Comparative analysis of multi-angle structural alterations and cold-water solubility of kudzu starch modifications using different methods.

Kudzu, a plant known for its medicinal value and health benefits, is typically consumed in the form of starch. However, the use of native kudzu starch is limited by its high pasting temperature and low solubility, leading to a poor consumer experience. In this study, kudzu starch was treated using six modification techniques: ball milling, extrusion puffing, alcoholic-alkaline, urea-alkaline, pullulanase, and extrusion puffing-pullulanase. The results of the Fourier transform infrared spectrum showed that the intensity ratio of 1047/1022 cm-1 for the modified starches (1.02-1.21) was lower than that of the native kudzu starch (1.22). The relative crystallinity of modified kudzu starch significantly decreased, especially after ball milling, extrusion puffing, and alcoholic-alkaline treatment. Furthermore, scanning electron microscopy and confocal laser scanning microscopy revealed significant changes in the granular structures of the modified starches. After modification, the pasting temperature of kudzu starch decreased (except for the urea-alkaline treatment), and the apparent viscosity of kudzu starch decreased from 517.95 Pa·s to 0.47 Pa·s. The cold-water solubility of extrusion-puffing and extrusion puffing-pullulanase modified kudzu starch was >70 %, which was significantly higher than that of the native starch (0.11 %). These findings establish a theoretical basis for the potential development of instant kudzu powder.

The isolation, characterization and biological activities of the non-glucan polysaccharides from the high-starch-content plant Pueraria mirifica.

The dried root of Pueraria mirifica (P. mirifica) is an edible foodstuff widely used in Asian countries. P. mirifica is known for its high starch content. The isolation of polysaccharides from high-starch plant parts is challenging due to the interference of starch. Therefore, this study aimed to develop a technique for isolating and investigating the structure and activity of non-glucan polysaccharides from P. mirifica (PMP). An effective starch removal process was developed using α-amylase hydrolysis and thorough membrane dialysis. Four non-glucan polysaccharides were isolated, and PMP-2 was subjected to structural elucidation. The results indicated that PMP-2 has a molecular weight of 124.4 kDa and that arabinose and galactose are the main components, accounting for 27.8 % and 58.5 %, respectively. Methylation and NMR analysis suggested that PMP-2 is an Arabinogalactan composed of 1,6-linked Galp and 1,4-linked Galp as the main chain, with arabinan and rhamnose as side chains. Furthermore, PMP-C and PMP-2 exhibited concentration-dependent antioxidant activities against DPPH, ABTS, and hydroxyl radicals and certain immunomodulatory activities related to the release of NO, TNF-α and IL-6. These findings suggest that PMP-2 has potential therapeutically active ingredient in functional foods. The developed method successfully removed starch and isolated non-glucan polysaccharides from the high-starch content plant P. mirifica and can be applied to other high-starch plants.

Deep eutectic solvent extraction combined with magnetic bead ligand fishing for identification of α-glucosidase inhibitors from Pueraria lobata.

In this study, a deep eutectic solvent (DES) extraction combined with a magnetic bead ligand affinity analytical method was developed and used for α-glucosidase inhibitor identification from Pueraria lobata. Several critical parameters affecting the analysis performance, including the type of DES, molar ratio, water amount, pH, salt concentration, and volume of DES, were investigated. The selected analytical sample preparation conditions were as follows. The composition of DES is choline chloride-1,4-butanediol (1:3), the water content is 40%, pH is 7.0 and the volume of extraction solution is 2 mL. The obtained sample extraction solution was analyzed directly using α-glucosidase immobilized magnetic beads (GMBs). Three α-glucosidase inhibitors in Pueraria lobata, including puerarin, daidzin, and daidzein, were identified. Luteolin was used as a positive control to evaluate the method's selectivity. Results showed it could selectively bond to the GMBs in the DES. As the affinity analysis was performed directly in a DES, the solution-removing process could be avoided. The intra-day and inter-day precisions of the method are 5.21% and 6.38%, respectively. The solvent amount was 1/50-1/2000 of that used in traditional methods.

Characterization of a Putative New Member of the Genus Potyvirus from Kudzu (Pueraria montana var. lobata) in Mississippi.

