Effect of extraction methods and UHT treatment conditions on the level of isoflavones during soymilk manufacture

The objective of this study was to compare the effect of hot and cold grinding as well as the effect of direct and indirect ultra high temperature (UHT) treatment conditions on the level of isoflavones during the manufacture of soymilk. Soymilks were manufactured from dehulled soybeans by hot grinding or cold grinding processes. After inactivation of lipoxygenase at 85 °C, the resulting slurries were decanted and supernatants were held at 120 °C for 80 s to inactivate the trypsin inhibitor. The decanted soya bases were cooled and subjected to different temperature/time regimes by direct and indirect UHT treatments. Samples were drawn at different points in the processing operation and a reversed phase high performance liquid chromatography method was used to determine the concentration of isoflavones. Results showed that hot grinding caused a higher extraction of isoflavones into the soymilk than the cold-grinding process. However, direct or indirect heating in the UHT process did not significantly influence the concentration of isoflavones.     

Isoflavone Profiles and Kinetic Changes during Ultra‐High Temperature Processing of Soymilk

Isoflavone profile is greatly affected by heating process. However, kinetic analyses of isoflavone conversion and degradation using a continuous industry processing method have never been characterized. In this study, Proto soybean was soaked and blanched at 80 °C for 2 min and then processed into soymilk, which underwent UHT (ultra‐high temperature) at 135 to 150 °C for 10 to 50 s with a pilot plant‐scale Microthermics processor. The isoflavone profile was determined at different time/temperature combinations. The results showed that all isoflavone forms exhibited distinct changing patterns over time. In the soymilk under UHT conditions, the degradation (disappearance) of malonyldaizin and malonylgenistin exhibited first‐order kinetics with activation energies of 59 and 84 kj/mole, respectively. At all UHT temperatures, malonylgenistin showed higher rate constants than malonyldaidzin. However, malonylglycitin changed irregularly under these UHT temperatures. The increase of genistin, daidzin, glycitein and acetlydaidzin during heating demonstrated zero‐order kinetics and the rate constants increased with temperature except for the conditions of 145 to 150 °C for 50 s. Overall, genistein series exhibited higher stability than daidzein series. Under all UHT conditions, total isoflavone decreased from 12% to 24%.     

Content composition and antioxidant activity of isoflavones in commercial and homemade soymilk and tofu

BACKGROUND: Isoflavones, found in soymilk and tofu, are one of the phytochemicals in soy‐based products that may promote good health. Homemade tofu and various homemade soymilk samples were made using different soaking, grinding, and cooking methods. The homemade samples were compared to commercial tofu and soymilk for total isoflavone content and composition as well as their antioxidant capacity. All samples were freeze‐dried and extracted with a 58% acetonitrile solution which was subsequently used to determine the isoflavone content by reverse‐phase high‐performance liquid chromatography. The antioxidant activity of extracts was determined using a modified 2,2‐azino‐bis‐(3‐ethylbenzthiazoline‐6‐sulfonic acid) method and total antioxidant capacity was reported as ascorbic acid equivalents. RESULTS: The total isoflavone, aglycone, and antioxidant levels were significantly higher in homemade soymilk and tofu (1571 µg) than in commercial samples. Homemade soymilk made by the extended boiling method yielded the highest total isoflavone (2567 µg) and glucoside (1525 µg) content. A strong positive correlation was observed between the total isoflavone, aglycone conjugates, and genistein series concentration and antioxidant capacity of soymilk. CONCLUSION: Increased moist heating time yielded the highest concentration of total isoflavones as well as aglycone conjugates and the genistein series. Increasing the duration of boiling can increase the isoflavone content of both homemade and commercial soymilk and tofu. Copyright © 2007 Society of Chemical Industry     

