The chiron approach to pironetins: Synthesis of the δ-lactonic fragment and modified building blocks from D-glucal

Article abstract of DOI:10.1080/07328300600735090

The synthesis of the δ-lactonic fragment of pironetin (1), a natural product with outstanding antimitotic properties, is reported. The synthesis was achieved from commercially available tri-O-acetyl-D-glucal (6) that was employed as starting material for

Full text of DOI:10.1080/07328300600735090

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The Chiron Approach to Pironetins: Synthesis
of the δ‐Lactonic Fragment and Modified
Building Blocks from D‐Glucal
Francisco Sarabia a , Miguel García‐Castro a , Samy Chammaa a & Antonio
Sánchez‐Ruiz a
a Department of Biochemistry, Molecular Biology and Organic Chemistry,
Faculty of Sciences, University of Malaga, Malaga, Spain
Published online: 17 Aug 2006.
To cite this article: Francisco Sarabia , Miguel García‐Castro , Samy Chammaa & Antonio Sánchez‐Ruiz (2006):
The Chiron Approach to Pironetins: Synthesis of the δ‐Lactonic Fragment and Modified Building Blocks from
D‐Glucal, Journal of Carbohydrate Chemistry, 25:2-3, 267-280
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Journal of Carbohydrate Chemistry, 25:267–280, 2006
Copyright # Taylor & Francis Group, LLC
ISSN: 0732-8303 print 1532-2327 online
DOI: 10.1080/07328300600735090
The Chiron Approach to
Pironetins: Synthesis of the
d-Lactonic Fragment and
Modified Building Blocks
from D-Glucal
´
Francisco Sarabia, Miguel Garcıa-Castro, Samy Chammaa,
´
and Antonio Sanchez-Ruiz
Department of Biochemistry, Molecular Biology and Organic Chemistry,
Faculty of Sciences, University of Malaga, Malaga, Spain
The synthesis of the d-lactonic fragment of pironetin (1), a natural product with out-
standing antimitotic properties, is reported. The synthesis was achieved from com-
mercially available tri-O-acetyl-D-glucal (6) that was employed as starting material
for the preparation of ethyl ketone 4, through a synthetic sequence that included a
Ferrier rearrangement of 6 and suitable functional group manipulations of the resulting
O-glycoside 7 to obtain the 4-ethyl glycoside 11, together with a series of 4-C-alkyl
modified derivatives. The completion of the synthesis of 4 was performed via chain
elongation at C-6 by the introduction of a nitrile group and subsequent reduction,
nucleophilic attack with ethyl magnesium bromide, and, finally, oxidation of the result-
ing alcohol 20.
Keywords Pironetin, Ferrier rearrangement, Antimitotic agents, Chiron approach
INTRODUCTION
Isolated by two independent research groups from culture broths of
Streptomyces sp. NK10958[1] and Streptomyces prunicolor PA-48153,[2] pirone-
tin (1, PA-48153C) belongs to the privileged class of natural products with
Received November 22, 2005; accepted March 2, 2006.
Address correspondence to Francisco Sarabia, Department of Biochemistry, Molecular
Biology and Organic Chemistry, Faculty of Sciences, University of Malaga, 29071,
Malaga, Spain. E-mail: frsarabia@uma.es
267

F. Sarabia et al.
268
antimitotic properties in virtue of its capacity of interacting with tubulines,
producing inhibition of their assembly to microtubules.[3] Initially, pironetin
was identified as a plant growth regulator, inducing shortening of the plant
height of rice,[4] and later, as a potent immunosuppressant,[5] with a potency
similar to the well-known immunosuppressive agents cyclosporine A and
FK-506.[6] Particularly intriguing and stimulating was, however, the disclos-
ure of its antitumoral properties, by induction of microtubule disassembly
in cells type 3Y1 and inhibition glutamate-induced tubulin assembly in a
10-mM range concentration, which elicited a great deal of interest in the
scientific community.[7] The result of this biological action was the arrest of
cellular growth at the M-phase and, like other microtubule inhibitors, pirone-
tin seems to induce the phosphorylation of Bcl-2, which triggers apoptosis.[8]
The binding mode of pironetin to tubuline was initially surmised in a vinblas-
tine-like mode due to its capacity of inhibiting tubulin binding of vinblastine.
In fact, although effective doses of pironetin were slightly superior than those
of vinblastine, against HL-60 cells, its Kd value was 10-fold lower, as an indi-
cator of a greater affinity for tubulin than that of vinblastine. In contrast to
these exciting findings, recent biological assays[9] have revealed that the
binding site is located on the surface of a-tubulin, the Lys352 residue being
responsible for a covalent binding with pironetin through a Michael addition
of the terminal amino group of this amino acid to the a,b-unsaturated
d-lactone moeity.[10] In fact, structure-activity relationship studies of pironetin
analog have reflected the importance of the 2-pyranone ring system as well as
the hydroxyl group at C-7 in its antitumor properties.[11] All these outstanding
biological properties exhibited by pironetin renders it as an enticing synthetic
endeavor. Thus, several total syntheses of pironetin (1) have been reported,
including linear asymmetric[12] and chiron approaches[13] to total syntheses.
Despite its relatively small size, pironetin offers both a challenge and an oppor-
tunity for the development of a new class of novel anticancer agents, which
captured our attention and encouraged us to initiate a program directed
toward its total synthesis, according to a synthetic strategy capable of generat-
ing analogs by a convergent and flexible route. To this end, we devised, in ret-
rosynthetic terms, a dissection at the C-8/C-9 bond of compound 2, obtained
after the appropiate functional group transformations of 1, via a retro-aldol
reaction, to produce aldehyde 3 and ketone 4 as advanced precursors of 1.
The synthesis of the d-lactonic fragment, contained in 4, was planned accord-
ing to an appropiate chain elongation method at C-6 and the incorporation of
an ethyl group at C-4, leading to glycoside 5 as a potential and valuable pre-
cursor, whose synthesis could be achieved via a Ferrier rearrangement from
the commercially available tri-O-acetyl-D-glucal (6). Herein we wish to
report the synthesis of the d-lactonic derivative 4 together with 4-C-alkyl
modified analog according to the delineated retrosynthetic analysis depicted
in Scheme 1.

