Synthesis of phosphono analogues of dihydroxyacetone phosphate and glyceraldehyde 3-phosphate

Article abstract of DOI:10.1016/S0968-0896(99)00065-6

The present paper describes the synthetic routes of six phosphono analogues of dihydroxyacetone phosphate and five phosphono analogues of glyceraldehyde 3-phosphate through α-, β- and γ-hydroxyphosphonate esters precursors containing a protected carbonyl

Full text of DOI:10.1016/S0968-0896(99)00065-6

Bioorganic & Medicinal Chemistry 7 (1999) 1403±1412
Synthesis of Phosphono Analogues of Dihydroxyacetone
Phosphate and Glyceraldehyde 3-Phosphate
Patrick Page, Casimir Blonski* and Jacques Perie
Â
Groupe de Chimie Organique Biologique, UMR 5623, Universite Paul Sabatier, Bat II R1, 118 route de Narbonne, 31062 Toulouse
Ã
Cedex, France
Received 5 November 1998; accepted 18 January 1999
AbstractÐThe present paper describes the synthetic routes of six phosphono analogues of dihydroxyacetone phosphate and ®ve
phosphono analogues of glyceraldehyde 3-phosphate through a-, b- and g-hydroxyphosphonate esters precursors containing a
protected carbonyl group. In some situations, depending on the sequence used for the deprotection of the phosphonate and carbonyl
groups, the aldol/ketol rearrangement allowed the synthesis of either dihydroxyacetone phosphate or glyceraldehyde 3-phosphate
analogues from the same precursors. All these analogues are of interest both as active-site probes and as potential substrates for
glycolytic enzymes such as fructose 1,6-diphosphate aldolases (EC 4.1.2.13). # 1999 Elsevier Science Ltd. All rights reserved.
Introduction
phosphorus-containing nucleophiles in presence of
BF3 OEt2,13,14 or the addition of the latter to an aldehyde
.
Glycolytic enzymes fructose-1,6-diphosphate aldolases
reversibly catalyze the production of d-fructose-1,6-
diphosphate (FDP) from d-glyceraldehyde 3-phosphate
(G-3-P) and dihydroxyacetone phosphate (DHAP)
(Scheme 1).1,2 The enzyme from rabbit muscle (EC
4.1.2.13), currently in use for carbohydrate synthesis,3
accepts a large variety of aldehydes as substrates, but is
rather selective toward DHAP since only limited struc-
tural modi®cations are accepted.4 In addition to the
synthetic purposes in organic chemistry, aldolases are
also considered as targets for the development of new
antiparasitic drugs since glycolysis is the only source of
energy for parasites such as trypanosomes (bloodstream
form).5 Among aldolase inhibitors,6 those correspond-
ing to substrate analogues are of interest as active-site
probes of the enzyme and for the design of a new class
of inhibitors.7±10
derivative, allowing the formation of a carbon±phos-
phorus bond of the a-, b- and g-hydroxy phosphonate
ester precursors. Furthermore, considering that DHAP
and G-3-P are isomeric, their phosphono analogues are
quantitatively prepared from the same precursor by
taking advantage of the a-ketol rearrangement.15
Besides their speci®c interest for the study of aldolases
and other glycolytic enzymes (triose phosphate iso-
merase, glycerol 3-phosphate dehydrogenase...), these
analogues are potential substrates of aldolases, hence
leading, in addition, to other analogues.16±18
Results
Syntheses of DHAP analogues (7a,b and 9a,b) from
phosphonate esters 4a and 4b
Phosphono analogues of natural phosphates display
modi®ed chemical and biological properties, and are of
high interest to biology and medicine.11,12 Along these
lines, the synthesis of six phosphono analogues of
DHAP and ®ve phosphono analogues of G-3-P can be
reported. The two key reactions consist of a regio-
speci®c ring opening of two appropriate epoxides by
As shown in Scheme 2, ketalization,19,20 dehydro-
halogenation,20,21 followed by quantitative epoxida-
tion22 of the starting 2-chlorobutanone gave epoxide 3.
The ring opening of the latter compound by phosphite
.
or phosphonate anion in presence of BF3 OEt2 yielded
b- and g-hydroxyphosphonate esters 4a and 4b. This
reaction also produced the formation of the by-products
40a or 40b, resulting from solvent (THF) insertion,13,14
more especially with the phosphite anion, where 40a
represents about 50% of the products.23 Oxidation of 4a
and 4b according to the P®tzner±Mo€at method24 pro-
duced ketones 5a and 5b.25 Catalytic hydrogenation
Key words: Phosphonic acids and derivs; isosteres; substituent e€ects;
enzymes and enzyme reactions.
* Corresponding author. Tel.: 33-5-6155-6486; fax: 33-5-6125-1733.
0968-0896/99/$ - see front matter # 1999 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(99)00065-6

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P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
Scheme 1. Reaction catalyzed by aldolase.
Scheme 2. Synthetic scheme for the synthesis of compounds 9a,b and 7a,b. (a) HOCH2CH2OH, TsOH,benzene, 82%; (b) KOH, HOCH2CH2OH,
130 ꢀC (82%); (c) m-CPBA, CH2Cl2 (98%); (d) (BnO)2P(O)H or (BnO)2P(O)CH3, n-BuLi, Et2O BF3, THF, 80 ꢀC (n=1: 40%, n=2: 75%); (e)
.
