Parallel solution-phase synthesis of a 2-aminothiazole library including fully automated work-Up

Article abstract of DOI:10.2174/138620711794474024

A straightforward and effective procedure for the solution phase preparation of a 2-aminothiazole combinatorial library is described. Reaction, work-up and isolation of the title compounds as free bases was accomplished in a fully automated fashion using the Chemspeed ASW 2000 automated synthesizer. The compounds were obtained in good yields and excellent purities without any further purification procedure. 2011 Bentham Science Publishers Ltd.

Full text of DOI:10.2174/138620711794474024

104
Combinatorial Chemistry & High Throughput Screening, 2011, 14, 104-108
Parallel Solution-Phase Synthesis of a 2-Aminothiazole Library Including
Fully Automated Work-Up
Hans-Peter Buchstaller* and Uwe Anlauf
MS-DTC-MD1B Med Chem Darmstadt 1B1, Merck Serono, Merck KGaA, Frankfurter Strasse 250, D-64293 Darmstadt,
Germany
Abstract: A straightforward and effective procedure for the solution phase preparation of a 2-aminothiazole
combinatorial library is described. Reaction, work-up and isolation of the title compounds as free bases was accomplished
in a fully automated fashion using the Chemspeed ASW 2000 automated synthesizer. The compounds were obtained in
good yields and excellent purities without any further purification procedure.
Keywords: Aminothiazole, combinatorial chemistry, solution phase synthesis, automation.
INTRODUCTION
thioureas, first reported in 1887 [6], is a robust and well
established procedure and was already used several times for
the solution-phase preparation of 2-aminothiazoles in a
combinatorial fashion. On the one hand, dedicated libraries
of this scaffold for medicinal chemistry programs were
reported [7]. On the other hand, papers appeared in the
literature dealing with the general applicability of this
reaction to parallel solution phase synthesis. Bailey et al. [8]
described a convenient procedure to gain such compounds,
but they quenched the reaction with 10% dimethylamine in
ethanol which remained as hydrobromide in the final
product. Polymer-supported reagents were used for either the
reaction itself [9] or the liberation of the products from their
salts [10], but the use of this technology can become costly
when applied to the preparation of large libraries. Also
recently a microwave system was used to accelerate the
condensation reaction [11]. Given their widespread
occurrence in biological active molecules and the robustness
of their syntheses from readily available substrates, we
explored the possibility of an automated solution phase
preparation of a library of 2-aminothiazoles for our screening
collection.
The design and synthesis of small molecules that are
precious therapeutic agents is one of the main objectives of
organic and medicinal chemistry. Many examples verify that
substituted heterocyclic compounds can offer a high degree
of structural diversity and have proven to be broadly useful
as therapeutic agents. The preparation of heterocyclic
scaffolds even in an automated fashion has gained enormous
interest in combinatorial synthesis, and the development of
strategies for library generation has generated a great deal of
research. A huge number of publications have been
accumulated since the mid-nineties, but the vast majority of
reported papers have employed solid phase methods. In
contrast, the use of conventional solution phase chemistry
for library synthesis has received scant attention. The effort
required to purify the products has been probably the
primary reason for this bias. However, the huge number of
transformations available from extensive solution phase
literatures gives great flexibility in the synthetic design of
the molecules in a library, especially if no subsequent
purification step is required. Herein, we report the adaptation
of the Hantzsch synthesis of 2-aminothiazoles to allow the
automated solution phase preparation and isolation of
discrete 2-aminothiazoles with special emphasis on the
avoidance of any further purification step.
H
S
H
Br
R2
R2
N
N
S
i, ii, iii
R1
R1
+
N
NH2
O
R3
R3
2-Aminothiazole is a commonly occurring structural unit
in drug molecules and their biological activity has been well
documented [1]. Several reviews about the application of 2-
aminothiazoles in the treatment of a diverse range of
different diseases like Alzheimer’s disease [2], chronic pain
[3], cancer [4] and Huntington’s disease [5] appeared in the
scientific literature over the last two years.
Scheme 1. Solution-phase synthesis of 2-aminothiazolesa.
aReagents and conditions: i) THF or dioxane, 55-70°C, 8-48 h; ii)
Et3N; iii) liquid-liquid extraction.
The goal was to synthesize a library of discrete 2-
aminothiazoles as a free base in a fully automated fashion
using the Chemspeed ASW 2000 automated synthesizer
[12]. Since the reaction of ꢀ-haloketones and thioureas
leading to the desired compounds is a robust reaction and
can therefore easily be transferred to automated systems, we
expected the work-up procedure to be by far the more
challenging part of the adaptation. Although we wanted to
perform the reaction in solution, in principle due to the
versatility of the synthesizer - the system allows the use of
solution- and solid-phase reactors - different options for the
isolation of the final products were conceivable. However,
RESULTS AND DISCUSSION
The Hantzsch synthesis of the title compounds is outlined
in Scheme 1. The cyclocondensation of ꢀ-haloketones and
*Address correspondence to this author at the MS-RTC-MedChem DA35,
Merck Serono, Merck KGaA, Frankfurter Strasse 250, D-64293 Darmstadt,
Germany; Tel: +49 6151 72-4092; Fax: +49 6151 72-3129;
E-mail: Hans-Peter.Buchstaller@merck.de
1386-2073/11 $58.00+.00
© 2011 Bentham Science Publishers Ltd.

