Formation of the bisulfite anion (HSO3 -, m/z 81) upon collision-induced dissociation of anions derived from organic sulfonic acids

Article abstract of DOI:10.1002/jms.2975

In the negative-ion collision-induced dissociation mass spectra of most organic sulfonates, the base peak is observed at m/z 80 for the sulfur trioxide radical anion (SO₃ ^-·). In contrast, the product-ion spectra of a few sulfonates, such as cysteic acid, aminomethanesulfonate, and 2-phenylethanesulfonate, show the base peak at m/z 81 for the bisulfite anion (HSO₃ ^- ). An investigation with an extensive variety of sulfonates revealed that the presence of a hydrogen atom at the β-position relative to the sulfur atom is a prerequisite for the formation of the bisulfite anion. The formation of HSO₃ ^- is highly favored when the atom at the β-position is nitrogen, or the leaving neutral species is a highly conjugated molecule such as styrene or acrylic acid. Deuterium-exchange experiments with aminomethanesulfonate demonstrated that the hydrogen for HSO₃ - formation is transferred from the β-position. The presence of a peak at m/z 80 in the spectrum of 2-sulfoacetic acid, in contrast to a peak at m/z 81 in that of 3-sulfopropanoic acid, corroborated the proposed hydrogen transfer mechanism. For diacidic compounds, such as 4-sulfobutanoic acid and cysteic acid, the m/z 81 ion can be formed by an alternative mechanism, in which the negative charge of the carboxylate moiety attacks the α-carbon relative to the sulfur atom. Experiments conducted with deuterium-exchanged and deuterium-labeled analogs of sulfocarboxylic acids demonstrated that the formation of the bisulfite anion resulted either from a hydrogen transfer from the β-carbon, or from a direct attack by the carboxylate moiety on the α-carbon. Copyright

Full text of DOI:10.1002/jms.2975

Research Article
Received: 3 December 2011
Revised: 31 January 2012
Accepted: 11 February 2012
Published online in Wiley Online Library
(wileyonlinelibrary.com) DOI 10.1002/jms.2975
–
Formation of the bisulfite anion (HSO , m/z 81)
3
upon collision-induced dissociation of anions
derived from organic sulfonic acids
Freneil B. Jariwala, Ryan E. Wood, Upul Nishshanka and Athula B. Attygalle*
In the negative-ion collision-induced dissociation mass spectra of most organic sulfonates, the base peak is observed at m/z 80
–
3
ꢀ
for the sulfur trioxide radical anion (SO ). In contrast, the product-ion spectra of a few sulfonates, such as cysteic acid,
–
aminomethanesulfonate, and 2-phenylethanesulfonate, show the base peak at m/z 81 for the bisulfite anion (HSO
3
). An
investigation with an extensive variety of sulfonates revealed that the presence of a hydrogen atom at the b-position relative
–
3
to the sulfur atom is a prerequisite for the formation of the bisulfite anion. The formation of HSO is highly favored when
the atom at the b-position is nitrogen, or the leaving neutral species is a highly conjugated molecule such as styrene or
acrylic acid. Deuterium-exchange experiments with aminomethanesulfonate demonstrated that the hydrogen for
–
3
HSO
formation is transferred from the b-position. The presence of a peak at m/z 80 in the spectrum of 2-sulfoacetic acid,
in contrast to a peak at m/z 81 in that of 3-sulfopropanoic acid, corroborated the proposed hydrogen transfer mechanism.
For diacidic compounds, such as 4-sulfobutanoic acid and cysteic acid, the m/z 81 ion can be formed by an alternative
mechanism, in which the negative charge of the carboxylate moiety attacks the a-carbon relative to the sulfur atom.
Experiments conducted with deuterium-exchanged and deuterium-labeled analogs of sulfocarboxylic acids demonstrated
that the formation of the bisulfite anion resulted either from a hydrogen transfer from the b-carbon, or from a direct attack
by the carboxylate moiety on the a-carbon. Copyright © 2012 John Wiley & Sons, Ltd.
Keywords: anions; sulfonates; fragmentation; CID; gaseous ions; bisulfite
–
3
[20,28]
(HSO
, m/z 81).
For example, in the collision-induced dissocia-
INTRODUCTION
tion (CID) mass spectra of fluorotelomer sulfonates, the base peak is
observed at m/z 81 indicating the formation of the bisulfite anion.
