Keiji Kakita

Gel chromatographic separation of human C-peptide and proinsulin

The immunoreactivity of circulating C-peptide is separated into two main peaks on a Bio-Gel column; the faster peak should not be proinsulin but an associated C-peptide without a covalent bond. Proinsulin is in fact eluted in the fraction prior to the faster eluting peak of C-peptide immunoreactivity with 1 M acetic acid as the eluting buffer. Therefore the use of gel chromatography to study C-peptide and proinsulin needs to be carefully re-evaluated, although the method has been established as one of the standard methods.

Analysis of immunoreactive insulins in man in relation to the effects of aging

In order to elucidate the effects of aging on insulin content in human pancreas, the immunoelectrophoretic analysis of insulin was applied. The pancreas of senile humans appears to contain less total immunoreactive insulin, because of the decrement of proinsulin fraction after overnight fasting. The results suggest the involvement of insulin biosynthesis according to the aging process.

Insulin and C-peptide secretion from B cells in human subjects stimulated with glucose

Two groups of immunoreactive insulin in human sera were reported by Kakita et al. (4), using gel chromatography after acid-alcohol extraction. These analogs were noted not only in circulating human sera but also in incubation medium and incubated human pancreas. The release of these insulin analogs was discussed in a previous report (5). The circulating C-peptide immunoreactivity was separated into two groups on a Bio-Gel column, and the early peak should not be proinsulin but an associated C peptide (6). These analogs of insulin were separated by the methods of ion-exchange chromatography, isoelectric focusing, gel electrophoresis, and gel chromatography. Immunoreactive insulin was also separated into two major bands by standard polyacrylamide gel electrophoresis. The fast migrating band corresponds to the rat insulin II position, and the slower corresponds to rat insulin I, which has one more basic amino acid residue in comparison with rat insulin II. Further studies have been performed in five healthy adults in order to elucidate the physiological relationship between analogs of insulin and C-peptide peak substances in human serum; the results are reported in this paper with a consideration of the mechanism of insulin secretion.