Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid)
A method for assaying glutathione reductase (GSH; EC 1.6.4.2) in crude plant extracts is described. The method is based on the increase in absorbance at 412 nm when 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) is reduced by GSH. The effects of the following parameters on the assay were tested: various...
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| Veröffentlicht in: | Analytical biochemistry 1988-12, Vol.175 (2), p.408-413 |
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| container_title | Analytical biochemistry |
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| creator | Smith, Ivan K. Vierheller, Thomas L. Thorne, Carol A. |
| description | A method for assaying glutathione reductase (GSH; EC 1.6.4.2) in crude plant extracts is described. The method is based on the increase in absorbance at 412 nm when 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) is reduced by GSH. The effects of the following parameters on the assay were tested: various buffers, pH, buffer concentration, compounds commonly present in enzyme preparations, thiols, and the presence of another NADPH-dependent enzyme. The assay is more sensitive and less subject to interference than the widely used assay where NADPH oxidation is monitored. In particular, the specificity of DTNB allows assay of glutathione reductase in the presence of other NADPH-dependent enzymes and common protein extract contaminants. |
| doi_str_mv | 10.1016/0003-2697(88)90564-7 |
| format | Article |
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The method is based on the increase in absorbance at 412 nm when 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) is reduced by GSH. The effects of the following parameters on the assay were tested: various buffers, pH, buffer concentration, compounds commonly present in enzyme preparations, thiols, and the presence of another NADPH-dependent enzyme. The assay is more sensitive and less subject to interference than the widely used assay where NADPH oxidation is monitored. In particular, the specificity of DTNB allows assay of glutathione reductase in the presence of other NADPH-dependent enzymes and common protein extract contaminants.</description><identifier>ISSN: 0003-2697</identifier><identifier>EISSN: 1096-0309</identifier><identifier>DOI: 10.1016/0003-2697(88)90564-7</identifier><identifier>PMID: 3239770</identifier><identifier>CODEN: ANBCA2</identifier><language>eng</language><publisher>San Diego, CA: Elsevier Inc</publisher><subject>5,5'-dithiobis(2-nitrobenzoic acid) ; Analytical, structural and metabolic biochemistry ; Biological and medical sciences ; Dithionitrobenzoic Acid ; Enzymes and enzyme inhibitors ; Fundamental and applied biological sciences. Psychology ; glutathione ; glutathione reductase ; Glutathione Reductase - analysis ; Glycine max ; Kinetics ; Oxidoreductases ; plant systems ; Plants - enzymology ; spectrophotometry ; Spectrophotometry - methods</subject><ispartof>Analytical biochemistry, 1988-12, Vol.175 (2), p.408-413</ispartof><rights>1988</rights><rights>1989 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c483t-2abd13a71d05716cd804e3567068ede6dd858a46ac0b0f61c75b5e06f34c6d933</citedby><cites>FETCH-LOGICAL-c483t-2abd13a71d05716cd804e3567068ede6dd858a46ac0b0f61c75b5e06f34c6d933</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/0003-2697(88)90564-7$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=7188098$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/3239770$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Smith, Ivan K.</creatorcontrib><creatorcontrib>Vierheller, Thomas L.</creatorcontrib><creatorcontrib>Thorne, Carol A.</creatorcontrib><title>Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid)</title><title>Analytical biochemistry</title><addtitle>Anal Biochem</addtitle><description>A method for assaying glutathione reductase (GSH; EC 1.6.4.2) in crude plant extracts is described. The method is based on the increase in absorbance at 412 nm when 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) is reduced by GSH. The effects of the following parameters on the assay were tested: various buffers, pH, buffer concentration, compounds commonly present in enzyme preparations, thiols, and the presence of another NADPH-dependent enzyme. The assay is more sensitive and less subject to interference than the widely used assay where NADPH oxidation is monitored. In particular, the specificity of DTNB allows assay of glutathione reductase in the presence of other NADPH-dependent enzymes and common protein extract contaminants.