protein synthesis and mRNA translation initiation following postexercise ingestion of various protein-containing meals. Treadmill-acclimated rats (n = 6-8 per group) were randomly designated as nonexercised controls (NEX) or run for 2h at ~75%VO2max and then orally administered one of four isocaloric meals: EC, purely carbohydrate; ESI, carbohydrate plus 20% soy protein isolate; ESC, carbohydrate plus 20% soy protein concentrate (~3.5X elevated phytochemical content compared to ESI); EW, carbohydrate plus 20% whey protein concentrate. One hour following exercise, all protein-containing meals promoted the fractional rate of muscle protein synthesis to a similar extent above EC. Additionally, all protein-containing meals reduced association of the mRNA cap binding protein, eIF4E with the translational repressor, 4E-BP1, and promoted formation of the eIF4E-eIF4G active mRNA cap binding complex similarly. In contrast, phosphorylation of 4E-BP1 and the 70kD ribosomal protein S6 kinase (S6K1), a factor involved in regulating cell size but whose exact role in protein synthesis is unclear, was highest in EW. Insulin concentrations among postexercise-fed rats were similar, but serum concentrations of isoleucine, leucine, methionine, and threonine were highest in EW. Thus, short-term recovery of protein synthesis and formation of the eIF4 active complex is promoted in muscle by both soy and whey protein in exercised rats. However, acute phosphorylation of proteins downstream of the mammalian target of rapamycin (mTOR) kinase is greatest in response to whey protein, which is likely related to its higher branchedchain amino acid content. • This project was funded by Solae, Inc. Soy Protein Intake Has Broad Positive Interactions with Exercise. R. DiSilvestro, Ohio State University, Columbus, OH, USA. Previously, our laboratory has shown that in men, intake of high isoflavone soy protein can impact biochemical indices of exercise recovery, and promote exercise training-induced lean body mass gain. More recently, these results have been extended by finding the following actions of soy protein intake: • In aerobically, recreationally trained males, antioxidant functional improves without depression of serum testosterone; • There is improvement of aerobic performance recovery in some, though not all of a group of recreationally trained males; • A lowering of cortisol responses to resistance exercise occurs in recreationally resistance exercise trained men; • In recreationally resistance exercise trained women, an enhancement is produced in recovery from a moderately intense resistance exercise session (based on a number of parameters); • In recreationally resistance exercise trained women, a depression is produced in oxidant stress between exercise sessions. These results show that soy can exert a variety of positive interactions with exercise in both genders without a previously perceived adverse effect in males (testosterone lowering). In addition, this work shows that soy antioxidant effects can extend beyond immediate recovery, and that soy protein intake can impact not just biochemical indices, but also physical performance. However, much more research is needed in this last area. Part of this work was supported by research gifts from Solae. Cancer Phytoestrogen Intake Prior to Diagnosis is Associated with Improved Indicators of Breast Cancer Survival in a Group of Newly-Diagnosed Australian Women. J. Ha1, P. Lyons-Wall*2, D. Moore3, D. Tattam3, J. Boyages4, O. Ung4, and R. Taylor1, 1School of Public Health, The University of Sydney, Australia, 2School of Public Health, Queensland University of Technology, Australia, 3Faculty of Pharmacy, The University of Sydney, Australia, 4New South Wales Breast Cancer Institute, Westmead Hospital, Australia. Breast cancer incidence is lower and survival is higher in Asian compared to Caucasian women and dietary phytoestrogens have been examined as a possible reason for the disparity. The aim of this analytical cross-sectional study was to examine the associations between phytoestrogen intake prior to diagnosis and indicators of breast cancer survival. Subjects were 128 women aged 40-79 yr, newly-diagnosed with invasive breast cancer at the New South Wales Breast Cancer Institute, Westmead Hospital, Sydney. Intakes of dietary and supplemental phytoestrogens (isoflavonoids and lignans) were obtained using a specially-designed 112-item frequency questionnaire that estimated intake over the previous year. Outcome variables relating to tumour characteristics, collected from pathology reports, included estrogen and progesterone receptor status, tumour size, histological grade and degree of spread; lymphovascular invasion and positive axillary lymph nodes were used as proxy indicators for breast cancer survival. Results showed that after controlling for confounding factors, the odds of being diagnosed with: a positive progesterone receptor increased by a factor of 2.42 [95%CI 1.03-5.68] (p=0.04), a positive lymphovascular invasion decreased by 0.32 [0.12-0.85] (p=0.02), or smaller size tumour decreased by 0.50 [0.26-0.94] (p=0.03), for each unit increase in lignan consumption. The odds of being diagnosed with: any stage of cancer other than stage 1 decreased by 0.83 [0.70-0.98] (p=0.03) or a positive progesterone receptor increased by a factor of 1.22 [1.00-1.49] (p=0.05), for each unit increase in isoflavonoid consumption. These results suggest that higher phytoestrogen intake prior to diagnosis is associated with improved prognosis in women with breast cancer. 13 2005 Abstracts Objective: To investigate the effects of genistein (Gen) on the proliferation, DNA synthesis, nitric oxide (NO) production, nitric oxide synthase (NOS) activity, induced nitric oxide synthase (iNOS) gene transcription and expression of human gastric carcinoma cell line SGC-7901. Methods: MTT method, 3H-TdR incorporation, DNA electrophoreses, spectrophotometric measurement, immunohistochemistry method, western blot method, and RT-PCR were used to evaluate the possible mechanism of cell growth inhibition. Results: Gen at 20 µg/mL significantly inhibited the proliferation of human gastric carcinoma cell line SGC-7901, inhibited DNA synthesis of SGC-7901 and induced cell apoptosis. Exposure of SGC-7901 cells to Gen resulted in statistically significant increase in iNOS mRNA transcription, accompanied with increased iNOS protein expression, NO production and NOS activity in the dose-dependent and time-dependent manners. Conclusions: The increased expression of iNOS may contribute to the growth inhibitory as well as apoptotic effects of Gen on gastric carcinoma cells. The results provide evidence for the potential usefulness of Gen in the prevention and treatment of human gastric cancer. This work was supported by research grants from the National Nature Science Foundation of China. The in vitro Metabolism of Estrogens by Human Fecal Bacteria: A Comparison of Equol-Producers and NonProducers. C. Atkinson1, S. Berman2, W. Thomas1, and J. Lampe1, 1Fred Hutchinson Cancer Research Center, Seattle, WA, USA, 2Bastyr University, Kenmore, WA, USA. Determinants of circulating levels of estrogens and estrogen metabolism are among the primary risk factors for breast cancer. Hormone metabolism in humans involves several steps including intestinal bacterial metabolism. Thus, inter-individual differences in intestinal bacteria may contribute to variation in disease susceptibility through effects on metabolism and subsequent exposure to estrogens. One-third to onehalf of the population harbor bacteria that metabolize daidzein to equol. Studies suggest that equol production may be associated with reduced breast cancer risk, but mechanisms for this relationship are unknown. Our aim was to determine whether fecal bacterial metabolism of estrogens differs according to equol-producer phenotype. Frozen fecal samples from 34 women were available. According to previously analyzed urine samples (collected after soy consumption) 20 were equol-producers (EP), 12 were equol non-producers (ENP), and 2 had trace amounts of equol in their urine. Thawed feces was incubated anaerobically in media for 5d at 37°C with daidzein or the following estrogens: estrone, estradiol, estriol, 2-hydroxyestrone, 16alpha-hydroxyestrone, and 2-methoxyestrone. After incubation, estrogens and isoflavones were extracted and quantified by gas chromatography-mass spectrometry. EP, compared to ENP, metabolized significantly more estrone to estradiol, and 16alpha-hydroxyestrone to estriol (p≤0.05). Although differences between EP and ENP in the amount of 2-hydroxyestrone metabolized to 2-hydroxyestradiol were non-significant, EP were more likely than ENP to metabolize 2-hydroxyestrone to 2-hydroxyestradiol (χ2=4.1, p=0.04). All other differences were non-significant. These preliminary findings suggest that differences in fecal bacterial estrogen metabolism may exist between EP and ENP. This work was supported by a Department of Defense Breast Cancer Research Program Concept Award (DAMD17-03-1-0604), and Fred Hutchinson Cancer Research Center. Phytoestrogens do not Affect the Growth of Breast Cancer Tumors in Mice. D. Gallo1, C. Ferlini1, M. Fabrizi1, S. Prislei1, A. Riva2, P. Morazzoni2, E. Bombardelli2, and G. Scambia1, 1Department of Obstetrics and Gynecology, Catholic University of the Sacred Heart, Rome, Italy, 2INDENA S.p.A., Milan, Italy. Consumption of soy-derived products has been encouraged for post-menopausal women as a natural alternative to hormone replacement therapy. One group of women most in need of and who frequently use alternatives consists of breast cancer patients; however, the estrogenic effects of isoflavones observed in some experimental systems have recently led to considerable controversy among health professionals over the use of soy by breast cancer patients. The present study was designed to investigate the effects of a standardized soy extract (Soyselect®, SSE) on the growth of estrogen dependent (MCF-7) and estrogen-unresponsive (MDA-MB-231) human breast cancer xenografts in athymic mice. Results obtained provided evidence that MCF-7 tumors did not grow over the treatment period (five weeks) in ovariectomized females receiving 50 or 100 mg/kg/day SSE (oral route); administration of SSE also did not affect the 17β-estradiol-sustained growth of MCF-7 tumors in mice. Similarly, no effect on tumor growth was observed in SSE-treated mice bearing MDA-MB-231 xenografts. Data from pS2, PR and cyclin D1 mRNA expression in tumors suggest that SSE was able to induce a moderate estrogenic effect in MCF-7 cells without increasing cellular proliferation and tumor growth. Besides, when used in association with low dose 17β-estradiol, it displayed anti-estrogenic activity. The expression of other genes involved in tumor progression and angiogenesis, such as Thrombospondin 1 and Transforming Growth Factor β2, was also evaluated in tumor samples, results showing a decrease in mRNA expression upon SSE treatment. Biological effects will be also discussed in relationship with plasma levels of isoflavones. Prospective Survey Evaluating the Use of Soy Products in Women with Breast Cancer. J. Franciose, C. Lammersfeld, J. Grutsch, P. Vashi, and S. Walker, Cancer Treatment Centers of America, Zion, IL, USA. Introduction: Many studies have investigated the potential role of soy in the prevention of breast cancer. A number of components in soy appear to have anticancer properties, including the isoflavones genistein and daidzein. The use of soy by women with breast cancer is currently being questioned because of the estrogen-like effects of isoflavones and possible interactions with tamoxifen. Clinicians providing nutrition counseling for these women are concerned because the availability of soy foods and supplements has increased dramatically in the past few years. The goal of this study was to quantify the intake of isoflavones in women with breast cancer to determine if their intake is of clinical significance. Methods: We evaluated a consecutive case series of 100 women with breast cancer treated at Cancer Treatment Centers of America between 09/03 and 02/04. After obtaining informed consent, each patient was asked to complete a soy food frequency questionnaire (FFQ) that was scored by Fred Hutchinson Cancer Research Center, the institute that developed and validated the FFQ. Results: The mean age was 50.5 years (std. dev. = 9.4; range 3170) and the mean BMI was 27.3kg/m2 (std. dev. = 6.75; range 17-59). Poster Abstracts Effect of Genistein on iNOS Expression and its Relation to Proliferatory Inhibition of Gastric Carcinoma Cells. D.F. Song1,2, S.K.C. Chang1, and H.B. Cui2, 1Department of Cereal & Food Sciences, North Dakota State University, 376 IACC, Fargo, ND, USA, 2Department of Food Nutrition & Hygiene, Public Health College, Harbin Medical University, Harbin, Heilongjiang Province, China. 6th International Symposium on the Poster Abstracts 14 Role of Soy in Preventing and Treating Chronic Disease Genistein and daidzein consumption was limited to 65 (65%) patients with a mean intake of 81.5 mg/week (std. dev. = 153; range 0.02-682) and 52.9 mg/week (std. dev. = 98.8; range 0.02-482) respectively. Conclusions: Our study suggests that the isoflavone intake of breast cancer patients treated at our hospital was quite variable. Thirtyfive patients (35%) reported no soy intake. The mean daily intake translates into 11.6 mg genistein and 7.4 mg daidzein, which is the equivalent of less than 1⁄4 cup of tofu per day. This amount is slightly higher than has been previously reported in non-Asian American women. Dietary Soy Isoflavones have no Adverse Effects on the Non-Human Primate Prostate, Testis, or Mammary Gland. D. Perry1, J. Spedick1, M. Adams1, A. Franke2, S. Walker1, and J. Cline1, 1Department of Pathology, Section on Comparative Medicine Wake Forest University School of Medicine, Winston-Salem, NC, USA, 2Cancer Research Center of Hawaii, Honolulu, HI, Honolulu, HI, USA. In mice, dietary isoflavone supplementation at an equivalent of 120 mg/person/day has produced marked prostatic atrophy. In human prostatic cancer cell lines, isoflavones have induced antiproliferative and apoptotic effects. Some Asian diets contain 50 mg/day isoflavones compared to some traditional Western diets which deliver 2 to 3 mg/day. It has been suggested that this may explain why Asian men have approximately one tenth the incidence of prostate cancer. Yet, no evidence of prostatic atrophy or reduced reproductive function is seen in these populations. We assessed the effects of dietary soy isoflavones on mammary, prostatic, and testicular morphology and function in age matched, adult male cynomolgus monkeys. Diets differed only by protein source: 1) a soy-free, casein/lactalbumin based diet or 2) and 3) soy protein isolate diet containing an equivalent of 75 or 150 mg/person/day soy-derived isoflavones. Total serum isoflavones were 67+/-23 (soy-free diet), 799+/44 (75 mg/person/day equivalent diet), and 1458+/-80 nM (150 mg/person/day equivalent diet) (mean+/-sem, p<0.0001). Diet did not alter serum estradiol and testosterone concentrations, epididymal or testicular sperm counts nor prostate, testicular, epididymal, or seminal vesicle weights. Mammary gland morphometric and histopathologic analysis revealed no gynecomastia. We conclude there was no morphologic evidence of adverse reproductive effects of soy isoflavones at doses equivalent to 75 to 150 mg/person/day in adult male monkeys. Beneficial Effects of Regular Consumption of Whole Soy Foods on Serum Lipids, Lipid Peroxidation, High Sensitivity CRP and Blood Pressure in Perimenopausal Women: A Randomized, Controlled, Crossover Trial. S. Songchitsomboon1, K. Chanda2, D. Danboonchant2, J. Manonai3, J. Hong4,And S. Komindr5, 1Research Center, Faculty of Medicine, Ramatibodi Hospital, Mahidol University, Bangkok 10400, Thailand, 2Graduate Program of Nutrition, Faculty of Medicine, Ramatibodi Hospital, Mahidol University, Bangkok 10400, Thailand, 3Department of ObstetricGynecology, Faculty of Medicine, Ramatibodi Hospital, Mahidol University, Bangkok10400, Thailand, 4School of Food and Life Science, Inje University, Korea, 5Department of Medicine, Faculty of Medicine, Ramatibodi Hospital, Mahidol University, Bangkok 10400, Thailand. Background: Hypercholesterolemia, menopausal symptoms and osteoporosis are the major health problems in middle-aged Thai women and elsewhere. Meanwhile, little is known about the effects of commonly consumed soy foods on cardiovascular risk factors in Thai people. Objective: To determine whether incorporating reasonable amount of soy foods in a self selected diet, low in saturated fat and cholesterol would reduce markers of cardiac risk in perimenopausal women Method: Thirty-seven perimenopausal women aged 40 - 59 years participated in a randomized crossover trial with two 12-wk diets and 4-wk washout period before and between treatments. The study diets consisted of a control diet (soy-free diet) and an isocaloric soy-rich diet with their own recipes (25 g soy protein as soymilk, tofu, texture soy protein and sausage contained more than 50 mg isoflavones daily). Subjects were asked to fill-in daily eaten soy food lists. Blood levels of lipid profile, lipid hydroperoxides, conjugated dienes, high sensitivity c-reactive protein (hs CRP), antioxidant capacity in water- and lipid-soluble substances, glucose, and isoflavones were measured at the start and end of each diet. Results: It was noted that at the beginning of the study, general characteristics of both groups were similar. Good compliance to the diet was shown by the significant elevation of serum levels of daidzein and genistein during the soy-rich diet period. Compared with baseline, significant improvement in the blood levels of the following FBS, LDL-C, HDL-C, total cholesterol/HDL-C, LDL-C/HDL-C, total antioxidant capacity of water-soluble substances and diastolic blood pressure were observed after both diets. Moreover, a soy-rich diet resulted in significantly lower total cholesterol, lipid hydroperoxides, hs CRP and systolic blood pressure. In addition, the changes of total cholesterol and 10-yr coronary heart disease risk in the soy-rich diet differ significantly from the changes in the control diet. Conclusions: This study provides evidence that long-term ingestion of soy foods has beneficial effects on blood lipid and CRP concentrations. This study was support in part by a grant from the Thai Health Promotion Foundation. The analysis of isoflavones in food and blood samples were supported by NutraTech Co, Ltd, Thailand and Unifood Tech Laboratory, Korea. Using Biomarkers to Assess Phytoestrogen Intake in Breast Cancer Patients During a Dietary Intervention Study. T. Rawjee1, G. Spahn2, C. Kennemann2, A. Blake3, J. Mackinnon1, G. Dobos2, and M. Ritchie1, 1Bute Medical School, University of St. Andrews, Fife, Scotland, 2Dept. of Internal and Integrative Medicine, University Duisburg-Essen, Essen, Germany, 3Scottish Crop Research Institute, Dundee, Scotland. OBJECTIVE: To determine phytoestrogen (PE) intake by urine analysis in 20 German breast cancer patients following either A) a Mediterranean diet within a 10 week structured multimodal day clinic programme or B) an home based exercise programme without nutritional intervention. MATERIALS AND METHODS: Urine samples (24 hour urine collections as well as timed urine samples) were collected before and after each intervention. Group A attended a nutrition day within the Dept. of Integrative Medicine which included a single meal high in soy and other PE rich foods. Analysis for the isoflavones (genistein