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CONTRIBUTION OF ION MOBILITY FOR STRUCTURAL ANALYSIS AND ANALYTICAL CHEMISTRY: THE USE OF PROBE LIGANDS AND SELECTIVE IMS SHIFT REAGENTS C. Kune , J. Far, C. Delvaux, G. Eppe, E. De Pauw 1
CONTRIBUTION OF ION MOBILITY FOR STRUCTURAL ANALYSIS AND ANALYTICAL CHEMISTRY:
THE USE OF PROBE LIGANDS AND SELECTIVE IMS SHIFT REAGENTS
C. Kune, J. Far, C. Delvaux, G. Eppe, E. De Pauw
1
Plan:
1/ Concept of specific coordination complex formation with probe ligands
2/ Specificity depending on the presence of chemical function
• Proline/Valine model
• Contaminant of Selenomethionine – intramolecular oxidation
3/ Specificity depending on the steric hindrance o DAN isomers – Crown ether model
o Isomers ratio determination of two isobaric selenium compounds
4/ Specificity depending on the polarity – differential induce folding
o DAN isomers – Cyclodextrin model
2
Plan:
1/ Concept of specific coordination complex formation
2/ Specificity depending on the presence of chemical function
• Proline/Valine model
• Contaminant of Selenomethionine – intramolecular oxidation
3/ Specificity depending on the steric hindrance o DAN isomers – Crown ether model
o Isomers ratio determination of two isobaric selenium compounds
4/ Specificity depending on the polarity – differential induce folding
o DAN isomers – Cyclodextrin model
3
Structural elucidation
• Structural elucidation by ion mobility mass spectrometry (IMMS):
• Structural elucidation by computational chemistry
Not optimized
3D structure
Structure 1
Structure 2
Structure 3
CCS 1
CCS 2
CCS 3
Comparison Structure determination
IMS Quad. TOF MS
MSMS spectra Fragmentation CCS determination:
Information about 3D structure
m/z determination: Empirical formula
4
Structural elucidation with probe ligands
• Probe ligands : Molecules which interact with target ion to form specific complexes depending on their physicochemical properties
• Confirmation of physicochemical properties:
Such as the presence of chemical moeities
Such as steric hindrance
Such as polarity
• Formation of complexes with different CCS
Application as Selective Shift Reagent
5
Plan:
1/ Concept of specific coordination complex formation
2/ Specificity depending on the presence of chemical function
• Proline/Valine model
• Contaminant of Selenomethionine – intramolecular oxidation
3/ Specificity depending on the steric hindrance o DAN isomers – Crown ether model
o Isomers ratio determination of two isobaric selenium compounds
4/ Specificity depending on the polarity – differential induce folding
o DAN isomers – Cyclodextrin model
6
Time (ms) 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00
%
0
100 5.70
2.12
5.21
Ligand used:
18-crown-6 ether
≡
m/z 112 116 120
%
0
100
117.0746
116.0739
m/z 414 418 422
%
0
100 418.1847
420.1987 421.2030
Selective to carboxylic
acid
%
m/z 378 382 386
0
100 382.2555
383.2563
Selective to priminary
amino group
Specific coordination complex depending on the chemical functions:
Proline Valine model
Controlled ligand Specificity: Important IMS shift provided by crown ether complexation Selective shift Reagent (SSR)
Crown ether Selective to primary amino group
Crown ether + K+
Selective to carboxylate group
K+
7
m/z 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240
%
0
100 198,0047
180,9789
152,0004 108,9567 134,9710
C H 3 Se
O H
O
N H 3 +
Mass spectrum of SeMet (5ppm) (Synapt G2 as "QTOF MS ")
Exact mass: 198.0033uma Δm = 7.07ppm
Detection of isobaric compounds
m/z 195,960 196,000 196,040
100 196.0050
195.9897
%
0
Specific coordination complex depending on the chemical functions:
Application as SSR Se isotope pattern
…confirmed by high resolution mass spectrometry
0
%
100
C5H12NO278Se+
196,00434
195,97 195,99 196,01 196,03 m/z
195,98797
78Se isotope of SeMet
8
SeMet Contaminant
…with Se isotopic pattern
Exact mass univoque empiric formula for m/z = 196 ion Higher Double Bound Equivalent (DBE) value Double bound or ring formation ?
