Hydrogen Tunneling above Room Temperature Evidenced by Infrared Ion Spectroscopy

Article abstract of DOI:10.1021/jacs.6b10348

While hydrogen tunneling at elevated temperatures has, for instance, often been postulated in biochemical processes, spectroscopic proof is thus far limited to cryogenic conditions, under which thermal reactivity is negligible. We report spectroscopic evi

Full text of DOI:10.1021/jacs.6b10348

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Article
Hydrogen Tunneling Above Room Temperature
Evidenced by Infrared Ion Spectroscopy
Mathias Schäfer, Katrin Peckelsen, Mathias Paul, Jonathan Martens, Jos
Oomens, Giel Berden, Albrecht Berkessel, and Anthony J. H. M. Meijer
J. Am. Chem. Soc., Just Accepted Manuscript • DOI: 10.1021/jacs.6b10348 • Publication Date (Web): 10 Mar 2017
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Hydrogen Tunneling Above Room Temperature Evidenced by
Infrared Ion Spectroscopy
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Mathias Schäfer†*, Katrin Peckelsen†, Mathias Paul†, Jonathan Martens‡, Jos Oomens‡,§*, Giel Ber-
den‡, Albrecht Berkessel†*, and Anthony J. H. M. Meijer∥*
†Department of Chemistry, Organic Chemistry, University of Cologne, Greinstraße 4, 50939 Cologne, Germany.
‡Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7c, 6525 ED Nijmegen, The
Netherlands.
§van’t Hoff Institute for Molecular Sciences, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Nether-
lands.
∥Department of Chemistry, University of Sheffield, Sheffield S3 7HF, UK.
KEYWORDS: carbenes; tunneling; mass spectrometry; infrared ion spectroscopy; DFT calculations
ABSTRACT: While hydrogen tunneling at elevated temperatures has, for instance, often been postulated in biochemi-
cal processes, spectroscopic proof is thus far limited to cryogenic conditions, under which thermal reactivity is negligi-
ble. We report spectroscopic evidence for H-tunneling in the gas phase at temperatures around 320-350K observed in
the isomerization reaction of a hydroxycarbene into an aldehyde. The charge-tagged carbene was generated in situ in a
tandem mass spectrometer by decarboxylation of oxo[4-(trimethylammonio)phenyl]acetic acid upon collision induced
dissociation. All ion structures involved are characterized by infrared ion spectroscopy and quantum chemical calcula-
tions. The charge-tagged phenylhydroxycarbene undergoes 1,2-H-shift to the corresponding aldehyde with an half-life
of about 10 s, evidenced by isomer-selective two-color (IR-IR) spectroscopy. In contrast, the deuterated (OD) carbene
analogue showed much reduced 1,2-D-shift reactivity with an estimated half-life of at least 200 seconds under the ex-
perimental conditions, and provides clear evidence for hydrogen atom tunneling in the H-isotopologue. This is the first
spectroscopic confirmation of hydrogen atom tunneling governing 1,2-H-shift reactions at non-cryogenic temperatures,
which is of broad significance for a range of (bio)chemical processes, including enzymatic transformations and or-
ganocatalysis.
respective hydroxycarbenes. Characterization of the
I. INTRODUCTION
trapped hydroxycarbenes by infrared (IR) spectroscopy
in an argon matrix at 11 K,6-8 in combination with
Rates and selectivities of chemical reactions are classi-
cally defined by energy barriers, but can be significant-
ly biased by quantum-mechanical tunneling.1,2 Indeed,
hydrogen tunneling is known to play a crucial role in
chemical reactivity at cryogenic temperatures, such as
in astrochemistry, and at room temperature in bio-
chemistry, organic chemistry, and catalysis. However,
the effect of tunneling processes at room temperature
are, until now, mainly recognized on the basis of kinet-
ic isotopic effect measurements3,4 and direct spectro-
scopic proof for tunneling control of chemical reac-
tions is thus far limited to cryogenic conditions, where
thermal reactivity is negligible.5-10
high-level computations, established that the isomeri-
zation of hydroxycarbenes to their respective alde-
hydes and enols, proceeds by a tunneling controlled
1,2-hydrogen shift reaction. This conclusion is con-
sistent with the predominant formation of the thermo-
dynamic product through hydrogen tunneling under the
higher kinetic barrier at cryogenic temperatures.6-9
In addition, O’Hair demonstrated in his extensive
work on the fundamentals of the Pesci decarboxylation
