Synthesis and biological testings as inhibitors of HMGCoA reductase of the seco-acid of tuckolide and its C-7 epimer

Article abstract of DOI:10.1016/S0968-0896(99)00020-6

The seco-acid of the natural macrolactone, tuckolide (decarestrictin D) and the C-7 epimer have been prepared in enantiomerically pure form from d-gluconolactone and poly(3-hydroxy butyric acid). The key steps are Horner-Emmons olefination and stereoselective reduction of the resulting enone to provide both epimers at C-7. None of the seco-acids inhibit microsomal HMGCoA reductase of pea or rat liver. It may be concluded that the cholesterol biosynthesis inhibiting effect of tuckolide is unlikely to proceed via HMGCoA reductase inhibition. Copyright (C) 1999 Elsevier Science Ltd.

Full text of DOI:10.1016/S0968-0896(99)00020-6

Bioorganic & Medicinal Chemistry 7 (1999) 1049±1057
Synthesis and Biological Testings as Inhibitors of HMGCoA
Reductase of the Seco-acid of Tuckolide and its C-7 Epimer
a
a
a,
b
Ste
Â
phane Colle, Claude Taillefumier, Yves Chapleur, * Rex Liebl
and Arthur Schmidtb
aGroupe SUCRES, UMR 7565 CNRS-Universite Henri PoincareÂ-Nancy I, Case 79, BP 239,
F-54506 Nancy-Vandoeuvre, France
b
Dow AgroSciences, 9330 Zionsville Road, Indianapolis, IN 46268, USA
Received 15 July 1998; accepted 9 October 1998
AbstractÐThe seco-acid of the natural macrolactone, tuckolide (decarestrictin D) and the C-7 epimer have been prepared in
enantiomerically pure form from d-gluconolactone and poly(3-hydroxy butyric acid). The key steps are Horner±Emmons ole®na-
tion and stereoselective reduction of the resulting enone to provide both epimers at C-7. None of the seco-acids inhibit microsomal
HMGCoA reductase of pea or rat liver. It may be concluded that the cholesterol biosynthesis inhibiting e€ect of tuckolide is unlikely
to proceed via HMGCoA reductase inhibition. # 1999 Elsevier Science Ltd. All rights reserved.
Introduction
Decarestrictin D, a macrolactone recently isolated from
4
a
Penicillium corylophilum and P. simplicissimum, and
tuckolide, the same compound isolated from the fungi
Polyporus tuberaster,4b proved to be inhibitors of choles-
The control of cholesterol blood level is of considerable
interest for the control of coronary diseases which are
responsible for about 40% of morbidity in developed
countries. Ecient drugs are now on the market and
most of these compounds, known as statines or mevinic
acids, are more or less related to a family of lactonic
compounds derived from the lead compounds compac-
tin and mevinolin. They are inhibitors of the rate-
limiting enzyme of cholesterol biosynthesis, 3-hydroxy-
-methyl-glutaryl coenzyme A reductase (HMGR),
5
terol biosynthesis in vitro in the HEP-G2 cells assay. These
data were con®rmed in vivo with an activity at 10 mg/kg
in rats equivalent to clo®brate at 100 mg/kg. Moreover
no antibacterial and antifungal activities were detected.
1
,2
Tuckolide/decarestrictin D should be an interesting lead
compound for the discovery of new cholesterol lowering
agents. This interest was recently reinforced by the dis-
closure by Andrus and Shih of a total synthesis of this
compound in optically active form using Sharpless
dihydroxylation as the key step.6 In this paper the
suggestion that tuckolide would be an inhibitor of
HMGR on the basis of structural similarities between
tuckolide and mevinolin attracted our attention. These
similarities rely on the presence of a lactone moiety and
the presence of an hydroxy group in b position of the
carbonyl group. However the lipophilic part, present in
mevinolin, which is essential for inhibition cannot be
found in tuckolide. The lactone function of `mevinic
3
which is responsible for the double reduction of 3-hy-
droxy-3-methyl-glutaryl coenzyme A into mevalonic
acid. It is known that the control of this enzyme is e-
,7
3
cient in the lowering of plasma cholesterol (Fig. 1).
The biologically active form of mevinic acids is the open
chain hydroxy-acid which mimics the natural substrate
-hydroxy-3-methyl-glutaryl coenzyme A. At the mole-
cular level the key feature of mevinic acids is the pre-
sence of a dihydroxy-acid moiety associated with a
lipophilic part which is quite intricate in the natural
inhibitors such as compactin, mevinolin and pravasta-
tin. The search for more simple compounds resulted in
the preparation of aromatic derivatives associated with
the hydroxy-acid part.2
3
acids' is formed upon isolation and it is well known that
the free acid is the biologically active form.8
±10
This
suggested that the seco-acid of tuckolide might be the
real inhibitor of HMGR. In that case the open-chain
acid of tuckolide would be a lead compound en route to
a new class of `hydrophilic' inhibitors with a rather
simple structure.11 On this basis we decided to prepare
the seco-acid of tuckolide and to evaluate its biological
properties as inhibitor of HMGR.
Key words: Cholestereol biosynthesis; HMGCoA reductase; inhibi-
tion; tuckolide; synthesis.
Corresponding author. Tel.: 33-3-83-91-23-55; fax: 33-3-83-91-24-79.
*
0968-0896/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(99)00020-6

1
050
S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
ꢀ
with lithiotrimethylphosphonate at low temperature to
at � 10 C led to amide 4 (86%), which was condensed
prevent any epimerisation. This led reproducibly to the
expected phosphonate 5.1
5±18
At this stage the extent of
epimerisation, if any, cannot be determined. However,
in the subsequent step, no detectable formation of dia-
stereoisomers which may be formed from the S-epimer
of 5 was observed.
Figure 1.
Synthesis of the aldehydic moiety
The presence in d-glucono-1,5-lactone of all the chiral
centers needed in synthon II made it an ideal precursor
provided that the carboxylic acid could be eciently
transformed into an aldehyde. We started from the
known ester 6 prepared in a single step from d-glucono-
lactone in 50% yield.19 The alcohol group of 6 was
eciently removed under radical conditions by forma-
tion of a phenylthiocarbonate or better by use of imi-
dazolyl thionocarbonate 7 followed by treatment with
tributyltin hydride. The deoxy compound 8 was
Results
Our retrosynthetic analysis of hydroxy-acid 24 is based
on an ole®nic coupling between two enantiomerically
pure building blocks. The phosphonate moiety I could
be obtained by condensation of a phosphonate anion on
a suitable derivative of 3-hydroxy butyric acid. The
aldehydic part II could be obviously prepared from d-
gluconic acid 1,5-lactone, a cheap and readily available
starting material.
1
9
obtained routinely in 60±65% for the two steps.
In order to achieve the reduction of the ester group into
the required aldehyde function, we turned once again to
Weinreb amide, the reduction of which is well documented
and provide almost pure aldehyde upon treatment with
lithium aluminium hydride. Treatment of ester 8 with
the magnesium salt of N,O-dimethylhydroxylamine led
to the amide 9 in 70% yield, together with a small
amount of the corresponding isopropyl ketone (25%)
because of the competing reaction of 8 or 9 with iso-
propyl magnesium chloride. The amide 9 was then sub-
jected to lithium aluminium hydride reduction. Total
consumption of the starting material was evident from
TLC analysis, but to our surprise, hydrolysis of the
Synthesis of the phosphonate moiety
Poly [(R) 3-hydroxy butyric acid] 1 was depolymerized
according to Seebach procedure to provide ester 2 in
1
2
7
1% yield. The hydroxyl group of 2 was protected as a
silyl ether using conventionnal procedure to provide
ester 3 in 95% yield. The next crucial step was the
homologation of ester 3 to the ketophosphonate 5.