Kudzu (Pueraria montana var. lobata), a plant native to Southeastern Asia, has become a major noxious weed covering millions of hectares in the Southern United States. A kudzu patch displaying virus-like symptoms located in Ackerman, northeastern Mississippi (MS), was used as a source for virus isolation and characterization involving mechanical and vector transmission, ultrastructural observation, surveys, Sanger and high-throughput genome sequencing, and sequence analyses. The results revealed the presence of a new potyvirus in infected kudzu, closely related to wisteria vein mosaic virus (WVMV) and provisionally named kudzu chlorotic ring blotch virus (KudCRBV). Genome features and pairwise comparison with six WVMV genomes currently available in GenBank and three additional isolates from MS sequenced in this work suggest that KudCRBV is likely a member of a new species in the genus Potyvirus. Furthermore, under experimental conditions, KudCRBV was successfully transmitted by cotton and potato aphids and mechanically to soybean and beans. A state-wide survey revealed several kudzu patches infected by the virus in northern MS.

[Effect and mechanism of Puerariae Lobatae Radix in alleviating insulin resistance in T2DM db/db mice based on intestinal flora].

This study aimed to examine the effect and underlying mechanism of Puerariae Lobatae Radix on insulin resistance in db/db mice with type 2 diabetes mellitus(T2DM) based on the analysis of intestinal flora. Fifty db/db mice were randomly divided into a model group(M group), a metformin group(YX group), a high-dose Puerariae Lobatae Radix group(YGG group), a medium-dose Puerariae Lobatae Radix group(YGZ group), and a low-dose Puerariae Lobatae Radix group(YGD group). Another 10 db/m mice were assigned to the normal group(K group). After continuous administration for eight weeks, body weight and blood sugar of mice were measured. Enzyme linked immunosorbent assay(ELISA) was used to detect glycosylated serum protein(GSP) and fasting serum insulin(FINS), and insulin resistance index(HOMA-IR) was calculated. The histopathological changes in the pancreas were observed by HE staining. Tumor necrosis factor(TNF)-α expression in the pancreas was detected using immunohistochemistry. The structural changes in fecal intestinal flora in the K, M, and YGZ groups were detected by 16S rRNA. Western blot was used to detect the expression of farnesoid X receptor(FXR) and takeda G protein-coupled receptor 5(TGR5) in the ileum, cholesterol 7α-hydroxylase(CYP7A1) and sterol 27α-hydroxylase(CYP27A1) in the liver, and G protein-coupled receptors 41(GPR41) and 43(GPR43) in the colon. Compared with the K group, the M group showed increased body weight, blood sugar, serum GSP, fasting blood glucose(FBG), and FINS, increased HOMA-IR, inflammatory infiltration of islet cells, necrosis and degeneration of massive acinar cells, unclear boundary between islet cells and acinar cells, disturbed intestinal flora, and down-regulated FXR, TGR5, CYP7A1, CYP27A1, GPR41, and GPR43. Compared with the M group, the YX, YGG, YGZ, and YGD groups showed decreased body weight, blood sugar, serum GSP, FBG, and FINS, islet cells with intact and clumpy morphology and clear boundary, necrosis of a few acinar cells, and more visible islet cells. The intestinal flora in the YGZ group changed from phylum to genus levels, and the relative abundance of intestinal flora affecting the metabolites of intestinal flora increased. The protein expression of FXR, TGR5, CYP7A1, CYP27A1, GPR41, and GPR43 increased. The results show that Puerariae Lobatae Radix can improve the inflammatory damage of pancreatic islet cells and reduce insulin resistance in db/db mice with T2DM. The mechanism of action may be related to the increase in the abundance of Actinobacteria, Bifidobacterium, and Bacteroides in the intestinal tract and the protein expression related to metabolites of intestinal flora.

Ultrasonic effects on the degradation kinetics, structural characteristics and protective effects on hepatocyte lipotoxicity induced by palmitic acid of Pueraria Lobata polysaccharides.

