Charge-remote and charge-proximate fragmentation processes in alkali- cationized fatty acid esters upon high-energy collisional activation. A new mechanistic proposal

Article abstract of DOI:10.1002/(SICI)1096-9888(199807)33:7<631::AID-JMS668>3.0.CO;2-M

The effect of the metal ion on the high-energy collision-induced dissociation (CID) of alkali metal-cationized n-butyl and methyl ester derivatives of palmitic and oleic acid was examined. The results show that the alkali metal ion has a pronounced effect and does not act as a mere 'spectator' ion with respect to the fragmentation process. While C-H cleavage is a dominant process for [M + Li]⁺ as well as [M + Na]⁺ precursor ions, C- C cleavage is also significant for the [M + Na]⁺ ions. Homolytic mechanisms involving the formation of a transient biradical cation are proposed which enable us to rationalize in a straightforward manner all product ions formed by both charge-remote and charge-proximate fragmentations. The mechanistic proposal is discussed in view of available knowledge on electron impact, CID and related processes. In order to predict how the alkali metal ion could affect the reactivity of the postulated biradical state formed following electronic excitation of the alkali metal-cationized molecules, quantum chemical calculations were performed on methyl and n-butyl acetate as model substances. The decreased spin density at the carbonyl oxygen atom in the biradical state may provide an explanation for the greater tendency towards C-C cleavage reactions of the sodium-cationized fatty acid esters relative to the corresponding lithium complexes.

Full text of DOI:10.1002/(SICI)1096-9888(199807)33:7<631::AID-JMS668>3.0.CO;2-M

JOURNAL OF MASS SPECTROMETRY
J. Mass Spectrom. 33, 631È643 (1998)
Charge-remote and Charge-proximate
Fragmentation Processes in Alkali-cationized Fatty
Acid Esters upon High-energy Collisional
Activation. A New Mechanistic Proposal
Magda Claeys,* Liberata Nizigiyimana, Hilde Van den Heuvel, Irina Vedernikova and Achiel
Haemers
University of Antwerp (UIA), Department of Pharmaceutical Sciences, Universiteitsplein 1, B-2610 Antwerp, Belgium
The e†ect of the metal ion on the high-energy collision-induced dissociation (CID) of alkali metal-cationized
n-butyl and methyl ester derivatives of palmitic and oleic acid was examined. The results show that the alkali metal
ion has a pronounced e†ect and does not act as a mere “spectatorÏ ion with respect to the fragmentation process.
While C–H cleavage is a dominant process for [M + Li]‘ as well as [M + Na]‘ precursor ions, C–C cleavage is
also signiÐcant for the [M + Na]‘ ions. Homolytic mechanisms involving the formation of a transient biradical
cation are proposed which enable us to rationalize in a straightforward manner all product ions formed by both
charge-remote and charge-proximate fragmentations. The mechanistic proposal is discussed in view of available
knowledge on electron impact, CID and related processes. In order to predict how the alkali metal ion could a†ect
the reactivity of the postulated biradical state formed following electronic excitation of the alkali metal-cationized
molecules, quantum chemical calculations were performed on methyl and n-butyl acetate as model substances. The
decreased spin density at the carbonyl oxygen atom in the biradical state may provide an explanation for the
greater tendency towards C–C cleavage reactions of the sodium-cationized fatty acid esters relative to the corre-
sponding lithium complexes. ( 1988 John Wiley & Sons, Ltd.
KEYWORDS: fatty acids, charge-remote fragmentation, collision-induced dissociation, alkali metal ion, diradical
mass spectrometric techniques to determine structural
INTRODUCTION
features in biomolecules such as fatty acids was recog-
nized by Gross and co-workers (for reviews, see Refs 4
and 5). They reported that, in closed-shell ions contain-
ing a stable charge center, structurally informative frag-
mentation, termed charge-remote fragmentation (CRF),
occurs at sites remote from the charge. Because similar
fragmentation was observed for both positively and
negatively charged closed-shell precursor ions, the
logical assumption was made that the charge is not
involved in CRF reactions. In the present study, atten-
tion was focused on some new insights into the mecha-
nism of CRF in the hope that further progress can be
made towards a physico-chemical understanding of this
process and, more generally, of high-energy CID pro-
cesses of even-electron ions.
A great number of studies have dealt with fundamen-
tal aspects of charge-remote fragmentation and its ana-
lytical application to biomolecules. The mechanism of
CRF has been investigated for molecular ions of satu-
rated fatty acids ([M [ H]~ and [M [ H ] 2Li]`) and
functionalized alkanes, but has remained a subject of
Since the development of soft ionization techniques
capable of producing protonated, deprotonated or
cationized molecules, many studies have been devoted
to collision-induced dissociation (CID) processes of
even-electron molecular ions. Early studies by Rollgen
and co-workers1h3 dealing with high-energy CID reac-
tions of small organic molecules and monosaccharides,
which were cationized with an alkali metal ion (Li`,
Na` or K`) under Ðeld desorption or surface ionization
conditions, demonstrated that the alkali metal ion acts
predominantly as a charge carrier in the qualiÐcation of
a “spectatorÏ ion with respect to the fragmentation
process. It was also noted that molecules cationized
with lithium behave to some extent as odd-electron
molecular ions which are formed in electron ionization
and as excited neutral molecules.1 Investigation of the
CID of even-electron molecular ions enjoyed a revival
when the potential of CID in combination with tandem
controversy.5 A concerted 1,4-H -elimination mechan-
2
ism resulting in terminally unsaturated product ions
* Correspondence to: M. Claeys, University of Antwerp (UIA),
Department of Pharmaceutical Sciences, Universiteitsplein 1, B-2610
Antwerp, Belgium.
E-mail: claeys=uia.ua.ac.be
Contract/grant sponsor: Fund for ScientiÐc Research (FWOÈ
Flanders); Contract/grant number: G.0082.98.
has been formulated by Jensen et al.,6 while a homolytic
CÈC cleavage mechanism has been proposed by
Wysocki and Ross.7 In recent studies, evidence for an
alternative homolytic mechanism involving an initial
CÈH cleavage as a product-determining reaction has
CCC 1076-5174/98/070631È13 $17.50
Copyright ( 1998 John Wiley & Sons, Ltd.
Received 13 January 1998
Accepted 9 March 1998

