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To explore the importance of equol on health outcomes in future studies, it was necessary to develop a method to reduce equol production. Female monkeys (n = 22) fed a soy diet were treated twice daily with vehicle (control; n = 4), doxycycline (2.5 mg/kg; n = 4), metronidazole (125 mg/d; n = 3), kanamycin (1000 mg/d; n = 4), vancomycin (100 mg/d; n = 3) or kanamycin+vancomycin (n = 4). Plasma samples were collected 4 h postfeeding at baseline, after 4 wk of treatment and 8 wk after the end of treatment and analyzed for isoflavonoid concentrations. Fecal swabs were collected at baseline and at the end of antibiotic treatment for analysis of Gram(+) and Gram(-) bacterial growth. Equol concentrations were reduced (P < 0.05) compared with baseline by 80, 93, 98 and 99% after treatment with metronidazole (955 [+ or -] 164 vs. 193 [+ or -] 53 nmol/L), kanamycin (545 [+ or -] 211 vs. 37.1 [+ or -] 17.6 nmol/L), vancomycin (607 [+ or -] 163 vs. 8.9 [+ or -] 8.2 nmol/L) and kanamycin+vancomycin (721 [+ or -] 169 vs. 17.4 [+ or -] 17.3 nmol/L), respectively. Daidzein concentrations were increased (P < 0.05) compared with baseline by treatment with doxycycline (336 [+ or -] 87 vs. 576 [+ or -] 76 nmol/L), kanamycin (168 [+ or -] 67 vs. 374 [+ or -] 15 nmol/L), and kanamycin+vancomycin (166 [+ or -] 35 vs. 384 [+ or -] 78 nmol/L). Similar increases (P < 0.05) in dihydrodaidzein were observed after treatment with kanamycin (31.2 [+ or -] 6.2 vs. 479 [+ or -] 188 nmol/L) and metronidazole (56.0 [+ or -] 27.9 vs. 414 [+ or -] 212 nmol/L). Isoflavonoid concentrations returned to baseline values after antibiotic treatment was terminated. Gram(+) bacterial growth was reduced by all treatments, including Control, compared with baseline. In conclusion, treatment with antibiotics resulted in a marked reduction in plasma equol concentrations and altered plasma isoflavonoid patterns in cynomolgus monkeys. KEY WORDS: * cynomolgus monkeys * soy * isoflavonoids * equol, antibiotics

The pharmacokinetics of isoflavones in 10 healthy women were determined from serum appearance/ disappearance concentration profiles and urinary excretions after single-bolus ingestion of 10, 20 or 40 g of soy nuts delivering increasing amounts of the conjugated forms of daidzein (6.6, 13.2 and 26.4 mg) and genistein (9.8, 19.6 and 39.2 mg). Peak serum daidzein and genistein concentrations were attained after 4-8 h, and elimination half-lives were 8.0 and 10.1 h, respectively. There were no differences in the pharmacokinetics of daidzein and genistein between pre- and postmenopausal women, indicating absorption and disposition of isoflavones to be independent of age or menopausal status. A curvilinear relationship was observed between the bioavailability of daidzein and genistein, apparent from the area under the curve to infinity (AU[C.sub.inf]) of the serum concentration-time profiles and the amount of isoflavones ingested. The mean fraction of the isoflavones excreted in urine decreased with increasing intake when expressed as a percentage of the administered dose (63.2 [+ or -] 8.0, 54.4 [+ or -] 8.1 and 44.0 [+ or -] 4.3%, respectively, for daidzein, and correspondingly, 25.2 [+ or -] 5.3, 13.4 [+ or -] 2.1 and 15.8 [+ or -] 2.7% for genistein), underscoring the trend toward nonlinear pharmacokinetics. Equol was identified as a metabolite in 30% of women; it was present consistently in urine and blood from the same subjects. Its delayed appearance was consistent with colonic synthesis. On the basis of the pharmacokinetics, optimum steady-state serum isoflavone concentrations would be expected from modest intakes of soy foods consumed regularly throughout the day rather than from a single highly enriched product. J. Nutr. 133: 1027-1035, 2003. KEY WORDS: * phytoestrogen * isoflavone * pharmacokinetics * soy foods * humans

We investigated whether the bioavailability of isoflavones could be enhanced by enzymatic hydrolysis of glycosides to aglycones before consumption of a nonfermented soy food. Two drinks were formulated with an enriched isoflavone extract from soy germ (Fujiflavone P10), one of which was hydrolyzed enzymatically with [beta]-glucosidase to produce aglycones. In a randomized, double-blinded, cross-over study, six European, post-menopausal women consumed each soy drink at a 1-wk interval at a concentration of 1 mg total isoflavones/kg body. The plasma and urinary pharmacokinetics of daidzein, genistein and glycitein did not differ after consumption of the two beverages. Plasma total isoflavone concentrations reached 4-5 [micro]mol/L. The pharmacokinetics of glycitein were similar to those of daidzein. The isoflavone secondary metabolites detected were dihydrodaidzein in plasma and O-desmethylangolensin, equol, and dihydrogenistein in urine. The ratios of individual isoflavones to one another were not conserved from food to plasma to urine, indicating that the individual isoflavones do not have the same absorptions and body retentions. In conclusion, previous hydrolysis of glycosides to aglycones does not enhance the bioavailability of isoflavones in humans. KEY WORDS: * isoflavones * soy * bioavailability * excretion * glycosides * plasma * humans

Genistein, a soy isoflavone, has in vitro immunosuppressive properties. We investigated whether genistein or dietary soy protein containing isoflavones could influence the outcome of rat cardiac allografts. Lewis rats were fed a diet with protein from high isoflavone soy protein fraction (HIS), casein (CAS) or casein with isoflavones added (CI) starting 1 wk before heart transplants from Wistar Furth donors, and continuing throughout the study. HIS-fed rats had significantly prolonged time to rejection compared with CAS- and Cl-fed recipients (10.8 [+ or -] 2.62 vs. 7.18 [+ or -] 0.75 and 7.22 [+ or -] 0.44 d, P < 0.001). Intravenous genistein [20mg/(kg * d) for 14 d] significantly prolonged heart survival compared with controls and dissolvent-treated recipients (23.2 [+ or -] 7.4 vs. 8.4 [+ or -] 1.3 and 11.4+/3.6 d, P < 0.0005), and had an additive effect when given to heart recipients also receiving low dose cyclosporine for 7 d (30.8 [+ or -] 2.3 vs. 23.4 [+ or -] 2.4 d, P < 0.005). Concanavalin A-stimulated lymphocytes, isolated from Lewis rats given intraperitoneal genistein for 7 d, had decreased production of interferon [gamma] compared with controls or dimethyl sulfoxide-treated groups (22.6 [+ or -] 9.9 vs 149 [+ or -] 105 and 154 [+ or -] 103 [micro] g/L, P < 0.05). In conclusion, a high isoflavone soy diet and intravenous genistein, but not isoflavone extract alone, delay rejection of rat cardiac allografts, with an additive effect in cyclosporine-treated rats. In addition, intraperitoneal genistein has immunosuppressive properties in vivo. KEY WORDS: * soy * genistein * allograft * rejection * isoflavone * rats

