Characterization of phenolic substances and antioxidant properties of food soybeans grown in the North Dakota-Minnesota region

Phenolic profiles and antioxidant properties of a total of 30 soybean samples, including 27 grown in the North Dakota-Minnesota region and three soybeans from the other regions, were investigated. The total phenolic content (TPC), total flavonoids content (TFC), phenolic acids, flavonols, anthocyanins, and isoflavones were quantified. Antioxidant properties of soybean extracts were assessed using 2-diphenyl-1-picryhydrazyl free radical scavenging activity (DPPH), ferric reducing antioxidant power (FRAP), and oxygen radical absorbance capacity (ORAC) methods. Results showed that black soybean cultivars possessed significantly higher TPC, TFC, DPPH, FRAP, and ORAC values than all yellow soybean cultivars. However, black soybean cultivars did not exhibit significantly higher individual phenolic contents (except for anthocyanins), such as phenolic acids and isoflavones, than the yellow soybean cultivars. The isoflavone profiles of North Dakota soybean cultivars were similar to those of South Dakota, but average values of total isoflavone (TI) contents were higher than soybeans grown in the other states and Korea and Japan according to the U.S. Department of Agriculture-Iowa State University Database on the isoflavone contents of foods. Correlation assays showed that TPC, TI, total phenolic acids, daidzin, genistin, malonyldaidzin, daidzein, genistein, and trans-cinnamic acid significantly ( r = 0.73, 0.62, 0.49, 0.68, 0.59, 0.59, 0.56, 0.47, and 0.76, respectively, p < 0.0001) correlated with ORAC values of yellow soybeans. Both isoflavones and phenolic acids contributed to the ORAC values of yellow soybeans. These data suggest that some selected soybean cultivars may be used as high-quality food-grade soybeans for providing high phenolic phytochemicals and antioxidant activities.     

Isoflavone composition and antioxidant capacity of modified-lipoxygenase soybeans grown in Maryland

Maryland-grown soybean lines modified for low lipoxygenase-1 (LOX-1) content and a traditional nonmodified cultivar were analyzed for fatty acid composition, total phenolic content (TPC), isoflavone composition, relative DPPH? scavenging capacity (RDSC), and hydroxyl radical scavenging capacity (HOSC). Soybean lines included black, brown, and yellow soybeans. TPC of all soybean lines ranged from 2.84 to 4.74 mg gallic acid equiv (GAE)/g flour. Total isoflavones were between 2.78 and 8.66 μmol/g flour. RDSC of all lines was between 0.48 and 14.62 μmol Trolox equiv (TE)/g flour, and HOSC ranged from 53.57 to 135.52 μmol TE/g flour. Some modified-LOX genotypes demonstrated antioxidant capacity and/or isoflavone content similar to or higher than those of the nonmodified cultivar (P < 0.05). Black soybeans demonstrated higher TPC and RDSC than most yellow soybean lines, although they did not have higher isoflavone content. The results demonstrate that modification of the LOX trait did not necessarily alter the antioxidant capacity or chemical composition of the experimental soybean lines when compared with a nonmodified cultivar. These soybean lines may be studied further for nutraceutical properties and use in functional foods.     

Correlations of oil and protein with isoflavone concentration in soybean [Glycine max (L.) Merr.

Twelve isoflavones were detected by high-performance liquid chromatography in seeds of 17 soybean [Glycine max (L.) Merrill] cultivars grown at three locations. 6' '-O-Malonyldaidzin and 6' '-O-malonylgenistin together constituted 71-81% of total isoflavones, which ranged in concentration from 2038 to 9514 microg/g and averaged 5644 microg/g across locations and cultivars. The total as well as several individual isoflavones had a moderate negative correlation with oil across locations and cultivars. Six cultivars had a moderate or strong negative correlation of total isoflavones with oil. Five cultivars had a moderate or strong positive correlation of total isoflavones with protein. These results suggest that judicious selection of germplasm for soybean breeding may facilitate development of soybean lines with desirable isoflavone concentrations.     

