Extraction and Quantification of Recombinant Bovine Somatotropin from Oleaginous Vehicle

Abstract

A non-denaturing assay for the determination of potency of a recombinant bovine somatotropin (somidobove) formulated in oleaginous vehicles was developed. In this paper, the optimal conditions for extraction of somidobove from oleaginous vehicle were investigated. Results demonstrated that somidobove monomer, oligomers, and soluble aggregates were quantitatively extracted by borate - ethylenediaminetetraacetate (EDTA) pH 9.5 buffer solution in a VanKel dissolution system at 40°C for 3 hours. The extracted analytes, somidobove monomer, dimer, and soluble aggregates, were separated and determined by using a high performance size exclusion chromatographic system consisting of a TSK G3000SW column and a borate - EDTA buffer (pH 7.3) mobile phase. This method was validated for the effects of buffer type, extraction temperature, extraction time and stirring speed. The average CV of intra- and inter-day precision obtained in this method were 2.3%. The recovery of somidobove monomer was 93.7% with a coefficient of variance (CV) of 3.71%. The CV for precision generated in three different labs ranged from 1.3 - 3.9%. This method was linearly correlated with the hypophysectomized rat body weight gain assay, regression coefficient = 0.953.