The effect of soy protein, soy isoflavones and saponins on paraquat (PQ)-induced oxidative stress was investigated in rats. Rats were fed experimental diets containing casein (CAS), soy protein (SPI), and casein with soy isoflavones and saponins (CAS + IS). The diets were supplemented or not with 0.025% paraquat (CAS + PQ, SPI + PQ, and CAS + IS + PC)). The protective effects of soy protein, soy isoflavones, and saponins on paraquat-induced oxidative stress were examined. Ingestion of soy protein generally mitigated the lung enlargement (P = 0.076), loss of body weight (P = 0.051) and oxidation of liver lipid (P = 0.043) and glutathione (P = 0.035) induced by paraquat, although soy isoflavones and saponins did not. To determine whether soy protein exerted its antioxidative effects by preventing paraquat absorption from digestive organs, rats were fed CAS or SPI diets and orally administered a 12.5 g/L paraquat solution. Plasma, urine, and fecal paraquat concentrations did not differ between the two groups, indicating that soy protein did not prevent paraquat absorption. The present study suggests that intake of soy protein itself, but not soy isoflavones and saponins, reduces paraquat-induced oxidative stress in rats, although this effect was not due to reduced absorption of paraquat from digestive organs. KEY WORDS: * rats * soy protein * paraquat * oxidative stress * isoflavone * saponin

We used a previously developed compartmental model to assess the postprandial distribution and metabolism of dietary nitrogen (N) in the splanchnic and peripheral areas after the ingestion of a single mixed meal containing either [sup.15]N-labeled milk or soy purified protein. Although the lower whole-body retention of dietary N from soy protein was measured experimentally, the splanchnic retention of dietary N was predicted by the model not to be affected by the protein source, and its incorporation into splanchnic proteins was predicted to reach ~35% of ingested N at 8 h after both meals. However, dietary N intestinal absorption and its appearance in splanchnic free amino acids were predicted to be more rapid from soy protein and were associated with a higher deamination, concomitant with a higher efficiency of incorporation of dietary N into proteins in the splanchnic bed. In contrast, soy protein was predicted to cause a reduction in peripheral dietary N uptake, as a consequence of both similar splanchnic retention and increased oxidation compared with milk protein. In addition, protein synthesis efficiency was reduced in the peripheral area after soy protein intake, leading to dietary N incorporation in peripheral proteins that fell from 26 to 19% of ingested N 8 h after milk and soy protein ingestion, respectively. Such a model thus enables a description of the processes involved in the differential metabolic utilization of dietary proteins and constitutes a valuable tool for further definition of the notion of protein quality during the period of protein gain. KEY WORDS: * interorgan amino acid metabolism * milk protein * soy protein * postprandial period * parameter estimation H

The mechanisms by which dietary soy favorably influences lipoprotein metabolism and inhibits atherosclerosis are uncertain. Studies of blood mononuclear cells and cultured hepatocytes have indicated that certain soy peptides (i.e., 7S globulins) stimulate expression of LDL receptors. This pathway represents a hypothetical mechanism by which soy's hypocholesterolemic and antiatherosclerotic effects may be mediated. However, direct evidence supporting this hypothesis is lacking. To address this, we compared effects of dietary soy protein isolate in two genetically engineered mouse models of atherosclerosis. One mouse [LDL receptor -/- apolipoprotein (apo) B transgenic] is devoid of LDL receptors and overproduces apolipoprotein B, whereas the other (apoE -/-) has a normal complement of LDL receptors but does not produce apolipoprotein E. Male (n = 10-12/group) and ovariectomized female (n = 10-12/group) mice were studied. There were three treatment groups, which differed principally by the source of the protein component of the diet: 1) casein/lactalbumin (no isoflavones), 2) alcohol-washed soy protein isolate (total isoflavones = 0.04 mg/g), and 3) intact soy protein isolate (total isoflavones = 1.72 mg/g). Atherosclerosis was assessed by quantifying the aortic content of esterified cholesterol. Atherosclerosis was inhibited (relative to the casein/lactalbumin group) by both alcohol-washed (45 and 31%) (P < 0.05) and intact (65 and 41%) (P < 0.05) soy protein isolate in LDL receptor -/- and apoE -/-mice, respectively. There was no sex difference. In a two-way analysis, there were significant effects of type of soy isolate and type of mouse. The antiatherosclerosis effect was enhanced in LDL receptor -/- mice (P < 0.001) and diminished in mice fed alcohol-washed soy protein isolate (P < 0.001). Furthermore, inhibitory effects of soy on atherosclerosis were unrelated to plasma LDL, VLDL or HDL cholesterol concentrations. The results represent direct evidence for the existence of LDL receptor- and plasma lipoprotein-independent pathways by which dietary soy protein isolate inhibits atherosclerosis. KEY WORDS: * atherosclerosis * mice * soy protein * isoflavones * lipoproteins

We examined the biological fate of 7S globulin from soybean in a hepatoma cell line (Hep G2) and in human skin fibroblasts (HSF) to gain new insights into the 7S globulin cell process, the final effect of which is an enhanced expression of the LDL-receptor. The ability of 7S globulin to bind and to be internalized and degraded by both cell types was investigated under different experimental conditions. In all cases, specific uptake (binding + internalization) and degradation of 125I-7S globulin were curvilinear functions of substrate concentration at 37 [degrees] C. The two processes were saturated at around 80 mg/L, a concentration at which an up-regulation of LDL-receptor was previously reported. The specific uptake of 125I-7S globulin at 37 [degrees] C was a curvilinear function of time, and achieved equilibrium after 6 and 12 h in HSF and Hep G2 cells, respectively. Binding experiments, conducted at 4 [degrees] C in Hep G2 cells, showed a specific and saturable association of 7S globulin to the cell membrane. Linear Scatchard analysis demonstrated a single population of binding sites. The amount of 7S globulin bound at saturation ([B.sub.max]) was about 2.73 mg/L, with an apparent [K.sub.d] of 21/[micro]mol/L, assuming 175 kDa as the 7S globulin molecular weight. SDS-PAGE of Hep G2 membrane proteins incubated with 125I-7S globulin revealed a specific interaction of 7S globulin with a cell protein component with molecular weight between 14 and 21 kDa. Further studies are needed to ascertain whether this interaction is directly or indirectly related to the observed stimulation of the LDL-receptor. INDEXING KEY WORDS: 75, 11S soybean globulins, Hep G2 cells, human skin fibroblasts, cholesterol reduction

Possible associations between the intake of soy products and reduced cancer risk are discussed. These spring to mind because of the identification, cloning, and characterization of a potent antimitotic peptide from soybeans.

