Synthesis and biological evaluation of cruentaren a analogues

Article abstract of DOI:10.1002/chem.200701673

The complex macrolide cruentaren A is a highly selective and potent inhibitor of F-ATPase (F-type adenosine triphosphatase). As it shows some resemblance to benzolactone enamides like apicularen A, it was of interest to perform some structure-activity studies to delineate the key functional groups that are responsible for the activity. Building upon our previously developed route to cruentaren A, which is based on a ring-closing alkyne metathesis reaction (RCAM), several cruentaren analogues were prepared. Replacement of the 3-hydroxy hexanoic part with acids that lack the hydroxy group function resulted in a significant drop in cytotoxicity and F-ATPase inhibition. Furthermore, two enamide analogues 23 and 50 were synthesized. However, these compounds were only cytotoxic in the micromolar range. Under the conditions for cleavage of the C3 aromatic methyl ether, the enamide function was transformed to the corresponding oxazinanone, resulting in analogues 25 and 52.

Full text of DOI:10.1002/chem.200701673

FULL PAPER
DOI: 10.1002/chem.200701673
Synthesis and Biological Evaluation ofCruentaren A Analogues
Viktor V. Vintonyak,[a] Marcellino Calà,[a] Frank Lay,[a] Brigitte Kunze,[b]
Florenz Sasse,[c] and Martin E. Maier*[a]
Abstract: The complex macrolide
cruentaren A is a highly selective and
potent inhibitor of F-ATPase (F-type
adenosine triphosphatase). As it shows
some resemblance to benzolactone en-
amides like apicularen A, it was of in-
terest to perform some structure–activ-
ity studies to delineate the key func-
tional groups that are responsible for
the activity. Building upon our previ-
ously developed route to cruentaren A,
which is based on a ring-closing alkyne
metathesis reaction (RCAM), several
cruentaren analogues were prepared.
Replacement of the 3-hydroxy hexano-
ic part with acids that lack the hydroxy
group function resulted in a significant
drop in cytotoxicity and F-ATPase in-
hibition. Furthermore, two enamide an-
alogues 23 and 50 were synthesized.
However, these compounds were only
cytotoxic in the micromolar range.
Under the conditions for cleavage of
the C3 aromatic methyl ether, the en-
amide function was transformed to the
corresponding oxazinanone, resulting
in analogues 25 and 52.
Keywords: cruentaren · cyclization
reactions
lactones
products
·
enzyme inhibitors
·
·
metathesis Natural
·
Introduction
Recently, Hçfle and co-workers described the structure of
the macrolide cruentaren A (1).[1,2] This unique natural
product was isolated form the myxobacterium Byssovorax
cruenta (Scheme 1). In a cellular assay with the L929 cell
line, cruentaren A showed powerful cytotoxicity with an
IC50 value of 1.2 ngmLꢀ1. Further studies revealed that on a
molecular level, cruentaren A inhibits mitochondrial F-
ATPase (F-ATPase=F-type adenosine triphosphatase).[3,4]
These membrane-bound proteins are crucial for a living cell
as they use a proton gradient to power the synthesis of ATP.
Key structural features of cruentaren A include a 12-mem-
Scheme 1. Structure of cruentaren A.
bered macrolactone with a Z double bond. The side chain
extending from C15 contains a stereotetrad and an (Z)-ally-
lamide terminus. Interestingly, cruentaren A does not show
much similarity to other polyketide inhibitors of F-ATPase,
such as apoptolidin or oligomycin.[5]
[a]Dipl.-Chem. V. V. Vintonyak, Dipl.-Chem. M. Calà,
Dipl.-Chem. F. Lay, Prof. Dr. M. E. Maier
Institut für Organische Chemie, Universität Tübingen
Auf der Morgenstelle 18, 72076 Tübingen (Germany)
E-mail: martin.e.maier@uni-tuebingen.de
Other natural products with an allylamide include leucas-
candrolide[6,7] (2) (Scheme 2), neopeltolide,[8] callipelto-
side[9,10] and ajudazol A (3).[11] Except for ajudazol A, which
was reported to be an inhibitor of mitochondrial electron
transport, the mode of action of the other mentioned com-
pounds still remains unclear. Furthermore, a similarity of
cruentaren A to the benzolactone enamides,[12] like apicular-
en A (4) was noted.[2] However, cruentaren A does not in-
hibit V-ATPase (V-type ATPase),[3,13] the target of the ben-
[b]Dr. B. Kunze
Arbeitsgruppe Mikrobielle Kommunikation
Helmholtz-Zentrum für Infektionsforschung
Inhoffenstrasse 7, 38124 Braunschweig (Germany)
[c]Dr. F. Sasse
Abteilung Chemische Biologie
Helmholtz-Zentrum für Infektionsforschung
Inhoffenstrasse 7, 38124 Braunschweig (Germany)
Supporting information for this article is available on the WWW
under http://www.chemeurj.org/ or from the author.
Chem. Eur. J. 2008, 14, 3709 – 3720
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3709

Scheme 2. Structures of some natural products that resemble cruentaren
A.
zolactone enamides. Therefore, it would be of interest to
identify some key structural elements that are decisive for
the biological activity of cruentaren A.
In previous papers, we outlined an efficient synthetic
strategy towards cruentaren A.[14,15] The macrolactone ring
was formed through a ring-closing alkyne metathesis reac-
tion[16,17] on the ester 5 by using the Schrock catalyst 6
(Scheme 3). To prevent an unwanted translactonization to
the six-membered lactone, extension of the side chain was
done on the cyclic alkyne 7. Thus, the aldehyde derived
from 7 was converted to the alkyne 8 through the Best-
mann–Ohira reaction. Extension of the terminal alkyne with
formaldehyde allowed for the formation of the key proparg-
yl amine 9 through the Mitsunobu reaction. Condensation
of the amine 9 with the protected 3-hydroxy acid 10 led to
the amide 11. Finally, cleavage of the C3 OMe ether, the sil-
icon protecting groups and Lindlar hydrogenation complet-
ed the total synthesis of cruentaren A.[15] Recently, another
ring-closing alkyne metathesis (RCAM)-based synthesis of
cruentaren A was achieved by Fürstner et al.[18]
With regard to the design of analogues, we wanted to use
the available stereotetrad building blocks[14] and stick to the
proven RCAM reaction. We intended to answer the follow-
ing questions: How important is the carboxylic acid part of
the amide? How important is the free OH at C3? Can we
make enamides instead of (Z)-allylamides? Although quite
speculative, it could be that the allylamide isomerizes to an
enamide that then might form a highly electrophilic acylimi-
nium ion upon protonation (Scheme 4).[19]
Scheme 3. Key steps in the synthesis of cruentaren A; DMB=3,4-dime-
thoxybenzyl, TBS=tert-butyldimethylsilyl, TIPS=triisopropylsilyl.
Scheme 4. Possible isomerization of the allyamide to an enamide.
Results and Discussion
Synthesis: We began with the preparation of the 3-O-methyl
ether of cruentaren. As outlined in Scheme 3, the synthesis
of 1 passed through the diyne 11. Omitting the cleavage of
the C3 methyl ether and instead treating the lactone 11 with
the HF·pyridine complex led to the triol 12 (Scheme 5). A
final Lindlar reduction delivered 3-OMe cruentaren 13.
Another key intermediate of the total synthesis, the prop-
argyl amine 9, presented itself for derivatization reactions.
Accordingly, the amine 9 was condensed with the acids 14a–
d by using 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluroni-
um hexafluorophosphate (HBTU) in the presence of 1-hy-
droxy-1H-benzotriazole (HOBt) and Hünigꢁs base in N,N-
dimethylformamide (DMF; Scheme 6). These acylation re-
actions proceeded in quite good yields (Table 1). For these
derivatives, we chose to generate the original aromatic part
Herein, we describe the synthesis together with the bio-
logical evaluation of several cruentaren A analogues.
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Investigation of Cruentaren A Analogues
FULL PAPER
Scheme 5. Synthesis of 3-OMe cruentaren 13 from the amide 11. Py=
pyridine.
with the 3-OH group. Selective ether cleavage on the lac-
tones 15a–d by using boron trichloride furnished the corre-
sponding 3-hydroxy compounds 16a–d, again in excellent
yields. The Lindlar reduction of the diynes 17b and 17c pro-
ceeded as expected to give the analogues 18b and 18c.
From the reduction of diyne 17a, the analogue 18a was ob-
tained, but we also isolated the dihydro compound 18e, re-
sulting from hydrogenation of the cinnamoyl double bond.
In the case of the hept-2-en-4-ynamide 17d, only the prod-
uct 18 f resulting from complete hydrogenation was ob-
served. The internal Z double bond survived as in the other
amide analogues.
As a further branching point for the synthesis of the ana-
logues, we identified the lactone 8 with a propynyl terminus.
We thought that the derived vinyl iodide might be useful for
the synthesis of enamide derivatives. With this in mind,
diyne 8 was subjected to hydrozirconation with the Schwartz
reagent followed by addition of iodine to the intermediate
vinylmetal species (Scheme 7).[20] Thereafter, a copper-cata-
lysed cross-coupling reaction of vinyl iodide 19 with the
amide 20 under Buchwald conditions[21–26] was performed,
resulting in enamide 21 in high yield. Owing to the expected
sensitivity of the enamide to harsh acidic conditions, the de-
methylation step was omitted. Nevertheless, the enamide
survived the conditions (HF·pyridine complex) for global
deprotection of the silyl ethers. A Lindlar reduction on
diyne 22 completed the synthesis of enamide analogue 23.
Scheme 6. Preparation of various amide analogues of cruentaren A.
HBTU=O-(1H-benzotriazol-1-yl)-N,N,N’,N’-tetramethyluronium hexa-
fluorophosphates.
Table 1. Yields for the various steps for the synthesis of amide analogues
18 of cruentaren A.
Acid
used
Transformation
BCl3 HF·py
acylation
[%]
Lindlarꢁs
catalyst
[%]
[%]
[%]
14a
14b
14c
14d
87
88
91
85
92
86
83
89
95
93
85
92
74[a]
93
87
73[b]
[a]By-product dihydro derivative 18e. [b]Only the saturated heptanoyl
derivative 18 f was formed under the Lindlar conditions.
If however, the enamide 21 was treated with boron tri-
chloride to cleave the C3 O-methyl ether, followed by
global silyl removal with HF·pyridine complex, a compound
1
(24), which lacked the enamide signals in the H NMR spec-
trum was isolated (Scheme 8). According to LC–MS analy-
sis, the mass was the same as expected for the enamide. The
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M. E. Maier et al.
Scheme 8. Formation of the oxazinanone system during methyl ether
cleavage on enamide 21 with BCl3 resulting in analogue 25.
which originated from a Marshall–Tamaru reaction,[14,27] was
extended to the propargyl alcohol 27. A hydrogenation reac-
tion provided the propanol derivative 28. Protection of the
hydroxyl group function with dimethoxybenzyl imidate[28] to
give 29 was followed by cleavage of the isopropylidene
group by using aqueous copper(ii) chloride[29] to provide diol
30. The 1,2-diol was converted to the epoxide 32 by using
the arylsulfonyl derivative 31.[30] The stage was now set for
epoxide opening with lithium trimethylsilylacetylide in the
presence of BF3·OEt2.[31,32] Silylation of 33 and removal of
the acetylenic silyl group from 34 furnished alkyne 35. In
preparation for the RCAM reaction, the terminal alkyne 35
was converted to the inner alkyne 36 by using nBuLi fol-
lowed by MeI. The subsequent treatment of bis-silyl ether
36 with tetrabutylammonium fluoride (TBAF) delivered
diol 37. A possible shortcut from epoxide 32 to the alkyne
36 was attempted by direct opening of the epoxides with
propynyllithium. Unfortunately, the major product in this
reaction by using propynyllithium prepared in situ from 1-
bromopropene[33] turned out to be the corresponding bro-
mohydrin.