In this study, a high-molecular-weight Pueraria lobata polysaccharide (PLP) with a molecular weight of 273.54 kDa was degraded by ultrasound, and the ultrasonic degradation kinetics, structural characteristics and hepatoprotective activity of ultrasonic degraded PLP fractions (PLPs) were evaluated. The results showed that the ultrasonic treatment significantly reduced the Mw and particle size of PLP, and the kinetic equation of ultrasonic degradation of PLP followed to the midpoint fracture model (the fist-order model). The monosaccharide composition analysis, FT-IR, triple helix structure and XRD analysis all indicated that the ultrasound degradation did not destroy the primary structure of PLP, but the thermal stability of degraded fractions improved. Additionally, the scanning electron microscopy analysis demonstrated that the surface morphology of PLP was altered from smooth, flat, compact large flaky structure to a sparse rod-like structure with sparse crosslinking (PLP-7). The degraded PLP fractions (0.5 mg/mL) with lower Mw exhibited better antioxidant activities and protective effects against palmitic acid-induced hepatic lipotoxicity, which may be due to the increased exposure of active groups such as hydroxyl groups of PLP after ultrasound. Further investigation showed that PLPs not only increased Nrf2 phosphorylation and its nuclear translocation, thereby activating Nrf2/Keap1 signaling pathway, but also enhanced HO-1, NQO-1, γ-GCL gene expressions and promoted superoxide dismutase and catalase activities, which protected hepatocytes against PA-induced oxidative stress and lipotoxicity. Overall, our research might provide an in-depth insight into P. Lobata polysaccharide in ameliorating lipid metabolic disorders, and the results revealed that ultrasonic irradiation could be a promising degradation method to produce value-added polysaccharide for use in functional food.

Extraction, structural-activity relationships, bioactivities, and application prospects of Pueraria lobata polysaccharides as ingredients for functional products: A review.

Pueraria lobata (Willd.) Ohwi is an important resource with dual functions in medicine and food since ancient times. Polysaccharides are the main bioactive component of P. lobata and have various bioactivities, such as antidiabetic, antioxidant, immunological activities, etc. Due to the distinctive bioactivity of P. lobata polysaccharides (PLPs), the research on PLPs is booming. Although a series of PLPs have been isolated and characterized, the chemical structure and mechanism are unclear and need further study. Here, we reviewed recent progress in isolation, identification, pharmacological properties, and possible therapeutic mechanisms of PLPs to update awareness of these value-added natural polysaccharides. Besides, the structure-activity relationships, application status, and toxic effects of PLPs are highlighted and discussed to afford a deeper understanding of PLPs. This article may provide theoretical insights and technical guidance for developing PLPs as novel functional foods.

Immunoaffinity separation of miroestrol and deoxymiroestrol from Pueraria candollei var. mirifica (Airy Shaw & Suvat.) Niyomdham using fragment antigen-binding antibody produced via Escherichia coli.

INTRODUCTION: Miroestrol and deoxymiroestrol are potent phytoestrogens and are oestrogen markers of Pueraria candollei var. mirifica. However, purifying these compounds is difficult because they only exist in trace amounts. OBJECTIVES: Active fragment antigen-binding (Fab) antibodies were produced via Escherichia coli SHuffle® T7 and used to selectively separate these compounds. MATERIALS AND METHODS: Two immunoaffinity separation approaches were developed, namely the immunoaffinity column (IAC) and a cell-based method. Group-specific Fab antibodies against miroestrol and deoxymiroestrol (anti-MD Fab) were used as biological binding reagents for selective separation. RESULTS: The Fab-based IAC effectively separated miroestrol and deoxymiroestrol (0.65 and 2.24 µg per 2 mL of resin, respectively) from P. mirifica root extract. When P. mirifica extract was added to E. coli cultures during Fab expression via a cell-based method, the target compound accumulated in intracellular compartments and, thus, were separated from E. coli cells after the removal of other compounds. A yield of 1.07 µg of miroestrol per gram of cell pellet weight was obtained. Miroestrol and deoxymiroestrol were successfully purified from P. mirifica extract using anti-MD Fab via the IAC and an intracellular cell-based method. CONCLUSION: The proposed methods can simplify the miroestrol and deoxymiroestrol extraction process and provide a basis for applications utilising recombinant antibodies to separate target compounds.

Spectrum-effect relationship study to reveal the pharmacodynamic substances in Flos Puerariae-Semen Hoveniae medicine pair for the treatment of alcohol-induced liver damage.

