Soy Isoflavone Metabolites Research Articles

Clinical effect of equol supplementation in the treatment of desquamative gingivitis with 1-year follow-up.

Desquamative gingivitis (DG) is characterized by desquamative erosion, edematous erythema, and vesicle formation on the gingiva. Because of its prevalence in women during the pre- and postmenopausal period, its potential association with female hormones has been suggested. Equol is a soy isoflavone metabolite with a chemical structure similar to estrogen. Scientific evidence suggests that equol helps in alleviating menopausal symptoms. This study evaluated the clinical effect of a 12-month equol supplementation as a substitute for estrogen to alleviate DG symptoms. The study enrolled 16 women with DG who regularly visited Nihon University School of Dentistry Dental Hospital. Urinary equol levels, periodontal tissue examination, O'Leary's plaque control record, stimulated saliva flow rate, and gingival pain-related questionnaires were evaluated before and after the 12-month daily intake of 10 mg equol supplement. Equol supplementation led to a statistically significant improvement in bleeding on probing, visual findings, and reductions in the frequency and severity of gingival pain. Urinary equol testing and equol supplementation may be novel treatment options for female patients with DG.

Advances in the Metabolic Mechanism and Functional Characteristics of Equol.

Equol is the most potent soy isoflavone metabolite and is produced by specific intestinal microorganisms of mammals. It has promising application possibilities for preventing chronic diseases such as cardiovascular disease, breast cancer, and prostate cancer due to its high antioxidant activity and hormone-like activity. Thus, it is of great significance to systematically study the efficient preparation method of equol and its functional activity. This paper elaborates on the metabolic mechanism of equol in humans; focuses on the biological characteristics, synthesis methods, and the currently isolated equol-producing bacteria; and looks forward to its future development and application direction, aiming to provide guidance for the application and promotion of equol in the field of food and health products.

Soy isoflavone metabolite equol inhibits cancer cell proliferation in a PAP associated domain containing 5-dependent and an estrogen receptor-independent manner

Isoflavone is a species of polyphenol found mainly in soy and soy products. Many studies have demonstrated its estrogen receptor (ER)-dependent action. Equol is an intestinal metabolite of a major soy isoflavone daidzein. We aimed to elucidate the mechanism for ER-independent actions of equol. Equol has been shown to inhibit proliferation of HeLa human cervical cancer cells and mouse melanoma B16 cells in an ER-independent manner. Using functional genetic screening, PAP associated domain containing 5 (PAPD5), which is a non-canonical poly(A) polymerase, was identified as an essential molecule in the ER-independent action. While peroral administration of equol inhibited tumor growth of control B16 cells subcutaneously inoculated in mice, it had little effect on the growth of PAPD5-ablated B16 cells. Intriguingly, equol progressed tumor growth of the PAPD5-ablated human breast cancer MCF-7 cells, which have high ERα expression. Equol has been found to induce polyadenylation of snoRNAs in a PAPD5-depdendent manner. Furthermore, peroral equol administration increased microRNA miR-320a expression in tumors. Together, these results suggest that equol may have a dual effect on ER-positive cancer cells, acting with, antiproliferative activity through PAPD5 and exhibiting proliferative activity via ERα and the former could be associated with miR-320a.

Effects of Soy Isoflavones, Resistant Starch and Antibiotics on Polycystic Ovary Syndrome (PCOS)-Like Features in Letrozole-Treated Rats.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in reproductive-aged women. Recently, various dietary interventions have been used extensively as a novel therapy against PCOS. In the present study, we show that soy isoflavone metabolites and resistant starch, together with gut microbiota modulations, were successful in decreasing the severity of PCOS-like reproductive features while increasing the expression of gut barrier markers and butyric acid in the gut. In the letrozole-induced PCOS model rats, the intake of both 0.05% soy isoflavones and 11% resistant starch, even with letrozole treatment, reduced the severity of menstrual irregularity and polycystic ovaries with a high concentration of soy isoflavones and equol in plasma. Antibiotic cocktail treatment suppressed soy isoflavone metabolism in the gut and showed no considerable effects on reducing the PCOS-like symptoms. The mRNA expression level of occludin significantly increased with soy isoflavone and resistant starch combined treatment. Bacterial genera such as Blautia, Dorea and Clostridium were positively correlated with menstrual irregularity under resistant starch intake. Moreover, the concentration of butyric acid was elevated by resistant starch intake. In conclusion, we propose that both dietary interventions and gut microbiota modulations could be effectively used in reducing the severity of PCOS reproductive features.