Although the direct condensation of lithiophosphonates
on esters is possible, Weinreb amide was preferred.13
Ester 3 was transformed into amide 4 using an ecient,
recently reported, procedure.14 Treatment of 3 with the
magnesium salt of N,O-dimethylhydroxylamine in THF
Scheme 1. Retrosynthetic analysis of tuckolide-seco-acid 24.

S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
1051
Scheme 2. Reagents: (i) H
ꢀ
2
SO
, 1,2-dichlorethane:MeOH, re¯ux, 72 h; (ii) TBDMSCl, CH Cl
4 2
2
, NEt
3
, DMAP; (iii) LiCH
2
P(O)OMe
2
, THF,
�
100 C.
Scheme 3. Reagents: (i) DMP, Acetone, PTSA; (ii) (Imid) CS, 2 equiv, pyridine/CH
2
2
ꢀ
Cl
2
, rt, 18 h; (iii) Bu SnH, 1.2 equiv, AIBN, degased toluene,
3
.
ꢀ
6
Et
h, 60±65%; (iv) MeNH(OMe) HCl 1.6 equiv THF then i-PrMgCl, 3.2 equiv, � 20 C; (v) LiAlH
4
, 1.2 equiv THF, 0 C, 89%; (vi) 5, 1.5 equiv,
.
O, LiOH H
2
2
O, 2.6 equiv then 10, Et
2
O, 82.
reaction mixture led to only small amounts of the
expected aldehyde 11. The main compound was isolated
and identi®ed as the hemi-aminal resulting from simple
reduction of the carbonyl group. This was evident from
spectral data which showed the presence of an hydroxyl
and the N,O-dimethyl groups and the presence of two
diastereoisomers at the newly created chiral center. This
stability can be explained by a possible hydrogen bond
formation between the OH group and the neighboring
groups which preclude the decomposition of the adduct
in neutral medium. Assuming that the hemi-aminal 10
should decompose easily in slightly basic medium, we
attempted the Horner±Emmons ole®nation between 5
and 10. To our delight, using Blackwell conditions
desilylated (TBAF, THF, rt) and the resulting diols
were treated with dimethoxypropane and p-toluene-
sulfonic acid in acetone to give the corresponding
acetonides 18 or 19 in 95% yield. The absolute con®g-
uration at C-8 of each acetonide was determined using
the 13C NMR method developed by Rychnovsky. The
di€erence of chemical shifts of the methyl groups of the
acetonide are characteristic of the anti or syn con®g-
uration. Accordingly we were able to attribute the anti
con®guration to diol 14 and the syn con®guration to
diol 15.
21
We also explored the stereoselective reduction, taking
advantage of the presence of the hydroxy group at C-10
to direct the reduction at C-8. Such stereoselective
reductions of b-hydroxy-ketones have ample precedents
in the literature, however the reduction of a b-hydroxy-
enone under such conditions is by far less documented.
In order to direct the reduction of the carbonyl group at
C-8, the hydroxyl group at C-10 must be desilylated to
ensure complexation to the boron reagent. However all
attempts to e€ect conventional desilylation (TBAF,
THF, rt) failed to give the expected alcohol probably
due to undesired reaction of the enone in basic medium.
We turned to acid hydrolysis and found that under
2
0
(LiOH-H O, ether), a clean reaction occurred giving
2
only the trans ole®n in 82% yield.
Stereoselective reduction of the carbonyl group
The last key step of the synthesis was the reduction of
the carbonyl group at C-8 of enone 12. In a ®rst series
of experiments this enone was reduced stereo-randomly
using Luche conditions. A 1:1 mixture of the two C-8
epimers 14 and 15 was obtained in 85±90%. The two
compounds were easily separated by column chromato-
graphy. In order to determine the absolute con®gura-
tion at C-8 of 14 and 15, each of them was separately
carefully controlled conditions (AcOH, THF, H O, rt, 4
2
days), it was possible to isolate the desired hydroxy-

1
052
S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
Scheme 4. Reagents: (i) NaBH
4
, CeCl , MeOH, 90%; (ii) TBAF, 1.1 equiv, THF; (iii) DMP, Acetone, PTSA cat., 95% 2 steps; (iv) AcOH:-
3
H
2
4 3 3
O:THF, 3:7:20, 4 days, rt; (v) Me NBH(OAc) , 5 equiv, CH CN:AcOH, 1:1.
ketone 13 in fair yield together with some starting
material. The formation of a saturated product resulting
probably from 1,4-addition on the enone system was
also observed but was not further investigated. Evans
method was chosen to reduce the carbonyl group to
obtain the anti con®guration as in the natural product.22
Treatment of ketone 13 with tetramethylammonium
triacetoxy borohydride in acetonitrile provided a 9:1
mixture of the diols 16 and 17. The anti con®guration of
compound 16 was con®rmed by transformation into the
acetonide 18 previously prepared. The observed stereo-
selectivity of the reagent for the anti con®guration was
in agreement with literature data. It is interesting to
note that no double bond reduction is observed. Due to
the diculty to remove the silyl protecting group to
achieve chelation controlled reduction, no attempts
were made to prepare the syn isomer by a similar route.
We preferred to use the non-stereoselective reduction
and subsequent chromatographic separation of the dia-
stereoisomers.
isopropylidene group and oxidative cleavage of the
resulting 1,2-diol. All attempts to use Wu's method
24
(H IO , ether) to perform both reactions in a single
5
6
step failed. A stepwise procedure was explored. We
found that careful acid hydrolysis (AcOH:H O:THF,
2
ꢀ
9:1:5, 55 C) led to the tetrols 20 and 21, respectively, in
about 80% yield and fully deprotected material in about
10% yield. Each diol was then successively treated with
sodium periodate to carry out the diol cleavage and the
resulting aldehyde was oxidized without puri®cation
(NaClO , tBuOH, 2-methyl-2-butene) to provide the
2
corresponding carboxylic acids 22 (88%) and 23 (68%),
respectively. The ®nal step was the removal of the last
2
3
protecting group by acid hydrolysis (AcOH:H O:THF,
2
ꢀ
droxide to yield the target compounds 24 and 25.
1:1:2, 60 C) followed by treatment with sodium hy-
Biological evaluation
As was previously mentioned, in animal and plant cells
HMGR is a highly regulated control point in the bio-
synthesis of a vast array of isoprenoids and prenyl-
lipids. In plants the isoprenoid pathway has additional
branches that lead to photosynthetic pigments, growth
regulators (abscisic acid, gibberellins, and some cyto-
kinins) and phytoalexins. Therefore, inhibition of
HMGR-linked isoprenoid biosynthesis would have wide
Diol cleavage and ®nal deprotection
The last steps of the synthesis were performed on com-
pounds 14 and 15. Desilylation under standard condi-
tions led to diols 16 and 17, respectively. The remainder
of the synthesis consisted in selective removal of one
ꢀ
Scheme 5. Reagents: (i) AcOH:H
NaH PO , 4.5 equiv in H
2
O:THF, 9:1:5, 55 C, 80%; (ii) NaIO , 1.5 equiv, MeOH:H
4
2
O; 2-methyl-2-butene, tBuOH then NaClO
2
, 6 equiv,
ꢀ
2
4
2
O; (iv) AcOH:H
2
O:THF, 1:1:2, 60 C, 20 h then NaOH.

