Recovery Of Isoflavones Research Articles

Improved Food-Processing Techniques to Reduce Isoflavones in Soy-Based Foodstuffs.

Soy is a growing protein source; however, the isoflavones it contains are of concern, as they exhibit estrogenic activities whose toxicological limits might be exceeded. Reducing their concentrations to safe levels while preserving nutritional quality in soy foodstuffs is therefore a matter of public health. The main objective of this paper is to develop at pilot scale a process for isoflavones' extraction from soybeans, and to show its feasibility and efficiency. The study was conducted by first optimizing the previously obtained laboratory treatment key factors. These data were then transposed to the pilot level. Finally, the process was adjusted to technical constraints which appeared at pilot scale: the mandatory use of drenching and the exploration of granulometry analysis. The involved steps were validated by monitoring the genistein and daidzein content variations through statistical analysis of the data of an ELISA and a Folin-Ciocalteu assay. Additionally, isoflavones' recovery from treatment waters for their valorisation and the water cleaning by means of filtration, centrifugation and resin adsorption were carried out. The results showed that the most successful pilot treatment developed involved soybean dehulling, drenching, washing and drying and almost halved isoflavones while preserving the main nutritional characteristics. A combination of techniques led to almost complete recovery of isoflavones from process waters.

An eco-friendly approach to enhance the extraction and recovery efficiency of isoflavones from kudzu roots and soy molasses wastes using ultrasound-assisted extraction with natural deep eutectic solvents (NADES)

The extraction efficiency, recovery, quantification, and stability of isoflavones (daidzein, genistein and puerarin) from the wastes /by-products of kudzu roots and soy molasses were evaluated by using natural deep eutectic solvents (NADESs) coupled with ultrasound-assisted extraction. The NADES extraction was compared with the conventional Soxhlet extraction technique, and the extracted isoflavones were quantified using HPLC-UV/Vis. Results showed extractability of puerarin to be significantly increased (p < 0.0001) in proportion to the increased water concentration of NADES (9.7 ± 0.2, 12.6 ± 0.2, and 14.8 ± 0.3 mg/g in NADESs containing 10%, 20%, and 30% water, respectively). Despite having less isoflavone content, kudzu roots and soy molasses extracts obtained using NADES exhibited higher antioxidant activity when compared with methanolic extracts. Further, NADES extracts also markedly reduced the degradation degree of isoflavones. The results of this investigation support the use of green extraction techniques to obtain value-added components like isoflavones from wastes/by-products like that of kudzu roots and soy molasses, which are envisaged to find future potential applications in the food and pharmaceutical industries.

Enzymatic pretreatment in the extraction process of soybean to improve protein and isoflavone recovery and to favor aglycone formation

This research aimed to evaluate the pretreatment of soybean with the carbohydrase multi-enzyme complex “Viscozyme L”, during the extraction process; in order to improve the recovery of proteins and isoflavones in soybase, and reduce the loss of these compounds through the okara residue. Three concentrations of enzyme were studied at 50 °C, along with an experiment carried out without enzyme addition (control experiment). The results were also evaluated in relation to standard soybase processing. In comparison to the standard and control processes, the enzymatic pretreatment reduced up to 85% the total amount of okara residue. Due to the action of the multi-enzyme complex, protein and total isoflavone recovery increased from 42% to 83% and from 59% to 93%, respectively. The application of Viscozyme L also favored the conversion of conjugated forms of isoflavone to aglycone in the soybase, representing up to 50% of the total isoflavones. The enzymatic pretreatment of soybean with carbohydrase improved the nutritional quality of the soybase, while at the same time reducing residue generation; showing that the proposed food process can be considered environmentally friendly method.

