Protective Effects of Ascorbic Acid and α-Tocopherol on the In Vitro Oxidation of Equine Erythrocytes Caused by Extracts of Wilted Red Maple Leaves

• We examined whether ascorbic acid and α-tocopherol would reduce oxidative stress in equine erythrocytes.
• Percentage hemolysis and methemoglobin were used to determine oxidative damage caused by red maple.
• Dietary supplements effectively reduced oxidative damage caused by red maple leaves.
• Hemolysis and methemoglobin reverted to baseline levels when antioxidant supplementation was terminated.

Red maple leaf toxicosis in horses is a debilitating condition. Horses which ingest wilted leaves of red maple, as well as those of silver and sugar maple, experience depression, lethargy, pale mucous membranes, and brown discoloration of plasma and urine due to severe oxidation of erythrocytes. The objective of this study was to evaluate the protective effect of equine dietary supplementation with the antioxidants ascorbic acid and α-tocopherol on hemolysis and methemoglobin production of erythrocytes when challenged by extracts of wilted red maple leaves in bioassays. Horses were assigned to one of four groups which received either (1) no dietary supplement, (2) ascorbic acid, (3) α-tocopherol, or (4) both ascorbic ascorbic acid and α-tocopherol for 12 weeks. At weeks 1, 12, and 24 after starting supplementation, blood samples were collected from horses and incubated with either 0 or 75 μL of leaf extract. Hemolysis and methemoglobin production were determined based on spectrophotometry. After 12 weeks of ascorbic acid supplementation, percentage hemolysis and methemoglobin production were significantly reduced in blood samples incubated with 75 μL of leaf extract. In contrast, only percentage hemolysis was significantly reduced after 12 weeks of α-tocopherol supplementation. After 24 weeks, values for percentage hemolysis and methemoglobin had returned to their baseline levels after termination of supplementation at week 12. The combination of both supplements did not significantly reduce hemolysis or methemoglobin production. These results suggest that daily dietary supplements of ascorbic acid (and to a lesser degree α-tocopherol) have the potential to lessen the effects of erythrocyte oxidation which occurs during red maple leaf toxicosis.