reaction11 that charged carboxylate metal complexes
expel neutral CO2 to form metal-ion carbene complex-
es upon collision induced dissociation (CID) in the gas
phase of a mass spectrometer (MS). The tandem MS
experiments were conducted at room temperature in a
linear quadrupole ion trap (QIT) with subsequent struc-
ture analysis of the organometallic ions by UV ion
From 2008 onwards, Schreiner and Allen et al. have
demonstrated that α-ketocarboxylic acids such as gly-
oxylic acid, phenylglyoxylic and pyruvic acid, can be
decarboxylated by flash-vacuum pyrolysis to form the
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spectroscopy. Furthermore, this set-up allows investi-
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II. RESULTS AND DISCUSSION
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gations of the intrinsic reactivity of the in situ formed
analytes via ion−molecule reactions (IMR). To investi-
gate the decarboxylation reaction of α-ketocarboxylic
acids, which is of central importance for e.g. thiamine
dependent enzyme catalysis with pyruvate decarboxyl-
ases,12 we synthesized a charge-tagged phenylglyoxylic
acid derivative. As illustrated in Scheme 1, the fixed-
charge α-ketocarboxylic acid precursor ions are trans-
ferred into the gas-phase by electrospray ionization
(ESI) for analysis in a QIT mass spectrometer.13
II.1 Characterization of hydroxy[4-(trimethylam-
monio)phenyl]carbene (2-H) by infrared ion spec-
troscopy and theory
Decarboxylation of 1-H/1-D can result in both 2-H/2-
D and 3-H/3-D product ions (m/z 164/165) which have
been characterized here by infrared ion spectroscopy.13,
15 The infrared ion spectra of the isobaric product ions
2-H/3-H at m/z 164 (Figure 1 a-c), and of 2-D/3-D at
m/z 165 (Supplementary Figure 6a and 6c), result from
the monitoring of the depletion of the respective pre-
cursor ion and their photo-fragment ions upon IRMPD.
The predominant fragmentation process in all cases is
the loss of a methyl radical, along with the loss of
C2H4 and CH4 and combinations of these (see Supple-
mentary Table 3, SI for details). These results are pre-
sented in Fig. 1 in which panel (a) displays the infrared
spectrum (black) of product ions isolated at m/z 164
compared to the computed IR spectrum of the singlet
trans-hydroxycarbene, 2-Ht (see Supplementary Fig. 3
for comparison of the computed IR spectrum of the
singlet cis-hydroxycarbene, 2-Hc). The presence of 2-
Ht in the ion population at m/z 164 is immediately
identified by the presence of an OH stretching vibra-
tion, vO-H, at 3553 cm-1 and a C-O stretch mode vC-O at
1248 cm-1, both matching the calculated bands for 2-
Ht. In panel (b), the experimental (black) and calculat-
ed (red) IR spectra of the synthesized aldehyde 3-H
model compound (4-(trimethylammonio)benzalde-
hyde) are presented, showing an absence of intensity in
the region of 1250 cm-1 and between 3500-3600 cm-1
where the vO-H and vC-O resonances are found for the
hydroxycarbene 2-Ht. As well, the characteristic alde-
hyde vC=O stretch for 3-H above 1700 cm-1 in panel (b)
identifies the corresponding band at the same position
in the experimental spectrum in panel (a), unaccounted
for the calculated IR spectrum of 2-Ht. This clearly
indicates a mixture of 2-Ht and 3-H in the ion popula-
tion at m/z 164 (see Supplementary Fig. 4 for compari-
son with other constitutional isomers and Supplemen-
tary Table 1).8 A horizontal dashed line in panel (b)
indicates a threshold below which IR resonances are of
insufficient intensity to absorb the required number of
photons to reach the dissociation limit of the molecule,
so that these resonances remain unobserved in the
experimental spectrum. While this threshold limits the
number of bands observed experimentally for the alde-
hyde species (in both panels (b) and (c) of Fig 1.), we
see that this is not the case for the hydroxycarbene
resonances in panel (a), where most of even the weak-
est bands still appear in the experimental spectrum.
This apparent lower dissociation threshold of the hy-
droxycarbene 2-Ht is attributed to isomerization of the
hydroxycarbene to the aldehyde during IR photodisso-
ciation measurements, where the energy released by
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Scheme 1. Gas-phase formation of a hydroxycar-
bene from an α-ketocarboxylic acid precursor in a
QIT at room temperature.
