J. Fransson et al., SOLVENT EFFECTS ON THE SOLUBILITY AND PHYSICAL STABILITY OF HUMAN INSULIN-LIKE-GROWTH-FACTOR-I, Pharmaceutical research, 14(5), 1997, pp. 606-612
Purpose. The solubility and physical stability of human Insulin-like G
rowth Factor I (hIGF-I) were studied in aqueous solutions with differe
nt excipients. Methods. The solubility of hIGF-I was determined by UV-
absorption and quantification of light blocking particles. The physica
l stability of hIGF-I was studied with differential scanning calorimet
ry (DSC) and circular dichroism (CD) spectroscopy. Results. Human IGF-
I precipitated at low temperature in the presence of 140 mM benzyl alc
ohol and 145 mM sodium chloride. CD data showed that the tertiary stru
cture of hIGF-I during these conditions was perturbed compared to that
in 5 mM phosphate buffer. In the presence of benzyl alcohol 290 mM ma
nnitol stabilized hIGF-I. Sodium chloride or mannitol by themselves ha
d no effect on either the solubility or the tertiary structure. Benzyl
alcohol was attracted to hIGF-I, whereas sodium chloride was preferen
tially excluded. The attraction of benzyl alcohol was reinforced by so
dium chloride leading to salting-out of hIGF-I. The CD-data indicated
interactions of benzyl alcohol with phenylalanine in hIGF-I. Thermal d
enaturation of hIGF-I occurred in all solutions with sodium chloride,
whereas mannitol or benzyl alcohol had no effect on the thermal stabil
ity. The thermal stability of hIGF-I was thus decreased in 145 mM sodi
um chloride although it was excluded from hIGF-I. Conclusions. The sel
f-association and thermal aggregation of hIGF-I is driven by hydrophob
ic interactions. Benzyl alcohol is attracted to hIGF-I and induces cha
nges in the tertiary structure causing hydrophobic attraction of the p
rotein at low temperatures.