Rapid Conventional and Microwave-Assisted Decarboxylation of L-Histidine and Other Amino Acids via Organocatalysis with R-Carvone under Superheated Conditions

Article abstract of DOI:10.1080/00397911.2015.1100745

This article reports a new methodology taking advantage of superheated chemistry via either microwave or conventional heating for the facile decarboxylation of alpha amino acids using the recoverable organocatalyst, R-carvone. The decarboxylation of amino acids is an important synthetic route to biologically active amines, and traditional methods of amino acid decarboxylation are time consuming (taking up to several days in the case of L-histidine), are narrow in scope, and make use of toxic catalysts. Decarboxylations of amino acids including L-histidine occur in just minutes while replacing toxic catalysts with green catalyst, spearmint oil. Yields are comparable to or exceed previous methods and purification of product ammonium chloride salts is aided by an isomerization reaction of residual catalyst to phenolic carvacrol. The method has been shown to be effective for the decarboxylations of a range of natural, synthetic, and protected amino acids.

Full text of DOI:10.1080/00397911.2015.1100745

Synthetic Communications
An International Journal for Rapid Communication of Synthetic Organic
Chemistry
ISSN: 0039-7911 (Print) 1532-2432 (Online) Journal homepage: http://www.tandfonline.com/loi/lsyc20
Rapid Conventional and Microwave-Assisted
Decarboxylation of L-Histidine and Other Amino
Acids via Organocatalysis with R-Carvone Under
Superheated Conditions
Douglas M. Jackson, Robert L. Ashley, Callan B. Brownfield, Daniel R.
Morrison & Richard W. Morrison
To cite this article: Douglas M. Jackson, Robert L. Ashley, Callan B. Brownfield, Daniel
R. Morrison & Richard W. Morrison (2015): Rapid Conventional and Microwave-
Assisted Decarboxylation of L-Histidine and Other Amino Acids via Organocatalysis
with R-Carvone Under Superheated Conditions, Synthetic Communications, DOI:
10.1080/00397911.2015.1100745
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Date: 03 December 2015, At: 21:30

Synthetic Communications1, 0: 1–10, 2015
Copyright © Taylor & Francis Group, LLC
ISSN: 0039-7911 print/1532-2432 online
DOI: 10.1080/00397911.2015.1100745
RAPID CONVENTIONAL AND MICROWAVE-ASSISTED
DECARBOXYLATION OF L-HISTIDINE AND OTHER
AMINO ACIDS VIA ORGANOCATALYSIS WITH
R-CARVONE UNDER SUPERHEATED CONDITIONS
Douglas M. Jackson, Robert L. Ashley, Callan B. Brownfield,
Daniel R. Morrison, and Richard W. Morrison
Department of Chemistry, University of Georgia, Athens, Georgia, USA
GRAPHICAL ABSTRACT
Abstract This article reports a new methodology taking advantage of superheated chemistry
via either microwave or conventional heating for the facile decarboxylation of alpha amino
acids using the recoverable organocatalyst, R-carvone. The decarboxylation of amino
acids is an important synthetic route to biologically active amines, and traditional methods
of amino acid decarboxylation are time consuming (taking up to several days in the case
of L-histidine), are narrow in scope, and make use of toxic catalysts. Decarboxylations of
amino acids including L-histidine occur in just minutes while replacing toxic catalysts with
green catalyst, spearmint oil. Yields are comparable to or exceed previous methods and
purification of product ammonium chloride salts is aided by an isomerization reaction of
residual catalyst to phenolic carvacrol. The method has been shown to be effective for the
decarboxylations of a range of natural, synthetic, and protected amino acids.
Keywords Amino acids; decarboxylation; green chemistry; microwave chemistry;
organocatalysis
INTRODUCTION
Biogenic amines are ubiquitous in the body, regulating a variety of functions
ranging from neurotransmission to cellular signaling to various physiological
Received August 13, 2015.
Address correspondence to Douglas M. Jackson and Richard W. Morrison, Department of
Chemistry, University of Georgia, 140 Cedar St., Athens, GA 30602, USA. E-mail: dmjackson@
uga.edu; rwm@uga.edu
Color versions of one or more of the figures in the article can be found online at www.tandfonline.
com/lsyc.
1