Scheme 7. Preparation of the enamide analogue 23 of cruentaren A.
1
signal at d=5.02 ppm in the H NMR spectrum pointed to
the presence of the 1,3-oxazinan-4-one. The formation of
this heterocyclic ring system can easily be explained by the
corresponding acyliminium ion. Although we were not able
to unambiguously assign the stereochemistry at the aminal
carbon, we assume a 2,6-cis-configuration (oxazinone-4-
numbering). Force-field calculations using Chem3D on
2,5,6-trimethyl-1,3-oxazinan-4-one showed the cis-2,6-diaste-
reomer to be 5.65 kJmolꢀ1 more stable than the correspond-
ing 2,6-trans isomer. Lindlar reduction of the triple bond led
to the oxazinan-4-one analogue 25. It can be assumed that
oxazinanone formation occurs upon treatment of the enam-
ide 21 with BCl3 as the HF·pyridine complex does not seem
to affect the enamide as could be seen with the deprotection
of 21 to enamide 22.
To further probe the potential biological relevance of a
cruentaren A enamide, the homologated enamide derivative
of analogue 25 was targeted. In this case, we could have
used macrolactone 8, with a propyne terminus, as the start-
ing material, but instead we began with the stereotetrad-
containing building block[14] 26 (Scheme 9). This compound,
As we had outlined in the synthesis of the core structure,
esterification of benzoic acid 38 with building block 37 was
best performed with the diol itself. After conversion of the
acid to the carbonylimidazolide 39, esterification with the
sodium alcoholate of 37 went smoothly and in a regioselec-
tive manner (Scheme 10).[34] Silylation of the free hydroxy
group function of 40 gave rise to the ester 41, the substrate
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Investigation of Cruentaren A Analogues
FULL PAPER
Scheme 9. Synthesis of the alkynediol 37 from terminal alkyne 26. CDI=
N,N’-carbonyl diimidazole, PPTS=p-toluenesulfonate.
for the alkyne metathesis reaction. Based on the diol 37, the
yield for the ester 41 amounted to 67%. The RCAM[16,35] re-
action of 41 with the Schrock catalyst[36] 6 proceeded in ex-
cellent chemical yield, furnishing lactone 42. To set up an
(E)-vinyl iodide at the side-chain terminus, the DMB pro-
tecting group was removed under oxidative conditions.[37]
The resulting primary alcohol 43 was oxidized to the alde-
hyde 44. Extension of the aldehyde 44 to the alkyne could
be accomplished with the Bestmann–Ohira reagent[38] 45 in
the presence of K2CO3. Finally, hydrozirconation and iodina-
tion provided the vinyl iodide 47.
As outlined before, a cross-coupling reaction of vinyl
iodide 47 with the amide 20 was used to set up the enamide
functionality (Scheme 11). Global silicon ether cleavage on
48 and Lindlar reduction of the triple bond furnished ana-
logue 50.
Scheme 10. Synthesis of the macrolactone 47 featuring a 4-iodobutenyl
side chain. DDQ=2,3-dichloro-5,6-dicyano-1,4-benzoquinone.
oxazinanones have been generated by condensation of aro-
matic aldehydes with alicyclic 2-hydroxy-1-carboxamides.[39]
For such oxazinanone derivatives, a 2,6-cis-configuration
was observed. The synthesis for 5-phenylthio-1,3-oxazinan-
4-ones is based on the hetero Diels–Alder reaction between
an azadiene and an aldehyde.[40] Other 1,3-oxazinanones are
known as well.[41,42,43]
If the enamide 48 was treated with BCl3, cleavage of the
C3 OMe ether was accompanied by the formation of the ox-
azinan-4-one 51 (Scheme 12). The analogue 52 was obtained
through silyl group removal and the Lindlar reduction.
Characteristic peaks for the oxazinanone part of 52 are as
follows: d=4.71 (2-H; 22-H), 6.40 ppm (N-H); 13C NMR:
d=83.8 (C2; C22), 174.6 (C4; C23), 38.8 (C5; C24), 72.9
(C6; C25) ppm.
Biological testing: The described analogues as well as the
diyne 53 were tested for cytotoxicity against the L929 cell
line and the inhibitory efficacy on F-ATPase in mitochondri-
al preparations of bovine heart. The obtained IC50 values in
the cell-culture assay as well as the percentage of F-ATPase
inhibition of the compounds at a concentration of 0.1 and
1.0 mm, respectively, are listed in Table 2. The analogues are
ordered according to increasing IC50 values against the L929
cell line.
The formation of 1,3-oxazinan-4-ones from enamides con-
taining a b-hydroxy acid seems to be unprecedented. Similar
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M. E. Maier et al.
Table 2. Biological activity of cruentaren A (cru) and the analogues.
Entry Compound IC50 IC50 Inhibition of
[mgmLꢀ1
ative with a side chain of seven
carbons can be considered as
highly cytotoxic, but it showed
only low inhibitory efficacy in
the F-ATPase assay (seven-
carbon, entry 3). Then there are
compounds of intermediate cy-
totoxicity and F-ATPase inhibi-
tion, namely the cruentaren de-
rivatives with a modified car-
boxylic part in the side chain
(18e, 18b, 18 f; Table 2, en-
tries 5–7). In particular 18c and
18b make clear that the OH
group of the carboxylic acid
part is extremely important. Fi-
nally, there are compounds that
are essentially nontoxic, starting
Description
[a]
A
]
[mm]F-ATPase activity [%]
1 mm
0.1 mm
1
2
3
4
5
6
7
8
9
1
13
52
0.00042ꢁ0.00005 0.00071ꢁ0.00008 94
78
42
2
30
32
27
48
8
15
30
10
12
cru (synthetic)[b]
3-OMe-cru
7C-oxazinanone-cru
isobutanoyl-cru
dihydro-cinnamoyl-cru
hexanoyl-cru
0.017ꢁ0.004
0.085ꢁ0.02
0.3ꢁ0.01
2.4ꢁ0.1
0.028ꢁ0.007
0.14ꢁ0.03
0.56ꢁ0.02
4.0ꢁ0.2
80
34
44
47
47
67
47
51
40
21
18
18c
18e
18b
18 f
50
23
18a
53
2.5ꢁ0.1
4.5ꢁ0.2
2.9ꢁ0.3
5.0ꢁ0.5
heptanoyl-cru
3.0ꢁ0.4
5.0ꢁ0.7
7C-enamide-cru
6C-enamide-cru
cinnamoyl-cru
diyne-cru
6C-oxazinanone-cru
3.0ꢁ1.1
5.1ꢁ1.9
10
11
12
6.1ꢁ0.7
10.3ꢁ1.2
11.1ꢁ0.7
13.0ꢁ1.6
6.5ꢁ0.4
25
7.5ꢁ0.9
[a]The inhibition values are the mean values of at least two independent assays. The deviations did not
exceed a range of ꢁ10% inhibition. [b]The natural cruentaren A displayed a slightly lower activity (IC 50
(0.002ꢁ0.0019) mM, F-ATPase inhibition=93% at 1 mm). This might be attributed to different purity values.
The value for the synthetic product does lie within the deviation of the natural material.
=
Scheme 11. Synthesis of the enamide analogue 50.
As can be seen in Table 2, there are some highly effective
compounds. In most cases, cytotoxicity and inhibitory activi-
ty against F-ATPase in vitro run parallel. However, there
are some exceptions in this regard. For example, 18c and
the three analogues 18e, 18b and 18 f differ in their cellular
activity by a factor of almost 10, but display similar effects
on the F-ATPase. This might be explained by differences in
cellular uptake. The highest effective compound is cruentar-
en A itself (Table 2, entry 1), followed by 3-OMe cruentaren
(13) (Table 2, entry 2). Furthermore, the oxazinanone deriv-
Scheme 12. Synthesis of the oxazinan-4-one analogue 52 and the diyne
cruentaren 53.
with compound 18a. Surprisingly, both enamides show nei-
ther a high cytotoxicity, nor significant inhibition of F-
ATPase. One hypothesis in the design of the enamide ana-
logues was that with a structural resemblance to typical V-
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Investigation of Cruentaren A Analogues
FULL PAPER
matography (petroleum ether/EtOAc, 10:1!4:1) to give 20.5 mg (87%)
of amide 15a as a colourless amorphous solid. TLC (petroleum ether/
EtOAc, 4:1): Rf =0.39; [a]2D0 =ꢀ13.1 (c=1.4, CH2Cl2); 1H NMR
ATPase inhibitors like apicularen A or salicylihalamide A,
these analogues would show corresponding activity. As it
could be assumed that a V-ATPase inhibitor would be
highly cytotoxic, this shows that the enamide side is not suf-
ficient to convert the F-ATPase inhibitor cruentaren A into
a V-ATPase inhibitor. The most puzzling observation is the
relatively high cytotoxicity of the oxazinanone 52, which
shows only low inhibition of F-ATPase. We also checked for
inhibitory effects on V-ATPase with PtK2 potoroo cells.
However, when we investigated treated cells by fluorescent
techniques, we did not observe the characteristic changes in
the endoplasmatic reticulum that are typical for V-ATPase
inhibitors. One explanation for the cytotoxicity of 52 could
be that the heterocyclic ring is opened to an electrophilic
acyliminium ion when taken up by the cells. The lack of ac-
tivity for diyne cruentaren 53 can be attributed to conforma-
tional effects.
(400 MHz, CDCl3): d=0.04 (s, 3H, Si
N
A
0.87–1.02 (m, 39H, 16-CH3, 18-CH3, 10-CH3, Si(C
ACHTREUNG
AHCTREUNG
2.20 (m, 1H, 11-H), 2.22–2.30 (m, 1H, 10-H), 2.33–2.55 (m, 4H, 8-H, 14-
H, 11-H), 3.60–3.65 (m, 1H, 17-H), 3.73 (s, 3H, OCH3), 3.75–3.83 (m,
4H, OCH3, 8-H), 4.00–4.18 (m, 3H, 9-H, 22-H), 5.47–5.59 (m, 1H, 15-H),
6.02 (br s, 1H, NH), 6.31 (d, J=2.0 Hz, 1H, 6-H), 6.40–6.46 (m, 2H, 4-H,
25-H), 7.29–7.34 (m, 3H, o-, p-CH of Ph), 7.41–7.47 (m, 2H, m-CH of
Ph), 7.62 ppm (d, J=15.7 Hz, 1H, 24-H); 13C NMR (100 MHz, CDCl3):
d=ꢀ3.9 (Si
(CH(CH3)2), 18.1 (CH
(C11), 26.1 (Si(C(CH3)3)), 30.0 (C22), 37.2 (C18), 37.3 (C16), 38.6 (C8),
A
N
A
ACHTREUNG
AHCTREUNG
40.3 (C10), 55.2 (OCH3), 55.7 (OCH3), 75.0 (C17), 76.6 (C9), 77.2 (C15),
79.6 (C=C), 81.7 (C=C), 83.2 (C=C), 96.6 (C4), 108.2 (C6), 118.0 (C2),
120.2 (C24), 127.8 (C4’), 128.7 (C3’), 129.6 (C2’), 134.8 (C1’), 139.4 (C7),
141.2 (C25), 157.4 (C5), 160.4 (C3), 165.4 (C1), 167.6 ppm (C23); HRMS
(ESI): [M+Na]+ calcd for C51H77NNaO7Si2 894.51308, found 894.51309.