and daidzein) was carried out by Liquid Chromatography-Mass Spectrometry RESULTS: Dietary PE intake increased significantly in breast cancer patients ingesting the PE-rich meal (from a mean intake of 11.7mg/d to a mean intake of 28.7mg/d). No difference in PE intake was observed between the exercise group and the multimodal intervention group in the day clinic at the end of the study. CONCLUSIONS: This study shows clearly that a diet rich in PE could raise urinary PE concentrations threefold if breast cancer patients would 15 2005 Abstracts Are Phytoestrogens Beneficial in Breast Cancer Patients? Use of a Newly Validated Biomarker to Assess Phytoestrogen Intake in Women With Breast Cancer and Controls. J. Mackinnon1, T. Rawjee1, A. Blake3, G. Spahn2, G. Dobos2, and M. Ritchie1, 1Bute Medical School, University of St. Andrews, Fife, Scotland, 2Dept. of Integrative Medicine, University Duisburg-Essen, Essen, Germany, 3Scottish Crop Research Institute, Dundee, Scotland. OBJECTIVES: To compare phytoestrogen (PE) intake in women with non-metastatic breast cancer, women with metastatic breast cancer and healthy controls. METHODS: Timed urine samples were obtained from German breast cancer patients with non-metastatic disease who had completed treatment, and age matched healthy Scottish controls. Urinary PE concentrations are valid biomarkers of PE intake and LC-MS urinalysis was used to determine dietary PE intake. Patients with metastatic breast cancer are currently being recruited and results of urinalysis are expected by Sept 2005. RESULTS: Urinalysis to date indicates mean PE intake in breast cancer patients was significantly higher than mean PE intake in age-matched controls (p<0.0001). Mean PE intake in breast cancer patients (n=17) was 10.46mg/day compared with 2.76mg/day in controls (n=11). DISCUSSION: This is the first study to use a validated biomarker to compare PE intake in different populations and patient groups. Despite dietary variation between countries, the use of validated biomarkers enables the influence of specific dietary phytochemicals on disease risk and disease characteristics to be identified. Although the data are preliminary, this study supports the hypothesis that increasing PE intake may not prevent the occurrence of breast cancer in adult women. However, it also supports the hypothesis that a higher PE intake may help to restrain tumour progression and metastasis, since breast cancer patients with a higher PE intake than the general population did not show disease progression. Comparison with PE intake in women with metastatic breast cancer will provide further evidence to confirm or refute this hypothesis. This study was funded by the Carstens Foundation, Essen, Germany and the Melville Trust for the Care and Cure of Cancer, Edinburgh, Scotland. Phytoestrogen-Gene Associations with Sex Hormone Levels Among Postmenopausal Women in EPIC-Norfolk. Y. Low1, A. Dunning2, M. Dowsett3, and S. Bingham1, 1MRC Dunn Human Nutrition Unit, Cambridge, United Kingdom, 2Cancer Research UK - Department of Oncology, Cambridge, United Kingdom, 3Institute of Cancer Research, London, UK. Phytoestrogens have been hypothesised to stimulate sex hormone-binding globulin (SHBG) production and modulate sex hormone levels through inhibiting enzymes involved in estrogen metabolism. In this large cross-sectional study, we investigated the relationship between phytoestrogen exposure and plasma sex hormones and SHBG levels in postmenopausal women and their relation with polymorphisms in genes involved in estrogen signalling and metabolism. Plasma estradiol, testosterone, androstenedione, estrone and SHBG were measured in 1968 free-living healthy postmenopausal women in the Norfolk arm of the European Prospective Investigation into Cancer and Nutrition (EPIC) study. We have previously shown that urinary phytoestrogen levels correlated strongly with serum phytoestrogen levels (R=0.81 to 0.91, p<0.001) and can serve as suitable dietary biomarkers (1). Urinary levels of seven phytoestrogens (daidzein, genistein, glycitein, O-desmethylangolensin (O-DMA), equol, enterodiol and enterolactone) were measured and used as biomarkers for dietary phytoestrogen intake. Nineteen single nucleotide polymorphisms (SNPs) in the HSD17B1, CYP19, SHBG, ESR1 and NR1I2 genes were genotyped using end-point Taqman assays. After adjusting for potential confounders, O-DMA was positively correlated with plasma estradiol (R=0.059, p=0.009), equol was negatively correlated with plasma estradiol (R= -0.050, p=0.029) and enterodiol was negatively correlated with estrone (R= -0.076, p=0.013) and testosterone levels (R= -0.057, p=0.016). Genistein and enterolactone were positively correlated with plasma SHBG (R=0.046, p=0.047 and R=0.110, p=0.0000015 respectively). The relationships between phytoestrogens and plasma sex hormone levels were significantly different among women with different genotypes for selected polymorphisms in genes involved in estrogen signalling and metabolism, suggesting phytoestrogen-gene interactions. Even in Western populations with low habitual soy consumption, phytoestrogen exposure exerts modest effects on plasma sex hormone levels in postmenopausal women and diet-gene interactions may be involved. Reference 1. Grace PB, Taylor JI, Low Y-L, Luben RN, Mulligan A, Botting NP, et al. Phytoestrogen concentrations in serum and spot urine as biomarkers for dietary phytoestrogen intake and their relation to breast cancer risk in European Prospective Investigation of Cancer and Nutrition-Norfolk. Cancer Epidemiol Biomarkers Prev 2004;13:698-708. Grant support: UK Medical Research Council, Cancer Research UK and the Agency for Science, Technology and Research, Singapore. Microarray Analysis of Gene Expression Regulation By Equol in MCF-7 Cells. J.S. Kim1, J.H. Kim1, J.R. Kim1, C.H. Jang1, H.A. Lim1, S.J. Lee3, and D.Y. Kwon2, 1Department of Animal Science & Biotechnology, Kyungpook National University, Daegu, S. Korea, 2Korea Food Research Institute, Sungnam, Kyunggi, Republic of Korea, 3Division of Food Science, Korea University, Seoul, Republic of Korea. Equol, a metabolite produced from daidzein by gut fermentation, drew much attention due to the stronger potential in preventing chronic diseases than parent compound. We examined the effect of equol on the gene expression in mammary tumor MCF-7 cells using microarray. The expression of 1122 genes out of total 33766 genes probed appeared to be changed by more than 2-fold. We also performed Kmeans clustering for 1122 genes whose expression was significantly modulated, and also assigned each of those genes to one of 10 kinds of biological processes according to their function. Twelve clusters in which each cluster had unique expression pattern were obtained. Eighteen genes involved in apoptosis changed significantly their expression and most of them were upregulated by equol treatment in MCF-7 cells. Forty six genes among the significantly regulated 1122 genes were associated with cell cycle. A large proportion of genes whose expression Poster Abstracts continuously adhere to such a diet. At present, no such recommendation can be made, but if beneficial effects of PE should be substantiated for breast cancer patients, the study shows the value of a nutritional intervention. A diet rich in fruit and vegetables which is generally recommended and followed by health conscious German breast cancer patients conveys a significantly higher PE content than in age-matched healthy controls from Scotland. The study was funded by the Carstens foundation, Essen, Germany and the Melville Trust for Care and Cure of Cancer, Edinburgh, Scotland. 6th International Symposium on the Poster Abstracts 16 Role of Soy in Preventing and Treating Chronic Disease was changed belonged to the family of signal transduction, nucleoside, nucleotide and nucleic acid metabolism. Half of genes involved in cell proliferation and differentiation were upregulated while 68 percent of genes involved in amino acid metabolism were increased in MCF-7 cells treated with 100 uM equol. In conclusion, equol appears to have potential to cause cell growth inhibition by upregulating genes associated with apoptosis in mammary tumor cells. This study was supported by a grant of the Korea Health 21 R&D Project, Ministry of Health & Welfare, Republic of Korea (A050376). Isoflavones Discrepancy Between Self-Claimed and Actual Soy Intakes. M. Sagara1, M. Mori1, H. Mori2, and Y. Yamori*3, 1Research Institute for Production Development, Kyoto, Japan, 2Institute for Health Restoration, Nishinomiya, Japan, 3International Center for Research on Primary Prevention of Cardiovascular Diseases, Kyoto, Japan. Objective: Recent reports indicate the importance of soy isoflavones for the risk reduction of lifestyle-related diseases. This knowledge is becoming popular among the general public in Japan. To investigate a discrepancy between self-claimed and actual soy intakes, we carried out a questionnaire survey asking the frequency of soy intake and the depth of concern in soy, while we measured 24-hour urinary isoflavone excretion to estimate the dietary intake. Design and Methods: One thousand Japanese in total, 500 of each gender and 250 of each decade at the age range from 20 to 59, were randomly selected from Eastern and Western Japan to respond to the questionnaire through the Internet in Japan, 2005. After obtaining informed consent, we collected 24-hour urine for the measurement of isoflavone excretion from 49 men and 56 women living in the same districts in Japan as the respondents to the questionnaire. Results: According to the analysis of the questionnaire, 62.8% of the respondents thought they should take soy products every day and 45.7% believed they were taking soy products every day. 84.8% of the respondents like soy products and 97.0% thought soy products were good for health, but only 20.3% knew about the effects of soy isoflavones. From the analysis of 24-hour urinary isoflavone excretion, however, 66.0% of the whole volunteers for the urine collection and 38.1% of those who claim to take soy products every day were taking less than 40mg/day of isoflavones, which should be recommended for the prevention of lifestyle related-diseases from our world-wide survey on the association of 24-hour urinary isoflavone excretion with the mortality rates of coronary heart diseases and breast and prostate cancers as well as isoflavone effects on risk reduction by our nutritional intervention studies. Conclusions: The significant proportion of Japanese are not taking desirable amount of soy products which are recommended for the prevention of lifestyle-related diseases, although the large portion of Japanese know the importance of soy intake and believe they are taking soy products every day. Genetic Manipulation of Soybean Seed Isoflavones. V. Lozovaya, O. Zernova, A. Lygin, and J. Widholm, Dept. of Crop Sciences, University of Illinois at Urbana-Champaign, Urbana, IL, USA. A number of soybean transgenic lines has been generated via bombardment of embryogenic cultures (cv. Jack) with mixtures of cassettes, harboring the important genes of the phenylpropanoid pathway, such as PAL5 (phenylalanine ammonia- lyase), CHS6 (chalcone synthase) , IFS( isoflavone synthase) under the seed specific lectin promoter and a selectable marker HPT gene under 35 S promoter. The segregation of genes in the T1 generation was found in 3 out of 8 lines analyzed. One line transformed with PAL5, CHS6, IFS and HPT has been followed in 4 generations until homozygous. This line showed the expression of the target genes in RT- PCR tests of immature seeds and had a reduced total isoflavone level, a higher proportion of glycitein and a lower proportion of daidzein in mature seeds compared to the untransformed control. We found that the cotyledons of untransformed seeds contain about 70-80% of the isoflavones, and embryos, that account for about 3% of the whole seed weight, contain 20-30% of the total seed isoflavones. The main isoflavones of cotyledons are genistein and daidzein and only trace amounts of glycitein were detected in cotyledons. However, embryos contain a high glycitein proportion. Interestingly, the expression of phenolic genes (PAL5, CHS6, IFS) resulted in decreased total isoflavone content in cotyledons only, while there were no changes in total isoflavone content in embryos of seeds collected from transgenic plants compared to controls. This indicates that the lectin promoter drives genes in cotyledons and not in embryos. This project was carried out with funds from the United Soybean Board, Illinois Soybean Program Operating Board, and Soybean Disease Biotechnology Center. Effect of Ipriflavone on the Growth Performance and Related Physiological Function in Rats. H. Ma, Z. Han*, G. Wang, and S. Zou, Key Lab of Animal Physiology & Biochemistry, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, Jiangsu, China. Ipriflavone is a synthetic daidzein-derived compound able to modulate the oxidative phosphorilation and to exert a direct inhibitory effect on osteoclast activity and possibly to stimulate the osteoblast activity. At present, ipriflavone is mainly used in the treatment of osteoporosis in clinic. The aim of the present investigation was to evaluate the effect on growth performance. The experiment was conducted with 20 rats of 28 days old fed ipriflavone at dosage of 5 mg/kg in diet for 5 weeks. Compared with those of male control group, the body weight gain increased by 38.82 (p<0.01), the average daily body weight gain increased by 38.65% (p<0.01) and feed intake/daily body weight gain was lowed 62.17 %; The relative percentage to the body weight of muscle increased by 56.84% (p<0.05), fat relative percentage was lowed 5.71% (p>0.05); The serum urea-N was low 10.17 % (p<0.05), but the activity of glutamic pyruvic transaminase and alkaline phosphatase and the level of testoterone increased by 33.67 % (p>0.05) and 43.76 % (p<0.05) and 773.53% (p<0.01), respectively. The ratio of RNA/DNA of muscle increased by 10.40 %. Compared with those of female control group, there were no significant effect of ipriflavone on the average daily body gain and feed intake/daily body weight gain. The relative percentage to the body weight of muscle increased by 2.22% (p>0.05), fat relative percentage was lowed 16.92% (p<0.05); The activities of glutamic pyruvic transaminase and alkaline phosphatase, the levels of serum urea-N and estrogen were decreased. Effects of Isoflavone on Aromatase Activity and Muscle Growth in Male Rats. H. Ma, Z. Han*, G. Wang, and S. Zou, Key Lab of Animal Physiology & Biochemistry, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, Jiangsu, China. 17 2005 Abstracts Mechanism of Soybean Isoflavone on Regulation of Testosterone Secretion in Rat Leydig Cell. H. Ma, Z. Han*, S. Zou, and G. Wang, Key Laboratory of Animal Physiology & Biochemistry, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, Jiangsu, China. Isoflavones belong to one kind of phytoestrogens occurring naturally in many plants. Isoflavones have structural similarities to human 17β-estradiol and can bind to the oestrogen receptor in many tissues and exert weak estrogenic activity. The weakly oestrogenic diphenols formed influence sex-hormone production, metabolism and biological activity. Also, isoflavone would affect on some of the most important steroid biosynthetic enzymes may result in beneficial modulation of hormone concentration and action in body. In our laboratory, a series of research shows that daidzein could enhance the concentration of testosterone in serum and development of muscle cockerel and rat. In the present study we have investigated the effects of soybean isoflavones on the regulation of testosterone secretion in rat Leydig cell by assaying the concentrations of cAMP, the activities of PKA and aromatase. Result showed that 20~100ng/mL of daidzein and geistein decreased 3H-TdR incorporation in DNA of rat Leydig cell. Also, daidzein, genistein and estradiol could promote cAMP accumulation and activate the activity of PKA and inhibited the activity of aromatase thereby improved testosterone secretion in rat Leydig cell. Daidzein and genistein have no effect on the activity of adenylate cyclase , but estradiol could improve the activity of adenylate cyclase. Temperature and Soil Moisture Effects on Soybean Seed Isoflavones. A.V. Lygin1, V.V. Lozovaya1, A.V. Ulanov1, R.L. Nelson2, and J. Daide3, 1Dept. of Crop Sciences, University of Illinois, Urbana, IL, USA, 2USDAAgricultural Research Service, University of Illinois at Urbana-Champaign, , Urbana, IL, USA, 3Ecole Supérieure d’Agriculture de Purpan, Toulouse Cédex 03, France. The objective of our research was to evaluate the effects of air temperature and soil moisture conditions during soybean seed fill on seed isoflavone concentrations of soybean cultivars with known differences in isoflavone concentration. Two French cvs., Imari and Queen, and three U.S. cvs., Dwight, Jack and Loda, all in maturity group II were studied. Soybean plants were grown in the greenhouse under intermediate (18/28°C), 9.5 hour night/14.5 hour daytime temperatures with high soil moisture conditions. Beginning at the R6 growth stage plants were subjected to either intermediate (18/28°C), low (13/23°C) or high (23/33°C) 9.5 hour night/14.5 hour daytime temperatures with either low or high soil moisture conditions. The overall results show that low temperatures and high soil moisture conditions produced the highest seed isoflavone concentrations with changes in temperature having the larger effect. The changes in daidzein and genistein concentrations were similar to changes in total isoflavones but the glycitein concentration was much less affected. All cultivars showed a 3-fold increase in total isoflavone concentrations at the low temperature regime compared to the high temperature regime. Soil moisture and air temperature have clear effects on the total isoflavone concentration, but the ranking of cultivars by average isoflavone concentration remained consistent. This shows that isoflavone potential is under strong genetic control. This project was carried out with funds from the Illinois-Missouri Biotechnology Alliance, the United Soybean Board, the College of ACES Value-Added Program, the Illinois Soybean Program Operating Board, the Illinois Agricultural Experiment Station and the USDA Agricultural Research Service. Determination of Major Isoflavone Components Based on HPLC Technology among Southern Soybean Varieties in China. J.-M. Sun, F.-X. Han, and A.