Best Practices for OINDP Pharmaceutical Development
Programs
Leachables and Extractables
IV. Analysis of Leachables and Extractables
PQRI Leachables & Extractables Working Group
September 2006
PQRI Training Course
September 20-21, 2006
Washington,
PQRI Training Course DC
1
Key Points to Consider
►Identification of leachables and extractables is a
problem in Trace Organic Analysis.
► “Trace Organic Analysis” can be defined as the
qualitative and quantitative analysis of a
complex mixture of trace level organic
compounds contained in a complex matrix.
►Other similar problems include:
 Analysis of pollutants in environmental matrices
 Organic geochemical analysis
 Metabolite profiling in biological matrices
September 2006
PQRI Training Course
2
Key Points to Consider (continued)
►Trace Organic Analysis problems require
(in general):
 Some knowledge of the chemical nature of
the analyte mixture and matrix (Supplier
Information).
 Removal/extraction of the complex mixture of
organic compounds from the matrix.
 Separation of the complex mixture of organic
compounds into individual chemical entities.
 Compound specific detection of the individual
chemical entities within the complex mixture.
September 2006
PQRI Training Course
3
Compound Specific Detectors
(from the ridiculous to the sublime)
►Single-crystal X-ray Spectrometer
►FTIR (Fourier Transform Infrared
Spectrophotometer)
►NMR (Nuclear Magnetic Resonance
Spectrometer)
►Mass Spectrometer
 GC/MS
 LC/MS
September 2006
PQRI Training Course
4
September 2006
PQRI Training Course
5
What is a Mass Spectrometer???????
► Sample
inlet system
► Ion Source
► Mass analyzer
► Detector
September 2006
PQRI Training Course
6
September 2006
PQRI Training Course
7
Sample Inlet Systems
► Direct
Insertion Probes (EI/CI/FAB)
► Batch Inlet Systems (AGIS/heated septum
inlets/”targets”)
► Gas Chromatographs
► Liquid Chromatographs (with or without
separation in an HPLC column)
September 2006
PQRI Training Course
8
Ionization Processes
►
►
►
►
►
►
►
►
►
EI CI FI
FAB LSIMS TSP ESI APCI MALDI -
September 2006
electron ionization
chemical ionization
field ionization
fast atom bombardment
liquid secondary ion
thermospray
electrospray
atmospheric pressure CI
matrix assisted laser desorption
PQRI Training Course
9
Mass Analyzers
► Magnetic
sector (high resolution)
► Quadrupole filter (“benchtop”/Triple
Quad.)
► Time-of-flight (scan rate/mass
range/accurate mass measurements)
► Quadrupole ion trap (MSn)
► Ion cyclotron resonance (FTMS/ultrahigh resolution/mass range/accurate
mass measurements)
September 2006
PQRI Training Course
10
GC/MS – Circa Late 1980s
Circa 2006
September 2006
PQRI Training Course
11
GC/MS – Electron Ionization
September 2006
PQRI Training Course
12
GC/MS Extractables Profile of an Elastomer
A b u n d a n c e
T IC : 0 2 0 3 0 3 0 1 .D
9 0 0 0 0 0 0
8 5 0 0 0 0 0
antioxidant
Paraffin related
8 0 0 0 0 0 0
7 5 0 0 0 0 0
7 0 0 0 0 0 0
6 5 0 0 0 0 0
6 0 0 0 0 0 0
5 5 0 0 0 0 0
5 0 0 0 0 0 0
4 5 0 0 0 0 0
4 0 0 0 0 0 0
3 5 0 0 0 0 0
Coumarone-indene
resin related
3 0 0 0 0 0 0
2 5 0 0 0 0 0
2 0 0 0 0 0 0
1 5 0 0 0 0 0
1 0 0 0 0 0 0
5 0 0 0 0 0
0
5 .0 0
1 0 .0 0
1 5 .0 0
2 0 .0 0
2 5 .0 0
3 0 .0 0
3 5 .0 0
T im e -->
September 2006
PQRI Training Course
13
GC/MS Extractables Profile of Polypropylene
A b u n d a n c e
T I C :
0 7 1 5 0 3 0 3 . D
1 3 0 0 0 0 0
antioxidant
1 2 0 0 0 0 0
1 1 0 0 0 0 0
1 0 0 0 0 0 0
9 0 0 0 0 0
Polypropylene oligomers
8 0 0 0 0 0
7 0 0 0 0 0
6 0 0 0 0 0
5 0 0 0 0 0
4 0 0 0 0 0
3 0 0 0 0 0
2 0 0 0 0 0
1 0 0 0 0 0
5 . 0 0
T im
1 0 . 0 0
1 5 . 0 0
2 0 . 0 0
2 5 . 0 0
3 0 . 0 0
3 5 . 0 0
e -->
September 2006
PQRI Training Course
14
Interpreting a Mass Spectrum
►Confirm molecular weight!!!!!!!!!!!
