Undergraduate Honors Thesis Projects

Date of Award

Spring 4-2021

Document Type

Honors Paper

Degree Name

Biochemistry and Molecular Biology-BS

Department

Biology & Earth Science

Advisor

Jeffrey Lehman

First Committee Member

Louise Captein

Second Committee Member

Brandon Sinn

Keywords

Strong Oxidants, Erythrocytes, Equines, Acer spp., Methemoglobin, Hemolysis

Subject Categories

Animal Diseases | Animals | Comparative and Laboratory Animal Medicine | Disorders of Environmental Origin | Large or Food Animal and Equine Medicine | Medicinal and Pharmaceutical Chemistry | Natural Products Chemistry and Pharmacognosy | Other Medicine and Health Sciences | Other Pharmacy and Pharmaceutical Sciences | Veterinary Preventive Medicine, Epidemiology, and Public Health | Veterinary Toxicology and Pharmacology

Abstract

Maple toxicosis is a deadly condition affecting equines after they have ingested wilted leaves of the genus Acer (i.e., maple species). It is characterized by hemolytic anemia and methemoglobinemia. Although the toxins responsible for causing toxicosis have not been identified, they are thought to be strong oxidants. The objective of this study is to determine the effect of eight compounds (galloyl derivatives) found in Acer spp. on hemolysis and methemoglobin production in bioassays with equine erythrocytes. Seven of the compounds are known constituents of Acer spp. that are commercially available. One of the compounds--acertannin--was isolated and characterized from leaf extracts of red maple. Equine erythrocytes were incubated with each compound at concentrations of 0, 0.2, 0.4, 0.8 and 1.6 mg/ml. Toxicity was determined spectrophotometrically based on percentage hemolysis of erythrocytes and on the production of methemoglobin. Experiments were repeated three times. Of the eight compounds, only tannic acid and gallic acid showed a significant effect on hemolysis. Tannic acid caused a higher level of hemolysis than did gallic acid. Concentrations of tannic acid > 0.4 mg/ml caused significantly more hemolysis than the control. The amount of hemolysis caused by the highest concentration (1.6 mg/ml) was roughly 3 times higher than that of the control. For gallic acid, only the most concentrated treatment of 1.6 mg/ml resulted in a significant increase in hemolysis relative to the control. Pyrogallol had the greatest effect on the production of methemoglobin. As little as 0.2 mg/ml of pyrogallol significantly increased methemoglobin production (0.9% versus 56.7% for 0 mg/ml and 0.2 mg/ml, respectively). Quercetin, methyl gallate, gallic acid, and tannic acid significantly affected methemoglobin production, in most cases with as little as 0.2 mg/ml of compound. Acertannin had no effect on hemolysis or on methemoglobin. The results of this study show that maple toxicosis can be largely explained by the presence of tannic acid and pyrogallol which cause hemolysis and the production of methemoglobin, respectively.

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