Fermentation of soybean meal improves isoflavone metabolism after soy biscuit consumption by adults

Urinary excretion of isoflavones and their metabolites after the consumption of soybean meal (SBM) and fermented soybean meal (FSBM) biscuits was investigated in humans in a randomized double-blinded crossover clinical trial. Aglycones were the most abundant metabolites in urine in both treatments. After FSBM consumption, total urinary excretion of aglycones and metabolites was 54% higher when compared to SBM biscuits. All metabolites were more rapidly excreted after FSBM biscuit consumption when compared to SBM biscuits. Urinary recovery of isoflavones was higher after FSBM biscuit consumption, reaching 67%. Women excreted higher amounts of colonic metabolites and lower amounts of aglycones than men. The prevalence of O-demethylangolensin-producer, equol-producer and nonproducer metabotypes was 56%, 11% and 36% after SBM consumption and 72%, 11% and 17% after FSBM consumption, respectively. Together, our findings indicate that fermentation improves isoflavone bioavailability and possibly reduces the impact of gut microbiota on the metabolism of isoflavones. © 2020 Society of Chemical Industry.

Recovery of Antimicrobials and Bioaccessible Isoflavones and Phenolics from Soybean (Glycine max) Meal by Aqueous Extraction.

Soybeans display strategic potential in food security as a source of protein and functional bioactives for human consumption. Polyphenols and other bioactive compounds can be recovered after an aqueous extraction from soybean meal, a byproduct of soy oil refining. The objective of the present study was to compile and quantify compounds from soybean oil refinery by-products, providing information about valuable bioactive phytochemicals, their bioaccessibility and potential bioactivities. Genistin, daidzin, glycitin and malonylgenistin were the predominant isoflavones, and the overall bioaccessibility of their glycosidic forms was of nearly 75%. Sixteen phenolics were identified and caffeic acid, 5-caffeoylquinic chlorogenic acid and hesperidin were the most predominant. Approximately 30% of gallic acid, syringic acid, vanillic acid and myricetin were released and the antioxidant capacity of aqueous extract was enhanced after simulated in vitro gastro intestinal digestion. The ability of aqueous soybean meal extract to inhibit lipid peroxidation was higher than natural and synthetic food antioxidants. Antimicrobial activity against several foodborne pathogens and antitumoral activity towards human glioblastoma cell line were also observed, but the aqueous extract showed no cytotoxicity to healthy murine cells. Compounds derived from the aqueous soybean meal extract have the potential to be used as health promoting agents.

An Optimized Method for the Quantification of Isoflavones in Dry Soy Extract Containing Products

Isoflavones, a class of flavonoids with estrogen-like activity, are present in the human diet mainly derived from soy-based foods. Soy isoflavones (namely genistein, daidzein, and glycitein and their derivatives) and concentrates have primarily been investigated for their ability to prevent sex hormone-dependent cancers and cardiovascular problems. Several studies demonstrated that many of the biological effects of soy are due to isoflavones. The aim of our study was to develop a simple method for the determination of isoflavones in dry soy extract containing products. Since biological effects of soy isoflavones depend upon aglycone forms, extracts were hydrolyzed to gain isoflavone aglycones. To optimize the extraction and hydrolysis of isoflavones, the effect of HCl concentration (1.5–6 N), hydrolysis time (25–210 min), and temperature (30–100 °C) on total isoflavone aglycone content was studied using an RP-HPLC-DAD method. By mathematical fitting and optimization methods, optimum hydrolysis conditions for maximizing the quantification of isoflavones were determined (t = 96 min, cHCl = 4.9 N, T = 80 °C). The experimentally verified model has a good coefficient of R 2 (0.9928), and the recovery of isoflavones was 97.81–102.76 %. The developed method allows reliable and maximum determination of isoflavones in dry soy extract containing products. Based on this method, six soy extract containing products were analyzed. Our experiments revealed significant variations in the isoflavone content and composition. These results highlight the necessity to clearly specify the isoflavone composition of soy products.