Soy isoflavone supplementation in healthy men prevents NF-kappaB activation by TNF-alpha in blood lymphocytes. Davis JN, Kucuk O, Djuric Z, Sarkar FH. Free Radic Biol Med 2001;30:1293-1302. Dietary intake of [...]

Lack of effect of soy isoflavone on thyroid hyperplasia in rats receiving an iodine-deficient diet. Son HY, Nishikawa A, Ikeda T, et al. Jpn J Cancer Res 2001;92:103-108. We have [...]

Genistein and daidzein are the most abundant isoflavonoids in soybeans, soy meal and tofu. These two molecules exhibit anticancer chemopreventive activities. The significantly lower incidence of breast and prostate cancer in populations consuming high amounts of food containing genistein and daidzein has been attributed to their ability to bind to the estrogen receptor and compete with endogenous estrogens. Furthermore, genistein is an inhibitor of tyrosine kinase and decreases breast cancer cell proliferation. A few reports also suggest that genistein is inhibits endothelial cell proliferation and could potentially inhibit angiogenesis. We examined the effect of genistein and daidzein on both the cell cycle and apoptosis of human umbilical vein endothelial cells (HUVEC). We demonstrate that these two molecules inhibit, in a dose-dependent fashion, HUVEC proliferation by blocking the cell cycle at the phases G2/M and G1 and also significantly induce their apoptosis. Genistein and daidzein abolish the sprouting of capillaries from rat aortic rings and inhibit in vivo angiogenesis in the chicken chorioallantoic membrane assay. Interestingly, the combination of both isoflavonoids increases the antiangiogenic effect of these compounds. Our data demonstrate that genistein and daidzein are angiogenesis inhibitors that could be included in the antiangiogenic arsenal currently being developed to fight cancer and other angiogenesis-dependent diseases.

The activation of LDL receptors was described recently in a human hepatoma cell line (Hep G2) exposed both to [Alpha] + [Alpha]' subunits from 7S soy globulin and to [Croksoy.sup.R]70, a commercial isoflavone-poor soy concentrate. To assess the final identity of the peptide(s) putatively responsible for the biochemical effect, experiments were performed in Hep G2 cells, exposed either to synthetic peptides corresponding to specific sequences of 7S soy globulin or to peptides from the in vitro digestion of [Croksoy.sup.R]70. Moreover, the ability of the whole 7S globulin, its subunits and whole [Croksoy.sup.R]70 to interfere in the apolipoprotein B (apo B) secretion in the medium as well as in sterol biosynthesis was evaluated in the same model. Increased [sup.125]I-LDL uptake and degradation vs. controls were shown after Hep G2 incubation with a synthetic peptide ([10.sup.-4] mol/L, MW 2271 Da) corresponding to positions 127-150 of the 7S globulin. Cells exposed to [Croksoy.sup.R]70 enzyme digestion products showed a more marked up-regulation of LDL receptors vs. controls, compared with vs. Hep G2 cells incubated with undigested [Croksoy.sup.R]70. Among soy-derived products, only the 7S globulin inhibited apo B secretion and [sup.14]C-acetate incorporation when tested in Hep G2 cells at a concentration of 1.0 g/L. These findings support the hypothesis that if one or more peptides can reach the liver after intestinal digestion, they may elicit a cholesterol-lowering effect. Moreover, the protein moiety, devoid of isoflavone components, is likely to be responsible for this major biochemical effect of soy protein. J. Nutr. 130: 2543-2549, 2000. KEY WORDS: * soy proteins * LDL receptors * apolipoprotein B * cholesterol homeostasis * Hep G2 cells

Soy phytoestrogens have been shown to increase plasma levels of HDL cholesterol and apolipoprotein (apo) A-I, its major protein component, in animal studies and in some human studies. The human hepatoma cell line Hep G2 was used to study the effect of the phytoestrogens genistein and daidzein on apo A-I secretion and gene expression in liver cells. Both genistein and daidzein increased apo A-I secretion in a dose-dependent fashion. Apo A-I concentration in the media of treated cells was increased approximately fivefold by 10/[micro]mol/L genistein (P [is less than] 0.001) and approximately onefold by 10/[micro]mol/L daidzein (P [is less than] 0.001) compared with control cells. The effect of genistein on apo A-I secretion was similar to that observed with 17-[Beta]-estradiol. Treatment of cells with genistein for 24 h increased the transcriptional activity of the apo A-I gene as measured by nuclear run-on assay. Transfection experiments with plasmids containing regulatory regions of the apo A-I gene cloned in front of the luciferase reporter gene indicated that the 5' region of the apo A-I gene contained between nucleotides -256 and -41 is responsible for the increased expression of this gene by genistein. J. Nutr. 130: 2489-2492, 2000. KEY WORDS: * phytoestrogens * genistein * daidzein * apolipoprotein A-I * gene expression

To examine the kinetics of carnitine palmitoyl-transferase-I (CPT-I) and the influence of dietary variables, young pigs (18 kg, n = 20) were fed corn-soybean meal diets supplemented with 40 g soy oil/kg and containing either 136 or 180 g crude protein/kg and either 0 or 500 mg/kg L-carnitine (2 x 2 factorial design). Diets were offered for 10 d (85% of ad libitum); CPT-I activities in liver and skeletal muscle mitochondria were determined, and enzyme kinetic constants ([V.sub.max] and [K.sub.m] for carnitine) were estimated. Kinetics of CPT-I in muscle were not affected by diet (P [is greater than] 0.1; carnitine [K.sub.m] = 480 [+ or -] 44 [micro]mol/L). In contrast, the [K.sub.m] for carnitine in liver was increased from 164 to 216 [+ or -] 20 [micro]mol/L by dietary L-carnitine supplementation (P [is less than] 0.01) and from 169 to 211 [+ or -] 20 [micro]mol/L by high protein feeding (P [is less than] 0.05). Dietary L-carnitine increased muscle and liver free carnitine concentrations by 72 and 158% over control concentrations (770 and 80 [micro]mol/kg wet muscle and liver, respectively). Because tissue carnitine concentrations were within the range of the respective [K.sub.m] for both liver and muscle tissue, it is inferred that alteration of tissue carnitine concentrations via dietary supplementation could modulate CPT-I activity in young pigs. J. Nutr. 130: 2467-2470, 2000. KEY WORDS: * pigs * carnitine * protein * carnitine palmitoyltransferase-I * liver * muscle