[28]
Sulfonic acids and their derivatives play an important role in
pharmaceutical and industrial chemical manufacturing. For
example, aliphatic and aromatic sulfonates are frequently utilized
as anionic surfactants in detergents.
aliphatic sulfonates are also widely used in the chemical industry
[
1]
In addition, we observed that the base peak in the CID mass
spectrum of cysteic acid is at m/z 81. However, in a previous pub-
lication, the formation of an ion of m/z 80 had been attributed as
[2–6]
Substituted aromatic and
[
25,26]
the major fragmentation pathway of cysteic acid.
In this
[
7–10]
for the production of pharmaceuticals and dyes.
Conjugate
paper, we report the results about our investigations on the
fragmentation mechanisms of organic sulfonates.
bases of sulfonic acids, such as benzenesulfonate (besylate),
methanesulfonate (mesylate), ethanesulfonate (esylate), and
camphorsulfonate (camsylate), provide suitable anionic counter-
ions, which are sometimes necessary for salt formation of certain
pharmacologically active ingredients (APIs).
sulfonic acid esters are genotoxic; at least one drug (nelfinavir
EXPERIMENTAL
[
11,12]
However, many
Materials
mesylate) has been withdrawn from the market because of such
Methanol [high-performance liquid chromatography (HPLC)
grade], acetonitrile (HPLC grade), hydrochloric acid (ACS grade),
formic acid (88%, ACS grade), and ammonium hydroxide (ACS
grade) were purchased from Pharmco-AAPER (Brookefield, CT,
USA). Methanesulfonic acid (1), ethanesulfonic acid (2), benzenesul-
fonic acid (5), p-toluenesulfonic acid (6), p-aminobenzenesulfonic
[13–16]
complications.
In organic chemistry, low-molecular weight
sulfonic acids, such as methanesulfonic acid, trifluoromethanesulfo-
nic acid, and p-toluenesulfonic acid, are often used as acid catalysts
[
17–19]
for esterification, alkylation, and condensation reactions.
The anionic nature of sulfonates renders them ideal candidates
for analysis by negative-ion electrospray ionization mass spec-
trometry. Although studies have been carried out on fragmenta-
tion pathways of simple sulfonates, the intricate details of their
* Correspondence to: Athula B. Attygalle, Center for Mass Spectrometry, Department
of Chemistry, Chemical Biology, and Biomedical Engineering, Stevens Institute of
Technology, Hoboken, NJ 07030, USA. E-mail: aattygal@stevens.edu
[
20–28]
fragmentation mechanisms are not very well understood.
Negative-ion mass spectra of alkyl and aryl sulfonates indicate that
the major product ion produced by collision-induced fragmenta-
Center for Mass Spectrometry, Department of Chemistry, Chemical Biology,
and Biomedical Engineering, Stevens Institute of Technology, Hoboken, NJ,
07030, USA
–
3
ꢀ
[20–28]
tion is the sulfur trioxide radical anion (SO , m/z 80).
On the
other hand, some sulfonates are known to give the bisulfite anion
J. Mass. Spectrom. 2012, 47, 529–538
Copyright © 2012 John Wiley & Sons, Ltd.

F. B. Jariwala et al.
acid (7), p-bromobenzenesulfonic acid (8), sulfamic acid (12), ami-
nomethanesulfonic acid (13), 2-aminoethanesulfonic acid (taurine,
14), 3-aminopropanesulfonic acid (15), L-homocysteic acid (23),
1-bromopentane, 1-bromoheptane, benzyl bromide, 2-phenylethyl
bromide, mesitylene, bromoacetic acid, 3-bromopropanoic acid, 4-
bromobutanoic acid, 6-bromohexanoic acid, L-cysteine, L-cysteine
2
ethyl ester, and [ H
4
]ammonium deuteroxide were purchased from
Sigma-Aldrich Co. (Milwaukee, WI, USA). Anhydrous sodium sulfite,
fuming sulfuric acid (20% SO ), and hydrogen peroxide (30%) were
purchased from Fisher Chemical Co. (Fairlawn, NJ, USA). L-[3,3- H
Cysteine was purchased from Cambridge Isotope Laboratories, Inc.
