关键词: 超高效液相色谱-四极杆-飞行时间质谱, 数据非依赖性采集, 靶向筛查, 芪玉三龙汤

Abstract:

Qi-Yu-San-Long decoction (QYSLD) is a classic traditional Chinese medicine prescription consisting of ten types of herbal medicines, including Astragali Radix, Polygonati Odorati Rhizoma, Scolopendra, Pheretima, Solanum nigrum L., Hedyotis diffusa Willd., Coicis Semen, Euphorbia helioscopia L., Curcumae Rhizoma, and Fritillariae Cirrhosae Bulbus, combined in a ratio of 15∶5∶3∶3∶10∶10∶10∶3∶5∶3 by weight. QYSLD has been used to treat non-small cell lung cancer (NSCLC) for over 20 years in clinical practice, and its curative effect is considered credible. However, the chemical constituents of QYSLD have not been revealed because of their complexity, which has significantly hindered the systematic clarification of the efficacy of the materials and quality evaluation. In this study, a reliable strategy based on the data-independent acquisition (DIA) technology of ultra high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) combined with a targeted screening method was established to investigate the chemical components of QYSLD. A 2-μL aliquot from each vial was injected into a Waters ACQUITY UPLC BEH C₁₈ column (100 mm×2.1 mm, 1.7 μm) to separate complex components. The temperature of the column was 35 ℃, and the flow rate was set at 0.2 mL/min. The mobile phase consisted of 0.1% formic acid aqueous solution and acetonitrile. Detection was conducted using an Xevo G2-XS QTOF-MS with a LockSpray capable-electrospray interface. The data for complex components in QYSLD were collected by full-information tandem mass spectrometry (MS ^E) in the positive and negative ion modes. In the MS^E mode, data acquisition was performed using a mass spectrometer by rapidly switching from a low-collision-energy (CE) scan to a high-CE scan during a single LC run. Thus, accurate precursor and fragment ions were collected in a single run, which was helpful for the structural elucidation of multiple components in QYSLD. In addition, systematic information on isolated chemical compounds was collected and distinguished from the ten individual herbs in QYSLD using databases such as China Academic Journals Full-text database (CNKI), PubMed, Web of Science, Medline, and ChemSpider. Accordingly, a self-building library of QYSLD, including the component name, molecular formula, and structure of the components from the herbs, was established. Subsequently, the raw MS^E data of the collected samples and the self-building chemical composition library were imported into a natural product post-processing screening (UNIFI) platform for targeted screening of the chemical components in QYSLD. The parameters for UNIFI platform were as follows: the retention time deviation was ±0.1 min; an error margin of no more than 5×10 ^-6 for the identified compounds was allowed; positive adducts, including [M+H]⁺and [M+Na]⁺, were selected; and negative adducts, including [M-H]^- and [M+HCOO]^-, were selected. The results showed that a total of 166 compounds were initially identified, including 22 saponins, 13 alkaloids, 27 flavonoids, 32 terpenes, 20 amino acids, 16 phenylpropanoids, 9 organic acids, 6 sterols, 6 anthraquinones, and 15 other components. Among them, sixteen components were confirmed unambiguously with the reference substances. To better understand the chemical contribution of individual herbs to the entire decoction, the attributes of each component were summarized. This study provides a foundation for exploring the pharmacodynamic substances of QYSLD.

Key words: ultra high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS), data independent acquisition (DIA), targeted screening strategy, Qi-Yu-San-Long decoction (QYSLD)