柱切换HPLC法分析尿中吗啡和O^6—单乙酰吗啡的研究

目的建立用柱切换HPLC技术分析尿中吗啡和06-单乙酰吗啡的方法.方法尿样用硼砂缓冲液(pH9.2)稀释后进入预处理柱(YWG-C18,33mm×5.0mm,10μm),用H2O洗去杂质,再用CH3OHH2O(6040)将被分析组分洗脱进入分析柱(Lichrospher(R)100CN,125mm×4.0mm,5μm),分析流动相为CH3OH磷酸盐缓冲液(pH6.86)=2278.紫外检测器波长为286nm.结果尿中吗啡和06--单乙酰吗啡的线性范围分别为50～1 600n/ml和100～1 600n/ml.吗啡和O6--单乙酰吗啡的精密度均小于4%.吗啡和O6-单乙酰吗啡的检测限均为40n/ml.结论用CSHPLC测定尿中吗啡和O6-单乙酰吗啡,方法准确、灵敏、快速、简便.

Column-switching high-performance liquid chromatographic method for the determination of morphine and O6-monoacetylmorphine in urine

OBJECTIVE: To develop a column-switching high-performance liquid chromatographic method for the determination of morphine and O6-monoacetylmorphine in urine. METHODS: Urine samples (1.0 ml) were spiked with 1.0 ml borate buffer, after centrifugation, 1.0 ml of supernate were injected directly into an extraction column (YWG C18 33 mm x 5.0 mm, 10 microns). After a washing step with the extraction mobile phase, the retained morphine and O6-monoacetylmorphine were flushed into the analytical column (Lichrospher 100 CN 125 mm x 4.0 mm, 5 microns) with the mobile phase CH3OH-H2O (60:40). The analytical mobile phase is CH3OH-phosphate buffer (pH6.86) (22:78). The UV detector was set at lambda 286 nm. RESULTS: The method shows excellent linearity from 50 to 1,600 ng/ml for morphine and from 100 to 1,600 ng/ml for O6-monoacetylmorphine. The linear correlation coefficients were > 0.999. The relative standard deviations were < 4%. The limits of detection were 40 ng/ml for both morphine and O6-monoacetylmorphine. CONCLUSION: The method described is sensitive, rapid, reproducible, and simple.