A Study on Isolation of Chemical Constituents from Sophora Flavescens Ait. and Their Anti-Glioma Effects

Background

Sophora flavescens Ait. is a traditional Chinese medicine with a long history in China. It is mainly used in the treatment of heat dysentery and similar ailments in the clinical. The objective of this paper was to isolate, purify and identify alkaloids from Sophora flavescens Ait. and to explore their inhibitory effects on C6 glioma cells.

Materials and Methods

Column chromatography, extraction and NMR spectroscopy were used to structurally identify the isolated compounds. MTT assay and flow cytometry were used to detect the inhibitory effect of matrine on C6 cells.

Results

Three compounds were isolated from Sophora flavescens Ait., namely matrine, oxymatrine and lupeol. Different concentrations of matrine solution all had inhibitory effects on growth of C6 cell lines, which showed apparent dose-effect relationship. Compared with the control group, proportion of G0/G1 phase cells increased in each matrine concentration group to a maximum of 79.8%; proportion of S phase cells reduced, and proportion of G2/M phase cells declined slightly to a minimum of 6.3%, suggesting that after the action of matrine proliferation of C6 cells was significantly inhibited and the cells were arrested in the G1 phase.

Conclusion

We concluded that Sophora flavescens Ait. has an inhibitory effect on C6 cell proliferation.     

Extraction and Isolation of Alkaloids of Sophora Alopecuroides and Their Anti-Tumor Effects in H22 Tumor-Bearing Mice

Background

Alkaloids of Sophora alopecuroides have good biological activity, and are widely used in clinical settings, which not only have pharmacological activities of anti-cancer, cancer suppression, as well as the inhibition, and killing of various microorganisms; but also possess extensive pharmacological effects on immune system, nervous system and cardiovascular system. The objective of this paper was to extract and isolate total alkaloids of Sophora alopecuroides (TASA), and to study their anti-tumor effects in H22 tumor-bearing mice.

Materials and Methods

TASA were extracted and isolated using thin-layer chromatography, and column chromatography; and the isolated compounds were analyzed using nuclear magnetic resonance. The inhibitory effects of TASA on tumor in H22-bearing mice were determined by MTT assay.

Results

Three compounds were isolated from Sophora alopecuroides L., which were matrine, oxymatrine and sophoridine, respectively. Meanwhile, mouse H22 sarcoma model was established and different doses of TASA apparently inhibited solid H22-tumor in mice; it inhibited the thymus, and spleen to some extent; the degree of inhibition was more obvious for the spleen.

Conclusion

TASA has an anti-tumor effect in H22 tumor-bearing mice.     

海桐花蕾挥发油的GC-MS分析

目的:提取海桐花蕾中的挥发性成分。方法:采用固相微萃取法和水蒸气蒸馏法,并用GC-MS测定其化学成分,分别鉴定了67种和41种成分,并用归一化法测定其相对百分含量。结果:与水蒸气蒸馏法相比,固相微萃取法得到的十一烷、壬烷、α-荜澄茄油烯等的含量增加;同时提取出了水蒸气蒸馏法不能蒸馏出来的一些成分,如:α-胡椒烯、α-金合欢烯、β-月桂烯、马鞭草烯等。结论:采用固相微萃取法和水蒸汽蒸馏法,提取挥发油的时间明显缩短     

苦豆子总碱舒张家兔离体主动脉的研究

目的:研究苦豆子总碱(total alkali Sophora alopecuroids L,TASa)舒张家兔离体主动脉血管的作用机制.方法:采用离体恒温灌流浴槽,以去甲肾上腺素(noradrenaline,NA)预收缩后,给予TASa(5,10,20,40 mg·L~(-1))观察其舒张血管的量效关系;将标本分为对照组、TASa组、TASa+吲哚美辛(1×10-5mol·L~(-1))、TASa+普萘洛尔(1×10~(-5) mol·L~(-1))、TASa+左旋硝基精氨酸(1×10~(-4)mol·L~(-1))和TASa+去内皮细胞6组探讨TASa舒张血管的机制;用TASa(40 mg·L~(-1))温育标本后,观察TASa对NA(1×10~(-8)～1×10~(-5) mol·L~(-1)),KCl(6.3～100 mmol·L~(-1))和CaCl_2,(1×10~(-5)～1×10~(-2) mol·L~(-1))量效收缩曲线的影响.结果:TASa能舒张主动脉血管,有量效依赖性(r=0.94,P<0.01);去内皮、左旋硝基精氨酸、吲哚美辛和普萘洛尔对TASa舒张血管的作用无明显影响;TASa能压低NA,KCl和CaCl_2的量效收缩曲线.结论:TASa可能是抑制内质网Ca~(2+)释放和拮抗Ca~(2+)通道,降低胞浆Ca~(2+)而使血管舒张.     