Comparison of isoflavone concentrations in soybean (Glycine max (L.) Merrill) sprouts grown under two different light conditions

We determined and compared the composition and content of isoflavones in the cotyledon, hypocotyl, and root of 17 soybean sprout varieties grown under dark and light conditions. The total average isoflavone concentrations in 17 soybean sprout varieties were 2167 microg g(-1) (green sprout) and 2538 microg g(-1) (yellow sprout) in cotyledons, 1169 microg g(-1) (green sprout) and 1132 microg g(-1) (yellow sprout) in hypocotyls, and 2399 microg g(-1) (green sprout) and 2852 microg g(-1) (yellow sprout) in roots. There were no significant differences in total isoflavone concentrations between the green and yellow sprouts. However, significant differences in total isoflavone amounts were observed among the three organs, with roots exhibiting the highest total isoflavone concentrations followed by cotyledons and hypocotyls. Total daidzin concentrations of green (775 microg g(-1)) and yellow (897 microg g(-1)) sprouts increased to more than 4 times that in seeds (187 microg g(-1)). Yellow sprouts contained the highest (1122 microg g(-1)) total genistin concentrations, and green (155 microg g(-1)) and yellow (155 microg g(-1)) sprouts had more total glycitin concentrations than seeds. In cotyledons of green and yellow sprouts, genistin, daidzen, and glycitin constituted more than 67%, more than 28%, and less than 4% of the total isoflavone contents, respectively. In hypocotyls, total daidzin represented more than 45% of the total isoflavones, and total glycitin was higher than in cotyledons and roots. Malonylglycoside concentrations were highest in cotyledons, whereas glycoside concentrations were highest in hypocotyls and roots. The high accumulation of isoflavones in roots is consistent with isoflavones serving as signal molecules in the induction of microbial genes involved in soybean (Glycine max) nodulation.     

Isoflavone characterization and antioxidant activity of ohio soybeans

Seventeen Ohio soybeans were screened for isoflavone content and antioxidant activity. Isoflavone content was determined by C(18) reversed phase high-performance liquid chromatography coupled with a photodiode array detector. Antioxidant activities of soybean extracts were measured using 2,2-diphenyl-1-picryl-hydrazyl (DPPH) free radical and photochemiluminescence (PCL) methods. The highest and lowest total isoflavone contents were 11.75 and 4.20 micromol/g soy, respectively, while the average was 7.12 micromol/g soy. Antioxidant activities of soybean extracts ranged from 7.51 to 12.18 micromol butylated hydroxytoluene (BHT) equivalent/g soy using the DPPH method. Lipid and water soluble antioxidant activities of soybean extracts ranged from 2.40 to 4.44 micromol Trolox equivalent/g soy and from 174.24 to 430.86 micromol ascorbic acid equivalent/g soy, respectively, using the PCL method.     

Effects of the environment, cultivar, maturity, and preservation method on red clover isoflavone concentration

Red clover (Trifolium pratense L.) contains isoflavones that are of interest because of their benefits for human health as well as their adverse effects on the fertility of farm animals. A series of field experiments was conducted in Sainte-Anne-de-Bellevue, QC, Canada, to determine the effects of the environment, cultivar, plant maturity, plant part, and preservation method on the concentration of the two predominant isoflavones in red clover, formononetin and biochanin A. In a multi-year, multisite trial, the total isoflavone concentration in 10 cultivars ranged between 8,923 and 12,753 microg g(-1) of DM averaged across sites, harvests, and years. Despite strong environmental effects, the cultivar "Start" consistently had the lowest isoflavone concentrations, with few differences observed among other cultivars. Across stages of maturity, leaves were found to have the highest isoflavone concentration followed by stems and inflorescences (11,970, 4,896, and 3,297 microg g(-1) of DM, respectively). Changes in isoflavone concentrations with increasing maturity varied depending on the plant part. Overall, highest isoflavone concentrations were found in leaves and stems during the vegetative stages, with the formononetin concentration declining until plants initiated flowering, especially in stems, with concentrations then stabilized in both parts. Upon initiation, inflorescences contained similar isoflavone concentrations than leaves, but concentrations decreased rapidly during flower development to fall even below those observed in stems. Inflorescences then had isoflavone concentrations that were as much as 11 times lower than leaves. Fresh herbage contained higher formononetin and total isoflavone concentrations than did silage and hay (14,464, 12,200, and 11,604 microg g(-1) of DM, respectively). The isoflavone concentration in field-grown red clover is thus high but can be affected by a range of agronomic factors.     

