Contributing mechanisms for cysteine excitotoxicity in cultured cerebellar granule cells

The purpose of this research was to develop new in vitro methodology for measuring release from petrolatum-based semisolids and to determine whether two ointments, both of which contained betamethasone dipropionate, 0.05%, but with different formulations, could be distinguished by release measurements. Several receptor media were explored to optimize the procedure utilizing Franz-type cells. Analysis was by HPLC. The release slope was 1.5 to 6 times greater from the ointment than the "augmented" ointment (which had greater clinical potency). Release was highest with a receptor consisting of a 5% solution of hexane in acetonitrile. Even so, it was necessary to subject samples of receptor from the augmented ointment to evaporation followed by reconstitution with a smaller volume of mobile phase to bring corticosteroid concentrations up to quantifiable levels. In another series of experiments, the HPLC mobile phase was used as the receptor and a relatively large volume (100 microliters) was injected onto the column. With the second approach, measured concentrations were lower but more reproducible. Quantifiable levels of betamethasone dipropionate were obtained for both formulations beginning from the first data point (at 1 hr), with satisfactory linearity of plots of amount released per unit area of membrane versus the square root of time. Using this methodology, it was possible to distinguish the effect of formulation differences in two ointments containing the same drug in the same concentration.