 Adduct ions/multiply charged ions (LC/MS)
 Alternate ionization techniques (EI/CI and
APCI/ESI)
 Note features of the molecular ion
(obvious heteroatoms/nitrogen rule)
►Rationalize significant fragmentation
processes:
 Structures of fragment ions
 Mechanisms for fragment ion formation
September 2006
PQRI Training Course
15
“Good” Library Search Result
A b u n d a n c e
S c a n
3 8 8 1
( 1 5 .8 9 9
m
in ) : 0 7 1 1 0 3 0 1 . D
1 6 7
9 0 0 0
8 0 0 0
7 0 0 0
6 0 0 0
5 0 0 0
4 0 0 0
3 0 0 0
2 0 0 0
6 9
1 0 0 0
8 3
9 6
1 0 9
1 2 3
5 0
0
2 0
4 0
1 3 5
1 5 1
6 0
8 0
1 0 0
1 2 0
1 4 0
2 0 7
1 6 0
1 8 0
2 0 0
2 3 1
2 2 0
2 4 0
2 6 8
2 6 0
m /z - - >
A b u n d a n c e
# 5 1 9 2 3 : 2 ( 3 H ) - B e n z o t h ia z o le t h io n e
( C A S )
1 6 7
$ $
B E N Z O T H IA Z O L E - 2 -
9 0 0 0
8 0 0 0
7 0 0 0
6 0 0 0
5 0 0 0
4 0 0 0
3 0 0 0
2 0 0 0
6 9
1 0 0 0
2 8
4 5
9 1
1 0 9
1 2 3
5 8
1 4 0
0
2 0
m
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 2 0
2 4 0
2 6 0
/z - - >
September 2006
PQRI Training Course
16
“Good” Library Search Result
A
b
u
n
d
a
n
c
e
S
8
9
0
0
0
8
0
0
0
7
0
0
0
6
0
0
0
5
0
0
0
4
0
0
0
3
0
0
0
2
0
0
0
1
0
0
c
1
0
5
/ z - - >
u n d a
n
c
0
4
6
( 2
1
. 4
6
8
m
2
i n
) :
0
7
1
1
0
3
0
1
. D
0
8
0
1
0
1
0
0
3
1
:
5
1
8
0
1
0
4
0
8
4
2
2
0
i n
c
3
5
1
4
1
4
9
0
1
1
6
6
0
7
1
8
0
2
0
1
2
0
0
1
6
2
2
2
3
0
1
2
2
4
0
5
2
2
7
6
0
2
8
8
0
9
3
0
0
3
2
0
3
4
3
6
0
3
6
0
0
3
6
0
6
e
1
8
8
9
0
0
0
8
0
0
0
7
0
0
0
6
0
0
0
5
0
0
0
4
0
0
0
3
0
0
0
2
0
0
0
1
0
0
0
Z
,
b
i s
( d
i m
e
t h
y
l c
a
r b
a
m
o
d
i t h
i o
a
t o
- S
, S
' ) - ,
3
1
4
7
4
2
0
1
3
0
6
0
8
8
0
- 4
) -
(
1
0
0
1
2
0
7 1
3
1
4
0
5
1
2
6
0
1
8
4
6
2
1
0
4
2
1
4
( T
8
0
/ z
4
3
6
6
#
m
5
1
7
4
b
n
3
0
m
A
a
8
0
2
0
0
2
2
0
3
1
2
2
4
0
5
2
2
7
6
0
2
8
8
0
9
3
0
0
3
2
0
3
4
- - >
September 2006
PQRI Training Course
17
“Questionable” Library Search Result
A
b u n d a n c e
S
c a n
5 4 4 6
( 2 1 . 4 6 8
m
in ) :
0 7 1 1 0 3 0 1 . D
8 8
9 0 0 0
8 0 0 0
7 0 0 0
3 0 4
6 0 0 0
5 0 0 0
4 0 0 0
3 0 0 0
1 2 0
1 8 4
2 0 0 0
2 1 6
7 3
1 0 0 0
5 6
1 4 9 1 6 7
1 0 5
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
2 3 1
2 0 1
0
1 8 0
2 0 0
2 2 0
2 4 0
2 8 9
2 5 7
2 6 0
2 8 0
3 6 6
3 0 0
3 2 0
3 4 0
3 6 0
m / z - - >
A b u n d a n c e
# 1 1 7 0 2 1 :
8 8
Z
in c ,
b is ( d im
e t h y lc a r b a m
o d it h io a t o - S
, S
') - ,
( T
- 4 ) -
9 0 0 0
8 0 0 0
7 0 0 0
6 0 0 0
5 0 0 0
4 0 0 0
4 4
3 0 0 0
2 0 0 0
1 0 0 0
1 2 0
7 2
3 0 4
1 8 4
1 5
0
2 0
m
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 2 0
2 4 0
2 6 0
2 8 0
3 0 0
3 2 0
3 4 0
3 6 0
/ z - - >
September 2006
PQRI Training Course
18
“Questionable” Library Search Result
A b u n d a n ce
S c a n 7 7 4 2 ( 2 9 .6 3 9 m i n ) : 0 7 1 5 0 3 0 3 .D
2 8 7
8 0 0 0
5 7
6 0 4
1 1 7
6 0 0 0
1 4 7
4 0 0 0
3 9 9
2 1 5
8 3
1 7 5
2 0 0 0
3 3 5
2 5 5
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
3 0 0
4 4 1
3 6 9
3 5 0
4 0 0
4 5 0
5 4 7
5 0 0
5 5 0
6 0 0
m /z - - >
A b u n d a n ce
# 2 6 9 0 5 7 : d e lta .- ( 2 - F o r m y lv in y l) m e s o p y r o p h e o p h o r ib id e a M e th y
6 0 4
8 0 0 0
6 0 0 0
4 0 0 0
5 1 7
4 8 0
2 0 0 0
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
3 0 0
3 5 0
4 0 0
4 5 0
5 0 0
5 5 0
6 0 0
m /z - - >
September 2006
PQRI Training Course
19
Example of Chemical Ionization (ammonia
reagent gas)
T071703005 1173 (21.561) Cm (1171:1177-(1164:1168+1201:1214))
88
100
Scan CI+
4.35e3
CI+ (ammonia)
100
74
[M+NH4]+
%
130
386
208
86
70
61
73
101
84
98
191
192
177
128
163
175 178
135
114
161
152
193
124
0
F07013003 1111 (21.557) Cm (1109:1112-1116:1122)
100
209
218
229 237
252 257
272 274 276 293
313
312
314
330
331
387
352353 367
368 370
377
388 397
409
420
425
MS2 EI+
4.86e4
191
EI
163
.
175 178
M+ (???)