ETHNOPHARMACOLOGICAL RELEVANCE: Alcoholic liver disease (ALD) is the most serious and irreversible liver damage associated with alcohol consumption. Flos Puerariae and Semen Hoveniae are traditional Chinese medicines (TCM) for dispelling the effects of alcohol. Many studies have shown that the combination of two medicinal materials has the enhanced effect of treating ALD. AIM OF THE STUDY: The aim of this study is to assess the pharmacological effects of Flos Puerariae-Semen Hoveniae medicine pair, to elucidate its action mechanism in the treatment of alcohol-induced BRL-3A cells, and to reveal the active ingredients in the medicine pair that exerted pharmacological effects by spectrum-effect relationship study. MATERIALS AND METHODS: Firstly, MTT assays, ELISA, fluorescence probe analysis, and Western blot were employed to study the underlying mechanisms of the medicine pair in alcohol-induced BRL-3A cells by examining pharmacodynamic indexes and related protein expression. Secondly, HPLC method was established for chemical chromatograms of the medicine pair with different ratios and the sample extracted by different solvents. Then, principal component analysis, pearson bivariate correlation analysis and grey relational analysis were applied for development of the spectrum-effect correlation between pharmacodynamic indexes and HPLC chromatograms. Moreover, prototype components and their metabolites in vivo were identified by the HPLC-MS method. RESULTS: Flos Puerariae-Semen Hoveniae medicine pair remarkably increased cell viability, decreased the activity of ALT, AST, TC and TG, reduced the generation of TNF-α, IL-1ß, IL-6, MDA and ROS, increased the activity of SOD and GSH-Px, reduced protein expression of CYP2E1, compared with alcohol-induced BRL-3A cells. The medicine pair modulated the PI3K/AKT/mTOR signaling pathways by up-regulating the levels of phospho-PI3K, phospho-AKT and phospho-mTOR. Also, the results of the spectrum-effect relationship study showed that P1 (chlorogenic acid), P3 (daidzin), P4 (6″-O-xylosyl-glycitin), P5 (glycitin), P6 (unknown), P7 (unknown), P9 (unknown), P10 (6″-O-xylosyl-tectoridin), P12 (tectoridin) and P23 (unknown) can be considered as the main components of the medicine pair in the treatment of ALD. Furthermore, 6″-O-xylosyl-tectoridin, tectoridin, daidzin, 6″-O-xylosyl-glycitin and glycitin can be absorbed into the blood and showed clear metabolic and excretion behaviors in rats. CONCLUSION: In this study, the hepatoprotective effects and the pharmacology mechanism of Flos Puerariae-Semen Hoveniae medicine pair in alcohol-induced BRL-3A cells were initially investigated and revealed. Through the spectrum-effect relationship study, the potential pharmacodynamic constituents such as daidzin, 6″-O-xylosyl-glycitin, 6″-O-xylosyl-tectoridin, glycitin, and tectoridin exert pharmacological effects on alcohol-induced oxidative stress and inflammation by modulating the PI3K/AKT/mTOR signaling pathways. This study provided experimental basis and data support for revealing the pharmacodynamic substance basis and pharmacology mechanism in the treatment of ALD. Moreover, it provides a robust mean of exploring the primary effective components responsible for the bioactivity of complicated TCM.

The chemistry and activity-orientedcharacterization of isoflavones difference between roots of Pueraria lobata and P. thomsonii guided by feature-based molecular networking.

Isoflavones are important chemical components in Pueraria species with various biological activities. This study proposed an integrated strategy combining feature-based molecular networking (FBMN), chemometrics and activity evaluation for isoflavone analysis in the roots of P. lobate (PLR) and P. thomsonii (PTR). Based on the strategy, a total of 68 isoflavones were annotated in the two Pueraria species, and 11 significant difference isoflavones between PLR and PTR were identified by chemometric methods. Additionally, the correlation coefficient between the characteristic isoflavones and hypoglycemic activity were calculated, and 7 isoflavones were further confirmed as bioactive marker compounds. This approach provided guidance for the discovery of active markers among different products.

Spatially resolved metabolomics combined with bioactivity analyses to evaluate the pharmacological properties of two Radix Puerariae species.