Potential Protective Effects of Equol (Soy Isoflavone Metabolite) on Coronary Heart Diseases-From Molecular Mechanisms to Studies in Humans.

Equol, a soy isoflavone-derived metabolite of the gut microbiome, may be the key cardioprotective component of soy isoflavones. Systematic reviews have reported that soy isoflavones have no to very small effects on traditional cardiovascular disease risk factors. However, the potential mechanistic mode of action of equol on non-traditional cardiovascular risk factors has not been systematically reviewed. We searched the PubMed through to July 2021 by using terms for equol and each of the following markers: inflammation, oxidation, endothelial function, vasodilation, atherosclerosis, arterial stiffness, and coronary heart disease. Of the 231 records identified, 69 articles met the inclusion criteria and were summarized. Our review suggests that equol is more lipophilic, bioavailable, and generally more potent compared to soy isoflavones. Cell culture, animal, and human studies show that equol possesses antioxidative, anti-inflammatory, and vasodilatory properties and improves arterial stiffness and atherosclerosis. Many of these actions are mediated through the estrogen receptor β. Overall, equol may have a greater cardioprotective benefit than soy isoflavones. Clinical studies of equol are warranted because equol is available as a dietary supplement.

Ischemic-time associated reductions in equol monosulfate plasma levels in a mouse model of ischemic stroke: support the existence of a 'brain-gut axis'.

Interest is growing in the role played by intestinal flora in the pathogeneses of diseases and in the possibility of treating disease by altering intestinal flora compositions. Recent studies have focused on the relationship between the intestinal microbiome and brain function as proposed by the brain-gut axis hypothesis. To investigate the relation between ischemic stroke and plasma equol monosulfate levels (a soy isoflavone metabolite) in a middle cerebral artery occlusion (MCAO) mouse model. Mice (C57BL/6) were subjected to MCAO for various times (30 min to 24 h), and degrees of cerebral damage were assessed using total infarction volumes, brain edema severities and neurological deficit scores. Hematoxylin and eosin and cresyl violet staining were used to observe morphological changes in ischemic brains. Levels of equol monosulfate in plasma and the relationships between these and degree of brain injury were investigated. Infarction volumes, brain edema severity and neurological deficit scores were significantly correlated with ischemic time, and morphological deteriorations of brain neuronal cells also increased with ischemic duration. Equol monosulfate contents were ischemic-time dependently lower in MCAO treated animals than in sham-operated controls. Ischemic stroke may time-dependently reduce plasma levels of equol monosulfate by lowering the metabolic rate of equol in MCAO-induced mice. This study provides indirect support of the brain-gut axis hypothesis.

A Systematic Review of the Effects of Equol (Soy Metabolite) on Breast Cancer.

Equol is a soy isoflavone metabolite that can be produced by intestinal bacteria. It is lipophilic and resembles natural oestrogens with an affinity to oestrogen receptors. This review is focused on how equol affects breast cancer, as evidenced by in vivo and in vitro studies. Equol is considered chemoprotective in specific endocrine-related pathologies, such as breast cancer, prostate cancer, cardiovascular diseases, and menopausal symptoms. In humans, not everyone can produce equol from gut metabolism. It is postulated that equol producers benefit more than non-equol producers for all the endocrine-related effects. Equol exists in two enantiomers of R-equol and S-equol. Earlier studies, however, did not specify which enantiomer was being used. This review considers equol’s type and concentration variations, pathways affected, and its outcome in in vivo and in vitro studies.

Formononetin - An isoflavone metabolite found in the liver of rats fed with soy protein isolate

Soy protein isolate, containing genistein, daidzein and glycitein, was used to study the profiles of soy isoflavone metabolites in Sprague-Dawley rats. After 50 days, rat liver tissues, serum and urine were analyzed by HPLC-MS and HPLC-ECD. HPLC-APCI-MS results showed that daidzein, equol, genistein and O-DMA were detected. Interestingly, the aglycone and the conjugated forms of formononetin, which is a methylated derivative of daidzein found in many plants, were also observed in rat liver and serum, but not in urine. The respective concentrations of these two forms of formononetin were 0.056 ± 0.022 and 0.621 ± 0.116 nmoL/g (mean ± SD) in liver tissue, and were 0.005 ± 0.001 and 0.035 ± 0.002 nmoL/mL in rat serum. Formononetin in conjugated forms were 90.8% in liver tissue and 85.7% in serum by calculation. Incubation of daidzein with liver homogenate, microsomes, and catechol 4′-O-methyl transferase (COMT) resulted in a 7∼ 10% conversion of daidzein to the aglycone form of formononetin. Formononetin is proposed to be a metabolite of O-methylation of daidzein in rat liver and exists mainly in conjugated forms.

