Synthesis of the (E)-dehydrobutyrine-thiazoline-proline-leucine fragment of vioprolides B and D

Article abstract of DOI:10.1016/j.tetlet.2013.04.017

A procedure is reported for the introduction of both the thiazoline and (E)-dehydrobutyrine components into a tetrapeptide-derived fragment of vioprolides B and D. The (E)-dehydrobutyrine is introduced first but, as the carbon-carbon double-bond of the dehydrobutyrine appeared incompatible with an adjacent thiol, the thiazoline was assembled by dehydration of a serine containing thioamide not by dehydration of a cysteinyl analogue.

Full text of DOI:10.1016/j.tetlet.2013.04.017

Tetrahedron Letters 54 (2013) 3150–3153
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Tetrahedron Letters
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Synthesis of the (E)-dehydrobutyrine–thiazoline–proline–leucine fragment
of vioprolides B and D
⇑
Hao Liu, Eric J. Thomas
School of Chemistry, University of Manchester, Manchester M13 9PL, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 27 February 2013
Revised 4 April 2013
Accepted 5 April 2013
Available online 13 April 2013
A procedure is reported for the introduction of both the thiazoline and (E)-dehydrobutyrine components
into a tetrapeptide-derived fragment of vioprolides B and D. The (E)-dehydrobutyrine is introduced first
but, as the carbon–carbon double-bond of the dehydrobutyrine appeared incompatible with an adjacent
thiol, the thiazoline was assembled by dehydration of a serine containing thioamide not by dehydration
of a cysteinyl analogue.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Peptolides
[E]-dehydrobutyrine
Thiazoline
Vioprolides
The vioprolides A–D 1–4 are a small group of peptolides iso-
lated from Cystobacter violaceus.1 They possess antifungal activity
and are highly cytotoxic to mammalian cells with vioprolide D
showing the least cytotoxicity and maximum antifungal activity.
Of particular interest is the observation that they have a synergistic
effect on the murine type 1 inferferon signalling pathway leading
to more than a three-fold increase in activity when stimulated by
viral infection.2 However, further studies into their biological activ-
ity are being delayed because of the extremely limited supply of
these natural products.
Structural features of interest in the vioprolides are the 4-
methylazetidinecarboxylic acid (MAZ), the (E)-dehydrobutyrine
and the thiazoline. This was shown during the chemical degrada-
tion of the natural products to facilitate epimerisation of the neigh-
bouring MAZ or proline-derived fragment. Two conformers were
detected for vioprolide A 1 attributed to atropisomers about the
N-methylvaline–threonine amide bond.1
To date, the only studies on synthetic approaches to the viopro-
lides are a non-stereoselective synthesis of MAZ reported during
their isolation1 and a synthesis of the azetidinyl-thiazoline compo-
nent of vioprolides A 1 and C 2.3 We here report studies concerned
with the introduction of both the thiazoline and (E)-dehydrobuty-
rine components into a fragment of the vioprolides.
Procedures are available for the dehydration of threonine con-
taining peptides to give either (Z)- or (E)-dehydrobutyrine contain-
ing fragments4,5 and cysteine-derived thiazolines can be accessed
by dehydration of thioamides adjacent to a serine residue6 or of
cysteine containing peptides.7 The latter approach was selected
for the initial studies since several, fairly recently developed, pro-
cedures are available for this transformation.
S-Trityl-N-Fmoc-protected
lyl ester 6 and Fmoc-removal gave the amine 7. This was coupled
with N-nosyl protected
-proline9 to give the protected dipeptide
L
-cysteine 58 was converted to its al-
L
8, see Scheme 1. Nosyl protection for the proline was used as the
1H NMR spectra of the corresponding Boc-protected dipeptides
were complicated by the presence of rotamers. Triflic anhydride
and triphenylphosphine oxide7a effected thiazoline formation from
the dipeptide 8 and gave a good yield of the thiazoline 9 as a single
diastereoisomer (1H NMR). However, during attempts to continue
with the synthesis by conversion of the allyl ester 9 into the corre-
⇑
Corresponding author. Tel.: +44 (0) 161 275 4613; fax: +44 (0) 161 275 4939.
E-mail address: e.j.thomas@manchester.ac.uk (E.J. Thomas).