We investigated the ability of genistein and daidzein, two soybean isoflavones, compared with that of 17 [Alpha] -ethinylestradiol, to prevent bone loss in ovariectomized rats, a model for postmenopausal osteoporosis. Female Wistar rats (n = 65; 12 mo old) were either sham-operated (SH; n = 13) or ovariectomized (OVX; n = 52). On d 0, OVX rats were randomly assigned to groups as follows: 13 received genistein [G; 10 [micro] g/(g body weight * d)], 13 were treated with daidzein [D; 10 [micro] g/(g body weight * d)], 13 received 17 [Alpha] -ethinylestradiol [E.sub.2; 30 [micro] g/kg body weight * d)] and 13 were untreated (OVX). Compounds were mixed with a soy protein-free powdered semipurified diet and given orally for 3 mo. On d 90, the bone mineral density (BMD) in lumbar vertebrae, femur and its metaphyseal and diaphyseal zones (rich in cancellous and cortical bone, respectively) was lower in OVX than in SH (P [is less than] 0.01). In D or E.sub.2, the four BMD were not different from SH, whereas in G, only the diaphyseal BMD was not different from SH. Image analysis performed in the distal femur metaphysis revealed that the cancellous bone area was lower in OVX than in SH (P [is less than] 0.01). Only the area in D was not different from that in SH. Finally, the bone turnover, which was higher in OVX than in SH (P [is less than] 0.005 and P [is less than] 0.05 for plasma osteocalcin concentration and urinary deoxypyridinoline excretion, respectively), was not different in G, D or E.sub.2 compared with SH. Therefore, consumption of 17 [Alpha] -ethinylestradiol or daidzein was more efficient than genistein in preventing ovariectomy-induced bone loss in rats. J. Nutr. 130: 1675-1681, 2000. KEY WORDS: * genistein * daidzein * prevention * bone * rats

In mammals, protein ingestion increases the glomerular filtration rate (GFR), an effect which has been incriminated as a risk factor in progression of renal disease. Some studies suggest that a postprandial increase in GFR is absent or mild with vegetable proteins compared to animal proteins. The objective of this experiment was to determine whether vegetable (soy) protein had different effects than animal protein on GFR in dogs with normal or reduced renal function. A trial was conducted in which GFR was measured in four dogs with normal kidney function and seven dogs with reduced renal mass before and after administering protein. Normal dogs were fed four protein sources (casein, soy meal, soy flakes and purified soy protein). Dogs with reduced renal mass were fed three protein sources (casein, purified soy protein and pork liver). All proteins significantly (P [is less than] 0.05) increased the GFR in both groups except for casein (P = 0.066) in normal dogs. Proteins did not differ significantly in the magnitude of the increase in GFR that was induced. This study indicates that soy proteins in dogs have the same effect on GFR as animal-source proteins, which is contrary to reports of effects in humans. J. Nutr. 130: 745-748, 2000. KEY WORDS: * glomerular filtration rate * soy protein * dogs * renal disease

There is a general agreement that soy isoflavones can be beneficial to health in adults. However, isoflavones have acted as endocrine-disrupting chemicals. Therefore, some consideration should be given to the possibility that soy foods may adversely affect the reproductive system in infants. On d 5 of pregnancy, we fed Sprague-Dawley rats one of three diets containing daidzein and genistein and having a total isoflavone concentration of 0, 0.5 and 1.0 g/kg. Although the body weight and serum triiodothyronine concentration of dams in the 1.0-g/kg group were lower than those in O-g/kg group, no significant differences were observed among the suckling pups in the three groups. We detected both daidzein and genistein in the blood of dams and pups and the stomach contents (milk) of pups. However, small amounts of isoflavones transferred from mother to suckling pups.

A dose range finding study was conducted by feeding genistein to Sprague-Dawley rats at 0, 5, 25, 100, 250, 625 and 1250 ppm in a soy-free diet, from gestation d 7 to postnatal d 50. Plasma levels ranged from ~l0 nmol/L to 10 [micro]mol/L. Adverse effects included hypertrophy and hyperplasia of the mammary ducts and acini in males at 25 ppm and in females at 625 ppm. Hypospermia at the head of the epididymis, inflammation of the dorsal prostate and asynchronous cycles of the uterus and vagina were observed at 625 ppm. Degeneration of the ovaries and seminiferous tubules was seen at 1250 ppm. There was a dose-dependent decrease in thyroid peroxidase activity at 25-1250 ppm. Thyroid peroxidase activity is known to be inhibited in vitro by genistein, and the prevalence of autoimmune thyroiditis was reported to be increased in children fed soy infant formula. In a separate study, genistein (1-100 [micro]g) was injected into neonatal mice daily on postnatal d 1-5 and the mice were killed at 18 mo. Polyovular multioocyte) follicles, uterine epithelial hyperplasia, hypoplastic uteri and uterine adenocarcinoma were observed. These were similar to the lesions reported earlier to be induced by diethylstilbestrol in mice by the use of the same treatment protocol. In addition to genistein, 45 other phyto- or mycoestrogens were assayed in a uterine estrogen receptor competitive binding assay. Twenty-nine of these competed and had a wide range of relative binding affinities. The number of naturally occurring chemicals that can bind to the estrogen receptor suggests that more intense or additional effects may be expected from exposures to multiple estrogenic chemicals from plants. These findings show a significant number of adverse effects from genistein in estrogen target organs, including malignancies. Some of these effects occur at low doses, consistent with findings from other estrogens. Additionally, genistein inhibition of thyroid peroxidase suggests the possibility that this mechanism may be responsible for goiter and autoimmune thyroiditis. The results of these studies have been used to design an ongoing multi-generation study with genistein that will examine reproductive fitness, among other endpoints. [Supported by USEPA #R825299-01-0.]

Important among estrogen's benefits for postmenopausal women is the maintenance of cognitive function (brain agonist effects). As a part of our efforts to determine the optimal role of soy phytoestrogens in postmenopausal hormone therapy, we used rat models to determine the effectiveness of soy phytoestrogens relative to 17 [Beta]-estradiol ([E.sub.2]) in regulating biomarkers important to cognitive function and to address whether soy phytoestrogens attenuate the beneficial effects of [E.sub.2] on working memory. We examined the effects of oral [E.sub.2] and soy phytoestrogen extract on the mRNA levels of choline acetyltransferase (CHAT), nerve growth factor and brain-derived neurotrophic factor (BDNF) in ovariectomized young and retired breeder rats. Within each age category, 15 bilaterally ovariectomized rats were randomly divided into three groups, i.e., untreated control, [E.sub.2] and soy phytoestrogen groups, and were treated for 8 wk. Both [E.sub.2] and soy phytoestrogens up-regulated ChAT and BDNF mRNA in the frontal cortex of ovariectomized retired breeder rats. In addition, [E.sub.2] significantly up-regulated nerve growth factor mRNA in the hippocampus of young rats. The nerve growth factor mRNA level in the hippocampus of the soy phytoestrogen group was intermediate between the other two groups. In a separate study, we examined whether soy phytoestrogens could improve working memory and whether they would attenuate the beneficial effects of [E.sub.2] in ovariectomized retired breeder rats; we used a radial-arm maze test. Bilaterally ovariectomized retired breeder rats (n = 84) were randomly divided into 12 groups (in a 3 x 4 factorial design) and were treated for 10 mo. There were four levels of [E.sub.2] (0, 0.5, 1.0 and 2.0 mg/1800 kcal; women's equivalent doses), three levels of soy with phytoestrogens (0, 72 and 144 mg/1800 kcal), and all combinations of these doses of [E.sub.2] and soy phytoestrogens. Rats in groups given only [E.sub.2] or soy phytoestrogens showed a dose-dependent improvement in test performance. In addition, there was no attenuation of the beneficial effects of [E.sub.2] by soy phytoestrogens.

