Development and Application of a Modified Method to Determine the Encapsulation Efficiency of Proteins in Polymer Matric
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Research Article Development and Application of a Modified Method to Determine the Encapsulation Efficiency of Proteins in Polymer Matrices Abdulkareem A. Alanezi,1 Steven H. Neau,1,2 and Anil P. D’mello1
Received 2 May 2020; accepted 17 August 2020 Abstract. A modified method to determine protein encapsulation efficiency in polymer matrices has been developed and applied to two proteins and two polymers to demonstrate its wide range of applicability. This study was pursued due to the wide variation in reported protein encapsulation efficiency of polymer-based microcapsules, even when the protein, the polymer, and the microcapsule manufacturing method were consistent. Hemoglobin (Hb) and bovine serum albumin (BSA) were chosen as model proteins and ethylcellulose and poly(lactic-co-glycolic acid) (PLGA) as model polymers. The polymer of the microcapsule was dissolved in dichloromethane/ethanol or dichloromethane/ethyl acetate for ethylcellulose or PLGA microcapsules, respectively. Liberated proteins were simultaneously precipitated, pelleted by centrifugation, isolated by decanting the polymer solution, redissolved in 10% w/ v sodium dodecyl sulfate in 0.8 N sodium hydroxide, and quantified using a modified Lowry assay. Blank microcapsules and exogenously added proteins demonstrated ≥ 93.8% recovery of proteins. The mean encapsulation efficiency of ethylcellulose or PLGA microcapsules was 52.4 or 76.9% for Hb and 86.4 or 74.7% for BSA, respectively. This demonstrates the effective use of centrifugation and the importance of an appropriate cosolvent system in the measure of encapsulation efficiency where one solvent dissolves the polymer while the other solvent quantitatively precipitates the liberated protein. It is evident that an alkaline solution of sodium dodecyl sulfate is efficient at quantitatively dissolving precipitated proteins. Remediation of problems observed with current methods and high reproducibility suggest that this modified method is generally applicable to the measure of protein encapsulation efficiency of polymer microcapsules. KEY WORDS: microcapsule; spray drying; polymer; encapsulation efficiency; protein precipitation.
Summary of Key Findings Encapsulation efficiency of proteins in polymer matrices was measured by first dissolving the polymeric coat and simultaneously precipitating the released protein in an organic cosolvent system. Each sample was centrifuged to isolate the precipitated protein as a pellet. The polymer solution was decanted to minimize its interference in protein quantification. The isolated protein was dissolved and quantified. The method to measure the protein encapsulation efficiency is highly reproducible and overcomes limitations encountered with methods commonly described in the literature. This suggests that the proposed method is applicable to the measure of the encapsulation efficiency of proteins in polymer matrices involving polymers with limited solubility in water or that are insoluble in water. 1
Department of Pharmaceutical Sciences, University of the Sciences in Philadel
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