Segment solid-phase total synthesis of the anthelmintic cyclooctadepsipeptides PF1022A and emodepside

Article abstract of DOI:10.1002/ejoc.201101421

Cyclodepsipeptides of the enniation, PF1022 and verticilide families represent a diverse class of highly interesting natural products with respect to their manifold biological activities. However, until now, no practicable solid-phase syntheses of these compounds have been accomplished, probably due to the problematic combination of N-methyl amino acids and hydroxycarboxylic acids. We report herein an efficient synthesis of the anthelmintic PF1022A and its commercial analogue emodepside on Kaiser and Wang resins. Our protocol provides the basis for the solid-phase synthesis of cyclodepsipeptide libraries with a high probability of anthelmintic, antibacterial or insecticidal activity.

Full text of DOI:10.1002/ejoc.201101421

FULL PAPER
DOI: 10.1002/ejoc.201101421
Segment Solid-Phase Total Synthesis of the Anthelmintic
Cyclooctadepsipeptides PF1022A and Emodepside
Jürgen Scherkenbeck,*[a] Sebastian Lüttenberg,[a] Monika Ludwig,[a] Karin Brücher,[a] and
Andreas Kotthaus[a]
Keywords: Natural products / Solid-phase synthesis / Total synthesis / Amino acids / Peptides
Cyclodepsipeptides of the enniation, PF1022 and verticilide
families represent a diverse class of highly interesting natural
products with respect to their manifold biological activities.
However, until now, no practicable solid-phase syntheses of
these compounds have been accomplished, probably due to
the problematic combination of N-methyl amino acids and
hydroxycarboxylic acids. We report herein an efficient syn-
thesis of the anthelmintic PF1022A and its commercial ana-
logue emodepside on Kaiser and Wang resins. Our protocol
provides the basis for the solid-phase synthesis of cyclodepsi-
peptide libraries with a high probability of anthelmintic, anti-
bacterial or insecticidal activity.
Introduction
Nematode infections cause tremendous health problems
in livestock and domestic animals. In humans they are a
major cause of morbidity and loss of disability-adjusted life
years.[1,2] Around two billion people are thought to have
active nematode infections. According to the World Health
Organization, parasitic worm infections are one of the most
important classes of neglected tropical diseases.[3] Since the
early 1960s three different types of broad-spectrum anthel-
mintics have become available, the benzimidazoles, the nico-
tinic acetylcholine receptor agonists and the macrocyclic
lactones.[4] In particular, in the livestock industry, anthel-
mintic treatment has become problematic due to the growth
in resistance against these commonly used anthelmintics.
The 24-membered cyclooctadepsipeptide PF1022A (1), a
metabolite of Mycelia sterilia (Rosselinia sp.) originally iso-
lated from the leaves of Camellia japonica, has been estab-
lished as a novel, resistance-breaking anthelmintic with low
toxicity in animals (Figure 1).[5,6] A semi-synthetic analogue
of PF1022A, emodepside (2), has been introduced into the
market recently for the treatment of parasitic helminth in-
fections in companion animals (Figure 1).[7]
Figure 1. Structures of PF1022A (1) and emodepside (2).
published, but not a total synthesis of PF1022A itself or
its commercial analogue emodepside, although solid-phase
synthesis is the method of choice not only for the prepara-
tion of peptides but also, with some limitations, of depsi-
peptides.[11,12]
However, solid-phase depsipeptide synthesis turns out to
be considerably more difficult than conventional peptide
couplings due to the generally lower state-of-the-art meth-
odology in protecting-group chemistry and the lack of high-
yielding coupling reagents for the formation of ester bonds.
The solid-phase total synthesis of valinomycin by Kuisle et
al. is one of the rare examples in which all the ester bonds
were formed on the resin.[13] A more frequent strategy re-
stricts the couplings on the resin to the formation of amide
bonds by using depsipeptide building blocks prepared in
solution.[14]
A prerequisite for the development of a PF1022A-based
anthelmintic against human worm infections is an efficient
procedure that allows the synthesis of a large number of
PF1022 analogues for biological screening. Despite several
published total syntheses in solution and an in vitro synthe-
sis approach, only a limited number of PF1022A derivatives
have become available.[5,8–10] Remarkably, until now only a
partial solid-phase synthesis of PF1022 analogues has been
Also to be taken into account is the fact that couplings
of N-methyl-substituted amino acids generally afford lower
yields and increased racemization due to the steric hin-
drance caused by the bulky methyl groups.[15] That is espe-
cially true for syntheses on solid supports, as the reactivity
[a] Department of Chemistry, University of Wuppertal,
Gaußstraße 20, 42119 Wuppertal, Germany
Fax: +49-202-439-3464
E-mail: Scherkenbeck@uni-wuppertal.de
1546
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2012, 1546–1553

Total Synthesis of Anthelmintic Cyclooctadepsipeptides
of resin-bound secondary amines towards activated acids is Boc-l-MeLeu-d-morphPhLac-OH) are required. These are
even lower than in solution. Although this problem has available in high yields by solution synthesis.
been known for decades, there is no single method of
choice. In solid-phase syntheses of the cyclopeptides cyclo-
sporine and omphalotine A, triphosgene has been em-
ployed successfully to couple two adjacent N-methylvalines
whereas in the solution synthesis of thiocoraline the more
conventional HATU/HOAt (see the Exp. Sect. for the defi-
nitions of reagents) reagent afforded good yields.[16–18] Tri-
azine-based coupling reagents introduced recently provide
a new concept as they form a cyclic intermediate that
tethers the two reacting centres and thus facilitates the cou-
pling through an intramolecular pathway.[19]
An instructive example of the problems arising from N-
methyl amino acids can be found in a paper by Lee describ-
ing the synthesis of conformationally restricted PF1022 an-
alogues on Kaiser oxime. Starting with the Pro-Lac resin,
already the removal of the first protecting group and cou-
pling with Boc-MeLeu-PhLac-OH resulted in extensive for-
mation of the Pro-Lac morpholinedione accompanied by
immobilized Boc-MeLeu-PhLac and some tetradepsi-
peptide.[11,12] The problem of morpholinedione formation
was “solved” by attaching a complete tetradepsipeptide to
the resin, which reduces the on-resin chemistry to only two
elongation steps. Also, from another aspect this synthesis
cannot be regarded as a general route for the solid-phase
synthesis of natural PF1022A or emodepside because all
analogues contained a turn-inducing building block, which
facilitates the macrocyclization reaction considerably.
Results and Discussion
Synthesis of the Didepsipeptide Building Blocks
The didepsipeptides 8, 9 and 15–17, used as starting ma-
terials for the solid-phase synthesis, were prepared in good-
to-excellent yields (Scheme 1) from Boc- or Fmoc-protected
N-Me-Leu and the benzyl-protected lactates 5, 10 and 11
following literature procedures.[20–23] Using the inexpensive
natural (S)-lactic acid enantiomer, Mitsunobu conditions
were applied to establish the correct R configuration during
the coupling reaction of (S)-benzyl lactate (5) with N-Me-
leucines 3 and 4 to afford the coupling products 6 and 7.
