Isoflavone Extract Research Articles

Abstract 4037: Combining tyrosine kinase inhibitors and isoflavones to target metastatic activity in prostate cancer cells

Abstract Background: Efforts to impede early steps in the metastatic cascade have led to the development of promising small molecule targeting agents. The purpose of this study was to evaluate the use of Src tyrosine kinase inhibitors known to interfere with cell migration and invasion, in combination with a cytostatic soy isoflavone extract (ISFs) on metastatic activity in prostate cancer cells. Methods: PC-3ML cells (a gift from A. Fatatis, Drexel University), and LNCaP cells treated with a Src inhibitor, ISFs, or a combination, were evaluated for clonogenic cell survival, cell cycle progression, and transwell migration and invasion activity. Results: Src inhibitors and ISFs induced an accumulation of cells in the G1 and G2/M phases of the cell cycle, respectively, with little change in cell viability or survival. Invasion studies with PC-3 ML and LNCaP cells suggest that ISFs can reduce metastatic activity by 39% and 35%, respectively. In addition, ISFs enhance the effectiveness of the Src inhibitors to reduce invasiveness. This may indicate that including ISFs in treatment regimens may allow a lower dose of the targeting agent to be used to achieve optimal response and decrease toxicity. Conclusions: In vitro studies suggested that combining a Src inhibitor and ISFs resulted in greater inhibition of metastatic activity than either alone. Citation Format: Lori P. Rice, Christine Pampo, Sharon Lepler, Dietmar W. Siemann. Combining tyrosine kinase inhibitors and isoflavones to target metastatic activity in prostate cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4037. doi:10.1158/1538-7445.AM2014-4037

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.

Optimization of ionic liquid-based microwave-assisted extraction of isoflavones from Radix puerariae by response surface methodology

Objective: The aim of this work was to develop a based microwave-assisted approach for extraction of isoflavonoids from the herbal medicines.Background: Room temperature ionic liquids (RTILs) as a design green solvent are regarded as an attractive alternative or replacement to conventional volatile organic solvents. The microwave-assisted extraction has been applied widely to extract natural products. But the extracting solvents used often include organic solvents, which have strong volatility and toxicity and can cause serious pollution issues. The extraction technique that combines microwave extraction technology with ionic liquids for application in the field of extraction and separation constitutes a new approach.Method: [bmim]Br was selected as the solvent. The microwave-assisted extraction parameters, including extraction time, extraction temperature, microwave power, and the concentration of [bmim]Br had been concerned. Response surface methodology was used to develop predictive models for simulation and optimization of the extraction process. The multivariate quadratic regression equation was obtained by Central Composite Design. The response surfaces were determined by using the yields of total isoflavonoids as response values. HPLC and SEM were performed to compare the ionic liquid based microwave extraction with the traditional alcohol MAE.Results: The optimum extraction conditions were: extraction temperature of 70oC, extraction time of 8 min, microwave power of 400 W, and the concentration of [bmim]Br of 1.2 mol/L. The extraction yield of total isoflavonoids was 10.09%, which is well in close agreement with the value predicted by the model.Conclusion: Compared to conventional organic solvents, the ionic liquid based MAE could provide higher extraction yields and take much shorter extraction time.

Abstract P423: Effects of Green Tea and Soy Isoflavones on Plasma Lipids In Healthy Subjects

