甘草次酸类化合物的制备和抗肿瘤活性研究

目的制备18α-和18β-甘草次酸类复合物,并对体外抗癌活性进行研究。方法以18β-甘草次酸(Ⅰ)为起点,进行构型转化得到其光学异构体18α-甘草次酸(Ⅱ),再经C30-位羧基的甲酯化分别得到18β-甘草次酸甲酯(Ⅲ)和18-α-甘草次酸甲酯(Ⅳ),并与环磷酰胺的抗癌活性体内代谢产物——二氯磷酰氮芥偶联而得2个抗癌复合物18β-甘甲磷氮芥(Ⅴ)和18α-甘甲磷氮芥(Ⅵ);用四大光谱法(NMR、MS、IR、UV)对各化合物的化学结构进行鉴定分析;利用人类肝癌细胞株BEL-7402为体外实验模型,用MTT法观察各化合物对人肝癌细胞增殖的抑制活性;抗肿瘤药物顺铂作为阳性对照物。结果对化合物的合成工艺进行优化;18α-甘草次酸的构象转化得率为45%;目标化合物18β-甘甲磷氮芥(Ⅴ)和18α-甘甲磷氮芥(Ⅵ)的产率分别为35%和25%。所制备的甘草次酸衍生物中,化合物Ⅱ和Ⅵ对BEL-7402肿瘤细胞的增殖显示明显强的抑制活性,浓度在5～500μg/mL范围内,对肿瘤细胞增殖的抑制率分别为2.6%～86%及1.3%～50.1%;在相同条件下,对照物顺铂的抑制率为20.0%～73.41%。结论目标化合物的制备中磷酰酯化反应的条件控制(无水、物料比1∶1.25,温度65℃和反应时间3～4h)是合成关键;化合物Ⅱ和Ⅵ在中高浓度时,与顺铂具有较类似的抑制肝癌细胞增殖活性。甘草次酸类化合物的构型转化及与抗癌基团偶联可能提高其抗癌潜力。

Preparation and anticancer property of Glycyrrhetinic Acid Compounds

Objective Two anticancer prodrugs-18α and 18β-Methy Glycyrrhetinic Acid-Phosphorus Nitrogen Mustard Ester(18α and 18β-MGA-PNME,compound Ⅵ and Ⅴ) were prepared and structurally characterized.Methods Firstly,18β-Glycyrrhetinic Acid(18β-GA,compound Ⅰ) was converted into its optical isomer 18α-Glycyrrhetinic Acid(18α-GA,compound Ⅱ,yield 45%),transformed into methyl ester of 18α-GA(Ⅳ,18α-MGA,yield 58%) and of 18β-GA(Ⅲ,18β-MGA,yield 46%).18α-MGA and 18β-MGA were combined respectively with dichloro-phosphorus nitrogen mustard(DPNM),the active metabolite of anticancer drug cyclophosphamide,for preparing target compounds 18α-MGA-PNME(Ⅵ,yield 25%) and 18β-MGA-PNME(Ⅴ,yield 35%).Their chemical structures were determined by four routine spectrometric methods(NMR,MS,IR,UV).The in vitro antitumor activities of these compounds were studied using human hepatocarcinoma cell line Belle 7402.The MTT method was used for evaluating the capacity to inhibit tumor cell proliferation.Cis-platin was used as positive reference.Results Compounds Ⅱ-Ⅵ were obtained by above procedures;their preparation technologies were optimized;The initial biological data showed that the compound Ⅱ and Ⅵ exhibited relatively strong antitumor activities,at a concentration range of 5～500 μg/mL,they inhibited cancer cell proliferation by 2.6～86% and 1.3%-50.1%,respectively.In the same conditions,the reference compound cis-platin inhibited cell proliferation by 20～73.41%.Conclusion The optimal reaction conditions of phosphorus esterification,such as the anhydride condition and the ratio of reactants(1∶1.25,65℃/3-4 h),were the key points for obtaining the target compounds.The biological results suggest that,the compounds Ⅱ and Ⅵ have similar cancer cell inhibiting activities as cis-platin at the middle and higher concentration.Furthermore,the optical conversion and binding with antitumor moiety can significantly increase the antitumor potency of glycyrrhetinic acid derivatives.Their hepato-targeting property should be evaluated with further in vivo animal study.