不同品种大豆异黄酮组分及体外抗氧化活性分析

为研究不同品种大豆异黄酮组分和含量以及抗氧化活性,测定了50个特色大豆品种的百粒重、色泽（L*、a*、b*）,通过高效液相色谱法（HPLC）测定异黄酮组分,评估不同品种大豆甲醇提取物的DPPH、ABTS自由基清除能力,并用相关性分析、主成分分析及聚类分析法对样品进行分析。结果表明:大豆的百粒重范围为7.05~47.46 g。不同种皮色大豆L*、a*和b*呈现显著差异（P<0.05）。HPLC分析结果发现糖苷型异黄酮含量是大豆中主要的异黄酮组分,占总异黄酮含量的90%以上。其中染料木苷含量最高。安豆115品种的大豆异黄酮的总含量最高,为2500.78 μg/g;靖江丝瓜青品种的大豆异黄酮含量最低,为888.86 μg/g。安豆115品种的DPPH和ABTS自由基清除能力较强,分别为（16.11±0.25）和（8.12±0.04）μmol V_C/g。糖苷型异黄酮含量与DPPH和ABTS自由基清除能力有较高的线性相关性（R2分别为0.903和0.867）。主成分分析表明6项指标可用2个主成分来表示（累计贡献率达66.3%）。聚类分析将50个品种的大豆分为4类:第一类中黄豆黄素和黄豆黄苷含量较高;第二类中大豆苷、大豆苷元、染料木苷和染料木素含量较高;第三类和第四类中其大豆异黄酮含量基本处于中等水平和较低含量。综上所述,不同大豆品种的异黄酮组成、含量和抗氧化能力存在较大差异,可为其进一步综合开发利用提供一定理论基础。     

Chromatographic fingerprints and quantitative analysis of isoflavones in Tofu-type soybeans

Tofu-type soybeans can differ from conventional soy varieties in seed size, seed composition, flavour and nutrition. Using HPLC coupled with ESI-MS and PDA detection, a total of 19 isoflavones were detected and identified from Tofu-type soybean seeds, more than previously reported forms from many other soybeans. An HPLC/UV fingerprint study was performed for qualitative evaluation, which enabled the isoflavone profile of Tofu-type soybeans to be characterised and differentiated from other vegetative soybeans by similarity comparison. To meet the frequent quantitative application for isoflavone contents, a simple, precise and reliable method using HCl hydrolysis during sample extraction and LC/UV for the detection was developed and validated to quantitate total isoflavones in soybeans, and then applied to determine the total isoflavone contents of different Tofu-type soy varieties grown in different field locations over two growing seasons. A range of total isoflavone contents for Tofu-type soybeans was established to confirm the use of this analytical approach for quality control applications.     

INFLUENCE OF HEAT TREATMENT AND GRAIN GERMINATION ON THE ISOFLAVONE PROFILE OF SOY MILK

Soy foods have attracted much attention for their possible effects on human health because of their phytochemical content, mainly isoflavones. Twelve forms of isoflavones have been identified in soybeans and soy products. Technological processes like heat treatment or fermentation can change the isoflavone profile of soy products. In this study, the effects of heat treatment and grain germination on isoflavone profile of soy milk prepared from BRS 213 soybean variety were evaluated. Isoflavone forms were separated and quantified by reversed‐phase high‐performance liquid chromatography. Heat treatment by autoclaving for 5 min did not affect the total isoflavone content, but led to a 90% decrease in malonyl forms, whereas β‐glycosides increased by 70%. Autoclaving for 15 min led to a 20% loss of total isoflavones. Germination for 3 days did not change the total isoflavone content, but changed the distribution profile (15% increase of malonyl forms and 30% decrease of β‐glycosides). Germination for 7 days increased the bioavailability of aglycone isoflavones in soy milk by seven times.     

Isoflavone profiles,phenol content,and antioxidant activity of soybean seeds as influenced by cultivar and growing location in Ohio

Isoflavone levels and isoflavone chemical composition in soybeans vary between planting locations although the exact factors which control isoflavone biosynthesis are unclear. We compared levels of 12 isoflavones in soybean seeds of six cultivars grown in four different locations in Ohio in 2002 as determined by high‐performance liquid chromatography. Antioxidant activity contained in plant‐based foods can improve food oxidative stability and phenolics and isoflavones have proven active in food systems. Radical scavenging activity was assessed using the 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical. Total phenolics (TPCs) were determined by using Folin–Ciocalteu reagent. Total isoflavones (TIs) varied five‐fold (1573–7710 nmol g^?1) between seeds from the various location–cultivar combinations. One location (Wooster, Ohio) produced seeds containing half the isoflavones as the other locations tested apparently due to poor growing conditions. The cultivars could be divided into two groups based on TI, one having approximately 50% more isoflavones. Surprisingly, across the entire data set, with increasing TI, the proportion of isoflavones accounted for by the daidzein family increased due primarily to malonyl daidzin. DPPH scavenging did not differ significantly by location or cultivar (P > 0.05) and did not correlate with TPC or TI. Profiling soybean isoflavones could help elucidate how isoflavone biosynthesis is regulated and lead to better disease resistance of soybean crops and soy foods with greater health benefits. Copyright © 2007 Society of Chemical Industry     

