Synthesis of (R)-2-methyl-4-deoxy and (R)-2-methyl-4,5-dideoxy analogues of 6-phosphogluconate as potential inhibitors of 6-phosphogluconate dehydrogenase.

Article abstract of DOI:10.1039/b210606j

The synthesis of (2R)-2-methyl-4,5-dideoxy and (2R)-2-methyl-4-deoxy analogues of 6-phosphogluconate is described. The synthetic strategy relies on the Evans aldol reaction for the installation of the chiral centres in the 2- and 3-positions. The selective phosphorylation at the primary alcohol function of (2R,3S)-3,6-dihydroxy-2-methylhexanoic acid benzyl ester (5) and (2R,3S,5S)-3,5,6-trihydroxy-2-methylhexanoic acid benzyl ester (20) was achieved with dibenzyl phosphochloridate and dibenzyl phosphoiodinate respectively, working at low temperature. (2R,3S)-3-Hydroxy-2-methyl-6-phosphonoxyhexanoic acid (9) was obtained in 25% overall yield from 4-benzyloxybutanol and (2R,3S,5S)-3,5-dihydroxy-2-methyl-6-phosphonoxyhexanoic acid (28) in 10% overall yield from L-malic acid.

Full text of DOI:10.1039/b210606j

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Synthesis of (R)-2-methyl-4-deoxy and (R)-2-methyl-4,5-dideoxy
analogues of 6-phosphogluconate as potential inhibitors of
6-phosphogluconate dehydrogenase†
Christophe Dardonville and Ian H. Gilbert*
Welsh School of Pharmacy, Cardiff University, Redwood Building, King Edward VII Avenue,
Cardiff, UK CF10 3XF
Received 28th October 2002, Accepted 25th November 2002
First published as an Advance Article on the web 14th January 2003
The synthesis of (2R)-2-methyl-4,5-dideoxy and (2R)-2-methyl-4-deoxy analogues of 6-phosphogluconate is
described. The synthetic strategy relies on the Evans aldol reaction for the installation of the chiral centres in
the 2- and 3-positions. The selective phosphorylation at the primary alcohol function of (2R,3S)-3,6-dihydroxy-
2-methylhexanoic acid benzyl ester (5) and (2R,3S,5S)-3,5,6-trihydroxy-2-methylhexanoic acid benzyl ester (20)
was achieved with dibenzyl phosphochloridate and dibenzyl phosphoiodinate respectively, working at low
temperature. (2R,3S)-3-Hydroxy-2-methyl-6-phosphonoxyhexanoic acid (9) was obtained in 25% overall
yield from 4-benzyloxybutanol and (2R,3S,5S)-3,5-dihydroxy-2-methyl-6-phosphonoxyhexanoic acid (28)
in 10% overall yield from -malic acid.
Introduction
The parasitic infection Human African Trypanosomiasis (sleep-
ing sickness) is a major health problem in sub-Saharan Africa
causing an estimated 66 000 deaths per year.1 The treatment of
African Trypanosomiasis is unsatisfactory, with most of the
drugs in use giving rise to serious side effects, increasing
problems of resistance and, in some cases, poor blood-brain
barrier permeability.2 Thus, there is an urgent need for new
drugs to treat sleeping sickness.
Bloodstream forms of Trypanosoma brucei (T. brucei), the
causative organisms of sleeping sickness, exclusively produce
energy (ATP) through glycolysis, making the inhibition of any
of the glycolytic enzymes a potential therapeutic approach.3
In our search for new anti-trypanosomal agents, we decided
to target the enzyme 6-phosphogluconate dehydrogenase
(6PGDH), the third enzyme of the pentose phosphate pathway
(PPP).4 The PPP is responsible for generation of NADPH
(a biosynthetic intermediate used in protection against oxid-
ative stress) and ribulose 5-phosphate (used in the biosynthesis
of nucleotides) (Fig. 1).
A number of reasons suggest that 6PGDH could be a good
target for chemotherapy5 and inhibitors have been found that
are selective for 6PGDH of T. brucei. In particular, 2-deoxy-
Fig. 1
6-phosphogluconate (Fig. 1) is a selective inhibitor of the
T. brucei enzyme.6
Crystallographic studies of the sheep liver7 as well as the
T. brucei8 enzyme have shown differences at the active site that
it should be possible to exploit for selective drug design. A
particular point of interest is a larger binding pocket for the
trypanosome enzyme in the region of the 2-OH group of
the substrate. Moreover, molecular modelling studies9 on the
interaction between 6PG and the T. brucei enzyme revealed a
number of important functionalities involved in the interaction.
In particular, the 4-OH group does not appear to have strong
interactions with the enzyme and we wanted to probe the
importance of the 5-OH in determining activity and selectivity.
These findings were used to design a series of (2R)-2-methyl-
4,5-dideoxy and (2R)-2-methyl-4-deoxy analogues of 6PG
(Fig. 1) that will allow us to carry out Structure Activity
Relationship studies to probe the active site of 6PGDH enzyme.
Results and discussion
General synthetic approach
The synthetic strategy relies on the Evans aldol reaction for the
installation of the chiral centres in the 2- and 3-positions. The
use of boron Z-enolates of chiral oxazolidin-2-one derivatives
allows high diastereoselectivity in the reaction and formation of
almost exclusively the syn aldol product.10 The second step is
the exchange of the chiral auxiliary with a benzyl ester pro-
tecting group that can be removed in very mild conditions
(i.e. catalytic hydrogenolysis). Finally, introduction of the phos-
phate moiety with protected phosphorus() reagents is achieved
selectively at the primary alcohol working at low temperature.
† Electronic supplementary information (ESI) available: experimental
procedure and spectroscopic data (1H NMR, 13C NMR, DEPT) for
compounds 2, 12, 13, 14, 15 and 21b and the previous synthetic
approach tried for the synthesis of (2R)-2-methyl-4,5-dideoxy ana-
logues. See http://www.rsc.org/suppdata/ob/b2/b210606j/
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
T h i s j o u r n a l i s © T h e R o y a l S o c i e t y o f C h e m i s t r y 2 0 0 3
552

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A previous approach11 in which we tried to remove the chiral
auxiliary at the end of the synthesis just before the deprotection
of the phosphate turned out to be unsuccessful because of the
sensitivity of the phosphate to the reaction conditions used
(LiOH–H2O2). The phosphate group was cleaved using these
reaction conditions and the main product obtained was a seven-
member ring lactone that resulted from the intramolecular
attack of the terminal 5-OH to the carbonyl at C1.
isolation of the benzyl ester 5 in only low to moderate yield (13–
50%). Since we thought that the presence of the free hydroxy
groups could be in part responsible for the low to moderate
yields, diol 4 was first protected as a THP ether and the crude
THP-protected 4 reacted with three equivalents of lithium ben-
zyloxide. In this reaction, excess benzyl alcohol, generated as a
by-product, has the same Rf value as the THP-protected benzyl
ester product, which makes the purification of the intermediate
difficult. Hence, the crude reaction mixture was treated with
Dowex acidic resin to remove the THP protecting groups, and 5
could be isolated by chromatography (overall yield for the three
steps from 4–5 is 56–77%).
Synthesis of (2R)-2-methyl-4,5-dideoxy analogues (Scheme 1)
Swern oxidation12 of 4-benzyloxybutanol 1 yielded 4-benzyl-
oxybutyraldehyde13 2 in 87% yield. Aldehyde 2 was used in the
boron aldol reaction14 with (4R)-4-benzyl-3-propionyloxazol-
idin-2-one15 and freshly prepared dibutylboron triflate ‡16 to
afford the syn aldol product (2R,3S)-3 in 70% yield.
