Human oral drugs absorption is correlated to their in vitro uptake by brush border membrane vesicles.

Brush border membrane vesicles (BBMV) were prepared from the rabbit small intestine for testing drug absorption potency through the enterocyte's apical membrane, which is an important compartment for drug oral absorption. Some modifications have been made to the traditional vesicle assay for adapting it to the 96-well plate format. The accumulation of 23 reference drugs was measured, and the data showed a good correlation with human oral absorption with a correlation coefficient R=0.

Two-step mechanism of phlorizin binding to the SGLT1 protein in the kidney.

The relationships between phlorizin binding and Na+-glucose cotransport were addressed in rabbit renal brush-border membrane vesicles. At pH 6.0 and 8.

Kinetic mechanisms of inhibitor binding: relevance to the fast-acting slow-binding paradigm.

Although phlorizin inhibition of Na+-glucose cotransport occurs within a few seconds, 3H-phlorizin binding to the sodium-coupled glucose transport protein(s) requires several minutes to reach equilibrium (the fast-acting slow-binding paradigm). Using kinetic models of arbitrary dimension that can be reduced to a two-state diagram according to Cha's formalism, we show that three basic mechanisms of inhibitor binding can be identified whereby the inhibitor binding step either (A) represents, (B) precedes, or (C) follows the rate-limiting step in a binding reaction. We demonstrate that each of mechanisms A-C is associated with a set of unique kinetic properties, and that the time scale over which one may expect to observe mechanism C is conditioned by the turnover number of the catalytic cycle.

Sugar transport heterogeneity in the kidney: two independent transporters or different transport modes through an oligomeric Protein? 1. Glucose transport studies.

The kinetics of Na+/d-glucose cotransport (SGLT) were reevaluated in rabbit renal brush border membrane vesicles isolated from the whole kidney cortex using a fast-sampling, rapid-filtration apparatus (FSRFA, US patent #5,330,717) for uptake measurements. Our results confirm SGLT heterogeneity in this preparation, and both high (HAG) and low (LAG) affinity glucose transport pathways can be separated over the 15-30 degrees C range of temperatures. It is further shown that: (i) Na+ is an essential activator of both HAG and LAG; (ii) similar energies of activation can be estimated from the linear Arrhenius plots constructed from the Vmax data of HAG and LAG, thus suggesting that the lipid composition and/or the physical state of the membrane do not affect much the functioning of SGLT; (iii) similar Vmax values are observed for glucose and galactose transport through HAG and LAG, thus demonstrating that the two substrates share the same carrier agencies; and (iv) phlorizin inhibits both HAG and LAG competitively and with equal potency (Ki = 15 microM).