Sophoricoside, a genistein glycoside from Fructus Sophorae, promotes hair growth via activation of M4 muscarinic AChR in dermal papilla cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Alopecia, or hair loss, refers to ongoing decline of mature hair on the scalp or any other region of the body. Fructus Sophorae, a fruit from Sophora japonica L., contains various phytochemicals, e.g., sophoricoside, that exhibit a broad range of pharmacological effects. The potential functions of herbal extracts deriving from Fructus Sophorae and/or its major phytochemical, sophoricoside, in treating alopecia are probed here. AIM OF STUDY: The objective was to determine the ability of Fructus Sophorae extract and sophoricoside in promoting hair growth and it signalling mechanism. METHODS: Molecular docking studies were conducted to measure the binding affinities between sophoricoside and M4 mAChR in the allosteric binding site. The mechanism of Fructus Sophorae and sophoricoside in activating the signalling involving Wnt/ß-catenin and muscarinic AChR was evaluated by using immortalized human dermal papilla cell line (DPC), as well as their roles in promoting hair growth. The activity of pTOPflash-luciferase in transfected DPCs was used to examine the transcriptional regulation of Wnt/ß-catenin-mediated genes. RT-PCR was applied to quantify mRNA expressions of the biomarkers in DPCs responsible for hair growth. The phosphorylated protein levels of Wnt/ß-catenin and PI3K/AKT in DPC were revealed by using Western blot analysis. The culture of ex vivo mouse vibrissae hair follicle was used to evaluate the hair growth after the treatments. RESULTS: The ethanol extract of Fructus Sophorae and sophoricoside activated Wnt/ß-catenin signalling. The result of molecular docking showed a high binding affinity between sophoricoside and M4 mAChR. The effect of sophoricoside was blocked by specific inhibitor of M4 mAChR, but not by other inhibitors of mAChRs. Sophoricoside promoted hair growth in cultured ex vivo mouse vibrissae hair follicle by acting through M4 mAChR. CONCLUSION: The ethanol extract of Fructus Sophorae and sophoricoside activated Wnt/ß-catenin signalling via activation of M4 mAChR. The results suggested beneficial functions of Fructus Sophorae and sophoricoside as a potential candidate in treating alopecia.

Sophoricoside ameliorates methicillin-resistant Staphylococcus aureus-induced acute lung injury by inhibiting Bach1/Akt pathway.

BACKGROUND: The lack of effective treatments for methicillin-resistant Staphylococcus aureus (MRSA) infection, which often leads to severe acute lung injury (ALI), poses a grave threat to human life. Sophoricoside (SOP), an isoflavone glycoside abundant in the fruit of traditional Chinese herbal Sophora japonica l., showed anti-inflammatory effects against atopic dermatitis, allergic inflammation, and lipopolysaccharide-induced ALI. However, its effect and underlying mechanism on MRSA-induced ALI remain unclear. PURPOSE: The aim of this study is to assess the protective effect of SOP in MRSA-induced ALI and elucidate its underlying molecular mechanisms. METHODS: In vivo experiments were conducted using wild-type mice to establish MRSA-induced ALI mouse model, and the effects of SOP on ALI were evaluated by hematoxylin-eosin staining, flow cytometry, quantitative real-time polymerase chain reaction, and several biochemical indicators. Adoptive transfer experiments and BTB and CNC homology 1 knockout (Bach1-/-) mice were also utilized in this study. In vitro studies employed murine macrophages RAW264.7 cells, primary bone marrow-derived macrophages (BMDMs), and primary lung macrophages to explore the underlying molecular mechanisms. RESULTS: The administration of SOP ameliorated MRSA-induced ALI by improving pulmonary histological damages, reducing neutrophil infiltration, suppressing oxidative stress levels, and decreasing the expression of inflammatory cytokines. In isolation experiments with ALI mouse lung macrophages and macrophage adoptive transfer experiments, SOP prevented macrophage activation, thereby reducing the production of proinflammatory cytokines. In vitro experiments demonstrated that SOP decreased the expression of inflammatory mediators in lipoteichoic acid (LTA)-stimulated RAW264.7 cells, BMDMs, and primary lung macrophages. Additionally, SOP inhibited protein kinase B (Akt) phosphorylation and treatment with MK2206-a specific inhibitor of Akt-eliminated SOP's ability to suppress LTA-stimulated macrophage inflammation. Furthermore, stimulation with LTA or MRSA up-regulated Bach1 expression; however, deletion of Bach1 abolished the inhibitory effect of SOP on p-Akt activation as well as inflammation and ALI development. CONCLUSION: This study provides the first evidence that SOP effectively mitigates MRSA-induced ALI via suppressing macrophage activation through the inhibition of Bach1/Akt pathway. These findings highlight the potential of SOP as a novel therapeutic agent for treating MRSA-induced ALI.