DCC, pyridine, DMSO, TFA (n=1: 79%, n=2: 85%); (f) 1. H2, Pd/C, MeOH; 2. Ba(OH)2 (n=1: 92%, n=2: 97%); (g) DOWEX 50WX8 (H+),
H2O; (h) 1. DOWEX 50WX8 (H+), H2O; 2. Ba(OH)2 (n=1: 97%, n=2: 97%).

P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
1405
removed the benzyl protecting group to give the corres-
ponding phosphonic acids 6a, 6b, 8a and 8b, which
were isolated as barium salts. Subsequently, deketaliza-
tion led to a-hydroxyketone phosphonates 7a and 7b,
and a-diketophosphonic acids 9a and 9b. The attempts
to isolate the two latter compounds as barium or
sodium salts failed, and gave decomposition products
due likely to possible reaction of the formed phospho-
nate anion on one of the two dicarbonyl groups. How-
ever, 9a and 9b could be stored without decomposition
in an aqueous acidic medium.
deketalization (step g) yielded a-hydroxyaldehyde
phosphonates 16a±c. In contrast, the deprotection of
the carbonyl group of 13b and 13c carried out in more
acidic conditions (step i) gave a-hydroxyketonephos-
phonate esters 19a and 19b analogues of DHAP.33±35
The reversibility of this reaction has been previously
demonstrated33 and can be rationalized in terms of an
a-ketol rearrangement.15 In its turn, deprotection of the
phosphonate esters 19a and 19b provided the corres-
ponding phosphonic acid analogues of DHAP.33
Relative reactivities of G-3-P and DHAP analogues with
DHAP and G-3-P, respectively
Analogues of G-3-P (16a±c and 18a,b) and of DHAP
(19a,b) from phosphonate esters 13a±d
Phosphonic analogues of G-3-P (16a±c and 18a,b) and
DHAP (7a,b and 19a,b) have been tested as substrates
with rabbit muscle aldolase. The progress of the aldose-
catalyzed condensation was followed as previously
described4,18 and the initial relative rates (Vrel) for the
di€erent reactions expressed as a percentage of the
initial relative velocity with the natural substrates G-3-P
or DHAP, are given in Table 1.
Several methods were examined to prepare key epoxide
(‹) glycidaldehyde diethyl acetal 11 (vide infra) from
acroleine diethyl acetal (see Scheme 3): (a) oxidation by
m-chloroperbenzoic acid24 was not quantitative, due
likely to the poor stability of epoxide 11 in acidic med-
ium, (b) in contrast, the reaction carried out in neutral
conditions with hydrogen peroxide in presence of ben-
zonitrile26 gave a fair yield epoxide, (c) this epoxide
could also be obtained in two steps with diol 10 as an
intermediate,27 (d) through 3-O-mesylate27 or 10, or (e)
using the Mitsunobu reaction.18,28
Acceptable relative rates are obtained with the aldehyde
derivatives and these substrates are applicable to a
large-scale synthesis. In contrast, the data obtained with
the DHAP analogues show that only the phosphonate
isostere of DHAP (19b) is substrate for aldolase.17
These results represent new examples that con®rm the
high selectivity of muscle aldolase toward the DHAP
structure for the enzyme-catalyzed aldol condensation
as previously suggested by other works.4,9,16
As shown in Scheme 4, the b- and g-hydroxypho-
sphonate esters 13a and 13b were obtained by ring
opening of the epoxide 11, respectively, by phosphite
13,14
.
and phosphonate anion catalyzed by BF3 OEt2.
As
for compound 4a, the 13a yield was signi®cantly low-
ered because of a side reaction with the solvent. Never-
theless, its parent phosphonate ester 13c could be
quantitatively obtained in another way using aldehyde
12 (easily formed from glyoxal29) and lithium diethyl
methylphosphonate carbanion. Additionally, this alde-
hyde reacted also with diethylphosphite anion to give
the a-hydroxyphosphonate ester 13d also in high yield.
Oxidation (step d), deprotection of the phosphonate
moiety with trimethylsilyl bromide (step f),30 followed
by deketalization (step h) of phosphonate esters 13a and
13b yielded G-3-P analogues 18a and 18b.31 Deprotec-
tion of the phosphonate group of compounds 13b±d
(step e) in basic conditions32 gave the corresponding
phosphonic acids 15a±c as lithium salts; subsequently
Discussion
Whereas the access to DHAP analogues through diazo-
ketones is well documented,8,9,27 the syntheses of phos-
phonate analogues is more limited. The present work
supplements this route with, as a key step, the formation
of a-, b- and g-hydroxyphosphonate ester intermediates
(4a,b and 13a±d), which allow both DHAP and G-3-P
analogues.
Regiospeci®c ring opening of epoxides 3 and 11 by
phosphite or methane phosphonate anions in presence
.
of BF3 OEt2 furnished most of the intermediates. How-
ever, we observed solvent participation in the progress
of the reaction, more especially when phosphite anion
was used, and consequently 4a and 13a yields were
lowered. Conversely, we found the use of aldehyde
derivative 12 to be superior to that of epoxide 11 as a
nucleophile acceptor leading to the formation of inter-
mediates 13c and 13d, quantitatively.
The scope of these syntheses is enlarged by the oppor-
tunity given with the a-ketol rearrangement that allows,
depending on the deprotection conditions, access either
to DHAP or to G-3-P analogues from the same 13b and
13c intermediates. Moreover, some DHAP (7a, 9a and
19a) and G-3-P (16a, 16c and 18a) analogues described
here are only possible or stable as phosphonate and
have no equivalent in the phosphate series (e.g. 7a,
Scheme 3. Synthetic scheme for the synthesis of glycidaldehyde diethyl
acetal 11. (a) m-CPBA, CH2Cl2, 0 ꢀC to rt (5±10%); (b) H2O2, PhCN,
KHCO3, MeOH (70%); (c) KMnO4, H2O (54%); (d) 1. CH3SO2Cl,
pyridine; 2. NaOH, H2O (56%); (e) DEAD, Ph3P, benzene (77%).

