A total synthesis of the ammonium ionophore, (-)-enniatin B

Article abstract of DOI:10.1016/j.tetlet.2012.05.110

A nine-step (longest linear) batch total synthesis of the cyclic hexadepsipeptide (-)-enniatin B is described. The synthesis minimizes precipitation during reaction conditions for adaptability to flow synthesis. The route was used to prepare >100 mg of th

Full text of DOI:10.1016/j.tetlet.2012.05.110

Tetrahedron Letters 53 (2012) 4077–4079
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Tetrahedron Letters
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A total synthesis of the ammonium ionophore, (À)-enniatin B
⇑
Dennis X. Hu, Max Bielitza, Peter Koos, Steven V. Ley
Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW, United Kingdom
a r t i c l e i n f o
a b s t r a c t
Article history:
A nine-step (longest linear) batch total synthesis of the cyclic hexadepsipeptide (À)-enniatin B is
described. The synthesis minimizes precipitation during reaction conditions for adaptability to flow syn-
thesis. The route was used to prepare >100 mg of the natural product.
Received 13 March 2012
Revised 14 May 2012
Accepted 23 May 2012
Available online 7 June 2012
Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
Keywords:
(À)-Enniatin B
Macrolactamization
Cyclic hexadepsipeptide
Ammonium ionophore
Precipitation-free
(À)-Enniatin B (1, Fig. 1) is a cyclohexadepsipeptide fungal
product known to possess various antibiotic,1 cytotoxic,2 and ion-
transport3 activities. It has also been investigated for its potential
as an anti-HIV4 and anti-cancer agent,5 the latter due to its recently
discovered new mechanisms of pro-apoptotic activity.6 Addition-
ally, as a chemically stable mycotoxin, enniatin B serves as an indi-
cator of fungal contamination of food grains,7 and is thus required
as an analytical standard worldwide.8 Small quantities of enniatin
B for such diverse research purposes are commercially available
through fermentation, albeit though at high cost (ꢀ$200 USD per
milligram). A simple chemical synthesis of enniatin B to provide
both material and potential analogs is therefore desirable.
Previous syntheses of enniatin B were completed simulta-
neously in 1963 by Shemyakin18,19 and Plattner20 using PCl5 and
HBr-mediated couplings. Another synthesis of enniatin B was
reported in 2006 employing Mitsunobu reactions to form the
required ester bonds and a silver-mediated amide trimethylation
process for the final step.21 Neither route is ideally suited for flow
chemistry, due to precipitation and difficult intermediate purifica-
tion steps. We have therefore developed an improved batch
synthesis, disclosed herein.
In initial studies, several attempts to couple fragments 2 and 3
failed to provide desired tetramer 4, evidently due to rapid forma-
tion of oxazolonium cation 5 (Scheme 1).
In our laboratories, we have been investigating methods for the
flow chemical synthesis of natural products.9–11 Despite their sim-
ple appearance, the syntheses of cyclodepsipeptides such as enni-
atin B are known to be difficult to automate with solid-phase
chemistry:12 depsipeptide bonds require several equivalents of
the expensive hydroxyester component to drive on-resin chain
extension to completion,13 N-methyl amines are known to be very
poor nucleophilic coupling partners,14 and N-methyl amides are
known to cause difficulties such as epimerization during chain
extension due to the rapid formation of oxazolones.14 We therefore
wished to apply newly developed flow technologies to automate
the synthesis of enniatin-type cyclodepsipeptides in the solution
phase. To study the application of our flow technologies, we first
needed to define a batch synthesis that would minimize the risk
of in-line precipitation events throughout the sequence.15–17
We thus opted to reverse the order of bond formation to pre-
vent amide-induced deactivation of the necessary acid coupling
components. EDCI coupling of acid 822 and alcohol 7 cleanly
provided depsipeptide 9 (Scheme 2). In the coupling step, it was
important to use stoichiometric DMAP to minimize the formation
⇑
Corresponding author. Tel.: +44 1223 336398; fax: +44 1223 336442.
E-mail address: svl1000@cam.ac.uk (S.V. Ley).
Figure 1. (À)-Enniatin B (1), a fungal cyclohexadepsipeptide.
0040-4039/$ - see front matter Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2012.05.110

