Efficient S-alkylation of cysteine in the presence of 1,1,3,3- tetramethylguanidine

Article abstract of DOI:10.1016/j.tetlet.2010.08.097

The synthesis of S-alkylated cysteine derivatives was carried out successfully in the presence of 1,1,3,3-tetramethylguanidine. Alkylation proceeded in high yields on unprotected amino acids and peptides containing a sulfhydryl group.

Full text of DOI:10.1016/j.tetlet.2010.08.097

Tetrahedron Letters 51 (2010) 5977–5979
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Efficient S-alkylation of cysteine in the presence of 1,1,3,3-tetramethylguanidine
⇑
Marek Włostowski , Sylwia Czarnocka, Piotr Maciejewski
Laboratory of Technological Processes, Faculty of Chemistry, Warsaw University of Technology, ul. Noakowskiego 3, 00-664 Warsaw, Poland
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis of S-alkylated cysteine derivatives was carried out successfully in the presence of 1,1,3,3-
tetramethylguanidine. Alkylation proceeded in high yields on unprotected amino acids and peptides con-
taining a sulfhydryl group.
Received 21 April 2010
Revised 19 August 2010
Accepted 31 August 2010
Available online 9 September 2010
Ó 2010 Elsevier Ltd. All rights reserved.
The isoprene unit located at the C-terminal of cysteine has been
found to be crucial for membrane association and transforming the
activity of oncogenic ras proteins.1–5 The need for this post-trans-
1. 2 equiv TMG
2. RX 2
3. H+
SH
S
R
HO (R)
HO (R)
NH2
NH2
O
O
lational modification of L-cysteine residue, the necessity for alkyl-
1
3
ation of sulfhydryl groups in peptide mapping and protein
identification,6 and the wide spectrum of biological activity of mol-
ecules with a cysteine moiety in their structure7–11 have inspired
research on the S-alkylation problem in cysteine chemistry.
The reported conditions for S-alkylation of cysteine12–15 with
primary alkyls (dilute solutions: NaOH or KOH in water and alco-
hol, or EtONa in ethanol) which were useful in substitution reac-
tions with primary alkyl halides, failed in the case of long chain
lipophilic halides. Alkylation of the sulfhydryl group in proteins
with farnesyl bromide in alcohol/water mixture in the presence
of MgO has been reported.16,17 Unprotected and protected cyste-
ines were alkylated in DMSO/DMF/MeCN using a fourfold excess
of farnesyl bromide in the presence of N,N-diisopropylethylamine
(DIEA).18 Brown et al. prepared alkylated cysteine derivatives in
an ammonia/methanol solution or in liquid ammonia;19 these
procedures were applied for the synthesis of molecules in anti-
leukemia drug research,20,21 lipidated peptides,22 and S-farnesylat-
Scheme 1.
We observed that treatment of L-cysteine in lower alcohols
(methanol, ethanol, propanol) with two equivalents of TMG
resulted in an exothermic reaction leading to a very reactive and
highly soluble di-salt. In fact, preparation of solutions up to a
concentration of 40% was possible. Addition of alkylating agents to
the homogeneous, concentrated solution resulted in exothermic S-
alkylations. In order to complete the reaction, the mixture was
heated at 50 °C, then (after TLC or GC analysis35) the solvent was
evaporated; water and an equivalent of acetic acid or HCl was added
to the residue and the resulting S-substituted L-cysteine precipitate
was filtered, washed with water, (if insoluble in water) and recrys-
tallized from ethanol or dilute ethanol—Method A. For products sol-
uble in water an equivalent of acetic acid or HCl (in methanol or
ethanol) and small amount of methanol or other alcohol to initiate
of crystallization were added—Method B. TMG could be recovered
from the reaction mixture by concentration, basification to pH 12
with 30% NaOH at a temperature of 5–10 °C, extraction with chloro-
form, washing with brine, drying over MgSO4, concentration by
evaporation, and distillation under reduced pressure to afford the
pure base which could be recycled in >83% yield.
ed-N-acylated-L L
-cysteine.23 S-alkylated -cysteines24 containing a
protected carboxyl or amino group were prepared in the presence
of a fresh solution of sodium ethoxide in absolute ethanol. Unsat-
isfactory yields, poor selectivity, problems with solubility and
purification, formation of side-products, racemisation etc. were
among the difficulties encountered.
