Action of l-Ascorbic Acid upon the Isolated Frog Heart. ∗

Abstract

It was recently reported that hydrogen peroxide is the cause of the positive inotropic and of the irreversible systolic effect upon the frog heart of ascorbic acid solutions. 1 The evidence available in the literature indicates that the hydrogen peroxide formation in ascorbic acid solutions is related to the dehydrogenation of the molecule, and, more specifically, to the catalytic action of copper in this process. 2 It is known that the catalytic action of copper can be prevented by the formation of complexes between the metal and suitable substances like protein 3 or diethyl-dithiocarbamate. 4 Serum globulin and sodium diethyldithiocarbamate were therefore used to determine whether copper inhibitors were able to diminish or prevent the action of l-ascorbic acid solutions upon the frog heart.

Method. 50 to 100 mg of l-ascorbic acid were dissolved in one cc of distilled water; and this solution, after neutralization with sodium bicarbonate, was diluted to an initial concentration of approximately 1:10,000 with bicarbonate buffer solution (NaCl 85.4; KCl 1.9; CaCl₂ 0.99; NaHCO₃ 27.0 mM/L) saturated with 95% oxygen and 5% carbon dioxide. The pH of the solution was between 7.5 and 7.8. The room temperature varied between 20 and 25°C. In the experiments with inhibitor, this was dissolved in the bicarbonate buffer solution previous to the addition of the l-ascorbic acid. The serum globulin was prepared in the laboratory of Dr. E. J. Cohn and consisted of 90% gamma-globulin and 10% beta-globulin.

The frog hearts were isolated as previously described, 1 and the bicarbonate buffer solution was continuously perfused through the cannula at a rate of 2 to 2.5 cc per minute, with a gas mixture of 95% oxygen and 5% carbon dioxide bubbling through the fluid in the cannula.

The ascorbic acid determinations were made with the 2,6-dichlorindophenol method.