recent in the in identification and quantification of · recent advances in the identification and...
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Recent advances in the identification and quantification of metallated compounds using chromatography and
mass spectrometry
Recent advances in the identification and quantification of metallated compounds using chromatography and
mass spectrometry
Laurent Ouerdane
and Ryszard Lobinski
CETAMA Seminar : "Nuclear analytical chemistry : New trends and future challenges", 24‐26 May 2011, Montpellier, France
Laboratoire de Chimie Analytique Bio‐inorganique et Environnement (LCABIE),
UMR 5254, Université
de Pau et des Pays de l’Adour / CNRS, Pau, France.
Sn
Trace elements in biology and medecine
● Transition metals act as cofactors
in some enzymes, binding substrates
or stabilize protein structure
● Studies of human and model organisms: definition of some molecular details of metal metabolization
● Challenge: get a systematic view of metal content, speciation, localization and use within organisms and ecosystems
Predict potential dispersion and bioaccumulation in the environment
of released radioisotopes (of studied elements or similar to)
DefinitionDefinition: Study of the metallome, interactions and functional connections of metal ions and their species
with genes, proteins, metabolites and other biomolecules in biological systems (IUPAC definition, 2010).
«
Metallomics
»
Genome and Transcriptome
Proteome
Metabolome
ARNm A
ARNm B
ARNm C
(γGlu‐Cys)n
‐Gly
N
COOH COOH
NH NH2
COOH
nicotianamine
COOH
COOH
OHHOOC
phytochelatins
citric acid
Phytochelatine synthaseCitrate synthaseNicotianamine synthase
ARNm D
Metal(‐complexes)
transporters (Yellow
Stripe Like …)
interactions
Mn+
Mn+
Mn+Mn+
Mn+
“Understanding the functional connections between genes, proteins, metabolites and mineral
ions is one of biology greatest challenges in the post‐genomic era” Lahner et al., Nature, 2003
Introduction to analytical issue
‐ Numerous metals and metalloids with high biological significance (=> accumulation)
‐
Unusual & unstable
compounds (complexes, element‐specific chemistry, redox
changes…)
‐ High diversity: from small polar metabolites to
proteins, covalent to non‐covalent
‐ Low
concentrated to trace
levels
‐ Case to case approach, no general method
⇒ Need for the development of a systematic approach by MS
HPLC, 1D GE, 2D GEHPLC, 1D GE, 2D GE……‐ Decrease amount of sample matrix
‐ Preconcentrate (multidimensional LC)
Elemental MSElemental MS‐ Tracking elements of interest
‐ Evaluate (pre)concentration
‐ Evaluate degradation (sample prep.)
Molecular MSMolecular MS
‐ Identify m/z of searched compounds
‐ Fragmentation and structure elucidation
‐ Screening in raw sample
Need sensitive
high resolution
MS instrumentNeed sensitive
and element
specific MS instrument
Analytical protocol
ICP‐Q‐MS with collision cell ESI‐LTQ Orbitrap MS
• LOW CONCENTRATED COMPOUNDS
• COMPLEX BIOLOGICAL MATRIX
Mass spectrometric techniques for metallomics
Elemental ions MSICP MS
Se+H+
O+
N+
C+ S+P+
(Pseudo)molecular ions MSElectrospray MS
http://zenobi.ethz.ch
4149.0 4153.8 4158.6 4163.4 4168.2m/z, amu
0
50
100
% In
tensity
4160.612
4159.613
4158.640
4161.598
4157.639
4162.630
4156.650
4163.614
4155.635
4164.616
4164.816
4154.616
4153.668
4152.643
Redundancy phenomena common (adducts)Ionization suppression by salts and co‐eluted ionsQuantification is difficult
