Analysis of the anti-T2DM components in dichloromethane fraction of Schisandra sphenanthera and its mechanism

Background:The type 2 diabetes mellitus(T2DM)pharmacodynamic study of various parts of Schisandra sphenanthera was conducted in the previous stage,and it was found that dichloromethane extracted part(SDP)had a significant hypoglycemic effect.Therefore,the components of SDP were analyzed,and the specific mechanism of its anti-T2DM was explored.Methods:We used a high-fat,high-sugar diet in combination with streptozotocin to induce a T2DM rat model,and the model rats were divided into two groups according to body weight and blood glucose.Triglyceride,oral glucose tolerance test,fasting blood glucose,low density lipoprotein cholesterol,superoxide dismutase,insulin,glycated hemoglobin,total cholesterol,nonesterified free fatty acids,alanine aminotransferase,high-density lipoprotein cholesterol,aspartate aminotransferase,malondialdehyde,and glutathione peroxidase were measured,organ indices were calculated,and pathological sections of pancreas and liver were observed.The 16S rRNA V3–V4 region of intestinal flora was sequenced to explore the effect of SDP on biochemical indicators and intestinal flora.Based on the above indicators,the anti-T2DM mechanism of SDP in Schisandra sphenanthera was analyzed.Results:After six weeks of administration,the biochemical indices of diabetic rats were diminished compared to the control group.And SDP could significantly increase the gut microbialα-diversity index,resulting in significant changes in the flora of T2DM rats,with increased richness and diversity,reduced harmful flora,and significantly back-regulated the levels of acetic acid,propionic acid,and butyric acid.Conclusion:SDP can improve the symptoms associated with elevated blood glucose,dyslipidemia,elevated fasting insulin levels,and damaged glucose tolerance in rats.SDP against T2DM may be through the control of intestinal flora to normalize and exert anti-diabetic effect;its main active components may be lignans and terpenoids.

Chemical constituents from Schisandra sphenanthera

The chemical constituents of the stems of Schisandra sphenanthera are described for the first time. This investigation has resulted in the isolation of a new phenolic glycoside （1）, along with seven known compounds. The structure of 1 was assigned by using spectroscopic techniques, including 2D NMR spectra.

Schisanlactone G,a new 3,4-seco-lanostane triterpenoid from Schisandra sphenanthera

A new 3,4-seco-lanostane triterpenoid, schisanlactone G （1）, was isolated from the fruits of Schisandra sphenanthera. Its structure was established on the basis of extensive spectroscopic analysis.

Schisandra sphenanthera extract(Wuzhi Tablet)protects against chronic-binge and acute alcohol-induced liver injury by regulating the NRF2-ARE pathway in mice

Alcohol abuse leads to alcoholic liver disease and no effective therapy is currently available.Wuzhi Tablet(WZ), a preparation of extract from Schisandra sphenanthera that is a traditional hepatoprotective herb, exerted a significant protective effect against acetaminophen-induced liver injury in our recent studies, but whether WZ can alleviate alcohol-induced toxicity remains unclear. This study aimed to investigate the contribution of WZ to alcohol-induced liver injury by using chronic-binge and acute models of alcohol feeding. The activities of ALT and AST in serum were assessed as well as the level of GSH and the activity of SOD in the liver. The expression of CYP2E1 and proteins in the NRF2-ARE signaling pathway including NRF2, GCLC, GCLM, HO-1 were measured, and the effect of WZ on NRF2 transcriptional activity was determined. We found that both models resulted in liver steatosis accompanied by increased transaminase activities, but that liver injury was significantly attenuated by WZ. WZ administration also inhibited CYP2E1 expression induced by alcohol, and elevated the level of GSH and the activity of SOD in the liver. Moreover, the NRF2-ARE signaling pathway was activated by WZ andthe target genes were all upregulated. Furthermore, WZ significantly activated NRF2 transcriptional activity. Collectively, our study demonstrates that WZ protected against alcohol-induced liver injury by reducing oxidative stress and improving antioxidant defense, possibly by activating the NRF2-ARE pathway.

