UPLC-MS/MS法同时测定肾康注射液中7个有效成分

目的：建立一种UPLC-MS/MS法同时测定肾康注射液中7种成分（丹参素、羟基红花黄色素A、毛蕊异黄酮、黄芪甲苷、大黄酸、丹参酮Ⅱ_A、大黄素）的含量。方法：采用Phenomenex Kinetex XB-C₁₈色谱柱（2.1 mm×50 mm，2.6 μm），以甲醇（A）-0.05%甲酸水（B）溶液为流动相进行梯度洗脱（0~3 min，8% A→65% A；3~5 min，65% A→85% A），流速0.3 mL·min^-1，柱温为30℃，进样量20 μL；采用电喷雾离子源进行负离子模式监测，多反应监测模式（MRM）用于定量分析。结果：在5 min内，肾康注射液的7种成分（丹参素、羟基红花黄色素A、毛蕊异黄酮、黄芪甲苷、大黄酸、丹参酮Ⅱ_A、大黄素）被完全分离，峰面积与浓度有良好的线性关系，相关系数（r）均不小于0.998 9。实验精密度、重复性和稳定性良好；平均加样回收率为96.24%~101.2%。5批样品中丹参素、羟基红花黄色素A、毛蕊异黄酮、黄芪甲苷、大黄酸、丹参酮Ⅱ_A和大黄素的含量测定结果分别为12.46~16.70、8.104~10.97、0.092 6~0.115 3、23.19~29.91、9.869~11.22、1.984~2.736和0.324 0~0.372 3 μg·mL^-1。结论：经方法学验证，本方法可用于肾康注射液中丹参素、羟基红花黄色素A、毛蕊异黄酮、黄芪甲苷、大黄酸、丹参酮Ⅱ_A、大黄素的定量测定。除大黄酸和大黄素外，其他5种成分均为肾康注射液中首次测定。

Objective: To develop a method for the simultaneous determination of the danshensu,hydroxysafflor yellow A,calycosin-7-O-β-D-glucoside,astragaloside Ⅳ,rhein,tanshinone Ⅱ_A,and emodin in Shenkang injection by UPLC-MS/MS.Methods: The samples were separated on a Phenomenex Kinetex XB-C₁₈ column(2.1 mm×50 mm,2.6 μm).The mobile phase was composed of methanol(A)and 0.05% aqueous formic acid(B)with gradient elution(0-3 min,8%A→65%A;3-5 min,65%A→85%A).The flow rate was 0.3 mL·min^-1,the column temperature was maintained at 30℃,and the injection volume was 20 μL.Multiple-reaction monitoring(MRM)scanning was employed for quantification with switching electrospray ion source polarity in negative mode.Results: Within 5 min,the seven compounds of Shenkang injection(danshensu,hydroxysafflor yellow A,calycosin-7-O-β-D-glucoside,astragalosideⅣ,rhein,tanshinone Ⅱ_A,emodin) were completely separated.The regression equations showed linear relationships between the peak area and content of each compound.The correlation coefficient(r)was above 0.998 9.The precision,repeatability,and stability of the method were good for all the seven components.The average recovery ranged from 96.24% to 101.2%.The contents of danshensu,hydroxysafflor yellow A,calycosin-7-O-β-D-glucoside,astragaloside Ⅳ,rhein,tanshinone Ⅱ_A,and emodin in 5 samples were 12.46-16.70,8.104-10.97,0.092 6-0.115 3,23.19-29.91,9.869-11.22,1.984-2.736 and 0.324 0-0.372 3 μg·mL^-1,respectively.Conclusion: The established method can be used for the quantitative determination of 7 compounds in Shenkang injection.Among these constituents,danshensu,hydroxysafflor yellow A,calycosin-7-O-β-D-glucoside,astragaloside Ⅳ,tanshinone Ⅱ_A are quantified in the Shenmai injection for the first time.

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