In the case of phenyllactic acid the R enantiomer was used
directly due to its lower cost. Thus, coupling of the (R)-
phenyllactic acids 10 and 11 with DCC afforded the dide-
psipeptides 12–14. The benzyl protecting group was re-
moved by hydrogenolysis with Pd/C or preferably with the
Pearlman catalyst to give the corresponding acids in excel-
lent yields.[24] No epimerizations were observed in either the
coupling or the hydrogenolysis steps.
Solid-Phase Synthesis on Kaiser Resin
The Kaiser oxime was used as the resin of choice because
We wish to report herein an efficient solid-phase synthe- it allows a cyclizative cleavage and can be reused without
sis of PF1022A and emodepside based on didepsipeptide regeneration, an important prerequisite for solid-phase syn-
coupling reactions. Following this strategy, the synthetic thesis on a multigram scale.[25] However, piperazinedione
challenge is reduced to on-resin amide formation between formation is a notorious side-reaction of the Kaiser oxime
an N-methyl amino acid and a hydroxycarboxylic acid. Due because the oxime ester can be cleaved even with weak nu-
to the C2 symmetry of PF1022A and emodepside, only two cleophiles. Extensive studies have demonstrated the critical
different didepsipeptides (Boc-l-MeLeu-d-Lac-OH and role of the first and second amino acid as well as the cou-
Boc-l-MeLeu-d-PhLac-OH or Boc-l-MeLeu-d-Lac-OH, pling conditions. Bulky amino acids such as Val, Leu or
Scheme 1. Preparation of the didepsipeptide segments in solution. Reagents and conditions: (a) TPP, DEAD, THF, 0 °C for 2 h, then
room temp. for 1 h; (b) H2, Pd/C (10%), EtOH, room temp., 4 h for 8 and 15–17; H2, Pd(OH)2/C (20%), EtOH, room temp., 4 h for 9;
(c) DCC, HOBt, DMAP, DCM, 0 °C, 30 min, then room temp. for 24 h.
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J. Scherkenbeck et al.
FULL PAPER
Phe in the first position and N-methyl amino acids in the reaction. In addition, the liberated oxime resin reacted im-
second position, exactly the situation in the PF1022A syn- mediately with the second didepsipeptide Boc-MeLeu-Lac-
thesis, generally facilitate piperazinedione formation.[26–28]
OH to form a deletion sequence. However, the cleavage re-
As expected, the yields for the coupling of the didepsi- action was strongly dependent on the type of coupling rea-
peptides 8, 15 and 16 to the Kaiser resin were strongly de- gent, the solvent and auxiliary base. Fortunately, HATU/
pendent on the didepsipeptide used as the first building DIEA, already used successfully for immobilizing the first
block, the coupling reagents and the solvents (Table 1). The building block (Boc-MeLeu-PhLac-OH), also proved the
best yields, determined by picric acid titration after depro- best coupling reagent for chain elongation to the tetradepsi-
tection, were obtained with the didepsipeptide Boc-MeLeu- peptide 24 (Table 2). In general, the use of HATU in the
PhLac (15) by using HATU as the coupling reagent synthesis of both PF1022A and emodepside led to coupling
(Table 1, entry 6).[29,30] DIC, which was employed by Lee, yields in the range of 75–90%, which can be regarded as
gave in our experiments only poor yields (19%) with both more than satisfactory for solid-phase couplings of N-
didepsipeptides 8 and 15.[11,12] In general, the didepsi- methyl amino acids (Scheme 2). However, we were not able
peptide Boc-MeLeu-Lac (8) provided lower yields for the to suppress the cleavage of the N-terminal didepsipeptides
coupling to the resin (Table 1, entry 4) and tended to exten- completely.
sive diketopiperazine formation in the subsequent coupling
to the tetradepsipeptide 21. The yields for the coupling of
the morpholino-substituted didepsipeptide 16 to the Kaiser
oxime were similar.
During chain elongation to the tetradepsipeptide Me-
Leu-Lac-MeLeu-PhLac, cleavage of the didepsipeptide Me-
Leu-PhLac from the solid support with the formation of
the corresponding dioxomorpholine became a major side-
A particular feature that characterizes the Kaiser resin is
the anchoring oxime ester bond, which, as a polymeric
active ester, can be cleaved by solvolysis or aminolysis to
afford peptide esters or amides.[31–33] The reactivity of the
oxime ester towards nucleophiles also allows cyclizative
cleavage, which releases the cyclodepsipeptide directly into
solution without the need for an additional cyclization reac-
tion. A limited loading of the resin (0.3 mmol/g resin) pro-
vides the high-dilution environment needed for the macro-
cyclization reaction. Cyclizative cleavages of cyclopeptides,
but not of cyclodepsipeptides, have been accomplished un-
der a variety of reaction conditions.[34–36] After some exper-
imentation (Table 2) we found a weakly acidic solution of
DIEA (2.2 equiv.) and HOAc (5.5 equiv.) in DCM best
suited for the cyclizative cleavage of the linear octadepsi-
peptides 27 and 28 (Scheme 2), whereas neutral (ethyl acet-
ate) or weakly basic (pyridine) reaction conditions afforded
drastically reduced yields (10 and 27%, respectively).[27]
The unusually high yield (75%, based on the loading of
the linear precursor 27) of the ring-closing reaction to give
PF1022A (1) can be attributed to a cis-amide bond between
one leucine and one lactic acid residue. This positions the
N- and C-terminus in close proximity and thus facilitates
the macrocyclization reaction.[37] The raw PF1022A (1) had
a purity of at least 80% and can be further purified by flash
chromatography to afford an overall yield of 30%, which
is around three times more than the yield of the solution
synthesis.[37] Note, the macrocyclization yield declined sig-
nificantly when Kaiser resins with loadings of 1 mmol/g
resin or more were used, probably due to intermolecular
coupling reactions with the resin.[34]
Table 1. Yields for the coupling of didepsipeptides 8 and 15 to the
Kaiser resin under different conditions.
Entry
1
Didepsipeptide
(amount [equiv.]) couplings
Number of Conditions
Solvent Yield
[%]
8 (2)
1
2 equiv. HATU, DMF
65
2 equiv. HOBt,
5 equiv. DIEA
3 equiv. DIC,
3 equiv. DMAP
3 equiv. HATU, DMF
3 equiv. HOBt,
2
3
8 (2)
8 (3)
1
1
DMF
19
70
8 equiv. DIEA
4
5
6
7
8
9
8 (2)
15 (1)
15 (2)
15 (1)
15 (3)
15 (3)
1
2
1
2
1
1
2 equiv. HATU
3 equiv. DIPEA
1 equiv. TBTU,
6 equiv. NMM
2 equiv. HATU, DCM
3 equiv. DIEA
1 equiv. TBTU,
3 equiv. NMM
3 equiv. HATU, DMF
6 equiv. DIEA
DCM
DMF
62
53
87
58
48
19
DMF
3 equiv. DIC,
3 equiv. HOBt,
6 equiv. DIEA
DMF
Table 2. Chain elongation to the tetradepsipeptide Boc-MeLeu-Lac-MeLeu-PhLac-resin (24).
Entry
Lac-Leu [equiv.]
Coupling reagent
(amount [equiv.])
Base (amount [equiv.])
