J. Fransson et al., OXIDATION OF HUMAN INSULIN-LIKE GROWTH-FACTOR-I IN FORMULATION STUDIES - KINETICS OF METHIONINE OXIDATION IN AQUEOUS-SOLUTION AND IN SOLID-STATE, Pharmaceutical research, 13(8), 1996, pp. 1252-1257
Purpose. The aim of this work was to study the kinetics of oxidation o
f methionine in human Insulin-like Growth Factor I (hIGF-I)(I) in aque
ous solution and in the solid state by the aid of quantification of ox
ygen. Methods. The oxidized form of hIGF-I was characterized by trypti
c peptide analysis, RP-HPLC and FAB-MS and quantified by RP-HPLC. The
oxygen content was quantified polarographically by a Clark-type electr
ode. Results, Second-order kinetics with respect to amount of protein
and dissolved oxygen was found to be appropriate for the oxidation of
methionine in hIGF-I. The rate constants ranged from 1 to 280 M(-1) mo
nth(-1) and had an activation energy of 95 (+/- 4) kJ/mole. Light expo
sure, storage temperature and oxygen content were found to have a cons
iderable impact on the oxidation rates. No significant difference in r
eaction rates was found for the oxidation of NGF-I in aqueous solution
or in the solid state. A method for decreasing the oxygen content in
aqueous solution without purging is described. Conclusions. Polarograp
hic quantification of dissolved oxygen makes it possible to establish
the kinetics for oxidation of proteins. The oxidation of methionine in
hIGF-I appears to follow second-order kinetics.