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目的:研究微透析探针的体外回收率与灌流速度和盐酸小檗碱浓度之间的关系,确定回收率的校正方法。方法:利用高效液相色谱法测定透析液中盐酸小檗碱含量,采用Agilent Eclipse XDB-C18(4.6 mm×250 mm,5μm)色谱柱,流动相为乙腈-0.05 mol·L~(-1)磷酸二氢钠溶液(磷酸调节pH为3.0)(35∶65),流速1 m L·min~(-1),检测波长346 nm,柱温30℃。计算不同灌流速度、不同盐酸小檗碱浓度时,正、反透析法测得的探针体外回收率。同时测定固定流速下,不同药物浓度时探针的体内回收率。结果:在体外回收率实验中,将盐酸小檗碱质量浓度定为5.0μg·m L~(-1),灌流速度分别为1、2和3μL·min~(-1)时,透析法测得的回收率依次为(73.6±5.4)%、(60.4±5.8)%和(35.2±6.3)%;反透析法测得的回收率依次为(70.2±7.0)%、(62.6±4.7)%和(32.5±8.3)%。在体内回收率实验中,将灌流速度定为2μL·min~(-1),盐酸小檗碱灌流液浓度分别为0.1、0.5和5.0μg·m L~(-1)时,体内回收率依次为(51.4±3.5)%、(53.2±2.7)%和(52.6±2.4)%。探针回收率与盐酸小檗碱浓度无关,与流速成反比。体外回收率实验中,透析法与反透析法测得的回收率相近。结论:在用微透析法对盐酸小檗碱生物样品取样时,可采用反透析法对探针回收率进行校正。
OBJECTIVE: To study the relationship between the recovery rate of microdialysis probe in vitro, the perfusion rate and the concentration of berberine hydrochloride, and to determine the recovery method. Methods: The content of berberine hydrochloride in dialysate was determined by high performance liquid chromatography. The mobile phase was acetonitrile -0.05 mol·L -1 with Agilent Eclipse XDB-C18 (4.6 mm × 250 mm, 5 μm) Sodium dihydrogen phosphate solution (adjusted to pH 3.0 with phosphoric acid) (35:65) at a flow rate of 1 m L · min -1 with a detection wavelength of 346 nm and a column temperature of 30 ℃. Calculation of different perfusion rates, different berberine concentrations, positive and reverse dialysis measured probe in vitro recovery. Simultaneous determination of in vivo recovery of probes at different drug concentrations at constant flow rates. Results: In the experiment of in vitro recovery, the concentration of berberine hydrochloride was 5.0 μg · mL -1 and the perfusion rates were 1, 2 and 3 μL · min -1, The recoveries were (73.6 ± 5.4)%, (60.4 ± 5.8)% and (35.2 ± 6.3)%, respectively. The recoveries of dialysis were (70.2 ± 7.0)% and (62.6 ± 4.7)% And (32.5 ± 8.3)% respectively. In vivo recovery experiments, the perfusion rate was set at 2μL · min -1, berberine hydrochloride perfusate concentrations were 0.1,0.5 and 5.0μg · m L -1, the recovery rate in turn (51.4 ± 3.5)%, (53.2 ± 2.7)% and (52.6 ± 2.4)%, respectively. The probe recovery rate has nothing to do with the concentration of berberine hydrochloride, inversely proportional to the flow rate. In vitro recovery experiments, dialysis and reverse dialysis measured recoveries similar. CONCLUSIONS: The probe recovery can be corrected by dialysis when samples of berberine hydrochloride are taken by microdialysis.