An efficient and expeditious Fmoc protection of amines and amino acids in aqueous media

Article abstract of DOI:10.1039/c1gc15868f

A new and environmentally friendly Fmoc protection of a variety of aliphatic and aromatic amines, amino acids, amino alcohols and amino phenols is reported in aqueous media under mild and catalyst-free conditions. The reaction proved to be chemoselective in presence of ambident nucleophiles.

Full text of DOI:10.1039/c1gc15868f

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Green Chemistry
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An efficient and expeditious Fmoc protection of amines and amino acids in
aqueous media†
Manoj B. Gawande* and Paula S. Branco*
Received 18th July 2011, Accepted 23rd September 2011
DOI: 10.1039/c1gc15868f
A new and environmentally friendly Fmoc protection of
a variety of aliphatic and aromatic amines, amino acids,
amino alcohols and amino phenols is reported in aqueous
media under mild and catalyst-free conditions. The reac-
tion proved to be chemoselective in presence of ambident
nucleophiles.
considered a milder substitute to the more classical Boc strategy.8
Also, Fmoc protected amines and amino acids have applicability
in the solid phase synthesis of chiral peptide nucleic acid,9
solid-phase synthesis of hydrazinopeptides,10 the synthesis of
Fmoc protected b-hydroxyl amino acids11 and in many other
circumstances. Thus the synthesis of F-moc protected amines,
amino alcohols and amino acids under catalyst-free conditions
in aqueous media is desirable as the tight legislation on the
maintenance of greenness in synthetic pathways and processes
demand us to prevent waste, avoid use of hazardous solvents (e.g.
halogenated and high-boiling solvents) auxiliary substances,
additional reagents and expensive catalysts, and minimize energy
requirements.12 This is not the case of several reported examples
of Fmoc protection of amines that makes use of acid and base
reagents usually in stoichiometric quantity as also, expensive
reagents or catalysts and organic/toxic solvents.13 There is no
report of catalyst-free Fmoc protection of amines in aqueous
medium. In continuation of our programme for catalyst-free
reactions as well as for environmentally benign protocols for the
synthesis of important raw materials,14 herein, we report a very
simple and highly efficient strategy for the protection of amines
with Fmoc under catalyst-free conditions and in a very friendly
solvent - water.
Introduction
It is widely accredited that there is an increasing importance
for more environmentally viable methods in the fine chemical
synthesis, pharmaceutical industries and small heterocyclic
synthesis. This progressive development known as ‘Green
Chemistry’1 or ‘Sustainable Technology’ necessitates a paradigm
shift from traditional concepts of process efficiency, that focus
largely on yield of product, to one that assigns economic
value, eliminating waste and avoiding the use of toxic and/or
hazardous substances. Organic reactions in water without use
of any harmful organic solvents are of great current interest,
because water is an easily available, economical, safe and
environmentally benign solvent.2,3 In addition the remarkable
properties seen in water as a result of its chemical and physical
properties are very useful for selectivity/reactivity that cannot
be attained in organic solvents and make it an attractive solvent
for many organic reactions, not just for biochemical processes.4,5
In the field of organic chemistry such as peptide, nucleoside,
or combinatorial synthesis, easily accessible amine protecting
groups are frequently required, which are stable under a broad
range of reaction conditions and are easily and selectively
cleavable.6 The 9-fluorenylmethoxycarbonyl (Fmoc) group has
grown extremely in reputation since its introduction as a protect-
ing group for primary and secondary amines and especially for
amino acids. It is employed widely in most contemporary solid
and solution phase peptide synthesis due to its stability to acid
and liability to base, being the chloroformate ester (Fmoc-Cl)
traditionally employed for its incorporation.7 Fmoc when used
in conjunction with acid-labile side-chain protecting groups is
Results and discussion
We began our investigation with a model reaction of aniline
(1a) with 9-fluorenylmethoxycarbonyl chloride (2) (1 : 1.2 mmol)
in water (1.5 mL) under catalyst-free conditions to yield N-(9-
fluorenylmethoxycarbonyl)aniline (3a) (Scheme 1).‡
REQUIMTE, Departamento de Qu´ımica, Faculdade de Cieˆncias e
Tecnologia, FCT, Universidade Nova de Lisboa, Portugal.
E-mail: mbgawande@yahoo.co.in, psb@dq.fct.unl.pt
† Electronic supplementary information (ESI) available. See DOI:
10.1039/c1gc15868f
Scheme 1 Synthesis of N-(9-fluorenyl mehoxycabonyl)aniline (3a).