Chiron Approach to Pironetins
269
Scheme 1: Structure of pironetin (1) and retrosynthetic analysis.
RESULTS AND DISCUSSION
The synthesis of the subtarget molecule 4 commenced with the Ferrier
rearrangement[14] of commercially available tri-O-acetyl-D-glucal (6) (Tri-O-
acetyl-D-Glucal (6) was purchased from Aldrich.) by reaction with isopropyl
alcohol in the presence of boron trifluoride to afford O-glycoside 7 in 95%
yield as a 9:1 mixture of a:b anomers. Methanolysis of 7 by reaction with
sodium methoxide, followed by selective silylation of the resulting diol 5,
provided silyl ether 8, which was treated with methanesulphonyl chloride in
the presence of triethyl amine to obtain the mesylate 9, a product that was con-
sidered very suitable to attempt the introduction of the ethyl group contained
at C-4 of the d-lactonic fragment. In a first attempt, 9 was reacted with
Et2CuLi,[15] a reagent that was generated in situ by treatment of two equiva-
lents of ethyllithium (Ethyllithium was purchased from Aldrich as a 0.5-M
solution in benzene/cyclohexane 90/10.) with CuI. However, the result was
not satisfactory because of the recovery of starting material. In a similar
reaction, however, 9 reacted smoothly with Me2CuLi to obtain the 4-methyl
derivative 10, albeit in a low 30% yield. Gratifyingly, the treatment of 9 with
either EtMgBr/CuCN[16] or Et2Zn/CuCN[17] furnished the desired 4-ethyl
derivative 11 in an 80% yield for both cases. Combining NMR spectroscopic
methods with theoretical calculations (Theoretical calculations of both confor-
mers A and B, corresponding to the a-anomer, were performed with
HyperChem 5.0, preoptimizing with MMþ as the force field with a gradient
˚
limit of 0.001 kcal/(A.mol), followed by full optimization with AM1, obtaining
energy values of –115927,88 and –115925,83 kcal/mol for conformers A and

F. Sarabia et al.
270
B respectively. Coupling constants were determined by the Karplus equation
J ¼ 4.22 – 0.5.cos a þ 4.5.cos 2a. Similar conformational studies were under-
taken for the b-anomer of 11 and for the a/b anomers of the 4-(S) diastereoi-
somer, but in any case the resulting calculated J were in accordance with the
experimental value of J4,5 for compound 11.[18]) let us to confirm the 4-(R) con-
figuration of product 11 as a result of a SN2 process (Sch. 2). This successful
result prompted us to prepare a family of 4-C-alkyl analog, which could be of
interest for future biological evaluations, taking into account the apparent
key importance of the 2-pyranone system in their binding to tubulines. In
addition, this synthesis represents useful convergent approaches, particularly
to related natural products that contain the same 4-C-alkyl-a,b-unsaturated
d-lactonic fragment.[19] Then, exposure of 9 to the action of different alkyl mag-
nesium bromides in the presence of CuCN afforded the corresponding 4-C-alkyl
derivatives 10–14 in a wide range of yields (20–80%) (Sch. 2).
For the completion of the synthesis of the coveted ethyl ketone 4, compound
11 was desilylated by treatment with TBAF, and the resulting alcohol 15 was
activated as its O-mesyl derivative 16, which was subjected to the action of
NaCN in refluxing DMF[20] to afford, after 24 h, the nitrile derivative 17 in a
95% yield. Our strategy toward 4 was set forth on the notion that nitrile 17
might be amenable to a nucleophilic attack of ethyl magnesium bromide to
provide 4 directly. However, our unrelenting efforts to effect this direct trans-
formation were thwarted by the fragile nature of the glycosidic bond, which
was highlighted during the acidic work-up of the imine intermediate 18, occur-
ring the cleavage of such glycosidic bond. Different attempts to unmask the
ketone group from imine 18 under a wide variety of milder acidic conditions[21]
were similarly unsuccessful. These results forced us to overcome the lability of
the glycosidic bond by a sequence entailing reduction of nitrile 17, nucleophilic
Scheme 2: Synthesis of 4-C-alkyl glycoside derivatives 10–14.