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P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
.
Scheme 4. Synthetic scheme for the synthesis of compounds 16a±c, 18a,b and 19a,b. (a) (EtO)2P(O)H or (EtO)2P(O)CH3, n-BuLi, Et2O BF3, THF,
80 ꢀC (n=1: 46%, n=2: 80%); (b) n-BuOH, hexane, DOWEX 50WX8 (H+), re¯ux (62%); (c) (EtO)2P(O)CH3 or (EtO)2P(O)H, n-BuLi, THF,
80 ꢀC (n=1: 93%, n=0: 77%); (d) DCC, pyridine, DMSO, TFA, benzene (n=1: 76%, n=2: 89%); (e) LiOH, NaBH4, H2O, 120 ꢀC, 2 Bar (n=1:
90%, n=2: 96%, n=0: 92.5%); (f) 1. Me3SiBr, 24 h, 30 ꢀC; 2. H2O, Ba(OH)2 (n=1: 92%, n=2: 96%); (g) 1. DOWEX 50WX8 (H+), H2O; 2.
NaOH or LiOH (n=1: 96%, n=2: 96%, n=0: 91%); (h) DOWEX 50WX8 (H+), H2O; (i) HCl 0.1 M, 45 ꢀC (n=1: 54%, n=2: 70%).
which presents a speci®c interest by its reversed struc-
ture, compared to that of DHAP). This is of particular
signi®cance for synthetic purposes, as well as for
mechanistic studies of FDP-aldolases and possibly other
glycolytic enzymes that operate with DHAP or G-3-P as
substrates. The same routes can be used to prepare
chiral compounds starting from chiral epoxides such as
d-glycidaldehyde and diethylacetal obtained from fruc-
tose18,26 or d-3-chloro-2-hydroxypropanal.36
Experimental
Materials and methods
Chemicals and solvents were reagent grade. The NMR
spectra were recorded in CDCl3 or D2O on a Bruker
1
AC80 (80-MHz H NMR) or a Bruker AC200 (200-
1
MHz H NMR, 50-MHz 13C NMR and 81-MHZ 31P
NMR) spectrometer. All chemical shifts are reported in

P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
1407
17.2 Hz, 1H); 13C NMR (CDCl3) d 24.6, 64.5, 107.4,
114.8, 136.4. Anal. calcd for C6H10O2: C, 63.1; H, 8.83;
O, 28.1. Found: C, 62.9; H, 8.88; O, 28.5.
Table 1. Relative reactivities of G-3-P and DHAP analogues with
DHAP and G-3-P, respectively, in rabbit muscle aldolase-catalyzed
aldol condensations
a
Compounds
Vrel
Substrate classb
(‹)-2-Methyl-2-oxiranyl-1,3-dioxolane (3). A mixture of
2 (3.1 g, 0.27 mmol) and m-chloroperbenzoic acid (25 g,
0.144 mmol) in 200 mL of CH2Cl2 was stirred at room
temperature for 5 days. Saturated NaHCO3 (100 mL)
was added to the mixture which was cooled to 0 ꢀC. The
aqueous layer was separated and extracted with CH2Cl2
(3Â200 mL). The organic fractions were combined,
washed with 3% Na2S2O3, saturated NaHCO3 and
water, dried over anhydrous MgSO4, and evaporated
under reduced pressure. The remaining residue was
puri®ed by ¯ash distillation (80 ꢀC, 5 mm Hg) to yield 3
as a colourless oil (3.45 g, 98%). 1H NMR (CDCl3)
d 1.29 (s, 3H), 2.59 (d, J=3.4 Hz, 2H), 2.93 (t, J=
3.4 Hz, 1H), 3.8±3.9 (m, 4H); 13C NMR (CDCl3) (21.8,
43.75, 54.9, 65.5, 66.0, 106.6. Anal. calcd for C6H10O3:
C, 55.3; H, 7.74; O, 36.9. Found: C, 55.3; H, 7.6; O,
36.7.
G-3-P
16a
16b
16c
18a
100c
36
41
7
26
28
+++
+++
+++
+
+++
+++
18b
DHAP
7a
7b
19a
19b
100
+++
Ð
Ð
Ð
+
<0.01
<0.01
<0.01
6.1 (10)d
a Reactivities were measured in 0.1 M triethanolamine bu€er (pH 7.0,
25 ꢀC) containing both substrates at 50 mM concentration.
b Substrate class according to the scale previously used: (+++) Vrel
>25; (++) 25>Vrel>1; (+) 10>Vrel>1; (Ð) Vrel<1.
c Vrel for the D stereoisomer.
d Value from literature.4,16
1
parts per million with respect to TMS for H and 13C
(‹)-Dibenzyl [2-(2-methyl-1,3-dioxolan-2-yl)-2-hydroxy-
ethyl]-phosphonate (4a). A 1.6 M solution of n-BuLi in
hexane (19 mL, 30.4 mmol) was added dropwise to a
stirred solution of dibenzylphosphite (6.0 g, 23 mmol) in
dry THF (60 mL) at 80 ꢀC. After 30 min of stirring,
the mixture was added dropwise to a stirred solution of
3 (1 g, 7.68 mmol) in dry THF (40 mL) at 80 ꢀC. After
spectra and H2PO4 for 31P as internal standards. Ele-
mental analyses were performed by the Ecole Nationale
Superieure de Chimie de Toulouse. Flash chromato-
graphy was performed on a Merck Geduran SI 60
(0.040±0.063 mM).