632
M. CLAEYS ET AL .
been obtained for fatty acid esters cationized with
lithium.8h10 It was demonstrated that this mechanism
not only applies to carbon positions remote from the
charge but also holds equally well for proximate posi-
tions.9 The latter mechanism involving CÈH cleavage
was originally proposed in a study by Claeys and Van
den Heuvel11 of the CRF behavior of long-chain alk-
enylsalicylic acids. Whereas for saturated fatty acid
esters CÈH cleavage appears to be a major pathway,
allylic CÈH and CÈC cleavages are found to be pre-
dominant pathways for monounsaturated and 1,4-diun-
saturated fatty acid derivatives.8,10 It is worth
mentioning that CÈH cleavage has also been reported
to be important in the CID of Li`-cationized tri-
peptides by Leary et al.,12 more speciÐcally, in the for-
matrix, m-nitrobenzyl alcohol, was saturated with LiI or
NaI, respectively.
The fragmentation yield was determined by express-
ing the sum of the abundances of the C , C and @C
2
x
x
(3 O x O n [ 1, where n is the number of carbon atoms
in the fatty acyl chain) ions as a percentage of the sum
of the abundances of these ions and the [M ] Li]` or
[M ] Na]` precursor ion, under experimental condi-
tions such that the precursor ion signal did not become
saturated.
Ion nomenclature
The nomenclature for the fragmentation of fatty acid
molecular ions in CID reactions introduced by Griffiths
et al.14 and modiÐed by Claeys et al.8 has been fol-
lowed. For clarity, the rules relevant to this study are
brieÑy summarized as follows: (i) the italicized capital
letters C, A and H are used to describe the nature of the
bond broken with the charge retained on the carbox-
ylate group; C refers to a bond in a saturated part
whereas A and H refer to an allylic and a homoallylic
bond respectively; (ii) a subscript to the right of the
letter indicates the number of carbon atoms remaining
mation of [A ] Li [ H]`-type ions.
n
In the present study we compared the fragmentation
of Li` and Na` cationized methyl and n-butyl palmi-
tate and oleate with the aim of examining the e†ect of
the alkali metal ion on CID processes. The results show
that the alkali metal ion has an e†ect and does not act
as a mere “spectatorÏ ion: while CÈH cleavage is a domi-
nant process for [M ] Li]` and for [M ] Na]` ions,
CÈC cleavage is also signiÐcant for the [M ] Na]`
ions. Mechanisms involving a transient biradical cation
are proposed which enable us to rationalize in a
straightforward manner all product ions formed. They
account for the di†erences observed between the frag-
mentation of [M ] Li]` and [M ] Na]` ions of satu-
rated fatty acid esters and also provide an explanation
for the peculiar CID behavior of cationized mono-
unsaturated fatty acid derivatives. In order to predict
how the alkali metal ion could a†ect the reactivity of
the postulated excited transition state, quantum chemi-
cal calculations were performed on methyl and n-butyl
acetate as model substances. Some of the results
obtained in this study were presented at the 14th Inter-
national Mass Spectrometry Conference (25È29 August
1997, Tampere, Finland) and a brief preliminary
account will be published in the proceedings of that
conference.13
in the fatty acyl part, e.g. C ; and (iii) a prime to the left
n
of the letter indicates that the product ion is deÐcient in
one hydrogen compared with that product ion which
would be formed by homolytic fragmentation at the
same point in the precursor molecular ion.
It is pointed out that the last rule is a variation of
that introduced by Griffiths et al.,14 which was adapted
in order to make the nomenclature more general and
also applicable to product ions formed from precursor
molecular ions other than [M [ H]~ ions such as
[M ] Li]` and [M [ H ] 2Li]` ions.8 The C ions are
formed by cleavage of a CÈC bond and can be regarded
as odd-electron (OE) ions. The terminally unsaturated
ions @C are generated by CÈH cleavage at various posi-
n
tions in the alkyl chain and subsequent radical-induced
CÈC cleavage and correspond to even-electron (EE)
ions.
EXPERIMENTAL
Materials
Palmitic acid and oleic acid were purchased from Sigma
Mass spectrometry
(St Louis, MO, USA). [9,9-2H ]Palmitic acid was
2
obtained from Merck, Sharp and Dohme (Montreal,
High-energy CID spectra were obtained on
a
Canada).
VG70SEQ hybrid mass spectrometer of EBqQ design
equipped with a cesium ion source (Micromass, Man-
chester, UK). Cesium ions with an impact energy of
D18 keV and a beam Ñux of 0.3 lA were used as the
ionization beam. The accelerating potential in the
source was 8 kV. The helium pressure in the collision
cell of the Ðrst Ðeld-free region was adjusted until the
mass-selected ion beam was reduced to D50% of its
original value. The product ion spectra were acquired
by linked scanning at constant B/E at a scan rate of 8 s
per decade and were obtained in triplicate. The samples
were dissolved in dichloromethane (2.5 mg ml~1) and 1
ll of the solution was mixed with 2 ll of the liquid
matrix on the stainless-steel probe tip. To produce
[M ] Li]` or [M ] Na]` precursor ions, the liquid
Derivatization
Methyl esters were prepared using diazomethane and
n-butyl esters were obtained following the method
reported by Greeley.15 BrieÑy, n-butyl esters were pre-
pared by dissolving 2.5 mg of fatty acid in 95 ll of
methanol and adding 2 mg of tetramethylammonium
hydroxide in 5 ll of methanol and 10 ll of 1-
iodobutane. The mixture was allowed to react at room
temperature for 15 min and then Ðltered in order to
remove tetramethylammonium iodide. The Ðltrate was
evaporated under nitrogen and the residue was dis-
solved in 1 ml of dichloromethane.
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