-L. Ding, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, P.R. China. The contents of major isoflavone components in two hundred forty-nine soybean varieties from southern six provinces of China were analyzed by HPLC technology. Results showed that six major components of isoflavone could be determined by HPLC in soybean seeds, including daidzin (D), glycitin (GL), genistin (G), malonyldaidzin (MGD), malonylglycitin (MGL) and malonylgenistin (MGG). The component with the highest content was malonyl-isoflavone (61.2%), and correlations among components were statistically significant. A great variation among varieties was presented in 249 varieties, and the variation coefficient was 49.6%. Total isoflavone content of cultivar Haimenhonghuangdouyi was the highest (4932.3µg/g), that of cultivar Baoyingdengxifeng was the lowest (367.1µg/g). There was significant difference on isoflavone contents in different varieties among various provinces. The average content of isoflavone in Zhejiang province was the highest (2717.2µg/g), and the lowest content (1181.8µg/g ) in Anhui province. There was a greatly significant positive correlation between isoflavone content and date of growth (r = 0.319***), a significant positive correlation between isoflavone content and weight of 100 seeds, and two negative correlations between isoflavone content and oil content (r = -0.45***) and protein content (r = -0.136) in soybean seeds. Intervention Study of Soy Isoflavone Supplement for Menopausal Women. N. Ishiwata1, S. Watanabe2, Y. Omori3, M. Murayama3, W. Mohara3, A. Yamada3, and T. Wada3, 1Atomi Junior College, Bunkyo-ku, Tokyo, Japan, 2National Institute of Health and Nutrition, Shinjyuku-ku, Tokyo, Japan, 3Tokyo University of Agriculture, Setagayaku, Tokyo, Japan. OBJECTIVES: Double-blind, cross-over clinical intervention study by isoflavone (IF) tablet were carried out on 111 peri-menopausal Japanese women. DESIGN: 122 peri-menopausal women participated in the study after mail invitation to 656 mothers of graduates of women’s college. Finally 111 women came to the baseline survey, which included questionnaire for job, dietary habit, life habit, menopausal symptoms, menstrual condition, and physical examination including biochemistry. Poster Abstracts An experiment was conducted with 20 Sprague-Dawley rats of 28 days of age, supplemented with daidzein, ipriflavone and 7-hydroxyisoflavone 5mg • Kg-1, respectively and 0 mg • Kg-1 in the control group in diet to investigate the mechanism of daidzein, ipriflavone and 7hydroxyisoflavone on muscle growth. Compared with those of control group, daidzein, ipriflavone and 7-hydroxyisoflavone can increased the ratio of RNA/DNA in the leg muscle by 19.09%, 9.09% and 15.45%, respectively. Meanwhile, the blood level of testosterone was increased and the level of estradiol decreased with aromatase activity alleviated. These results suggested that daidzein, ipriflavone and 7-hydroxyisoflavone could promote the animals growth by inhibiting the activity of aromatase which reduce the conversion of testosterone to estradiol, thereby increase the content of testosterone in the blood. 6th International Symposium on the Poster Abstracts 18 Role of Soy in Preventing and Treating Chronic Disease Women were divided to 3 groups; placebo (PL), IF tablet 40 mg a day (IF-L) and 80 mg a day (IF-H). Participants were allowed to take up to 20mg IF from daily ordinary meals. RESULT: At baseline survey, 49 women were pre-menopausal and 62 were post-menopausal. After 3 months of intervention, serum biochemistry revealed lowering effects of total cholesterol and triacylglycerol. In hormonal response, post-menopausal women showed a different attitude from pre-menopausal women. There was no significant difference in the menopausal symptom score among PL, IF-L and IF-H groups at baseline. As compared to PL, IF-L in the pre-menopausal group significantly altered specific symptom category score of motornerve system. The IF-L pre-menopausal group reported a significant improvement in chill (p=0.05), shortness of breath (p=0.05), excitation (p=0.02), worry (p=0.04), tiredness (p=0.03) and joint aches and pains (p<0.01). Nervousness (p=0.02) decreased in the IF-L post-menopausal group. We would show the final cross-over results at the Conference. Genistein and Daidzein Reduces Level of Total Serum Cholesterol in Orhidectomized Middle-Aged Rats. B. Sosic-Jurjevic1, D. Brkic*2, B. Filipovic1, V. Ajdzanovic1, and M. Sekulic1, 1Institute for Biological Research, Belgrade, Serbia and Montenegro, 2Crown Agents, Belgrade, Serbia and Montenegro. Soybean consumption during menopause or andropause is associated with potential health benefits, such as prevention of atherosclerosis progression and bone preservation. In this study we examined the effects of chronic genistein and daidzein treatment on serum level of total cholesterol in orhidectomized middle-aged rats. Moreover, we compared the effects of soy isoflavones with corresponding chronic estradiol and testosterone treatments. Male Wister rats, 14.5 months old, were orhidectomized (ORH) under Ketamine anesthesia (0.015 mg/kg b.w.). Two weeks after the surgery animals were divided into 5 groups (n = 8), which were subcutaneously treated with genistein (30 mg/kg b.w.), daidzein (30 mg/kg b.w./every day), estradiol dipropionate (0.625 mg /kg b.w) every day for 3 weeks, or 2 times a week with depotestosterone enanthate (4.76 mg/kg b.w.).The control ORH group received the vehicle (sterile olive oil) alone. The animals were sacrificed 24h after the last treatment, sera were separated from trunk blood after decapitation. The total cholesterol level was determined by CHOD-PAP method. In comparison to control rats, soy isoflavones genistein and daidzein significantly decreased the level of serum cholesterol (by 23% and 15%, p<0.05, respectively). However, this decrease was smaller in comparison to the effects of chronic testosterone treatment, which lowered the same parameter by 27%, p< 0.01. Interestingly, estradiol did not change the level of serum cholesterol in ORH males. These results indicate that both soy isoflavones, genistein and daidzein have a positive effect on cholesterol status in middle-aged ORH rats, but genistein was more efficient than daidzein in decreasing the serum cholesterol. regard to sensitivity, having a quantitative detection limit of 2-10 nM for isoflavones and their metabolites from a 1 ml serum sample. In the present study, analysis of isoflavones has been carried out on an ABI-Sciex 4000 Qtrap mass spectrometer using heated nebulizer atmospheric pressure chemical ionization (HN-APCI)-MRM-MS. Sera from normal subjects and sera spiked with known amounts of isoflavones and their metabolites and internal standards were treated with βglucuronidase/sulfatase and extracted with ether. Extracts were analyzed by gradient reverse-phase HPLC-HN-APCI-MRM-MS. RESULTS: Although sensitivity differences existed between individual isoflavones, the quantitative detection limit of each isoflavone was 10 fmol or better. This translates to a concentration limit of as little as 0.1 nM from a 1 ml serum sample, or 1 nM from 0.1 ml sample. Unlike previous applications, this analysis was performed using 15 different channels, enabling a wide range of isoflavones, polyphenols and their metabolites to be examined simultaneously. Chrysin (5,7-dihydroxyflavone) was superior to apigenin as an internal standard for this analysis. CONCLUSIONS: Analysis of isoflavones in a modern triple quadrupole mass spectrometer by HN-APCI-MRM-MS permits their accurate and specific quantification even in sera from subjects on non-soy control diets, or from small animals. This research was supported by the Purdue-UAB Botanicals Center for Age-Related Disease (P50 AT00477) and a NIH/NCRR Shared Instrumentation Grant (S10 RR19231). Isoflavone Metabolism Absorption of Soybean Isoflavones in Isolated Rat Small Intestine. H. Ma, Z. Han*, S. Zou, and G. Wang, Key Lab of Animal Physiology & Biochemistry, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, Jiangsu, China. The present study aims at investigating absorption mechanism of soybean isoflavones, daidzein and genistein, in the small intestine of male rats in vitro. The content of daidzein and genistein in and out of the reverted small intestinal sacs and in the mucous using reverted small intestinal sacs were determined by RP-HPLC, at the same time the activity of ATPase in the mucous was assayed to study the absorption of daidzein and genistein. Results showed that the time of the highest absorption of daidzein and genistein were 2~3 h and 3~4 h in the reverted small intestinal sacs, respectively. The absorption rate was gradually decreased with the concentration of isoflavone increasing. The concentration of isoflavone in the mucous exhibited distinct dosedependent and time-dependent. Daidzein and genistein could significantly enhance the activity of total ATPase in small mucous. This indicates that the absorption of soybean isoflavone in rat small intestine in vitro was mainly by active transportation. High Sensitivity, Quantitative LC-MS Analysis of Isoflavones and Their Metabolites in Physiological Fluids. K. Jones, R. Moore, S. Barnes, University of Alabama at Birmingham, Birmingham, AL,USA. Comparison of the in vitro Metabolism of Isoflavones by Fecal Flora from Human Flora-Associated Mice and Human. M. Tamura1 and H. Saitoh2, 1National Food Research Institute, Tsukuba, Ibaraki, Japan, 2Biotechonology and Food Research Institute, Fukuoka Industrial Technology Center, Kurume, Fukuoka, Japan. Quantitative analysis of isoflavones and their metabolites in physiologic samples from clinical and small animal studies is an important research endpoint. Although LC-electrospray ionization multiple reaction ion monitoring mass spectrometry (LC-ESI-MRM-MS) has been a gold standard for analysis of many xenobiotics and is very specific, it is inferior to radioimmunoassay and electrochemical detection with There has recently been great interest in the biological activity of equol, especially with regards to the prevention of osteoporosis and prostate and breast cancer. In this study, we investigated the in vitro metabolism of daidzein, daidzin, and genistein by fecal flora, in human flora-associated mice (HFA mice) and a human donor. The in vitro incubation of feces with isoflavones demonstrated that we had successful- 19 2005 Abstracts Assessment of Dietary Isoflavone Intake by the 24hour Urinary Excretion in Japanese Women. M. Mori1, M. Sagara1, H. Mori2, and Y. Yamori1,3, 1Research Inst. for Production Development, Kyoto, Japan, 2Institute for Health Restoration, Nishinomiya, Hyogo, Japan, 3International Center for Research on Primary Prevention of Cardiovascular Diseases, Kyoto, Japan. Objective: Recent reports indicate the importance of soy Isoflavones(ISO) is the risk reduction of lifestyle related-diseases. Mostly the amounts of ISO taken form diets are estimated from nutritional survey or by 24-hour urinary ISO excretion. We measured 24-hour urinary ISO excretion in 137 healthy volunteers given different doses of ISO tablets (ISO glycosides : 40-150 mg) supplemented in their daily diets in order to clarify the relationship between the dietary intake and urinary excretion of ISO. Design and Methods: After obtaining informed consent 137 healthy Japanese pre-and-post menopausal volunteer women (40-63 years old) were asked to restrict the daily intake of common soybean diets such as tofu, soymilk and fermented soybeans for 7 weeks. From the 3rd to 7th week different doses of ISO tablets from 40 mg to 150 mg of ISO glycosides (20.9% daidzein, 4.5% genistein, 10.5% glycitein) were given for 4 weeks. Questionnaires about dietary customs were asked and health surveys including blood sampling and 24-hour urine collection were carried out before and after the ISO tablet administration. Results: The amount of urinary Isofravone excretion were confirmed to be 6.78±10.98, 21.82±11.62, 42.77±28.41, 62.59±37.98, and 69.25±38.7 umol/day in the participants given 0, 40, 75, 100 and 150 mg of dietary ISO supplements, respectively. Therefore, the association of the urinary ISO excretion (UI) with the amount ISO intake (DI) was : DI (mg)=2.22UI (umol/day). Conclusions: Our study indicates that the daily dietary intake of ISO (DI) can be estimated by analyzing the amount of 24-hour urinary ISO excretion(UI) from the equation : DI(mg)=2.22UI (umol/day). Bioavailability of Soy Isoflavones as Affected by Gender, Age, and Food Matrix in Rats. E. Sepehr*1,5, P. Robertson1, G. S. Gilani1, G. M. Cooke2,5, B. P-Y Lau3, And J. Fournier4, 1Nutrition Research Division, Health Product and Food Branch, Health Canada, Banting Research Centre, Ottawa, ON, Canada; 2Toxicology Research Division, Health Product and Food Branch, Health Canada, Banting Research Centre, Ottawa, ON, Canada, 3Food Research Division, Health Product and Food Branch, Health Canada, Banting Research Centre, Ottawa, ON, Canada, 4Animal Resources Division, Health Product and Food Branch, Health Canada, Banting Research Centre, Ottawa, ON, Canada, 5Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, ON, Canada. There is considerable interest in consumption of plant-based foods rich in phytoestrogens, more specifically the isoflavones and their derivatives from soy, due to their reported beneficial effects such as their potential to prevent breast and prostate cancer as well as prevent coronary heart disease and osteoporosis. However, there is still insufficient information regarding the pharmacokinetics of the absorption and excretion of soy isoflavones. Moreover, data on the effects of sex, age and of the source of isoflavones on their bioavailability are limited. The present study was conducted to obtain information on the effects of sex, age and source of soy isoflavones on their bioavailability in a rat model. Three sources of isoflavones; Novasoy (extracted isoflavones supplement), a mixture of synthetic aglycosides (daidzein, genistein and glycitein) and a mixture of synthetic glycosides (daidzin, genistin and glycitin) were administered once only by oral gavage at 20 or 40 mg/kg body weight. The first phase of the animal project has been completed, plasma samples were collected at 0, 10, 30 min and 1, 2, 8, 24, 48hr, and urine and fecal samples at 0-2,2-8,8-24,24-48hr post dosing. The second phase of the animal project will measure the concentrations of isoflavones and their metabolites in plasma following a single intravenous injection of synthetic aglycosides (daidzein, genistein and glysitein) and synthetic glycosides (daidzin, genistin and glysitin). The extent of absorption will be determined for each respective dosage form by comparing the area under the curve (AUC) of the plasma-concentration time curve after intravenous (iv) administration with that following oral administration. The extent of bioavailability will then be calculated as: F =AUCoral/ AUCiv x 100. Primarily results showed a significant gender difference in uptake of isoflavones in male and female Sprague Dawley rats. No equol production was observed within the first 48hr post dosing in male rats. However, female rats started producing equol after 24hr post dosing. Uptake of isoflavones was lower in old rats (20 months) compared with young rats (3 months)and old rats were able to produce equol as early as 8hr post dosing. The bioavailability data will assist in assessing safety, nutritional quality and health benefits of dietary phytoestrogens. Intra-Individual Variability of Phytoestrogen Excretion During Three Months Soy Consumption. S. Rebello1, A. Duncan2, K. Wangen1, W. Thomas1, and M. Kurzer1, 1University of Minnesota, St Paul, Minnesota, USA, 2University of Guelph, Ontario, Canada. Adaptation to long-term soy exposure has not been well characterized. To better understand this, we studied changes in urinary isoflavonoid excretion in 11 postmenopausal women (7 equol excretors and 4 non-excretors) who consumed soy protein isolate containing 65 mg of isoflavones per day for 3 months. Urinary concentrations of Odesmethylangiolensin (ODMA), daidzein, equol, genistein and glycitein were measured by GC/MS at baseline and at the end of months 1, 2 and 3. Excretion of all phytoestrogens increased from baseline to month 1 (p < 0.05). ODMA and daidzein excretion at month 2 tended to be higher than at month 1 (p < 0.20). At month 3, genistein excretion tended to be higher (p=0.148) and glycitein excretion was significantly higher (p < 0.05) than at month 1. When the group was divided into equol excretors and non-excretors, the trends toward increased ODMA and glycitein excretion were only observed in non-excretors (p < 0.20). In Poster Abstracts ly transferred the potential for equol production from human feces to HFA mice. There was no significant difference in the concentration of equol produced by incubation of feces from HFA mice in daidzein or daidzin. The average equol concentrations were, however, higher in feces from HFA mice than in those from their donor. In feces from both HFA mice and their human donor, the ratio equol/daidzein was significantly higher in the suspensions incubated with daidzin than with daidzein. The amount of genistein produced by incubation with genistein was significantly higher in fecal suspensions from HFA mice than human. The present study is the first to compare the equol production by fecal flora from human feces with those from HFA mice in vitro. Our results suggest that degree of daidzein metabolism by intestinal flora might be different from the degree of genistein metabolism by intestinal flora. The HFA mice produced in our experiment might also be useful for investigating the biological activity of equol in detail. 6th International Symposium on the Poster Abstracts 20 Role of Soy in Preventing and Treating Chronic Disease equol excretors, there was a trend toward an increase in equol excretion throughout the study. One subject did not excrete equol at month one although she became an equol-excretor at months 2 and 3. Changes in equol excretion tended to be inversely related to changes in ODMA excretion in equol excretors (R= -0.64; p=0.119). Within-subject coefficients of variation tended to be low for ODMA (11%), daidzein (7%) and genistein (11%), and higher for glycitein (19%) and equol (20% in non-excretors and 39% in excretors). Thus, equol excretion in particular is highly variable, in both excretors and non-excretors. Food Matrix-Assisted Control of Urinary Isoflavone Metabolite Profiles in Humans. K. Riedl1, T. Bohn1, M. Rogers2, Y. Vodovotz1, S. Clinton1, and S. Schwartz1, 1The Ohio State University, Columbus, OH, USA, 2University of Michigan, Ann Arbor, MI, USA. Health benefits due to soy consumption have been suggested to result from biotransformation of isoflavones by gut bacteria to biologically active components. Daidzein can be metabolized in sequence to dihydrodaidzein (dhD) and equol or o-desmethylangolensin (ODMA). We hypothesized that the food matrix could affect isoflavone metabolism and designed a soy bread and soy-tomato juice to test this hypothesis in a human feeding trial. Subjects (n=18) consumed 3-4 slices of soy bread (75-100 mg of isoflavone/d) for 3 days collecting a 24 h urine on the third day and 11 subjects consumed 2 x 177 mL cans of tomatosoy juice (81 mg of isoflavones/d). Isoflavone conjugates in 2 mL urine (with internal standard 2’,4’ dihydroxy 2-phenyl acetophenone) were digested enzymatically and extracted twice with diethyl ether. Extracts were pooled, dried and redissolved in 350 µL MeOH then analyzed by LC-MS/MS calibrating with authentic standards (genistein, glycitein, daidzein, ODMA, dhD, equol, dihydrogenistein, 6-OH ODMA). When comparing the two matrices there was a dramatic difference in the profile of urinary daidzein metabolites: consumers of the tomato-soy juice showed only 24% accounted for by equol and ODMA in contrast to 58% for consumers of soy bread. In subjects consuming the juice, a portion of daidzein was metabolized to dhD and both compounds readily absorbed. The bread matrix apparently delayed daidzein absorption such that more reached the distal gut microflora and was converted to equol and ODMA. This study demonstrates that the extent of daidzein metabolism can be modulated by altering the food matrix. Funding was provided by USDA-grant No M-01-RR00034 and the University of Michigan General Clinical Research Center, grant #M01RR00042. Pharmacokinetic Characteristics of Bonistein™ (Synthetic Genistein) in Humans. U. Ullmann1, J. Metzner2, and H. Oberwittler3, and J. Elliott1, 1DSM Nutritional Products, Kaiseraugst, Switzerland, 2Galmed, Halle/Saale, Germany, 3Institute for Clinical Pharmacology, Bobenheim, Grunstadt, Germany. Bonistein™ is a novel product consisting of 99.4% synthetic genistein. It is a dietary ingredient being developed for bone health. Up to now, Bonistein™ pharmacokinetics (PK) were assessed in three clinical phase I trials in young healthy men and women and in postmenopausal women. Single doses of 30, 60, 150 and 300 mg (bolus) and repeated dosing OID (up to 2 weeks, steady state (SS)) of 30, 60 and 120 of Bonistein™ were investigated. Plasma genistein (fraction unbound) and its conjugates (total genistein) were determined by a standardized LC/MS analytical method using D4-genistein as an internal standard. Orally ingested Bonistein™ is rapidly absorbed from the gut. In doses up to 150 mg, AUCs and Cmax values increased dose-proportionally. At higher doses the intestinal absorption seems to be limited and the plasma increase is less than dose-proportional. Average elimination half-lives (t1/2) were calculated to be 8.4 h (bolus) and 10.8 h (SS), respectively. Subgroup analysis (ANOVA) showed a slightly better bioavailability of Bonistein™ in females. In postmenopausal women Bonistein™ was more bioavailable and had a prolonged terminal elimination half-life. In all studies the volunteers ingested Bonistein™ after an overnight fast and remained fasted for several hours p.a. In conclusion, Bonistein™ reaches PK steady state after 4 to 5 days and behaves dose-proportional up to 150 mg. No systemic accumulation occurs. Bonistein™ was safe and well tolerated. Further Development of a Biomarker of Isoflavone Intake. J. Mackinnon1, E. Vink1, A. Blake2, T. Rawjee1, A. Riches1, and M. Ritchie1, 1Bute Medical School, University of St. Andrews, Fife, Scotland, 2Scottish Crop Research Institute, Invergowrie, Dundee, Scotland. OBJECTIVES: To validate 24-hour urine collections (24hU), timed spot urine samples and timed blood samples as biomarkers of isoflavone intake up to 100mg/day. METHODS: Healthy volunteers consumed 50mg/d or 100mg/d soy isoflavones (ISF) or placebo (one week of each) in a randomised, double-blind, cross-over study. 