Molecular mechanisms of apoptosis in hepatocellular carcinoma cells induced by ethanol extracts of Solanum lyratum Thumb through the mitochondrial pathway

AIM To explore the induction effects and mechanism of Solanum lyratum Thumb(ST) on human hepatocellularcarcinoma SMMC-7721 cells through the mitochondrial pathway.METHODS The experiments were conducted on three groups: an experimental group (with ST ethanol extracts' concentration being 2.5, 5 and 10 mg/L), a negative control group (with only nutrient solution, 0 mg/L ST ethanol extracts), and a positive control group (2.5 mg/L DDP). The inhibition rate of cell proliferation was checked by using the methyl thiazolyl tetrazolium method, and cell apoptosis was tested by TUNEL method. Furthermore, RT-PCR was used to examine m RNA expression of Fas, Fas L, caspase-8, caspase-3, p53 and Bcl-2 genes.RESULTS Compared with the negative control group, the inhibition and apoptosis rates of the experimental group with different concentrations of ST extracts on human hepatocellular carcinoma SMMC-7721 cells significantly increased(P<0.05). Besides, the m RNA expression of Fas L and Bcl-2 significantly decreased(P<0.05) while the m RNA expression of Fas, caspase-8, caspase-3 and p53 increased significantly. When compared with the positive control group, the experimental groups with 5 mg/L ST ethanol extracts showed effects similar to the positive control group.CONCLUSION ST ethanol extracts induced the apoptosis of hepatocellular carcinoma SMMC-7721 cells through up-regulated Fas, caspase-8, caspse-3 and p53, and down-regulated Fas L and Bcl-2 in the mitochondrial pathway.

RP-HPLC method for the simultaneous determination of daidzein,genistein and formonetin in Solanum Lyratum Thunb

A rapid method for the simultaneous determination of daidzein,genistein and formonetin in solanum Lyratum Thunb by high performance liquid chromatography(HPLC)was developed.Separation was achieved on a Diamonsil C18 column(250 mm×4.6 mm,5 μm)with isocratic elution,using a mobile phase of methanol-tetrahydrofuran-water(44∶3∶53,v/v).The wavelength was set at 260 nm and column was maintained at 35 ℃.The linear ranges of daidzein,genistein and formonetin were 1.0-40.0,0.1-4.0 and 0.1-4.0 μg/mL,respectively.The average recoveries were between 98.4% and 101.3%.This method could be used for the quality control of Solanum lyratum Thunb due to its simplification,reliability,rapidity and excellent precision.

Phytochemical and pharmacological studies on Solanum lyratum:a review

Solanum lyratum is one of the temperate plants,broadly distributed in Korea,China,Japan,India,and South-East Asia and well-documented in those oriental ethnic medicine systems for curing cancers,jaundice,edema,gonorrhea,cholecystitis,phlogosis,rheumatoid arthritis,etc.This review systematically summarized the research progress on S.lyratum respecting the botany,traditional uses,phytochemistry,pharmacology,and toxicology to increase people’s in-depth understanding of this plant,by data retrieval in a series of online or off-line electronic databases as far as we can reach.Steroidal saponins and alkaloids,terpenoids,nitrogenous compounds,and flavonoid compounds are the main chemical constituents in S.lyratum.Among them,steroidal alkaloids and saponins are the major active ingredi-ents ever found in S.lyratum,exerting activities of anti-cancer,anti-inflammation,anti-microbial,anti-allergy,and anti-oxidation in vivo or in vitro.As a result,S.lyratum has been frequently prescribed for the abovementioned therapeutic purposes,and there are substantial traditional and modern shreds of evidence of its use.

A review:Chemical constituents and pharmacological activities of Solanum lyratum Thunb.

Solanum lyratum Thunb.belonging to the family Solanaceae,is a common Chinese herbal medicine in folk.It has been used for the treatment of colds,fever,jaundice hepatitis,cholecystitis,cholelithiasis,nephritis,edema and cancer.Previous phytochemical studies on S.lyratum have led to the identification of many chemical constituents,including steroidal saponins,alkaloids,terpenoids,flavonoids,quinonoids,phenylpropanoids,organic acids and several other kinds of compounds,which showed diverse bioactivities,such as anticancer,anti-inflammatory and antioxidant activities.This paper summarized the primary chemical constituents and pharmacological effects of S.lyratum to provide reference for its further development and utilization.