Detection of conjugated soy metabolites in urinary and tissue samples after methanol extraction

Glucuronide conjugates of soy isoflavone metabolites genistein, daidzein, glycitein and equol in urinary samples from women, female pigs and female rats were pretreated with methanol instead of solid phase extraction or liquid-liquid extraction after enzymatic digestion. The samples were analyzed using an LC-ESI-MS/MS ion trap system. Analysis of rat liver sample using this procedure was also attempted. By comparison the elution sequences with results from previous studies, glucuronide conjugates including 7-gluc-Dai, 7-gluc-Gly, 4′-gluc-Dai, 7-gluc-Gen, then 4′-gluc-Gen were identified sequentially in women urine. However, 5-gluc-Gen was also observed in female porcine urine. In rat urine, sul-Dai was observed after 7-gluc-Dai, while 7-gluc-Equ and 4′-gluc-Equ were observed after 7-gluc-Gen. The metabolite profiles in rat liver tissue samples were found with 7-gluc-Dai and 7-gluc-Gen. However, glucuronide conjugates of equol and sulfate conjugate of daidzein were only found in rat urine. Interestingly, quantification studies performed by LC-APCI/MS and LC-ECD showed that the aglycone and conjugated glycitein were both found in porcine and rat urine, but only conjugated glycitein was found in woman urine.

Nutrikinetic study of fermented soybean paste (Cheonggukjang) isoflavones according to the Sasang typology.

BACKGROUND/OBJECTIVESIn Oriental medicine, certain foods may be beneficial or detrimental based on an individual's constitution; however, the scientific basis for this theory is insufficient. The purpose of this study was to investigate the effect of body constitution, based on the Sasang type of Korean traditional medical classification system, on the bioavailability of soy isoflavones of Cheonggukjang, a quick-fermented soybean paste.SUBJECTS/METHODSA pilot study was conducted on 48 healthy Korean men to evaluate the bioavailability of isoflavone after ingestion of food based on constitution types classified by the Sasang typology. The participants were classified into the Taeeumin (TE; n = 15), Soyangin (SY; n = 15), and Soeumin (SE; n = 18) groups. Each participant ingested 50 g of Cheonggukjang per 60 kg body weight. Thereafter, blood was collected, and the soy isoflavone metabolites were analyzed by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry. Ntrikinetic analysis of individual isoflavone-derived metabolites was performed.RESULTSOur nutrikinetic analysis identified 21 metabolites derived from isoflavones in the blood samples from 48 healthy Korean men (age range, 21-29 years). Significant differences were observed in the time to maximum concentration (Tmax) and elimination half-life (t1/2) for nine metabolites among the three groups. The Tmax and t1/2 of the nine metabolites were higher in the SE group than in the other groups. Moreover, the absorption rates, as determined by the area under the plasma-level curve (AUC) values of intact isoflavone, were 5.3 and 9.4 times higher in the TE group than in the SY and SE groups, respectively. Additionally, the highest AUC values for phase I and II metabolites were observed in the TE group.CONCLUSIONSThese findings indicate that isoflavone bioavailability, following Cheonggukjang insgestion, is high in individuals with the TE constitution, and relatively lower in those with the SE and SY constitutions.

S-equol inhibits proliferation and promotes apoptosis of human breast cancer MCF-7 cells via regulating miR-10a-5p and PI3K/AKT pathway