The reaction was completed in 2 h at 60 ◦C and the
corresponding Fmoc-protected amine obtained in 90% yield. We
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have studied the Fmoc protection of aniline in distilled water as
well as in tap water. No considerable difference was observed
in terms of yield. Paying attention to the need to develop
economical sustainable methods we carried out all reactions
in tap water. To understand the role of water on the Fmoc
protection of amines, we have tested the reaction of aniline with
Fmoc chloride in solvent-free conditions at 60 ◦C. It should
be noted that in solvent-free conditions and after 2 h, 3a was
obtained in 20% yield whereas after 24 h it was observed to
increase to only 40%. All the attempts to perform the reaction at
room temperature (in solvent- and catalyst-free conditions) were
fruitless even after 48 h. The aqueous media showed a significant
improvement of yield as well as reaction time when compared
to the solvent-free reaction (Fig. 1).
20 min to 4 h. Note that a reduction in the amount of Fmoc
reagent led to a decrease in the yield of the reaction (Table 1,
entries a, g, l, and v). Excellent chemoselectivity was observed
in case of 3-amino phenol, ethanol amine, and dopamine
chloride (Table 1, entry c, p and s respectively), no bis Fmoc
derivatives or mixture of O/N Fmoc-derivatives was observed.
After successful completion of Fmoc protection of amines and
in view of their importance in solid phase synthesis as well
as in nucleoside, or combinatorial synthesis we have further
elaborated this study for the development of Fmoc protection
of important amino acid which are useful in peptide synthesis
and other some important organic reactions (Table 1, entry t, u
and v).15 The integrity of the stereocenter for the chiral N-Fmoc
a-amino acids is maintained as determined by HPLC (less than
1% of racemization, Table 1, entries u and v).
A proposed mechanism for the Fmoc protection of amines
in aqueous media is presented in Scheme 2. Water as a solvent
can provide polar and other interactions needed to stabilize
the transition state without causing a rate penalty because of
dilution by solvent.3,16 Due to hydrogen bonding established
by the molecule of water (Scheme 2, structure 4 and 5) the two
compounds, the chloroformate ester and the amine will be in the
vicinity of each other and the outcome will be an enhancement
of the electrophilicity of the carbonyl group and increased
nucleophilicity of the amine nitrogen. This double character that
water can endeavour during the catalyst-free Fmoc protection
of amines has already been proposed by Chankeshwara and
Chakraborti on the N-tert-butyloxycarbonylation of amines.3
Fig. 1 Kinetic curve of Fmoc protection of aniline in water (red line)
and in solvent-free conditions (blue line) at 60 ◦C.
Also the amount of water is crucial for the elapse of the
reaction. When using 0.5 mL of water, 3a was obtained in 42%
after 2 h of reaction. Increasing the amount of water to 1.0 mL
the yield slightly increased to 55%. The best result was attained
in the presence of 1.5 mL (90%, Fig. 2).
Fig. 2 Effect of the quantity of water on the yield of N-(9-
fluorenylmethoxycarbonyl)aniline (3a).
Scheme 2 Proposed mechanism for the Fmoc protection of amines.
