An improved synthesis of Fmoc-N-methyl serine and threonine

Article abstract of DOI:10.1016/j.tetlet.2007.05.112

An improved method for the synthesis of Fmoc-N-methyl serine and threonine has been developed, which involves formation and subsequent reduction of the corresponding oxazolidinone with a Lewis acid under mild conditions, with improved yields and shorter reaction times.

Full text of DOI:10.1016/j.tetlet.2007.05.112

Tetrahedron Letters 48 (2007) 5003–5005
I
An improved synthesis of Fmoc-N-methyl serine and threonine
*
Rajesh H. Bahekar, Pradip A. Jadav, Dipam N. Patel, Vijay M. Prajapati, Arun A. Gupta,
Mukul R. Jain and Pankaj R. Patel
Zydus Research Center, Cadila Healthcare Ltd., Sarkhej-Bavala N.H. 8A, Moraiya, Ahmedabad 382 210, India
Received 12 April 2007; revised 7 May 2007; accepted 18 May 2007
Available online 24 May 2007
Abstract—An improved method for the synthesis of Fmoc-N-methyl serine and threonine has been developed, which involves for-
mation and subsequent reduction of the corresponding oxazolidinone with a Lewis acid under mild conditions, with improved yields
and shorter reaction times.
ꢀ
2007 Elsevier Ltd. All rights reserved.
1
. Introduction
sis of oxazolidinones from N-protected amino acids and
paraformaldehyde was originally devised by Ben-
Ishai. Subsequently, Freidinger et al. developed a
1
13
Structural conformations and biological activity of syn-
thetic peptides change drastically with the incorporation
of N-methylated amino acids. Peptides containing N-
methylated amide bonds often exhibit higher proteolytic
stability, increased membrane permeability, enhanced
duration of action and offer conformational rigidity.
method for reducing oxazolidinones with Et SiH and
3
1
4,15
TFA.
Based upon this method, Luo et al. reported
the preparation of N-Cbz and N-Fmoc N-methyl serine
2
3
and threonine with TBDMS protection of the side-chain
4
,5
16
alcohol. The (O-TBDMS) N-protected serine and
A review by Fairlie et al. discusses many aspects of the
biological activity of peptides including several examples
of N-methylation in natural products and therapeutic
threonine were reacted with paraformaldehyde and
reduction of the corresponding oxazolidinone gave the
desired N-methyl derivative with concomitant removal
of the TBDMS protecting group. However, introduction
of the TBDMS protecting group to Cbz serine or threo-
nine required 2–3 days and further reduction of the oxa-
6
agents. N-Methylated peptides are generally synthe-
sized by incorporation of Boc- or Fmoc-protected N-
methylated amino acids, either by solution or solid
phase peptide synthesis approaches.
zolidinones with Et SiH/TFA required 3–5 days stirring
3
at rt. Also, this method resulted in a partial loss of the
Fmoc protecting group, which had to be reintroduced.
Various methods have been developed for the synthesis
of optically active N-methyl amino acids, both with
7
,8
functionalized and unfunctionalized side chains.
Aurelio et al. reported the synthesis of N-benzyloxycar-
bonyl protected N-methyl serine and threonine along
with other naturally occurring a-amino acids through
However, very few procedures exist for the synthesis
of N-methylated b-hydroxy amino acids (Ser and Thr),
which mainly give dehydrated derivatives, b-eliminated
by-products or products which undergo partial racemi-
One of the mildest and most general proce-
dures for the synthesis of N-methyl amino acids is
1
7
5-oxazolidinone reduction. The b-hydroxyl group of
the Ser or Thr side chain was protected by O-acetyl-
ation. Furthermore, reductive cleavage with TFA gave
the benzyloxycarbonyl protected N-methyl serine/threo-
nine. This method has been highly successful in generat-
ing novel N-benzyloxycarbonyl protected N-methyl
amino acid derivatives. However, reductive cleavage
with TFA again required a long reaction time.