ETHNOPHARMACOLOGICAL RELEVANCE: P. lobata and P. thomsonii are medicinal plants with similar pharmacological functions but different therapeutic effects. A novel method is presented herein to investigate metabolites in terms of their distribution and qualification, quantification is necessary to elucidate the different therapeutic effects of the two Puerariae species. AIM OF THE STUDY: The aim of the present study was to perform spatially resolved metabolomics combined with bioactivity analyses to systematically compare the metabolite differences in P. lobata and P. thomsonii by distribution, qualification, quantification, and biological activity to evaluate their pharmacological properties. MATERIALS AND METHODS: Air flow-assisted desorption electrospray ionization-mass spectrometry imaging (AFADESI-MSI) was performed to characterize the differences in the metabolite distributions of P. lobata and P. thomsonii. Further qualitative and quantitative analyses of the differential metabolites were performed using liquid chromatography-mass spectrometry (LC-MS). Biological activities correlated with the differences in the metabolites were validated by MTT assays. RESULTS: Some metabolites showed complementary distributions of the phloem and xylem in the two species, saccharide, vitamin, and inosine levels were higher in the phloem of P. thomsonii but higher in the xylem of P. lobata. The 3'-hydroxyl puerarin level was higher in the xylem of P. thomsonii but higher in the phloem of P. lobata. Qualitative and quantitative analyses of the metabolites revealed a total of 52 key differential metabolites. MTT assays showed that daidzein, daidzin, puerarin, ononin, genistin, formononetin, 3'-hydroxy puerarin, 3'-methoxy puerarin, mirificin, and 3'-methoxy daidzin exerted protective effects on H9c2 cells against hypoxia/reoxygenation injury. P. lobata extracts exhibited a significantly better protective efficacy than P. thomsonii extracts. CONCLUSIONS: In this study, AFADESI-MSI combined with LC-MS and biological activities comprehensively elucidated metabolite differences in the distribution, qualification, quantification, and pharmacological properties of P. lobata and P. thomsonii. The results of this study could provide a novel strategy for species identification and quality assessment of similar Chinese herbal medicines.

Immunochromatographic assay for miroestrol and deoxymiroestrol, its cross-reactivity, and application in Pueraria mirifica (white Kwao Krua) analysis.

INTRODUCTION: Miroestrol (Mi) and deoxymiroestrol (Dmi) are trace, yet potent, phytooestrogens found in white Kwao Krua [Pueraria candollei var. mirifica (Airy Shaw & Suvat.) Niyomdham, PM]. However, the analysis of these substances is difficult because of complex matrix effects and their various analogues. In addition, alteration in the cross-reactivity of a gold nanoparticle (AuNP)-based immunochromatographic assay (ICA) resulting from the electrostatic adsorption between antibodies and AuNPs has not yet been evaluated. OBJECTIVES: This study aims to develop, characterise, and validate ICA with a monoclonal antibody exhibiting similar reactivity against Mi and Dmi (MD-mAb). MATERIALS AND METHODS: The ICA performance was validated for cross-reactivity and performance in comparison with those of indirect competitive enzyme-linked immunosorbent assays (icELISAs) with MD-mAb and mAb exhibiting specificity against Mi (Mi-mAb). RESULTS: The ICA showed a limit of detection (LOD) at 1 and 16 µg/mL for Mi and Dmi, respectively. The cross-reactivity of the ICA with Dmi was lower (6.25%) than that observed with the icELISA (120%). Cross-reactivity of ICA against other compounds of the PM was also correlated with those of icELISA; no false-positive/negative results were observed. The repeatability and reproducibility of the ICA were confirmed. The results obtained using ICA in samples of PM are correlated with the concentrations determined through icELISAs. CONCLUSION: An ICA with MD-mAb was constructed and validated. However, direct conjugation via the electrostatic adsorption of mAb-AuNPs was expected to alter the cross-reactivity of ICA, especially that of the analyte analogue Dmi.

Comparative stability and analytical performance of anti-miroestrol recombinant antibody in different cassettes.