2-Hydroxy-4-methoxybenzoate (16a): A solution of amide 15a (18 mg,
0.02 mmol) in CH2Cl2 (3 mL) was treated with BCl3 (80 mL, 1.0m in
CH2Cl2, 0.08 mmol, 4 equiv) at ꢀ808C. The reaction was stirred for 2 h at
ꢀ808C before a saturated solution of NaOAc (3 mL) was added. After
separation of the layers, the aqueous phase was extracted twice with
CH2Cl2. The combined organic layers were washed with H2O followed by
saturated NaCl solution, dried over MgSO4, filtered and concentrated in
vacuo. Flash chromatography (petroleum ether/EtOAc, 4:1) afforded
phenol 16a (15.8 mg, 92%) as a slightly yellow oil. TLC (petroleum
ether/EtOAc, 4:1): Rf =0.4; [a]2D0 =+17.0 (c=0.7, CH2Cl2); 1H NMR
Conclusion
By using the RCAM strategy that led to the total synthesis
of cruentaren A (1), a range of cruentaren analogues were
prepared. Replacing the 3-hydroxy-hexanoic acid gave ana-
logues 18a, 18b, 18c, 18e, 18 f, however, with the exception
of the truncated isobutanoyl analogue 18c, none of the ana-
logues were highly active. Furthermore, the two enamide an-
alogues 23 and 50 were prepared via cross-coupling (amina-
tion) of the corresponding vinyl iodides 19 and 47, respec-
tively. As the enamides did not survive the conditions
(BCl3) of the cleavage of the aromatic methyl ether, we pre-
pared the 3-OMe derivatives. Even though this methyl ether
is important (see 1 and 13), the complete lack of activity for
the two enamides 23 and 50 is somewhat surprising. Upon
cleavage of the 3-OMe ether with BCl3, the enamide func-
tion of 21 and 49 reacted with the hydroxyl group function
of the carboxylic acid to give an unusual oxazinanone het-
erocycle. Among the two oxazinanone analogues, compound
52, which might be a metabolite of 1, was quite active and
showed an IC50 value of 140 nm. This work also constitutes a
novel synthesis of 1,3-oxazinan-4-ones from enamides.
(400 MHz, CDCl3): d=0.05–0.08 (m, 6H, Si
ACHTREUNG
16-CH3, 18-CH3, 10-CH3, Si(C(CH3)3), Si(CH
A
ACHTREUNG
16-H, 19-H), 1.94–2.20 (m, 4H, 19-H, 18-H, 11-H), 2.23–2.35 (m, 2H, 10-
H, 14-H), 2.44–2.52 (m, 1H, 8-H), 2.55–2.67 (m, 1H, 14-H), 2.85–2.97 (m,
1H, 8-H), 3.56–3.62 (m, 1H, 17-H), 3.77 (s, 3H, OCH3), 4.10–4.15 (m,
2H, 22-H), 4.17–4.25 (m, 1H, 9-H), 5.19–5.28 (m, 1H, 15-H), 5.81 (br s,
1H, NH), 6.33–6.42 (m, 3H, 6-H, 4-H, 25-H), 7.32–7.39 (m, 3H, o-, p-CH
of Ph), 7.47–7.53 (m, 2H, m-CH of Ph), 7.64 (d, J=15.7 Hz, 1H, 24-H),
11.22 ppm (br s, 1H, 3-OH); 13C NMR (100 MHz, CDCl3): d=ꢀ4.0 (Si-
A
N
A
ACHTREUNG
AHCTREUNG
(C17), 75.6 (C9), 76.9 (C=C), 77.2 (C15), 82.7 (C=C), 83.2 (C=C), 98.9
(C4), 104.2 (C6), 119.7 (C2), 120.0 (C24), 127.8 (C4’), 128.8 (C3’), 129.8
(C2’), 134.7 (C1’), 141.6 (C7), 143.3 (C25), 163.4 (C3), 164.6 (C5), 165.3
(C1), 171.1 ppm (C23); HRMS (ESI): [M+Na]+ calcd for
C50H75NO7Si2Na 880.49743, found 880.49810.
Deprotected macrolactone (17a): HF·pyridine complex (70% HF,
0.3 mL) was added dropwise to a stirred solution of the phenol 16a
(14 mg, 0.016 mmol) in THF (0.4 mL, in a plastic test tube) at ꢀ808C.
The reaction mixture was allowed to warm to ꢀ58C. After 2 h, the mix-
ture was partioned between an ice-cooled mixture of EtOAc (20 mL)
and a saturated aqueous NaHCO3 solution (20 mL). The organic layer
was separated and the H2O layer extracted with EtOAc (2”20 mL). The
combined organic layers were dried over MgSO4, filtered and concentrat-
ed in vacuo. The residue was purified by flash chromatography (CH2Cl2/
MeOH, 95:5!9:1) to give 9.0 mg (95%) of triol 17a. TLC (CH2Cl2/
MeOH, 9:1): Rf =0.56; [a]2D0 =+11.7 (c=0.8, CH2Cl2); 1H NMR
(400 MHz, CDCl3): d=0.91–0.96 (m, 6H, 16-CH3, 18-CH3), 1.02 (d, J=
6.8 Hz, 3H, 10-CH3), 1.74–1.85 (m, 2H, 16-H, 18-H), 2.00–2.10 (m, 2H,
19-H, 18-H), 2.15–2.27 (m, 4H, 19-H, 11-H, OH), 2.28–2.37 (m, 2H, 11-
H, 8-H), 2.38–2.46 (m, 1H), 2.57–2.65 (m, 1H, 14-H), 2.77–2.85 (m, 1H,
14-H), 2.88–2.98 (m, 1H, 10-H), 3.63–3.69 (m, 1H, 17-H), 3.72–3.76 (m,
1H, 8-H), 3.91–3.99 (m, 1H, 9-H), 4.11–4.16 (m, 2H, 22-H), 5.32–5.39
(m, 1H, 15-H), 5.97 (br s, 1H, NH), 6.34–6.43 (m, 3H, 6-H, 4-H, 25-H),
7.33–7.40 (m, 3H, o-, p-CH of Ph), 7.46–7.52 (m, 2H, m-CH of Ph), 7.63
(d, J=15.7 Hz, 1H, 24-H), 11.00 ppm (br s, 1H, 3-OH); 13C NMR
(100 MHz, CDCl3): d=8.5 (16-CH3), 14.1 (24-CH3), 16.2 (18-CH3), 16.5
(10-CH3), 21.1 (C19), 22.5 (C14), 23.0 (C11), 29.9 (C22), 35.7 (C26), 36.7
Experimental Section
General details are included in the Supporting Information. The experi-
mental details for Scheme 5, part of Scheme 6, Scheme 8 and Scheme 9
are covered in the Supporting Information as well. The pH 7 buffer was
prepared by dissolving KH2PO4 (85 g, 0.625 mol) and NaOH (14.5 g,
0.3625 mol) in water (1 L).
Cinnamic acid amide (15a): (E)-cinnamic acid (6.4 mg, 0.043 mmol,
1.6 equiv), HBTU (20.5 mg, 0.054 mmol, 2 equiv), HOBt (7.3 mg,
0.054 mmol, 2 equiv), and N,N-diisopropylethylamine (48 mL, 0.27 mmol,
10 equiv) were added to a solution of amine[15] 9 (20 mg, 0.027 mmol,
1 equiv) in dry DMF (2 mL). After the mixture was stirred at room tem-
perature for 4 h, H2O (5 mL) was added and the obtained emulsion was
extracted with Et2O (3”15 mL). The combined organic layers were
washed with saturated NaHCO3 solution (10 mL), dried over MgSO4, fil-
tered and concentrated in vacuo. The residue was purified by flash chro-
Chem. Eur. J. 2008, 14, 3709 – 3720
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
3715

M. E. Maier et al.
(C18), 37.3 (C16), 38.4 (C8), 55.4 (OCH3), 73.8 (C17), 75.3 (C9), 77.2
(C15), 77.6 (C=C), 79.3 (C=C), 81.8 (C=C), 83.3 (C=C), 99.5 (C4), 106.5
(C2), 111.5 (C6), 120.0 (C24), 127.8 (C4’), 128.8 (C3’), 129.8 (C2’), 134.7
(C1’), 141.7 (C7), 143.2 (C25), 163.7 (C5), 164.5 (C3), 165.5 (C1),
170.7 ppm (C23); HRMS (ESI): [M+Na]+ calcd for C35H41NaNO7
610.27752, found 610.27802.
H), 2.35–2.54 (m, 5H, 8-H, 14-H, 23-H, 11-H), 3.45–3.49 (m, 1H, 17-H),
3.72–3.78 (m, 8H, OCH3, 24-H, 8-H), 3.97–4.03 (m, 1H, 9-H), 4.92–5.01
(m, 1H, 20-H), 5.38–5.51 (m, 1H, 15-H), 6.31 (d, J=2.0 Hz, 1H, 6-H),
6.41 (d, J=2.0 Hz, 1H, 4-H), 6.73 (dd, J=14.0 Hz, 10.6 Hz, 1H, 21-H),
8.08 ppm (br d, J=10.6 Hz, 1H, NH); 13C NMR (100 MHz, CDCl3): d=
ꢀ4.6 (Si
(16-CH3), 12.4 (23-CH3), 13.0 (CH
17.9 (CH(CH3)2), 17.9 (Si(C(CH3)3)), 18.1 (CH
19.4 (C26), 23.3 (C14), 23.7 (C11), 25.9 (Si(C(CH3)3)), 26.1 (Si(C
A
G
U
(CH3)2), 11.3
AHCTREUNG
Cinnamoyl cruentaren (18a) and diyhdrocinnamoyl cruentaren (18e):
Lindlarꢁs catalyst (5 wt% Pd on CaCO3, poisoned with lead, 4.2 mg,
A
N
ACHTREUNG
N
ACHTREUNG
100 wt%) was added to
a stirred solution of diyne 17a (4.2 mg,
32.6 (C19), 34.7 (C25), 37.6 (C18), 38.1 (C16), 38.6 (C8), 40.7 (C10), 45.6
(C23), 55.2 (OCH3), 55.7 (OCH3), 74.9 (C17), 76.2 (C9), 77.2 (C24), 79.7
(C=C), 81.2 (C=C), 96.6 (C4), 108.3 (C6), 110.9 (C20), 118.2 (C2), 123.4
(C21), 139.4 (C7), 157.2 (C5), 160.2 (C3), 167.4 (C1), 170.9 ppm (C22);
HRMS (ESI): [M+Na]+ calcd for C54H97NaNO8Si3 994.64142, found
994.64053.
0.007 mmol) in EtOAc (2 mL) containing quinoline (1.5 mg, 0.01 mmol).
