Studying of anti-inflammatory and antioxidant effects of tectorigenin in ovalbumin-induced asthma mice models.

BACKGROUND: Allergic asthma is an important respiratory system problem characterized by airway inflammation, breathlessness, and bronchoconstriction. Allergic asthma and its outcomes are triggered by type 2 allergic immune responses. Tectorigenin is a methoxy-isoflavone with anti-inflammatory effects. In this study, we investigated the effects of tectorigenin on the pathophysiology of allergic asthma in an animal model. METHODS: Asthmatic mice were treated with tectorigenin. Then airway hyperresponsiveness (AHR), eosinophil percentage, levels of interleukin (IL)-33, IL-25, IL-13, IL-5, IL-4, total and ovalbumin (OVA)-specific immunoglobulin (Ig)E, and lung histopathology were evaluated. RESULT: Tectorigenin significantly (P 〈 0.05) reduced eosinophil infiltration (41 ± 7%) in the broncho-alveolar lavage fluid (BALF), serum IL-5 level (41 ± 5, pg/mL), and bronchial and vascular inflammation (scores of 1.3 ± 0.2 and 1.1 ± 0.3, respectively) but had no significant effects on AHR, serum levels of IL-33, -25, -13, and -4 (403 ± 24, 56 ± 7, 154 ± 11, and 89 ± 6 pg/mL, respectively), total and OVA-specific IgE (2684 ± 265 and 264 ± 19 ng/mL, respectively), goblet cell hyperplasia, and mucus production. CONCLUSION: Tectorigenin could control inflammation and the secretion of inflammatory mediators of asthma, so it can be regarded as a potential antiasthma treatment with the ability to control eosinophilia-related problems.

Antiviral Rotenoids and Isoflavones Isolated from Millettia oblata ssp. teitensis.

Three new (1-3) and six known rotenoids (5-10), along with three known isoflavones (11-13), were isolated from the leaves of Millettia oblata ssp. teitensis. A new glycosylated isoflavone (4), four known isoflavones (14-18), and one known chalcone (19) were isolated from the root wood extract of the same plant. The structures were elucidated by NMR and mass spectrometric analyses. The absolute configuration of the chiral compounds was established by a comparison of experimental ECD and VCD data with those calculated for the possible stereoisomers. This is the first report on the use of VCD to assign the absolute configuration of rotenoids. The crude leaves and root wood extracts displayed anti-RSV (human respiratory syncytial virus) activity with IC50 values of 0.7 and 3.4 µg/mL, respectively. Compounds 6, 8, 10, 11, and 14 showed anti-RSV activity with IC50 values of 0.4-10 µM, while compound 3 exhibited anti-HRV-2 (human rhinovirus 2) activity with an IC50 of 4.2 µM. Most of the compounds showed low cytotoxicity for laryngeal carcinoma (HEp-2) cells; however compounds 3, 11, and 14 exhibited low cytotoxicity also in primary lung fibroblasts. This is the first report on rotenoids showing antiviral activity against RSV and HRV viruses.

Natural Amino Acid-Bearing Carbamate Prodrugs of Daidzein Increase Water Solubility and Improve Phase II Metabolic Stability for Enhanced Oral Bioavailability.

Daidzein (DAN) is an isoflavone, and it is often found in its natural form in soybean and food supplements. DAN has poor bioavailability owing to its extremely low water solubility and first-pass metabolism. Herein, we hypothesized that a bioactivatable natural amino acid-bearing carbamate prodrug strategy could increase the water solubility and metabolic stability of DAN. To test our hypothesis, nine amino acid prodrugs of DAN were designed and synthesized. Compared with DAN, the optimal prodrug (daidzein-4'-O-CO-N-isoleucine, D-4'-I) demonstrated enhanced water solubility and improved phase II metabolic stability and activation to DAN in plasma. In addition, unlike the passive transport of DAN, D-4'-I maintained high permeability via organic anion-transporting polypeptide 2B1 (OATP2B1)-mediated transport. Importantly, D-4'-I increased the oral bioavailability by 15.5-fold, reduced the gender difference, and extended the linear absorption capacity in the pharmacokinetics of DAN in rats. Furthermore, D-4'-I exhibited dose-dependent protection against liver injury. Thus, the natural amino acid-bearing carbamate prodrug strategy shows potential in increasing water solubility and improving phase II metabolic stability to enhance the oral bioavailability of DAN.

