甘草“解百药毒”的胃肠道效应及机制——以四逆汤/调味承气汤为例

Licorice (Glycyrrhizae radix et rhizoma) has been widely utilized to the compatibility of prescriptions in traditional Chinese medicine (TCM) due to its detoxifying effect. The deintoxication mechanism of Licorice through decoction process has been the focus of many studies, however, it is still poorly understood due to the absence of reliable knowledge on intestinal first-pass metabolism. It is known that intestinal first-pass metabolism plays a key role for the orally administered drugs owning to the metabolic enzymes, efflux transporter expressed in epithelial cells, as well as intestinal bacteria. The nature of orally administrationed TCM decoction has prompted a systematic research on intestinal first-pass metabolism of the detoxifying mechanism of licorice. In this proposal, we intend to investigate licorice's attenuation effect of toxicity on intestinal first-pass metabolism in order to understand the scientific basis of deintoxication mechanism of Licorice. Specifically, we will evaluate in vitro and in vivo effects of licorice on the enzymes in phasees I & II metabolism, those of intestinal bacteria and efflux transporter P-glycoprotein (P-gp). Such evaluation will based on the analysis of toxic components in decoctions of Si-Ni Tang (SNT) and Tiaowei-Chengqi Tang (TWCQT), with or without adding licorice. We expect this proposed research to provide insights into the drug-drug interaction between licorice and toxic components, the mechanism of detoxifying effect, and ultimately, provide scientific basis to the compatibility of prescriptions in TCM theory. In addition, this proposed research will demonstrate a novel approach to the exploration of TCMs as well as a means of rational drug use in TCM clinical practice.

甘草因"解百药毒"之功在方剂配伍中广泛应用，其解毒奥秘为现代研究者所共同关注；虽对其进行了大量研究，但缺乏在肠道首过代谢方面的系统研究。由于肠道上皮细胞内代谢酶、药物外排蛋白及肠道菌对药物代谢、转运的特别贡献，使得口服类药物的肠道首过代谢研究受到重视。本研究依据中药汤剂口服用药的特点，重点阐明甘草在"胃肠道受过代谢"中的减毒作用，力求阐释甘草"解百药毒"的科学实质。采用体外模拟、体内验证的研究手段，以四逆汤和调胃承气汤为例，以方中毒烈成分为检测指标，系统研究甘草对肠道主要I、II相代谢酶、肠道菌代谢酶、P-gp的影响以及配伍甘草后毒烈成分的代谢、转运变化，以期阐释甘草在"胃肠道首过代谢"中与配伍中药产生药物间相互作用之"解百药毒"的机制，为中药配伍理论提供科学证据；为中药系统研究提供新的思路；同时也为中医临床合理用药提供参考。

甘草因“解百药毒”之功在方剂配伍中广泛应用。为探索其在胃肠道代谢的减毒作用，本研究依据中药汤剂口服特点，以四逆汤、调胃承气汤为例，对附子/大黄单味药及配伍炙甘草水煎液在人工胃/肠液、人源肠道菌及肠道微粒体、Caco-2细胞模型进行体外模拟胃肠道代谢、转运研究，及瞬时转染人源CAR的LS 174T细胞体外模型验证，首次考察毒性中药配伍炙甘草后目标毒烈性成分在胃肠道代谢、转运的变化。研究发现四逆汤中制附子的苯甲酰乌头碱(benzoylaconine)、苯甲酰中乌头碱(benzoylmesaconine)、苯甲酰次乌头碱(benzoylhypaconine)、次乌头碱(hypaconitine)，干姜烈性成分[6]-姜酚([6]-gingerol)，以及调味承气汤中大黄游离型蒽醌芦荟大黄素(aloe-emodin)、大黄酸(rhein)、大黄素(emodin)、大黄酚(chrysophanol)①在人工胃/肠液中稳定，且配伍炙甘草不影响其稳定性；②炙甘草分别与制附子、干姜配伍，不影响二者毒烈性成分在人源肠道菌中代谢；炙甘草配伍大黄后可抑制游离型蒽醌的肠道菌代谢，同时促进结合型蒽醌番泻苷A(sennoside A)、B(sennoside B)代谢为游离型而增强致泻作用/烈性。③所有毒烈性成分受肠道上皮细胞中CYP3A催化发生I相代谢，生成极性较大产物利于排泄而解毒；除制附子毒性成分外，其他成分受肠道上皮细胞UGTs的催化发生II相代谢，生成极性产物而利于排泄解毒。由于炙甘草经肠道菌代谢后生成的苷元可抑制CYP3A表达，而使毒烈药味配伍炙甘草后受CYP3A催化I相代谢有所抑制而不利于解毒。④肠道上皮细胞外排型转运蛋白P-gp、MRP2参与毒烈性成分转运。炙甘草及其肠道菌代谢产物可提高P-gp蛋白表达而利于毒性成分外排；大黄酸配伍炙甘草可促进大黄酸外排而利于解毒；但制附子配伍炙甘草却促进目标毒性成分吸收，表明可能有摄入型转运蛋白参与制附子毒性成分吸收。上述研究证明，四逆汤、调胃承气汤中制附子、大黄与炙甘草配伍后经口服，炙甘草在肠道菌中的代谢既可抑制肠道CYP3A表达又抑制肠道菌对毒烈性成分的代谢，但同时又促进毒烈性成分的外排而减毒。本研究旨在阐释甘草在“胃肠道首过代谢”中与配伍中药产生药物间相互作用之“解百药毒”的科学实质，为中药配伍理论及中医临床用药提供科学证据和参考。