ICP MS allows high‐throughput multielemental detection
Response virtually independent of matrix and analyte
Signal intensity is a linear function of element quantity
From ICP MS:
‐ Obtain precise chromatographic retention times of metalated compounds
‐ Estimate compound concentration
to evaluate feasibility for ESI MS analysis
From ESI LTQ Orbitrap MS:
‐ Search compounds with unusual mass defect
‐ Search element specific isotopic profile
‐
Search specific mass differences
between analogue
compounds (S Se) or between
metalated and non‐metalated forms
‐ Search specific inter‐isotopic mass differences
of a same
species
‐ Apply also these tools to
MSn
spectra
Data treatment
52Cr/50Cr
48Ti/46Ti
56Fe/54Fe
88Sr/86Sr
68
1012141618202224
60Ni/58Ni
34S/32S
66Zn/64Zn
37Cl/35Cl26Mg/24Mg
81Br/79Br
78Se/76Se
41K/39K
65Cu/63Cu
94Mo/92Mo
68Zn/66Zn
74Ge/72Ge
80Se/78Se
96Mo/94Mo
112Cd/110Cd
118Sn/116Sn
0
0,5
1
1,5
2
2,5
3
1,994 1,995 1,996 1,997 1,998 1,999 2 2,001 2,002 2,003 2,004 2,005
Ratio (m
+2X / mX)
Mass difference (m+2X ‐ mX)
Cu
Hv
Nw
Ox
Py
Naz
Mw
↑(13C ↑)
‐ Great help for compounds below 1000‐1500 amu
‐ Several “ratio‐Δm”
specific to one element
‐ Applicable for MSn
spectra
Δm = 2
Data treatment: “inter‐isotopic Δm »
Case studies
●
Identification of metal‐protein (metallothionein) complexes
●
Identification of metal‐metabolite complexes in plant fluids
●
Detection and characterisation of selenoproteins
●
Detection and characterisation of selenometabolites
Combined ICP MS and electrospray MS/MS detection in metallomics
exposure to CdS nanoparticles10 µm CdS
pig kidney cell line (LLC‐PK1)
preparation of
cytosol
Molecular response of renal cell lines to cadmium nanoparticles
ICP MS
ESI MS of
ESI MS
ESI MS/MS
Direct
After on‐line post‐column acidification (demetallation)
0 10 20Time, min
Intensity
114 Cd, cps
control
control spiked with Cd
sample
Reversed‐phase µHPLC
ICP‐MS‐assisted top‐down proteomics characterisation of bioinduced metallothionein
Metallothionein
(MT)
is
a
family
of
cysteine‐rich,
low
molecular weight (5‐10 kDa) proteins. MTs have the capacity to bind physiological (such as Zn, Cu) and
xenobiotic
(such
as
Cd,
Hg,
Ag)
heavy
metals
through
the
thiol
group of its cysteine residues)
Size‐exclusion fractionation
Identification of MT and their metal complexes
0 10 20 30 40 50Time (min)
0
20
40
60
80
100
Relative abun
dance
12
3
4
5
67
8
0
20
40
60
80
0
1
2
3
4
5
0 10 20 30 40 50 60
Intensity
114Cd
, cou
nts x104
12
3
4
5
6 7
8
ICP MS
100
Relative abun
dance
12
3
4
5
67
8ES MS
ES MSwith post‐column acidification
Ac‐M1
D2
P3
N1
C20
S9
A6
T3
G6
K6
R1
Q1
I1
V1
1202.0 1202.5 1203.0 1203.5 1204.0 1204.5 1205.0m/z
0
20
40
60
80
100Re
lative abun
dance
1203.242521203.04360
1203.64074
1202.844301203.84037
1204.040301202.64455
1204.24064
1204.441141202.444251204.64181
Mr
(theoretical): 6011.1772 Mr
(measured): 6011.1934δ
= 2.6 ppm
Difference‐7Cd +14H+
1354 1355 1356 1357 1358 1359 1360 13610
20
40
60
80
100
Relative abun
dance
1357.88285
1358.482321357.48310
1357.08321 1358.88223
1359.282501356.68374
1359.682381356.28382
1360.082271355.88514
1355.48591 1360.68003
Mr
(theoretical): 6784.3848 Mr
(measured): 6784.3752δ
= 1.4 ppm
Amino acid composition:
Mounicou, S., Ouerdane, L., l'Azou, B., Passagne, I., Ohayon‐Courtès, C., Szpunar, J., Lobinski, R. (2010) Analytical Chemistry, 82
(16), pp. 6947‐6957.
