The glycosaminoglycans synthesized by 5 corneal explants with macular corneal dystrophy were analyzed by sequential degradation using heparitinase (and/or butyl nitrite), chondroitin ABC lyase, chondroitin AC lyase, and keratan sulfate endo-beta-galactosidase. The tissue was first pulse-labeled by incubation in phosphate-buffered saline (pH 7.3) containing 100 muCi/ml 3H-glucosamine and approximately 1000 muCi/ml 35S-sulfate for 1 hour and chased with Eagle's minimum essential medium containing 10% fetal calf serum. Notable differences were detected between the products of these corneal explants with macular corneal dystrophy and controls that were prepared and analyzed in the same manner. As assayed by its susceptibility to keratan sulfate endo-beta-galactosidase degradation, corneal tissue with macular corneal dystrophy incorporated less 35S-sulfate, and usually less 3H-glucosamine, into keratan sulfate than normal corneas. The corneal explants with macular corneal dystrophy also synthesized prominent fractions which eluted from DEAE-Sephacel columns in 0.05 M tris(hydroxymethyl)aminomethane buffer (pH 7.2) with 0.15 M and/or 0.25 M lithium chloride after pronase digestion. Since normal corneas produced small quantities of material with a similar elution profile after preparation and analysis under identical conditions it is possible that the fractions are normal synthetic products of the cornea. The significance of these findings in the pathogenesis of macular corneal dystrophy remains to be determined.