Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1969 Aug;113(5):879–884. doi: 10.1042/bj1130879

Studies on protein–polysaccharides from pig laryngeal cartilage. Extraction and purification

C P Tsiganos 1, Helen Muir 1
PMCID: PMC1184779  PMID: 4241780

Abstract

1. Protein–polysaccharides of chondroitin sulphate were extracted from fresh laryngeal cartilage at pH6·8 by two procedures. Procedure I consisted of brief low-speed homogenization in 0·15m (iso-osmotic) sodium acetate and procedure II consisted of longer homogenization followed by prolonged extraction in 10% calcium chloride solution. 2. The protein–polysaccharides in both extracts were isolated and purified by precipitation with 9-aminoacridine hydrochloride. They were free from serum proteins, collagen and nucleic acids and also of degradative enzymes. The absence of such enzymes was shown by viscosity measurements on solutions of protein–polysaccharides incubated for up to 24hr. at pH4 and 6·8. 3. Mannose, glucose or fucose were not detected by paper chromatography and only traces of sialic acid were present. 4. The yield with procedure II was twice that with procedure I and the products differed in their protein and glucosamine contents. 5. Hyaluronic acid was unlikely to have been precipitated at an acid pH, so the glucosamine was attributed to keratan sulphate, as serum proteins were absent. There was no free keratan sulphate in the preparation. 6. Both preparations were heterogeneous in the ultracentrifuge, showing at least three components.