Sophoricoside improved Crohn's disease-like colitis by inhibiting intestinal epithelial cell apoptosis through PI3K/AKT signaling.

BACKGROUND AND AIMS: Increased apoptosis of intestinal epithelial cells (IECs) is a significant cause of intestinal barrier dysfunction in Crohn's disease (CD). Sophoricoside (SOP) is an isoflavone glycoside known for its anti-apoptotic properties. The aim of this study was to investigate the effects of SOP on mice with CD-like colitis and to understand the underlying mechanisms. METHODS: Mice treated with 2,4,6-trinitrobenzene sulfonic acid (TNBS) were used to examine the therapeutic effect of SOP on CD-like colitis and intestinal barrier damage. To further explore SOP's impact on IECs apoptosis and intestinal barrier protection, an in vitro colonic organoid apoptosis model induced by TNF-α was utilized. Network pharmacology was employed to predict the relevant pathways and molecular processes associated with SOP in the treatment of CD. RESULTS: Treatment with SOP significantly improved colitis symptoms in TNBS mice, as demonstrated by reductions in the Disease Activity Index (DAI), weight loss, colon shortening, macroscopic scores, colonic tissue inflammatory scores, and the expression of pro-inflammatory factors. Our experiments confirmed that SOP protects the intestinal barrier by counteracting IECs apoptosis. Additionally, this study established that SOP reduced IECs apoptosis by inhibiting the PI3K/AKT signaling pathway. CONCLUSIONS: SOP can reduce IECs apoptosis through the inhibition of the PI3K/AKT signaling pathway, thereby protecting the intestinal barrier. This study is the first to illustrate how SOP ameliorates colitis and protects the intestinal barrier, suggesting SOP has potential clinical application in treating CD.

Sophoricoside ameliorates cerebral ischemia-reperfusion injury dependent on activating AMPK.

AIMS: Ischemic stroke accounts for 87% of all strokes, and its death and disability bring a huge burden to society. Brain injury caused by ischemia-reperfusion (I/R) is also a major difficulty in clinical treatment and prognosis. Sophoricoside (SOP) is an isoflavone glycoside isolated from the seed of medical herb Sophora japonica L. Previously, SOP was found to be effective in anti-inflammation and glucose-lipid metabolism-related diseases. In order to investigate whether SOP has a regulatory effect on cerebral I/R injury, we conducted this study. METHODS: Here, by application of SOP into MCAO (transient middle cerebral artery occlusion)-induced mice and OGD/R (oxygen glucose deprivation/reperfusion)-induced primary neurons, the regulation effects of SOP was analyzed by detecting neurological score of post-stroke mice, phenotypes of brains and brain sections, cell viabilities, and apoptosis- and inflammation-regulation. RNA sequencing and molecular biology experiments were performed to explore the mechanism of SOP regulating cerebral I/R injury. RESULTS: SOP administration decreased the infarct size, neurological deficit score, neuronal cell injury, inflammation and apoptosis. Mechanistically, SOP exerted its protective effect by activating the AMP-activated protein kinase (AMPK) signaling pathway. CONCLUSION: SOP inhibits cerebral I/R injury by promoting the phosphorylation of AMPK.

Sophoricoside attenuates neuronal injury and altered cognitive function by regulating the LTR-4/NF-κB/PI3K signalling pathway in anaesthetic-exposed neonatal rats.

