New Organocatalyst scaffolds with high activity in promoting hydrazone and oxime formation at neutral pH

Article abstract of DOI:10.1021/ol503372j

The discovery of two new classes of catalysts for hydrazone and oxime formation in water at neutral pH, namely 2-aminophenols and 2-(aminomethyl)benzimidazoles, is reported. Kinetics studies in aqueous solutions at pH 7.4 revealed rate enhancements up to 7-fold greater than with classic aniline catalysis. 2-(Aminomethyl)benzimidazoles were found to be effective catalysts with otherwise challenging aryl ketone substrates.

Full text of DOI:10.1021/ol503372j

Letter
pubs.acs.org/OrgLett
New Organocatalyst Scaffolds with High Activity in Promoting
Hydrazone and Oxime Formation at Neutral pH
Dennis Larsen,† Michael Pittelkow, Saswata Karmakar, and Eric T. Kool*
,‡
‡
†
,†
†
Department of Chemistry, Stanford University, Stanford, California 94305-5017, United States
‡
Department of Chemistry, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark
S Supporting Information
*
ABSTRACT: The discovery of two new classes of catalysts for
hydrazone and oxime formation in water at neutral pH, namely 2-
aminophenols and 2-(aminomethyl)benzimidazoles, is reported.
Kinetics studies in aqueous solutions at pH 7.4 revealed rate
enhancements up to 7-fold greater than with classic aniline catalysis.
2-(Aminomethyl)benzimidazoles were found to be effective catalysts
with otherwise challenging aryl ketone substrates.
1
ioconjugation is an essential tool in chemical biology.
B
Chemical transformations that are fast, reliable, and
biocompatible are key to enable important tasks such as site-
1a
selective functionalization of proteins and attachment of
2
reporter molecules to biomacromolecules.
For this reason, numerous bioconjugation techniques have
1
b−d
emerged.
One classical bond-forming reaction is the
formation of hydrazone or oxime linkages to aldehyde or ketone
moieties. However, the relatively slow rate of these reactions at
1c,3
biological pH has limited the usefulness of this approach. In
recent years, several groups have used nucleophilic catalysts to
Figure 1. Top: Proposed reaction mechanism for hydrazone and oxime
formation under nucleophilic catalysis by an amine at neutral pH.
Bottom: Suggested transition states for the rate-determining dehy-
dration step using catalysts from earlier studies and from this study.
11,12
4
facilitate these reactions, including the use of aniline and its
5
simple substituted derivatives. Nucleophilic catalysis of
hydrazone or oxime formation has also found use within other
fields of chemistry, such as in dynamic combinatorial
1
4
4
a,6
7
catalysts have a pK near solution pH. We have suggested that
chemistry, polymer chemistry, and the functionalization of
a
5e,f,8
intramolecular protonation of the intermediate hemiaminal via
an 8-membered ring in the proposed transition state (TS)
facilitates the limiting dehydration step (Figure 1, bottom).
We hypothesized that catalysts based on a smaller scaffold
could potentially lead to increased reaction rates by invoking a
more favorable 7-membered intramolecular proton transfer in
the TS (Figure 1, bottom). To test this, we investigated
potential catalysts with three covalent bonds between the
nucleophilic amino group and the proton donor and report here
the discovery of two new highly active catalyst scaffolds.
To test catalysts, the reaction between p-chlorobenzaldehyde
and phenylhydrazine in phosphate-buffered saline at pH 7.4
PBS) with 10% DMF was followed using UV/vis spectroscopy
(Scheme 1). We included the organic cosolvent to ensure
solubility of all components, although later tests showed that it
was unnecessary, at least in some cases (see below). Nonlinear
regression was used to obtain (pseudo)-second-order rate
carbohydrates via the nonreducing end.
In recent studies we
have focused on elucidating the electronic factors that affect the
rate of formation of these linkages and on developing more
efficient organocatalysts for these reactions by appending proton
9c
9
donor/acceptor moieties to aniline scaffolds. To further
improve the utility of hydrazone and oxime conjugation, new
15
catalyst scaffolds would be useful because aniline is cytotoxic and
10
is a relatively inefficient catalyst.
