Study of reproducibility of spin trapping results in the use of C-phenyl-N-tert-butyl nitrone (PBN) for trichloromethyl radical detection in CCl4 metabolism by rat liver microsomal dispersions. Biological spin trapping I.
The well-known metabolism of CCl4 to trichloromethyl radicals in rat liver microsomal dispersions has been reinvestigated with the goal to determine the repeatability and reproducibility of the EPR signal intensity of the EPR spectrum of the CCl3 adduct of PBN. It was found that at least eight repeat experiments were needed under identical conditions to obtain an average value with an error of +/- 10%. When the effect of changing the concentrations of CCl4, PBN or NADPH-generating system was investigated, the plots of EPR signal intensity vs. the variable selected showed initial smooth increases in signal strength with respect to an increase in concentrations of CCl4, PBN or NADPH-generating system. However, considerable scatter was found after the initial slope and only general trends could be recognized. It is concluded that with CCl4, no increase in EPR signal is found after 10 mM concentration. For PBN, the optimum concentration is about 30 mM. The signal strength seems to increase with increased amounts of NADPH generating system although with diminishing slope.
Janzen
E G
EG
National Biomedical Center for Spin Trapping and Free Radical, Oklahoma Medical Research Foundation, Oklahoma City 73104.
Poyer
J L
JL
West
M S
MS
Crossley
C
C
McCay
P B
PB
D000818:Animals; D002247:Carbon Isotopes; D002251:Carbon Tetrachloride / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism*; D002374:Catalase / Q000494:pharmacology; D003497:Cyclic N-Oxides; D004121:Dimethyl Sulfoxide; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D009589:Nitrogen Oxides; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D012995:Solubility; D013113:Spin Labels
D002247:Carbon Isotopes; D003497:Cyclic N-Oxides; D009589:Nitrogen Oxides; D013113:Spin Labels; C025280:trichloromethyl free radical; C029217:phenyl-N-tert-butylnitrone; D009249:NADP; D002251:Carbon Tetrachloride; D002374:Catalase; D004121:Dimethyl Sulfoxide