Introduction: This study examined the protective effects of sophoricoside on neuronal injury and cognitive dysfunction in anaesthetic-exposed neonatal rats. Material and methods: Neuronal injury was induced in rat pups by exposure to isoflurane (0.75%) with 30% oxygen for 6 h on P7. The protective effects of sophoricoside were evaluated by assessing cognitive function using the neurological score and Morris water maze. Neuronal apoptosis was assessed in hippocampus tissue using a TUNEL assay. The cytokine and macrophage inflammatory protein levels were assessed by ELISA. Western blot assays and RT-PCR were performed to assess the expression of NF-κB, TLR-4, Akt, and PI3K proteins in neuronal tissues. Immunohistochemical and histopathological changes were observed in the brain tissues of isoflurane-induced neuronal injury rats. Results: The sophoricoside treatment improved cognitive and neuronal function in rats exposed to isoflurane. Cytokine and MIP levels in the brain tissues of isoflurane-exposed rats decreased. However, sophoricoside treatment attenuated the expression of TLR-4, PI3K, and Akt protein in the brain tissues of isoflurane-exposed rats. The histopathology improved in the sophoricoside-treated isoflurane-exposed rats. Conclusions: Sophoricoside treatment protects against neuronal injury and reduced cognitive function in isoflurane-induced neuronal injury rats by regulating TLR-4 signalling.

Oral administration of sophoricoside (SOP) inhibits neuronal damage and neuroinflammation to curb neurodegeneration in Parkinson's disease.

Neuronal apoptosis and neuroinflammation are key factors involved in the pathological changes of Parkinson's disease (PD). Sophoricoside (SOP) has shown anti-inflammatory and anti-apoptosis effects in various diseases. However, the role of SOP in PD has not been reported. In this experiment, we found that oral administration of SOP alleviated weight loss and motor symptoms in 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-injected mice. Further studies revealed that SOP inhibited inflammatory responses and neuronal apoptosis in the midbrain region of MPTP-injected mice. In vitro mechanistic study, we found that SOP exerts neuroprotective effects through a two-sided action. On the one hand, SOP inhibits Lipopolysaccharide (LPS)-induced inflammatory responses in microglia by inhibiting the Nuclear factor kappa-B(NF-κB) pathway. On the other hand, SOP inhibits 1-methyl-4-phenylpyridinium (MPP+)-induced neuronal apoptosis by regulating the Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) signaling pathway. Thus SOP is expected to be a potential therapeutic agent for PD by targeting neuroinflammation and neuronal apoptosis.

Sophoricoside attenuates autoimmune­mediated liver injury through the regulation of oxidative stress and the NF­κB signaling pathway.

The prevalence of autoimmune hepatitis (AIH) is increasing, yet specific pharmacotherapies remain to be explored. The present study aimed to investigate the effects of sophoricoside (SOP), a bioactive component of medical herbs, on AIH and to elucidate the underlying mechanisms. Bioinformatic approaches were used to predict the potential targets and underlying regulatory mechanisms of SOP on AIH. The effects of SOP on AIH were evaluated by determining the expression levels of inflammatory cytokines, histological liver injury and hepatic fibrosis in an improved chronic cytochrome P450 2D6 (CYP2D6)­AIH mouse model and in a model of concanavalin­A (ConA)­induced acute immune­mediated liver injury. The antioxidant activity of SOP was detected in in vivo and in vitro experiments. The selected signal targeted by SOP in AIH was further confirmed using western blot analysis and immunofluorescence staining. The results of bioinformatic analysis revealed that the targets of SOP in AIH were related to oxidative stress and the NF­κB gene set. The NF­κB transcription factor family is a key player that controls both innate and adaptive immunity. The activation of the NF­κB signaling pathway is often associated with autoimmune disorders. In the animal experiments, SOP attenuated CYP2D6/ConA­induced AIH, as evidenced by a significant reduction in the levels of hepatic enzymes in serum, inflammatory cytokine expression and histological lesions in the liver. The oxidative response in AIH was also significantly inhibited by SOP, as evidenced by a decrease in the levels of hepatic malondialdehyde, and elevations in the total antioxidant capacity and glutathione peroxidase levels. The results of the in vivo and in vitro experiments revealed that SOP significantly reduced the enhanced expression and nuclear translocation of phosphorylated p65 NF­κB in the livers of mice with AIH and in lipopolysaccharide­stimulated AML12 cells. On the whole, the present study demonstrates the protective role of SOP in AIH, which may be mediated by limiting the oxidative response and the activation of the NF­κB signaling pathway in hepatocytes.