Introduction: Green tea and soy products are widely consumed in daily life and are considered to have beneficial effects on reducing cardiovascular risks. The flavonoids present in soya bean and tea leaves are believed to be responsible for the proposed benefits. Hypothesis: We assessed the hypothesis that epigallocatechin gallate (EGCG), the most abundant active component of green tea and soy isoflavonoids may reduce the plasma lipid levels in healthy volunteers. Methods: Healthy Chinese male subjects were given green tea extract containing EGCG 800 mg once daily for 14 days and soy isoflavones extract 80 mg once daily for 14 days with a wash-out of at least 4-weeks between the two treatments. Sitting blood pressure and fasting plasma lipids were measured at baseline, after green tea and after soy isoflavones, respectively. Results: In 18 healthy male subjects (mean age 26.6 ± 6.0 years, weight 61.7 ± 6.3 kg), consumption of green tea extract, but not soy isoflavones, was associated with significant reductions in plasma low-density lipoprotein cholesterol (LDL-C) (8.1% [95% confidence interval: -2.0, -14.3%], P<0.01) and total cholesterol (4.8 % [0.4,-10.1%], P<0.05) compared to baseline (Table 1). Reduction in LDL-C with green tea was observed in 15 out of 18 subjects and the change in LDL-C was not affected by the baseline levels. Neither product influenced the plasma high-density lipoprotein cholesterol (HDL-C) or triglyceride levels or blood pressure in this normotensive group of subjects (Table 1). Conclusion: This study showed that short term treatment with a high dose of green tea extract, which is equivalent to about 7-10 cups of green tea, reduced plasma LDL-C which may be beneficial for reducing cardiovascular risk.

Isoflavone extraction from okara using water as extractant

We here report on the use of water as a ‘green’ extraction solvent for the isolation of isoflavones from okara, a by-product of soymilk production. At a low liquid-to-solid ratio of 20 to 1 and 20°C, 47% of the isoflavones that can be extracted with 70% aqueous ethanol were extracted. The malonyl-glucosides were fully recovered with a ratio of 20 to 1, while β-glucosides were recovered with an increased liquid-to-solid ratio of 40 to 1. The extraction of aglycones was better at higher ratios, but leveled off before reaching a 100% yield. Temperature hardly affected the total amount of isoflavones. At a 20 to 1 ratio, 20°C, and pH 10, there was no significant difference (p>0.05) between isoflavone extraction in water and in 70% aqueous ethanol. The results suggest that water may be used as a green alternative for separation of isoflavones from okara.

Simultaneous extraction of oil and soy isoflavones from soy sauce residue using ultrasonic-assisted two-phase solvent extraction technology

Simultaneous extraction of oil and soy isoflavones from soy sauce residue (SSR) was investigated by means of the two-phase solvent extraction intensified by ultrasonication. A single factor test was first carried out to study the effects of ultrasonic time, ethanol concentration, ratio of ethanol/water phase to raw material, ratio of hexane phase to raw material, and ultrasonic power on the extraction rates of oil and isoflavones, then response surface methodology was applied to further optimize and simulate four key factors. The results showed that the extraction rates of oil and isoflavones of 92.07% and 92.53%, respectively, could be obtained when ethanol concentration was 74.88%, ratio of ethanol/water phase to raw material 15:1, ratio of hexane phase to raw material 8.64:1, extraction time 20min, and ultrasonic power 160W. The predictive rates were well matched with the experimental ones. The quality analysis of oil and soy isoflavones from SSR showed that the extracted oil with high acid value could be a potential raw material for biodiesel production instead of cooking. The extracted soy isoflavones existed mainly in aglycones form, which might afford higher bioactivity than that from the other soybean products.

Can soy intake affect serum uric acid level? Pooled analysis from two 6-month randomized controlled trials among Chinese postmenopausal women with prediabetes or prehypertension

Hyperuricemia is a recognized risk factor for cardiovascular diseases. Soy foods contain a moderate amount of purine and may predispose to raised serum uric acid (UA). However, no study has examined the long-term effect of soy intake on UA levels. We examined whether consumption of soy foods and isoflavone extracts for 6 months altered serum UA. The analysis included two randomized controlled trials (soy protein trial and whole soy trial) among total 450 postmenopausal women with either prehypertension or prediabetes. We conducted a pooled analysis by combining participants from both the soy flour and soy protein groups (combined soy foods group), participants from both the isoflavone and daidzein groups (combined isoflavone group) and participants from both milk placebo groups. Fasting venous samples were obtained at baseline and the end of the trial for serum UA analysis. In the pooled data, 417 subjects completed the study according to protocol. The baseline serum UA levels were comparable among the three combined groups. There was a lower decrease in UA levels among women in the combined soy foods group compared with women in the other two groups (p = 0.028 and 0.026). The net decrease and % decrease in UA were 14.5 μmol/L (95 % CI 1.93-25.6, p = 0.023) or 4.9 % (95 % CI 1.3-8.5 %, p = 0.023) between the combined soy foods group and placebo group. Among Chinese postmenopausal women with either prehypertension or prediabetes, soy intake did not increase urate levels.