S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
1053
Table 1. Lemna and HMGR inhibition by 24 and 25
France) using silica gel (Merck 60 40±60 m). Optical
rotations were measured on a Perkin±Elmer 141 polari-
meter at 20 C. Melting points were measured in capil-
Lemna I50 (ppm)a
HMGR I50a
ꢀ
Rat
Pea
lary tubes and are uncorrected. The elementary analyses
were performed by the Service Central de Microanalyses
du CNRS at Vernaison, France. Mass spectra were
obtained on a Nermag R10-10C in the EI mode or VG
Plattform in the ES mode. Tetrahydrofuran was dis-
tilled prior to use from sodium-benzophenone.
6
� 7
Mevastatin
0.1
1.1Â10�
M
M
M
4Â10
M
M
>50b
� 4
� 4
b
b
� 4
b
b
2
2
4
5
>7.5Â10
>7.5Â10
>7.5Â10
>7.5Â10
>50b
� 4
M
a
I
50 concentration which causes 50% inhibition of the control values.
No inhibition at the speci®ed concentration.
b
(R)-Methyl 3-O-(tert-butyldimethylsilyl)-butanoate (3).
To a stirred solution of 2 (10.8 g, 91.2 mmol) in
12
CH Cl2 (300 mL) under a nitrogen atmosphere were
2
ranging e€ects on plant growth and development.
Mevastatin was a potent inhibitor of pea and rat liver
microsomal HMGR activity. 24 and 25 did not inhibit
either the plant or mammalian enzymes. These results
correlated with biological activity where mevastatin was
a potent inhibitor of Lemna growth (GR50 0.1 ppm).
However, 24 and 25 were not active on Lemna.
added
successively
tert-butylchlorodimethylsilane
(13.7 g, 91.2 mmol), Et N (20 mL), DMAP (1.10 g,
3
9.1 mmol) and Et N (18 mL). The reaction mixture was
3
stirred at room temperature for two days and then
washed successively with 3 N HCl (90 mL) and water.
The organic layer was dried (MgSO ) and evaporated
under vacuum. Flash chromatography on silica gel
a€orded the silyl ether 3 (20.1 g, 95%) which can be
4
used in the next step. R 0.74 (hexane:ethyl acetate, 8:2);
f
1
H NMR: d 0.05 (s, 3H, SiCH ), 0.07 (s, 3H, SiCH ),
3
3
Conclusion
0
.87 (s, 9H, SiC(CH ) ), 1.21 (d, 3H, H C-
3 3 3
The present work has provided two new compounds
corresponding to the open form of the macrolactone
tuckolide and its C-6 epimer starting from two enantio-
merically pure natural products. The key feature of
these syntheses are the ecient and highly stereo-
controlled Horner±Emmons coupling between a phos-
phonate and an hemi-aminal resulting from the
reduction of a Weinreb amide. Subsequent reduction of
the resulting b-hydroxy-enone is possible with high ste-
reocontrol using Evans conditions without reduction of
the double bond. None of the ®nal products have shown
interesting inhibitory activity toward HMGR. Because
it has been postulated that the tight binding of HMGR
inhibitors is the result of the compounds ability to
simultaneously interact with the HMGR binding
domain of the enzyme and an adjacent hydrophobic
pocket, which does not appear to be utilized in substrate
binding, it is not surprising that the absence of a lipo-
philic moiety, such as the decalin ring present on mevi-
nic acids, on 24 and 25 likely limits the binding of these
compounds to HMGR. Our results show that the
reported inhibition of cholesterol biosynthesis by tuck-
olide is likely not related to the inhibition of HMGR by
the open-chain hydroxy-acid but is probably related to
a subsequent step of the biosynthesis.
CHOTBDMS, J=6 Hz), 2.38 (dd, 1H, CH COOCH ,
2 3
J=3, Jgem=9 Hz), 3.68 (s, 3H, COOCH ), 4.28 (m, 1H,
3
CHOTBDMS).
(R)-3-O-(tert-Butyldimethylsilyl)-N-methoxy-N-methyl-
butanamide (4). The ester 3 (5.0 g, 21 mmol) and N-
methoxy-N-methylamine
hydrochloride
(2.92 g,
30 mmol) were suspended in 40 mL of THF cooled to
ꢀ
propyl magnesium chloride in THF (25 mL, 2.4 M) was
� 10 C under nitrogen atmosphere. A solution of iso-
ꢀ
added dropwise below � 10 C. The mixture was stirred
ꢀ
NH Cl and diluted with ether. The organic layer was
for 2 h at � 10 C, quenched with saturated aqueous
4
washed with water, dried over MgSO4 and con-
centrated. Puri®cation on silica gel a€orded the amide 4
as a clear oil (5.5 g, 86%). R 0.25 (hexane:ethyl acetate,
f
9:1). 1H NMR: d 0.02 (s, 3H, SiCH ), 0.07 (s, 3H,
3
SiCH3), 0.88 (s, 9H, SiC(CH ) ), 1.20 (d, 3H, H ,
J3,4=6 Hz), 2.35 (dd, 1H, H , Jgem=14, J2,3=5.5 Hz),
3
3
4
2
0
(s, 3H, OCH ), 4.35 (ddd, 1H, H ); MS (m/z): 246
2.77 (dd, 1H, H , J
2
0
=7 Hz), 3.18 (s, 3H, NCH ), 3.70
2 ,3 3
3
3
+
(M� 15) . Anal. calcd for C H NO Si: C, 55.14; H,
12
27
3
10.42; N, 5.36. Found: C, 55.56; H, 10.11; N, 5.30.
(R)-4-O-(tert-Butyldimethylsilyl)-1-(dimethoxyphosphinyl)-
pentan-2-one (5). A 100-mL round-bottomed ¯ask
equipped with a rubber septum was charged with n-
BuLi (10.9 mL, 1.25 M in hexane) under an argon
Experimental
ꢀ
atmosphere and cooled to � 80 C. Dimethyl methyl-
1
H NMR spectra were recorded with a Bruker AC 250
1
phosphonate (1.5 mL, 13.9 mmol) was added dropwise
by syringe. During addition, a mixture of THF (2 mL)
and Et O (2 mL) was added to aid in stirring. The reac-
3
operating at 250 MHz and 63 MHz for the C, using
deuteriochloroform as solvent. Assignments were con-
2
®
rmed by double irradiation or two dimensional spec-
troscopy. Chemical shifts are reported relative to
tion mixture was stirred 15 min after the completion of
the addition and cooled to � 110 C, a solution of 4
ꢀ
(2.81 g, 10.8 mmol) in a mixture of THF (1 mL) and
internal SiMe . TLC was performed on silica gel plates
4
(
(
Merck 60F254). Column chromatography used silica gel
Merck 60 70±23 mesh). Preparative high pressure liquid
Et O (1 mL) was added. The mixture was stirred at
2
ꢀ
� 110 C for 15 min and allowed to warm slowly to
ꢀ
mixture of 1 M H PO (20 mL) and Et O (60 mL). The
chromatography was performed on 40 mm diameter
stainless steel column (Prochrom, Champigneulles,
� 70 C over 20 min. The reaction was quenched with a
3
4
2

1
054
S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
layers were separated and the aqueous layer was
extracted twice with EtOAc (100 mL). The combined
extracts were washed with saturated aqueous NaHCO3,
Et O (65 mL) was stirred under nitrogen at room tem-
2
perature during 0.5 h. A solution of compound 10 (4.5 g,
14.7 mmol) in Et O (25 mL) was then added dropwise
2
brine and dried over MgSO . The crude product was
4
over 15 min. After stirring during 0.5 h, this mixture was
hydrolysed with satured aqueous NH Cl (10 mL). The
chromatographed on silica gel to a€ord 5 as a clear oil
ꢀ
4
(
0
2.70 g, 77%). R 0.51 (ethyl acetate); [a] � 30.5 (c
organic layer was washed with water (2Â10 mL), dried
f
d
�
1 1
.91, CHCl ); IR: 1715 cm ; H NMR: d 0.01 (s, 3H,
over MgSO and concentrated under reduced pressure.