Total phenolics, phenolic acids, isoflavones, and anthocyanins and antioxidant properties of yellow and black soybeans as affected by thermal processing

The effects of boiling and steaming processes on the phenolic components and antioxidant activities of whole yellow (with yellow seed coat and yellow cotyledon) and black (with black seed coat and green cotyledon) soybeans were investigated. As compared to the raw soybeans, all processing methods caused significant (p < 0.05) decreases in total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), monomeric anthocyanin content (MAC), DPPH free radical scavenging activity (DPPH), ferric reducing antioxidant power (FRAP), and oxygen radical absorbing capacity (ORAC) in black soybeans. Pressure steaming caused significant (p < 0.05) increases in TPC, CTC, DPPH, FRAP, and ORAC in yellow soybeans. The steaming resulted in a greater retention of TPC, DPPH, FRAP, and ORAC values in both yellow and black soybeans as compared to the boiling treatments. To further investigate the effect of processing on phenolic compounds and elucidate the contribution of these compounds to changes of antioxidant activities, phenolic acids, isoflavones, and anthocyanins were quantitatively determined by HPLC. The pressure steaming treatments caused significant (p < 0.05) increases in gallic acid and 2,3,4-trihydroxybenzoic acid, whereas all treatments caused significant (p < 0.05) decreases in two predominant phenolic acids (chlorogenic acid and trans-cinnamic acid), and total phenolic acids for both yellow and black soybeans. All thermal processing caused significant (p < 0.05) increases in aglucones and beta-glucosides of isoflavones, but caused significant (p < 0.05) decreases in malonylglucosides of isoflavones for both yellow and black soybeans. All thermal processing caused significant (p < 0.05) decreases of cyanidin-3-glucoside and peonidin-3-glucoside in black soybeans. Significant correlations existed between selected phenolic compositions, isoflavone and anthocyanin contents, and antioxidant properties of cooked soybeans.     

Variation in isoflavone of soybean cultivars with location and storage duration

Fifteen soybean [Glycine max (L.) Merrill] cultivars were grown in Seoul, Suwon, and Kyongsan, Korea, in 1998, 1999, and 2000, and their isoflavone contents were assessed. After harvest, the beans were stored for 3 years at room temperature. Soybean isoflavones were analyzed using high-performance liquid chromatography (HPLC) within each crop year and after storage. Total isoflavone contents ranged from 188.4 to 685.6 mg 100 g(-1) in 1998, from 218.8 to 948.9 mg 100 g(-1) in 1999, and from 293.1 to 483.0 mg 100 g(-1) in 2000. The year x variety, and year x location x variety interactions were significantly different in 1998, the year x location, year x variety, and year x location x variety interactions were significantly different in 1999, and the year x variety interaction was significantly different in 2000 for total and individual isoflavone contents. Total isoflavone contents of soybeans stored for 1 year were only slightly higher than those of soybeans stored for 2 or 3 years. However, the concentrations of individual isoflavones, especially 6' '-O-malonyldaidzin and 6' '-O-malonylgenistin, decreased markedly in soybeans stored for 2 or 3 years. These data suggest that it may be feasible to improve soybean cultivars with higher antioxidative substances.     

Evaluation of soyasaponin, isoflavone, protein, lipid, and free sugar accumulation in developing soybean seeds

A combination of analytical techniques was used to examine and quantify seed compositional components such as protein, lipid, free sugars, isoflavones, and soyasaponins during soybean development and maturation in two Korean soybean cultivars. Protein accumulation was rapid during reproductive stages, while lipid content was only relatively moderately increased. The major carbohydrate saccarides sucrose and stachyose constantly increased during the reproductive stage. Previously published results suggest that the free sugar and lipid content reached their maximal concentrations at a relatively early stage of seed development and remain constant in comparison to other chemical components. The malonylglucosides were the predominant isoflavone form followed by the glucosides, acetyl glucosides, and aglycone forms. As soybean seed matures, total soyasaponin concentration was constantly decreased until the R8 stage. Soyasaponin beta(g) was the major soyasaponin in DDMP-conjugated group B soyasaponins, followed by the non-DDMP counterpart soyasaponin I and soyasaponin A1. The ratio of total isoflavone to total soyasaponin in the developing soybean increased from 0.06 to 1.31. Protein, lipid, and free sugar contents in the developing soybean seeds showed significant positive correlations with conjugated isoflavones and total isoflavone concentration, while the lipid contents showed a negative correlation with the isoflavone aglycone. Protein, lipid, and free sugar contents showed a negative correlation with total group A and B soyasaponins and total soyasaponins; however, only the soyasaponin A content was significantly negatively correlated with free sugar content. Total soyasaponin content was negatively correlated with isoflavone content (r = -0.828 at p < 0.01).     