Solvent
Fragments[a]
1
2
3
4
5
6
7
2
5
5
5
2
3
3
TBTU (2)
TBTU (5)
TBTU (5)
TBTU (5)
BTFFH (5)
HATU (2)
HATU (2)
NMM (6)
DMF
50% B, 45% AB
100% B
100% B
NMM (6.6)
NMM (6.6)
NMM (6.6)
DIEA (10)
DIEA (6.5)
DIEA (6.5)
DMSO
nitrobenzene
acetone
DMF
DMF
DCM
100% B
80% B, 20% AB
90% AB, 10% B
70% AB, 30% B
[a] HPLC–MS of test cleavages with morpholine; B: morpholide of MeLeu-Lac; AB: morpholide of MeLeu-Lac-MeLeu-PhLac.
1548 Eur. J. Org. Chem. 2012, 1546–1553
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© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Total Synthesis of Anthelmintic Cyclooctadepsipeptides
Scheme 2. Chain elongation steps and cyclization reactions. Reagents and conditions: (a) for 21, 22: i. 25% TFA/DCM, room temp.,
30 min; ii. Boc-MeLeu-Lac-OH (8), HATU, DIEA, DMF, room temp., 16 h; iii. 25% TFA/DCM, room temp., 30 min; for 23: i. 25%
piperidine/THF, room temp., 30 min; ii. Fmoc-MeLeu-Lac-OH (9), HATU, DIEA, THF, room temp., 16 h; iii. 25% piperidine/THF,
room temp., 30 min; (b) for 24: i. Boc-MeLeu-PhLac-OH (15), HATU, DIEA, DMF, room temp., 16 h; ii. 25% TFA/DCM, room temp.,
30 min; for 25: i. Boc-MeLeu-morphPhLac-OH (16), HATU, DIEA, DMF, room temp., 16 h; ii. 25% TFA/DCM, room temp., 30 min;
for 26: i. Fmoc-MeLeu-PhLac-OH (17), HATU, DIEA, THF, room temp., 16 h; ii. 25% piperidine/THF, room temp., 30 min; (c) for 27,
28: i. Boc-MeLeu-Lac-OH (8), HATU, DIEA, DMF, room temp., 16 h; ii. 25% TFA/DCM, room temp., 30 min; for 29: i. Fmoc-MeLeu-
Lac-OH (9), HATU, DIEA, THF, room temp., 16 h; ii. 25% piperidine/THF, room temp., 30 min; (d) with Kaiser oxime: DIEA
(2.2 equiv.), AcOH (5 equiv.), DCM, room temp., 16 h; with Wang resin: i. 50% TFA/DCM, room temp., 1 h; ii. BOPCl, DIEA, room
temp., 48 h.
Remarkably, emodepside (2) was obtained in an even lated cyclodepsipeptides because excellent protocols for
higher yield (40–45%) under the coupling and cyclizative solution cyclizations exist.[37,38]
cleavage conditions developed for PF1022A. The overall
In contrast to the Kaiser oxime, a DIC/DMAP/HOBt
yield for emodepside complies with a coupling yield of coupling reagent combination was found to give the highest
more than 90% for each step, including the critical macro- yields (93%) in the anchoring step, but the HATU/DIEA
cyclization reaction.
system proved best again for the amide couplings. However,
the yields of the amide-forming reactions were generally
lower on the Wang resin than on the Kaiser resin. Although
the yield of the tetradepsipeptide Fmoc-MeLeu-Lac-Me-
Solid-Phase Synthesis on Wang Resin
A solid-phase synthesis on Wang resin with a comple- Leu-PhLac was still in the range of 80%, increasing
mentary, base-labile protecting group strategy (Fmoc) pro- amounts of failure and deletion sequences accompanied by
vides access to PF1022 analogues that are not accessible by minor side-products resulting from cleavage of the ester
the Kaiser oxime route. In addition, the Wang resin has a bonds were identified in the couplings to hexadepsipeptide
higher loading and the anchor linkage is more stable, which 26 and, in particular, to octadepsipeptide 29, which was ob-
reduces the formation of dioxomorpholine and premature tained in only around 50% yield from the hexadepsipeptide.
cleavage from the resin during the chain-elongation steps. Cleavage of the octadepsipeptide 29 from the resin with
On the other hand, the more stable benzyl ester linkage TFA gave a raw material with a linear octadepsipeptide
renders a cyclizative cleavage unfeasible. This is, however, content in the range of 40–50%. Subsequent solution cycli-
unproblematic in the particular case of PF1022A and re- zation (62% yield) of the crude octadepsipeptide with
Eur. J. Org. Chem. 2012, 1546–1553
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J. Scherkenbeck et al.
FULL PAPER
phenylphosphane (6.30 g, 24.0 mmol) were dissolved in THF
(12 mL). At 0 °C a solution of DEAD (3.78 mL, 24.0 mmol) in
THF (12 mL) was added dropwise through a syringe pump during
2 h. The solution was stirred for 1 h at room temperature, diluted
with ethyl acetate and washed twice with a saturated NaHCO3
solution (18 mL each) and 0.5 m citric acid (18 mL each). The or-
ganic phase was dried with Na2SO4 and the solvents evaporated.
Flash chromatography (cyclohexane/ethyl acetate, 8:2) of the resi-
BOPCl according to
a literature procedure afforded
PF1022A in an overall yield of 20–25% (Scheme 2) after
chromatographic purification.[37]
Conclusion
We have developed an efficient solid-phase synthesis of
the anthelmintic cyclooctadepsipeptide PF1022A and its due afforded the protected didepsipeptide 6 (6.02 g, 74%) as a
colourless oil. 1H NMR (400 MHz, CDCl3): δ = 0.91–0.97 (m, 6
H, CδH3-Leu), 1.45 (s, 9 H, CH3-tBu), 1.48–1.54 (2 m, 4 H, CγH-
Leu, CβH3-Lac), 1.62–1.73 (m, 2 H, CβH-Leu), 2.69–2.81 (m, 3 H,
commercial analogue emodepside by segment coupling re-
actions. After careful optimization of the oxime ester for-
mation, the N-methylleucine couplings and the cyclizative
NCH3), 4.68–4.80 (m, 1 H, CαH-Leu), 4.90–5.04 (m, 1 H, CαH-
cleavage, overall yields of 30–45% were obtained on Kaiser
Lac), 5.09–5.24 (m, 2 H, CH2-Bn), 7.29–7.39 (m, 5 H, Ar-H) ppm.
oxime. An analogous synthesis with complementary pro-
13C NMR (CD3CN): δ = 16.86, 23.19, 24.53, 24.80, 28.33, 29.81,
tecting group chemistry on Wang resin afforded somewhat
lower yields in the range of 20–30%. Nevertheless, the stan-
dard conditions used for the Wang synthesis should allow
unproblematic adaptation to an automated synthesizer, al-
30.23, 37.28, 37.63, 55.82, 56.75, 67.02, 69.06, 79.81, 80.14, 128.23,
128.44, 128.59, 135.24, 156.31, 170.12, 171.89 ppm. IR (film): ν =
˜
1693 (C=O), 1742 (C=O) cm–1. MS (ESI): m/z (%) = 837 (5), 408
(31), 352 (34), 308 (100). HRMS (ESI): calcd. for C22H33NNaO6
lowing the preparation of cyclodepsipeptide libraries for [M + Na]+ 430.2200; found 430.2204.
screening against human pathogenic worms and also for in-
Fmoc-L-MeLeu-D-Lac-OBn (7): Benzyl l-lactate (5; 7.00 g,
secticidal and antibacterial activities.