Selective phosphorylation of 5 at the primary alcohol group
was accomplished by adding 1.3 eq. of dibenzyl phosphochlor-
idate (freshly prepared from dibenzyl phosphate and sulfuryl
chloride in CCl4)18 at 0 ЊC to a solution of 5 in pyridine. That
phosphorylation occurred on the primary alcohol was estab-
Hydrogenolysis of the benzyl protecting group with 10% Pd–
C followed by lithium benzyloxide transesterification17 allowed
1
lished by the diagnostic downfield H NMR chemical shift of
the CH2 protons at C6 (4.06 ppm compared to 3.70 ppm for 5)
and the unaffected CH proton at C2 (3.9 ppm).
To probe the necessity of the 3-OH, the 3-hydroxy was
derivatised as follows: phosphate 6 was acetylated with acetic
anhydride and a catalytic amount of DMAP in pyridine to
afford 7. The methylated derivative 8 was obtained by reaction
of 5 with silver() oxide and MeI in CH3CN at 60 ЊC for 3 days.
André et al.19 reported that among the numerous procedures of
alkylation of hydroxy groups described in the literature, it
appeared that the one using MeI and Ag2O was the most
efficient for substrates possessing an ester or a lactone group.20
However, the main products we obtained in this reaction were
the transesterification product 8b (47%) and recovered starting
material 6 together with 20% of the expected 8a (39% con-
version). An attempt at methylation of 6 with MeI and NaH
failed and no expected product was detected in the reaction
mixture. Finally, the free phosphate derivatives 9, 10 and 11
were obtained quantitatively by hydrogenolysis of the benzyl
protecting groups.
Synthesis of (2R)-2-methyl-4-deoxy analogues (see Scheme 3)
The strategy used for the preparation of the 4-deoxy analogues
was similar to that used for the 4,5-dideoxy analogues, using the
aldehyde (3S)-1519,21 in the boron aldol reaction. Enantio-pure
(3S)-15 was prepared in four high-yielding steps from -malic
acid (Scheme 2).22
Scheme 1
‡ Triflate = trifluoromethanesulfonate.
Scheme 2
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
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Scheme 3
Diastereoselective boron aldol reaction with aldehyde 15 and
(R)-4-benzyl-3-propionyloxazolidin-2-one afforded a mixture
of the expected (2R,3S)-acetonide 16 (60%) and the depro-
tected triol (2R,3S)-17 (18%, Scheme 3). The latter could be
protected again by reaction with acetone and catalytic toluene-
p-sulfonic acid. Direct removal of the chiral auxiliary by BnOLi
transesterification17 gave a mixture of the expected benzyl ester
19 and a by-product arising from the opening of the oxazol-
idinone ring and an elimination process. Moreover, BnOH has
the same Rf value as the desired product 19, making its purifi-
cation by chromatography rather difficult. Since the direct
transesterification reaction did not give good results, the benzyl
ester 19 was prepared in two steps: the chiral auxiliary was first
removed with LiOH–H2O2,17 affording the pure acid 18 in good
yield (86%) by acid-base work-up. Acid 18 was subsequently
esterified with benzyl bromide and K2CO3 in DMF affording 19
in 78% yield (67% for two steps).
Scheme 4
progression of the reaction to avoid bis-phosphorylation of the
product (this behaviour was observed when 2 eq. of reagent
were added at once).
Acetylation of 21 yielded 23 (90%) whereas methylation with
MeO3BF4–2,6-di-tert-butyl-4-methylpyridine (DTBMP) in
CH2Cl2 afforded a mixture of the mono and bismethylated
products 24 and 25 respectively. These two compounds were
isolated by flash chromatography with 21% and 62% yield
respectively. The last step of the synthesis, consisting of hydro-
genolysis of the benzyl protective group with Pd–C, was
successfully carried out with 23. However, in the case of 21,
which contains two free hydroxy groups, hydrogenolysis led to a
mixture of two phosphorus compounds, possibly due to the
migration of the phosphate to the adjacent free hydroxy group.
Phosphate migration during hydrogenolysis is a well-known
phenomenon, in inositols for instance,26 where migration of
fully protected phosphate around the ring can occur. On the
contrary, migration of phosphate in fully deprotected inositol
1-phosphate only occurs in strong acid or strong base. More-
over, cyclic phosphate formation is a potential side reaction
where vicinal diols are present.27 Some authors have success-
fully prepared some -glycero--manno-heptoside phosphate
derivatives by catalytic hydrogenolysis of their fully benzyl-
protected precursors (no migration was observed in this case).28
Those facts suggested that the protection of the 5-hydroxy of
21 as benzyl ether should reduce the tendency to phosphate
migration during hydrogenolysis. Protection of 21 with benzyl
trichloroacetimidate and catalytic triflic acid in CH2Cl2 was
attempted, affording only a moderate yield of 22 (< 25%) in all
Removal of the isopropylidene protective group is not as
straightforward as it seems because of the presence of the
benzyl ester functionality and therefore the risk of debenzyl-
ation during acid hydrolysis or transesterification when using
alcohol as solvent. This behaviour was observed when Dowex
50WX2 in MeOH was used to hydrolyse the acetonide and the
methyl ester derivative was obtained as a major by-product.
i
Eventually, CuCl2ؒ2H2O in PrOH was used according to the
procedure of Iwata et al.23 to afford the triol 20 in good yield
(69%).
Selective phosphorylation at the primary hydroxy group was
first attempted with dibenzyl phosphochloridate giving only
low yield (< 20%) of the phosphate 21. The fact that starting
material 20 was always recovered in these attempts prompted us
to use the more reactive dibenzyl phosphoiodinate (prepared
from (BnO)3P and I2 in CH2Cl2)24 as the phosphorylating agent.
Working at Ϫ78 ЊC allowed the preparation of 21 in 54% yield.
However, higher temperatures (i.e. Ϫ10 ЊC to ϩ15 ЊC over 2 h)
resulted in the lactonisation of the product to 21b (Scheme 4).25
Furthermore, although two equivalents of dibenzyl
phosphoiodinate are used to drive the reaction, the second
equivalent should be added in small portions, controlling
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
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the attempts carried out. Thus, the 1,3-diol 21 was protected as
its benzylidene acetal derivative with benzaldehyde dimethyl
acetal and PPTS in DMF to afford 26 (71%).
Unfortunately, hydrogenolysis of either 22 or 26 led to a
mixture of two phosphate regioisomers. To circumvent this
problem, 21 was protected as a THP ether 27 in high yield
(81%) and then subjected to hydrogenolysis30 followed by mild
acidic hydrolysis to remove the THP protecting groups, and
afford the phosphate 28 (70%).
(1H, m, CHMe); 3.55 (2H, m); 3.3 (1H, dd, J 3.3 and 13.5,
CH2Ph); 2.85 (1H, dd, J 9.5 and 13.5, CH2Ph); 1.9–1.6 (5H, m);
1.3 (3H, d, J 7, CH3); δC (CDCl3) 177.6 (s); 153.5 (s); 138.7 (s);
135.5 (s); 129.9 (d); 129.4 (d); 128.8 (d); 128.2 (d); 128.14 (d);
128.1 (d); 128.0 (d); 127.9 (d); 73.4 (t); 71.9 (d); 70.6 (t); 66.6 (t);
55.6 (d); 42.8 (d); 38.2 (d); 31.5 (t); 26.8 (t); 11.1 (q); m/z (ESϩ)
434 (M ϩ Na); m/z (ES) 412.2117 (M ϩ H.C24H30NO5 requires:
412.2124).