S-equol is the exclusive enantiomeric form of the soy isoflavone metabolite produced by human intestinal bacterial flora, which has strong anti-cancer activity. Based on this, the purpose of this study was to investigate the anti-breast cancer mechanism of S-equol. We examined the effects of S-equol on proliferation and apoptosis of MCF-7 cells by cell counting kit-8 assay and flow cytometry. Screening for microRNAs and predicting their target genes using the starBase and Targetscan website, respectively. Protein expression was detected by Western blot. The microRNA level were quantified by real-time PCR. The results showed that S-equol inhibited the proliferation of breast cancer MCF-7 cells in a time- and dose-dependent manner and promoted apoptosis of MCF-7 cells. The expression of miR-10a-5p was significantly decreased in breast cancer tissues and breast cancer cell lines, and the expression of miR-10a-5p was negatively correlated with the proliferation of MCF-7 cells. Luciferase reporter experiments demonstrated that miR-10a-5p directly targets PIK3CA 3′UTR to function. It was further found that S-equol exerts an anti-breast cancer effect by up-regulating miR-10a-5p and inhibiting the PI3K/AKT pathway. Our study revealed the mechanism of S-equol against breast cancer, and miR-10a-5p may be a potential target for the treatment of breast cancer.

Effects of Lactic Acid Bacteria-Fermented Soymilk on Isoflavone Metabolites and Short-Chain Fatty Acids Excretion and Their Modulating Effects on Gut Microbiota.

Lactobacillus rhamnosus strain ASCC 1520 with high soy isoflavone transformation ability was used to ferment soymilk and added to the diet of mice. The impact of L. rhamnosus fermentation on soy isoflavone metabolites and intestinal bacterial community, in conjunction with fecal enzyme activity and short-chain fatty acids (SCFA) excretion was evaluated. Antibiotics intervention resulted in a decrease in fecal enzyme activities and SCFA. Although long-term intake of soymilk or L. rhamnosus-fermented soymilk did not affect the fecal β-glucuronidase and β-galactosidase activities, it improved the β-glucosidase activity when antibiotics were concomitantly administered. Soymilk or fermented soymilk administration increased the isoflavone metabolites (O-DMA and equol) excreted in urine. Antibiotics decreased the daidzein excretion and its metabolites but showed little effect on glycitein and genistein excretion. Principal coordinates analysis (PCoA) of the 16s rRNA gene sequencing data found a remarkable shift in gut microbiota after soymilk administration and antibiotics treatment. Matastats test of the relative abundance of bacterial taxa revealed Odoribacter (Bacteroidales family), Lactobacillus (Lactobacillales order), and Alistipes (Rikenellaceae family) were enriched in soymilk while bacterial taxa from Bacteroides and Lactobacillus were enriched in L. rhamnosus-fermented soymilk. Furthermore, there was less decrease in bacterial taxa with fermented soymilk group even when antibiotics were concomitantly administered. Overall, this study revealed that the gut microbiota of a healthy host is enough for the whole isoflavone metabolism under normal conditions. Feeding mice with L. rhamnosus-fermented soymilk improved fecal enzyme activity and kept the balance of the gut mirobiota when antibiotics were used. PRACTICAL APPLICATION: Feeding mice with L. rhamnosus-fermented soymilk improved fecal enzyme activity and kept the balance of the gut mirobiota when antibiotics were used.

Reduction of soy isoflavones by use of Escherichia coli whole-cell biocatalyst expressing isoflavone reductase under aerobic conditions

Soy isoflavone metabolites are currently receiving much attention due to the stronger and wider bioactivities than that of isoflavones. Therefore, biosynthesis of isoflavone metabolites by isolated isoflavone biotransforming bacteria is important. However, the biosynthesis process must be under obligate anaerobic conditions due to the reduction reactions catalysed by isoflavone biotransforming bacteria. In this study, we cloned the daidzein and genistein reductase gene (dgr) from Slackia sp. AUH-JLC159. The recombinant Escherichia coli (E.coli) whole-cell was used for the first time as the biocatalyst for aerobic biosynthesis of dihydrodaidzein (DHD) and dihydrogenistein (DHG) from soy isoflavones daidzein and genistein. Our results indicated that the recombinant E.coli whole-cell was able to reduce daidzein and genistein to DHD and DHG under aerobic conditions, while the maximal concentration of the substrate daidzein or genistein that the E.coli whole-cell was able to convert efficiently was only 0·4mmoll(-1) . Under the optimized conditions, the maximal concentration of daidzein or genistein that the E.coli whole-cell was able to convert efficiently was increased to 1·4mmoll(-1) . Our results demonstrated that E.coli whole-cell is an efficient biocatalyst for biosynthesis of isoflavone metabolites under aerobic conditions. Soy isoflavone metabolites, which are more biologically active than their precursor isoflavones, are currently receiving much more attention. However, the non-natural isoflavone metabolites are synthesized or biosynthesized under obligate anaerobic conditions. Here, we describe a new approach to the reduction of soy isoflavones daidzein and genistein under aerobic conditions by use of the recombinant Escherichia coli whole-cell expressing isoflavone reductase. Our study provides the first evidence that isoflavone metabolites, such as dihydrodaidzein and dihydrogenistein, are able to be produced efficiently under aerobic conditions.

