Glutathione peroxidase activity in selenium-deficient rat liver☆

Publisher Summary This chapter presents a procedure for the preparation of glutathione peroxidase, which is regarded as a major protective system against endogenously and exogenously induced lipid peroxidation. Two types of methods are used for determining the activity of glutathione peroxidase. One involves a direct measurement of unconsumed glutathione (GSH) at fixed time periods by polarographic GSH analysis' (Method 1), or by the dithionitrobenzoic acid method (Method 2). The second approach takes advantage of the capability of glutathione reductase, with nicotinamide adenine dinucleotide phosphate (NADPH), to regenerate GSH from oxidized GSH. The decrease in NADPH is continuously measured spectrophotometrically, while the GSH concentration in the enzymatic cycle remains essentially constant (Method 3). A convenient source for the preparation of glutathione peroxidase is bovine blood including the following steps: hemolysate; organic solvent precipitation; phosphate precipitation; absorption to phenyl-sepharose; and washing on diethylaminoethyl (DEAE)–sephadex, S-300 sephacryl, and hydroxylapatite column.

[44] Glutathione peroxidase

Bearing the public image of a deadly "gas of rotten eggs," hydrogen sulfide (H2S) can be generated in many types of mammalian cells. Functionally, H2S has been implicated in the induction of hippocampal long-term potentiation, brain development, and blood pressure regulation. By acting specifically on KATP channels, H2S can hyperpolarize cell membranes, relax smooth muscle cells, or decrease neuronal excitability. The endogenous metabolism and physiological functions of H2S position this gas well in the novel family of endogenous gaseous transmitters, termed "gasotransmitters." It is hypothesized that H2S is the third endogenous signaling gasotransmitter, besides nitric oxide and carbon monoxide. This positioning of H2S will open an exciting field-H2S physiology-encompassing realization of the interaction of H2S and other gasotransmitters, sulfurating modification of proteins, and the functional role of H2S in multiple systems. It may shed light on the pathogenesis of many diseases related to the abnormal metabolism of H2S.

Two’s company, three’s a crowd: can H2S be the third endogenous gaseous transmitter?

Publisher Summary Glutathione (GSH) is the most important nonprotein thiol in living systems and is of widespread occurrence in the intracellular milieu of animals, plants, and microorganisms GSH was isolated and named by the English biochemist Frederick Gowland Hopkins This chapter discusses GSH status, the biologically relevant chemistry of GSH, the forms in which GSH can be present within the cell, along with the GSH content of cells and the methods for analysis of this substance GSH-related biochemical reactions and the biological roles of GSH are discussed in the chapter The use of perturbations in GSH status as a means for investigating GSH-related phenomena and an analysis of the consequences of perturbation are presented A short summary of genetic lesions related to GSH is also included Like chemically induced perturbations in GSH status, genetic lesions provide valuable insights into the role of GSH in normal functions and processes in cells The chapter concludes with some brief comments about the future of the relationship of GSH status to cellular processes

The Glutathione Status of Cells

A good number of abstracts and research articles (in total 74) published, so far, for evaluating antioxidant activity of various samples of research interest were gone through where 407 methods were come across, which were repeated from 29 different methods. These were classified as in vitro and in vivo methods. And those are described and discussed below in this review article. In the later part of this review article, frequency of in vitro as well as in vivo methods is analyzed with a bar diagram. Solvents are important for extracting antioxidants from natural sources. Frequency of solvents used for extraction is also portrayed and the results are discussed in this article. As per this review there are 19 in vitro methods and 10 in vivo methods that are being used for the evaluation of antioxidant activity of the sample of interest. DPPH method was found to be used mostly for the in vitro antioxidant activity evaluation purpose while LPO was found as mostly used in vivo antioxidant assay. Ethanol was with the highest frequency as solvent for extraction purpose.

Review on in vivo and in vitro methods evaluation of antioxidant activity.

Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase

A severe deficiency of a red cell pyrimidine 5'-nucleotidase was found to be associated with hereditary hemolytic anemia in four members of three kindreds. The syndrome was characterized by marked increases above normal in red cell basophilic stippling, total nucleotides, and GSH and by a fairly severe deficiency of ribosephosphate pyrophosphokinase (EC 2.7.6.1.). Patient erythrocytes uniquely contained large amounts of pyrimidine 5'-ribonucleotides. In earlier studies, these were erroneously considered to be adenosine phosphates, since all previous investigations of the nucleotides of human red cells and reticulocytes have shown 97% or more to contain adenine. Total nucleotides in patient cells were present in amounts 3-6 times greater than normal, and approximately 80% contained pyrimidine. The ultraviolet spectral curves of deproteinized red cell extracts exhibited a shift in maximum absorbance from the usual 256-257 nm to approximately 266-270 nm, and absorbance at 250, 270, 280, and 290 nm, expressed as a ratio of that at 260 nm, differed greatly from normal. The spectral characteristics of extracts provide the basis of a readily performed screening procedure, which does not require enzyme assay. The nucleotidase activity in deficient red cells assayed less than 14%, and usually less than 10%, of normal and much less in terms of reticulocyte-rich blood, where it was consistently found to be increased. The enzyme has a pH optimum of 7.5-8.0, is inhibited by EDTA, and does not utilize purine 5'-ribonucleotides or beta-glycerophosphate as substrates. While comparatively few family members have been available thus far for study, initial data are compatible with an autosomal, recessive mode of transmission of the deficiency. The pyrimidine 5'-ribonucleotides are presumably derived from RNA degradation and, not being diffusible, accumulate when the enzyme catalyzing their dephosphorylation is deficient. It is postulated that the prominent basophilic stippling results from retarded ribosomal RNA degradation secondary to accumulation of degradation products, namely pyrimidine 5'-ribonucleotides. Ribosephosphate pyrophosphokinase deficiency is considered to be an epiphenomenon. The mechanism responsible for increased red cell GSH is unknown.

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Hereditary Hemolytic Anemia with Human Erythrocyte Pyrimidine 5′-Nucleotidase Deficiency

Lead intoxication is accompanied by an acquired deficiency of erythrocyte pryimidine-specific, 5'-nucleotidase. Genetically determined deficiency of this enzyme is associated with chronic hemolysis, marked basophilic stippling of erythrocytes on stained blood films, and unique intraerythrocytic accumulations of pyrimidine-containing nucleotides. The present report documents that lead-induced deficiency when sufficiently severe gives rise to findings similar to the hereditary disorder. Whereas pyrimidine-containing nucleotides are virutally absent in the erythrocytes of normal and reticulocyte-rich blood, 12% of erythrocyte nucloetides in the blood of a patient with lead intoxication contained cytidine. Nucleotidase activity was about 25% that in normal erythrocytes and 15% or less of that expected in comparable reticulocyte-rich blood. The distribution of nucleotidase activity in patient erythrocytes is unknown, and much more severe deficiency could have been present in subsets of the cell populations analyzed. The findings indicate that the hemolytic anemia and increased basophilic stippling characteristic of certain cases of lead intoxication may share a common etiology with essentially identical features of the genetically determined disorder.

https://dm5migu4zj3pb.cloudfront.net/manuscripts/108000/108545/JCI76108545.pdf

Lead poisoning: association with hemolytic anemia, basophilic stippling, erythrocyte pyrimidine 5'-nucleotidase deficiency, and intraerythrocytic accumulation of pyrimidines.

/pdf/studies-on-chromated-erythrocytes-effect-of-sodium-chromate-33jwybuwlg.pdf

Studies on Chromated Erythrocytes. Effect of Sodium Chromate on Erythrocyte Glutathione Reductase

Cooley's anemia, 1 or thalassemia, 2 is a rare but well known disease. This fatal disorder is characterized by a chronic, progressive, hypochromic and microcytic type of anemia, peripheral erythroblastosis, increased number of target and oval red blood cells, decreased fragility of erythrocytes, splenomegaly, deposition of pigment in the viscera and changes in the bones. The condition was early noted to have a familial incidence and to be virtually restricted to the Mediterranean peoples. In 1937, Angelini (quoted from Wintrobe and associates 3 ) observed that in some instances the erythrocytes of apparently healthy parents and of siblings of patients suffering from this anemia showed decreased fragility. Caminopetros 4 independently confirmed this observation, finding that 22 of 30 parents tested showed reduced fragility. He also reported slight changes in the bones in some parents. A few years later, Wintrobe, Matthews, Pollack and Dobyns 3 described the occurrence in several Italian families

William N. Valentine

Papers

Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase

Hereditary Hemolytic Anemia with Human Erythrocyte Pyrimidine 5′-Nucleotidase Deficiency

Lead poisoning: association with hemolytic anemia, basophilic stippling, erythrocyte pyrimidine 5'-nucleotidase deficiency, and intraerythrocytic accumulation of pyrimidines.

Studies on Chromated Erythrocytes. Effect of Sodium Chromate on Erythrocyte Glutathione Reductase

Hematologic and genetic study of the transmission of thalassemia: (cooley's anemia; mediterranean anemia)