FRAGMENTATION OF ALKALI-CATIONIZED FATTY ACID ESTERS
633
proÐle: in addition to the ions of the @C series observed
for the fragmentation of the n-butyl and methyl
[M ] Li]` ions, the enhanced relative abundance of
C-type ions, corresponding to CÈC cleavage in the acyl
chain, is a striking feature of the [M ] Na]` spectra. It
is pointed out that the ion due to the loss of 1-butene is
absent from the n-butyl palmitate [M ] Na]` spec-
trum, suggesting that the alkali metal ion also has an
e†ect on the latter fragmentation reaction.
Theoretical calculations
All geometries were optimized with ab initio calculation
using the STO-3G and 6È31G* basis sets (GAMESS-
UK molecular orbital package). The calculations were
performed as follows: in a Ðrst step, the geometry of the
uncomplexed ester (R) was optimized; in a second step,
the geometry optimization was done for the alkali metal
ion complex (RMe`); and subsequently, quantum
chemical properties of the RMe` complex biradical
species were calculated at the unrestricted and restricted
HartreeÈFock (HF) SCF level. The calculations at the
restricted HF level were performed following those at
the unrestricted HF level to determine the Ðnal
transition-state energy. The convergence criterion was
that the norm of the gradient is less than 0.000 75 and of
the average gradient is less than 0.0005.
Two speciÐc sites of Me` attachment to methyl and
n-butyl acetate were found by total optimization using
the STO-3G basis set. The lowest total energies of dif-
ferent conformers formed by Li`/Na` attachment were
determined. In the case of Li` methyl acetate com-
plexes, the structures of the complexes were also fully
optimized at the HF/6È31G* level and the geometries
were reÐned at the MP2/6È31G* level. In addition, the
zero-point vibrational energy (ZPVE) correction was
added for the lowest vibrational level of a ground elec-
tronic state. Calculation at the MP2/6È31G* level also
allowed the deÐnition of two stable conformers for the
Li` methyl acetate complexes.
With respect to the formation of C-type ions, it is
worth mentioning that this fragmentation behavior has
also been reported for fatty acid carboxylate ions14,16
and for triacylglycerols cationized with NH `.17
4
Bambagiotti-Alberti et al.16 observed C-type ions upon
high-energy CID of fatty acid carboxylate ions formed
by dissociative electron capture of fatty acid methyl
esters, while Griffiths et al.14 noted C-type ions employ-
ing an intermediate collision energy (400 eV, laboratory
frame) for electrospray-generated fatty acid carboxylate
ions. Pittenauer et al.17 reported similar homolytic frag-
mentation behavior for electrospray-generated ammon-
iated triacylglycerols which were collisionally activated
on high-energy CID. Furthermore, C-type ions formed
by distal cleavages in monounsaturated and 1,4-diunsa-
turated chains have recently also been shown to be of
high diagnostic value in the structure characterization
of isomeric 5-alk(adi)enylresorcinols by Suzuki et al.18
for determining the locations of double bonds in posi-
tional isomers present in a mixture.
It has been demonstrated in previous studies that
CÈH cleavage in the acyl chain at both charge-remote
and charge-proximate positions is a decisive product-
determining cleavage when the [M ] Li]` ion of
n-butyl palmitate is collisionally activated.8,9 These
former results together with observations on Li`-and
Na`-cationized fatty acid n-butyl and methyl esters
made in the present study, which indicate that the alkali
metal ion inÑuences the fragmentation, prompted us to
RESULTS AND DISCUSSION
CID of n-butyl and methyl palmitate [M + Li]‘ and
[M + Na]‘ ions
consider mechanisms involving
a biradical cation
The CID spectra of the [M ] Li]` and [M ] Na]`
ions of n-butyl and methyl palmitate are illustrated in
Figs 1 and 2. The fragmentation of the n-butyl palmi-
tate [M ] Li]` ion has been discussed in detail in pre-
vious studies8,9 but is brieÑy summarized here in order
to facilitate the discussion of the results obtained in the
present study for the n-butyl palmitate [M ] Na]` ion
and methyl palmitate [M ] Li]` and [M ] Na]` ions.
Product ions formed by high-energy homolytic cleavage
reactions and also a low-energy rearrangement reaction
are present in the high-energy CID product ion spec-
trum of the n-butyl palmitate [M ] Li]` ion [Fig.
1(A)]. The ion at m/z 263 is formed by a low-energy
rearrangement reaction, more speciÐcally, a McLa†erty-
formed by excitation of the alkali-cationized molecule.
Pathways for the high-energy CID fragmentation of
n-butyl palmitate [M ] Li]` and [M ] Na]` ions are
given in Schemes 1 and 2. The proposed pathways not
only account for the formation of @C-type ions (pathway
a/b/c), but also enable us to formulate the C ion and
2
the other ions of the C series in a logical manner
(pathways d and e/f). The proposed pathway (a/b/c) for
the formation of @C-type ions takes into account the
Ðnding by Cordero and Wesdemiotis19 that alkenes are
eliminated during high-energy CID of lithium-
cationized fatty acids. However, with regard to sodium-
cationized fatty acid derivatives, the latter process needs
to be conÐrmed. Previous deuterium labeling results
type rearrangement in the n-butyl ester part. The C ion
obtained
for
lithium-cationized
n-butyl
[9,9-
a is product-
2
and the ions of the @C series correspond to ions charac-
2H ]palmitate indicate that step
2
teristic of high-energy CID, which can all be rational-
ized by initial CÈH cleavage reactions. The
fragmentation of the [M ] Li]` ion of methyl palmi-
tate [Fig. 2(A)] is similar to that of n-butyl palmitate,
except that the ion due to the loss of 1-butene corre-
sponding to a McLa†erty-type rearrangement in the
ester part is absent from the spectrum. Compared with
the [M ] Li]` spectra, the [M ] Na]` spectra [Figs
1(B) and 2(B)] clearly reveal a di†erent product ion
determining.8 Formation of the @C ion may compete
5
with that of the C ion, providing a reasonable explana-
2
tion for the decreased relative abundance of the @C ion
5
compared with other ions of the @C series, which is a
typical feature of the high-energy CID product ion
spectra. The enhanced relative abundance of the C ion
2
and ions of the C-type in the [M ] Na]` spectra may
be explained by a decreased spin density at the carbonyl
O atom of the biradical cation (see below) formed upon
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