Detailed studies by Jencks of the mechanism of semicarbazone
and oxime formation under aniline catalysis showed that the rate-
limiting part of the reaction is condensation of the substrate with
the catalyst to form an activated imine, which is subsequently
attacked by the α-nucleophile in a rapid transimination
(
1
1
reaction. Under neutral conditions, Jencks found that the
specific rate-determining step in forming the activated imine is
1
2,13
dehydration of the intermediate hemiaminal (Figure 1, top).
Anilines with ortho-proton donors such as carboxylic acid or
constants (k ) from the data (Tables 1 and 2).
2
phosphonic acid groups give considerable rate enhancements
9c,d
over aniline itself in both hydrazone and oxime formation.
Evidence strongly supports the notion of nucleophilic catalysis
combined with general acid/base catalysis, where the best
Received: November 20, 2014
©
XXXX American Chemical Society
A
DOI: 10.1021/ol503372j
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Letter
Scheme 1. Model Reaction for Catalyst Screening
aminophenols were investigated (entries 6−12), and it was
found that electron-withdrawing groups para to the hydroxy
group (e.g., NO , CN, COOR) enhanced the catalytic effect.
2
This supports the hypothesis that the system is under general
acid/base catalysis, along with the expected nucleophilic
catalysis, as adding electron-withdrawing groups to 2-amino-
1
6
Table 1. Aromatic Amines, Aminophenols, and Aliphatic
Amines As Catalysts for Hydrazone Formation
phenol (pK 9.75) brings the phenol pK closer to the pH of the
a
a
a
medium. A broader range of other aminophenols were also tested
see Supporting Information (SI)), but p-nitro and p-
(
methoxycarbonyl derivatives (entries 6, 11) yielded the fastest
rates. Indeed, the p-methoxycarbonyl derivative (11) yielded rate
enhancements 7 times that of aniline and 4-fold better than that
of 1,3-diaminobenzene (13), described recently as “highly
5b
efficient”.
At pH 7.4, simple aliphatic amines, with pK ’s of ca. 9−11, are
a
expected to be poor catalysts in this context due to the non-
nucleophilic nature of their ammonium ions. Inspired by the
3
e,17
work of Hine on imine formation of protonated diamines,
2-
1
8
dimethylaminoethylamine (14) with a pK value of 7.13 was
a
investigated, but only a small rate enhancement was observed. All
proteinogenic amino acids in principle possess the structural
motif sought after in this study, as they all have three covalent
bonds between the amine nucleophile and the proton donor
(
carboxylic acid OH-group). Although α-amino acids are
generally zwitterionic at pH 7.4, a small selection was screened.
L-Alanine (15) showed practically no catalytic activity (k = 1.1)
rel
while L-proline (16) showed a slightly higher activity (k = 1.3),
rel
19
possibly due to its higher nucleophilicity. However, consid-
erable catalytic activity was found with L-histidine (17) (krel
.0) and L-histidine methyl ester (18) (krel = 3.7). Since
=
2
esterification of the carboxylic acid leads to an increase in
reaction rate, we hypothesized that the imidazolium moiety was
responsible for the high activity of L-histidine compared to other
aliphatic amine compounds. The imidazolium moiety in the
a
Second-order rate constants are given as mean values ± standard
deviations (based on triplicate measurements or better). Conditions:
1
sodium and potassium phosphates (pH 7.4). Concentrations of
reactants and catalyst as per Scheme 1.
20
:9 DMF/water with 123 mM NaCl, 2.4 mM KCl, and 10.7 mM
methyl ester of histidine has a pK value of 7.33, consistent with
a
the importance of protein histidine units in general acid/base
21
catalysis at physiological pH.