[Engineering the 182 site of cyclodextrin glucosyltransferase for glycosylated genistein synthesis].

Genistein and its monoglucoside derivatives play important roles in food and pharmaceuticals fields, whereas their applications are limited by the low water solubility. Glycosylation is regarded as one of the effective approaches to improve water solubility. In this paper, the glycosylation of sophoricoside (genistein monoglucoside) was investigated using a cyclodextrin glucosyltransferase from Penibacillus macerans (PmCGTase). Saturation mutagenesis of D182 from PmCGTase was carried out. Compared with the wild-type (WT), the variant D182C showed a 13.42% higher conversion ratio. Moreover, the main products sophoricoside monoglucoside, sophoricoside diglucoside, and sophoricoside triglucoside of the variant D182C increased by 39.35%, 56.05% and 64.81% compared with that of the WT, respectively. Enzymatic characterization showed that the enzyme activities (cyclization, hydrolysis, disproportionation) of the variant D182C were higher than that of the WT, and the optimal pH and temperature of the variant D182C were 6 and 40℃, respectively. Kinetics analysis showed the variant D182C has a lower Km value and a higher kcat/Km value than that of the WT, indicating the variant D182C has enhanced affinity to substrate. Structure modeling and docking analysis demonstrated that the improved glycosylation efficiency of the variant D182C may be attributed to the increased interactions between residues and substrate.

Engineering the 182 site of cyclodextrin glucosyltransferase for glycosylated genistein synthesis / 生物工程学报

Genistein and its monoglucoside derivatives play important roles in food and pharmaceuticals fields, whereas their applications are limited by the low water solubility. Glycosylation is regarded as one of the effective approaches to improve water solubility. In this paper, the glycosylation of sophoricoside (genistein monoglucoside) was investigated using a cyclodextrin glucosyltransferase from Penibacillus macerans (PmCGTase). Saturation mutagenesis of D182 from PmCGTase was carried out. Compared with the wild-type (WT), the variant D182C showed a 13.42% higher conversion ratio. Moreover, the main products sophoricoside monoglucoside, sophoricoside diglucoside, and sophoricoside triglucoside of the variant D182C increased by 39.35%, 56.05% and 64.81% compared with that of the WT, respectively. Enzymatic characterization showed that the enzyme activities (cyclization, hydrolysis, disproportionation) of the variant D182C were higher than that of the WT, and the optimal pH and temperature of the variant D182C were 6 and 40℃, respectively. Kinetics analysis showed the variant D182C has a lower Km value and a higher kcat/Km value than that of the WT, indicating the variant D182C has enhanced affinity to substrate. Structure modeling and docking analysis demonstrated that the improved glycosylation efficiency of the variant D182C may be attributed to the increased interactions between residues and substrate.

HPLC-UV/HRMS methods for the unambiguous detection of adulterations of Ginkgo biloba leaves with Sophora japonica fruits on an extract level.

CONTEXT: Ginkgo biloba L. (Ginkgoaceae) leaf extract is one of the most frequently sold herbal extracts. There have been reports on poor quality and adulteration of ginkgo leaf extracts or the powdered plant material with extracts or powder of Styphnolobium japonicum (L.) Schott (Fabaceae) (syn. Sophora japonica L.) fruits, which is rich in flavone glycosides. OBJECTIVE: The study investigates whether ginkgo leaves genuinely contain genistein and sophoricoside and whether these two substances could be used as markers to detect adulterations with sophora fruits. MATERIALS AND METHODS: A total of 33 samples of dried ginkgo leaves were sourced from controlled plantations in China, the USA, and France. After extraction, the samples were analyzed using two high-performance liquid chromatography (HPLC) coupled with UV/HRMS methods for the detection of genistein and sophoricoside, respectively. Chromatograms were compared to standard reference materials. RESULTS: In none of the tested ginkgo samples, neither genistein nor sophoricoside could be detected. The applied method was designed to separate genistein from apigenin. The latter is a genuine compound of ginkgo leaves, and its peak may have been previously misidentified as genistein because of the same molecular mass. The method for the detection of sophoricoside allows identification of the adulteration with sophora fruit without prior hydrolysis. By both HPLC methods, it was possible to detect adulterations of ≥2% sophora fruits in the investigated ginkgo extract. CONCLUSION: The methods allow unambiguous detection of adulterations of ginkgo leaves with sophora fruits, using genistein and sophoricoside as marker compounds.

