Department of Chemical Pathology and Endocrinology, Armed Forces Institute of Pathology, Rawalpindi, Pakistan.
Watim Medical & Dental College, Rawalpindi, Pakistan.
Anal Biochem. 2022 Feb 1;638:114496. doi: 10.1016/j.ab.2021.114496. Epub 2021 Nov 25.
LC-MS/MS has recently emerged as the best practice for simultaneous analysis of 2, 4, 6 Trinitrotoluene (TNT) and its metabolites. We have developed and validated an LC-MS/MS method for simultaneous quantification of 2, 4, 6 Trinitrotoluene (TNT) and its metabolites 4-ADNT, 2-ADNT, 2,4-DNT, and 2,6-DNT in urine samples. These four metabolites were acid hydrolyzed using 1 mL of urine followed by extraction using n-Hexane and ethyl acetate as an extracting solvent. Separation was achieved by centrifugation, and the supernatant was dried under nitrogen, reconstituted with water and acetonitrile, and then filtered. Chromatographic separation was achieved on Agilent Poroshel 120 EC-C18 column (2.1 mm × 75 mm × 2.7 μm) utilizing two mobile phases 0.1% formic acid in water and 0.1% formic acid in acetonitrile in gradient flow. The validated AMR of TNT and its metabolites was 7.8-1000 ng/mL. The method showed an excellent correlation (>0.99) for TNT and its metabolites. Accuracy and within/between day precision of TNT and its metabolites were within ±15%. The integrity of diluted samples was maintained for each dilution factor. The method was found stable after storage and freeze-thaw cycle. The presented method can be used for TNT screening in occupationally exposed ordnance factory workers.
LC-MS/MS 最近已成为同时分析 2,4,6-三硝基甲苯（TNT）及其代谢物的最佳实践。我们已经开发并验证了一种 LC-MS/MS 方法，用于同时定量尿液样品中的 2,4,6-三硝基甲苯（TNT）及其代谢物 4-ADNT、2-ADNT、2,4-DNT 和 2,6-DNT。这四种代谢物使用 1 毫升尿液进行酸水解，然后使用正己烷和乙酸乙酯作为提取溶剂进行提取。通过离心实现分离，上清液在氮气下干燥，用水分和乙腈重新配制，然后过滤。通过 Agilent Poroshel 120 EC-C18 柱（2.1mm×75mm×2.7μm）利用两种流动相 0.1%甲酸在水中和 0.1%甲酸在乙腈中的梯度流实现色谱分离。TNT 及其代谢物的验证 AMR 为 7.8-1000ng/mL。该方法对 TNT 及其代谢物表现出极好的相关性（>0.99）。TNT 及其代谢物的准确度和日内/日间精密度均在±15%以内。每个稀释因子都保持了稀释样品的完整性。该方法在储存和冻融循环后发现稳定。所提出的方法可用于职业暴露于弹药厂工人的 TNT 筛选。
