Polarographic Studies of Proteins and their Degradation Products. V. Products from Tryptic Digests of Serum Albumin and Development of a Sensitive Polarographic Trypsin Assay.*

Plasma protein fractions show characteristic differences in their polarographic behavior after being subjected to alkaline digestion. Serum albumin (fraction V) in particular is unique in that it produces a catalytic double-wave in divalent cobalt buffer which increases markedly during the first 10 20 minutes of digestion and then decreases gradually during the following hour. During this same period of digestion no sulfosalicylic acid-soluble split products are formed which produce catalytic polarographic waves in trivalent cobalt buffer. When the same albumin is subjected to tryptic digestion instead of alkaline denaturation, the results are quite different. Not only does the doublewave in divalent cobalt buffer increase without a decay reaction (at least during 5 hours) as the digestion progresses, but also split products are formed that are sulfosalicylic acid-soluble and give a catalytic wave in trivalent cobalt buffer. This catalytic wave is peculiar in that it shows only the first peak of the catalytic double wave". This finding led to a more detailed study of the proteolytic action of trypsin which involved an investigation of the stability of trypsin preparations5' and a comparison of polarographic data with known methods for the assay of tryptic activity6', as far as possible. It has also led to the development of a sensitive and rapid polarographic method for assaying the activity of trypsin which is presented here. Trypsin and its proteolytic action have been studied polarographically on numerous occasions. It was found that trypsin itself, like other proteins, produces a catalytic double wave in divalent cobalt buffer and that the relationship between trypsin concentration and wave height resembles a Langmuir adsorption isotherm. The wave height of trypsin is much smaller than that of plasma at corresponding concentrations and the second wave