O
OH/D
O
O
H/D
MS2, CID
–CO2
OH/D
H/D
R
tunneling
O
Me3N
Me3N
2-H (m/z = 164)
2-D (m/z = 165)
Me3N
1-H (m/z = 208)
1-D (m/z = 209)
3-H (m/z = 164)
3-D (m/z = 165)
ESI-MS
Infrared ion spectroscopy
The molecular ions of oxo[4-(trimethylammonio)-
phenyl]acetic acid (1-H at m/z 208) and its mono-
deuterated derivative (1-D at m/z 209) are selected and
dissociated by collisional activation. The loss of CO2
via CID delivers a pair of isobaric product ions (2-H,
3-H at m/z 164 from precursor 1-H, and 2-D, 3-D at
m/z 165 from 1-D, respectively) in a charge-remote
fragmentation process.14 Samples of (4-(trimethylam-
monio)benzaldehyde) (3-H) and its isotopomer 3-D
were also prepared and characterized spectroscopically
as reference data sets for comparison (see SI for de-
tails). For the identification and characterization of
individual isomers in a mixture of isobaric product
ions, i.e. the charge-tagged hydroxycarbene 2-H and
the respective aldehyde 3-H, and isotopologues 2-D
and 3-D, we used infrared ion spectroscopy.15 Wave-
length-tunable radiation from a free electron laser
(FEL) and optical parametric oscillator (OPO) laser
sources was used for the irradiation of ions stored in a
spherical QIT mass spectrometer.13 As the density of
ions stored in a QIT is far too low for direct IR absorp-
tion spectroscopy, the extent of precursor ion depletion
and product ion formation is monitored as the energy
of the photons used for activation is tuned (FEL: from
600 to 1800 cm-1 and OPO: 2500-3700 cm-1). In infra-
red ion spectroscopy, the energy of tens to hundreds of
resonantly absorbed photons is turned into vibrational
excitation of all oscillators of the activated ion due to
intramolecular vibrational redistribution (IVR), which
ultimately leads to internal energies where one or more
dissociation pathways become accessible. Infrared ion
spectroscopy combined with a computational analysis
of ion structures has proven to be a powerful analytical
strategy to elucidate ion structures.15
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Figure 1. Experimental and calculated infrared spectra. (a) The experimental IR spectrum of the ion population at m/z
164 generated by CO2 loss from 1-H (black; see Supplementary Table 3; SI) is compared to the calculated IR spectrum of the
charge-tagged trans-hydroxycarbene, 2-Ht (blue). (b) The experimental IR spectrum of the synthesized aldehyde (4-
(trimethylammonio)benzaldehyde) model compound (black; see Supplementary Table 3; SI) is compared with the calculated
IR spectrum (red) for 3-H. (c) The experimental IR spectrum (green) of the remaining ions at m/z 164 in an isomer-selective
two-colour IR-IR experiment (see (c) inset) is compared to the experimental IR spectrum of reference compound 3-H. (d)
The experimental IR spectrum of the •CH3 loss product (m/z 149) recorded after isomer-selective photodissociation of hy-
droxycarbene 2-Ht species is compared to the experimental IR spectrum of the CID generated •CH3 loss product (m/z 149)
from the 3-H model compound. The structures of 2-Ht and 3-H shown in the right of panels (a) and (b) have been computed
at the dispersion-corrected B3LYP-GD3BJ/cc-pVTZ level of theory. Frequencies are scaled by 0.97 (500-2000 cm-1)15 and
by 0.95 (2800-3800 cm-1).16
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isomerization assists the system in reaching its dissocia-
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a
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tion threshold (fewer photons must be absorbed). Panel
(c) of Fig. 1 presents an isomer-selective IR spectrum
(green) of the aldehyde 3-H component of the ion popu-
lation at m/z 164. This spectrum is generated in a novel
two-colour IR-IR approach. Here, the ion population at
m/z 164 is first irradiated at 3553 cm-1 using a 10W cw-
OPO source. At this frequency only the hydroxycarbene
isomer is resonant (vO-H), but the aldehyde is transparent.
Hence, we selectively photo-dissociate the hydroxycar-
bene 2-Ht ions in the population.8 In a second step, the
remaining ions at m/z 164, those without a vO-H reso-
nance at 3553 cm-1, are re-isolated and probed with
tuneable IR light to measure their infrared spectrum. The
clear match between this isomer-selective IR spectrum
and the reference spectrum measured for the 3-H model
compound (black, panels (b) and (c) of Fig. 1) confirms
the assignment of 3-H as the only additional component
besides 2-Ht of the ion population at m/z 164.
