HPLC法检测兔胫骨及骨髓中的左氧氟沙星

Determination of levofloxacin concentration in rabbit tibia and bone marrow by HPLC

目的:建立反相高效液相色谱仪(high performance liquid chromatography,HPLC)检测兔胫骨及骨髓中左氧氟沙星的方法,用于评价纳米微球载左氧氟沙星可控降解骨修复材料中的左氧氟沙星在兔局部骨组织及骨髓中的缓释特点。方法:分别将不同浓度左氧氟沙星加入空白胫骨、骨髓,以环丙沙星作为内标,采用Sepax BR-C18(250 mm×4.6 mm 5μm)色谱柱,流动相为:0.01 mol/L磷酸二氢钾—甲醇—0.5 mol/L四丁基溴化铵(75∶25∶4),流速:1.0 ml/min,柱温:40℃,紫外检测波长290 nm。评价指标为:专属性、检测线、定量线、回收率、稳定性、精密度。结果:当胫骨中的左氧氟沙星浓度在0.05μg/ml^300μg/ml范围内线性关系良好(r=0.999);低、中、高3个浓度的方法回收率分别为(108.81±1.14)%、(96.85±3.64)%、(114.38±0.27)%;提取回收率分别为(110.92±0.42)%、(110.48±0.97)%、(80.39±0.08)%;日内精密度的RSD分别为1.30%、0.23%、5.51%;日间精密度的RSD分别为:7.09%、3.71%、2.55%,样品稳定性良好。骨髓中的左氧氟沙星浓度在0.01μg/ml^300μg/ml范围内线性关系良好(r=0.999);低、中、高3个浓度的方法回收率分别为(91.67±3.73)%、(101.77±0.26)%、(87.72±0.72)%;提取回收率分别为(102.97±0.81)%、(87.00±0.62)%、(99.97±1.15)%;日内精密度的RSD分别为1.49%、1.06%、0.11%;日间精密度的RSD分别为:5.61%、2.56%、2.37%,样品稳定性良好。结论:本方法操作简便、灵敏、稳定、回收率高,适用于检测新型载药骨修复材料中的左氧氟沙星在兔局部骨组织中的分布。

Objective：To establish a high performance liquid chromatography（HPLC）method for the determination of levofloxacin in the rabbit tibia and bone morrow;to evaluate the release characteristics of levofloxacin in nanoparticles contained levofloxacin degradation-controllable bone repair materials in rabbit local bone tissue and bone marrow. Methods：Different concentrations of levofloxacin were added in blank tibia and bone marrow respectively. Ciprofloxacin was used as an internal standard. Sepax BR-C18（250 mm×4.6 mm 5 μm）column was employed;the mobile phase was：0.01 mol/L potassium dihydrogen phosphate-methanol-0.5 mol/L tetrabutylammonium bromide（75∶25∶4）;flow rate was 1.0 ml/min;column temperature was 40 ℃;detection wavelength was 290 nm. Evaluation indexes were linear range,recoveries,extraction recoveries,intra-day precision,inter-day precision,sample stability and accuracy. Results：When the tibia of levofloxacin concentrations in 0.05-300 μg/ml,good linear relationship was achieved（r=0.999）;low,medium and high concentrations of three recoveries were（108.81±1.14）%,（96.85±3.64）%,（114.38±0.27）%;the extraction re coveries were（110.92±0.42）%,（110.48±0.97）%,（80.39±0.08）%;intra-day precision RSD were 1.30%,0.23%,5.51%;inter-day precision RSD were as follows：7.09%,3.71%,2.55%,a good sample stability. When the bone marrow levofloxacin concentrations in 0.05-300 μg/ml,a good linear relationship was achieved（r=0.999）;low,medium and high concentrations of three recoveries were（91.67±3.73）%,（101.77±0.26）%,（87.72±0.72）%;the extraction recoveries were（102.97±0.81）%,（87.00±0.62）%,（99.97±1.15）%;intra-day precision RSD were 1.49%,1.06%,0.11%;inter-day recision RSD were as follows：5.61%,2.56%,2.37%,a good sample stability. Conclusion ：With the advantages of simple,sensitive,stable,high recovery rate,this method is applicable to the detection of distribution of levofloxacin in rabbit local bone tissue in a new bone repair materials.