38.1 mmol), N-Fmoc-N-methyl-l-leucine (4; 13.99 g, 38.1 mmol)
and triphenylphosphane (11.98 g, 45.7 mmol) were dissolved in
THF (100 mL). A solution of DEAD (8.04 g, 45.7 mmol) in THF
(50 mL) was added at 0 °C over 2 h. The reaction was stirred for
1 h at room temperature, then diluted with ethyl acetate and
washed with a saturated NaHCO3 solution (50 mL), 0.5 m citric
acid (50 mL) and water (50 mL) twice each. Standard work-up and
chromatographic purification (cyclohexane/ethyl acetate, 9:1)
yielded the coupling product 7 as a viscous yellow oil (17.6 g, 87%).
1H NMR (400 MHz, CDCl3): δ = 0.81 and 0.91 (2 d, J = 6.7 Hz,
3 H, CδH3-Leu), 0.96 (pseudo-t, J = 6.0 Hz, 3 H, CδH3-Leu), 1.47–
1.74 (m, 6 H, CβH2-Leu, CγH-Leu, CβH3-Lac), 2.85 and 2.87 (2 s,
3 H, NCH3), 4.24 and 4.30 (2 m, 1 H, CH-Fmoc), 4.38 (m, 1 H,
CH2-Fmoc), 4.50 (m, 1 H, CH2-Fmoc), 4.66 and 5.04 (2 m, 1 H,
CαH-Leu), 5.07 and 5.14 (2 m, 1 H, CαH-Lac) 5.20 (m, 2 H, CH2-
Bn), 7.31 (m, 8 H, Ar-H), 7.57 (m, 3 H, Ar-H), 7.76 (m, 3 H, Ar-
H), ppm. 13C NMR (100 MHz, CDCl3): δ = 16.9, 21.4, 23.3, 24.7,
30.7, 37.2, 47.3, 56.5, 67.1, 67.7, 69.2, 120.0, 124.8, 125.0, 125.1,
127.0, 127.7, 128.2, 128.5, 128.6, 141.3, 143.9, 144.1, 156.9, 170.0,
Experimental Section
General: The starting materials (R)-3-phenyllactic acid, benzyl (R)-
3-phenyllactate, N-Fmoc-N-methyl-l-leucine and N-Boc-N-methyl-
l-leucine were either purchased or prepared by standard literature
procedures.[21–23] All reactions except the hydrogenations were per-
formed in dried solvents. Dichloromethane, DIEA and piperidine
were heated at reflux for 1 h over calcium hydride and distilled.
THF was heated at reflux for several hours over LiAlH4 and then
distilled. Toluene was dried by filtration through basic alumina.
The following instruments were used: 1H NMR: Bruker Avance
400, Bruker Avance III 600; 13C NMR: Bruker Avance 400, Bruker
Avance III 600. FTIR: Nicolet PROTÉGÉ 460 E.S.P. MS: Bruker
micrOTOF; ESI-MS: Varian IT 500-MS Iontrap. LC: preparative
low-pressure liquid chromatography (LPLC) was performed by
using silica gel 60 μm (230–400 mesh, Macherey–Nagel), Büchi
pump. TLC: silica gel 60 F254 (Merck). For preparative HPLC, a
Spectra Physics Analytical Spectra System AS 3000 instrument
with UV detection was used. The progress of the solid-phase reac-
tions was monitored by using either the Bromophenol Blue colour
test, determination of the resin-loading by UV analysis of the Fmoc
group cleavage product or by test cleavages and HPLC-MS analy-
sis.[39]
171.0 ppm. IR (KBr): ν = 1745 (C=O), 1704 (C=O) cm–1. MS
˜
(ESI): m/z (%) = 308 (14), 530 (100), 547 (50). HRMS (ESI): calcd.
for C32H35NNaO6 [M + Na]+ 552.2357; found 552.2356.
Boc-L-MeLeu-D-Lac-OH (8): A suspension of didepsipeptide 6
(4.08 g, 10.0 mmol) and Pd/C (10%, 0.5 g) in ethanol (60 mL) was
hydrogenated (1 bar) overnight at room temperature. After that
time the catalyst was filtered off and washed with ethanol. After
evaporation of the combined organic phases Boc-l-MeLeu-d-Lac-
OH (8; 3.17 g, 100%) was obtained as a light-grey oil. 1H NMR
(400 MHz, CDCl3): δ = 0.89–0.97 (m, 6 H, CδH3-Leu), 1.45 (s, 9
H, CH3-tBu), 1.52 (d, J = 7.0 Hz, 3 H, CβH3-Lac), 1.54–1.63 (m,
1 H, CγH-Leu), 1.62–1.79 (m, 2 H, CβH-Leu), 2.80 (s, 3 H, NCH3),
4.68–4.77 and 4.77–4.86 (2 m, 1 H, CαH-Leu), 5.10 (q, J = 7.0 Hz, 1
H, CαH-Lac), 8.39 (br. s, 1 H, COOH) ppm. 13C NMR (100 MHz,
CDCl3): δ = 16.78, 16.84, 21.09, 21.44, 23.09, 23.16, 24.55, 24.84,
28.31, 29.93, 30.99, 37.38, 37.54, 68.64, 68.71, 80.44, 155.86,
Abbreviations: ACN, acetonitrile; Bn, benzyl; Boc, tert-butyloxy-
carbonyl; BOPCl, N,NЈ-bis(2-oxo-3-oxazolidinyl)phosphinic chlor-
ide; DCC, N,NЈ-dicyclohexylcarbodiimide; DCM, dichlorometh-
ane; DEAD, diethyl azodicarboxylate; DIC, N,NЈ-diisopropyl-
carbodiimide; DIEA, diisopropylethylamine; DMAP, 4-dimethyl-
aminopyridine; DMF, dimethylformamide; DMSO, dimethyl sulf-
oxide; Fmoc, 9-fluorenylmethoxycarbonyl; HATU, N,N,NЈ,NЈ-tet-
ramethyl-O-(7-azabenzotriazol-1-yl)uranium hexafluorophosphate;
HOAc, acetic acid; HOAt, 1-hydroxy-7-azabenzotriazole; HOBt, 1-
hydroxybenzotriazole; MeOH, methanol; MTBE, methyl tert-butyl
ether; TEA, triethylamine; TFA, trifluoroacetic acid; THF, tetra-
hydrofuran; TPP, triphenylphosphane.
156.31, 171.67, 174.49, 174.72 ppm. IR (film): ν = 1668 (C=O),
˜
1697 (C=O), 1742 (C=O) cm–1. MS (ESI): m/z (%) = 655 (8), 317
(4), 316 (23), 244 (100), 170 (41). HRMS (ESI): calcd. for
C15H27NNaO6 [M + Na]+ 340.1731; found 340.1733.
Preparation of Building Blocks and Didepsipeptide Segments in
Solution
Boc-L-MeLeu-D-Lac-OBn (6): Benzyl l-lactate (5; 3.60 g,
Fmoc-L-MeLeu-D-Lac-OH (9): A suspension of benzyl ester 7
20.0 mmol), N-Boc-N-methyl-l-leucine (4.91 g, 20.0 mmol) and tri-
(19.71 g, 37.2 mmol) and Pd(OH)2/C (20%, 1.05 g) in ethanol
1550
www.eurjoc.org
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2012, 1546–1553

Total Synthesis of Anthelmintic Cyclooctadepsipeptides
(200 mL) was hydrogenated (1 bar H2) at room temperature for 4 h.