Concentration of phytoestrogens in soybeans and soybean products in Korea

Concentrations of total isoflavones in soybean varieties of different origins and soybean products were determined by high performance liquid chromatography. Wild soybeans showed the trend to have more isoflavone content than cultivars or landraces, showing mean value of 1528, 1265 and 1249 mg kg⁻¹, respectively. There were no meaningful differences in the ratio of genistein and daidzein among the modern variety groups. Korean varieties including South Korea and North Korea seemed to have total isoflavone content more than Chinese varieties, although samples of Chinese varieties were limited. In a dried basis of soyfoods, total isoflavone contents were in order of soybean sprout, chungkukjang, soymilk, soybean curd, denjang, kochujang, respectively. Five soybean foods contributed 96% of total genistein and daidzein intake in Korea: soybean sprout, soybean curd, denjang, chungkukjang and soybean seed itself. According to a Korean National Survey Report and our data, the average Korean daily intake of isoflavone from soybean foods can be estimated as 21 mg day⁻¹. © 2000 Society of Chemical Industry     

发酵对黑豆营养及活性成分的影响研究

本试验采用百粒重为12克的黑豆为原料，对黑豆、蒸煮黑豆及发酵黑豆的营养及活性成分进行了哆究，黑豆经发酵处理后糖苷型大豆异黄酮在β-葡萄糖苷酶的作用下，可转化为游离型大豆异黄酮，使游离型大豆异黄酮含量明显提高。蛋白质、多肽、可溶性糖、游离氨基酸的含量也有所提高。     

Change in isoflavone concentration of soybean (Glycine max L.) seeds at different growth stages

This study was conducted to determine the fluctuating levels of isoflavone in soybean (Glycine max L.) seeds of six varieties at different development stages. The average concentration of total isoflavone in the fully matured seed was 669 µg g^?1, whereas those in the seeds of the R5 (beginning), R6 (green full‐sized) and R7 (onset of physiological maturity) stages were 145, 296 and 611 µg g^?1, respectively. Sowonkong variety was highest in total isoflavone content of the fully matured and the R7 seed among all the varieties. In all six varieties, total isoflavone levels at the R7 stage were higher than those of R5 and R6 stages. Total isoflavone concentrations rapidly increased as growth stages transitioned from R5 to R7, as indicated by a positive correlation (r = 0.80***) between the total isoflavone and growth stages. Overall, total isoflavone levels were similar between R7 and the full maturity stages. Of the various isoflavone derivatives, acetylglucoside and aglycon were not significantly correlated with total isoflavone concentrations. However, glucoside (r = 0.97***), malonylglucoside (r = 0.99***), TGIN (genistein + genistin + malonylgenistin + acetylgenistin, r = 0.93***) and TDIN (daidzein + daidzin + malonyldaidzin + acetyldaidzin, r = 0.99***) were very highly correlated with total isoflavone concentrations. As a result, they constituted most of the total isoflavones in the soybean seeds at R5, R6, R7, and full maturity stages. Copyright © 2007 Society of Chemical Industry     

Research progress in tofu processing: From raw materials to processing conditions

As a traditional soybean product with good quality and a healthy food with many functional components, tofu is increasingly consumed in people's daily life. Traditional tofu processing consists of numerous steps, including the soaking and grinding of soybean seeds, heating of the soybean slurry, filtering, and addition of coagulants, and others. The properties of soybean seeds, processing scale, soaking and heating conditions, type and concentration of coagulant, and other factors collectively impact the processing steps and the final tofu quality. The generation of whole soybean tofu with more nutritive value comparing with traditional tofu has been successfully reported by several studies. As one of the most important functional component, isoflavones and their presence in tofu are also influenced by the above-mentioned factors, which influence the nutritive value of tofu. Research investigating the influence of tofu processing conditions on the quality and isoflavone profiles of tofu are the subject of this review. Issues that should be further studied to investigate the influence of processing conditions on the quality and nutritive value of tofu are also introduced.     

净作和套作条件下大豆籽粒异黄酮积累规律比较

以我国西南地区3个常用大豆品种为材料,比较净作和套作条件对大豆籽粒发育及后熟过程中异黄酮积累规律的影响。结果表明：籽粒成熟期按总异黄酮积累速率可分为初始积累期和快速积累期;受田间小气候影响,套作条件下大豆较晚进入快速积累期;田间采收时,贡选1号品种净作总异黄酮含量极显著高于套作,南豆16和贡秋豆3号净作与套作之间差异不显著。3个品种籽粒后熟期套作条件下总异黄酮的积累量均极显著高于净作,总异黄酮含量套作均显著高于净作;后熟期不同品种异黄酮各组分及总异黄酮的积累量均存在显著差异。本研究发现净套作对3个大豆品种的异黄酮积累动态及总含量均有显著影响,品种间籽粒异黄酮的积累对种植条件的反应不同。相对净作而言,后熟过程更利于套作大豆籽粒异黄酮的进一步积累。     