Top‐down sequencing of metallothioneins
90.3% of protein coverage 93.5% of protein coverageLow ppm mass accuracy
Y ionsZ=5
Z=4Z=3
Z=2Z=1
CID on m/z
= 1225.80
400 600 800 1000 1200 1400 1600 1800 2000m/z
0
20
40
60
80
100
Relative abun
dance
1168.23231z=5
1280.47564z=4
1381.26176z=4
1459.78848z=4
1706.96435z=3
603.19024z=1
762.25476z=1
944.32762z=1
1510.06344z=2
1104.43958z=2
487.14268z=1
1854.43436z=1
HCD on m/z
= 1225.80
400 600 800 1000 1200 1400 1600 1800 2000m/z
0
20
40
60
80
100 1298.23364z=4
1706.96475z=3
1168.23205z=5
1446.51663z=2
603.19037z=1
1494.56948z=3
944.32787z=1
402.14414z=1
647.22786z=1
1754.32243z=3
1104.43933z=2
1923.23152z=2
Relative abun
dance
M-D-P-N-C-S-C-A-A-A-G-D-S-C-T-C-A-N-S-C-T-C-K-A-C-K-C-T-S-C-K-K-S-C-C-S-C-C-P-P-G-C-A-K-C-A-Q-G-C-I-C-K-G-A-S-D-K-C-S-C-C-A
Form N°
Time, min
m/z, z=5, measured*
molecular mass,
measured
molecular mass,
theoretical
difference, ppm
m/z, z=5, measured*
molecular mass,
measured
molecular mass,
theoretical
difference, ppm
Mass difference (apo-
complexed)
1 33.8 1189.03184 5940.12023 5940.1279 1.3 M1D2P3N1C20S9A7T3G6K6R1Q1I1 1343.67156 6713.35380 6713.3428 1.6 7Cd - 14H+
2 35 1189.83748 5944.14843 5944.1592 1.8 M1D2P2N1C20S10A6T3G6K7R1V2 1344.6784 6718.38800 6718.3736 2.1 7Cd - 14H+
3 37.4 1197.43692 5982.14563 5982.1385 -1.2 Ac-M1D2P3N1C20S9A7T3G6K6R1Q1I1 1352.07300 6755.36100 6755.3534 1.1 7Cd - 14H+
4 38.1 1194.84007 5969.16135 5969.1593 -0.03 M1D2P3N1C20S9A6T3G6K6R1Q1I1V1 1349.47504 6742.37120 6742.3743 0.5 7Cd - 14H+
5 38.5 1198.24285 5986.17528 5986.1698 -0.09 Ac-M1D2P2N1C20S10A6T3G6K7R1V1 1353.07920 6760.39200 6760.3848 1.1 7Cd - 14H+
6 40.5 1203.24648 6011.19343 6011.1772 0.6 Ac-M1D2P3N1C20S9A6T3G6K6R1Q1I1V1 1357.88285 6784.37525 6784.3849 1.4 7Cd - 14H+
7 41 1203.23964 6011.15923 6011.1449 0.9 Ac-M1D2P3N1C20S9A7T3G6K5R2Q1I1 1357.87658 6784.34390 6784.3597 2.3 7Cd - 14H+
8 44.3 Ac-M1D3P2N1C20S8A7T2G6K8Q1I1V1 1272.16000 6355.79600 6355.7928 0.5 Cd2Cu2 - 8H+
Apo MTs
Amino acid composition
Complexed MTs
Expressed MTs identified in the LLC‐PK1 cell line
0
1
2
3
4
5
0 10 20 30 40 50 60
Inte
nsi
ty 1
14C
d, c
ou
nts
x104
12
3
4
5
6 7
8
RP HPLC -
ICP MS
Time, min
Case studies
●
Identification of metal‐protein (metallothionein) complexes
●
Identification of metal‐metabolite complexes in plant fluids
●
Detection and characterisation of selenoproteins
●
Detection and characterisation of selenometabolites
Combined ICP MS and electrospray MS/MS detection in metallomics
Metal speciation in post‐phloem of pea, Pisum sativum
SEC ICP-MS
365.0 365.2 365.4 365.6 365.8 366.0 366.2 366.4 366.6 366.8 367.0 367.2m/z