Herein we present a very simple and effective method for the
S-alkylation of L-cysteine in the presence of 1,1,3,3-tetramethyl-
Using this procedure37 the alkylation of
L-cysteine with a series
guanidine (TMG) which is free from the above mentioned prob-
lems (Scheme 1).
TMG is a strong, non-ionic base which is widely applied in
various base-mediated organic reactions.25,26 However, to our
knowledge, its application in reactions of amino acids27–31 and
the S-alkylation of thiols32–34 is very limited.
of alkyl halides was carried out. The results are given in Table 1.
The yields of isolated products were very high, even for trans,-
trans-farnesyl, trans-geranyl or dodecyl bromides, and in some
cases yields were almost quantitative. A lower yield was observed
for highly soluble S-tert-butyloxy carbonylmethyl-L-cysteine entry
m. No O- or N-alkylated side-products were detected in the reac-
tion mixture if strictly stoichiometric quantities of reagents were
used and the homogeneity of cysteine di-salt solution prior to
alkylation was assured.
⇑
Corresponding author. Tel.: +48 22 234 5504; fax: +48 22 625 5317.
E-mail address: Marek.Wlostowski@ch.pw.edu.pl (M. Włostowski).
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.08.097

5978
M. Włostowski et al. / Tetrahedron Letters 51 (2010) 5977–5979
Table 1
Preparation of S-alkylated sulfides 3a–q in the presence of TMG and comparison with the best results from the literature
Entry
Thiol 1
Alkylation halide 2
Mp [°C]
Yielda [%]
Lit. yield36 [%]
Lit. conditions
a
b
c
Methyl iodide
219–220
253–254
216–218
210–211
154–156
178–179
192–194
170-172
155-157
214–215
173–175
94–96
93f
89f
91e
95e
99e
85e
88e
91f
92f
89e
79e
93e
53f
93f
91f
84e
99f
8836a
7536b
9415
EtONa/EtOH
NaNH2/NH3(liq)
NaOH(aq)/EtOH
Et3N/EtOH/H2O
NaOH(aq)/1,4-dioxane
Ba(OH)2(aq)/EtOH
NaNH2/NH3(liq)
NH3(g)/MeOH
NaNH2/NH3(liq)
NaOH(aq)/EtOH
—
L
L
L
L
L
L
L
L
L
L
L
L
L
L
-Cysteine
-Cysteine
-Cysteineb
-Cysteine
-Cysteine
-Cysteine
-Cysteineb
-Cysteine
-Cysteine
-Cysteine
-Cysteineb
-Cysteine
-Cysteine
-Cysteine
Ethyl bromide
Benzyl chloride
36c
d
e
f
p-Methylbenzyl chloride
p-Nitrobenzyl chloride
Propargyl chloride
Butyl iodide
—
6036d
8536e
7536b
9136f
7536g
8013
—
g
h
i
trans,trans-Farnesyl bromide
trans-Geranyl bromide
Dodecyl bromide
1,4-Dichloro-2-butynec
2-Bromoacetophenone
t-Butyl chloroacetate
Chloroacetonitrile
j
k
l
8036h
KOH(aq)/EtOH/H2O
—
m
n
o
p
q
179–180
160–162 dec.
176-179
163–165
199–200
—
—
—
2-Bromomethylnaphthalene
Benzyl chloride
—
—
D
-Penicillamine
-Penicillamine
7936i
7436j
NH3(liq)
D
L
-Glutathione redd
p-Nitrobenzyl chloride
NaOH(aq)/EtOH/H2O
a
b
c
d
e
f
Yield refers to isolated products—not optimized.