Immunoassays are efficient for the phytochemical analysis of various matrices. However, producing an appropriate recombinant antibody for small molecules is challenging, resulting in costly analyses. In this study, we aimed to develop recombinant fragment antigen-binding (Fab) antibodies against miroestrol, a potent phytoestrogen marker of Pueraria candollei. Two expression cassettes of Fab were established for the production of active Fab antibodies using SHuffle® T7 Escherichia coli cells. The orientation of variable fragment heavy chain (VH) and variable fragment light chain (VL) in the expression vector constructs influences the reactivity, stability, and binding specificity of the resultant Fab. Stability testing of antibodies demonstrated that Fab is a more stable form of recombinant antibody than a single-chain variable fragment (ScFv) antibody in all conditions. Based on the obtained Fab, the ELISA specifically detected miroestrol in the range of 39.06-625.00 ng/mL. The intra- and inter-assay precisions were 0.74-2.98% and 6.57-9.76%, respectively. The recovery of authentic miroestrol spiked into samples was 106.70-110.14%, and the limit of detection was 11.07 ng/mL. The results for P. candollei roots and products determined using our developed ELISA with Fab antibody and an ELISA with anti-miroestrol monoclonal antibody (mAb) were consistent (R2 = 0.9758). The developed ELISA can be applied for the quality control of miroestrol derived from P. candollei. Therefore, the appropriate expression platform of Fab resulted in the stable binding specificity of the recombinant antibody and was applicable for immunoassays.Key points• ELISAs with Fab has higher sensitivity than that with ScFv.• Fab is more stable than ScFv.• Fab-based ELISA can be used for miroestrol determination of Pueraria candollei.

Therapeutic potential of puerarin against cerebral diseases: From bench to bedside.

The incidence of cerebral diseases is rapidly increasing worldwide, and they have become an important challenge for modern medicine. Most of the available chemical drugs used in the treatment of cerebral diseases are highly toxic and single-targeted. Therefore, novel drugs from natural resources have attracted much attention for their potential to manage cerebral diseases. Puerarin is a natural isoflavone isolated from the roots of Pueraria species such as P. lobata (Willd.) Ohwi, P. thomsonii, and P. mirifica. Several authors have reported the beneficial effects of puerarin in cerebral ischemic disease, intracerebral hemorrhage, vascular dementia, Alzheimer's disease, Parkinson's disease, depression, anxiety, and traumatic brain injury. This review summarizes the brain pharmacokinetics, brain drug delivery system, clinical use (in cerebral diseases), toxicity, and the adverse clinical reactions of puerarin. We have systematically presented the pharmacological actions and the molecular mechanisms of puerarin in various cerebral diseases to provide a direction for future researches on the therapeutic use of puerarin in cerebral diseases.

Preparation, Structural Analysis and Antioxidant Activity of Polysaccharides and Their Derivatives from Pueraria lobata.

Pueraria lobata polysaccharides (PLPs) were obtained by hot water extraction using Pueraria lobata as raw material. Structural analysis revealed that PLPs may have a repetitive backbone units of →4) -α-D-Glcp (1→4-α-D-Glcp (1→. Phosphorylated Pueraria lobata polysaccharides (P-PLPs), carboxymethylated Pueraria lobata polysaccharides (CM-PLPs) and acetylated Pueraria lobata polysaccharides (Ac-PLPs) were obtained by chemical modifications of PLPs, respectively. The physicochemical properties and antioxidant activities of these four Pueraria lobata polysaccharides were studied in comparison. In particular, the clearance rate of P-PLPs exceeded 80 %, and was expected to achieve the same effect as Vc . The results showed that the effects of different chemical modifications on the antioxidant activity of PLPs varied greatly.

Combined systematic pharmacology and urine metabonomics to study the therapeutic mechanism of type 2 diabetic treated with the herbal pair of Salvia miltiorrhiza Bunge and Pueraria montana var. lobata (Willd.) Sanjappa & Pradeep.

BACKGROUND: The herbal pair of Salvia miltiorrhiza Bunge and Pueraria montana var. lobata (Willd.) Sanjappa & Pradeep (DG) is commonly used in the treatment of type 2 diabetes (T2DM) in traditional Chinese medicine (TCM). The drug pair DG was designed by Dr. Zhu chenyu to improve the treatment of T2DM. AIM: This study combined with systematic pharmacology and urine metabonomics to explore the mechanism of DG in the treatment of T2DM. METHODS: The therapeutic effect of DG on T2DM was evaluated by fasting blood glucose (FBG) and biochemical indexes. Systematic pharmacology was used to screen the active components and targets that may be related to DG. Metabonomics was established to find urinary metabolites and pathways that may be induced by DG. Finally, integrate the results of these two parts for mutual verification. RESULTS: FBG and biochemical indexes showed that DG could reduce FBG and adjust the related biochemical indexes. Metabolomics analysis indicated that 39 metabolites were related to DG for T2DM treatment. In addition, systematic pharmacology showed compounds and potential targets which were associated with DG. Finally, 12 promising targets were selected as targets for T2DM therapy by integrating the results. CONCLUSION: The combination of metabonomics and systematic pharmacology based on LC-MS is feasible and effective, which provides strong support for exploring the effective components and pharmacological mechanism of TCM.

