Local minimum in fragility for trehalose/glycerol mixtures: implications for biopharmaceutical stabilization.

Approximately a decade ago it was observed that adding a small amount (5 wt %) of glycerol to trehalose could substantially improve the stability of enzymes stored in these glasses even though the final glass transition temperature (Tg) was reduced by ∼20 K. This finding inspired great interest in the fast dynamics of dehydrated trehalose/glycerol mixtures, leading to the observation that suppression of fast dynamics was optimal in the presence of ∼5 wt % of glycerol. It was also recognized that the fast dynamics should, in theory, be related to the fragility of these glass formers, but experimental confirmation of this hypothesis has been lacking for trehalose/glycerol mixtures or any other mixtures of this nature. In the present study a dynamic mechanical analyzer (DMA) was used to determine both the Tg and the kinetic fragility index (m) of trehalose/glycerol mixtures within the mass fraction range of 80-100 wt % of trehalose. It was found that the fragility index correlated with the mass fraction of trehalose in a nonmonotonic manner, with a local minimum between 87.5 and 95 wt % of trehalose, whereas the composition dependence of Tg was found to follow a Gordon-Taylor-like relationship, with no local minimum. The composition of 5-12.5 wt % glycerol in trehalose thus yielded a matrix that maximized the strong glass-forming contribution of glycerol, while minimizing its Tg lowering effect. This quantitative evidence supports speculation about the fragility characteristics of these mixtures that has been ongoing for the past decade. The DMA-based Tg and fragility determination method developed in this study represents a new approach for identifying optimal compositions for preservation of biologics.