A Highly Sensitive and Rapid Colloidal Gold Immunoassay for Puerarin Detection.

Radix Puerariae is a traditional Chinese medicinal material with a rich history of use in East and Southeast Asia. Puerarin, a unique component of the Pueraria genus, serves as a quality control marker for herbal medicines like Pueraria lobata and Pueraria thomsonii in China, displaying diverse pharmacological properties. This study developed puerarin colloidal gold immunoassay dipsticks utilizing an anti-puerarin monoclonal antibody, resulting in a fast and sensitive detection method with a limit of 500-1000 ng·mL-1. Evaluation using tap water-extracted P. lobata and P. thomsonii samples showed consistent results compared to LC-MS analysis. Cross-reactivity assessments of puerarin analogs revealed minimal interference, affirming the dipstick's reliability for distinguishing between the two species.

Exploring simple ways to avoid collecting highly 137Cs-contaminated Aralia elata buds for the revival of local wild vegetable cultures.

Collection and cooking of wild vegetables have provided seasonal enjoyments for Japanese local people as provisioning and cultural ecosystem services. However, the Fukushima Daiichi Nuclear Power Plant accident in March 2011 caused extensive radiocesium contamination of wild vegetables. Restrictions on commercial shipments of wild vegetables have been in place for the last 10 years. Some species, including buds of Aralia elata, are currently showing radiocesium concentrations both above and below the Japanese reference level for food (100 Bq/kg), implying that there are factors decreasing and increasing the 137Cs concentration. Here, we evaluated easy-to-measure environmental variables (dose rate at the soil surface, organic soil layer thickness, slope steepness, and presence/absence of decontamination practices) and the 137Cs concentrations of 40 A. elata buds at 38 locations in Fukushima Prefecture to provide helpful information on avoiding collecting highly contaminated buds. The 137Cs concentrations in A. elata buds ranged from 1 to 6,280 Bq/kg fresh weight and increased significantly with increases in the dose rate at the soil surface (0.10-6.50 µSv/h). Meanwhile, the 137Cs concentration in A. elata buds were not reduced by decontamination practices. These findings suggest that measuring the latest dose rate at the soil surface at the base of A. elata plants is a helpful way to avoid collecting buds with higher 137Cs concentrations and aid in the management of species in polluted regions.

A Gnotobiotic Mouse Model with Divergent Equol-Producing Phenotypes: Potential for Determining Microbial-Driven Health Impacts of Soy Isoflavone Daidzein.

The implications of soy consumption on human health have been a subject of debate, largely due to the mixed evidence regarding its benefits and potential risks. The variability in responses to soy has been partly attributed to differences in the metabolism of soy isoflavones, compounds with structural similarities to estrogen. Approximately one-third of humans possess gut bacteria capable of converting soy isoflavone daidzein into equol, a metabolite produced exclusively by gut microbiota with significant estrogenic potency. In contrast, lab-raised rodents are efficient equol producers, except for those raised germ-free. This discrepancy raises concerns about the applicability of traditional rodent models to humans. Herein, we designed a gnotobiotic mouse model to differentiate between equol producers and non-producers by introducing synthetic bacterial communities with and without the equol-producing capacity into female and male germ-free mice. These gnotobiotic mice display equol-producing phenotypes consistent with the capacity of the gut microbiota received. Our findings confirm the model's efficacy in mimicking human equol production capacity, offering a promising tool for future studies to explore the relationship between endogenous equol production and health outcomes like cardiometabolic health and fertility. This approach aims to refine dietary guidelines by considering individual microbiome differences.

Genistin: A Novel Estrogen Analogue Targeting ERß to Alleviate Thrombocytopenia.

