The water replacement theory discussed in Section 3.1 stated the importance of stoichiometry of sugar to protein in providing adequate hydrogen bonding interaction for protein stabilization. The effectiveness of sugars as stabilizers for proteins can be subject to the ratio of sugar to protein in the mixture. Sugars might not provide an adequate stabilization effect during the process if the ratio of excipient to protein is not sufficient. The final mass ratio of sugar to protein determines the stabilization power for protein protection during drying. recent study has demonstrated that a 360:1 molar ratio which is about 1:1 w/w of sugar to protein was essential to ensure good storage stability of protein. In general, the optimal weight ratio of 3:1 to 5:1 of sugar to protein would be ideal for inhibiting lyophilization-induced unfolding. However, the optimal ratio between sugar and protein for protein stabilization can be sugar-type dependent. Lu et al. found that the denature temperature gradually increased with an increase sucrose to protein ratio while decreasing with an increase in trehalose, xylitol, and lactitol to protein ratio. Later on, a more comprehensive study have been done to investigate the impact of sucrose levels on protein stability from both thermodynamic and kinetic aspects. The study found a sucrose to protein ratio saturation point that the extent of five protein's structure protection from FTIR by sucrose reached a plateau after reaching a mass ratio of 1:1, though the stabilization effect limiting the aggregation behavior is sucrose concentration dependent. Interestingly, the fast local dynamic measured NS as <u 2 > demonstrated linear correlation with the log scale of aggregation rate constant over the entire sucrose range studied. As such, it is critical to evaluate the sugar to protein ratio on the protein's physical stability from both structure preservation and fast dynamic changes during the formulation composition optimization.