The statistical analysis confirmed that both temperature and time exerted highly significant main effects on the CQAs of the VLP vaccine ( p < 0.0001), and a significant interaction was identified between the two factors ( p < 0.0001).
At 25 °C, over 28 days, the relative content of basic variants showed a decreasing trend (from 12.07% to 4.22%), and a shift exceeding 1.5% in the relative content of charge variants was observed by day 3, with the isoelectric point trending toward the acidic direction ( Figure 1 B).
In the groups supplemented with trehalose and sucrose, both T m and T agg showed an increasing trend of 2–4 °C, with trehalose increasing T m from 70.6 °C to 72.9 °C and T agg from 64.0 °C to 66.5 °C, sucrose increasing T m from 70.6 °C to 74.1 °C and T agg from 64.0 °C to 66.8 °C.