The reaction was placed under H2 atmosphere and stirred for 1 h. The
mixture was filtered through a pad of celite and the filtrate concentrated
in vacuo. Flash chromatography of the residue (CH2Cl2/MeOH, 95:5!
9:1) afforded cinnamide 18a (3.1 mg, 74%) and phenylpropionamide 18e
(1.0 mg, 24%).
18a: TLC (CH2Cl2/MeOH, 9:1): Rf =0.64; [a]2D0 =+8.3 (c=0.3, CH2Cl2);
1H NMR (400 MHz, CDCl3): d=0.80 (d, J=6.8 Hz, 3H, 18-CH3), 0.92 (d,
J=7.1 Hz, 3H, 16-CH3), 0.97 (d, J=6.8 Hz, 3H, 10-CH3), 1.70–1.79 (m,
2H, 18-H, OH), 1.93–2.06 (m, 3H, 10-H, 11-H, 16-H), 2.16–2.26 (m, 2H,
14-H, 19-H), 2.28–2.40 (m, 3H, 11-H, 8-H, 19-H), 2.79–2.96 (m, 1H, 14-
H), 3.51 (dd, J=9.4 Hz, 1.8 Hz, 1H, 17-H), 3.61–3.67 (m, 1H, 9-H), 3.72–
3.74 (m, 1H, 8-H), 3.75–3.80 (m, 4H, OCH3, OH), 3.87–3.95 (m, 1H, 22-
H), 4.05–4.14 (m, 1H, 22-H), 5.27–5.35 (m, 1H, 15-H), 5.42–5.52 (m, 3H,
21-H, 12-H, 13-H), 5.53–5.63 (m, 1H, 20-H), 6.02 (br s, 1H, NH), 6.30 (d,
J=2.6 Hz, 1H, 6-H), 6.35–6.41 (m, 2H, 4-H, 25-H), 7.32–7.39 (m, 3H, o-,
p-CH of Ph), 7.46–7.51 (m, 2H, m-CH of Ph), 7.62 (d, J=15.7 Hz, 1H,
24-H), 11.50 ppm (br s, 1H, 3-OH); HRMS (ESI): [M+Na]+ calcd for
C35H45NaNO7 614.30882, found 614.30923.
18e: TLC (CH2Cl2/MeOH, 9:1): Rf =0.58; [a]2D0 =+6.4 (c=0.1, CH2Cl2);
1H NMR (400 MHz, CDCl3): d=0.77 (d, J=6.8 Hz, 3H, 18-CH3), 0.90 (d,
J=7.1 Hz, 3H, 16-CH3), 1.00 (d, J=6.6 Hz, 3H, 10-CH3), 1.63–1.77 (m,
1H, 18-H), 1.90–2.50 (m, 12H, 10-H, 11-H, 16-H, 14-H, 19-H, 8-H, 25-H,
OH), 2.78–2.90 (m, 1H, 14-H), 2.91–2.99 (m, 2H, 24-H), 3.43–3.50 (m,
1H, 17-H), 3.61–3.67 (m, 1H, 9-H), 3.72–3.82 (m, 5H, 8-H, 22-H, OCH3),
3.85–3.97 (m, 1H, 22-H), 5.25–5.36 (m, 2H, 15-H, 21-H), 5.40–5.59 (m,
3H, 12-H, 13-H, 20-H), 5.7 (br s, 1H, NH), 6.28–6.32 (m, 1H, 6-H), 6.34–
6.39 (m, 1H, 4-H), 7.15–7.22 (m, 3H, m-, p-CH of Ph), 7.24–7.31 (m, 2H,
o-CH of Ph), 11.50 ppm (br s, 1H, 3-OH); HRMS (ESI): [M+Na]+ calcd
for C33H47NO8Na 616.32447, found 616.32467.
Deprotected enamide macrolactone (22): The HF·pyridine complex
(70% HF, 0.3 mL) was added dropwise to a stirred solution of the enam-
ide 21 (10 mg, 0.01 mmol) in THF (0.4 mL, in a plastic test tube) at
ꢀ808C. The reaction mixture was allowed to warm to ꢀ108C. After 2 h,
the mixture was partioned between an ice-cooled mixture of EtOAc
(20 mL) and saturated aqueous NaHCO3 solution (20 mL). The organic
layer was separated and the H2O layer extracted with EtOAc (2”
20 mL). The combined organic layers were dried over MgSO4, filtered
and concentrated in vacuo. The residue was purified by flash chromatog-
raphy (CH2Cl2/MeOH, 95:5!9:1) to give triol 22 (4.4 mg; 75%). TLC
(CH2Cl2/MeOH, 9:1): Rf =0.47; [a]2D0 =ꢀ3.3 (c=0.4, CH2Cl2); 1H NMR
(400 MHz, CDCl3): d=0.81 (d, J=6.8 Hz, 3H, 18-CH3), 0.90–0.98 (m,
6H, 27-H, 10-CH3), 1.10 (d, J=7.1 Hz, 3H, 16-CH3), 1.16 (d, J=7.3 Hz,
3H, 23-CH3), 1.28–1.39 (m, 2H, 25-H, 26-H), 1.41–1.51 (m, 2H, 25-H, 26-
H), 1.58–1.67 (m, 2H, 19-H, OH), 1.90–2.02 (m, 4H, 18-H, 19-H, 11-H,
OH), 2.05–2.14 (m, 1H, 16-H), 2.29–2.40 (m, 2H, 8-H, 10-H), 2.46–2.63
(m, 3H, 14-H, 11-H), 2.71–2.79 (m, 1H, 23-H), 2.85 (br s, 1H, OH), 3.29–
3.26 (m, 1H, 8-H), 3.45–3.49 (m, 1H, 17-H), 3.78 (s, 3H, OCH3), 3.80 (s,
3H, OCH3), 3.83–3.90 (m, 2H, 9-H, 24-H), 5.10–5.19 (m, 1H, 20-H),
5.46–5.52 (m, 1H, 15-H), 6.35 (d, J=2.0 Hz,1H, 6-H), 6.41 (d, J=
2.0 Hz,1H, 4-H), 6.75 (dd, J=14.2, 10.4 Hz, 1H, 21-H), 7.52 ppm (d, J=
10.4 Hz, 1H, NH); HRMS (ESI): [M+Na]+ calcd for C33H49NaNO8
610.33504, found 610.33433.
Enamide analogue (23): Lindlarꢁs catalyst (5 wt% Pd on CaCO3, pois-
oned with lead, 4.0 mg, 100 wt%) was added to a stirred solution of
diyne 22 (4.0 mg, 0.007 mmol) in EtOAc (2 mL) containing quinoline
(1.4 mg, 0.011 mmol). The reaction was placed under H2 atmosphere and
stirred for 1 h. The reaction mixture was filtered through a pad of celite
and the filtrate concentrated in vacuo. The residue was purified by flash
chromatography (CH2Cl2/MeOH, 95:5!9:1) to give enamide 23 as a col-
(E)-Vinyl iodide (19): [Cp2Zr(H)Cl](31 mg, 0.12 mmol) was added to a
solution of alkyne[15] 8 (44 mg, 0.06 mmol) in THF (1.5 mL) at 08C and
the resulting mixture was stirred for 2 h at 08C.
A solution of I2
(0.24 mL, 0.5m in THF, 0.12 mmol) was then added dropwise and stirring
was continued for 2 h. The reaction was quenched with saturated aque-
ous Na2S2O3 solution (5 mL). The mixture was repeatedly extracted with
Et2O. The combined organic layers were dried over MgSO4, filtered and
evaporated, and the residue was purified by flash chromatography (petro-
leum ether/EtOAc, 4:1) to give (E)-vinyl iodide 19 as an amorphous
solid (52 mg, 92%), which was used directly in the next step. TLC (petro-
leum ether/EtOAc, 4:1): Rf =0.74.
ourless oil (3.4 mg, 85%). TLC (CH2Cl2/MeOH, 9:1): Rf =0.52; [a]D20
=
ꢀ4.3 (c=0.3, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=0.83 (d, J=
6.8 Hz, 3H, 18-CH3), 0.93 (t, J=6.8 Hz, 3H, 27-H), 0.98 (d, J=6.8 Hz,
3H, 16-CH3), 1.05 (d, J=6.8 Hz, 3H, 10-CH3), 1.16 (d, J=7.3 Hz, 3H,
23-CH3), 1.28–1.40 (m, 2H, 25-H, 26-H), 1.42–1.71 (m, 4H, 25-H, 26-H,
19-H, OH), 1.88–1.99 (m, 4H, 18-H, 19-H, 11-H, 14-H), 2.07–2.21 (m,
1H, 16-H), 2.27–2.37 (m, 2H, 8-H, 10-H), 2.37–2.50 (m, 1H, 11-H), 2.70–
3.00 (m, 4H, 23-H, 14-H, 8-H, OH), 3.45–3.52 (m, 1H, 17-H), 3.71–3.81
(m, 7H, OCH3, 24-H), 3.85–3.91 (m, 1H, 9-H), 5.09–5.19 (m, 1H, 20-H),
5.40 (dd, J=9.9, 4.0 Hz, 1H, 15-H), 5.48–5.54 (m, 2H, 12-H, 13-H), 6.33–
6.36 (m, 2H, 6-H, 4-H), 6.74 (dd, J=14.2, 10.4 Hz, 1H, 21-H), 7.52 ppm
(d, J=10.4 Hz, 1H, NH); HRMS (ESI): [M+Na]+ calcd for
C33H51NaNO8 612.35069, found 612.35104.
Enamide 21: A Schlenk tube was charged with CuI (10.5 mg, 0.055 mmol,
1 equiv), amide[44] 20 (28.5 mg, 0.11 mmol, 2 equiv) and Cs2CO3 (46 mg,
0.14 mmol, 2.5 equiv). The tube was evacuated and backfilled with argon.
N,N’-Dimethylethylenediamine (12.0 mL, 0.11 mmol, 2 equiv), vinyl
iodide 19 (52 mg, 0.055 mmol) and THF (1.0 mL) were added under
argon. The Schlenk tube was closed and immersed in an oil bath, which
was preheated to 608C. The mixture was stirred for 14 h. After the result-
ing pale-blue suspension was allowed to reach room temperature, ethyl
acetate (5 mL) was added. The reaction mixture was filtered through a
pad of celite and the filtrate concentrated in vacuo. The residue was puri-
fied by flash chromatography (petroleum ether/EtOAc, 10:1!4:1) to
give enamide 21 as an amorphous solid (44 mg, 82%). TLC (petroleum
ether/EtOAc, 4:1): Rf =0.66; [a]2D0 =ꢀ23.5 (c=1.6, CH2Cl2); 1H NMR
A
N
benzyl)oxy]-1,3-dimethylhexyl}pent-3-ynyl 2,4-dimethoxy-6-{(2R,3S)-3-
methyl-2-[(triisopropylsilyl)oxy]hept-5-ynyl}benzoate (41): A solution of
diol 37 (470 mg, 1.24 mmol) in anhydrous DMF (2.5 mL) was stirred in
the presence of sodium hydride (60% wt in mineral oil, 124 mg,
3.1 mmol, 2.5 equiv) at 08C for 10 min and for a further 1 h at room tem-
perature. CDI (390 mg, 2.4 mmol) was added to a solution of acid 38
(917 mg, 2.0 mmol) in anhydrous DMF (3.5 mL) in a separate flask and
the reaction mixture was then allowed to stir for 4 h at 508C. Then, the
solution of the imidazolide derivative 39 (analysed by LC–MS) was
(400 MHz, CDCl3): d=0.03–0.07 (m, 6H, Si
Si(CH3)2), 0.84–0.98 (m, 48H, 24-CH3, 27-H, 16-CH3, Si(C
Si(CH(CH3)2)3), 1.02–1.10 (m, 6H, 18-CH3, 10-CH3), 1.16–1.26 (m, 1H,
ACHTREUNG
A
ACHTREUNG
ACHTREUNG
26-H), 1.31–1.50 (m, 3H, 25-H, 26-H), 1.65–1.75 (m, 2H, 18-H, 16-H),
1.76–1.84 (m, 1H, 11-H), 1.90–1.99 (m, 1H, 10-H), 2.10–2.27 (m, 2H, 19-
3716
www.chemeurj.org
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2008, 14, 3709 – 3720

Investigation of Cruentaren A Analogues
FULL PAPER
cooled to 08C and added to the above solution of the disodium salt of
diol 37 at 08C. The mixture was allowed to warm to room temperature
and stirred for three days. After the addition of saturated NH4Cl solu-
tion, the mixture was extracted with EtOAc (3”30 mL). The combined
organic extracts were washed with 1m HCI, saturated NaHCO3 and satu-
rated NaCl solution, dried over MgSO4, filtered and concentrated in
vacuo to provide 1.01 g of crude hydroxyester 40, which was used in the
next step without further purification.