-Cysteine hydrochloride monohydrate and 3 equiv of TMG were used.
L0Á5 equiv of 2k were used.
3 equiv of TMG were used.
Isolation method A.
Isolation method B.
Acknowledgments
SH
HO(S)
NH2
´
The authors are grateful to Professor R. Gawronski for his kind
support and Professor M. Ma˛kosza for helpful discussions. This
work was partly financially supported by Warsaw University of
Technology (503R/1021/1209/000).
O
D-penicillamine
SH
H
O
O
O
(R) N
(S)
HO
N
H
OH
References and notes
NH2
O
L-glutathione
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There was no analytical evidence for racemisation occurring
during the alkylation. The same results were obtained when
commercially available -cysteine hydrochloride was used but
one additional equivalent of TMG had to be added. Due to the
mild reaction conditions a wide range of alkylating agents con-
taining labile functional groups such as esters, ketones or nitriles
could be used. Thus TMG is an ideal basic reagent for the dis-
crimination between the three nucleophilic centers of the cys-
teine di-anion and leads to selective alkylation of the
sulfhydryl group. The S-alkylation reaction was extended suc-
L
12. Hwan, D.-R.; Helquist, P.; Shekhani, M. S. J. Org. Chem. 1985, 50, 1264–1271.
13. Frankel, M.; Gertner, D.; Jacobson, H.; Zilkha, A. J. Chem. Soc. 1960, 1390–1393.
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Tsuchiya, E.; Abe, K.; Fukui, S. Experimentia 1979, 35, 1275–1276.
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Chem. Soc. 1991, 113, 3176–3177.
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116, 5963–5966.
cessfully to
D
-penicillamine (Fig. 1) (entries o and p) and a more
-glutathione (en-
complex compound, the important tripeptide,
L
try q); in this case, 3 equiv of TMG were used. The yields for
17 examples using the TMG method are compared with the best
yields obtained under other conditions. In all but one case (entry
c) the TMG methodology was proved to be superior or equal to
the known methods.
In conclusion, 1,1,3,3-tetramethylguanidine (TMG) has been
successfully applied in the S-alkylation of
L-cysteine, D-penicilla-
mine, and -glutathione (with unprotected NH2 and COOH groups)
L
with various alkyl halides. Since no racemisation of the amino
acids occurred during the reaction, the application of TMG can be
considered a general route to activation of the SH group in amino
acids and peptides.

M. Włostowski et al. / Tetrahedron Letters 51 (2010) 5977–5979
5979
23. Delson, J. L.; Hodges, H. B.; MacDouglas, D. D.; Henriksen, B. S.; Hrycyna, C. A.;
Gibbs, R. A. Bioorg. Med. Chem. Lett. 2006, 16, 4420–4423.
24. Perrey, D. A.; Uckun, F. M. Tetrahedron Lett. 2001, 42, 1859–1861.
25. Marianoff, C. A.; Leadbeater, N. E. In e-EROS Encyclopedia of Reagents for Organic
Synthesis; Crich, D.; Ed.; John Wiley & Sons, Ltd., 2007.
Method B: stoichiometric quantity of AcOH or HCl in MeOH (or EtOH) was
added to the reaction mixture and more MeOH (or EtOH) to initiate
crystallization of the product which was then recrystallized from alcohol or
diluted alcohol.
Compounds 3a–q are enantiomerically pure and were fully characterized on
26. Ishikawa, T.; Kumamoto, T. Synthesis 2006, 737–752.
the basis of spectral (1H, 13C, IR) and CHN data.
27. Ali, A.; Fahrenholtz, F.; Weinstein, B. Angew. Chem., Int. Ed. Engl. 1972, 11, 289.
28. Steglich, W.; Hinze, S. Synth. Commun. 1976, 399–401.
29. Włostowski, M.; Rowicki, T.; Synoradzki, L. Tetrahedron: Asymmetry 2004, 15,
2333–2338.
