Abstract
Activities of the anti-oxidative enzymes, superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase were studied in rat tissues to determine the ability of detergents both to solubilize the enzymes and also to stabilize enzyme activity. Rat brain, heart and liver were homogenized in 0.1M KCl, 0.1% sodium dodecyl sulfate, 0.1% lubrol, or 0.1% cetyl-trimethylammonium bromide. In general lubrol was more effective than the other solutions in solubilizing GPx and catalase. Lubrol and 0.1M KCl were equally effective in solubilizing SOD. The highest enzyme activities were (1) SOD: 2484 ng/mg (brain), 2501 ng/mg (heart), and 5586 ng/mg (liver); (2) GPx: 224 mU/mg (brain), 1870 mU/mg (heart), and 7332 mU/mg (liver); (3) catalase: 2.8 mU/mg (brain), 10.6 mU/mg (heart), and 309 mU/mg (liver). While cetyl trimethylammonium bromide is marginally better than sodium dodecyl sulfate in solubilizing active enzyme, neither ionic detergent has any advantage over lubrol or 0.1M KCl. For catalase and GPx, enzyme activity loss with time is biphasic. After initial, rapid activity loss (1–5 days for GPx and 7–10 days for catalase) the differences noted among the homogenizing solutions disappear and very little if any activity loss is noted over the next 2–3 weeks. For catalase and GPx, only baseline enzyme activity from t = 0 – 3 weeks is found in the most chaotropic solution, 0.1% sodium dodecyl sulfate while biphasic activity loss is most pronounced in 0.1% lubrol. These results may indicate active GPx and catalase species stabilized by a lipid-like environment. Correlatingin vitro catalase or GPx measurements within vivo anti-oxidative protection may underestimate tissue defences.
Similar content being viewed by others
References
Kloc M, Mailer K, Stepkowski S: Superoxide dismutase decrease in cardiac transplants. Transplantation 41: 794–796, 1986
Clara RW, Knowles R: Superoxide dismutase, catalase, and peroxidase in ammonium-grown and nitrogen fixing Azospirillium brasilense. Can J Microbiol 30: 1222–1228, 1984
Del Maestro R, McDonald W, Anderson R: Superoxide dismutase, catalase, and glutathione peroxidase in experimental and human brain tumors. In RA Greenwald and G Cohen (eds) Oxy Radicals and Their Scavenger Systems. VII. Elsevier Biomedical, New York, 1983, p 28–35
Hazelton GA, Lang CA: Glutathione peroxidase and reductase activities in the aging mouse. Mech Aging and Dev 29: 71–81, 1985
Paynter DI, Caple IW: Age-related changes in activities of the superoxide dismutase enzymes in tissues of the sheep and the effect of dietary copper and manganese on these changes. J Nutr 114: 1909–1916, 1984
Currie RW, Karmazyn M, Kloc M, Mailer K: Heat shock response is associated with enhanced post-ischemic recovery. Circ Res 63: 543–549, 1988
Murakoshi M, Osamura Y, Yoshimura S, Watanabe K: Correlation between intramitochondrial localization of glutathione peroxidase and steroidogenesis in the rat adrenocortical cells. Acta Histochem Cytochem 17: 491–498, 1984
Capel ID, Smallwood AE: Sex differences in the glutathione peroxidase activity of various tissues of the rat. Res Commun Chem Pathol Pharmacol 40: 367–378, 1983
Julicher KHM, Sterrenberg L, Haenen G, Bast A, Noordhoek J: Sex differences in the cellular defence system against free radicals from oxygen or drug metabolites in rats. Arch Toxicol 56: 83–86, 1984
Ono T, Okada S: Unique increase of superoxide dismutase level in brains of long living mammals. Expt Geront 19: 349–354, 1984
Brawn K, Fridovich I: Superoxide radical and superoxide dismutases: threat and defense. Acta Physiol Scand Suppl 492: 9–19, 1980
Marklund S, Marklund G: Involvement of the superoxide anion radical in the autooxidation of pyrogallol and a convenient assay for superoxide dismutase. Eur J Biochem 47: 469–474,1974
Paglia DE, Valentine WM: Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J Lab Clin Med 70: 158–169, 1967
Del Rio LA, Ortega MG, Lopez AL, Gorge JL: A more sensitive modification of the catalase assay with the Clark Oxygen Electrode. Anal Biochem 80: 409–415, 1977
Harboe M: A method for determination of Hemoglobin in plasma by near-ultraviolet spectrophotometry. Scand J Clin Lab Invest 11: 66–70, 1959
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–275, 1951
Stein TN, Keen CL, Lonnerdal B, Hurley LS: Effect of sample preparation on analysis of superoxide dismutase activity and isozymes. J Inorg Biochem 16: 71–77, 1982
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Mailer, K., Del Maestro, R.F. Stability of the anti-oxidative enzymes in aqueous and detergent solution. Mol Cell Biochem 107, 47–54 (1991). https://doi.org/10.1007/BF02424575
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02424575