.
15 min of stirring, BF3 OEt2 (3.8 mL, 30.9 mmol) was
Enzymatic aldol condensations and kinetic measurements
slowly introduced. The reaction mixture was stirred for
2 h at 70 ꢀC, then overnight at room temperature.
Saturated NH4Cl (20 mL) was added, the solvents were
removed under reduced pressure and the remaining
residue dissolved in 100 mL of ethyl acetate. The
organic solution was washed with brine, dried over
MgSO4, then concentrated and puri®ed by ¯ash chro-
matography (CH2Cl2:MeOH, 98:2 to 96:4) to provide
4a as a colourless oil (1.2 g, 40%). 31P NMR (CDCl3) d
Enzymatic assays and kinetic measurements were car-
ried out by the procedure previously described.4,18
Chemical syntheses
(‹)-2-(1-Chloro-ethyl)-2-methyl-1,3-dioxolane (1). To a
round-bottomed ¯ask equiped with a Dean±Stark system
and containing 30 g (0.24 mol) of 3-chloro-2-butanone in
400 mL of benzene, were added 15 g (0.325 mol) of eth-
ylene glycol and 1 g (5.74 mmol) of p-toluene sulfonic
acid monohydrate. The mixture was re¯uxed and stirred
for 20 h and the solvent removed under reduced pres-
sure. The residue was dissolved in CH2Cl2 (250 mL) and
the solution washed with saturated NaHCO3 and brine.
The organic layer was dried (Na2SO4) and concentrated.
The remaining oil was distilled at 43 ꢀC (1 mm Hg) to
give 1 as a colourless oil (30.1 g, 83%). 1H NMR
(CDCl3) d 1.39 (s, 3H), 1.48 (d, J=6.8 Hz, 3H), 3.94 (q,
J=6.8 Hz, 1H) 3.98 (s, 4H); 13C NMR (CDCl3) d 20.0,
20.1, 60.9, 65.4, 65.6, 110.1. Anal. calcd for C6H11O2Cl:
C, 47.8; H, 7.36; O, 21.2. Found: C, 47.6; H, 7.4; O,
20.9.
1
32.0; H NMR (CDCl3) d 1.27 (s, 3H), 1.5±1.9 (m, 2H),
3.1 (br s, 1H, D2O exchangeable), 3.8±4.0 (m, 5H), 4.9±
5.2 (m, 4H), 7.33 (s, 10H); 13C NMR (CDCl3) d 19.7,
28.7 (d, 1JCP=142 Hz), 65.1, 65.5, 67.4, 67.6, 70.2, 110.8
3
(d, JCP=19.2 Hz), 128.4, 128.7, 136.3. Anal. calcd for
C20H25O6P: C, 61.2; H, 6.38. Found: C, 61.4; H, 6.3.
(‹)-Dibenzyl [3-(2-methyl-1,3-dioxolan-2-yl)-3-hydroxy-
propyl]-phosphonate (4b). Compound 4b was prepared
from 3 (1 g, 7.68 mmol) in 75% yield (2.34 g) by follow-
ing the same procedure described for 4a, using diben-
zylmethylphosphonate (6.4 g, 23 mmol) in place of
1
dibenzylphosphite. 31P NMR (CDCl3) (34.1; H NMR
(CDCl3) d 1.24 (s, 3H), 1.5±2.3 (m, 4H), 2.5 (m, 1H,
D2O exchangeable), 3.5 (m, 1H), 3.92 (m, 4H), 4.9±5.2
(m, 4H), 7.33 (s, 10H); 13C NMR (CDCl3) d 19.58, 22.7
2-Methyl-2-vinyl-1,3-dioxolane (2). Compound 1 (5 g,
33.2 mmol) was added dropwise to a stirred solution of
KOH (12 g, 0.214 mol) in ethylene glycol (24 mL) at
125±130 ꢀC. After 5 h of warming, the mixture was pur-
i®ed by distillation at 110±112 ꢀC (760 mm Hg) to yield
2 as a colourless oil (3.1 g, 82%). IR (®lm) n(CˆC)
1
(d, JCP=142 Hz), 24.2, 65.1, 65.3, 67.2, 74.9 (d,
3JCP=14.3 Hz), 110.2, 127.6, 128.2, 136.5. Anal. calcd
for C21H27O6P: C, 62.06; H, 6.65; O, 23.6. Found: C,
62.4; H, 6.62; O, 23.8.
