All sample measurements were carried out in triplicate. Two hundred microlitres of the toluenized cells were placed in a fresh 1·5 ml microcentrifuge tube and 20 μl of 0·5 mol l−1 ACC was added to the suspension, briefly vortexed and then incubated at 30°C for 15 min. Following the addition of 1 ml of 0·56 mol l−1 HCl, the mixture was vortexed and centrifuged for 5 min at 12 000 g at room temperature. One millilitre of the supernatant was vortexed together with 800 μl of 0·56 mol l−1 HCl. Then, 300 μl of the 2,4-dinitrophenyl hydrazine reagent was added to the glass tube, the contents vortexed and then incubated for 30 min at 30°C. Following the addition and mixing of 2 ml of 2 N NaOH, the absorbance was measured at 540 nm