4
3
SiCH ), 0.045 (s, 3H, SiCH ), 0.85 (s, 9H, SiC(CH ) ),
1.15 (d, 3H, H , J=6 Hz), 2.61 (dd, 1H, H , Jgem=15.5,
Column chromatography (hexane:ethyl acetate, 9:1)
a€orded 12 (5.39 g, 17.7 mmol, 82%). R 0.55 (hexane:
3
3
3 3
5
3
f
ꢀ
J3,4=5 Hz), 2.78 (dd, 1H, H3
H1
J=1 Hz), 3.78 (s, 3H, OCH ), 4.26 (m, H ); C NMR:
0
, J
0
=7 Hz), 3.05 (d, 1H,
ethyl acetate, 8:2); [a] � 21.3 (c 1.1, CHCl ); IR: 1674,
3 ,4
d
3
�
1 1
0
), 3.14 (d, 1H, H , Jgem=3 Hz), 3.73 (d, OCH ,
1634, 1472, 1371 cm ; H NMR: d 0.04 (s, 3H, SiCH ),
1
3
3
13
0.01 (s, 3H, SiCH ), 0.82 (s, 9H, SiC(CH ) ), 1.15 (d,
=6 Hz), 1.32 (s, 3H, C(CH ) ), 1.36 (s,
10,11 3 2
3
4
3
3 3
d 4.99±4.55 (Si(CH ) ), 18.00 (SiC(CH ) ), 23.88 (C ),
3H, H , J
11
3
2
3 3
5
3
1
2
5.76 (3C, SiC(C)H ) ), 41.57, 43.61, 53.30; P NMR: d
3H, C(CH ) ), 1.38 (s, 3H, C(CH ) ), 1.39 (s, 3H,
3 2 3 2
3
3
2
3.57 (PO(OCH ) ).
C(CH ) ), 1.73 (ddd, 1H, H , Jgem=15.5, J3,4=8.5,
3 2 3
3
2
J4a,5=5 Hz), 1.86 (ddd, 1H, H , J
0
=8, J2,3
2.49 (dd, 1H, H , Jgem=15.5, J9,10=5 Hz), 2.79 (dd, 1H,
0
=4 Hz),
10
3 ,4
4
-Deoxy-2,3:5,6-di-O-isopropylidene-D-xylo-N-methoxy-
1
9
9
methylhexonamide (9). To a solution of ester 8 (5.2 g,
8.9 mmol) and N-methoxy-N-methylamine hydrochlo-
ride (2.95 g, 30 mmol) in anhydrous THF (50 mL) under
H9
0
,
J
J1,2=6.5 Hz), 3.83 (ddd, 1H, H , J3,4=3.5 Hz), 4.05 (dd,
0
=7 Hz), 3.54 (dd, 1H, H , Jgem=8.5,
9 ,10 1
1
4
1H, H1
(m, 1H, H3
0 0
, J =6 Hz), 4.10±4.22 (m, 2H, H , H ), 4.30
0
1 ,2 2 5
ꢀ
nitrogen at � 20 C was added slowly over 1 h a solution
), 6.34 (dd, 1H, H , J
7
=16, J5,7=1.5 Hz),
trans
13
of isopropyl magnesium chloride (30 mL, 2 M in Et O,
2
6.73 (dd, 1H, H , J5,6=5.5 Hz); C NMR: d � 4.53,
6
60 mmol). The reaction was monitored by TLC
� 4.88 (Si(CH ) ), 18.19 (SiC(CH ) ), 24.21 (C ), 25.77
3
2
3 3
11
(
hexane:ethyl acetate, 7:3) and after 1 h, was quenched
by saturated aqueous NH Cl (8 mL). The aqueous layer
(3C, SiC(CH ) ), 26.75 (C(CH ) ), 27.03, 27.26
3 3 3 2
(C(CH ) ), 37.75 (C ), 50.17 (C ), 65.73 (C ), 69.78
3 2 3 9 10
4
was extracted with Et O and the combined organic lay-
2
(C ), 73.39 (C ), 77.49 (C ) 80.67 (C ), 108.95, 109.71
1 2 4 5
ers were dried over MgSO and concentrated. The crude
4
(C(CH ) ), 131.46 (C ), 141.70 (C ), 198.75 (C ), MS
3 2 7 6 8
+
product was puri®ed by column chromatography on
silica gel (hexane:ethyl acetate, 7:3) to provide the
expected amide 9 (3.91 g, 12.9 mmol, 68%). R 0.80
(m/z): 427 (M� 15) . Anal. calcd for C H O Si: C,
23
42
6
62.40; H, 9.60. Found: C, 62.06; H, 9.61.
f
ꢀ
(
hexane:ethyl acetate, 7:3); [a] � 1.6 (c 1.6, CHCl );
(2S,4S,5S,6E,10R)-10-Hydroxy-1,2:4,5-di-(isopropylidene-
dioxy)-6-undecen-8-one (13). To a stirred solution of the
silyl ether 12 (0.52 g, 1.17 mmol) in THF (20 mL) was
added a mixture of water (7 mL) and acetic acid (3 mL).
After being stirred for 4 days at room temperature, the
solution was neutralised with solid sodium carbonate.
The aqueous layer was extracted with EtOAc and com-
bined organic layers were washed with saturated aqu-
d
3
�
1
1
IR: 1670, 1457, 1380 cm ; H NMR: d 1.27 (s, 3H,
C(CH )), 1.34 (s, 3H, C(CH )), 1.40 (s, 6H, C(CH )),
3
3
3
1.82 (ddd, 1H, H , Jgem=14, J4,5=5, J3,4=7 Hz), 1.93
4
(
ddd, 1H, H4
0
, J
.51 (dd, 1H, H , Jgem=8.5 Hz, J5,6=6.5 Hz), 3.70 (s,
0
=7, J3,4
0
=5 Hz), 3.18 (s, 3H, NCH3),
4 ,5
3
3
1
2
6
6
H, OCH ), 4.03 (dd, 1H, H , J5,6 =6 Hz), 4.12 (m,
0 0
6
H, H ), 4.40±4.51 (m, 2H, H , H ); C NMR d 25.60,
5.95, 26.82, 27.29 (4C, C(CH ) ), 37.41 (C ), 61.47,
6.67 (OCH , NCH ), 69.33, 69.64, 73.31, 77.46
3
13
5
2
3
eous NaCl, dried over Na SO4 and the solvent
2
3
2
4
evaporated under vacuum. Puri®cation by HPLC (hex-
ane:ethyl acetate gradient from 90:10 to 50:50) a€orded
the b-hydroxyketone 13 (0.191 g, 0.58 mmol, 50%)
accompanied with some starting material (0.103 g,
3
3
(
(
4C, C , C , C , C ), 108.58, 110.38 (C(CH ) ), 169.98
2 3 5 6 3 2
C1).
-Deoxy-2,3:5,6-di-O-isopropylidene-D-xylo-hexose (11).
