Moisture-induced aggregation of lyophilized proteins in the solid state.
A critical problem in the storage and delivery of pharmaceutical proteins is their aggregation induced by moisture. A model system has been elaborated and investigated to elucidate the mechanism of this phenomenon. When 10 mg of bovine serum albumin lyophilized from an aqueous solution of pH 7.3 are wetted with just 3 muL of a buffered physiological saline solution and incubated in the solid state at 37 degrees C, the protein progressively loses its solubility in water; e.g., after a 24 h incubation 97% of the protein becomes insoluble. This moisture-induced aggregation of albumin has been discovered to be due to an intermolecular S-S bond formation via the thiol-disulfide interchange reaction. The dependence of the extent of the solid-state aggregation on the amount and mode of addition of moisture and the atmosphere, additives, temperature, and history of the protein powder have been investigated. The moisture-induced solid-state aggregation has also been established and studied for three other lyophilized proteins: ovalbumin, glucose oxidase, and beta-lactoglobulin. In all cases, the loss of solubility is caused by thiol-disulfide interchange either alone or in combination with a conformational (noncovalent) process. The aggregation can be minimized by lyophilizing the proteins from acidic aqueous solutions, by adding inorganic salts, by co-lyophilizing the proteins with water-soluble polymers, or by controlling the moisture content at optimal levels.
Liu
W R
WR
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Langer
R
R
Klibanov
A M
AM