</description><subject>5,5'-dithiobis(2-nitrobenzoic acid)</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Biological and medical sciences</subject><subject>Dithionitrobenzoic Acid</subject><subject>Enzymes and enzyme inhibitors</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>glutathione</subject><subject>glutathione reductase</subject><subject>Glutathione Reductase - analysis</subject><subject>Glycine max</subject><subject>Kinetics</subject><subject>Oxidoreductases</subject><subject>plant systems</subject><subject>Plants - enzymology</subject><subject>spectrophotometry</subject><subject>Spectrophotometry - methods</subject><issn>0003-2697</issn><issn>1096-0309</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1988</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU1uFDEQhS1EFCaBG4DkBUKJlIZyu_3TG6QoCj9SJDawttx29cSopx1c3UhhxZk4EidJDzOaJVnVor73qvQeYy8FvBUg9DsAkFWtW3Nm7XkLSjeVecJWAlpdgYT2KVsdkGfshOg7gBCN0sfsWNayNQZWrL8k8vc893w9zJOfblMekReMc5g8IU8jD2WOyKdENCO_zZu8xtFPSHymNK65ulB_f_-pYtpqu0RndTWmqeQOx185Be5DiufP2VHvB8IX-3nKvn24_nr1qbr58vHz1eVNFRorp6r2XRTSGxFBGaFDtNCgVNqAthhRx2iV9Y32ATrotQhGdQpB97IJOrZSnrI3O9-7kn_MSJPbJAo4DH7EPJMz1tRS6sdBoaRebqkFbHZgKJmoYO_uStr4cu8EuG0Pbhuy24bsrHX_enBmkb3a-8_dBuNBtA9-2b_e7z0FP_TFjyHRATPCWmjtgr3fYbiE9jNhcRQSjgFjKhgmF3P6_x8P_Lik1A</recordid><startdate>19881201</startdate><enddate>19881201</enddate><creator>Smith, Ivan K.</creator><creator>Vierheller, Thomas L.</creator><creator>Thorne, Carol A.</creator><general>Elsevier Inc</general><general>Elsevier</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>M81</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>19881201</creationdate><title>Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid)</title><author>Smith, Ivan K. ; Vierheller, Thomas L. ; Thorne, Carol A.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c483t-2abd13a71d05716cd804e3567068ede6dd858a46ac0b0f61c75b5e06f34c6d933</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1988</creationdate><topic>5,5'-dithiobis(2-nitrobenzoic acid)</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Biological and medical sciences</topic><topic>Dithionitrobenzoic Acid</topic><topic>Enzymes and enzyme inhibitors</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>glutathione</topic><topic>glutathione reductase</topic><topic>Glutathione Reductase - analysis</topic><topic>Glycine max</topic><topic>Kinetics</topic><topic>Oxidoreductases</topic><topic>plant systems</topic><topic>Plants - enzymology</topic><topic>spectrophotometry</topic><topic>Spectrophotometry - methods</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Smith, Ivan K.</creatorcontrib><creatorcontrib>Vierheller, Thomas L.</creatorcontrib><creatorcontrib>Thorne, Carol A.</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>Biochemistry Abstracts 3</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Analytical biochemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Smith, Ivan K.</au><au>Vierheller, Thomas L.</au><au>Thorne, Carol A.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid)</atitle><jtitle>Analytical biochemistry</jtitle><addtitle>Anal Biochem</addtitle><date>1988-12-01</date><risdate>1988</risdate><volume>175</volume><issue>2</issue><spage>408</spage><epage>413</epage><pages>408-413</pages><issn>0003-2697</issn><eissn>1096-0309</eissn><coden>ANBCA2</coden><abstract>A method for assaying glutathione reductase (GSH; EC 1.6.4.2) in crude plant extracts is described. The method is based on the increase in absorbance at 412 nm when 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) is reduced by GSH. The effects of the following parameters on the assay were tested: various buffers, pH, buffer concentration, compounds commonly present in enzyme preparations, thiols, and the presence of another NADPH-dependent enzyme. The assay is more sensitive and less subject to interference than the widely used assay where NADPH oxidation is monitored. In particular, the specificity of DTNB allows assay of glutathione reductase in the presence of other NADPH-dependent enzymes and common protein extract contaminants.</abstract><cop>San Diego, CA</cop><pub>Elsevier Inc</pub><pmid>3239770</pmid><doi>10.1016/0003-2697(88)90564-7</doi><tpages>6</tpages></addata></record> |
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| ispartof | Analytical biochemistry, 1988-12, Vol.175 (2), p.408-413 |
| issn | 0003-2697 1096-0309 |
| language | eng |
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| subjects | 5,5'-dithiobis(2-nitrobenzoic acid) Analytical, structural and metabolic biochemistry Biological and medical sciences Dithionitrobenzoic Acid Enzymes and enzyme inhibitors Fundamental and applied biological sciences. Psychology glutathione glutathione reductase Glutathione Reductase - analysis Glycine max Kinetics Oxidoreductases plant systems Plants - enzymology spectrophotometry Spectrophotometry - methods |
| title | Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid) |
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