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D. X. Hu et al. / Tetrahedron Letters 53 (2012) 4077–4079
(nine steps, longest linear, 15% yield overall). Notably, only two
chromatography steps were necessary in the entire sequence
beginning from -valine and no precipitation from reaction solu-
L
tion was observed during the reaction sequence. Our efforts to
harness this sequence using flow technology and biological inves-
tigations on the natural product will be disclosed in due course.
Acknowledgments
We thank the BP Endowment (to S.V.L.) for financial support of
this work. D.X.H. thanks the Winston Churchill Foundation for a
Churchill Scholarship, P.K. thanks Georganics Ltd, and M.B. thanks
the Industrieclub Düsseldorf for a grant.
Supplementary data
Supplementary data (detailed experimental procedures, com-
pound characterization, and original 1H and 13C NMR spectra of
all compounds) associated with this article can be found, in the on-
line version, at http://dx.doi.org/10.1016/j.tetlet.2012.05.110.
Scheme 1. Initial coupling partners 2 and 3 failed to provide product 4 due to rapid
formation of oxazalonium cation 5; anhydride 6 resulted from a similar interaction
with the Boc-protecting group in acid 8.
References and notes
1. (a) Shemyakin, M. M.; Ovchinnikov, Y. A.; Ivanov, V. T.; Kiryushkin, A. A.; Zhdanov,
G. L.; Ryabova, I. D. Experientia 1963, 19, 566–568; (b) Bahdury, P.; Mohammad, B.
T.; Wright, P. C. J. Ind. Microbiol. Biot. 2005, 33, 325–337; (c) Meca, G.; Soriano, J. M.;
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Font, G.; Ruiz, M. J. Toxicol. Mech. Method 2011, 21, 503–512; (d) Fornelli, F.;
Minervini, F.; Logrieco, A. J. Invertebr. Pathol. 2004, 85, 74–79.
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Zachariasova, M.; Hajslova, J. J. Agric. Food Chem. 2011, 59, 12990–12997;
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9. Baxendale, I. R.; Griffiths-Jones, C. M.; Ley, S. V.; Tranmer, G. K. Synlett 2006,
427–430.
Scheme 2. Total synthesis of (À)-enniatin B. Reagents and conditions: (a) EDCI,
DMAP (1.3 equiv), CH2Cl2 (82%); (b) H2, Pd/C, EtOAc (100%) (c) 4 M HCl/1,4-dioxane
(100%), then vacuum; (d) 10, Ghosez’s reagent, DIPEA, CH2Cl2 (91%); (e) H2, Pd/C,
EtOAc (100%); (f) 11, Ghosez’s reagent, DIPEA, CH2Cl2 (69%, 86% based on recovered
starting material); (g) H2, Pd/C, EtOAc (88%); (h) 4 M HCl/1,4-dioxane, then vacuum
(100%); (i) Ghosez’s reagent, then DIPEA, 0.005 M CH2Cl2 (56%). Ghosez’s
reagent = 1-chloro-N,N,2-trimethyl-1-propenylamine, DMAP = 4-dimethylamino-
pyridine, DIPEA = diisopropylethylamine.
10. Baxendale, I. R.; Deeley, J.; Griffiths-Jones, C. M.; Ley, S. V.; Saaby, S.; Tranmer,
G. K. Chem. Commun. 2006, 2566–2568.
11. (a) Baumann, M.; Baxendale, I. R.; Brasholz, M.; Hayward, J. J.; Ley, S. V.; Nikbin,
N. Synlett 2011, 1375–1380; (b) Hodgkinson, J. T.; Galloway, W. R. J. D.; Saraf,
S.; Baxendale, I. R.; Ley, S. V.; Ladlow, M.; Welch, M.; Spring, D. R. Org. Biomol.
Chem. 2011, 9, 57–61.
12. Scherkenbeck, J.; Lüttenberg, S.; Ludwig, M.; Brücher, K.; Kotthaus, A. Eur. J. Org.
Chem. 2012, 1546–1553.
of anhydride 6 (Scheme 1). Hydrogenolysis and acid-catalyzed
deprotection of 9 provided acid 10 and amine salt 11 respectively,
which were then coupled using Ghosez’s chloroenamine re-
agent23,24 to provide tetradepsipeptide 12. Tetradepsipeptide 12
was subjected to hydrogenolysis and coupled to a second equiva-
lent of amine salt 11 to provide hexadepsipeptide 13. Global
deprotection of 13 by hydrogenolysis and acidification provided a
linear amino acid HCl salt25 which was cyclized with Ghosez’s
reagent with the assistance of a tertiary amine base to provide
synthetic (À)-1, which was identical in all respects to a sample of
enniatin B.26
13. Albericio, F.; Burger, K.; Ruiz-Rodriguez, J.; Spengler, J. Org. Lett. 2005, 7, 597–
600. and references cited therein..
14. Teixido, M.; Albericio, F.; Giralt, E. J. Pept. Res. 2005, 65, 153–166.
15. Slurries and bulk solids can be handled in laboratory flow reactors with
)
agitating reactors or ultrasound, (see Ref.16 or monoliths (see Ref.17);
precipitation events which generate small particles out of homogenous
solution, however, bypass filters, damage pumps, and cause clogging in thin
flow lines during multi-step synthesis.
16. (a) Sedelmeier, J.; Ley, S. V.; Baxendale, I. R.; Baumann, M. Org. Lett. 2010, 12,
3618–3621; (b) Noel, T.; Naber, J. R.; Hartman, R. L.; McMullen, J. P.; Jensen, K.
F.; Buchwald, S. L. Chem. Sci. 2011, 2, 287–290; (c) Browne, D. L.; Deadman, B. J.;
Ashe, R.; Baxendale, I. R.; Ley, S. V. Org. Proc. Res. Dev. 2011, 15, 693–697.
In summary, a convenient batch total synthesis of (À)-enniatin
B from (2R)-2-hydroxy-3-methylbutanoic acid has been described