HPLC法测定苦参生物碱部位中4种成分含量

目的建立苦参生物碱部位中4种成分氧化苦参碱、氧化槐果碱、苦参碱和槐果碱的HPLC含量测定方法。方法采用Agilent Zorbax C18柱(4.6 mm×150 mm,5μm),系统A为10 mmol/L醋酸铵水溶液(0.1%氨水,pH=9.2),系统B为20 mmol/L醋酸铵的甲醇-乙腈(1∶1)混合溶液,梯度洗脱,洗脱流速为0.8 mL/min,检测波长220 nm。结果氧化苦参碱、氧化槐果碱、苦参碱、槐果碱平均回收率分别为101.43%、99.08%、102.59%和102.06%,RSD分别为1.30%、0.94%、1.69%和1.10%。结论3批样品测定结果表明,该方法简便、准确,可用于苦参生物碱部位中4种生物碱成分的含量测定。     

白刺花生物碱的体外抑瘤作用

报道了从白刺花种子中分得的６种生物碱单体对人体１１种癌瘤株体外抑瘤作用。结果表明，该类生物碱对白血病瘤株有较强的抑制作用，对表皮癌瘤株也有一定的抑制作用，而且抑瘤作用呈现出较强的选择性。     

苦参总碱抗柯萨奇B病毒作用机理的研究

在证实苦参具有抗柯萨奇Ｂ病毒（ＣＶＢ）作用的实验基础上，作者以纯化ＣＶＢ３作为病毒蛋白质对照，通过空斑测定及病毒蛋白质Ｌ－３５Ｓ－甲硫氨酸标记等方法，了解到纯化苦参总碱抗ＣＶＢ的作用机理是不影响病毒吸附、穿入，但它能进入细胞内影响病毒的生物合成，主要表现为抑制病毒蛋白质的合成。     

苦参抗白血病作用及其机制

采用CFU GM和白血病CFU L集落培养、NBT/MTT还原试验和DNA的片段化分析法 ,观察了苦参对正常人骨髓、脐带血粒 巨噬系造血祖细胞 (CFU GM)和人急性髓系白血病HL 6 0细胞增殖、分化、凋亡的影响。结果表明 :5 ,10 ,15 ,2 0 μg·μl-1苦参均能明显抑制HL 6 0细胞增殖并诱导其凋亡 ,呈浓度 效应关系 ;其中 15 μg·μl-1苦参诱导HL 6 0细胞分化作用明显 ,且 5～ 15 μg·μl-1苦参对正常人造血祖细胞无明显的细胞毒作用。提示苦参在一定浓度范围内有选择性抗白血病效应。此结果为进一步研究和开发苦参并应用于肿瘤临床提供了实验依据。     

阳离子交换树脂酸水洗脱法纯化苦参总碱的工艺优选

目的优选阳离子交换树脂不同浓度盐酸溶液洗脱纯化苦参总碱的最佳工艺条件。方法以苦参碱、氧化苦参碱和总生物碱的洗脱率为考察指标,以盐酸溶液作为洗脱剂,优选阳离子交换树脂纯化苦参总碱的最佳工艺和参数条件。结果苦参总碱阳离子交换树脂纯化洗脱条件为:以5%盐酸溶液作为洗脱剂,以6 BV.h-1的流速洗脱,苦参碱和氧化苦参碱、苦参总碱洗脱率均可达85%。结论采用阳离子交换树脂酸水洗脱法纯化苦参总碱效率高,工艺简便,可用于苦参总碱的提取纯化。     

HPLC法测定苦参总碱中苦参碱与氧化苦参碱的含量

目的:采用高效液相色谱法测定苦参总碱中苦参碱和氧化苦参碱的含量。方法:采用 Elite Hypersil NH_2柱(5μm,250mm×4.6 mm),乙腈-无水乙醇-3%磷酸溶液(81:10:9)为流动相,流速:1.0 mL·min~(-1),检测波长220 nm,柱温:25℃,进样量:10 μL。结果:苦参碱、氧化苦参碱的线性范围分别为5.32～53.2μg·mL~(-1)(r=0.9997),65.7～525.6μg·mL~(-1)(r=0.9995)。苦参碱平均加样回收率为99.5%,RSD=1.0%(n=6);氧化苦参碱的平均加样回收率为99.7%,RSD=0.6%(n=6)。结论:本方法操作简便、快速、可靠,可用于控制苦参总碱质量。