New steroidal alkaloid and furostanol glycosides isolated from Solanum lyratum with cytotoxicity

Two previously undescribed steroidal compounds, 16, 23-epoxy-22, 26-epimino-cholest-22(N), 23, 25(26)-trien-3β-ol-3-O-β-D-glucopyranosyl-(1→2)-β-D-glucopyranosyl-(1→4)-β-D-galactopyranoside(1) and 26-O-β-D-glucopyranosyl-(25 R)-5α-furost-20(22)-en-3β, 26-diol(2), together with 7 known ones including 26-O-β-D-glucopyranosyl-(25 R)-5, 20(22)-dien-furost-3β, 26-diol(3),(25 R)-5-en-spirost-3β-ol-O-β-D-glucopyranosyl-(1→4)-[α-L-rhmanopyranosyl-(1→2)]-β-D-galactopyranoside(4), funkioside D(5), aspidistrin(6), tigogenin-3-O-β-D-lucotrioside(7), desglucolanatigonin II(8), and degalactotigonin(9), were isolated from Solanum lyratum Thunb. Their cytotoxic activities were tested in two cancer cell lines by MTT method. One of the steroidal glycosides(6) showed significant cytotoxic activity against gastric cancer SGC7901 and liver cancer BEL-7402 cells.

Lignanoids from Solanum lyratum

In the present study,we aimed to investigate the chemical constituents from Solanum lyratum.The constituents were separated by column chromatography on silica gel,Sephadex LH-20 and preparative HPLC.Their structures were elucidated by spectroscopic means.A total of 13 compounds were isolated from S.lyraturn and identified as syringin(1),(+)-isolariciresinol(2),(+)-syringaresinol(3),leptolepisol D(4),(-)-secoisolariciresinol(5),(-)-epi-syringaresinol(6),aviculin(7),zhebeiresinol(8),ciwujiatone(9),(-)-(7′S,8S,8′R)-4,4′-dihydroxy-3,3′,5,5′-tetramethoxy-7′,9-epoxylignan-9′-ol-7-one(10),(+)-lariciresinol(11),(+)-pinoresinol(12)and(+)-medioresinol(13).All the compounds were isolated from S.lyratum for the first time.

Two new cytotoxic sesquiterpenoids from Solanum lyratum

Two new sesquiterpenoids,attributable to eudesmane-type（1 and 2,named solajiangxins F and G）,were isolated from the whole plant of Solanum lyratum.Their structures were elucidated on the basis of integrated spectroscopic techniques.In vitro,compounds 1 and 2 were found to show signifcant cytotoxic activity against selected cancer cells,including P-388,HONE-1 and HT-29.

Three previously undescribed steroidal glycoalkaloids from Solanum lyratum and their anti-tumor activities

In the present study,three previously undescribed steroidal glycoalkaloids(compounds 1–3)were isolated from Solanum lyratum.Their structures were elucidated based on comprehensive spectroscopic data.Their anti-angiogenesis and anti-metastatic activities were evaluated by MTT and wound-healing assays,respectively.Tumor-derived vascular endothelial cells(TdECs),obtained by co-culture of A549 and human umbilical vein endothelial cells(HUVECs),were treated with compounds 1–3.Results showed that compounds 1–3 significantly inhibited the migration of TdECs at 25μM despite the weak cytotoxic activities,which indicated that the compounds exerted anti-tumor activities by inhibiting metastasis,rather than directly inhibiting the proliferation of TdECs.

白毛藤化学成分及其抗肿瘤活性研究

目的研究白毛藤化学成分及其抗肿瘤活性。方法白毛藤提取物采用硅胶柱、SephadexLH-20柱进行分离纯化,根据理化性质及波谱数据鉴定所得化合物的结构。MTT法测定其抗肿瘤活性。结果从中分离得到21个化合物,分别鉴定为royleanone(1)、邻苯二甲酸二正丁酯(2)、邻苯二甲酸二正戊酯(3)、sativan(4)、2′,4′-二羟基查尔酮(5)、isopetasin(6)、balanophonin(7)、10-eicosenoicacid(8)、4,6,7-三甲氧基-5-甲基-香豆素(9)、hydroxydihydrobovolide(10)、axillarin(11)、柳穿鱼黄素(12)、magnoflorine(13)、委陵菜酸(14)、芹菜素(15)、木犀草素(16)、7-ketositosterol(17)、pinoresinol(18)、bavachinine(19)、bisbenzopyran(20)、isotachioside(21)。化合物6~7、11、18~19IC50值在(28.37±3.71)~(73.26±8.61)μmol/L之间。结论所有化合物均为首次从该植物中分离得到。化合物6~7、11、18~19具有一定的抗肿瘤活性。

Two new glycoalkaloids from Solanum lyratum Thunb.

