Synthesis of ten members of the maradolipid family; Novel diacyltrehalose glycolipids from caenorhabditis elegans

Article abstract of DOI:10.1055/s-0030-1260318

The synthesis of ten members of the maradolipid family is described using a direct route starting from trehalose. Georg Thieme Verlag Stuttgart · New York.

Full text of DOI:10.1055/s-0030-1260318

2482
LETTER
Synthesis of Ten Members of the Maradolipid Family; Novel Diacyltrehalose
Glycolipids from Caenorhabditis elegans
Synthesis of
T
en M
l
ember
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s
of the
M
arado
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lipid Familye Pässler,a Margit Gruner,a Sider Penkov,b Teymuras V. Kurzchalia,*b Hans-Joachim Knölker*a
a
Department Chemie, Technische Universität Dresden, Bergstr. 66, 01069 Dresden, Germany
Fax +49(351)46337030; E-mail: hans-joachim.knoelker@tu-dresden.de
b
Max Planck Institute for Molecular Cell Biology and Genetics, Pfotenhauer Str. 108, 01307 Dresden, Germany
Received 25 July 2011
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Abstract: The synthesis of ten members of the maradolipid family
is described using a direct route starting from trehalose.
R2
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R1
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Key words: acylation, carbohydrates, lipids, natural products,
regioselectivity
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1 maradolipids
On exposure to difficult environmental conditions, for ex-
ample overcrowding or starvation, Caenorhabditis ele-
gans leaves the reproductive life cycle by generating
specific dauer larvae.1 These larvae have their own mor-
phology and enhanced stress metabolism to sustain them-
selves under unfavorable conditions. The biology of dauer
larva formation has been extensively investigated, howev-
er, the chemistry that supports their survival under harsh
conditions is not understood. During our studies on the
transition of C. elegans from reproductive larvae to dauer
larvae,2 we have recently identified a novel class of gly-
colipids called maradolipids (from maradi, Georgian for
enduring or dauer).3 These maradolipids represent a gen-
uine lipid component that is specific for dauer larvae. Ex-
tensive mass and NMR spectroscopic studies led to the
structural assignment of the maradolipids as 6,6¢-di-O-
acyltrehaloses 1 (Figure 1). It was found that the marado-
lipids are composed of a mixture of at least 60 derivatives
of trehalose 2, which differ in the two fatty acid acyl side
chains at the 6- and 6¢-positions. The fatty acid side chains
can be identical (R1 = R2, symmetrical maradolipids),
however, in the majority they differ (R1 π R2, unsymmet-
rical maradolipids). The structure of the maradolipids is
similar to glycolipids in Mycobacterium tuberculosis.4
However, to the best of our knowledge, this was the first
isolation of diacyltrehaloses from animals.
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2 trehalose
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About two thirds of the maradolipid mixture isolated from
C. elegans contained at least one branched fatty acid side
chain and one third of the maradolipids contained only
straight and cyclopropane-containing fatty acid side
chains. The maradolipid mixture found in C. elegans con-
tains about 7 to 8% of 6-O-(13-methylmyristoyl)-6¢-O-
oleoyltrehalose (1a; Mar 15:0/18:1) as the major compo-
nent. A selective access to the individual compounds of
the maradolipid mixture was required in order to confirm
the structural assignment and to identify their biological
HO
1a Mar 15:0/18:1
Figure 1 Structures of the maradolipids (1; R1COOH, R2COOH = fat-
ty acids), trehalose (2) and the most abundant maradolipid: Mar 15:0/
18:1 (1a).
function. Considering the structural diversity of the mara-
dolipids due to the different fatty acid side chains, we
have developed a synthetic strategy that provides the indi-
vidual members of this class of glycolipids in pure form.
Our preliminary results are reported here.
SYNLETT 2011, No. 17, pp 2482–2486
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Advanced online publication: 19.09.2011
DOI: 10.1055/s-0030-1260318; Art ID: B14311ST
© Georg Thieme Verlag Stuttgart · New York
Using an excess of N,O-bis(trimethylsilyl)acetamide and
catalytic amounts of tetrabutylammonium fluoride