Targeted bile acids and gut microbiome profiles reveal the hepatoprotective effect of WZ tablet(Schisandra sphenanthera extract) against LCA-induced cholestasis

Cholestasis is caused by the obstacle of bile formation or secretion and can develop into severe liver diseases. We previously reported the ethanol extract of Schisandra sphenanthera(Wuzhi tablet, WZ) can significantly protect against lithocholic acid(LCA)-induced intrahepatic cholestasis in mice, partially due to the activation of PXR pathway and promotion of liver regeneration.However, the effect of WZ on the bile acids profile and gut microbiome in cholestastic mice remain unknown. In this study, the effect of WZ against LCA-induced liver injury was evaluated and its effect on the bile acids metabolome and gut microbiome profiles in cholestastic mice was further investigated. Targeted metabolomics analysis was performed to examine the change of bile acids in the serum, liver, intestine and feces. The change of intestinal flora were detected by the genomics method. Targeted metabolomics analysis revealed that WZ enhanced the excretion of bile acids from serum and liver to intestine and feces. Genomics analysis of gut microbiome showed that WZ can reverse LCA-induced gut microbiome disorder to the normal level. In conclusion, WZ protects against LCAinduced cholestastic liver injury by reversing abnormal bile acids profiles and alteration of gut microbiome.

Comparing chloroplast genomes of traditional Chinese herbs Schisandra sphenanthera and S.chinensis

Objective:Schisandra sphenanthera and S.chinensis are the two important medicinal plants that have long been used under the names of"Nan-Wuweizi"and"Wuweizi",respectively.The misuse of"NanWuweizi"and"Wuweizi"in herbal medical products calls for an accurate method to distinguish these herbs.Chloroplast(cp)genomes have been widely used in species delimitation and phylogeny due to their uniparental inheritance and lower substitution rates than that of the nuclear genomes.To develop more efficient DNA markers for distinguishing S.sphenanthera,S.chinensis,and the related species,we sequenced the cp genome ofS.sphenanthera and compared it to that of S.chinensis.Methods:The cp genome of S.sphenanthera was sequenced at the lllumina HiSeq platform,and the reference-guided mapping of contigs was obtained with a de novo assembly procedure.Then,comparative analyses of the cp genomes of S.sphenanthera and S.chinensis were carried out.Results:The cp genome of S.sphenanthera was 146853 bp in length and consisted of a large single copy(LSC)region of 95627 bp,a small single copy(SSC)region of 18292 bp,and a pair of inverted repeats(IR)of 16467 bp.GC content was 39.6%.A total of 126 functional genes were predicted,of which 113 genes were unique,including 79 protein-coding genes,30 transfer RNA(tRNA)genes,and four ribosomal RNA(rRNA)genes.Five tRNA,four protein-coding genes,and all rRNA were duplicated in the IR regions.There were 18 intron-containing genes,including six tRNA genes and 12 protein-coding genes.In addition,45 SSRs were detected.The whole cp genome of S.sphenanthera was 123 bp longer than that of S.chinensis.A total of 474 SNPs and 97 InDels were identified.Five genetic regions with high levels of variation(Pi>0.015),trnS-trnG,ccsA-ndhD,psbI-trnS,trnT-psbD and ndhF-rpl32 were revealed.Conclusion:We reported the cp genome ofS.sphenanthera and revealed the SNPs and InDels between the cp genomes of S.sphenanthera and S.chinensis.This study shed light on the species identification and further phylogenetic study within the genus of Schisandra.

基于生态位模型的南、北五味子潜在适生区预测及其影响因子研究

基于184条南五味子(Schisandra sphenanthera)地理分布记录、129条北五味子(Schisandra chinensis)地理分布记录以及23个环境因子数据,采用MaxEnt生态位模型预测南、北五味子潜在适宜分布区,评估了制约其地理分布的主要环境因子。结果表明,北五味子和南五味子MaxEnt模型训练集和验证集曲线下面积(AUC)均大于0.90,建立的模型预测结果准确。北五味子最佳生境主要分布在辽宁省和吉林省的长白山区域以及北京市;南五味子的最佳生境主要分布在秦巴山区内,以及湖南省武陵山脉,郴州市、株洲市和衡阳市,浙江省金华市等地区。限制北五味子生长的重要环境因子变量分别是最湿月份降水量、最热月份最高温度、土壤酸碱度、最暖季平均降水量、最冷季平均降水量和昼夜温差月均值;限制南五味子分布的环境因子变量分别是最冷月份最低温温度、年平均降水量、温度变化方差和降水量变化方差。南、北五味子的潜在适生区呈区域化分布。