Chiron Approach to Pironetins
271
attack of the resulting aldehyde, and subsequent oxidation. Thus, reduction to
aldehyde 19 of nitrile 17 was accomplished by treatment with DIBAL-H in 60%
yield. The resulting aldehyde 19 was then reacted with ethyl magnesium
bromide to provide alcohol 20 in an 85% yield as a 1:1 mixture of diastereo-
isomers, which was finally transformed into the ketone 4 by reaction with
TPAP[22] in 88% yield (Sch. 3).
Having successfully synthesized the subtarget compound 4 in an efficient
manner, our next objective will be to achieve the total synthesis of pironetin
1, according to the retrosynthetic blueprint showed in Scheme 1, which
might offer us, in addition, the possibility of providing analog by modification
at C-4 and at C-7 positions for future biological evaluations as antimitotic
agents. These synthetic efforts are currently in progress and will be reported
in due course.
EXPERIMENTAL
General Techniques
All reactions were carried out under an argon atmosphere with dry, freshly
distilled solvents under anhydrous conditions, unless otherwise noted. Tetra-
hydrofuran and diethyl ether were distilled from sodium benzophenone and
methylene chloride and methanol from calcium hydride. Yields refer to chro-
matographically and spectroscopically (1H NMR) homogeneous materials,
unless otherwise stated. All solutions used in work-up procedures were satu-
rated unless otherwise noted. All reagents were purchased at highest commer-
cial quality and used without further purification unless otherwise stated.
All reactions were monitored by thin-layer chromatography carried out on
0.25 mm E. Merck silica gel plates (60F-254) using UV light as visualizing
agent and 7% ethanolic phosphomolybdic acid or p-anisaldehyde solution and
Scheme 3: Synthesis of ketone 4.

F. Sarabia et al.
272
heat as developing agents. E. Merck silica gel (60, particle size 0.040–
0.063 mm) was used for flash column chromatography. Preparative thin-layer
chromatography (PTLC) separations were carried out on 0.25, 0.50, or 1 mm
E. Merck silica gel plates (60F-254).
NMR spectra were recorded on a Bruker Avance-400 instrument and cali-
brated using residual undeuterated solvent as an internal reference. The fol-
lowing abbreviations were used to explain the multiplicities: s, singlet; d,
doublet; t, triplet; q, quartet; m, multiplet; band, several overlapping signals;
1
b, broad. H NMR assignments were undertaken based on two-dimensional
COSY experiments (cosygp experiment). Optical rotations were recorded on a
Perkin-Elmer 241 polarimeter. High-resolution mass spectra (HRMS) were
recorded on a Kratos MS 80 RFA mass spectrometer under fast atom bombard-
ment (FAB) conditions.
Isopropyl
4,6-Di-O-acetyl-2,3-dideoxy-a-D-erythro-hex-2-enopyrano-
side (7). To a solution of tri-O-acetyl D-glucal (6) (10 g, 36.76 mmol, 1.0
i
equiv) in CH2Cl2 (70 mL) was added PrOH (14.1 mL, 183.8 mmol, 5.0 equiv)
.
and BF3 OEt2 (9.21 mL of a 48% solution, 47.79 mmol, 1.3 equiv) at 08C.
After stirring for 3 h, the reaction mixture was warmed to rt, treated with
a saturated aqueous NaHCO3 solution, and diluted with CH2Cl2 (30 mL).
After separation of both layers, the aqueous phase was extracted with
CH2Cl2 and the combined organic solution was washed with water and brine,
dried (MgSO4), filtered, and concentrated under reduced pressure. The result-
ing crude product was subjected to purification by flash column chromato-
graphy (silica gel, 30% EtOAc in hexanes) to obtain O-glycoside 7 as an
unseparable 9:1 mixture of a:b anomers (9.51 g, 95%): Yellow oil; Rf ¼ 0.55
1
(silica gel, 30% EtOAc in hexanes); H NMR (400 MHz, CDCl3) d (ppm): 1.12
(d, J ¼ 5.9 Hz, 3H), 1.19 (d, J ¼ 6.4 Hz, 3H), 2.02 (s, 3H), 2.04 (s, 3H), 3.93
(ddd, J ¼ 5.4, 5.9, 6.4 Hz, 1H), 4.06-4.12 (m, 1H), 4.12–4.22 (m, 2H), 5.07 (bs,
1H), 5.24 (dd, J ¼ 1.6, 9.7 Hz, 1H), 5.75 (dt, J ¼ 2.1, 10.2 Hz, 1H), 5.81 (bd,
J ¼ 10.2 Hz, 1H); 13C NMR (100 MHz, CDCl3) d (ppm): 20.7, 21.1, 23.6, 62.8,
64.3, 70.4, 71.2, 92.5, 127.0, 133.2, 170.3.
Isopropyl 2,3-Dideoxy-a-D-erythro-hex-2-enopyranoside (5). A solution
of O-glycoside 7(9.51 g, 34.92 mmol, 1.0 equiv) in MeOH (60 mL) was treated
with NaOMe (377 mg, 6.98 mmol, 0.2 equiv) at 258C. After stirring for 1 h,
the crude mixture was concentrated under reduced pressure, and the resulting
crude product was purified by flash column chromatography (silica gel, 90%
EtOAc in hexanes ! EtOAc) to obtain diol 5 (6.51 g, 99%) as a white solid:
1
Rf ¼ 0.13 (silica gel, 50% EtOAc in hexanes); H NMR (400 MHz, CDCl3) d
(ppm): 1.12 (d, J ¼ 5.9 Hz, 3H), 1.19 (d, J ¼ 6.4 Hz, 3H), 3.68 (dt, J ¼ 4.5,
9.0 Hz, 1H), 3.75–3.86 (m, 2H), 3.89 (sep, 1H), 4.15 (bd, J ¼ 4.5 Hz, 1H), 5.04
(bs, 1H), 5.68 (dt, J ¼ 2.1, 10.2 Hz, 1H), 5.88 (bd, J ¼ 10.2 Hz, 1H); 13C NMR
(100 MHz, CDCl3) d (ppm): 21.8, 23.6, 62.8, 64.3, 70.4, 71.2, 92.5, 127.0, 133.2.

