中空纤维离心超滤-HPLC法测定人血浆中阿莫西林的浓度
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摘要
目的建立一种中空纤维离心超滤-高效液相色谱(HPLC)法分析人血浆中阿莫西林的浓度,并用于阿莫西林的药代动力学研究。方法血浆样本经中空纤维离心超滤技术预处理后,取超滤液20μL直接进行HPLC分析,用Hypersil BDS C_(18)柱(150 mm×4. 6 mm,5μm),流动相:0. 05 mol·L~(-1)磷酸二氢钾(2 mol·L~(-1)KOH将pH调至5. 0)-乙腈=97. 5∶2. 5;流速:1. 0 mL·min~(-1);柱温:30℃;检测波长:220 nm。结果人血浆中阿莫西林在0. 10～20. 3μg·mL~(-1)线性关系良好(r~2=0. 999),检测线为2. 54×10~(-2)μg·mL~(-1),方法回收率为91. 0%～100. 6%,绝对回收率为83. 1%～86. 8%,批内、批间RSD均不大于2. 9%,在室温放置8h、冻融循环3次和冰冻21 d的稳定性良好。成功用于我院人血浆中阿莫西林的药代动力学分析。结论本研究建立的方法简便、准确、灵敏,适用于人血浆中阿莫西林浓度分析。
Objective To establish a hollow fiber centrifugal ultrafiltration-high performance liquid chromatography( HPLC) method for the analysis of concentration of amoxicillin in human plasma and its application in pharmacokinetic study. Methods The plasma samples were prepared by hollow fiber centrifugal ultrafiltration,and filtrate 20 μL was directly injected into the HPLC for analysis. Hypersil BDSC_(18) column( 150 mm × 4. 6 mm,5 μm) was used. The mobile phase consisted of acetonitrile( 2. 5%) and 50 mmol·L~(-1) potassium dihydrogen phosphate( 2 mol·L~(-1) KOH adjusted to pH 5. 0,97. 5%) at the flow rate of 1. 0 mL·min~(-1). The column temperature was maintained at 30 ℃. The detected wavelength was 220 nm. Results The good linear relationship were both obtained between the free concentration of amoxicillin from 0. 10-20. 3 μg·mL~(-1)( r~2= 0. 999). The limit of detection for the analysis of free amoxicillin was 2. 54 × 10~(-2)μg·mL~(-1). The method recovery rates of free amoxicillin were 91. 0%-100. 6%. The absolute recovery rates were 83. 1%-86. 8%,respectively. The intra-day and inter-day precision( RSD) were all less than 2. 9%. The amoxicillin was all stable at the conditions of maintaining at room temperature for 8 h,after freeze-thaw cycle and keeping at -80 ℃ for 21 d. The method was successfully applicated in amoxicillin pharmacokinetic study in our hospital. Conclusion The present method is simple,accurate and sensitive,which is suitable for the analysis of amoxicillin concentration in human plasma.
Objective To establish a hollow fiber centrifugal ultrafiltration-high performance liquid chromatography( HPLC) method for the analysis of concentration of amoxicillin in human plasma and its application in pharmacokinetic study. Methods The plasma samples were prepared by hollow fiber centrifugal ultrafiltration,and filtrate 20 μL was directly injected into the HPLC for analysis. Hypersil BDSC_(18) column( 150 mm × 4. 6 mm,5 μm) was used. The mobile phase consisted of acetonitrile( 2. 5%) and 50 mmol·L~(-1) potassium dihydrogen phosphate( 2 mol·L~(-1) KOH adjusted to pH 5. 0,97. 5%) at the flow rate of 1. 0 mL·min~(-1). The column temperature was maintained at 30 ℃. The detected wavelength was 220 nm. Results The good linear relationship were both obtained between the free concentration of amoxicillin from 0. 10-20. 3 μg·mL~(-1)( r~2= 0. 999). The limit of detection for the analysis of free amoxicillin was 2. 54 × 10~(-2)μg·mL~(-1). The method recovery rates of free amoxicillin were 91. 0%-100. 6%. The absolute recovery rates were 83. 1%-86. 8%,respectively. The intra-day and inter-day precision( RSD) were all less than 2. 9%. The amoxicillin was all stable at the conditions of maintaining at room temperature for 8 h,after freeze-thaw cycle and keeping at -80 ℃ for 21 d. The method was successfully applicated in amoxicillin pharmacokinetic study in our hospital. Conclusion The present method is simple,accurate and sensitive,which is suitable for the analysis of amoxicillin concentration in human plasma.
引文
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[8]卓海通,曹晓梅,曹文.用HPLC-荧光检测同时测定阿莫西林和克拉维酸钾的浓度[J].中国临床药理学杂志,2000,16(2):111-113.
[9] DONG W C,ZHANG J F,HOU Z L,et al. The influence of volume ratio of ultrafiltrate to sample on the analysis of non-protein binding drugs in human plasma[J]. Analyst,2013,138(24):7369-7375.
[10] LI J M,LI C,JIANG Y. Pretreatment of plasma samples by a novel hollow fiber centrifugal ultrafiltrate device for the determination of cefaclor concentrations in human plasma[J]. J Chromatogr A,2010,1217(44):6824-6828.
[2] HOIZEY G,LAMIABLE D,FRANCES C,et al. Simultaneous determination of amoxicillin and clavulanic acid in human plasma by HPLC with UV detection[J]. J Pharm Biomed Anal,2002,30(3):661-666.
[3] WIBAWA J D,FOWKES D,SHAW P N,et al. Measurement of amoxicillin in plasma and gastric samples using high-performance liquid chromatography with fluorimetric detection[J]. J Chromatogr B Analyt Technol Biomed Life Sci,2002,774(2):141-148.
[4] SINGH S S,JAIN M,SHAH H,et al. Direct injection,column switching-liquid chromatographic technique for the estimation of rabeprazole in bioequivalence study[J]. J Chromatogr B Analyt Technol Biomed Life Sci,2004,813(1-2):247-254.
[5] YUAN Z,RUSSLIE H Q,CANAFAX D M. Sensitive assay for measuring amoxicillin in human plasma and middle ear fliud using solid-phase extraction and reversed-phase high-performance liquid chromatography[J]. J Chromatogr B Analyt Technol Biomed Life Sci,1995,674(1):93-99.
[6] IKEDA K,IKAWA K,IKEDA A,et al. A simple and rapid determination of biapenem in plasma by high-performance liquid chromatography[J]. J Chromatogr B Analyt Technol Biomed Life Sci,2006,844(1):148-152.
[7]沈铮,张亮,冯艳. HPLC-MS同时测定人血浆中阿莫西林和克拉维酸的浓度[J].中国药科大学学报,2002,33(1):24-27.
[8]卓海通,曹晓梅,曹文.用HPLC-荧光检测同时测定阿莫西林和克拉维酸钾的浓度[J].中国临床药理学杂志,2000,16(2):111-113.
[9] DONG W C,ZHANG J F,HOU Z L,et al. The influence of volume ratio of ultrafiltrate to sample on the analysis of non-protein binding drugs in human plasma[J]. Analyst,2013,138(24):7369-7375.
[10] LI J M,LI C,JIANG Y. Pretreatment of plasma samples by a novel hollow fiber centrifugal ultrafiltrate device for the determination of cefaclor concentrations in human plasma[J]. J Chromatogr A,2010,1217(44):6824-6828.