Li Xue-Yan, Liu Yang, Liu Fang, Chen Hong-Jiao, Yang Wen-Ning, Yang Hai-Yang, Jiang Xiao-Quan, Sen Mu-Li, Wang Guo-Peng, Wang Jing, Pan Yan-Li
School of Chinese Materia Medica, Beijing University of Chinese Medicine Beijing 102488, China.
Zhongcai Health(Beijing) Biological Technology Development Co., Ltd. Beijing 101503, China.
Zhongguo Zhong Yao Za Zhi. 2021 Apr;46(7):1822-1831. doi: 10.19540/j.cnki.cjcmm.20201230.501.
Based on the target occupancy mathematical model, the binding kinetic process of potential active ingredients of lowering uric acid in Chrysanthemum morifolium with xanthine oxidase(XOD) was evaluated. The potential active ingredients of lowering uric acid in Ch. morifolium were screened by UPLC-Q-Exactivems MS technology, reference substance identification and in vitro enzymatic kinetics experiments. The binding kinetic parameters of xanthine oxidase and potential inhibitor in Ch. morifolium were determined by surface plasma resonance(SPR). The verified mathematical model of the XOD target occupancy evaluated the kinetic binding process of inhibitors and xanthine oxidase in vivo. According to UPLC-Q-Exactive MS and reference substance identification, 39 potential uric acid-lowering active ingredients in Ch. morifolium extracts were identified and the inhibitory activities of 23 compounds were determined. Three potential xanthine oxidase inhibitors were screened, namely genistein, luteolin, and apigenin. whose IC_(50 )were 1.23, 1.47 and 1.59 μmol·L(-1), respectively. And the binding rate constants(K_(on)) were 1.26×106, 5.23×105 and 6.36×105 mol·L(-1)·s(-1), respectively. The dissociation rate constants(K_(off)) were 10.93×10(-2), 1.59×10(-2), and 5.3×10(-2 )s(-1), respectively. After evaluation by different administration methods, the three selected compounds can perform rapid and sustained inhibition of xanthine oxidase in vivo under combined administration. This study comprehensively evaluated the target occupancy process of three effective components in different ways of administration in vivo by UPLC-MS, concentration-response method, SPR technology and xanthine oxidase target occupancy model, which would provide a new research idea and method for screening active ingredients in traditional Chinese medicine.
基于目标占有率数学模型，评估了菊花中潜在降尿酸活性成分与黄嘌呤氧化酶（XOD）的结合动力学过程。采用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱联用（UPLC-Q-Exactivems MS）技术、对照品鉴定和体外酶动力学实验，对菊花中潜在的降尿酸活性成分进行筛选。通过表面等离子体共振（SPR）技术测定菊花中黄嘌呤氧化酶与潜在抑制剂的结合动力学参数。经验证的XOD目标占有率数学模型评估了抑制剂与黄嘌呤氧化酶在体内的动力学结合过程。根据UPLC-Q-Exactive MS和对照品鉴定，鉴定出菊花提取物中39种潜在的降尿酸活性成分，并测定了23种化合物的抑制活性。筛选出3种潜在的黄嘌呤氧化酶抑制剂，即染料木黄酮、木犀草素和芹菜素，其半数抑制浓度（IC₅₀）分别为1.23、1.47和1.59 μmol·L⁻¹。结合速率常数（Kₒₙ）分别为1.26×10⁶、5.23×10⁵和6.36×10⁵ mol·L⁻¹·s⁻¹。解离速率常数（Kₒբ）分别为10.93×10⁻²、1.59×10⁻²和5.3×10⁻² s⁻¹。经不同给药方式评估，所选的3种化合物联合给药时可在体内对黄嘌呤氧化酶进行快速且持续的抑制。本研究通过UPLC-MS、浓度-效应法、SPR技术和黄嘌呤氧化酶目标占有率模型，以不同方式全面评估了3种有效成分在体内不同给药方式下的目标占有率过程，为中药活性成分筛选提供了新的研究思路和方法。