Sophoricoside from Styphnolobium japonicum improves experimental atopic dermatitis in mice.

BACKGROUND: Abnormal immune responses, specifically excessive differentiation of Th2 cells, are associated with the development of atopic dermatitis (AD). Sophoricoside, the genistein-4'-ß-D-glucoside isolated from Styphnolobium japonicum, has previously demonstrated anti-inflammatory and immunosuppressive effects along with IL-3 and IL-5 inhibitory activities. Therefore, we speculated that sophoricoside could regulate AD by regulating abnormal immune responses. PURPOSE: To investigate the role of sophoricoside on AD-like allergic skin inflammation induced by ovalbumin (OVA) or 2,4,6-trinitrochlorobenzene (TNCB) in mouse models. METHODS: Sophoricoside was isolated from the 70% ethanol extract of S. japonicum dried mature seeds. After being submitted to a purification process, its purity was assessed by high-performance liquid chromatography (HPLC). The effects of sophoricoside were determined in vivo by OVA- and TNCB-induced AD-like allergic skin inflammation in mice. Skin tissues were subjected with hematoxylin-eosin (H&E), Giemsa and toluidine blue staining. In vitro CD4+ T cell differentiation was performed and the levels of serum immunoglobulins, cytokines, and genes related to CD4+ T cell differentiation were determined by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time PCR. Cytokine bioassay, mixed lymphocytes reaction and cell viability assay were performed. RESULTS: Topical application of sophoricoside decreased the symptoms of AD-like allergic skin inflammation, including elevated hypertrophic scars with spongiotic epidermis, epidermal hyperplasia, hyperkeratosis, infiltration of immune, and mast cells, dermal thickness, amounts of immunoglobulins, and pro-inflammatory cytokines, and the mast cell population in the skin. Sophoricoside also decreased T cell antigen receptor (TCR)-mediated immune responses. In particular, sophoricoside suppressed the differentiation of naïve CD4+ T cells into Th cell subsets, including Th1, Th2, and Th17, by inhibiting the expression of their subset-specific master transcription factors, leading to suppression of the expression and production of these cell subset-specific cytokines. CONCLUSION: Sophoricoside can improve AD-like allergic skin diseases mainly by inhibiting pathogenic CD4+ T cell differentiation and immune responses.

Sophoricoside from Sophora japonica ameliorates allergic asthma by preventing mast cell activation and CD4+ T cell differentiation in ovalbumin-induced mice.

Asthma is a chronic inflammatory lung disorder with continuously increasing prevalence worldwide. Novel strategies are needed to prevent or improve asthma. The aim of this study was to investigate the effects of sophoricoside from Sophora japonica on allergic asthma. The mature seeds of S. japonica contain a large amount of sophoricoside. Sophoricoside reduced allergic and asthmatic symptoms by suppressing airway inflammation and antibody-antigen reaction in mouse models. In particular, sophoricoside suppressed immune cell recruitment into the airway lumens of the lungs and production of pro-inflammatory cytokines in the bronchoalveolar lavage fluid (BALF) of ovalbumin (OVA)-induced mice. It also decreased the amounts of histamine and arachidonic acid metabolites released in OVA-induced mice and antibody-antigen stimulated mast cells. In addition, sophoricoside decreased differentiation of naïve CD4+ T cells into T helper type 1 (Th1), Th2, and Th17 cells. Overall, we demonstrated that sophoricoside improved allergic asthma by suppressing mast cell activation and CD4+ T cell differentiation.