In light of these results, we examined other aminoalkyl
imidazoles (Table 2), with the simplest derivative, 2-(amino-
methyl)imidazole (19), giving a 6.8-fold rate enhancement. 2-
Table 2. Aminoalkyl-Substituted N-Heterocycles As Catalysts
for Hydrazone Formation
a
22
(
Aminomethyl)pyridine (20) with a pK of 8.6 also showed
a
good catalytic activity for an aliphatic amine, with rate
enhancements similar to that of L-histidine methyl ester (entry
18 in Table 1). Significantly, an almost 10-fold increase in
reaction rate was seen with 2-(aminomethyl)benzimidazole
23
(
21). With a pK of 7.85, this catalyst falls near the ideal range
a
for general acid/base catalysis at pH 7.4.
Based on the latter finding, some derivatives of 2-(amino-
methyl)benzimidazole were investigated to elucidate the
electronic and structural requirements for this new catalyst
scaffold (entries 22−29). While solubility issues prevented the
proper measurement of derivatives bearing methyl or chloro
substituents in the 5-position (entries 23, 24), the 5-methoxy
derivative (22) gave a slightly higher rate constant than the
parent compound (21). Studies of the two N-methyl derivatives
(25 and 26) suggest that the mechanism of action for 2-
(aminomethyl)benzimidazole catalysts involves imine formation
on the amino group, rather than iminium formation on the
imidazole moiety, since methylation of the imidazole-N has only
a small effect on the rate. Employing a more nucleophilic
secondary amine, as in the 2-pyrrolidinyl derivative (27), did
improve the catalyst slightly compared to the N-methyl
derivative (26), but it did not fully restore the reactivity seen
a
Second-order rate constants are given as mean values ± standard
deviations (based on triplicate measurements). Conditions as per
Table 1. Relative to the uncatalyzed reaction (entry 1 in Table 1).
b
Aniline (2) was screened as a reference (Table 1), and it
yielded a 1.8-fold increase in reaction rate at 1.0 mM. Proceeding
to new scaffolds, first we studied aminophenols. 4-Aminophenol
(
(
3) proved to be unstable, but it was evident that 2-aminophenol
5) is an interesting scaffold. A range of substituted 2-
B
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Letter
in the parent compound (21). Adding an isopropyl group to the
bridging methylene lowers the activity only slightly (entry 28).
Using an ethylamino group instead of the methylamino group
confers more flexibility to the catalyst, but this also lowers the
activity (entry 29), consistent with the notion that smaller ring
sizes are favorable for intramolecular catalysis.
The finding of these new, active catalyst scaffolds prompted us
to investigate their reaction scope with a range of aldehydes and
ketones. We tested the best catalysts identified here with 10
different electrophiles, employing phenylhydrazine as the
standard nucleophile (Scheme 2). Rate constants for all
In contrast to aldehydes, ketones react at similar rates whether
they are aromatic or aliphatic, and efficient catalysts for these
substrates have previously failed to emerge, either from studies of
9c,d,24
the classic aniline catalyst or more recent investigations.
We
reasoned that employing primary aliphatic amine nucleophiles
such as 2-(aminomethyl)benzimidazoles might solve this
problem by presenting less steric bulk. In fact, 2-(amino-
methyl)-5-methoxybenzimidazole (22) gives a 5.4-fold increase
of the reaction rate of phenylhydrazine with acetophenone at 1.0
mM catalyst concentration, a hitherto unseen rate enhancement
for the catalysis of hydrazone formation on ketones under
2
4
physiological conditions. With ketone substrates, the 2-
aminomethyl)benzimidazoles 21 and 22 are better than or
equivalent to the previously reported phosphonic acid catalyst
Scheme 2. General Reaction for Scope Investigation
(
(
PA). This is significant because aryl ketones are common
1
b−d,2a,25
reactants in bioconjugations.
In order to test the applicability of these catalysts in purely
aqueous conditions without cosolvent, the reaction between
benzaldehyde and phenylhydrazine in phosphate-buffered saline
at pH 7.4 was performed under identical conditions to Scheme 1
without addition of DMF. Rate enhancements for the three
employed catalysts, aniline (2) (2.2-fold), 2-amino-4-nitro-
phenol (6) (4.8-fold), and 2-(aminomethyl)benzimidazole
substrates were measured without a catalyst and under aniline
catalysis for comparison (Table 3). We also measured rate
constants with 2-amino-5-methylphenylphosphonic acid (de-
noted PA in Table 3), a catalyst previously reported by our
9
d
group.
