【作者】 张初航；

【导师】 孙启时； 王峥涛；

【作者基本信息】 沈阳药科大学 ， 中药学， 2009， 博士

【摘要】 锦灯笼为茄科酸浆属植物Physalis alkekengi L.var.franchetii（Mast.）Makino,具有清热解毒,利咽,化痰,利尿等功效,在我国许多地区都有分布,是常用的清热解毒药,在临床上有很好的疗效。但锦灯笼的质量标准目前尚不完善。本文在综述了酸浆属植物化学成分和锦灯笼药理活性研究的基础上,运用现代色谱分离技术对锦灯笼的化学成分进行了系统的分离,并对其中的酸浆苦味素类成分进行了体外抗肿瘤活性和构效关系的研究。在此基础上,采用HPLC法同时测定了锦灯笼中2种酸浆苦味素类主要成分的含量;同时研究了酸浆苦味素D单体在锦灯笼药材中的稳定性。1锦灯笼化学成分研究利用多种色谱分离技术,从锦灯笼宿萼的乙醇提取物中共分离得到37个化合物,经理化和光谱数据鉴定出其中31个化合物。其中酸浆苦味素类9个,黄酮类10个,酚类5个,苯丙素类3个,其他类型的化合物4个。在已鉴定的31个化合物中,有1个新化合物,即5α-羟基-25,27-二氢-4,7-二去氢-7-脱氧新酸浆苦味素A（5α-hydroxy-25,27-dihydro-4,7-didehydro-7-deoxyneophysalin A）,11个为酸浆属内首次分离得到,1个为种内首次分得。对从锦灯笼中分离得到的酸浆苦味素类化合物进行了体外抗肿瘤活性研究,结果显示:酸浆苦味素类化合物对人肝癌细胞系Bel 7402具有一定的生长抑制作用。其中,化合物PA-9对人肝癌细胞系Bel 7402的抑制率最高,化合物PA-12最低。但是化合物PA-9对人肝正常细胞系LO-2的细胞毒性也较大。利用MTT法,通过用人前列腺癌细胞LNCAP,PC-3进行测试,发现从锦灯笼中分离得到的1个新化合物在不同浓度下都具有细胞生长抑制作用,IC₅₀值分别为:5.266,0.7285μg ml^-1。2锦灯笼的质量评价研究建立了HPLC-DAD法同时测定锦灯笼中2种酸浆苦味素类成分含量的方法,并且测定了20批锦灯笼药材、4批苦蘵、1批天泡果,以及锦灯笼药材不同部位、苦蘵药材不同部位、天泡果药材不同部位和锦灯笼鲜果及干果中2种酸浆苦味素的含量。实验结果表明:锦灯笼药材中酸浆苦味素D是主要的酸浆苦味素类成分,20批锦灯笼药材中酸浆苦味素D的平均含量为2.025mg/g（即0.202%）,该成分可认定是锦灯笼的特征性成分,因此建议药典中应该相应的以这种成分为指标进行质量控制,是比较合理的;苦藏中酸浆苦味素A为主要成分,天泡果中未检出这2种成分。锦灯笼不同部位,宿萼中这2种成分的含量最高;整株植物中,酸浆苦味素D的含量最高,进一步说明锦灯笼中酸浆苦味素D为主要的酸浆苦味素类成分。苦蘵药材中仅检出酸浆苦味素A,且果实和宿萼中的含量最高;天泡果药材中2种酸浆苦味素均未被检出。锦灯笼鲜果中酸浆苦味素D的含量略高于干果。3锦灯笼化学成分的稳定性研究采用化学动力学方法研究了温度、pH值和光照等因素对酸浆苦味素D的化学稳定性的影响。实验结果表明,酸浆苦味素D在酸性、中性、弱碱性条件中均较为稳定,而随着pH值的增大,t_1/2骤然减小,酸浆苦味素D的降解速度加快,即其在强碱性条件下很不稳定。在加热试验中,根据阿累尼乌斯经验公式计算得到酸浆苦味素D的活化能＞100 kJ·mol^-1,说明酸浆苦味素D在12小时内几乎不受到温度的影响。酸浆苦味素D在自然光中的半衰期要小于在紫外光照射下,说明酸浆苦味素D在自然光照射下的降解速度更快,且不适宜长时间光照。因此,建议酸浆苦味素D对照品应保存在避光的环境下,对照品溶液应现配现用。同时对锦灯笼药材中酸浆苦味素D的稳定性进行了研究。实验结果表明,锦灯笼药材30%甲醇提取液中的酸浆苦味素D在酸性、中性、弱碱性溶液中均较为稳定,而在pH＞12.0的溶液中,则完全被破坏,溶液颜色由棕黄色迅速变为亮黄色。95℃以下的加热温度对酸浆药材中的酸浆苦味素D几乎无影响,降解反应活化能Ea为6722.2kJ·mol^-1,说明降解反应很慢,在此条件下锦灯笼药材中的酸浆苦味素D很稳定。自然光及紫外光对锦灯笼药材中的酸浆苦味素D的稳定性无影响。更多还原

