复方黄柏胶囊定性定量方法的研究

目的建立复方黄柏胶囊定性定量方法。方法采用薄层色谱法对复方黄柏胶囊中所含黄柏、栀子、姜黄、蒺藜进行了定性鉴别研究。采用高效液相色谱法(HPLC)测定复方黄柏胶囊中盐酸小檗碱的含量,色谱柱kromatosil C18,流动相为乙腈-0.03mol/L磷酸二氢钠-磷酸(35:65:0.08),紫外检测波长为265nm,流速为1.0mL/min。结果薄层色谱鉴别方法专属性强,斑点分离良好。复方黄柏胶囊中盐酸小檗碱HPLC测定样品平均回收率为96.5%,RSD=1.6%(n=9)。结论所建立的方法可作为复方黄柏胶囊的质量控制方法。     

左金胃漂浮缓释片中小檗碱、巴马汀、吴茱萸碱和吴茱萸次碱的体外释放规律和机制研究

目的探讨左金胃漂浮缓释片体外多成分释放的规律和机制。方法采用转篮法以人工胃液为介质,以HPLC法测定左金胃漂浮缓释片中小檗碱、巴马汀、吴茱萸碱和吴茱萸次碱在8 h内的体外累积释放率,通过相似因子比较法、Higuchi方程,零级、一级释放方程等释放模型拟合法以及Peppas方程研究左金胃漂浮缓释片多成分释放的规律。结果左金胃漂浮缓释片中4种生物碱的平均释放曲线相互之间的相似因子均大于80%,均以Higuchi释放模型为最佳拟合模型,释放机制均为扩散协同骨架溶蚀作用。结论左金胃漂浮缓释片中生物碱类成分的释放具有均衡缓释性。     

RP-HPLC法测定中药材黄柏中小檗碱的含量

用反相高效液相色谱法,安定为内标,ZorbaxC₈柱,含30mmol/L。磷酸的乙腈-水(35:65)为流动相,于紫外346nm处测定了黄柏中小檗碱的含量。排除了巴马汀对小檗碱的干扰。平均回收率为101.7%,RSD为1.83%(n=20)。     

Oxygen activation by photoexcited protoberberinium alkaloids from Mahonia aquifolium

Protoberberinium salts, i.e. berberine (I), palmatine (II) and jatrorrhizine (III) prepared from Mahonia aquifolium (Pursh) Nutt. belong to isoquinoline alkaloids possessing interesting biological activity (e.g. antibacterial, antimalarial, antitumor). The characteristic UV/Vis absorption band maxima of I-III iodide salts were found in regions 350 and 425 nm in dimethylsulfoxide (DMSO) and ethanol solvents, and were only negligibly influenced by substitution changes on the C-2 and C-3 positions. The fluorescence intensity of protoberberinium salts monitored in ethanol solutions was significantly lowered by iodide counter-ions, and decreased in the order berberine > palmatine > jatrorrhizine. EPR spectroscopy supplied evidence of the formation of super-oxide anion radicals and singlet oxygen upon irradiation of berberine in oxygenated DMSO solvent. The photochemical generation of O(2) (.-) and (1)O(2) in DMSO solutions of palmatine and jatrorrhizine was substantially lower, and probably reflected the replacement of a photolabile methylenedioxy group at C-2 and C-3 positions in the berberine molecule by two methoxy groups in palmatine, and methoxyl (C-2) and hydroxyl (C-3) substitution in jatrorrhizine. Additionally, the powder EPR spectra of protoberberinium iodides I-III measured at 290 K revealed the presence of single-line EPR signals (g(eff) = 2.0044), which were attributed to hydroperoxidic structures produced by the autoxidation process. The photochemical reactions of protoberbenium salts producing reactive oxygen species after UVA excitation should be integrated in biological activity investigations, as well as in their applications in skin disorder treatment.     

黄芪多糖和小檗碱对3T3-L1脂肪细胞糖代谢及细胞分化的影响

目的比较黄芪多糖和小檗碱对脂肪细胞糖代谢和细胞分化的影响,并分析其改善糖代谢的可能机制。方法检测黄芪多糖和小檗碱干预的脂肪细胞对^3H-葡萄糖的摄取,对黄芪多糖和小檗碱干预分化的细胞进行油红O染色并通过比色定量分析脂肪细胞分化程度。逆转录多聚酶联反应(RT—PCR)检测脂肪细胞分化相关基因过氧化物体增殖剂活化受体γ(PPARγ)、CAAT／增强子结合蛋白α(C／EBPα)mRNA的表达。结果黄芪多糖和小檗碱组葡萄糖摄取率分别为正常组的109．3％和182．7％;黄芪多糖明显促进分化及其PPARγmRNA表达,与对照组比较,差异有显著性(P<0．01);小檗碱则明显抑制细胞分化及PPARγ,C／EBPα mRNA表达,与对照组比较,差异有显著性(P<0．01)。结论黄芪多糖促进脂肪细胞葡萄糖摄取及细胞分化和PPARγ mRNA表达,而小檗碱促进葡萄糖摄取但抑制脂肪细胞分化和PPARγ及C／EBPα mRNA的表达。     

Cytotoxic effects of Coptis chinensis and Epimedium sagittatum extracts and their major constituents (berberine, coptisine and icariin) on hepatoma and leukaemia cell growth

