An in-depth study on <i>Z</i> / <i>E</i> -methoxime isomers in gas chromatography-quadrupole mass spectrometry analysis of C6-keto-opioids in human urine as their methoxime- and acyl-derivatives

C6-keto-opioids, such as hydrocodone, hydromorphone, oxycodone, and oxymorphone, are a group of semi-synthetic morphine-like analgesics with extensive applications in clinical settings and high potential for abuse and misuse. Therefore, they have become targets of workplace forensic urine drug testing (UDT) for years. Due to the undesired C6-C7 keto-enol tautomerization, the C6 ketone often needs to be deactivated prior to further derivatization for GC-MS analysis. Although it has been over two decades since the method of converting the C6 ketone to its methoxime-derivative was initially reported, little information has been published regarding the resulting Z/E-methoxime-derivative isomers' formation mechanism, stereo-configurations, or relative kinetic or thermodynamic features. Mixed Z/E-methoxime-derivative isomers create a potential peak resolution issue for GC-MS-based C6-keto-opioids identification and quantification, since the two isomers are often difficult to be completely separated by GC and they share common fragmentation pathways. We here provided the first detailed report and qualitative conformational analyses of the Z/E-methoxime-derivative isomers of C6-keto-opioids and their isomerization under the non-aqueous Brønsted-Lowry acidic conditions. By in-depth studying the C6-keto-opioids Z/E-methoxime-derivative isomers, we were able to gain important insights into potential reaction condition optimization with an attempt to reduce the formation of the minor methoxime-derivative isomer, thus, to minimize the potential interferences caused by co-existing of the two isomers and further improve the method's limit of detection (LOD) and/or limit of quantification (LOQ). Our report offered valuable information that could facilitate other laboratories using the similar derivatization procedures for GS-MS-based C6-keto-oipoids testing to improve their testing method sensitivity and enhance their analysis product quality.