Then the catalyst was filtered and washed with ethanol. LPLC (cy-
clohexane/ethyl acetate, 8:2, + 0.1% acetic acid) of the crude prod-
uct, obtained after evaporation of the solvent, gave Fmoc-l-Me-
Leu-d-Lac-OH (9) as a white foam (11.76 g, 72%). 1H NMR
(400 MHz, CDCl3): δ = 0.79 and 0.91 (2 d, J = 7.0 Hz, 2 H, CδH3-
Leu), 0.94 (m, 4 H, CδH3-Leu), 1.51–1.78 (m, 6 H, CβH2-Leu, CγH-
Leu, CβH3-Lac), 2.87 and 2.89 (2 s, 3 H, NCH3), 4.25 and 4.29 (2
m, 1 H, CH-Fmoc), 4.37 and 4.51 (2 m, 2 H, CH2-Fmoc), 4.61 and
4.99 (2 m, 1 H, CαH-Leu), 5.02 and 5.13 (2 m, 1 H, CαH-Lac),
7.29 (m, 2 H, Ar-H), 7.40 (m, 2 H, Ar-H), 7.62 (m, 1 H, Ar-H),
7.79 (m, 2 H, Ar-H), 8.08 (br. s, 1 H, COOH) ppm. 13C NMR
(100 MHz, CDCl3): δ = 21.2, 23.0, 24.6, 30.4, 37.2, 47.2, 56.5, 67.7,
68.9, 120.0, 124.8, 125.1, 125.1, 127.1, 127.6, 127.7, 128.5, 141.4,
484 (43), 384 (100). HRMS (ESI): calcd. for C28H37NNaO6 [M +
Na]+ 506.2513; found 506.2520.
Boc-L-MeLeu-D-morphPhLac-OBn (13): MorphPhLac depsipept-
ide 13 was prepared following the protocol for compound 12. After
LPLC (cyclohexane/ethyl acetate, 1:1, + 0.1% TEA) a colourless
1
oil was obtained in a yield of 92%. H NMR (400 MHz, CDCl3):
δ = 0.90 (d, J = 7.9 Hz, 6 H, CδH3-Leu), 1.44 and 1.49 (2 s, 9 H,
CH3-tBu), 1.53–1.65 (m, 3 H, CγH-Leu, CβH-Leu), 2.64 and 2.66
(2 s, 3 H, NCH3), 2.99–3.09 (m, 2 H, CβH-PheLac), 3.08–3.13 (m,
4 H, NCH2-morpholine), 3.82–3.87 (m, 4 H, OCH2-morpholine),
4.71 and 4.99 (2 dd, J = 4.0, 5.0 Hz, 1 H, CαH-PhLac), 5.12 (m, 2
H, CH2-Bn), 5.17–5.26 (m, 1 H, CαH-Leu), 6.79 (d, J = 8.0 Hz, 2
H, Ar-H), 7.06 (d, J = 8.0 Hz, 2 H, Ar-H), 7.20–7.37 (m, 5 H, Ar-
H) ppm. 13C NMR (100 MHz, CDCl3): δ = 21.09, 21.34, 23.18,
24.45, 24.74, 28.40, 29.79, 30.08, 36.44, 37.65, 49.35, 60.32, 66.89,
67.11, 73.57, 79.75, 115.62, 126.72, 128.39, 128.53, 130.20, 130.32,
143.8, 156.4, 157.2, 171.5, 175.1, 177.1 ppm. IR (KBr): ν = 3160
˜
(O–H), 1746 (C=O), 1714 (C=O), 1667 (C=O) cm–1. MS (ESI): m/z
(%) = 179 (17), 218 (34), 440 (100), 457 (39). HRMS (ESI): calcd.
for C25H29NNaO6 [M + Na]+ 462.1887; found 462.1888.
135.00, 150.30, 169.00, 171.74 ppm. IR: ν = 1669 (C=O), 1752
˜
(C=O) cm–1. MS (ESI): m/z (%) = 591 (2), 570 (35), 569 (100), 513
(35). HRMS (ESI): calcd. for C32H44N2NaO7 [M + Na]+ 591.3041;
found 591.3052.
Benzyl (R)-2-Hydroxy-3-(4-morpholinophenyl)propanoate (11): A
suspension of (R)-3-(4-morpholino)phenyllactic acid (4.02 g,
16.0 mmol), benzyl alcohol (5.19 g, 48.0 mmol) and p-toluenesul-
fonic acid (3.31 g, 19.2 mmol) in toluene (60 mL) was heated for 4 h
at reflux until the theoretical amount of water had been collected in
a Dean–Stark trap. The reaction was cooled to room temperature,
diluted with methyl tert-butyl ether and washed with a saturated
NaHCO3 solution. The aqueous layer was extracted with methyl
tert-butyl ether (3ϫ). The combined organic phases were washed
with a saturated NaCl solution and dried with Na2SO4. The solvent
was removed in vacuo and the residue purified by LPLC (cyclohex-
ane/ethyl acetate, 1:1, + 0.1% triethylamine) to afford benzyl ester
Fmoc-L-MeLeu-D-PhLac-OBn (14): DCC (6.38 g, 30.8 mmol) in
DCM (100 mL), HOBt (4.71 g, 30.8 mmol) and DMAP (3.76 g,
30.8 mmol) were added to an ice-cold solution of benzyl d-3-phen-
yllactate (10; 7.30 g, 28.5 mmol) and N-Fmoc-N-methyl-l-leucine
(4; 10.47 g, 28.5 mmol) in DCM (200 mL). After stirring for 24 h
at room temperature, the solvent was evaporated and the residue
was taken up in ethyl acetate (100 mL) and filtered. The solution
was washed with a NaHCO3 and a NaCl solution, dried with
Na2SO4, filtered and the solvents evaporated. LPLC (cyclohexane/
ethyl acetate, 8:2) afforded the coupling product 14 (2.156 g, 89%)
as a white foam.[23] 1H NMR (600 MHz, CDCl3): δ = 0.75 and 0.86
(2 d, J = 7.0 Hz, 2 H, CδH3-Leu), 0.94 (pseudo-t, J = 6.0 Hz, 4 H,
CδH3-Leu), 1.45–1.64 (m, 3 H, CβH2-Leu, CγH-Leu), 2.76 (s, 3 H,
NCH3), 3.15 and 3.20 (2 m, 2 H, CβH2-PhLac), 4.19 and 4.31 (m,
1 H, CH-Fmoc), 4.43–4.50 (m, 2 H, CH2-Fmoc), 4.62 and 5.04 (m,
1 H, CαH-Leu), 5.13–5.18 (m, 2 H, CH2-Bn), 5.23 and 5.31 (m, 1
H, CαH-PhLac), 7.14 (m, 2 H, Ar-H), 7.20 (m, 2 H, Ar-H), 7.29
(m, 3 H, Ar-H), 7.36 (m, 5 H, Ar-H), 7.45 (m, 2 H, Ar-H), 7.50
and 7.55 (d, J = 7.6 Hz, 1 H, Ar-H), 7.65 (t, J = 6.6 Hz, 1 H, Ar-
H), 7.78 (d, J = 7.6 Hz, 1 H, Ar-H), 7.82 (d, J = 7.6 Hz, 1 H, Ar-
H) ppm. 13C NMR (150 MHz, CDCl3): δ = 21.7, 23.0, 24.5, 30.1,
37.2, 37.6, 47.2, 56.4, 67.2, 67.7, 73.3, 120.0, 124.8, 125.0, 127.0,
127.6, 127.7, 128.4, 128.6, 129.0, 129.4, 135.0, 135.4, 141.4, 143.9,
1
11 (4.22 g, 77%) as a white solid. H NMR (400 MHz, CDCl3): δ
= 2.69 (s, 1 H, OH), 2.92 (dd, J = 6.0, 14.0 Hz, 1 H, CβH2), 3.05
(dd, J = 5.0, 14.0 Hz, 1 H, CβH2), 3.09–3.17 (m, 4 H, NCH2-
morpholine), 3.81–3.90 (m, 4 H, OCH2-morpholine), 4.45 (m, 1 H,
CαH), 5.18 (s, 2 H, CH2-Bn), 6.79 (d, J = 9 Hz, 2 H, Ar-H), 7.06
(d, J = 9.0 Hz, 2 H, Ar-H), 7.30–7.43 (m, 5 H, Ar-H-benzyl) ppm.