Chiron Approach to Pironetins
273
Isopropyl
6-O-tertButyldiphenylsilyl-2,3-dideoxy-a-D-erythro-hex-2-
enopyranoside (8). To a solution of diol 5(6.51 g, 34.57 mmol, 1.0 equiv) in
DMF (40 mL) was added TBDPSCl (10.62 mL, 41.48 mmol, 1.2 equiv) and imida-
zole (3.06 g, 44.94 mmol, 1.3 equiv) at 08C. After stirring for 2 h at this tempera-
ture, the reaction mixture was quenched by addition of MeOH, diluted with Et2O
(50 mL), and followed by addition of a saturated aqueous NH4Cl solution. After
separation of both layers, the aqueous phase was extracted with ether and the
combined organic solution was washed with water and brine. The organic
solution was then dried (MgSO4) and filtered, and, after concentration under
reduced pressure, the resulting crude product was subjected to purification by
flash column chromatography (silica gel, 20% EtOAc in hexanes) to obtain silyl
ether 8 (13.26 g, 90%) as a colorless oil: Rf ¼ 0.31 (silica gel, 20% EtOAc in
hexanes); 1H NMR (400 MHz, CDCl3) d (ppm): 1.07 (s, 9H), 1.15 (d, J ¼ 5.9 Hz,
3H), 1.16 (d, J ¼ 6.4 Hz, 3H), 3.82-4.06 (m, 4H), 4.22 (dd, J ¼ 1.6, 8.1 Hz, 1H),
5.04 (bs, 1H), 5.72 (dt, J ¼ 2.1, 10.2 Hz, 1H), 5.94 (bd, J ¼ 10.2 Hz, 1H), 7.34–
7.48 (m, 6H), 7.61-7.73 (m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.1,
25.2, 26.7, 66.2, 68.2, 107.0, 110.1, 127.7, 129.7, 132.8, 135.4, 141.9.
Isopropyl 6-O-tertButyldiphenylsilyl-2,3-dideoxy-4-O-methylsulpho-
nyl-a-D-erythro-hex-2-enopyranoside (9). To a solution of silyl ether 8
(7.2 g, 16.89 mmol, 1.0 equiv) in CH2Cl2 (50 mL) was added TEA (4.69 mL,
33.78 mmol, 2.0 equiv), followed by a dropwise addition of methanesulphonyl
chloride (1.94 mL, 25.34 mmol, 1.5 equiv) at 08C. After stirring for 1.5 h, the
reaction mixture was warmed at rt, and a saturated aqueous NH4Cl solution
and ether (50 mL) were added. After decantation of both layers, the aqueous
phase was extracted with ether and the combined organic solution was
washed with water and brine, dried (MgSO4), filtered, and concentrated
under reduced pressure. The resulting crude product was purified by flash
column chromatography (silica gel, 20% EtOAc in hexanes) to obtain
compound 9 (7.2 g, 84%) as a colorless oil: Rf ¼ 0.35 (silica gel, 20% EtOAc in
hexanes); 1H NMR (400 MHz, CDCl3):
d (ppm): 1.07 (s, 9H), 1.16
(d, J ¼ 5.9 Hz, 3H), 1.17 (d, J ¼ 6.4 Hz, 3H), 2.92 (s, 3H), 3.87 (d, J ¼ 3.2 Hz,
2H), 3.92–4.02 (m, 2H), 5.14 (bs, 1H), 5.29 (dd, J ¼ 1.6, 9.7 Hz, 1H), 5.86 (dt,
J ¼ 2.1, 10.2 Hz, 1H), 6.08 (bd, J ¼ 10.2 Hz, 1H), 7.34–7.46 (m, 6H), 7.65–
7.74 (m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.1, 23.5, 25.2, 26.6, 39.0,
62.3, 70.3, 112.7, 127.5, 129.6, 135.5, 135.8, 140.6. FAB HRMS (NBA) m/e
527.1893, M þ Naþ calcd for C26H36O6SSi 527.1899.
Isopropyl
4-C-Alkyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-a-D-
threo-hex-2-enopyranosides 10-14. Procedure A: To a suspension of
CuCN (89 mg, 0.993 mmol, 2.5 equiv) in THF (7 mL) was added the Grignard
reagent (2.00 mmol, 5.0 equiv) at 2408C and stirred for 5 min, prior to the
addition of a solution of mesylate 9 (200 mg, 0.397 mmol, 1.0 equiv) in THF
(4 mL) at 2408C. The reaction mixture was left to reach 2188C, and stirred

