HPLC-UV法测定大鼠体内S-腺苷蛋氨酸浓度及其药动学研究

目的: 建立HPLC-UV法测定大鼠血浆中S-腺苷蛋氨酸(S-Adenosylmethionine, SAMe)浓度,并用此法研究静脉给药后的大鼠体内药代动力学特征。方法: 色谱柱:大连依利特Kromasil C₁₈ 柱(4.6 mm×200 mm, 5 μm),流动相:甲醇∶0.3%三氟乙酸溶液(含1%磷酸二氢钠盐)=2∶98,等浓度洗脱,紫外检测波长:260 nm,流速:0.6 mL/min,柱温:30 ℃。大鼠(n=6)经静脉给予 140 mg/kg SAMe并测定血浆药物浓度,采用DAS 2.0 软件进行药动分析。结果: SAMe能完全分离,线性范围:10~2000 μg/mL,回归方程为Y=30241X+256204,r=0.9991, 日内精密度分别为 2.6%,3.6%,3.9%;日间精密度分别为 6.3%,4.9%,7.9%;回收率分别为107%,101%,98.3%。静脉给予 140 mg/kg SAMe药动学参数,血浆药物浓度-时间曲线下面积AUC_(0-t)为(569.52±174.87) mg·L^-1·h,半衰期(t_1/2)为(2.03±0.96) h,清除率(CL)为(0.21±0.13)L·h^-1·kg^-1,表观分布容积(V_d)为(0.53±0.31) L/kg。结论: 本实验方法专属性好,灵敏度高,可靠性强,可用于SAMe药动学分析。

Determination and pharmacokinetics of S-Adenosylmethionine in rats plasma by HPLC-UV

AIM: To establish an HPLC-UV assay for determining plasma concentration of S-Adenosylmethionine (SAMe) and to study its pharmacokinetics in rats. METHODS: The chromatographic conditions were as follows: a Kromasil C₁₈ column (4.6 mm×200 mm , 5 μm) ,a flow rate of 0.6 mL/min for the mobile phase of methanol/0.3% trifluoroacetic acid (V/V) (dissolve 1% phosphate buffer) (2∶98) , an isocratic elution mode was carried out at column temperature (30 ℃) and the UV detection at the wavelength of 260 nm. 140 mg/kg SAMe was administrated by intravenous and plasma concentration were determined by HPLC-UV assay. RESULTS: SAMe was successfully separated. The calibration curve showed a good linear regression within the range of 10-2000 μg/mL. The regression equation was Y=30241X+256204, r=0.9991. The precision of intra-assay and inter-assay were 2.6%, 3.6%, 3.9% and 6.3%, 4.9%, 7.9%, respectively. The average extraction recovery was 107%, 101%, 98.3%, respectively. The pharmacokinetic parameter were AUC_(0-t) (569.52±174.87) mg·L^-1·h,t_1/2 (2.03±0.96) h, CL (0.21±0.13) L·h^-1·kg^-1, V_d (0.53±0.31) L/kg, respectively.CONCLUSION: This method was proved to be simple and efficient with excellent specificity, sensitivity, precision, recovery and can be applied to pharmacokinetic investigation of SAMe.