超高效液相色谱法监测重症脑损伤患者血浆中丙泊酚的血药浓度

Monitoring the plasma concentration of propofol in intensive care unit patients with severe brain injury by UPLC

目的:建立测定重症脑损伤患者血浆中丙泊酚浓度的超高效液相色谱-荧光检测方法。方法:血浆样品经乙腈沉淀蛋白后,取上清液进样分析,色谱条件为采用Thermo Acclaim C18色谱柱(150mm×4.6mm,5μm),流动相:左泵-水(含1‰甲酸)-乙腈(V∶V)=40∶60,流速1.0mL·min-1,右泵:0～1min:水-乙腈(V∶V)=90∶10,流速:1mL·min-1;2～4min:水-乙腈(V∶V)=70∶30,流速:1mL·min-1,6～10min:水-乙腈(V∶V)=10∶90,流速:0.3mL·min-1;10.01～16min:水-乙腈(V∶V)=90∶10,流速:0.3mL·min-1;柱温40℃;检测波长276nm;发射波长:310nm;进样量:100μL,sensitivity:6;lamp mode:longlife。采用肝素采血管收集血液样品,血液样品分别于给药前0h,开始给药后1,3,5,10,15,30,60,70,80,90min,停药后1,3,5,10,15,30,60,90,120min从患者给药部位对侧上肢静脉采样1mL。结果:丙泊酚血药浓度在0.025～2μg·mL-l内线性关系良好,R2=0.999 7,回归方程为Y=2.113 5 X-0.016 7,最低检测限为0.01μg·mL-1。低、中、高(0.025,0.5,2μg·mL-1)3个浓度的日内RSD分别为5.7%、1.37%和2.85%,日间RSD分别为14.15%、11.25%和9.21%,平均方法回收率分别为103.46%、96.16%和99.5%。监测10例重症脑损伤患者丙泊酚血药浓度,稳态浓度范围为0.25～0.81μg·mL-1。结论:本方法操作简单快速、灵敏、准确度高,无介质效应影响,所需血浆样本量少,可作为患者丙泊酚治疗浓度的常规监测方法。通过监测重症脑损伤患者丙泊酚血药浓度发现,重症脑损伤患者单纯镇静时,给药速度为20mg·h-1均可满足临床需要。

OBJECTIVE To establish a sensitive method for monitoring the plasma concentration of propofol in intensive care unit patients with severe brain injury by ultra performance liquid chromatography with fluorescence detection. METHODS The separation was performed with a Thermo Acclaim C18 column (150 mm×4.6 mm, 5 μm) using a binary gradient elution of a mixed solution water and acetonitrile, 0-1 min (90:10, V:V), flow velocity was 1 mL·min^-1; 2-4 min(70:30, V:V), flow velocity was 1 mL·min^-1; 6-10 min (10:90, V:V), flow velocity was 0.3 mL·min^-1; 10.01-16 min (90:10, V:V), flow velocity was 0.3 mL·min^-1, and the detection and emission wavelength of detector was set at 276 nm and 310 nm. Left pump:water (including 1‰ formic acid):acetonitrile (V:V)=40:60, flow velocity was 1 mL·min^-1. The column temperature was set at 40℃, injected sample volume was 100 μL, lamp mode was set at longlife. Venous blood samples were collected before dosing and 1, 3, 5, 10, 15, 30, 60, 70, 80, 90 min after the beginning of infusion and 1, 3, 5, 10, 15, 20, 30, 60, 70, 80 min after the termination of propofol infusion. Blood was collected into heparinized tubes and centrifuged immediately. Plasma was stored at 4℃. RESULTS A good linearity was obtained over the range of 0.025-2.00 μg·mL^-1, the regression equation was Y=2.113 5X-0.016 7, R2=0.999 7. The inter-and intra-day RSDs of 0.025, 0.5, 2 μg·mL^-1 were 5.7%, 1.37%, 2.85% and 14.15%, 11.25%, 9.21% respectively. The average recovery rates were 103.46%, 96.16% and 99.5%, respectively. The steady concentration of propofol in 10 patients with severe brain injury was 0.25-0.81 μg·mL^-1. CONCLUSION The method provides a sensitive, rapid and specific analytical procedure for the therapeutic drug monitoring of propofol in clinical and phamacokinetic studies. It conforms to ethical requirements and only 200 μl of blood is needed. The study shows that the patient maintains calm when the propofol speed is 20 mg·h^-1.