F. Sarabia et al.
274
until depletion of starting material (ca 20 min). Then the reaction mixture was
treated with MeOH and diluted with ether (15 mL) and a saturated aqueous
NH4Cl solution added. After decantation of both layers, the aqueous phase
was separated and extracted with ether and the combined organic extracts
were sequentially washed with water and brine, dried (MgSO4), filtered, and
concentrated under reduced pressure. The obtained crude product was sub-
jected to purification by flash column chromatography (silica gel, 5% EtOAc
in hexanes) to obtain the corresponding 4-C-alkyl glycoside. Procedure B:
To a suspension of CuCN (1.775 g, 19.83 mmol, 2.0 equiv) in THF (25 mL)
was added Et2Zn (19.8 mL of a 1.0 M solution, 19.83 mmol, 2.0 equiv) at 08C.
After stirring for 10 min, a solution of mesylate 9 (5.0 g, 9.91 mmol, 1.0 equiv)
in THF (35 mL) was dropwise added. The reaction mixture was stirred for
20 min at 08C, and then it was quenched with MeOH. Dilution with ether
(30 mL) was followed by addition of a saturated aqueous NH4Cl solution, the
phases were separated, and the aqueous layer was extracted with ether. The
combined organic solutions were washed with water and brine, dried
(MgSO4), filtered, and concentrated under reduced pressure. Purification by
flash column chromatography (silica gel, 5% EtOAc in hexanes) furnished
compound 11 (3.48 g, 80%) as a colorless oil.
Isopropyl
4-C-Methyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-a-D-
threo-hex-2-enopyranoside (10). (Colorless oil, 30%): Rf ¼ 0.79 (silica gel,
20% EtOAc in hexanes); 1H NMR (400 MHz, CDCl3) d (ppm): 0.95
(d, J ¼ 7 Hz, 3H), 1.07 (s, 9H), 1.15 (d, J ¼ 6.4 Hz, 3H), 1.24 (d, J ¼ 6.4 Hz,
3H), 1.90–1.97 (m, 1H), 3.71(dd, J ¼ 5.9, 10.2 Hz, 1H), 3.79 (dd, J ¼ 5.9,
10.2 Hz, 1H), 3.98 (dt, J ¼ 12.4, 5.9 Hz, 1H), 4.23–4.31 (m, 1H) 4.22–4.32
(m, 1H), 4.88 (bs, 1H), 5.70–5.79 (m, 1H), 5.86 (bd, J ¼ 10.2 Hz, 1H), 7.34–
7.46 (m, 6H), 7.67–7.73 (m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.2,
20.5, 23.6, 26.8, 30.6, 45.6, 66.5, 68.7, 96.7, 99.1, 127.7, 129.6, 133.6, 135.7.
Isopropyl
4-C-Ethyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-a-D-
threo-hex-2-enopyranoside (11). (Colorless oil, 80%): Rf ¼ 0.75 (silica gel,
20% EtOAc in hexanes); 1H NMR (400 MHz, CDCl3): d (ppm): 0.96
(t, J ¼ 7.5 Hz, 3H), 1.09 (s, 9H), 1.17 (d, J ¼ 6.4 Hz, 3H), 1.24 (d, J ¼ 6.4 Hz,
3H), 1.38–1.58 (m, 2H), 1.90–1.97 (m, 1H), 3.71 (dd, J ¼ 5.9, 10.2 Hz, 1H),
3.79 (dd, J ¼ 5.9, 10.2 Hz, 1H), 3.98 (dt, J ¼ 12.4, 5.9 Hz, 1H), 4.23–4.31
(m, 1H), 4.83 (bs, 1H), 5.78 (d, J ¼ 10.2 Hz, 1H), 5.83 (dd, J ¼ 10.2, 3.4 Hz,
1H), 7.36–7.46 (m, 6H), 7.69–7.74 (m, 4H); 13C NMR (100 MHz, CDCl3) d
(ppm): 11.3, 19.2, 21.5, 23.5, 26.4, 26.8, 41.0, 66.5, 68.7, 69.3, 97.6, 125.7,
127.4, 127.6, 129.6, 133.5, 135.6.
Isopropyl
4-C-Allyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-a-D-
threo-hex-2-enopyranoside (12). (Colorless oil, 79%): Rf ¼ 0.87 (silica gel,
1
20% EtOAc in hexanes); H NMR (400 MHz, CDCl3) d (ppm): 1.08 (s, 9H),

