Evaluation of the stability and cross-reactivity of zopiclone and eszopiclone using immunoassay kits.

Zopiclone (ZOP) and eszopiclone (EZOP) are hypnotics belonging to the cyclopyrrolone class, which are metabolized to zopiclone-N-oxide (ZOPNO) and N-desmethylzopiclone (NDZOP), respectively, and finally to 2-amino-5-chloropyridine (2A5C) after long-term storage, freezing, and conservation. This study aimed to examine the stability of ZOP and EZOP using a commercially available immunoassay urine screening kit for ZOP. ZOP and EZOP, as well as their metabolites and 2A5C, were analyzed in urine using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Spiked urine samples were used to confirm cross-reactivity for the immunoassay test and to compare the quantitative results of the clinical urine samples. The results indicate that urine samples stored for an extended period or altered postmortem may lead to false-negative results for ZOP, providing important information for interpreting results in cases involving ZOP.