Thrombocytopenia, a prevalent hematologic challenge, correlates directly with the mortality of numerous ailments. Current therapeutic avenues for thrombocytopenia are not without limitations. Here, we identify genistin, an estrogen analogue, as a promising candidate for thrombocytopenia intervention, discovered through AI-driven compound library screening. While estrogen's involvement in diverse biological processes is recognized, its role in thrombopoiesis remains underexplored. Our findings elucidate genistin's ability to enhance megakaryocyte differentiation, thereby augmenting platelet formation and production. In vivo assessments further underscore genistin's remedial potential against radiation-induced thrombocytopenia. Mechanistically, genistin's efficacy is attributed to its direct interaction with estrogen receptor ß (ERß), with subsequent activation of both ERK1/2 and the Akt signaling pathways membrane ERß. Collectively, our study positions genistin as a prospective therapeutic strategy for thrombocytopenia, shedding light on novel interplays between platelet production and ERß.

Genistein effect in cultured ovine ovarian granulosa cells.

Genistein, an isoflavone has the potential to mimic, augment, or dysregulate the steroid hormone production pathways. We hypothesized that genistein affects the granulosa cell (GCs) functions through a series of biochemical, molecular, and genomic cascades. The present study was conducted to evaluate the impact of genistein exposure on GCs viability, apoptosis, and steroidogenesis. The present study involved 3/5 days of exposure to genistein on GCs collected from abattoir-derived ovine ovaries at doses of 0, 1, 10, 25, 50, and 100 µM. The harvested GCs were used for growth, cytotoxicity, and gene expression studies related to apoptosis, growth, and steroidogenesis. We observed that genistein had both stimulatory at 10 and 25 µM levels as well as inhibitory effects at 50 and 100 µM levels on the growth and proliferation of GCs. Genistein significantly decreased the levels of 17ß-estradiol at higher exposure (50 and 100 µM), whereas the progesterone level increased significantly as the genistein exposure increased. Additionally, genistein could also alter the mRNA expression of the steroidogenic receptor, enzymes, proteins, and growth-related genes suggesting that genistein could potentially alter the steroidogenic pathways. We conclude that genistein can interfere with cell survival and steroidogenesis by exhibiting a dose-dependent biphasic response on the viability, growth-related parameters, and the synthesis of 17ß-estradiol in the cultured GCs.

The effect of soy isoflavones on non-alcoholic fatty liver disease and the level of fibroblast growth factor-21 and fetuin A.

A two-arm randomized open labeled controlled clinical trial was conducted on 50 patients with non-alcoholic fatty liver disease (NAFLD). Subjects were randomized to either receive two tablets of soy isoflavone (100 mg/day) or placebo. At week 12, the serum levels of alanine amino transferase (ALT), aspartate amino transferase (AST) and controlled attenuation parameter (CAP) score were significantly decreased only in the soy isoflavone group (P < 0.05). A significant decline in the gamma glutamyl transferase (GGT) level was observed only in the placebo group (P = 0.017). A significant increase in the serum level of fetuin A was shown in both groups at the end of the trial with a significantly greater increment in the soy isoflavone group compared to the placebo group (P < 0.05). The changes in the serum level of FGF-21 were not significant in any of the two groups. Steatosis grade significantly improved only in the soy isoflavone group (P = 0.045). There was no significant change in the fibrosis grade in the groups. Soy isoflavone intake led to a decrease in ALT, AST, CAP score, steatosis grade and an increase in the level of fetuin A. However, no significant changes were observed in the fibrosis grade and serum levels of GGT and FGF-21.

Daidzein protects endothelial cells against high glucose-induced injury through the dual-activation of PPARα and PPARγ.