(Ar of DMB), 139.3 (C7), 148.4 (Ar of DMB), 148.9 (Ar of DMB), 157.2
(C5), 160.2 (C3), 167.3 ppm (CO2R); HRMS (ESI): [M+Na]+ calcd for
C50H82NaO9Si2 905.53896, found 905.53829.
Alcohol 43: DDQ (194 mg, 0.85 mmol, 1.4 equiv) was added to a cooled
(08C) solution of DMB ether 42 (540 mg, 0.61 mmol) in a mixture of
CH2Cl2/pH 7 phosphate buffer solution (20:1, 32 mL). The mixture was
allowed to warm to room temperature and stirred for 40 min. Then it was
treated with saturated NaHCO3 solution and the layers were separated.
The aqueous layer was extracted with CH2Cl2. The combined organic ex-
tracts were washed with saturated NaHCO3 and saturated NaCl solution,
dried over MgSO4, filtered and concentrated in vacuo. Flash chromatog-
raphy (petroleum ether/EtOAc, 4:1) afforded alcohol 43 (435 mg, 97%)
as an amorphous solid. TLC (petroleum ether/EtOAc, 4:1): Rf =0.42;
A solution of the crude hydroxyester 40 (1.01 g, 1.23 mmol) in CH2Cl2
(10 mL) was cooled to ꢀ508C, then 2,6-lutidine (0.58 mL, 4.9 mmol) fol-
lowed by tert-butyldimethylsilyltriflate (TBSOTf; 0.46 mL, 2.0 mmol)
were added. The reaction mixture was allowed to warm to room temper-
ature and stirred for 2 h before it was treated with water. After separa-
tion of the layers, the aqueous layer was extracted with CH2Cl2. The com-
bined organic extracts were washed with 1m HCl, saturated NaHCO3 and
saturated NaCl solution, dried over MgSO4, filtered and concentrated in
vacuo. Flash chromatography (petroleum ether/EtOAc, 10:1!4:1) af-
forded ester 41 (0.778 g, 67% for 2 steps, based on diol 37) as a colour-
less oil. Besides ester 41, some unreacted imidazolide derivative 39
(295 mg, 30%) was isolated. TLC (petroleum ether/EtOAc, 4:1): Rf =
0.46, [a]2D0 =+24.8 (c 2.4, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=
1
[a]2D0 =ꢀ28.0 (c=4.0, CH2Cl2); H NMR (400 MHz, CDCl3): d=0.02, 0.06
(2 s, 6H, Si
N
N
Si(CH(CH3)2)3), 1.02 (d, J=7.1 Hz, 3H, 10-CH3), 1.05–1.10 (m, 1H, 20-
C
H), 1.38–1.51 (m, 2H, 20-H, 19-H), 1.55–1.73 (m, 3H, 19-H, 11-H, OH),
1.76–1.82 (m, 1H, 11-H), 1.86–1.97 (m, 1H, 16-H), 2.10–2.18 (m, 1H, 18-
H), 2.34–2.51 (m, 4H, 8-H, 14-H, 10-H), 3.50 (dd, J=4.3, 4.3 Hz, 1H, 17-
H), 3.55 (dd, J=6.3, 6.3 Hz, 2H, 21-H), 3.70–3.78 (m, 7H, 8-H, OCH3),
3.76 (s, 3H, OCH3), 3.96–4.02 (m, 1H, 9-H), 5.37–5.50 (m, 1H, 15-H),
6.31 (d, J=2.3 Hz, 1H, 6-H), 6.40 ppm (s, J=2.3 Hz, 1H, 4-H); 13C NMR
ꢀ0.01, 0.02 (2 s, 6H, Si
C
(CH3)3), 3’’’-
ACHTREUNG
(100 MHz, CDCl3): d=ꢀ4.0 (Si
12.9 (CH(CH3)2), 16.7 (10-CH3), 17.8 (Si(C
18.3 (18-CH3), 23.2 (C14), 23.4 (C11), 26.1 (Si(C
A
ACHTREUNG
(m, 1H, 5’’’-H), 1.45–1.58 (m, 3H, 4’’’-H, 5’’’-H), 1.66 (t, J=2.3 Hz, 3H,
C=CCH3), 1.71 (t, J=2.3 Hz, 3H, C=CCH3), 1.80–1.91 (m, 1H, 3’-H),
1.96–2.05 (m, 2H, 3’’’-H, 4’-H), 2.07–2.20 (m, 2H, 1’’’-H, 4’-H), 2.49–2.71
(m, 4H, 2’’-H, 1’-H), 3.34–3.40 (m, 2H, CH2ODMB), 3.50–3.54 (m, 1H,
G
A
ACHTREUNG
AHCTREUNG
(C19), 37.4 (C16), 37.4 (C18), 38.6 (C8), 40.3 (C10), 55.1 (OCH3), 55.8
(OCH3), 62.9 (C21), 76.0 (C15), 76.7 (C17), 77.3 (C=C), 79.6 (C9), 81.2
(C12), 96.7 (C4), 108.3 (C2), 118.1 (C6), 139.4 (C7), 157.2 (C5), 160.2
(C3), 167.4 ppm (C1); HRMS (ESI): [M+Na]+ calcd for C41H72NaO7Si2
755.47088, found 755.47080.
CH
ACHTREUNG
OCH3 of DMB), 4.21–4.36 (m, 1H, CH
AHCTREUNG
DMB), 4.93–5.00 (m, 1H, 1’’-H), 6.24 (d, J=2.3 Hz, 1H, 5-H), 6.43 (d,
J=2.3 Hz, 3-H), 6.74–6.85 ppm (m, 3H, aryl H of DMB); 13C NMR
Aldehyde 44: A solution of Dess–Martin periodinane (15% wt, 1.02 mL,
0.49 mmol) was added to a cooled (08C) solution of alcohol 43 (198 mg,
0.27 mmol) in CH2Cl2 (6 mL). After stirring for 0.5 h at 08C and for 2 h
at room temperature, the reaction mixture was concentrated, loaded on a
flash column and eluted with petroleum ether/EtOAc (4:1) to give
188 mg (95%) of aldehyde 44, which was used directly in the next reac-
tion. TLC (petroleum ether/EtOAc, 4:1): Rf =0.69.
(100 MHz, CDCl3): d=ꢀ3.7 (Si
3.5 (C=CCH3), 10.3 (1’’’-CH3), 12.9 (CH
(CH3)3), 18.0 (CH(CH3)2), 18.2 (CH(CH3)2), 18.5 (3’’’-CH3), 22.1 (C=
CCH2), 22.1 (C=CCH2), 26.1 (Si(C(CH3)3)), 28.0 (C5’’’), 28.7 (C4’’’), 36.2
A
N
AHCTREUNG
A
N
ACHTREUNG
AHCTREUNG
(C1’), 37.5 (C3’’’), 37.8 (C1’’’), 38.8 (C3’), 55.2 (OCH3), 55.6 (OCH3), 55.8
(OCH3), 55.9 (OCH3), 70.7 (C6’’’), 72.8 (CH2 of DMB), 74.6 (CH3C=C),
74.9 (CH3C=C), 75.2 (C2’), 76.0 (CH2C=C), 76.6 (CH2C=C), 77.9 (C2’’’),
78.1 (C1’’), 96.7 (C3), 107.0 (C5), 110.8 (Ar of DMB), 111.0 (Ar of
DMB), 117.9 (C1), 120.2 (Ar of DMB), 131.2 (Ar of DMB), 139.0 (C6),
148.4 (Ar of DMB), 148.9 (Ar of DMB), 157.8 (C4), 160.7 (C2),
167.7 ppm (CO2R); HRMS (ESI): [M+Na]+ calcd for C54H88NaO9Si2
959.58591, found 959.58513.