9
–12
zation.
based on the reduction of 5-oxazolidinones. The synthe-
Keywords: Fmoc-N-methyl serine; Fmoc-N-methyl threonine; Oxazoli-
dinones; Lewis acid; b-hydroxy amino acids.
q
As a result of these limitations, Zhang et al. developed a
more efficient method for the reduction of the intermedi-
ate Fmoc-oxazolidinones of non-functionalized and
ZRC communication No. 211.
*
Corresponding author. Tel.: +91 2717 250801; fax: +91 2717
50606; e-mail: rajeshbahekar@zyduscadila.com
2
0040-4039/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2007.05.112

5004
R. H. Bahekar et al. / Tetrahedron Letters 48 (2007) 5003–5005
O
O
R
N
O
R
O
i
Fmoc
Fmoc
O
N
H
COOH
O
1
1
a R=H
b R=CH3
2
a: 92%
2b: 92%
ii
O
R
OH
R
N
O
iii
Fmoc
N
COOH
Fmoc
COOH
4
4
a: 88%
b: 88%
3a: 90%
3b: 91%
Scheme 1. Reagents and conditions: (i) paraformaldehyde, p-TsOH, toluene, 1 h reflux; (ii) AlCl
2 M), 60 ꢁC, 12 h heating.
3 3
, (iPr) SiH, DCM, 25 ꢁC, 4 h; (iii) dioxane and HCl
(
functionalized side chain-containing amino acids, using
was achieved using AlCl and triisopropylsilane ((iPr) -
3 3
1
8
a Lewis acid catalyst such as AlCl or ZnBr . Further,
SiH) giving Fmoc-N-methyl-O-acetyl amino acids 3a–
b. Finally, deprotection of the hydroxyl group was
achieved using 2 M HCl giving Fmoc-N-methylated
3
2
it was reported that the AlCl reaction needed 4 h for
3
completion whilst that with ZnBr required 22 h, with
2
2
2
negligible side product formation. However, when we
attempted this reaction for the synthesis of Fmoc-N-
methyl-Ser(OH)–OH and Fmoc-N-methyl-Thr(OH)–
OH without side chain protection, no N-methylated
product was obtained using either AlCl3 or ZnBr2.
Chruma reported the synthesis of N-methyl amino acids,
including Ser and Thr, by reductive amination of
b-hydroxyl amino acids 4a–b in very good yields.
Typically, the reaction time for the formation and sub-
sequent reduction of the corresponding oxazolidinone
was less than 1 h and the crude product obtained after
work-up was sufficiently pure to be used in solid-phase
peptide synthesis. Using this protocol, 500 g of both
Ser and Thr Fmoc-protected N-methylated b-hydroxyl
amino acids were prepared in good yields.
O’Donnell’s Schiff base amino esters with NaBH CN
3
1
9
and the appropriate aldehyde in THF. This method
was found to be inadequate for the large scale produc-
tion of N-methylated Ser and Thr, due to the difficulty
in preparing the Schiff bases of the parent amino acids.
In conclusion, Fmoc N-methylated b-hydroxyl amino
acids were prepared in excellent yields. This method al-
lows large-scale preparation of Fmoc-protected N-meth-
ylated b-hydroxyl amino acids, with simple extraction
yielding sufficiently pure compound. This process uti-
lizes less expensive reagents and offers a shorter route
for the synthesis of Fmoc-protected N-methyl-Ser and
Thr amino acid derivatives.
In the present Letter we report an efficient method for
the synthesis of Fmoc-N-methyl serine and threonine.
Among various functionalized amino acids, efforts were
directed towards the synthesis of Fmoc-N-methyl serine
and threonine because, in general, N-methylation of
b-hydroxy amino acids is difficult, mainly due to b-elim-
ination or dehydration and also, Fmoc-protected
N-methylated b-hydroxyl amino acids can be easily
utilized for solution or solid phase peptide synthesis.
