Utilization of 2-(4-Nitrophenylsulfonyl)ethoxycarbonyl (Nsc) as a substitute for 9H-fluoren-9-ylmethoxycarbonyl (Fmoc) in liquid phase chemistry

Article abstract of DOI:10.1002/adsc.200404016

2-(4-Nitrophenylsulfonyl)ethoxycarbonyl (Nsc) is a useful substitute for the Fmoc group. It is easily removed not only with secondary amines but with tris(aminoethyl)amine (TAEA) and with resin-bound TAEA, thus allowing for a simplified work-up: the side products of the deprotection are removed either by extraction with phosphate buffer or by filtration.

Full text of DOI:10.1002/adsc.200404016

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Utilization of 2-(4-Nitrophenylsulfonyl)ethoxycarbonyl (Nsc) as a
Substitute for 9H-Fluoren-9-ylmethoxycarbonyl (Fmoc) in Liquid
Phase Chemistry
Thomas C. Maier, Joachim Podlech*
Institut f¸r Organische Chemie der Universit‰t Karlsruhe (TH), Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany
Fax: (þ49)-721-608-7652, e-mail: joachim.podlech@ioc.uka.de
Received: January 16, 2004; Accepted: May 7, 2004
th
Dedicated to Joe Richmond on the occasion of his 60 birthday.
Abstract: 2-(4-Nitrophenylsulfonyl)ethoxycarbonyl
Nsc) is a useful substitute for the Fmoc group. It
substrate was neither possible with conventional nor
with reverse-phase chromatography and purification
by crystallization led to heavy loss of material. The
DBFoligomer is hardly removable even by repeated fil-
trations and the amine adduct is not separable by extrac-
tion since it shows similar basicity as compared to the de-
protected product. As Carpino et al. have shown, the
latter problem can be faced by the use of tris(amino-
ethyl)amine (TAEA) as cleaving base (50 equivs.,
1.5 M in CH Cl ) leading to an amine adduct which
(
is easily removed not only with secondary amines
but with tris(aminoethyl)amine (TAEA) and with
resin-bound TAEA, thus allowing for a simplified
work-up: the side products of the deprotection are
removed either by extraction with phosphate buffer
or by filtration.
Keywords: amino acids; polymer-bound Lewis bases;
2
2
protecting groups
can be effectively extracted with phosphate buffer
[8]
(
pH 5.5). Nevertheless, with this, the formation of
DBF oligomers is still a significant obstacle.
We found that the 2-(4-nitrophenylsulfonyl)ethoxy-
[
1]
First reported by Carpino and Han in 1970, the 9H-flu- carbonyl group (Nsc) which has been occasionally
[
5d,9]
oren-9-ylmethoxycarbonyl group (Fmoc) has become used in solid-phase peptide synthesis
and in nucleo-
[
10]
one of the most frequently used a-amino protecting side chemistry is especially useful for peptide syn-
groups in solid-phase peptide synthesis. Its usefulness thesis in solution. Nsc-protected amino acids are com-
was immediately recognized with the observation that mercially available or easily prepared in four steps
[
6,11,12]
Fmoc cleavage is readily achieved by amines like piper- from 4-nitrochlorobenzene.
Like Fmoc, the Nsc
[2]
a
idine. The real break-through of this type of N -protec- group is cleaved under basic conditions via an E1cb
tion occurred with its application in solid-phase syn- mechanism, but ± unlike dibenzofulvene ± the primary
[
3]
thesis by Chang and Meienhofer and Atherton product of Nsc cleavage, 4-nitrophenyl vinyl sulfone,
[4]
et al. The relatively mild, versatile deprotection proce- does not tend to polymerize and is trapped virtually irre-
dure with amines is operationally simple, liberating the versibly by piperidine and other amines (Scheme 1).
free amino function and the by-product dibenzofulvene
We tested the applicability of the Nsc group by inves-
(
DBF), which is easily removed from the solid support tigating the behavior of the dipeptide Nsc-Leu-Phe-
by repeated washings. The chemistry and usefulness of OMe under varying reaction conditions: We could con-
this protecting group have been extensively re- firm that this protecting group is cleavable by mild bases
viewed.
[5,6]
While in solid phase synthesis using base- (Table 1) with no formation of interfering polymeric
promoted Fmoc cleavage, DBF and its amine-adducts material and with irreversible formation of the vinyl sul-
are simply removed by filtration and washings, the fone-amine adduct. When TAEA was used as cleaving
lack of volatility and the reactivity of the DBF by-prod- base only a small excess was necessary to achieve com-
uct cause difficulties in solution phase synthesis render- plete deprotection. With three equivalents of TAEA
ing the Fmoc group less attractive for solution synthesis only small traces of starting material were left (Entry 4)
[6,7]
(
Scheme 1). This communication summarizes our ef- while full conversion was achieved with higher excess of
forts in solving this specific problem. the amine (Entry 3). Excess base and the vinyl sulfone-
During deprotection studies of Fmoc-protected sub- amine adduct could be removed by washing with phos-
strate 1 we encountered significant problems arising phate buffer. Additionally we tested a solid phase-sup-
[
13]
from the side products DBF, its gelatinous oligomer ported TAEA (¾±TAEA) as a cleaving base which
and the respective amine adducts. A purification of the is commercially available but has not previously been
Adv. Synth. Catal. 2004, 346, 727±730
DOI: 10.1002/adsc.200404016
¹ 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
727