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b
Panel (d) of Fig. 1 presents a second two-color IR-IR
experiment where the IR spectrum of the photofragment
at m/z 149, generated selectively from the hydroxycar-
bene (orange), is presented (see experimental section for
details). Similar to the two-color approach in panel (b),
hydroxycarbene ions at m/z 164 are first selectively
photodissociated by irradiation at 3553 cm-1. However,
in the second step, rather than re-isolating the remaining
ions at m/z 164, fragment ions at m/z 149 (•CH3 loss) are
isolated and probed using tunable IR light to generate
their IR spectrum. The resulting spectrum features a
sharp band at 1700 cm-1, indicative of a vC=O stretch,
strongly favoring the assignment of an aldehyde struc-
ture and not a hydroxycarbene structure. Additionally,
the IR spectrum of the •CH3 loss product (m/z 149) from
the 3-H aldehyde model compound was measured
(black) and matches very closely with the m/z 149 frag-
ment generated selectively from the hydroxycarbene.
Supplementary Fig. 5 (Supporting Information) presents
calculated spectra for the •CH3 loss product from 2-Ht
and 3-H, clearly demonstrating that the orange spectrum
in panel (d) cannot be accounted for by a fragment ion
with a hydroxycarbene structure (7-Ht), but matches
well with the calculated IR spectrum of the correspond-
ing, and energetically favored, aldehyde fragment (6-H).
These observations directly implicate isomerization of
the hydroxycarbene 2-Ht, to the aldehyde 3-H, during IR
photodissociation. Our calculations also show that the
energy needed to form 7-Ht from 2-Ht and to form 6-H
from 3-H are 125.8 kJ mol-1 and 220.3 kJ mol-1, respec-
tively. These results are in agreement with the higher
stability for 3-H found in the experiment.
Figure 2. Reaction energy profiles.§ (a) Formation of 2-
Ht from 1-Hc, and 3-H from 1-Ht. (b) Conversion of 2-Ht
into 3-H as well as for the CO-loss reaction from 2-Hc,
respectively. All structures have been calculated at the
dispersion-corrected B3LYP-GD3BJ/cc-pVTZ level of
theory. The red line indicates the corresponding zero-point
corrected CCSD(T)-F12b energies. Reaction Gibbs free
energy profiles are provided in Supplementary Figs. 9 and
10.
zero-point corrected CCSD(T) energies calculated by
Schreiner and Allen et al. for phenylhydroxycarbene in
§
We have chosen to focus on single-molecule events here and are there-
fore plotting the internal energy. This allows for comparison with the
internal energies used for the low temperature (T=10K) experiments of
Schreiner et al.8 where all molecules will most likely be in a single quan-
tum state.19 If we were to consider that in the trap the ions will be in a
thermalized (above room temperature) distribution over quantum states
before, during, and after the CID and IRMPD processes,18a,18b then the
reaction profiles for the Gibbs energy from the SI are appropriate.
To further explore the energetics and pathways for the
formation and reactions of the hydroxycarbene 2-Ht and
the aldehyde 3-H, the zero-point corrected reaction en-
ergy profiles are presented in Figure 2 (the correspond-
ing Gibbs energy reaction profiles are given in Supple-
mentary Figs. 9 and 10). Figure 2 matches closely to the
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terms of relative energies, multiplicity and transition
states8 and demonstrates that the charge-tag does not
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significantly influence the energy profiles for 2-Ht and
3-H.17 As well, CCSD(T)-F12b calculations (see SI for
details) on 2-Ht, 3-H and the barrier between them, as
well as on the neutral analogues, confirm the validity of
the DFT calculations (see also Supplementary Table 2).
Panel (a) of Fig. 2 furthermore shows that 1-H exists as
two nearly isoenergetic conformers, 1-Hc (cis) and 1-Ht
(trans), the latter of which is also found in the crystal
structure of [4-(dimethylamino)phenyl]oxoacetic acid
(S-2-H, see Supplementary Figs. 11 and 12 for overlays
of structures).
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The energy profiles in Fig. 2 indicate that the energy
needed to form 2-Ht is significantly less than the energy
barrier for forming the aldehyde 3-H (see Supplemen-
tary Table 2). Nevertheless, the aldehyde tautomer (3-H)
is formed and experimentally identified as shown in Fig.
1 and discussed above. While the initial formation of the
hydroxycarbene product ion 2-Ht from precursor ion 1-
Ht is clearly favored as shown in panel (a) of Fig. 2,
CID experiments do not exclude energetically demand-
ing fragmentation channels, making the direct formation
of the aldehyde 3-H possible.18a,c,d Finally, the computed
energetics in Fig. 2 show that the formation of the triplet
state of 2-Ht is thermodynamically unlikely, as its ener-
gy lies 53.6 kJ mol-1 above the singlet ground state.8
Moreover, the process is kinetically inhibited due to the
generally slow singlet to triplet intersystem crossing for
molecules without heavy atoms. In line with this, no
evidence of the hydroxycarbene triplet 2-Ht appears in
the IR spectrum of the ions at m/z 164 (see Supplemen-
tary Fig 3). A similar argument holds for CO-loss from
2-Hc.
