甘草制远志减缓“戟人咽喉”毒性与增效作用机理研究

Glycyrrhiza decoction boiled Polygalae Radix can reduce the toxicity in throat, benefit intelligence and calm the nerve system, but the processing mechanism is not clear. Our previous study has drawn three conclusions. First of all, in Glycyrrhiza decoction boiled Polygalae Radix, the contents of many kinds of oligosaccharide ester glycosides and saponins have reduced, but the contents of glomeratose A, sinapinic acid and tenuifolin have increased significantly. Secondly, the steam produced during the process of preparing contains benzoic acid, a toxic and irritant hydrolysate. At last, benzoic acid can inhibit the absorption of sinapinic acid in the stomach and intestine of rats. Based on the conclusions above, we deduce that in the process of preparing for Glycyrrhiza decoction boiled Polygalae Radix, the oligosaccharide ester glycosides and saponins can be hydrolyzed into their secondary glycosides or aglycones. The active hydrolysis products are more beneficial to be absorbed, and the benzoic acid which can inhibit the absorption of active components may volatilize in the process. This project plans to obtain the characteristic constituents which are changed during the process of Glycyrrhiza decoction boiled in terms of both quality and quantity by HPLC fingerprint tracking and modern chromatographic methods, then elucidate their structures by spectroscopic methods such as MS and NMR . Furthermore, we are going to study the changing regulation and transforming mechanisms of these obtained components during the process of preparing, and the absorption and metabolic differences in rats in vivo, using the HPLC, HPLC-MS/MS，analog processing technology and so on to. Additionally, we will explore the influence of benzoic acid on the absorption of active components of Glycyrrhiza decoction boiled Polygalae Radix in vivo. Based on the studies above, we may reveal the mechanism on toxicity attenuating and effect enhancement for Glycyrrhiza decoction boiled Polygalae Radix and provide the scientific evidences for the processing technics and quality standard of Polygalae Radix.

甘草制远志可减缓其“戟人咽喉”毒性，增强安神益智作用，但其炮制机理尚未阐明。我们前期研究发现，远志经甘草煮制后，多种寡糖酯苷和皂苷成分含量降低，而球腺糖A、芥子酸及细叶远志皂苷含量升高；甘草煮远志产生的蒸汽中含有毒性、刺激性成分苯甲酸；苯甲酸可抑制芥子酸在大鼠胃和小肠的吸收。据此推测“甘草煮远志可使其部分寡糖酯苷和皂苷类成分水解，生成其次级苷或者苷元；活性水解物更利于机体吸收，而苯甲酸能抑制活性成分的吸收但其在煮制过程中升华”。本课题拟运用HPLC指纹图谱跟踪及多种柱色谱分离技术，制备远志甘草煮制前后质变与量变特征成分，经MS、NMR等技术鉴定其结构；采用HPLC、HPLC-MS/MS及模拟炮制等技术，研究该类成分在甘草煮制过程中的变化规律、转化机制及在大鼠体内吸收与代谢差异；探讨苯甲酸对远志活性成分体内吸收的影响；揭示甘草制远志减毒增效的机理，为制定远志炮制工艺和质量标准提供科学依据。

甘草制远志可减缓其“戟人咽喉”毒性，增强安神益智作用，但其炮制机理尚未阐明，致使其炮制工艺和质量标准缺乏专属性科学依据。本项目采用HPLC-TOF/MS、GC-MS、柱色谱分离、HPLC及模拟炮制等技术，研究远志甘草汁煮制过程中质量与量变特征成分的实质、变化规律及转化机制，分析远志炮制质变特征化合物在大鼠胃和小肠中的代谢物；采用HPLC-MS/MS 法研究远志甘草汁煮制质变与量变特征化合物的生物利用度，探讨苯甲酸对远志活性有机酸生物利用度的影响；分析远志不同化学部位生制品的成分差异，研究远志不同化学部位生制品及苯甲酸对豚鼠的毒性和咽喉刺激性。结果从远志中分离了11种化合物，鉴定了其结构；远志经甘草汁煮制后寡糖酯（糖酯、寡精）和皂苷类成分产生质变，有机酸类成分产生量变；远志中含有酯苷键的寡糖酯类成分性质不稳定，经甘草汁煮制后可使其酯苷键水解生成其次级苷和（或）苷元，水解生成的小分子有机酸（芥子酸、阿魏酸、3,4,5-三甲氧基肉桂酸）具有较强的生物活性，且分子量变小，较其原苷生物利用度提高，更易于机体吸收，从而达到炮制增效的目的；炮制水解的寡糖酯类成分在大鼠体内也可代谢为其次级苷或者苷元；远志皂苷和寡糖酯部位为远志毒性和咽喉刺激性部位，二者经模拟炮制后毒性和刺激性均降低；远志寡精和部分皂苷类成分经模拟炮制后可水解生成苯甲酸，苯甲酸具有毒性和咽喉刺激性，可抑制阿魏酸和3,4,5-三甲氧基肉桂酸的吸收，但苯甲酸具有升华的性质，可在煮制过程中升华。远志炮制后毒性和刺激性降低分析主要是由于在炮制过程中远志寡精类成分水解生成苯甲酸，苯甲酸在煮制过程中升华及皂苷类成分水解所致。本课题创新性地从毒性/活性成分转化、吸收代谢及药理学角度揭示了甘草制远志减毒增效机理，丰富了中药炮制减毒增效理论，为规范远志的炮制工艺和制定专属性远志饮片质量标准提供了科学依据，并且对远志的临床应用具有重要的指导意义。