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The Use of Gas Chromatography Mass
Spectrometry in the Evaluation of the Synthesis
of 3-dimethy laminomethyl-2 (3',4'dichlorophenyl)bicyclo[2.2.2.1octane
-
A Novel Drug with Potential Antidepressant Activity
David N. B. Mallen* and Julie M. Smith
Lilly Research Centre Limited, Erl Wood Manor, Windlesham, Surrey GU20 6PH, UK
Gas chromatography mass spectrometry was used to characterize compounds observed throughout the synthesis
of a cis-fused 3',4'-dichlorophenyl bicyclo-octane,a potential CNS acting drug substance. An ortho chloro-effect
identified the isomeric 2',4'-dichlorophenyl impurities.
INTRODUCTION
~~
3-Dimethylaminomethy1-2-(3',4'-dichlorophenyl)bicyclo[2.2.2.]octane (1) was one of a series of cis-fused
phenylvyclo[2. 2.2 .]octanes which showed CNS
activity on initial pharmacological studies. The stereospecific five-stage synthesis of 1, developed from the
amalgamation of two alternative
required improvement prior to the production of further
material.
During the further development of the synthesis, gas
chromatography mass spectrometry (GC/MS) was
used to characterize all impurities, especially isomeric
by-products, prior to their quantification by gas chromatography for yield improvement purposes. It was
particularly desirable to minimize the level of the
trans-isomer in 1. Hence, all raw materials, intermediates and final product material were subject to a
careful quality control.
A specific impurity observed at each synthetic stage
was the corresponding 2,4-dichloro-isomer which originated from impure starting material. In this publication
the characteristic electron impact mass spectra of 1 and
its synthetic precursors are discussed with reference to
the isomeric 2,4-dichloro-analogues.
hexanol (200 pl). The corresponding esters (7a-7f, 8a8f and 9a-9f) of the Diels-Alder adduct isomers were.
prepared by the addition of the appropriate alcoholl
(200 p1) and pyridine (1 drop) to the reaction mixture:
(20 pl) containing 7, 8 and 9. The resultant solutions
were heated at 80 "C for 30 min and analysed directly
by GC/MS.
Gas chromatography mass spectrometry
Gas chromatographic mass spectrometric analyses were
carried out using an LKB 9000s mass spectrometer
linked to a VG 2040L data system. The cinnamate ester
samples were introduced by gas chromatography on .a
2 m x 4 mm i.d. glass column packed with 2% butane1,4-diol succinate on Gas Chrom Q (100/120 mesh) alt
180°C. Products from all other synthetic stages were
analysed on a 2 m x 4 mm i.d. 2% Apiezon J column sit
210 "C or 230 "C. Helium, at a flow rate of 30 ml min-',
was used as carrier gas. Mass spectra were recorded
using an ion accelerating voltage of 3.5 kV, an electron
voltage of 20 eV and source and separator temperatures
of 270 "C.
RESULTS AND DISCUSSION
EXPERIMENTAL
Preparation of esters of cinnamic acid and
Diels-Alder adduct
Compounds 5a and 6a were prepared by reaction of 3
or 4 (1-10 mg), respectively, with excess ethereal
diazomethane solution. After 10 min the resultant solution was blown to dryness. An aliquot of the acid
chloride reaction solution (20 pl) containing 5 and 6
was evaporated to dryness under nitrogen. Compounds
5a-5f and 6a-6f were prepared by adding methanol,
ethanol, n-propanol, n -butanol, n-pentanol or n-
* Author to whom correspondence should be addressed.
-
The five-stage synthesis of 1 is shown in Scheme 1. The
need to characterize appropriate chromatographic
peaks and estimate levels of the 2,4-dichloro-isomer at
each synthetic stage became apparent when it was
observed that trans- 3,4-dichlorocinnamic acid (3) ra.w
material often contained significant levels of trans- 2,4dichlorocinnamic acid (4). Minimal levels of the cis- 3,4isomer were observed. Conversion of 3 to the acid
chloride (5) and reaction with 1,3-cyclohexadiene gave
the corresponding Diels-Alder adduct (7). Two major
isomeric peaks were observed, which were identified as
the trans-2,4-isomer (8) and the cis-3,4-isomer (9).
The presence of 9 in 7, which was at a greater level
than that of the corresponding isomeric impurity in balth
CCC-0306-042X/82/0009-0125$02.50
@ Heyden & Son Ltd, 1982
BIOMEDICAL MASS SPECTROMETRY, VOL. 9, NO. 3, 1982 1125
D. N. B. MALLEN AND J. M. SMITH
c1
H
I
3 3-chloro
1
9
4 2-chloro
5 3-chloro
I
6 2-chloro
p.