To study the alkaloids of Solanum lyratum Thunb.ODS and silica gel column chromatography combined with HPLC were used to separate and purify the alkaloids.Seven alkaloids were identified via UHPLC-ESI-Q-TOF-MS,1 D and 2 D NMR,respectively 15β-ethoxy-(3β,5α,25 R)-16,23-epoxy-23,24-imino-cholestan-16,20,23(N)-triene-3β-ol-3-O-β-D-glucopyranosyl-(1→2)-β-D-glucopyranosyl-(1→4)-β-D-galactopyran-oside(1),15β-hydroxyl-(3β,25 R)-16,23-epoxy-23,24-imino-cholestan-5,16,20,23(N)-tetraene-3β-ol-3-O-β-D-glucopyranosyl-(1→2)-β-D-glucopyranosyl-(1→4)-β-D-galactop-yranoside(2),Solalyraine A(3),Solalyraine B(4),16,23-epoxy-22,26-imino-cholest-22(N),23,25(26)-trien-3β-ol-3-O-β-D-glucopyranosyl-(1→2)-β-D-glucopyranosyl-(1→4)-β-D-galactopyranoside(5),N-(4-amino-butyl)-3-(3-hydroxy-4-methoxy-phenyl)-E-acrylamide(6)and magnoflorine(7).Among them,compounds 1 and 2 were two new steroidal glycoalkaloids that has not been reported in the literature,and compound 7 was isolated from S.lyratum for the first time.

正交设计法优化白英中总生物碱的提取工艺

目的:探究白英中总生物碱提取工艺,并用正交设计法进行优化。方法:以士的宁为标准对照品,用酸性染料比色法测定总生物碱含量,采取单因素对照实验,考查提取温度、提取时间、乙醇浓度和固液比4个变量,在此基础上再进行正交试验,确定最佳提取工艺条件。结果:当提取温度为70℃,提取时间为6 h,乙醇浓度为90%,固液比为1∶60(g/mL)时,提取效果最佳,提取率为12.19%。结论:该方法优选了白英总生物碱的最佳提取工艺,为白英的进一步开发利用提供了依据。

白英总皂苷的抗炎作用及机制研究

利用脂多糖刺激RAW 264.7细胞(小鼠单核巨噬细胞系)产生的炎性反应,检测了白英总皂苷对RAW264.7细胞的TNF-α、MD2和TLR4蛋白表达的影响,探讨了白英总皂苷在炎性反应中的作用及分子机制.结果表明:白英总皂苷对RAW264.7细胞无毒性作用,但对TNF-α蛋白表达有抑制作用,且这一过程可通过抑制MD2和TLR4蛋白的表达实现.

白英生物碱抗肿瘤作用的机制研究概述

白英具有清热祛湿、消肿解毒的功效,是临床常用的具有抗肿瘤活性的中药之一,对多种恶性肿瘤具有明显抑制作用,尤其对肝恶性肿瘤、肺恶性肿瘤、胃恶性肿瘤、宫颈恶性肿瘤等效果显著。本文通过总结近十年白英生物碱(solanum lyratum thunberg alkaloid,STA)抗肿瘤作用的国内外文献,阐述其抗肿瘤的作用机制,明确STA在临床抗肿瘤治疗的意义。白英具有多种化学成分,如皂甙、有机酸、生物碱等,但STA是白英发挥抗肿瘤作用的主要有效成分。本文将从以下几个方面进行总结:STA通过调控细胞凋亡相关的生存素(survivin)、天冬氨酸特异性半胱氨酸蛋白酶(caspase)、Fas、B淋巴细胞瘤-2(B-cell lymphoma-2,bcl-2)基因和蛋白的表达,发挥促进肿瘤细胞凋亡的作用;通过破坏细胞周期的完整性,抑制肿瘤细胞的增殖;通过调控信号通路,影响肿瘤的发生发展;通过抑制肿瘤新生血管的形成,抑制肿瘤的增殖、侵袭与转移;通过提高细胞的免疫功能,抑制肿瘤的发展进程。

响应面法优化白英中总黄酮提取工艺研究

目的:探究白英中总黄酮提取量的影响因素,优选白英中总黄酮的提取工艺条件。方法:以乙醇浓度、液料比、提取时间为条件进行单因素实验,并采用响应面法进行提取条件的优化。结果:提取工艺最佳条件为乙醇浓度73%,提取时间2.1 h,液料比1∶69,提取率为4.89%。结论:该方法优选了白英总黄酮的最佳提取工艺,为白英的进一步开发利用提供了依据。