13C NMR (100 MHz, CDCl3): δ = 39.61, 49.41, 66.92, 67.31, 71.40,
115.70, 127.37, 128.56, 128.63, 130.27, 135.12, 150.20, 174.05 ppm.
IR: ν = 1716 (C=O), 3526 (OH) cm–1. MS (ESI): m/z (%) = 705
˜
(6), 342 (100). HRMS (ESI): calcd. for C20H24NO4 [M + H]+
342.1700; found 342.1706.
Boc-
21.6 mmol) in DCM (40 mL) was added dropwise at 0 °C over
20 min to solution of benzyl 3-phenyllactate (10; 5.13 g,
L-MeLeu-D-PhLac-OBn (12): A solution of DCC (4.46 g,
144.1, 156.8, 171.2, 173.7 ppm. IR (KBr): ν = 1743 (C=O), 1701
˜
a
(C=O) cm–1. MS (ESI): m/z (%) = 606 (70), 623 (100), 628 (35).
HRMS (ESI): calcd. for C38H39NNaO6 [M + Na]+ 628.2670; found
628.2671.
20.0 mmol), N-Boc-N-methyl-l-leucine (4.91 g, 20.0 mmol), HOBt
(2.92 g, 21.6 mmol) and DMAP (2.64 g, 21.6 mmol). The reaction
was warmed to room temperature and stirred for another 24 h.
Then the solvent was removed in vacuo and the residue was dis-
solved in ethyl acetate and filtered. The filtrate was washed twice
with a saturated NaHCO3 and NaCl solution, dried with Na2SO4
and the solvents evaporated. The residue was purified by LPLC
(cyclohexane/ethyl acetate, 7:1) to yield the coupling product 12
(9.03 g, 93%) as a white crystalline solid. M.p. 50–51 °C. 1H NMR
(400 MHz, CDCl3): δ = 0.89 (d, J = 6.0 Hz, 6 H, CδH3-Leu), 1.43
Boc-L-MeLeu-D-PhLac-OH (15): Hydrogenolysis of didepsipeptide
12 (3.87 g, 8.0 mmol) was performed in the same way as described
for compound 17 (12 h, room temp., see below). The didepsipeptide
acid 15 was obtained in quantitative yield as a colourless oil
(3.15 g). 1H NMR (400 MHz, CDCl3): δ = 0.82–092 (d, J = 6.0 Hz,
6 H, CδH3-Leu), 1.43 and 1.45 (2 s, 9 H, CH3-tBu), 1.48–1.70 (m,
3 H, CγH-Leu, CβH-Leu), 2.73 and 2.75 (2 s, 3 H, NCH3), 3.12
(s, 9 H, CH3-tBu), 1.49–1.61 (m, 3 H, CγH-Leu, CβH-Leu), 2.61 and 3.26 (2 dd, J = 4.0, 14.0 Hz, 2 H, CβH2-PhLac), 4.62–4.73 (m,
and 2.66 (2 s, 3 H, NCH3), 3.07–3.24 (m, 2 H, CβH-PhLac), 4.61–
1 H, CαH-Leu), 5.26 and 5.31 (2 dd, J = 4.8 Hz, 1 H, CαH-PhLac),
4.99 (2 m, 1 H, CαH-Leu), 5.04–5.19 (m, 2 H, OCH2Ph), 5.27 and 7.18–7.34 (m, 5 H, Ar-H) ppm. 13C NMR (100 MHz, CDCl3): δ =
5.29 (2 dd, J = 5.0 Hz, 1 H, CαH-PhLac), 7.12–7.39 (2 m, 10 H, 21.11, 21.48, 23.02, 23.12, 24.46, 24.78, 28.29, 28.33, 29.97, 31.40,
Ar-H) ppm. 13C NMR (100 MHz, CDCl3): δ = 21.08, 21.31, 23.15,
24.45, 24.75, 28.30, 28.38, 29.78, 30.04, 37.28, 37.64, 55.64, 56.69,
67.13, 67.21, 73.26, 73.34, 127.04, 128.38, 128.43, 128.48, 128.57,
37.18, 37.26, 37.50, 37.58, 56.71, 58.45, 72.99, 127.07, 128.43,
128.50, 129.30, 129.55, 135.70, 155.54, 155.66, 171.62, 173.01 ppm.
IR (film): ν = 1671 (C=O), 1742 (C=O) cm–1. MS (ESI): m/z (%)
˜
129.38, 129.55, 135.04, 135.54, 168.96, 171.74, 210.94 ppm. IR: ν
= 804 (21), 394 (20), 338 (38), 294 (100). HRMS (ESI): calcd. for
˜
= 1692 (C=O), 1740 (C=O) cm–1. MS (ESI): m/z (%) = 501 (52),
C21H31NNaO6 [M + Na]+ 416.2044; found 416.2049.
Eur. J. Org. Chem. 2012, 1546–1553
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
1551