Alkyne 46: Diethyl-1-diazo-2-oxopropylphosphonate[38] (45) (124 mg,
0.52 mmol, 2 equiv) was added to a solution of aldehyde 45, which was
obtained in the previous step (188 mg, 0.26 mmol), and K2CO3 (122 mg,
0.88 mmol, 3.4 equiv) in MeOH (5 mL) followed by stirring of the mix-
ture for 12 h at room temperature. The reaction mixture was diluted with
Et2O (50 mL) and washed with an aqueous solution (5%) of NaHCO3
(20 mL). The layers were separated and the organic layer dried over
Na2SO4. After filtration and evaporation of the solvent, the residue was
purified by flash chromatography (EtOAc/petroleum ether, 1:10) to give
179 mg (97%) of alkyne 46 as an amorphous solid. TLC (petroleum
ether/EtOAc, 4:1): Rf =0.78; [a]2D0 =ꢀ32.2 (c 2.0, CH2Cl2); 1H NMR
(3S,8S,9R)-3-{(1R,2R,3R)-2-{[tert-Butyl(dimethyl)silyl]oxy}-6-[(3,4-di-
A
methoxybenzyl)oxy]-1,3-dimethylhexyl}-12,14-dimethoxy-8-methyl-9-
[(triisopropylsilyl)oxy]-5,6-didehydro-3,4,7,8,9,10-hexahydro-1H-2-ben-
zoxacyclododecin-1-one (42):
A solution of (tBuO)3W=CCMe3 (6)
(32.8 mg, 0.069 mmol) in toluene (1.0 mL) was added to a solution of
ester 41 (650 mg, 0.69 mmol) in toluene (81 mL) and the mixture was
stirred at 858C for 3 h. For workup, the solvent was evaporated and the
residue purified by flash chromatography (petroleum ether/EtOAc, 10:1)
to give macrolactone 42 as an amorphous solid (553 mg, 90%). TLC (pe-
troleum ether/EtOAc, 4:1): Rf =0.63; [a]2D0 =ꢀ19.0 (c=2.4, CH2Cl2);
(400 MHz, CDCl3): d=0.03–0.07 (m, 6H, Si
A
16-CH3, 18-CH3, Si(C(CH3)3), Si(CH(CH3)2)3), 1.04 (d, J=7.1 Hz, 3H,
A
ACHTREUNG
10-CH3), 1.20–1.32 (m, 1H, 20-H), 1.61–1.71 (m, 1H, 20-H), 1.75–1.84
(m, 1H, 16-H), 1.87–1.98 (m, 3H, 19-H, C=CH), 2.02–2.28 (m, 3H, 10-H,
11-H, 18-H), 2.36–2.47 (m, 4H, 8-H, 14-H, 11-H), 3.52 (dd, J=4.3,
4.3 Hz, 1H, 17-H), 3.71–3.79 (m, 7H, 8-H, OCH3), 3.78 (s, 3H, OCH3),
3.97–4.02 (m, 1H, 9-H), 5.40–5.52 (m, 1H, 15-H), 6.31 (d, J=2.0 Hz, 1H,
6-H), 6.41 ppm (s, J=2.0 Hz, 1H, 4-H); 13C NMR (100 MHz, CDCl3): d=
1H NMR (400 MHz, CDCl3): d=0.01–0.06 (m, 6H, Si
ACHTREUNG
(m, 36H, 18-CH3, 10-CH3, Si(C
A
ACHTREUNG
6.8 Hz, 3H, 16-CH3), 1.40–1.52 (m, 2H, 20-H), 1.60–1.85 (m, 4H, 19-H,
11-H, 16-H), 1.89–1.98 (m, 1H, 18-H), 2.09–2.19 (m, 1H, 11-H), 2.35–2.51
(m, 4H, 14-H, 8-H, 10-H), 3.37 (dd, J=6.4, 6.4 Hz, 2H, 21-H), 3.48 (dd,
J=4.3, 4.3 Hz, 1H, 17-H), 3.71–3.79 (m, 7H, 8-H, OCH3), 3.84–3.87 (m,
6H, OCH3 of DMB), 3.98–4.02 (m, 1H, 9-H), 4.40 (s, 2H, CH2 of DMB),
5.35–5.50 (m, 1H, 15-H), 6.30 (d, J=2.0 Hz, 1H, 6-H), 6.40 (d, J=2.0 Hz,
1H, 4-H), 6.78–6.88 ppm (m, 3H, Ar of DMB); 13C NMR (100 MHz,
ꢀ4.0 (Si
(10-CH3), 16.5 (C20), 17.8 (CH
23.2 (C14), 23.3 (C11), 26.1 (Si(C
A
G
U
A
ACHTREUNG
AHCTREUNG
(C16), 38.6 (C8), 40.4 (C10), 55.1 (OCH3), 55.7 (OCH3), 68.4 (C22), 74.8
(C=C), 75.6 (C15), 77.2 (C17), 79.6 (C=C), 81.2 (C=C), 84.5 (C9), 96.6
(C4), 108.2 (C2), 118.1 (C6), 139.4 (C7), 157.2 (C5), 160.2 (C3),
167.3 ppm (C1); HRMS (ESI): [M+Na]+ calcd for C42H70NaO6Si2
749.46031, found 749.46039.
CDCl3): d=ꢀ4.0 (Si
(CH3)2), 16.7 (10-CH3), 17.8 (CH
CH3), 23.3 (C=CCH2), 23.5 (C=CCH2), 26.1 (Si(C
A
ACHTREUNG
G
A
ACHTREUNG
AHCTREUNG
(E)-Vinyl iodide 47: [Cp2Zr(H)Cl](38 mg, 0.14 mmol) was added to a so-
lution of alkyne 46 (51 mg, 0.07 mmol) in THF (1.5 mL) at 08C and the
resulting mixture was stirred for 2 h at that temperature. A solution of I2
(0.28 mL, 0.5m in THF, 0.14 mmol) was then added dropwise and stirring
28.4 (C19), 37.4 (2C, C16, C18), 38.5 (C8), 40.3 (C10), 55.1 (OCH3), 55.7
(OCH3), 55.7 (OCH3), 55.8 (OCH3), 70.3 (C21), 72.7 (CH2 of DMB), 76.0
(C15), 77.2 (C17), 79.6 (C9), 81.2 (CH2C=C), 96.6 (C3), 108.2 (C6), 110.8
(Ar of DMB), 110.9 (Ar of DMB), 118.1 (C2), 120.1 (Ar of DMB), 131.1
Chem. Eur. J. 2008, 14, 3709 – 3720
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
3717

M. E. Maier et al.
was continued for 2 h. The reaction was quenched with saturated aque-
ous Na2S2O3 solution (5 mL), and the mixture was repeatedly extracted
with Et2O. The combined organic layers were dried (MgSO4), filtered
and evaporated. The residue was purified by flash chromatography (pe-
troleum ether/EtOAc, 4:1) to give (E)-vinyl iodide 47 as an amorphous
3H, 24-CH3), 1.24–1.50 (m, 5H, 26-H, 27-H, 20-H), 1.55–1.77 (m, 4H, 19-
H, 20-H, OH), 1.89–2.02 (m, 3H, 18-H, 19-H, 11-H), 2.05–2.20 (m, 3H,
16-H, 24-H, 11-H), 2.24–2.33 (m, 1H, 8-H), 2.46–2.62 (m, 3H, 14-H, 10-
H, OH), 2.73–2.82 (m, 1H, 14-H), 3.09 (br s, 1H, OH), 3.27 (dd, J=
13.8 Hz, 2.7 Hz, 1H, 8-H), 3.38–3.44 (m, 1H, 17-H), 3.75–3.89 (m, 8H,
OCH3, 9-H, 25-H), 5.01–5.10 (m, 1H, 21-H), 5.42–5.50 (m, 1H, 15-H),
6.34 (d, J=2.0 Hz,1H, 6-H), 6.40 (d, J=2.0 Hz,1H, 4-H), 6.70 (dd, J=
14.2, 10.4 Hz, 1H, 22-H), 7.68 ppm (d, J=10.4 Hz, 1H, NH); HRMS
(ESI): [M+Na]+ calcd for C34H51NNaO8 624.35069, found 624.35092.
solid (66 mg, 95%). TLC (petroleum ether/EtOAc, 4:1): Rf =0.77; [a]D20
=
ꢀ21.0 (c=3.6, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=0.03–0.07 (m,
6H, Si
G
N
A
1.43–1.54 (m, 1H, 19-H), 1.64–1.72 (m, 1H, 20-H), 1.74–1.82 (m, 1H, 16-
H), 1.87–1.98 (m, 2H, 19-H, 18-H), 2.02–2.20 (m, 2H, 10-H, 11-H), 2.37–
2.48 (m, 4H, 8-H, 14-H, 11-H), 3.52 (dd, J=4.3, 4.3 Hz, 1H, 17-H), 3.72–
3.81 (m, 7H, 8-H, OCH3), 3.97–4.02 (m, 1H, 9-H), 5.39–5.44 (m, 1H, 15-
H), 5.90 (d, J=14.4 Hz, 1H, 22-H), 6.33 (d, J=2.0 Hz, 1H, 6-H), 6.36–
6.45 ppm (m, 2H, 4-H, 21-H); 13C NMR (100 MHz, CDCl3): d=ꢀ4.0 (Si-
Enamide analogue 50: A 5-mL round-bottom flask was charged with
diyne 49 (3.0 mg, 0.005 mmol) and stirred with a stir bar. EtOAc (2 mL)
and quinoline (1.0 mg, 0.08 mmol) were added with stirring. This was fol-
lowed by the addition of Lindlarꢁs catalyst (5 wt% Pd on CaCO3, pois-
oned with lead, 3 mg, 100 wt%). The reaction was placed under H2 at-
mosphere and stirred for 1 h. The reaction mixture was filtered through a
pad of celite and the filtrate concentrated in vacuo. The residue was puri-
fied by flash chromatography (CH2Cl2/MeOH, 95:5!9:1) to give enam-
ide 50 as a colourless oil (2.7 mg, 90%). TLC (CH2Cl2/MeOH, 9:1): Rf =
A
G
R
A
ACHTREUNG
ACHTREUNG
(C16), 38.6 (C8), 40.0 (C10), 55.2 (OCH3), 55.8 (OCH3), 74.5 (C22), 75.4
(C15), 77.2 (C17), 79.6 (C=C), 81.4 (C9), 96.6 (C4), 108.4 (C2), 118.0
(C6), 139.4 (C7), 146.6 (C21), 157.3 (C5), 160.3 (C3), 167.3 ppm (C1);
HRMS (ESI): [M+Na]+ calcd for C42H71INaO6Si2 877.37261, found
877.37329.
1
0.41; [a]2D0 =ꢀ5.6 (c=0.2, CH2Cl2); H NMR (400 MHz, CDCl3): d=0.84–
0.89 (m, 6H, 28-H, 18-CH3), 1.01–1.06 (m, 6H, 10-CH3, 16-CH3), 1.14 (d,
J=7.1 Hz, 3H, 24-CH3), 1.21–1.31 (m, 4H, 26-H, 27-H, 20-H), 1.34–1.49
(m, 2H, 26-H, 27-H), 1.57 (br s, 1H, OH), 1.63–1.75 (m, 2H, 18-H, 19-
H), 1.81–2.07 (m, 5H, 10-H, 11-H, 16-H, 19-H, 8-H), 2.15–2.24 (m, 1H,
14-H), 2.25–2.40 (m, 1H, 24-H), 2.58 (br s, 1H, OH), 2.74–2.86 (m, 1H,
11-H), 2.92–3.00 (m, 1H, 8-H), 2.82 (dt, J=14.3, 11.5 Hz, 1H, 14-H),
3.29–3.39 (m, 2H, 17-H, OH), 3.69–3.76 (m, 4H, 25-H, OCH3), 3.77–3.83
(m, 4H, 9-H, OCH3), 5.10–5.19 (m, 1H, 21-H), 5.36 (dd, J=10.2, 3.2 Hz,
1H, 15-H), 5.41–5.56 (m, 2H, 12-H, 13-H), 6.32–6.36 (m, 2H, 6-H, 4-H),
6.76 (dd, J=14.2 Hz, 10.6 Hz, 1H, 22-H), 7.69 ppm (d, J=10.4 Hz, 1H,
NH); HRMS (ESI): [M+Na]+ calcd for C34H53NO8Na 626.36689, found
626.36672.
Enamide 48:
A Schlenk tube was charged with CuI (14.0 mg,
0.075 mmol, 1 equiv), amide 20 (39 mg, 0.15 mmol, 2 equiv) and Cs2CO3
(62 mg, 0.19 mmol, 2.5 equiv). The tube was evacuated and backfilled
with argon. N,N’-Dimethylethylenediamine (16.0 mL, 0.15 mmol, 2 equiv),
vinyl iodide 47 (64 mg, 0.075 mmol) and THF (1.0 mL) were added under
argon. The Schlenk tube was closed with a glass stopper, immersed in a
preheated to 608C oil bath and the reaction mixture was stirred for 14 h.
After the resulting pale-blue suspension was allowed to reach room tem-
perature, ethyl acetate (5 mL) was added and the reaction mixture was
filtered through a pad of celite and the filtrate concentrated in vacuo.