Parallel Solution-Phase Synthesis of a 2-Aminothiazole Library
Combinatorial Chemistry & High Throughput Screening, 2011, Vol. 14, No. 2 105
after the first set of experiments we quickly realized that our
attempts to isolate the products as hydrobromide salts by
simple filtration using the solid phase reaction blocks would
fail, because either the products did not precipitate
sufficiently, or were difficult to redissolve for the subsequent
extraction step. Therefore, we changed our set-up and
switched to the solution phase reactors. But also with this
equipment the identification of a suitable procedure was not
straightforward. A work-up protocol starting with the
evaporation of the reaction solutions failed because the
residues spattered at the end of the drying process and could
hardly be redissolved without considerable loss of product.
Attempts to dissolve the precipitated products in water or 2N
sodium hydroxide solution followed by an extraction were
unsuccessful, because either a too large volume was required
to completely dissolve some of the residues, or the transfer
needle clogged sometimes during aspiration of the organic
layer after the first extraction. Due to these observations it
was clear that the avoidance of precipitates at any stage of
the work-up procedure is essential. For this reason we tried
to liberate the products from the hydrobromide salts by
adding a slight excess of an organic base, which either could
be removed via extraction or was volatile enough to be
removed from the final products simply by evaporation.
While potassium tert-butylate was a too strong base causing
side reactions in some cases, triethylamine turned out to be
the reagent of choice for the successful release of the
solubility of further building blocks selected for library
synthesis in THF, a certain amount of DMF had to be added
for the preparation of these stock solutions. Moreover,
because of occasional large differences in reactivity of some
of the building blocks, fine tuning of the reaction conditions
was necessary prior to library synthesis. To this end, by
changing the solvent from THF to dioxane, and incubating at
an elevated temperature (70°C), the reaction could be driven
to completion sometimes even in a shorter period of time.
In order to demonstrate the versatility of this procedure,
20 diverse ꢀ-bromoketones 1-20 and thioureas A-T (Fig. 1)
were selected, thus generating a 400-member library. The
preparation of the compounds was performed in the solution
phase reaction vessels (48 vessels, 13 ml volume) equipped
with reflux condensers. Stock solutions of the thioureas as
well as ꢀ-bromoketones were prepared (for information
regarding the applied solvents for each reagent please see
Fig. 1) and aliquots (0.3 mL, 0.15 mmol) of 0.5 M solutions
of both reagents were dispensed into the individual vessels
via the liquid handling tool. The resulting solutions were
shaken and heated at 55-70°C for 8-48 h. Release of the
products from their hydrobromide salt was achieved by
adding a 0.5 M solution of triethylamine in chloroform (0.4
mL, 0.2 mmol). For the subsequent extraction chloroform
(1.5 mL) and water (1 mL) were added and the vessels were
shaken at 50°C for a couple of minutes. The lower layer (1.4
mL) was transferred to pre-weighed vials, the extraction of
the aqueous layer was repeated 2 times (1 mL of chloroform
each) and the organic phase collected in the corresponding
vials. Following the last transfer (1.6 mL) an aliquot of the
product solution was set aside in a microtiter plate for
analyses. The combined product solutions were evaporated
without further drying procedure, yielding 30-50 mg (51-
99%) of the desired compounds. Analyses of all compounds
(Fig. 2) illustrate the high quality of the material afforded by
this fully automated library preparation. Only the reactions
with unsubstituted thiourea A resulted in product mixtures
products. In addition, the dissolution and extraction process
was facilitated by heating the reaction vessels to 50°C. Since
the ASW 2000 synthesizer is not provided with an automatic
boundary detection system, chloroform was used as a
solvent, because the lower layer could be easily separated by
aspiration of
a distinct volume. Using this work-up
procedure the desired 2-aminothiazoles were obtained after
incubation of a 1:1 mixture of substituted thioureas and ꢀ-
bromoketones at 55°C in THF in excellent purities and
yields, at least for those building blocks which were initially
selected for the optimization process. Due to the low
O
O
O
O
O
O
O
O
NO2
O
O
Br
Br
O
Br
Br
F2C
Br
Br
Br
Br
Br
Br
Cl
F
O2N
NC
O
O
O
Br
O
7
8
9
10
3
5
6
1
2
4
O
O
O
O
O
O
O
O
O
O
Cl
N
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
O
O
Cl
F3C
Cl
O
N
H
19
13
14
11
12
15
16
17
18
20
CN
F
Cl
F3C
N
F
S
S
S
S
S
S
S
S
S
S
N
CF3 H2N
N
H
H2N
N
H2N
N
N
H2N
N
H
H2N
N
H
H2N
N
H2N
N
H
H2N
H2N
NH2
H2N
H
H
H
H
H
G
H
I
J
C
E
F
A
B
D
O
CF3
CN
O
O
O
O
S
S
N
S
S
S
S
S
S
S
S
N
O
H2N
N
H
H2N
N
H
N
N
H2N
N
H2N
N
H
H2N
N
H
H2N
H2N
N
H2N
N
H2N
H2N
H
H
H
H
H
H
T
P
Q
R
S
K
L
M
N
O
Fig. (1). ꢀ-Bromoketonesa and thioureasb for a 400-member library. aStock solutions of ꢀ-bromoketones were prepared in THF (1-5, 12, 18,
20), in dioxane (1-15, 17-20), in DMF (16). bStock solutions of thioureas were prepared in THF or dioxane (B, D, G, H, K), in THF/DMF -
2/1 (C, E, I, J), in dioxane/DMF - 2/1 (C, E, G-Q, S, T), in dioxane/DMF - 1/1 (A, F), in DMF/THF - 3/2 (R).