2
D. M. JACKSON ET AL.
responses.[1–4] Given the physiological sensitivity to biogenic amines, the ability to
synthesize these compounds or unnatural derivatives for pharmaceutical purposes
is attractive. Of the biogenic amines, histamine is one of the most widely dispersed
in the body and is known for its importance in the regulation of many bodily
processes.[5–10]
Many biogenic amines including histamine are derived from their respective
a-amino acids via a variety of decarboxylase enzymes.[6,11–17] For example,
study of L-histidine decarboxylase knockout (HDC KO) mice has given much
insight into the effects of histamine in the body.[5,18] These mice lack the ability to
produce histamine because their HDC encoding gene has been “knocked out” or
removed during the mouse cloning process. Studies show that these HDC KO mice
are deficient in natural killer T-cell production[19] and suffer from decreased wound
healing rates.[20] It follows that current pharmaceutical uses of histamine are pri-
marily related to the activation of dendritic cells via the H4 receptor and subsequent
immune response,[21,22] making it clear that an efficient production of high-purity
histamine is immediately important.
Enzymatic decarboxylation of amino acids occurs in many organisms and has
long been reported as a synthetic option for the decarboxylation of the amino acid
histidine.[23,24] Many amino acids have also been shown to undergo decarboxylation
upon reflux in a high boiling solvent, such as cyclohexanol, in the presence of a
ketone[25–27] catalyst with cyclohex-2-en-1-one[28] or acetophenone derivatives[29]
being the most recently reported catalysts. For syntheses of some of the more biolo-
gically relevant amines, previously reported procedures are slow at best and are com-
plicated by difficult purification protocols required to remove by-products and high
boiling solvents. In recent years two processes for the removal of product free amines
by distillation from a high boiling solvent[30,31] have been reported. These methods
assist with the problem of solvent removal for lower boiling amines; however, the
reaction times were still quite long and the successful reports of difficult decarboxy-
lations such as histidine are unsubstantiated. Most especially for free amines that
exhibit high boiling points, an alternative method of isolation is needed to prevent
thermal degradation.
RESULTS AND DISCUSSION
Solvent Optimization
To address the problem of long reaction times in the decarboxylations of many
amino acids such as histidine, it was envisioned that chemistry at temperatures above
the reflux temperature of cyclohexanol (∼160 °C) may provide a solution. However,
as previously noted, a significant amount of effort must be devoted to the removal of
high-boiling solvents at the expense of yield and efficiency. One advantage to using
cylcohexanol or polyethylene glycol reported by all previous researchers was the
solubility of the amine product and insolubility of amino acids, allowing visual deter-
mination of reaction completion.
Rather than employing an even higher boiling solvent system, which would
involve the same difficulties as previous methods, the possibility of a pressurized
reaction system using a solvent with a lower normal boiling point was investigated.

