Post on 28-Mar-2015
M. Triquigneaux, B. Tuccio, R. Lauricella et L. Charles
Laboratoire Chimie Provence, UMR 6264, Equipe Spectrométries Appliquées à la Chimie Structurale, Campus de St Jérôme, 13397 Marseille cedex 20.
EPR/MS combination:
Mechanism elucidation applied
to thiol compounds
Electronic Paramagnetic Resonance
Specificity paramagnetic species only
Sensitivity concentration 10-7-10-8 mol/L
Structure g factor, hyperfine coupling constants…
R1 N
O
R2
Nitroxide
EPR/MS Combination
1
How to access the global structure of nitroxide ?
EPR/MS Combination
aN ~ 15.5 G
aH ~ 1.8 GN
O
CH
NH2
NH
O
O
HN
HN
NH
NH2
O
OH
N
O
CH
OHaN aH
2
EPR/MS Combination
Mass spectrometry
Detection of nitroxide radical species
Detection of derived redox species
Nitroxide Oxoammonium Hydroxylamine
m/z m/z – 1 m/z + 1
Tuccio B., Lauricella R., Charles L., Int. J. Mass Spec., 252 (2006), 47-53.
El Hassan I., Charles L., Lauricella R., Tuccio B., New J. Chem., 32 (2008), 680-688.
H+
R1 N
O
R2 R1 N+
O
R2 R1 N
OH
R2+2
H+
3
Glutathione
Aim of the study: reactivity of glutathione toward MNP
O
HO
NH2
O
NH
O
HN
O
OH
SH
R.
antioxidant
physiological substrate
toxic by-products
S-transferase MNPxenobiotics
N
O
?
MNP
4
1) Glutathione MNP
2) Oxidant
S-adduct
O
HO
NH2
O
NH
O
HN
O
OH
S
NO
aN = 18.2 G
EPR Study
Experimental conditions
5
1) Glutathione MNP Oxidant
2) Photo-irradiation
EPR Study
O-adduct
NO O
O
HO
NH2
O
NH
O
HN
O
OH
S
NOO
aN = 27.1 G
Experimental conditions
6
No Detection of S- and O-adducts as nitroxides
Detection of derived redox species
MS Study
S-adduct
O-adduct
m/z 393.1 m/z 407.1
m/z 409.1
ESI-MS in negative mode
GS
NOCH3
- H
-- H
-
GNSHO
- H
-
GSN
OHO
7
MS/MS Study
S-adductESI-MS/MS in negative mode
128.1143.1
179.1210.1
254.1
272.1
393.1
25 eV393.1
m/z
Glutathione product ions
Dieckhaus C., Fernandez-Metzler C., King R., Krolikowski P., Baillie T., Chem. Res. Toxicol., 18 (2005), 630-638.
O
HO
NH2
O
NH
O
HN
O
OH
S
NOH
- H
-
O
HO
NH2
O
NH
O
HN
O
OH
S
NOH
- H
-
8
MS/MS Study
O-adduct
ESI-MS/MS in negative mode
128.1143.1
179.1210.1
254.1
272.1
409.1 20 eV409.1Glutathione product ions
321.1304.1
m/z
- H-
O
HO
NH2
O
NH
O
HN
O
OH
S
NOHO
- H-
O
HO
NH2
O
NH
O
HN
O
OH
S
NOHO
9
MS/MS Study
m/z 409.1
m/z 409.1
m/z 321.1
m/z 304.1
N
OH
-
N
OH
-
OH
O
HO
NH2
O
NH
O
HN
O
OH
S
NOHO
- H
-
-
O
HO
NH2
O
NH
O
HN
O
OH
S
O- H
-- H
O
HO
NH2
O
NH
O
HN
O
OH
S
O
-- H
O
HO
NH2
O
NH
O
HN
O
OH
S
NOHO
H
-- H
O
HO
NH2
O
NH
O
HN
O
OH
S
10
MS/MS Study
Methylated product of S-adduct
ESI-MS/MS in negative mode
128
143254
Glutathione product ions
m/z 407.1 m/z 272.1
-- H
O
HO
NH2
O
NH
O
HN
O
OH
S
NOCH3
-- H
O
HO
NH2
O
NH
O
HN
O
OH
S
NOCH3 -
- HO
HO
NH2
O
NH
O
HN
O
OH
11
Inverted spin trapping
Forrester-Hepburn
-NO + NuH -N(OH)-Nu -N(O .) -Nuoxidation
-NO -.+NO -N(O.)-Nu oxidation
hν
NuH
Detection of reaction intermediates
Distinct experimental conditions
Mechanism elucidation
12
Mechanism elucidation
Forrester-Hepburn
Inverted spin trapping
MS RPE
N
O
N
OH
SG N
O
SGGSH oxidation
N+
O
N
O
OSGoxidation
h
GSH
GSOH
oxidation
S-adduct
O-adduct13
MNP/GSH/oxidant Stable S-adduct
New O-adduct
Derivatization methylated product
Detailed reactivity of glutathione toward MNP
EPR/MS Structural characterization
Mechanism elucidation
Efficient method for radical study
Conclusion
14
SACS teamProvence University,Laboratory LCP,Marseille
Spectropole FEDER OBJ2142-3341
Organization committee SMAP 2009
Acknowledgments