Andover, MA, USA). The silica gel (particle size: 32–63 mm) used for
flash chromatography was obtained from Selecto Scientific Inc.
Suwanee, GA, USA).
3
2
2
]
(
(
Figure 2. Product-ion spectra of anions derived from benzenesulfonic
acid (m/z 157) (A), p-toluenesulfonic acid (m/z 171) (B), 2-mesitylenesulfonic
acid (m/z 199) (C), p-aminobenzenesulfonic acid (m/z 172) (D), p-bromoben-
zenesulfonic acid (m/z 235) (E), and an overlay of the m/z 81 peak from the
five aforementioned compounds (F) recorded on an API-3000 tandem mass
spectrometer at laboratory-frame collision energy of 25 eV.
Scheme 1. Fragmentation mechanism for the formation of the sulfur
–
3
ꢀ
).
trioxide radical anion (m/z 80, SO
Scheme 2. Proposed mechanism for the formation of the bisulfite anion
–
(
m/z 81, HSO
3
) upon fragmentation of deprotonated alkyl sulfonic acids
2
–4.
Synthesis of alkyl sulfonic acids
Figure 1. Product-ion spectra of anions derived from methanesulfonic acid
Pentanesulfonic acid (3), heptanesulfonic acid (4), phenylmethane-
sulfonic acid (10), and 2-phenylethanesulfonic acid (11) were
synthesized from 1-bromopentane, 1-bromoheptane, benzyl
(
(
m/z 95) (A), ethanesulfonic acid (m/z 109) (B), pentanesulfonic acid (m/z 151)
C), and heptanesulfonic acid (m/z 179) (D) recorded on an API-3000 tandem
mass spectrometer at laboratory-frame collision energy of 25 eV.
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J. Mass. Spectrom. 2012, 47, 529–538

Dissociation of organic sulfonates
Scheme 4. Proposed mechanism for the formation of the bisulfite anion
–
–
(
3
m/z 81, HSO ) upon fragmentation of 2-phenylethanesulfonate ([11 – H] ).
Synthesis of sulfocarboxylic acids
-Sulfoacetic acid (16), 3-sulfopropanoic acid (17), 4-sulfobutanoic
Scheme 3. Proposed mechanism for the formation of the bisulfite anion
–
(
m/z 81, HSO
3
) upon fragmentation of deprotonated aromatic sulfonic
2
acids 5–8.
acid (18), and 6-sulfohexanoic acid (19) were also synthesized
[
29,30]
by the Strecker sulfite alkylation.
Bromoacetic acid (50 mg,
0
.360 mmol), 3-bromopropanoic acid (50 mg, 0.327 mmol), 4-
bromide, and 2-phenylethyl bromide, respectively, by Strecker
bromobutanoic acid (50 mg, 0.299 mmol), or 6-bromohexanoic
[29,30]
sulfite alkylation.
1-Bromopentane (50 mg, 0.331 mmol),
acid (50 mg, 0.256 mmol), was refluxed with an aqueous solution
ꢁ
1
0
-bromoheptane (50 mg, 0.279 mmol), benzyl bromide (50 mg,
.292 mmol), or 2-phenylethyl bromide (50 mg, 0.270 mmol), was
of sodium sulfite (1.0 M; 10 ml) at 110 C for 12 h. The reaction
mixture was then acidified with conc. HCl to a pH of 1.0, and the
resulting sulfocarboxylic acids were purified by flash chromatogra-
phy (32–63 mm of silica gel) using methanol as the eluting solvent.
refluxed with an aqueous solution of sodium sulfite (1.0M; 10 ml)
ꢁ
at 110 C for 12h. The reaction mixture was then acidified with
conc. HCl to a pH of 1.0, and the resulting sulfonic acids were
purified by flash chromatography (32–63 mm of silica gel) using
methanol as the eluting solvent. The sulfonic acids in methanol
The methanolic extracts were dried under a stream N
compounds obtained were crystalline in nature.
2
, and the
were dried under a stream N and were crystalline in nature.
2
Oxidation of cysteine and its analogs to cysteic acid
2
L-Cysteine, L-[3,3- H
2
]cysteine, and L-cysteine ethyl ester were
Synthesis of 2-mesitylenesulfonic acid
2
oxidized by performic acid to L-cysteic acid (20), L-[3,3- H
2
]
2
-Mesitylenesulfonic acid (9) was synthesized from mesitylene
cysteic acid (21), and L-cysteic acid ethyl ester (22), respec-
[
31]
[32]
by aromatic sulfonation.