ORGANOCATALYTIC DECARBOXYLATION OF AMINO ACIDS
3
Both microwave-promoted and hot oil bath systems were employed using a sealed
15-bar maximum pressure reaction vessel. While many solvents satisfy the criterion
of greater volatility, the search was narrowed to a series of short-chain alcohol
solvents to promote microwave absorption. Among the short-chain alcohol solvents,
n-butanol, n-pentanol, and isopropanol proved to absorb microwaves insufficiently
while ethanol, methanol, and water dissolve the reactant amino acid at the optimum
reaction temperatures of >160 °C, hindering visual determination of reaction
completion and failing to facilitate decarboxylation of histidine. n-Propanol was
determined to be the optimum solvent for visual inspection of reaction completion
that could reach the maximum safe temperature and be easily removed after reaction
completion. This solvent achieves a maximum temperature in the instrument
(1200 W) of 190 °C (calibrated Æ2 °C) with a vapor pressure of 15 bar as determined
by the Clausius-Claperyon equation. It should also be noted that reactions
performed neat resulted in poor to no yield and aprotic solvents failed to promote
decarboxylation even upon heating to 190 °C in an oil bath.
Catalyst Optimization
Another factor affecting the reaction rate and overall ease of purification of
the product mixture is the identity and load of the catalyst. Previously 1% v=v of
cyclohex-2-en-1-one has been reported by Hashimoto and coworkers for a wide
range of amino acid decarboxylations.[28] Significant amounts of impurities were
observed in the resulting reaction mixture in attempts to reproduce these experi-
ments. Alternatively, it was reported that when acetophenone and derivatives were
used at 20 mol% for decarboxylation of histidine, modest success was observed after
>54 h. In catalyst optimization experiments, cyclohex-2-en-1-one proved to provide
a greater catalytic effect at 20 mol% than acetophenone.
It was envisioned that the “enone” R-carvone, the natural product spearmint
oil, may retain the catalytic advantage over acetophenone while providing an alter-
native method of removal of the catalyst based on the known isomerization reaction
to phenolic carvacrol[32] (Scheme 1). R-Carvone provides the added advantage of
replacing cyclohex-2-en-1-one, a relatively expensive catalyst having acute human
toxicity.[33] It was observed during preliminary experiments that the rate of reaction
significantly increased at greater catalyst load than the 1 mol% load previously
reported by Hashimoto. As a general rule, the catalytic effect becomes appreciable
at about 0.1 mol equivalents for both cyclohex-2-en-1-one and R-carvone and peaks
at about 2 mol equivalents for the amino acid decarboxylation in this study.
The reaction times in minutes of a series of microwave-assisted decarboxylations
of phenylalanine in n-propanol at 190 °C with varying load of R-carvone are given
in Fig. 1.
Scheme 1. Isomerization of R-carvone to phenolic carvacrol.

4
D. M. JACKSON ET AL.
Figure 1. Decarboxylation times of phenylalanine with variable molar ratio of R-carvone.
Decarboxylations of a series of amino acids were performed at the 2 equivalent
catalyst load, comparing the catalytic abilities of cyclohex-2-en-1-one, acetophenone,
both carvone enantiomers, and the simple ketone acetone under solvent-optimized
conditions. Five min were allowed for the microwave to achieve maximum reaction
temperature of 195 °C. Additional reflux time at the maximum temperature was
allowed if necessary. Reaction completion was determined by the transformation
of amino acid slurry to clear solution as shown in Fig. 2. A summary of catalyst
optimization experiments may be viewed in Table 1.
In these reactions R-carvone showed reactivity similar to that of previous cat-
alysts. Decarboxylations with S-carvone as catalyst were also effective, though dec-
arboxylation of L-histamine required 50 min reflux as opposed to 25 min. Given
these results, decarboxylations of D-amino acids with R-carvone as catalyst may also
be performed as desired. With catalytic effect confirmed, it remained to be seen
whether the decarboxylations occur cleanly with minimal by-products. Isolation of
products using the other catalysts under the higher temperature conditions and
catalyst load proved impossible due to decomposition of products and=or catalysts.
Catalyst Removal and Purification
Given that decarboxylations of amino acids with ketone catalysts is thought to
occur through an imine intermediate[26,27] and the observed rise in impurities as a
result of increasing the catalyst load following the Hashimoto procedure, it became
Figure 2. Generic decarboxylation for determination of catalytic effect.

ORGANOCATALYTIC DECARBOXYLATION OF AMINO ACIDS
5
Table 1. Summary of catalyst optimization: Reaction times given in minutes
R-Carvone
S-Carvone
Amino
acid
Phe
His
Trp
Tyr
5
25
5
5
25
5
5
25
5
7
50
5
5
—
7
a
10
6
20
16
40
aCompleted in ∼72 h in an oil bath with decomposition of product.
apparent that the fate of the decarboxylated imine should be investigated. Gas
chromatography–mass spectrometry (GC-MS) analysis of the product mixture after
the decarboxylation of phenylalanine at the 2 mol equivalent catalyst load shows all
decarboxylated imine, no free amine, and minimal volatile impurities. The reaction
mixture was then diluted with aqueous hydrochloric acid, excess R-carvone was
removed via ether wash, and then the free-based product was partitioned to the
organic phase via aqueous sodium hydroxide wash. A significant degree of hydroly-
sis was expected, however, in GC-MS analysis of the organic extract the imine of the
decarboxylated product is quite stable and persists as shown by the chromatogram
and mass spectrum in Fig. 3.
Figure 3. GC=MS analysis of product mixture after acidic workup.

