UPLC-MS/MS法同时测定苦黄注射液中7种成分的含量

Simultaneous Determination of 7 Components in Kuhuang Injection by UPLC-MS/MS

目的:建立同时测定苦黄注射液中苦参碱、槐果碱、大黄素、大黄酸、绿原酸、柴胡皂苷a、芦荟大黄素含量的方法。方法:采用超高效液相色谱-串联质谱法。色谱柱为Phenomenex Kinetex C_(18),流动相为甲醇-0.1%甲酸(梯度洗脱),流速为0.5 mL/min,柱温为20℃,进样器温度为10℃,平衡时间为4 min,进样量为5μL。离子化模式为电喷雾电离,源喷射电压分别为5 500、-4 500 V,雾化气压力为4.14×10~5Pa,加热气压力为4.48×10~5Pa,帘气压力为1.72×10~5Pa,离子源温度为600℃,工作模式为多反应监测模式。结果:苦参碱、槐果碱、大黄素、大黄酸、绿原酸、柴胡皂苷a、芦荟大黄素检测质量浓度线性范围分别为1.25~80.0 ng/mL(r=0.999 3)、1.10~70.0 ng/mL(r=0.999 5)、0.16~10.5 ng/mL(r=0.999 3)、2.61~168 ng/mL(r=0.999 3)、1.50~96.0 ng/mL(r=0.999 3)、1.48~94.5 ng/mL(r=0.999 6)、6.11~391 ng/mL(r=0.999 1);定量限分别为0.061、0.109、0.041、1.313、0.500、0.492、3.055 ng/mL,检测限分别为0.025、0.054、0.016、0.656、0.150、0.148、1.528 ng/mL;精密度、稳定性、重复性试验的RSD≤3%;加样回收率分别为95.45%~99.45%(RSD=1.43%,n=6)、97.50%~101.00%(RSD=1.50%,n=6)、95.67%~101.73%(RSD=2.85%,n=6)、97.17%~100.57%(RSD=1.16%,n=6)、95.19%~98.90%(RSD=1.71%,n=6)、95.38%~103.85%(RSD=3.39%,n=6)、95.50%~101.17%(RSD=1.20%,n=6)。结论:该方法简便、高效、准确、可靠,适用于同时测定苦黄注射液中7种有效成分的含量。

OBJECTIVE： To establish a method for the content determination of matrine, sophocarpine, emodin, rhein, chloro- genic acid, sarkosaponin a, aloe-emodin in Kuhuang injection.METHODS： UPLC-MS/MS was adopted. The determination was per- formed on Phenomenex Kenetix C18 column with mobile phase consisted of methanol-0.1% formic acid （gradient elution） at the flow rate of 0.5 mL/min. The column temperature was set at 20 ℃, and injector temperature was set at 10℃. The equilibrium time was 4 min and sample size was 5 μL. Ionization mode was electrospray ionization with spray voltage of 5 500 V and --4 500 V, atomizing air pressure of 4.14× 105 Pa, heater pressure of 4.48 × 105 Pa, curtain air of 1.72× 105Pa and ion source temperature of 600 ℃ The work mode was multiple reaction monitoring mode. RESULTS： The linear range were 1.25-80.0 ng/mL for matrine （r=0.999 3）, 1.10-70.0 ng/mL for sophocarpine （r=0.999 5）, 0.16-10.5 ng/mL for emodin （r=0.999 3）, 2.61-168 ng/mL for rhein （r=0.999 3） , 1.50-96.0 ng/mL for chlorogenic acid （r=0.999 3） , 1.48-94.5 ng/mL for sarkosaponin a （r=0.999 6） and 6.11-391 ng/mL for aloe-emodin （r=-0.999 1）. The limits of quantification were 0.061,0.109,0.041,1.313,0.500,0.492,3.055 ng/ mL, limits of detection were 0.025,0.054,0.016,0.656,0.150,0.148,1.528 ng/mL. RSDs of precision, stability and reproducibility tests were all no more than 3%. The recoveries of them were 95.45%-99.45% （RSD=1.43%, n=6）, 97.50%-101.00% （RSD= 1.50%,nm-6）, 95.67%-101.73%（RSD=2.85%,n=6）, 97.17%-100.57% （RSD=1.16%,n=6）,95.19%-98.90%（RSD=1.71%, n=6）, 95.38%-103.85% （RSD=3.39% ,n=6）, 95.50%-101.17% （RSD=1.20% ,n=6）, respectively. CONCLUSIONS： The method is simple, effective, accurate, reliable and suitable for simultaneous determination for 7 components in Kuhuang injection.