0.000.050.100.150.200.250.300.350.400.450.500.550.600.650.700.750.800.850.900.951.001.051.101.151.201.251.301.351.40
Rel
ativ
e A
bund
ance
365.06416 367.06264
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30Time (min)
0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
75
80
85
90
95
100
13.17
12.7414.04 16.17
18.69
18.48
m/z 869.845
m/z 811.840
m/z 753.835m/z 437.972
m/z 366.063
m/z 365.064
SEC ESI‐MS
808.0 809.0 810.0 811.0 812.0 813.0 814.0 815.0 816.0m/z
05
101520253035404550556065707580859095
100
Rel
ativ
e A
bund
ance
811.84007
812.84314
809.84495
813.84450810.84770
m/z 811.8403 Fe(III)cit2 mal2
Δm = 1.9951 amu (theor. 1.9953)
Δm = 1.9985 amu
(theor. 1.9982)
m/z 365.064Cu + nicotianamine
56Fe
54Fe
63Cu
65Cu
Rela
tive
abu
ndan
ce
FeFe33 --(citrate)(citrate)2 2 (malate)(malate)22 CC2020 HH2020 OO1919 --FeFe33
HILIC HILIC --
ICP MSICP MS(mass balance)(mass balance)
XylemXylem
FeFe
x105
4
8
Inte
nsit
y, c
ps
0 10 20 30
Time, min
50
100
Rela
tive
Abun
danc
e
40
m/z m/z 811,8420811,8420
XICXICPositive Positive ion modeion mode
810 811.0 812.0m/z
0
50
100
Rela
tive
Abu
ndan
ce
811.8420
809.8470
Zoom of ESI MS spectrum Zoom of ESI MS spectrum withwith
Fe patternFe patternIsotopic pattern of iron
100 200 300 400 500 600 700 800
m/z
0
50
100
Rela
tive
Abu
ndan
ce
577.8376
635.8428 679.8326
MS/MS spectrum (positive ion mode)
Validation of iron speciation in xylem (Pisum Sativum) by hydrophilic interaction chromatography (HILIC) with
the parallel ICP MS and electrospray MS/MS
1 3 Search for the elemental isotopic pattern
2 Confirm the retention time matching 4 Confirm the structureRe
lati
ve A
bund
ance
54 55 56 57 58m/z
50
100 56FeD = 0,0000
54FeD = -1,9953
57FeD = 1,0005
58FeD = 1,9983
Case studies
●
Identification of metal‐protein (metallothionein) complexes
●
Identification of metal‐metabolite complexes in plant fluids
●
Detection and characterisation of selenoproteins
●
Detection and characterisation of selenometabolites
Combined ICP MS and electrospray MS/MS detection in metallomics
Human selenoproteome
Adapted from Chavatte et al. 2004
Genetically coded specific incorporation to form SeCys (21st amino acid)
Important biological functions
Low abundance
Low stability during standard analytical procedures
Selenoproteins
25 selenoproteins were predicted by
bioinformatics on the basis of genomic
sequences; one third of them have never been detected in
vivo
and their functions are unknown
challenges:‐
sensitivity ‐
stability: conversion of selenocysteine residues to dehydroalanine
‐
high matrix protein load in real samples
