Department of Pharmacology and Toxicology, National Veterinary Research Institute, 24-100 Pulawy, Poland.
Molecules. 2020 May 24;25(10):2445. doi: 10.3390/molecules25102445.
A simple, rapid, and accurate HPLC-UV method was developed for the determination of creatinine in pig urine. Usually, it is determined in urine in biomonitoring of xenobiotics to correct for variations in dilutions of urine samples. The colorimetric method (based on Jaffe reaction), which was mainly used for this purpose in mycotoxin biomonitoring, is not a reliable approach for pig urine. Therefore, a novel and accurate HPLC method for creatinine determination was developed. The sample preparation was based on the dilute and shoot approach. An HPLC separation was performed with a porous graphitic carbon column with an aqueous mobile phase to achieve satisfactory retention time for creatinine. The method has been successfully validated, applied for the determination of creatinine in pig urine, and compared with other methods commonly used for that purpose-a colorimetric method based on Jaffe reaction and commercial ELISA test. The developed HPLC method shows the highest precision and accuracy for pig urine samples. Finally, the method was applied as a normalization tool in LC-MS/MS mycotoxin biomarkers analysis. The standardization to a constant creatinine level (0.5 mg/mL) enables similar matrix effects for eleven mycotoxin biomarkers for pig urine samples with different creatinine levels.
建立了一种简单、快速、准确的 HPLC-UV 法测定猪尿中的肌酐。通常，在检测外源性化学物质的生物监测中，肌酐用于校正尿液样品稀释度的变化。基于 Jaffe 反应的比色法（主要用于霉菌毒素的生物监测）在此目的上并不适用于猪尿。因此，建立了一种用于肌酐测定的新型准确 HPLC 方法。该方法的样品前处理基于稀释后进样法。采用多孔石墨碳柱，以水相作为流动相进行 HPLC 分离，可获得令人满意的肌酐保留时间。该方法已成功验证，并应用于猪尿中肌酐的测定，同时与其他常用于该目的的方法（基于 Jaffe 反应的比色法和商业 ELISA 检测）进行了比较。该开发的 HPLC 方法显示出对猪尿样具有最高的精密度和准确度。最后，该方法作为 LC-MS/MS 霉菌毒素生物标志物分析的归一化工具进行了应用。肌酐水平标准化至 0.5mg/mL 时，可使肌酐水平不同的猪尿样中 11 种霉菌毒素生物标志物具有相似的基质效应。