(
21) (9.0-fold), were very similar to those reported in Tables 1
Comparing reaction rates of aliphatic vs aromatic aldehydes,
and 2 (see SI for details). This establishes that the cosolvent
DMF does not dramatically influence the catalytic effect and is
not needed for the solubility of these catalysts. This suggests their
further use in biological applications.
we confirmed previous findings that aliphatics react much more
9
a,b
rapidly. Even bulky aliphatic aldehydes react much faster than
aromatic aldehydes, and rate enhancements when adding
catalysts are relatively low (up to 2.8-fold), suggesting that the
catalysts have less impact when background hydrazone
formation is fast. Significantly higher rate enhancements are
seen for the aromatic aldehydes (up to 19-fold for new catalysts).
The best new catalysts are roughly on par with the previously
reported phosphonic acid catalyst (PA) when using aliphatic
aldehyde substrates, but for aromatic aldehydes the phosphonic
acid derivative is generally better (1.6−3.7-fold faster than the
best new catalyst).
Oxime formation is considerably slower than hydrazine
3a,4a,9b
formation,
but oximes are more resistant to hydrolysis
3d
than hydrazones. Accordingly, there is a demand for catalysts of
this related reaction. Therefore, we investigated our most
efficient new scaffold as a catalyst for the oxime formation
between 2-(dimethylamino)ethoxyamine and 4-nitrobenz-
aldehyde. We found that 2-(aminomethyl)benzimidazole (21)
was effective in catalyzing this reaction, giving a 5.2-fold rate
a
Table 3. Scope of Catalysis of Hydrazone Formation between Phenylhydrazine and a Range of Aldehydes and Ketones
a
Second-order rate constant (M s−1) given as mean value ± standard deviation (based on triplicate measurements). Conditions as per Table 1 and
−1
b
Scheme 2. Values in parentheses are normalized to 1.0 mM catalyst concentration by extrapolation from lower catalyst concentrations to avoid
detector saturation caused by spectral overlap (see SI). From duplicate measurements. n.d., no data.
c
C
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Organic Letters
Letter
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see SI for details).
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1
hydrazone and oxime formation by introducing two new
subclasses of catalyst scaffolds, namely the 2-aminophenols and
the 2-(aminomethyl)benzimidazoles. Both new scaffolds are
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■
*
S
(
Full list of tested aminophenol catalysts, supporting figures and
procedures, equations, and examples of data fits used for kinetics.
This material is available free of charge via the Internet at http://
pubs.acs.org.
(
1
(
AUTHOR INFORMATION
Corresponding Author
(
■
(
(
12) Cordes, E. H.; Jencks, W. P. J. Am. Chem. Soc. 1962, 84, 832.
13) Results show first-order dependence on hydrazine under the
*E-mail: kool@stanford.edu.
conditions applied here. We hypothesize that this can be explained by a
quasi-steady-state approximation (see SI).
Notes
(
14) Anslyn, E. V.; Dougherty, D. A. Modern Physical Organic
The authors declare no competing financial interest.
Chemistry; University Science Books: Sausalito, CA, 2006.
(
(
15) Galli, C.; Mandolini, L. Eur. J. Org. Chem. 2000, 3117.
16) Vandenbelt, J. M.; Henrich, C.; Vanden Berg, S. G. Anal. Chem.
ACKNOWLEDGMENTS
■
D.L. is thankful for travel grants for this work by the Danish
Chemical Society and the Lundbeck, Oticon, and Augustinus
foundations. We thank the Lundbeck Foundation for a Young
Group Leader fellowship (M.P.) and a PhD fellowship (D.L.).
E.T.K. acknowledges support from the U.S. National Institutes of
Health (GM068122 and GM110050).
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