Exergy driven process synthesis for isoflavone recovery from okara

Isoflavones, found in soybeans and other members of the fabaceae family, are considered bioactive components of high economic value. An opportunity would be to separate isoflavones from okara, the by-product of the soymilk and tofu production. Such a process would not only valorise that side-stream but also, and maybe more importantly, reduce the waste of high quality bioactive compounds. Extraction is an important part during the recovery of isoflavones from okara and was conceptually designed in this work. Due to environmental constraints, ethanol and water were the only solvents considered in this work for extraction of isoflavones. Different process scenarios were established and assessed by solvent footprinting, energy use, and exergy analysis. Simulation of the various process scenarios showed that distillation and the loss of ethanol in the spent okara represent the largest inefficiencies regarding exergy waste and energy usage. Furthermore, even though the use of ethanol leads to a higher recovery, water is in most cases the preferred solvent due to the high exergetic cost of losing some ethanol in the spent okara and during distillation.

Recovery of Isoflavones from the Soy Whey Wastewater Using Two-Stage Batch Foam Fractionation

The feasibility of recovering bioactive substances without surface activity from their aqueous solutions was studied using biosurfactants as collectors. For recovering the isoflavones from the soy whey wastewater, a two-stage batch foam fractionation was developed using the soy proteins as collectors. The results showed that when the conditions of the first stage were temperature of 60 °C, pH of 5.0, volumetric air flow rate 80 of mL/min, and loading liquid volume of 400 mL, a high isoflavones enrichment ratio of 4.05 was obtained and the residual solution could be used as the feeding solution of the second stage. When the conditions of the second stage were temperature of 25 °C, pH of 5.0, and volumetric air flow rate of 150 mL/min, the soy proteins concentration in the residual solution could decrease to 622 mg/L. By reusing the foamate of the second stage as the feeding solution of the first stage, the total isoflavones recovery percentage reached 87.72%.

Polymorphisms in genes involved in the metabolism and transport of soy isoflavones affect the urinary metabolite profile in premenopausal women following consumption of a commercial soy supplement as a single bolus dose

Genetic variation in relevant enzymes and transporters may contribute to discordant observations concerning health outcomes of dietary isoflavone consumption, so we examined the association of the UGT1A1*28 promoter polymorphism and of other SNPs with isoflavone metabolites in urine. We genotyped prospectively for polymorphisms in UGT1A1 (UGT1A1*28), LPH (666G>A), CBG (1368T>A), ABCG2 (421C>A), and ABCC2 (1249G>A) to select 100 women (18-50 years) to receive a commercial soy supplement as a single dose and collect all urine over 24 h for analysis by RP-HPLC. We observed large differences in isoflavone recovery (mean 39%, eightfold variation) and metabolites. Glucuronides were the major metabolites (72% of total). UGT1A1*28 was associated only with percentage of glycitein as sulphate (positive; p = 0.046), but excluding five participants with both minor alleles of CBG and ABCG2 uncovered additional associations with percentage of glycitein as glucuronide (negative; p = 0.028), combined isoflavones as sulphate (positive; p = 0.035) and sulphate-to-glucuronide ratio for combined isoflavones (positive; p = 0.036). CBG1368T>A, ABCG2 421C>A, and ABCC2 1249G>A were also associated with differences in isoflavone metabolites in urine. Genetic variation in UGT1A1, CBG, ABCG2, and ABCC2 influences isoflavone metabolism so may affect benefits of dietary consumption.

Method development and optimization for isoflavone quantification in dry soy extract containing products

Isoflavones are secondary metabolites belonging to flavonoids found in a variety of plants, especially soy (Glycine max L.). Due to the estrogen-like activity of these compounds several soy products are on the market and applied to relieve menopausal symptoms. The aim of our study was to develop a simple method for the determination of isoflavones in soy extract containing products. The main isoflavones of soybean are genistin, glycitin, daidzin and their respective acetyl, malonyl and aglycone forms. Quantitative measurement of each individual isoflavone is difficult because some of the reference compounds are not widely commercially available [1]. Keeping in mind that the biological effects of soy isoflavones depend upon aglycone form, hydrolysis of glycosides is a practical method for the quantitative determination of total isoflavones and assessment of biological value of extracts [2]. The objective of our investigation was to establish an acid hydrolysis method for the quantitative HPLC determination of total isoflavones including daidzein, genistein and glycitein. To optimize the extraction and hydrolysis of isoflavones, the effect of HCl concentration (1.5–6 N), hydrolysis time (25–210min) and temperature (30–100°C) on total isoflavone aglycone content was studied using an RP-HPLC-DAD method. By mathematical fitting and optimization methods optimum hydrolysis conditions for maximizing the quantification of isoflavones were determined (t=94min, cHCl=5.09 N, T=80°C). The experimentally verified model has a good coefficient of R2 (0,9928) and the recovery of isoflavones was 97.81–102.76%. The developed method allows a reliable and maximum determination of isoflavones in dry soy extract containing products.