(Bio)-synthesis of Au NPs from soy isoflavone extracts as a novel assessment tool of their antioxidant capacity

In this work, (bio)-synthesis of Au NPs using selected isoflavone classes has been studied. To this end, relevant aglycones (genistein and daidzein) and their glycosides (genistin and daidzin) as well as complex soy-extracts with a well-documented composition have been explored. A sigmoidal curve described the (bio)-synthesis of Au NPs which was used to evaluate the antioxidant activity and to carry out a reliable comparison of the antioxidant activity of soy isoflavone extracts towards the assessment of two relevant parameters: the concentration value at which the absorption reaches its half-value; X50c, and Au NPs production, KAuNPs. Although both isoflavone classes generated Au NPs; however, genistein exhibited a marked bio-synthesis efficiency with the lowest X50c = 70 mg L−1 and the highest KAuNPs = 0.200 mg−1 L values. In addition, complex soy isoflavone extracts with high total isoflavone aglycone content exhibited higher bio-synthesis efficiency than those found with high total glycoside content. Interestingly, excellent correlations between X50c and total phenolics (TP) (from Folin–Ciocalteu (FC) assay as truly antioxidant capacity assay) (r = 0.93, p-value < 0.05) and KAuNPs and TP (r = 0.85, p-value < 0.10) were also obtained confirming the superior antioxidant activity of aglycones isoflavone class. UV-vis and TEM characterization revealed that both genistein standard and soy isoflavone extracts with a high aglycone content produced red colloidal solutions of Au NPs. In contrast, glycoside standards as well as soy isoflavone extracts with a high glycoside content, generated blue colloidal solutions. The overall results obtained from our studies allowed us to hypothesize that the high antioxidant activity found in soy isoflavone extracts could have a significant direct relationship with the (bio)-synthesis of Au NPs with a small diameter and with the strong red colour exhibited at 540 nm. All of this opens new avenues in a very important research field such as the bio-synthesis of Au NPs as a novel tool for evaluation of antioxidant activity of target extracts.

Comparative Study of the Methodology Used in the Extraction of Isoflavones from Legumes Applying a Modified QuEChERS Approach

Isoflavones are phytochemicals of great interest because of their association with a large variety of positive effects on human health. The major sources of isoflavones in the diet are plants of the Leguminosae family, especially soybeans, although many other legumes more widely consumed in the Mediterranean diet have also been reported to contain these compounds. In previous work we extracted isoflavones from samples using a modified QuEChERS (Quick Easy Cheap Effective Rugged Safe) methodology. To compare different methods for placing the sample and the solvent in contact to optimise the extraction of isoflavones from legumes (chickpeas, lentils and white beans) using a modified QuEChERS methodology. Five different approaches to sample agitation were tested: vortex agitation, thermostatted stirring agitation and thermostatted tray shaking, and a thermostatted ultrasound bath and an ultrasound probe. To evaluate the different methodologies a modified QuEChERS approach was used as the extraction method. The separation and quantification of isoflavones was carried out using liquid chromatography-triple quadrupole/mass spectrometry (LC-MS/MS). The best methods were found upon using a thermostatted shaking tray for the extraction of chickpeas and white beans and the ultrasound probe for lentil samples. These methods were chosen based on the highest amount of analytes obtained as well as the best recovery values. Determination of isoflavones in foods may be affected by the different methods used to place the sample and the solvent in contact in the extraction step. The main advantages of the proposed extraction procedures are their simplicity, speed, reliability and low cost.