%
135
368
141
57
91
65
0
60
77
69
79
119
105
103
147 155
133
117
September 2006
100
121
120
192
312
179
180
85
80
169
140
160
180
193 202
200
256
215
227 237
220
240
255
257 267 281 295
260
PQRI Training Course
280
297 311 313 321
300
320
369
339
353 354
340
360
370
380
m/z
400
420
20
Leachables Profile – 1 Week Timepoint
Expanded Section
A b u n d a n c e
T IC : 1 1 1 0 0 3 0 3 .D
5 0 0 0 0 0
4 5 0 0 0 0
4 0 0 0 0 0
3 5 0 0 0 0
3 0 0 0 0 0
2 5 0 0 0 0
2 0 0 0 0 0
1 5 0 0 0 0
1 0 0 0 0 0
5 0 0 0 0
0
1 3 .5 0
1 4 .0 0
1 4 .5 0
1 5 .0 0
1 5 .5 0
1 6 .0 0
1 6 .5 0
1 7 .0 0
T im e -->
September 2006
PQRI Training Course
21
Library Search Identification of
A b u n d a n c e
S c a n
4 0 9 1
(1 6 .6 4 6
m in ): 1 1 1 0 0 3 0 3 . D
5 7
9 0 0 0
7 1
8 0 0 0
7 0 0 0
8 5
6 0 0 0
5 0 0 0
4 0 0 0
3 0 0 0
2 4 6
9 9
2 0 0 0
1 0 0 0
1 2 3
1 4 1 1 5 5
1 1 1
1 6 9 1 8 2
1 9 7
2 1 1
2 8 2
2 2 5
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 2 0
2 4 0
2 6 0
2 8 0
m /z -->
A b u n d a n c e
# 1 6 3 8 7 9 : E ic o s a n e
4 3
9 0 0 0
(C A S ) $ $
n -E ic o s a n e
5 7
8 0 0 0
7 1
7 0 0 0
6 0 0 0
8 5
5 0 0 0
4 0 0 0
2 9
3 0 0 0
2 0 0 0
9 9
1 0 0 0
1 1 3
1 2 7
1 4 1
1 5 5
1 6 9
1 8 3
1 9 7
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 1 1
2 2 5
2 2 0
2 3 9
2 4 0
2 5 3
2 6 7
2 6 0
2 8 2
2 8 0
m /z -->
September 2006
PQRI Training Course
22
Leachables Profile – 1 Week Timepoint
Expanded Section (2)
A bundance
T IC : 1 1 1 0 0 3 0 3 .D
120000
110000
100000
90000
80000
70000
60000
50000
40000
30000
20000
10000
0
1 7 .8 01 8 .0 01 8 .2 01 8 .4 01 8 .6 01 8 .8 01 9 .0 01 9 .2 01 9 .4 01 9 .6 01 9 .8 02 0 .0 02 0 .2 02 0 .4 02 0 .6 0
T im e -->
September 2006
PQRI Training Course
23
Library Search Identification of
A b u n d a n c e
S c a n 4 5 7 9 ( 1 8 .3 8 3 m in ) : 1 1 1 0 0 3 0 3 .D
8 3
9 7
6 1
(-)
8 0 0 0
6 0 0 0
1 1 1
4 0 0 0
1 2 5
2 0 0 0
1 3 9
1 5 31 6 6 1 8 1
2 2 4
2 5 2
2 0 7
2 9 7
0
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 2 0
2 4 0
2 6 0
2 8 0
3 0 0
m /z - - >
A b u n d a n c e
# 1 8 7 5 0 7 : A c e tic a c id , o c ta d e c y l e s te r $ $ O c ta d e c y l a c e ta te $
4 3
8 0 0 0
6 0 0 0
5 7
8 3
4 0 0 0
1 1 1
7 0
2 0 0 0
0
9 7
1 2 5
2 9
1 3 9 1 5 3
1 6 7 1 8 1 1 9 6 2 0 9 2 2 4
1 5
2 0
4 0
6 0
8 0
1 0 0
1 2 0
1 4 0
1 6 0
1 8 0
2 0 0
2 2 0
2 5 2
2 4 0
2 6 0
2 9 7
2 8 0
3 0 0
m /z - - >
September 2006
PQRI Training Course
24
Key Points to Consider
► EI
is under “kinetic control”. Therefore, EI
spectra are reproducible and spectral libraries
can be employed (remember limitations of
libraries!!!!!!!).
► CI is under “thermodynamic control”. It is useful
for molecular weight confirmation but no
libraries.
► GC/MS provides both a chromatographic
retention time and mass spectrum (spectra)
which can be compared with an authentic
reference material (Good luck finding it).
