Medvedovici Andrei, Udrescu Stefan, Albu Florin, Tache Florentin, David Victor
University of Bucharest, Department of Analytical Chemistry, Bucharest - 050663, Romania.
Bioanalysis. 2011 Sep;3(17):1935-47. doi: 10.4155/bio.11.148.
Liquid-liquid extraction of target compounds from biological matrices followed by the injection of a large volume from the organic layer into the chromatographic column operated under reversed-phase (RP) conditions would successfully combine the selectivity and the straightforward character of the procedure in order to enhance sensitivity, compared with the usual approach of involving solvent evaporation and residue re-dissolution. Large-volume injection of samples in diluents that are not miscible with the mobile phase was recently introduced in chromatographic practice. The risk of random errors produced during the manipulation of samples is also substantially reduced.
A bioanalytical method designed for the bioequivalence of fenspiride containing pharmaceutical formulations was based on a sample preparation procedure involving extraction of the target analyte and the internal standard (trimetazidine) from alkalinized plasma samples in 1-octanol. A volume of 75 µl from the octanol layer was directly injected on a Zorbax SB C18 Rapid Resolution, 50 mm length × 4.6 mm internal diameter × 1.8 µm particle size column, with the RP separation being carried out under gradient elution conditions. Detection was made through positive ESI and MS/MS. Aspects related to method development and validation are discussed.
The bioanalytical method was successfully applied to assess bioequivalence of a modified release pharmaceutical formulation containing 80 mg fenspiride hydrochloride during two different studies carried out as single-dose administration under fasting and fed conditions (four arms), and multiple doses administration, respectively. The quality attributes assigned to the bioanalytical method, as resulting from its application to the bioequivalence studies, are highlighted and fully demonstrate that sample preparation based on large-volume injection of immiscible diluents has an increased potential for application in bioanalysis.
与常规的溶剂蒸发和残渣重新溶解方法相比，从生物基质中液 - 液萃取目标化合物，然后将大量有机层注入反相（RP）条件下操作的色谱柱中，能成功结合该方法的选择性和简便性，以提高灵敏度。最近在色谱实践中引入了在与流动相不混溶的稀释剂中进行大体积进样。在样品处理过程中产生随机误差的风险也大幅降低。
一种用于评估含芬司匹利药物制剂生物等效性的生物分析方法，其样品前处理程序是从碱化血浆样品中用1 - 辛醇萃取目标分析物和内标（曲美他嗪）。从辛醇层取75 μl直接进样到Zorbax SB C18快速分离柱上，该柱长50 mm，内径4.6 mm，粒径1.8 µm，在梯度洗脱条件下进行反相分离。通过正离子电喷雾电离和串联质谱进行检测。讨论了与方法开发和验证相关的方面。
该生物分析方法已成功应用于评估含80 mg盐酸芬司匹利的缓释药物制剂在两项不同研究中的生物等效性，这两项研究分别在禁食和进食条件下单剂量给药（四个组）以及多剂量给药情况下进行。强调了该生物分析方法在生物等效性研究中的应用所赋予的质量属性，充分证明了基于不混溶稀释剂大体积进样的样品前处理在生物分析中的应用潜力有所增加。