Recent evidence indicates that a diet containing soy isoflavones can have beneficial effects on the cardiovascular system, including improving the plasma lipid profile. Effects on blood pressure and vascular reactivity, however, are controversial. We examined the effects of a soy diet on blood pressure and skin blood flow reactivity in men and postmenopausal women with moderate levels of hypertension. Subjects were admitted to the study when they exhibited an average diastolic blood pressure of at least 90 mm Hg over a 12-h period after they had stopped taking antihypertensive medications for at least 2 wk. The protocol was a randomized, double-blind, crossover design with a 2- to 4-wk run-in period, a 4-wk treatment period in which subjects ate soy food either with (Soy+) or without (Soy-) isoflavones, a 2-wk washout period and a second 4-wk treatment period with the other soy treatment. Women ate soy muffins daily that contained 38 g soy protein (73 mg total isoflavone, 43 mg aglycone isoflavone). Men consumed soy muffins and drinks daily with 35 g soy protein (162 mg total isoflavone, 95 mg aglycone isoflavone). Blood pressure measurements were made weekly for 12 h/d. In addition, skin blood flow responses to a 3-min occlusion of the arm were routinely measured with a laser Doppler flow monitor. Systolic, diastolic and mean arterial blood pressures, heart rate and pulse pressure are presented in Table 1 (means [+ or -] SEM). After occlusion of the arm for 3 min, skin blood flow increased transiently and returned to near-normal levels within 3 min but rose initially to a higher level during Soy+ treatment. Although Soy+ treatment may alter some vascular responses, our findings indicate that a short-term diet containing soy isoflavone has no overall effect on blood pressure in moderately hypertensive subjects during a 1-mo diet period. [Supported by a grant from the South Dakota Soybean Research and Promotion Council.]

Results from studies in the hypercholesterolemic hamster model as well as a brief review of the literature with respect to saponins as hypocholesterolemic agents will be presented. Sixty male and 60 female Golden Syrian hamsters were randomly assigned to six different groups (n = 10) and fed for 10 wk. The experimental diets contained 0.1% cholesterol and 10% coconut oil to produce hypercholesterolemia. The six treatments were casein control, isolated soy protein (ISP), ISP-, daidzein, soy germ (SoyLife) and soy-germ extract. Diets provided a total isoflavone content of 1.3 mmol/kg diet (except for casein and ISP- groups). The ISP- diet contained 0.013 mmol isoflavone/kg diet. All hamsters fed ISP, ISP-, daidzein, soy germ and soy-germ extract had significantly lower plasma total cholesterol (16-28% lower), LDL cholesterol (15-50% lower) and ratios of LDL to HDL cholesterol than did the control group (P [is less than] 0.01). For male hamsters, there were no differences among treatments in plasma HDL concentrations. For females, the ISP and ISP -- groups had significantly higher HDL levels (20-30% higher) (P [is less than] 0.01) than did the casein groups. There were no differences in triglyceride levels among treatments for both sexes. These data support the cholesterol-lowering effects in hamsters of soy protein with or without isoflavone, the soy isoflavone daidzein and soy germ and suggest that a plateau of maximal hypocholesterolemia was achieved by all of these treatments. A hamster study of similar design is planned to compare effects of the following five treatments vs. a casein control: ISP, ISP-, ISP extract containing both isoflavone and saponins, purified daidzein and genistein, and purified saponins in amounts comparable to their amounts in intact ISP.

Previous studies established that soy protein has a cholesterol-lowering effect, but many questions remain. For example, does soy protein affect the lipid profile of men and women in a similar matter and are the isoflavones responsible for this lipid effect? Numerous concerns about soy protein have also been raised. We performed three studies to examine these questions in some detail. Study 1 involved 54 postmenopausal women with a total cholesterol level (TC) [is greater than] 5.5 mmol/L after a 4-wk cholesterol-lowering diet. Subjects were randomly assigned to receive normal soy protein (ISP+) containing 65 mg isoflavone or soy protein with isoflavone alcohol-extracted (ISP-) for 12 wk. Subjects were reassessed 4 wk after (off phase) the end of the 12-wk treatment phase. The following were measured at intervals: TC, triglyceride (TG), HDL, LDL, luteinizing hormone (LH), follicle-stimulating hormone (FSH), sex hormone--binding globulin (SHBG), thyroid-stimulating hormone (TSH), bone markers (bALP, osteocalcin, and urinary deoxypyridinoline), as well as glucose and insulin response to a glucose load. The mean age of subjects was 57 y; mean baseline cholesterol was 7.3 mmol/L. There was no weight change over the study period. There were five dropouts, leaving 49 for analysis. The mean lipid reductions were 4.3% for TC and 6.1% for LDL. No difference was seen between ISP+ and ISP--, implying that some component other than isoflavones was responsible for the lipid-lowering effect. No effect was seen on glucose metabolism, TSH, bone markers (apart from a mild effect on urinary deoxypyridinoline), FSH or Greene menopause scores. A mild but significant effect was seen for ISP+ on serum levels of LH, and a significant reduction was seen for SHBG in the ISP+ group. Interestingly the levels of SHBG returned to normal during the off phase. Study 2 was an uncontrolled study of 27 men with a TC level [is greater than] 5.5 mmol/L after a 4-wk cholesterol-lowering diet. During the 12-wk study period, all subjects were given 28 g soy protein containing 65 mg isoflavone. Subjects were reassessed 4 wk after (off phase) the end of the 12-wk treatment phase. The following were measured: lipid profile, LH, FSH, SHBG, TSH and androgen profile. The mean age of subjects was 52 y; mean baseline TC was 6.8 mmol/L. The average increase in HDL cholesterol was 9.5%; this value returned to baseline during the off phase. There was no effect on TSH, testosterone, androstenedione, LH or FSH. As was seen in the study with women, there was a [is greater than] 10% decrease in SHBG and a nonsignificant increase in DHEA (P = 0.06). Study 3 involved women using estrogen patches for a total of 12 wk, but during the middle 4 wk they were also given soy protein to see whether this interfered with their estrogen levels. The average estradiol level before soy was 200 pmol/L and during soy was 212 pmol/L. Thus no effect was seen. In conclusion, these three studies demonstrated that men and women appear to have a different response to soy protein in terms of their lipid profile. In the studies both with men and women, a significant reduction in SHBG was demonstrated. This is a novel finding because previous studies using soy and cereal typically showed an elevation in SHBG.