J. Scherkenbeck et al.
FULL PAPER
Boc-
L
-MeLeu-
D
-morphPhLac-OH (16): Hydrogenolysis of didepsi-
Cyclizative Cleavage Ϫ PF1022A (1): A suspension of the resin-
bound octadepsipeptide 27 (50 mg) in DCM (0.5 mL) containing
DIEA (2.2 equiv.) and acetic acid (5.0 equiv.) was agitated for 12 h
at room temperature. Then the resin was filtered off and washed
several times with DCM. The filtrate and rinses were combined
peptide 13 was performed in the same way as described for com-
pound 17 (12 h, room temp., see below). 1H NMR (400 MHz,
CDCl3): δ = 0.87–0.94 (m, 6 H, CδH3-Leu), 1.44 (s, 9 H, CH3-tBu),
1.47–1.77 (m, 3 H, CβH-Leu, CγH-Leu), 2.74 and 2.83 (2 s, 3 H,
NCH3), 3.01–3.09 and 3.14–3.23 (2 m, 2 H, CβH-PhLac), 3.10– and the solvents evaporated. Preparative HPLC purification (ACN/
3.15 (m, 4 H, NCH2-morpholine), 3.82–3.87 (m, 4 H, OCH2-
morpholine), 4.37–4.47 and 4.63–4.74 (2 m, 1 H, CαH-Leu), 5.18–
5.25 and 5.25–5.33 (2 m, 1 H, CαH-PhLac), 6.84 (d, J = 8.0 Hz, 2
H, Ar-H), 7.13 (d, J = 8.0 Hz, 2 H, Ar-H) ppm. 13C NMR
H2O gradient) of the residue afforded PF1022A (1; 8.1 mg) in a
yield of 75% (based on the loading of the support with the linear
precursor 27) and a purity of 99.5%. 1H NMR (400 MHz,
[D6]DMSO): δ = 0.67–0.98 (m, 21 H, CH3-Leu, CβH3-Lac), 1.06–
(100 MHz, CDCl3): δ = 21.09, 21.34, 23.10, 23.18, 24.48, 24.79, 1.76 (m, 15 H, CβH2-Leu, CγH-Leu), 1.28 (d, J = 6.7 Hz, 3 H,
28.33, 29.99, 31.22, 36.39, 37.46, 37.63, 49.56, 56.56, 56.77, 58.40, CH3-Lac), 2.68, 2.77, 2.80, 2.85 and 2.91 (5 s, 12 H, NCH3), 3.05
66.76, 73.42, 115.98, 127.62, 130.19, 130.31, 150.06, 156.46, 171.64, (m, 2 H, CβH2-PhLac), 4.41, 5.11 and 5.22 (3 m, 4 H, CαH-Leu),
172.60 ppm. IR (KBr): ν = 1613 (C=O), 1691 (C=O), 1783 5.01, 5.32 and 5.42 (3 m, 2 H, CαH-Lac), 5.52, 5.68 and 5.71 (3
˜
(C=O) cm–1. MS (ESI): m/z (%) = 957 (28), 480 (27), 479 (100),
423 (88). HRMS (ESI): calcd. for C25H39N2O7 [M + H]+ 479.2752;
found 479.2755.
m, 2 H, CαH-PhLac), 7.21–7.33 (m, 10 H, Ar-H) ppm. 13C NMR
(100 MHz, [D6]DMSO): δ = 15.5, 16.3, 16.8, 20.4, 20.6, 20.8, 20.8,
20.9, 21.0, 23.0, 23.1, 23.2, 23.2, 23.4, 23.9, 24.2, 24.3, 24.3, 24.4,
28.9, 30.1, 30.1, 30.2, 30.3, 30.6, 35.7, 36.3, 36.5, 36.6, 36.7, 36.7,
36.8, 37.1, 37.3, 53.0, 53.2, 53.3, 56.4, 66.7, 67.5, 67.7, 70.1, 70.9,
71.0, 126.7, 126.7, 126.8, 128.1, 128.2, 128.2, 129.5, 135.1, 135.2,
135.9, 169.0, 169.0, 169.3, 169.5, 169.7, 170.2, 170.3, 170.7,
Fmoc-L-MeLeu-D-PhLac-OH (17): A suspension of benzyl ester 14
(18.1 g, 30.0 mmol) and Pd/C (10%, 0.3 g) in ethanol (200 mL) was
hydrogenated (1 bar) at room temperature for 2 h. Then the catalyst
was filtered off and the solvent evaporated. LPLC (cyclohexane/
ethyl acetate, 8:2, + 0.1% acetic acid) afforded Fmoc-l-MeLeu-d-
170.9 ppm. IR (KBr): ν = 1739 (C=O), 1660 (C=O) cm–1. MS
˜
(ESI): m/z (%) = 949 (100), 966 (33). HRMS (ESI): calcd. for
Phlac-OH (17) as
a
white foam (11.20 g, 73%). 1H NMR
C52H76N4NaO12 [M + Na]+ 971.5352; found 971.5351.
(400 MHz, CDCl3): δ = 0.75 and 0.87 (2 d, J = 7.0 Hz, 2 H, CδH3-
Leu), 0.94 (pseudo-t, J = 7.0 Hz, 4 H, CδH3-Leu), 1.45–1.64 (m, 3
H, CβH2-Leu, CγH-Leu), 2.81 (s, 3 H, NCH3), 3.12 and 3.25 (2 m,
2 H, CβH2-PhLac), 4.16 and 4.30 (m, 1 H, CH-Fmoc), 4.48 (m, 2
H, CH2-Fmoc), 4.59 and 4.93 (m, 1 H, CαH-Leu), 5.20 and 5.32
(m, 1 H, CαH-PhLac), 7.20–7.36 (m, 8 H, Ar-H), 7.43 (m, 2 H, Ar-
H), 7.63 (m, 1 H, Ar-H), 7.80 (m, 2 H, Ar-H), 9.72 (br. s, 1 H,
COOH) ppm. 13C NMR (100 MHz, CDCl3): δ = 21.0, 23.0, 24.5,
30.3, 37.2, 37.5, 47.2, 56.4, 67.7, 73.0, 120.0, 124.8, 125.1, 125.1,
127.1, 127.6, 127.7, 128.5, 128.6, 129.0, 129.4, 129.8, 135.5, 141.4,
Cyclizative Cleavage Ϫ Emodepside (2): The cyclizative cleavage re-
action was performed as described for PF1022A (1). After HPLC
purification (ACN/H2O), pale-yellow emodepside (2; 9.3 mg, 45%
over all) was obtained, which was identical to a reference sample
1
with respect to all the spectroscopic and biological data. H NMR
(400 MHz, CDCl3): δ = 0.77–1.05 (m, 30 H, CδH3-Leu, CβH3-Lac),
1.20–1.83 (m, 18 H, CβH2-Leu, CγH-Leu, CH3-Lac), 2.73, 2.74,
2.80, 2.83 and 3.00 (5 s, 12 H, NCH3), 2.85–3.07 (m, 4 H, CβH2-
morphPhLac), 3.08–3.15 (m, 8 H, N-CH2-morpholine), 3.81–3.88
(pseudo-t, 8 H, OCH2morpholine), 4.47, 5.08, 5.19 5.34 and 5.38–
5.53 (5 m, 6 H, CαH-Leu, CαH-morphPhLac), 5.54–5.67 (m, 2 H,
CαH-Lac), 6.77–6.86 (m, 4 H, Ar-H), 7.09–7.17 (m, 4 H, Ar-
H) ppm. 13C NMR (100 MHz, CDCl3): δ = 15.8, 17.1, 20.9, 21.1,
21.2, 23.4, 23.5, 23.5, 23.6, 24.2, 24.7, 24.9, 29.4, 30.5, 31.2, 36.2,
36.7, 37.2, 37.5, 38.1, 49.9, 54.0, 57.1, 66.5, 66.6, 66.9, 68.6, 70.8,
71.3, 116.1, 116.9, 130.4, 130.6, 141.7, 169.8, 170.2, 170.4, 170.6,
143.8, 156.4, 157.1, 171.2, 173.7 ppm. IR (KBr): ν = 3156 (O–H),
˜
1732 (C=O), 1668 (C=O) cm–1. MS (ESI): m/z (%) = 294 (16), 516
(100), 533 (35), 538 (1). HRMS (ESI): calcd. for C31H33NNaO6 [M
+ Na]+ 538.2200; found 538.2201.