Mesitylene (50 mg, 0.417 mmol)
tively.
The performic acid reagent was prepared by admixing
was vigorously shaken with conc. fuming sulfuric acid
0.5 ml) until the organic layer had completely disappeared
H O (30%, 10 ml) and formic acid (88%, 90 ml). The solution was
kept at room temperature for 30 min to maximize performic acid
2 2
(
and a yellowish solution resulted. This solution was then
poured into 10 ml of ice water. After an hour at room temper-
ature, vacuum filtration was used to collect the precipitated 2-
mesitylenesulfonic acid crystals. The white crystalline product
formation. L-Cysteine (50 mg, 0.413 mmol), L-cysteine ethyl ester
2
(50 mg, 0.336 mmol), or L-[3,3- H ]cysteine (5 mg, 0.041 mmol),
2
was dissolved in the performic acid solution, and the mixture
was kept at room temperature for 30 min. The reaction mixture
ꢁ
W
was washed with 10 ml of cold water (~5 C) and 10 ml of cold
was dried in an Eppendorf Vacufuge concentrator (Eppendorf,
ꢁ
ꢁ
methanol (~5 C). The resulting product was air-dried and used as
Hauppauge, NY, USA) at 60 C for 2 h, and crystalline cysteic acids
obtained.
were recovered.
Figure 3. Product-ion spectra of anions derived from phenylmethanesulfonic acid (m/z 171) (A), and 2-phenylethanesulfonic acid (m/z 185) (B)
recorded on an API-3000 tandem mass spectrometer at laboratory-frame collision energy of 25 eV.
J. Mass. Spectrom. 2012, 47, 529–538
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F. B. Jariwala et al.
Mass spectrometry
The negative-ion CID mass spectra were acquired on an Applied
Biosystems API 3000 (Concord, ON, Canada) triple quadrupole
W
mass spectrometer equipped with a TurboIonSpray source.
ꢁ
The source temperature was held at 50 C. Both the nebulizer
gas (N ) and curtain gas (N ) flow rate settings were at 15 (arbi-
2
2
trary units). The drying gas (N ) was maintained at a flow rate
2
of 1 l/min at 60 psi. The ionspray voltage was held at ꢂ4500 V,
and a declustering potential of ꢂ50 V was used. The collision
2
gas (N ) pressure in the collision cell was maintained at
–
5
2
.1 ꢃ 10 Torr, and the collision energy was varied between 20
and 30 eV. Each of the compounds 1–23 was diluted to a concen-
tration of 1 mg/ml in a 1 : 1 mixture of acetonitrile-water with 1%
v/v ammonia. For deuterium-exchanged experiments, samples
were dissolved in a 1 : 1 mixture of acetonitrile-deuterium oxide
with 1% v/v deuteriated ammonia. Samples were introduced into
the source by infusion at a flow rate of 10 ml/min.
RESULTS AND DISCUSSION
Aliphatic and aromatic sulfonates are ideal candidates for
negative-ion electrospray ionization mass spectrometry be-
cause of their acidic nature and the ability to readily form
the corresponding conjugate bases in solution. Thus, anions
derived from sulfonic acids have been widely used for their
[
28,33–38]
qualitative and quantitative determinations.
The prod-
uct-ion mass spectra of anions derived from aliphatic and ar-
omatic sulfonic acids (1–9) show an intense signal at m/z 80,
–
3
ꢀ
representing the sulfur trioxide radical anion (SO ) resulting
from a homolytic cleavage of the precursor (Figs 1 and 2,
Scheme 1). However, the mass spectra of aliphatic sulfonates
with alkyl chains containing two or more carbons also show
–
3
a minor peak at m/z 81 for the bisulfite anion (HSO
) (Fig. 1
Figure 4. Product-ion spectra of anions derived from sulfamic acid (m/z 96)
(
(
A), aminomethanesulfonic acid (m/z 110) (B), 2-aminoethanesulfonic acid
m/z 124) (C), 3-aminopropanesulfonic acid (m/z 138) (D), sulfamic acid in
(B,C, and D)). Although the relative intensity of the peak for
the m/z 81 ion is only about 2%, the presence of this peak
is intriguing because these anions do not bear any exchange-
able hydrogen atoms. Because the peak at m/z 81 is only ob-
served in the mass spectra of sulfonates with alkyl chains
containing two or more carbons, we concluded that the frag-
mentation process may be best represented as a cyclic syn-
elimination mechanism (Scheme 2), similar to the Cope amine
D
2
O (m/z 98*) (E), aminomethanesulfonic acid in D
aminoethanesulfonic acid in D O (m/z 126*) (G), and 3-aminopropanesulfonic
acid in D O (m/z 140*) (H) recorded on an API-3000 tandem mass spectrom-
eter at laboratory-frame collision energy of 25 eV. (*Note that isotopologues
2
O (m/z 112*) (F), 2-
2
2
34
bearing either two deuterium atoms or one S atom are isobaric; therefore,
ions isolated at unit-resolution should be composites).