September 2006
PQRI Training Course
25
LC/MS Interfaces/Ionization Processes
► Moving
belt/wire (transport device)
► Thermospray (unique ionization process)
► Continuous flow FAB (transport device)
► Particle-beam (transport device)
► Electrospray (unique ionization process)
► APCI (transport device/ionization process)
September 2006
PQRI Training Course
26
A “Modern” LC/MS System
September 2006
PQRI Training Course
27
“Bench-top” LC/MS Systems
September 2006
PQRI Training Course
28
Schematic of an Orthogonal API Source
September 2006
PQRI Training Course
29
September 2006
PQRI Training Course
30
Features of
Electrospray
► Often
reflects solution chemistry
(multiply charged ions observed)
► Operates at atmospheric pressure
► Uses a strong electric field
► Flow-rate (optimum performance at
uL/min)
► Soft-ionization process
► Extremely rugged
September 2006
PQRI Training Course
31
Features of APCI
► Gas
phase ionization process
► Operates at atmospheric pressure
► Flow-rate (optimum performance at
mL/min)
► Uses a corona discharge
► Soft-ionization process
► Extremely rugged
September 2006
PQRI Training Course
32
APCI LC/MS Extractables Profile(s)
T072303003
100
280
An1
1.54e5
2.93
1.90
17.36
UV @280 nm
3.59
%
4.53
1.58
5.08 5.54
8.85
1.53
0
T072303003
100
16.16
17.28
Scan AP+
TIC
1.33e6
17.95 18.21 18.93 19.57
TIC (APCI+)
12.19
%
1.65 1.95
0
2.00
September 2006
4.00
6.00
8.00
10.00
12.00
PQRI Training Course
14.00
16.00
18.00
Time
20.00
33
Mass Chromatograms (Diphenylamine Acetone
Condensate)
T072303003
100
m/z 210 (dimethyldihydroacridine)
%
1.52
3.73
3.12 3.17
1.95
1.68
2.43
2.19
Scan AP+
210
3.01e3
3.65
2.67 2.80 2.88
3.01
4.08
3.89
3.39
4.51 4.56
4.27
0
T072303003
100
4.88 4.99 5.09 5.15
5.39 5.47 5.57
4.67
Scan AP+
212
7.17e3
4.56
m/z 212 (isopropyldiphenylamine)
%
2.99
2.27
1.87
3.12
2.45
0
T072303003
100
3.31
3.97
3.65
5.01 5.12
4.27
5.33 5.47
Scan AP+
170
5.31e3
2.96
m/z 170 (diphenylamine)
%
1.49
1.68
1.76 1.92 2.05 2.19 2.29
0
T072303003
100
2.51 2.59
3.23
3.36 3.49
2.80
3.81 3.92
4.24
4.40
4.45
4.99 5.09
4.61
5.28 5.39
5.55
280
An1
1.54e5
2.93
1.90
UV
%
3.65
3.59
4.53
1.58
5.08
5.54
1.53
0
Time
1.60
1.80
September 2006
2.00
2.20
2.40
2.60
2.80
3.00
3.20
3.40
3.60
3.80
PQRI Training Course
4.00
4.20
4.40
4.60
4.80
5.00
5.20
5.40
5.60
34
APCI+ Mass Spectrum of
Isopropyldiphenylamine
[M+H+AcCN]+
T072303003 170 (4.559) Cm (166:174-153:164)
100
[M+H]+
Scan AP+
4.45e3
253.1
212.1
%
269.1
144.9
175.1 180.0
166.0
187.2
150.0
199.1
251.1
213.1
211.0
222.0
229.1232.1
239.1
254.2
267.1
282.1
261.1
285.1
292.1
294.2 305.1
0
m/z
150
September 2006
160
170
180
190
200
210
220
230
240
PQRI Training Course
250
260
270
280
290
300
35
Irganox 1010
T072303003
100
Scan AP+
1199.5
1.27e4
17.41
m/z 1199.5
%
17.09
18.00
0
T072303003
100
280
An1
1.33e5
17.36
UV
%
0
T072303003
100
16.96
16.16
17.28
17.57
17.79
17.95
Scan AP+
TIC
1.33e6
18.03 18.21
TIC
%
0
Time
15.60
15.80
September 2006
16.00
16.20
16.40
16.60
16.80
17.00
17.20
PQRI Training Course
17.40
17.60
17.80
18.00
18.20