Group B saponins in soy products in the U.S. Department of Agriculture--Iowa State University isoflavone database and their comparison with isoflavone contents

Isoflavones in soy protein foods are thought to contribute to the cholesterol-lowering effect observed when these products are fed to humans. The group B saponins are another ethanol-soluble phytochemical fraction associated with soy proteins and isoflavones and have also been associated with cholesterol-lowering abilities. We measured the group B soyasaponin concentrations in a variety of soy foods and ingredients in the U.S. Department of AgricultureIowa State University Isoflavone Database. We compared the isoflavone and soy saponin concentrations and distributions in intact soybeans, soy ingredients, and retail soy foods. Group B saponins occur in six predominant forms. There appears to be no correlation between saponin and isoflavone concentrations in intact soybeans ranging from 5 to 11 mumol isoflavones/g soybean and from 2 to 6 mumol saponin/g soybean. Depending upon the type of processing, soy ingredients have quite different saponins/isoflavones as compared to mature soybeans. In soy foods, the saponin:isoflavone ration ranges from 1:1 to 2:5, whereas in soy protein isolates, the ratio is approximately 5:3. Ethanol-washed ingredients have very low saponins and isoflavones. These very different distributions of saponins and isoflavones in soy products may affect how we view the outcome of feeding trials examining a variety of protective effects associated with soy consumption.     

Isoflavone content and its potential contribution to the antihypertensive activity in soybean Angiotensin I converting enzyme inhibitory peptides

A soybean angiotensin I converting enzyme (ACE) inhibitory peptide fraction was reported to have antihypertensive activity in a rat study. The purpose of the present study was to examine if the presence of isoflavones in the soybean ACE inhibitory peptide fraction may contribute to the blood-pressure-lowering property. The isoflavone concentration in soybean samples was analyzed on a C 18 reverse-phase column using a two-step gradient solvent system. Producing soybean hydrolysate led to a nearly 40% loss of isoflavones compared with the original soybean flour, but the isoflavone composition did not change and the dominant isoflavone chemicals remained as 6'-O-malonylgenistin and 6'-O-malonyldaidzin. Ion exchange chromatography affected significantly both the content and the composition of the isoflavones. The dominant isoflavones in the ion-exchanged fraction were aglycones and nonacylated isoflavones, accounting for 95.8% of the total amount of 987.7 microg/g. It was calculated that the isoflavone content in the soybean ACE inhibitory peptide fraction was 25 times less than the minimal effective isoflavone dose reported. In vitro study also showed that adding isoflavones into both soybean flour hydrolysate and soybean ACE inhibitory peptide samples to a concentration of as high as 31.5% (w/w) did not affect ACE inhibitory activity (IC 50 values). The findings along with previously published results indicated that the contribution of isoflavones in soybean ACE inhibitory peptide fraction to the blood-pressure-lowering property in a short-term feeding study might be negligible.     

Isoflavone conjugates are underestimated in tissues using enzymatic hydrolysis

Many health effects of soy foods are attributed to isoflavones. Isoflavones upon absorption present as free form, glucuronide, and sulfate conjugates in blood, urine, and bile. Little is known about the molecular forms and the relative concentrations of soy isoflavones in target organs. Acid hydrolysis or enzymatic hydrolysis (glucuronidases and sulfatases) was used to study isoflavone contents in the heart, brain, epididymis, fat, lung, testis, liver, pituitary gland, prostate gland, mammary glands, uterus, and kidney from rats fed diets made with soy protein isolate. The heart had the lowest isoflavone contents (undetectable), and the kidney had the highest (1.8 +/- 0.6 nmol/g total genistein; 3.0 +/- 1.1 nmol/g total daidzein). Acid hydrolysis released 20-60% more aglycon in tissues than enzymatic digestion (p < 0.05), and both hydrolysis methods gave the same level of isoflavones in serum. Approximately 28-44% of the total isoflavone content within the liver was unconjugated aglycon, and the remainder was conjugated mainly as glucuronide. The subcellular distribution of total isoflavones was 55-60% cytosolic and 13-16% in each of the nuclear, mitochondrial, and microsomal fractions. These results demonstrated that (1) soy isoflavones distribute in a wide variety of tissues as aglycon and conjugates and (2) the concentrations of isoflavone aglycons, which are thought to be the bioactive molecules, are in the 0.2-0.25 nmol/g range, far below the concentrations required for most in vitro effects of genistein or daidzein.     