0040-4039/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2013.04.017

H. Liu, E. J. Thomas / Tetrahedron Letters 54 (2013) 3150–3153
3151
Scheme 1. Initial studies on thiazoline formation; Reagents and conditions: (i) CH2=CHCH2Br, NaHCO3, DMSO, rt, 16 h (83%); (ii) piperidine, DMF, r.t., 16 h (94%); (iii) N-
nosyl-L-proline, HATU, HOBt, DMF, add 7 then EtNiPr2, rt, 16 h (77%); (iv) Tf2O, Ph3PO, DCM, 0 °C, 10 min, add 8, rt, 2 h (68%); (v) Pd(Ph3P)4 (10%), N-methylaniline, THF, rt, 3 h
(95%).
sponding acid, epimerisation of the proline occurred and a mixture
of the epimeric acids 10 was obtained, see Scheme 1.
peting reactions between the thiol formed on deprotection of the
trityl thio-ether 15 and the neighbouring double-bond of the dehy-
drobutyrine. This suggested that, although the thiazoline will have
to be introduced as late as possible in the synthesis to minimise
epimerisation of the neighbouring amino acid residue, free-thiol
containing intermediates would have to be avoided to prevent
complications due to the presence of the neighbouring dehydrobu-
tyrine. The alternative procedure for the introduction of cysteinyl-
derived thiazolines into peptides based on the dehydration of ser-
ine-derived thioamides was therefore investigated.6
Although the epimerisation of amino acid residues adjacent to
thiazolines in peptide derivatives is well known10 and indeed
was observed during attempts to degrade the vioprolides into their
constituent amino acids,1 it was not reported during the synthesis
of the azetidinyl-thiazoline fragment of the vioprolides.3 In the
case of the thiazoline 10 the rapid epimerisation may be due to
hydrogen bonding between the nitrogen of the thiazoline and the
carboxylic acid, see structure 11. This could facilitate imine–enam-
ine tautomerisation thus leading to epimerisation.
Only low yields of the O-TBS-L-threonine-O-TIPS-L-serine dipep-
Next, procedures for the introduction of the (E)-dehydrobuty-
rine residue were investigated. Deallylation of the protected dipep-
tide 8 and coupling the resulting acid 12 with the allyl ester of O-
tert-butyldimethylsilyl protected L-threonine gave the protected
tripeptide 13. This was desilylated to give the alcohol 14 ready
for the dehydration to the corresponding alkene. (E)-Dehydrobuty-
rines are available by dehydration of threonine containing peptides
using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and
copper(II) chloride5 and indeed this procedure gave a good yield
of the required dehydrobutyrine-containing peptide 15, the geom-
etry of the double-bond being assigned at this stage by analogy
with the literature.5 However, attempts to effect a subsequent
dehydration to form the required thiazoline using the triflic anhy-
dride and triphenylphosphine oxide procedure7 were unsuccessful,
complex mixtures of products being obtained, see Scheme 2.
The problems encountered during the attempted formation of
the thiazoline 16 from the thio-ether 15 were attributed to com-
tide 19 were obtained in our hands if O-TIPS-protected serine was
coupled with the allyl ester of O-TBS-protected threonine, and so
the protected L-threonine was coupled with the Fmoc-protected L-
serine 17 with a free hydroxyl group, to give the protected dipeptide
18 that was silylated to give the bis-silylated dipeptide 19, see
Scheme 3. Following removal of the Fmoc-group, coupling the
resulting amine 20 with the Fmoc-protected thioacylating agent
2111 gave a thioamide that was converted into the corresponding
amine 22 by treatment with piperidine under standard conditions.
This was now coupled with Boc-protected D-leucine to give the mod-
ified tetrapeptide 23. Selective removal of the secondary TBS-group
in the presence of the primary TIPS-group proved difficult and so
both of the silyl protecting groups were removed using tetrabutyl-
ammonium fluoride, and the primary serine-derived alcohol was
resilylated to give the monosilyl ether 24. Dehydration of this using
EDC and copper(II) chloride under conditions known to be selective
for formation of (E)-dehydrobutyrines5 gave a product identified as
Scheme 2. Initial studies on thiazoline formation in the presence of a dehydrobutyrine; Reagents and conditions: (i) Pd(Ph3P)4 (10%), N-methylaniline, THF, rt, 3 h (95%); (ii)
12, HATU, HOBt, DMF, add O-TBS-
DCM, 0 °C, 10 min, add 15, rt.