Endothelial damage caused by persistent glucose and lipid metabolism disorders is the main reason of diabetic vascular diseases. Daidzein exerts positive effects on vascular dysfunction. Peroxisome proliferator-activated receptors (PPARs) regulate critically glucose and lipid metabolism. However, the interaction of daidzein to PPARs is still insufficiently explored. In this study, the cell proliferation was detected by EdU. The intrinsic activity and binding affinity of daidzein for human PPARs (hPPARs) were estimated by transactivation reporter gene test and HPLC-UV method, respectively. Daidzein significantly reversed high glucose (HG, at 30 mmol/l)-induced injury in HUVECs, which was inhibited by both PPARα and PPARγ antagonist, but no PPARß antagonist. Daidzein selectively activated hPPARα and hPPARγ1, but weakly hPPARß. Additionally, daidzein also bound to both hPPARα and hPPARγ1. The findings suggested that daidzein may be a PPARα and PPARγ dual-agonist. The amelioration of daidzein on HUVECs from hyperglycemia may be mediated by the activation of PPARα and PPARγ receptors.

ROS Scavenging Effect of Selected Isoflavones in Provoked Oxidative Stress Conditions in Human Skin Fibroblasts and Keratinocytes.

Isoflavones, belonging to polyphenolic compounds, show structural similarity to natural estrogens, and in this context, they have been extensively studied. Some of them are also applied as cosmetic additives; however, little is known regarding their effects on skin cells. In this investigation, common isoflavones, including genistein, daidzein, glycitein, formononetin, and biochanin A, as well as coumestrol, were evaluated for antioxidant activity and their impact on human skin fibroblasts and keratinocytes. Antioxidant effects were assessed using DPPH, ABTS, and FRAP tests, and the ability to scavenge reactive oxygen species (ROS) was tested in cells with H2O2-provoked oxidative stress. The impact on the activity of antioxidant enzymes (SOD, CAT, GSH) and lipid peroxidation (MDA) was also explored. As shown by Alamar Blue and neutral red uptake assays, the compounds were not toxic within the tested concentration range, and formononetin and coumestrol even demonstrated a stimulatory effect on cells. Coumestrol and biochanin A demonstrated significant antioxidative potential, leading to a significant decrease in ROS in the cells stimulated by H2O2. Furthermore, they influenced enzyme activity, preventing depletion during induced oxidative stress, and also reduced MDA levels, demonstrating protection against lipid peroxidation. In turn, genistein, daidzein, and glycitein exhibited low antioxidant capacity.

Chickpea Sprouts as a Potential Dietary Support in Different Prostate Disorders-A Preliminary In Vitro Study.

BACKGROUND: Prostate cancer (PC) and benign prostatic hyperplasia (BPH) are common health problems in the aging male population. Due to the unexplored and unconfirmed impact of food containing isoflavones, like sprouts, on the development of the management of BPH and prostate cancer, we decided to extend the knowledge in this area. RESULTS: We have demonstrated for the first time that chickpea sprouts may play an important role in the chemoprevention of prostate disorders. However, attention should be paid to the isoflavone content in the sprouts, as in our study, chickpea sprouts with a moderate concentration of the compounds, harvested in natural light conditions (CA10L) and blue LED light (CA7B), showed the best scores in terms of their potential towards prostate disorders. METHODS: Chickpea seeds were grown in LED chambers. The methanol extracts from sprouts were quantitatively defined using the HPLC system. Experiments such as the determination of PSA, 5-α-reductase, and dihydrotestosterone were performed on PNT2 and LNCaP cells. For anti-inflammatory assays (determination of NO, IL-6, and TNF-alpha release), murine RAW264.7 macrophages were used. CONCLUSIONS: The role of legume products as a diet element should be deeply evaluated for the development of future dietary recommendations for prostate cancer and BPH prevention.

Effect of Tempeh and Daidzein on Calcium Status, Calcium Transporters, and Bone Metabolism Biomarkers in Ovariectomized Rats.