2. General experimental procedures
Synthesis of oxazolidinone 2a: Compound 1a (100 g,
0.271 mol), paraformaldehyde (50.0 g) and p-TsOH
(2.79 g, 0.06 equiv) were suspended in anhydrous tolu-
ene (5.5 L) and slowly heated to dissolve most of
the starting material. The reaction mixture was then
refluxed for 1 h using a Dean–Stark apparatus. The
As shown in Scheme 1, the N-Fmoc-protected O-acetyl-
2
0,21
amino acids 1a–b
were refluxed with paraformalde-
hyde and a catalytic amount of p-TsOH in anhydrous
toluene to give the corresponding oxazolidinones 2a–b
in very good yields. Reduction of the oxazolidinone

R. H. Bahekar et al. / Tetrahedron Letters 48 (2007) 5003–5005
5005
reaction was cooled to rt and washed with 5% NaHCO3
and brine, dried over Na SO and evaporated to yield 2a
Acknowledgements
2
4
1
6
1
as a colourless oil (94.9 g, 92% yield).
H NMR
We are grateful to the management of the Zydus Group
for encouragement. We thank the analytical department
for providing analytical support.
(
(
(
CDCl , 300 MHz): d 7.76 (d, 2H, J = 7.17 Hz), 7.54
d, 2H, J = 7.31 Hz), 7.39 (t, 2H, J = 7.15 Hz), 7.30
t, 2H, J = 7.33 Hz), 5.22–5.10 (m, 2H), 4.83–4.81 (m,
3
1
1
1
H), 4.49–4.42 (m, 2H), 4.24 (br s, 2H), 4.04–3.85 (m,
H), 2.01 (s, 3H); IR (Nujol): 3068, 1714, 1421, 1355,
109, 758 cm ; FABMS: m/z 382 [M+1] ; Anal. Calcd
References and notes
À1
+
for C H NO : C, 66.13; H, 5.02; N, 3.67%. Found: C,
1. Vitoux, B.; Aubry, A.; Cung, M. T.; Marraud, M. Int. J.
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2
1
19
6
6
6.21; H, 5.03; N, 3.69%.
2
Synthesis of Fmoc-N-methyl-O-acetyl amino acid 3a: To
a solution of Fmoc-protected oxazolidinone 2a (90.0 g,
0
.23 mol) and anhydrous AlCl3 (62.8 g, 0.47 mol) in
3
dry DCM (2.7 L) was added (iPr) SiH (96.5 mL,
3
0
.47 mol). The reaction mixture was stirred at ambient
temperature until TLC showed the absence of starting
4. Ostresh, J. M.; Husar, G. M.; Blondelle, S.; Dorner, B.;
Weber, P. A.; Houghten, R. A. Proc. Natl. Acad. Sci.
U.S.A. 1994, 91, 11138.
material (ꢀ1 h). An additional amount of DCM
(
2.7 L) was added and the organic phase was washed
5
. Miller, S. M.; Simon, R. J.; Ng, S.; Zuckermann, R. N.;
with 1 M HCl (5.0 L). The organic phase was dried over
anhydrous Na SO and concentrated under vacuum to
Kerr, J. M.; Moos, W. H. Drug. Dev. Res. 1995, 35, 20.
6. Fairlie, D. P.; Abbenante, G.; March, D. R. Curr. Med.
Chem. 1995, 2, 654.
2
4
1
9 1
afford pure 3a (81.4 g, 90% yield). H NMR (CDCl3,
300 MHz): d 9.63 (br s, 1H), 7.72 (d, 2H, J = 7.21 Hz),
7.51 (d, 2H, J = 7.23 Hz), 7.36 (t, 2H, J = 7.24 Hz),
7.29 (t, 2H, J = 7.23 Hz), 5.21–5.13 (m, 2H), 4.83–4.82
7
. Aurelio, L.; Brownlee, R. T. C.; Hughes, A. B. Chem. Rev.
004, 104, 5823.