Identification of yeast α-glucosidase inhibitors from Pueraria lobata by ligand fishing based on magnetic mesoporous silicon combined with knock-out/knock-in technology.

In this study, a ligand fishing technique based on magnetic mesoporous silicon was established and used to screen α-glucosidase inhibitors from Pueraria lobata. To clarify quantity-activity relationships in a holistic view, the knock-out/knock-in technology was used to analyse the interactions of several active constituents in P. lobata. Magnetic mesoporous silicon with a large specific surface area and better biocompatibility was synthesised. Subsequently, α-glucosidase was immobilised on -NH2-modified magnetic mesoporous silicon, and the compounds in the crude extract of P. lobata were screened across enzyme binding. The structures of the ligands were elucidated using UPLC-Q-TOF-MS/MS, and their activities were verified by knock-out/knock-in experiments and molecular docking. Daidzein and puerarin showed α-glucosidase inhibitory activities with an IC50 of 0.088 ± 0.003 mg mL-1 and 0.414 ± 0.005 mg mL-1, respectively. Among them, puerarin, which accounted for more than 40% of the total content, showed synergistic effects with other components and was the main contributor to the α-glucosidase inhibitory activity of P. lobata.

Comparative Transcriptome Analysis of Pueraria lobata Provides Candidate Genes Involved in Puerarin Biosynthesis and Its Regulation.

Pueraria lobata is a traditional Chinese herb in which an isoflavone C-glucoside, namely puerarin, has received the utmost interest due to its medicinal properties. To date, the biogenesis of puerarin, especially its C-glucosyl reaction in the pathway, remains poorly understood. Moreover, the transcription factors (TFs) that regulate puerarin biosynthesis in P. lobata have not been reported. Here, we performed phytochemical studies on the different developmental stages of the root, stem, and leaf tissues of two P. lobata cultivars, which suggested that both the roots and stems of P. lobata were the sites of puerarin biosynthesis. RNA-sequencing was conducted with the root and stem tissues of the two cultivars under different stages, and the clean reads were mapped to the recently published genome of P. lobata var. thomsonii, yielding the transcriptome dataset. A detailed analysis of the gene expression data, gene coexpression network, and phylogeny proposed several C-GTs that likely participate in puerarin biosynthesis. The first genome-wide analysis of the whole MYB superfamily in P. lobata presented here identified a total of 123 nonredundant PlMYB genes that were significantly expressed in the analyzed tissues. The phylogenetic analysis of PlMYBs with other plant MYB proteins revealed strong PlMYB candidates that may regulate the biosynthesis of isoflavones, such as puerarin.

Oriental traditional herbal Medicine--Puerariae Flos: A systematic review.

ETHNOPHARMACOLOGICAL RELEVANCE: Pueraria Flos (PF), a traditional herbal medicine, is botanically from the dried flowers of Pueraria lobate (Willd.) Ohwi. (Chinese: ) or Pueraria thomsonii Benth. (Chinese: ). It has a long history of thousands of years in China for awakening the spleen, clearing the lungs, relieving alcohol. AIM OF THE REVIEW: This review aims to report the up-to-date research progress in ethnopharmacology, phytochemistry, pharmacology and toxicology, metabolism and therapeutic application of PF, so as to provide a strong basis for future clinical treatment and scientific research. MATERIALS AND METHODS: Relevant information on PF was collected from scientific literature databases including PubMed, CNKI and other literature sources (Ph.D. and M.Sc. dissertations and Chinese herbal classic books) by using the keyword "Puerariae". RESULTS: Briefly, phytochemical research report has isolated 39 flavonoids, 19 saponins and 25 volatile oils from PF. Flavonoids and saponins are the most important bioactive compounds, and most of the quality control studies focus on these two types of compounds. Modern pharmacological studies have revealed their significant biological activities in relieving alcoholism, hepatoprotective, anti-tumor, anti-inflammatory, and anti-oxidation, which provides theoretical support for the traditional use. CONCLUSIONS: Comprehensive analysis showed that pharmacological activity of most purified compounds from PF had not been reported. Kakkalide, tectoridin and their deglycosylated metabolites (irisolidone and tectorigenin) has been focused on excessively due to their higher content and better activities. This leads to low development and resources waste. Interestingly, PF made a breakthrough in the field of food. Many kinds of fat-lowering foods such as PILLBOX Onaka have been popular in Japan market, which received extensive attention. Therefore, we suggest that future research can be paid attention on the development of the plant's function in the field of food and medicine, as well as the transformation from experimental to clinical.