C5H10NO280Se+
Overlapping on the Se isotopic pattern Orthogonal separation required
0,02 a.m.u
Use the difference in chemical group Probe ligands
18-Crown-6 Ether Selective to primary amino group • Support the expected structure of 196 ion
• Remove the interferences from SeMet
Free interferences MS2 spectra of 196 ion
2,00 2,25 2,50 2,75 3,00 3,25 3,50
%
0
100
2,84
SeMet
Time (ms)
(Aire: 2,05e5)
2,56
196 ion
m/z 176 180 184 188 192 196 200 204
%
0
100 180,9789
195,9897
%
0
100
198,0047
176 180 184 188 192 196 200 204 m/z
dt = 2,56ms dt = 2,84ms
Separation by IMS-MS Separation by ion mobility spectrometry
intramolecular oxidation hypothesis
2,44
181 ion
Se N +
C H 3
O H
O
H XIC m/z 196
181 ion fragment (ammonium loss)
Unable to obtain MS2 spectra of 196 ion without
interference from SeMet or in-source fragment
Selenomethionine m/z: 196 ion
Specific coordination complex depending on the chemical functions:
Application as SSR for Selenomethionine
196,0043
9
Interferences free MS2 spectra of 196 ion
Time (ms)
2,55ms: Spectra of 196 ions without interferences
Time (ms)
0,00 1,00 2,00 3,00 4,00 5,00 6,00 7,00
%
0
100
5.05
6.02
4.01
1.95
2.82
3.52
4.23
Complexes with 18-crown-6 ether
(drift time of 196 ion) SeMet with low
concentration of 18-crown-6 ether (1:10)
0,00 1,00 2,00 3,00 4,00 5,00 6,00 7,00
%
0
100
2.88
2.44
4.28
6.02
(SeMet + 18-Crown-6 Ether)+ SeMet+
(181 ion)+
2,55ms: Spectra of 196 ions and Fragments of SeMet
(Free 18-Crown-6 Ether)+
(SeMet + 18-Crown-6 Ether)+ SeMet with High concentration of 18-crown-6 ether (1:50)
(Free 18-Crown-6 Ether)+
Specific coordination complex depending on the chemical functions: Application as SSR for Selenomethionine
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• Fragments of 196 ions: o Confirme the structure of 196 ion Formation of 5-membered ring compounds
o Differ from those of SeMet
%
0
100 167.9572
121.9495
94.9394
149.9448
195.9870
80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 m/z
240
Se N H +
O H
O
C H 3
Exact mass : 167.9564uma Δm = 4.76ppm
CH2
NH+Se
O
Exact mass : 149.9458uma Δm = 6.67ppm
Se N +
H
C H 2
Exact mass : 121.9509uma Δm = 11.5ppm Se
H
N+
H
Exact mass : 94.9352uma Δm = 44ppm
Se N +
C H 3
O H
O
H
Exact mass : 195.9877uma Δm = 3.57ppm
C H 3 Se
C H +
O H
O
C H 3 Se
N H 2 +
Se +
C H 2
C H 3
Se
C H 2 +
C H 3
Exact mass : 134.9713uma Δm = 2.2ppm
Exact mass : 151.9978uma Δm = 17.1ppm
Exact mass: 108.9556uma Δm = 10.1ppm
Exact mass : 180.9768uma Δm = 11.6ppm
m/z 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240
%
0
100 (4.74e5 cps)
198.0047
180.9789
152.0004 108.9567 134.9710
C H 3 Se
O H
O
N H 3 +
Exact mass : 198.0033uma Δm = 7.07ppm
Specific coordination complex depending on the chemical functions: Application as SSR for Selenomethionine
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Plan:
1/ Concept of specific coordination complex formation
2/ Specificity depending on the presence of chemical function
• Proline/Valine model
• Contaminant of Selenomethionine – intramolecular oxidation
3/ Specificity depending on the steric hindrance o DAN isomers – Crown ether model
o Isomers ratio determination of two isobaric selenium compounds
4/ Specificity depending on the polarity – differential induce folding
o DAN isomers – Cyclodextrin model
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0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00
%
0
100 9.28
3.09
5.43