24hU, timed spot urine samples and timed blood samples were obtained at baseline and at two points during each intervention. ISF content of the samples was assayed by LC-MS. Background dietary intake was assessed using food diaries and a validated database of the ISF content of foods. RESULTS: Significant correlation (r = 0.5083, t=3.686, n = 42, p <0.001) was observed between genistein concentration in timed spot urines and dietary ISF intake. The correlation between ISF intake and 24hour urinary genistein excretion was also significant (r = 0.59, t = 2.42, p <0.04). Average urinary excretion of genistein (in timed spot urines) corresponding to intakes of 100mg isoflavones/day was 1.8 times greater than that for 50mg isoflavones/day (p = 0.024). Plasma samples from this group and from a further 40 volunteers are awaiting analysis. CONCLUSIONS: 24hU, timed spot urine samples and timed blood samples have previously been validated as biomarkers of ISF intakes up to 60mg/d. This study demonstrates that dietary and/or supplemental intake of ISF up to 100mg/d can be assessed using timed spot urines and 24hU. These samples can therefore be used as an accurate, objective measure of high ISF intakes in future observational and intervention studies. This study was funded by The Melville Trust for the Care and Cure of Cancer, Edinburgh, Scotland. Isoflavone tablets were kindly supplied by Lamberts Healthcare Limited, Kent, England. Monkeys Exhibit Sex Differences in Behavioral and Physiological Responses to a High-Isoflavone, SoyBased Diet. J. Kaplan1, M. Adams1, N. Simon2, J. Wagner1, and A. Franke3, 1Wake Forest University School of Medicine, Winston-Salem, NC, USA, 2Lehigh University, Bethlehem, PA, USA, 3Cancer Research Center of Hawaii, Honolulu, HI, USA. We have shown previously that male monkeys deriving their dietary protein from soy are more aggressive and less affiliative than casein fed controls. Here we systematically compare the social and physiological responses of group-housed, soy (n = 31) and casein treated (n = 30) male monkeys to the responses of similarly housed pre- 21 2005 Abstracts Determination of the Factors that Influence the Ability of Equol Production. S. Vanhemmens, K. Decroos, and W. Verstraete, LabMET, University of Ghent, Belgium. Recent research reveals the key role of equol in the beneficial health effects of a soy diet. This metabolite, formed by colon bacteria from daidzein, has a superior antioxidative and estrogenic action and a better absorption compared to its precursor. Unfortunately, only 35% of the Western population is able to form this metabolite. The identity of the responsible bacteria, the microbial-ecological mechanisms and the nutritional factors that influence this conversion are part of our research at LabMET. We obtained a stable and transferable mixed culture of four bacteria (EPC4) transforming daidzein into equol. These strains were identified as Lactobacillus mucosae, Enterococcus faecium, Finegoldia magna and a yet unknown Veillonella sp. related strain. This mixed culture was used to evaluate the influencing factors during in-vitro equol production. It was found that hydrogen gas plays a crucial role. The speed of equol production was related to the hydrogen gas concentration in the headspace and in the absence of hydrogen gas there was no equol production anymore. Research with the SHIME (Simulator of the Human Intestinal Microbial Ecosystem) has shown that equol production takes place in the last part of the colon. However, methanogenic and sulfate reducing bacteria use also a lot of hydrogen gas and are also established in the last part of the colon. To investigate if there is a competition for hydrogen gas between equol producing bacteria and other hydrogen gas consuming bacteria, both in-vitro and in-vivo research is done at LabMET. The in-vitro research includes the incubation of pure cultures of methanogenic and sulfate reducing bacteria with EPC4 in different combinations. These results show that EPC4 has a higher affinity for hydrogen than methanogenic and sulfate reducing bacteria because the production of methane and hydrogen sulfide was reduced while equol production was the same when EPC4 was incubated with the methanogenic and sulfate reducing bacteria. When 1% of EPC4 was added to faecal samples from non-equolproducers with either methanogenic or sulfate reducing abilities, the same results were obtained. Our current in-vivo research evaluates these results on human population level. The faecal samples of hundred menopausal women, consuming soy milk (Alpro Soja) or isoflavone tablets (Acatris) during 4 days, are analysed by real time PCR in order to quantify the methanogenic and sulfate reducing bacteria. Also other important bacterial groups are quantified. Equol in urine samples is analysed by HPLC. This could reveal the relation between the diversity in colon bacteria and the production of equol. The in-vivo study was conducted in cooperation with Acatris Holding BV (Giessen, The Netherlands) and Alpro Soya (Wevelgem, Belgium). S.V. benefits from a FWO scholarship (Fund for Scientific Research) Bioavailability of Isoflavones and Flavones Correlates with Human Gut Microbial Degradation. A. Simons, M. Renouf, S. Lee, S. Hendrich, and P. Murphy*, Iowa State University, Ames, IA, USA. The relationship between flavonoid structure, gut microbial degradation in vitro and human bioavailability in vivo was determined. Gut microbial fermentations from 33 healthy subjects (20 men, 13 women) were conducted with 15 flavonoids, genistein, daidzein, glycitein, apigenin, naringenin, kaempferol, luteolin, quercetin, myricetin, hesperetin, chrysin, flavone, 5,4’-dihydroxyflavone, 6,4’-dihydroxyflavone, 7,4’dihydroxyflavone, & 5,3’-dihydroxyflavone. Flavonoids with 5, 7 and 4’ – hydroxyl groups, genistein, apigenin, naringenin, luteolin, kaempferol and quercetin, rapidly disappeared compared to the other flavonoids (k = 0.46 ± 0.10 h−1 vs. 0.07 ± 0.02 h−1, p < 0.0001). The methoxylated flavonoids hesperetin and glycitein were rapidly demethylated and disappeared with k = 0.24 ± 0.21 h−1. Human bioavailability of flavonoids was investigated in 10 volunteers (5 men, 5 wome). Subjects ingested 28 mg (104 µmol) genistein and 16 mg (62 µmol) daidzein from soymilk, 422 mg (1549 µmol) naringenin and 8 mg (26 µmol) hesperetin from grapefruit juice and 115 mg (381 µmol) quercetin from sautéed onions with 1 week washout in between foods. Peak plasma concentrations of flavonoids analyzed over the 24 hour period ranged from 0.01 – 1 µM. The bioavailability, calculated as urinary excretion as the percentage of ingested dose, was significantly lower for the rapidly degraded flavonoids, genistein (7.2 ± 4.6 %), hesperetin (7.3 ± 3.2 %), quercetin (5.6 ± 3.7 %) and naringenin (3.2 ± 1.7 %) compared to daidzein (43.4 ± 15.5 %, p = 0.02). These data reveal that the chemical structure of flavonoids affect their gut microbial degradation rate and, thus, their overall bioavailability. This work was supported by the Center for Designing Foods to Improve Nutrition, Iowa State University, USDA Special Grant No. 20030679, Project No. 3302 of the Iowa Agriculture and Home Economics Experiment Station. Identification of Human Fecal Microorganisms and Human Fecal Microbial DNA Sequences Influencing Isoflavone Degradation: Putative Bioavailability Biomarkers. M. Renouf, A. Simons, and S. Hendrich*, Iowa State University, Ames, IA, USA. High fecal isoflavone disappearance rate, which predicts low isoflavone bioavailability in humans, may coincide with distinct fecal bacterial species. Fecal DNA was extracted from 33 adults, bacterial 16S rDNA variable region sequences amplified by PCR and separated by denaturing gradient gel electrophoresis (DGGE). The four highest genistein and glycitein degraders, as measured by HPLC analysis of in vitro anaerobic fecal incubations, shared 5 DNA bands of greater intensity than in feces of the 4 lowest degraders. Some of these fecal microbial DNA sequences corresponded to known human gut microbes Poster Abstracts menopausal monkeys consuming the same soy (n = 46) or casein (n = 45) based diets for comparable periods of time (≥ 20 months). The soy protein contained 1.88 mg/isoflavones/g, approximately equivalent to a human consumption of 130 mg isoflavones/day. In contrast to the males, dietary soy had no significant effects on the aggressive behavior of females. Furthermore, while soy-fed males exhibited reductions in affiliation, soy consumption increased sociality among females. Males and females also exhibited disparate physiological responses to the diet. Hence, soy significantly increased insulin sensitivity in females while reducing it in males. Among the plasma lipids, soy increased HDL and reduced LDL cholesterol concentrations to a significantly greater degree in females than males. Notably, females also exhibited significantly lower plasma concentrations of genistein, daidzein and equol than males, despite consuming the same diet. In sum, a soy proteinbased diet relatively high in isoflavones had uniformly salutary effects on premenopausal monkeys but had effects that were either less beneficial or adverse to males. These sex differences may relate to interactions with ovarian or testicular hormones (which were unchanged by soy treatment), or to differences in isoflavone metabolism (as reflected by plasma concentrations). Supported in part by grants from the National Heart Lung and Blood Institute (HL045666, HL079421) and the National Cancer Institute (CCSG CA71789). Soy protein supplied by Solae Corporation. 6th International Symposium on the Poster Abstracts 22 Role of Soy in Preventing and Treating Chronic Disease (Bacteroides, Prevotella and Clostridium groups. But several fecal microbial DNA sequences differing between high and low isoflavone degraders were not identifiable using BLAST (NIH system for DNA sequence identification). In nutrient-rich and nutrient-poor in vitro fecal incubation systems, Bacteroides ovatus and acidifaciens, Eubacterium ramulus, Clostridium orbiscindens and Tannerella forsythensis were human gut microbial species that increased human fecal isoflavone degradation compared with control under both conditions. These species may be partly responsible for human gut microbial disappearance of isoflavones in both high and low isoflavone degraders. Prevotella pallens and oralis, Bacteroides eggerthii, fragilis and uniformis significantly increased fecal isoflavone disappearance only in nutrient-poor conditions, which may mimic increased fecal isoflavone degradation in humans with relatively slow gut transit time, as previously observed. Microbial species such as B. eggerthii, as well as several DNA sequences we discovered that correspond with currently unknown microbial species, may be useful biomarkers to distinguish subjects of low isoflavone bioavailability. Supported by Iowa State University Bailey Career Research Development Award. Varying Isoflavone Content of Soy Protein Affects Plasma Isoflavones but not Plasma Lipids in Surgically Postmenopausal Monkeys. J. Kaplan1, T. Clarkson1, M. Anthony1, T. Badger2, and H. Chen1, 1Wake Forest University School of Medicine, Winston-Salem, North Carolina, USA, 2Arkansas Children’s Hospital Research Institute, Little Rock, Arkansas, USA. Background: Soy protein compared to casein improves the plasma lipid profiles of monkeys, an effect often attributed to the isoflavones. In this study we fed monkeys diets containing equivalent amounts of soy protein that was either high or low in isoflavones to determine directly the effect of isoflavones on plasma lipids. Materials and Methods: 26 postmenopausal cynomolgus monkeys were used in a crossover study involving 2, 3-month treatment periods. During the first period animals consumed a diet deriving 37% of protein from soy and containing either 65 (Lo) or 141 (Hi) mg total isoflavones/1800 Cal. During the second treatment period animals crossed over to the opposite diet. Plasma HDLC, LDL+VLDLC and triglycerides (TG) were measured 3 times in each phase while plasma isoflavones were determined once in each phase. Results: Dietary isoflavone content significantly influenced plasma isoflavone concentrations (total isoflavones: Hi 553 vs. Lo 136 nmol/L, p<0.0001; genistein: Hi 98 vs. Lo 72, p=0.21; daidzein: Hi 112 vs. Lo 58, p=0.04; equol, Hi 333 vs. Lo 10, p<0.0001). In contrast, the two treatment diets were not associated with any significant differences in plasma lipid profiles (TG, Hi 44 vs. Lo 36 mg/dl, p=0.44; HDLC, Hi 47 vs. Lo 52, p=0.22; LDL+VLDLC, Hi 266 vs. Lo 293, p=0.13; TPC:HDLC, Hi 8.2 vs. Lo 8.3, p=0.90). Discussion: The results suggest that a doubling of dietary isoflavone content does not provide additional plasma lipid benefits, a negative outcome that was particularly surprising in view of the 30-fold difference in equol concentrations. In Vitro Study of Microbial Transformations of Daidzein in a Dynamic Model of the Gastrointestinal Tract. Karel Decroos1, Ellen Eeckhaut1, Steffi Vanhemmens1, Sam Possemiers1 and Willy Verstraete1, 1Laboratory of Microbial Ecology & Technology (LabMET), Department of Biochemical and Microbial Technology, Ghent University, Coupure Links, 653, B-9000 Gent, Belgium. Soy isoflavones get a lot of attention for their potential role in preventing chronic disease. An important issue with respect to bioavailability and bioactivity of dietary isoflavones is the metabolic fate after ingestion, in particular the microbial transformations in the colon, which are subjected to great interindividual variability. From the soy isoflavone daidzein about one third of the population forms the metabolite equol, with superior biological activities and better absorption. Up to date, very little is known about the environmental conditions in the colon, the dietary factors and the bacteria involved in this important microbial transformation. At the Laboratory of Microbial Ecology & Technology we developed a five-stage reactor system in which the human gastrointestinal tract is simulated (SHIME, Simulator of the Human Intestinal Microbial Ecosystem). This system consists of five reactors which each represent a particular part of the GIT, with the emphasis on the colonic part. Using a polyphasic approach we are able to monitor the transformations of food components (HPLC, LC-MS, IC, GC-MS) and the composition of the microbial community (PCR-DGGE, real-time PCR, flow cytometry). Previously, we isolated a unique microbial consortium (EPC4) from a human faecal sample efficiently transforming daidzein into equol (1). The microbial metabolism of soy isoflavonoids was examined in the SHIME with faecal inocula from different volunteers. The different daidzein-metabolizing phenotypes, observed from in vivo urinary excretion data, were confirmed in vitro. Supplementation of EPC4 to the SHIME, inoculated with a faecal sample from a non-producing individual and fed with a diet containing 3,5 g soygerm powder/d, led to high equol production (150µmol/L) in the distal parts of the colon. These results prove that the concept of using a probiotic for the stimulation of equol production is valid. Moreover, the SHIME was shown to be an excellent tool for the study of microbial transformations of dietary phytoestrogens. REFERENCES (1) K. Decroos, S. Vanhemmens, N. Boon & W. Verstraete (2005) Isolation and characterization of an equol-producing mixed microbial culture from a human faecal sample and its activity under gastrointestinal conditions. Arch. Micr. (183) 45-55 Effects of Isoflavone Supplements vs. Soy Foods on Blood Concentrations of Genistein and Daidzein. Christopher D. Gardner1, Lorraine M. Chatterjee1, Brian M. Oliveira1, and Adrian A. Franke2, 1Stanford Center for Research in Disease Prevention and the Department of Medicine, Stanford University Medical School, Stanford, CA, USA, 2Cancer Research Center of Hawaii, Honolulu, HI, USA BACKGROUND: Although interest in soy isoflavones and their possible health benefits have grown substantially, understanding of isoflavone pharmacokinetics is still limited. OBJECTIVE: The objective was to examine the pharmacokinetics of isoflavone concentrations over a 24-hour period among healthy adults consuming either soy foods or soy isoflavone tablets at different doses. DESIGN: Twelve generally healthy adults were randomly assigned to the order of three phases in a crossover trial. The three phases were isoflavone tablets at 1) 154 mg/day or 2) 308 mg/day, and 3) soy foods designed to provide a calculated 96 mg isoflavones/day, with daily intake divided equally among the three main meals (doses in aglycone equivalents). After 6 days on each of the study phases, plasma isoflavone concentrations were determined on the seventh day of dosing at 0, 4, 8, 10, 12 and 24 hours. RESULTS: Average levels of total isoflavone concentrations at the 23 2005 Abstracts Cardiovascular Disease Anti-Hypertensive Effects of Nicotianamine in Spontaneously Hypertensive Rat (SHR) and Tukuba Hypertensive Mice (THM). T. Sato1, A. Hayashi2, S. Tokutake1, A. Matsuyama1, M. Kikuchi1, and K. Kimoto2, 1R&D Division, Kikkoman Corporation, Noda City, Chiba, Japan, 2Dept. of Food and Nutrition, Tokyo Kasei University, Itabashi-ku, Tokyo, Japan. Nicotianamine (NA) is found in plants and has anti-hypertensive effect in vivo. However, the mechanism of its beneficial effect is not fully investigated. We examined the effects of NA administration on systolic blood pressure (SBP), NA serum concentration, and organ angiotensin I converting enzyme (ACE) activity using spontaneously hypertensive Rat (SHR) and Tukuba hypertensive mice (THM). The THM is a hypertensive animal model prepared by carrying human renin and angiotensinogen genes into C57BL/6 mouse. THM is known to cause hypertension by a single factor, an enhancement of the renin-angiotensin system (RAS). In SHR, Long-term and single administration of NA significantly decreased SBP. The serum concentration of NA has its peak at 1 h after oral administration, and there are no differences between feeding and 12-h fasting. In blood and lung, ACE activity was decreased at 2 and 6 h after NA administration. These data suggest that NA is absorbed directly soon after administration, is not influenced its absorption by diet, inhibits serum and lung ACE, and shows antihypertensive effects in SHR. We also measured NA concentration in the plasma of THM. It was confirmed that, after administration in the stomach, NA has been absorbed from the intestine and detected into the blood. Consequently, the blood pressure significantly descended until 6 hours after administration of NA. In THM, ACE activities in plasma, lung and kidney were also decreased by administration of NA. It was suggested that NA decreased SBP by inhibition of RAS in SHR and THM. It is expected that NA is used as functional compound to prevent and alleviate hypertension. Modulation of Hepatic Thyroid Hormone and Retinoic Acid Receptors May be a Novel Mechanism of Soy Hypolipidemic and Anticarcinogenic Effects. C.W. Xiao1,3, W. Huang1, C. Wood1, M.R. L’Abbé1, G.S. Gilani1, G.M. Cooke2,3, and I. Curran2, 1Nutrition Research Division, Food Directorate, Health Products and Food Branch, Health Canada, Ottawa, Ontario, Canada, 2Toxicology Research Division, Food Directorate, Health Products and Food Branch, Health Canada, Ottawa, Ontario, Canada, 3Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, Ontario, Canada. Thyroid hormones and retinoic acid (RA) are important regulators of growth, differentiation, tissue homeostasis and lipid metabolism. Thyroid hormone receptor (TR) and RA receptor (RAR) are involved in mediating carcinogenesis and anticarcinogenesis, respectively. Soy consumption has been shown to be hypolipidemic, and linked to lower incidence of certain types of cancers. The objective of this study was to examine the effect of dietary soy protein isolate (SPI) and soy-derived isoflavones (ISF) on TR and RAR content and function. Sprague-Dawley rats were fed diets containing either casein or increasing amounts of supplemental ISF (5-1250 mg/kg diet) or alcohol-washed SPI (5, 10, or 20%). Protein content and DNA binding abilities of TR and RAR were measured by Western blot and electrophoretic mobility shift assay, respectively. The content of TRβ1 and RARβ proteins in liver, but not in other tissues examined was significantly increased by dietary SPI in a dose-dependent manner compared with casein. Supplemental ISF had no effect. The content of other TR and RAR isoforms were not affected by SPI. In contrast, the binding abilities of hepatic TR and RAR to DNA were markedly inhibited by SPI. Additionally, plasma triglyceride levels and hepatic expression of TR-regulated genes for acetyl-CoA carboxylase-β and -β were suppressed by SPI compared with casein. Taken together, these results suggest that modulation of hepatic TRβ1 and RARβ, key regulators of gene expression involved in lipid metabolism and carcinogenesis, by soy proteins may be a novel mechanism by which soy components lower blood lipid level and prevent carcinogenesis. (Supported by Health Canada) Effect of Dietary GABA-Enriched Fermented Soybean (GABA-tempeh) on Blood Viscosity of Rats. Nakamichi Watanabe*1, Kenichi Nakatsugawa1, Kenshiro Fujimoto2, and Hideyuki Aoki3, 1Showa Women’s University, SetagayaKu, Tokyo, Japan, 2Koriyama Women’s University, Koriyama-Shi, Fukushima, Japan, 3Ikeda Tohka Industries Lab., Fukuyama-Shi, Hiroshima, Japan. Tempeh, a traditional soybean fermented food made by salt-free aerobic fermentation with Rhizopus microsporus, was originally developed in Central Java, Indonesia. The GABA-tempeh used in the present study was prepared by aerobic and successive anaerobic cultivation of boiled soybean with R. microsporus. The contents of free amino acids especially GABA (γ-amino butyric acid) and oligopeptides in the GABAtempeh were significantly higher than in the conventional tempeh. In this study, the blood viscosity of rats fed the GABA-tempeh, conventional tempeh or unfermented soybean were compared. Blood viscosity was measured using a micro channel array flow analyzer (MC-FAN) equipped with the same interval interstice siliconized gages, which are narrower than the average diameter of RBC much like a real capillary vessel. It has therefore been possible to determine whole blood viscosity with reliability and reproducibility. The blood viscosity of the rats being fed the GABA-tempeh was significantly lower than those given the conventional tempeh or unfermented soybean. However, there were no significant differences in blood ingredients such as hematocrit, triacylglycerol, total cholesterol, and glucose levels. The levels of lipid peroxidation products of serum and red blood cell were similar among the groups. GABA which is specifically rich in the GABA-tempeh may be responsible for the lower blood viscosity. Poster Abstracts peak time points of 8, 10 and 12 hours were >4 µmol/L for the soy food phase and for the higher dose tablet phase. Genistein concentrations were highest overall in the soy food vs. supplement phases of the study (p<0.05). When comparing plasma concentrations for the two doses of tablets, saturation appeared more evident for genistein than daidzein. CONCLUSIONS: Most previous pharmacokinetic studies have investigated a single daily bolus of isoflavones from just one type of source. These results contrasting source and dose of isoflavones, when taken three times daily, indicate important differences in the pharmacokinetics of genistein and daidzein and suggest that soy foods may lead to higher isoflavone bioavailability than the isoflavone supplements used in this investigation. 6th International Symposium on the Poster Abstracts 24 Role of Soy in Preventing and Treating Chronic Disease Cognitive Function Soy Foods Exposure Predicts Better Baseline Cognitive Function in Healthy Older Adults. S. Meade1,2, N. Lane1,2, T. Ohrt1,2, S. Asthana1,2, and C. Gleason1,2, 1University of Wisconsin, Dept. of Medicine, Sect. of Geriatrics, Madison, WI, USA, 2Madison VA GRECC, Madison, WI, USA. In this randomized, placebo-controlled, double-blind design clinical pilot study, the effects of soy isoflavones on cognition were studied in men and women over the age of 60. Data were collected from 30 cognitively healthy older adults (50% women). Subjects received either 100 mg/day of soy isoflavones or a placebo for 6 months. Neuropsychological testing was administered at baseline and at 1, 3, and 6 months after starting the treatment phase. In addition, Food Frequency questionnaires were administered to monitor the intake of soy foods and products throughout the study. The baseline Food Frequency questionnaires were used to determine whether the subjects were exposed to soy isoflavones before entering the treatment phase (soy-exposed), or not exposed to soy isoflavones (soy naïve). Soyexposed subjects are those having a weekly intake of soy foods between 6.58 and 4434 grams. Soy-naïve subjects are those having a weekly intake of soy foods equal to zero grams. There were no significant differences between the two groups in terms of age, education level, and Mini-Mental State Examination scores. On cognitive data, however, the soy-exposed group outperformed the soy-naïve group on tests of executive function and language. Specific findings include better baseline performance on the Trail Making Test (t=2.332, p=0.03), Letter Fluency test (t=2.539, p=0.017) and Category Fluency test (t=2.958, p=0.006). Interestingly, the soy-naïve group showed greater response to one month of isoflavone treatment than the soy-exposed group on Trail Making Test (F=3.277 p=0.053). It seems that soy exposure may impart cognitive benefits for older adults. NIA grant K23 AG24302. Research staff and study volunteers from the Wisconsin Comprehensive Memory Program. The General Clinical Research Center: grant M01 RR03186 from the National Center for Research Resources, National Institutes of Health. Madison GRECC of the Department of Veterans Affairs. Archer Daniels Midland provided Novasoy® brand isoflavones tablets and matching placebo Saponins Interaction Between Soybean Saponin and Protein, and Possible Functionality. M. Shimoyamada1, S. Ikedo2, R. Yamauchi2, and K. Watanabe3, 1Miyagi University, Sendai, Miyagi, Japan, 2Gifu University, Gifu, Gifu, Japan, 3Tokyo University of Agriculture, Atsuki, Kanagawa, Japan. Saponin, which is widely distributed in plant kingdom as well as soybean, is a kind of natural surfactants, and it is expected to have some affinity to proteins. So interaction between saponin and protein extracted from soybean seeds was evaluated by a modified equilibrium dialysis, gel filtration chromatography and protease hydrolysis. The equilibrium dialysis and gel filtration chromatography of soybean saponin and protein mixture demonstrated that a large proportion of saponin interacts weekly with soybean protein. Soybean acid precipitated protein (APP), which was prepared from water extract of soybean seeds, contained about 1 % of saponin. Water extract was subjected to a hydrophobic resin column and removed almost all of saponin (NS-APP). Three kinds of protein fraction, namely, APP, NS-APP and a mixture of NS-APP and saponin were incubated with chymotrypsin. As the result, hydrolysis level by chymotrypsin was shown to increase by saponin removal and reversibly decrease by saponin readdition. Similar results were shown in tryptic hydrolysis of saponinprotein mixture. As soybean was usually utilized after heat treatment, each APP fraction with or without saponin was heated, cooled and hydrolyzed by chymotrypsin. The effect of saponin on hydrolysis level of the heated APP was similar to the effect on the non-heated APP. These data may suggest that saponin controls digestion of soybean protein to change functionality of APP in gut, e.g. cholesterol-lowering activity. Development of a Saponin Rich Soybean: Relationship Between Saponin Content and Genes Controlling the Polymorphism of Saponin Composition. C. Tsukamoto1, I. Tayama1, Y. Takada2, M. Kamada3, and K. Kitamura3, 1The Graduate School of Agricultural Sciences, Iwate University, Morioka, Iwate, Japan, 2National Agricultural Research Organization, National Agricultural Research Center for Tohoku Region, Kariwano, Akita, Japan, 3The Graduate School of Agriculture, Hokkaido University, Sapporo, Hokkaido, Japan. The health claims of soybean saponins depend on the total amount of saponins and on the chemical structure of the individual saponin molecules. Although the variation in seed saponin composition is explained by combinations of genes controling the utilization of soyasapogenol glycoside substrate, factors affecting total saponin content have not been studied. The Sg-4 gene, which controls arabinosylation of the glucuronic acid residue attached at the C-3 position of sayasapogenols, is expressed in the seed cotyledons of 100% of G. max and G. soja lines we have tested to date. When we tested the same collections for Sg-4 expression in the hypocotyls, we found only 2% of the G. max lines expressing Sg-4 in the hypocotyls versus 63% for the G. soja lines. Saponin content in the hypocotyls of G. soja lines was much higher than that of G. max. To study the relationship between the total saponin content and the expression of Sg-4, recombinant inbred lines (RILs) were developed from crosses between lines with and without the expression of Sg-4 in the hypocotyls. Quantitative saponin analysis in the cotyledons and hypocotyls of 264 RILs (F7) showed that the total saponin content was not affected by the Sg-4 gene but that Sg-4 influenced the composition and quantity of individual saponins. Examination of total saponin content of two mutant lines, one possessing a new aglycone and the other lacking soyasapogenol A, showed the same tendency. Total saponin content in soybean seeds seems to be dependent on the supply of soyasapogenols. Soy Saponins Display High Content and Profile Variability in Isoflavones Enriched Dietary Supplements. M. Berger1, J. Hubert2,1, and J. Daydé1, 1Ecole Supérieure d’Agriculture de Purpan, Toulouse, France, 2Genibio Recherche, Lorp Sentaraille, France. Soybean presents a remarkable phytochemical profile, including many health-promoting polyphenolic and triterpenoid glycosilated compounds. An increasing number of isoflavone extracts are available and consumed as dietary supplements in western countries, and the process employed for the manufacturing of these products can also modulate their content in soy saponins. Using simple low cost HPLC/UV analyses, our objective was to investigate the group A and B soy saponin and isoflavone contents and profiles in various commercial soy-dietary sup- 25 2005 Abstracts Poster Session II Tuesday, November 1, 2005 .........7:30 a.m.–5:00 p.m. Wednesday, November 2, 2005....7:30 a.m.–12:00 p.m. Diabetes A Novel Functional Food Ingredient—Soy ProteinCatechins Complex. C. Kuo and S. Chen*, Food Industry Research & Development Institute, Hsinchu, Taiwan. The beneficial effects of both soy and tea are well studied, such as antiangeogenesis, antihyperglycemia, antiobesity, and lowering the risks of cardiovascular diseases. There have being numerous attempts to combine these two to provide synergistic effects. Although it has been shown that isoflavones from soy and green tea have synergistical antitumor effects, soy proteins readily precipitate with catechins. And, it’s generally believed that the formation of soy protein-catechins precipitates leads to less bioavailabilities/biofunctions and may even hinder nutrient absorption. In this study, we set out to characterize soy proteincatechins precipitates and their potential as a novel functional food ingredient. Soy milk, soy okara, or defatted soybean were mixed with tea under different pHs (4, 7, 9) and temperatures (25 or 80oC). Freeform catechins and caffeine contents in the supernatants were analyzed by HPLC, and protein patterns by SDS-PAGE. It’s found that more than 70% of EGCG and less than 50% of caffeine of tea solution co-precipitated with proteins when soy milk was mixed with tea catechins at 1:5 under pH 4. Temperature has no effects on both the amounts of freetype catechins and of soy protein-catechins precipitates. Only 10% of the precipitated EGCG could be extracted by water, suggesting that EGCG and soy protein precipitates co-precipitate to form complexes. Surprisingly, more than 70% of EGCG immediately dissociated from the precipitates when subjecting to simulated gastric fluid. No difference between the protein banding patterns of both soy milk and soy proteincatechins complexes was observed. These results suggest that (1) the complexes formation have little effects on the bioavailabilities of ether EGCG or soy proteins, (2) soy proteins can used as adsorbents for EGCG partial purification, reducing the usage of organic solvents, and (3) EGCG-soy proteins complexes may serve as a novel functional ingredient providing additive or synergistic beneficial effects. Glycemic Response to Selected Soy Foods in Selected Diabetics and Development of Software on Diabetes with Special Reference to Soy and Glycemic Index. R. Chithra and S. Deepa, PSG College of Arts & Science, Coimbatore-14, Tamil Nad, India. Anthropometric measurements, food and nutrient intake and other data related to diabetes were collected from 30 Type - II diabetics of Chennai city. Soy products incorporated foods like chapathi, pongal and pancakes were formulated and evaluated organoleptically. Nutritive value of the recipes and the quantities of recipes to be used for the study of glycemic response (GR) were ascertained. The GR to standard and soy foods were studied in the selected patients. User friendly software was developed and the programming was done in such a way that it gave detailed nformation about diabetes with special reference to types, etiology, signs, diagnosis, prevention, complications, dietary modifications, foods to be included and avoided etc. Energy expenditure for various activities, glycemic index of various foods, diabetic diets based on caloric needs, food exchange lists and importance of soy foods in glycemic control were also provided. Forty per cent of the diabetics had grade I obesity and 26 per cent grade II. Most of them were taking oral hypoglycemic drugs and also had control over their diet. The mean food and nutrient intake were lower than the recommendations of Indian Council of Medical Research. The formulated recipes were highly acceptable recording high protein and low carbohydrates. The mean incremental blood glucose level on ingestion of the soy recipes was significantly lower than that of the respective standards showing the superiority of soy products in glycemic control. Software developed was interactive for diet counseling and will be of great help to diabetics and dietitians. PSG COLLEGE OF ARTS & SCIENCE Proximate Composition, Glycemic Index, and Glycemic Load of Rice, Diabetic Diet Rice and Soy Incorporated Selected Recipes of India. R. Chithra, S. Chitralekha, and P. Gayathri, PSG College of Arts & Science, Coimbatore, Tamil Nadu, India. Diabetics are recommended foods with low lycemic index (GI). Soy bean has low GI. Specially processed rice called ‘diet rice’ is claimed to have undergone a treatment where starch is complexed to reduce the GI. Common rice based Indian recipes viz., idli, dosa, pancake, appam, adai, soy dosa, soy pongal, vegetable rice, mint rice, rice with dhal powder, kolaputtu and string hoppers were standardized and organoleptically evaluated. Nutritive value and portion sizes were determined. Fasting blood glucose levels were studied in 10 suitable volunteers. The reference food (Glucose) and test foods prepared from ordinary rice (IR20) and diet rice were administered on different days. Blood glucose levels after 30, 60, 90 and 120 minutes were registered. GI and GL were determined followed by statistical analysis. All the recipes were acceptable. The GI of vegetable rice made of ordinary rice was 51 and that made of diet rice reduced to 31 showing a maximum reduction of 39%. The GI of pancakes and soy dosa reduced by 33% and 31% respectively on diet rice substitution. All the other recipes also uniformly recorded considerable reduction in GI varying from 13 to 29%. GL also followed the same trend. Soy Dosa registered significantly lower GI and GL than ordinary Dosa showing the beneficial effect of soy bean on glycemic control. Many varieties of rice and more soy recipes may be studied taking all other interfering factors into consideration to arrive at a more valid conclusion. PSG COLLEGE OF ARTS AND SCIENCE, COIMBATORE AND CHAMAN LAL SETIA EXPORTS, HARYANA, INDIA Dietary Intake of Isoflavones and Lignans is Related to Lower Prevalence of Metabolic Syndrome and Improved Insulin Sensitivity. Yvonne van der Schouw, Poster Abstracts plements. Very high concentrations of saponins were observed in several products, from 5.5 µmol/g to 107.8 µmol/g. The saponin A/B ratio ranked from 0.3 to 8.6, reflecting the raw soy material employed during processing (germ vs cotyledon). Some products contained 4-fold higher amounts of isoflavones than saponins. Others contained comparable quantities of both. Interestingly, our results show that some products contained up to 2-fold higher amounts of saponins than isoflavones. These significant amounts of saponins in products which are purchased as isoflavone concentrates only have to be taken into account. Indeed, soy saponins may interact with isoflavone biological activities or contribute to many of the health benefits attributed to isoflavones. Both of these soy secondary metabolites have to be clearly determined when discussing the biological activity of soy- based dietary supplements. 