HPLC法测定大鼠血浆中呱氨托美丁及其代谢产物MED-5与托美丁的浓度

Determination of amtolmetin guacil and its active metabolite MED-5 and tolmetin in rat plasma by high-performance liquid chromatography

目的建立同时测定大鼠血浆中呱氨托美丁(MED-15)及其代谢产物MED-5与托美丁浓度的HPLC法。方法血浆样品用乙腈沉淀蛋白,采用HypersilODS C_(18)柱(250mm×4.6mm,5μm)进行分离,以甲醇-2%冰醋酸(60:40,V/V)为流动相,流速1.0mL·min^(-1);紫外检测波长313 nm。结果大鼠血浆中MED-15、MED-5和托美丁线性范围分别是0.050～5.6、0.042～5.4和0.039～5 mg·L^(-1)(r分别为0.995 1、0.995 7、0.999 0);定量下限分别为0.050、0.042和0.039mg·L^(-1);平均相对回收率分别是100.3%、99.4%和99.9%,平均绝对回收率分别88.9%、87.8%和88.7%;日内、日间精密度均<12%。血浆样品的稳定性符合要求。结论本研究所建立的大鼠血浆中MED-15、MED-5和托美丁浓度测定的HPLC方法灵敏、准确、精密、稳定,适用于大鼠体内MED-15的药动学研究。

AIM To develop a method for the determination of amtolmetin guacil（MED-15） and its active metabolite MED-5 and tolmetin in rat plasma by HPLC method.METHODS Plasma samples were extracted with acetonitrile. The separation was carried out by using a Hypersil ODS C_（18） column（250 mm×4.6 mm,5μm） with a mobile phase of methanol-2%glacial acetic acid（60：40,V/V） at a flow rate of 1.0 mL·min^（-1）,and detection wave length was set at 313 ran.RESULTS The linear ranges for MED-15,MED-5 and tolmetin were 0.050 - 5.6 mg·L^-1, 0.042-5.4mg·L^-1 and 0.039-5.0 mg·L^-1（r= 0.995 1,r=0.995 7,r = 0.999 0）,respectively.The limit of quantitation of MED-15,MED-5 and tolmetin was 0.05,0.042 and 0.039 mg·L^1,respectively.The relative recoveries of MED-15,MED-5 and tolmetin were 100.3%,99.4%and 99.9%,respectively.The absolute recoveries were 88.9%,87.8%and 88.7%,respectively.The intra-day RSD and inter-day RSD were both less than 12%.CONCLUSION The developed method for the quantification of MED-15,MED-5 and tolmetin in rat plasma is sensitive,accurate, precise and stable,and is suitable for the study on the pharmacokinetics of MED-15 in rats.