1
cm 1: 1675. H NMR (CDCl3) d 1.44 (s, 3H), 3.9±4.0
Dibenzyl [2-(2-methyl-1,3-dioxolan-2-yl)-2-oxo-ethyl]-
phosphonate (5a). To a stirred mixture of dicyclohexyl-
carbodiimide (0.95 g, 4.59 mmol) and dry pyridine
(m, 4H), 5.12 (dd, J=1.7 Hz, J=10.5 Hz, 1H), 5.45 (dd,
J=1.7 Hz, J=17.2 Hz, 1H), 5.8 (dd, J=10.5 Hz, J=

1408
P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
(0.18 mL, 2.22 mmol) in dry benzene (20 mL) was added
dropwise 4a (0.40 g, 1.02 mmol) in dry DMSO (3 mL)
under nitrogen atmosphere. Tri¯uoroacetic acid
(0.090 mL, 1.1 mmol) was added and the reaction mix-
ture was stirred overnight at room temperature. The
solvent was removed under reduced pressure and the
remaining residue dissoved in 60 mL of ether. The sus-
pension (dicyclohexylurea) was ®ltered o€ and the ®l-
trate washed with brine (3Â30 mL). The organic layer
was dried over MgSO4 and concentrated under reduced
pressure. The residue was chromatographed (CH2Cl2:
MeOH, 95:5) to yield 5a as a colourless oil (0.30 g,
(‹)-2-Hydroxy-3-oxo-butyl-1-phosphonic acid, barium
salt (7a). Salt 6a (0.050 g, 0.144 mmol) was treated with
0.5 mL of DOWEX 50 WX8 (H+) in 0.5 mL of water
for 10 min. The resin was discarded by centrifugation
and washed with water (3Â0.25 mL). The aqueous
solutions were combined and incubated for 4 days at
room temperature. The progress of hydrolysis was
monitored by 31P NMR. The pH was adjusted to 7.4
with saturated Ba(OH)2 and the solution was freeze-
dried. The resulting powder was washed with ether and
dried in vacuo to give 7a as a white powder (0.045 g,
1
97%). 31P NMR (D2O) d 18.2; H NMR (D2O) d 1.5±
75%). 31P NMR (CDCl3) d 21.3; H NMR (CDCl3) d
1.9 (m, 2H), 2.1 (s, 3H), 3.3±3.5 (m, 1H); 13C NMR
1
1.41 (s, 3H), 3.26 (d, 2JHP=22 Hz, 2H), 3.8±4.0 (m, 4H),
5.0±5.1 (m, 4H), 7.34 (s, 10H); 13C NMR (CDCl3) d
(D2O) d 30.9, 34.3 (d, JCP=131 Hz), 76.8, 209.0 (d,
1
3JCP=14.9 Hz). Anal. calcd for C4H7O5PBa: C, 15.8; H,
2.3; O, 26.4. Found: C, 15.6; H, 2.2; O, 26.8.
1
20.32, 35.88 (d, JCP=135 Hz) 65.62, 68.0, 107.6 (d,
3JCP=5 Hz), 128.4, 128.6, 136.0, 199.6. Anal. calcd for
C20H23O6P: C, 61.5; H, 5.89; O, 24.6. Found: C, 61.7;
H, 5.8; O, 24.8.
(‹)-3-Hydroxy-4-oxo-pentyl-1-phosphonic acid, barium
salt (7b). Compound 7b was obtained from 6b (0.100 g,
0.277 mmol) in 97% yield (0.090 g) by following the
same procedure described for 7a. 31P NMR (D2O) d
Dibenzyl [3-(2-methyl-1,3-dioxolan-2-yl)-3-oxo-propyl]-
phosphonate (5b). Compound 5b was prepared from 4b
(0.90 g, 2.2 mmol) in 78% yield (0.70 g) by following the
same procedure described as for 5a, using 1.37 g
(6.66 mmol) of dicyclohexylcarbodiimide and 4 mL of
dry DMSO. 31P NMR (CDCl3) d 32.7; 1H NMR
(CDCl3) d 1.28 (s, 3H), 1.8±2.1 (m, 2H), 2.6±2.8 (m, 2H)
3.7±4.0 (m, 4H), 4.8±5.1 (m, 4H) 7.25 (s, 10H); 13C
1
24.45; H NMR (D2O) d 1.5±2.0 (m, 4H), 2.08 (s, 3H),
1
3.2±3.5 (m, 1H); 13C NMR (D2O) d 26.31 (d, JCP
=
133 Hz), 28.2, 29.9, 79.9 (d, 3JCP=15.2 Hz), 217.5. Anal.
calcd for C5H9O5PBa: C, 18.9; H, 2.8; O, 25.2. Found:
C, 18.7; H, 2.78; O, 25.4.
[2-(2-Methyl-1,3-dioxolan-2-yl)-2-oxo-ethyl]-phosphonic
acid, barium salt (8a). Compound 8a was obtained
from 5a (0.200 g, 0.513 mmol) in 79% yield by following
the same procedure as described for 6a. 31P NMR
1
NMR (CDCl3) d 19.37 (d, JCP=144 Hz), 20.6, 29.5,
65.5, 67.4, 107.5, 127.9, 128.5, 136.2, 205.2 (d, JCP=
3
14.3 Hz). Anal. calcd for C21H25O6P: C, 62.37; H, 6.19;
O, 23.76. Found: C, 62.39; H, 6.25; O, 23.9.
1
(D2O) d 12.0; H NMR (D2O) d 1.46 (s, 3H), 3.1 (d,
2JPH=20 Hz, 2H) 3.8±4.1 (m, 4H); 13C NMR (D2O) d
22.6, 39.7 (d, JCP=134 Hz), 67.6, 111.0 (d, JCP=
1
3
(‹)-[2-(2-Methyl-1,3-dioxolan-2-yl)-2-hydroxy-ethyl]-phos-
phonic acid, barium salt (6a). To a suspension of Pd:C
(10%, 50 mg) in a solution of water:EtOH (1:1, 20 mL),
was added 4a (0.10 g, 0.255 mmol). The mixture was
degassed and hydrogenated for 12 h at atmospheric
pressure. The catalyst was ®ltered o€ and the pH was
adjusted to 7.6 with saturated Ba(OH)2. The solution
was then freeze-dried, and the residue dissolved in 8 mL
of distilled water. The suspension was discarded by
centrifugation, barium salt 6a was precipitated by addi-
tion of ethanol (24 mL) and the resulting mixture was
kept at 0 ꢀC for 3 h. The salt was collected by cen-
trifugation, washed twice with ethanol (80%, then
absolute), ethyl ether and dried in vacuo to yield 6a
(0.082 g, 92%). 31P NMR (D2O) d 21.2; 1H NMR (D2O)
d 1.31 (s, 3H), 1.5±1.9 (m, 2H), 3.8±4.0 (m, 1H), 4.02 (s,
4H); 13C NMR (D2O) d 21.9, 34.0 (d, 1JCP=135Hz), 67.6,
67.9, 73.7, 109.1. Anal. calcd for C6H11O6PBa: C, 20.75;
H, 3.17; O, 27.66. Found: C, 20.6; H, 3.26; O, 27.75.