11
10 2-chloro
3-chloro
~
/
.
/'.CHO
~~1
CH,NMe,
14
7 3-chloro
8 2-chloro
Coc1
''.\I/-----
12 3-chloro
13 2-chloro
CHNMez
'-H
CH,NMe2
H
1 3-chloro
2 2-chloro
Scheme 1. The synthesis of cis-fused 3-dimethylaminomethyl-2-(3',4'-dichlorophenyl)bicyclo[2.2.2.]octane.
3 and 5, can only be explained by isomerism or by
adverse fractionation of sample during the Diels-Alder
reaction. In the subsequent stages, the corresponding
2,4-dichloro-isomers were still observed. The purity of
final product material was also adversely affected by
formation of the cis-3,4-dichloro-isomer (14)during
reduction of the enamine (12).
The cinnamic acids (3 and 4) were methylated using
diazomethane prior to gas chromatography. Using
GC/MS the methyl ester of 4 could be readily distinguished from that of 3 by a significant loss of chlorine.
The acid chlorides (5 and 6) formed in the next stage
can be readily converted into a range of esters by simple
addition of the appropriate alcohol to a reaction
mixture. Hence it was possible to evaluate this
orfho-chlorine effect in a range of esters. The relative
abundance of the nrincipal ions in the mass spectra of
the methyl, ethyl, n-propyl, n-butyl, n-pentyl and nhexyl esters of 5 (5a-5f) and 6 (6a-6f), respectively, are
shown in Table 1.
Scheme 2 indicates the expected two parallel fragmentation pathways for 6a-6f. Loss of the ortho
chloro-atom (route A) is common to all the esters with
formation of a coumarin ion structme ( a ) . A similar
effect hqs been reported for 2-chlorocinnamic acid.3
Although for the methyl ester (6a), a is the base ion,
the ortho chloro-effect is minimized with increase in
length of the ester carbon chain. Ion a eliminates the
corresponding alkene by a McLaff erty rearrangement
to form ion b of m / z 181/183. The alternative fragmentation route B, involving simple scission of the ester
side-chain to form an acylium ion c is of less significance.
Table 1. Key ions in the electron impact mass spectra (20 eV) of the esters of truns-3,4-dichlorocinnamicacid and truns-2,4dichlorocinnamic acid
Methyl
Ion
rnlz
[MI"
a [M-CI]+
d [M-CnHznI'
c lM-OCnHzn+ilt
b [M-CI-C,HZ,I'
e [M-OC,H,,+, -COl+
21612181220
19912011203
1811183
17111731175
6a
5b
6b
5c
6r
96
5
<1
100
<1
37
40
100
<I
37
88
7
46
100
10
21
22
85
<1
46
45
<1
100
66
6
5
11
34
<1
29
100
5
126 BIOMEDICAL MASS SPECTROMETRY, VOL. 9, NO. 3, 1982
<I
34
n-Pentyl
5e
6e
n-fropyl
Ethyl
5s
100
16
42
<I
100
55
6
6
6
20
<1
16
100
3
24
<1
100
42
<1
<1
5
16
il
10
100
2
n-nexvt
5f
6f
17
<I
100
28
<I
<1
3
17
<I
11
100
<l
@ Heyden & Son Ltd, 1982
Route A*
I
Route C
CI
OC,Hzn+l
a II
I
m / z 199/201/203
C12
I
b
1
e
m l z 1811183
mlz 111/113/115
Scheme 2. Fragmentation of dichlorocinnamates.
Its abundance again decreases with increase in length
of the ester carbon chain. Ion c can eliminate [CO] to
form the styryl ion e.
In contrast to 6a-6f, 5a-5f all show minimal loss of
chlorine and increased fragmentation by route B. For
5a (methyl ester) and 5b (ethyl ester) ion c is the most
intense, but for 5c-5d, loss of the corresponding alkene
from the molecular ion to form ion d becomes dominant
(route C).
Following the Diels-Alder reaction, in addition to
the formation of the two trans-isomers (7 and S), a third
isomer, the cis- 3,4-dichloro analogue (9) was also observed. As in the previous stage, addition of the
appropriate n-alcohols gave three series of esters (7a-7f
for the trans-3,4-dichloro-, 8a-8f for the truns-2,4dichloro- and 9a-9f for the cis- 3,4- dichloro-isomers).
