多黏菌素B血药浓度的测定及其在重症患者中的应用

Determination of polymyxin B concentration in plasma and its application in critically ill patients

目的建立测定多黏菌素B血药浓度的方法并应用于临床。方法血浆样品经5%三氯乙酸溶液蛋白沉淀后,以多黏菌素E2为内标,采用超高效液相色谱-串联质谱(UPLC-MS/MS)法测定多黏菌素B1、B2的质量浓度。以BEH C18为色谱柱,以水(含0.1%甲酸)-乙腈(含0.1%甲酸)为流动相进行梯度洗脱,流速为0.5 mL/min,进样量为10μL。采用电喷雾离子源以多反应监测模式进行正离子扫描,用于定量分析的离子对分别为m/z 603.2→101.2(多黏菌素B1)、595.7→101.1(多黏菌素B2)、578.5→101.1(内标)。采用上述方法测定79例重症患者体内多黏菌素B的血药浓度,记录患者急性肾损伤(AKI)的发生情况并分析多黏菌素B血药浓度与AKI发生的相关性。结果多黏菌素B1、B2检测质量浓度的线性范围分别为200~20000、50~5000 ng/mL(r>0.995),定量下限分别为200、50 ng/mL;日内、日间精密度的RSD均不高于12.06%,平均提取回收率为103.04%~117.44%(RSD≤10.45%),基质效应、稳定性试验的RSD均不高于7.42%。79例患者的多黏菌素B稳态谷、峰浓度分别为(2.54±2.52)、(8.17±5.20)mg/L。在被纳入AKI评价的27例患者中,有18例患者(66.67%)发生AKI;未发生AKI患者的多黏菌素B峰浓度显著低于AKI患者(P<0.05),但两者谷浓度比较差异无统计学意义(P>0.05)。结论所建UPLC-MS/MS法操作简便、灵敏度高,可用于患者体内多黏菌素B血药浓度的检测;患者AKI的发生可能与体内多黏菌素B的峰浓度有关。

OBJECTIVE To establish a method for the determination of polymyxin B concentration in plasma and apply it to clinical practice.METHODS After precipitated with 5%trichloroacetic acid solution,using polymyxin E2 as internal standard,the concentrations of polymyxin B1 and B2 in plasma sample were determined by UPLC-MS/MS.The determination was performed on BEH C18 chromatographic column with water(0.1%formic acid)-acetonitrile(0.1%formic acid)as mobile phase(gradient elution)at the flow rate of 0.5 mL/min.The sample size was 10μL.The detection was accomplished with electrospray ionization operated in positive ion scanning by multi-reaction monitoring mode.The ion pairs for quantitative analysis were m/z 603.2→101.2(polymyxin B1),m/z 595.7→101.1(polymyxin B2)and m/z 578.5→101.1(internal standard).The plasma concentration of polymyxin B in 79 critically ill patients was measured by the above method,the occurrence of acute renal injury(AKI)was recorded and the relationship of polymyxin B concentration in plasma with AKI was analyzed.RESULTS The linear ranges of polymyxin B1 and polymyxin B2 were 200-20000,50-5000 ng/mL(r>0.995),and the lower limits of quantification were 200 and 50 ng/mL,respectively.RSDs of intra‐day and inter‐day precision tests were not higher than 12.06%,the average extraction recovery was 103.04%-117.44%,and RSDs of matrix effect test and stability test were all not higher than 7.42%.Steady state trough and peak plasma concentration were(2.54±2.52)and(8.17±5.20)mg/L for 79 clinical patients using polymyxin B.Eighteen patients out of 27 included patients developed AKI,with an incidence of 66.67%.The peak concentration of polymyxin B of patients without AKI was significantly lower than that of patients with AKI(P<0.05),but there was no significant difference in the trough concentration between two groups(P>0.05).CONCLUSIONS The established UPLC-MS/MS has the advantages of simple operation and high sensitivity,and can be used to monitor the plasma concentration of polymyxin B in patients.The occurrence of AKI is correlated with the peak concentration of polymyxin B.