10.0 Hz), 201.4. Anal. calcd for C6H9O6PBa: C, 20.87;
H, 2.6; O, 27.82. Found: C, 21.0; H, 2.62; O, 27.65.
[3-(2-Methyl-1,3-dioxolan-2-yl)-3-oxo-propyl]-phosphonic
acid, barium salt (8b). Compound 8b was obtained from
5b (0.190 g, 0.47 mmol) in 85% yield (0.150 g) by fol-
lowing the same procedure described for 6a. 31P NMR
1
(D2O) d 23.1; H NMR (D2O) d 1.48 (s, 3H), 1.5±1.7
(m, 2H), 2.65 (m, 2H), 3.85±4.1 (m, 4H); 13C NMR
1
(D2O) d 22.9, 24.4 (d, JCP=132 Hz), 34.6, 68.0, 110.6,
212.0 (d, JCP=14.0 Hz). Anal. calcd for C7H11O6PBa:
C, 23.4; H, 3.06; O, 26.76. Found: C, 23.6; H, 3.12; O, 26.9.
3
2,3-Dioxo-butyl-1-phosphonic acid (9a). Salt 8a (0.140 g,
0.405 mmol) was treated with 1 mL of DOWEX 50
WX8 (H+) in 1 mL of water for 10 min. The resin was
discarded and washed three times with water (0.50 mL).
The aqueous solutions were combined and incubated
for 7 days at 100 ꢀC. The progress of the hydrolysis was
monitored by 31P and 13C NMR. The title compound,
did not show decomposition, was used for enzymatic
studies without further puri®cation. 31P NMR
(D2O+H2O) d 16.1; 13C NMR (D2O+H2O) d 25.8,
38.5 (d, 1JCP=129 Hz, CH2-P), 90.6 (C2, hydrated form),
(‹)-[3-(2-Methyl-1,3-dioxolan-2-yl)-3-hydroxy-propyl]-
phosphonic acid, barium salt (6b). Compound 6b was
prepared from 4b (0.15 g, 3.69 mmol) in 90% yield
(0.120 g) by following the same procedure described for
1
6a. 31P NMR (D2O) d 23.5; H NMR (D2O) d 1.30 (s,
3H), 1.35±2.0 (m, 4H), 3.5 (m, 1H), 4.0 (s, 4H); 13C
208.2 (d, JCP=23.0 Hz).
3
1
NMR (D2O) d 21.75, 28.58 (d, JCP=131 Hz), 28.68,
67.4, 77.9 (d, JCP=14.9 Hz), 113.3. Anal. calcd for
3
3,4-Dioxo-pentyl-1-phosphonic acid (9b). Compound 9b
was prepared from 8b by following the same procedure
as described for 9a. 31P NMR (D2O+H2O) d 26.5; 13C
C7H13O6PBa: C, 23.37; H, 3.6; O, 26.6. Found: C, 23.4;
H, 3.55; O, 26.4.

P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
1409
NMR (D2O+H2O) d 22.7 (d, 1JCP=139 Hz), 26.3, 32.9,
98.9 (d, JCP=21.0 Hz, C3, hydrated form), 209.0.
added KHCO3 (3 g, 30 mmol), benzonitrile (20.4 g,
200 mmol) then 30% hydrogen peroxide (23.4 g,
206 mmol). The reaction mixture was stirred for 15 h at
room temperature then 4 h at 40 ꢀC. Water (75 mL) was
added and most of the methanol was removed under
reduced pressure. The aqueous mixture was extracted
with CH2Cl2 (4Â100 mL), the combined organic layers
dried over MgSO4 and concentrated under reduced
pressure. The resulting residue was extracted with
60 mL of hexane, the insoluble product (benzamide) ®l-
tered out and the solvent concentrated. The residue was
puri®ed by ¯ash distillation to give 11 (19.6 g, 70%). 1H
NMR (CDCl3) d 1.29 and 1.33 (2t, J=7 Hz, 2 CH3),
2.53 (m, 2H, CHCH2), 2.86 (m, 1H, CH O), 3.2±3.6
(m, 4H, CH2CH3) 4.1 (d, J=4.3 Hz, 1H, CH); 13C
NMR (CDCl3) d 15.0, 43.5, 51.6, 62.0, 62.6, 101.4.
Anal. calcd for C7H14O3: C, 57.52; H, 9.65; O, 32.87.
Found: C, 57.1; H, 9.82; O, 33.24.
3
(‹)-Glyceraldehyde diethyl acetal (10). To a stirred
solution of acroleine diethylacetal (10 g, 77 mmol) in
85 mL of distilled water was added dropwise 13 g
(84.6 mmol) of KMnO4 in 300 mL of water over 1.5 h.