Chiron Approach to Pironetins
275
1.15 (d, J ¼ 6.4 Hz, 3H), 1.23 (d, J ¼ 6.4 Hz, 3H), 2.08–2.26 (m, 3H), 3.71 (dd,
J ¼ 5.9, 10.2 Hz, 1H), 3.78 (dd, J ¼ 5.9, 10.2 Hz, 1H), 3.96 (dt, J ¼ 12.4,
5.9 Hz, 1H), 4.25–4.30 (m, 1H), 4.82 (bs, 1H), 5.04 (d, J ¼ 10.2 Hz, 1H), 5.07
(d, J ¼ 10.2 Hz, 1H), 5.75–5.85 (m, 3H), 7.35–7.46 (m, 6H), 7.67–7.73
(m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.2, 21.6, 23.5, 26.8, 37.7,
39.1, 66.4, 68.9, 69.4, 97.1, 116.4, 126.1, 127.1, 127.6, 129.6, 133.5, 135.6,
135.7, 136.1.
Isopropyl
threo-hex-2-enopyranoside (13). (Colorless oil, 77%): Rf ¼ 0.79 (silica gel,
20% EtOAc in hexanes); 1H NMR (400 MHz, CDCl3)
(ppm): 0.87
4-C-Isobutyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-a-D-
d
(d, J ¼ 6.4 Hz, 6H), 1.07 (s, 9H), 1.08–1.11 (m, 5H), 1.18–1.23 (m, 4H), 2.10
(d, J ¼ 7.0 Hz, 1H), 3.64–3.79 (m, 2H), 3.82–3.97 (m, 1H), 4.03 (dt, J ¼ 5.9,
12.4 Hz, 1H), 4.76 (bs, 1H), 5.75 (d, J ¼ 10.2 Hz, 1H), 6.03-6.11 (m, 1H), 7.32-
7.47 (m, 6H), 7.63 2 7.73 (m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.2,
23.5, 26.4, 26.4, 26.8, 39.6, 66.5, 68.7, 69.3, 97.6, 125.7, 127.4, 127.6, 129.6,
133.5, 135.6.
Isopropyl 4-C-Isopropenyl-6-O-tertbutyldiphenylsilyl-2,3,4-trideoxy-
a-D-threo-hex-2-enopyranoside (14). (Colorless oil, 20%): Rf ¼ 0.81 (silica
gel, 20% EtOAc in hexanes); 1H NMR (400 MHz, CDCl3) d (ppm): 1.04
(s, 3H), 1.06 (s, 9H), 1.15 (d, J ¼ 5.9 Hz, 3H), 1.22 (d, J ¼ 6.4 Hz, 3H), 2.67
(bs, 1H), 3.62–3.70 (m, 1H), 3.78 (dd, J ¼ 5.9, 10.2 Hz, 1H), 3.94 (dt,
J ¼ 12.4, 6.4 Hz, 1H), 4.23–4.30 (m, 1H), 4.79 (bs, 1H), 4.84 (bs, 2H), 5.63–
5.70 (m, 1H), 5.91 (d, J ¼ 10.2 Hz, 1H), 7.33 2 7.45 (m, 6H), 7.65–7.72
(m, 4H); 13C NMR (100 MHz, CDCl3) d (ppm): 19.2, 21.4, 23.5, 26.8, 47.7,
66.4, 68.5, 68.8, 96.9, 113.2, 125.5, 126.5, 127.6, 129.6, 133.5, 135.6, 144.7.
Isopropyl
4-C-Ethyl-2,3,4-trideoxy-a-D-threo-hex-2-enopyranoside
(15). A solution of compound 11 (3.48 g, 7.94 mmol, 1.0 equiv) in THF (40 mL)
was treated with TBAF (11.9 mL of a 1.0 M solution in THF, 11.91 mmol, 1.5
equiv) at 258C. After stirring for 20 min, a saturated aqueous NH4Cl solution
and ether (30 mL) were added. After decantation of both layers, the aqueous
phase was extracted with ether and the combined organic solution was
washed with water and brine, dried (MgSO4), filtered, and concentrated
under reduced pressure. The resulting crude alcohol was subjected to purifi-
cation by flash column chromatography (silica gel, 30% EtOAc in hexanes), pro-
viding pure alcohol 15 (2.32 g, 88%) as a colorless oil: Rf ¼ 0.25 (silica gel, 20%
1
EtOAc in hexanes); H NMR (400 MHz, CDCl3) d (ppm): 0.96 (t, J ¼ 7.5 Hz,
3H), 1.17 (d, J ¼ 5.9 Hz, 3H), 1.24 (d, J ¼ 6.4 Hz, 3H), 1.39–.56 (m, 2H),
1.90–1.98 (m, 1H), 3.60 (dd, J ¼ 6.4, 11.3 Hz, 1H), 3.72 (dd, J ¼ 3.2, 11.3 Hz,
1H), 3.98 (dt, J ¼ 5.9, 12.4 Hz, 1H), 4.26–4.32 (m, 1H), 4.85 (bs, 1H), 5.63
(d, J ¼ 10.2 Hz, 1H), 5.84–5.90 (m, 1H); 13C NMR (100 MHz, CDCl3) d (ppm):
11.4, 21.5, 23.5, 26.8, 41.0, 65.2, 68.6, 68.9, 97.5, 124.5, 128.7.