30. Luo, J.; Smith, M. D.; Lantrip, D. A.; Wang, S.; Fuchs, P. L. E. J. Am. Chem. Soc.
1997, 119, 10004–10013.
31. Kemp, D. S.; Wang, S.-W.; Rebek, J., Jr.; Mollan, R. C.; Banquer, C.;
Subramanyam, G. Tetrahedron 1974, 30, 3955–3967.
Spectral data for new compounds: compound 3k: from EtOH, mp 173–175 °C;
[
a
]
À5.6 (c 1.0, 0.1 M NaOHaq); dH (400 MHz, D2O) 3.97 (2H, dd, J = 3.2, 4.4),
D
3.28 (2H, dd, J = 14.8, 4.4), 3.15 (dd, 2H, J = 14.8, 7.6), 2.94 (4H, s); dC (100 MHz,
D2O) 173.61, 80.00, 54.04, 32.65, 19.89; mmax (KBr/cmÀ1) 3410, 3045, 1607,
1408, 1131, 541 cmÀ1
; MS (EI) silylated derivative: C22H48O4N2S2Si4, m/z
(%) = 581 (0.2) [M+], 463 (18), 317 (19), 316 (14), 315 (7), 232 (9), 220 (11), 219
(22), 218 (100), 147 (13), 73 (24). Elemental anal.: C10H16N2O4S2, calcd: C,
41.08; H, 5.52; N, 9.58; S, 21.93; obsd: C, 41.28; H, 5.41; N, 9.64; S, 22.14.
32. Williams, J. M.; Brands, K. M. J.; Skerlj, R. T.; Jobson, R. B.; Marchesini, G.;
Conrad, K. M.; Pipik, B.; Savary, K. A.; Tsay, F.-R.; Houghton, P. G.; Sidler, D. R.;
Dolling, U.-H.; DiMichele, L. M.; Nowak, T. J. J. Org. Chem. 2005, 70, 7479–7487.
33. Bera, S.; Langley, J. L.; Pathak, T. J. Org. Chem. 1998, 63, 1754–1760.
34. Włostowski, M.; Czarnocka, S.; Maciejewski, P. unpublished results.
Compound 3m: from methanol, mp 179–180 °C; [
a
]
À23.62 (c 0.1, 0.1 M
D
NaOHaq); dH (400 MHz, NaOD/D2O) 3.52 (1H, dd, J = 4.7, 7.5), 3.37 (1H, dd,
J = 5.6, 7.5), 3.05 (1H, dd, J = 4.7, 13.5), 2.84 (1H, m, J = 5.6, 13.5), 2.83 (2H, s),
2.70 (1H, dd, J = 7.5, 13.5), 1.19 (9H, s,); dC (100 MHz, NaOD/D2O) 181.57,
178.83, 70.34, 55.56, 40.08, 38.28, 30.21; mmax (KBr/cmÀ1) 3485, 3115, 2970,
35.
A
sample the mixture was concentrated, silylated with BSA [N,O-bis-
1728, 1610, 1410, 1300, 1126, 675 cmÀ1
C
;
MS (EI) silylated derivative:
(trimethysilyl)acetamide] to form the disilylated derivative and tested by
GC/MS or GC analysis on a chiral column.
15H33O4NS2Si4, m/z (%) = 379 (0.02) [M+], 234 (16), 190 (26), 146 (43), 134
(100), 116 (22), 75 (22), 73 (33), 57 (32), 43 (13); 28 (32). Elemental anal.:
C9H17NO4S, calcd: C, 45.94; H, 7.28; N, 5.95; S, 13.63; obsd: C, 46.01; H, 7.21; N,
5.98; S, 13.58.
