Virtual elimination of the interference of unstirred water layers on intestinal sugar transport kinetics by use of the tissue accumulation method at appropriate shaking rates

The intestinal transport of sugars and amino acids seems to follow Michaelis-Menten kinetics, but the presence of unstirred water layers at the outer face of the brush border membrane may distort kinetic measurements. According to current theory, the capacity parameter,J _mc ^max would not be affected, but theK _t would be increased to a higher value,K _t , in proportion to the thickness of the unstirred water layer,d.We reasoned that by increasing the shaking rate in the tissue accumulation method,d might drop to such small values thatK _t would fall to a constant level practically equal to the trueK _t .We measuredd-galactose influx into rings of everted hamster intestine as a function of both the substrate concentration and the shaking rate. Our results show that as the circular stirring rate increases from 0.38–6.2 Hz,J _mc ^max remains constant, as expected, butK _t first drops, then levels off to reach a plateau between 2 and 6.2 Hz. We conclude that the averageK _t values in this frequency range (K _t =7.4 mM) represent the true transportK _t . Furthermore, all previous kinetic work performed in our laboratory has been carried out under identical conditions, including shaking rates of 4 Hz. The validity of our preceding results is thus upheld.