F. Sarabia et al.
276
Isopropyl
4-C-Ethyl-6-O-methylsulphonyl-2,3,4-trideoxy-a-D-threo-
hex-2-enopyranoside (16). To a solution of alcohol 15 (1.0 g, 5.0 mmol, 1.0
equiv) in CH2Cl2 (20 mL) was added TEA (0.68 mL, 10 mmol, 2.0 equiv) and
methanesulphonyl chloride (0.58 mL, 7.5 mmol, 1.5 equiv) at 08C. After
stirring for 45 min at this temperature, the reaction mixture was diluted
with ether (35 mL), and a saturated aqueous NH4Cl solution was added. Separ-
ation of both layers was followed by extractions of the aqueous phase with
ether. The combined organic solution was washed with water and brine,
dried (MgSO4), filtered, and concentrated under reduced pressure. Purification
of the resulting crude product by flash column chromatography (silica gel, 40%
EtOAc in hexanes) provided mesylate 16 (1.34 g, 96%) as a colorless oil:
Rf ¼ 0.34 (silica gel, 40% ether in hexanes); 1H NMR (400 MHz, CDCl3) d
(ppm): 0.96 (t, J ¼ 7.5 Hz, 3H), 1.17 (d, J ¼ 5.9 Hz, 3H), 1.23 (d, J ¼ 6.4 Hz,
3H), 1.37–1.56 (m, 2H), 1.91–1.98 (m, 1H), 3.05 (s, 3H), 3.97 (dt, J ¼ 6.4,
12.4 Hz, 1H), 4.24 (dd, J ¼ 6.4, 10.7 Hz, 1H), 4.29 (dd, J ¼ 3.8, 10.7 Hz, 1H),
4.44–4.49 (m, 1H), 4.83 (bs, 1H), 5.62 (d, J ¼ 10.7 Hz, 1H), 5.89–5.95
(m, 1H); 13C NMR (100 MHz, CDCl3) d (ppm): 11.3, 21.5, 23.5, 26.4, 37.7,
40.9, 66.7, 69.0, 71.2, 97.5, 122.6, 129.9. FAB HRMS (NBA) m/e 301.1089,
M þ Naþ calcd for C12H22O5S 301.1086.
2-((2S,3R,6S)-3-Ethyl-3,6-dihydro-6-isopropoxy-2H-pyran-2-yl)-aceto-
nitrile (17). To a solution of mesylate 16 (1.34 g, 4.80 mmol, 1.0 equiv) in DMF
(50 mL) was added sodium cyanide (1.40 g, 28.8 mmol, 6.0 equiv) and the result-
ing suspension was heated at 658C. After stirring for 24 h, the reaction mixture
was warmed at rt, diluted with ether (50 mL), and washed with water. Both
layers were separated, the aqueous phase was extracted with ether, and the
combined organic extracts were washed with water and brine, dried
(MgSO4), filtered, and concentrated under reduced pressure. The resulting
crude product was purified by flash column chromatography (silica gel, 20%
EtOAc in hexanes) to afford nitrile 17 (957 mg, 95%) as a colorless oil:
1
Rf ¼ 0.50 (silica gel, 20% EtOAc in hexanes); H NMR (400 MHz, CDCl3) d
(ppm): 0.96 (t, J ¼ 7.5 Hz, 3H), 1.17 (d, J ¼ 5.9 Hz, 3H), 1.23 (d, J ¼ 6.4 Hz,
3H), 1.41–1.59 (m, 2H), 1.89–1.96 (m, 1H), 2.58 (dd, J ¼ 6.4, 10.7 Hz, 2H),
3.97 (dt, J ¼ 6.4, 12.4 Hz, 1H), 4.40–4.46 (m, 1H), 4.83 (bs, 1H), 5.65
(d, J ¼ 10.7 Hz, 1H), 5.92 (dd, J ¼ 4.3, 10.2 Hz, 1H); 13C NMR (100 MHz,
CDCl3) d (ppm): 11.4, 21.5, 23.4, 26.5, 40.8, 64.0, 69.1, 97.7, 117.1, 124.6, 129.6.
2-((2S,3R,6S)-3-Ethyl-3,6-dihydro-6-isopropoxy-2H-pyran-2-yl)-acet-
aldehyde (19). A solution of nitrile 17 (606 mg, 3.11 mmol, 1.0 equiv) in
CH2Cl2 (25 mL) was treated with DIBAL-H (4.4 mL of a 1.0 M solution in
toluene, 4.4 mmol, 1.4 equiv) at 2788C. After stirring for 4 h, the reaction
mixture was quenched with ethyl acetate at –788C, warmed to rt, and
treated with a saturated aqueous sodium-potassium tartrate solution for
30 min. After decantation of both layers, the aqueous phase was extracted