基于《德国药品法典》要求的南五味子质量控制方法研究

目的:依据《德国药品法典》(Deutscher Arzneimittel-Codex,DAC)的相关规定,建立符合DAC要求的南五味子薄层色谱鉴别方法和含量测定方法,并比较DAC与《中华人民共和国药典》(以下简称《中国药典》)2020年版对以上2项内容规定的异同点。方法:采用硅胶GF254薄层色谱板,以乙酸-乙酸乙酯-甲苯(1∶18∶46)为展开剂,以五味子乙素和五味子醇乙为系统适应性测试(SST)指标,以五味子酯甲和五味子甲素为检测指标建立了南五味子的薄层色谱鉴别方法。以HALO Biphenyl色谱柱(250 mm×4.6 mm,5μm)为固定相,乙腈-水为流动相进行梯度洗脱,检测波长为254 nm,流速为0.8 mL·min^(-1),柱温为35℃,以五味子酯甲和五味子酯乙的分离度为SST评价指标考察方法的可行性,建立了南五味子药材中五味子酯甲和五味子甲素的高效液相色谱法(HPLC)含量测定方法。结果:按照DAC相关要求进行了SST考察,结果显示,建立的薄层色谱鉴别方法具有良好的分离能力,可满足南五味子药材薄层色谱鉴别的要求;HPLC测定结果中五味子酯甲和五味子酯乙的分离度均大于3.5,符合SST要求,可用于南五味子含量测定;方法学考察结果均符合含量测定要求;31批五味子药材中五味子酯甲和五味子甲素的质量分数分别为0~0.7611%和0.0109%~0.7909%,且2个成分含量呈正相关。结论:建立的南五味子薄层色谱鉴别方法和HPLC含量测定方法均符合DAC要求,且操作简便、结果准确、重复性好,可用于南五味子的质量控制;在药材的薄层色谱鉴别及含量测定方法方面,DAC与《中国药典》2020年版大致相同,但在SST方面存在差异,应加以重视。

南五味子二氯甲烷部位化学成分结构鉴定及对HepG2和INS-1细胞的影响

目的探讨南五味子二氯甲烷部位(SDP)对HepG2细胞胰岛素抵抗和INS-1细胞高糖损伤的影响。方法采用1×10^(-6) mol·L^(-1)胰岛素建立HepG2细胞胰岛素抵抗和30 mmol·L^(-1)葡萄糖建立INS-1细胞高糖损伤模型,并使用不同浓度的南五味子二氯甲烷部位进行干预。检测HepG2细胞中三酰甘油(TG)、丙二醛(MDA)、超氧化物歧化酶(SOD)、细胞内活性氧(ROS)、腺苷三磷酸水平(ATP)、葡萄糖水平;检测INS-1细胞中MDA、SOD、过氧化物歧化酶(GSH)、ROS、基础葡萄糖(BIS)和高糖(GSIS)水平;并采用UPLC-MS/MS法分析SDP中的化学成分。结果经胰岛素诱导48 h的HepG2细胞,模型组TG、MDA、ROS葡萄糖升高(P<0.01),SOD、ATP降低,经SDP干预后,TG、MDA、ROS均有所回调,SOD、ATP显著升高;经高糖诱导48 h的INS-1细胞,模型组MDA、ROS升高(P<0.01),SOD、GSH-Px、BIS、GSIS降低(P<0.01),SDP干预后MDA、ROS回调(P<0.01),SOD、GSH-Px、BIS、GSIS上升(P<0.01)。从SDP样品中共成功鉴定出1824个化学成分,其中134个木脂素类、283个酚酸类、283个香豆素等其他类化合物,而木脂素类化合物相对含量最高。结论SDP能改善HepG2细胞胰岛素抵抗和INS-1细胞高糖损伤,分析了SDP化学成分,为防治T2DM提供实验依据。

南五味子药材UPLC指纹图谱及一测多评法研究

建立南五味子药材超高效液相色谱(UPLC)指纹图谱和6种木脂素类成分一测多评法(QAMS),为南五味子药材质量标准提高提供参考。采用UPLC法建立南五味子药材指纹图谱,运用聚类分析(HCA)和正交偏最小二乘法-判别分析(OPLS-DA)对指纹图谱进行分析;以五味子酯甲为内参物,建立五味子酯丙、五味子酯乙、安五脂素、五味子甲素和五味子酯丁的一测多评法,并与外标法(ESM)测定结果对比,判断方法的准确性。14批南五味子药材指纹图谱确定了15个共有峰,通过对照品比对指认了其中7个成分,分别为原儿茶酸、五味子酯丙、五味子酯甲、五味子酯乙、安五脂素、五味子甲素、五味子酯丁,其指纹图谱相似度均在0.95以上;通过聚类分析(HCA)和正交偏最小二乘法-判别式分析(OPLS-DA)可以将南五味子药材分为4类,峰1(原儿茶酸)、峰2、峰4、峰6、峰7(五味子酯乙)、峰8、峰9、峰12、峰15共9个成分是导致产地差异性的主要标志物。一测多评法与外标法测定的五味子酯丙、五味子酯乙、安五脂素、五味子甲素和五味子酯丁含量结果无显著差异。所建立的方法简便、可靠,为南五味子药材质量评价提供参考。