D. X. Hu et al. / Tetrahedron Letters 53 (2012) 4077–4079
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17. Baumann, M.; Baxendale, I. R.; Ley, S. V.; Nikbin, N.; Smith, C. D. Org. Biomol.
Chem. 2008, 6, 1587–1593.
18. Shemyakin, M. M.; Ovchinnikov, Y. A.; Kiryushkin, A. A.; Ivanov, V. T. Izv. Akad.
Nauk SSSR: Ser. Khim. 1963, 9, 1623–1630.
19. Shemyakin, M. M.; Ovchinnikov, Y. A.; Kiryushkin, A. A.; Ivanov, V. T.
Tetrahedron. Lett. 1963, 14, 885–890.
20. Plattner, P. A.; Vogler, K.; Studer, R. O.; Quitt, P.; Keller-Schierlein Helv. Chim.
Acta 1963, 98, 927–935.
22. Kogan, T. P.; Somers, T. C.; Venuti, M. C. Tetrahedron 1990, 46, 6623–6632.
23. Ghosez, L.; Haveuaux, B.; Viehe, H. G. Angew. Chem., Int. Ed. 1969, 8,
454–455.
24. Ghosez, L.; George-Koch, I.; Patiny, L.; Houtekie, M.; Bovy, P.; Nshimyumukiza,
P.; Phan, T. Tetrahedron 1998, 54, 9207–9222.
25. The use of the linear amino acid’s HCl salt in the macrocyclization step was
important in protecting the N-terminus from premature reaction with
Ghosez’s reagent.
21. Nakatsu, S.; Yamada, K.; Oku, H.; Katakai, R. Proceedings of the International
Conference of the 43rd Japanese Peptide Symposium and 4th Peptide
Engineering Meeting 2006, 66–67.
26. One milligram from Sigma–Aldrich Corporation, from Gnomonia errabundia;
see Supplementary data.