Erythromycin and other antibiotics have been used empirically in the treatment of patients with chronic obstructive pulmonary disease (COPD). We studied whether this empirical role of antibiotics might not be related to a possible direct effect on respiratory glycoconjugate (RGC) secretion. The effect of erythromycin on RGC secretion and hypersecretion was studied in an in vitro preparation of human airways that were secreting [3H]glucosamine respiratory glycoconjugate (RGC), and on a human endometrial adenocarcinoma cell line secreting a glycoconjugate (tumor glycoconjugate = TGC) chemically similar to the TRC secreted by the airways. Erythromycin at 10-5 M reduced TRC secretion by 35 ± 4% (n = 9, p < 0.001) in both human airways and the adenocarcinoma cells, and was increasingly active in the pharmacologic range of 10-7 to 10-4 M. The inhibitory effect of erythromycin was maximal within 16 h and was still evident 34 h after incubation. Erythromycin was noted to reduce both spontaneous (baseline) and stimulated RGC secretion (by histamine and methacholine) from airways in culture. The blocking effect appeared to be more selective for histamine than methacholine. These effects were not associated with any toxicity to the tissues and were not associated with the inhibition of protein synthesis. Dexamethasone also inhibited RGC release in both assay systems and exhibited dose-related effects in the physiologic ranges (10-9 to 10-5 M). When administered together, erythromycin and dexamethasone had an additive inhibitory effect on RGC secretion (68.0 ± 3.0%, n = 7, p < 0.001). Other antibiotics such as penicillin, ampicillin, tetracycline, and cephalosporin, in the range of 10-7 to 10-2 M, did not have any effect on RGC secretion from either the airways or the tumor cells.