Metabolomics of the anti-inflammatory effect of Pueraria lobata and Pueraria lobata var. Thomsonii in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Pueraria lobata (Willd.) Ohwi and Pueraria lobata var. Thomsonii (Benth.) Maesen are essential medicinal and edible homologous plants widely cultivated in Asian countries. Therefore, P. lobata and P. thomsonii are widely used in the food, health products and pharmaceutical industries and have significant domestic and international market potential and research value. P. lobata and P. thomsonii have pharmacological effects in the clinic, such as antipyretic, analgesic, anti-inflammatory and antioxidant effects. These plants are commonly used in the treatment of inflammatory diseases and other related diseases. However, the potential mechanisms of the anti-inflammatory effects of P. lobata and P. thomsonii have not been elucidated. AIM OF THE STUDY: This study aimed to confirm the anti-inflammatory effects of P. lobata and P. thomsonii on inflammatory model diseases and to investigate the mechanism of their anti-inflammatory effects from the perspective of plasma metabolomics. MATERIALS AND METHODS: First, P. lobata and P. thomsonii were identified by high‒performance liquid chromatography (HPLC). Second, we established the following three inflammation models: an acute inflammation model of auricular swelling in mice induced by xylene, an acute inflammation model of foot swelling in rats induced by carrageenan gum, and a chronic inflammation model of cotton ball granuloma in rats. Then we examined the weight and swelling rate of auricular swelling in mice; the residence time, contact area, and mean contact pressure in rats on the gait meter; and the weight of granulomas in rats and the content of IL-1ß and TNF-α in plasma to investigate the anti-inflammatory pharmacodynamics of P. lobata and P. thomsonii. Third, we used LC‒MS‒based plasma metabolomics techniques to obtain potential biomarkers of P. lobata and P. thomsonii related to inflammation. Then, the potential biomarkers were enriched by MetaboAnalyst and KEGG metabolomics analysis tools to obtain metabolic pathways related to inflammation. Finally, we tested the indicators of COX-2, 5-LOX, GSH, GSSG and γ⁃GCL in rat plasma from the granuloma model by enzyme-linked immunosorbent assays (ELISAs) to verify the inflammation-related metabolic pathway. RESULTS: The experimental results showed that P. lobata and P. thomsonii could reduce the swollen weight and swelling rate of the auricle in mice, and could increase the residence time, contact area and mean contact pressure in rats on the gait meter. Moreover, P. lobata and P. thomsonii could inhibit the growth of granulomas and reduce the content of IL-1ß and TNF-α in plasma in rats. The above results preliminarily verified that P. lobata and P. thomsonii have different anti-inflammatory effects. We identified eighteen plasma biomarkers associated with P. lobata and sixteen plasma biomarkers related to P. thomsonii in regulating inflammation by a plasma metabolomics analysis. The following two major metabolic pathways were further screened and enriched: arachidonic acid metabolism and glutathione metabolism. Then we noted that P. lobata and P. thomsonii could reduce the COX-2, 5-LOX and GSSG levels and increase the GSH, GSH/GSSG and γ⁃GCL levels based on the ELISA results, which demonstrated that P. lobata and P. thomsonii affect the anti-inflammatory mechanism through arachidonic acid metabolism and glutathione metabolism. CONCLUSIONS: The results of this study further elucidate the anti-inflammatory mechanism of action of P. lobata and P. thomsonii, providing a scientific basis for developing new drugs for the treatment of inflammation-related diseases and laying a foundation for the development of herbal resources, such as P. lobata and P. thomsonii.