目的 基于超高效液相色譜-軌道阱高分辨質(zhì)譜（UPLC-HR-Orbitrap-MS）和親和超濾技術(shù)，篩選柴胡中的神經(jīng)氨酸酶抑制劑，尋找具有神經(jīng)氨酸酶抑制活性的單體并進(jìn)行體外驗(yàn)證。方法 建立柴胡提取物的UPLC-HR-Orbitrap-MS分析方法，結(jié)合親和超濾實(shí)驗(yàn)篩選柴胡中能夠與神經(jīng)氨酸酶特異性結(jié)合的活性單體成分，通過質(zhì)譜解析及對(duì)照品驗(yàn)證確定成分，對(duì)篩選的活性單體進(jìn)行體外神經(jīng)氨酸酶抑制檢測(cè)，并采用分子對(duì)接分析潛在的對(duì)接位點(diǎn)。結(jié)果 在柴胡中共篩選出能夠與神經(jīng)氨酸酶特異性結(jié)合的單體化合物10個(gè)，包括皂苷類、黃酮類和酚酸類，其中6''-O-柴胡皂苷A等的神經(jīng)氨酸酶體外抑制活性為49.53%～78.07%，分子對(duì)接結(jié)果與體外神經(jīng)氨酸酶結(jié)果存在差異。結(jié)論 親和超濾-液質(zhì)聯(lián)用技術(shù)能夠快速篩選中藥中特異性活性成分，為中藥活性標(biāo)志物的發(fā)現(xiàn)提供新思路。

Objective Based on ultrahigh performance liquid chromatography Orbitrap high resolution mass spectrometry (UPLC-HR-Orbitrap-MS) and affinity ultrafiltration technology, the monomers with neuraminidase inhibitory activity in Bupleuri Radix were screened, and then to verify in vitro. Methods The UPLC-HR-Orbitrap-MS analysis method of the extract of Bupleuri Radix was established. The active monomers in it which can specifically bind to neuraminidase were screened by affinity ultrafiltration. The components were determined by mass spectrometry and control product verification and the neuraminidase inhibitions of the selected active monomers were detected in vitro, and the potential docking sites were analyzed by molecular docking. Results A total of ten monomers, including saponins, flavonoids and phenolic acids, were screened for specific binding to neuraminidase in Bupleuri Radix. Among them, the neuraminidase inhibitory activities of 6''-O- saikosaponin A and others ranged from 49.53% to 78.07% in vitro, and the molecular docking results were different from those of neuraminidase in vitro. Conclusion Affinity ultrafiltration-LC-MS can rapidly screen the specific active components in traditional Chinese medicine, which provides a new idea for the discovery of active markers of traditional Chinese medicine.

R284.1

國家自然科學(xué)基金面上項(xiàng)目（81774173）;山東省高等學(xué)校青創(chuàng)人才引育計(jì)劃（活性天然產(chǎn)物發(fā)現(xiàn)與開發(fā)研究團(tuán)隊(duì)）;山東省中醫(yī)藥科技發(fā)展計(jì)劃（2019-0019，2019-0021）