6th International Symposium on the Poster Abstracts 26 Role of Soy in Preventing and Treating Chronic Disease Majon Muller, and Diederick Grobbee, UMC Utrecht, Utrecht, The Netherlands. Background: The effect of phytoestrogens on insulin sensitivity and the metabolic syndrome is largely unknown. Methods: We studied the association between dietary isoflavone and lignan intake and characteristics of the metabolic syndrome in a population-based cross-sectional study comprising 400 independently living men between 40 and 80 yr of age. Dietary intakes of isoflavones and lignans was calculated from a validated food frequency questionnaire. The metabolic syndrome was defined according to the National Cholesterol Education Program definition, and insulin sensitivity was calculated by use of the quantitative insulin sensitivity check index. We used logistic and linear regression analysis adjusted for age, smoking, alcohol intake and physical activity. Results: Increasing isoflavone intake was associated with decreased prevalence of the metabolic syndrome [odds ratio (OR) for fourth versus first quartile 0.46; 95% confidence interval (CI), 0.23-0.92]. Increasing lignan intake was also associated with decreased prevalence of the metabolic syndrome [OR for fourth versus first quartile 0.39; 95% CI, 0.19-0.79]. Additional adjustment for insulin attenuated the associations with both isoflavones and lignans. Increasing isoflavone intake was associated with increased insulin sensitivity [difference (D) between fourth and first quartile 0.014; SD 0.005]. Increasing lignan intake was also associated with increased insulin sensitivity [difference (D) between fourth and first quartile 0.019; SD 0.005]. Conclusions: Higher dietary intake of isoflavones as well as lignans in aging males are independently associated with a higher insulin sensitivity and a reduced prevalence of the metabolic syndrome. The pathway for this effect might be through decreasing insulin levels. Genistein Inhibits High Glucose-Induced MonocyteEndothelial Cell Interaction Through a CampDependent Protein Kinase Pathway. W. Zhen, H. Si, and D. Liu*, Virginia Polytechnic Institute and State University, Blacksburg, VA, USA. Atherosclerosis is the major vascular complication in diabetic patients. Genistein, a soy isoflavone, has anti-diabetic and anti-atherosclerotic effects, but the mechanisms involved are still unclear. Adhesion of Monocytes to the vascular endothelium is one of the key steps in the development of atherosclerosis. We investigated the hypothesis that genistein, at physiological concentrations, inhibits leukocyte-endothelial interaction under hyperglycemic condition. Following treatment of human aortic endothelial cells (HAEC) with glucose, release of IL-8 and monocyte chemotactic protein-1 (MCP-1) was measured by ELISA, and mononuclear cell adhesion was determined by U937 cell binding. Protein kinase A (PKA) activity was determined by measuring kemptide phosphorylation. High glucose (25 mM) induced IL-8 and MCP-1 production by 1.3 and 1.5 fold over the normal glucose (5.5 mM) (n=4, p<0.01). Genistein (0.01-10 micromole) does dependently inhibited high glucose-induced IL-8 and MCP-1 production. Genistein also inhibited high glucose-induced U937 cell adhesion at physiologically relevant concentrations (0.1- 1 micromole) (n=4, p<0.01). This effect was not blocked by antagonism of estrogen receptor. In addition, genistein at 15 micromole did not change tyrosine kinase activity. Treatment of cells with selective inhibitors for protien kinase A partially ablated the inhibitory effect of genistein on IL-8 and MCP-1 production and monocyte adhesion (n=4, p<0.01). Exposure cells to genistein for 30 min increased the protein kinase A activity 1.7 fold over control (n=4, p<0.01). These results demonstrate that genistein inhibits high glucoseinduced chemotactic cytokine production and mononuclear cell adherence to human endothelial cells, at least in part, through activation of PKA, suggesting a novel anti-inflammatory mechanism for genistein and protective role in hyperglycemia-induced endothelial dysfunction. The Glycemic Index and Insulin Index of Selected Soy Foods. Robert M. Blair1, Aaron Tabor1, E.C. Henley2, 1Physicians Pharmaceuticals Inc., Kernersville, North Carolina, USA, 2EC Henley Consulting, Athens, Georgia, USA. Foods with a low glycemic index (GI) may provide a variety of health benefits. The objective of the present study was to measure the GI and insulin index (II) of selected soy foods. The study was conducted in two parts with “low-carb” products being tested separately. In Experiment 1, subjects averaged 23.2 years of age with BMI=22.0, while subjects in Experiment 2 averaged 23.9 years of age with BMI=21.6. The reference (glucose) and test foods were served in portions containing 10 grams of carbohydrates in Experiment 1 and 25 grams of carbohydrates in Experiment 2. Subjects consumed the reference food twice and each test food once. For each test, subjects were instructed to consume a fixed portion of the reference food or test food together with 250 grams of water within 12 minutes. Blood samples were collected before each test and at 15, 30, 45, 60, 90, and 120 minutes after consumption of reference or test foods to quantify glucose and insulin. Two-hour blood glucose and plasma insulin curves were constructed and areas under the curves were calculated. GI and II values for each subject and test food were calculated. In Experiment 1, both “low-carb” soy foods were shown to have significantly (P<0.05) lower GI and II values than the reference food. In Experiment 2, three of the four test foods had significantly (P<0.05) lower GI and II values than the reference food. In conclusion, all but one of the soy foods tested had a low GI (i.e. <55). Immune Function Genistin at the Concentration Present in Soy-Based Infant Formula Inhibits Rotavirus Infectivity in vitro Through Inhibition of Protein Tyrosine Kinase Activity. A. Andres1, S.M. Donovan1, T.B. Kuhlenschmidt2, and M.S. Kuhlenschmidt1,2, 1Div. Nutritional Sciences, University of Illinois, Urbana, IL, USA, 2Dept. Veterinary Pathobiology, University of Illinois, Urbana, IL, USA. Rotavirus (RV) is the most common cause of infantile diarrhea, with 111 million episodes each year worldwide. No RV vaccine is currently available, thus, it is critical to identify other means to reduce the incidence and severity of RV infection. Isoflavones have anti-viral activity and we previously evaluated the anti-RV activity of aglycone, glycoside and acetyl-glycoside forms of genistin, daidzin, and glycitin, as well as the MIX of all isoflavones at soy formula concentrations and found that genistin alone and the MIX were effective. Herein, the anti-RV activity of the MIX, genistin and genistein were further investigated by a focus forming unit assay in MA104 cells. Genistin (14 µg/ml) and MIX (42 µg/ml) decreased (p<0.05) RV infectivity vs. control over a 16-fold dose of RV concentrations. In contrast, the MIX without genistin did not inhibit RV infection, implicating genistin as the active component. Further, genistin and genistein at the same concentration were equally inhibitory, suggestion bioconversion of genistin to genistein and leading us to investigate the cellular actions of genistein as potential mechanisms of action. A protein tyrosine kinase inhibitor (Herbimycin), a topoisomerase II inhibitor (Ectoposide), and an estrogen agonist (17-β estradiol) were tested. Only Herbimycin decreased (p<0.05) RV infectivity vs. control. In summary, genistin at the concentration found soy infant formula inhibits 27 2005 Abstracts Menopausal Symptoms Soybean Composite Functional Factors Enhanced the Serum Estradiol Concentration and Superoxide Dismutase Activity in Postmenopausal Women. Dan Li*1,2, Changcheng Gao2, Xing Chen2, Yuzhen Huang2, and Xiaolei Li1, 1Dept of Food Sci.& Eng. in Changchun University,Changchun 130022, P.R. China, Changchun, Jilin Province, P.R. China, 2National Reseach and Popularize Center for Soybean Refined Processing, Changchun 130022,P.R. China, Changchun, Jilin Province, P.R. China. Soybean composite functional factors (SCFF), containing 0.72% isoflavones, 1.06% saponin, 24.0% oligosaccharide, 5.2% protein and 7.2% nucleic acid, were extracted using membrane technology. Oral administration of SCFF at 30mg/day was carried out in 10 volunteers from postmenopausal women. Radio immune and chemical illuminance assay were employed respectively to determine the serum estradiol (E2) concentration and superoxide dismutase (SOD) activity. After 20 days, the increase of 43.5% for E2 concentration and 55.6% for SOD activity were found. The maximum tolerance dose of SCFF was 75g/kg body weight. As a result, SCFF was considered as a dietary supplement for the reduction of menopause symptoms. The composition of soybean composite functional factors ingredients unit value isoflavone mg/g 7.2 saponin mg/g 10.6 oligosaccharide % 24.00 protein % 5.2 nucleic acid % 7.2 folic acid Ìg/g 0.057 nicotinic acid Ìg/g 340 thiamin Ìg/g 17 riboflavin Ìg/g 1870 carotene Ìg/g 45.8 calcium mg/kg 629.48 zinc mg/kg 149.50 iron mg/kg 73.9 selenium mg/kg 0.0133 Authors would like to give our many thanks to Changchun People pharmacy Co. Ltd for their support. Effects of a Novel Extract of Daidzein-Rich Isoflavone Aglycone on Dehydroepiandrosterone Production in Japanese Menopausal Woman. Yoshiaki Sato*1 And Weijun Pan2,3, 1Sophia Ladies Clinic, Kanagawa 229-0033, JAPAN, 2Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA, 3Nichimo Co., Ltd., Tokyo 140-0002, Japan. Dehydroepiandrosterone sulfate (DHEA-S) level in blood decline sharply with aging, it may play a protective role in many chronic diseases. The effects of soy isoflavone on DHEA-S production in menopausal women are unclear. We determine the effect of a novel of daidzein-rich isoflavone aglycone extract from soy germ fermentation with Koji fungus (AglyMax), an effective antioxidant on DHEA-S production in 40 Japanese menopausal women in relieving menopausal symptoms. An 8-week openlabel feasibility study included 40 Japanese menopausal. All subjects are taking with a supplement of 20-40 mg/day of AglyMax for 8 weeks. Subjects recorded their menopausal symptoms score activity weekly by Koyama’s Simplified Menopause Index (SMI). Routine blood count, chemistry, blood levels of sex hormones and DHEA-S were measured on baseline and 8-week. Blood fluidity were measured by MC-FAN (Micro Channel Array Flow Analyzer) both baseline and 8-week. Forty subjects completed the study. AglyMax improved vasomotor symptoms. The score of SMI decreased from 68 ± 20 at baseline to 40 ± 12 at 8 weeks (p <0.05). DHEA-S was increased from 890 ± 112 ng/ml at baseline to 2,430 ± 210 ng/ml at 8 weeks (p <0.01). There were no significant changes in the measured blood chemistries or other hormone levels. 78.1% of the subjects were improved their blood fluidity after 8 week. The study suggests that daidzein-rich isoflavone aglycone (AglyMax) supplement may be improve DHEA-S production that may overlap with weak estrogenic effect of soy isoflavone to preventing treating for menopausal symptoms. Impact of Supplementation of Soybean Chikki as a Source of Isoflavin on Plasma Calcium Levels of Menopause Women. Ch. Kavitha and K. Krishna Kumari*,ANGR Agricultural University, Hyderabad, Andhra Pradesh, India. Women during menopause stage undergo physiological and emotional stress and 72% of Indian women (above 40 years) suffer from osteoporosis. The isoflavins, exclusively present in soybean are found to relieve menopausal symptoms and considered as an important component for bone health. An investigation has been made to assess the impact of supplementation of soybean chikkias a source of isoflavin on bone health of menopause women. The plasma calcium level was selected as a parameter to assess the bone health. A traditional recipe chikki was prepared with puffed soybean and jaggery at 1: 1 proportion. Chikki 100g contains 19.2 g protein, 8g fat,155mg calcium,370 Kcal energy and 190 mg isoflavins. Soybean chikki (50g) containing 95 mg isoflavins had been supplemented daily to two groups (50-55 years and 55-60 years) of menopause women with 15 subjects in each group for a period of 90 days through careful monitoring. The effect of supplementation was assessed through food and nutrient intake, anthropometry, menopausal symptoms and plasma calcium levels of subjects in the beginning of the study and after 45 and 90 days of supplementation. It was observed that the menopausal symptoms like backache and irritability disappeared in the subjects after supplementation. There was no significant change in body weight and BMI of subjects. Among the nutrients the intake of energy was sufficient in both the groups, while the intake of protein (63 & 67g), fat (46 & 49g) and calcium (885 & 894mg) were more than RDA in both the groups in the beginning and the levels were further increased towards the end of supplementation. The intake of isoflavins were 95mg from soybean chikki throughout the study period, though it was nil in the beginning. The initial plasma calcium level (6.78 & 7.43mg%)of both the groups were found below normal (8.7mg%) in spite of the diet appears to supply double the RDA of calcium. The levels were increased from 6.78 to 7.74mg% in group I and 7.43to 8.33 mg% in group II by 45 days of supplementation. The levels were further increased to 8.65&9.32 mg% respectively by 90 days of supplementation. It is interesting to observe that in group II (55-60 years), the plasma calcium levels have Poster Abstracts RV infection in vitro across a continuum of severity in part via inhibition of the protein tyrosine kinase signaling pathways. (Supported by AG 03-34505-13320 and the Illinois Soybean Program Operating Board) 6th International Symposium on the Poster Abstracts 28 Role of Soy in Preventing and Treating Chronic Disease reached almost the normal levels by 45 days of supplementation. This increase in calcium level might have been due to the enhanced absorption or altered metabolism of calcium through soy protein and isoflavins consumed from soybean chikki. The authors wish to acknowledge the ANGR Agricultural University,Hyderabad,India for providing the facilities to carry out this study. Obesity Effects of Dietary Soy on Adipose Tissue, Adipocytokines, and Insulin Sensitivity. K. Ingram, J. Kaplan, K. Kavanagh, L. Zhang, and J. Wagner, Wake Forest University School of Medicine, Winston-Salem, North Carolina, USA. Soy supplements are used widely for a number of health benefits. We have shown previously that soy protein containing isoflavones improves insulin sensitivity in female monkeys. Our objective in this study was to determine if changes in adipose tissue mediated the improvement in insulin sensitivity. Premenopausal female cynomolgus monkeys consumed either a soy protein diet (n=46) with isoflavones at human equivalent dose of 130 mg isoflavones/d or a casein/lactalbumin (n=45) diet for 2 years. Abdominal subcutaneous and visceral fat biopsies were collected for cell size determinations (cellular triglyceride and DNA quantification). Plasma leptin, adiponectin, and active and desacyl ghrelin were determined by ELISA. Body fat, as measured by DEXA, body weight, and thyroid hormone concentrations (T3 and T4) were also measured. There was no change in body weight or body fat over the two years. However, soy fed monkeys had fewer (p=0.03) and larger (p=0.06) subcutaneous adipocytes but no change in visceral adipocytes. Consistent with the larger adipocytes, soy decreased adiponectin (p=0.01) and increased leptin (p=0.02). Active (p<0.001) and desacyl (p=0.03) ghrelin were increased with soy as were thyroid hormones (T4, p<0.05, T3, p<0.10). Ghrelin is associated with increased appetite, adipogenesis, and growth hormone release, which elevates resting metabolic rate independent of changes in body composition. Leptin and thyroid hormones also regulate energy expenditure, by increasing uncoupling protein 3 in muscle. We propose that soy may affect metabolic efficiency not by changes in adipose tissue, but by effects on other insulin sensitive tissues such as muscle to improve insulin sensitivity. Osteoporosis Effect of Soymilk With and Without Isoflavones on Insulin-Like Growth Factor 1. Results from a Two-Year Clinical Study. E. Lydeking-Olsen1, A. Juul2, N.E. Skakkebaek3, K.D.R. Setchell4, and J.