18.40
36
APCI+ Mass Spectrum Irganox 1010
T072303003 648 (17.306) Cm (648:655-(638:646+661:670))
100
[M+Na]+
1199.5
Scan AP+
5.92e3
1215.5
[M+K]+
1216.5
O
HO
(CH2)2
O CH2
C
%
1217.5
4
1194.5
0
973.5
945.1 963.1
1001.5 1007.3
940
960
September 2006
980
1044.2 1062.3
1089.8
1129.8 1143.5 1152.2
1193.5
1218.5
1219.5
1259.3 1284.6
1261.1
1291.4
m/z
1000 1020 1040 1060 1080 1100 1120 1140 1160 1180 1200 1220 1240 1260 1280
PQRI Training Course
37
APCI- Mass Spectrum Irganox 1010
(no induced fragmentation)
T072303005 651 (17.394) Cm (648:655-(637:646+658:669))
100
[M-H]-
1175.5
Scan AP2.02e4
%
1176.7
1177.6
112.9
0
100
175.0 212.9 233.1
397.2
277.2 367.3
200
September 2006
300
400
521.5
500
584.8
607.5
600
969.7
700
800
PQRI Training Course
900
1175.0
992.5
1000
1178.7
1180.8 1289.9
m/z
1100
1200
38
APCI- Mass Spectrum Irganox 1010
(induced fragmentation in the ion source)
T072303004 651 (17.401) Cm (650:653-(643:649+657:663))
205.1
100
Scan AP1.80e3
739.4
957.5
[M-H]-
521.3
%
958.5
740.4
522.3
206.2
126.9191.1
0
100
200
September 2006
1175.5
259.2 303.2
479.2
389.2
304.2
408.7
300
400
500
1177.5
697.4
533.2 607.4
600
959.5
741.4
769.7
625.1
700
800
PQRI Training Course
915.4
900
1178.6
969.4
901.0
1102.8
1000
1100
1263.4
m/z
1200
39
Structure and Fragmentation of Irganox
1010 in Negative Ion APCI
O
HO
O CH2
(CH2)2
C
4
O
HO
(CH2)2
O
O CH2
C
CH2
O
HO
-
-
CH2
3
m/z 957
September 2006
PQRI Training Course
m/z 205
40
APCI- Mass Spectrum of Stearic Acid
T072303005 455 (12.161) Cm (452:461-(415:443+498:530))
100
283.2
Scan AP1.73e5
[M-H]-
%
284.2
103.0 117.0 129.0
0
100
120
September 2006
140
157.0
160
173.0 185.0 197.1
180
200
219.1
239.1
220
240
255.2
260
PQRI Training Course
281.2 285.3
280
309.2
300
325.3 329.0 348.9
320
340
369.3 373.7
m/z
360
380
41
Triple Quadrupole Mass Spectrometer
September 2006
PQRI Training Course
42
MS/MS Spectrum of Irganox 1076
(ESI+; products of m/z 531)
T081903005 1 (0.502) Sm (Mn, 2x0.75)
100
475.4
531.5
[M+H-56]+
Daughters of 531ES+
1.31e7
[M+H]+
%
[M+H-2X56]+
419.3
0
200
220
240
September 2006
260
280
300
320
340
360
380
400
420
PQRI Training Course
440
460
480
500
520
540
560
580
m/z
600
43
Structure and Fragmentation of Irganox
1076 in Positive Ion APCI (MS/MS)
m/z 419
O
HO
(CH2)2
+H+
O C18H37
m/z 475
September 2006
PQRI Training Course
44
MS/MS Spectrum of Tetramethylthiuram
Disulfide
(ESI+; products of m/z 241)
T081903002 7 (0.153) Sm (Mn, 2x0.75); Cm (4:18)
88
100
Daughters of 241ES+
3.13e5
120
[M+H]+
241
%
0
m/z
40
September 2006
60
80
100
120
140
160
PQRI Training Course
180
200
220
240
260
45
Structure and Fragmentation of
Tetramethylthiuram Disulfide in Positive Ion
APCI (MS/MS)
+H+
S+
S
N
S
m/z 241
S
N
S S
m/z 196
+
m/z 120
S +
N+
N
m/z 88
September 2006
PQRI Training Course
m/z 44
46
September 2006
PQRI Training Course
47
ESI+ TOF Mass Spectrum of TMTMS
[M+Na]+
[2M+Na]+
[M+H]+
September 2006
PQRI Training Course
48
Expanded ESI+ TOF Mass Spectrum of TMTMS
+TOF MS: 0.220 to 0.293 min from tmms01.wiff Agilent
Max. 1.3e6 counts.
