THE STABILITY OF I$sup 131$-INSULIN IN SOLUTION AND ITS USE IN STUDIES OF THE FOREARM UPTAKE OF INSULIN IN MAN
Insulin was iodinated with /sup 131/I, and the excess /sup 131/I was removed by an Amberlite IRA-400 ion-exchange resin column in the chloride phase. This method gives, using 10 mC thiosulfate-free /sup 131/iodide with 6 mg insulin, specific activities of 0.2--0.7 mC /sup 131/I/mg protein. This represents much less than 0.1 atom I/molecule of insulin. In undiluted solution (1 mg/ml), insulin was not detectably removed by the column, but more dilute (1: 25 and 1: 250) solutions were considerably depleted of protein-bound counts. These counts could not be washed off the column by buffer or by dilute protein solutions. Paper electrophoresis showed that over 90% of the radioactivity migrated with insulin. The stability of a 1 mg/ml iodoinsulin solution at 4 deg C in glycine buffer at pH 9 was studied, to assess how soon before use it need be prepared. Samples of iodoinsulin were kept in the refrigerator for varying times of up to six weeks. Storage for a period as short as 30 or 54 hr resulted in the appearance of large percertages of nortrichloroacetic acid-precipitable material with altered electrophoretic mobility. By use of butanol/acetic acid chromatography on specimens of monthold insulin, the altered material was shown to correspond neither with iodide, monoiodotyrosine, diiodotyrosine, nor with a ninhydrin-staining band of breakdown products. The trichloroacetic acid supernatant contained counts only precipitated very slowly by certrifugation at 3000 rpm for 1 hr, in contrast to the quick precipitation on centrifugation of insulin-bound /sup 131/I after trichloroacetic acid treatment. Presumably, therefore, these breakdown products represert medium-sized iodopeptides. Protein, either albumin or plasma protein, added to the insulin solution inhibited the breakdown but because of the intra-arterial studies to be performed, the addition of albumin or other foreign protein was avoided. Three samples, frozen-dried, were found to contain 91, 94, and 95% insulin at the end of 6 weeks. This suggested that insulin was being damaged by ionized water rather than direct irradiation damage. This was supported by the even more rapid disirtegration of the insulin on keeping, when the forearm uptake of insulin after intra-arterial injection in humans showed the erroneous results obtained when aged, degraded samples of insulin-/sup 131/I were used. (BBB)
- Research Organization:
- Guy's Hospital, London
- NSA Number:
- NSA-18-017436
- OSTI ID:
- 4051615
- Journal Information:
- Guy's Hosp. Rept., Journal Name: Guy's Hosp. Rept. Vol. Vol: 111
- Country of Publication:
- Country unknown/Code not available
- Language:
- English
Similar Records
PREPARATION AND PURIFICATION OF HIGH SPECIFIC ACTIVITY INSULIN-131IODINE
Retroendocytosis of insulin in rat adipocytes
Related Subjects
ALBUMINS
AMBERLITE
BIOCHEMISTRY
BIOLOGY AND MEDICINE
BLOOD CIRCULATION
BLOOD PLASMA
BLOOD VESSELS
BUTANOL
CENTRIFUGATION
CHLORIDES
CHLORINATED HYDROCARBONS
CHROMATOGRAPHY
DECOMPOSITION
DIFFUSION
DISTRIBUTION
ELECTROPHORESIS
ERRORS
EXTRACTION COLUMNS
HORMONES
INSULIN
IODIDES
IODINATED HYDROCARBONS
IODINE 131
ION EXCHANGE MATERIALS
IONIZATION
IRRADIATION
LABELLED COMPOUNDS
LOW TEMPERATURE
MAN
METABOLISM
MOLECULES
NUMERICALS
PAPER
PERFORMANCE
PRECIPITATION
PREPARATION
PRESERVATION
PROTEINS
QUALITATIVE ANALYSIS
RADIATION EFFECTS
RADIOACTIVITY
REACTION KINETICS
RESINS
SAMPLING
SOLUTIONS
STABILITY
STORAGE
SULFA