Although relatively rare, retinopathy based on a disturbed metabolism of the retinal pigment epithelium (RPE), with ensuing degeneration of photoreceptors, is a known complication of treatment with the 4-aminoquinolones, chloroquine (CQ) and hydroxychloroquine (HCQ), in autoimmune diseases. The reported frequency of retinopathy, however, is much lower for HCQ than for CQ (less than 0.08% versus 1-2%). To test whether the difference in toxicity between the two lysosomotropic drugs is related to different lysosomal influence, we exposed confluent RPE cell cultures to CQ or HCQ for 2 weeks. To induce lipofuscin (LF) formation, known to be accelerated by increased lysosomal pH and intra-lysosomal oxidation during degradation of auto-/heterophagocytosed material, such treatment was combined with feeding of cells with photoreceptor outer segments (POS) and hyperoxia (40% ambient oxygen). HCQ was found to be a less potent enhancer of lipofuscinogenesis compared to CQ, apparently due to its less effective inhibition of lysosomal degradative capacity (evaluated by vital staining of lysosomes with Lyso Tracker Red, and periodic acid-Schiff reaction). This conclusion is supported by the fact that NH4Cl, a non-fluorescent substance which acts similarly to 4-aminoquinolones, induced an increase in LF fluorescence paralleled by increased periodic acid-Schiff reactivity of RPE cells.