Pól J. - Kterak toxikologický screening s použitím Orbitrapu odhalil

Kterak toxikologický screening s použitím
Orbitrapu odhalil nekalé praktiky
pražského podsvětí
Monika Židková, 1. LF UK, VFN
Jaroslav Pól. Thermo Fisher Scientific
The world leader in serving science
LC-MS metoda
Database
TIC
m/z and RT
XIC
Isotopic
Pattern
Full Scan
Full Scan
Identification or confirmation based on
m/z, RT, isotopic pattern
Spectral
Library
MS2
MS2 Scan
Identification or confirmation through
comparison with a spectral library
Hmotnostní rozlišení pro kvantifikaci
Hmotnostní rozlišení pro kvantifikaci
The Importance of Databases and Spectral Libraries
Database and Spectral
Library of compounds of
interest to the clinical
research and forensic
toxicology communities:
• Prescription drugs
• Natural and industrial
toxins
• Drugs of abuse
• Drugs of interest for
monitoring research
• Performance enhancing
drugs
Compound
Name
Clozapine N-oxide
Cocaethylene
Cocaine
Codeine
Colchicine
Coniine
Convallatoxin
Cortisone
Chemical
Formula
Extracted
Mass
Adduct
RT
Fragment
1
Fragment
2
Fragment
3
Fragment
4
Fragment
5
C18H19ClN4O
C18H23NO4
C17H21NO4
C18H21NO3
C22H25NO6
C8H17N
C29H42O10
C21H28O5
343.132
318.17
304.1543
300.1594
400.1755
128.1434
551.2851
361.201
M+H
M+H
M+H
M+H
M+H
M+H
M+H
M+H
5.9
5.3
4.8
3.4
5.9
3.1
5.6
6.1
243.0553
196.1328
182.1177
215.1061
358.1642
69.07058
85.02897
163.1117
256.063
82.06554
82.06519
243.1009
310.1194
55.05502
71.04983
121.065
244.0631
105.0336
105.0336
199.0749
282.1245
111.1171
351.1953
105.0703
227.0366
150.091
150.0915
225.0904
341.1377
83.08612
369.2058
145.1012
299.1051
108.0809
119.0493
58.06576
295.096
57.07066
323.2004
93.07037
The Importance of Databases and Spectral Libraries
Database and Spectral
Library of compounds of
interest to the clinical
research and forensic
toxicology communities:
• Prescription drugs
• Natural and industrial
toxins
• Drugs of abuse
• Drugs of interest for
monitoring research
• Performance enhancing
drugs
An updated version will be available later this year
with > 4500 compounds and metabolites
The Importance of Databases and Spectral Libraries
https://www.mzcloud.org/
A novel mass library/database
of finally cured fragmentation mass
spectra (MS/MS and MSn spectra)
Structural info for compounds
even if they are not represented in the
library through identification of
substructures
4,954 compounds - 8,024 trees - 1,335,070 spectra
Open consortium to establish a large
public domain library which our
software will link to
MzVault now available
Local based spectral library and compound database version of
MzCloud
Kazuistika
2 muži (40 a 36 let), požití neznámého bílého prášku
dezorientace
halucinace
křeče končetin
obličejové grimasy, vykřikování slabik
deficit vědomí
hypertenze 190/70
nalezený sáček PPS Powder s údajným obsahem
pyrilidinu n-propyl sulfobetainu
(„leštidlo“ pro galvanické niklování)
immunoscreening pozitivní fencyklidin
Metoda
Screening: full MS/AIF
Konfirmace: full MS/ddA
1 alikvot pH 3 - 4
L/L extrakce EtOAc
1 alikvot pH 8-9
Moč
precipitace ACN
Sérum
ředění 1:1 MF
Výsledky
Total ion chromatogram a extrahované chromatogramy vzorků moči a prášku „PPS“
RT: 4.00 - 20.01 SM: 5G
NL: 6.87E9
TIC F: FTMS + p
ESI Full ms
[100.00-600.00]
MS 04
8.64
100
TIC
80
60
9.22
40
20
4.67 4.76
6.16
6.83
7.51
04 #2239 RT: 8.64 AV: 1 NL: 3.87E9
8.40
9.54
10.44
12.75
T: FTMS
+ p ESI Full
ms [100.00-600.00]
11.15
11.68
100
8.65
XIC-moč 1
80
274.2160
13.03
[M+H]+
13.23 14.01
15.25 15.70 16.21
17.09
18.01 18.42
19.97
NL: 4.14E9
m/z=
274.21513274.21787 F: FTMS
+ p ESI Full ms
[100.00-600.00]
MS 04
95
90
85
60
80
40
75
20
70
7.19 7.59
0
100
8.33
8.54
9.27 9.71 10.16 10.71 11.28 11.86 12.43 13.19 13.67 14.16 14.91 15.53 15.88 16.63
65
XIC-moč 2
80
Relative Abundance
Relative Abundance
0
100
5.70
60
40
20
17.45 17.74
18.69 19.04
NL: 5.59E9
m/z=
274.21513274.21787 F: FTMS
+ p ESI Full ms
[100.00-600.00]
MS 05
60
55
50
45
40
274.0947
35
4.69
0
100
5.46
6.12
6.83 7.29 7.54
9.32 9.67 10.13 10.71 11.31 12.00 12.43 13.00 13.60 14.41 14.78 15.36 15.93 16.57 17.11 17.65 18.21
NL: 2.13E9
m/z=
274.21513274.21787 F: FTMS
+ p ESI Full ms
[100.00-600.00]
MS 06
25
XIC-oplach sáčku
80
19.41
30
8.69
275.2192
20
15
60
10
40
5 274.1174
20
0
4.01 4.76
0
5.37
6.25
7.09 7.52 7.79 8.04
274.2676 274.5785
275.0972
276.2223
275.2708
274.5
275.0 13.64 14.23
275.5
276.0
9.49 9.72 274.0
10.18 10.80 11.31
12.03 12.62 13.10
15.02 15.36 16.05
277.2257
276.5 18.15 18.50 277.0
17.05 17.50
19.33
277.5
m/z
4
5
6
7
8
9
10
11
12
Time (min)
13
14
15
16
17
18
19
20
278.0
Výsledky – Thermo Scientific FreeStyle SW
Výsledky
Molekulový vzorec:
C18H27NO
Přesná hmota:
m/z = 274.2165
Navržená látka – 2 izomery:
3(4)-MeO-PCP
4-MeO-PCP
Výsledky – ESI/HRMS full scan spektra
274.2160
Vzorek moči
•
•
189.1271
121.0648
50
75
100
125
150
175
200
225
250
275
274.2160
75
100
125
150
175
200
225
250
275
Ref. standard 4-MeO-PCP
189.1272
121.0650
274.2161
50
75
100
125
Ref. standard 3-MeO-PCP
189.1271
121.0648
50
Izolační okno 2 amu
Přesnost hmoty < 1.8 ppm
150
175
200
225
250
275
Výsledky – MS2 spektra potencionálních metabolitů
XIC 290.2146
3-MeO-PCHP
0
5
10
cis/trans-3-MeO-PPC
15
3-MeO-PCHP
cis/trans-MeO-PPC
86.0970
102.0918
189.1273
187.1116
121.0650
121.0650
75
100
125
150
175
200
225
75
250
100
275
125
150
175
200
225
250
275
Výsledky – TraceFinder „Target screning“
Výsledky – TraceFinder „Target screning“
3-MeO-PCP