PURPOSE: Feasibility of solid-state 1H NMR for determining the mobility of protein molecules in lyophilized cakes was considered. The mobility in cakes with various levels of water content was studied in relation to aggregation-susceptibility. METHODS: Spin-spin relaxation time T2) of protons in lyophilized bovine serum albumin (BSA) and gamma-globulin (BGG) was measured as a function of hydration level by solid state 1H NMR using a 'solidecho' pulse sequence. Moisture-induced aggregation of the lyophilized proteins was also determined by high performance size exclusion chromatography. RESULTS: Lyophilized BSA and BGG became susceptive to aggregation when water content exceeded about 0.2 g/g of protein. T2 of protein protons in the lyophilized cakes started to increase at lower water contents. The increase in aggregation susceptibility observed with increasing water content appears to follow the increase in T2 of protein protons. For lyophilized BGG, both aggregation and T2 of protein protons decreased at water contents above 0.5 g/g protein. CONCLUSIONS: Mobility of protein molecules in lyophilized cakes was successfully determined by solid-state 1H NMR. The aggregation susceptibility of proteins was strongly related to their molecular mobility as indicated by T2.
PURPOSE: Feasibility of solid-state 1H NMR for determining the mobility of protein molecules in lyophilized cakes was considered. The mobility in cakes with various levels of water content was studied in relation to aggregation-susceptibility. METHODS: Spin-spin relaxation time T2) of protons in lyophilized bovineserum albumin (BSA) and gamma-globulin (BGG) was measured as a function of hydration level by solid state 1H NMR using a 'solidecho' pulse sequence. Moisture-induced aggregation of the lyophilized proteins was also determined by high performance size exclusion chromatography. RESULTS: Lyophilized BSA and BGG became susceptive to aggregation when water content exceeded about 0.2 g/g of protein. T2 of protein protons in the lyophilized cakes started to increase at lower water contents. The increase in aggregation susceptibility observed with increasing water content appears to follow the increase in T2 of protein protons. For lyophilized BGG, both aggregation and T2 of protein protons decreased at water contents above 0.5 g/g protein. CONCLUSIONS: Mobility of protein molecules in lyophilized cakes was successfully determined by solid-state 1H NMR. The aggregation susceptibility of proteins was strongly related to their molecular mobility as indicated by T2.