Biochemical studies on experimental congenital malformations: flavin nucleotides and folic acid in fetuses and livers from normal and riboflavin-deficient rats.

In 1940, Warkany and Nelson observed that congenital malformations could be induced in the offspring of Sprague-Dawley rats by feeding the mothers a deficient, but poorly characterized, diet (2). These developmental anomalies, primarily of skeletal nature, were subsequently shown to result from a specific deficiency of riboflavin (3,4). Prolonged ingestion of the riboflavindeficient diet, for 4 to 5 weeks before pregnancy, as well as throughout the critical part of the gestation period, was necessary for the development of anomalies. Supplementation of the diet with the riboflavin antagonist, galactoflavin (5), provided a more acutely teratogenic milieu and simplified the experimental procedure considerably. Malformations could be induced by instituting the riboflavin-deficient diet on the first day of pregnancy if galactoflavin was added throughout the gestation period or on selected days of pregnancy when differentiation of specific systems was proceeding (6, 7). Moreover, use of the vitamin antagonist increased the number of abnormal young and extended the types of malformations. Precisely how an acute riboflavin deficiency influences differentiating embryonic tissue and misdirects its normal pathways is not yet understood. A reasonable working hypothesis would be that specific end products of the action of certain flavin enzymes are essential for normal differentiation and development. According to this concept, if the flavin concentration of the embryo, or certain areas of the embryo, falls below a required critical level, anomalies will result (4). The purpose of the present study is to test this hypothesis by attempting to establish the critical level of flavin nucleotide concentration. The results obtained indicate that the total riboflavin and flavin adenine dinucleotide concentration of differentiating embryos can be reduced as much as 30% without ill effects, but that a 60% reduction in flavin adenine dinucleotide content is teratogenic. The experimental congenital malformations induced in the offspring of pregnant rats made severely deficient in riboflavin have certain features in common with malformations induced by