[
39–42]
N-oxide elimination.
In a previous publication, it has been
conclusively demonstrated that the formation of the bisulfate
–
anion (HSO
via a similar mechanism.
In the case of aromatic sulfonates, the neutral loss of an SO
4
, m/z 97) upon CID of organic sulfates also occurs
[
43,44]
2
[
24]
(
Fig. 2(A–E)) has been studied in detail by Binkley et al. Further-
more, the m/z 81 peak is also observed in the mass spectra of
–
aromatic sulfonates, such as benzenesulfonate ([5 – H] , Fig. 2(A)),
–
p-toluenesulfonate ([6 – H] , Fig. 2(B)), p-aminobenzenesulfonate
–
–
(
[7 – H] , Fig. 2(D)), and p-bromobenzenesulfonate ([8 – H] , Fig. 2(E)).
A mechanism, similar to that given in Scheme 2, whereby the
hydrogen atom at the ortho-position is transferred via the cyclic
syn-elimination, to eliminate a benzyne derivative is envisaged for
the formation of the bisulfite anion from aromatic sulfonates
(Scheme 3). This cyclic syn-elimination mechanism was supported
by the observation that the m/z 81 peak was not observed in the
–
mass spectrum of 2-mesitylenesulfonate ([9 – H] , Fig. 2(C)). From
the few examples we have investigated that the properties of
Scheme 5. P–roposed mechanism for the formation of the bisulfite anion
(
m/z 81, HSO
3
) upon fragmentation of deprotonated aminoalkylsulfonic
acids 13–15.
the substituent group at the para position appear to have an effect
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Dissociation of organic sulfonates
Figure 5. Product-ion spectra of anions derived from 2-sulfoacetic acid (m/z 139) (A), 3-sulfopropanoic acid (m/z 153) (B), 4-sulfobutanoic acid (m/z
1
67) (C), 6-sulfohexanoic acid (m/z 195) (D), 2-sulfoacetic acid in D
in D O (m/z 168) (G), and 6-sulfohexanoic acid in D O (m/z 196) (H) recorded on an API-3000 tandem mass spectrometer at laboratory-frame col-
lision energy of 25 eV.
2 2
O (m/z 140) (E), 3-sulfopropanoic acid in D O (m/z 154) (F), 4-sulfobutanoic acid
2
2
on the formation of the bisulfite anion (Fig. 2(F)). Moreover, the
m/z 81 peak was not observed in the mass spectrum of phenyl-
methanesulfonate ([10 – H] , Fig. 3(A)), further validating the
seems that the elimination of a neutral styrene molecule is a
highly favorable pathway, similar to the results described by
Cope et al. for the thermal decomposition of amine N-oxide
–
[
40]
cyclic syn-elimination mechanism, because the b-carbon, rela-
tive to the sulfur atom, does not have any hydrogens. On the
other hand, the m/z 81 peak is the base peak in the spectrum
elimination.
Further support for the cyclic syn-elimination mechanism was
obtained by the CID experiments conducted with aminometha-
–
–
of 2-phenylethanesulfonate ([11 – H] , Fig. 3(B), Scheme 4). It
nesulfonate ([13 – H] , Fig. 4(B), Scheme 5). The spectrum of
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F. B. Jariwala et al.
–
3
Scheme 6. Proposed mechanism for the formation of the bisulfite anion (m/z 81, HSO
acids 17–19.