231.0051
1.3e6
1.2e6
1.1e6
[M+Na]+
I n t e n s it y , c o u n t s
1.0e6
9.0e5
8.0e5
7.0e5
6.0e5
5.0e5
4.0e5
3.0e5
233.0006
2.0e5
232.0065
231.2400
1.0e5
0.0
230.5
September 2006
231.0
231.5
232.0
232.5
m/z, amu
PQRI Training Course
233.0
233.5
234.0
49
Elemental Composition Report for TMTMS
from ESI+ TOF-MS
Single Mass Analysis - displaying only valid results
Tolerance = 10.0 PPM / DBE: min = -0.5, max = 20.0
Monoisotopic Mass, Odd and Even Electron Ions
43463 formula(e) evaluated with 3 results within limits (all results (up to 1000) for each
mass)
Minimum:
-0.5
Maximum:
2.0
10.0
20.0
Mass
Calc. Mass
mDa
PPM
DBE
Formula
231.0052 231.0052
0.0
0.2
8.0
12C4 1H2 14N9 23Na 32S
231.0051
0.1
0.6
9.5
12C11 1H7 14N2 32S2
231.0060
-0.8
-3.6
1.5
12C6 1H12 14N2 23Na 32S3
September 2006
PQRI Training Course
50
Expanded ESI+ TOF Mass Spectrum of Irganox
1010
+TOF MS: 7.699 to 7.976 min from Irgonax 1010 2.wiff Agilent
Max. 2.5e5 counts.
1199.7727
2.5e5
[M+Na]+
2.4e5
2.2e5
1200.7765
2.0e5
In te n s ity , c o u n ts
1.8e5
1.6e5
[M+NH4]+
1.4e5
1194.8173
1.2e5
1195.8213
1.0e5
1201.7807
8.0e4
6.0e4
1196.8258
4.0e4
1202.7833
2.0e4
0.0
1197.8268
1195.3564
1192
September 2006
1194
1196
1198
1200.3141
1200
m/z, amu
PQRI Training Course
1201.3150
1202
1203.7858
1204
1206
1208
51
Elemental Composition Report for Irganox
1010 from ESI+ TOF-MS
Single Mass Analysis
Tolerance = 5.0 PPM / DBE: min = -0.5, max = 40.0
Monoisotopic Mass, Odd and Even Electron Ions
289500 formula(e) evaluated with 3 results within limits (all results (up to 1000) for
each mass)
Minimum:
-0.5
Maximum:
2.0
5.0
40.0
Mass
Calc. Mass
mDa
PPM
DBE
Formula
1199.7727 1199.7738
-1.1
-1.0
19.5
12C73 1H108 16O12 23Na
1199.7763
-3.6
-3.0
22.5
12C75 1H107 16O12
1199.7680
4.7
3.9
28.5
12C80 1H104 16O7 23Na
September 2006
PQRI Training Course
52
My Philosophy with LC/MS
► For
CMC impurity profiling studies use APCI
as a primary technique.
► Reproduce the UV chromatogram.
► Use every trick in the book to make APCI
work (mobile phase additives, alternate
acquisition techniques, etc.).
► Use electrospray to complement and
confirm APCI.
September 2006
PQRI Training Course
53
IPAC-RS Proposed Identification Categories
Table 2. Identification Categories for Structure Elucidation of Extractables
and Leachables by GC/MS and LC/MS
Category
September 2006
Identification Data
A
Mass spectrometric fragmentation behavior
B
Confirmation of molecular weight
C
Confirmation of elemental composition
D
Mass spectrum matches automated library or literature
spectrum
E
Mass spectrum and chromatographic retention index
match authentic specimen
PQRI Training Course
54
Identification Categories
►A
Confirmed identification means that
identification categories A, B (or C), and D (or E)
have been fulfilled.
► A Confident identification means that sufficient
data to preclude all but the most closely related
structures have been obtained.
► A Tentative identification means that data have
been obtained that are consistent with a class of
molecule only.
September 2006
PQRI Training Course
55