The residue was purified by flash chromatography on silica gel (petro-
leum ether/EtOAc, 10:1!4:1) to give enamide 48 as an amorphous solid
Oxazinane-4-one 51: a) ortho-Demethylation: A solution of enamide 49
(18 mg, 0.018 mmol) in CH2Cl2 (2 mL) was treated with BCl3 (72 mL,
1.0m in CH2Cl2, 0.072 mmol, 4 equiv) at ꢀ808C. The reaction was stirred
for 2 h at ꢀ808C before a saturated solution of NaOAc (5 mL) was
added. After separation of the layers, the aqueous phase was extracted
twice with CH2Cl2. The combined organic layers were washed with H2O,
saturated NaCl solution, dried over MgSO4, filtered and concentrated in
vacuo to give 17 mg of crude 2-hydroxy-4-methoxybenzoate which was
used in the next step without further purification.
(63 mg, 85%). TLC (petroleum ether/EtOAc, 4:1): Rf =0.72; [a]D20
=
ꢀ23.9 (c=1.3, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=0.02–0.07 (m,
6H, Si
G
E
28-H, 16-CH3, Si(C
N
N
10-CH3), 1.16–1.27 (m, 3H, 20-H, 27-H), 1.30–1.52 (m, 5H, 26-H, 27-H,
18-H, 19-H), 1.74–1.83 (m, 1H, 16-H), 1.86–1.99 (m, 2H, 10-H, 11-H),
2.04–2.19 (m, 2H, 19-H, 11-H), 2.36–2.55 (m, 5H, 8-H, 14-H, 24-H),
3.43–3.48 (m, 1H, 17-H), 3.71–3.79 (m, 7H, OCH3, 25-H), 3.97–4.03 (m,
1H, 9-H), 4.94–5.03 (m, 1H, 21-H), 5.35–5.49 (m, 1H, 15-H), 6.31 (d, J=
2.0 Hz, 1H, 6-H), 6.41 (d, J=2.0 Hz, 1H, 4-H), 6.73 (dd, J=14.2,
10.6 Hz, 1H, 22-H), 8.08 ppm (br d, J=10.6 Hz, 1H, NH); 13C NMR
b) Global deprotection: HF·pyridine complex (70% HF, 0.6 mL) was
added dropwise to a stirred solution of the crude 2-hydroxy-4-methoxy-
benzoate (17 mg) in THF (0.8 mL, in a plastic test tube) at ꢀ808C drop-
wise. The reaction mixture was allowed to warm to ꢀ108C. After 2 h, the
mixture was partioned between an ice-cooled mixture of EtOAc (20 mL)
and saturated aqueous NaHCO3 solution (20 mL). The organic layer was
separated and the H2O layer extracted with EtOAc (2”20 mL). The
combined organic layers were dried over MgSO4, filtered and concentrat-
ed in vacuo. The residue was purified by flash chromatography (CH2Cl2/
MeOH, 95:5!9:1) to give 9.2 mg (87% for 2 steps) of triol 51. TLC
(CH2Cl2/MeOH, 9:1): Rf =0.38; [a]2D0 =+14.0 (c=0.4, CH2Cl2); 1H NMR
(400 MHz, CDCl3): d=0.82 (d, J=6.6 Hz, 3H, 18-CH3), 0.91–0.97 (m,
6H, 28-H, 16-CH3), 1.04 (d, J=7.1 Hz, 3H, 10-CH3), 1.19 (d, J=7.3 Hz,
3H, 24-CH3), 1.27–1.75 (m, 10H, 19-H, 26-H, 27-H, 20-H, 21-H), 1.95–
2.21 (m, 4H, 18-H, 16-H, 11-H, OH), 2.28–2.36 (m, 3H, 8-H, 24-H, OH),
2.54–2.73 (m, 3H, 14-H, 10-H, 11-H), 2.78–2.86 (m, 1H, 14-H), 3.53 (d,
J=8.8 Hz, 1H, 17-H), 3.69–3.78 (m, 2H, 25-H, 8-H), 3.80 (s, 3H, OCH3),
3.91–3.99 (m, 1H, 9-H), 4.79 (t, J=5.6 Hz, 1H, 22-H), 5.37–5.43 (m, 1H,
15-H), 6.38–6.40 (m, 2H, 4-H, 6-H), 6.76 (br s, 1H, NH), 11.1 ppm (br s,
1H, 3-OH); 13C NMR (100 MHz, CDCl3): d=7.9 (16-CH3), 12.0 (24-
CH3), 14.0 (C28), 16.4 (18-CH3), 16.7 (10-CH3), 18.8 (C27), 20.9, 21.7
(C14), 22.3 (C11), 29.7 (C20), 32.5 (C19), 33.3 (C21), 35.6 (C26), 36.0,
36.3 (C18), 37.8 (C16), 38.2 (C8), 40.0 (C24), 55.4 (OCH3), 71.0 (C25),
75.1 (C9), 76.2 (C15), 77.2 (C=C), 79.0 (C=C), 83.0 (C22), 84.0, 99.4 (C4),
110.2 (C6), 142.0 (C7), 162.0 (C5), 163.0 (C3), 170.2 (C1), 175.0 (C23);
HRMS (ESI): [M+Na]+ calcd for C33H49NaNO8 610.33504, found
610.33543.
(100 MHz, CDCl3): d=ꢀ4.6 (Si
ꢀ3.7 (Si(CH3)2), 11.2 (16-CH3), 12.6 (24-CH3), 13.0 (CH
(C28), 16.5 (10-CH3), 17.9 (Si(C(CH3)3)), 17.9(CH(CH3)2), 18.1 (CH-
(CH3)2), 18.4 (18-CH3), 19.3 (C27), 23.3 (C14), 23.6 (C11), 25.9 (Si(C-
(CH3)3)), 26.2 (Si(C(CH3)3)), 27.7 (C20), 32.5 (C19), 34.7 (C26), 37.0
A
G
A
G
ACHTREUNG
A
ACHTREUNG
ACHTREUNG
A
ACHTREUNG
(C18), 37.6 (C16), 38.6 (C8), 40.6 (C10), 45.6 (C24), 55.2 (OCH3), 55.7
(OCH3), 75.0 (C17), 76.1 (C9), 76.8 (C15), 77.2 (C25), 79.6 (C=C), 81.2
(C=C), 96.7 (C4), 108.3 (C6), 112.2 (C21), 118.2 (C2), 122.6 (C22), 139.4
(C7), 157.2 (C5), 160.2 (C3), 167.4 (C1), 171.0 ppm (C23); HRMS (ESI):
[M+Na]+ calcd for C55H99NNaO8Si3 1008.65707, found 1008.65733.
Deprotected macrolactone 49: HF·pyridine complex (70% HF, 0.3 mL)
was added dropwise to a stirred solution of the enamide 48 (12 mg,
0.012 mmol) in THF (0.4 mL, in a plastic test tube) at ꢀ808C. The reac-
tion mixture was allowed to warm to ꢀ108C. After 2 h the mixture was
partioned between an ice-cooled mixture of EtOAc (20 mL) and saturat-
ed aqueous NaHCO3 solution (20 mL). The organic layer was separated
and the H2O layer extracted with EtOAc (2”20 mL). The combined or-
ganic layers were dried over MgSO4, filtered and concentrated in vacuo.
The residue was purified by flash chromatography (CH2Cl2/MeOH, 95:5
! 9:1) to give 6.1 mg (85%) of triol 49. TLC (CH2Cl2/MeOH, 9:1): Rf =
0.38; [a]2D0 =ꢀ4.2 (c=1.2, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=0.84
(d, J=6.8 Hz, 3H, 18-CH3), 0.91 (t, J=7.1 Hz, 3H, 28-H), 0.99 (d, J=
7.1 Hz, 3H,10-CH3), 1.10 (d, J=7.1 Hz, 3H, 16-CH3), 1.14 (d, J=7.1 Hz,
3718
www.chemeurj.org
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Chem. Eur. J. 2008, 14, 3709 – 3720

Investigation of Cruentaren A Analogues
FULL PAPER
Oxazinan-4-one 52: A 10-mL round-bottom flask was charged with
alkyne 51 (9.0 mg, 0.015 mmol) and stirred with a stir bar. EtOAc (5 mL)
and quinoline (3.0 mg, 0.024 mmol) were added with stirring. This was
followed by the addition of Lindlarꢁs catalyst (5 wt% Pd on CaCO3, pois-
oned with lead, 9 mg, 100 wt%). The reaction was placed under H2 at-
mosphere and stirred for 1 h. The reaction mixture was filtered through a
pad of celite and the filtrate concentrated in vacuo. The residue was puri-
fied by flash chromatography (CH2Cl2/MeOH, 95:5!9:1) to give ana-
logue 52 as a colourless oil (8.4 mg, 93%). TLC (CH2Cl2/MeOH, 9:1):
Rf =0.38; [a]2D0 =ꢀ9.4 (c=0.8, CH2Cl2); 1H NMR (400 MHz, CDCl3): d=
0.78 (d, J=6.6 Hz, 3H, 18-CH3), 0.89–0.96 (m, 6H, 28-H, 16-CH3), 1.01
(d, J=6.8 Hz, 3H, 10-CH3), 1.17 (d, J=7.3 Hz, 3H, 24-CH3), 1.27–1.60
(m, 10H, 19-H, 26-H, 27-H, 20-H, 21-H), 1.82 (br s, 1H, OH), 1.93–2.10
(m, 3H, 18-H, 11-H, 16-H), 2.17–2.42 (m, 5H, 10-H, 14-H, 11-H, 8-H),
2.78–2.89 (m, 1H, 24-H), 3.38–3.47 (m, 1H, 17-H), 3.62–3.75 (m, 3H, 9-
H, 8-H, 25-H), 3.79 (s, 3H, OCH3), 4.71 (t, J=4.6 Hz, 1H, 22-H), 5.23
(dd, J=11.0, 4.2 Hz, 1H, 15-H), 5.38–5.54 (m, 2H, 12-H, 13-H), 6.30 (d,
J=2.5 Hz, 1H, 6-H), 6.36 (d, J=2.5 Hz, 1H, 4-H), 6.40 (br s, 1H, NH),
11.6 ppm (br s, 1H, 3-OH); 13C NMR (100 MHz, CDCl3): d=9.0 (16-
CH3), 12.0 (24-CH3), 14.0 (C28), 14.2 (18-CH3), 15.8 (10-CH3), 18.9
(C27), 20.8 (C20), 29.4 (C14), 29.7 (C11), 31.6 (C19), 32.0 (C21), 33.3
(C26), 36.3, 36.7, 37.1 (C18), 37.8 (C16), 38.3 (C8), 38.8 (C24), 40.1
(C10), 55.4 (OCH3), 72.9 (C25), 75.5 (C9), 78.0 (C15), 83.8 (C22), 99.7
(C4), 104.8 (C2), 112.4 (C6), 125.6 (C13), 132.3 (C12), 143.6 (C7), 163.6
(C5), 165.9 (C3), 171.5 (C1), 174.6 (C23); HRMS (ESI): [M+Na]+ calcd
for C33H51NaNO8 612.35069, found 612.35076.
[3]B. Kunze, F. Sasse, H. Wieczorek, M. Huss, FEBS Lett. 2007, 581,
3523–3527.
[4]For a review about ATP Synthase, see: P. D. Boyer, J. Biol. Chem.
2002, 277, 39045–39061.
[5]a) A. R. Salomon, D. W. Voehringer, L. A. Herzenberg, C. Khosla,
Proc. Natl. Acad. Sci. USA 2000, 97, 14766–14771; b) A. R. Salo-
mon, D. W. Voehringer, L. A. Herzenberg, C. Khosla, Chem. Biol.