15kDaDI1DI2DI3GPx1GPx2GPx3GPx4GPx6Sel HSel ISelKSelMSelNSelOSelPSel RSelSSel TSelVSelWSPS2TR1TR2TR3
Name Selenoprotein structure
Iodothyronine deiodinases
Glutathione peroxidases
Thioredoxin reductases
SelenoPhosphate Synthase 2
Internal organs
1‐5 ppmMuscles
0.2‐0.5 ppmAnimal/human blood
0.1‐0.2 ppmAnimal/human serum
0.05‐0.15 ppm
Intensity
signal, cps
Time, min0 5 10 15 20
0
1000
1000
1000
1000
1000
1000
1000
100 RP HPLC – ESI MS (XIC of some selenopeptides)
RP HPLC ‐
ICPMS (78Se) of SelP tryptic digest
Intensity
, relative abun
dance
T39‐40, m/z 796.321 (1+)
T33, m/z 544.142 (2+)
T41‐42m/z 990.233 (1+)
T42m/z 862.138 (1+)
T35m/z 587.227 (2+)
T37m/z 772.781 (2+)
T36m/z 791.333 (2+)
T38 glycolysatedm/z 1050.771 ( 3+)
Selenoprotein
P
(SelP)
is
the
main
selenoprotein
present in human blood (more than 55%
of Se).
SelP is thought to be present in multiple isoforms
(glycosylated
at
different
degrees,
with
1
to
10
SeCys).
Time, sec
0
0,005
0,01
0,015
0,02
0,025
0
0.5
1.0
1.5
2x104
0 500 1000 1500 2000
78Se
UV 254 nm
GSHPxSe‐albumin
SelP
Intensity
signal, cps
UV, Absorbance un
its
IMAC‐Co UV/ICP‐MS of human serumpH gradient
: 7 to 4.1 with ammonium acetate
Characterization of selenocysteines from selenoprotein P
m/z
772.78357
771.78395
773.78531771.28638770.78484
T37
Case studies
●
Identification of metal‐protein (metallothionein) complexes
●
Identification of metal‐metabolite complexes in plant fluids
●
Detection and characterisation of selenoproteins
●
Detection and characterisation of selenometabolites
Combined ICP MS and electrospray MS/MS detection in metallomics
Preparative scale SEC
0
1
2
P1P2
P3
P4
P5P6
P7
P8
3
0 50 100 150 200 250 300 350 400 450 500Elution volume, ml
Intensity
, cps
x105
Fractionation by ion‐exchange LC
ICP MS in selenometabolomics
Desalting by reversed‐phase LC
0
1
2
3
4
0 10 20 30 40 50
x104
Intensity
, cps
Time, min
0
0.4
0.8
1.2
1.6
2.0
0 10 20 30 40 50
x103
0
0.5
1.0
1.5
2.0
2.5
0 10 20 30 40 50
x105
0
2
4
6
8
10x104
0 5 10 15 20 25Time, min
0
2
4
6
8
10 x104
0 5 10 15 20 25Time, min
Intensity
, cps etc….. ESI MS/MSn
Dernovics and Lobinski, Metallomics 1 (4), pp. 317‐329
Se‐metabolites in yeast:State‐of‐the‐art of knowledge
Selenoglutathiones Selenoadenosyl species
Selenoamino acids derivatives
(m/z, [M+H]+) (m/z, [M+H]+)
Se
COOH
NH2CO
HN
OC
NH
HOOC
COOH
H2NOC
NH
C
S
OOH
S
SeN
N
OSeR1
OH OH
N
N
NHR3
HNHOOC
Se
3HOOC
Se
+ HNHOOC
Se
3HOOC
Se
+
R2
H
O
O
HH
H
OH
OH
H OH
H
OH
O
Se
HH
H
OH
OH
H
H
CH3
New Se‐species in plants by 3D HPLC‐FTMSn
Se‐carboxymethyl‐selenohomocysteine
O
OH
NH3+
SeOH
O
Selenoamino acids in Thlaspi arvense L.