) upon fragmentation deprotonated sulfocarboxylic
–
aminomethanesulfonate ([13 – H] ) showed the base peak at m/
is driven by the charge on the carboxylate, which attacks the
a-carbon resulting in the elimination of a neutral b-propiolactone
molecule (Scheme 6(B)). Because the intensity of the m/z 81 peak
is so large compared with that of m/z 82 (Fig. 5(F)), it is evident
that the sulfonate charge-mediated pathway (Scheme 6(A)) is
highly favored. The base peak of the spectra of both 3-sulfopropa-
z 81. When the same compound was sprayed as a D O solution,
a peak was observed at m/z 82, which was attributed to the
DSO anion (Fig. 4(F)). This observation indicated that the
hydrogen transfer occurs from the b-position for the bisulfite
anion formation. However, if the amino group is moved further
away from the sulfonate moiety, then the intensity of the m/z 81
peak is reduced to 2%, and the m/z 80 peak once again
becomes the base peak (Fig. 4(C and D)). In addition, the m/z
2
–
3
–
–
noic acid ([17 – H] ) and 4-sulfobutanoic acid ([18 – H] ) is m/z 81
(Figure 5(B and C)). However, the CID mass spectrum of
deuterium-exchanged 4-sulfobutanoic acid showed the base
peak at m/z 82, which revealed that the carboxylate charge-
8
(
1 peak is not observed in the mass spectrum of sulfamate
[12 – H] , Fig. 4(A)). Although a peak at m/z 82 (~10%) is also
–
–
mediated pathway is favored for 4-sulfobutanoic acid ([18 – H] )
observed in the spectra of deuteriated sulfamic acid (Fig. 4(E)),
deuteriated 2-aminoethanesulfonic acid (Fig. 4(G)), and deu-
teriated 3-aminopropanesulfonic acid (Fig. 4(H)), this peak was
(Fig. 5(G)). In this case, the m/z 82 peak is much more intense
than that at m/z 81, indicating that the formation and elimi-
nation of the g-butyrolactone is the favored pathway
(Scheme 6(B)), although the syn-elimination mechanism also
plays a role (Scheme 6(A)). As the carboxylate moiety is placed
farther away from the sulfonate, for example in the case of 6-
3
4
–ꢀ
–
3 3
attributed to the SO ion and not to a DSO , because the
intensity of the m/z 81 peak, in the spectra (Fig. 4(A, C, and D)) of
the non-deuteriated analogs, is very low (Note that the m/z 98,
–
1
26, and 140 ions isolated at unit-resolution are composites of
sulfohexanoic acid ([19 – H] ), the formation of the sulfur triox-
34
isotopologues bearing either two deuterium atoms or one
atom).
S
ide radical anion becomes the primary fragmentation pathway
(Fig. 5(D)). The CID mass spectrum of the anion derived from
deuterium-exchanged 6-sulfohexanoic acid shows peaks at
both m/z 81 and 82, indicating that both pathways are equally
favored in this case (Fig. 5(H)). Other fragmentation pathways
for anions derived from sulfocarboxylic acids consist of a loss
Further information about the hydrogen transfer mechanism
was obtained from the investigations carried out with sulfocar-
boxylic acids (16–19). As shown in Fig. 5(A), peaks for two major
product ions (m/z 95 and m/z 80) are observed in the mass spec-
–
trum of deprotonated 2-sulfoacetic acid ([16 – H] ). The m/z 95
2 2
of a neutral H O or CO molecule probably proceed via frag-
peak represents an ion that originates from a decarboxylation
mentation mechanisms similar to those detailed by Grossert
et al. for dicarboxylic acids.
Cysteic acid (20) is the oxidized form of the amino acid cyste-
–
[45]
of the anion of 2-sulfoacetic acid ([16 – H] ), and the m/z 80 peak
–
3
ꢀ
represents the sulfur trioxide radical anion (SO
). Compared with that
–
of 2-sulfoacetic acid ([16 – H] ), the spectrum of 3-sulfopropanoic
acid ([17 – H] ) shows that base peak is at m/z 81 (Fig. 5(B)). In fact,
ine. It is an intermediate in the metabolism of cysteine and is a
–
[32]
precursor of taurine (14).
The mass spectrometric fragmenta-
–
the addition of an extra methylene group between the carboxylic
tion pathways of anions derived from cysteic acid ([20 – H] ) have
been previously discussed by Smith et al.
