A COMPARISON OF ALTERNATIVE METHODS FOR THE DETERMINATION OF THE LEVELS OF PROTEINS ENTRAPPED IN POLY(LACTIDE-CO-GLYCOLIDE)MICROPARTICLES

Two alternative techniques, one involving extraction into dichloromethane (DCM) and the other polymer hydrolysis by sodium hydroxide (NaOH), and extraction by sodium dodecyl sulphate (SDS), were assessed for their ability to determine the levels of three model proteins, bovine serum albumin (BSA), ovalbumin and lysozyme, entrapped in poly(lactide-co-glycolide) microparticles. In addition, the two techniques were compared with two more widely used techniques for the quantitation of proteins and peptides, polyacrylamide gel electrophoresis (SDS-PAGE) followed by scanning densitometry, and amino acid analysis (AAA). As a control, BSA was radiolabelled to allow an accurate determination of the total level of protein entrapped in microparticles. The results showed that NaOH/SDS extraction resulted in accurate determinations of the levels of proteins entrapped in microparticles, but that extraction into DCM resulted in underestimations of the total amount of protein present. AAA also resulted in accurate determinations of the levels of entrapped proteins, but SDS-PAGE resulted in underestimations.