As expected the reaction is faster with aliphatic amines due
to their better nucleophilicity (Table 1, entries m to q) although
secondary cyclic amines (entries j to l) being stronger bases gave
a slightly faster reaction which results from stronger hydrogen
bonds. Here we can’t rule out that the hydrophobic effect which is
the tendency of nonpolar species to aggregate in water solution
so as to decrease the hydrocarbon-water interfacial area can
play its part. Some rather simple organic reactions can show
hydrophobic effects when they are carried out in water solution
Further increasing the quantity of water had the opposite
effect and the yield of product decreased which could be
related to the dispersion of the reagents, when large quantity
of water is used. With the established optimal conditions, the
scope of the reaction was probed. As highlighted in Table 1
the Fmoc protection of various amines (aromatic, aliphatic,
cyclic and heterocyclic), amino alcohols and amino phenols
were carried out in excellent yields and reduced time, between
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Table 1 Catalyst-free Fmoc protection of amines in aqueous mediaa
Entry
a
Amine and amino acids substrate (1)
Time (h)
2
3, Isolated yield (%)
3a 90
77d
b
c
d
e
f
R = 4-Me
R= 3-OH
R = 3-Cl
R = 4-Br
R = 4-F
2
3b 83
3c 81
3d 87
3e 80
3f 84
3g 88
2.5
2.5
1
1
1
g
70d
3h 87
3i 80
3j 80b
h
i
j
R = 4-Me
R = 4-OMe
0.5
0.5
0.5
k
l
0.33
0.33
3k 82b
3l 84b
68d
m
n
1
3m 80b
0.5
3n 90b
o
p
1
3o 82
0.5
3p 92b
q
r
1
3
3q 87
3r 80
s
4
3s 75
t
4
4
3t 85c
u
3u 86c,e
v
1
3v 89c,f
72d
a Reaction conditions: Amine (1 mmol), FmocCl (1.2 mmol), water 1.5 mL, 60 ◦C. b Reaction carried out at room temperature. c Solvent, water:ethanol
(3 : 1). d 1 mmol of FmocCl used for reaction. HPLC analysis of 3u and 3v on Chiralpak-IA, mobile phase 10% ispropanol/hexane (v/v) containing
0.1% TFA; flow rate = 1 mL min-1; detector 254 nm; e Enantiomeric purity (D:L = 1 : 99); f Enantiomeric purity (D:L = 0.5 : 99.5).
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Table 2 Comparison of the synthesis of N-(9-fluorenylmethoxy-carbonyl)aniline with reported methods
Entry
Solvent and conditions
70 ◦C, Toluene18
Time (h)
Isolated yield (%)
E-factor
Mass intensity
1
2
3
4
5
5
24
1
7
2
78
68
94
90
90
14.34
527.54
0.5
14.2
5.05
16.81
666.00
4.6
150.56
6.35
19
RT, CH2Cl2
RT, Benzene or reflux, phenyl isocyanate17
23 ◦C, pyridine, and THF20
No catalyst, 60 ◦C, H2O
RT – room temperature
if nonpolar segments of the reactants are brought together in the
transition states.4 A slower reaction for the phenylethylamine
substrates (entries r and s) can be explained due to unfavourable
interactions that destabilize the transition state (5). To under-
stand the efficiency of our protocol with respect to others for the
synthesis of N-(9-fluorenylmethoxycarbonyl) aniline, the results
are collected and depicted in Table 2. The results clearly indicates
that the catalyst-free Fmoc protection of aniline in aqueous
media reported here is the most efficient, clean and green process
(Table 2, entry 5). The method reported by Carpino and Han17
(Table 2, entry 3) also shows a comparable yield and also related
E-factor and mass intensity for N-(9-fluorenylmethoxycarbonyl)
aniline but the main drawback of this method is related to the
use of benzene as solvent or phenyl isocyanate under reflux
conditions. For specific reaction conditions see corresponding
literature and ESI† for calculation.
Fmoc-amino acids. Enantiomeric purity was obtained by HPLC analysis
(see ESI†).
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Conclusions
In conclusion, a very simple methodology for the catalyst-
free Fmoc protection of amines is reported. The method
is highly efficient and environmentally benign owing to the
mild conditions, high yields, excellent chemoselectivity, ease of
product isolation and purification which fulfil the criterion for a
green chemistry practice. This methodology can be applicable for
the protection of amino acids, a crucial step in solution and/or
solid phase synthesis as well as in the synthesis of important
drug intermediates.
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Acknowledgements
Manoj B. Gawande thanks FCT for the award of postdoctoral
grant SFRH/BPD/64934/2009 and financial assistance.
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Notes and references
‡ General procedure for the Fmoc protection of amines: To Fmoc chloride
(1.2 mmol) was added the amine (1 mmol) and water 1.5 mL and the
reaction mixture stirred at 60 ◦C. The reaction was monitored by thin
layer chromatography using ethyl acetate and hexane as eluent (3 : 7).
After consumption of the amine the reaction product was filtered,
washed with water and recrystallized from hot ethanol to afford the
pure product.
General procedure for the Fmoc protection of amino acids To the mixture
of the two solids the amino acid (1 mmol) and Fmoc chloride (1.2 mmol)
it was added 1.5 mL of water:ethanol (3 : 1). The reaction mixture
was stirred at 60 ◦C for particular time. After completion of reaction
monitored by TLC the solution was acidified with HCl (1 M) untill
pH 4–5 at O ◦C and extracted with EtOAc (3 X10 mL). The combined
organic layers were dried (Na2SO4) and evaporated affording the pure
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