Compound 3n: from H2O, mp 160–162 °C decomp. [
NaOHaq); dH (400 MHz, D2O) 4.01 (1H, dd, J = 6.6, 5.6), 3.60 (2H, s), 3.28 (2H,
m); dC (100 MHz, D2O) 172.82, 116,67, 53.71, 33.21, 17.35; mmax (KBr/cmÀ1
3426, 2976, 2247, 1587, 1509, 1397, 1344, 1300, 865, 546; MS (EI) silylated
derivative: C11H24N2O2SSi2, m/z (%) = 304 (0.14) [M+], 219 (20), 218 (100), 188
(14), 187 (92), 147 (35), 146 (14), 144 (34), 100 (18), 75 (14), 73 (81).
Elemental anal.: C5H8N2O2S, calcd: C, 37.49; H, 5.03; N, 17.49; S, 20.02; obsd: C,
37.52; H, 5.06; N, 17.53; S, 20.08.
36. (a) Hwang, D.-R.; Helquist, P.; Shekhani, M. S. J. Org. Chem. 1985, 50, 1264–
1271; (b) Armstrong, L.; Lewis, J. D. J. Org. Chem. 1951, 16, 749–751; (c)
Erickson, B. W.; Merrifield, R. B. J. Am. Chem. Soc. 1973, 95, 3750–3756; (d)
Berse, C.; Boucher, R.; Piche, L. J. Org. Chem. 1957, 22, 805–807; (e) Marrone, L.;
Siemann, S.; Beecroft, M.; Viswanatha, T. Bioorg. Chem. 1996, 24, 401–416; (f)
Cini, E.; Lampariello, L. R.; Rodriquez, M.; Taddei, M. Tetrahedron 2009, 65, 844–
848; (g) Joullie, M.; Lakah, L.; Maillard, G.; Muller, P. Ger. Patent 2329819,
1974; Chem. Abstr. 1974, 80, 83651.; (h) Tang, G.; Ji, T.; Hu, A.-F.; Zhao, Y.-F.
Synlett 2008, 1907–1909; (i) Moynihan, H. A.; Roberts, S. M. J. Chem. Soc., Perkin
Trans. 1 1994, 797–806; (j) Vince, R.; Daluge, S.; Wadd, W. B. J. Med. Chem.
1971, 14, 402–404.
a] 34.19 (c 0.1, 0.1 M
D
)
Compound 3o: from MeOH, mp 176–179 °C. [a]D À85.11 (c 0.1, 0.1 M NaOHaq),
37. Procedure for the synthesis of S-alkylated cysteine and related compounds 3a–q: A
solution of TMG (21 mmol) in 5 ml of MeOH (or other alcohol) was added
dH (400 MHz, NaOD/D2O) 6.89 (4H, m), 6.46 (3H, m), 3.13 (2H, s), 2.97 (1H, s),
1.16 (3H, s), 0,84 (3H, s); dC (100 MHz, NaOD/D2O) 178.71, 135.44, 133.53,
132.54, 128.58, 127.94, 127.71, 126.30, 125.90, 63.63, 51.33, 32.75, 27.71,
dropwise to a stirred (under argon) mixture of L-cysteine (10 mmol) in 10 ml of
MeOH (or other alcohol). To the homogeneous solution was added alkylation
reagent 2a–q (10 mmol) in THF dropwise and the reaction mixture was stirred
1 h at 50 °C. The mixture was evaporated and after addition of 10 ml of H2O,
acidified with a stoichiometric quantity of AcOH or HCl (1:1). The separated
precipitate was recrystallized from alcohol, dilute alcohol or H2O—Method A.
22.94; m
max (KBr/cmÀ1) 3504, 3151, 3046, 2966, 1622, 1508, 1410, 1370, 1132,
884, 836, 756, 564; MS (EI) silylated derivative: C13H28N2O2SSi2, m/z (%) = 433
(0.08) [M+], 220 (10); 219 (44), 218 (41), 215 (12), 142 (15), 141 (100), 115
(11), 73 (31), 32 (15), 28 (64). Elemental anal.: C16H19NO2S, calcd: C, 66.41; H,
6.62; N, 4.84; S, 11.08; obsd: C, 66.59; H, 6.60; N, 4.78; S, 11.01.