目的 采用超高效液相色譜串聯(lián)四極桿靜電場(chǎng)軌道阱質(zhì)譜法（UPLC-Q-Exactive Orbitrap-MS/MS）對(duì)五神湯中的化學(xué)成分進(jìn)行鑒定，進(jìn)而對(duì)五神湯中京尼平苷酸、秦皮甲素、咖啡酸、木犀草苷、β-蛻皮甾酮和毛蕊花糖苷 6種成分進(jìn)行含量測(cè)定。方法 UPLC-Q-Exactive Orbitrap-MS/MS 法鑒定五神湯中的化學(xué)成分，采用 ACQUIFY UPLCTMBEH C₁₈ 色譜柱（100 mm×2.1 mm，1.7 μm）進(jìn)行分離，以0.1%甲酸-乙腈為流動(dòng)相進(jìn)行梯度洗脫，檢測(cè)波長(zhǎng)為 254 nm，柱溫30 ℃；電噴霧離子源（ESI）全掃模式；根據(jù)一級(jí)質(zhì)譜中的分子離子，推測(cè)化合物的相對(duì)分子質(zhì)量和元素組成；再利用二級(jí)質(zhì)譜信息，與對(duì)照品及文獻(xiàn)報(bào)道中化合物信息庫(kù)進(jìn)行比對(duì)，推測(cè)五神湯的體外化學(xué)成分。HPLC 法檢測(cè)五神湯中京尼平苷酸、秦皮甲素、咖啡酸、木犀草苷、β-蛻皮甾酮和毛蕊花糖苷，Aglient Eclipse XDB-C₁₈色譜柱（150 mm×4.6 mm，5 mm），流動(dòng)相為0.1%磷酸水（A）-乙腈（B），梯度洗脫，檢測(cè)波長(zhǎng)254 nm，體積流量1.0 mL·min^-1，柱溫30 ℃，進(jìn)樣量10 μL。結(jié)果 共檢測(cè)到五神湯中的71種化學(xué)成分，主要包括黃酮類、酚酸類、三萜皂苷類以及香豆素類；6種成分在濃度范圍內(nèi)線性關(guān)系良好，平均加樣回收率為89.6%～93.0%。京尼平苷酸、秦皮甲素、咖啡酸、木犀草苷、β-蛻皮甾酮和毛蕊花糖苷在10批樣品中的平均質(zhì)量濃度依次為 2.92、2.48、1.72、1.38、2.08、1.77 mg·g^-1。結(jié)論 建立的 UPLC-Q-Exactive Orbitrap-MS/MS、HPLC法可用于五神湯化學(xué)成分的鑒定和含量測(cè)定。

Objective To establish a method for the identification of chemical constituents in Wushen Decoction (WSD) by UPLC-QExactive orbitrap-MS/MS, then, establish a method for the determination of six components of WSD: geniposidic acid, esculin, caffeic acid, luteoloside, β -ecdysterone and verbascoside by HPLC.Methods Identification of chemical components in WSD by UPLC-Q-Empirical Orbitrap MS/MS method, the separation was performed on an ACQUIFY UPLCTMBEH C₁₈ column (100 mm×2.1 mm, 1.7 μm), the detection wavelength was 254 nm and the column temperature was 30 ℃ . Mass spectrum conditions: electrospray ion source (ESI) full sweep mode. The mobile phase was eluted by gradient elution with 0.1% formic acid water and acetonitrile at 254 nm. Infer the relative molecular weight and elemental composition of the compound based on the molecular ions in the primary mass spectrometry. By utilizing the secondary mass spectrometry information and comparing it with the reference material and the compound information library in literature, the in vitro chemical composition of WSD can be inferred. HPLC method was used to detect geniposidic acid, aesculin, caffeic acid, luteoloside, β -ecdysterone and verbascoside, Aglient Eclipse XDB-C₁₈ column (150 mm×4.6 mm, 5 mm), mobile phase consisting of 0.1% phosphoric acid water (A) - acetonitrile (B), gradient elution, detection wavelength of 254 nm, volume flow rate of 1.0 mL·min^-1, column temperature of 30 ℃, injection volume of 10 μL.Results A total of 71 chemical components were detected in WSD, including flavonoids, phenolic acids, triterpenoid saponins and coumarins, according to primary, secondary and control samples by mass spectrometry. The linear relationships of the six components were good in the concentration range, and the average recoveries were 89.6%—93.0%. The average contents of geniposidic acid, aesculin, caffeic acid, luteoloside, β-ecdysterone and verbascoside in ten batches of samples were 2.92, 2.48, 1.72, 1.38, 2.08, and 1.77 mg·g^-1, respectively.Conclusion The established UPLC-Q-Exactive Orbitrap-MS/MS and HPLC method are stable, reliable and can be used for the identification and content determination of the chemical constituents of WSD.

R286.02

遼寧省自然科學(xué)基金面上項(xiàng)目（2021-MS-361）