(4R)-4-Benzyl-3-[(2R,3S)-3,6-dihydroxy-2-methylhexanoyl]-
oxazolidin-2-one (4)
Conclusion
A solution of 3 (572 mg, 1.4 mmol) was dissolved in MeOH
(50 cm3) and hydrogenated for 2.5 h with 5% Pd–C (200 mg)
and 30 Psi H2 pressure. The catalyst was filtered off and the
solvent evaporated to yield 4 as a yellowish oil (434 mg, 97%);
Rf: 0.2 (25% hexane in EtOAc); [α]2D0 = Ϫ81.8 (c 0.11, MeOH);
υmax(neat)/cmϪ1 3400; 1780; 1696; δH (CDCl3) 7.5–7.2 (5H, m);
4.85 (1H, m, CHN); 4.35 (2H, m, OCH2CH); 4.15 (1H, m,
CHOH); 4.0–3.75 (3H, m); 3.4 (1H, dd, J 3.3 and 13.5, CH2Ph);
2.9 (1H, dd, J 9.4 and 13.5, CH2Ph); 2.6 (2H, br s, 2 × OH );
1.85 (2H, quint, CH2CH2CH2); 1.75 (2H, m); 1.4 (3H, d, J 7,
CH3); δC (CDCl3) 177.3 (s); 153.7 (s); 135.5 (s); 129.9 (d); 129.4
(t); 127.8 (t); 72.2 (d); 66.7 (t); 62.8 (t); 55.6 (d); 43.2 (d); 38.1 (t);
31.6 (t); 29.8 (t); 11.5 (q); m/z (ESϩ) 344 (M ϩ Na).
An efficient synthetic route to (2R)-2-methyl-4-deoxy and
(2R)-2-methyl-4,5-dideoxy analogues of 6-phosphogluconate
is reported, starting from 4-benzyloxybutanol 1 and -malic
acid respectively and using boron aldol methodology for the
incorporation of the (2S,3R) chiral centres. This general
approach is currently being used to prepare more 2-substituted-
4-deoxy analogues of 6-phosphogluconate.29
Experimental section
Where applicable, all glassware was oven dried overnight and all
reactions were carried out under nitrogen. All dry solvents were
purchased from Aldrich or Fluka in sure seal bottles. All reac-
tions were monitored by Thin Layer Chromatography (TLC)
using silica gel 60 F254 plates (Merck). Column chromatography
was performed on Uetikon silica gel C-560 Act (0.035–0.070
mm). The infrared spectroscopy (IR) of the compounds was
recorded on a Perkin Elmer 1600 series FTIR spectrometer as
films on sodium chloride discs (thin film). [α]D values were
measured on a ADP220 polarimeter (Bellingham & Stanley
Ltd) and are given in 10Ϫ1 deg cm2 gϪ1. 1H NMR, 13C NMR and
31P NMR spectra were recorded on a Bruker Avance DPX 300
MHz spectrometer at 300, 75 and 121 MHz respectively, using
the solvent residual peak as internal reference. J values are
given in Hz. Elemental analyses were determined on a Perkin-
Elmer 240C elemental analyser. Low resolution mass spectra
(MS) were recorded on a Fisons VG Platform II spectrometer
using electrospray ionisation (ES) technique either in positive
or negative ion modes. Mobile phases were acetonitrile–water
1 : 1 or methanol (ES). High-resolution mass spectra (HRMS)
in chemical ionisation (CI) and fast atom bombardment
(FAB) were determined by the EPSRC Mass Spectroscopy
Centre, Swansea, UK.
(2R,3S)-3,6-Dihydroxy-2-methylhexanoic acid benzyl ester (5)
A solution of 4 (453 mg, 1.41 mmol), 3,4-dihydro-2H-pyran
(DHP) (0.77 cm3, 8.5 mmol) and PPTS (38 mg, 0.14 mmol) in
CH2Cl2 (6 cm3) was stirred at rt for 16 h. The reaction was
partitioned between CH2Cl2 and half-saturated brine. The
organic phase was dried (MgSO4) and concentrated to give a
colourless oil (702 mg). The crude THP-protected diol 4 was
dissolved in THF (8 cm3) and treated at Ϫ5 ЊC with a BnOLi
solution [prepared by addition at 0 ЊC of nBuLi (4.9 mmol)
to a solution of benzyl alcohol (0.58 cm3, 5.6 mmol) in THF
(3 cm3)]. After 2 h at 0 ЊC, the reaction was quenched with
saturated NH4Cl solution (2 cm3). The mixture was partitioned
between water and EtOAc and the aqueous phase was extracted
with EtOAc (3 × 20 cm3). Combined organic extracts were
washed with brine, dried (MgSO4) and concentrated to give a
yellowish oil (1.23 g). This oil was dissolved in 50 cm3 of MeOH
and treated with wet Dowex 50X8 resin for 18 h. The resin was
filtered off and rinsed with MeOH. The methanolic solution
was concentrated in vacuo and the residue was partitioned
between EtOAc and brine. The organic phase was dried
(MgSO4) and concentrated to give a crude yellow oil. Chrom-
atography with EtOAc–hexane (3 : 1) afforded the pure ester 5
as a colourless oil (200 mg, typical yield: 56–77%); Rf: 0.22
(EtOAc–hexane: 3 : 1); [α]2D3 = Ϫ10 (c 0.1, MeOH); δH (CDCl3)
7.45 (5H, m); 5.2 (2H, s, OCH2Ph); 3.95 (1H, m, CHOH); 3.70
(2H, m, CH2OH); 3.20 (1H, br, OH ); 2.7 (1H, m, CHMe); 2.4
(1H, br, OH ); 1.75 (2H, m); 1.6 (2H, m); 1.3 (3H, d, J 7, CH3);
δC (CDCl3) 176.3 (s); 136.1 (s); 129.0 (d); 128.8 (d); 128.6 (d);
72.3 (d); 66.9 (t); 63.1 (t); 45.2 (d); 31.6 (t); 30.0 (t); 11.5 (q); m/z
(ESϩ) 275 (M ϩ Na); m/z (ES) 253.1442 (M ϩ H. C14H21O4
requires: 253.1440).
(4R)-4-Benzyl-3-[(2R,3S)-6-benzyloxy-3-hydroxy-2-methyl-
hexanoyl]oxazolidin-2-one (3)
Freshly prepared dibutylboron triflate 16 (2.35 cm3, 9.3 mmol)
was added to a solution of (3R)-3-propionyl-4-benzyloxazol-
idin-2-one (1.67 g, 7.2 mmol) in dry CH2Cl2 (15 cm3) at 0 ЊC.
Et3N (1.3 cm3, 9.3 mmol) was then added keeping the internal
temperature below ϩ3 ЊC. The resulting yellow solution was
cooled to Ϫ78 ЊC and aldehyde 2 (1.53 g, 8.6 mmol) was added
dropwise. The clear yellow solution was stirred for 1 h 15 min
at 0 ЊC. The reaction was quenched with 7 cm3 of phosphate
buffer (pH 7.2) and 20 cm3 of MeOH. Then, 10 cm3 of a 2 : 1
solution of MeOH–H2O2 (30% in water) was added dropwise,
keeping the internal temperature below ϩ10 ЊC. This mixture
was stirred for 1 h at 0 ЊC. The solvent was removed in vacuo
and the resulting oil was partitioned between water and EtOAc.
The aqueous phase was extracted with EtOAc (2 × 50 cm3).