LETTER
Synthesis of Ten Members of the Maradolipid Family
2483
HO
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Scheme 1 Synthesis of the symmetrical maradolipids (1). Reagents and conditions: (a) N,O-bis(trimethylsilyl)acetamide (8.9 equiv), TBAF
(0.06 equiv), DMF, 15 °C to r.t., 45 min; (b) K2CO3 (1.0 equiv), MeOH, 0 °C, 140 min, 88% over two steps; (c) RCOOH (4 equiv; fatty acid),
EDC⋅HCl (3 equiv), cat. DMAP, toluene, 50 °C, 4 d, 73–92%; (d) 50 equiv TFA–THF–H2O (1:4:6), 20 °C, 3 min, then evaporation, 81–90%.
(TBAF),5 trehalose (2) was transformed into the corres- For the synthesis of the unsymmetrical maradolipids, a
ponding octakis(trimethylsilyl)trehalose (3), which was differentiation between the two primary hydroxy groups
converted into the hexakis(trimethylsilyl)trehalose 4 in of trehalose (2) had to be achieved. Thus, a modified ap-
situ by chemoselective cleavage of the silyl ethers at the proach that could be used for selective monoacylation was
primary alcohol functions (Scheme 1).6 For the synthesis required (Scheme 2). The fatty acids were commercially
of the symmetrical maradolipids, compound 4 was treated available except for the cyclopropano-fatty acid for the
with an excess of the corresponding fatty acid and 1-ethyl- synthesis of the maradolipid 1j, which was easily pre-
3-(3-dimethylaminopropyl)carbodiimide (EDC) hydro- pared by cyclopropanation of oleic acid using Charette’s
chloride in the presence of catalytic amounts of 4-(N,N- procedure.7 The first fatty acid 6 (oleic acid or 13-meth-
dimethylamino)pyridine (DMAP) to afford the diacyl de- ylmyristic acid) was treated with two equivalents of
rivative 5. Cleavage of the silyl ether protecting groups hexakis(trimethylsilyl)trehalose 4 in the presence of 2.5
using a large excess of trifluoroacetic acid (TFA) and sub- equivalents of EDC hydrochloride and catalytic amounts
sequent purification by HPLC led to the symmetrical ma- of DMAP to provide the monoacyl derivative 7. The
radolipids 1b–e in four steps and 55–73% overall yield second acylation was achieved by reaction with an excess
based on trehalose (2) (Figure 2).
of the second fatty acid (R2COOH) and EDC hydrochlo-
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Scheme 2 Synthesis of the unsymmetrical maradolipids (1). Reagents and conditions: (a) 4 (2 equiv), EDC⋅HCl (2.5 equiv), cat. DMAP,
toluene, 50 °C, 4 d, 54%; (b) R2COOH (3 equiv), EDC⋅HCl (2.0 equiv), cat. DMAP, toluene, 50 °C, 4 d, 97–100%; (c) 50 equiv TFA–THF–
H2O (1:4:6), 20 °C, 3 min, then evaporation, 73–84%.
Synlett 2011, No. 17, 2482–2486 © Thieme Stuttgart · New York

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U. Pässler et al.
LETTER
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1b Mar 14:0/14:0
1c Mar 15:0/15:0
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1e Mar 18:1/18:1
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1j Mar 18:1/19:1
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Figure 2 Maradolipids 1a–j obtained by synthesis from trehalose (2). The symmetrical maradolipids 1b–e were synthesized following the
route shown in Scheme 1; the unsymmetrical maradolipids 1a and 1f–j were synthesized following the route shown in Scheme 2.
ride in the presence of DMAP to provide the diacyl deriv- cal maradolipids 1a and 1f–j in five steps and 34–40%
ative 8 almost quantitatively. Finally, deprotection with overall yield based on trehalose (2) (Figure 2).
TFA and purification by HPLC afforded the unsymmetri-
Synlett 2011, No. 17, 2482–2486 © Thieme Stuttgart · New York

