Quantitative Determination of Polypeptides by Gradient Elution High Pressure Liquid Chromatography

Biemond, M. E. F.; Sipman, W.A.; Olivié, J

doi:10.1080/01483917908060146

Cited by 40 publications

(2 citation statements)

References 18 publications

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“…These include ion exchange chromatography in buffers containing urea and preparative isoelectric focusing, both already discussed. Partition chromatography (HRUBY and GROGINSKY 1971) and reverse-phase high pressure liquid chromatography (BIEMOND et al 1979;HANCOCK et al 1978;BURGUS and RIVIER 1976) are two other powerful techniques that have been applied successfully to glucagon.…”

Section: B Isolation and Purificationmentioning

confidence: 99%

Chemical Characteristics of Glucagon

Bromer¹

1983

Glucagon I

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The object of this chapter is to describe the covalent chemistry of pancreatic glucagon to provide a background for subsequent chapters. Modifications of the primary structure are considered with respect to biologic activity, but not immunologic activity which is discussed in Chap. 9. Conformation and function are considered in Chap. 3. B. Isolation and PurificationGlucagon is effectively extracted from animal pancreas by any of the acidic ethanol methods commonly employed to extract insulin (e.g., SUTHERLAND and DEDuVE 1948;KENNY 1955). Such methods have been successfully used with mammalian, avian, and piscine pancreas (for details see Chap. 8). The glucagon content of freshly collected bovine or porcine pancreas is about 5-10 J.tg/g wet tissue. RHOTEN (1976) found as much as 5 mg/g glucagon in splenic pancreas from lizards. In a typical insulin purification scheme, glucagon and insulin are purified together to the point of insulin crystallization, where most of the glucagon is found in the mother liquor (SUNDBY and MARKUSSEN 1971). Even at this stage the glucagon fraction is less than 2% pure. The first successful purification of glucagon by STAUB et al. (1955) employed repeated precipitations culminating in crystallization, which was in itself an excellent purification step. In more recent studies, purification was often achieved by separations based more directly on molecular size and on charge. For example, in the purification of chicken glucagon, POLLOCK and KIMMEL (1975) used ion exchange chromatography on columns of CM-Sephadex and QAE-Sephadex. A particulary elegant procedure applicable to small-scale purification was developed by SUNDBY and MARKUSSEN (1971) for rat glucagon and was applied subsequently to glucagon from several other species: (a) the mother liquor fraction from the citrate crystallization of insulin was brought to about half-saturation with NaCl; (b) the resulting precipitate was dissolved in acidic urea solution and gel-filtered in dilute acetic acid; ( c) glucagon fractions (4 % pure) were lyophilized and fractionated by preparative isoelectric focusing in polyacrylamide gels; (d) the glucagon fraction was eluted and crystallized. If desirable, the isoelectric focusing step may most likely be replaced by ion exchange chromatography using both anion and cation exchange media.The first glucagon crystals contained about 10% of a minor acidic component which was separable by electrophoresis (STAUB et al. 1955); reelectrophoresis of the P. J. Lefèbvre (ed.), Glucagon I

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Section: B Isolation and Purificationmentioning

confidence: 99%

Chemical Characteristics of Glucagon

Bromer¹

1983

Glucagon I

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“…The solvent was prepared according to the method of Biemond et al [6]: 0.025 mol/l tétraméthylammo nium hydroxide was added to a methanol solution and the pH of this mixture was adjusted with phosphoric acid to 3.0. The ratio of water to methanol mixture was 20:80 for solvent A and 50:50 for solvent B.…”

Section: Isolation O F Carbamylated Insulinmentioning

confidence: 99%

Carbamylation of Insulin and Its Biological Activity

Oimomi

Hatanaka

Yoshimura

et al. 1987

Nephron

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A nonspecific binding reaction between cyanic acid formed from urea and protein or peptide is called carbamylation. In the present study, insulin, a peptide hormone, was subjected to carbamylation and the activity of carbamylated insulin was determined. Both immunological and biological activities of insulin changed on carbamylation. The decrease in biological activity in respect to glucose oxidation of fat cells or receptor-binding capacity of rat hepatocytes was greater than that in immunological activity.

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Effect of Mobile Phase Composition on Selectivity in Preparative Hydrophobic Interaction Chromatographic Purification of Glucagon

Angelova

Dimov

1996

Biomed. Chromatogr.

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Quantitative Determination of Polypeptides by Gradient Elution High Pressure Liquid Chromatography

Cited by 40 publications

References 18 publications

Chemical Characteristics of Glucagon

Chemical Characteristics of Glucagon

Carbamylation of Insulin and Its Biological Activity

Effect of Mobile Phase Composition on Selectivity in Preparative Hydrophobic Interaction Chromatographic Purification of Glucagon

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