Early Detection of Degraded A₁₄-¹²⁵I-Insulin in Human Fibroblasts by the Use of High Performance Liquid Chromatography

We studied the metabolism of A₁₄-¹²⁵I-Insulin in intact human fibroblasts using high performance liquid chromatography (HPLC) to detect and separate its early degradation products. The high resolving power of HPLC enabled us to separate what has been considered “intact insulin” by Sephadex G-50 chromatography or TCA precipitability into two additional peaks that had decreased biochemical properties with respect to immunoprecipitability and receptor binding but not decreased TCA precipitability. We conclude that human fibroblast is capable of metabolizing insulin within 2 min at 37°C into intermediate molecules that can be detected by HPLC but not by TCA precipitability or molecular sieve chromatography.