634
M. CLAEYS ET AL .
Figure 1. High-energy CID spectra of n-butyl palmitate (A) ÍM ½Li˽ and (B) ÍM ½Na˽ ions. The peaks marked with … refer to C-type
ions.
collisional activation as a Ðrst excited transition state,
favoring CÈC cleavage at positions all along the acyl
chain (pathways d and e/f), which can be regarded as
competitive pathways. Deuterium labeling results
of methyl palmitate, the fragmentation efficiency is 6.4%
for the [M ] Li]` ion vs. 4.3% for the [M ] Na]` ion.
This result can also be rationalized by the lower reacti-
vity of the postulated biradical species in the case of
Na` cationization. As alternative mechanisms to those
depicted in Schemes 1 and 2, direct hydrogen and alkyl
radical release, which is expected to be energetically
more demanding, could also be considered.
obtained for the n-butyl and methyl [9,9-2H ]palmitate
2
[M ] Na]` ions (not shown) are consistent with alkyl
migration because a kinetic isotope e†ect could only be
detected for the @C ion. It can also be noted that the
10
fragmentation efficiency is higher for the [M ] Li]`
Furthermore, comparison of the results obtained for
the n-butyl and methyl palmitate [M ] Li]` and
ions than the [M ] Na]` ions. For example, in the case
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

FRAGMENTATION OF ALKALI-CATIONIZED FATTY ACID ESTERS
635
Figure 2. High-energy CID spectra of methyl palmitate (A) ÍM ½Li˽ and (B) ÍM ½Na˽ ions. The peaks marked with … refer to C-type
ions.
[M ] Na]` ions indicates that the ester alkyl group
ment (FAB)-generated fatty acid carboxylate ions
mainly @C-type ions are formed, employing an interme-
diate collision energy (400 eV, laboratory frame)
electrospray-generated ions also result in a regular
series of C-type ions, suggesting that the internal energy
of the precursor ions has an e†ect on CID fragmenta-
tion processes.20 The formation of a homologous series
of odd-electron ions, which can be compared with
C-type ions, was discussed in detail by Wysocki and
Ross7 for protonated 4-pentadecylpyridine and tri-
methyloctadecylammonium ions which were col-
lisionally activated by low-energy surface-induced
dissociation. While FAB-generated protonated 4-
pentadecylpyridine mainly gave rise to terminally
also has an e†ect on high-energy CID processes. The C
2
ion and the other ions of the C series have an increased
relative abundance in Li` cationized methyl palmitate
compared with Li`-cationized n-butyl palmitate. With
respect to the behavior of Na`-cationized methyl and
n-butyl palmitate, the e†ect of the ester alkyl group can
only be noted for the C ion. These results suggest that
2
the ester alkyl group may in addition to the alkali metal
ion also inÑuence the reactivity of the postulated excited
transition state (see below).
As mentioned already, formation of C-type ions has
been demonstrated for electrospray-generated fatty acid
carboxylate ions.14 Whereas for fast atom bombard-
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