Menopause marks a critical life stage characterized by hormonal changes that significantly impact bone health, leading to a heightened susceptibility to bone fractures. This research seeks to elucidate the impact of daidzein and tempeh on calcium status, calcium transporters, and bone metabolism in an ovariectomized rat model. Forty female Wistar rats, aged 3 months, participated in a two-phase experiment. The initial phase involved inducing a calcium deficit, while the second phase comprised dietary interventions across five groups: Sham (S) and Ovariectomy (O) with a standard diet, O with bisphosphonate (OB), O with pure daidzein (OD), and O with tempeh (OT). Multiple parameters, encompassing calcium levels, calcium transporters, bone histopathology, and serum bone metabolism markers, were evaluated. The findings revealed that the OT group showcased heightened levels of bone turnover markers, such as pyridinoline, C-telopeptide of type I collagen, bone alkaline phosphatase, and procollagen type I N-terminal propeptide, in contrast to S and O groups, with statistical significance (p < 0.05). Histopathologically, both the OD and OT groups exhibited effects akin to the OB group, indicating a decrease in the surface area occupied by adipocytes in the femoral bone structure, although statistically non-equivalent, supporting the directionally similar trends. Although TRPV5 and TRPV6 mRNA expression levels in the jejunum and duodenum did not display statistically significant differences (p > 0.05), the OD and OT groups exhibited increased expression compared to the O group. We hypothesized that obtained results may be related to the effect of isoflavones on estrogen pathways because of their structurally similar to endogenous estrogen and weak estrogenic properties. In conclusion, the daily consumption of pure daidzein and tempeh could potentially improve and reinstate calcium status, calcium transport, and bone metabolism in ovariectomized rats. Additionally, isoflavone products demonstrate effects similar to bisphosphonate drugs on these parameters in ovariectomized rats.

Multifunctional thiolated chitosan/puerarin composite hydrogels with pH/glutathione dual responsiveness for potential drug carriers.

Puerarin (PUE), a natural and biologically active isoflavone extracted from Chinese medicine Pueraria lobata, can self-assemble to form a hydrogel without other chemical modifications. However, although PUE hydrogel has pH responsivity, but it is difficult to adapt to the changeable pathological environment. Therefore, thiolated chitosan (TCS) is synthesized and hybridized with PUE hydrogel to prepare TCS10/PUE composite hydrogel. The results of rheological measurement showed that the resultant composite hydrogels inherited the low loss performance of TCS hydrogel, which means that they have stronger elasticity. Transmission electron microscopy (TEM) and scanning electron microscopy (SEM) images displayed that TCS10/PUE composite hydrogel has a fibrous-network structure. X-Ray Diffractometer (XRD) and Fourier transform infrared spectroscopy (FT-IR) proved the existence of hydrogen bonds and disulfide bonds in the formation of composite hydrogel. Degradation experiment showed that TCS10/PUE composite hydrogels have pH and glutathione (pH/GSH) dual sensitivity. Furthermore, TCS10/PUE composite hydrogels exhibited multi-functionality including thixotropy, cytocompatibility, antibacterial and anti-inflammatory properties. Berberine chloride hydrate (BCH) was further used as a model drug for in vitro release study. BCH and PUE could be released cooperatively under pH/GSH dual responsivity. These results indicated that the resultant composite hydrogel has eminent pH/GSH dual responsivity and could act as a potential new intelligent drug carrier.

Effect of soy isoflavone supplementation on blood pressure: a meta-analysis of randomized controlled trials.

BACKGROUND: Previous experimental studies have suggested that the consumption of soy isoflavones may have a potential impact on lowering blood pressure. Nevertheless, epidemiological studies have presented conflicting outcomes concerning the correlation between soy isoflavone consumption and blood pressure levels. Consequently, a comprehensive meta-analysis of all eligible randomized controlled trials (RCTs) was conducted to explore the influence of soy isoflavones on systolic blood pressure (SBP) and diastolic blood pressure (DBP) in adults. METHODS: A thorough search of PubMed, Embase, and the Cochrane Library for relevant literature up to April 30, 2023 was conducted. RCTs involving adults that compared soy isoflavone supplementation with a placebo (the same matrix devoid of soy isoflavone) were included. The combined effect size was presented as the weighted mean difference (WMD) along with 95% confidence interval (CI), employing a fixed-effects model. RESULTS: Our meta-analysis included a total of 24 studies involving 1945 participants. The results revealed a significant reduction in both SBP and DBP with soy isoflavone supplementation. Subgroup analyses suggested more pronounced reductions in SBP and DBP for interventions lasting ≥6 months, in individuals receiving mixed-type soy isoflavone, and among patients with metabolic syndrome or prehypertension. However, we did not detect significant nonlinear associations between supplementation dosage and intervention duration concerning both SBP and DBP. The overall quality of evidence was deemed moderate. CONCLUSIONS: The current meta-analysis revealed that supplementation with soy isoflavones alone effectively reduces blood pressure. Additional high-quality studies are required to investigate the efficacy of blood pressure reduction through supplementation with an optimal quantity and proportion of soy isoflavone.