Fecal bacterial community changes associated with isoflavone metabolites in postmenopausal women after soy bar consumption.

Soy isoflavones and their metabolism by intestinal microbiota have gained attention because of potential health benefits, such as the alleviation of estrogen/hormone-related conditions in postmenopausal women, associated with some of these compounds. However, overall changes in gut bacterial community structure and composition in response to addition of soy isoflavones to diets and their association with excreted isoflavone metabolites in postmenopausal women has not been studied. The aim of this study was to determine fecal bacterial community changes in 17 postmenopausal women after a week of diet supplementation with soy bars containing isoflavones, and to determine correlations between microbial community changes and excreted isoflavone metabolites. Using DGGE profiles of PCR amplified 16S rRNA genes (V3 region) to compare microbial communities in fecal samples collected one week before and one week during soy supplementation revealed significant differences (ANOSIM p<0.03) before and after soy supplementation in all subjects. However, between subjects comparisons showed high inter-individual variation that resulted in clustering of profiles by subjects. Urinary excretion of isoflavone (daidzein) metabolites indicated four subjects were equol producers and all subjects produced O-desmethylangolensin (ODMA). Comparison of relative proportions of 16S rRNA genes from 454 pyrosequencing of the last fecal samples of each treatment session revealed significant increases in average proportions of Bifidobacterium after soy consumption, and Bifidobacterium and Eubacterium were significantly greater in equol vs non-S-(-)equol producers. This is the first in vivo study using pyrosequencing to characterize significant differences in fecal community structure and composition in postmenopausal women after a week of soy diet-supplementation, and relate these changes to differences in soy isoflavones and isoflavone metabolites.Trial RegistrationClinicaltrials.gov NCT00244907

Pharmacokinetics of Equol, a Soy Isoflavone Metabolite, Changes with the Form of Equol (Dietary versus Intestinal Production) in Ovariectomized Rats

Recent findings indicate that soy isoflavones and their metabolites may play a role in mitigating postmenopausal bone loss. Equol, a metabolite of the soy isoflavone daidzein produced by intestinal bacteria, has shown some potential, but only 30–50% of the U.S. population is capable of converting dietary daidzein to equol. There are limited data on the pharmacokinetics of dietary racemic equol and its metabolites. This study was conducted to assess the levels of equol and its conjugates in plasma for a 24 h period resulting from oral administration of dietary daidzein and racemic equol in ovariectomized Sprague–Dawley rats. Plasma samples were analyzed for conjugated and free forms of equol using LC-MS/MS. The maximum plasma concentration (Cmax) and time to reach it (tmax) for total equol (conjugated and unconjugated) were 8815 ± 2988 nmol/L and 2.17 ± 2.91 h and 3682 ± 2675 nmol/L and 20.67 ± 4.67 h, for dietary equol and daidzein, respectively. Although the majority of equol metabolites present were glucuronide conjugates (≥99%), there were low levels of equol monosulfate present. The changes in equol metabolism, specifically equol conjugates, due to the form of equol may play a role in the potential health benefits of equol.

Abstract 162: Reduced pro-inflammatory cytokines and immunosuppressive cells in patients with prostate cancer following consumption of soy isoflavone enriched bread.