Simulation, scale-up and economics of adsorption and membrane based processes for isoflavones recovery

Laboratory scale adsorption and membrane based processes for the recovery of isoflavones from red clover flowers were reported earlier ( Xu et al., 2005, 2006). However, studies on scale-up and simulation for commercialization for both the methods have not been studied. In present study the economic feasibility of isoflavones recovery based on these two methods was investigated with a commercial bio-process simulator (SuperPro Designer ®). Laboratory scale separation data from previously published work was used in the simulations. Preliminary simulation studies confirmed the requirement of effluent streams recycling in order to reduce the production costs. Different flowsheets incorporating solvent recovery and recycle operations were designed and simulated to compare the economics of operation. Modified adsorption and membrane based processes incorporating recycling of waste streams were found to be economically more attractive than that of respective standard processes reported in the literature. The membrane process with solvent recycling had the lowest production cost of US$618/kg for isoflavones supplement. The adsorption-recycle process was found to be more expensive at US$1116/kg, while it had a higher content of isoflavones.

Recovery of isoflavones from red clover flowers by a membrane-based process

Isoflavones in red clover flowers were extracted and recovered using a new process that mainly comprised of ethanol extraction, membrane processing, micelle formation and drying. To obtain maximum isoflavone extraction an ethanol concentration between 40% and 50% in water was found to be optimal. The extracted isoflavones were processed by ultrafiltration for preliminary purification, and then concentrated by reverse osmosis. As ethanol was removed by evaporation, micelles were formed in the reverse osmosis retentate, which was dried to yield an isoflavone-enriched product. This product contained about 9% isoflavone highlighting its potential use as a direct nutraceutical supplement. Industrial relevance Isoflavones present in agricultural biomass are utilized as source of functional food supplements. Extraction and refining of isoflavones involves several steps and the process is very energy intensive. This work reports an interesting approach for extraction and refining of isoflavones by developing a new process, which is energy efficient and gives a final product, which contains sufficiently high amounts of isoflavones for consumer applications.

Bioavailability of isoflavone phytoestrogens in postmenopausal women consuming soya milk fermented with probiotic bifidobacteria

We investigated the effects of consuming an isoflavone aglycone-enriched soya milk containing viable bifidobacteria on urinary isoflavone excretion and percentage recovery. Sixteen postmenopausal women were randomly divided into two groups to consume either fermented or non-fermented soya milk. Each group participated in a double-blind, crossover study with three 14 d supplementation periods, separated by a 14 d washout. Subjects ingested three daily dosages of isoflavone via the soya milk and collected four 24 h pooled urine specimens per supplementation period. Soya milks were prepared with soya protein isolate and soya germ, followed by fermentation with bifidobacteria. Isoflavone levels were quantified using HPLC. Non-fermented soya milks at 20, 40 and 80 mg isoflavone/200 ml contained 10 %, 9 % and 7 % aglycone, respectively, with their fermented counterparts containing 69 %, 57 % and 36 % aglycone (P<0.001). A trend to a greater percentage urinary recovery of daidzein and glycitein was observed among women consuming fermented soya milk at a dosage of 40 mg isoflavone (P=0.13). A distinct linear dose response for the fermented soya milk group (R2=0.9993) compared with the non-fermented group (R2=0.8865) suggested less interindividual variation in isoflavone absorption. However, total urinary isoflavone excretion was similar for both groups (P>0.05), with urinary isoflavone recovery at approximately 31 %. Increasing the isoflavone dosage correlated positively with its urinary excretion, but urinary percentage recovery of isoflavone was inversely related to dosage level. Hence, a modest dosage ranging from 20 to 30 mg/d may provide the most bioavailable source of isoflavone, regardless of whether it is via an aglycone-rich fermented soya milk or a glucoside-rich soya milk.

