Lipid peroxidation in porcine irises: Dependence on pigmentation

Purpose. Pigmentation of the iris is caused by varying amounts of melanin p igment granula in a constant number of melanocytes in the superficial strom a. Melanin has been shown to act as antioxidant. We have now investigated l ipid peroxidation in dependence on stromal pigmentation in isolated porcine irises. Methods. The same number of lightly and heavily pigmented porcine irises (v isual selection) were homogenized (1:20w/v) in buffer (50 mmol/l phosphate buffer and 4 mmol/l sodium azide). 500 mul homogenate were incubated at 37 degreesC in duplicate for 5, 10, 20 and 40 min in absence and presence of F e2+ as inducer of lipid peroxidation. The amount of lipid peroxidation was assayed by the thiobarbituric acid (TBA) test. The results are expressed as nmol of TEA reactive material (TBAR) produced/mg protein. Fe2+ concentrati on of the supernatant was determined spectrophotometrically with 1,10 ortho phenanthroline. Concentrations of D-glucose and D-fructose in iris tissue h omogenates were determined spectrophotometrically by enzymatic bioanalysis. Results. 70, 180 and 360 mu mol/l Fe2+ induced lipid peroxidation. A platea u region was reached after 20 min. The amount of lipid peroxidation differe d in dependence on stromal pigmentation in porcine irises. The effect was m ost significant at 180 mu mol/l Fe2+, which induced 1.373 +/- 0.138 nmol TB AR/mg protein in lightly compared to 0.491 +/- 0.125 nmol TBAR/mg protein i n heavily pigmented irises after 10 min incubation (p < 0.0001, n = 4). Sim ilar effects (factor 2-3) were also measured after 20 and 40 min incubation . On the other hand the content of Fe2+ in the supernatant was the same wit hin error. Sugar concentrations (D-glucose and D-fructose) did not differ s ignificantly for the two differently pigmented iris tissues. Conclusions. There is a stronger induction of lipid peroxidation in lightly compared to heavily pigmented porcine irises. This effect may be related t o the difference in stromal melanin content and its antioxidant activity.