Solid-Phase Synthesis on Kaiser Oxime
Coupling of the First Segment to the Resin: Kaiser oxime (100 mg,
0.068 mmol) was suspended in DCM (1.0 mL) and agitated for
30 min. Then Boc-MeLeu-PhLac-OH (15; 53.5 mg, 0.136 mmol),
HATU (51.7 mg, 0.136 mmol) dissolved in DCM (1 mL) and
DIEA (29.0 μL, 0.204 mmol) were added. After shaking for 12 h
the resin was filtered off, washed with DCM and DCM/EtOH (1:1)
three times and dried in vacuo.
171.0, 171.2, 171.7 ppm. IR (KBr): ν = 1744 (C=O), 1665 (C=O),
˜
1614 (C=O) cm–1. MS (ESI): m/z (%) = 374 (28), 560 (85), 1119
(69), 1141 (100). HRMS (ESI): calcd. for C60H90N6NaO14 [M +
Na]+ 1141.6407; found 1141.6414.
Solid-Phase Synthesis on Wang Resin
End-Capping: Acetic anhydride (0.064 mL, 0.68 mmol) and DIEA
(58.0 μL, 0.34 mmol) were added to the resin (100 mg) suspended
in DCM (0.5 mL). The mixture was shaken for 2 h. Then the resin
was filtered off, washed three times alternating with MeOH (10 mL
for each washing) and DCM (10 mL for each washing) and dried
under high vacuum until a constant mass was reached.
Coupling of the First Segment to the Resin: Wang resin was sus-
pended in THF (1.0 mL/100 mg resin) and agitated for 30 min to
achieve a good swelling. The solvent was removed and a solution
of didepsipeptide 17 (3 equiv.), DIC (3 equiv.), HOBt (3 equiv.) and
DMAP (3 equiv.) in THF (1.0 mL/100 mg resin) was added. The
suspension was agitated for 16 h at room temperature. Then the
resin was washed three times with DCM (10 mL), acetone (10 mL)
and DCM (10 mL) and dried under high vacuum until a constant
mass was reached. The whole coupling and washing procedure was
repeated once.
Cleavage of the Boc Group: The Kaiser oxime was suspended in a
TFA/DCM solution (25%, 1.3 mL/100 mg resin) and shaken for
30 min. The resin was filtered off and washed with DCM (3ϫ),
EtOH (1ϫ), DCM (2ϫ), EtOH (3ϫ), DMF (3ϫ) and DCM (1ϫ).
Amide Coupling Reactions: A solution of the depsipeptide acid
(3 equiv.) in DMF (1.0 mL), HATU (2 equiv.) and DIEA
(6.5 equiv.) were added to a suspension of the resin (100 mg) in
DMF (1.5 mL). After shaking for 12 h the resin was filtered off,
washed with DMF (3ϫ10 mL), DCM (3ϫ10 mL), DMF
(3ϫ10 mL) and DCM (3ϫ10 mL) and dried under high vacuum
until a constant mass was reached.
Cleavage of the Fmoc-Group: The resin was swollen in THF for
30 min. After removal of the solvent a solution of piperidine in
THF (25%, 10 mL) was added and the suspension was shaken for
45 min at room temperature. Then the resin was drained and
washed three times with DCM (10 mL), acetone (10 mL) and
DCM (10 mL) and dried under high vacuum. Aliquots from the
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Eur. J. Org. Chem. 2012, 1546–1553

Total Synthesis of Anthelmintic Cyclooctadepsipeptides
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combined rinses were used to determine the loading of the resin,
as described in the General section.
Amide Couplings: A solution of the didepsipeptide acid (3 equiv.),
HATU (2 equiv.) and DIEA (3 equiv.) in THF (1.0 mL/100 mg
resin) were added to the swollen resin. The suspension was stirred
for 16 h at room temperature. Then the solvent was filtered off and
the resin washed three times with DCM (10 mL), acetone (10 mL)
and DCM (10 mL). The whole coupling and washing procedure
was repeated once.
H-
L-MeLeu-D-Lac-L-MeLeu-D-PhLac-L-MeLeu-D-Lac-L-MeLeu-
D-PhLac-OH (29): The octadepsipeptide-resin (712 mg) was treated
with TFA/DCM (1:1; 10 mL/500 mg resin) and shaken for 1 h. The
resin was filtered and washed eight times with DCM (10 mL each).
The cleavage solution and the washings were combined and evapo-
rated to yield a mixture of depsipeptides containing around 40%
(114 mg) of octadepsipeptide 29 (23% overall yield). The crude ma-
terial was used in the cyclization reaction without further purifica-
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Cyclooctadepsipeptide PF1022A (1): DIEA (65.3 mg, 0.504 mmol)
and BOPCl (61.6 mg, 0.242 mmol) were added at 0 °C to a solution
of the crude octadepsipeptide 29 (221.7 mg, corresponds to
0.092 mmol pure substance) in DCM. The reaction was stirred for
24 h and then the same amounts of BOPCl and DIEA were added
and stirring was continued for another 24 h at room temperature.
The solution was washed with a saturated NaHCO3 solution, dried
with Na2SO4 and the solvents evaporated. Chromatographic purifi-
cation (toluene/2-propanol, 20:1) of the residue afforded PF1022A
(87.8 mg, 100%) as a white solid, which was identical to a natural
sample with respect to all the spectroscopic and biological data. 1H
NMR (400 MHz, [D6]DMSO): δ = 0.68–0.98 (m, 21 H, CδH3-Leu,
CβH3-Lac), 1.15–1.75 (m, 14 H, CβH2-Leu, CγH-Leu), 1.28 (d, J =
6.8 Hz, 3 H, CH3-Lac), 2.68, 2.77, 2.80, 2.85 and 2.91 (5 s, 12 H,
NCH3), 3.05 (m, 2 H, CβH2-PhLac), 4.41, 5.11 and 5.22 (3 m, 4
H, CαH-Leu), 5.01, 5.32 and 5.42 (3 m, 2 H, CαH-Lac), 5.52, 5.68
and 5.71 (3 m, 2 H, CαH-PhLac), 7.23–7.32 (m, 10 H, Ar-H) ppm.
13C NMR (100 MHz, [D6]DMSO): δ = 15.5, 16.3, 16.8, 20.4, 20.6,
20.8, 20.8, 20.9, 21.0, 23.0, 23.1, 23.2, 23.2, 23.4, 23.9, 24.2, 24.3,
24.3, 24.4, 28.9, 30.1, 30.1, 30.2, 30.3, 30.6, 35.7, 36.3, 36.5, 36.6,
36.7, 36.7, 36.8, 37.1, 37.3, 53.0, 53.2, 53.3, 56.4, 66.7, 67.5, 67.7,
70.1, 70.9, 71.0, 126.7, 126.7, 126.8, 128.1, 128.2, 128.2, 129.5,
135.1, 135.2, 135.9, 169.0, 169.0, 169.3, 169.5, 169.7, 170.2, 170.3,
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˜
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Generous financial support by Bayer Animal Health GmbH is
gratefully acknowledged. We thank Dr. Werner Hallenbach, Bayer
CropScience, for providing a sample of morpholinophenyllactic
acid and Dr. Gerd Kleefeld for reference samples of PF1022A and
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Published Online: January 24, 2012
Eur. J. Org. Chem. 2012, 1546–1553
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
1553