Soy stanols and sterols act in the intestine to block cholesterol absorption and reduce LDL cholesterol. However, more work is needed to establish the safety of these agents for general use in foods. Although absorption of phytosterols in humans is less than that for cholesterol, the exact level is not known, especially under conditions of intended use. Concern has been expressed about reports of appreciable absorption of campestanol and campesterol, which comprise about 25% of soy sterols. The current study was designed to measure the systemic absorption of lecithin-emulsified soy sterols and stanols using a double stable isotope technique in which one label was given orally and another intravenously. [2,2,4,4,6-[sup.2][H.sub.5]] Soy sterols (pentadeuterated) were prepared and an aliquot was reduced to the corresponding 5[alpha]-stanols using platinum-charcoal catalyst. [3,4-[sup.2][H.sub.5]]Soy sterols (dideuterated) were treated similarly. Ten normal subjects underwent two single-meal absorption tests in random order 2 wk apart, one with intravenous dideuterated sterols and oral pentadeuterated sterols and the other with the corresponding labeled stanols. A standard test breakfast was given including a lemonade drink containing 625 mg emulsified labeled oral tracer and an equal amount of enzyme-modified soy lecithin. Plasma tracer levels were measured by negative ion mass spectrometry 8 h and 1,2,3, and 4 d later. The ratio of labeled oral and intravenous tracers became constant in plasma after 2 d, reflecting absorption. The absorption of sitosterol from mixed soy sterols was 0.51% [+ or -] 0.04% and that of campesterol was 1.89 [+ or -] 0.27%. The absorption of sitostanol from mixed soy stanols was 0.044% [+ or -] 0.004% and campestanol absorption was 0.16% [+ -] 0.02%. The absorption of lecithin-emulsified soy stanols and sterols is very small and less than predicted from older work. Stanols have 1/10 the absorbability of sterols.

Soy isoflavones have antioxidant activities in vitro and have been reported to contribute to increasing LDL ex vivo oxidation resistance. LDL oxidative damage is implicated in the pathogenesis of atherosclerosis. The objective of this project was to investigate the effects in postmenopausal women of a dietary soy protein supplement containing isoflavones (Soy+) compared with soy protein without isoflavones (Soy-) and a control casein protein (Cas). The primary endpoint was brachial artery reactivity (reported elsewhere). Secondary endpoints were the effect of the three protein supplements on ex vivo LDL oxidation susceptibility and whole plasma oxidation. A double-blind, randomized, crossover design was used to compare the three proteins, which were consumed daily for 6 wk each, separated by a 4-wk washout. Resistance of LDL to copper ion-induced oxidation was determined. The study population consisted of 28 healthy, normolipidemic, postmenopausal women with mean age 54.9 y (range 46-69 y) and body mass index 24.6 [+ or -] 0.6. The proteins contributed 100 kcal and total isoflavone aglycone units of 107, 2, and 0 mg for the Soy+, Soy-, and Cas supplements, respectively. Dietary intake in this free-living population was assessed by 3-d food records at baseline and remained relatively constant during the study. Plasma lipids at baseline were 191.7 [+ or -] 3.6 mg/dL (4.97 [+ or -] 0.09 mmol/L) for total cholesterol, 104.1 [+ or -] 3.8 mg/dL (2.70 [+ or -] 0.10 mmol/L) for LDL cholesterol, 61.1 [+ or -] 2.9 mg/dL (1.58 [+ or -] 0.08 mmol/L) for HDL cholesterol, and 91.0 [+ or -] 7.5 mg/dL (1.03 [+ or -] 0.08 mmol/L) for triglycerides. Both soy treatments resulted in significant (P < 0.05) decreases in total cholesterol and LDL cholesterol of 2.0-2.4% and 0.5-1.5%, respectively, compared with baseline. In a subset (n = 12) with consistent intake of [alpha]-tocopherol equivalents (SD < 1 between treatments) there was a significant increase (P [less than or equal to] 0.05) in mean conjugated diene lag time of 1-5% after consumption of both soy protein supplements. There was no significant difference in the resistance of whole plasma to oxidation between treatment groups or compared with baseline.

In this study, the effects of several additives such as cad 2, sulfur, [H.sub.2][O.sub.2], and lipoxygenase on the quality of soy protein (concentrated and texturized) were investigated. The change in the amount of phytochemicals in the modified soy protein was noticeable. The main isoflavones in soy are genistein, daidzein, and glycitein. LDL concentration was lowered by 30% if diets contained soy protein three times per week (75 g/wk). In this paper, the effect of different amounts of soy protein in the daily diet are reviewed, The results show that 5-7 g soy protein per serving decreases the concentration of LDL and thus decreases the risk of coronary heart disease.

Some recent studies have reported that hormone (estrogen) replacement therapy (HRT) reduces the risk of cardiovascular disease in postmenopausal women. However, HRT is associated with carcinogenic side effects in women. Soybean isoflavones are phytoestrogens and are not associated with carcinogenesis. Therefore, we evaluated the preventive effects of isoflavone aglycone-rich fermented soybean extract (IFA-FSE) on hypercholesterolemia and atherosclerosis induced by a cholesterol diet in aged female Japanese quails. Twenty-four 15mo-old female Japanese quails were housed in separate cages in a comfortable environment and randomly assigned to three experimental groups: a control group (CON) consumed a control diet, a cholesterol group (CHO) consumed a control diet with 2% cholesterol, and a cholesterol and IFA-FSE treatment group consumed a control diet with 2% cholesterol with IFA-FSE given by gavage at 100 mg/kg body weight. IFA-FSE contained 30% isoflavone aglycones by weight (70% daidzein, 10% genistein, and 20% glycitein) and was provided by Nichimo Co. (Tokyo, Japan). The experiment lasted 2 mo. The serum total cholesterol was 132.8 [+ or -] 18.2 mg/dL (3.44 [+ or -] 0.47 mmol/L) in CON and was significantly increased to 521.8 [+ or -] 54.5 mg/dL (13.51 [+ or -] 1.41 mmol/L) in CHO (P < 0.001). IFA-FSE treatment significantly decreased serum total cholesterol to 339.0 [+ or -] 49.7 mg/dL (8.78 [+ or -] 1.29 mmol/L; P < 0.01 compared with CHO). Atherosclerotic lesion area was measured with an Olympus DP50 image analyzer and expressed as a percentage of the total surface area of the aortic intima. Atherosclerotic lesion area of aortic arch was 35.0% [+ or -] 1.3% in CON and was significantly increased to 75.7% [+ or -] 7.6% in CHO (P < 0.001). This cholesterol-induced atherosclerotic lesion was significantly inhibited by IFA-FSE treatment by 23% (58.5% [+ or -] 3.3%, P < 0.05). In comparison with CON (8.4% [+ or -] 1.5%), cholesterol also significantly increased the atherosclerotic lesion area of thoracic aorta (40.4% [+ or -] 5.2%, P < 0.001), and this increase was significantly inhibited by IFA-FSE treatment (23.1% [+ or -] 3.3%, P < 0.05). Our studies suggest that IFA-FSE may prevent atherosclerotic cardiovascular diseases by its hypolipidemic activity. This is the first report on the preventive effect of isoflavone aglycone on the development of atherosclerosis in a Japanese quail model.

