Unusually large deuterium discrimination during spore photoproduct formation

Abstract

The deuterium-labeling strategy has been widely used and proved highly effective in mechanistic investigation of chemical and biochemical reactions. However, it is often hampered by the incomplete label transfer, which subsequently obscures the mechanistic conclusion. During the study of photoinduced generation of 5-thyminyl-5,6-dihydrothymine, which is commonly called the spore photoproduct (SP), the Cadet laboratory found an incomplete (∼67%) deuterium transfer in SP formation, which contrasts to the exclusive transfer observed by the Li laboratory. Here, we investigated this discrepancy by re-examining the SP formation using d3-thymidine. We spiked the d3-thymidine with varying amounts of unlabeled thymidine before the SP photochemistry is performed. Strikingly, our data show that the reaction is highly sensitive to the trace protiated thymidine in the starting material. As many as 17-fold enrichment is detected in the formed SP, which may explain the previously observed one-third protium incorporation. Although commercially available deuterated reagents are generally satisfactory as mechanistic probes, our results argue that attention is still needed to the possible interference from the trace protiated impurity, especially when the reaction yield is low and large isotopic discrimination is involved. © 2014 American Chemical Society.