Incorporation of tritium-labeled leucine into proinsulin and insulin was used to indicate proinsulin synthesis and insulin formation. In these experiments the incorporation of labeled leucine into proinsulin and that into insulin were closely related. This incorporation was stimulated by addition of glucose and mannose but not by fructose. The glucose- and mannose-stimulated incorporation was inhibited by mannoheptulose. Inhibition by mannoheptulose was not relieved by addition of pyruvate. Pyruvate, with the addition of small amounts of fumarate and glutamate, failed to stimulate proinsulin and insulin synthesis even though insulin secretion was stimulated by tolbutamide or ribose. It was concluded that neither free glucose, nor pyruvate derived from glucose, acts as a signal to initiate biosynthesis. Possibly some more immediate metabolite of glucose or some enzyme or cofactor associated with its metabolism may be involved. Ribose and xylitol failed to stimulate the biosynthesis of proinsulin, and insulin. This, taken together with the reported stimulating effect of ribose and xylitol on insulin secretion, supports, for the islets, the postulate of Krass and LaBella for posterior pituitary hormones, that the pentose phosphate pathway is concerned with hormone storage and secretion rather than with hormone synthesis.