QuEChERS-超高效液相色谱-串联质谱法测定血液中4种乌头类生物碱

UHPLC-MS/MS Determination of 4 Aconitum Alkaloids in Blood with QuEChERS

移取1.00mL血液样品,加入2mL乙腈和50mg氯化钠,振荡10min后,在-4℃下以8 000r·min^(-1)转速离心10min,取上清液,加入15mg N-丙基乙二胺和25mg十八烷基硅烷,振荡5min,重复上述离心操作,取上清液,过0.22μm有机微孔膜,采用超高效液相色谱-串联质谱法测定滤液中次乌头碱、新乌头碱、乌头碱和滇乌头碱等4种乌头类生物碱的含量。以Agilent ZORBAX Eclipse Plus C_(18)色谱柱为固定相,以不同体积比的0.1%(质量分数)氨水和甲醇的混合液为流动相进行梯度洗脱,串联质谱分析中采用电喷雾正离子源和多反应监测模式。4种乌头类生物碱的质量浓度在一定范围内与其对应的峰面积呈线性关系,方法的检出限(3S/N)为0.010～0.035μg·L^(-1)。以空白样品为基体进行加标回收试验,所得回收率为96.3%～109%,回收量的日内相对标准偏差(n=6)为2.3%～4.2%,日间相对标准偏差(n=6)为0.70%～6.7%。

The blood sample(1.00 mL) was taken,and then 2 mL of acetonitrile and 50 mg of sodium chloride were added.After oscillation for 10 min,the above mixture was centrifugated for 10 min at the speed of 8 000 r·min-1 at -4℃.PSA(15 mg) and C18(25 mg) were added in the supernatant.After oscillation for 5 min,the above centrifugal operation was repeated.The supernatant was filtered on 0.22 μm organic microporous membrane.UHPLC-MS/MS was applied to the determination of 4 aconitum alkaloids,i.e.hypaconitine,mesaconine,aconitine and yunaconitine in the filtrate.Agilent ZORBAX Eclipse Plus C18 chromatographic column was used as stationary phase,and the mixture of 0.1%(mass fraction) ammonia and methanol mixed in different ratios was used as mobile phase in gradient elution.ESI+and multi-reactions monitor mode were adopted in MS/MS.Linear relationships between values of peak area and mass concentration of the 4 aconitum alkaloids were kept in definite ranges.Detection limits(3S/N) of the method ranged from 0.010 to 0.035μg·L-1.On the base of blank sample,test for recovery was made by standard addition method;values of recovery found were in the range of 96.3%-109%,with intra-day RSD(n=6) and inter-day RSD(n=6) of recovery amounts in the ranges of 2.3%-4.2% and 0.70%-6.7%.