To a solution of Weinreb amide 9 (3.75 g, 12.4 mmol)
4
0.23 mmol, 20%). R 0.34 (hexane:ethyl acetate, 5:5);
f
ꢀ
J10,11=6 Hz), 1.33 (s, 3H, C(CH ) ), 1.38 (s, 3H,
1
[a] 33.8 (c 2.06, CHCl ); H NMR: d 1.18 (d, 1H, H ,
d
3
11
ꢀ
in anhydrous THF (120 mL) at 0 C under nitrogen was
added lithium aluminium hydride (0.635 g, 15 mmol) in
3 2
C(CH ) ), 1.40 (s, 3H, C(CH ) ), 1.42 (s, 3H, C(CH ) ),
3
2
3 2
3 2
3
portions over 10 min. After stirring for 5 min, hydro-
lysis was carefully carried out by adding water (0.7 mL),
0% aqueous solution of NaOH (1.4 mL) and ®nally
water (15 mL). The aqueous layer was extracted with
1.78 (ddd, 1H, H , Jgem=13, J=5 Hz, J=8.5 Hz), 1.86
3
(ddd, 1H, H3
H9 ), 3.55 (dd, 1H, H , Jgem=8, J1,2=7 Hz), 3.86 (ddd,
1H, H , J=8.5 Hz), 4.07 (dd, 1H, H1
0
, J=4, J=7 Hz), 2.55±2.80 (m, 2H, H9,
3
0
1
0
, J =6 Hz),
0
1 ,2
4
2
Â40 mL of Et O. The combined organic layers were
4.12±4.31 (m, 4H, H , H , H , H ), 6.35 (dd, 1H, H ,
2 5 8 10 7
2
dried over MgSO4 and solvents were evaporated.
Puri®cation by column chromatography on silica gel
J
trans=16, J5,7=1 Hz), 6.80 (dd, 1H, H , J5,6=5 Hz);
6
1
3
C NMR: d 22.34 (C ), 25.67, 26.65, 26.96, 27.17 (4C,
11
(
hexane:ethyl acetate, 5:5) gave the hemi-aminal 10
C(CH ) ), 36.76 (C ), 48.25 (C ), 63.82 (C ), 69.70 (C ),
73.35 (C ), 77.97 (C ), 80.55 (C ), 108.98, 109.80
2 4 5
(C(CH ) ), 130.55 (C ), 142.58 (C ), 200.33 (C ).
3 2 7 6 8
3 2 3 9 10 1
(3.39 g, 11.1 mmol, 89%) as a diastereoisomeric mixture
that was used in coupling with phosphonate 5 without
further puri®cation.
Compounds 14 and 15. To a solution of ketone 12
.
(1.50 g, 3.39 mmol) and CeCl 7H O (1.26 g, 3.39 mmol)
3 2
in 30 mL of MeOH at 0 C was added NaBH (0.128 g,
4
3.39 mmol) in one portion. A vigorous gas evolution
took place. Stirring was continued for a few minutes
(
2S,4S,5S,6E,10R)-10-O-(tert-Butyldimethylsilyl)-1,2:4,5-
ꢀ
di-(isopropylidenedioxy)-6-undecen-8-one (12). A solu-
tion of phosphonate 5 (7.10 g, 21.9 mmol) and lithium
hydroxide monohydrate (0.93 g, 38.7 mmol) in anhydrous

S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
1055
before the pH was adjusted to 7 with diluted aqueous
HCl. MeOH was evaporated under vacuum and the
yellow residue was diluted with Et O and washed with
2
(2S,4S,5S,8R,10R)-8,10-Dihydroxy-1,2:4,5-di-(isopropylid-
enedioxy)-6-undecene (17). Compound 17 was prepared
in quantitative yield from 15 according to the above
H O and brine. The organic layer was dried over
2
MgSO and concentrated in vacuo. The crude material
4
was puri®ed by preparative high pressure liquid chro-
matography (hexane:ethyl acetate, 85:15) to give 15
mentioned procedure. R 0.15 (hexane:ethyl acetate,
f
ꢀ
� 1
5:5); [a] 7.2 (c 0.9, CHCl ); IR: 3396, 1457, 1370 cm
;
d
3
1
H NMR: d 1.10 (d, 3H, H , J10,11=6 Hz), 1.29 (s, 3H,
11
C(CH ) ), 1.32 (ls, 9H, C(CH ) ), 1.44� 1.69 (m, 3H,
3
2
3 2
(
3
0.78 g, 1.77 mol, 52%) and 14 (0.56 g, 1.28 mmol,
8%).
H , H
9
0
, H ), 1.79 (ddd, 1H, H
3
0
, J =12.5, J
gem
0
=7.5,
2,3
9
3
J3 =2.5 Hz), 3.51 (dd, 1H, H , Jgem=9, J1,2=7 Hz),
0
,4 1
3
.70 (ddd, 1H, H , J3,4=10.5, J4,5=7.5 Hz), 3.91 (dd,
4
(2S,4S,5S,8S,10R)-10-O-(tert-Butyldimethylsilyl)-8-hy-
droxy-1,2:4,5-di-(isopropylidenedioxy)-6-undecene (14).
1H, H , J =7.5 Hz), 3.85� 4.05 (m, 1H, H ), 4.02 (dd,
5
5,6
10
1H, H1 , J
3.60� 4.20 (2H, 2OH), 4.29 (ddd, 1H, H , J7,8=5.5, J
0
0
=6 Hz), 4.15 (dddd, 1H, H , J =5.5 Hz),
2
1 ,2
2,3
ꢀ
R 0.55 (hexane:ethyl acetate, 7:3); [a] � 8.6 (c 1.33,
f
d
8
8,9
1
CHCl ); H NMR: d 0.08 (s, 6H, Si(CH ) )), 0.90 (s,
J
0
=12 Hz), 5.58 (dd, 1H, H , J
6
=15, J5,6=7.5 Hz),
trans
3
3 2
8,9
13
9
H, SiC(CH ) )), 1.23 (d, 3H, H , J10,11=6.5 Hz), 1.34
5.76 (dd, 1H, H , J7,8=5.5 Hz); C NMR: d 23.88 (C1),
3
3
11
7
(
3
J3
s, 3H, C(CH ) )), 1.38 (s, 9H, C(CH ) )), 1.52±1.73 (m,
3
25.73 (C(CH ) ), 26.98 (C(CH ) ), 27.27 (C(CH ) ),
36.28 (C ), 44.54 (C ), 67.98 (C ), 69.78 (C ), 72.03
3 9 10 1
(C ), 73.78 (C ), 77.74 (C ), 82.00 (C ), 108.72, 109.02
8 2 4 5
2
3 2
3 2
3 2
3 2
H, H , H , H
0
0
), 1.90 (ddd, 1H, H3
0
, Jgem=14, J2,3
0
=7,
,4=2.5 Hz), 3.37 (d, 1H, OH, J8,OH=2 Hz), 3.58 (dd,
3
9
9
1H, H , Jgem=8.5, J1,2=7 Hz), 3.79 (ddd, 1H, H4,
J3,4=10, J4,5=9.5 Hz), 3.98 (dd, 1H, H , J5,6=6.5 Hz),
(C(CH ) ), 126.36 (C ), 137.88 (C ). MS (m/z): 315
3 2 6 7
1
+
(M� 15) . Anal. calcd for C H O : C, 61.8; H, 9.2.
5
17 30
6
4
(
.08 (dd, 1H, H1
0
, J
m, 1H, H8), 5.68 (ddd, 1H, H6,
0
=5.5 Hz), 4.21 (m, 1H, H ), 4.50
2
Found: C, 61.38; H, 9.22.