6
D. M. JACKSON ET AL.
Figure 4. Isomerization reaction of R-carvone to carvacrol.
It was observed that the desired imine hydrolysis occurs readily only after heat-
ing in aqueous acid at >50 °C. Even after treatment of the reaction mixture with
many times the reaction volume of 2.0 M HCl, it proved difficult to adequately
remove all traces of the imine at system equilibrium. However, 5 min reflux at
190 °C in 2 M HCl accomplished both hydrolysis of the imine and isomerization
of R-carvone to carvacrol as shown in Fig. 4. Carvacrol is then easily removed via
ether extraction. It should be noted that gentle reflux at 80 °C allows the imine to
hydrolyze in equilibrium and ∼80% of R-carvone catalyst may be recovered via
three sequential refluxes and extractions with ether. Alternatively, near quantitative
recovery is possible via soxhlet extraction with warm toluene. If R-carvone is recov-
ered via the three quick extractions, a final high-temperature reflux should be per-
formed to isomerize residual R-carvone to carvacrol. All methods of amino acid
decarboxylation in the literature fail to account for the quantity and reactivity of
imine that may remain, perhaps necessitating the extensive further purification
required by these authors to achieve pharmaceutical purity. Product amines are iso-
lated as hydrochloride salts by rotary evaporation of water under reduced pressure
and further drying overnight in a vacuum oven.
RESULTS AND SCOPE
A generic representation of the organocatalytic decarboxylations of a range of
alpha amino acids is given in Scheme 2 for both microwave and conventional heating
procedures. The reaction times for the decarboxylation of selected amino acids of
interest are reported in Table 2 along with isolated yields of the amine hydrochloride
or dihydrochloride salts for the optimized reaction conditions highlighted in
Scheme 2. While the range of applicability for decarboxylations using this procedure
was quite good, the procedure proved ineffective for the natural amino acids Arg,
Asp, L-DOPA, Glu, Ser, Asn, Gln, Cys, and Met, given the sensitivity of specific
functional groups to the reaction conditions.

ORGANOCATALYTIC DECARBOXYLATION OF AMINO ACIDS
7
Scheme 2. Summary of (a) microwave and (b) conventional heating one-pot decarboxylation procedures.
The slight differences in the overall reaction times reported between
microwave (MW) heating and oil bath heating are the result of the necessary
differences in experimental protocols. In the microwave reactor initial heating
occurs over a 5-min period and temperature is computer controlled by an infrared
thermometer in a continuous feedback system to Æ2 °C. Conventional heating was
performed in a preheated bath with observed temperature oscillations of no more
than Æ5 °C. Figure 5 shows a representative 1H NMR for the product amine
hydrochloride salts in D2O. The solvent peak arising from acidic proton exchange
1
was suppressed. Note that no organic impurities are observed in H NMR of the
hydrochloride salts using dimethylsulfoxide (DMSO-d6) as solvent.
Further decarboxylations were investigated with unnatural amino acids
and protected amino acids. Results are summarized in Table 3. Note that the
procedure allows for the simultaneous decarboxylation and deprotection of the
protected amino acids investigated. It should be noted that decarboxylated
imines are present with protecting groups in tact in GC-MS chromatograms after
the initial step.
Table 2. Optimized decarboxylations of natural amino acids
MW
Reaction time (min)a
Oil bath
Reaction time (min)
Amino
acid
Yield (%)
Yield (%)
Ala
Gly
His
Ile
Leu
Lys
Phe
Pro
Thrb
Trp
Tyr
Val
5
13
25
9
5
12
5
5
5
20
20
5
60
86
87
69
72
73
76
80
59
53
53
79
38
40
12
12
5
17
5
5
12
9
40
9
74
59
92
76
69
93
78
48
41
72
67
55
aReaction times represent total programmed time including 5 min initial heating period.
bRequires 80 °C hydrolysis with soxhlet extraction.