Soybean concentrated extract counteracts oxidative stress in the uterus of rats

Objective To evaluate the effects of soy isoflavone extract in the pro-oxidant/antioxidant balance in the uterus of ovariectomized rats.Methods Twenty 3-month-old adult female Wistar rats were divided into four equal groups: GI: sham-operated (estrous phase); GII: control ovariectomized rats; GIII: ovariectomized rats treated with genistein (50 μg/kg/day) by gavage; GIV: ovariectomized rats subcutaneously treated with estrogen (10 μg/kg/day). After 30 consecutive days of treatment, the rats were euthanized and the uterus removed. The distal thirds of the uterine horns were processed for histomorphometric analyses of endometrial and myometrial thicknesses and glandular area. Other regions of the uteri were kept in liquid nitrogen and subsequently processed for analysis of reactive species quantification (DCF), total antioxidant capacity (TAC) and lipid oxidation status (TBARS). Data were statistically analyzed by one-way ANOVA, complemented by the Tukey–Kramer test (p < 0.05).Results GII and GIII exhibited lower endometrial thickness, glandular area and myometrial thickness than GI and GIV, while a higher myometrial thickness was observed in GIV compared with the other groups. Moreover, the isoflavone-treated group showed lower DCF and TBARS compared to GII, and also an improvement of TAC compared to GI and GIV. Despite the significant decrease in TBARS, no significant difference in DCF nor a decrease in TAC were observed in GIV when compared to GII.Conclusion Our data show that isoflavones improve antioxidant status and counteract oxidative stress, without promoting a trophic effect in the uterus of rats.

The potential of crude okara for isoflavone production

This study describes the extraction of isoflavones from crude okara, a by-product from soymilk production, using industrially relevant conditions. Ethanol and water were chosen as environmentally friendly and non-toxic solvents. A wide range of ethanol concentrations was tested (0–90% ethanol) for extraction at room temperature. It was shown that the extraction of isoflavones was possible from crude okara. This creates opportunities for a more attractive extraction process regarding industrial processing, as the energy-consuming step of drying could be omitted. The optimal concentration of isoflavones in the extract was reached with ethanol concentrations between 50% and 70%. Ethanol concentrations above 60% required an elevated liquid-to-solid ratio due to the high moisture content in okara. Increased ethanol concentrations lowered the protein content, which corresponded to an increase in purity. A high water content in the solvent resulted in co-extraction of a larger amount of other components.

Effects of soy isoflavones on mammographic density and breast parenchyma in postmenopausal women

This study aims to evaluate the effects of soy isoflavones on breast tissue in postmenopausal women. In this randomized, double-blind, placebo-controlled study, 80 women (aged ≥ 45 y and with amenorrhea >12 mo) with vasomotor symptoms were randomized to receive either 250 mg of standardized soy extract corresponding to isoflavone 100 mg/day (n = 40) or placebo (n = 40) for 10 months. Breasts were evaluated through mammographic density and breast parenchyma using ultrasound (US) at baseline and 10-month follow-up. Independent t test, analysis of variance, Mann-Whitney U test, and χ2 trend test were used in statistical analysis. Baseline clinical characteristics showed no significant differences between the isoflavone group and the placebo group, with mean (SD) age of 55.1 (6.0) and 56.2 (7.7) years, mean (SD) menopause duration of 6.6 (4.8) and 7.1 (4.2) years, and mean (SD) body mass index of 29.7 (5.0) and 28.5 (4.9) kg/m2, respectively (P > 0.05). The study was completed by 32 women on isoflavone and 34 women on placebo. The groups did not differ in mammographic density or breast parenchyma by US (P > 0.05). Within each group, the baseline and final moments did not differ in mammography or US parameters significantly (P > 0.05). The use of soy isoflavone extract for 10 months does not affect breast density, as assessed by mammography and US, in postmenopausal women.