Sophoricoside attenuates lipopolysaccharide-induced acute lung injury by activating the AMPK/Nrf2 signaling axis.

Sophoricoside (SOP), an isoflavone glycoside isolated from seed of Sophora japonica L., has been reported to have various pharmacological activities, including anti-cancer, anti-allergy and anti-inflammation. However, the effect of SOP on lipopolysaccharides (LPS)-acute lung injury (ALI) is completely unclear. Here, we found that SOP pretreatment significantly ameliorated LPS-induced pathological damage, tissue permeability, neutrophil infiltration and the production of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) in a murine model of ALI. Besides, SOP reduced the production of pro-inflammatory mediators such as iNOS, NO and inflammatory cytokines including TNF-α, IL-1ß and IL-6 in LPS-stimulated RAW264.7 cells and bone marrow derived macrophages. Interestingly, treatment with SOP exhibited no effect on the activation of NF-κB and MAPKs in macrophages but prominently accelerated the expression and nuclear translocation of Nrf2. By using ML385, a specific Nrf2 inhibitor, we found that inhibition of Nrf2 abolished the inhibitory effect of SOP on LPS-induced iNOS expression, NO production as well as pro-inflammatory cytokine generation. SOP also activated AMPK, an upstream protein of Nrf2, under LPS stimuli. Furthermore, we demonstrated that the accelerated expression of Nrf2 induced by SOP was reversed by interference with the AMPK inhibitor Compound C. Taken together, our results suggested that SOP attenuated LPS-induced ALI in AMPK/Nrf2 dependent manner and indicated that SOP might be a potential therapeutic candidate for treating ALI/ARDS.

Quality Analysis and Suggestion for Ginkgo Folium Tablets and Ginkgo Folium Capsules / 中国实验方剂学杂志

Objective:To conduct quality evaluation of Ginkgo Folium preparations by analyzing the national evaluation sampling test results， analyze the quality differences， and put forward suggestions for the improvement of quality standards and market supervision. Method:The contents of total flavonol glycosides and terpene lactones in Ginkgo Folium tablets and Ginkgo Folium capsules were determined according to the methods of determination in the 2015 edition of Chinese Pharmacopoeia （the first volume）， and the contents of free flavonoids （quercetin， kaempferide and isorhamnetin） and sophoricoside in Ginkgo Folium preparations were determined according to related supplementary testing method of Ginkgo Folium tablets and Ginkgo Folium capsules issued by National Medical Products Administration. The quality differences of Ginkgo Folium preparations from different batches and different manufacturers were compared according to the contents of total flavonol glycosides， terpene lactones， free flavonoids and sophoricoside in 328 batches of Ginkgo Folium tablets and Ginkgo Folium capsules manufactured by 48 enterprises. Result:Quality of 328 batches of Ginkgo Folium tablets and Ginkgo Folium capsules was in accordance with the standard， but the contents of terpene lactones and total flavonol glycosides were all distributed in a wide range， and the quality of samples varied greatly among different enterprises. Conclusion:It is recommended that each enterprise should optimize the production process and strictly control the raw materials to ensure the consistency between different batches of samples.

Sophoricoside ameliorates cardiac hypertrophy by activating AMPK/mTORC1-mediated autophagy.

AIM: The study aims to evaluate protective effects of sophoricoside (Sop) on cardiac hypertrophy. Meanwhile, the potential and significance of Sop should be broadened and it should be considered as an attractive drug for the treatment of pathological cardiac hypertrophy and heart failure. METHODS: Using the phenylephrine (PE)-induced neonatal rat cardiomyocytes (NRCMs) enlargement model, the potent protection of Sop against cardiomyocytes enlargement was evaluated. The function of Sop was validated in mice received transverse aortic coarctation (TAC) or sham surgery. At 1 week after TAC surgery, mice were treated with Sop for the following 4 weeks, the hearts were harvested after echocardiography examination. RESULTS: Our study revealed that Sop significantly mitigated TAC-induced heart dysfunction, cardiomyocyte hypertrophy and cardiac fibrosis. Mechanistically, Sop treatment induced a remarkable activation of AMPK/mTORC1-autophagy cascade following sustained hypertrophic stimulation. Importantly, the protective effect of Sop was largely abolished by the AMPKα inhibitor Compound C, suggesting an AMPK activation-dependent manner of Sop function on suppressing pathological cardiac hypertrophy. CONCLUSION: Sop ameliorates cardiac hypertrophy by activating AMPK/mTORC1-mediated autophagy. Hence, Sop might be an attractive candidate for the treatment of pathological cardiac hypertrophy and heart failure.