C6
H12
NO4
Se+
Hexose‐Pentose‐SeCH3
Fragment of longer sugar chains and/or «
trapping
»
molecule for reactive and volatile compounds
Selenocarbohydrates in Oenanthe decumbens, Allium sativum
and Buddleia lindleyana
C12
H22
O9
Se
237 238 239 240 241 242 243 244 245m/z
0
50
100
Relative Ab
undance
241.99266
239.99353
243.99273237.99531238.99608
383 384 385 386 387 388 389 390 391 392 393 394m/z
0
50
100
Rel
ativ
e Ab
unda
nce
389.03576
387.03665
385.03865391.03600386.03933
m/zexp
= 241.99266m/zthéo
= 241.99261
Δ
= 0.21 ppm
m/zexp
= 389.03576m/zthéo
= 389.03571
Δ
= 0.13 ppm
Selenolipids in Se‐enriched yeast
C27
H57
NO9
PSe+
N+
CH3
OP
O
CH3
CH3
O
OH
OH
O
O
CH3
Se
OH
CH3 O
Formation of selenohydrin
Selenohydrin of C18:1 lyso‐phosphatidylcholine
Loss of selenohydrin=> m/z = 520.33962104
184 337
258
86
SEC – LTQ Orbitrap MS analysis of
yeast extract purified by SPE
Oxidation of lipids driven by
methaneselenic acid
100 150 200 250 300 350 400 450 500 550 600m/z
0
50
100
Relativ
e Abu
ndance
184.07319
520.33962
632.28241502.32919
258.11001337.27364
104.10697
‐H2O ‐H2O‐H2O
86.09637
645 646 647 648 649 650 651 652 653 654m/z
0
50
100
Relative Abu
ndance
650.29651
648.29745
651.29982646.29932
652.29670647.30009 649.30063653.29989
650.29
651
Se speciation in human urine at basal levels (ppb)
0
20
40
60
80
100
0 10 20 30 40
Fraction 5
Intensity
80Se
Fraction 4
0
50
100
150
200
0 10 20 30 40
Intensity
80Se
MS (zoom x 65) MS (zoom x315)
80Se
78Se
82Se
77Se76Se
Fraction 5MS spectrum at 20.25 minR = 30 000 at m/z 400
> 16 species
within 0.5 amu
Inter‐isotopic Δm matching
288 289 290 291 292 293 294m/z
291.13758
292.05523289.15056
288.10076
294.05525
293.05862
287.9 288.0 288.1 288.2 288.3 288.4m/z
288.10076
288.05793288.14318
288.16547288.27826
288.21633287.97292
287.90761
288.35442287.84037
150 500 1000m/z
229.15437
0
20
40
60
80
100
0 10 20 30 40
0 10 20 30 40Time (min)
0
50
100
Relative Ab
undance
at m/z 292.056(± 10 ppm)
Fraction 5
Intensity
80Se
HILIC – ESI MS (XIC at the mass of the
indentified Se‐compound
HILIC – ICP MS (80Se)
Fraction 4
0
50
100
150
200
0 10 20 30 40
at m/z 300.034(± 10 ppm)
0 10 20 30 40Time (min)
0
50
100
Relativ
e Abu
ndance
296.0 297.0 298.0 299.0 300.0 301.0 302.0m/z
0
50
100
Relativ
e Abu
ndance
300.03459
298.03540
302.03496296.03756297.03878
HILIC – ESI MS(mass spectrum at the RT of Se elution)
288.0 289.0 290.0 291.0 292.0 293.0 294.0m/z
0
50
100
Relativ
e Abu
ndance
292.05563
290.05624294.05572
288.05786
289.05844
Δm = 1.9994 amu
(theor. 1.9992)Δm = 1.9992 amu
(theor. 1.9992)
O
HH
OH
HOH
H NH
Se
OHCH3
O
CH3
H++
SeGalNAc
Identification of selenium compounds by high resolution MS
??