L
-threonine allyl ester, EtNiPr2, rt, 16 h (67%); (iii) Et3N(HF)3, THF, rt, 48 h (62%); (iv) EDC, CuCl2, tol., 80 °C, 30 min (62%); (v) Tf2O, Ph3PO,

3152
H. Liu, E. J. Thomas / Tetrahedron Letters 54 (2013) 3150–3153
Scheme 3. Synthesis of the dehydrobutyrine–thiazoline–proline–leucine fragment of vioprolides B and D. Reagents and conditions: (i) HATU, HOBt, DMF, add O-TBS-
threonine allyl ester, EtNiPr2, rt, 16 h (97%); (ii) TIPSOTf, 2,6-lutidine, DCM, rt, 16 h (56%); (iii) piperidine, DMF, rt, 16 h (89%); (iv) (a) compound 21, THF, rt, 6 h (b) piperidine,
DMF, rt, 16 h (40%); (v) Boc-
30 min (65%); (viii) TBAF, THF, rt, 16 h (62%); (ix) DAST, DCM, ꢁ15 °C, 1 h (75%); (x) DAST, pyridine, DCM, 0 °C, 2 h (31%).
D
-leucine, HATU, HOBt, DMF, add 22, EtNiPr2, rt, 16 h (85%); (vi) (a) TBAF, THF, rt, 16 h (b) TIPSCI, imid., THF (30%); (vii) EDC, CuCL2,tol., 80 °C,
the required (E)-isomer 25 on the basis of precedent. To check that
the (E)- and (Z)-isomers could be distinguished, the alcohol 24 was
also dehydrated using diethylaminosulfur trifluoride (DAST) under
conditions known to be selective for formation of (Z)-dehydrobuty-
rines.4 Although only a modest yield of alkene was obtained in the
latter case, the alkene that was isolated was different by 1H NMR
from that obtained using EDC-copper(II) chloride and was identified
as the (Z)-isomer 28 [25, =CHCH3, dH (CDCl3) 1.98; 28, =CHCH3, dH
(CDCl3) 1.72], see Scheme 3. Indeed following comparison of the
1H NMR spectra of the two isomers, a small amount of the (Z)-iso-
mer 28 was detected in the (E)-isomer 25; (E)-25: (Z)-28 = 95:5.
It now remained to check that the formation of a thiazoline by
dehydration of the serine-thioamide was compatible with the (E)-
dehydrobutyrine component. Deprotection of the tri-isopropylsilyl
ether 25 using tetrabutylammonium fluoride gave the correspond-
ing alcohol 26 and this on treatment with DAST6 was dehydrated
to give a major product isolated in 75% yield. This was identified
as the required thiazoline 27 containing ca. 20% of its epimer due
to partial racemisation of the proline-derived residue as indicated
in the 1H NMR spectrum of 27 by some doubling of peaks, for
example of the doublets attributed to the diastereotopic methyl
groups present in the leucine.
This work has developed procedures for the assembly of both an
(E)-dehydrobutyrine and an adjacent thiazoline into peptides re-
lated to fragments of the vioprolides.12 Because of its tendency to
promote epimerisation of an adjacent amino acid residue,10 the
thiazoline should be introduced as late as possible in any synthetic
approach to a vioprolide. However, the compatibility of the intro-
duction of the thiazoline from a thioamide using DAST with the
(E)-dehydrobutyrine suggests that this procedure may be appro-
priate for the synthesis of a natural product. Some optimisation
may be necessary to minimise epimerisation at the proline- or
MAZ-derived stereogenic centre, although a depsipeptide with a
proline derived residue next to the thiazoline has been prepared
using this procedure without any racemisation being observed.6
Acknowledgments
We thank Novartis for a studentship (to H.L.).
References and notes
1. Schummer, D.; Forche, E.; Wray, V.; Domke, T.; Reichenbach, H.; Höfle, G.
Leibigs Ann. 1996, 971.
2. Bollati-Fogolín, M.; Oggero, M.; Mirazo, S.; Frank, R.; Kratje, R.; Muller, W. Cells
and Culture, ESACT Proceedings 4, 485; Springer, 2010, doi: 10.1007/978-90-
481-3419-9_86.
3. Chopin, N.; Couty, F.; Evano, G. Lett. Org. Chem. 2010, 7, 353.
4. (a) Ogura, H.; Sato, O.; Takeda, K. Tetrahedron Lett. 1981, 22, 4817; (b)
Andruszkiewicz, R.; Czerwínski, A. Synthesis 1982, 968; (c) Berti, F.; Ebert, C.;
Gardossi, L. Tetrahedron Lett. 1992, 33, 8145; (d) Goodall, K.; Parsons, A. F.
Tetrahedron Lett. 1995, 36, 3259; (e) Yokokawa, T.; Shioiri, T. Tetrahedron Lett.