Effects of Solvent Polarity and Acidity on the Extraction Efficiency of Isoflavones from Soybeans (Glycine max)

Soy isoflavones have been correlated with beneficial health effects. The predominant chemical forms of isoflavones present may affect their biological activities. Choosing the solvent system that can accurately quantify the amounts of individual isoflavones present in these products is paramount. Our objectives were to compare frequently used solvent systems and to evaluate the effects of polarity and acidity on the recovery of isoflavones from soybeans. Isoflavones were extracted from pulverized Manokin soybeans using six solvent systems, which are the combinations of three polarity levels (83% acetonitrile, 80% methanol, and 58% acetonitrile) and two acidity levels (nonacidified and acidified). The pulverized soybean was stirred for 2 h in each solvent system before filtration and concentration using rotary evaporation. The extract was resuspended in 16% acetonitrile and analyzed by high-performance liquid chromatography. Recoveries of pure standards were evaluated with all solvent systems. Solvents with a higher polarity extracted a significantly higher amount of total isoflavones. For individual isoflavones, 58% acetonitrile (highest polarity) extracted either the highest amounts or no less than other solvents, while 83% acetonitrile (lowest polarity) extracted either the lowest amounts or no more than other solvents except for the aglycone form. Acidification significantly reduced the recovery of the malonylglucoside form and the total isoflavones. The recovery study revealed that acidification favored the chemical transformations of isoflavones during the extraction. Among the six solvent systems examined, 58% acetonitrile aqueous solution without acid was the best for extraction of isoflavones from soybeans.

Effects of Fermentation Temperature on the Content and Composition of Isoflavones and β-Glucosidase Activity in Sufu

Sufu is a popular fermented tofu product in China. The low quality of sufu produced in the hot summer is a big problem in sufu manufacture, so we prepared sufu at two different temperatures, 26 degrees C as normal and 32 degrees C as high temperature, and the effects of temperature on isoflavones and beta-glucosidase activity were investigated. Fermentation temperature did not cause significant differences in the recovery of isoflavones, but resulted in a different redistribution of isoflavone isomers in sufu. Sufu fermented at 26 degrees C was richer in isoflavone aglycones than at 32 degrees C; the enrichment of isoflavone aglycones might have the advantage of enhancing the physiological function. No 6''-O-malonyl-glucosides were detected in sufu fermented at 26 degrees C, whereas some 6''-O-malonyl-glucosides were found at 32 degrees C. A fermentation temperature of 26 degrees C benefited the beta-glucosidase production by fungi, which contributed to valid conversion from beta-glucosides to aglycones. It was also found that beta-glucosidase converted beta-glucosides more effectively than 6''-O-malonyl-glucosides and 6''-O-acetyl-glucosides into aglycones.

Enhancement of tofu isoflavone recovery by pretreatment of soy milk with koji enzyme extract.

Isoflavones are novel nutraceutical constituents of soybeans, but considerable amounts are lost in the whey during conventional tofu manufacturing. In this study, in a small-scale process, 2 mL of koji enzyme extract (soybean koji/deionized water, 1/3, w/v) was combined with 600 mL of soy milk, and 30 mL aliquots were incubated at 35 degrees C for 0, 30, 60, 120, and 300 min, for enzyme pretreatment. After each treatment time, soy milk was heated to 85 degrees C, CaSO4 was added to aggregate protein, and the mixture was centrifuged to separate the solids (tofu) from the whey. The tofu yield and moisture contents from soy milk treated for 30 or 60 min were higher than those from soy milk treated for 0 (control), 120, or 300 min. The protein content of freeze-dried tofu varied in a limited range, and native PAGE and SDS-PAGE patterns revealed slight quantitative and qualitative variations among products. Soy milk daidzein and genistein contents increased while daidzin and genistin contents decreased as the time of enzyme pretreatment of the soy milk increased. After 30 min of pretreatment, daidzin, genistin, daidzein, and genistein contents recovered in tofu products were higher than those of the control. In a pilot-scale process, aliquots (3 L) of soy milk were enzyme-treated for 30 min, aggregated with CaSO4, and hydraulically pressed to remove the whey. As in pretreatments, soy milk daidzein and genistein contents increased while daidzin and genistin contents decreased. In a comparison of the control and enzyme-treated tofu products, the total recoveries of daidzin, genistin, daidzein, and genistein in the tofu products increased from 54.9% to 64.2%. When the tofu products were subjected to a sensory panel test, both products were judged acceptable.