南五味子多糖对高糖损伤INS-1细胞及糖尿病小鼠的改善作用

目的结合体内外实验研究南五味子多糖(SSP)对高糖损伤大鼠胰岛细胞瘤细胞系INS-1细胞及糖尿病小鼠的影响,以期为防治糖尿病提供理论支持。方法SSP干预高糖损伤INS-1细胞,采用CCK8法检测各组细胞活力,ELISA法测定细胞上清液胰岛素含量,比色法检测SOD及GSH-Px活性与MDA含量,DCFH-DA荧光探针法检测ROS水平。建立糖尿病小鼠模型,随机分为NC组、Mod组、Met组、SSP-Low组、SSP-Hig组。给药21 d,记录小鼠体重变化。给药结束后,检测小鼠口服葡萄糖耐量、SOD活性和胰岛素水平以及胰腺组织病理变化。结果对于高糖损伤的INS-1细胞,SSP能够提高胰岛素分泌水平(P<0.01),使SOD及GSH-Px活性显著上升(P<0.01),MDA含量和ROS水平显著下降(P<0.01);对于糖尿病小鼠,SSP能够升高体重,改善糖耐量异常,使胰岛素水平和SOD活性增加(P<0.05),胰岛内细胞数量增多,改善胰岛形态。结论南五味子多糖对高糖损伤的INS-1细胞和糖尿病小鼠具有改善作用,值得进一步深入研究。

南五味子果酒工艺优化及品质分析

以酒精度为指标,采用单因素试验和正交试验确定南五味子果酒最佳发酵工艺,并进行品质分析。结果表明,南五味子果酒最佳发酵工艺为发酵温度32℃,发酵时间8 d,初始糖度26%,菌种接种量1.2%。在此条件下发酵出来的南五味子果酒酒精度为8°,还原糖质量浓度为23 mg/mL,感官评价良好。

套作玉米对华中五味子幼苗生长及根际土壤微生物群落的影响

玉米套作如何影响华中五味子(Schisandra sphenanthera Rehd.et Wils)幼苗生长及根际微生物群落结构尚不清楚。本研究通过大田试验将华中五味子与玉米套作,探究套作对华中五味子植株生物量的影响,并使用高通量测序技术对华中五味子根际土壤的细菌、真菌群落结构进行测定。结果表明,套作增加华中五味子生物量(茎长、地径、地上部分干质量、地下部分干质量)。此外,和单作相比,与玉米套作显著改变了华中五味子土壤微生物群落结构,且提高了部分与华中五味子生物量呈正相关根际促生菌如Pseudomonas(假单胞菌)、Gaiella、Kotlabaea等的丰度。且这些与华中五味子呈正相关的细菌、真菌受氮、磷、铝等及部分微量元素调控。本研究将为华中五味子幼苗管理提供候选方案及相关理论依据。

南北五味子油的成分分析及其生物活性研究

为了研究南北五味子油的成分及其生物活性,通过超临界CO_(2)萃取法提取南北五味子油,采用GC-MS和HPLC法分析其成分,利用HepG2细胞胰岛素抵抗模型测定其降糖活性,DPPH自由基清除实验测定其抗氧化活性。结果表明北五味子萃取物中五味子醇甲、五味子乙素和脂肪酸含量明显高于南五味子,南五味子萃取物减轻胰岛素抵抗作用强于北五味子,北五味子萃取物DPPH自由基清除率高于南五味子。因此,南五味子降糖作用较好,北五味子抗氧化效果较好。

