Cone voltage induced in-source dissociation of glucuronides in electrospray and implications in biological analyses

Glucuronidation is a common mechanism in drug metabolism. In-source dissociation of glucuronides in electrospray generates fragment ions identical to those of the precursor ions of the original drugs. The effect of experimental parameters on the process was investigated in the present study using both commercially available compounds and glucuronides generated from microsomal glucuronidation incubations. It was found that cone voltage was the most critical parameter contributing to in-source fragmentation of both O- and N-glucuronides, whereas both the desolvation temperature and the source temperature had little effect. Additionally, the extent of in-source dissociation varied for different glucuronides and could be minimized by lowering cone voltage. As demonstrated in real examples, minimizing in-source dissociation can lead to higher sensitivity in detecting glucuronides in biological samples. In addition, product ions resulting from in-source dissociation of glucuronides potentially interfere with accurate determinations of corresponding drug levels if chromatographic separation is not adequate. For cases in which chromatographic separation of glucuronides from the original drugs is not readily achieved or high-throughput analyses are desired, interference caused by in-source dissociation can usually be eliminated simply by using lower cone voltage. This approach has been proven to be effective in the analysis of more than 100 glucuronides generated from in vitro microsomal incubations.