白毛藤多糖的分离、纯化和鉴定

目的分离、纯化和鉴定白毛藤S olanum ly ra tum中的多糖的成分。方法经脱脂、沸水抽提、乙醇沉淀、酶-Sevag法脱蛋白得到的粗多糖,再经DEAE-Sepharose fast flow阴离子交换和Sephadex G-200葡聚糖凝胶柱色谱分离得到纯化的白毛藤多糖(SLPS),用红外光谱和紫外光谱分析鉴定该多糖,凝胶渗透色谱(GPC)法测定其相对分子质量,运用薄板色谱(TLC)和纸色谱(PC)法初步测其结构组成。结果红外光谱分析具有典型的多糖特征吸收峰,紫外光谱分析未见蛋白质(280 nm)与核酸(260 nm)的特征吸收峰。结论本实验所提取的白毛藤多糖为单一组分的多糖。

白英乙醇提取物抗肿瘤作用初步研究

目的:研究白英乙醇提取物体内外抗肿瘤作用。方法:采用MTT法观察白英乙醇提取物体外对人肝癌BEL-7402细胞及人胃腺癌SGC-7901细胞的增殖抑制作用;以小鼠S180肉瘤及H22肝癌为模型,观察白英乙醇提取物在体内对肿瘤生长的抑制作用。结果:白英乙醇提取物对人肝癌BEL-7402细胞及人胃腺癌SGC-7901细胞有增殖抑制作用,且均呈现良好的浓度-效应依赖关系,作用48 h的IC50值分别为(287.40±5.84),(176.14±5.18)μg.mL-1;白英乙醇提取物在体内对小鼠S180肉瘤和H22肝癌肿瘤的生长均有显著的抑制作用,且呈现良好的剂量-效应关系,高剂量组的抑瘤率分别达到(41.15±4.54)%和(45.00±7.37)%。结论:白英乙醇提取物具有抗肿瘤作用。

白英提取液对Hela细胞凋亡及Fas/FasL基因表达的影响

目的:探讨白英提取液对Hela细胞凋亡及Fas/FasL基因表达的影响。方法:采用MTT法检测细胞增殖抑制率,流式细胞仪检测凋亡,二步法免疫组化检测Fas蛋白表达,半定量RT-PCR检测Fas和FasL mRNA表达。结果:白英提取液对Hela细胞增殖有较强的抑制作用,且这种作用呈一定的剂量-时间依赖性;细胞凋亡率明显升高;Fas基因mRNA和蛋白表达显著上升,FasL mRNA表达明显下降。结论:白英提取液通过上调Fas基因和下调FasL基因表达,诱导细胞凋亡,从而抑制Hela细胞增殖。

白英总苷抗肿瘤作用初步研究

目的研究白英总苷体内外抗肿瘤作用。方法采用MTT法考察白英总苷体外对人肝癌BEL-7402细胞及人胃腺癌SGC-7901细胞的增殖抑制作用;以小鼠S180肉瘤及H22肝癌为模型,考察白英总苷体内对肿瘤生长的抑制作用。结果白英总苷对人肝癌BEL-7402细胞及人胃腺癌SGC-7901细胞有显著地增殖抑制作用,且均呈现良好的浓度-效应依赖关系,作用48h的IC50值分别为(180.22±6.32)μg/ml和(114.89±4.89)μg/ml;白英总苷对小鼠S180肉瘤有显著的抑制作用,且呈现良好的剂量-效应关系,三次重复实验,高剂量组的抑瘤率达到33.76%±3.24%;白英总苷各剂量对小鼠H22肝癌抑制作用均不显著。结论白英总苷体内外有一定的抗肿瘤作用。

HPLC法测定中药白英中薯蓣皂苷元的含量

目的 用HPLC法测定不同来源自英中薯蓣皂苷元的含量。方法 选用Apollo C18色谱柱（4．6mm×250mm．5μm），流动相为乙腈-水（体积比为90：10），检测波长为210nm，柱温为室温。结果薯蓣皂苷元的质量浓度在0．04～0．20g·L^-1。内与峰面积呈良好线性关系，方法 的平均回收率为104．5％（n=9）；不同来源的白英药材中薯蓣皂苷元的含量有较大差异。结论 方法操作简便、准确、灵敏度高，为白英药材质量控制提供了定性和定量依据。

白英中总黄酮的含量测定

目的：建立白英中总黄酮的含量测定方法。方法：以芦丁为对照，三氯化铝显色，采用分光光度法，在419nm处测定吸光度值，计算白英中总黄酮含量。结果：芦丁的质量浓度在5．48～21．920．g／mL范围内与吸光度值呈良好线性关系（r=0．9996），方法的平均回收率为96．7％（n=9），RSD为1．1％；不同来源的白英药材中总黄酮含量差异较大。结论：本方法操作简便、准确、灵敏、重现性好，可用于白英药材的质量控制。