Chiron Approach to Pironetins
277
with EtOAc, and the combined organic solutions were washed with water and
brine, dried (MgSO4), filtered, and concentrated under reduced pressure. The
resulting crude product was subjected to purification by flash column chrom-
atography (silica gel, 20% EtOAc in hexanes) to obtain aldehyde 19 (396 mg,
1
60%) as a colorless oil: Rf ¼ 0.70 (silica gel, 30% EtOAc in hexanes); H NMR
(400 MHz, CDCl3) d (ppm): 0.95 (t, J ¼ 7.5 Hz, 3H), 1.17 (d, J ¼ 5.9 Hz, 3H),
1.24 (d, J ¼ 6.4 Hz, 3H), 1.36–1.53 (m, 2H), 1.88–1.95 (m, 1H), 2.56–2.60
(m, 2H), 3.97 (dt, J ¼ 6.4, 12.4 Hz, 1H), 4.68–4.74 (m, 1H), 4.78 (bs, 1H), 5.67
(d, J ¼ 10.7 Hz, 1H), 5.82 (dd, J ¼ 4.3, 10.2 Hz, 1H), 9.81 (t, J ¼ 2.7 Hz, 1H);
13C NMR (100 MHz, CDCl3) d (ppm): 11.4, 21.5, 23.4, 26.5, 40.8, 64.0, 69.1,
97.7, 124.6, 129.6, 202.3.
(2S,3R,6S)-3-Ethyl-3,6-dihydro-2-((2R,S)-20-hydroxybutyl)-6-isopropoxy-
2H-pyran (20). To a solution of aldehyde 19 (396 mg, 1.87 mmol, 1.0 equiv) in
THF (20 mL) was added EtMgBr (3.4 mL of a 1.0 M solution in THF, 3.36 mmol,
1.8 equiv) at 08C. After stirring for 1 h, the reaction mixture was quenched with
MeOH at 08C and was warmed to rt. Then, a saturated aqueous NH4Cl solution
was added and extracted with ether (20 mL). After decantation of both layers,
the aqueous phase was extracted with ether, and the combined organic solution
was washed with water and brine, dried (MgSO4), filtered, and concentrated
under reduced pressure. Purification by flash column chromatography (silica
gel, 25% EtOAc in hexanes) afforded alcohol 20 (385 mg, 85%) as a 1:1 diaster-
eomeric mixture: Colorless oil; Rf1 ¼ 0.20 and Rf2 ¼ 0.27 (silica gel, 25% EtOAc
in hexanes); 1H NMR (400 MHz, CDCl3) d (ppm): 0.91–0.99 (m, 6H), 1.17
(d, J ¼ 5.9 Hz, 3H), 1.25 (d, J ¼ 6.4 Hz, 3H), 1.38–1.61 (m, 4H), 1.67–1.73
(m, 2H), 1.86–1.94 (m, 1H), 3.75–3.82 (m, 1H), 3.97 (dt, J ¼ 5.9, 12.4 Hz,
1H), 4.37–4.44 (m, 1H), 4.50–4.55 (m, 1H), 4.81 (bs, 1H), 5.61 (dd,
J ¼ 10.2 Hz, 1H), 5.70–5.84 (m, 1H); 13C NMR (100 MHz, CDCl3) d (ppm):
9.8, 11.4, 21.5, 23.5, 26.6, 26.9, 30.1, 40.2, 40.8, 66.3, 68.7, 70.1, 73.2, 97.7,
126.3, 127.2. FAB HRMS (NBA) m/e 265.1782, M þ Naþ calcd for C14H26O3
265.1779.
1-((2S,3R,6S)-3-Ethyl-3,6-dihydro-6-isopropoxy-2H-pyran-2-yl)-2-butanone
˚
(4). To a solution containing alcohol 20 (385 mg, 1.59 mmol, 1.0 equiv), 4 A
molecular sieves (1.6 g), and NMO (278 mg, 2.38 mmol, 1.5 equiv) in CH2Cl2
(20 mL) was added TPAP (56 mg, 0.16 mmol, 0.1 equiv) at 08C. After stirring
for 10 min at this temperature, the reaction mixture was stirred for an
additional hour at rt. Then, the reaction mixture was diluted with ether
(20 mL) and a saturated aqueous NH4Cl solution was added. The aqueous
phase was separated and extracted with ether and the combined organic
solution was washed with water and brine, dried (MgSO4), filtered, and
concentrated under reduced pressure. The resulting crude product was
purified by flash column chromatography (silica gel, 15% EtOAc in hexanes),
to obtain ketone 4 (336 mg, 88%) as a colorless oil: Rf ¼ 0.47 (silica gel, 25%

F. Sarabia et al.
278
1
EtOAc in hexanes); H NMR (400 MHz, CDCl3) d (ppm): 0.94 (t, J ¼ 7.5 Hz,
3H), 1.06 (t, J ¼ 7.5 Hz, 3H), 1.14 (d, J ¼ 5.9 Hz, 3H), 1.25 (d, J ¼ 6.4 Hz,
3H), 1.36–1.53 (m, 2H), 1.84–1.92 (m, 1H), 2.49 (q, J ¼ 7.5 Hz, 2H), 2.51
(dd, J ¼ 4.8, 15.6 Hz, 1H), 2.67 (dd, J ¼ 8.6, 15.6 Hz, 1H), 3.96 (dt, J ¼ 5.9,
12.4 Hz, 1H), 4.63–4.70 (m, 1H), 4.74 (bs, 1H), 5.63 (bd, J ¼ 10.2 Hz, 1H),
5.75 (dd, J ¼ 5.9, 10.2 Hz, 1H); 13C NMR (100 MHz, CDCl3) d (ppm): 7.5, 11.4,
21.4, 23.4, 26.6, 29.7, 30.3, 36.8, 40.8, 47.5, 64.6, 68.4, 97.3, 127.0, 127.5,
209.2. FAB HRMS (NBA) m/e 263.1625, M þ Naþ calcd for C14H24O3 263.1623.
ACKNOWLEDGMENTS
´
´
This work was financially supported by Fundacion Ramon Areces and the
´
´
´
´
Direccion General de Investigacion y Cientıfica Tecnica (ref. CTQ2004-08141)
´
´
and fellowships from Fundacion Ramon Areces (M. G. and S. C.) and Ministerio
´
´
de Educacion y Ciencia (A. S.). We thank Unidad de Espectroscopıa de Masas
de la Universidad de Granada for exact mass spectroscopic assistance.
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