Our recent finding of the co-localization of 11-cis retinyl esters and 11-cis retinyl ester hydrolase (11-cis REH) activity in bovine retinal pigment epithelium (RPE) plasma membrane (PM) has led us to explore the possibility that the PM may provide 11-cis retinal for rhodopsin regeneration. In the RPE, visual chromophore is synthesized via a membrane associated 11-cis retinol dehydrogenase (11-cis RD). Accordingly, bovine RPE membranes enriched with either endoplasmic reticulum (ER) or plasma membrane (PM) enzyme markers were prepared and assayed for visual cycle enzyme activities. Pronounced 11-cis RD activity was associated with both ER- and PM-enriched membrane fractions. In contrast, 11-cis REH activity was mostly recovered in PM-enriched fractions while LRAT activity was found only in ER-enriched membranes. The finding that both 11-cis retinol and 11-cis retinal can be produced at the PM of the bovine RPE strongly suggests that 11-cis retinyl esters at this subcellular locale serve as a precursor of visual chromophore for pigment regeneration.