1 ,2
Jtrans=15.5,
3
1
J6,8=1 Hz), 5.84 (dd, 1H, H , J7,8=5 Hz); C NMR: d
(2S,4S,5S,8S,10R)-1,2:4,5:8,10-Tri-(isopropylidenedioxy)-
6-undecene (18). To a stirred solution of 14 (0.172 g,
0.38 mmol) in THF was added TBAF (0.40 mmol, 1M
solution in THF) at room temperature. After being
7
�
5.01, � 4.43 (Si(CH ) ), 22.85 (C ), 25.74 (1C,
3
2
11
0
C(CH ) ), 25.77 (3C, C CH ) ), 26.99 (C(CH ) ), 27.27
3
2
3 2
3 2
(
C ), 69.85 (C ), 73.77 (C ), 77.80 (C ), 82.19 (C5),
1C, C(CH ) ), 36.31 (C ), 44.18 (C ), 67.26, 68.39 (C ,
3 2 3 9 8
stirred for 1 h, the mixture was diluted with Et O,
2
1
0
1
2
4
1
08.76, 108.87 (C(CH ) ), 125.97 (C ), 138.36 (C ). MS
7
washed successively with saturated aqueous NH Cl,
4
NaCl, dried (MgSO ) and the solvent removed in vacuo.
4
3
2
6
+
(
6
m/z): 329 (M� 15) . Anal. calcd for C H O Si: C,
2
3
44
6
2.12; H, 9.97. Found: C, 62.39; H, 10.04.
Without further puri®cation the resulting oil was dis-
solved in a mixture of acetone (5 mL) and 2,2-di-
methoxypropane (0.5 mL) and a catalytic amount of
APTS was added. After a few minutes the reaction was
®nished and the mixture was neutralised with solid
sodium carbonate, ®ltered through a pad of Celite and
evaporated under vacuum. The residue was puri®ed by
chromatography on silica to a€ord compound 18 in
95% yield. R 0.11 (hexane:ethyl acetate 5:5); [a]
(2S,4S,5S,8R,10R)-10-O-(tert-Butyldimethylsilyl)-8-hydr-
oxy-1,2:4,5-di-(isopropylidenedioxy)-6-undecene (15). R
0
CHCl3).
f
.48 (hexane:ethyl acetate, 7:3). [a]d � 19.7ꢀ (c 0.28,
(
2S,4S,5S,8S,10R)-8,10-Dihydroxy-1,2:4,5-di-(isopropylid-
enedioxy)-6-undecene (16). Compound 14 (1.87 g,
.21 mmol) was dissolved in 20 mL of anhydrous THF
f
d
ꢀ
1
4
� 30.9 (c 0.5, CHCl ); H NMR: d 1.18 (d, 1H, H ,
3
11
under nitrogen at room temperature. Tetra-
butylammonium ¯uoride (1 M solution in THF, 4.3 mL,
4
tion was quickly washed with water, dried with sodium
sulfate and concentrated under reduced pressure. Rapid
column chromatography (hexane:ethyl acetate, 5:5)
J
6.5 Hz), 1.33 (s, 6H, C(CH ) ), 1.35 (s, 3H,
10,11 3 2
C(CH ) ), 1.39 (s, 9H, C(CH ) ), 1.54� 1.80 (m, 3H, H ,
3
2
3 2
9
.3 mmol) was added. After stirring for 5 min, the solu-
H
J2,3
0
, H ), 1.91 (ddd, 1H, H
0
0
, J =14.0, J 3.0,
0
9
3
3
gem
3 ,4
=6.5 Hz), 3.58 (dd, 1H, H , Jgem=9, J1,2=6.5 Hz),
1
0
3.77 (ddd, 1H, H , J=8, J =10 Hz), 3.90� 4.10 (m, 2H,
4
H , H ), 4.07 (dd, 1H, H , J =5 Hz), 4.20 (m, 1H,
0 0
5
10
1 1 ,2
gave the pure product 16 (1.37 g, 4.1 mmol, 98%). R
ꢀ
H , J =8 Hz) , 4.36 (m, 1H, H , J7,8=5 Hz), 5.61 (dd,
2 2,3 8
f
0
.85 (dichloromethane:methanol, 9:1); [a] 10.6 (c 1.6,
1H, H , Jtrans=15, J5,6=7.0, J6,8=1.0 Hz), 5.85 (dd,
6
d
1
13
CHCl ); H NMR: d 1.13 (d, 3H, H , J10,11=5.5 Hz ),
1H, H7); C NMR: d 21.68, 24.92 (2C), 25.70 (2C),
3
11
1
.26 (s, 3H, C(CH ) ), 1.30 (s, 6H, C(CH ) ), 1.33 (s,
0
26.94, 27.21 (7C, C , C(CH ) ), 36.31, 39.11 (C , C ),
62.44, 66.65, 69.81, 73.82, 77.71, 82.18 (6C, C , C , C ,
1 2 4
C , C , C ), 100.17, 108.62, 108.87 (3C, C(CH ) ),
5 8 10 3 2
3
2
3 2
11
3 2
3
9
3
H, C(CH ) ), 1.42� 1.75 (m, 3H, H , H
, H ), 1.80
3
3
2
9
9
(
ddd, 1H, H =7, J
dd, 1H, H , Jgem=9, J1,2=6.5 Hz), 3.74 (ddd, 1H, H4,
0
, J =12.5, J
0
0
=3 Hz), 3.50
3
gem
2,3
3 ,4
(
J3,4=10, J4,5=7 Hz), 3.88 (dd, 1H, H , J5,6=7.5 Hz),
126.50, 136.05 (C , C ).
6 7
1
5
3
J
4
.80� 4.03 (m, 1H, H ), 4.02 (dd, 1H, H1
0
,
(2S,4S,5S,8R,10R)-1,2:4,5:8,10-Tri-(isopropylidenedioxy)-
6-undecene (19). Compound 19 was prepared from 17 in
95% yield according to the procedure described for the
synthesis of 18. R 0.25 (hexane:ethyl acetate 3:7); [a]
1
0
0
=5.5 Hz), 4.15 (m, 1H, H
.20 (m, 1H, H ), 5.54 (dd, 1H, H , J
0
), 3.50� 4.40 (2H, 2OH),
1
,2
2
=15.5,
C
8
6 trans
13
J5,6=7.5 Hz), 5.77 (dd, 1H, H , J7,8=5.5 Hz);
7
f
d
ꢀ
1
NMR: d 24.28 (C ), 25.93 (C(CH ) ), 27.08 (C(CH ) ),
+5.7 (c 1.2, CHCl ); H NMR: d 1.18 (d, 1H, H ,
1
3 2
3 2
3
11
2
7
1
7.37 (C(CH ) ), 36.68 (C ), 44.74 (C ), 68.12 (C ),
0.53 (C ), 74.24 (C ), 75.10 (C ) 78.36 (C ), 82.20 (C ),
J10,11=7 Hz), 1.35 (s, 3H, C(CH ) ), 1.37 (s, 3H,
C(CH ) ), 1.39 (s, 6H, C(CH ) ), 1.41 (s, 3H, C(CH ) ),
3
2
3
9
10
3 2
1
2
8
4
5
3 2
3 2
3 2
08.95, 109.42 (C(CH ) ), 126.84 (C ), 138.10 (C ). MS
3
1.46 (s, 3H, C(CH ) ), 1.48� 1.70 (m, 3H, H , H
0
, H3),
2
6
7
3 2
9
9
+
(m/z): 315 (M� 15) . Anal. calcd for C H O : C, 61.8;
1.89 (ddd, 1H, H3
3.58 (dd, 1H, H , Jgem=8, J1,2=7.5 Hz), 3.76 (ddd, 1H,
0
, J =14, J =3, J
0
=6.5 Hz),
17
30
6
gem
3,4
2,3
H, 9.2. Found: C, 61.42; H, 9.30.