II.2 Isomerization kinetics of hydroxy[4-(trimethyl-
ammonio)phenyl]carbene (2-H/2-D) measured by
isomer-selective infrared ion spectroscopy
Figure 3. Time-dependent IRMPD measurements of
deuterated and non-deuterated m/z 164/165 ions. Hy-
droxycarbene-selective photodissociation yields were
measured at 1300 cm-1 where hydroxycarbene species are
resonant and give fragmentation, and aldehyde species are
transparent and not detected. Black squares represent the
normalized precursor ion signal and blue circles represent
the sum of the normalized fragment signals, which are
shown in Supplementary Table 3 (IR yield). Three meas-
urements were taken at each delay time (open symbols) and
are averaged (closed symbols).
Prior to the analysis of the isomerization kinetics, the
deuterated isotopologue 1-D was decarboxylated upon
CID and the resulting product ions at m/z 165 were sub-
jected to IR ion spectroscopy. The respective IR spec-
trum is in convincing agreement with the computed one
of 2-D, but also delivers evidence for the concomitant
presence of the aldehyde tautomer 3-D on the basis of
the clearly detected carbonyl stretching mode (see Sup-
plementary Figure 6, SI).
To investigate the time-dependence of the isomeriza-
tion of the hydroxycarbenes 2-Ht and 2-Dt to their re-
spective aldehydes 3-H and 3-D, IR photodissociation
experiments were completed placing a delay (0-20 s)
between generation of m/z 164/165 by CID and probing
by IR photodissociation. Fig. 3 displays the normalized
photofragment signal vs. the normalized precursor ion
signal as a function of the delay. Using a wavelength of
1300 cm-1, where 2-Ht and 2-Dt are both resonant, while
cold 3-H and 3-D are transparent (see Fig. 1 and Sup-
plementary Fig. 6), the isomerization of the hydroxycar-
bene species to aldehyde can be inferred from a decreas-
ing photofragment signal with increasing delay. Panel
(a) of Fig. 3 shows that the maximum fragment ion sig-
nal from m/z 164 is indeed observed at t=0 s and has
almost completely decayed after 20 s, indicating a loss
of resonance at 1300 cm-1 due to isomerization to the
transparent aldehyde species. Conversely, under the
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same conditions, m/z 165 shows essentially no decay in
attributed to hydrogen tunneling.3,24 It shall finally be
mentioned that studies of Gronert24 and O’Hair25 suggest
that ions stored in spherical QITs at RT at a He pressure
of about ∼10−3 mbar, are quickly (in milliseconds)26 and
effectively cooled by numerous collisions with the He
buffer gas, and ultimately thermalize to an effective
temperature of about 320-350 K.18a,18b At a Teff of 350 K
thermal isomerization is virtually impossible for 2-Ht,
due to the substantial energy barrier of about 128 kJ mol-
1, providing further support for the assumption that the
observed isomerization of hydroxycarbene 2-Ht to 3-H
proceeds via hydrogen tunneling.
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fragmentation signal over the full range of 20 s, indicat-
ing that no isomerization takes place and m/z 165 re-
mains resonant with photons at 1300 cm-1.
Together with the identification of the aldehyde 3-H
structure (see Fig. 1, above) as the minor component of
the ion population at m/z 164 (see Fig. 3a at 0s), these
results show that the decay of 2-Ht is accompanied by
the progressive formation of aldehyde 3-H. This isom-
erization reaction of 2-Ht to 3-H and the absence of the
corresponding isomerization of 2-Dt to 3-D represents a
direct observation of hydrogen tunneling at room tem-
perature and above.