2002, 43, 8673; (f) Wojciechowska, H.; Pawlowitcz, R.; Andruszkiewicz, R.;
Grzybowska, J. Tetrahedron Lett. 1978, 19, 4063; (g) Miller, M. J. J. Org. Chem.
1980, 45, 3131; (h) Srinivasan, A.; Stephenson, R. W.; Olsen, R. K. J. Org. Chem.
1977, 42, 2256; (i) Yokokawa, F.; Shioiri, T. Tetrahedron Lett. 2002, 43, 8679.
5. Sai, H.; Ogiku, T.; Ohmizu, H. Synthesis 2003, 201.
6. McKeever, B.; Pattenden, G. Tetrahedron 2003, 59, 2701.
7. (a) You, S.-L.; Razavi, H.; Kelly, J. W. Angew. Chem., Int. Ed. 2003, 42, 83; (b) Ma,
D.; Zou, B.; Cai, G.; Hu, X.; Lin, J. O. Chem. Eur. J. 2006, 12, 7615.
8. Commercially available from Sigma–Alrich.
9. Prepared by reaction of
L-proline with 2-nitrobenzenesulfonyl chloride,
NaHCO3, H2O, rt, 16 h (85%).
10. (a) North, M.; Pattenden, G. Tetrahedron 1990, 45, 8267; (b) Wipf, P.; Fritch, P.
C. Tetrahedron Lett. 1994, 35, 5397.
11. Prepared from Fmoc-protected proline i (see Ref. 10).
Prep. of thioacylating agent 21: (i), 2-amino-4-nitroaniline, iBuOCOꢀCl, THF,
ꢁ20 °C, add i, rt, 16 h (72%); (ii), P4S10, NaHCO3, THF, 1 h, add ii, 0 °C, ꢁrt, 3 h
(89%); (iii), NaNO2, AcOH, 5% H2O, 0 °C, 30 min (99%).
12. (E)-Dehydrobutyrine 25: A solution of the alcohol 24 (40 mg, 0.05 mmol), EDC
(17 mg, 0.11 mmol) and copper(II) chloride (1 mg, 0.01 mmol) in anhydrous
toluene (2 mL) was stirred at 80 °C under an atmosphere of nitrogen for

H. Liu, E. J. Thomas / Tetrahedron Letters 54 (2013) 3150–3153
3153
30 min. Water was added and the mixture was extracted with ethyl acetate
(5 mL). The organic extracts were dried (Na2SO4) and concentrated under
reduced pressure. Chromatography of the residue eluting with ether gave the
title compound 25 (23 mg, 65%) as a colourless gum, Rf 0.6 (light petroleum:
cmꢁ1 3321, 1702, 1682, 1643, 1510, 1425, 1366, 1343, 1250, 1162, 1015, 988,
932, 854 and 780 cmꢁ1; dH (400 MHz, CDCl3) major epimer 0.87 and 0.91 (each
3H, d, J 6.6, 4000-CH3 and 5000-H3), 1.35 [9H, s, C(CH3)3], 1.41 (2H, m, 3000-H2), 1.64
(1H, m, 4000-H), 1.85-2.00 (2H, m, 400-H2), 2.03 (3H, d, J 7.6, 4-H3), 2.05–2.15 (2H,
m, 300-H2), 3.40-3.60 (2H, m, 30-H2), 3.88 (2H, m, 500-H2), 4.44 (1H, dt, J 4.8, 9.6,
2000-H), 4.67 (2H, m, OCH2CH=), 4.84 (1H, dd, J 4.0, 9.0, 20-H), 5.03 (2H, m, 200-H
and NH), 5.20 and 5.30 (each 1H, m, HCH=CH), 5.90 (1H, m, CH2=CH), 6.96 (1H,
q, J 7.8, 3-H) and 8.62 (1H, br s, NH); minor epimer 0.74 and 0.80 (each 3H, d, J
6.6, 4000-CH3 and 5000-H3), 4.19 (1H, dt, J 4.8, 9.6, 2000-H) and 7.13 (1H, q, J 7.8, 3-
H); m/z (ES+) 559 (M++23, 100%).
ether = 30: 70),
½
a 2D0
ꢂ
ꢁ37.5 (c
2
in CHCl3) (Found: M++H, 711.4189.