Soy-Based Tempeh as a Functional Food: Evidence for Human Health and Future Perspective.

Tempeh is an Indonesian traditional food made from fermented soybeans, which offers wide culinary use in East Asian countries. Similar to all fermented foods, its preparation offers the purpose of food preservation. However, preclinical studies have highlighted that microbial action leads to a modification in the nutritional composition of the food's matrix. Although there is a wide availability of data on the beneficial effect of soy, tempeh remains relatively unexplored, perhaps due to its limited diffusion in the world, which limits its research availability. However, available data suggest that tempeh may confer beneficial health effects due to the high bioavailability of nutrients and phytochemicals, showing ameliorative action on oxidative stress, glycaemic control, and blood lipid levels. Furthermore, the high biological value of tempeh means it can be used to optimize protein and caloric intake in athletes, vegetarians, and children. Moreover, the microbial fermentation used in the production of tempeh, in addition to improving the bioavailability of minerals, proteins, fibre, vitamins, and isoflavones, produces biopeptides whose biological effect is currently of great interest. Tempeh can be employed in traditional preparations as well as second-generation foods, such as plant-based meat substitutes, to provide functional and nutritional properties and a higher eco-friendly option compared to animal foods. This review aims to provide an overview of tempeh's properties, regarding human data and future research perspectives.

Anti-Obesity Effect and Signaling Mechanism of Potassium Poly-γ-Glutamate Produced by Bacillus subtilis Chungkookjang in High-Fat Diet-Induced Obese Mice.

The purpose of this work was to examine the effects of potassium poly-γ-glutamate (PGA-K) on mice fed a high-fat diet consisting of 60% of total calories for 12 weeks. PGA-K administration reduced the increase in body weight, epididymal fat, and liver weight caused by a high-fat diet compared to the obese group. The triglyceride, low-density lipoprotein cholesterol and high-density lipoprotein cholesterol levels, which are blood lipid indicators, were significantly increased in the obese group but were significantly decreased in the PGA-K-treated group. The administration of PGA-K resulted in a significant inhibition of pro-inflammatory cytokines, including tumor necrosis factor α and interleukin 6. Moreover, the levels of leptin and insulin, which are insulin resistance indicators, significantly increased in the obese group but were significantly decreased in the PGA-K-treated group. These results suggest that PGA-K exhibits a protective effect against obesity induced by a high-fat diet, underscoring its potential as a candidate for obesity treatment.

Calycosin (CA) inhibits proliferation, migration and invasion by suppression of CXCL10 signaling pathway in glioma.

