First, the porous structures observed under simulated acidic conditions can house mRNA payloads inside the water-filled pores, as observed in our in silico self-assembly simulations. Incorporation of mRNA payloads into these porous structures under acidic conditions could explain the high loading efficiency observed with bleb structures that form under acidic conditions. Specifically, it has recently been reported that very high (~90%) loading and in vivo transfection efficiencies are observed for LNP/mRNA systems when formulated at buffered pH 4 using Na-citrate. 42 b Determined from a linear fit between the points that span the value where the number of protonated and deprotonated lipids are equal. c Determined from a sigmoidal fit of the data. d Determined using a Henderson-Hasselbach fit of the data At the same time, previous work suggests that blebs with higher loading efficiency correspond to polar regions of the LNPs, 72, which agrees with the highly protonated nature of the systems simulated in acidic conditions. Lastly, the formation of bleb structures is known to be dependent on the species of the ionizable lipid used in their formulation, 40 which is inherently related to the pKa of an ionizable lipid, the apparent pKa of the LNP, and the environment-dependent distributions of pKa values within an LNP. Overall, we anticipate that the ability to predict such interactions computationally represents a powerful tool, which will be able to assist in advancing the basic understanding of the complex structure-activity relationships involved in LNP formulations, while also aiding and abetting in the design of new ionizable lipids that could further improve loading efficiency and/or to theoretically screen for optimal formulation conditions in the future.