TY - JOUR
T1 - Analysis of biological thiols
T2 - Quantitative determination of thiols at the picomole level based upon derivatization with monobromobimanes and separation by cation-exchange chromatography
AU - Fahey, Robert C.
AU - Newton, Gerald L.
AU - Dorian, Randel
AU - Kosower, Edward M.
N1 - Funding Information:
This research was supported by the USPHS under Grant GM22122 from the National Institute of General Medical Sciences and Grant CA25009 from the National Cancer Institute. We are indebted to Dr. Thomas Ved-vick for providing samples of human red blood cells. E.M.K. is grateful to the European Research Office, U. S. Army for support.
PY - 1981/3/1
Y1 - 1981/3/1
N2 - A method for quantitative determination of biological thiols is presented. Thiols are converted to fluorescent derivatives by reaction with monobromobimane or monobromotrimethylammoniobimane. The derivatives are separated by ion-exchange chromatography and detected by fluorometry. Thiols that can be separated and quantitated by combined use of mBBr and qBBr derivatives include N-acetylcysteine, coenzyme A, coenzyme M, cysteine, cysteamine, cysteinylglycine, ergothioneine, ethanethiol, glutathione, γ-glutamylcysteine, homocysteine, 2-mercaptoethanol, methanethiol, pantetheine, 4′-phosphopantetheine, thiosulfate, thiouracil, and the mono- and diderivatives of dithiothreitol. Most thiols can be detected at the 1-pmol level and quantitated when present at the 10-pmol or higher level. Some, such as ergothioneine, exhibited low fluorescent yields and can be measured only at levels an order of magnitude higher. The method was applied to human red blood cell where the main thiols were found to be glutathione (2.4 mm) and ergothioneine (120 μm), in accord with earlier reports.
AB - A method for quantitative determination of biological thiols is presented. Thiols are converted to fluorescent derivatives by reaction with monobromobimane or monobromotrimethylammoniobimane. The derivatives are separated by ion-exchange chromatography and detected by fluorometry. Thiols that can be separated and quantitated by combined use of mBBr and qBBr derivatives include N-acetylcysteine, coenzyme A, coenzyme M, cysteine, cysteamine, cysteinylglycine, ergothioneine, ethanethiol, glutathione, γ-glutamylcysteine, homocysteine, 2-mercaptoethanol, methanethiol, pantetheine, 4′-phosphopantetheine, thiosulfate, thiouracil, and the mono- and diderivatives of dithiothreitol. Most thiols can be detected at the 1-pmol level and quantitated when present at the 10-pmol or higher level. Some, such as ergothioneine, exhibited low fluorescent yields and can be measured only at levels an order of magnitude higher. The method was applied to human red blood cell where the main thiols were found to be glutathione (2.4 mm) and ergothioneine (120 μm), in accord with earlier reports.
UR - http://www.scopus.com/inward/record.url?scp=0019472699&partnerID=8YFLogxK
U2 - 10.1016/0003-2697(81)90573-X
DO - 10.1016/0003-2697(81)90573-X
M3 - ???researchoutput.researchoutputtypes.contributiontojournal.article???
AN - SCOPUS:0019472699
SN - 0003-2697
VL - 111
SP - 357
EP - 365
JO - Analytical Biochemistry
JF - Analytical Biochemistry
IS - 2
ER -