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Thomas C. Maier, Joachim Podlech
Table 1. Deprotection of an Nsc-protected dipeptide derivative.
Entry
Base
Equivalents (concentration)
Solvent
CH Cl
Time
12 h
0.5 h
0.5 h
15 h
15 h
15 h
50 h
Result
1
2
3
4
5
6
7
Et NH
50 (1.0 M)
50 (1.5 M)
50 (1.5 M)
3 (0.06 M)
2 (0.04 M)
1 (0.02 M)
8 (ꢀ)
complete conversion
complete conversion
complete conversion
traces of starting material
starting material left
starting material left
complete conversion
2
2
2
2
TAEA
TAEA
TAEA
TAEA
TAEA
¾-TAEA
CH Cl
2
MeOH
CH Cl
2
2
2
2
2
CH Cl
2
CH Cl
2
CH Cl
2
Nsc-protected substrates bearing a proline moiety as
N-terminal amino acid could in our case be deprotected
quantitatively with TAEA within a few minutes (En-
try 1). However, even after prolonged reaction times
(
95 h) with polymer-supported TAEA deprotection
was not complete, though only traces of starting material
could be detected by TLC (Entries 6 and 7). It is very
likely that the required reaction times and/or the re-
quired excess of immobilized TAEA strongly depend
on the quality/age of the cleaving agent. We tested ami-
nomethyl-polystyrene resin (¾-CH NH ) as a some-
2
2
what cheaper reagent but found that cleavage rates
were very low with this reagent (Entry 8).
In conclusion, the Nsc group together with the herein
presented deprotection methods (TAEA and supported
TAEA, respectively) fulfills perfectly the prerequisites
for a useful protecting group in liquid peptide synthesis.
It is simply introduced, it is resistant towards usually ap-
plied reaction conditions and it is removed without
traces simply by washing or filtration procedures. With
these features it is similarly useful as the acid-labile
tert-butoxycarbonyl group (Boc) to which it is complete-
ly orthogonal. Since Nsc-protected amino acids are now
commercially available, this protecting group is a useful
substitute for Fmoc and other groups in specific cases.
Scheme 1.
used as cleaving base for this or similar protecting
groups to the best of our knowledge. It was similarly ap-
plied and after use separated together with the corre-
sponding vinyl sulfone adduct (irreversibly bound to
the solid phase) simply by filtration. An excess of about
eight equivalents led to a complete deprotection of the
Nsc group (Entry 7). Both methods led to deprotected
Experimental Section
products which contained no significant amounts of im- General Remarks
purities.
Nsc-protected amino acids were purchased from Hyundai
Pharm (Bucheon, Korea) or were prepared from 2-(nitrophe-
We applied comparable reaction conditions to more
complex substrates and found that Nsc is again a useful
alternative to the Fmoc group. Proline, a secondary
amine is ± mostly, but not only for reasons of steric hin-
[6,11]
nyl)sulfonyl]ethyl chloroformate (Nsc-Cl)
by standard pro-
Tris(2-aminoethyl)amine (TAEA, Aldrich),
tris(2-aminoethyl)amine resin (¾-TAEA, Advanced Chem-
[6,11]
cedures.
drance ± usually more difficult to deprotect, which often Tech) and aminomethyl-polystyrene resin (¾-CH NH , Bach-
2
2
results in a somewhat longer reaction time (Table 2). em) were commercially available.
728
¹ 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
Adv. Synth. Catal. 2004, 346, 727±730

2-(4-Nitrophenylsulfonyl)ethoxycarbonyl (Nsc) as a Substitute for Fmoc
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Table 2. Deprotection of peptidomimetics bearing an N-terminal praline.
Entry
Substrate
Base
Equivalents (concentration)
Solvent
CH Cl
Time
Result
1
2
3
4
5
6
7
8
3
3
2
2
2
2
2
2
TAEA
TAEA
50 (1.5 M)
20 (0.6 M)
30 (0.2 M)
1.1 (0.02 M)
8 (ꢀ)
0.2 h
0.5 h
15 h
50 h
50 h
95 h
95 h
80 h
complete conversion
complete conversion
complete conversion
starting material left
2
2
2
2
CH Cl
2
Et NH
CH Cl
2
2
piperidine
¾-TAEA
¾-TAEA
¾-TAEA
THF
CH Cl
starting material left
2
2
2
15 (ꢀ)
CH Cl
traces of starting material
traces of starting material
essentially starting material
2
15 (ꢀ)
MeCN
CH Cl
2
¾-CH NH
8 (ꢀ)
2
2
2
General Procedure for the Deprotection of Fmoc- or References andNotes
Nsc-Protected Amino Acids or Oligopeptides with
TAEA
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5
To the compound to be deprotected (0.50 mmol) in CH Cl
2
2
[
[
[
(
5 mL) TAEA (25.0 mmol) was added at room temperature
3
and the solution was stirred for about 10 min (as monitored
by TLC). The mixture was diluted with CH Cl , washed succes-
2
2
1
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2
4
2
4
2
2
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porator (408C, 100 mbar). The resulting product is usually
pure enough for further reactions.
4
5
37±539.
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[
4
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(
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2
2
(
0
10 mL) was added to polymer-bound TAEA (1 g, loading ca.
.70 mmol/g) and stirred at room temperature. The reaction
was monitored by TLC; after about 70 h no starting material
was detected (reaction time depends on the excess of immobi-
lized TAEA and on its quality). The solid material was re-
moved via a glass sinter and the solvents were removed at
the rotary evaporator. The resulting product is usually pure
enough for further reactions.
[
[
7] L. Andersson, L. Blomberg, M. Flegel, L. Lepsa, B. Nils-
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Adv. Synth. Catal. 2004, 346, 727±730
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¹ 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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730
¹ 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
Adv. Synth. Catal. 2004, 346, 727±730