Glycolytic pH oscillations in a flow reactor

A new type of flow reactor (UCSTR) has been developed that uses anisotropic ultrafiltration membranes in a continuous flow stirred tank reactor (CSTR) to facilitate the study of nonlinear enzyme catalyzed reactions. The design allows the study of enzymes with subunit molecular weights > or = 9000 dalton and protein concentrations up to at least 2 mg/ml under flow conditions with a residence time of 3 min or more, in a reactor of volume 1.67 ml. The UCSTR allows continuous potentiometric or spectrophotometric measurement without design change. Calibration of reactor performance was carried out by reproducing pH oscillations in the ferrocyanide-hydrogen peroxide reaction. Experimental verification of oscillatory glycolysis in the UCSTR was carried out with extract of rat skeletal muscle. Input feeds were fructose-6-phosphate and ATP with low concentrations of phosphate as buffer. Oscillations in pH, sustained for over eight hours, were observed. A six-step mechanism, including product activation and substrate inhibition, seven concentration variables, and four enzymes sufficed simulate the pH oscillations observed in the UCSTR.