1

1
056
S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
H , J
4
0
=9, J4,5 =9 Hz), 3.92� 4.05 (m, 2H, H , H ),
1.78 mmol) was dissolved in a mixture of tert-butanol
(26 mL) and 2-methyl-2-butene (6.5 mL). A solution of
sodium chlorite (1 g, 11 mmol) and sodium dihy-
drogenophosphate (1 g, 8.5 mmol) in water (10 mL) was
added dropwise over 10 min at room temperature.
Reaction was monitored by TLC (dichloromethane:
methanol 85:15) and after stirring for 20 min, the sol-
vents were removed under reduced pressure. The crude
mixture was diluted with CH Cl (30 mL) and ethanol
3 ,4
5
10
4
5
5
1
2
7
.08 (dd, 1H, H1
.5 Hz), 4.36 (ddd, 1H, H , J7,8=5, J8,9=2, J8,9 =12 Hz),
.65 (dd, 1H, H , Jtrans=15.5, J5,6=7.5 Hz), 5.79 (dd,
H, H7); C NMR: d 20.08, 22.48, 26.09, 27.36 (2C),
7.58, 30.54 (7C, C , C(CH ) ), 36.73, 38.93 (C , C ),
0.22 (C ), 65.14, 69.11, 74.20, 78.11, 82.38 (5C, C , C ,
, J =6 Hz), 4.21 (m, 1H, H , J =
0 0
1 ,2 2 2,3
0
8
6
1
3
11
3 2
3
9
1
2
4
C , C , C ), 98.94, 108.95, 109.21 (3C, C(CH ) ),
3 2
5
8
10
+
1
27.06, 135.93 (C , C ); MS (m/z): 355 (M� 15) . Anal.
6
7
2
2
calcd for C H O : C, 64.74; H, 9.20. Found: C, 64.47;
20 34 6
(10 mL). The salts were ®ltered o€ under reduced pres-
sure and carefully rinsed with ethanol. Flash column
chromatography on silica gel (dichloromethane:methanol
85:15) a€orded acid 23 (0.43 g, 1.57 mmol, 88%). Rf
0.20 (dichloromethane:methanol 85:15); IR: 1646,
H, 9.36.
(
2S,4S,5S,8S,10R)-1,2,8,10-Tetrahydroxy-4,5-(isopropylid-
enedioxy)-6-undecene (20). The diol 16 (1.36 g,
.12 mmol) was dissolved in THF (5 mL) and a mixture
�
1 1
4
1719 cm ; H NMR: d 1.18 (d, 3H, H , J9,10=6 Hz),
10
of water (1 mL) and acetic acid (9 mL) was added. The
ꢀ
1.41 (s, 3H, C(CH ) ), 1.42 (s, 3H, C(CH ) ), 1.55±1.68
0
(m, 2H, H , H8 ), 2.52 (dd, 1H, H , Jgem=15.5,
8 2
3 2 3 2
solution was heated at 55 C and the reaction monitored
by thin layer chromatography. To avoid total deprotec-
tion of 16, reaction was stopped after 5 h. Solvents were
evaporated under reduced pressure and the residue was
co-evaporated with toluene. Column chromatography
on silica gel (dichloromethane:methanol 95:5) a€orded
the expected product 20 (0.95 g, 1.87 mmol, 80%) to-
gether with totally deprotected compound (0.127 g,
J2,3=4.5 Hz), 2.66 (dd, 1H, H2
0
, J
(m, 3H, H , H , H ), 4.38 (dd, H , J =4.5, J
0
=5 Hz), 3.95� 4.13
2 ,3
=
3
4
9
7 6,7 7,8
13 Hz), 5.68 (dd, 1H, H , Jgem=16, J4,5=6.5 Hz), 5.88
5
(dd, 1H, H6); 13C NMR: d 23.88 (C ), 26.94, 27.07
10
(C(CH ) ), 44.13 (C8), 58.81 (C ), 68.16 (C ), 71.65 (C ),
79.92, 81.50 (C , C ), 109.01 (C(CH ) ), 125.87 (C ),
3
2
2
9
7
3
4
3 2
6
137.97 (C ), 173.59 (C ).
5
1
0
9
.4 mmol, 10%). R 0.14 (dichloromethane:methanol
f
�
1 1
:1); IR: 3357, 1654 cm , H NMR: d 1.16 (d, 3H, H ,
(3S,4S,7S,9R)-7,9-Dihydroxy-3,4-(isopropylidenedioxy)-
5-decenoic acid (22). Compound 22 was prepared in
68% yield from 20 according to the procedure men-
tioned above: Rf 0.15 (dichloromethane:methanol
11
J10,11=6 Hz), 1.40 (s, 6H, C(CH ) ), 1.52� 1.73 (m, 4H,
3 2
H , H , H , H ), 3.42 (dd, 1H, H , J =11.5, J =
0 0
3
3 9 9 1 gem 1,2
6
(
.5 Hz), 3.53 (dd, 1H, H1
0
, J
0
=3.5 Hz), 3.72� 4.50
1 ,2
�
1 1
4H, 4 OH), 3.70� 3.85 (m, 1H, H ), 3.85� 4.03 (m, 2H,
85:15); IR: 3416, 1719, 1649 cm ; H NMR: (400 MHz,
C D O): d 1.16 (d, 3H, H , J9,10=6 Hz), 1.35 (s, 3H,
4
0
=15.5, J =9 Hz),
H , H ), 4.05� 4.18 (m, 1H, H ), 4.29 (ddd, 1H, H , J J
2
0
5
10
8
3
6
10
0
J =6 Hz), 5.65 (dd, 1H, H , J
6
C(CH ) ), 1.36 (s, 3H, C(CH ) ), 1.55 (ddd, 1H, H ,
trans
5,6
3 2 3 2 8
1
3
0 00 0
=14, J =4, J =8 Hz), 1.61 (ddd, 1H, H , J =4,
0
gem 8
5
2
6
8
.89 (dd, 1H, H , J7,8=6 Hz); C NMR: d 23.76 (C11),
J
7
00
J =8 Hz), 2.50 (dd, 1H, H , Jgem=15.5, J2,3=8.5 Hz),
2
7.20, 27.45 (C(CH ) ), 35.38 (C ), 44.33 (C ), 64.89,
9.21, 69.50 (3C, C , C , C ), 66.79 (C ), 77.68 (C ),
3
2
3
9
2.60 (dd, 1H, H2
0
, J
0
=3.5 Hz), 2.81� 2.95 (m, 2H,
2
8
10
1
8
2 ,3
2.08 (C ), 109.06 (C(CH ) ), 126.78 (C ), 138.24 (C ).
