E. P. Cannon

Antagonism of cyanide intoxication with murine carrier erythrocytes containing bovine rhodanese and sodium thiosulfate

Murine carrier erythrocytes containing bovine rhodanese and sodium thiosulfate are being explored as a new approach to antagonize the lethal effects of potassium cyanide in mice. Prior studies indicated that these carrier erythrocytes persist in the vascular system for the same length of time as normal erythrocytes and can enhance metabolism of cyanide to thiocyanate. The present studies demonstrate the ability of these carrier red blood cells containing rhodanese and thiosulfate to antagonize the lethal effects of cyanide either alone or in various combinations with sodium nitrite and/or sodium thiosulfate. Potency ratios are compared in groups of mice treated with sodium nitrite, sodium thiosulfate, and carrier erythrocytes containing rhodanese and sodium thiosulfate either alone or in various combinations prior to the administration of potassium cyanide. These results indicate that the administration of carrier erythrocytes containing rhodanese and thiosulfate alone can provide significant protection against the lethal effects of cyanide. These carrier erythrocytes potentiate the antidotal effect of sodium thiosulfate alone or the combination of sodium nitrite and sodium thiosulfate. The mechanisms of cyanide antagonism by these carrier erythrocytes and their broader conceptual significance to the antagonism of other chemical toxicants are discussed.

In vivo studies on rhodanese encapsulation in mouse carrier erythrocytes

Resealed erythrocytes containing sodium thiosulfate and rhodanese (CRBC) are being employed as a new approach in the antagonism of cyanide intoxication. In earlier in vitro studies, the behavior of red blood cells containing rhodanese and sodium thiosulfate was investigated with regard to their properties and their capability of metabolizing cyanide to thiocyanate. The present studies are concerned with the properties of these rhodanese-containing carrier erythrocytes in the intact animal. These carrier erythrocytes were administered intravenously and the survival of the encapsulated enzyme was compared with the administration (iv) of free exogenous enzyme. Also, the amount of leakage of the encapsulated rhodanese from the red blood cell was determined. The survival of the carrier red blood cell. prepared by hypotonic dialysis, was found to be characterized by a biphasic curve. There was an initial rapid loss of approximately 40 to 50% of the carrier cells with a t1/2 = 2.5 hr. Subsequently the remaining resealed annealed carrier erythrocytes persisted in the vascular system with a t1/2 = 8.5 days. When free exogenous rhodanese was administered directly into the vascular system, it was rapidly eliminated with a t1/2 = 53 min. Red blood cells containing sodium thiosulfate and rhodanese apparently are effective in vivo in the biotransformation of cyanide. In animals pretreated with encapsulated rhodanese and sodium thiosulfate, blood cyanide concentrations are appreciably decreased with a concomitant increase in thiocyanate ion, a metabolite of cyanide. When erythrocytes, which contained no rhodanese or sodium thiosulfate, were subjected to hypotonic dialysis, cyanide was not metabolized to any appreciable extent. Furthermore, carrier erythrocytes containing rhodanese and sodium thiosulfate were found to increase the protection against the lethal effects of cyanide by approximately twofold. The ability of these carrier erythrocytes alone to metabolize cyanide and to antagonize the lethal effects of cyanide reflects the potential of this new antidotal approach in the antagonism of chemical toxicants.

Organophosphorus Antagonism by Resealed Erythrocytes Containing Recombinant Paraoxonase

Parathion has the highest incidence of poisonings among the agricultural pesticides. An alternative conceptual approach was employed to prevent intoxication of paraoxon, the active metabolite of parathion, by using resealed carrier erythrocytes containing a highly purified recombinant paraoxonase (CRBC). The CRBC were found to be effective alone and synergistic with 2-PAM and atropine to protect over 1,000 lethal doses of paraoxon. This striking protection greatly exceeds any antidotal regimen ever reported against chemical toxicants. The present drug carrier model provides a general conceptual approach to develop specific antidotes against many other chemical toxicants for which no antidotes are presently available.

Determination of Organophosphorus Acid Anhydrase in Blood

In an attempt to develop a new conceptual approach to paraoxon antagonism by encapsulating organophosphorus acid anhydrase (OPA anhydrase) into carrier erythrocytes, it necessitated the spectral determination of OPA anhydrase activity in blood. OPA anhydrase determination usually is conducted in serum or plasma. This presents a problem, because blood contains various substances that interfere with the determination of OPA anhydrase. This enzyme used paraoxon as the substrate, liberating p-nitrophenol and diethylphosphate. OPA anhydrase has a broad substrate specificity and is capable of hydrolyzing other organophosphorus compounds, i.e., pesticides, nerve gases, etc. Blood OPA anhydrase activity was determined with paraoxon as the substrate. This reaction was terminated by adding hydrochloric acid to the reaction mixture. The p-nitrophenol then was extracted with dichloromethane, and p-nitrophenol was determined at 400 nm. The sensitivity of this method can be greatly enhanced by increasing the sample siz...