Bioavailability of Isoflavone Metabolites After Korean Fermented Soybean Paste (Doenjang) Ingestion in Estrogen‐Deficient Rats

Doenjang (DJ), a fermented soybean product used in soups, stews, and sauces, contains high quality proteins, fats, vitamins, minerals, and other functional ingredients, including isoflavones and saponins. This study investigated whether DJ improves the bioavailability of isoflavones compared to boiled soybean (BS) in sham‐operated or ovariectomized (OVX) rats. We also examined the effects of ovariectomy on the differences in bioavailability of isoflavones. BS and DJ were administered in sham‐operated and OVX rats, and blood samples were collected. Twenty‐six isoflavone‐derived metabolites were identified. Pharmacokinetic analysis revealed that T_1/2 values of the individual isoflavone metabolites were most different in sham and OVX rats, even after the same sample treatment; however, T_max values were significant different in a few metabolites such as daidzein 4′‐glucuronide, daidzein 4′‐sulfate, 2‐(4‐hydroxyphenyl)propionic acid, and benzoic acid. For most of the individual metabolites, C_max was higher in both sham and OVX rats administered BS than those administered DJ. The AUC was generally lower in OVX rats than in sham rats. The AUC of daidzein and genistein in BS‐fed sham rats was approximately 1.7‐fold higher than those administered DJ, whereas glycitein was detected only in the DJ group. No significant differences in AUC of daidzein and genistein were observed between BS and DJ administration in OVX rats, although the total isoflavone content of DJ was lower; thus, DJ‐mediated isoflavone bioavailability was more effective in OVX rats. Similar tendencies were observed for phase II and gut‐mediated metabolites. These results suggested that DJ enhanced isoflavone bioavailability under estrogen deficiency, even when the total isoflavone content was decreased by fermentation.     

大豆异黄酮分离特性及其检测的初步探讨

对大豆异黄酮在大豆加工过程中的变化和大豆异黄酮组分的分离检测等方面的研究现状进行了阐述 ,并针对大豆异黄酮在大豆加工与分离中组分间的转化提出利用和控制的可能途径和方法     

Tracking isoflavones: From soybean to soy flour,soy protein isolates to functional soy bread

Soybean seeds with three different levels (low, intermediate and high) of isoflavones were processed to soy flour and soy protein isolates (SPIs) and developed into functional soy breads. The effect of factors involved in all steps of the process was investigated by tracking the composition and concentration of native forms of isoflavones. The total isoflavone contents were 8033.3, 10570.1 and 15169.0 nmol/g DM (dry matter) in the three soybeans; 13201.5, 20034.4 and 26014.3 nmol/g DM in defatted soy flours; 9113.2, 13274.6 and 17918.3 nmol/g DM in the SPI; 2782.7, 4081.4 and 5590.3 nmol/g DM in soy breads, respectively. The bread making processes did not affect the total isoflavone content, but changed glucosides/acetylglucosides to aglycones. Malonylglucosides were stable prior to baking but degraded to acetylglucosides and further to glucosides during baking. Our results provide critical information for the production of functional soy breads that contain varying amounts of soy isoflavones.     

Innovative Soaking and Grinding Methods and Cooking Affect the Retention of Isoflavones,Antioxidant and Antiproliferative Properties in Soymilk Prepared from Black Soybean

This study's objective was to characterize the effect of traditional and 3 newly devised (soaking+grinding) methods combined with cooking on the content and composition of phenolic substances, antioxidant, and antiproliferative properties of soymilk prepared from black soybean. Phenolic substances and antioxidant profile were characterized and antiproliferation of prostate cancer DU145 cells was conducted using a cell culture assay. Results indicated Grinding Method 4 produced significantly (P < 0.05) higher total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), and total isoflavone content in both raw and cooked black soymilk as compared to Method 1. Cooking soymilk reduced 23% to 38% of total phenolic substances. Raw black soymilk produced by Method 4 displayed the highest antioxidant capability, which was determined using ORAC, FRAP, and DPPH assays, and a higher antiprostate cell proliferation ability. Cooking only slightly reduced the potency to inhibit DU145 prostate cancer cells as IC₅₀ value was increased from the average of about 4.0 mg/mL of raw soymilk extracts to 5.5 mg/mL of cooked soymilk extracts of all grinding methods. Overall, total isoflavone content was the only component that was negatively correlated with IC₅₀ value (r = –0.93, P < 0.05) which indicates the ability to inhibit prostate cancer cell is associated with the increase in total isoflavone content, not with any other phenolic substances or antioxidant properties.     

