THE CHEMICAL BASIS OF THE PERIODIC ACID SCHIFF REACTION OF COLLAGEN FIBERS WITH REFERENCE TO PERIODATE CONSUMPTION BY COLLAGEN AND BY INSULIN

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Abstract
1. Oxidation of collagen fibers in tissue sections or of Standard Hide Powder (hide collagen) in the testtube by an acid solution of periodate (pH 1.9) resulted in a product which gave a positive (pink to red) Schiff reaction. Oxidation by an alkaline solution of periodate (pH 8.2) gave an equivocal (faint pink) reaction. 2. The reaction between the insoluble oxidation product in collagen and the Schiff reagent was blocked completely, both in tissue sections and in the testtube, by an aqueous solution of aniline chloride, phenylhydrazine, or dimedone at room temperature, but not by the aqueous solvent alone, indicating that free insoluble aldehyde groups were produced as the result of acid periodate oxidation. 3. The acid periodate Schiff reaction of collagen fibers was not due to the presence of lipid or glycogen and did not depend upon chemical fixation of the tissue. 4. Periodate cleavage oxidation of the ε-amino-δ-hydroxy grouping of hydroxylysine in collagen did not seem to play a significant role in the acid periodate Schiff reaction. 5. Periodate cleavage oxidation of the serine and threonine constituents of collagen probably does not occur. 6. The largest fraction of carbohydrate in the dry hide powder was of nonglucosamine polysaccharide, calculated as 0.55% (expressed as glycogen). The quantity of hexosamine and uronic acids in the dry, defatted hide powder was 0.047% and less than 0.039% respectively. By indirect evidence it is deduced that the nonglucosamine polysaccharide complex conjugated with collagen most likely was responsible for the acid periodate Schiff reaction. 7. A large fraction of the periodate consumed by hide collagen could not be accounted for by oxidation of the carbohydrate content, by oxidation of vicinal hydroxy amino groups in the protein, or by nonspecific adsorption of –IO4. Evidence based upon the experiments with purified insulin indicates that periodate may be consumed by protein without the production of any aldehydes or free ammonia. This is suggested as the fate of periodate not otherwise accounted for in oxidized hide collagen.