EFFECT OF SIZE AND CHARGE ON THE PASSIVE DIFFUSION OF PEPTIDES ACROSSCACO-2 CELL MONOLAYERS VIA THE PARACELLULAR PATHWAY

Purpose. To evaluate the effect of size and charge on the permeation c haracteristics of peptides across the intestinal mucosa. Methods. The lipophilicities of neutral, positively and negatively charged capped a mino acids (Asn, Lys, Asp), tripeptides (Ac-Gly-X-Ala-NH2; X = Asn, Ly s, Asp) and hexapeptides (Ac-Trp-Aia-Gly-Gly-X-Ala-NH2; X = Asn, Lys, Asp) were estimated using an immobilized artificial membrane. The diff usion coefficients used to calculate the molecular radii were measured by NMR. The transport characteristics of the model peptides were dete rmined across Caco-2 cell monolayers. Results, When model compounds ha ving the same charge were compared, permeation was highly size-depende nt (capped amino acids > tripeptides > hexapeptides), suggesting trans port predominantly via the paracellular route. For example, the flux o f the negatively charged Asp amino acid (P-app = 10.04 +/- 0,43 x 10(- 8) cm/s) was 3 times greater than that observed for the Asp-containing hexapeptide (P-app = 3.19 +/- 0.27 x 10(-8) cm/s). When model compoun ds of the same size were compared, permeation across the cell monolaye r was charge-dependent (negative < positive less than or equal to neut ral). For example, the neutral, Asn-containing tripeptide (P-app = 25. 79 +/- 4.86 x 10(-8) cm/s) was substantially more able to permeate the Caco-2 cell monolayer than the negatively charged Asp-containing trip eptide (P-app = 7.95 +/- 1.03 x 10(-8) cm/s) and the positively charge d Lys-containing tripeptide (P-app = 9.86 +/- 0.18 x 10(-8) cm/s). The permeability of the cell monolayer to peptides became less sensitive to net charge as the size of the peptides increased. Conclusions. A po sitive net charge of hydrophilic peptides enhances their permeation ac ross the intestinal mucosa via the paracellular pathway. With increasi ng molecular size, molecular sieving of the epithelial barrier dominat es the transport of peptides, and the effect of the net charge becomes less significant.