Abstract Soybeans are a rich source of bioactive phytochemcials. Among the phytochemicals, the isoflavones (primarily genistein and daidzein) have received the greatest attention regarding cancer prevention. Indeed, we and others have reported dietary soy has measureable benefit in experimental tumor models of prostate cancer (PCa). Thus, we and others are actively investigating the potential of diets rich in soy components to prevent or slow human PCa progression, particularly in the early hormone sensitive state. Isoflavones are hypothesized to act through various pathways to impact PCa progression, such as inhibition of tumor growth factor signaling, anti-angiogenesis, and cell cycle inhibition. However, one area that has received very little focus is the impact of soy components on the immune system. We hypothesized that dietary soy may benefit prostate cancer (PCa) patients by modulating the balance of pro-inflammatory cytokines and immunosuppressive cells. A randomized, phase II trial was conducted in 32 PCa patients with asymptomatic biochemical failure (rising prostate specific antigen/PSA). Patients were randomized to 3 slices of soy bread (33 mg isoflavones/slice) or soy bread containing almond extract daily as a source of β-glucosidase. Flow cytometry was used to study cellular phenotype and bioplex assay to measure cytokines in peripheral blood on days 0 (baseline) and 56. Soy isoflavone metabolites in patient urine were assessed by LC-MS/MS. Adequate blood samples were available from 25 of 32 men completing the &amp;gt;56 day protocol and evaluated. Phenotypic analysis of Day 0 and 56 blood revealed no change in percentage of CD8+ T cells, but showed increased CD56+ NK cells (p=0.038) and reduced CD4+ T cells (p=0.04). Cells with a natural T regulatory cell phenotype (CD4+CD25+FoxP3+) were significantly decreased after 56 days consuming either type of soy bread (p=0.035). A significant decrease in cells with a monocytic (CD33+HLADRlowCD14+) and granulocytic (CD33+HLADR-CD11b+) MDSC phenotype was observed in patients consuming both types of soy bread (p=0.0064 and p=0.049). Plasma cytokine and chemokine levels were significantly decreased on Day 56 compared to baseline. Subgroup analysis indicated reduction in both Th1 type cytokines (p=0.0349) and MDSC associated cytokines (p=0.0284). Th2 and Th17 cytokines were not significantly altered as compared to baseline. Exploratory analyses using a K-means clustering approach revealed patients who metabolized soy isoflavones into dihyrdodaidzein and equol had significantly lower MDSC after soy intervention (p's&amp;lt;0.02). Finally, patients with prolonged or no change in PSA doubling time had fewer T regulatory cells or MDSC on day 56 versus baseline. Our data suggests that soy bread modulates immune suppressor cell biology in humans and may enhance the efficacy of anti-cancer immune responses. Citation Format: Gregory Brian Lesinski, Thomas A. Mace, Patrick Reville, Jennifer Ahn-Jarvis, Matthew Bill, Courtney Nicholas, Yael Vodovotz, Elizabeth Grainger, Kenneth Riedl, Gregory Young, Steven Schwartz, Steven Clinton. Reduced pro-inflammatory cytokines and immunosuppressive cells in patients with prostate cancer following consumption of soy isoflavone enriched bread. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 162. doi:10.1158/1538-7445.AM2013-162

Abstract LB-188: Isoflavone pharmacokinetics and metabolism after consumption of soy and soy-almond bread in men with asymptomatic prostate cancer.

Abstract Background: Soy phytochemicals, particularly isoflavones, are hypothesized to have anti-prostate cancer activity. Soy isoflavones are present predominantly as glucosides but when combined with β-glucosidase activity in almond meal, a greater conversion to aglycones has been observed. We hypothesized that soy bread will serve as an excellent source of soy polyphenols and that an aglycone-rich soy-almond bread (SAB) compared to a glycoside-rich soy bread (SB) will demonstrate greater bioavailability of soy isoflavones. Methods: SB and SAB were fed in a 20 week randomized phase II cross-over trial involving 32 men with asymptomatic metastatic prostate cancer and escalating prostate specific antigen (PSA). Isoflavones were quantitated by HPLC in soy breads and isoflavone metabolites in 24h urine. HPLC-tandem mass spectrometry evaluated isoflavone pharmacokinetics in plasma. Biomarkers were determined by chemiluminescence-based assays. Results: The aglycone composition in the SB and SAB was 18% and 78%, respectively. Compliance was 95±14%, toxicity rare and limited to grade I GI complaints (NIH_NCI criteria). Urinary isoflavone patterns over 24hours revealed that isoflavonoid metabolites were significantly greater with soy bread consumption. Plasma pharmacokinetics following a test meal showed that daidzein (p=0.028) and genistein (p=0.010) absorption was significantly faster from the aglycone-rich SAB. Interestingly, PSA doubling time was prolonged compared to pre-enrollment, in 60% of men (n=30) with soy intervention. Conclusions: Bread is an excellent vehicle for the delivery of soy phytochemicals with outstanding compliance. Modification of soy bread with a source of β-glucosidase may enhance the initial absorption rates of isoflavones. Food products, such as SAB, with consistent and stable composition of phytochemicals can be developed that are appropriate for long-term chemoprevention studies in humans or as an adjunct to therapy. Citation Format: Jennifer H. Ahn-Jarvis, Steven K. Clinton, Elizabeth M. Grainger, Kenneth M. Riedl, Steven J. Schwartz, Yael Vodovotz. Isoflavone pharmacokinetics and metabolism after consumption of soy and soy-almond bread in men with asymptomatic prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-188. doi:10.1158/1538-7445.AM2013-LB-188