【Abstract】 Jindenglong,Calyx seu fructus physalis,a commonly used Chinese herb,is the dried calyxes of Physalis alkekengi L.var.franchetii（Mast.） Makino（Family Solanaceae）.Its action is considered as clearing away heat and toxic material, relieving sore throat,dissipating phlegm,diureses and so on.It is distributed in most areas of China and is used widely as a treatment for different illnesses.Based on the review on the chemical constituents and pharmacological research of Physalis,the chemical constituents of Physalis alkekengi var.francheti and their antitumor activities were researched.On the basis of systematic research on its chemical constituents,the thesis focused on the quality-related topics of Jindenglong, such as（1）quantitative determination of 2 major physalins simultaneously in Jindenglong using HPLC method,（2） the stability study on physalin D.The results were summarized as below.1 Chemical Constituents of Physalis alkekengi var.franchetiBy means of many different chromatographic methods such as silica gel, sephadex LH-20 and ODS column chromatography,37 compounds have been separated from the calyxes of Physalis alkekengi var.francheti and 31 of them were identified by chemical and spectral methods,including 9 physalins,10 flavonoids,5 phenols,3 coumarins and 4 other miscellaneous compounds.Among them,there is one new compound identified as 5α-hydroxy-25,27-dihydro-4,7-didehydro-7-deoxyneophysalin A.11 compounds were isolated from the Physaliphorous for the first time,and 1 compound was found from this plant for the first time.The antitumor activities of the new neophysalin and eight known physalins isolated from the calyxes of Physalis alkekengi var.francheti were tested.It was found that all physalins inhibited cell growth against Bel 7402 cell,in which compound PA-9 was the most potent and PA-11 was the most inpotent.However, compound PA-9 can also inhibite the normal human liver cells LO-2 seriously.The cytotoxic activities of the new compound were evaluated according to MTT assay against the PC-3（human prostate cancer） and LNCaP（human prostate cancer） cell lines.The cytotoxicity data(IC₅₀ values) are 5.266 and 0.7285μg ml^-1, respectively.2 Quality assessments on Physalis alkekengi var.franchetiA novel method has been developed for quantitative determination of 2 major physalins simultaneously by HPLC method in 20 samples of P.alkekengi var. francheti as well as 4 samples of P.angulata,1 sample of P.minima,different parts of P.alkekengi var.francheti、P.angulata and P.minima.3 Stability and kinetics study of physalin DThe chemical dynamics of physalin D was investigated by using HPLC with DAD detection.The influence factors of pH value,temperature and light on the stability of physalin D in solution were carried out respectively.The results indicated that physalin D was degraded when pH value exceeded 10.The optimal stability condition was at normal temperature,pH＜10 and away from light.更多还原