高效液相色谱法测定小鼠血清中磺胺间甲氧嘧啶及其N4-乙酰化代谢物含量

Determination of Sulfamonomethoxine and Its N4-acetyl Metabolite in Mice Serum by HPLC

关键词:  磺胺间甲氧嘧啶  N4-乙酰化磺胺间甲氧嘧啶  高效液相色谱  二极管阵列检测器

Key words:  sulfamonomethoxine  N₄-acetyl sulfamonomethoxine  high performance liquid chromatography  diode array detector

摘要:

磺胺间甲氧嘧啶（SMM）于ICR小鼠哺乳期暴露后，测定其血清中SMM和N4-乙酰化磺胺间甲氧嘧啶（ACSMM）含量。小鼠哺乳期以0、10、50、200 mg/(kg·d)灌胃给药直至21 d，于第22天麻醉，摘眼球取血。血清样品经乙腈沉淀蛋白，超声辅助提取，采用反相 C18（250 mm×4.6 mm，5 μm）色谱柱，以乙腈：水（含 0.2%冰乙酸）为流动相，流速为1 mL/min，柱温35 ℃，高效液相色谱耦合二极管阵列检测器对目标物检测，SMM和ACSMM检测波长分别为273 nm和267 nm，外标法峰面积定量。SMM和ACSMM血清加标回收率为85.8%~110.2%，精密度（RSD%）均小于8.3%，在0.05 ~2.5 μg/mL和5~30 μg/mL（适用母鼠体内SMM）范围内与峰面积呈良好的线性关系，r2> 0.990；最低检出限分别为0.023 和0.042 μg/mL。本法灵敏、快速、准确，可用于血清中SMM和ACSMM的测定。

Abstract:

Determination of sulfamonomethoxine (SMM) and N4-acetyl metabolite (ACSMM) in ICR mice exposed to SMM during lactation. The lactational mice were orally administered with SMM (0, 10, 50, 200 mg/(kg·d) for 21 d, the mice were anesthetized and then taken blood by removalling eyeball. Following serum protein precipitation with acetonitrile, the ultrasonic- assisted extraction of the targets, separation of them were carried out on a reversed-phase C18 (250 mm×4.6 mm, 5 μm) column, the mobile phase was involved in acetonitrile and 0.2% acetic acid in water, the flow rate was 1 mL/min and the column temperature was 35 ℃, the high performance liquid chromatography was coupled with a diode array detector, detection wavelength of SMM and ACSMM were 273 and 267 nm respectively, and quantified with external standard method. The recoveries of SMM and ACSMM spiked in serum were 85.8% ~110.2%, RSDs less than 8.3% were obtained, the linear range was 0.05~2.5 μg/mL, but 5~30 μg/mL suitable for SMM in the dams, and showed a good linear relationship (r2>0.990). The limit of quantitation (LOQ) was 0.023 and 0.042 μg/mL respectively. This method was sensitive, rapid, accurate, can be used for the determination of SMM and ACSMM in serum.