Organic extracts were washed with NaHCO3, brine, and dried
(MgSO4). Removal of the solvent afforded a yellow oil that was
chromatographed (40% EtOAc–hexane) to give 3 as a colour-
(2R,3S)-6-(Bis(benzyloxy)phosphoryloxy)-3-hydroxy-2-methyl-
hexanoic acid benzyl ester (6)
Sulfuryl chloride (0.12 cm3,1.5 mmol) was added with a syringe
to a flask containing a solution of dibenzyl phosphite (0.33 cm3,
1.5 mmol) in CCl4 (2 cm3). After 1 h at rt, the volatiles were
removed in vacuo and the product was used directly without
purification. (BnO)2P(O)Cl (320 mg, 1.3 eq.) was added at 0 ЊC
to the alcohol 5 dissolved in pyridine (2 cm3) under a N2 atmos-
phere. The reaction was stirred for 2 h at 0 ЊC and 1 h at rt. The
solvent was removed in vacuo and the crude yellow oil was
chromatographed with EtOAc–hexane (5 : 2) to give 6 (247 mg,
less oil (2.06 g, 70%); Rf: 0.37 (50% EtOAc in hexane); [α]2D2
=
Ϫ40 (c 0.1, CH2Cl2); υmax(neat)/cmϪ1 3442; 1777; 1696; δH
(CDCl3) 7.5–7.25 (10H, m); 4.75 (1H, m, CHN); 4.6 (2H, s,
OCH2Ph); 4.25 (2H, m, OCH2CH); 4.1 (1H, m, CHOH); 3.85
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
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60%) as a colourless oil; Rf: 0.47 (EtOAc–hexane, 3 : 1); [α]2D3
=
47.4 (d); 32.6 (t); 28.65 (d, J 7.5, CH2); 12.9 (q); δP (CD3OD)
ϩ1.37; m/z (ESϪ) 241 (M Ϫ H); m/z (ESϪ) 241.0478 (M Ϫ H.
C7H14O7P requires 241.0477).
Ϫ4.3 (c 0.23, MeOH); δH (CDCl3) 7.4 (15H, m); 5.2 (2H, s,
OCH2Ph); 5.1 (2H, d, J 3.3, POCH2Ph); 5.08 (2H, d, J 3.3,
POCH2Ph); 4.06 (2H, m, CH2CH2OP); 3.9 (1H, m, CHOH);
2.86 (1H, d, J 4.9, OH ); 2.58 (1H, qd, J 7.2 and 3.8, CHMe);
1.85 (1H, m); 1.7 (1H, m); 1.45 (2H, m); 1.23 (3H, d, J 7.2,
CH3); δC (CDCl3) 176.2 (s); 136.3 (2 × s); 136.1 (s); 129.1 (d);
129.0 (2 × d); 128.8 (d); 128.6 (d); 128.4 (d); 71.7 (d); 69.6 (d,
J 7.5, CH2); 68.2 (d); 66.9 (t); 45.0 (d); 30.2 (t); 27.4 (d, J 7.5,
CH2); 11.3 (q); δP (CD3OD) ϩ 0.5; m/z (ESϩ) 535 (M ϩ Na);
m/z (ES) 513.2047 (M ϩ H. C28H34O7P requires: 513.2042).
(2R,3S)-3-Acetoxy-2-methyl-6-phosphonoxyhexanoic acid (10)
A suspension of 7 (25 mg, 0.045 mmol) and 5% Pd–C in MeOH
(5 cm3) was hydrogenated for 1 h to yield 10 as a colourless oil
(16 mg, 100%); [α]2D4 = Ϫ6.2 (c 0.16, MeOH); δH (CD3OD) 5.2
(1H, br m); 4.0 (2H, br); 2.7 (1H, m); 2.05 (3H, s, CH3CO);
1.8–1.5 (4H, br m); 1.15 (3H, d, J 6.9, CHCH3); δC (CD3OD)
177.82 (s); 172.84 (s); 75.59 (d); 67.33 (t); 44.75 (d); 30.0 (t);
28.22 (t); 21.28 (q); 12.87 (q); δP (CD3OD) ϩ1.42; m/z (ESϪ) 283
(M Ϫ H); m/z (ESϩ) 307.0554 (M ϩ Na. C9H17NaO8P requires
307.0559).
(2R,3S)-3-Acetoxy-6-bis(benzyloxy)phosphoryloxy-2-methyl-
hexanoic acid benzyl ester (7)
A solution of 6 (80 mg, 0.156 mmol), acetic anhydride (16 µl,
0.17 mmol) and DMAP (4 mg, 0.03 mmol) in pyridine (2 cm3)
was stirred for 18 h at rt. Pyridine was removed in vacuo and the
crude oil was chromatographed with 50% EtOAc–hexane to
afford 7 as a colourless oil (48 mg, 56%); Rf: 0.34 (50% EtOAc–
hexane); [α]2D2 = Ϫ4 (c 0.25, MeOH); δH (CDCl3) 7.4 (15H, m);
5.2 (1H, m, CHOAc); 5.13 (2H, s, OCH2Ph); 5.1 (2H, m,
POCH2Ph); 5.05 (2H, m, POCH2Ph); 4.0 (2H, m, CH2CH2OP);
2.75 (1H, m, CHMe); 2.0 (3H, s, CH3CO); 1.65 (4H, m); 1.2
(3H, d, J 7.1, CHCH3); δC (CDCl3) 173.8 (s); 170.89 (s); 136.3
(s); 136.21 (s); 136.16 (s); 129.03 (d); 129.0 (d); 128.84 (d);
128.76 (d); 128.39 (d); 73.86 (d); 69.73 (t); 69.66 (t); 67.7 (J 5.9,
CH2); 67.03 (t); 43.48 (d); 28.35 (t); 26.8 (d, J 7.3, CH2);
21.2 (q); 12.1 (q); δP (CDCl3) ϩ 0.32; m/z (FAB/3-nitrobenzyl
alcohol (NOBA) matrix) 555 (M ϩ H); m/z (FAB) 555.2143
(M ϩ H. C30H36O8P requires: 555.2148).
(2R,3S)-3-Methoxy-2-methyl-6-phosphonoxyhexanoic acid (11)
A suspension of 8a (45 mg) and 5% Pd–C in MeOH (4 cm3) was
hydrogenated for 22 h at rt to yield 11 as a yellowish oil.
δH (CD3OD) 4.0 (2H, br); 3.7 (1H, br d); 3.55 (1H, br m); 3.4
(3H, s, OCH3); 2.62 (1H, m); 1.9–1.6 (4H, m); 1.15 (3H, d, J 7.2,
CHCH3); δC (CD3OD) 179 (s); 83.6 (d); 67.6 (t); 58.6 (q); 44.4
(d); 29.4 (t); 28.0 (t); 12.4 (q); δP (CD3OD) ϩ1.33; m/z (ESϪ) 255
(M Ϫ H); m/z (ESϪ) 255.0632 (M Ϫ H. C8H16O7P requires
255.0638).