[
25,26]
acid and the sulfonic acid moieties results in the exclusive formation
Their results were
–
3
of the bisulfite anion (HSO
). However, when the 3-sulfopropanoic
based on mass-analyzed ion kinetic energy spectra obtained
from fast atom bombardment MS/MS experiments. According
–
acid ([17 – H] ) is sprayed in D O, a peak (~15%) for the
deuteriated bisulfite anion (DSO ) is also observed (Fig. 5(F)).
2
–
–
to their results, cysteic acid ([20 – H] ) fragments similar to other
3
Therefore, two separate mechanisms are expected to participate
sulfonates and a peak at m/z 80 for the sulfur trioxide radical an-
–
ꢀ
[25,26]
in the formation of the bisulfite anion (Schemes 6(A and B)). For
ion (SO ) had been reported.
However, our low-energy
3
–
3
-sulfopropanoic acid ([17 – H] ), the primary mechanism is a
collision-induced fragmentation spectra recorded from cysteic
–
cyclic syn-elimination wherein the charge on the sulfonate oxy-
gen attacks the hydrogen attached to the b-carbon, resulting in
the elimination of a neutral acrylic acid molecule (Scheme 6(A)).
acid ([20 – H] ) showed only a peak at m/z 81 in that region
(Fig. 6(A)). A detailed evaluation of CID mass spectra of the deu-
terium-exchanged cysteic acid indicated that the two separate
fragmentation pathways proposed for the sulfocarboxylic acids
–
The secondary mechanism for 3-sulfopropanoic acid ([17 – H] )
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Dissociation of organic sulfonates
2
Figure 6. Product-ion spectra of anions derived from cysteic acid (m/z 168) (A), [3,3- H
2
]-cysteic acid (m/z 170) (B), cysteic acid ethyl ester
]-cysteic acid in D O (m/z 173) (F), cysteic acid ethyl ester in
2
O (m/z 185) (H) recorded on an API-3000 tandem mass spectrometer at laboratory-frame collision
2
(
D
m/z 196) (C), homocysteic acid (m/z 182) (D), cysteic acid in D
O (m/z 198) (G), and homocysteic acid in D
energy of 25 eV.
2
O (m/z 171) (E), [3,3- H
2
2
2
(
Scheme 6) are also valid for cysteic acid (Fig. 6). In this case, the
carboxylate-mediated charge-directed attack on the a-carbon
Scheme 7(B)) is more favorable than the sulfonate-based
charge-mediated attack on the hydrogen on the b-carbon
group exclusively attacks the b-carbon to form the bisulfite an-
ion (Fig. 6(B and F)). The CID mass spectra of cysteic acid ethyl
–
(
ester ([22 – H] ) and deuterium-exchanged cysteic acid ethyl es-
ter further corroborated the sulfonate charge-mediated syn-
elimination mechanism (Fig. 6(C and G)). In both these spectra,
the base peak is observed at m/z 81, indicating that the sulfo-
nate charge-mediated pathway is the only accessible pathway
(
[
[
Scheme 7(A)). The CID mass spectra of anions derived from
2
–
3,3- H ]cysteic acid ([21 – H] ) and deuterium-exchanged
2
2
3,3- H ]cysteic acid support the conclusion that the sulfonate
2
J. Mass. Spectrom. 2012, 47, 529–538
Copyright © 2012 John Wiley & Sons, Ltd.
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F. B. Jariwala et al.
–
3
Scheme 7. Proposed mechanism for the formation of the bisulfite anion (m/z 81, HSO
) upon fragmentation of deprotonated cysteic acid (20) and
2
2
[3,3- H ]cysteic acid (21).
CONCLUSIONS
Although the major product ion upon fragmentation of many sul-
–
ꢀ
3
, m/z 80), in certain
–
fonates is the sulfur trioxide radical anion (SO
cases, the major product ion is the bisulfite anion (HSO
3
, m/z 81).
The data are summarized in Table 1. In fact, in the negative-ion
CID mass spectra of anions derived from 2-phenylethanesulfonic
–
–
acid ([11 – H] ), aminomethanesulfonic acid ([13 – H] ), 3-
–
–
sulfopropanoic acid ([17 – H] ), 4-sulfobutanoic acid ([18 – H] ),
and cysteic acid ([20 – H] ), the base peak is observed at m/z 81.