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Reaction mechanisms that do not involve tunneling
imply small primary kinetic isotope effects (KIEs) for
reactions where the hydrogen transfer proceeds via a
non-linear TS, a criterion that clearly applies to the 1,2-
H-shift in 2-Ht, as Scheme 1 and Figs. 2 and 3 illus-
trate.20 In contrast, large primary kinetic isotope effects
(KIEH/D = kH / kD) are found when tunneling is involved
(KIE ˃ 6.4 at 300 K)3,4,21 and competitive reaction iso-
tope effects can be excluded.22
III. Conclusions
In summary, we present the first spectroscopic evidence
for hydrogen tunneling at temperatures above 300K
using two-color tautomer selective vibrational spectros-
copy of gaseous, isolated molecular ions. A charge-
tagged phenylhydroxycarbene was generated by the
decarboxylation of oxo[4-(trimethylammonio)phenyl]-
acetic acid in an ion trap mass spectrometer and its
structure and reactivity probed by infrared ion spectros-
copy using an infrared free electron laser (FELIX) in
combination with quantum chemical modelling.15 The
hydroxycarbene undergoes 1,2-H-shift to its correspond-
ing aldehyde, which can be clearly distinguished by
isomer-selective two-color infrared-infrared depletion
experiments. The deuterated carbene analogue shows
negligible 1,2-D-shift reactivity, providing clear evi-
dence that the hydrogen atom rearrangement proceeds
via tunneling. This observation is the first spectroscopic
confirmation of hydrogen tunneling at non-cryogenic
temperatures and provides a firm basis for the interpreta-
tion of tunneling-controlled processes such as enzymatic
reactivity3-5 and organocatalysis.27-29 Additionally, these
results open up new avenues for further investigation of
the intrinsic reactivity of hydroxycarbenes at room tem-
perature in ion trap mass spectrometers via ion molecule
reactions.11,30
Isomerization of hydroxycarbene 2-Ht to aldehyde 3-
H occurs with a t1/2 of approximately 10 s. In the case of
the 1,2-H-shift in 2-Ht, kD is close to zero as the deuter-
ated hydroxycarbene 2-Dt does not appreciably isomer-
ize over the time frame of the experiments (20 s), indi-
cating a major primary KIE. A quantitative analysis of
the data is presented in Supplementary Figs. 7 and 8 and
suggests a KIE ˃ 20. If 2-Ht were to isomerize to 3-H
without involving tunneling, the Eyring equation shows
that the KIE should be approximately 1.2 at 298 K based
on the calculated Gibbs free energies of activation for 2-
Ht and 2-Dt. We note that the substantially shorter half-
life of about 9.5 ± 0.3 s in our experiments above room
temperature (see Supplementary Figure 7, SI), compared
to 2.5 h at 11 K determined by Schreiner et al.8,23 for
phenylhydroxycarbene, suggests a reduced effective
barrier width for the isomerization. This must be due to
the elevated temperature, given that the computed barri-
ers for the neutral phenylhydroxycarbene and the charge
tagged derivative investigated here are similar. The ele-
vated temperature (and associated vibrational excitation)
means that the height of the barrier is smaller relative to
the internal energy of the molecule and that the barrier is
narrower (see Supplementary Fig. 13).8 In addition,
lower-energy vibrational modes will be more extensive-
ly populated in our experiment compared to the one by
Schreiner. Taken together, this may lead to a lower bar-
rier due to geometric and/or electronic changes in the
molecule, whereby the anharmonicity of those low-
energy vibrations is likely to play a role as well.
EXPERIMENTAL SECTION
Mass spectrometry
The compounds 1-H, 1-D, 3-H and 3-D were freshly
synthesized for this study (see Supporting Information
for details) and characterized by positive ion mode elec-
trospray ionization mass spectrometry ((+)ESI-MS).18b
Compounds 1-H and 3-H were dissolved (c ~ 10−5 M) in
CH3OH, compounds 1-D, 3-D were analyzed in
CD3OD/CH3OH. All (+)ESI-MS and MS2 experiments
as well as accurate ion mass measurements were con-
ducted on an LTQ-Orbitrap XL instrument (Ther-
moFisher, Bremen Germany); see Supplementary Fig-
ures 1 and 2. Product ion spectra were acquired in the
linear ion trap (LTQ) part of the LTQ-Orbitrap instru-
ment by CID with the He bath gas present (P = 2 × 10−5
Based on the large KIE observed for the gas-phase
isomerization reaction of 2-Ht to 3-H, we conclude that
the 1,2-H shift reaction of the hydroxycarbene 2-Ht
probed at temperatures above RT can dominantly be
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Torr) and the product ions were analyzed in the orbitrap (each an average of four mass spectra) were triply meas-
1
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4
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8
(see Supplementary Figure 2). Accurate ion masses were
determined in the orbitrap analyser with a resolution of
30000 FWHM with external calibration (Δm < 3 ppm)
or with addition of internal standards (Δm < 2 ppm).
