反相高效液相色谱法检测小鼠血清中丹皮酚浓度

Determination of paeonol in mice serum by reversed-phase high performance liquid chromatography

目的 :建立反相高效液相色谱法 (RP HPLC)测定血清中丹皮酚 ( paeonol,Pae)浓度。 方法 :血清样品用氯仿单步萃取 ,萃取物以氮气流吹干后 ,用甲醇溶解用于色谱分析。色谱条件 :采用LichrospherC18柱 ( 4 .6mm× 2 5 0mm ,5 μm ) ;以甲醇 乙腈 水 ( 30∶4 0∶30 )为流动相 ;流速为 0 .8mL·min-1;检测波长为 2 74nm ;柱温为 2 5℃ ;进样量为 2 0 μL。 结果 :Pae最低检测浓度为 0 .0 1mg·L-1;血清样品浓度在 0 .0 2～ 12 .80mg·L-1范围内呈良好线性关系 ,回归方程为Y =2 712 .1836X -5 .7137,r =0 .9999,n =5 ;相对回收率为 ( 10 1.2± 1.2 ) % ,平均萃取回收率为 ( 79.7± 3.0 ) % ;日内RSD与日间RSD均小于10 .0 % (n =5 )。结论 :该方法灵敏、简便、快速、重复性好 。

OBJECTIVE A simple assay method for paeonol in mice serum by reversed-phase high performance liquid chromatography (RP-HPLC) was established.METHODS Serum sample was extracted one time with chloroform for RP-HPLC analysis. The RP-HPLC assay was conducted using Lichrospher C 18 column( 4.6 mm×250 mm, 5 μm) with methanol-acetonitrile-water(30∶40∶30 ) as mobile phase at a flow rate of 0.8 mL·min -1 . 20 μL of sample solution were injected into the C 18 column where the temperature was 25℃.The detection wavelength was 274 nm.RESULTS The limit of detection for paeonol was 0.01 mg·L -1 in serum sample.The linearity range for paeonol was obtained from 0.02 mg·L -1 to 12.80 mg·L -1 . The linear regression equation was Y= 2712.1836 X- 5.7137 ,r= 0.9999 ,n=5. The recoveries of paeonol were ( 101.2 ± 1.2 )%. The average recovery of paeonol in samples was ( 79.7 ± 3.0 )%. The relative standard deviations of the intra-day and inter-day were less than 10%(n=5).CONCLUSION The method is sensitive, simple, rapid, and accurate, which may be used for blood concentration monitoring and pharmacokinetics study for paeonol.