The fragmentation of all these types of molecules
(Scheme 3) was dominated by a retro Diels-Alder rearrangement. In consequence, the differential behaviour
of the 2,4-dichloro- and 3,4-dichloro-isomers was less
apparent. Although the loss of the chlorine atom to
give ion f was only a minor fragmentation pathway, it
still retained a characteristic presence for the 2,4dichloro-isomers. The loss of the alkyl moiety was now
only apparent for the butyl, pentyl and hexyl esters,
compounds 9d, 9e and 9f showing the highest abundance of the resultant ion g.
The loss of an alkoxy group to form acylium ion h
was similarly diminished. An important fragmentation
not observed for the dichloro-cinnamates was elimination of the ester group with formation of the
phenylbicyclo-octadiene ion j . cis- Stereochemistry and
3,4-dichloro-substitutionfavoured this elimination. The
relative abundance of fragment ions k, 1 and m indicated
that the dominant retro Diels-Alder rearrangement was
favoured by trans-stereochemistry.
Once the bicyclo-octene ring has been reduced to
the corresponding bicyclo-octane, as in the aldehyde
(lo),the enamine (12)and the dimethylamine product
@ Heyden & Son Ltd, 1982
(l),
the retro Diels-Alder rearrangement is'minimized
and the ortho- chloro elimination effect is more clearly
observed.
Both 10 and the 2,4-dichloro-aldehyde (11)show a
significant molecular ion (100 o/' for 10) and primary
fragments corresponding to the loss of ethylene and the
aldehydic moiety (Fig. 1).The intensity of the [M - C1]+
ion is, however, characteristically different (100 '/o for
100
t
P
282
(a)
100
200
300
100
47
+
0
[L
a'
2 82
80
I
60
201
40
159
I
1
20
0
100
200
300
m /I
Figure 1. Mass spectra of (a) 3-formyl-2-(3',4'-dichlorophenyl)bicyclo[2.2.2.]octane and (b) 3-formyl-2-(2',4'-dichlorophenyl)bicyclo[2.2.2.]octane.
BIOMEDICAL MASS SPECTROMETRY, VOL. 9,
NO. 3, 1982 127
D. N. B. MALLEN AND J. M. SMITH
\
h
+
CEO
m/z279/281/283
'co;
g
m / z 29512971299
L
I
Scheme 3. Fragmentation of 3-carboxylate-2-(dichlorophenyl)bicyclo[2.2.2loctanes.
11 but 21 '/o for 10). The enamines show even less
tendency to fragment. For both the 3,4-dichloroenamine (12) and the 2,4-dichloro-enamine (13)the
molecular ion is the base ion. Compound 12 loses the
dialkylamine group whereas 13 is characterized by loss
of chlorine. Compound 1 and the 2,4-dichloro-isomer
(2)both show an even simpler spectrum, characterized
by the molecular ions ( m / z 311/313/315) and the m / z
58 ion [CH,=N+(CH&]. The 2,4-dichloro-isomer
shows an additional [M - Cl]' ion of similar abundance
to that of the molecular ion. This characteristic mass
spectral behaviour of 11, 13 and 2 is probably due to
the ease of formation of the cyclic fused ions n, p and
q respectively, analogous to that of ion a for the 2,4dichlorocinnamates. As compounds 1 and 14 were not
easily resolved by packed gas chromatography column,
the impurity level of 14 in 1 was determined after
conversion of both isomers to their respective carbamates.4 As expected, no significant loss of chlorine is
seen from either of these carbamate derivatives. Hence,
a characteristic mass spectral difference is observed for
all the corresponding 2,4-dichloro- and 3,4-dichloro128 BIOMEDICAL MASS SPECTROMETRY, VOL. 9,
NO.3,1982
4
\
synthetic intermediates. Identification of gas chromatograph peaks of the appropriate chloro-isomers was
achieved without the need to collect mg quantities
for further spectroscopic characterization. The 2,4dichloro-isomer always eluted before the corresponding
3,4-dichloro-compound.
@ Heyden & Son Ltd, 1982
REFERENCES
1. C. H. Cashin, J. Fairhurst, D. C. Howell, I. A. Pullar, S.Sutton,
G. H. Tirnms, E. Wildsmith and F. Wright. Eur. J. Med. Chem.
Chirn. Ther. 13,495 (1978).
2. D. C. Howell and G. H. Tirnrns, Synth. Commun. 9,223 (1979).
3. 6. Schaldach and H-Fr. Grutzrnacher, Org. Mass. Specrrorn.
15, 175 (1980).
@ Heyden & Son Ltd, 1982
4. R. C. Harden, D. N. B. Mallen, D. J. Osborne and J. M. Smith,
J. Chrornatogr. 206, 83 (1981).
Received 14 September 1981
@ Heyden & Son Ltd, 1982
BIOMEDICAL MASS SPECTROMETRY, VOL. 9, NO. 3, 1982 129
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