Time (ms) 0.00 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00
%
0
100 5.43
2.98
9.33
Time (ms)
1,5
-DA
N
1,8
-DA
N
SSR used:
Dibenzo 24-crown-8 ether
≡
Controlled Stoechiometry: Application as Selective shift Reagent (SSR)
2 priminary amino groups available
Only one primary amino group available
Specific coordination complex depending on the steric hindrance:
Diaminonaphtalen isomers / Crown Ethers model
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Specific coordination complex:
100
m/z 668 672 676
%
0
667.2
129
671.1
658
669.1
625
dt
= 1
1.1
8m
s
0
100
50 100 150 200 250 300 350 m/z
359.0510
181.9814
271.0294 176.0618
%
m/z 355 359 363
%
0
100 355.0919
359.0
408
357.0
305
dt
= 6
.07m
s
0
100
50 100 150 200 250 300 350
135.9698 181.9814
336.1176 MS
/MS
%
0
2
4
6
8
10
12
14
0 2 4 6 8 10 12 14
Complexed isomer B
11.18
6.07 Isomer A
13%
87%
Good agreement with theoretical values obtained by computational chemistry and isomer ratio determined from raw data by Dernovics and coworkers
Travelling wave ion mobility did not successfully separate the native isomers of 2,3-DHP-selenocystathionine The use of a nitrobenzo 15-crown-5 ether as SSR allowed to perform the separation and quantification of the isomer ratio (87% - 13% ).
MS
/MS
Far and coworkers, Anal. Chem (2014) Vol.86, Issue 22, : 11246-11254
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Plan:
1/ Concept of specific coordination complex formation
2/ Specificity depending on the presence of chemical function
• Proline/Valine model
• Contaminant of Selenomethionine – intramolecular oxidation
3/ Specificity depending on the steric hindrance o DAN isomers – Crown ether model
o Isomers ratio determination of two isobaric selenium compounds
4/ Specificity depending on the polarity – differential induce folding
o DAN isomers – Cyclodextrin model
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Specific coordination complex depending on the polarity: DAN-Cyclodextrin model
%
0
100 2.09
15.21
2.00 6.00 10.00 14.00 18.00 Time (ms)
m/z 1290 1294 1298
1293.4863
1297.4518
%
0
100
m/z 155 159 163
%
0
100 159.0936
160.0932
1,5
-DA
N
%
0
100 x 50 2.20
15.44
2.00 6.00 10.00 14.00 18.00 Time (ms)
m/z 1290 1294 1298
%
0
100 1297.4518
1293.4863
m/z 155 159 163
%
0
100 159.0936
160.0988
1,8
-DA
N
Time (ms)
%
0
100 2.32
14.44
2.00 6.00 10.00 14.00 18.00
m/z 1290 1294 1298
1293.4863
1297.4518
%
0
100
m/z 155 159 163
%
0
100 159.0936
160.0988
2,3
-DA
N
Controlled folding: Application as Selective Shift Reagent (SSR)
2,3-DAN: Exclusion 1,5-DAN:
Inclusion
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Conclusion and perspectives
• Confirmation of hypothetical structure o Probe physicochemical properties
• As chemical functions, steric hindrance, polarity, pi stacking
• Use as Selective Shift Reagent o Allow a control of the arrival time distribution of ion
• Separation of isomers
• Obtention of interference free MS2 spectra
• Perspectives: o Use ligands to probe the three dimensional structural of larger (bio)molecules
• As peptide, protein, DNA
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Aknowledgement:
• Laboratory of Mass Spectrometry
o Docteur Johann Far Professeur Gauthier Eppe
o Cédric Delvaux Professeur Edwin De Pauw
• Laboratory of Theoritical Physical Chemistry o Professeur Françoise Remacle
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Thanks for your attention
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