-E. Beck Jensen4, 1Institute for Optimum Nutrition, Denmark, 2Copenhagen University Hospital, Rigshospitalet, Denmark, 3Childrens Hospital and Medical Center, USA, 4Copenhagen University Hospital, Hvidovre, Denmark. Background: The insulin-like growth factor (IGF) system has been linked to the process of bone acquisition through epidemiological analyses of cohorts and in vitro studies of bone cells. Earlier, short-term or epidemiological studies have revealed conflicting results with respect to the effect of soy and its isoflavones on IGF 1 levels, as studies in males have revealed that 3 mo consumption of soy+ (88 mg iso) increased IGF1 significantly, compared to milk protein (Khalil 2002). An epidemiological study demonstrated an association between soy / isoflavone consumption and increasing levels og IGF1 in males, but not in females (Probst-Hensch 2003). In postmenopausal women, one 3 mo interven- tion study have found that if not on HRT, soy+ (88 mg total isoflavones) increased IGF-1 97% (p<0,05, Arjmandi 2003) and another, that increasing levels of soy isoflavone intake, decreased levels of IGF -1 (Wangen 2000). No long term studies exist. Aim: The aim of the study was to compare the long-term effects of soymilk, with or without isoflavones, on levels of insulin-like growth factor 1, after 2 years and relate this to response to treatment effect on BMD in postmenopausal women. Methods: Postmenopausal, Caucasian women with established osteoporosis or at least 3 risk-factors for osteoporosis, were randomly assigned, double-blind to two treatment-groups: soymilk containing 76 mg isoflavones as aglycones (soy+, n=23) or isoflavone-poor soymilk, (control, soy÷ n=21). All subjects received comparable intakes of calcium, minerals and vitamins. Serum IGF-1 were analysed by RIA and bone mineral content (BMC) and density (BMD) were measured in lumbar spine and hip by using dual-energy X-ray absorptiometry (DEXA) at baseline and after 2 years. Findings: IGF-1 levels are shown in table, demonstrating an inverse relationship between soy - and soy +, as soy - increases and soy + decreases IGF-1 levels, respectively. The percentage change in lumbar spine BMD and BMC respectively, did not differ from zero in the soy+ group (+1,1%, +2,0%) but significant bone loss occurred in the soy/control-group (÷·4,2%, ÷·4,3%).No significant changes occurred for femoral neck BMD or BMC. No correlation was found between change in IGF-1 and BMD response in either treatment group. Interpretation: Daily intake of two glasses of soymilk containing 76 mg isoflavones decreased IGF-1 levels, and soymilk without isoflavones increased IGF1 levels, but no correlation was found in relation to BMD response. Effect of soy milk with and without isoflavones on serum-IGF 1, after 2 years treatment* Soy -, control, n=21 Soy+, n=23 p-value** IGF 1, start, (mcg/L) 190,9±10,7 203,4±14,8 0,50 IGF 1, 2 y, (mcg/L) 204,4±16,9 188,9±13,3 0,47 Difference, (mcg/L) 13,6±8,3 -14,5±7,8 0,02 Relative difference, % 5,4 -5,1 0,05 *means values SEM **t-test Phytoestrogen Excretion is Associated with Improved Markers of Bone Health in Australian Women. K. Hanna1, J. Wong2, G. Eaglesham3, C. Patterson1, S. O’Neill2, and P. Lyons-Wall*1, 1School of Public Health, Queensland University of Technology, Australia, 2Betty Byrne Henderson Centre, Royal Brisbane & Women’s Hospital, Australia, 3Pathology and Scientific Services, Queensland Health, Australia. Phytoestrogens may protect against osteoporosis by exerting estrogenic actions on bone cells in postmenopausal women with low serum estrogen concentration. The aim was to examine associations between phytoestrogens and biomarkers of osteoporosis. Subjects were a cross-section of 141 Australian women aged 40-60 y and participating in the Brisbane Longitudinal Assessment of Ageing in Women (LAW). Phytoestrogen status was estimated by measuring excretion of nine isoflavonoids and four lignans in three 24-hr urines, utilizing HPLCMS/MS. Bone mineral density (BMD) of femur, total hip and lumbar spine was measured by dual energy x-ray absorptiometry. Bone formation was assessed by serum bone alkaline phosphatase (bone ALP) and osteocalcin (OC); resorption was assessed by urinary-N-terminal crosslinking-telopeptide of type-I-collagen (NTX) and deoxypyridinoline (DPD). Analyses were conducted in the total group and sub-groups with 29 2005 Abstracts Effects of Genistein, Polyunsaturated Fatty Acids, and Vitamins D3 and K1 on Bone Metabolism in the OVXRat Model and the OVX-Dog Model of Osteoporosis. S. Krammer2, U. Wehr1, W. Rambeck1, and P. Weber2, 1Institute of Animal Physiology, Ludwig-MaximiliansUniversity Munich, Munich, Germany, 2Animal Nutrition and Health R&D, DSM Nutritional Products, Basle, Switzerland. Osteoporosis and low bone mass are currently estimated to be a major public health concern. Research is focusing on various nutrients and their effects in the prevention and treatment of osteoporosis. Trials with laboratory animals have played a major role in recent osteoporosis research. As no animal model represents human conditions in total, the use of more than one animal species is advisable to get a broader view. The aim of the two studies was to evaluate the effects of a combination of Genistein (15 mg Genistein/kg BW), PUFA (5 % of DM), and Vitamins D3 (1500 IU Vitamin D3/kg BW) and K. (2000 µg Vitamin K1/kg BW) on preventing the subsequent loss of bone mass in rats and dogs induced by ovariohysterectomy (OHX). The dog’s bone resorption measured as urinary pyridinoline excretion doubled in the control group (A) in the weeks after OHX (see Figure). This strong increase was clearly prevented by the administration of the combination of the substances (group B). Similar effects were observed in rats (urinary pyridinoline excretion in nmol/mmol Crea in week 12 after OHX: 303.7 ± 42.5 versus 417.8 ± 58.2 for treated and control animals respectively). *: Statistically significant difference (p<0.05) between the two groups at one time point Figure: Urinary pyridinoline excretion in dogs The results showed that the combination of the tested substances reduced a high bone turnover and therefore can have a protective effect on osteoporotic bone. The reproducibility of the results in two different models of osteoporosis was demonstrated. Supplement of Soy Isoflavone and/or Calcium Can Enhance Bone Density in Growing Rats. Yuling Lin*, Hsuching Cheng, and Jimmy Tsai, Department of Bioscience Technology, Chung Yuan University, Chung Li 32023, Taiwan. The aim of this study was to investigate the effect of dietary soy isoflavone extract and/or calcium on bone growth and density in young growing rats. Fifty-six 3-weeks-old Wistar female growing rats were randomly divided into 7 groups, 8 each. Animals were fed 0.18% Ca (1/4Ca), 0.36% Ca (1/2Ca), 0.72% Ca (AIN93G-mineral mix, Ca), 0.18% Ca + 0.15% soy extract (0.027% Isoflavone, 1/4CaS), 0.36% Ca + 0.15% soy extract (1/2CaS), or 0.72% Ca + 0.15% soy extract (CaS) diet for 3 months. Animals in one of the groups were ovariectomized at the beginning of trial and then fed CaS diet. After feeding the experimental diets, bone mineral density (BMD) of femurs and tibiae were measured using dual energy x-ray absorptiometry (DEXA). Data demonstrated that increasing intake of Ca and/or soy isoflavone enhanced the BMD of femurs and tibiae (p<0.05). Results also showed that supplement of soy isoflavone could enhance the numbers of osteoblasts and the thickness of growth plate in tibiae (p<0.05). Trabecular mineral apposition rates (MAR) showed a similar trend as BMD (p<0.05). Data also revealed that ovariectomy in growing rats reduced BMD significantly (p<0.05). In conclusion, both Ca and soy isoflavone intakes could enhance bone growth and BMD in growing rats. Effects of Isoflavone and Calcium on Bone Cell Activities and Their Biomarkers in Growing Rats. Hisu-Ching Cheng* and Jimmy Tsai, Department of Bioscience Techonology, Chun-Yuan Christian University, Chungli 32023, Taiwan. Calcium (Ca) is one of the fundamental elements for bone growth and metabolism. Soy-isoflavone (S) has been reported to act as an estrogen-like compound to prevent bone loss. Effects of various levels of Ca and S combinations on the activities of osteoblasts (OB) and osteoclasts (OC) in growing rats were investigated. Fifty-six 3 weeks old female Wistar rats were housed individually in stainless steel cages. Forty-eight of them were assigned into 6 dietary groups randomly, 8 each, and fed 1/4Ca, 1/2Ca, Ca (AIN-93G recommendation), or with 0.027% isoflavone in the diets (1/4CaS, 1/2CaS, CaS). Eight of them were ovariectomized and fed CaS diet (CaSOx). All rats were fed for 12 weeks. Two bone formation markers, serum bone-specific alkaline phosphatase (BAP) and C-terminal propeptide of Type I collagen (CICP), and 2 bone resorption markers, serum pyrinodiline (Pyd) and N-terminal telopeptide of bone collagen (NTx) were determined. Bone sections by histomorphometry were also employed to determine the numbers of OB and/or OC. Results demonstrated that increasing Ca intakes enhanced bone formation and reduced bone resorption significantly. Data of analyses revealed that supplement of S promoted the activities of OB and suppressed the activities of OC. The bone sections also showed that S supplement increased OB numbers and decreased OC numbers alone the surface of trabecular bone significantly, especially in CaS group comparing with others. It was also observed that ovariectomized rats had less OB activities and higher OC activities comparing with normal rats fed same diet (CaS). The conclusion is that dietary S supplement can enhance Ca bioavailabilities and bone formation, and reduce bone resorption. Poster Abstracts body mass index (BMI) <25 kg/m2 or osteopenia/osteoporosis. After adjusting for confounders, results showed a negative association between isoflavonoids and NTX in women with lower BMI (r=0.33, P<0.05); a positive association between lignans and bone ALP in the total sample (r=0.21, P<0.05) and women with osteopenia/osteoporosis (r=0.41, P<0.05); and a trend towards a positive association between lignans and OC (r= 0.30, P=0.06). There were no associations between phytoestrogens and BMD. Inverse associations between isoflavonoids and NTX, and positive associations between lignans and bone ALP or OC, support a mechanism whereby phytoestrogens act to suppress bone resorption while stimulating formation. Findings are not consistent with observed actions of hormone therapy and endogenous estrogens, which suppress overall turnover by reducing both resorption and formation. 6th International Symposium on the Poster Abstracts 30 Role of Soy in Preventing and Treating Chronic Disease A Novel Extract of Fermented Soybean Germs (AglyMax) Promoted Bone Growth in Ovariectomized Mice Fed a High-Fat Diet. C. Wang1, W. Pan2,3, L. Huang1, and J. Zhou2, 1Human Nutrition Program, Kentucky State University, Frankfort, KY 40601, USA, 2Nutrition/Metabolism Lab, Beth Israel Deaconess Medical Center at Harvard Medical School, 330 Brookline Ave. Burlington-5, Boston, MA 02215, USA, 3Biotics R&D Division, Nichimo Co., Ltd., Higashi-Shinagawa, Shinagawa-ku, Tokyo 140-0002, Japan. Daidzein is one of the most abundant isoflavones present in soy and it is unique as it can be further metabolized to equol, a compound with greater estrogenic activity than other isoflavones. The aglycone forms of daidzein may be more potent biologically than its conjugated form. Our preliminary study showed that a novel extract of fermented soybean germs (AglyMax) that is rich in daidzein aglycones promoted bone growth in male mice. Seventy percent of the extract by weight was isoflavones with daidzein:genistein:glycitein aglycones in the ratio of 7:1:2. The objective of this study was to determine the effects of AglyMax on bone in ovariectomized mice. The mice (6 weeks old) were randomly assigned into the treatment groups. A baseline group was killed at the beginning of the experiment. Group A was sham-operated but fed the high-fat diet with 60% of the energy from fat; Groups B, C and D were all ovariectomized and fed the high fat diet containing 0, 1.2 or 5 g of AglyMax per kg diet, respectively. The food intake was kept similar for all the groups. At the end of the 12-week experiment, femur bones were analyzed for bone mineral content and density by dualenergy X-ray densitometry (Lunar DPX-IQ). Each bone was also evaluated by a three-point bending test for its breaking strength. Groups A and D had significantly higher bone mineral content, bone density, breaking strength, bone ash and calcium content than Groups B and C (all at P≤0.01). The baseline group was lowest in all the indicators measured. These results suggest that AglyMax at 0.5% of the high fat diet promoted bone growth in ovariectomized mice. Soy Protein Isolate and Moderate Exercise Independently and Additively Impact Bone Turnover but not Bone Mineral Density in Postmenopausal Women. Ellen M. Evans, Susan B. Racette, John O. Holloszy, and Dennis T. Villareal, Division of Geriatrics and Gerontology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO, USA Dietary soy protein and exercise are lifestyle approaches used to manage menopause-related osteoporosis; however the interaction of these interventions on bone health is unknown. The aim of this study was to assess the independent and additive effects of soy protein isolate (SOY) and moderate-intensity exercise (EX) on bone turnover and bone mineral density (BMD). A 2 (SOY vs. milk protein isolate; MLK) by 2 (EX vs. control) study design was used, with 61 women randomized and 43 (62±5 y) completing the 9 mo intervention (SOY; n = 10: MLK; n = 12: SOY+EX; n = 11: MLK+EX; n = 10). BMD was measured by DXA. Serum CTX and BSAP were measured as markers of bone resorption and formation, respectively. Although no main effects of SOY or EX were evident at 6 mos; SOY reduced CTX (-13.3±15.3% vs. -1.5±21.0%; P = 0.023) and BSAP (-4.7±14.7% vs. 6.5±17.7%; P = 0.016) at 9 mos. EX attenuated the reduction in S-CTX (-1.9±21.6% vs. -12.4±15.3%; P = 0.038); however, no EX effects were apparent in BSAP at 9 mos (2.8±16.1% vs. -1.0±18.3%; P = 0.28). Neither SOY or EX impacted BMD at any site; however, change in BMD was related to the change in fat mass (r = 0.40, P < 0.05). The data suggests that 1) SOY appears to reduce bone turnover with no impact on BMD over 9 mos and 2) the potential benefits of moderate exercise on bone turnover and BMD may be modulated by change in fat mass. Other Isoflavones Do Not Show Astringent Taste in Soy Foods. M. Abdullah Al, C. Tsukamoto, and T. Ono, The United Graduate School of Agricultural Sciences, Iwate University, Japan, Ueda 3-18-8, Morioka 020-8550, Iwate, Japan. Health beneficial effect of soybean rich in isoflavones have been well studied. The normal daily requirement of soy isoflavone for health beneficial effect is 50-90mg / day. Normally, isoflavone concentrations (mg/100g of fresh wt) in soymilk and tofu have 20-40mg. Therefore, more consumption of isoflavone rich soy beverage/food should require as daily basis. Soymilk and tofu have been consumed as a traditional food in East Asian countries and is gradually spreading to the Western world. Unfortunately, soymilk is not well accepted for its bean flavor and astringent taste. The improvement of the bean flavor was successfully achieved, nevertheless, factors that impact astringent flavor have remained. Many studies have revealed that soybean isoflavones are responsible for the astringent taste in soyfoods. But no consistency was found between an astringent taste and isoflavone contents. Isoflavone-enriched extract showed no astringency. Soybean foods having large amounts of isoflavones showed less astringency. Meanwhile, there is no report considering why tofu curd is less astringent. Phytic acid combines with protein and calcium during tofu curd formation. Therefore, phytates might have effect of trapping the astringent components to prevent them from interacting with taste cells of the mouth and throat. A 0.1% potassium phytate solution at pH 7 showed the very same astringency as soymilk; however, calcium phytate at the same concentration and pH showed no astringency. Thus soluble phytic ions in soymilk may cause astringency. The strategy of utilization of soybean for food should be increased of health beneficial components and decrease of unfavorable taste. Study on the Development and Characteristics of Chinese Soybean’s Health Effects. X. Mingzhong, Xichang College, Xichang, Sichuan, P.R. China. The health effects of soybean is a result of the development of Chinese people’s food culture. It dates back to ancient China’s Zhou, Qin, Tang and Song Dynasties, and reaches its peak after 1949. There are three characteristics to Chinese soybean’s health effects. One is the government’s support, another is rich content, the third is the sophisticated techniques. There are two aspects in its health effects: relieving hunger and clinical effects. It also features simple and inexpensive processing methods, extensive application and regional differentiation. According to traditional Chinese medicine, soybean has a quieting and comforting effect. It can help spleen function and diuresis, produce blood and add to strength, balance dry and moisture, reduce heat and detoxicate. Thus, soybean is effective for symptoms such as a weak spleen and lack of energy, a weak body and poor appetite, inadequate blood and lack of strength, oedema and difficulty in urinating, poisoning, during pregnancy and skin ulcer. Black soybean is effective for dizziness, a faint eyesight, early whitening of hair, diarrhea and abdomen on pain, drug poisoning and drunkenness. Therefore, soybean is called “the king among beans” and “vegetable meat”. In this paper, the author gives an account of some clinical foods made of soybean and other ingredients which can be used to cure or prevent some common diseases. These foods can improve the function 31 2005 Abstracts Suitable Control Diets for Use in the Study of Phytoestrogens in, or Derived from, Soybeans. C. Benton1, J. Odum2, and G. Tobin1, 1Harlan Teklad Madison, Wisconsin,, USA, 2Syngenta Central Toxicology Laboratory, Macclesfield, Cheshire, UK. In recent years there has been considerable interest in the role of the soybean isoflavones and their metabolites on mammalian physiological systems. Among these effects is the stimulation of uterine growth in immature or ovariectomized animals and the advance of puberty in females that occurs when laboratory rodents are fed on diets containing soybean meal. To eliminate the contribution of soy isoflavones, it seems reasonable to use a standard purified diet such as AIN-76A as a control for studies on phytoestrogens or other estrogens. Our studies have shown that in practice, this may not be a satisfactory alternative. We, and others, have shown that the uterotrophic response and timing of vaginal opening are affected as much by a high-energy intake as by high levels of phytoestrogens. Most standard purified diets such as AIN76-A and AIN-93G contain a high level of energy, and in a form that leads to rapid absorption. Thus control animals fed on such diets may show a pseudo-estrogenic response before the application of an estrogenic treatment. This may not be recognized by the researcher, and may mean that the true estrogenic effects of the treatment are underestimated, and perhaps even obscured, in such circumstances. Such concerns may be assuaged by the use of low-energy purified diets or low-energy natural-ingredient diets that contain little or no soybean meal as a control or base diet. In our studies both these approaches resulted in minimal stimulation of uterus growth and prevented an abnormal advance in puberty in female animals. Isoflavones Protect Mice from Radiation-Induced Weight Loss. M. Landauer, J. Kramer, and V. Srinivasan, Armed Forces Radiobiology Research Institute, Bethesda, MD, USA. There is a need to develop medical countermeasures to protect first-responders, remediation workers, and patients undergoing radiotherapy from the harmful effects of ionizing radiation. We previously demonstrated that the isoflavones genistein and daidzein protect mice from lethal doses of ionizing radiation when administered 24 hours before irradiation. In a clinical setting, radiation is administered at sublethal doses to destroy tumor cells. A frequent side effect of sublethal irradiation in mammals is a reduction of body weight. Moreover, weight loss has been demonstrated to be a prognostic indicator of reduced survival rates. Therefore, the amelioration of radiation-induced weight loss is an important objective when using radiation therapy. In the present study, we characterized the effects of the isoflavones genistein and daidzein on gamma radiation-induced weight loss in CD2F1 male mice. A single injection of genistein or daidzein (5 mg/mouse) or vehicle was subcutaneously administered 24 hr before either sham irradiation or a sublethal dose of cobalt-60 gamma irradiation (7 Gy at 0.6 Gy/min). Mice were divided into six groups: (1) vehicle + sham irradiation, (2) genistein + sham irradiation, (3) daidzein + sham irradiation, (4) vehicle + 7-Gy irradiation, (5) genistein + 7-Gy irradiation and (6) daidzein + 7-Gy irradiation. Animals were weighed daily for 30 days after irradiation. We found that mice treated with vehicle or isoflavones that were sham-irradiated exhibited normal weight gain over the 30 days of the experiment. In contrast, a significant reduction in body weight for all groups of irradiated animals was observed. Beginning on day 3 after irradiation, both the genistein/irradiation and daidzein/irradiation groups exhibited decreased weight loss compared with the vehicle/irradiation group. By day 20 after irradiation, the body weight of the isoflavone/irradiation groups had returned to control levels. However, the body weight of irradiated mice that did not receive genistein or daidzein remained significantly below that of the other groups. These results demonstrate that genistein and daidzein can mitigate weight loss in mice receiving a sublethal dose of gamma radiation. Soybean Extracts Enhance Elastin in Skin. R. Zhao1, J.C. Liu2, Ch. Bertin3, J.P. Ortonne4, M. Seiberg1, and V. IotsovaStone*1, 1Skin Research Center, Johnson & Johnson Consumer Products Worldwide, Skillman , NJ, USA, 2Global Skin Care Growth Platform, Johnson & Johnson Consumer & Personal Products Worldwide, Skillman , NJ, USA, 3Scientific affairs, J&J Consumer Europe, les Moulineaux, , France, 4CPCAD , Hopital l’Archet II, Nice, France. Elastic fibers are essential extracellular matrix components of the skin, which contribute to its resilience and elasticity. During chronological aging, synthesis of elastic fibers is reduced, and the existing elastic fibers are degraded by elastases. Soybean extracts were found to protect the elastic network from degradation, and to restore elastic fiber synthesis. Using a luciferase reporter assay, soybean extracts were shown to induce elastin promoter activity in a dose-dependent manner. Soybean extracts inhibited the activity of human leukocyte elastase in vitro, and protected elastic fibers of fibroblast cultures from degradation by exogenous elastase. Human facial skins were treated with non-denatured soy extracts for 8 and 12 weeks, respectively, in two independent clinical trials. Monthly cutometer measurements demonstrated statistically significant increase in skin firmness as early as 4 weeks, and sustained to the end of treatment in each study. These data suggest the possible use of soybean extracts in skin care, to improve skin elasticity. Streamlining the Qualified Health Claim Process. A.S. Persad and R.A. Isbrucker, Burdock Group,Washington, DC, USA. Research on soy-based products has revealed a broad spectrum of potential health benefits, making qualified health claims (QHCs) a valuable tool to the soy industry. Although soy protein is generally recognized as safe (GRAS), this status is not a marker of its suggested efficacy. A QHC allows for the establishment of a relationship between a substance and its ability to reduce the risk of a disease or health-related condition. These claims, based on the available evidence and suffixed with tiered qualifiers issued by FDA, allow manufacturers to provide beneficial health information to their consumers without having to meet the stringent requirements of “Significant Scientific Agreement.” Notably, petitions for a QHC, such as the pending claim for soy protein and certain cancers, are not automatically granted; they must meet the ever-evolving FDA’s qualifying criteria. We propose a strategy to streamline QHC petitions that involve expert panel review, in which these experts determine if a threshold of credible evidence is met. This proposed system would parallel the current expert panel GRAS process and alleviate the potential bottleneck of submitted petitions to the Agency. The strategy requires: (1)reinforcing the credibility of the panel and supporting data (including meta-analyses) by the Agency; (2) preserving Poster Abstracts of liver and kidney, produce blood, help urinating, remove swelling, cure arthritics, high lipemia and cancer. In treating and preventing the abovelisted diseases, the following are discussed: the recipes of clinical foods, preparation, ways of applying, function and effects, indiction and points of attention. 6th International Symposium on the Poster Abstracts 32 Role of Soy in Preventing and Treating Chronic Disease proprietary data; and (3) public recognition of findings. The soy industry is challenged with navigating the regulatory requirements to establish efficacy of their products. Understanding these processes and developing this proposed streamlined approach would greatly aid both the soy industry and the FDA. Preparation of Extruded Snack Food from Green Gram Broken and Sawan Blends. Daya S. Singh, Aarti Patel, Krishna Tiwari, and S.K. Garg, Department of Post Harvest Process and Food Engineering, College of Agricultural Engineering, JNKVV, Jabalpur (M.P.) India. Studies on Antiradical Action of Herbal Extracts From Seasoning and Their Effect on Foods and Human Health. Boris Herskowitz*1, Rina Reznik2, Cathie Ioudkevitch2, Rodica Segal3, and Dana Moraru3, 1Soglowek Food Industries, Nahariya , Israel, 2Rad Natural Technologies Ltd., Petah Tikva, Israel, 3University “Dunarea de Jos “, Galati, Romania. Pulses are the inseparable part of India diet especially for vegetarian and are the major source of protein, when blended with cereals in certain proportion provides with a good protein diet among the consumer. Green gram (Vigna radita L.) is a major pulse of Madhya Pradesh which yields well under un-irrigated condition and on cloddy land. Sawan (Echinochloa frumentacea L.) is good source of starch and when blended with pulses, provides amino acids that are lacking in pulses. Therefore, the protein quality of cereals – pulses combination is thus betters than that of cereals/pulses alone. Extrusion cooking is a high temperature short time cooking process which could be used for processing of starchy as well as proteinaceous materials and is accomplished through the application of heat either directly by stream injection or indirectly through jacket and by dissipation of mechanical energy through shearing occurring within the blend. The results of cooking of ingredients during the extrusion process are the gelatinization of starch, the denaturation of protein, the destruction of naturally occurring toxic substance and the diminishing of microbial counts in the final product. The present study was undertaken to find the effects of extrusion cooking parameters and blending ratio on the quality of the extruded products and to study the overall acceptability through texture of the extrudates. The extrusion parameters were moisture content of the green gram broken and temperature of barrel. The extrusion cooking was done at Department of Food Science and Technology, JNKVV, Jabalpur. The extruder used was BTPL – Lab. Twin Screw Extruder with dia opening off 5.0 mm and the screw was driven with a 6 HP electric motor. Screw speed was kept constant at 100 rpm. Extrusion was carried out at three levels of moisture content (10%, 14% and 18%) of raw material and at 5 levels of percentage of green gram broken (10%, 13%, 16%, 19% and 22%) and at 3 levels of temperature (80, 90 and 100oC). The properties of the products obtained were evaluated and prediction models were developed. After analyzing all the attributes for physical parameters like Sectional Expansion Index, Longitudinal Expansion Index, Volumetric Expansion Index, Average diameter and bulk density; Texture Characteristics like Crispness, Hardness and Cutting strength, it was concluded that best quality of extrudates (products) were obtained at 14% moisture content, 80oC barrel temperature and 19% blending ratio followed by 10% moisture content, 80oC barrel temperature and 13% blending ratio. Meat Analog Schnitzels (MAS), containing Soy proteins, are subject to oxidative quality deterioration, expressing itself in Warmed-over Flavor (WOF), as result of exposure to high pre-frying and pre-cooking temperatures. Three stages were performed. In the first stage DPPH (1,1-Diphenyl-2-picryl hydrazyl) Free Radical Scavenging Activity was performed. In the second stage we made fried and cooked meat analog schnitzel (FCS) using Soy protein SUPRO 620 (P2). HS-GC analysis were performed in order to study the hexanal area of the product and to find the antioxidant power of the protein (P2-FCS= 477550) . In the third stage trying to improve the antioxidant capacity of P2 we have tested the anti-oxidative effect of adding a natural, herbal extract (OriganoxWS) (500ppm) RAD NATURAL to Soy proteins used in MAS (P2+WS-FCS=272865). The addition of 500 ppm of Origanox WS delayed oxidation of MAS by 43 %, as was measured by Hexanal method using Head Space GC, enhanced taste of food products eliminating WOF, prolongation of shelf life, giving to the final product a natural and healthy image. Change of Isoflavone Content During Manufacturing of Chunggukjang, a Traditional Korean Fermented Soyfood. J.S. Kim1, C.H. Jang1, J.K. Lim1, J.H. Kim2, C.S. Park3, and D.Y. Kwon4, 1Department of Animal Science & Biotechnology, Kyungpook National University, Daegu, S. Korea, 2Department of Food Science & Technology, Gyeonsang National University, JinJu, S. Korea, 3Department of Food Science & Technology, Kyunghee University, YoungIn, S. Korea, 4Korea Food Research Institute, SungNam, S. Korea. Chunggugjang is one of popular Korean traditional fermented soyfoods. It is manufactured by inoculating steamed soy with Bacillus subtillus or traditional method in which steamed soy is exposed to rice straw normally rich in Bacilllus species. Chunggukjang made from traditional method was found to have B. subtillus and B. licheniformis. We analyzed isoflavone composition of Chunggukjang collected at various fermentation times. Daidzin and genistin, major glycoside forms of isoflavones present in soy, were decreased by ~50% during 43 hrs. But the content of glycitein-O- glycoside showed a limited reduction. The contents of aglycones genistein and glycitein showed dramatic increase during first 30 hrs after fermentation started, with 5- and 1.7-folds increase in concentration, respectively. In conclusion, short-term fermentation of steamed soy with Bacillus species caused significant accumulation of aglycone forms of isoflavones, suggesting that Chunggukjang manufactured by traditional method might be a good source of readily absorbable isoflavones. This study was supported by Special Research and Development Grant from KOSEF (2005). Preparation of Nutritious Extruded Snacks from SoySorghum Blends to Solve the Problem of Malnutrition in Trabal Belt of India. Daya S. Singh and Duda Kalpana, Department of Post Harvest Process and Food Engineering Faculty Of Agricultural Engineering, J.,N. Agricultural University, Jabalpur-482004 (M.P.), India. Soybean (Glycine Max L) which belongs to the family leguminaceae, is considered as a cheapest source of high quality protein and oil content. In India, soybean is grown in 6.22 million hectares of land with total annual production of 5.86 million tonnes. Sorghum (sorghum bicolour L) belongs to the family poaceae, is most important cereal crop and cheapest source of nutrition for poor. Madhya Pradesh has distinguished it self as major soybean and sorghum growing state by covering 4.32 and 5.14 million hectare of land with 3.62 and 3.99 million tonnes production per annum (Anon. 2001-2002) respectively. Soybean 33 2005 Abstracts Skin Health Sunscreen Active Derived from Soybean Oil and Ferulic Acid: Synthesis and Applications. J. Laszlo1, D. Compton1, and R. Willis2, 1USDA, ARS, NCAUR, Peoria, IL, USA, 2iSoyTechnologies Corp., Cary, IL, USA. Ferulic acid is a natural plant phenol widely heralded for its ability to serve as a potent antioxidant. However, the low solubility of ferulic acid in lipids prevents it from protecting lipid membranes and associated structures. In its esterified forms, ferulic acid strongly absorbs light within the UVB and UVA regions from 290 to 370 nm, which suggests that it may serve as a natural sunscreen agent. This research describes the biocatalytic formation of stable (covalent) esters of ferulic acid with soybean oil. The UV absorbing soybean oil is under investigation for potential use as an all natural replacement for synthetic sunscreen active ingredients, as a natural UV protectant for insecticidal biocontrol agents, as an anti-oxidant in food frying, and as a bio-based industrial lubricant. Many potential human health applications have yet to be explored. We welcome inquiries regarding research collaborations. Soy Peptides Silicon Accumulation in Soybean Plants in Different Rhizosphere pH Conditions. L. Oliveira1,2, E. Oliveira1, G. Korndorfer1, and S. Tsai2, 1Universidade Federal de Uberlândia, Uberlândia, MG, Brazil, 2Centro de Energia Nuclear - USP, Piracicaba, SP, Brazil. The legumes, such as soybean, have the capacity of acidifying the rhizosphere soil, and this fact could explain the low ability of Si accumulation by this species, since Si absorption is dependent of soil pH. In acidic conditions Si forms polymers, which are little available for the plant. This work analyzed rhizosphere acidification with Si accumulation by soybean. The experiment was installed in the green house in a 5x2 factorial scheme, applying in the plots (vases) 200mg kg-1 N in the following proportions of N-NO3- and N-NH4+: 100% N-NO3-; 75% NNO3- + 25% N-NH4+; 50% N-NO3- + 50% N-NH4+; 25% N-NO3- + 75% N-NH4+; 100% N-NH4+; besides the N treatments there was a treatment with 200 mg kg-1 (silicon tetrachloride). The sources of nitrogen used were calcium nitrate and ammonium sulfate plus a nitrification inhibitor - nitrapirin, applied in a Ustoxic Quartzipsamment soil. The use of nitrate as N source caused an increase in rhizosphere and non-rhizosphere pH, leading to a greater Si availability in the soil. In the rhizosphere soil the pH values were lower than those found in non-rhizosphere soil. Silicon contents in the soybean above ground parts did not present any relationship with the availability of this element in the soil, however, in the root this relationship was positive, that is, the higher the pH, the greater the Si accumulation in the root. Silicon accumulation in soybean is proportionally greater in the root than in the aboveground parts. Glycine max, silicon, nitrate, ammonium. Enzymatic Production of Soybean Peptides with Potential Anti-Cancer Activity. E. de Mjia and W. Wang, University of Illinois, USA. Soybean proteins can be sources of bioactive peptides with potential anti-cancer activity. Cancer cells proliferate faster and have higher topoisomerases expression making this enzyme an ideal anti-cancer target. Thus, topoisomerase inhibition is an indicator of anticancer activity. The objective of this research was to determine the anti-topoisomerase potential of soy protein isolate (SPI) hydrolysates produced by commercially available proteases. Five proteases (S, P, N, A, and M, Amano Enzyme Inc.) were chosen for SPI hydrolysis. DNA human anti-Topo I and Topo II were carried out with TopoGen human drug screening assays. Identification and characterization of bioactive peptides were determined by SDS-PAGE, gel filtration, HPLC and mass spectrometry. The results showed that hydrolysates (0.14 mg/ml) produced by proteases P, S and N inhibited Topo II (38%, 16%, 21%, respectively). Subsequent in vitro digestion with pepsin and pancreatin, simulating gastrointestinal conditions, maintained 50% of the observed Topo II activity. At 2.45 mg/ml, all five SPI hydrolysates showed only 10% Topo I inhibition. Protease P hydrolysate contained the most active peptides with catalytic Topo II inhibition and broad MW (300-3000 Da) with hydrophilic and hydrophobic characteristics. Under in vitro gastric conditions and after 47% SPI hydrolysis as equivalents of L-leucine, small hydrophilic peptides (< 2000 Da) with Topo II inhibition activity (IC50=0.13 mg/ml) were produced. The results suggest anti-cancer potential of SPI hydrolysates formed by proteases that are available to the food industry. Furthermore, soybean derived peptides may play an important role, particularly in the prevention of chronic diseases, such as cancer. Poster Abstracts is rich in lysine but deficient in sulfer bearing amino acids. Sorghum, which contains adequate quantity of sulfer bearing amino acids is deficient in lysine (Bonyasirikool et al. 1986). Hence the combination of these sources of protein and starch can be effectively combined into snacks of high nutritive value, which may be regularly consumed during tea time/breakfast, will certainly solve the mal nutrition problem not only in India but in the entire Asiatic Countries. Protein in just 250 grams of soybean is equivalent to protein in 3 liters of milk or 1 kg of mutton or 24 eggs. The quality of soy protein is virtually equivalent in quality to that of milk and egg protein. In addition to containing rich nutrients, soy bean has other beneficial compounds such as, phytosterols, lecithins, etc. soy protein has a number of health benefits such as, cancer prevention, cholesterol reduction, combating asteoporosis and menopause regulation. Human studies suggest that as littel as one serving of soy foods each day may be protective against many types of cancers. The health claim states, “consumption of 25 grams of soy protein per day with a diet low in saturated fat may lower the risk of heart diseases”. Extrusion has become an important processing technique in an increasing variety of food processes. The result of cooking of ingredients during the extrusion process are the gelatinization of starch, the denaturation of protein, the destruction of naturally occurring toxic substances and diminishing of microbial counts in the final product. Once cooked the product is forced through the die at the extruder discharge end where it expands rapidly with some loss in moisture. Experiments were conducted to study the effect of processing parameter i.e. barrel temperature (80, 85, 90, 95 and 100oC), moisture content of blend (15, 20 and 25% wb) and blending ratio of soy in sorghum (5:95, 10:90, 15:85, 20:80 and 25:75), on the physical properties like, sectional expansion index, longitudinal expansion index, volumetric expansion index, bulk density; textural properties like crispness, hardness and cutting strength; protein content and organoleptic test of the extruded snacks. After evaluation and analysis of all the parameters for physical, textural and organoleptic characteristics, it was concluded that snacks produced at 15% moisture content, 85oC barrel temperature and 5% blend ratio was found best followed by product produced at 15% moisture content, 90oC barrel temperature and 5% blend ratio. 6th International Symposium on the Poster Abstracts 34 Role of Soy in Preventing and Treating Chronic Disease \Are Undigested Soy Peptides the Major Hypocholesterolemic Components of Soy Protein? J.W. Anderson and K. Patterson, University of Kentucky, Lexington, KY, USA. Soy proteins have many bioactive components including protein subunits, peptides and isoflavones. Emerging research in animals and humans indicate that peptides containing 5-20 amino acids (AA) are the most potent hypocholesterolemic components of soy protein. Isolated soy protein (ISP) hydrolysates (containing peptides of 5-20 AA) have substantially greater hypocholesterolemic effects than ISP in animals and humans. Possible mechanisms include decreased cholesterol absorption and LDL receptor upregulation. Feeding the α' subunit of the soy 7S globulin to rats has about five-fold greater hypocholesterolemic potency than the 7S globulin itself. An α' subunit peptide of ~17 AA upregulates LDL receptors in vitro. Oral administration of 10 mg/kg body weight of 7S globulin peptide produced significant hypocholesterolemia in rats. Large soy peptides (43 AA) are absorbed intact and retain biologic activity when extracted from blood or liver. Animal consumption of soy peptides (3-20 AA) exerts distinct effects on lipid metabolism, blood pressure, energy expenditure and appetite regulation. In humans, Wang et al. (1995) reported that soy protein hydrolysates decreased serum LDL-cholesterol by 24% more than ISP. Hori et al. (2001) reported that 3 g/d of soy peptides decreased serum LDL-cholesterol from baseline by 27% and 6 g/d decreased values by 43%. Thus, small amounts of soy peptides exert dramatic hypocholesterolemic effects in humans. Thus, the recent decline in hypocholesterolemic effects of ISP reported in humans may relate to fragmentation of bioactive peptides during soy protein isolation or food processing. As recent studies indicate, providing 3-6 grams of bioactive peptides may have dramatic hypolipidemic effects.
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