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{{Chembox |
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| verifiedrevid = 414452086 |
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| Reference=<ref> at ]</ref> |
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| Reference =<ref> at ]</ref> |
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| ImageFile = DTNB.png |
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| ImageFile_Ref = {{chemboximage|correct|??}} |
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| ImageSize = 200px |
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| ImageFile = DTNB.svg |
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| IUPACName = 5-(3-Carboxy-4-nitrophenyl)disulfanyl-2-nitrobenzoic acid |
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| ImageSize = |
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| OtherNames = Dithionitrobenzoic acid; 5,5'-Dithiobis(2-nitrobenzoic acid) |
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| PIN = 5,5′-Disulfanediylbis(2-nitrobenzoic acid) |
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| Section1 = {{Chembox Identifiers |
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| OtherNames = 3,3′-Disulfanediylbis(6-nitrobenzoic acid)<br />5-(3-Carboxy-4-nitrophenyl)disulfanyl-2-nitrobenzoic acid<br />Dithionitrobenzoic acid<br />5,5′-Dithiobis(2-nitrobenzoic acid) |
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|Section1={{Chembox Identifiers |
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| CASNo = 69-78-3 |
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| CASNo_Ref = {{cascite|correct|CAS}} |
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| Abbreviations = DTNB |
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| Abbreviations = DTNB |
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| ChemSpiderID_Ref = {{chemspidercite|correct|chemspider}} |
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| ChemSpiderID = 6018 |
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| ChemSpiderID = 6018 |
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| ChEBI_Ref = {{ebicite|correct|EBI}} |
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| ChEBI = 86228 |
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| ChEMBL_Ref = {{ebicite|correct|EBI}} |
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| ChEMBL = 395814 |
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| EC_number = 200-714-4 |
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| UNII_Ref = {{fdacite|correct|FDA}} |
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| UNII = 9BZQ3U62JX |
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| PubChem = 6254 |
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| InChI = 1/C14H8N2O8S2/c17-13(18)9-5-7(1-3-11(9)15(21)22)25-26-8-2-4-12(16(23)24)10(6-8)14(19)20/h1-6H,(H,17,18)(H,19,20) |
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| InChI = 1/C14H8N2O8S2/c17-13(18)9-5-7(1-3-11(9)15(21)22)25-26-8-2-4-12(16(23)24)10(6-8)14(19)20/h1-6H,(H,17,18)(H,19,20) |
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| InChIKey = KIUMMUBSPKGMOY-UHFFFAOYAD |
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| InChIKey = KIUMMUBSPKGMOY-UHFFFAOYAD |
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| StdInChI_Ref = {{stdinchicite|correct|chemspider}} |
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| ChEMBL = 395814 |
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| StdInChI = 1S/C14H8N2O8S2/c17-13(18)9-5-7(1-3-11(9)15(21)22)25-26-8-2-4-12(16(23)24)10(6-8)14(19)20/h1-6H,(H,17,18)(H,19,20) |
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| StdInChI = 1S/C14H8N2O8S2/c17-13(18)9-5-7(1-3-11(9)15(21)22)25-26-8-2-4-12(16(23)24)10(6-8)14(19)20/h1-6H,(H,17,18)(H,19,20) |
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| StdInChIKey_Ref = {{stdinchicite|correct|chemspider}} |
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| StdInChIKey = KIUMMUBSPKGMOY-UHFFFAOYSA-N |
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| StdInChIKey = KIUMMUBSPKGMOY-UHFFFAOYSA-N |
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| CASNo = 69-78-3 |
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| PubChem = 6254 |
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| SMILES = c1cc(c(cc1SSc2ccc(c(c2)C(=O)O)(=O))C(=O)O)(=O) |
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| SMILES = c1cc(c(cc1SSc2ccc(c(c2)C(=O)O)(=O))C(=O)O)(=O) |