Sophoricoside is a selective LXRß antagonist with potent therapeutic effects on hepatic steatosis of mice.

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease characterized by the accumulation of triglycerides and associated with obesity, hyperlipidemia and insulin resistance. Currently, there is no therapy for NAFLD. Emerging evidences suggest that the inhibition of liver X receptor (LXR) activity may be a potential therapy for hepatic steatosis. Here, we identified that sophoricoside is a selective antagonist of LXRß. Sophoricoside protected against obesity and glucose tolerance, and inhibited lipid accumulation in the liver of high-fat diet-induced obesity (DIO) mice and methionine and choline-deficient diet-induced nonalcoholic steatohepatitis mice. Furthermore, sophoricoside inhibited malondialdehyde, and increased superoxide dismutase and glutathione in the liver of the mice. In HepG2 cells, pretreatment with sophoricoside rescued GSH concentration decrease induced by H2 O2 treatment. Our data suggest that sophoricoside is a novel LXRß selective antagonist and may improve glucose and lipid dysfunction, and attenuate lipid accumulation in the liver of DIO mice via anti-oxidant properties, which may be developed as a therapy for NAFLD.

Engineering of Cyclodextrin Glycosyltransferase Reveals pH-Regulated Mechanism of Enhanced Long-Chain Glycosylated Sophoricoside Specificity.

Sophoricoside glycosylated derivatives, especially long-chain glycosylated sophoricosides (LCGS), have greatly improved water solubility compared with sophoricoside. Here, cyclodextrin glycosyltransferase from Paenibacillus macerans (PmCGTase) was employed for sophoricoside glycosylation. Saturation mutagenesis of alanine 156, alanine 166, glycine 173, and leucine 174 was performed due to their nonconservative properties among α-, ß-, and γ-CGTases with different product specificities. Variants L174P, A156V/L174P, and A156V/L174P/A166Y greatly improved the product specificity for LCGS. pH significantly affected the extent of glycosylation catalyzed by the variants. Further investigations revealed that the pH-regulated mechanism for LCGS synthesis mainly depends on a disproportionation route at a lower pH (pH 4) and a cyclization-coupling route at a higher pH (pH 8) and equivalent effects of cyclization-coupling and disproportionation routes at pH 5. Whereas short-chain glycosylated sophoricosides (SCGS) are primarily produced via disproportionation of maltodextrin at pH 4 and secondary disproportionation of LCGS at pH 8. At pH 5, SCGS synthesis mainly depends on a hydrolysis route by the wild type (WT) and a secondary disproportionation route by variant A156V/L174P/A166Y. Kinetics analysis showed a decreased Km value of variant A156V/L174P/A166Y. Dynamics simulation results demonstrated that the improved LCGS specificity of the variant is possibly attributed to the enhanced affinity to long-chain substrates, which may be caused by the changes of hydrogen bond interactions at the -5, -6, and -7 subsites. Our results reveal a pH-regulated mechanism for product specificity of CGTase and provide guidance for engineering CGTase toward products with different sugar chain lengths.IMPORTANCE The low water solubility of sophoricoside seriously limits its applications in the food and pharmaceutical industries. Long-chain glycosylated sophoricosides show greatly improved water solubility. Here, the product specificity of cyclodextrin glycosyltransferase (CGTase) for long-chain glycosylated sophoricosides was significantly affected by pH. Our results reveal the pH-regulated mechanism of the glycosylated product specificity of CGTase. This work adds to our understanding of the synthesis of long-chain glycosylated sophoricosides and provides guidance for exploring related product specificity of CGTase based on pH regulation.

