Infrared ATR spectroscopy of substrates in aqueous solution using cryoenrichment and its application in enzyme-activity assays

Assays of enzyme activity in aqueous solutions were carried out with the use of the cryoenrichment method for infrared spectroscopy. A calibration method that used partial least-squares (PLS) for the measurement of enzyme activity was investigated. In this method, the enzyme activity was estimated by measuring the change in the concentration of a substrate or a product based on enzyme catalysis using the cryoenrichment method for infrared spectroscopy. The correlation coefficients between the activity levels of lactate dehydrogenase, amylase, and creatine kinase and the concentration changes of the substrate or the product were over 0.93. However, the correlation coefficient for alkaline phosphatase was only 0.78. Our findings from this study confirm that enzyme activity can be measured by infrared spectroscopy.