2001, 8, 71–80; c) P. A. Wender, O. D. Jankowski, K. Longcore,
E. A. Tabet, H. Seto, T. Tomikawa, Org. Lett. 2006, 8, 589–592;
d) P. T. Daniel, U. Koert, J. Schuppan, Angew. Chem. 2006, 118,
886–908; Angew. Chem. Int. Ed. 2006, 45, 872–893.
[6]a) M. D ’Ambrosio, A. Guerriero, C. Debitus, F. Pietra, Helv. Chim.
Acta 1996, 79, 51–60; b) M. D’Ambrosio, M. Tato, G. Pocsfalvi, C.
Debitus, F. Pietra, Helv. Chim. Acta 1999, 82, 347–353.
[7]For recent total syntheses, see: a) Q. Su, L. A. Dakin, J. S. Panek, J.
Org. Chem. 2007, 72, 2–24; b) L. J. Van Orden, B. D. Patterson,
S. D. Rychnovsky, J. Org. Chem. 2007, 72, 5784–5793.
[8]A. E. Wright, J. C. Botelho, E. Guzmµn, D. Harmody, P. Linley, P. J.
McCarthy, T. P. Pitts, S. A. Pomponi, J. K. Reed, J. Nat. Prod. 2007,
70, 412–416.
[9]A. Zampella, M. V. D ’Auria, L. Minale, C. Debitus, C. Roussakis, J.
Am. Chem. Soc. 1996, 118, 11085–11088.
[10]For a summary, see: V. Wittmann, Nachr. Chem. Tech. Lab. 2002,
50, 841–845.
[11]a) R. Jansen, B. Kunze, H. Reichenbach, G. Hçfle, Eur. J. Org.
Chem. 2002, 917–921; b) B. Kunze, R. Jansen, G. Hçfle, H. Reichen-
bach, J. Antibiot. 2004, 57, 151–155.
[12]For reviews, see: a) J. A. Beutler, T. C. McKee, Curr. Med. Chem.
2003, 10, 787–796; b) L. Yet, Chem. Rev. 2003, 103, 4283–4306.
[13]For recent papers covering V-ATPases, see: a) M. Huss, F. Sasse, B.
Kunze, R. Jansen, H. Steinmetz, G. Ingenhorst, A. Zeeck, H. Wiec-
zorek, BMC Biochemistry 2005, 6, 13; b) K. W. Beyenbach, H. Wiec-
zorek, J. Exp. Biol. 2006, 209, 577–589.
Biological assays: The biological activity of the compounds was tested by
a growth-inhibition assay with fibroblast cells of the mouse cell line L929
(ACC2, DSMZ). The cells were cultivated in Dulbecoꢁs modified Eagle
medium with high glucose and 10% fetal calf serum at 378C and 10%
CO2. Aliquots of 120 mL of suspended cells (50,000 mLꢀ1) were given to
60 mL of serial dilutions of the compounds in 96-well microplates. After
five days, the reduction of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetra-
zolium bromide (MMT) was measured as a parameter of growth and
metabolic activity of the cells and related to control cells that were incu-
bated with the solvent only.
[14]V. V. Vintonyak, M. E. Maier, Org. Lett. 2007, 9, 655–658.
[15]V. V. Vintonyak, M. E. Maier, Angew. Chem. 2007, 119, 5301–5303;
Angew. Chem. Int. Ed. 2007, 46, 5209–5211.
[16]For a review, see: A. Fürstner, P. W. Davies, Chem. Commun. 2005,
2307–2320.
Selected compounds were checked for special phenotypic effects with
PtK2 potoroo cells (ATCC CCL-56). Cells grown on glass coverslips
(13 mm diameter) in four-well plates were incubated with the compounds
overnight, fixed with cold (ꢀ208C) acetone-methanol (1:1) for 10 min
and labelled for endoplasmatic reticulum (ER) with a primary antibody
against GRP-94 (1:1000; Affinity BioReagents) and a secondary goat
anti-rat IgG antibody conjugated with Alexa Fluor 488 (10 mgmLꢀ1; Mo-
lecular Probes).
[17]For some illustrative examples, see: a) A. Fürstner, G. Seidel,
Angew. Chem. 1998, 110, 1758–1760; Angew. Chem. Int. Ed. 1998,
37, 1734–1736; b) A. Fürstner, O. Guth, A. Rumbo, G. Seidel, J.
Am. Chem. Soc. 1999, 121, 11108–11113; c) A. Fürstner, C. Mathes,
C. W. Lehmann, Chem. Eur. J. 2001, 7, 5299–5317.
[18]A. Fürstner, M. Bindl, L. Jean, Angew. Chem. 2007, 119, 9435–
9438; Angew. Chem. Int. Ed. 2007, 46, 9275–9278.
ATPase assays with submitochondrial particles from beef heart were per-
formed in a final volume of 1000 mL and a pH of 8.0 at room tempera-
ture as described previously.[1] The samples contained 150 mg of bovine
protein, 50 mm Tris-HCl (Tris=tris(hydroxymethyl)aminomethane),
50 mm KCl and 2.5 mm MgCl2. The specific ATPase activity without in-
[19]The formation of acyliminium ions seems likely in the benzolactone
enamides, see: X.-S. Xie, D. Padron, X. Liao, J. Wang, M. G. Roth,
J. K. De Brabander, J. Biol. Chem. 2004, 279, 19755–19763.
[20]a) D. W. Hart, T. F. Blackburn, J. Schwartz, J. Am. Chem. Soc. 1975,
97, 679–680; b) S. L. Buchwald, Q. Fang, S. M. King, Tetrahedron
Lett. 1988, 29, 3445–3448.
hibitor was (0.5ꢁ0.13 m)molmgꢀ1 minꢀ1
.
[21]L. Jiang, G. E. Job, A. Klapars, S. L. Buchwald, Org. Lett. 2003, 5,
3667–3669.
[22]R. Nakamura, K. Tanino, M. Miyashita, Org. Lett. 2003, 5, 3583–
3586.
[23]R. Shen, J. A. Porco, Jr., Org. Lett. 2000, 2, 1333–1336.
[24]A. Fürstner, T. Dierkes, O. Thiel, G. Blanda, Chem. Eur. J. 2001, 7,
5286–5298.
Acknowledgements
Financial support by the Deutsche Forschungsgemeinschaft (grant Ma
1012/20–1) and the Fonds der Chemischen Industrie is gratefully ac-
knowledged. We also thank Graeme Nicholson (Institute of Organic
Chemistry) for measuring the HRMS spectra. In addition, a graduate fel-
lowship for V.V. Vintonyak of the state Baden-Württemberg (LGFG) is
acknowledged. We thank Birte Engelhardt and Bettina Hinkelmann for
excellent technical assistance.
[25]K. C. Nicolaou, D. W. Kim, R. Baati, A. OꢁBrate, P. Giannakakou,
Chem. Eur. J. 2003, 9, 6177–6191.
[26]Q. Su, J. S. Panek, J. Am. Chem. Soc. 2004, 126, 2425–2430.
[27]a) J. A. Marshall, G. M. Schaaf, J. Org. Chem. 2001, 66, 7825–7831;
b) Y. Tamaru, S. Goto, A. Tanaka, M. Shimizu, M. Kimura, Angew.
Chem. 1996, 108, 962–963; Angew. Chem. Int. Ed. Engl. 1996, 35,
878–880.
[28]a) K. Horita, T. Yoshioka, T. Tanaka, Y. Oikawa, O. Yonemitsu, Tet-
rahedron 1986, 42, 3021–3028; b) A. Dahan, M. Portnoy, J. Org.
Chem. 2001, 66, 6480–6482; c) C. Wang, C. J. Forsyth, Org. Lett.
2006, 8, 2997–3000.
[1]B. Kunze, H. Steinmetz, G. Hçfle, M. Huss, H. Wieczorek, H. Reich-
enbach, J. Antibiot. 2006, 59, 664–668.
[2]L. Jundt, H. Steinmetz, P. Luger, M. Weber, B. Kunze, H. Reichen-
bach, G. Hçfle, Eur. J. Org. Chem. 2006, 5036–5044.
Chem. Eur. J. 2008, 14, 3709 – 3720
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
3719

M. E. Maier et al.
[29]Z.-Y. Liu, Z.-C. Chen, C.-Z. Yu, R.-F. Wang, R.-Z. Zhang, C.-S.
Huang, Z. Yan, D.-R. Cao, J.-B. Sun, G. Li, Chem. Eur. J. 2002, 8,
3747–3756.
[30]a) D. R. Hicks, B. Fraser-Reid, Synthesis 1974, 203; b) R. D. Cink,
C. J. Forsyth, J. Org. Chem. 1995, 60, 8122–8123; c) A. B. Smith, III,
D.-S. Kim, J. Org. Chem. 2006, 71, 2547–2557.
[31]M. Yamaguchi, I. Hirao, Tetrahedron Lett. 1983, 24, 391–394.
[32]For a representative procedure, see: Y. Kobayashi, M. Asano, S.
Yoshida, A. Takeuchi, Org. Lett. 2005, 7, 1533–1536.
[37]I. Paterson, R. D. M. Davies, A. C. Heimann, R. Marquez, A.
Meyer, Org. Lett. 2003, 5, 4477–4480.
[38]a) S. Ohira, Synth. Commun. 1989, 19, 561–564; b) G. J. Roth, B.
Liepold, S. G. Müller, H. J. Bestmann, Synthesis 2004, 59–62; c) J.
Pietruszka, A. Witt, Synthesis 2006, 4266–4268.
[39]F. Fülçp, G. Bernµth, P. Sohµr, I. Pelczer, J. Chem. Soc. Perkin
Trans. 1 1984, 2043–2048.
[40]M. Panunzio, E. Tamanini, E. Bandini, E. Campana, A. D ’Aurizio,
P. Vicennati, Tetrahedron 2006, 62, 12270–12280.
[33]J. Suffert, D. Toussaint, J. Org. Chem. 1995, 60, 3550–3553.
[34]P. A. Stadler, B. S. Huegi, Helv. Chim. Acta 1985, 68, 1644–1646.
[35]For a recent example, see: A. Fürstner, A.-S. Castanet, K. Radkow-
ski, C. W. Lehmann, J. Org. Chem. 2003, 68, 1521–1528.
[36]a) J. H. Wengrovius, J. Sancho, R. R. Schrock, J. Am. Chem. Soc.
1981, 103, 3932–3934; b) R. R. Schrock, D. N. Clark, J. Sancho, J. H.
Wengrovius, S. M. Rocklage, S. F. Pedersen, Organometallics 1982, 1,
1645–1651; c) M. L. Listemann, R. R. Schrock, Organometallics
1985, 4, 74–83.
[41]F. J. P. Feuillet, D. G. Niyadurupola, R. Green, M. Cheeseman, S. D.
Bull, Synlett 2005, 1090–1094.
[42]T. L. Church, C. M. Byrne, E. B. Lobkovsky, G. W. Coates, J. Am.
Chem. Soc. 2007, 129, 8156–8162.
[43]T. Govender, P. I. Arvidsson, Tetrahedron Lett. 2006, 47, 1691–1694.
[44]See the Supporting Information.
Received: October 23, 2007
Published online: February 27, 2008
3720
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Chem. Eur. J. 2008, 14, 3709 – 3720