Nutritional aspects of second generation soy foods

Samples of 15 second generation soy-based products (n = 3), commercially available, were analyzed for their protein and isoflavone contents and in vitro antioxidant activity, by means of the Folin-Ciocalteu reducing ability, DPPH radical scavenging capacity, and oxygen radical absorbance capacity. Isoflavone identification and quantification were performed by high-performance liquid chromatography. Products containing soy and/or soy-based ingredients represent important sources of protein in addition to the low fat amounts. However, a large variation in isoflavone content and in vitro antioxidant capacity was observed. The isoflavone content varied from 2.4 to 18.1 mg/100 g (FW), and soy kibe and soy sausage presented the highest amounts. Chocolate had the highest antioxidant capacity, but this fact was probably associated with the addition of cocoa liquor, a well-known source of polyphenolics. This study showed that the soy-based foods do not present a significant content of isoflavones when compared with the grain, and their in vitro antioxidant capacity is not related with these compounds but rather to the presence of other phenolics and synthetic antioxidants, such as sodium erythorbate. However, they may represent alternative sources and provide soy protein, isoflavones, and vegetable fat for those who are not ready to eat traditional soy foods.     

Inhibition of reactive nitrogen species effects in vitro and in vivo by isoflavones and soy-based food extracts

Recent studies have shown that soy isoflavone inhibits inducible nitric oxide (NO) synthase activities and is reported to have peroxynitrite scavenging ability. Consequently, we investigated whether isoflavones (daidzein and genistein) and extracts from soy-based products (miso, soymilk, tofu, soy sprout, black soybean, soybean, and yuba) would inhibit the reactive nitrogen species (RNS) effect in vitro and in vivo. In the in vitro experiments [including the protection of cellular DNA from peroxynitrite or sodium nitroprusside damage, an inhibitory effect on nitric oxide production from lipopolysaccharide (LPS)-induced RAW 264.7 cells, and nitric oxide scavenging ability], extracts from soy-based foods showed a potent antioxidant activity and an inhibiting effect on RNS activity. These effects were correlated with total isoflavone content. In the in vivo experiments, rats were given isoflavones (4.0 mg/kg bw) or soy-based product extracts (1.0 g/kg bw) orally for 1 week and were injected with vehicle H(2)O (1 mL/kg bw) or LPS (10 mg/kg bw) on the day 7. Twelve hours after treatment, the rats were killed, and blood serum was collected for analysis. The intraperitoneal administration of LPS resulted in an increase in serum nitrite, nitrate, and nitrotyrosine concentrations. These are stable metabolite end products of nitric oxide, to 4-, 16-, and 5-fold levels, (4, 10 microM and 58 +/- 14 pmol/mL), of the placebo control, respectively. Results showed that oral administration of isoflavones and extracts from soy-based products significantly decreased serum nitrite, nitrate, and nitrotyrosine levels in LPS-induced rats. This study demonstrates that soy isoflavone supplementation may inhibit RNS-induced oxidation both in vitro and in vivo.     

Functional components in soybean cake and their effects on antioxidant activity

The antioxidant activities of four fractions of isoflavones from soybean cake were evaluated and compared with those of ISO-1 and ISO-2 fractions, five isoflavone standards, and mixtures of two or four isoflavone standards, as well as four commercial antioxidants, using DPPH, TEAC, reducing power, metal ion chelating, conjugated diene, and TBARS assays. Both malonylglucoside and glucoside fractions were isolated using preparative chromatography with Diaion HP-20 as adsorbent, whereas acetylglucoside and aglycone fractions were separated with silica gel as adsorbent. The other two fractions, ISO-1 and ISO-2, were soybean cake extracts containing 12 isoflavones for the former and a combination of 4 fractions for the latter. Both acetylglucoside and ISO-1 fractions exhibited the highest efficiency in scavenging DPPH free radicals, whereas all six fractions were effective in inhibiting conjugated diene formation. However, a low reducing power was observed for all six fractions and isoflavone standards. The aglycone fraction and genistein standard showed a pronounced increase of TEAC value and a moderate decrease of TBARs value. For chelating metal ions, both ISO-1 and ISO-2 fractions were the most efficient. Overall, the isoflavone fractions showed a better antioxidant activity than the isoflavone standards, probably caused by the presence of some other functional components such as saponin, flavonoid, and phenolic compounds in soybean cake.     