106 Combinatorial Chemistry & High Throughput Screening, 2010, Vol. 14, No. 2
Buchstaller and Anlauf
A
B
C
D
E
F
G
H
I
J
K
L
M
N
O
P
Q
R
S
T
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
ꢀ
ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ ꢀ
Fig. (2). Purities of a 400-member library. HPLC purity (220 nm): ꢀ > 90%, ꢀ 80-90%, ꢀ 70-80%. Equipment: La Chrom (Merck Hitachi)
system (interface L-7000, pump L-7100, auto sampler L-7200, diode array detector L-7450) using a LiChrospher 60 RP-select B (5 ꢁm)
column. Water (0.01% TFA) - acetonitrile (0.008% TFA) were used as eluent in mixtures (unlinear gradient) as follows: 0 min, 20% ACN; 7
min, 100% ACN; 10 min, 100% ACN; 10.5 min, 20% ACN; 12 min, 20% ACN; UV detection: 220 nm. HPLC/MS of the products were
recorded on an Agilent 1100 HPLC system (1100 high pressure gradient pump, 1100 diode array detector) interfaced to an Agilent 1100
mass spectrometer detector using a Chromolith SpeedROD RP 18e50-4.6 column. Water (0.01% TFA) - acetonitrile (0.008% TFA) were
used as eluent in mixtures as follows: 0 min, 20% ACN; 2.8 min, 100% ACN; 3.3 min, 100% ACN; 3.4 min, 20% ACN; 3.6 min, 20% ACN;
UV detection: 220 nm.
1
regardless of the applied conditions. However, these
products could be obtained by an alternative strategy. To this
optimization phase some derivatives were analyzed by H
NMR spectroscopy, and the spectra of the materials were all
in accordance with the desired 2-aminothiazoles (Fig. 3). In
case of derivatives bearing no substituent at position 5, the
exchangeable 5-H signal was characteristic.
end,
a
1:1 mixture of thiourea and substitutedꢀ-
bromoketones was heated in THF at 60°C for 18 h in the
presence of polymer-supported 1,5,7-triazabicyclo[4.4.0]dec-
5-ene (TBD-P) as base [9, 13]. In these cases the solid-phase
reaction vessels were employed and the products were
separated by filtration. It is noteworthy that in the course of
the library production several runs comprising reaction set-
up, reaction and extractive work-up were performed
completely unattended overnight.
The range of functional groups included in the ꢀ-
bromoketone and thiourea substrates for this library
demonstrate the versatility and scope of this procedure.
However, a considerable effort has to be invested in the
evaluation of building blocks prior to library synthesis and
some building blocks were excluded from library production.
The limited solubility in the reaction solvents turned out to
be the major difficulty, because increasing amounts of DMF,
which was required for the preparation of stock solutions of
the substrates, affected the work-up procedure and thus the
purity of the final compounds. In general, in our experience
the elaboration of suitable conditions is the most tedious part
of the adaptation of known reactions to automated synthesis.
HPLC analysis revealed material of high purity (Fig. 2)
and LC coupled mass spectrometry confirmed that in each
case the isolated product showed the expected molecular ion.
Evaporation of the taken aliquots in microtiter plates and
dissolution in ACN has thereby proven to be beneficial
providing chromatograms devoid of peaks from residual
reaction solvents. In addition, in the course of the

Parallel Solution-Phase Synthesis of a 2-Aminothiazole Library
Combinatorial Chemistry & High Throughput Screening, 2011, Vol. 14, No. 2 107
Fig. (3). 250MHz 1H NMR spectrum (d6-DMSO) of library sample 2B (spectra were recorded on a Bruker AM 250 spectrometer).
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Once a protocol is established library preparation proceeds
smoothly with reliable systems.
CONCLUSION
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[7]
In summary, we have developed a straightforward and
effective procedure for the solution phase preparation of
libraries of 2-aminothiazoles using the Chemspeed ASW
2000 automated synthesizer. Reaction, work-up via liquid-
liquid extraction and isolation of the title compounds was
accomplished in a fully automated fashion. The compounds,
each one in the form of its free base, were obtained in good
yields and excellent purities without any further purification
procedure.
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a novel class of muscarinic M3 selective
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Received: November 25, 2009
Revised: October 26, 2010
Accepted: October 27, 2010