636
M. CLAEYS ET AL .
Scheme 1. Mechanism proposed to rationalize the ¾C type ions formed at high-energy CID of methyl palmitate cationized with Li½ or Na½.
C–H cleavage (route a) occurs at positions all along the acyl chain.
unsaturated product ions, the isobutane chemical
ionization-generated precursor ions yielded an addi-
tional series of odd-electron ions at masses 1 unit
higher. Similar Ðndings were reported for tri-
methyloctadecylammonium ions, which were found to
result in a homologous series of odd-electron ions when
they were produced in a high-pressure FAB source
instead of in a regular FAB source. With respect to pos-
sible transitions in the carbonyl group, both n ] n and
n ] n transitions may be considered. The n,n excited
state, which requires a lower activation energy than the
n,n state, is probably responsible for the loss of 1-butene
in the ester part of lithium-cationized n-butyl palmitate,
which corresponds to a McLa†erty-type rearrangement
and is observed at a low collision energy (50 eV, labor-
atory frame).8 The latter fragmentation reaction may be
compared with the Norrish type 2 photochemical reac-
tion of ketones, which is known to involve an n,n
excited state capable of abstracting a hydrogen atom
from the c-position.21 Concerning the mechanism of the
McLa†erty-type rearrangement in even-electron ions, it
is relevant to mention here that Budzikiewicz and
Bold22 have proposed the formation of an excited
species, more speciÐcally, a transient biradical, for the
McLa†erty-type fragmentation of methyleneiminium
ions. Their proposal for the latter rearrangement,
Scheme 2. Mechanism proposed to rationalize the C type ions at high-energy CID of methyl palmitate cationized with Li½ or Na½ involving
C–C cleavage. It is pointed out that pathway e/f is mainly followed in the case of Na½ cationization,
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

FRAGMENTATION OF ALKALI-CATIONIZED FATTY ACID ESTERS
637
however, is di†erent from that formulated by Veith and
Gross,23 who advanced arguments in favor of a mecha-
nism involving ionÈneutral complexes.
in a previous study,8 but is brieÑy summarized here in
order to facilitate the discussion of the results obtained
in the present study for the n-butyl oleate [M ] Na]`
ion and methyl oleate [M ] Li]` and [M ] Na]` ions.
Comparison of the [M ] Li]` spectra with that of the
[M [ H ] 2Li]` molecular ion of oleic acid, reported
by Adams and Gross,24 shows the same homologous
ion series in each case. The gap of 54 units between the
ions at m/z 191 (@A ) and 245 (@A ), which is typical of
CID of n-butyl and methyl oleate [M + Li]‘ and
[M + Na]‘ ions
The CID spectra of the [M ] Li]` and [M ] Na]` ion
of n-butyl and methyl oleate are illustrated in Figs 3
and 4. The fragmentation of the n-butyl oleate
[M ] Li]` ion [Fig. 3(A)] has been discussed in detail
7
11
an unsaturation at the C(9) position, can be clearly dis-
cerned. The @A ion is accompanied by a signiÐcant
11
A
ion (m/z 246) of the OE type, formed by a distal
11
Figure 3. High-energy CID spectra of n-butyl oleate (A) ÍM ½Li˽ and (B) ÍM ½Na˽ ions.
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

638
M. CLAEYS ET AL .
Figure 4. High-energy CID spectra of methyl oleate (A) ÍM ½Li˽ and (B) ÍM ½Na˽ ions.
allylic cleavage. The @H ion (m/z 259) has an abun-
oleate, except that the ion due to the loss of 1-butene
corresponding to a McLa†erty-type rearrangement in
the ester part is absent from the spectrum.
12
dance comparable to those of the @A and @A ions, a
7
11
feature which is also typical of an unsaturation at the
C(9) position. The n-butyl oleate [M ] Li]` spectrum
shows the characteristics of a n-butyl ester derivative,
As found for the [M ] Na]` spectra of n-butyl and
methyl palmitate [Figs 1(B) and 2(B)], the [M ] Na]`
spectra of n-butyl and methyl oleate [Figs 3(B) and
4(B)] show a di†erent product ion proÐle: in addition to
the ions of the @C, @A and @H series observed for the
fragmentation of the n-butyl and methyl oleate
[M ] Li]` ions, the enhanced relative abundance of
some C-, A-and H-type ions, corresponding to CÈC
cleavage in the acyl chain, is a striking feature of the
[M ] Na]` spectra. In addition, as noted for the frag-
which include the loss of C H (m/z 289), the combined
loss of C H and H (m/z 287) and the formation of the
4 8
4 8
2
ion at m/z 79. In addition, an ion is present at m/z 160,
which can be explained by Li` adduct formation at the
double bond followed by homolytic C(7)ÈC(8) cleavage
and can be regarded as diagnostic of a C(9) double
bond. The fragmentation of the [M ] Li]` ion of
methyl oleate [Fig. 4(A)] is similar to that of n-butyl
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