New crystal forms and amorphous phase of sophoricoside: X-ray structures and characterization.

Sophoricoside, which is an isoflavone glycoside found in many plant species, has recently attracted attention because of its anti-fertility activity. One solvent-free form, two solvatomorphs and an amorphous phase of sophoricoside are reported for the first time. X-ray diffractometry, differential scanning calorimetry, thermal gravimetric analysis and Fourier-transform infrared spectroscopy were used to characterize the different forms. The results show that factors such as crystal symmetry, intermolecular arrangement, conformational flexibility, hydrogen-bonding interactions and solvent incorporation lead to different solid-state forms. An investigation of the transformations of the four forms showed that they can interconvert with each other under certain conditions. Amorphous phase and solvatomorphism were unstable but can improve the solubility of sophoricoside in water.

Hepatoprotective Effects of Sophoricoside against Fructose-Induced Liver Injury via Regulating Lipid Metabolism, Oxidation, and Inflammation in Mice.

The dried fruit of Sophora japonica L. is a traditional Chinese herb tea rich in sophoricoside that is an isoflavone glycoside. The aim of current study was to investigate the hepatic protective effect of sophoricoside in high fructose (HF) diet fed mice. Healthy male mice were fed 30% fructose water and treated 80 and 160 mg/kg·bw sophoricoside continuously for 8 wk. Our data showed that administration of sophoricoside at 80 and 160 mg/kg·bw observably decreased the body weight and liver weight in HF-fed mice. It was found that the treatment of sophoricoside decreased the hepatic cholesterol and triglyceride levels, and serum low-density lipoprotein-cholesterol and apolipoprotein-B levels, and elevated the serum high-density lipoprotein-cholesterol and apolipoprotein-A1 levels. Moreover, the administration of sophoricoside decreased the HF-caused elevations of hepatic malonaldehyde, interleukin-1 and tumor necrosis factor-α levels, while increased the HF-induced decreases of hepatic superoxide dismutase and glutathione peroxidase activities. Meanwhile, serum aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase activities were reduced by treatment of sophoricoside in HF-fed mice. Histopathology of hematoxylin and eosin (H&E) and oil red O staining of liver tissues also confirmed the beneficial effects of sophoricoside against liver injury induced by HF-diet in mice. These findings indicated that sophoricoside may be a novel natural isoflavone for alleviating HF-induced liver injury. PRACTICAL APPLICATION: Fruit of Sophora japonica L. is a traditional herb tea and it recently becomes popular in China. Sophoricoside is an isoflavone glycoside (Genistein-4'-O-ß-d-glucopyranoside) isolated from S. japonical L, and it possessed differential effects on the body health. The ingestion of sophoricoside or sophora fruit tea may be a novel strategy to prevent non-alcoholic fatty liver disease.

Identification of all herb medicines in Fuhuang Tablet / 中草药

Objective To establish the identification method of Typha angustifolia (stir-bake to black), Sophora japonica, Sanguisorba officinalis (stir-bake to black), and Rheum palmatum in Fuhuang tablets. Methods HPLC method were studied to identify typhaneoside and isorhamnetin-3-O-neoheptanoside in T. angustifolia, and sophoricoside in S. japonica. HPLC conditions were as follows: Agilent TC-C18 (2) column (250 mm × 4.6 mm, 5 μm) with a gradient elution at a temperature of 30 ℃; Phosphate buffer and acetonitrile were used as the mobile phase with a flow rate of 1.0 mL/min; The detection wavelength was 254 nm. The acid hydrolysis-HPLC method was explored to identify S. officinalis (stir-bake to black). TLC identification method was performed to identify R. palmatum. Results HPLC method can identify typhaneoside, isorhamnetin-3-O-neoheptanoside, and sophoricoside synchronously. Acid hydrolysis method can identify S. officinalis (stir-bake to black) by HPLC and TLC can identify R. palmatum. Conclusion The three methods are simple and accurate with high sensitivity and good specificity, which can be used to identify all herbal medicines in Fuhuang tablets.