目的:建立UPLC-MS/MS法同时测定天山茶藨茎14个酚类成分的含量,并测定其体外降血糖活性。方法:采用ACQUITY UPLC BEH Shield RP 18色谱柱(100 mm×3.0 mm,1.7 μm),以0.1%甲酸水溶液(A)-甲醇(B)为流动相,梯度洗脱,流速为0.3 mL·min-1,柱温为30 ℃;使用三重四极杆质谱仪,采用电喷雾电离源(ESI),负离子,以多反应监测模式(MRM)扫描,进行定量分析,测定其总酚、总黄酮含量及其抗糖尿病活性(α-葡萄糖苷酶、α-淀粉酶抑制活性)。结果:14个酚类化合物在测定的质量范围内具有良好的线性关系(R2≥0.991 7),稳定性、重复性、精密度试验的RSD≤4.9％,加样回收率范围在91.0%~102.0%(RSD≤4.6%);天山茶藨茎乙酸乙酯部位的总酚、总黄酮及咖啡酸、表没食子儿茶素、表儿茶素、原儿茶酸的含量最高,分别为(965.51±6.85) mg·g-1、(162.04±8.07) mg·g-1、10 452.00 μg·g-1、14 924.00 μg·g-1、3 922.50 μg·g-1、3 546.00 μg·g-1;并表现出最好的α-葡萄糖苷酶、α-淀粉酶抑制作用,IC50分别为(134.81±3.10) mg·L-1、(457.91±11.94) mg·L-1。结论:本实验建立的UPLC-MS/MS方法精确、高效、灵敏,可用于14个酚类成分的定量分析,结合降血糖活性筛选,可以为深入研究天山茶藨的化学成分和生物活性提供科学依据。
Objective: To establish a rapid UPLC-MS/MS method for simultaneous determination of 14 phenols of Ribes meyeri Maxim. stem, and screen their antidiabetic activities. Methods: The separation was performed by ACQUITY UPLC BEH Shield RP 18 column (100 mm×3.0 mm, 1.7 μm). The mobile phase was composed of 0.1% formic acid-water(A) and methanol(B) in gradient elution at a flow rate of 0.3 mL·min-1 and the column temperature was 30 ℃. A triple quadrupole mass spectrometer was equipped with an ESI source. The negative ionization mode was used for the analysis and determination. The multiple reaction monitoring (MRM) was used for quantitative analysis. Their total phenols, total flavonoids content and antidiabetic activity (α-glucosidase, α-amylase inhibitory activities) were determined. Results: The 14 phenols showed good linearity (R2≥0.991 7) in the tested ranges. The precision, repeatability and stability of the method were good for the 14 compounds (RSD≤4.9%). The average recovery varied between 91.0% and 102.0% (RSD≤4.6%). The ethyl acetate fraction had the highest total phenols content of (965.51±6.85) mg·g-1, total flavonoids content of (162.04±8.07) mg·g-1, caffeic acid content of 10 452.00 μg·g-1, epigallocatechin content of 14 924.00 μg·g-1, epicatechin content of 3 922.50 μg·g-1, protocatechuic acid content of 3 546.00 μg·g-1and it showed the highest α-glucosidase and α-amylase inhibitory activity, IC50: (134.81±3.10) mg·L-1 and (457.91±11.94) mg·L-1, respectively. Conclusion: The established UPLC-MS/MS method is accurate, efficient and sensitive. The 14 phenols can be rapidly quantified by this method. These results about quantitative analysis and antidiabetic activities can provide scientific basis for comprehensive study of chemical components and biological activity of Ribes meyeri Maxim..
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