7
2OH), 4.00� 4.10 (m, 1H, H ), 4.05 (ddd, 1H, H , J =
9 3 3,4
.5 Hz), 4.16 (dd, 1H, H , J =7 Hz), 4.20 (m, 1H, H ),
4 4,5 7
5
3 2
6
8
(
2S,4S,5S,8R,10R)-1,2,8,10-Tetrahydroxy-4,5-(isopropyli-
5.72 (ddd, 1H, H , J =1.5, J =14.5 Hz, J4,5=8 Hz),
5 5,7 5,6
denedioxy)-6-undecene (21). Compound 21 was prepared
in 78% yield from 17 according to the above mentioned
procedure. R 0.13 (dichloromethane:methanol 9:1); IR:
5.93 (dd, 1H, H , J =6 Hz); 13C NMR: d 24.75, 27.70
6
6,7
(C(CH ) ), 27.80 (C ), 37.59 (C ), 46.78 (C ), 65.14
(C ), 69.53 (C ), 78.53, 82.80 (C , C ), 109.63 (C(CH ) ),
9 7 3 4 3 2
3
2
10
2
8
f
�
1 1
3
415, 1651, 1374 cm ; H NMR: d 1.17 (d, 3H, H ,
126.71 (C ), 140.21 (C ), 172.22 (C ). Anal. calcd for
6 5 1
11
J10,11=6.5 Hz), 1.38 (s, 6H, C(CH ) ), 1.56� 1.82 (m,
3 2
C H O : C, 56.92; H, 8.08. Found: C, 55.75; H, 7.88.
13 22 6
4H, H , H
J1,2=7.5 Hz), 3.64 (dd, 1H, H1
0
, H , H
0
), 3.42 (dd, 1H, H , J =12,
1
3
3
9
9
gem
0
,
J
0
=3.5 Hz),
(3S,4S,7R,9R)-3,4,7,9-Tetrahydroxy-5-decenoic acid, sodium
salt (25). To a solution of carboxylic acid 23 (0.051 g,
0.18 mmol) in THF (2 mL) was added 2 mL of aqueous
1 ,2
3
.70� 4.75 (4H, 4 OH), 3.82� 4.08 (m, 4H, H , H , H ,
0
10 8 7,8 8,9 8,9
2
4
5
H ), 4.29 (ddd, 1H, H , J =5, J
J
=6 Hz), 5.69
ꢀ
(
dd, 1H, H , Jtrans=15.5, J5,6=7.5 Hz), 5.87 (dd, 1H,
6
acetic acid (1:1 v/v). The solution was heated at 60 C
1
3
H7); C NMR: d 24.05 (C ), 27.27, 27.52 (C(CH ) ),
for 36 h. Thin layer chromatography monitoring (tol-
uene 10:AcOEt 35:iPrOH 55:AcOH 2N 20) showed the
presence of less polar compounds resulting from lacto-
nisations. Solvents were removed under reduced pres-
sure. The crude mixture was diluted with 2 mL of water
and aqueous 1N NaOH was added to adjust the pH to
7±8. After lactonic compounds were no longer present
the solution was ®ltered. Lyophilisation a€orded 0.019 g
of the carboxylate 25 (0.07 mmol, 38%). 1H NMR:
(250 MHz, D O): d 1.21 (d, 3H, H , J9,10=6.5 Hz),
11
3 2
3
5.26 (C ), 44.73 (C ), 66.87 (C ), 67.70, 69.51, 71.34
3 9 1
(
(
3C, C , C , C ), 77.75 (C ), 81.85 (C ), 109.13
2 8 10 8 5
C(CH ) ), 126.39 (C ), 137.82 (C ).
3
2
6
7
(
3S,4S,7R,9R)-7,9-Dihydroxy-3,4-(isopropylidenedioxy)-
-decenoic acid (23). To a solution of syn diol 21
0.527 g, 1.82 mmol) in dry methanol (14 mL) was added
at room temperature a solution of sodium periodate
0.584 g, 2.73 mmol) in water (4 mL). The solution
5
(
(
2
10
turned cloudy. After being stirred for 10 min, the white
suspension was ®ltered and rinsed with methanol. Sol-
vents were removed under reduced pressure and the
expected crude aldehyde was used in the next step
without further puri®cation. The crude aldehyde (0.46 g,
1.59±1.85 (m, 2H, H , H8
0
), 2.29 (dd, 1H, H2,
8
Jgem=15.5, J =8.5 Hz), 2.45 (dd, 1H, H2 , J =4.5 Hz),
0 0
2,3
2 ,3
3.86±3.99 (m, 2H, H , H ), 4.07 (dd, 1H, H , J3,4=6,
3
9
4
J4,5=6 Hz), 4.31 (dd, 1H, H , J7,8=13, J6,7=5.5 Hz),
7
5.74 (dd, 1H, H , Jtrans=15.5 Hz), 5.83 (dd, 1H, H6);
5

S. Colle et al. / Bioorg. Med. Chem. 7 (1999) 1049±1057
1057
1
3
C NMR: d 25.30 (C ), 43.45 (C ), 47.56 (C ), 68.35
9 7 3 4 5
phate, 0.15 U glucose-6-phosphate dehydrogenase,
2 mM NADPH, test compounds in DMSO (3.3% ®nal
concentration) and 5±25 mg microsomal protein. After a
1
0
2
8
(C ), 72.73 (C ), 74.88 (C ), 77.82 (C ), 132.76 (C ),
1
2
38.35 (C ), 184.33 (C ); MS (m/z): ES negative mode:
6 1
33 (M� 23); ES positive mode: 279 (M+Na) , 257
+
ꢀ
10 min incubation at 30 C, the reaction was initiated by
+
14
(
M+H) , 242.
the addition of 0.45 mCi dl-3-[glutaryl-3- C]-HMG
CoA (DuPont-NEN, NEC-642) and conducted for
20 min. The assay was stopped by the addition of 30 mL
of 6 N HCl and vials were centrifuged to pellet protein.
(
3S,4S,7S,9R)-3,4,7,9-Tetrahydroxy-5-decenoic acid, sodium
salt (24). Compound 24 was prepared from 22 (0.1 g,
.36 mmol) in 22% yield (0.02 g, 0.08 mmol) according
to the above mentioned procedure. IR: 3346, 1664,
14
14
0
C-Mevalonic acid and
separated from the substrate,
C-mevalonolactone were
C-HMG-CoA, by
14
�
1 1
1
3
2
579, 1443 cm ; H NMR: (250 MHz, D O): d 1.22 (d,
HPLC on an Alltech Lichrosphere C18 column (5 u,
4.6Â250) using 75% water containing 0.5% phosphoric
acid and 25% methanol containing 0.5% phosphoric
acid. Radioactive peaks were detected using an INUS
B-Ram detector.
2
0
.28 (dd, 1H, H , Jgem=15.5, J2,3=9 Hz), 2.45 (dd, 1H,
H, H , J =7 Hz), 1.69 (dd, 2H, H , H
0
, J J =6.5Hz),
1
0
9,10
8
8
2
H2 , J =4.5 Hz), 3.86±3.99 (m, 2H, H , H ), 4.05 (dd,
1H, H , J3,4=12.5, J4,5=6.5 Hz), 4.32 (dd, 1H, H7,
J7,8=12.5, J6,7=6 Hz), 5.72 (dd, 1H, H , Jtrans=16 Hz),
5
4
1
0 0
2 ,3 3 9
4
5
.84 (dd, 1H, H6); 13C NMR: d 25.27 (C ), 43.63 (C ),
10
2
7.55 (C ), 67.23 (C ), 71.42 (C ), 74.74 (C ), 77.63 (C ),
31.74 (C ), 138.63 (C ), 182.86 (C ); MS (m/z): ES
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8
9
7
3
4
5
6
1
1. Chapleur, Y. In Recent Progress in the Chemistry of Anti-
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+
+
(
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3
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ꢀ
growth chamber at 26 C under continuous ¯uorescent
�
2
� 1
.
and incandescent light sources providing 300 mE m
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9:1) and a 50 mL aliquot combined with 2 mL of cul-
s
(
ture medium in a multi-well plate (Falcon, model 3043).
A single colony consisting of 4 fronds was added to each
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time visual injury ratings were obtained. By 7 days
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9
1
1
1
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ꢀ
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