Diphtheria toxin; a reinvestigation of the effect of iron on toxin and porphyrin production.

In diseases such as diphtheria and tetanus, almost all the clinical symptoms terminating in deat.h are due to specific soluble toxins produced in civo (or in vitro) by the diphtheria and t&anus bacilli respectively. Although diphtheria toxin has been isolated as an essentially pure protein which has been reasonably well characterized both chemically and physically (l-3), almost’ nothing is known of the primary mechanism by which this highly poisonous protein causes injury to the tissues of the susceptible host. The pathological changes brought about in animals and in man have been extensively studied, and recently attempts to demonstrate biochemical changes in intoxicated animals have been reviewed by Holmes (4). No enzyme system has as yet been discovered which is inhibited by diphtheria toxin, nor has it been possible to demonstrate enzymic activity with purified preparations of the toxin. It is our opinion that the gross lesions caused by the toxin, including the typical lesions of the adrenal glands and the metabolic changes which have been observed in intoxicated animals, represent secondary changes which follow by many hours or days a primary irreversible injury occurring a few moments after the toxin has been injected. It is the nature of this primary injury which the present research is designed to elucidate. It has occurred to us that, if the r&e which the toxin plays in the metabolism of the diphtheria bacillus itself can be determined, then it may well be possible to predict the manner by which it causes damage to the cells of higher animals. A promising. clue in this direction is provided by the well known inhibition of toxin production caused by the addition of small amounts of iron to the medium on which the diphtheria bacillus is cultivated (5, 6). At very low iron concentrations both growth and toxin production are poor. As iron is added, growth and toxin increase rapidly, the latter reaching a maximum biter in the culture filtrate when about 0.1 mg. of iron is present per liter of medium. As further iron is added, growth continues to increase slightly (10 to 20 per cent) but toxin production falls rapidly and practically none is found in culture filtrates from media con-