The reaction mixture was stirred for 3 h at room tem-
perature then re¯uxed for 1 h. After cooling, the pre-
cipitate was ®ltered o€ and K2CO3 (170 g) was added to
the ®ltrate and the aqueous solution was extracted with
ethyl acetate (3Â100 mL). The organic layer was dried
over MgSO4 and evaporated under reduced pressure.
The remaining residue was puri®ed by ¯ash chromato-
graphy (CH2Cl2:MeOH, 95:5) to give 10 as a colourless
1
oil (6.9 g, 54%). H NMR (CDCl3) d 1.17 (s, 6H), 2.6
(m, 2H, D2O exchangeable), 3.4±3.9 (m, 7H, CH O,
3
CH2O), 4.44 (d, JHH=7.0 Hz, 1H, O CH O); 13C
NMR (CDCl3) d 15.3, 62.5, 63.6, 64.2, 71.8, 103.4.
Anal. calcd for C7H16O4: C, 51.21; H, 9.82; O, 39.02.
Found: C, 51.17; H, 10.01; O, 39.20.
Di n-butoxy-2,2 ethanal (12). A mixture of 40% glyoxal
(36.25 g, 0.25 mol), 230 mL of butanol, 325 mL of hex-
ane and 25 g of DOWEX 50 WX8 (H+) was re¯uxed
for 2 h in a round-bottomed ¯ask equipped with a
Dean±Stark system. The mixture reaction was cooled to
room temperature, the resin ®ltered o€ and NaHCO3
(12.5 g) added. After standing for 30 min, the precipitate
was ®ltered o€ and the solvent removed under reduced
pressure. The resulting residue was puri®ed by ¯ash
distillation at 70 ꢀC (2 mm Hg) to yield 12 as a colour-
less oil (29.1 g, 62%). 1H NMR (CDCl3) d 0.89 (t,
J=7 Hz, 2 CH3), 1.5 (m, 8H, CH2), 3.6 (m, 4H, CH2O),
4.5 (d, J=2.1 Hz, 1H, O CH O), 9.4 (d, J=2.1 Hz,
1H, CHO); 13C NMR (CDCl3) d 13.8, 19.2, 31.8, 67.3,
101.2, 196.9. Anal. calcd for C10H20O3: C, 63.83; H,
10.64; O, 25.53. Found: C, 64.0; H, 10.72; O, 25.41.
(‹)-Glycidaldehyde diethyl acetal (11). Three di€erent
methods were used to obtain the title compound.
Method A: To a stirred solution of acetal 10 (2.25 g,
13.7 mmol) in dry pyridine (10 mL) cooled at 10 ꢀC
was added dropwise methane sulfonyl chloride (1.72 g,
15 mmol) over 20 min. The reaction mixture was kept at
0 ꢀC for 2 h, then at room temperature overnight. The
solvent was removed under reduced pressure and the
remaining residue dissolved in 20 mL of CH2Cl2. The
organic solution was washed with saturated NaHCO3
(3Â50 mL), brine (50 mL), dried over MgSO4, and con-
centrated under reduced pressure to yield the 3-O-
mesylate of 10 as a colourless oil (3.0 g, 90%), which
was used in the next step without further puri®cation;
(‹)-Diethyl (3,3-diethoxy-2-hydroxy-propyl)-1-phos-
phonate (13a). Compound 13a was prepared from the
acetal 11 (0.80 g, 5.48 mmol) in 46% yield (0.720 g) by
following the same procedure as described for 4a, using
diethyl phosphite in place of dibenzyl phosphite. 31P
1
the H and 13C NMR spectrum were consistent with
those reported in the literature.27 To a stirred solution
of the 3-O-mesylate of 10 (2.9 g, 12 mmol) in 20 mL of
water at 30 C containing phenolphtalein was added a
solution of 1 N NaOH over 90 mN to keep th pH of the
mixture slightly basic. The progress of the reaction was
monitored by TLC (CH2Cl2:MeOH, 98:2). The reaction
mixture was extracted with ethylacetate (3Â20 mL). The
organic layer was washed with brine, dried over MgSO4
and concentrated under reduce pressure. The resulting
residue was puri®ed by ¯ash distillation at 70 ꢀC (15±
16 mm Hg) to yield 11 as a colourless oil (1.10 g, 56%
based on glyceraldehyde diethyl acetal).
1
NMR (CDCl3) d 30.8; H NMR (CDCl3) d 1.18 (t, 6H,
CH3 acetal), 1.31 (t, 6H, CH3 ester), 1.9±2.1 (m, 2H,
CH2 P), 3.15 (s, 1H D2O exchangeable), 3.5±3.8 (m,
5H, CH O, OCH2 acetal), 4.0±4.15 (m, 4H, CH2O
ester), 4.37 (d, 1H, CH acetal); 13C NMR (CDCl3) d
1
15.3, 16.4, 28.0 (d, JCP=141 Hz, CH2 P), 61.8, 63.5,
3
64.0, 67.7, 104.1 (d, JCP=17.2 Hz, CH acetal). Anal.
calcd for C11H25O6P: C, 46.5; H, 8.80; O, 33.8. Found:
C, 46.6; H, 8.75; O, 33.6.
Method B: Diethyl azido dicarboxylate (2.40 g,
13.8 mmol) was added dropwise to a stirred mixture of
triphenylphosphine (3.11 g, 11.82 mmol) and acetal 10
(1.80 g, 11 mmol) in dry benzene (40 mL) under nitrogen
atmosphere. An exothermic reaction was observed. The
reaction mixture was stirred overnight at room tem-
perature. The benzene was removed under reduced
pressure and the remaining residue puri®ed by ¯ash
distillation to yield 11 (1.26 g, 78%).
