Molecular\nInvestigation of SNAC as an Oral Peptide\nPermeation Enhancer in Lipid Membranes via Solid-State NMR

Oral peptide therapeutics are increasingly favored in\nthe pharmaceutical\nindustry for their ease of use and better patient adherence. However,\nthey face challenges with poor oral bioavailability due to their high\nmolecular weight and surface polarity. Permeation enhancers (PEs)\nlike salcaprozate sodium (SNAC) have shown promise in clinical trials,\nachieving about 1% bioavailability. One proposed mechanism for enhancing\npermeation is membrane perturbation or fluidization, though direct\nexperimental proof and quantitative analysis of these effects are\nstill needed. This study employs solid-state NMR (ssNMR) to investigate\nhow SNAC interacts with hydrated DMPC liposomes, measuring enhancements\nin membrane fluidity across interfacial and transmembrane regions.\nThe methodology involves analyzing phosphate lipid headgroups and\nacyl chains using static <sup>31</sup>P chemical shift anisotropy\nand <sup>2</sup>H quadrupolar coupling measurements alongside <sup>1</sup>H and <sup>13</sup>C magic angle spinning NMR for motional\naveraging of <sup>1</sup>H–<sup>1</sup>H and <sup>1</sup>H–<sup>13</sup>C dipolar couplings. Our findings indicate an overall increase\nin the uniaxial motion of phospholipids with SNAC in a PE concentration-dependent\nmanner. It boosts lipid headgroup dynamics and enhancement plateaus\nat 25% between 24 and 72 mM concentrations. SNAC effectively enhances\nthe fluidity of the hydrophobic center by 43% at 72 mM PE concentration,\nmore significantly than the interfacial region. It is worth noting\nthat the extent of liposome dissolution and conversion to micelles\nincreases as SNAC concentration rises. Including a model peptide drug,\noctreotide, introduces a competitive equilibrium in this complex PE–lipid–peptide\nsystem, further influencing membrane dynamics for peptide permeation.\nInterestingly, the membrane enhancement does not show the expected\nplateau, and a less significant lipid mobility increase is observed\nin the presence of octreotide, suggesting a less substantial impact\ncompared to peptide-free systems, which is likely due to peptide–PE\ninteractions that consume monomeric SNAC, reducing its interaction\nwith the lipid membrane. This study provides the first quantitative\nand site-specific ssNMR measurements of membrane mobility influenced\nby one representative PE as a snapshot of PE lipid interaction in\na liposome model, demonstrating how peptide drugs modulate competitive\nequilibria and PE-induced lipid dynamics.