FRAGMENTATION OF ALKALI-CATIONIZED FATTY ACID ESTERS
639
mentation of the methyl palmitate [M ] Na]` ion, the
ion due to the loss of 1-butene is absent from the methyl
oleate [M ] Na]` spectrum.
The minimum basis set STO-3G was used to
compare the geometric and electronic properties of Li`
and Na` complexes because of the size of the com-
plexes. The calculation of shielding tensors of 13C and
17O in Li`/Na` complexes using STO-3G gave values
which are closer to experimental values than those
obtained with a higher level 6È31G** basis set.26 It was
also found that computation employing STO-3G
a†orded adequate geometric information in the case of
large Li clusters and allowed to predict basic trends
in their properties.27 The optimized geometric values
calculated for Li`/Na`-cationized methyl and n-butyl
acetate are given in Table 1. Two speciÐc sites of Me`
attachment to methyl and n-butyl acetate were found by
total optimization using the STO-3G basis set. Calcu-
lation at the MP2/6È31G* level also allowed us to
deÐne two stable conformers for the Li` methyl acetate
complexes. The main di†erences for the Li` and Na`
complexes (Table 1) relate to the Me`wO5xC3 region.
More speciÐcally, the Me`wO5 bond is higher in the
Na` complex, owing to the small metal chelation e†ect,
compared with the Li` complex [for example, in the
case of methyl acetate, the bond lengths are 2.161 Ó in
the Na` complex (3a) vs. 1.579 Ó in the Li` complex
(2a)], where it can be considered covalent-like.
It has been demonstrated by Antoine and Adams25
that the alkali metal ion has a pronounced e†ect on the
relative abundance of A-type ions corresponding to a
distal allylic cleavage for monounsaturated fatty acids
cationized with Li`, Na` and K`. In agreement with
the latter study, we also noted that the relative abun-
dance of the A ion is higher in the spectra of the
11
n-butyl and methyl oleate [M ] Na]` ion than in those
obtained for the [M ] Li]` ion (the [@A ][A ] ratios
11
11
are 2.6 and 1.9 in the [M ] Li]` spectra of n-butyl and
methyl oleate, respectively vs. 0.6 and 0.6 in the corre-
sponding [M ] Na]` spectra). A mechanism involving
direct C(11)ÈC(12) cleavage (Scheme 3) enables us to
formulate the @A and A ions; again, the decreased
11
11
spin density at the carbonyl O atom of the excited tran-
sition state formed upon CID (see below) allows us to
rationalize the increased relative abundance of the A
ion in the [M ] Na]` spectra.
11
Theoretical calculations
The calculated electronic properties of the Li`/Na`
complexes of methyl and n-butyl acetate are sum-
marized in Table 2. Based on the calculated total energy
Theoretical calculations were performed on Li`-and
Na`-cationized methyl and n-butyl acetate with the aim
of evaluating whether predicted electronic properties
such as the spin density could be correlated with experi-
mental data. In the case of Li`- and Na`-cationized
fatty acid esters, the metal ion behaves as an electron
acceptor for the n-bonded ester group. The complexes
are formed between electron-enriched metal atoms and
electron-poor atoms (O, C) and should therefore be
regarded as “inverseÏ alkali metal-bonded complexes.
using
a
higher basis set (6È31G* with ZPVE
correction), it could be conÐrmed that the a form of the
Li` complex, with the lithiumÈoxygen bond colinear
with the ester carbonyl bond, is more stable than the b
form, which is of the bidentate type. The linear a form
of the methyl acetateÈLi` complex, for example, is
favored over the bidentate b form by about 1.3 kcal
mol~1. The mode of bonding in Li` complexes has
Scheme 3. Mechanism proposed to rationalize the A and ¾A ions at high-energy CID of methyl oleate cationized with Li½ or Na½
involving C–C cleavage.
11
11
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

