A quantitative analysis of polysorbate 80 is an essential part of a characterization of therapeutic protein products. There are many methods to perform such type of analysis, including spectrophotometry and HPLC. However, there is no high-throughput method with high sensitivity and accuracy. In our work, we optimized conditions of polysorbate 80 hydrolysis to reduce the sample preparation time and developed a new isocratic reversed phase HPLC method for the quantification of oleic acid, the principal product of the hydrolysis. The validation of developed method shows that it is characterized by the repeatability less than 2.7% with the LOD about 25 ng (125 ppb), LOQ about 100 ng (500 ppb) within the analytical range of 0.025 to 1.5 mg/ml polysorbate 80 concentrations, accuracy between 92and108% and high precision (coefficient of variation less than3.2%). The total time needed for the analysis was about 1 h. The method could be routinely used for the quality control during therapeutic proteins manufacturing.
Keywords: HPLC; Hydrolysis; Quantitation; Validation; polysorbate 80.
Copyright © 2019 Elsevier B.V. All rights reserved.