The role of mucus glycoprotein, lipid and protein in the retardation of hydrogen ion diffusion by dog gastric mucus was investigated. The uniform layer of mucus (1.0 +/- 0.2mm thick), formed on the sintered glass support placed between equimolar (0.155M) solutions of HCl and NaCl, caused a 73% reduction in hydrogen ion diffusion when compared with a layer of NaCl. The rate of hydrogen ion diffusion through purified mucus glycoprotein was similar to that of native mucus, whereas a 30% increase in permeability of glycoprotein to hydrogen ion was observed following its delipidation. Preincubation of the purified mucus glycoprotein with serum albumin or IgA led to an increase in the ability of glycoprotein to retard the diffusion of hydrogen ion. This effect of albumin and IgA occurred at a physiological dose. About 70% decrease in hydrogen ion retardation was obtained following digestion of purified mucus glycoprotein with pepsin, and 74% following its digestion with pronase. The results show that the ability of gastric mucus to retard the diffusion of hydrogen ion depends upon the polymeric structure of undegraded mucus glycoprotein and the extent of interaction of this glycoprotein with lipids, serum albumin, and secretory IgA.