Studies of blood mononuclear cells and cultured hepatocytes have indicated that certain soy peptides (i.e., 7S globulins) stimulate expression of LDL receptors. This pathway represents a hypothetical mechanism by which soy's hypocholesterolemic and antiatherosclerosis effects may be mediated. However, direct evidence supporting this hypothesis is lacking. To address this, we compared effects of dietary soy protein isolate in two genetically engineered mouse models of atherosclerosis. One mouse (LDL receptor -/- + apoB transgenic) is devoid of LDL receptors and overproduces apoprotein B whereas the other (apoE -/-) has a normal complement of LDL receptors but does not produce apoprotein E. Male (n = 1012/group) and ovariectomized female (n = 10-12/group) mice were fed diets containing 20% protein, 64% carbohydrate, and 16% fat for 16 wk. Three treatment groups differed only in the source of the protein component of the diet: 1) casein/lactalbumin (no isoflavones), 2) alcohol-washed soy protein isolate (total isoflavone content 0.04 mg/g), and 3) intact soy protein isolate (total isoflavone content 1.72 mg/g). Atherosclerosis was assessed by quantifying the aortic content of esterified cholesterol. Atherosclerosis was inhibited (relative to the casein/lactalbumin group) by both alcohol-washed (45% and 31%, P < 0.05 for each) and intact (65% and 41%, P < 0.05 for each) soy protein isolate in LDL receptor -/- and apoE -/mice, respectively. There was no sex difference. In a two-way analysis, there were highly significant effects of type of soy isolate and type of mouse. The antiatherosclerosis effect was enhanced in LDL receptor -/- mice (P < 0.001) and diminished in mice fed alcohol-washed soy protein isolate (P < 0.001). Furthermore, inhibitory effects of soy on atherosclerosis were unrelated to plasma LDL, VLDL, or HDL cholesterol concentrations. The results represent direct evidence for the existence of LDL receptor-independent and plasma lipoprotein-independent pathways by which dietary soy protein isolate inhibits atherosclerosis.

Consumption of soy isoflavones is expected to decrease the risk of breast cancer, osteoporosis, coronary heart disease, and menopausal symptoms. In. Asian populations soy isoflavones are usually consumed as traditional foods. Although the issue of toxicity has been raised, the level of daily dietary soy isoflavone intake is considered to be safe. However, few studies have investigated the overdose safety issue of soy isoflavones taken as a supplement. The toxicity of soy isoflavones would be related to their plasma concentrations after ingestion. We therefore investigated the dose response of plasma concentration and urinary excretion of isoflavones after a single administration of soy isoflavone supplements. Eleven healthy young adults (5 female and 6 male) were given a tablet containing 38.3 mg soy isoflavones. Five doses (0, 3, 9, 15, and 21 tablets) were administered to the same individuals with a 1-wk wash out period between doses. Blood samples were collected at 0, 6, 8, and 24 h after a single administration of the test supplement. Urine samples were collected for 48 h. The plasma soy isoflavone concentration peaked 8 h (Cmax) after soy isoflavone supplementation. Cmax increased in a dose-dependent manner for up to 9 tablets. The Cmax values after ingestion of 9, 15, and 21 tablets were not different. However, urinary excretion of soy isoflavones increased in a dose-dependent manner for all doses. Biochemical and endocrinological markers in plasma were within normal ranges for all doses. The results indicate that the plasma concentration of soy isoflavones becomes saturated at high doses after a single administration of a soy isoflavone supplement. We conclude that toxicity of soy isoflavones is prevented by saturation of plasma isoflavone concentrations in an overdose situation.

HMF is an insoluble, high-molecular-weight fraction from the digestion product of soy protein isolate by a microbial protease. HMF effectively suppressed both colon and liver carcinogenesis that were induced by deoxycholate in rats. This effect can be explained on the assumption that HMF intake interrupts intestinal absorption and mucosal contact of deoxycholate (1,2). In this presentation we provide evidence that a resistant protein--an indigestible peptide residue derived from HMF--captured bile acids in the intestine and stimulated their fecal excretion. Feeding of HMF decreased serum bile acids concomitant with the increase of their fecal excretion in rats. There was good correlation between fecal content of nitrogen and bile acids. Amino acid analysis demonstrated that there were few free amino acids in feces, and fecal amino acids content determined after acid hydrolysis could explain most of the fecal nitrogen. These results indicated the existence of indigestible peptide, which we called a resistant protein, in the feces. However, attempts to extract the resistant protein from feces with several solvents failed, and most of the resistant proteins were insoluble even in buffer containing 8 mol urea/L. Therefore, we obtained peptide fragments by partial acid hydrolysis and sequenced the fragments. The results clearly showed that the resistant proteins were derived from regions of soybean [beta]-conglycinin rich in hydrophobic amino acids. The binding assay suggested that the resistant protein adsorbed bile acids by hydrophobic interaction. Finally, we prepared a residual fraction of HMF after in vitro peptic-pancreatic digestion of HMF. The residual fraction of HMF had a higher capacity to bind bile acids and could excrete more bile acids into feces than HMF did. The amino acid composition of the residual fraction of HMF was quite similar to that of feces of rats fed HMF.

Endogenous estrogens influence the growth, differentiation, and functioning of many tissues. They are mainly produced in the ovaries and testes. The physiological effects of these steroids are mediated by the estrogen receptors (ER[alpha], ER[beta]). When an estrogenic compound binds to the estrogen receptor, the estrogen receptor undergoes a conformational change, allowing the receptor to bind with high affinity to chromatin and to modulate the transcription of target genes. Human 17/[beta]-hydroxysteroid dehydrogenase type 1 (17[beta]-HSD1) plays an essential role in the formation of hormone steroids. This enzyme converts, for example, estrone to 17[beta]-estradiol, the most active estrogen, in the presence of the cofactor NADPH. 17[beta]-Estradiol is involved in the genesis and development of certain human breast cancers. Interest is growing in the use of dietary phytoestrogens (e.g., soy isoflavones) as potential alternatives to estrogen in hormone replacement therapy. Recently, a high intake of dietary phytoestrogens was associated with a lower incidence of breast cancer in women. Soy isoflavones are similar to endogenous estrogens and they can bind to ER[alpha] and ER[beta]. Estrogen receptors have been crystallized among other ligands with estradiol, raloxifene, or genistein ligands. All these ligands are bound with hydrogen bonds to the same three amino acids. In the case of 17[beta]-HSD1, hydrogen bonds to three amino acid residues are also essential for the optimal ligand binding. Genistein is completely buried within the hydrophobic core of the protein similar to estradiol and also binds similarly to the active site. In studying and designing the ER[alpha] and ER[beta] ligands, it is important to recognize the 17[beta]-HSD enzyme binding sites and possible interactions. The crystal structures of these three proteins (ER[alpha], ER[beta], and 17[beta]-HSD1) have opened new possibilities for structure-based analysis of dietary estrogens and drug discovery. Receptor-enzyme interactions by the Sprout de novo ligand design program will be presented.

