Activity and stability of caffeine demethylases found in Pseudomonas putida IF-3

Resting cell suspensions and cell-free extracts of Pseudomonas putida IF-3 were tested to assess their ability to degrade caffeine, and to determine their capacity to retain activity at different temperatures. A method to quantify cell lysis using optical density was developed in order to allow the comparison of cell free extract and resting cell caffeine degradation rates on the same basis. Caffeine degradation rates for cell free extracts were found to be 2.4 μmol g cells−1 min−1; this rate is 55 times faster than previously reported P. putida data. Resting cells degraded caffeine 12 times faster than cell free extracts, at 22 °C. However, both systems were equivalently active at 50 °C. Resting cells were significantly more stable than cell free extracts, retaining their ability to degrade caffeine even at elevated temperatures. Cell free extracts lost all activity after 15 min at 55 °C.