BSA Degradation Under Acidic Conditions: A Model For Protein Instability During Release From PLGA Delivery Systems

Abstract

Acidification of the internal poly(lactide-co-glycolide) (PLGA) microenvironment is considered one of the major protein stresses during controlled release from such delivery systems. A model protein, bovine serum albumin (BSA), was incubated at 37 degrees C for 28 days to simulate the environment within the aqueous pores of PLGA during the release phase and to determine how acidic microclimate conditions affect BSA stability. Size-exclusion high performance liquid chromatography (SE-HPLC), SDS-PAGE, and infrared spectroscopy were used to monitor BSA degradation. BSA was most stable at pH 7, but rapidly degraded via aggregation and hydrolysis at pH 2. These simulated degradation products were nearly identical to that of unreleased BSA found entrapped within PLGA 50/50 millicylinders. At pH 2, changes in BSA conformation detected by various spectroscopic techniques were consistent with acid denaturation of the protein. By contrast, at pH 5 and above, damage to BSA was insufficient to explain the instability of the protein in the polymer. Thus, these data confirm the hypothesis that acid-induced unfolding is the basis of BSA aggregation in PLGA and the acidic microclimate within PLGA is indeed a dominant stress for encapsulated BSA. To increase the stability of proteins within PLGA systems, formulations must protect against potentially extreme acidification such that native structure is maintained.

Adsorption and Conformation of Serum Albumin Protein on Gold Nanoparticles Investigated Using Dimensional Measurements and in Situ Spectroscopic Methods

“…[21][22][23][24] Furthermore, albumin continues to be used as a model protein for many biophysical and biochemical studies. 6,[25][26][27] Bovine serum albumin (BSA) is the most widely utilized serum protein due to its low cost, wide availability, and structural/ functional similarity to human serum albumin (HSA)-having 76% sequence homology 28 and nearly identical pH-dependent conformational transitions. 29 BSA is a water-soluble monomeric protein with a primary structure containing a single polypeptide chain with 583 amino acid residues and a molar mass of 66.4 kDa.…”

Untitled

“…The shape, size and charge of the particle before and after gammairradiation indicate that the bovine serum albumin (BSA) polymer that made up the matrix of the particle was not destroyed. The BSA polymer, a heat sensitive material, has been shown to undergo denaturation under chemical and heat conditions [25]. In this study, controlled gamma-irradiation of 25 kGy was sufficient to sterilize the particles while keeping the polymer intact.…”

A laboratory exercise illustrating the sensitivity and specificity of Western blot analysis

“…This corresponds to the molecular weight of BSA. In the lanes containing 1% BCS and FBS, there are weaker bands with much higher molecular weights suggesting the presence of BSA aggregates [13]. Several protein bands with molecular weights of 50-26 kDa are also observed.…”