–
In the absence of a secondary deprotonation site, the formation
of the bisulfite is facilitated by a site-specific hydrogen transfer
from the b-carbon via a cyclic syn-elimination. Furthermore, upon
introduction of a second deprotonation site, two separate path-
ways, the aforementioned syn-elimination pathway and a carbox-
ylate charge-mediated pathway, compete for the formation of
the bisulfite anion. In the case of the sulfonate charge-mediated
pathway, a hydrogen atom is exclusively abstracted from the b-
carbon for the formation of the bisulfite anion.
Scheme 8. Proposed mechanism for the formation of the bisulfite anion
–
(m/z 81, HSO
3
) upon fragmentation of deprotonated cysteic acid ethyl
ester (22).
(
unlike in the carboxylate group, the ester moiety cannot bear a
negative charge) (Scheme 8).
Homocysteic acid (23) is a gliotransmitter, known to function
[
46]
as an endogenous N-methyl-(D)-aspartic acid receptor agonist.
High concentrations of homocysteic acid may cause hyperho-
mocysteinemia, which can often lead to vascular and neuro-
Acknowledgements
[
46]
–
nal lesions.
In the case of homocysteic acid ([23 – H] ),
The authors would like to thank Bristol-Myers Squibb Co. for
donation of the Applied Biosystems API 3000 triple quadrupole
mass spectrometer. They also thank John Ressler for support in in-
stallation, maintenance, and operation of the API 3000 triple quad-
rupole mass spectrometer. The authors acknowledge support for
this research by funds from Stevens Institute of Technology.
the base peak is observed at m/z 80 with m/z 81 at 15%
relative intensity (Fig. 6(D)). However, the mass spectrum of the
deuterium-exchanged homocysteic acid shows a peak at m/z
82, revealing that the bisulfite anion is mostly a result of the car-
boxylate charge-mediated pathway (Figure 6(H), Scheme 9(B)).
–
3
Scheme 9. Proposed mechanism for the formation of the bisulfite anion (m/z 81, HSO
) upon fragmentation of deprotonated homocysteic acid (23).
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J. Mass. Spectrom. 2012, 47, 529–538

Dissociation of organic sulfonates
Table 1. A summary of the relative intensities of m/z 80 and 81 peaks in CID spectra of some organic sulfonates
–
Parent ion
[M – H] (m/z)
m/z 80 relative intensity (%)
m/z 81 relative intensity (%)
–
R = CH
R = CH
3
2
[1 – H] (m/z 95)
100
100
100
100
0
2
2
2
–
CH
3
[2 – H] (m/z 109)
–
R = n-Pentyl
R = n-Heptyl
[3 – H] (m/z 151)
–
[4 – H] (m/z 179)
–
R′ = H
[5 – H] (m/z 157)
100
100
100
100
100
0.5
1
0
0
R = H
–
R′ = CH
3
[6 – H] (m/z 171)
0
0
R = H
R′ = NH
–
2
[7 – H] (m/z 172)
2.5
4
0
0
R = H
7
9
–
R′ = Br
[8 – H] (m/z 235)
0
0
R = H
–
R′ = CH
3
[9 – H] (m/z 199)
0
0
0
R = CH
3
–
X = 1
X = 2
[10 – H] (m/z 171)
100
0
0
–
[11 – H] (m/z 185)
100
–
Y = 0
Y = 1
Y = 2
Y = 3
[12 – H] (m/z 96)
100
13
0
100
2
–
[13 – H] (m/z 110)
–
[14 – H] (m/z 124)
100
100
–
[15 – H] (m/z 138)
3
–
Z = 1
Z = 2
Z = 3
Z = 5
[16 – H] (m/z 139)
80
0
0
100
100
33
–
[17 – H] (m/z 153)
–
[18 – H] (m/z 167)
17
100
–
[19 – H] (m/z 195)
3
–
R = H
[20 – H] (m/z 168)
0
0
100
100
100
16
A = CH
2
3
–
R = H
[21 – H] (m/z 170)
A = CD
2
3
–
R = CH
2
CH
3
[22 – H] (m/z 196)
4
A = CH
2
3
–
R = H
[23 – H] (m/z 182)
100
2 2
A = CH CH
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