Typical (+)ESI-MS conditions: Flow rate: 5 µL min−1;
Capillary voltage: 3.20 kV; Sheath gas: 4.99 [arb. units];
Aux gas: 2.00 [arb. units]; Resolution: 30000 FWHM.
ured for these time-dependent measurements by increas-
ing the delay in steps of 2 s. The dissociation yield
(normalized total fragment ion intensity = IR yield) for
the non-deuterated species decreases from approximate-
ly 0.6 at t = 0 s, to approximately 0.14 at t = 20 s. For
the deuterated species, the IR yield is nearly constant
over the full range of delay times. We note that the dif-
ference in the dissociation yields between the non-
deuterated/deuterated species at 1300 cm−1 in their re-
spective IR spectra corresponds closely to the difference
in yields measured here at t = 0 s. The dissociation yield
at 1300 cm−1 is taken as a diagnostic for the fraction of
m/z 164 ions being in the carbene isomeric form (com-
pare experimental spectra in Fig. 1a and 1b).
9
Infrared ion spectroscopy
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A modified spherical 3D quadrupole ion trap mass spec-
trometer (Bruker, Amazon Speed ETD) was used for the
infrared ion spectroscopy experiments and has been
described in detail elsewhere.13 The 3D quadrupole ion
trap was operated at ambient temperature (∼320 K)
with a He pressure in the trap of ∼10−3 mbar to assure
optimal performance (e.g. effective collisional cooling,
signal to noise ratio, sensitivity etc.).13 Tunable radiation
for the photodissociation experiments was generated by
the Free Electron Laser for Infrared eXperiments
(FELIX)13,15,16 in the 600 – 1800 cm−1 range and a
pulsed OPO (LaserVision, USA) source in the 2800 –
3700 cm−1 range both operating at a repetition frequency
of 10 Hz. Pulse energies were approximately 20 – 50 mJ
per 5 µs long macropulse for the FEL and 10 – 20 mJ
per 5 ns long pulse for the OPO. The full width at half
maximum bandwidth of the FEL is approximately 0.4%
of the central wavelength and 3 cm−1 for the OPO. Pre-
cursor ions were formed by electrospray ionization from
solutions of 0.5 µM in methanol at a flow rate of 120 µL
hr−1. Ions were irradiated for 1 s, corresponding to inter-
action with 10 laser pulses. The IR spectra were con-
structed from a series of mass spectra recorded with the
laser scanning over the frequency range from 600 –
1800 cm−1, monitoring the intensity of the IR-induced
product ions (primarily methyl radical loss) and the
depletion of the precursor ion as a function of IR fre-
quency. Unimolecular dissociation results from the ab-
sorption of multiple IR photons (IRMPD). The IR yield
(ΣIfragment ions/ ΣIall ions) was determined after laser irradia-
tion at each frequency and was linearly corrected for
frequency-dependent variations in laser power. A grat-
ing spectrometer (wavemeter) was used to calibrate the
absolute frequency of the FEL (OPO).
Two color IR-IR isomer selective infrared ion spectros-
copy
The first laser set at a fixed frequency dissociates all
ions resonant at that selected frequency, here 3553 cm−1.
A 10 W cw-OPO (Argos 2400 BB) was used as the
“burn”-laser. A second, tunable laser is then used to
record the IR spectrum of the remaining ions (those that
were transparent to the frequency of the first laser), in an
analogous manner as the single laser experiments de-
scribed above. Specifically, CID product ions at m/z 164
(unknown mixture of carbene 2-Ht and aldehyde 3-H)
were generated from the m/z 208 precursor ion. We note
that after decarboxylation, the ions are collisionally
cooled before IR irradiation (10’s of ms separation be-
tween these steps).18,26 After isolation of m/z 164, the
ions were irradiated for 40 ms with the output from the
cw-OPO tuned to the carbene OH stretch at 3553 cm−1,
which depletes all ions in the trap absorbing at this fre-
quency, i.e. having a hydroxycarbene structure. The
aldehyde 3-H isomer has no resonances near this fre-
quency and remains unaffected. After 40 ms, further
depletion of the m/z 164 signal was not observed, indi-
cating that all hydroxycarbene ions were removed from
the population. The remaining ions at m/z 164 (those
transparent at 3553 cm−1) were again mass-isolated and
held in the trap for 1 s while being irradiated by the
FEL, which was tuned point-by-point over the 800 –
1800 cm−1 region. Thus, an isomer-selective IR spec-
trum was obtained.
