Castillo-García M L, Aguilar-Caballos M P, Gómez-Hens A
Analytical Chemistry Department, Institute of Fine Chemistry and Nanochemistry, Campus of Rabanales, Annex to Marie Curie Building, University of Córdoba, 14071 Córdoba, Spain.
J Chromatogr B Analyt Technol Biomed Life Sci. 2015 Mar 15;985:85-90. doi: 10.1016/j.jchromb.2015.01.026. Epub 2015 Jan 25.
A new method based on ultra high performance liquid chromatography (UPLC) with photometric and fluorometric detection for the determination of acetylsalicylic acid and its main metabolites, namely gentisic, salicylic and salicyluric acids, in bovine urine samples is reported. Photometric detection was used for acetylsalicylic acid determination, whereas the native fluorescence of the metabolites was monitored using fluorometric detection. The separation was performed under isocratic conditions, using acetonitrile-phosphate solution (3.5mM, pH 3.5) (26:74, v/v) as the mobile phase. The retention times of the four compounds were lower than 2min, which are shorter than those achieved using conventional HPLC. Under the optimum separation conditions, the dynamic ranges and detection limits (ngmL(-1)) were: 0.2-2500, 0.09 for gentisic acid; 0.2-2500, 0.08 for salicylic acid and 2.5-15,000, 1.1 for salicyluric acid, using fluorescence detection, and 10-25,000, 2.2 for acetylsalicylic acid, using UV detection. Intra-day and inter-day precision data were assessed at two levels of concentration of each analyte using both detection systems. The selectivity of the method was checked by assaying different drugs of veterinary use showing that most of them did not interfere with the determination of the analytes. The method has been applied to the analysis of bovine urine samples, which only required a simple clean up step of the samples prior to injection in the UPLC system. A recovery study was performed, which provided values in the range of 80-100%. This fact proves the practical usefulness of this method as an ultrafast analytical tool for the therapeutic control of acetylsalicylic acid administration in bovine animals intended for food production.
报道了一种基于超高效液相色谱（UPLC）并结合光度和荧光检测的新方法，用于测定牛尿液样本中的乙酰水杨酸及其主要代谢物，即龙胆酸、水杨酸和水杨尿酸。采用光度检测法测定乙酰水杨酸，而利用荧光检测法监测代谢物的天然荧光。分离在等度条件下进行，使用乙腈 - 磷酸盐溶液（3.5 mM，pH 3.5）（26:74，v/v）作为流动相。这四种化合物的保留时间均低于2分钟，比使用传统高效液相色谱法获得的保留时间更短。在最佳分离条件下，使用荧光检测时，龙胆酸的动态范围和检测限（ng mL⁻¹）分别为：0.2 - 2500，0.09；水杨酸为：0.2 - 2500，0.08；水杨尿酸为：2.5 - 15000，1.1；使用紫外检测时，乙酰水杨酸的动态范围和检测限为：10 - 25000，2.2。使用两种检测系统在每种分析物的两个浓度水平上评估日内和日间精密度数据。通过检测不同的兽用药物来检查该方法的选择性，结果表明大多数药物不会干扰分析物的测定。该方法已应用于牛尿液样本的分析，在将样本注入UPLC系统之前，只需要对样本进行简单的净化步骤。进行了回收率研究，回收率值在80 - 100%范围内。这一事实证明了该方法作为一种超快速分析工具在对用于食品生产的牛进行乙酰水杨酸给药治疗控制方面的实际实用性。