max/cmꢁ1 3294, 1682, 1645, 1507,
C
35H63N4O7SiS requires M, 711.4182);
m
1447, 1386, 1366, 1251, 1163, 1106, 1066, 1046, 1014, 994, 920, 881, 851, 778,
735 and 681 cmꢁ1; dH (400 MHz, CDCl3) 0.89 and 0.91 (each 3H, m, 4000-CH3 and
5000-H3), 0.99 [21H, m, 3ꢃ SiCH(CH3)2], 1.33 [9H, s, C(CH3)3], 1.36 (2H, m, 3000
-
H2), 1.64 (1H, m, 4000-H), 1.92 (1H, m, 400-H), 1.98 (3H, d, J 7.6, 4-H3), 2.04, 2.22
and 2.34 (each 1H, m, 2ꢃ 300-H or 400-H0), 3.51 (2H, m, 500-H2), 3.81 and 3.96
(each 1H, m), 4.21 (1H, dd, J 8.0, 12.0, 30-H), 4.34 (1H, m, 30-H0), 4.63 (2H, m,
OCH2CH=), 4.85 (1H, m), 5.05 (1H, m, NH), 5.18 and 5.28 (each 1H, m,
HCH=CH), 5.90 (1H, m, CH2=CH), 6.68 (1H, m, 3-H) and 8.13 and 8.62 (each 1H,
br m, NH); dC (125 MHz, CDCl3) 11.79, 14.17, 17.91, 21.93, 23.42, 24.36, 24.64,
28.32, 30.33, 32.37, 41.37, 47.71, 50.85, 62.26, 65.89, 79.99, 82.65, 103.73,
118.55, 126.11, 129.12, 131.96, 148.99, 155.72, 163.59 and 203.33; m/z (ES+)
733 (M++23, 100%).
(Z)-Dehydrobutyrine 28: Diethylaminosulfur trifuoride (7 mg, 0.04 mmol)
was added dropwise over 1 min to the alcohol 24 (20 mg, 0.027 mmol) and
pyridine (8 mg, 0.11 mmol) in DCM (0.5 mL) at 0 °C. The reaction was stirred at
0 °C for 2 h then saturated aqueous sodium hydrogen carbonate was added.
The organic extracts were washed with brine and dried (Na2SO4) then
concentrated under reduced pressure. Column chromatography of the
residue eluting with petrol–ether (30:70) gave the title compound 28 (6 mg,
31%) as a colourless gum, Rf 0.5 (30:70 petrol–ether), ½a D20
ꢁ11 (c 2.0 in CHCl3)
ꢂ
Thiazoline 27: Diethylaminosulfur trifluoride (10 mg, 0.06 mmol) was added
dropwise over 1 min to the thioamide 26 (11 mg, 0.02 mmol) in DCM (0.25 mL)
at ꢁ15 °C and the solution stirred for 1 h. Saturated aqueous sodium hydrogen
carbonate was added and the mixture allowed to warm to room temperature.
DCM (5 mL) and water (5 mL) were added and the mixture extracted with DCM
(10 mL). The organic extracts were dried (Na2SO4) and concentrated under
reduced pressure. Chromatography of the residue eluting with ether gave the
title compound 27 (8 mg, 75%) as a colourless gum, containing ca. 20% of its
(Found: M++H, 711.4183. C35H63N4O7SiS, requires M, 711.4182); mmax 3323,
1686, 1641, 1509, 1450, 1366, 1250, 1163, 1068, 1046, 1015, 994, 921, 882,
757 and 682 cmꢁ1; dH (400 MHz, CDCl3) 0.86 and 0.88 (each 3H, d, J 6.6, 4000
-
CH3 and 5000-H3), 0.99 [21H, m, 3ꢃ SiCH(CH3)2], 1.36 [9H, s, C(CH3)3], 1.42 (3H,
m, 3000-H2 and 4000-H), 1.72 (3H, d, J 7.1, 4-H3), 1.95 (2H, m, 400-H2), 2.26 (2H, m,
300-H2), 3.55 (2H, m, 500-H2), 3.70–3.95 (2H, m), 4.28 and 4.39 (each 1H, m), 4.59
(2H, m, OCH2CH=), 4.85 (1H, dd, J 4.0, 9.0, 200-H), 5.17 and 5.25 (each 1H, m,
HCH=CH), 5.84 (1H, m, CH2=CH), 6.79 (1H, q, J 7.3, 3-H), 7.45 (1H, br d, J 8.6,
NH), 7.97 (1H, s, NH) and 8.45 (1H, br d, J 7.3, NH); m/z (ES+) 733 (M++23,
100%).
epimer at the proline derived centre (1H NMR), Rf 0.5 (ether), ½a 2D0
ꢁ15.6 (c 2 in
ꢂ
CHCl3) (Found: M+ ꢁ H, 535.2596. C26H39N4O6S requires M, 535.2595); mmax
/