This ones on hIGF-1 thanks einstein1905

Pharmaceutical Research. Vol. 14, No.5. 1997

Solvent Effects on the Solubility and Physical Stability of Human Insulin- Like Growth Factor I

Purpose. The solubility and physical stability of human Insulin-like Growth Factor I (hIGF-l) were studied in aqueous solutions with differ- ent cxcipients.

Methods. The solubility of hlGP-r was determined by UV-absorption and quantification of light blocking particles. The physical stability of hlGF-r was studied with differential scanning caJorimetry (DSC) and circular dichroism (CD) spectroscopy.

Results. Human IGF-l precipitated at Jow temperature in the presence of J40 mM benzyl alcohol and 145 mM sodium chloride. CD data showed that the tertiary structure of hIGF-I during these condions was perturbed compared to that in 5 mM phosphate buffer. In the presence of benzyl alcohol 290 mM mannitol stabilized hIGF-I. Sodium chloride or mannitol by themselves had no effect on either the solubility or the tertiary structure. Benzyl alcohol was attracted to hIGF-l, whereas sodium chloride was preferentially excluded The attraction of benzyl alcohol WIlS reinforced by sodium chloride leading to salting-out of hlGF-l. The CD-data indicated interactions of benzyl alcohol with phenylalanine in hIGF-i. Thermal denaturation of hIGF-I occurred in all solutions with sodium chloride. whereas mannitol or benzyl alcohol had no effect on the thermal stability. The thermal stability of hIGF- I WIlS thus decreased in 145 mM sodium chloride although it WIlS excluded from hIGF-I.

Conclusions. The self-association and thermal aggregation of hIGF-I is driven by hydrophobic interactions. Benzyl alcohol is attracted to hIGF-I and induces changes in the tertiary structure causing hydropho- bic attraction of the protein at low temperatures.

Jonas Fransson,1,2,4 Dan Hallen,3 and Ebba Florin-Robertsson, 1

Received October 29, 1996; accepted February 19. 1997

1 Department of Pharmaceutical Technology, Pharmacia & upjohn, S-112 87 Stockholm, Sweden

2 Department of Pharmaceuticlas, Faculty of Pharmacy, Uppsala University, Box 570, S-751 23 Uppsala, Sweden

3 Department of Structural Chemistry, Parmacia & Upjohn, S-112 87 Stockholm, Sweden

4 To whom correspondence should be addressed. (e-mail: [email protected])

JohnnyB