Time-dependent measurements of the isomerization
reaction by selective IR photodissociation
A two-color IR-IR approach was also used to selec-
tively measure the IR spectrum of the fragmentation
product of the hydroxycarbene 2-Ht species resulting
Four steps were used for these experiments: 1) Isolation
and CID fragmentation of the precursor ions at m/z 208
(1-H) and m/z 209 (1-D), respectively, to induce CO2
loss and product ion formation at m/z 164 or 165, re-
spectively. 2) Isolation of the CID product ions at m/z
164 (mixture of carbene 2-H and aldehyde 3-H of un-
known ratio) from 1-H and analogously at m/z 165 from
1-D. 3) Variable delay of 0 to 20 s. 4) IRMPD at 1300
cm−1 of m/z 164 or m/z 165 ions for 1 s. 11 Data points
•
from CH3 loss (m/z 149). As described in the IR-IR
scheme above, CID product ions at m/z 164 (mixture of
carbene 2-Ht and aldehyde 3-H) were isolated and irra-
diated at 3553 cm−1 by the cw-OPO. This selectively
induces dissociation of the carbene species, but not of
the aldehyde species. In the next step, rather than isolate
the remaining ions at m/z 164 (which are assumed to be
aldehyde 3-H), the fragment ions from the hydroxycar-
bene 2-Ht at m/z 149 were instead mass-isolated. An
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IRMPD spectrum was then recorded using the FEL se-
lectively for this CH3-loss product of the carbene spe-
cies.
ACKNOWLEDGMENT
•
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8
The authors thank Professor Stefan Schlemmer, Institute of
Physics, University of Cologne and Professor Frank Tu-
recek, University of Washington, Seattle, USA for critical
discussion of the experiments and results, Dr. Jörg
Neudörfl, Department of Chemistry, University of Co-
logne, for x-ray crystallography, BSc Thomas Thomulka
for synthesizing 1-H, and BSc Rianne van Outersterp for
assistance with the time-dependent IR measurements. We
wish to thank Dr. J. Grant Hill of the University of Shef-
field for his help in running the CCSD(T) calculations. The
skillful assistance of the entire FELIX staff is gratefully
acknowledged. A license for the OpenEye tools, obtained
via the free academic licensing program, is gratefully
acknowledged.
Computations
Density functional theory (DFT) calculations were per-
formed using Gaussian09, version D.01.31 Gaussian was
compiled with Gaussian-supplied versions of BLAS and
ATLAS.32 The B3LYP32 functional was used throughout
this study with the GD3-BJ correction34 to account for
dispersion interactions, whereby it is noted that in this
case the correction did not significantly affect the results
in comparison to the bare B3LYP functional. The cc-
pVTZ basis35 set was used throughout with the ultrafine
setting for the integrals. This computational procedure is
an improvement on the procedure from our earlier work,
which already gave a good correlation with experi-
ments.36–38 All the calculations performed on these sys-
tems were done in vacuo. Intrinsic reaction coordinate
(IRC) calculations were performed with the calculation
of frequencies at every step of the profile. The calcula-
tion of the barrier penetration integral was done using
the IRC profile following the procedure used by
Schreiner et al. outlined in their supporting information,8
using atomic units throughout. CCSD(T)-F12b39 calcula-
tions were carried out with the MOLPRO package of ab
initio programs,40 using the cc-pVDZ-F12 basis set41
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ASSOCIATED CONTENT
Supporting Information
Experimental details of the synthesis of the carbene precursors
and reference aldehydes. Cartesian coordinates and energies of all
calculated structures, zero-point corrected energy profile of the
carbene formation and the isomerization to the respective alde-
hyde. Additional Figures, Tables, and X-ray data of 4-(tri-
methylammonio)benzaldehyde (3-H) and of [4-(dimethylamino)-
phenyl]oxo(O-2H)acetic acid (S-2-D).
AUTHOR INFORMATION
Corresponding Author
Correspondence and requests for materials should be ad-
dressed to M.S. (mathias.schaefer@uni-koeln.de), A.B.
(berkessel@uni-koeln.de), J.O. (j.oomens@science.ru.nl),
and A.J.H.M.M. (a.meijer@sheffield.ac.uk).
(16) Martens, J. K.; Grzetic, J.; Berden, G.; Oomens, J. Int. J. Mass
Spectrom. 2015, 377, 179-187.
Funding Sources
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No competing financial interests have been declared. Fund-
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new Figure 2a (R2)
189x134mm (300 x 300 DPI)
ACS Paragon Plus Environment

Journal of the American Chemical Society
Page 14 of 15
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new Figure 2b (R2)
177x141mm (300 x 300 DPI)
ACS Paragon Plus Environment

Page 15 of 15
Journal of the American Chemical Society
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Figure 3
215x279mm (300 x 300 DPI)
ACS Paragon Plus Environment