640
M. CLAEYS ET AL .
Table 1. Ab initio STO-3G optimized geometrical values (see Fig. 5 for deÐnition of bonds (in A) and angles in degrees))
Parameters
No.
Compound
C(1)—O(2)
O(2)—C(3)
C(3)—O(5)
O(5)—Me½
C(3)—C(4)
Me½—O(5)—O(3)
O(5)—C(3)—O(2)
O(2)—Me½
1
C H O
1.437
1.451
1.447
1.458
1.429
1.432
1.450
1.395
1.334
1.397
1.320
1.295
1.301
1.356
1.215
1.258
1.232
1.574
1.213
1.213
1.235
—
1.538
1.536
1.532
1.536
1.499
1.505
1.539
—
123.1
122.5
110.2
122.6
122.9
118.1
122.6
—
3
6 2
C H O Li½
2a
2b
2c
3a
3b
3c
1.579
1.830
1.574
2.161
2.132
1.982
174.9
91.6
3.766
1.838
3.754
4.339
3.590
4.243
3 6 2
C H O Li½
3
6 2
C H O Li½
12
C H O Na½
175.9
163.1
145.9
141.1
6
2
3
6 2
C H O Na½
3
6 2
C H O Na½
12
6
2
been evaluated for sulfones and sulfonates by Gal et
al.28 It is interesting to note that for these compounds,
two stable conformers were predicted by theoretical cal-
culations, a linear adduct with the lithiumÈoxygen bond
colinear with the sulfoxide bond and a bidentate form.
The nature of the bonding in the Li`/Na` complexes
may be evaluated using a Mulliken population analysis.
Table 2 shows that there is a very small charge transfer
in the Li` complex; for example, in the case of the
methyl acetateÈLi` complex, the Li` ion retains 0.64
Figure 5. Ab initio STO-3G optimized conformers of Li½ and Na½ methyl acetate complexes.
Table 2. Calculated spin densities and Mulliken charges for Me‘-cationized methyl and
n-butyl acetate (values discussed in the text are in bold)
Spin density
C(3)
Mulliken charge
No.
Compound
C(1)
O(2)
C(4)
O(5)
on the metal atom
2a
C H O Li½
(0.644)
(1.427)
(0.579)
(2.077)
(2.917)
(0.869)
3 6 2
C H O Li½
0.0023
0.0000
0.0023
0.0001
0.0016
0.0001
0.0002
0.0707
0.0041
0.0047
0.0306
0.0047
0.0013
0.0031
0.0035
0.0000
0.0055
0.0000
0.0008
0.0018
0.0084
0.0122
0.0084
0.0126
0.0529
0.0153
0.0659
0.0159
0.0296
0.0179
3 6 2
C H O Li½
2b
4a
3a
3b
5a
3
6 2
C H O Li½~~
3
6 2
C H O Li½
12
C H O Li½~~
12
C H O Na½
6
2
6
2
3
6 2
C H O Na½~~
C H O Na½
3
6 2
3
6 2
C H O Na½~~
3
6 2
C H O Na
12
C H O Na½~~
12
6
2
6
2
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

FRAGMENTATION OF ALKALI-CATIONIZED FATTY ACID ESTERS
641
electronic units of its positive charge in complex 2a and
1.43 units in complex 2b. The latter result indicates that
bonding in complex 2b is mostly due to electrostatic
interaction and that the major contribution is made by
ionÈdipole interactions. The calculated Mulliken popu-
lations for the Na` complexes reveal higher values for
the two conformers, due to a stronger electrostatic
interaction associated with the charge on Na.
posed mechanism for charge-remote fragmentation in
sodium-cationized fatty acid esters involving alkyl
migration to the cationized ester group, similarity exists
with EI fragmentation mechanisms of fatty acid methyl
esters, as, for example, with that proposed by Budzikie-
wicz et al.33 to rationalize the [M [ 43]` ion involving
a complex scission of an internal methylene part with a
hydrogen transfer and rearrangement of the carbon
skeleton.
The reactive sites for radical attack may be indicated
by the spin density. The reactive nature of the excited
transition state of the lithiated cation could be due to
the nature of the particular radical localization (Table
2). It is worth noting that there is a signiÐcant di†erence
in the spin density on the carbonyl O atom for the a
conformers of the Li` and Na` complexes of methyl
acetate (0.0529 vs. 0.0159). Furthermore, comparison of
the values for the a conformers of the Li` complex of
methyl and n-butyl acetate reveals a higher spin density
on the carbonyl O atom in the case of n-butyl acetate
(0.0529 vs. 0.0659). This result is in agreement with the
Ðnding that the nature of the ester alkyl group also has
an e†ect on CID processes and follows the experimental
trend that n-butyl esters show more pronounced CÈH
cleavage reactions relative to methyl esters. Hence it
appears that the high tendency of Li`-cationized fatty
acid esters to undergo CÈH cleavage following high-
energy collisional activation could be correlated with a
high radical localization on the carbonyl O atom in the
excited transition state.
As regards participation of the charge in high-energy
CID reactions, it should be mentioned that Tuinman
and co-workers34,35 observed a peculiar CID fragmen-
tation of alkyltrimethylammonium ions with a perdeu-
terated alkyl chain. They described mixed-site
fragmentation reactions, in which the Ðxed charge site
on the nitrogen atom interacts with backbone p-bonds,
as competitive reactions with CRF, resulting in a
second series of product ions accompanying the pre-
dicted CRF product ions, but at masses 3 units higher.
A
similar high-energy CID behavior of alkyl-
trimethylammonium ions with deuterated methyl
groups was also reported by Bambagiotti-Alberti et
al.36 Mixed-site fragmentation reactions were recently
investigated in more detail by Whalen et al.,37 who con-
sidered the involvement of a distonic diradical cation to
explain the CID fragmentation of alkyltrimethyl-
ammonium ions with a perdeuterated alkyl chain. The
latter distonic biradical cation may be formed following
an excitation, resulting in a transient excited species
with the active site located at an NwC bond, which can
induce cleavage of backbone CwC bonds. The regular
series of CRF product ions in alkyltrimethylammonium
ions (loss of C H ] H ) can be rationalized from the
How can the new mechanistic proposal be reconciled
with available knowledge on electron impact, CID and
related processes?
n 2n
2
same excited transition state, which can trigger CÈH
cleavage at carbon positions all along the alkyl chain.
With respect to the energetics, it has been demon-
strated by Wysocki et al.38 that CRF reactions are
already occurring at low excitation energies in the eV
range under surface-induced dissociation conditions. An
extreme case was reported by Cody,39 who examined
4-pentadecylpyridine and observed CRF reactions using
a Fourier transform ion cyclotron resonance spectro-
meter and collision energies as low as 2.5 eV. In con-
trast, a large activation energy ([10 eV) was needed to
observe CRF for the stearate anion and octadecyl-
ammonium ion under surface-induced dissociation con-
ditions.38 The energy required to generate an excited
transition state will be dependent on the nature of the
functional group participating in CRF and may either
concern an n ] n, n ] n or p ] p transition. The pro-
posed mechanism based on the formation of a Ðrst
excited transition state can, therefore, also be reconciled
with experimental Ðndings by Wysocki et al.38 that the
internal energy required to record spectra which show
CRF is compound dependent. Other observations rele-
vant to the energetics of CRF processes are those made
by Voinov et al.,40 who demonstrated that saturated
fatty acids form [M [ H]~ ions by electron capture in
two energy regions, 1.2 and 7 eV, and that those formed
by electron capture at 7 eV exhibit CRF behavior. In
contrast, the [M [ H]~ ions formed by electron
capture at 1.2 eV found to correspond to fatty acid car-
boxylate anions revealed a di†erent fragmentation
behavior (i.e. loss of H and H O, and formation of an
The proposed mechanism for charge-remote fragmenta-
tion in lithium-cationized fatty acid esters involving
CÈH cleavage corresponds to formal 1,4-eliminations of
H and can thus be reconciled with the simple mecha-
2
nism proposed earlier by Gross and co-workers.6 It is
also worth noting that the high-energy CID behavior of
lithium-cationized methyl and n-butyl palmitate and
oleate is very comparable to the EI behavior of the
picolinyl derivative of palmitic and oleic acid reported
by Harvey.29,30 The mechanism of the EI fragmentation
of the picolinyl derivatives of fatty acids relates to a
pyridinium radical cation-induced abstraction of a
hydrogen radical at positions all along the alkyl or
alkenyl chain and is similar to those proposed in the
present study for high-energy CID, in which radical
attack at various positions along the chain is triggered
by the excited transition state of the ester carbonyl
group. Furthermore, it is worth mentioning that EI of
fatty acid methyl esters gives rise to a homologous
series of carbomethoxy ions which were extensively
investigated by Dinh-Nguyen31 using deuterium label-
ing and could be partly rationalized by initial hydrogen
transfer to the carbonyl group followed by a radical-
induced CÈC cleavage in the acyl chain. This carbo-
methoxy ion series was also noted by Zirrolli and
Murphy32 on low-energy CID of EI-generated molecu-
lar ions of fatty acid methyl esters and was shown to be
of high diagnostic value for the structural determination
of branched-chain fatty acids. With respect to the pro-
2
2
( 1998 John Wiley & Sons, Ltd.
J. Mass Spectrom. 33, 631È643 (1998)

