The Preparation and Evaluation of Tritiated Polyalanyl Insulin Derivatives1

A series of polyalanyl insulin derivatives (PAI) have been synthesized and characterized. Their content of added alanine ranged from 0.49 to 16.6 moles per mole of insulin. PAI derivatives with an average of up to 3.75 moles of added alanine per mole of insulin retained full biological activity as measured by the epididymal fat pad assay. Data from phenylisothiocyanate degradation indicate that 87% of the molecules in such a derivative probably possess at least one added alanine, so it is concluded that insulin with minimal addition of alanine is biologically active. A preparation of high specific activity tritiated polyalanyl insulin has also been synthesized and shown to be fully biologically active by the epididymal fat pad assay. Preliminary autoradiographic studies using this insulin derivative demonstrate that the hormone is bound to the sarcolemmal membranes of both striated and cardiac muscle. 3H-polyalanyl insulin was also concentrated in the proximal tubules of the kidney, but barely detectable in the liver at seven minutes after intravenous injection. (Endocrinology90: 112, 1972)