(4R)-4-Benzyl-3-{(2R,3S)-4-[(1S)-2,2-dimethyl-[1,3]-dioxolan-
4-yl]-3-hydroxy-2-methylbutyryl}oxazolidin-2-one (16)
Dibutylboron triflate (3.55 cm3, 14 mmol) was added dropwise
to a solution of (3R)-3-propionyl-4-benzyloxazolidin-2-one
(2.995 g, 12.8 mmol) in dry CH2Cl2 (20 cm3) at 0 ЊC, followed by
Et3N (2.1 cm3, 15.4 mmol), keeping the internal temperature
below ϩ3 ЊC. The clear light yellow solution was then cooled to
Ϫ65 ЊC and a solution of aldehyde 15 (2.4 g, 16.7 mmol) in
CH2Cl2 (20 cm3) was added dropwise. The solution was stirred
for 40 min at Ϫ65 ЊC and 1 h at Ϫ5 ЊC. The reaction was
quenched with pH 7.2 phosphate buffer (20 cm3) and MeOH
(40 cm3). Then, 42 cm3 of a 2 : 1 solution of MeOH–H2O2 (30%
in water) was added dropwise, keeping the internal temperature
below ϩ10 ЊC. This mixture was stirred for 1 h at 0 ЊC. Solvent
was removed in vacuo and the resulting oil was partitioned
between water and EtOAc. The aqueous phase was extracted
with EtOAc (2 × 70 cm3). Organic extracts were washed with
5% NaHCO3, brine, and dried (MgSO4). Removal of the sol-
vent afforded an oil that was chromatographed. The aldol
product 16 (2.313 g, 60%) was eluted first (hexane–EtOAc,
50 100%) and then, the deprotected aldol product 17 (790 mg)
was eluted with EtOAc–MeOH 10%. 16 is a colourless oil that
(2R,3S)-6-Bis(benzyloxy)phosphoryloxy-3-methoxy-2-methyl-
hexanoic acid benzyl ester (8a)
A solution of 6 (100 mg, 0.195 mmol), Ag2O (68 mg, 0.29 mmol)
and MeI (0.5 cm3, 8 mmol) in CH3CN (2.5 cm3) was stirred
for 1 day at rt and 3 days at 60–70 ЊC. The reaction mixture
was diluted with CH2Cl2 and filtered on a path of Celite. Evap-
oration of the solvent gave an orange oil that was chromato-
graphed with 50% EtOAc–hexane. The expected product 8a (20
mg, 39% conversion) was eluted first, then the starting material
6 (50 mg, 50%) and finally the methyl ester by-product 8b (40
mg, 47%). [α]2D1 = Ϫ2.5 (c 2, MeOH); δH (CDCl3) 7.4 (15H, br s);
5.2 (2H, d, J 4.4, OCH2Ph); 5.12 (2H, m, POCH2Ph); 5.09 (2H,
m, POCH2Ph); 4.0 (2H, m, CH2CH2OP); 3.5 (1H, m, CHOH);
3.3 (3H, s, OCH3); 2.65 (1H, m, CHMe); 1.9–1.5 (4H, m); 1.2
(3H, d, J 7.2, CHCH3); δP (CDCl3) ϩ 0.45; m/z (ESϩ) 549 (M ϩ
Na); m/z (ES) 527.2195 (M ϩ H. C29H36O7P requires 527.2198).
solidified on standing. Rf: 0.4 (50% EtOAc in hexane); [α]2D3.7
=
Ϫ10.7 (c 0.28, CH2Cl2); δH (CDCl3) 7.45–7.2 (5H, m); 4.74 (1H,
m, CHN); 4.39 (1H, m); 4.33–4.12 (4H, m, OCH2CHN,
CH2CH(OH)CH2(O) and CH(OH)CH2OC); 3.86 (1H, qd, J 3
and 7, CHMe); 3.65 (1H, dd, J 7.4 and 8.0, CH(OH)CH2OC);
3.3 (1H, dd, J 3.3 and 13.4,CH2Ph); 3.24 (1H, br, OH ); 2.85
(1H, dd, J 9.4 and 13.4, CH2Ph); 1.9–1.65 (2H, m, CHCH2CH);
1.47 (3H, s, CCH3); 1.42 (3H, s, CCH3); 1.33 (3H, d, J 7,
CHCH3); δC (CDCl3) 177.6 (s); 153.5 (s); 135.4 (s); 129.9 (d);
129.4 (d); 127.9 (d); 109.2 (s); 73.9 (d); 70.4 (t); 69.1 (d); 66.6 (t);
55.5 (d); 42.9 (d); 38.2 (t); 38.0 (t); 27.4 (q); 26.1 (q); 11.3 (q);
m/z (ESϩ) 400 (M ϩ Na); m/z (ESϩ) 378.1910 (M ϩ H. C20H28-
NO6 requires 378.1916).
(2R,3S)-6-Bis(benzyloxy)phosphoryloxy-3-hydroxy-2-methyl-
hexanoic acid methyl ester (8b)
δH (CDCl3) 7.4 (10H, br s); 5.1 (2H, m, POCH2Ph); 5.05 (2H,
m, POCH2Ph); 4.1 (2H, m, CH2CH2OP); 3.9 (1H, m, CHOH);
3.75 (3H, s, COOCH3); 2.72 (1H, br d, J 4.7, OH); 2.54 (1H, qd,
J 3.5 and 7.2, CHMe); 1.9 (1H, m); 1.7 (1H, m); 1.5 (2H, m);
1.2 (3H, d, J 7.1, CHCH3); δP (CDCl3) ϩ 0.5; m/z (ESϩ) 459
(M ϩ Na); m/z (ESϩ) 437.1729 (M ϩ H. C22H30O7P requires:
437.1729).
(2R,3S)-3-Hydroxy-2-methyl-6-phosphonoxyhexanoic acid (9)
A solution of 6 (54 mg, 0.1 mmol) in MeOH (2 cm3) was hydro-
genated for 5 h with 5% Pd–C (12 mg). The catalyst was filtered
off and the solvent evaporated to yield a colourless oil (24 mg).
Et2O-mediated precipitation of the product dissolved in a little
MeOH yielded 9 as a very hygroscopic colourless solid (24 mg,
94%). [α]2D2 = Ϫ8.3 (c 0.24, MeOH); δH (CD3OD) 4.0 (2H, br m);
3.8 (1H, br m); 2.45 (1H, m); 1.9 (1H, br m); 1.8–1.4 (4H, br m);
1.2 (3H, d, J 7, CH3); δC (CD3OD) 179.3 (s); 73.5 (d); 68.0 (t);
(4R)-4-Benzyl-3-[(2R,3S)-3,5,6-trihydroxy-2-methylhexanoyl]-
oxazolidin-2-one (17)
White foam; Yield: 18%; Rf: 0.18 (EtOAc); [α]2D6 = Ϫ141.7
(c 0.24, MeOH); δH (CDCl3) 7.45–7.2 (5H, m); 4.8 (1H, m,
CHN); 4.4–4.2 (3H, m); 4.05 (1H, br m); 3.95–3.5 (4H, m); 3.3
(1H, dd, J 13.3 and 3.3, CH2Ph); 2.85 (1H, dd, J 9.3 and 13.3,
CH2Ph); 2.48 (1H, br s); 1.8–1.4 (3H, m); 1.3 (3H, d, J 7,
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
556

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CHCH3); δC (CDCl3) 176.9 (s); 154.0 (s); 135.5 (s); 129.8 (d);
129.3 (d); 127.8 (d); 69.48 (d); 68.88 (d); 66.94 (t); 66.79 (t); 55.7
(d); 43.7 (d); 38.0 (t); 37.7 (t); 12.0 (q); m/z (ESϩ) 360 (M ϩ Na).
as a colourless oil that solidified on standing (315 mg, 69%)
(Found: C, 62.10; H, 7.48. C14H20O5 requires C, 62.67; H,
7.51%); Rf: 0.28 (EtOAc); [α]2D4 = Ϫ22.2 (c 0.09, MeOH); δH
(CDCl3) 7.45 (5H, m); 5.25 (2H, s, OCH2Ph); 4.3 (1H, m,
CH2CH(OH)CH2); 4.1 (1H, m, CHCHOH); 3.7 (1H, m); 3.55
(1H, m); 3.5 (1H, br, OH ); 3.3 (1H, br, OH ); 2.8 (1H, br, OH );
2.72 (1H, m, CHMe); 1.75–1.5 (2H, m, CHCH2CH); 1.35 (3H,
d, J 7.1, CHCH3); δC (CDCl3) 176.0 (s); 138.0 (s); 129.0 (d);
128.6 (d); 69.5 (d); 69.0 (d); 67.3 (t); 66.9 (t); 45.8 (d); 37.4 (t);
12.2 (q); m/z (ESϩ) 269 (M ϩ Na); m/z (ES) 269.1388 (M ϩ H.