Comparison of nutritional components (isoflavone,protein, oil,and fatty acid) and antioxidant properties at the growth stage of different parts of soybean [<Emphasis Type="Italic">Glycine max</Emphasis> (L.) Merrill]

This is the first study to investigate antioxidant capacities of isoflavones prepared using microwave-assisted hydrolysis method from different parts (seeds, leaves, leafstalks, pods, stems and roots) of soybean at growth stages. In addition, the fluctuations in the isoflavone, protein, fatty acid, and oil contents in R6-R8 (R6: beginning; R7: beginning maturity; R8: full maturity) seeds were confirmed. The R7 seeds exhibited the most predominant contents of isoflavones (1218.1±7.3 μg/g) in the following order: daidzein (48%)>genistein (35%)>glycitein (17%). The second highest isoflavone content was found in the leaves (1052.1±10.4 μg/g), followed by R8 seeds>roots>R6 seeds>leafstalks> pods; the stems exhibited the lowest isoflavone content (57.2±1.7 μg/g). Interestingly, daidzein showed the highest individual isoflavone content with remarkable variations (57.2-766.8 μg/g), representing 46-100% of the total isoflavone content. R8 exhibited higher protein, fatty acid, and oil contents than R6 or R7. Moreover, the antioxidant capacities against two radicals in different parts of soybean plant showed considerable differences depending upon the isoflavone content. Our results suggested that soybean leaves and seeds might be useful materials for functional foods.     

发芽大豆异黄酮提取工艺条件的研究

首先采用大豆为原料制备发芽大豆,以异黄酮得率为指标,确定最佳发芽时间为96h.继而对发芽大豆中异黄酮的提取工艺条件进行研究,在单因素实验结果的基础上,进行正交实验确定最佳提取工艺条件,其结果为超声功率400W、超声波时间40min、乙醇浓度80%、回流时间1.5h,在此条件下异黄酮得率为3.774mg/g.     

Characterization of Isoflavones in Membrane-processed Soy Protein Concentrate

ABSTRACT: Aqueous soy flour solutions (5% w/w) were treated for 3 h at 45 to 50 °C with Crystalzyme®, without enzyme at 45 to 50 °C, or blanched at 80 °C. Solutions were ultra‐ and dia‐filtered to produce membrane soy concentrates (MSC). Total isoflavones of MSCs were 3.02 mg/g, 3.12 mg/g, or 3.42 mg/g, respectively, on a dry weight basis. Membrane processing contributed to approximately 18, 15, or 8% decreases, respectively, in total isoflavones of MSCs compared with soy flour (3.71mg/g). This represents at least an 82% recovery. There was no significant difference in total isoflavone content in any MSC, however the profile of isoflavones as glucosides or aglycones changed with treatment conditions.     

Comparison of isoflavones composition in seed,embryo, cotyledon and seed coat of cooked-with-rice and vegetable soybean (Glycine max L.) varieties

In order to study the content and composition of isoflavones retained in soybean seed component, obtained each component part the embryo, cotyledon and seed coat tissues of nine different soybean varieties were analyzed for 12 isoflavones using high performance liquid chromatography with photo diode array detector (HPLC-PDA) and were compared to each other. A total average concentration of isoflavone was 2887 μg g⁻¹ in embryo, 575 μg g⁻¹ in whole seed, 325 μg g⁻¹ in cotyledon, and 33 μg g⁻¹ in seed coat. With respect to each tissue of soybean varieties, isoflavone content was highest in Geomjeongkong 2 embryo (5701 μg g⁻¹), Geomjeongolkong whole seed (1321 μg g⁻¹), Heugcheongkong cotyledon (951 μg g⁻¹), and Keunolkong seed coat (56 μg g⁻¹). Isoflavone was least present in Keunolkong embryo (341 μg g⁻¹), Hwaeomputkong whole seed (175 μg g⁻¹), Seonheukkong cotyledon (81 μg g⁻¹), and Seoklyangputkong seed coat (5 μg g⁻¹). Overall, embryo and seed coat of all nine varieties contained isoflavones at the highest and lowest level, respectively. Isoflavones accumulated in the order of malonylglycoside, glycoside, acetylglycoside, and aglycon, among which malonylglycoside was the most abundant form ranging from 66% to 79% of the total isoflavone content in all three tissues. The embryo of cooked-with-rice soybean with black seed coat appears to be the best source of isoflavone.