An effective and green method for the extraction and purification of aglycone isoflavones from soybean

In this paper, an effective and green strategy was developed to obtain aglycone isoflavones from soybean through the extraction, hydrolysis, and purification steps. Firstly, a novel ethanol-alkaline extraction method was designed and optimized. The high extraction yield (94.34%) of isoflavones was achieved under the optimal extraction conditions of pH 9.0, 70°C, 60 min, ethanol concentration of 65%, and 1:15 of the solid to liquid ratio. Then, the cellulase (GC-220) was used instead of traditional β-glucosidase to hydrolyze the glycoside isoflavones into aglycone isoflavones which gave an excellent conversion of 95%. Finally, the crude aglycone isoflavones have been purified by an environmental-friendly procedure comprised of ethanol precipitation, ethyl acetate extraction, and water rinse. The final amount of aglycone isoflavones obtained was 1.279 mg for every gram of defatted soybean by this strategy, and the final recovery, and purity of aglycone isoflavones could achieve 80.38 and 87.21%, respectively.

Comparative Antioxidant Activities of Soybean and Tempeh Extract in Rat Brain

Soybean and local fermented soybean, tempeh have been known for their health benefits and memory enhancement. It has been reported that soybean has potential role in reducing the progression of neurodegenerative disorder. Reactive oxygen species (ROS) play a crucial role in ageing and age-related neurodegenerative changes. The present study highlighted the comparative studies between total isoflavone extract from soybean and tempeh on the neuronal antioxidant. The total isoflavone extracts were administered orally for 15 days with three selected doses (10, 20 and 40 mg/kg). Piracetam (400 mg/kg, p.o.) was used as standard drug. The measurement of brain catalase, superoxide dismutase (SOD), glutathione (GSH), glutathione reductase (GR), TBARS and nitric oxide (NO) were extended. Our results showed that soybean and tempeh extracts significantly improved antioxidant activities by significantly increased the levels of catalase, SOD, GR and GSH while decreased levels of TBARS and NO level in the rat brain. This suggests that soybean and tempeh extract might be useful in the age-related neurodegenerative changes including management and prevention of Alzheimer’s disease through its antioxidant activities.

Screening DNA Binders from Isoflavone Extracts of Red Clover by Centrifugal Ultrafiltration-LC-MSn

Centrifugal ultrafiltration and LC-MS methods were combined to screen DNA-binding agents from traditional Chinese medicine red clover isoflavone extracts. The results show that 10 red clover isoflavones have different binding degrees with DNA. The sample was analyzed by LC-MSn for identifying structures. Among these flavones studied, the strongest five potential DNA-binding agents are irilone-4(i)-O-beta-D-glucoside, glycitin, pratensein, irilone and biochanin A, respectively. A rapid centrifugal ultrafiltration-LC-MS platform for screening and identification of DNA binders from the herbal extract and other complex system was established.

Assessment of the estrogenic activities of chickpea (Cicer arietinum L) sprout isoflavone extract in ovariectomized rats

Chickpea (Cicer arietinum L) is a traditional Uighur herb. In this study we investigated the estrogenic activities of the isoflavones extracted from chickpea sprouts (ICS) in ovariectomized rats. Ten-week-old virgin Sprague-Dawley female rats were ovariectomized (OVX). The rats were administered via intragastric gavage 3 different doses of ICS (20, 50, or 100 mg·kg(-1)·d(-1)) for 5 weeks. Their uterine weight and serum levels of 17β-estradiol (E2), follicle stimulating hormone (FSH) and luteinizing hormone (LH) were measured. The epithelial height, number of glands in the uterus, and number of osteoclasts in the femur were histologically quantified, and the expression of proliferating cell nuclear antigen (PCNA) was assessed immunohistochemically. Bone structural parameters, including bone mineral density (BMD), bone volume/tissue volume (BV/TV), trabecular thickness (Tb.Th) and trabecular separation (Tb.Sp) were measured using Micro-CT scanning. Treatments of OVX rats with ICS (50 or 100 mg·kg(-1)·d(-1)) produced significant estrogenic effects on the uteruses, including the increases in uterine weight, epithelial height and gland number, as well as in the expression of the cell proliferation marker PCNA. The treatments changed the secretory profile of ovarian hormones and pituitary gonadotropins: serum E2 level was significantly increased, while serum LH and FSH levels were decreased compared with the vehicle-treated OVX rats. Furthermore, the treatments significantly attenuated the bone loss, increased BMD, BV/TV and Tb.Th and decreased Tb.Sp and the number of osteoclasts. Treatment of OVX rats with the positive control drug E2 (0.25 mg·kg(-1)·d(-1)) produced similar, but more prominent effects. ICS exhibits moderate estrogenic activities as compared to E2 in ovariectomized rats, suggesting the potential use of ICS for the treatment of menopausal symptoms and osteoporosis caused by estrogen deficiency.