基于HPLC-DAD法对护肝片中五味子掺伪情况研究

目的建立安五脂素的高效液相色谱-二极管阵列(HPLC-DAD)含量测定方法,以对护肝片中南五味子掺伪五味子的情况进行研究。方法采用Agilent ZORBAX Eclipse XDB-C18色谱柱(4.6 mm×250 mm,5μm),流动相为甲醇-水(70∶30),柱温30℃,流速1.0 mL/min,检测波长为280 nm,参比波长为360 nm,进样量为10μL。结果安五脂素在浓度在12.064~120.640μg/mL范围内线性关系良好,平均加样回收率为98.80%,RSD为1.07%。结论建立的安五脂素含量测定方法灵敏度高、专属性强、重现性好,可用于护肝片中南五味子掺伪五味子的快速鉴别。

二色五味子与南、北五味子抗炎活性比较研究

目的:比较二色五味子与南、北五味子的抗炎活性,探讨二色五味子作为中药五味子候补资源的可行性。方法:通过角叉菜胶致小鼠足肿胀实验、二甲苯致小鼠耳肿胀实验、醋酸致小鼠腹腔毛细血管通透性增加实验探究三种五味子的抗炎活性。结果:在三种急性小鼠炎症模型中,三种五味子均可减轻小鼠的足肿胀及耳肿胀度,降低腹腔毛细血管通透性增加及炎症部位肿瘤坏死因子α(TNF-α)、白介素-6(IL-6)水平。超高效液相色谱(UPLC)分析显示,南、北及二色五味子醇提物均含有五味子醇甲、五味子酯甲、五味子甲素和五味子乙素。结论:二色五味子与南、北五味子均具有抗炎作用,且二色五味子与南五味子抗炎活性相似。

南五味子不同溶剂提取物的抑菌活性研究

[目的]研究南五味子的乙醇提取物和水提取物的抑菌活性。[方法]分离南五味子提取物中的乙醇相物质和水相物质,用牛津杯法测定其对常见的致病菌和酵母菌的抑菌活性,用平板涂布法测定最低抑菌浓度(MIC)和最低杀菌浓度(MBC)。[结果]南五味子的醇相和水相物质对革兰氏阳性菌(G+)和革兰氏阴性菌(G-)均有明显的抑菌活性,对G+的抑菌活性均强于对G-的抑菌活性;醇相的抑菌活性优于水相的抑菌活性;对酵母菌的抑菌活性均不明显;醇相和水相的抑菌活性顺序均依次为:金黄色葡萄球菌>枯草芽孢杆菌>大肠杆菌>沙门氏菌>酵母菌。[结论]南五味子的乙醇提取物和水提取物均有一定的抑菌活性,值得进一步开发利用。

南五味子木脂素成分的HPLC含量测定及其变异规律

目的 :对南五味子的木脂素成分进行含量测定 ,探讨南五味子类药材木脂素成分的变异规律及其与地理分布的关系。方法 :高效液相色谱法。色谱柱 :SpherecloneLunaC1 8(2 ) (2 5 0× 4 6mm ,5 μm) ;流动相 :A .水 ,B .甲醇 ;梯度洗脱 :0～ 2 5min ,6 5 %～ 75 %B ,2 5～ 30min ,75 %B ,30～ 70min ,75 %～ 1 0 0 % ;检测波长 :2 5 4nm、2 80nm。结果与结论 :随着地理分布的不同 ,华中五味子果实木脂素成分的结构类型及其含量有显著差异。产于秦岭南侧、东侧、中条山及太行山南端的华中五味子果实含有主要的活性木脂素成分五味子酯甲、五味子甲素、安五脂素等 ,且总木脂素含量也较高 。

南、北五味子提取物对小鼠镇静、催眠作用的影响

目的:研究并比较南、北五味子不同溶剂提取物对小鼠镇静、催眠作用的影响。方法:采用系统溶剂分离法,对南、北五味子进行系统提取,观察各组分对小鼠自主活动的影响及协同戊巴比妥钠对小鼠的催眠作用。结果:南、北五味子的醚提物和醇提物能显著减少小鼠自主活动次数,增加阈下剂量戊巴比妥钠致小鼠睡眠只数,延长阈上剂量戊巴比妥钠致小鼠睡眠时间。结论:南、北五味子镇静、催眠有效成分主要分布于醚提物和醇提物中;南、北五味子相比,北五味子的镇静作用稍好,而催眠作用相当。

南、北五味子的红外光谱鉴别

对南五味子和北五味子药材的石油醚、乙醚和水的提取物进行红外光谱测定,结果南五味子和北五味子分别具有很好的光谱特征,根据其红外光谱可以准确地鉴别南、北五味子。