The strain‐promoted azide alkyne cycloaddition (SPAAC) is a powerful tool for forming covalent bonds between molecules even under physiological conditions, and therefore found broad application in fields ranging from biological chemistry and biomedical research to materials sciences. For many applications, knowledge about reaction kinetics of these ligations is of utmost importance. Kinetics are commonly assessed and studied by NMR measurements. However, these experiments are limited in terms of temperature and restricted to deuterated solvents. By using an inline ATR‐IR probe we show that the cycloaddition of azides and alkynes can be monitored in aqueous and even complex biological fluids enabling the investigation of reaction kinetics in various solvents and even human blood plasma under controlled conditions in low reaction volumes.
Taking a closer look: We show that azide cycloadditions can be monitored in complex reaction environments by using an inline ATR‐IR probe in a small‐volume reaction flask also facilitating full temperature control. This method enables following bioorthogonal azide cyclooctyne ligations even in biological fluids such as human plasma.