Monitoring of homocysteine in body fluids by means of MEKC and
on-column reaction with 2,2-dipyridyldisulfide

D 2002

Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide

ŠEVČÍKOVÁ, Petra a Zdeněk GLATZ

Základní údaje

Originální název

Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide

Autoři

ŠEVČÍKOVÁ, Petra a Zdeněk GLATZ

Vydání

Helsinki, Proceeding of the 13th International Symposium on Capillary Electroseparation Techniques ITP 2002, s. 78-78, 2002

Nakladatel

University of Helsinky, Finland

Další údaje

Jazyk

angličtina

Typ výsledku

Stať ve sborníku

Obor

10600 1.6 Biological sciences

Stát vydavatele

Finsko

Utajení

není předmětem státního či obchodního tajemství

Organizační jednotka

Přírodovědecká fakulta

Klíčová slova anglicky

homocysteine; derivatization

Štítky

derivatization, homocysteine

Změněno: 19. 5. 2009 18:44, prof. RNDr. Zdeněk Glatz, CSc.

Anotace

V originále

Ten years ago a new application for the evaluation of enzymatic reactions in capillary electrophoresis was proposed and developed, electrophoretically mediated microanalysis (EMMA). In this method, substrate(s) and enzyme are introduced into the capillary as distinct plugs, the first analyte injected being the one with the lower electrophoretic mobility. Upon the application of an electric field, the two zones interpenetrate due to the differences in their electrophoretic mobilities. Enzymatic reaction takes place and the resultant reaction product(s) and the unreacted substrate(s) are electrophoretically transported towards detector, where they are individually detected. Since its discovery by Bao and Regnier the EMMA methodology has been utilized in a number of biochemical systems - for assays of enzyme activities, determinations of substrates, Michaelis constants, etc. In this work, the EMMA method was applied to determine the kinetic parameters of rhodanese. Rhodanese (thiosulfate: cyanide sulfur transferase, EC 2.8.1.1) is responsible for the transfer of the sulfane sulfur of thiosulfate to an acceptor, which is likely to be cyanide under some physiological conditions: S2O32- + CN- = SCN- + SO32- Its physiological role has been debated for many years with proposals ranging from the detoxication of cyanide to suggestion that rhodanese is important in the bioenergetic oxidation of thiosulfate, in the generation of iron-sulfur protein complexes, in the lipoate metabolism and in the reactivation of nitrogenase. Several kinetic parameters of rhodanese was determined by means of EMMA - the Michaelis constant (Km) for thiosulfate and cyanide, Ki values of 2-oxoglutaric acid and the temperature optimum of the enzymatic reaction. Moreover, the kinetic mechanism of enzymatic reaction and types of inhibition of inhibitor were also elucidated. By best of our knowledge it is the first application of EMMA method for this purpose.

Návaznosti

GA525/00/0785, projekt VaV

Název: Acidifikační procesy v sulfidových odpadech

Investor: Grantová agentura ČR, Acidifikační procesy v sulfidových odpadech

Citovat

ŠEVČÍKOVÁ, Petra a Zdeněk GLATZ. Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide. In Proceeding of the 13th International Symposium on Capillary Electroseparation Techniques ITP 2002. Helsinki: University of Helsinky, Finland, 2002, s. 78.

@inproceedings{405933,
   author = {Ševčíková, Petra and Glatz, Zdeněk},
   address = {Helsinki},
   booktitle = {Proceeding of the 13th International Symposium on Capillary Electroseparation Techniques ITP 2002},
   keywords = {homocysteine; derivatization},
   language = {eng},
   location = {Helsinki},
   pages = {78-78},
   publisher = {University of Helsinky, Finland},
   title = {Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide},
   year = {2002}
}

TY  - JOUR
ID  - 405933
AU  - Ševčíková, Petra - Glatz, Zdeněk
PY  - 2002
TI  - Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide
PB  - University of Helsinky, Finland
CY  - Helsinki
KW  - homocysteine
KW  - derivatization
N2  - Ten years ago a new application for the evaluation of enzymatic reactions in capillary electrophoresis was proposed and developed, electrophoretically mediated microanalysis (EMMA). In this method, substrate(s) and enzyme are introduced into the capillary as distinct plugs, the first analyte injected being the one with the lower electrophoretic mobility. Upon the application of an electric field, the two zones interpenetrate due to the differences in their electrophoretic mobilities. Enzymatic reaction takes place and the resultant reaction product(s) and the unreacted substrate(s) are electrophoretically transported towards detector, where they are individually detected. Since its discovery by Bao and Regnier the EMMA methodology has been utilized in a number of biochemical systems - for assays of enzyme activities, determinations of substrates, Michaelis constants, etc. In this work, the EMMA method was applied to determine the kinetic parameters of rhodanese. Rhodanese (thiosulfate: cyanide sulfur transferase, EC 2.8.1.1) is responsible for the transfer of the sulfane sulfur of thiosulfate to an acceptor, which is likely to be cyanide under some physiological conditions: S2O32- + CN- = SCN- + SO32- Its physiological role has been debated for many years with proposals ranging from the detoxication of cyanide to suggestion that rhodanese is important in the bioenergetic oxidation of thiosulfate, in the generation of iron-sulfur protein complexes, in the lipoate metabolism and in the reactivation of nitrogenase. Several kinetic parameters of rhodanese was determined by means of EMMA - the Michaelis constant (Km) for thiosulfate and cyanide, Ki values of 2-oxoglutaric acid and the temperature optimum of the enzymatic reaction. Moreover, the kinetic mechanism of enzymatic reaction and types of inhibition of inhibitor were also elucidated. By best of our knowledge it is the first application of EMMA method for this purpose.
ER  -

ŠEVČÍKOVÁ, Petra a Zdeněk GLATZ. Monitoring of homocysteine in body fluids by means of MEKC and on-column reaction with 2,2-dipyridyldisulfide. In \textit{Proceeding of the 13th International Symposium on Capillary Electroseparation Techniques ITP 2002}. Helsinki: University of Helsinky, Finland, 2002, s.~78.

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