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| Section2 = {{Chembox Properties |
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|Section2={{Chembox Properties |
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| C=14 | H=8 | N=2 | O=8 | S=2 |
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| C=14 | H=8 | N=2 | O=8 | S=2 |
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| Appearance = |
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| Appearance = |
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| Density = |
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| Density = |
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| MeltingPt = 240-245 °C (dec.) |
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| MeltingPtC = 240 to 245 |
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| MeltingPt_notes = (decomposes) |
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| Section3 = {{Chembox Hazards |
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|Section3={{Chembox Hazards |
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| MainHazards = |
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| FlashPt = |
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| FlashPt = |
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| Autoignition = |
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| GHS_ref=<ref>{{cite web |title=5,5'-Dithiobis(2-nitrobenzoic acid) |url=https://pubchem.ncbi.nlm.nih.gov/compound/6254#section=Safety-and-Hazards |website=pubchem.ncbi.nlm.nih.gov |access-date=13 December 2021 |language=en}}</ref> |
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| RPhrases = {{R36/37/38}} |
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| SPhrases = {{S26}} {{S36}} |
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| GHSPictograms = {{GHS07}} |
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| GHSSignalWord = Warning |
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| HPhrases = {{H-phrases|315|319|335}} |
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| PPhrases = {{P-phrases|261|264|271|280|302+352|304+340|305+351+338|312|321|332+313|337+313|362|403+233|405|501}} |
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'''Ellman's reagent''' (5,5'-dithiobis-(2-nitrobenzoic acid) or '''DTNB''') is a chemical used to quantify the number or concentration of ] groups in a sample.<ref name = ellman>{{cite journal |author=Ellman GL |title=Tissue sulfhydryl groups |journal=Arch. Biochem. Biophys. |volume=82 |issue=1 |pages=70–7 |year=1959 |pmid=13650640 |doi=10.1016/0003-9861(59)90090-6}}</ref> |
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'''Ellman's reagent''' (5,5′-dithiobis-(2-nitrobenzoic acid) or '''DTNB''') is a ] chemical used to quantify the number or concentration of ] groups in a sample.<ref name = ellman>{{cite journal |author=Ellman GL |title=Tissue sulfhydryl groups |journal=Arch. Biochem. Biophys. |volume=82 |issue=1 |pages=70–7 |year=1959 |pmid=13650640 |doi=10.1016/0003-9861(59)90090-6}}</ref> It was developed by George L. Ellman. |
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==Preparation== |
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==Preparation== |
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==Ellman's test== |
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==Ellman's test== |
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Thiols react with this compound, cleaving the ] to give 2-nitro-5-thiobenzoate (NTB<sup>-</sup>), which ionizes to the NTB<sup>2-</sup> dianion in water at neutral and alkaline pH. This NTB<sup>2-</sup> ion has a yellow color. |
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Thiols react with this compound, cleaving the ] to give 2-nitro-5-thiobenzoate (TNB<sup>−</sup>), which ionizes to the TNB<sup>2−</sup> dianion in water at neutral and alkaline pH. This TNB<sup>2−</sup> ion has a yellow color. |