1. The present study was conducted to evaluate the cytotoxic effects of Coptis chinensis and Epimedium sagittatum extracts and their major constituents on hepatoma and leukaemia cells in vitro. 2. Four human liver cancer cell lines, namely HepG2, Hep3B, SK-Hep1 and PLC/PRF/5, and four leukaemia cell lines, namely K562, U937, P3H1 and Raji, were used in the present study. 3. Of the two crude drugs, C. chinensis exhibited the strongest activity against SK-Hep1 (IC50 = 7 microg/mL) and Raji (IC50 = 4 microg/mL) cell lines. The IC50 values for C. chinensis on HepG2, Hep3B and PLC/PRF/5 cell lines were 20, 55 and 35 microg/mL, respectively. The IC50 values for C. chinensis on K562, U937 and P3H1 cell lines were 29, 29 and 31 microg/mL, respectively. 4. With the exception of HepG2 and Hep3B, the E. sagittatum extract inhibited the proliferation of all cell lines (SK-Hep1, PLC/PRF/5, K562, U937, P3H1 and Raji), with IC50 values of 15, 57, 74, 221, 40 and 80 microg/mL, respectively. 5. Interestingly, the two major compounds of C. chinensis, berberine and coptisine, showed a strong inhibition on the proliferation of both hepatoma and leukaemia cell lines, with IC50 values varying from 1.4 to 15.2 microg/mL and from 0.6 to 14.1 microg/mL, respectively. However, icariin (the major compound of E. sagittatum) showed no inhibition of either the hepatoma or leukaemia cell lines. 6. The results of the present study suggest that the C. chinensis extract and its major constituents berberine and coptisine possess active antihepatoma and antileukaemia activities.     

春雨烧伤凝胶剂中盐酸小檗碱的含量测定

目的建立春雨烧伤凝胶剂中盐酸小檗碱含量测定的高效液相色谱法。方法色谱柱为Kromasil C18柱(4.6mm×250mm,5μm),流动相为乙腈-1%H3PO4-三乙胺(24∶76∶0.76),检测波长:345nm,流速:1.0mL/min,柱温:30℃。结果盐酸小檗碱在0.088～0.440μg(r=0.9997)之间具有良好的线性关系,平均回收率为98.13%,RSD=0.77%(n=6)。结论该法检测快速,精密度高,重现性好,可用于春雨烧伤凝胶剂的定量分析。     

黄连解毒汤有效成分对缺氧/复氧时脑微血管内皮细胞的保护作用

目的:研究栀子苷、黄芩苷和小檗碱对大鼠脑微血管内皮细胞的保护作用。方法:建立离体大鼠脑微血管内皮细胞缺氧/复氧损伤模型,用1.024,0.512,0.256,0.128,0.064,0.032,0.016,0.008μmol.mL^-1栀子苷,0.224,0.112,0.056,0.028,0.014,0.007,0.003μmol.mL^-1黄芩苷和0.192,0.096,0.048,0.024,0.012,0.006,0.003μmol.mL-1小檗碱分别作用于正常组和模型组细胞。细胞活性用MTT比色法检测。结果:0.128,0.064,0.032μmol.mL^-1栀子苷,0.028,0.014,0.007μmol.mL-1黄芩苷和0.024,0.012,0.006μmol.mL^-1小檗碱能够比较理想的保护缺氧4 h复氧12 h损伤的大鼠脑微血管内皮细胞。结论:适当浓度的栀子苷、黄芩苷和小檗碱等黄连解毒汤主要成分可以保护脑微血管内皮细胞。     

Agonistic effects of berberine hydrochloride on muscarinic receptors 1

用大鼠离体胸主动脉环和豚鼠离体气管条功能实验及放射配基受体结合实验研究盐酸小檗碱（Ｂｅｒ）对组织毒蕈碱受体（Ｍ受体）的作用．结果表明较高浓度的Ｂｅｒ（≥２０μｍｏｌ·Ｌ－１）对有内皮的动脉环具直接舒张作用;在去内皮或阻断Ｍ受体后,其舒张作用被完全抑制．Ｂｅｒ使豚鼠离体气管条产生浓度依赖性收缩,此作用可被阿托品拮抗,其ｐＡ２值为９．８．放射配基受体结合实验显示,Ｂｅｒ对大鼠的唾液腺,大脑皮质和心脏Ｍ受体均有中度亲和力,且使［３Ｈ］－二苯羟乙酸奎宁酯（ＱＮＢ）与３种组织Ｍ受体结合的Ｋｄ值增大,最大结合量不变．其Ｋｉ值分别为（７．６±１．３）,（１．６±０．９）和（０．５８±０．０７）μｍｏｌ·Ｌ－１．结果提示,Ｂｅｒ有Ｍ受体激动作用．     

Berberine modulates expression of mdr1 gene product and the responses of digestive track cancer cells to Paclitaxel.

Berberine is the major constituent of Coptis chinese and is commonly used in Chinese herbal medicine to treat patients with gastrointestinal disorders. In this study, using flow cytometry, we have found that a 24-h berberine treatment up-regulated the multidrug-resistant transporter (pgp-170) expression in two oral (KB, OC2), two gastric (SC-M1, NUGC-3) and two colon (COLO 205, CT 26) cancer cell lines. Decreased retention of rhodamine 123 was observed in berberine-treated cells as compared to vehicle control. To examine whether the berberine modulated pgp-170 expression in cancer cells is associated with changes in drug resistance, we determined the cytotoxicity, cell cycle progression and cell morphology of Paclitaxel-treated cells. Paclitaxel (1 nM-10 microM) treatment for 24 h induced cytotoxicity in OC2, SC-M1 and COLO 205 cells in a dose-dependent manner. Pretreatment of cells with 32 microM berberine for 24 h prior to Paclitaxel treatment resulted in increased viability as compared to that of Paclitaxel-treated cells. In addition, Paclitaxel-induced apoptosis and/or G2/M arrest in these three cancer cell lines. Pretreatment of cells with berberine prior to Paclitaxel blocked the Paclitaxel-induced cell cycle responses and morphological changes. These results together suggest that berberine modulated the expression and function of pgp-170 that leads to reduced response to Paclitaxel in digestive track cancer cells.