Rapid Characterization of Isotopic Purity of Deuterium‐Labeled Organic Compounds and Monitoring H/D Exchange Reaction by Electrospray High‐Resolution Mass Spectrometry

Deuterium-labeled organic compounds, reagents, drugs have been widely used in many scientific research fields. Isotopic purity as the feature parameter of deuterated compounds is of great importance. In this article, we used electrospray high-resolution mass spectrometry (ESI HR-MS) to study isotopic purity of deuterium-labeled organic compounds based on assigning and distinguishing the corresponding H/D isotopolog ions of deuterated compounds.We systematically considered the specific requirements of accuracy and resolution of ESI HR-MS when measuring isotope purity, and demonstrated some actual cases by ESI HR-MS and UPLC HR-MS. Meanwhile, ESI HR-MS/MS of deuterated compounds were performed to provide more information on deuterium-labeled position characterization.Two isotopic purity calculation methods base on the relative abundance in ESI HR-MS and UPLC HR-MS of H/D isotopolog ions (D0 ~Dn ), were compared, which gave consistent isotopic purity values and were all in good agreement with the certified isotopic purity values. We further studied and monitored the H/D exchange reaction of ethyl 3-(4-bromophenyl)-3-oxopropanoate (EBPO) by in situ evaluating the dynamic isotopic purity changes of EBPO-D2 in the H/D exchange reaction.The isotopic purity characterization methods by ESI HR-MS discussed in this article have some outstanding advantages: rapid, highly sensitive, very low sample consumption (even below ng level) and deuterated solvent-free. Thus, this low-impact analytical method is time and cost economic and might have good application potential for in-situ isotopic purity analysis and monitoring the H/D exchange reactions.