LC/UV/ESI-MS analysis of isoflavones in Edamame and Tofu soybeans

High-performance liquid chromatography coupled with ultraviolet and electrospray ionization mass spectrometry (HPLC/UV/ESI-MSD) was applied to the study of isoflavones in both Edamame and Tofu soy varieties, from which the immature fresh soybeans or the mature soybean seeds are consumed, respectively. Positive atmospheric pressure interface (API) MS and MS/MS were used to provide molecular mass information and led to the identification of a total 16 isoflavones, including three aglycones, three glycosides, two glycoside acetates, and eight glycoside malonates. The major isoflavones in soybean seeds were daidzein and genistein glycoside and their malonate conjugates. Trace levels of daidzein and genistein acetyl glycosides were found only in the mature dry soybean seeds. To facilitate quantitative analysis, acid hydrolysis during extraction of soy samples was selected to convert the various phytoestrogen conjugates into their respective isoflavone aglycones, allowing accurate quantitation of total phytoestrogens as aglycones. On the basis of HPLC combined with UV and MS detection, all three targeted soy isoflavone aglycones, daidzein, genistein and glycitein in hydrolyzed extracts were successfully quantified within 25 min with formononetin used as the internal standard. The standard curves of UV detection were fitted in the range of 14.16-29000 ng/mL for daidzein, 15.38-31500 ng/mL for genistein, and 11.72-24000 ng/mL for glycitein. For MS detection, the standard curves were established in the range of 3.54-1812.5 ng/mL for daidzein, 3.85-1968.75 ng/mL for genistein, and 2.93-1500 ng/mL for glycitein. Good linearities (r(2) > 0.999 for UV and r(2) > 0.99 for MS) for standard curves were achieved for each isoflavone. The accuracy and precision (RSD) were within 10% for UV detection and 15% for MS detection (n = 10). Using this method, the phytoestrogen levels of total isoflavone aglycones from 30 soybean seed varieties were then evaluated for confirmation of the technique. Total isoflavones ranged across the varieties from 0.02 to 0.12% in the Edamame varieties, which are harvested while the seeds are still immature, and from 0.16 to 0.25% in Tofu varieties, harvested when the seeds are physiologically mature. While the literature has focused on the isoflavone content of soy products and processing soy, this report provides a reliable analytical technique for screening of authenticated fresh immature Edamame soybeans and Tofu soybeans.     

Determination of 15 isoflavone isomers in soy foods and supplements by high-performance liquid chromatography

Soy isoflavone is the generic name for the isoflavones found in soy. We determined the concentrations of 15 soy isoflavone species, including 3 succinyl glucosides, in 22 soy foods and isoflavone supplements by high-performance liquid chromatography (HPLC). The total isoflavone contents in 14 soy foods and 8 supplements ranged from 45 to 735 μg/g and from 1,304 to 90,224 μg/g, respectively. Higher amounts of succinyl glucosides were detected in natto, a typical fermented soy product in Japan; these ranged from 30 to 80 μg/g and comprised 4.1-10.9% of the total isoflavone content. In soy powder, 59 μg/g of succinyl glucosides were detected, equivalent to 4.6% of the total isoflavone content. These data suggest that the total isoflavone contents may be underestimated in the previous studies that have not included succinyl glucosides, especially for Bacillus subtilis -fermented soy food products.     