Naturally occurring estrogen-like molecules in plants (phytoestrogens), present via soy, in animal diets exert many biological responses evoked by physiological estrogens. This study characterized the effects of dietary phytoestrogens on the expression of body, prostate, and adipose tissue weights as well as arterial blood pressure, plasma insulin and leptin, hypothalamic neuropeptide Y, and uncoupling protein-1 levels in brown adipose fat in adult Long-Evans rats. Using different experimental protocols animals, we fed rats either a phytoestrogen-rich diet (Phyto-600, containing 600 [micro]g phytoestrogens/g diet), which yielded similar plasma phytoestrogen levels to that of a human Asian diet, or a phytoestrogen low (Phyto-free) diet, which yielded similar plasma phytoestrogen levels to that of a human Western diet. Animals fed the Phyto-600 diet displayed significantly decreased arterial blood pressure and body, adipose tissue (males and females), and prostate weights compared with animals fed the Phyto-free diet. Plasma insulin levels in males fed the Phyto free diet were significantly increased compared with males fed the Phyto-600 diet whereas the opposite was found in females. No sex difference was found in plasma leptin levels. Plasma leptin levels in males and females fed the Phyto-free diet were significantly increased compared with males and females fed the Phyto-600 diet. Levels of hypothalamus neuropeptide Y (a molecule that stimulates feeding behavior) were significantly higher in Phyto-600 males than in Phyto-free males, suggesting a reason for significantly increased food and water intake in these animals. Furthermore, when we examined uncoupling protein-1 (a molecule that is associated with heat production) in brown adipose fat, males fed the Phyto-600 diet displayed significantly higher (twofold) values than Phyto-free rats. These data suggest that consumption of phytoestrogens via a soy diet can significantly alter the measured parameters as reported in this study. [Supported by National Science Foundation grant IBN-9507972 and Brigham Young University Graduate Dean's Fellowship in Neuroscience.].

Soy protein favorably alters serum lipids and lipoproteins in hypercholesterolemic individuals, thereby reducing cardiovascular disease risk. Our primary purpose was to determine the effect of soy protein (40 g/d) on circulating lipids, lipoproteins, and coagulation and fibrinolytic factors in normocholesterolemic and mildly hypercholesterolemic perimenopausal women. We also determined the contribution of coagulation, fibrinolytic, and other factors (e.g., body size and composition; serum estrogens, ferritin, and iron; dietary intake) to lipid profiles. Subjects were randomly assigned to treatment with isoflavone-rich soy (n = 24), isoflavone-poor soy (n = 24), or whey control (n = 21) protein. We measured circulating lipids and lipoproteins at baseline, week 12, and week 24 and coagulation and fibrinolytic factors at baseline and week 24. Coagulation and fibrinolytic factors were not adversely affected by treatment, as often occurs with estrogen or hormone replacement therapy. Treatment did not alter lipid profiles in mildly hypercholesterolemic subjects (n = 30) or in all subjects combined. Time had a significant (P < 0.001) effect on serum total cholesterol, triacylglycerol, LDL cholesterol, and HDL cholesterol concentrations. We could not attribute changes over time to various factors but at baseline accounted for 57% of the variability in HDL cholesterol (P < 0.0001) and for 50% in the total-HDL cholesterol ratio (P < 0.0001). Dietary vitamin E and percent energy from fat had positive effects whereas plasma plasminogen activator inhibitor-l, fibrinogen, body weight, and serum ferritin had negative effects on HDL and total-HDL cholesterol ratio. Isoflavone-rich or isoflavone-poor soy protein had no effect on lipid profiles or coagulation and fibrinolytic factors, whereas the effect of time suggested that the hormonal milieu during the menopausal transition may have overridden any detectable treatment effect on lipids. The relationship between coagulation factors and serum lipids should be further examined as indexes of cardiovascular disease risk in midlife women (1).

Soybean protein and its globulin fractions (11S and 7S) were suggested to decrease blood cholesterol and triglyceride concentrations. However, the unfavorable phytate is always bound to soy protein isolate during its preparation. This study examined the effects on plasma lipids in rats of soybean globulins from which phytate was mostly removed. Preparation of phytate-reduced soy protein isolate (SPI), [beta]-conglycinin, and glycinin was carried out by the novel method using phytase. Male Wistar young (age 6 wk) rats were fed high-cholesterol diets containing 20% casein, SPI, [beta]-conglycinin, or glycinin for 10 d. Blood samples were taken for analyses of plasma glucose, triglyceride, and total cholesterol at day 10. Liver and epididymal and perirenal fat pads were excised and weighed. The plasma triglyceride level was significantly lower in the SPI, 13-conglycinin, and glycinin groups than in the casein group and was lowest in the 13-conglycinin group. Plasma total cholesterol levels of the SPI, [beta]-conglycinin, and glycinin groups were significantly lower than that of the casein group. Relative liver weights of SPI, [beta]-conglycinin, and glycinin groups were significantly lower than that of the casein group. These results indicate that phytate-reduced [beta]-conglycinin may not only lower plasma total cholesterol level but also plasma triglyceride level and that its effects may be to decrease the fat content of liver.

Soy protein with isoflavones has been suggested for lipid lowering in men and women with high cholesterol, in part because of the beneficial effects of pharmaceutical estrogens. However, it is not known whether soy supplements with phytoestrogens enhance the effects of a low-fat diet or whether women on hormone replacement therapy (HRT+, n = 6) show the same benefits from dietary change as unmedicated postmenopausal women (HRT-, n = 12) and age-matched men (n = 15). We conducted a controlled feeding study that included a Step I diet (3 wk), a low-fat diet with 25 g soy protein and 90 mg isoflavones per day (6 wk), and a matched control diet with 25 g casein per day (6 wk). Subjects were their own control in this crossover design. As expected, HRT+ women exhibited significantly higher HDL cholesterol than did HRT- women and men (62, 54, and 48 mg/dL [1.61, 1.40, and 1.24 mmol/L, respectively]; P = 0.04) at study entry; therefore all analyses used pretreatment level as the covariate. In HRT- women and men only, the low-fat diets were associated with significant decreases in LDL cholesterol (average change = -30 [+ or -] 5 mg/dL [-0.78 [+ or -] 0.13 mmol/L]), total cholesterol (-42 [+ or -] 7 mg/dL [-1.09 [+ or -] 0.18 mmol/L]), and HDL cholesterol (-8 [+ or -] 3 mg/dL [-0.21 [+ or -] 0.08 retool/L]) relative to pretreatment values. These effects were not enhanced by soy, and there was no significant effect of protein type for any variable. In contrast, HRT+ women showed no significant changes in LDL, total, or HDL cholesterol during any of the dietary interventions. It is interesting that HRT is associated with preservation of HDL levels even in the presence of low dietary fat intake. This may illustrate a mechanism for the cardioprotective effects of HRT. However, the lack of LDL response in HRT+ subjects may be a result of selection bias by prescribers of HRT: they represent a group with particularly difficult to treat hypercholesterolemia and are good candidates for hormone therapy.