基于亲和超滤-液质联用结合相关性分析方法筛选青钱柳提取物中α-葡萄糖苷酶抑制剂

Screening ofα-Glucosidase Inhibitors in Cyclocarya paliurus Extracts Based on Affinity Ultrafiltration-Liquid Chromatography-Mass Spectrometry Combined with Correlation Analysis

目的:为快速筛选青钱柳提取物中的α-葡萄糖苷酶抑制剂,采用亲和超滤-液质联用技术结合相关性分析探究其有效成分与酶抑制活性间的关系。方法:运用大孔吸附树脂,建立青钱柳不同浓度乙醇洗脱液中相同化学成分量变化矩阵(X矩阵);分析各乙醇洗脱部位体外α-葡萄糖苷酶抑制作用,构建效应矩阵(Y矩阵);通过偏最小二乘回归法(Partialleastsquaresregression,PLS)预测潜在的活性成分与亲和超滤法(Affinity ultrafiltration,AUF)所筛选的化合物进行比对并通过分子对接进行验证。结果:青钱柳50%乙醇总提取物IC50值为14.8μg/mL,经大孔树脂吸附后的10%~80%浓度乙醇洗脱液IC50为5.4~204.1μg/mL。通过AUF法与相关性分析在正、负离子模式下共鉴定出10种青钱柳潜在的活性成分,其中鉴定出8种三萜类化合物、2种黄酮类化合物。进一步的分子对接结果表明,上述10种活性成分与α-葡萄糖苷酶对接的结合能小于-7.0 kcal·mol^(-1),其中熊果酸内酯的结合能最低(-10.0 kcal·mol^(-1))。结论:AUF与相关性分析相结合能快速筛选出青钱柳提取物中潜在的α-葡萄苷酶抑制剂,本研究结果可为进一步明确青钱柳活性成分奠定基础,为其活性成分质量控制研究、功能性食品开发的深入研究提供依据。

Objective:To rapidly screenα-glucosidase inhibitors in the extract of Cyclocarya paliurus,the affinity ultrafiltration-liquid chromatography-mass spectrometry technique combined with correlation analysis were used to explore the relationship between the active components and enzyme inhibitory activities.Methods:The change matrix(X matrix)of the same chemical component in ethanol elutes of different concentrations of Cyclocarya paliurus was established by using macroporous adsorption resin.The inhibitory effects ofα-glucosidase at each ethanol elution site in vitro were analyzed and the effect matrix(Y matrix)was constructed.Potential active ingredients were predicted by partial least squares regression(PLS)to be compared with compounds screened by affinity ultrafiltration(AUF)and validated by molecular docking.Results:The IC50 value of 50%ethanol extract of Cyclocarya paliurus was 14.8μg/mL,while the IC50 values of the ethanol elution of concentrations ranging from 10%to 80%after adsorption on microporous resin ranged from 5.4μg/mL to 204.1μg/mL.Through AUF combined with correlation analysis,a total of 10 potential active components of Cyclocarya paliurus were identified in both positive and negative ion modes.Among them,8 triterpenoids and 2 flavonoids were identified.Further molecular docking results showed that the binding energy of the 10 active ingredients toα-glucosidase was less than-7.0 kcal·mol^(-1),and the binding energy of ursolic acid lactone was the lowest(-10.0 kcal·mol^(-1)).Conclusion:The combination of AUF with correlation analysis can quickly and effectively screen potentialα-glucosidase inhibitors from Cyclocarya paliurus extracts.The results of this study can lay a foundation for further identifying the active ingredients of Cyclocarya paliurus and provide a basis for further research on the quality control of active ingredients and the development of functional food.