8
D. M. JACKSON ET AL.
Figure 5. 1H NMR of histamine dihydrochloride with solvent D2O suppressed.
Table 3. MW decarboxylations of selected unnatural amino acids
Amino acid
Reaction time (min)a
Yield (%)
4-Amino-Phe
4-Bromo-Phe
4-Methyl-Phe
4-Nitro-Phe
3,5-Dibromo-Tyr
3-Iodo-Tyr
Cycloleucine
8
5
5
5
5
5
16
5
5
5
99
47
99
74
41
77
40
86
74
81
76
64
N-Trityl-His[a]
N-Formyl-Trp[b]
O-t-Butyl-Tyr[c]
O-Acetyl-Tyr[c]
O-2,6-Diclorobenzyl-Tyr[c]
5
5
aIsolated product is histamine dihydrochloride.
bIsolated product is tryptamine hydrochloride.
cIsolated product is tyramine hydrochloride.
SUMMARY
Decarboxylation of L-histidine and other L-amino acids has been accomplished
via organocatalysis with R-carvone and subsequent one-pot hydrolysis under solvent
superheated conditions using both conventional heating and microwave irradiation.
Decarboxylation is more rapid than previous methods as the vessels are heated to
190 °C over 5 min. R-Carvone catalyst may be recovered via extraction with diethyl
ether or via soxhlet extraction with toluene if the hydrolysis is conducted at 80 °C;
however, to obtain the greatest purity it is necessary to conduct a high-temperature
hydrolysis to isomerize residual R-carvone to carvacrol. Isolated yields of amine
hydrochloride salts are comparable or improved over previous methods ranging from
1
60 to 90% with purity of hydrochloride salts estimated to be >99% by H NMR.

ORGANOCATALYTIC DECARBOXYLATION OF AMINO ACIDS
9
This process provides a more versatile and efficient option in the synthesis of
biologically active amines from amino acids given the demonstrated versatility with
not only natural amino acids but protected and synthetic derivatives as well.
EXPERIMENTAL
The 5-mmol scale microwave experiments were performed in Milestone 25-mL,
15-bar glass pressure reactors inside the Milestone StartSYNTH™ microwave reac-
tor with external infrared (IR) temperature control. Conventional heating experi-
ments were performed in silicone oil in identical reaction vessels. Solvents and
reagents were purchased from Sigma-Aldrich or Chem-Impex International and used
without additional purification. FT NMR experiments were recorded at 400 MHz in
D2O solvent.
General Decarboxylation Procedure
A magnetic stir bar, 3 mL of n-PrOH, 10mmol of R-carvone, and 5 mmol of
amino acid were charged to a pressure vessel. The vessel was heated from room tempera-
ture to 190 °C over 5 min with stirring. If necessary the reaction vessel was maintained at
190 °C for an additional time until the slurry became clear. The vessel was allowed to
cool to below the solvent boiling point, carefully vented to release evolved CO2, and
analyzed via GC-MS to verify the presence of decarboxylated imine. Ten mL of 2 M
HCl was then added in a one-pot fashion, and, in all cases but threonine, the reaction
vessel was heated to 190 °C over 5 min with stirring and then allowed to cool. In the case
of the temperature-sensitive threonine, soxhlet extraction with toluene was performed at
80°C to remove R-carvone from the reaction mixture. The aqueous reaction mixture
was washed three times with 25 mL of ether, and then water solvent was distilled off
from the hydrochloride salt. The hydrochloride salt was transferred to a vacuum oven
and dried overnight at 150 °C and 10Torr. The hydrochloride salt was then weighed,
melting point was obtained, and the salt was analyzed via NMR spectroscopy.
Additional spectra and experimental data may be viewed in supplementary information.
FUNDING
Funding by the UGA CURO Program and UGA STEM Institute are
gratefully acknowledged.
SUPPLEMENTAL MATERIAL
Supplemental experimental data for this article can be accessed on the
publisher’s website.
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