二维超高效液相色谱-串联质谱法测定尿液中8-羟基脱氧鸟苷

Two-Dimensional UHPLC-MS/MS Determination of 8-Hydroxydeoxyguanosine in Urine

尿液样品离心后经WATERS C18固相萃取柱净化,收集洗脱液。采用二维超高效液相色谱-串联质谱法测定样品溶液中8-羟基脱氧鸟苷的含量。以Waters BEH C18色谱柱(一维色谱)和Waters BEH HILIC色谱柱(二维色谱)为固定相,以0.1%(体积分数,下同)甲酸溶液与乙腈(体积比8∶2)组成的混合液(一维色谱)和0.1%甲酸溶液与乙腈(体积比9∶1)组成的混合液(二维色谱)为流动相,串联质谱分析中采用电喷雾离子源正离子模式和多反应监测模式。采用内标法定量。8-羟基脱氧鸟苷的线性范围为0.25~300.0 ng·g^(-1),检出限(3S/N)为0.10 ng·g^(-1)。以空白样品为基体进行加标回收试验,所得回收率为96.6%~103%,测定值的日内相对标准偏差(n=6)和日间相对标准偏差(n=4)依次为0.87%~3.3%,1.9%~4.9%。

After centrifugation, the urine sample was purified with WATERS C18 solid phase extraction column, and then the eluate was collected. Two-dimensional UHPLC-MS/MS was applied to the determination of 8-hydroxydeoxyguanosine in the sample solution. Waters BEH C18 chromatographic column(for one-dimensional chromatography) and Waters BEH HILIC chromatographic column(for two-dimensional chromatography) were used as stationary phase. The mixture of 0.1%(φ, the same below) formic acid solution and acetonitrile with volume ratio of 8∶2(for one-dimensional chromatography) and the mixture of 0.1% formic acid solution and acetonitrile with volume ratio of 9∶1(for two-dimensional chromatography) were used as mobile phase. ESI+ and multi-reactions monitor mode were adopted in MS/MS. Internal standard method was used for quantitative. Linearity range of 8-hydroxydeoxyguanosine was found in the range of 0.25-300.0 ng·g^(-1) with detection limit(3 S/N) of 0.10 ng·g^(-1). On the matrix of blank sample, test for recovery was made by standard addition method;values of recovery found were in the range of 96.6%-103%, with intra-day RSDs(n=6) and inter-day RSDs(n=4) of determined values in the ranges of 0.87%-3.3% and 1.9%-4.9% respectively.