Mamidi Rao N V S, Benjamin Biju, Ramesh Mullangi, Srinivas Nuggehally R
Bioanalysis, Drug Metabolism and Pharmacokinetics, Discovery Research, Dr Reddys Laboratories Ltd, Miyapur, Hyderabad 500-050, India.
Biomed Chromatogr. 2003 Sep;17(6):417-20. doi: 10.1002/bmc.272.
To the best of our knowledge, bioanalytical methods to determine rosiglitazone in human plasma reported in literature use internal standards that are not commercially available. Our purpose was to develop a simple method for the determination of rosiglitazone in plasma employing a commercially available internal standard (IS). After the addition of celecoxib (IS), plasma (0.25 mL) samples were extracted into ethyl acetate. The residue after evaporation of the organic layer was dissolved in 750 microL of mobile phase and 50 microL was injected on to HPLC. The separation was achieved using a Hichrom KR 100, 250 x 4.6 mm C(18) with a mobile phase composition potassium dihydrogen phosphate buffer (0.01 m, pH 6.5):acetonitrile:methanol (40:50:10, v/v/v). The flow-rate of the mobile phase was set at 1 mL/min. The column eluate was monitored by fluorescence detector set at an excitation wavelength of 247 nm and emission wavelength of 367 nm. Linear relationships (r(2) > 0.99) were observed between the peak area ratio rosiglitazone to IS vs rosiglitazone concentrations across the concentration range 5-1000 ng/mL. The intra-run precision (%RSD) and accuracy (%Dev) in the measurement of rosiglitazone were <+/-10.69 and <-12.35%, respectively across the QC levels (50-1000 ng/mL). The extraction efficiency was >80% for both rosiglitazone and IS from human plasma. The lower limit of quantitation of the assay was 5 ng/mL. In summary, the methodology for rosiglitazone measurement in plasma was simple, sensitive and employed a commercially available IS.
据我们所知，文献报道的用于测定人血浆中罗格列酮的生物分析方法所使用的内标并非市售可得。我们的目的是开发一种使用市售内标（IS）来测定血浆中罗格列酮的简单方法。加入塞来昔布（内标）后，将0.25 mL血浆样品用乙酸乙酯萃取。有机层蒸发后的残留物溶解于750 μL流动相中，取50 μL注入高效液相色谱仪。使用Hichrom KR 100、250×4.6 mm C(18)色谱柱进行分离，流动相组成为磷酸二氢钾缓冲液（0.01 m，pH 6.5）：乙腈：甲醇（40:50:10，v/v/v）。流动相流速设定为1 mL/min。通过荧光检测器监测柱流出物，激发波长设定为247 nm，发射波长设定为367 nm。在5 - 1000 ng/mL的浓度范围内，罗格列酮与内标的峰面积比与罗格列酮浓度之间呈现线性关系（r(2) > 0.99）。在质量控制水平（50 - 1000 ng/mL）下，罗格列酮测量的批内精密度（%RSD）和准确度（%Dev）分别<±10.69%和<-12.35%。罗格列酮和内标从人血浆中的萃取效率均>80%。该测定方法的定量下限为5 ng/mL。总之，血浆中罗格列酮的测定方法简单、灵敏且使用了市售内标。