The Covalent Binding of Genistein to the Non-prosthetic-heme-moiety of Bovine Lactoperoxidase Leads to Enzymatic Inactivation

Objective Genistein, a major soy isoflavone metabolite (SIF), inactivates oxidation activity of bovine lactoperoxidase (LPO). Modification of the heme moiety of LPO by nitrogen-containing compounds has been shown to inactivate LPO. In contrast, SIF mediated inactivation of LPO does not involve a heme modification and the mechanism of SIF inhibition is poorly understood. Methods After inactivation of LPO by genistein in the presence of H 2O 2, trypsin-digested LPO peptide fragments were collected and analyzed by MALDI-TOF-MS to characterize the chemical binding of genistein(s) to LPO. Results The heme moiety of LPO was not modified by genistein. A covalent binding study showed that 3H-genistein bound to LPO with a ratio of ∼12 to 1. After HPLC analysis and peak collection, trypsin-digested peptide fragments were analyzed by MALDI-TOF-MS. The 3H-genistein co-eluted peptide fragments (RT=24 min) were putatively identified as 199IVGYLDEEGVLDQNR214 with two bound genistein molecules or a genistein dimer (2 259 Da), 486TPDNIDIWIGGNAEPMVER504 with two bound genistein molecules or a genistein dimer (2 663 Da), and 161ARWLPAEYEDGLALPFGWTQR182 with three bound genistein molecules or a genistein trimer (3 060 Da). The fragment with a mass of 1 792 Da (RT=36 min) was identified as 132CDENSPYR139 with three genistein molecules or a genistein trimer. Conclusions The results suggest that LPO was inactivated by irreversible covalent binding of genistein or genistein polymers to particular peptide fragments constituting regions of the outward domain. No genistein interaction with the prosthetic heme moiety of LPO was observed.

Impact of perinatal exposure to equol enantiomers on reproductive development in rodents

There is now considerable interest in the intestinally derived soy isoflavone metabolite, equol, which occurs in the enantiomeric forms, S-(−)equol and R-(+)equol, both differing in biological actions. Little is known about effects of either enantiomer on reproductive development, yet such knowledge is fundamental because of the recent commercialization of S-(−)equol as a dietary supplement. S-(−)equol and R-(+)equol were therefore investigated to determine their effects on reproductive development and fertility in the Sprague–Dawley rat. Neither enantiomer affected fertility, number of litters produced, number of pups per litter, number of male and female pups born, birth weight, anogenital distance, testicular descent or vaginal opening. Histological analysis showed no major abnormalities in ovary, testis, prostate or seminal vesicle tissue with dietary exposure to S-(−)equol or R-(+)equol, but both enantiomers triggered hyperplasia of uterine tissue. With R-(+)equol this stimulatory effect subsided after exposure was discontinued, but the effect of S-(−)equol was prolonged.

Mammary gland differentiation by early life exposure to enantiomers of the soy isoflavone metabolite equol

The role of soy in reducing breast cancer risk has been suggested to be associated with early exposure to isoflavones, which alter mammary gland morphology. The objective of the study was to determine the effect of dietary exposure to the enantiomers of a key soy isoflavone metabolite, equol, on mammary gland development and later chemoprotection using the DMBA-induced animal model of breast cancer. Animals were exposed to S-(−)equol or R-(+)equol (250 mg/kg diet) during the neonatal (0–21 days) or prepubertal (21–35 days) periods only. Histological evaluation of the mammary glands showed that both enantiomers fed neonatally via the dam led to significant precocial mammary gland differentiation. By day 50, early S-(−)equol or R-(+)equol exposure resulted in a decrease in immature terminal end structures and an increase in mature lobules, suggesting an early ‘imprinting’ effect. Despite these morphological changes to the mammary gland, neonatal and prepubertal exposure to equol had no long-term chemoprevention against mammary tumors induced by DMBA, although for R-(+)equol there was a trend to delaying tumor formation. In summary, early exposure to equol was not chemopreventive, but neither did it increase tumor formation in response to DMBA, suggesting exposure in early life does not influence breast cancer risk.