2
8
. Freidinger, R. M. U.S. Patent 4,535,167, 1985; Chem.
Abstr. 1985, 104, 19819.
(
m, 1H), 4.49–4.42 (m, 2H), 4.04–3.85 (m, 1H), 3.02 (s,
3
1
H), 1.98 (s, 3H); IR (KBr): 2927, 1753, 1651, 1448,
110, 761 cm ; FABMS: m/z 384 [M+1] ; Anal. Calcd
9. Cheung, S. T.; Benoiton, N. L. Can. J. Chem. 1977, 55,
916.
10. Cheung, S. T.; Benoiton, N. L. Can. J. Chem. 1977, 55,
À1
+
for C H NO : C, 65.79; H, 5.52; N, 3.65%. Found: C,
2
1
21
6
9
06.
6
6.68; H, 5.53; N, 3.61%.
1
1
1. Olsen, R. K. J. Org. Chem. 1970, 35, 1912.
2. McDermott, J. R.; Benoiton, N. L. Can. J. Chem. 1973,
Synthesis of Fmoc-N-methyl amino acid 4a: Compound
a (40.0 g, 0.10 mol) was suspended in a mixture of diox-
5
1, 2555.
3
1
1
3. Ben-Ishai, D. J. Am. Chem. Soc. 1957, 79, 5736.
4. Freidinger, R. M.; Hinkle, J. S.; Perlow, D. S.; Arison, B.
H. J. Org. Chem. 1983, 48, 77.
ane and 2 M HCl (800 mL, 1:1) with stirring. The mix-
ture was then heated to 60 ꢁC for 12 h. After cooling,
the reaction mixture was diluted with water (4.0 L)
and extracted with ether (3 · 800 mL). The combined
organic phases were dried (Na SO ), filtered and evapo-
1
5. Aurelio, L.; Brownlee, R. T. C.; Hughes, A. B.; Sleebs, B.
E. Aust. J. Chem. 2000, 53, 425.
16. Luo, Y.; Evindar, G.; Fishlock, D.; Lajoie, G. A.
2
4
rated under reduced pressure to give 4a, which was iden-
Tetrahedron Lett. 2001, 42, 3807.
7. Aurelio, L.; Box, J. S.; Brownlee, R. T. C.; Hughes, A. B.;
Sleebs, M. M. J. Org. Chem. 2003, 68, 2652.
8. Zhang, S.; Govender, T.; Norstrom, T.; Arvidsson, P. I. J.
Org. Chem. 2005, 70, 6918.
1
1
1
tical in all respects with previously reported material
31.3 g, 88% yield).1 H NMR (CDCl , 300 MHz): d
8 1
(
3
9
7
.67 (br s, 1H), 8.24 (br s, 1H), 7.71 (d, 2H, J =
.18 Hz), 7.49 (d, 2H, J = 7.21 Hz), 7.38 (t, 2H,
9. Chruma, J. J.; Sames, D.; Polt, R. Tetrahedron Lett. 1997,
J = 7.16 Hz), 7.34 (t, 2H, J = 7.24 Hz), 4.51–4.48 (m,
3
8, 5085.
3
3
7
H), 4.46–4.42 (m, 2H), 4.04–3.85 (m, 1H), 3.07 (s,
H); IR (KBr): 3630, 2820, 1749, 1657, 1452, 1046,
57 cm ; FABMS: m/z 342 [M+1] ; Anal. Calcd for
2
2
0. Wilchek, M.; Patchornik, A. J. Org. Chem. 1964, 29, 1629.
1. Carpino, L. A.; Han, G. Y. J. Org. Chem. 1972, 37, 3404.
À1
+
22. The identity and the purity of the reaction products were
1
C H NO : C, 66.85; H, 5.61; N, 4.10%. Found: C,
established from their spectral ( H NMR, IR and MS)
1
9
19
5
6
6.81; H, 5.59; N, 4.08%.
data and by elemental analysis.