Determination of intracellular glutathione and cysteine using HPLC with a monolithic column after derivatization with monobromobimane.
An optimized high-performance liquid chromatography (HPLC) method is used to show that, as myoblasts differentiate into multinucleated muscle fibers, there is a shift to a more oxidized cell redox state. The HPLC method incorporated derivatization with monobromobimane for the determination of the reduced (GSH) and oxidized (GSSG) forms of glutathione and the reduced (Cys) and oxidized (CysSS) forms of cysteine. The derivatization was optimized to improve the sensitivity of the approach; the limits of detection for glutathione and cysteine were 3 x 10(-8) and 5 x 10(-8) M, respectively.
Conlan
Xavier A
XA
Institute for Technology Research and Innovation, Deakin University, Geelong, Victoria, Australia. xavier@deakin.edu.au <xavier@deakin.edu.au>
Stupka
Nicole
N
McDermott
Geoffrey P
GP
Francis
Paul S
PS
Barnett
Neil W
NW
D000368:Aged; D001643:Bridged Bicyclo Compounds; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003545:Cysteine / Q000032:analysis*; D003593:Cytoplasm / Q000737:chemistry; D005260:Female; D005978:Glutathione / Q000032:analysis*; D019803:Glutathione Disulfide / Q000032:analysis*; D006801:Humans; D042541:Intracellular Space / Q000737:chemistry*; D057230:Limit of Detection; D008297:Male; D018482:Muscle, Skeletal / Q000166:cytology*; D032446:Myoblasts / Q000737:chemistry; D010084:Oxidation-Reduction
D001643:Bridged Bicyclo Compounds; D005978:Glutathione; D003545:Cysteine; D019803:Glutathione Disulfide; C028381:monobromobimane