C14H21O5 requires 269.1389).
Acetonide protection of 17
A solution of 17 (1.5 g, 4.45 mmol) and TsOH (40 mg, 0.21
mmol) in acetone (150 cm3) was stirred for 2 h at rt. A few drops
of Et3N were added and the solvent removed in vacuo. The
residue was partitioned between EtOAc and 5% NaHCO3 and
the aqueous phase was extracted with EtOAc. The organic
extract was washed with brine, dried (MgSO4) and concentrated
to afford the pure acetonide 16 (1.34 g, 80%).
(2R,3S,5S)-6-[Bis(benzyloxy)phosphoryloxy]-3,5-dihydroxy-2-
methylhexanoic acid benzyl ester (21)
(2R,3S)-4-[(1S)-2,2-Dimethyl-[1,3]-dioxolan-4-yl]-3-hydroxy-2-
methylbutyric acid (18)
Iodine (575 mg, 2.26 mmol) was added at 0 ЊC to a solution of
tribenzyl phosphite 24 (852 mg, 2.4 mmol) in CH2Cl2 (7.5 cm3).
After 15 min at 0 ЊC and 10 min at rt, 4 cm3 of the clear solution
was added at Ϫ78 ЊC to a solution of 20 (345 mg, 1.29 mmol)
and pyridine (0.3 cm3, 3.0 mmol) in CH2Cl2 (7 cm3). The
reaction was followed by TLC and the rest of the dibenzyl
phosphoiodinate was added in 1 cm3 portions (to avoid diphos-
phorylation) approximately every 30 min. After a total of 5 h at
Ϫ78 ЊC, all the reagent (7.5 cm3, 1.8 eq.) had been added and
the reaction was diluted with CH2Cl2, filtered, washed with
water, dried (MgSO4) and concentrated in vacuo. The crude
yellow oil was chromatographed (hexane–EtOAc, 25 100%)
to give the phosphate 21 as a colourless oil (370 mg, 54%); Rf:
0.2 (EtOAc–hexane, 2 : 1); [α]2D2 = Ϫ12.5 (c 0.4, MeOH);
δH (CDCl3) 7.4 (15H, br s); 5.18 (2H, s, OCH2Ph); 5.12 (2H, m,
POCH2Ph); 5.08 (2H, m, POCH2Ph); 4.3–3.85 (4H, m); 3.45
(1H, br, OH ); 3.3 (1H, br d, J 4.2, OH ); 2.65 (1H, m, CHMe);
1.7–1.4 (2H, m, CHCH2CH); 1.25 (3H, d, J 7.2, CHCH3); δC
(CDCl3) 176.0 (s); 136.1 (2 × s); 136.0 (s); 129.1 (2 × d); 128.8
(d); 128.6 (d); 128.5 (d); 72.45 (CH2, J 7.5); 70.05 (CH2, J 7.5);
68.9 (d); 68.05 (CH, J 7.5); 66.8 (t); 45.3 (d); 36.4 (t); 11.9 (q); δP
(CDCl3) ϩ1.08; m/z (ESϩ) 551 (M ϩ Na); m/z (ES) 546.2269
(M ϩ NH4. C28H37NO8P requires 546.2257).
A 0.05 M solution of the aldol product 16 (410 mg, 1.1 mmol)
in 3 : 1 THF–H2O (20 cm3) was treated at 0 ЊC with H2O2 (30%
in water, 0.5 cm3) followed by lithium hydroxide monohydrate
(53 mg, 2.2 mmol). The resulting mixture was stirred for 1 h at
ice-bath temperature. Excess peroxide was quenched at 0 ЊC
with 1.5 M Na2SO3 (5 cm3) and buffered with 5% NaHCO3
(2 cm3). The chiral auxiliary was recovered by CH2Cl2 extrac-
tion (2 × 20 cm3). The aqueous phase was acidified (pH ∼ 2)
with 1 M HCl and extracted with EtOAc (3 × 20 cm3). Ethyl
acetate extracts were dried (MgSO4) and concentrated to give
the acid 18 as a colourless oil (200 mg, 86%); Rf: 0.2 (EtOAc);
[α]2D1 = Ϫ14.5 (c 2, CH2Cl2); δH (CDCl3) 7.5 (1H, br, CO2H ); 4.3
(1H, m, CH(OH)CH2O); 4.2–4.0 (1H, m, CHOH); 4.05 (1H,
dd, J 8.1 and 6.2, CH(OH)CH2O); 3.5 (1H, dd, J 8.1 and 7.3,
CH(OH)CH2O); 2.55 (1H, m, CHMe); 2.0 (1H, d, OH ); 1.6
(2H, m, CHCH2CH); 1.35 (3H, s, CCH3); 1.3 (3H, s, CCH3);
1.15 (3H, d, J 7.2, CHCH3); δC (CDCl3) 180.2 (s); 109.4 (s); 73.2
(d); 69.8 (t); 69.4 (d); 45.0 (d); 37.8 (t); 27.3 (q); 26.0 (q); 11.4
(q); m/z (ESϩ) 241 (M ϩ Na); m/z (ES) 236.1500 (M ϩ NH4.
C10H22NO5 requires 236.1498).
(2R,3S)-4-[(1S)-2,2-Dimethyl-[1,3]-dioxolan-4-yl]-3-hydroxy-2-
methylbutyric acid benzyl ester (19)
(2R,3S,5S)-3,5-Bis(benzyloxy)-6-bis(benzyloxy)phosphoryl-
A mixture of the acid 18 (737 mg, 3.38 mmol), benzyl bromide
(0.8 cm3, 6.8 mmol) and K2CO3 (938 mg, 6.8 mmol) in DMF
(15 cm3) was stirred for 16 h at rt. The reaction was diluted with
CH2Cl2 and inorganic insoluble salts were filtered off (Celite).
The filter cake was rinsed with CH2Cl2 and the filtrate was
concentrated in vacuo. Chromatography with hexane–EtOAc
(4 : 1) afforded 19 as a colourless oil (810 mg, 78%) (Found: C,
65.74; H, 8.07. C17H24O5ؒ0.1 H2O requires C, 65.82; H, 7.80%);
Rf: 0.36 (hexane–EtOAc, 2 : 1); [α]2D4 = Ϫ3.6 (c 1.66, CH2Cl2);
δH (CDCl3) 7.4 (5H, br s); 5.2 (2H, s, OCH2Ph); 4.34 (1H, m,
CH(OH)CH2O); 4.15 (1H, m, CHOH); 4.05 (1H, dd, J 8.1 and
6, CH(OH)CH2O); 3.55 (1H, dd, J 8.1 and 7.5, CH(OH)-
CH2O); 3.1 (1H, d, J 4.6, OH ); 2.65 (1H, m, CHMe); 1.8–1.6
(2H, m, CHCH2CH); 1.48 (3H, s, CCH3); 1.39 (3H, s, CCH3);
1.28 (3H, d, J 7.2, CHCH3); δC (CDCl3) 175.9 (s); 136.1 (s);
129.0 (d); 128.8 (d); 128.7 (d); 109.2 (s); 73.9 (d); 70.0 (t); 69.6
(d); 66.8 (t); 45.3 (d); 37.9 (t); 27.4 (q); 26.1 (q); 11.9 (q); m/z
(ESϩ) 331 (M ϩ Na).