Glioblastoma is the most common malignant tumor in the central nervous system and its occurrence and development is involved in various molecular abnormalities. C-X-C chemokine ligand 10 (CXCL10), an inflammatory chemokine, has been reported to be related to the pathogenesis of cancer while it has not yet been linked to glioma. Calycosin, a bioactive compound derived from Radix astragali, has demonstrated anticancer properties in several malignancies, including glioma. Nonetheless, its underlying mechanisms are not fully understood. This study explores CXCL10 as a potential therapeutic target for calycosin in the suppression of glioblastoma. We observed that CXCL10 expression correlates positively with glioma malignancy and inversely with patient prognosis, highlighting its potential as a glioblastoma treatment target. Furthermore, we found that calycosin inhibited proliferation, migration, and invasion in U87 and U251 glioma cells, and decreased CXCL10 expression in a dose-dependent manner, along with its downstream effectors such as NLRP3, NF-κB, and IL-1ß. Additionally, molecular docking experiments demonstrated that calycosin exhibits a notable binding affinity to CXCL10. Overexpression of CXCL10 counteracted the inhibitory effects of calycosin on cell proliferation, migration, and invasion, while CXCL10 knockdown enhanced these effects. Finally, we verified that calycosin inhibited glioma growth in a xenograft mouse model and downregulated CXCL10 and its downstream molecules. These findings suggest that targeting CXCL10 may be an effective strategy in glioblastoma treatment, and calycosin emerges as a potential therapeutic agent.

Influence of malting procedure on the isoflavonoid content of soybeans.

The goal of this study was to analyse, whether malting technique (consisting of seed hydration, germination and drying) can be used to modify concentration of various isoflavonoids in soybean seeds. Seeds of three soybean varieties were germinated by different lengths of time (from 24 to 120 h) and dried by two different methods, typically used to produce so-called 'light' and 'caramel' malts. It was determined, that malting decreases concentration of 7-O-ß-D-glucosides such as daidzin, genisitin and glycitin, while at the same time increasing concentration of aglycones (daidzein, genistein and glycitein). Increasing time of the germination period increased concentration of aglycones. 'Caramel' type malts were characterised with higher concentration of most of the isoflavonoids (daidzin, daidzein, genistin, genistein and glycitein) than 'light' type malts. Results of this study suggest that soybean malts can be an interesting substrate in the production of various food products with increased aglycone content.

Exploring functional genes' correlation with (S)-equol concentration and new daidzein racemase identification.

With its estrogenic activity, (S)-equol plays an important role in maintaining host health and preventing estrogen-related diseases. Exclusive production occurs through the transformation of soy isoflavones by intestinal bacteria, but the reasons for variations in (S)-equol production among different individuals and species remain unclear. Here, fecal samples from humans, pigs, chickens, mice, and rats were used as research objects. The concentrations of (S)-equol, along with the genetic homology and evolutionary relationships of (S)-equol production-related genes [daidzein reductase (DZNR), daidzein racemase (DDRC), dihydrodaidzein reductase (DHDR), tetrahydrodaidzein reductase (THDR)], were analyzed. Additionally, in vitro functional verification of the newly identified DDRC gene was conducted. It was found that approximately 40% of human samples contained (S)-equol, whereas 100% of samples from other species contained (S)-equol. However, there were significant variations in (S)-equol content among the different species: rats > pigs > chickens > mice > humans. The distributions of the four genes displayed species-specific patterns. High detection rates across various species were exhibited by DHDR, THDR, and DDRC. In contrast, substantial variations in detection rates among different species and individuals were observed with respect to DZNR. It appears that various types of DZNR may be associated with different concentrations of (S)-equol, which potentially correspond to the regulatory role during (S)-equol synthesis. This enhances our understanding of individual variations in (S)-equol production and their connection with functional genes in vitro. Moreover, the newly identified DDRC exhibits higher potential for (S)-equol synthesis compared to the known DDRC, providing valuable resources for advancing in vitro (S)-equol production. IMPORTANCE: (S)-equol ((S)-EQ) plays a crucial role in maintaining human health, along with its known capacity to prevent and treat various diseases, including cardiovascular diseases, metabolic syndromes, osteoporosis, diabetes, brain-related diseases, high blood pressure, hyperlipidemia, obesity, and inflammation. However, factors affecting individual variations in (S)-EQ production and the underlying regulatory mechanisms remain elusive. This study examines the association between functional genes and (S)-EQ production, highlighting a potential correlation between the DZNR gene and (S)-EQ content. Various types of DZNR may be linked to the regulation of (S)-EQ synthesis. Furthermore, the identification of a new DDRC gene offers promising prospects for enhancing in vitro (S)-EQ production.