Antioxidant capacity of seed coat, dehulled bean, and whole black soybeans in relation to their distributions of total phenolics, phenolic acids, anthocyanins, and isoflavones

Black soybeans have been used as an excellent dietary source for disease prevention and health promotion in China for hundreds of years. However, information about the distribution of health-promoting phenolic compositions in different physical parts of black soybean and the contribution of phenolic compositions to overall antioxidant capacity is limited. To elucidate the distribution of phenolic composition and their contribution to antioxidant activities in black soybean, the total and individual phenolic profiles, and antioxidant capacities of seed coat, dehulled and whole black soybean were systematically investigated. The seed coat exhibited much higher total phenolic indexes and antioxidant activities than whole and dehulled black soybean. Dehulled black soybean possessed similar levels of total phenolic content, total flavonoid content, 2-diphenyl-1-picryhydrazyl (DPPH) radical scavenging activity, ferric reducing antioxidant power (FRAP), and oxygen radical absorbance capacity (ORAC) activities as compared to whole yellow soybean. Cyanidin-3-glucoside, petunidin-3-glucoside, and peonidin-3-glucoside were detected in the seed coat but not in dehulled black soybean and yellow soybean. Among benzoic acid detected, caffeic and chlorogenic acid were the predominant phenolic acids. Whole black soybean and dehulled black soybean exhibited similar isoflavone contents in 7- O-beta-glucosides and malonylglucosides of daidzein and genistein. The seed coat possessed significantly ( p < 0.05) lower 7- O-beta-glucosides and malonylglucosides of daidzein and genistein, acetylglycitin, and total isoflavones than whole and dehulled black soybean. The contribution of phenolics in the seed coat to the antioxidant activity of black soybean parts depends on the assay methods. When measured with the DPPH and FRAP methods, the seed coat contributed 90% of the total antioxidant capacity of black soybean. However, when measured with the ORAC method, the seed coat and dehulled portion contributed approximately equally the total antioxidant capacity of black soybeans. The information generated from this study on the distribution and content of their active components is useful for the effective use of black soybeans as an ingredient for promoting health.     

Effect of temperature, elevated carbon dioxide, and drought during seed development on the isoflavone content of dwarf soybean [Glycine max (L.) Merrill] grown in controlled environments

The effects of elevated temperature, carbon dioxide, and water stress on the isoflavone content of seed from a dwarf soybean line [Glycine max (L.) Merrill] were determined, using controlled environment chambers. Increasing the temperature from 18 degrees C during seed development to 23 degrees C decreased total isoflavone content by about 65%. A further 5 degrees C increase to 28 degrees C decreased the total isoflavone content by about 90%. Combining treatments at elevated temperature with elevated CO(2) (700 ppm) and water stress to determine the possible consequences of global climate change on soybean seed isoflavone content indicated that elevated CO(2) at elevated temperatures could partially reverse the effects of temperature on soybean seed isoflavone content. The addition of drought stress to plants grown at 23 degrees C and elevated CO(2) returned the total isoflavone levels to the control values obtained at 18 degrees C and 400 ppm CO(2). The promotive effects of drought and elevated CO(2) at 23 degrees C on the 6' '-O-malonygenistin and genistin levels were additive. The individual isoflavones often had different responses to the various growth conditions during seed maturation, modifying the proportions of the principal isoflavones. Therefore, subtle changes in certain environmental factors may change the isoflavone content of commercially grown soybean, altering the nutritional values of soy products.     

Antioxidant properties of soybean isoflavone extract and tofu in vitro and in vivo

Isoflavones in soybean were extracted in the crude form using 80% food-grade ethanol at 80 degrees C for 6 h and followed by concentration and dehydration. The soy extract contained isoflavones primarily in the forms of glucosides. In vitro antioxidant activities of the soy extract containing 20-500 ppm isoflavones were conducted using a Rancimat method. The results showed that soy isoflavone extract had strong in vitro antioxidant activity. There was a dose-dependent response for the in vitro antioxidant activity at the lower concentrations but not at the higher concentrations. In vivo antioxidant property was determined by measuring the antioxidant enzymes, superoxide dismutase, and catalase in various organs of rats that were fed with diets containing partially oxidized oil and various levels of isoflavones for up to 24 weeks. Neither short-term (8 weeks) feeding nor low isoflavone content (50 ppm) induced changes in superoxide dismutase or catalase activities in rats. Only diets containing high isoflavone contents (150 and 250 ppm) showed obvious elevated enzymatic levels in various organs. In addition, a laboratory-prepared tofu containing approximately 50 ppm isoflavones had better effects than the soy extract with the 250 ppm isoflavone group, which indicated that molecules other than isoflavones may have a synergistic effect on in vivo antioxidant enzyme inductions of tofu.