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This reaction is rapid and stoichiometric, with the addition of one mole of thiol releasing one mole of NTB. The NTB<sup>2-</sup> is quantified in a ] by measuring the absorbance of visible light at 412 nm, using an ] of 14,150 ] cm<sup>−1</sup> for dilute buffer solutions,<ref>{{cite journal |author=Collier HB |title=Letter: A note on the molar absorptivity of reduced Ellman's reagent, 3-carboxylato-4-nitrothiophenolate |journal=Anal. Biochem. |volume=56 |issue=1 |pages=310–1 |year=1973 |pmid=4764694 |doi=10.1016/0003-2697(73)90196-6}}</ref><ref name=Riddles>{{cite journal |author=Riddles PW, Blakeley RL, Zerner B |title=Reassessment of Ellman's reagent |journal=Meth. Enzymol. |volume=91 |issue= |pages=49–60 |year=1983 |pmid=6855597 |doi=10.1016/S0076-6879(83)91010-8}}</ref> and a coefficient of 13,700 M<sup>−1</sup> cm<sup>−1</sup> for high salt concentrations, such as 6 M ] or 8 M urea.<ref name=Riddles/> Unfortunately the extinction coefficient for dilute solutions was underestimated in the original 1959 publication, as 13,600 M<sup>−1</sup> cm<sup>−1</sup>, and as noted in a recent article, this mistake has persisted in the literature.<ref name=Riener>{{cite journal |author=Riener CK, Kada G, Gruber HJ |title=Quick measurement of protein sulfhydryls with Ellman's reagent and with 4,4'-dithiodipyridine |journal=Anal Bioanal Chem |volume=373 |issue=4-5 |pages=266–76 |year=2002 |pmid=12110978 |doi=10.1007/s00216-002-1347-2}}</ref> Commercial DTNB may not be completely pure, so may require ] to obtain completely accurate and reproducible results.<ref name=Riddles/> |
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This reaction is rapid and stoichiometric, with the addition of one mole of thiol releasing one mole of TNB. The TNB<sup>2−</sup> is quantified in a ] by measuring the absorbance of visible light at 412 nm, using an ] of 14,150 ] cm<sup>−1</sup> for dilute buffer solutions,<ref>{{cite journal |author=Collier HB |title=Letter: A note on the molar absorptivity of reduced Ellman's reagent, 3-carboxylato-4-nitrothiophenolate |journal=Anal. Biochem. |volume=56 |issue=1 |pages=310–1 |year=1973 |pmid=4764694 |doi=10.1016/0003-2697(73)90196-6}}</ref><ref name=Riddles>{{cite book |vauthors=Riddles PW, Blakeley RL, Zerner B |title=Enzyme Structure Part I |chapter=Reassessment of Ellman's reagent |volume=91 |pages=49–60 |year=1983 |pmid=6855597 |doi=10.1016/S0076-6879(83)91010-8 |series=Methods in Enzymology |isbn=978-0-12-181991-0}}</ref> and a coefficient of 13,700 M<sup>−1</sup> cm<sup>−1</sup> for high salt concentrations, such as 6 M ] or 8 M urea.<ref name=Riddles/> Ellman's original 1959 publication estimated the molar extinction at 13,600 M<sup>−1</sup> cm<sup>−1</sup>, and this value can be found in some modern applications of the method despite improved determinations.<ref name=Riener>{{cite journal |vauthors=Riener CK, Kada G, Gruber HJ |title=Quick measurement of protein sulfhydryls with Ellman's reagent and with 4,4'-dithiodipyridine |journal=Anal Bioanal Chem |volume=373 |issue=4–5 |pages=266–76 |year=2002 |pmid=12110978 |doi=10.1007/s00216-002-1347-2|s2cid=30366479 }}</ref> Commercial DTNB may not be completely pure, so may require ] to obtain completely accurate and reproducible results.<ref name=Riddles/> |
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Ellman's reagent can be used for measuring low-molecular mass thiols such as ] in both pure solutions and biological samples, such as blood.<ref>{{cite journal |author=Sedlak J, Lindsay RH |title=Estimation of total, protein-bound, and nonprotein sulfhydryl groups in tissue with Ellman's reagent |journal=Anal. Biochem. |volume=25 |issue=1 |pages=192–205 |year=1968 |pmid=4973948 |doi=10.1016/0003-2697(68)90092-4}}</ref> It can also measure the number of thiol groups on proteins.<ref name=Riener/> |
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Ellman's reagent can be used for measuring low-molecular mass thiols such as ] in both pure solutions and biological samples, such as blood.<ref name=":0">{{Cite journal|last1=Riener|first1=Christian K.|last2=Kada|first2=Gerald|last3=Gruber|first3=Hermann J.|date=2002-07-01|title=Quick measurement of protein sulfhydryls with Ellman's reagent and with 4,4′-dithiodipyridine|journal=Analytical and Bioanalytical Chemistry|language=en|volume=373|issue=4–5|pages=266–276|doi=10.1007/s00216-002-1347-2|pmid=12110978|s2cid=30366479|issn=1618-2642}}</ref> It can also measure the number of thiol groups on proteins.<ref name=":0" /> |
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==References== |
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==References== |
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{{DEFAULTSORT:Ellman's Reagent}} |
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{{DEFAULTSORT:Ellman's Reagent}} |
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