UPLC-MS/MS simultaneous quantification of urinary circadian rhythm hormones and related metabolites: Application to air traffic controllers.

Civil aviation flight crew and civil aviation air traffic controllers are prone to circadian rhythm abnormalities, which can lead to a slew of other maladies. It could endanger people's health and provide a serious threat to the safety of civil aviation flights if it is not appropriately evaluated and addressed. Early detection of rhythm irregularities and prompt treatment for particular populations that are vulnerable to rhythm disorders are crucial for enhancing civil aviation safety. In general, monitoring of the classical circadian rhythm biomarkers (melatonin or cortisol) in plasma or saliva is an effective way to evaluate the rhythm status. Due to the challenging sample procedure and the trauma of plasma, urine sample testing has received an increasing amount of attention. While, urine circadian rhythm biomarkers have seldom been examined, and the relationship between urinary steroid hormones and melatonin is still poorly understood. In most cases, hormones are determined by immunoassays respectively, mainly enzyme-linked immunosorbent assay (ELISA) or radioimmunoassay (RIA). There are also reports describing the liquid chromatography with tandem mass spectrometry (LC-MS/MS) technique as a method of melatonin or few steroid hormones quantification, however, the simultaneous detection of multiple rhythmic hormones in human urine is rarely reported. For the quantification of the rhythmic hormones in human urine, an accurate approach using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was devised in this work. Nine endogenous hormones (melatonin, 6-hydroxymelatonin, 6-sulfatoxymelatonin, cortisol, corticosterone, cortisone, testosterone, epitestosterone and androsterone), in human overnight urine, were quantified after solid phase extraction (SPE). A reverse phase HSS C18 column was used for chromatographic separation with a 9-minute gradient elution and deuterated analogues of each analyte were applied as internal standards. This method was successfully applied to the analysis of 596 overnight urine samples (23:00-9:00) collected from 84 air traffic controllers in the Beijing area during shift work. This study's findings showed a clear correlation not only between melatonin and its metabolites; cortisol-related metabolites, but also between melatonin metabolites and endogenous metabolites upstream and downstream of cortisol, implying that these two categories of hormones can be used as potential biological rhythm indicators to provide circadian rhythm data support for future studies on circadian rhythm disorders.

Identification of characteristic heroin metabolites in urine based on data-mining technology and multivariate statistics analysis combined with a targeted verification approach for distinguishing heroin abusers.

A common phenomenon shows that ingestion of opium poppy shell-containing drugs can result in a "false-positive" urinalysis test result for mandatory or workplace heroin abuse screening. Owing to the short detection window (8 h in urine) of the characteristic heroin metabolite 6-monoacetylmorphine (6-MAM) confirmation or exclusion of heroin abusers still presents major challenges for toxicologists. In this work, we developed an ultra-performance liquid chromatography-time-of-flight mass spectrometry method (UPLC-TOF-MS) with online data acquisition and multiple post-data-mining technologies combined with a multivariate statistical and batch validation analysis workflow to assess the characteristic urine metabolites of heroin abusers. Based on the proposed methods, 28 characteristic metabolites were structurally identified, and their fragmentation patterns and metabolite pathways were also summarized. Correlation analysis was used to investigate the internal relationship and similarities among the identified metabolites, and seven representative metabolites were selected as "Target-metabolites". Multi-batch urine of samples of heroin abusers were certified based on the UPLC-MS/MS method for further validation of the practicability of using this method for routine analysis. Overall, the target-metabolites can be utilized as assistant "biomarkers" in workplace or mandatory drug screenings. This approach encourages further studies on the development of the "false-positive" identification system.