N+
O OH
CH3
CH3
CH3
N
NH
Se
CH3
NH
+N
Se
CH3
80 100 120 140 160 180 200 220 240 260 280 300 320m/z
0
50
100
Relativ
e Abu
ndance
188.99237
173.96898
248.0658294.05255
N+
CH3
CH3
CH3
N
NH
Se
CH3N
N +
NH
N
Se +
MS3
spectrum 292 → 248 →
‐
First time reported
‐
Ergothioneine family…
C10
H18
N3
O2
Se+
Identification of methylselenoine by MS/MS
292.05523 Fraction 5
0 5 10 15 20 25 30 35 40Time (min)
0
100
0
100
Relative Abu
ndance
20.82
22.46
MS3
335.06 → 291.06
→ 214.971
MS3
292.05 → 248.06
→ 188.991
Se‐methylselenoneine
Selenoneine
(derivatized)
0
1000
2000
3000
0 10 20 30 40
Se signal inten
sity, cps
Time (min)
0
100
0
10020.63
21.95MS3
287.10 → 243.10 → 167.027
MS3
244.10 → 200.11 → 141.047
Ergothioneine (derivatized)
S‐methylergothioneine
0 5 10 15 20 25 30 35 40
Bromine
interference
Time (min)
Relative Abu
ndance
Sulfur containing speciesSulfur containing species
Selenium Selenium
containing containing
speciesspecies
Chalcogenide histidine betaine compounds in human blood
ICP‐MSconfirmation:main seleno‐
metabolites
Presence confirmed in raw Presence confirmed in raw
filtered urine and in raw filtered filtered urine and in raw filtered
blood by HILIC ESI MSblood by HILIC ESI MS33
Klein M., Ouerdane L., Maïté
Bueno and Florence Pannier,
Metallomics, 2011, 3, 513‐520
Final results
•
The democratization of electrospray high resolution mass spectrometry opens
new perspectives in the analysis of individual selenium species,
metabolites
and proteins and their comprehensive high‐troughput monitoring.
•
The parallel detection by ICP MS is irreplaceable for the optimization of the
separation (control of the mass balance) and quantification.
•
Considerable method development work is still necessary to exploit the
synergy between ICP MS and ESI FT ion trap MSn. The dual MS is essential for
the successful speciation analysis
Conclusive combination of analytical techniques (multidim. HPLC, ICP MS & ESI HR MS) and data treatment methods
Collection of data relevant for biological or environmental studies
ICP‐MS + ESI HR MS: still an unexplored field…
Conclusive combination of analytical techniques (multidim. HPLC, ICP MS & ESI HR MS) and data treatment methods
Collection of data relevant for biological or environmental studies
ICP‐MS + ESI HR MS: still an unexplored field…
PERSPECTIVES
Ongoing development of parallel works‐
molecular biology
‐
localization/imaging (X‐ray)
‐
synthesis of the new identified compounds
Development of more customizable software for pattern screening Application to low mass elements by the use of ICP HR MS
Conclusions
Acknowledgements●
Nuclear
Toxicology
Programme
●
Agilent
●
Aquitaine Region
●
Agence Nationale de la Recherche
●
FEDER
B. Passagne, B. L’Azou, C. O’Hayon
Biochimie et Physiologie Moléculaire des PlantesAGRO‐M/INRA MontpellierS. Mari, C. Curie, L. Grillet
LCABIE, UPPA/ CNRS,
UMR 5254, Pau, France
Joanna SzpunarSandra Gil CasalSandra MounicouMarlène KleinKasia
Bierla
Paulina
Flis
Maité
Bueno
Florence PannierMihaly
Dernovics
Thanks for y
our
attention!