银杏叶提取物HPLC指纹图谱研究及共有峰的LC-MS鉴定

目的：建立银杏叶提取物的HPLC指纹图谱，为银杏叶提取物掺假鉴别提供依据。方法：采用HPLC梯度洗脱建立银杏叶提取物的指纹图谱，以0.1%甲酸溶液（A）-乙腈（B）为流动相，梯度洗脱（0~40 min，10% B→36% B；40~45 min，36% B→50% B；45~50 min，50% B→100% B；50~55 min，100% B；55~56 min，100% B→10% B；56~65 min，10% B），流速1 mL·min^-1，检测波长254 nm，柱温30℃。并运用“中药色谱指纹图谱相似度评价系统”对不同批次的银杏叶提取物进行相似度评价；采用LC-MS方法对共有峰进行鉴定。结果：建立了银杏叶提取物的特征指纹图谱，标定了20个共有峰，并结合LC-MS分析，确定了10个峰对应的化合物，分别为6-羟基犬尿喹啉酸、3，4-二羟基苯甲酸、芦丁、异槲皮素、山柰酚-3-O-芸香糖苷、异鼠李素3-O-芸香糖苷、槲皮苷、槲皮素、山柰酚和异鼠李素。疑似掺假提取物的色谱图与对照指纹图谱相比芦丁（峰6）响应低，而槲皮素（峰18）、山柰酚（峰19）、异鼠李素（峰20）响应值高，两者差异极大。结论：本文建立的银杏叶提取物指纹图谱对银杏叶提取物掺假评判具有指导意义。

Objective: To establish an HPLC method for the fingerprint analysis of Ginkgo biloba extracts (GBE).Methods: The analysis was carried on Agilent Eclipse XDB-C₁₈(4.6 mm×250 mm, 5 μm)column with mobile phase consisting of 0.1% formic acid-water solution(A), and the gradient elution(B)(0-40 min, 10%B → 36%B; 40-45 min, 36%B → 50%B; 45-50 min, 50%B → 100%B; 50-55 min, 100%B; 55-56 min, 100%B → 10%B;56-65 min, 10%B)was used.The flow rate was 1 mL·min^-1.The detection wavelength was set at 245 nm and the column temperature was 30℃.The HPLC fingerprints of GBE were established by analyzing 10 batches of GBE with "Similarity Evaluation System for Chromatographic Fingerprint of TCM", and the result was used to compare with the HPLC fingerprint of adulterated extracts.Meanwhile, the 10 common peaks were analyzed by LC-MS.Results: The mutual mode of GBE HPLC fingerprint was set up with 20 common peaks and the structural identification of 10 common peaks was performed by LC-MS.The 10 compounds were 6-hydroxy kynurenic acid, 3, 4-dihydroxybenzoic acid, rutin, isoquercetin, kaempferol-3-O-rutinoside, 3-O-rutinoside, quercitrin, quercetin, kaempferol and isorhamnetin.Compared with the reference fingerprint, the batches of adulterated extracts had low peak 6(rutin)response and high peak 18-20(quercetin,kaempferol,isorhamnetin), and there was significant difference between them.Conclusion: The established fingerprints of GBE have direct guiding significance for identification of adulterants in GBE.

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