LETTER
Synthesis of Ten Members of the Maradolipid Family
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Figure 3 Comparison of the 500 MHz 1H NMR spectrum (CD3OD) of the natural maradolipid mixture (top spectrum) with that of 6-O-(13-
methylmyristoyl)-6¢-O-oleoyltrehalose (1a; Mar 15:0/18:1; bottom spectrum).
The maradolipids 1a–j that have been synthesized in this found that dauer larvae of the type daf-2;DDtps, a double-
study were fully characterized from their spectroscopic mutant strain that cannot produce maradolipids due to in-
data (see the Supporting Information for complete 1H and hibition of trehalose biosynthesis,3 took up and accumu-
13C NMR spectra of 1a–j). A comparison of the original lated 1b (Figure 5).
1H NMR spectrum of the maradolipid mixture obtained
from C. elegans with that of 6-O-(13-methylmyristoyl)-
6¢-O-oleoyltrehalose (1a), obtained by the synthesis de-
scribed above, confirmed the assignment of the signals as
reported in our preceding paper (Figure 3).3
Figure 5 Left: 2D-TLC plate of the lipid extracts of the double-
mutant strain dauer larvae daf-2;DDtps; right: 2D-TLC plate of the li-
pid extracts of the double-mutant strain dauer larvae daf-2;DDtps fed
with the synthetic maradolipid 1b (Mar 14:0/14:0); the arrowhead indi-
cates the band of the maradolipid.
Figure 4 HMBC spectrum (150/600 MHz, CD3OD) of 6-O-(13-
methylmyristoyl)-6¢-O-oleoyltrehalose (1a; Mar 15:0/18:1).
In conclusion, we have developed a highly efficient syn-
thetic route to symmetrical as well as to unsymmetrical
The HMBC spectrum of pure synthetic 1a (150/600
maradolipids 1. Our present work provides broad access
MHz), clearly shows the difference between the chemical
to these novel glycolipids for the study of their biological
shifts of the two fatty acid carbonyl groups at the 6-O- and
function as well as their biophysical properties.
6¢-O-positions of trehalose (Figure 4; compare with
Figure 3b of our previous paper3).
Supporting Information for this article is available online at
http://www.thieme-connect.com/ejournals/toc/synlett.
When worms were fed with the synthetic maradolipid 1b
(Mar 14:0/14:0) to form dauer larvae at 25 °C, it was
Synlett 2011, No. 17, 2482–2486 © Thieme Stuttgart · New York

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U. Pässler et al.
LETTER
Theumer, G.; Riezmann, I.; Riezmann, H.; Knölker, H.-J.;
Kurzchalia, T. V. Dev. Cell 2009, 16, 833. (d) Martin, R.;
Entchev, E. V.; Däbritz, F.; Kurzchalia, T. V.; Knölker,
H.-J. Eur. J. Org. Chem. 2009, 3703. (e) For a review, see:
Martin, R.; Entchev, E. V.; Kurzchalia, T. V.; Knölker, H.-J.
Org. Biomol. Chem. 2010, 8, 739.
Acknowledgment
We are grateful to ESF EuroMembrane Network (DFG grant KN
240/13-1) for financial support. We thank P. Linning for HPLC
purifications.
(3) Penkov, S.; Mende, F.; Zagoriy, V.; Erkut, C.; Martin, R.;
Pässler, U.; Schuhmann, K.; Schwudke, D.; Gruner, M.;
Mäntler, J.; Reichert-Müller, T.; Shevchenko, A.; Knölker,
H.-J.; Kurzchalia, T. V. Angew. Chem. Int. Ed. 2010, 49,
9430; Angew. Chem. 2010, 122, 9620.
(4) Noll, H.; Bloch, H.; Asselineau, J.; Lederer, E. Biochim.
Biophys. Acta 1956, 20, 299.
(5) Johnson, D. A. Carbohydr. Res. 1992, 237, 313.
(6) Datta, A. K.; Takayama, K.; Nashed, M. A.; Anderson, L.
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(7) Charette, A. B.; Francoeur, S.; Martel, J.; Wib, N. Angew.
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References and Notes
(1) Antebi, A.; Culotti, J. G.; Hedgecock, E. M. Development
1998, 125, 1191.
(2) (a) Maytash, V.; Entchev, E. V.; Mende, F.; Wilsch-
Bräuninger, M.; Thiele, C.; Schmidt, A. W.; Knölker, H.-J.;
Ward, S.; Kurzchalia, T. V. PloS Biol. 2004, 2, 1561.
(b) Martin, R.; Schmidt, A. W.; Theumer, G.; Krause, T.;
Entchev, E. V.; Kurzchalia, T. V.; Knölker, H.-J. Org.
Biomol. Chem. 2009, 7, 909. (c) Hannich, J. T.; Entchev,
E. V.; Mende, F.; Boytchev, H.; Martin, R.; Zagoriy, V.;
Synlett 2011, No. 17, 2482–2486 © Thieme Stuttgart · New York

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