oxy)-2-methylhexanoic acid benzyl ester (22)
To a solution of diol 21 (104 mg, 0.2 mmol) and benzyl trichloro-
acetimidate (0.112 cm3, 0.6 mmol) in CH2Cl2 (2.5 cm3) was
added 10 drops of an ethereal solution of TfOH (3 drops neat
TfOH in 1 cm3 Et2O). The progress of the reaction was followed
by TLC and more ethereal TfOH solution was added after 1 h
(0.1 cm3), 1 h 25 min (0.3 cm3), 2 h (0.2 cm3 ϩ 1 drop of neat
TfOH). After 5 h, 2 drops of neat TfOH and CCl3CNHOBn
(0.02 cm3, 0.1 mmol) were added to drive the reaction to com-
pletion. After an additional 2 h stirring, the reaction was
quenched with 5% NaHCO3 (2 cm3) and partitioned between
water and CH2Cl2. The aqueous phase was extracted with
CH2Cl2 (2 × 15 cm3) and the combined organic extracts washed
with brine, dried (MgSO4) and concentrated to a crude white
solid. Chromatography (hexane–EtOAc, 25 50%) afforded a
mixture of 22 and trichloroacetimidate (same Rf as 22) as
shown by 1HNMR. This by-product was crystallised from
CHCl3 (white crystals) and removed by filtration. 22 was iso-
lated as a colourless oil (60 mg, 43%). [α]2D1 = Ϫ22 (c 1, CH2Cl2);
δH (CDCl3) 7.5 (25H, m); 5.17 (2H, s); 5.09 (2H, s); 5.06 (2H, s);
4.68 (1H, d, J 11.4); 4.52 (1H, d, J 11.2); 4.34 (2H, d, J 11.4);
4.18 (1H, d, J 11.2); 4.1–3.9 (3H, m); 3.81 (1H, m); 2.86 (1H,
m); 1.76 (2H, m); 1.24 (3H, d, J 7, CH3CH); δC (CDCl3) 174.81
(s); 138.66 (s); 138.59 (s); 136.29 (s); 136.16 (s); 129.01 (d);
128.97 (2 × d); 128.83 (d); 128.74 (d); 128.65 (d); 128.38 (2 × d);
128.35 (d); 128.15 (d); 128.02 (d); 77.20 (d); 74.68 (d, J 7.5,
CH); 72.37 (d, J 5.6, CH2); 69.74 (d, J 5.5, CH2); 69.52 (t);
69.44 (t); 66.81 (t); 43.34 (d); 35.56 (t); 12.14 (q); δP (CDCl3) ϩ
0.46; m/z (ESϩ) 709 (M ϩ H); m/z (ES) 709.2930 (M ϩ H.
C42H46O8P requires 709.2930).
(2R,3S,5S)-3,5,6-Trihydroxy-2-methylhexanoic acid benzyl
ester (20)
A solution of 19 (524 mg, 1.7 mmol) and CuCl2ؒ2H2O (1.45 g,
8.5 mmol, 5 eq.) in iPrOH (20 cm3) was stirred for 2 h at rt. The
reaction was quenched with 9.5 cm3 Na2CO3 1 M. When CO2
evolution had stopped, the reaction mixture was diluted with
MeOH and filtered on a path of Celite. The solvent was
removed in vacuo and the residue was partitioned between
EtOAc and water. The aqueous phase was extracted with
EtOAc (2 × 40 cm3). Organic extracts were dried (MgSO4) and
concentrated. Chromatography on silica with EtOAc yielded 20
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 5 5 2 – 5 5 9
557

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(2R,3S,5S)-3,5-Diacetoxy-6-bis(benzyloxy)phosphoryloxy-2-
methylhexanoic acid benzyl ester (23)
(CDCl3) 174.4 (s); 174.0 (s); 138.7 (s); 138.5 (s); 136.2 (s); 136.1
(s); 136.0 (s); 129.4 (d); 129.2 (2 × d); 129.1 (d); 129.0 (2 × d);
128.8 (2 × d); 128.6 (2 × d); 128.4 (4 × d); 126.6 (d); 126.5 (d);
95.8 (d); 95.0 (d); 74.0 (d); 73.6 (d); 71.4 (d, J 7.5, CH); 70.9 (d,
J 7.5, CH); 70.0 (t); 69.93 (t); 69.85 (t); 69.79 (t); 69.75 (t); 69.72
(t); 67.0 (t); 66.8 (t); 66.0 (d, J 6, CH2); 45.7 (d); 40.0 (d); 28.5
(t); 28.4 (t); 14.9 (q); 13.3 (q); δP (CDCl3) ϩ 0.71; ϩ 0.23; m/z
(ESϩ) 634 (M ϩ NH4); m/z (ES) 634.2572 (M ϩ NH4.
C35H41NO8P requires 634.2570).
A solution of 21 (70 mg, 0.13 mmol), acetic anhydride (40 µl,
0.4 mmol) and DMAP (3 mg, 0.026 mmol) in pyridine (2 cm3)
was stirred for 2 h at rt. The solvent was removed in vacuo and
the crude oil was partitioned between CH2Cl2 and water. The
organic phase was dried (MgSO4) and concentrated to give the
pure 23 as an oil (73 mg, 90%); Rf: 0.53 (50% EtOAc–hexane);
[α]2D3 = Ϫ20.3 (c 0.59, CH2Cl2); δH (CDCl3) 7.4 (15H, m); 5.25
(1H, m, CHOAc); 5.15 (2H, s, OCH2Ph); 5.15–5.0 (5H, m,
2 × POCH2Ph and CHOAc); 4.15 (1H, m, CHCH2OP); 3.97
(1H, m, CHCH2OP); 2.75 (1H, m, CHMe); 2.05 (3H, s,
CH3CO); 2.0 (3H, s, CH3CO); 1.85 (2H, m, CHCH2CH); 1.2
(3H, d, J 7.1, CH3CH); δC (CDCl3) 172.1 (s); 169.4 (s); 169.3 (s);
134.7 (s); 134.6 (s); 127.6 (d); 127.4 (d); 127.3 (d); 127.0 (d);
126.9 (d); 68.7 (d); 68.4 (2 × t); 67.15 (d, J 7.5, CH2); 66.75 (d,
J 7.5, CH); 65.6 (t); 42.2 (d); 31.0 (t); 19.8 (q); 19.7 (q); 11.1 (q);
δP (CDCl3) ϩ 0.35; m/z (ESϩ) 635 (M ϩ Na); m/z (ES) 630.2476
(M ϩ NH4. C32H41NO10P requires 630.2568).
(2R,3S,5S)-6-Bis(benzyloxy)phosphoryloxy-2-methyl-3,5-bis-
(tetrahydropyran-2-yloxy)hexanoic acid benzyl ester (27)
A solution of 21 (85 mg, 0.16 mmol), PPTS (20 mg, 0.08 mmol)
and 3,4-dihydro-2H-pyran (0.15 cm3,1.6 mmol) in CH2Cl2
(2.5 cm3) was stirred for 24 h at rt. Solvent was removed in vacuo
and the residue was chromatographed (0 70% EtOAc–hexane)
to afford 27 as a colourless oil (90 mg, 81%). δH (acetone-d6)
7.4–7.2 (15H, m); 5.05–4.95 (6H, m); 4.75–4.2 (2H, m); 4.2–3.65
(6H, m); 3.4–3.15 (2H, m); 2.67 (1H, m); 1.8–1.2 (14H, m); 1.05
(3H, m); δP (acetone-d6) ϩ0.63; ϩ0.53; ϩ0.41; ϩ 0.36; m/z
(ESϩ) 714 (M ϩ NH4); m/z (ES) 714.3401 (M ϩ NH4.