Changes in isoflavone contents and composition of sufu (fermented tofu) during manufacturing

Sufu is a fermented tofu product which is popular in China. Manufacturing procedures of sufu significantly affected the isoflavone contents and composition of sufu. The recovery of isoflavones in sufu from soybean was 16.9%. The loss of isoflavones was mainly attributed to the preparation of tofu and salting of pehtze (fresh bean curd overgrown with mould mycelia). The isoflavone composition was altered during sufu processing. The levels of aglycones increased, while the corresponding levels of glucosides decreased. “Former fermentation” corresponded to the fastest period of isoflavone conversion. The isoflavones in sufu, in the form of aglycones and in the form of glucosides, accounted for 99.7% and 0.3% of the total, respectively. The changes in the isoflavone composition were significantly related to the activity of β-glucosidase during sufu fermentation, which was affected by the NaCl content.

A membrane-based process for recovering isoflavones from a waste stream of soy processing

Based on a lab-scale aqueous process for making skimmed soymilk, a membrane intensive scheme was developed to recover isoflavones as a separate product from the waste stream while soymilk was prepared as the main product. It was shown that, despite the protein binding, most extracted isoflavones permeated through the selected ultrafiltration membranes. Therefore, instead of solvents or adsorbents, a combination of ultrafiltration and diafiltration was used to separate isoflavones from the proteins and other bean components, and isoflavones were then concentrated by reverse osmosis. A yield of approximately 50% of the total amount of isoflavones in soybeans was eventually obtained in the reverse osmosis retentate, which could be dried to make an isoflavone supplement. With minor modifications, the process was readily adaptable to isoflavone recovery from soy protein processing with defatted soy flour.

Extracting and purifying isoflavones from defatted soybean flakes using superheated water at elevated pressures

This paper focusses mainly on improving superheated water extraction conditions using the simulation optimization approach known as “steepest ascent”. The study examines the effect of operating conditions on the amount of five isoflavones extracted from defatted soybean flakes (DSF) using superheated water extraction (SWE), stirred solvent extraction, Soxhlet methanol extraction, and pressurized water extraction (PWE). Amberlite XAD16-HP adsorbent was used to purify five isoflavones obtained by reverse phase solid extraction. HPLC analysis was conducted to quantify these five isoflavones as proportions of the amount of total isoflavones (TIF). The steepest ascent design for SWE revealed that the optimum condition was 110 °C and 641 psig (4520 kPa) over 2.3 h of extraction. Under these condition, 3937 TIF μg/g DSF was produced. Solid phase purification, by Amberlite XAD16-HP absorption, yielded a solid with 37.0 purity and 49.0 de-sorption recovery of total isoflavones.

Extraction time for soybean isoflavone determination

Studies were carried out on the optimization of the extraction procedures for isoflavones determination in soybean by HPLC. Isoflavones were extracted from 100 mg samples of full fat soybean flour "Kinako" (milled toasted grains). Recovery of average total isoflavones was higher when extraction was performed with agitation(217.2 mg/100g), than without agitation(191.0 mg/100g). Isoflavone extraction without agitation for 1,4, 20 and 24 hours, were equally efficient for total and individual compounds. These results suggested that an efficient isoflavone recovery could be achieved with extraction for one hour with agitation.