642
M. CLAEYS ET AL .
ion at m/z 59). Structure investigation of the [M [ H]~
ion formed by electron capture at 7 eV employing
deuterium-labeled isotopomers suggests an elec-
tronically excited state of the molecule which has
retained the hydrogen atoms from the hydroxyl group
and the C(2) position.41 The energetics of charge-remote
fragmentation were recently studied in carbocyanine
dyes using collision- and surface-induced dissociation
mass spectrometry by Melnyk et al.42 This study clearly
showed di†erences in CRF dissociation processes with
varying internal energy deposition.
The mechanism of collisional energy transfer to 8 keV
macromolecular ions has been addressed by Uggerud
and Derrick,43 who questioned whether electronic exci-
tation is an important mechanism and proposed a
simple model for impulsive energy transfer between a
single atom in a macromolecular ion and a gas atom.
The experimental results obtained in the present study
for alkali-cationized fatty acid esters, however, favour
electronic excitation resulting in the formation of a
transient biradical located in the ester carbonyl group.
considerations have led to the proposal of new homo-
lytic mechanisms corresponding to formal 1,4-elimi-
nations of H and to eliminations of alkyl radicals.
2
The proposed mechanisms involve the formation of a
biradical cation as a Ðrst excited transition state with a
biradical site located in the ester carbonyl group, which
interacts with p-bonds of the fatty acyl chain at both
remote and proximate carbon positions. The results
obtained in the present study for charge-remote and
charge-proximate fragmentation in alkali-cationized
fatty acid esters are consistent with participation of the
alkali-cationized ester group in the fragmentation
process and point to electronic excitation resulting in a
transient biradical species during high-energy collisional
activation of these compounds. As a consequence, an
odd-electron ion behavior holds for the high-energy
CID fragmentation of their even-electron molecular
ions.
Acknowledgements
This work was supported by the Fund for ScientiÐc Research (FWOÈ
Flanders) through grant No. G.0082.98. M. Claeys is indebted to the
FWO as a Research Director. L. Nizigiyimana is on leave from the
University of Burundi and thanks the Belgian Ministry of Foreign
A†airs (ABOS) for a doctoral research fellowship. Professor F. W.
Rollgen is gratefully acknowledged for stimulating discussions con-
cerning alkali metal cationization.
CONCLUSIONS
Recent results obtained for the high-energy CID frag-
mentation of long-chain fatty acid methyl and n-butyl
esters cationized with Li` and Na` as well as other
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