Extraction and Concentration of Isoflavones from Soybean (Glycine max)

The extraction and concentration of isoflavones from defatted soy flour (DSF) is attempted by leaching, membrane process and liquid-liquid extraction. The optimized extraction conditions by response surface methodology (RSM) resulted in an isoflavone content of 3.2 mg/g of DSF. The extracted isoflavones are processed by ultrafiltration for preliminary purification resulting in 3.0 mg/g of DSF, and then concentrated by liquid-liquid extraction with diethyl ether (4.8 mg/g DSF). The final isoflavone content is 69 mg/g of dried extract with an overall recovery of 57.5%. The optimization of extraction conditions resulted in an about 2.5 fold enhancement of isoflavones and concentration step enriched the isoflavones by about 8.5 fold compared to initial extract.

Meltlets®of soy isoflavones: Process optimization and the effect of extrusion spheronization process parameters on antioxidant activity

In the current research work an attempt was made to develop “Melt in mouth pellets” (Meltlets®) containing 40% herbal extract of soy isoflavones that served to provide antioxidants activity in menopausal women. The process of extrusion–spheronization was optimized for extruder speed, extruder screen size, spheronization speed, and time. While doing so the herbal extract incorporated in the pellet matrix was subjected to various processing conditions such as the effect of the presence of other excipients, mixing or kneading to prepare wet mass, heat generated during the process of extrusion, spheronization, and drying. Thus, the work further investigates the effect of these processing parameters on the antioxidant activity of the soy isoflavone herbal extract incorporated in the formula. Thereby, the antioxidant activity of the soya bean herbal extract, Meltlets® and of the placebo pellets was evaluated using DPPH free radical scavenging assay and total reduction capacity.

QuEChERS method for the extraction of isoflavones from soy-based foods before determination by capillary electrophoresis-electrospray ionization-mass spectrometry

This article reports the development of a modified approach of the Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) methodology for the extraction of naturally occurring substances from solid soy-based products. The use of this methodology offers advantages such as simplicity and ease of use, especially taking into account the complexity of food matrices. Here, capillary electrophoresis coupled with electrospray ionization mass spectrometry (CE-ESI-MS) was applied for the separation and quantification of isoflavones in soy products. Because of the importance of the different forms of isoflavones as regards their biological activity, a method that would permit both aglycones and the glycoside forms of isoflavones to be determined was developed. The proposed method was successfully applied to the analysis of the glycosylated forms glycitin, daidzin and genistin, and the aglycones formononetin, biochanin A, glycitein, daidzein, and genistein in commercially available soy biscuit samples. The parameters evaluated for the validation of the method developed included the linearity of the calibration plots, the detection and quantification limits, the existence of matrix effect(s), repeatability, reproducibility and recovery. The results of the validation process revealed that the proposed method is a possible alternative in routine analysis of isoflavones in food matrices.

Purification of Soybean Isoflavones from Soy Sauce Residue by Ultrafiltration

The extraction of soybean isoflavones from the soy sauce residue which is the byproduct of soy sauce process is of great significance to improve the utilization of the soy sauce residue. Ultrafiltration (UF) membrane with molecular weight cutoff of 20 kDa was used to purify the extraction of isoflavones. The effects of ultrafiltration pressure, ultrafiltration temperature, feeding velocity, feed concentration and the feed pH on the membrane flux and rejection were analyzed. And then the membrane was cleaned. After purifying the crude soybean isoflavones from the soy sauce reside, the content of isoflavones in the freeze-dried powder was up to 6.27%.