C38H53NO10P requires 714.3407).
(2R,3S,5S)-6-Bis(benzyloxy)phosphoryloxy-3,5-dimethoxy-2-
methylhexanoic acid benzyl ester (24)
A solution of diol 21 (50 mg, 0.095 mmol), MeO3BF4 (70 mg,
0.47 mmol) and DTBMP (117 mg, 0.57 mmol) in CH2Cl2
(3 cm3) was stirred for 3 h at rt. The reaction was diluted with
EtOAc and washed successively with 5% NaHCO3 and brine.
The organic phase was dried (Na2SO4) and concentrated.
(2R,3S,5S)-3,5-Dihydroxy-2-methyl-6-phosphonoxyhexanoic
acid (28)
A suspension of 27 (55 mg) and 5% Pd–C30 (10 mg) in MeOH
(5 cm3) was hydrogenated for 30 min at rt. The catalyst was
filtered off and the solvent was removed in vacuo. The crude
residue was dissolved in THF (5 cm3) and 2 M aqueous HCl
(0.5 cm3) was added. The reaction was stirred for 2 h at rt and
the solvent was evaporated. The residue dissolved in MeOH
was treated with Et2O and the precipitate was allowed to settle
down overnight in the fridge. The solvent was removed with a
pasteur pipette and the oily residue dried in vacuo to afford 28
as a yellowish hygroscopic foam (14 mg, 70%); [α]2D4 = Ϫ14.28
(c 0.28, MeOH); δH (CD3OD) 4.1–3.7 (4H, m); 2.4 (1H, m,
CHMe); 1.65–1.18 (2H, m, CHCH2CH); 1.05 (3H, d, J 7.3,
CH3CH); δC (CD3OD) 177.4 (s); 72.2 (t); 70.2 (d); 68.6 (d, d);
47.8 (d); 39.4 (t); 12.5 (q); δP (CD3OD) ϩ 1.5 (br); m/z (ESϪ) 257
(M Ϫ H); m/z (ES) 281.0400 (M ϩ Na. C7H15O8NaP requires:
281.0402).
Chromatography (hexane–EtOAc, 33
66%) afforded 11 mg
of 24 (21%, Rf: 0.66, hexane–EtOAc, 1 : 2) and 33 mg of 25
(62%, Rf: 0.43, hexane–EtOAc, 1 : 2). [α]2D3 = Ϫ5 (c 0.2, CH2Cl2);
δH (CDCl3) 7.4 (15H, br s); 5.18 (2H, s, OCH2Ph); 5.11 (2H, m,
POCH2Ph); 5.08 (2H, m, POCH2Ph); 4.12 (1H, 1/2 ABX,
CHCH2OP); 3.94 (1H, 1/2 ABX, CHCH2OP); 3.73 (1H, m,
CHOMe); 3.53 (1H, m, CHOMe); 3.41 (3H, s, OCH3); 3.36
(3H, s, OCH3); 2.76 (1H, m, CHMe); 1.62 (2H, m, CHCH2CH);
1.2 (3H, d, J 7.1, CH3CH); δP (CDCl3) ϩ 0.46; m/z (ESϩ) 579
(M ϩ Na); m/z (ES) 557.2305 (M ϩ H. C30H38O8P requires
557.2304).
(2R,3S,5S)-6-Bis(benzyloxy)phosphoryloxy-5-hydroxy-3-meth-
oxy-2-methylhexanoic acid benzyl ester (25)
δH (CDCl3) 7.4 (15H, br s); 5.19 (2H, s, OCH2Ph); 5.12 (2H, m,
POCH2P); 5.09 (2H, m, POCH2P); 4.14 (2H, m, CHCH2OP
and CHOH); 3.98 (1H, m, CHCH2OP); 3.66 (1H, m, CHOMe);
3.43 (3H, s, OCH3); 2.6 (1H, m, CHMe); 1.57 (2H, m,
CHCH2CH); 1.25 (3H, d, J 7.2, CH3CH); δC (CDCl3) 176.3 (s);
136.4 (s); 136.3 (s); 129.2 (2 × d); 129.1 (d); 129.0 (d); 128.9 (d);
128.4 (d); 69.9 (2 × t); 69.0 (t); 68.9 (d); 67.0 (t); 59.0 (d); 58.8
(q); 45.5 (d); 36.0 (t); 11.8 (q); δP (CDCl3) ϩ 0.57; m/z (ESϩ) 565
(M ϩ Na); m/z (ES) 543.2153 (M ϩ H. C29H36O8P requires
543.2148).
(2R,3S,5S)-3,5-Diacetoxy-2-methyl-6-phosphonoxyhexanoic
acid (29)
Hydrogenolysis of 23 (34 mg, 0.055 mmol) with 10% Pd–C
(10 mg) for 16 h afforded 29 as a colourless oil. [α]2D0 = Ϫ16.7
(c 0.24, MeOH); δH (CD3OD) 5.25 (1H, br); 5.05 (1H, br); 4.0
(2H, br m); 2.65 (1H, br m, CHMe); 2.08 (3H, s, CH3CO); 2.03
(3H, s, CH3CO); 2.0 (2H, br, CHCH2CH); 1.15 (3H, d, J 6.9,
CH3CH); δC (CD3OD) 177.4 (s); 172.8 (s); 172.7 (s); 71.8 (d);
70.3 (d); 68.4 (t); 45.4 (d); 34.4 (t); 21.4 (q); 21.2 (q); 13.3 (q); δP
(CD3OD) ϩ 1.76; m/z (ESϪ) 341 (M Ϫ H); m/z (ESϪ) 341.0642
(M Ϫ H. C11H18O10P requires: 341.0638).
(2R,3S,5S)-3,5-(Benzylidenedioxy)-6-bis(benzyloxy)phosphoryl-
oxy-2-methylhexanoic acid benzyl ester (26)
A solution of 21 (90 mg, 0.17 mmol); PPTS (171 mg, 0.68
mmol) and benzaldehyde dimethyl acetal (0.25 cm3, 0.68 mmol)
in DMF (2 cm3) was stirred for 24 h at rt. At this time, the
reaction was not complete and was then heated at ca. 40 ЊC for
3 h. The reaction was diluted with EtOAc, washed successively
with 5% NaHCO3 and brine, then dried (MgSO4) and con-
centrated. Flash chromatography (gradient 0 33% EtOAc–
hexane) afforded 26 as a mixture of 2 diastereoisomers (colour-
less oil, 74 mg, 71%). Rf: 0.61 (50% EtOAc–hexane); δH (CDCl3)
d 7.55–7.3 (40H, m); 5.85 (1H, s, CHPh); 5.65 (1H, s, CHPh);
5.2 (4H, s, OCH2Ph); 5.15–5.05 (8H, m, POCH2Ph); 4.63 (1H,
m); 4.5–4.3 (2H, m); 4.3–3.97 (5H, m); 3.38 (1H, m, CHMe);
2.71 (1H, quint, J 7, CHMe); 2.2–1.95 (2H, m); 1.6–1.4 (2H, m);
1.41 (3H, d, J 6.9, CH3CH); 1.37 (3H, d, J 7, CH3CH); δC
Acknowledgements
We gratefully acknowledge the Wellcome Trust for financial
support and the EPSRC National Mass Spectrometry Service
Centre for accurate mass spectral data.
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