Full text

PDF
879

Images in this article

883Fig. 1.
on p.883

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. AMINOFF D. Methods for the quantitative estimation of N-acetylneuraminic acid and their application to hydrolysates of sialomucoids. Biochem J. 1961 Nov;81:384–392. doi: 10.1042/bj0810384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. ANTONOPOULOS C. A., BORELIUS E., GARDELL S., HAMNSTROM B., SCOTT J. E. The precipitation of polyanions by long-chain aliphatic ammonium compounds. IV. Elution in salt solutions of mucopolysaccharide-quaternary ammonium complexes adsorbed on a support. Biochim Biophys Acta. 1961 Dec 9;54:213–226. doi: 10.1016/0006-3002(61)90360-2. [DOI] [PubMed] [Google Scholar]
  3. BITTER T., MUIR H. M. A modified uronic acid carbazole reaction. Anal Biochem. 1962 Oct;4:330–334. doi: 10.1016/0003-2697(62)90095-7. [DOI] [PubMed] [Google Scholar]
  4. Bitter T., Muir H. Mucopolysaccharides of whole human spleens in generalized amyloidosis. J Clin Invest. 1966 Jun;45(6):963–975. doi: 10.1172/JCI105412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. CESSI C., PILIEGO F. The determination of amino sugars in the presence of amino acids and glucose. Biochem J. 1960 Dec;77:508–510. doi: 10.1042/bj0770508. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. CRUMPTON M. J. Identification of amino sugars. Biochem J. 1959 Jul;72:479–486. doi: 10.1042/bj0720479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. DINGLE J. T., LUCY J. A., FELL H. B. Studies on the mode of action of excess of vitamin A. 1. Effect of excess of vitamin A on the metabolism and composition of embryonic chick-limb cartilage grown in organ culture. Biochem J. 1961 Jun;79:497–500. doi: 10.1042/bj0790497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dunstone J. R., Franek M. D. Isolation of a glycoprotein from cartilage proteinpolysaccharide. Biochem Biophys Res Commun. 1967 Apr 7;27(1):39–45. doi: 10.1016/s0006-291x(67)80036-6. [DOI] [PubMed] [Google Scholar]
  9. Elson L. A., Morgan W. T. A colorimetric method for the determination of glucosamine and chondrosamine. Biochem J. 1933;27(6):1824–1828. doi: 10.1042/bj0271824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. GERBER B. R., FRANKLIN E. C., SCHUBERT M. Ultracentrifugal fractionation of bovine nasal chondromucoprotein. J Biol Chem. 1960 Oct;235:2870–2875. [PubMed] [Google Scholar]
  11. GREGORY J. D., LAURENT T. C., RODEN L. ENZYMATIC DEGRADATION OF CHONDROMUCOPROTEIN. J Biol Chem. 1964 Oct;239:3312–3320. [PubMed] [Google Scholar]
  12. HOFFMAN P., LINKER A., MEYER K. The acid mucopolysaccharides of connective tissue. III. the sulfate linkage. Biochim Biophys Acta. 1958 Oct;30(1):184–185. doi: 10.1016/0006-3002(58)90256-7. [DOI] [PubMed] [Google Scholar]
  13. HOLDEN M., PIRIE N. W. The preparation of ribonucleic acid from yeast, tobacco leaves and tobacco mosaic virus. Biochem J. 1955 May;60(1):46–53. doi: 10.1042/bj0600046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Heinegård D., Gardell S. Studies on protein-polysaccharide complex (proteoglycan) from human nucleus pulposus. I. Isolation and preliminary characterisation. Biochim Biophys Acta. 1967 Oct 9;148(1):164–171. doi: 10.1016/0304-4165(67)90292-9. [DOI] [PubMed] [Google Scholar]
  15. JEANLOZ R. W. The nomenclature of mucopolysaccharides. Arthritis Rheum. 1960 Jun;3:233–237. doi: 10.1002/art.1780030306. [DOI] [PubMed] [Google Scholar]
  16. LINDAHL U., RODEN L. THE ROLE OF GALACTOSE AND XYLOSE IN THE LINKAGE OF HEPARIN TO PROTEIN. J Biol Chem. 1965 Jul;240:2821–2826. [PubMed] [Google Scholar]
  17. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  18. Lindahl U., Rodén L. The chondroitin 4-sulfate-protein linkage. J Biol Chem. 1966 May 10;241(9):2113–2119. [PubMed] [Google Scholar]
  19. Luscombe M., Phelps C. F. The composition and physicochemical properties of bovine nasal-septa protein-polysaccharide complex. Biochem J. 1967 Jan;102(1):110–119. doi: 10.1042/bj1020110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. MALAWISTA I., SCHUBERT M. Chondromucoprotein: new extraction method and alkaline degradation. J Biol Chem. 1958 Jan;230(1):535–544. [PubMed] [Google Scholar]
  21. MATHEWS M. B. Isomeric chondroitin sulphates. Nature. 1958 Feb 8;181(4606):421–422. doi: 10.1038/181421a0. [DOI] [PubMed] [Google Scholar]
  22. MATHEWS M. B., LOZAITYTE I. Sodium chondroitin sulfate-protein complexes of cartilage. I. Molecular weight and shape. Arch Biochem Biophys. 1958 Mar;74(1):158–174. doi: 10.1016/0003-9861(58)90210-8. [DOI] [PubMed] [Google Scholar]
  23. MATHEWS M. B. The molecular weight of sodium chondroitin sulfate by light scattering. Arch Biochem Biophys. 1956 Apr;61(2):367–377. doi: 10.1016/0003-9861(56)90359-9. [DOI] [PubMed] [Google Scholar]
  24. MUIR H. The nature of the link between protein and carbohydrate of a chondroitin sulphate complex from hyaline cartilage. Biochem J. 1958 Jun;69(2):195–204. doi: 10.1042/bj0690195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Mathews M. B., Glagov S. Acid mucopolysaccharide patterns in aging human cartilage. J Clin Invest. 1966 Jul;45(7):1103–1111. doi: 10.1172/JCI105416. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Muir H., Jacobs S. Protein-polysaccharides of pig laryngeal cartilage. Biochem J. 1967 May;103(2):367–374. doi: 10.1042/bj1030367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. PARTRIDGE S. M., DAVIS H. F., ADAIR G. S. The chemistry of connective tissues. 6. The constitution of the chondroitin sulphate-protein complex in cartilage. Biochem J. 1961 Apr;79:15–26. doi: 10.1042/bj0790015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rodén L., Armand G. Structure of the chondroitin 4-sulfate-protein linkage region. Isolation and characterization of the disaccharide 3-O-beta-D-glucuronosyl-D-galactose. J Biol Chem. 1966 Jan 10;241(1):65–70. [PubMed] [Google Scholar]
  29. Rosenberg L., Johnson B., Schubert M. Proteinpolysaccharides from human articular and costal cartilage. J Clin Invest. 1965 Oct;44(10):1647–1656. doi: 10.1172/JCI105271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rosenberg L., Schubert M. The proteinpolysaccharides of bovine nucleus pulposus. J Biol Chem. 1967 Oct 25;242(20):4691–4701. [PubMed] [Google Scholar]
  31. Serafini-Fracassini A., Peters T. J., Floreani L. The protein-polysaccharide complex of bovine nasal cartilage. Studies on the protein core. Biochem J. 1967 Nov;105(2):569–575. doi: 10.1042/bj1050569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Tsiganos C. P., Muir H. Studies on protein-polysaccharides from pig laryngeal cartilage. Heterogeneity, fractionation and characterization. Biochem J. 1969 Aug;113(5):885–894. doi: 10.1042/bj1130885. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES