IMPORTANCE OF IN-VITRO EXPERIMENTAL CONDITIONS ON PROTEIN RELEASE KINETICS, STABILITY AND POLYMER DEGRADATION IN PROTEIN ENCAPSULATED POLY(D,L-LACTIC ACID-CO-GLYCOLIC ACID) MICROSPHERES

Two proteins, carbonic anhydrase (M(r) 31 000) and bovine serum albumin (M(r) 66 000), have been encapsulated as model protein drugs within microspheres composed of relatively fast degrading poly(D,L-lactic acid-co-glycolic acid, 50/50). The microspheres were incubated by employing two different in vitro experimental sets of the release system: polypropylene tube and dialysis bag. The protein stability problems, such as chemical degradation and aggregation, were investigated by analysis of released protein in the phosphate buffered saline medium and unreleased protein within the microspheres. Analysis of unreleased protein was performed after direct extraction from the microspheres by means of an electric field using a gel electrophoresis apparatus. Degradation of the microspheres incubated in the tube caused a significant pH decrease from pH 7.4 to below pH 3 after a one-month incubation period. This resulted in severe hydrolysis of both the released protein in the medium and the unreleased protein within the microspheres. However, the released protein in the medium and the unreleased protein within the microsphere exhibited less severe chemical degradation when the microspheres were incubated in the dialysis bag which permitted a constant pH. The microspheres in the tube demonstrated a significant decrease in polymer molecular weight and a pronounced morphological change over time, as compared to those in the dialysis bag. There was no apparent effect of encapsulated protein on the polymer degradation in either experimental set.