(‹)-Diethyl (4,4-diethoxy-3-hydroxy-butyl)-1-phosphonate
(13b). Compound 13b was prepared from acetal 11 (3 g,
20 mmol) in 76% yield (4.5 g) by following the same
procedure as described for 4a, using diethylmethyl
phosphonate (9.3 g, 61 mmol) instead dibenzyl phos-
1
phite. 31P NMR (CDCl3) d 30.8; H NMR (CDCl3) d
1.15 and 1.17 (2t, 6H, CH3 acetal), 1.27 (t, 6H, CH3
ester), 1.4±2.1 (m, 4H, CH2 CH2 P), 2.5 (m, 1H D2O
exchangeable), 3.3±3.8 (m, 5H, CH O, OCH2 acetal),
4.0±4.15 (m, 4H, CH2O ester), 4.21 (d, 1H, CH acetal);
1
Method C: To a stirred solution of acrolein diethyl
acetal (25 g, 192 mmol) in 100 mL of methanol was
13C NMR (CDCl3) d 15.4, 16.5, 21.8 (d, JCP=142 Hz,
CH2 P), 24.9, 61.6, 63.6, 71.5 (d, 3JCP=15.2 Hz, CH O),

1410
P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
104.8. Anal. calcd for C12H27O6P: C, 48.3; H, 9.0; O,
32.2. Found: C, 47.9; H, 9.1; O, 31.6.
1.17 mmol) in 89% yield (0.310 g) by following the same
procedure described as for 5a. 31P NMR (CDCl3) d
31.6; H NMR (CDCl3) d 1.23 (t, 6H, CH3 acetal), 1.32
1
(‹)-Diethyl (3,3-dibutoxy-2-hydroxy-propyl)-1-phos-
phonate (13c). A 1.6 M solution of n-BuLi in hexane
(20.6 mL, 33 mmol) was added dropwise to a stirred
solution of diethylmethyl phosphonate (5.0 g, 33 mmol)
in 25 mL of dry THF at 70 ꢀC under nitrogen atmo-
sphere. After 30 min of stirring at 70 ꢀC, acetal 12
(5.64 g, 30 mmol) in 50 mL of dry THF was added
dropwise. The reaction mixture was stirred for 1 h at
20 ꢀC then overnight at room temperature and quen-
ched with saturated NH4Cl (20 mL). The solvent was
removed under reduced pressure and the residue dis-
solved in 100 mL of ethyl acetate. The organic solution
was washed with brine, dried over MgSO4 and con-
centrated. The resulting residue was puri®ed by ¯ash
chromatography (CH2Cl2:MeOH, 96:4) to yield 13c as a
colourless oil (9.5 g, 93%). 31P NMR (CDCl3) d 30.8;
1H NMR (CDCl3) d 0.84 (t, 6H, CH3 acetal), 1.25 (t,
6H, CH3 ester), 1.3±1.6 (m, 8H, CH2), 1.65±1.95 (m,
2H, CH2 P), 3.12 (s, 1H D2O exchangeable), 3.4±3.7
(m, 4H, OCH2 acetal), 3.95 (m, 1H, CH O), 3.97±4.1
(m, 4H, CH2O ester), 4.29 (d, 1H, CH acetal); 13C
(t, 6H, CH3 ester), 1.8±2.2 (m, 2H, CH2 P), 2.8±3.0 (m,
2H, CH2CˆO), 3.5±3.75 (m, 4H, OCH2 acetal), 4.0±
4.15 (m, 4H, CH2O ester), 4.60 (s, 1H, CH acetal); 13C
1
NMR (CDCl3) d 15.1, 16.4, 18.9 (d, JCP=145 Hz,
CH2 P), 30.2, 61.6, 63.5, 102.4, 202.4 (d, JCP=
3
15.4 Hz, CˆO). Anal. calcd for C12H25O6P: C, 48.65; H,
8.44; O, 32.4. Found: C, 48.8; H, 8.2; O, 32.6.
(‹)-3,3-Dibutoxy-2-hydroxy-propyl-1-phosphonic acid,
dilithium salt (15a). A mixture of acetal 13c (2 g,
5.88 mmol), LiOH monohydrate (1.72 g, 41 mmol) and
NaBH4 (0.2 g, 5.1 mmol) in 30 mL of distilled water was
warmed for 12 h under pressure (2 Bar). Lithium salt
15a was precipitated by addition of ethanol (90 mL) and
the resulting mixture was kept at 0 ꢀC for 3 h. The salt
was collected by centrifugation, washed twice with eth-
anol (80%, then absolute), ethyl ether and dried in
vacuo to yield 15a as a white powder (1.56 g, 90%). 31P
NMR (D2O) d 20.0; 1H NMR (D2O) d 0.89 (t, 6H, CH3
acetal), 1.25±2.0 (m, 10H, CH2, CH2 P), 3.6±3.8 (m,
4H, OCH2 acetal), 3.8±4.0 (m, 1H, CH O), 4.38 (d, 1H,
O CH O); 13C NMR (D2O) d 15.8, 21.4, 33.2 (d,
1JCP=127 Hz, CH2 P), 33.8, 70.7, 71.1, 71.2, 108.0 (d,
3JCP=18.0 Hz, CH acetal). Anal. calcd for C11H23
O6PLi2: C, 44.59; H, 7.77; O, 32.4. Found: C, 44.82; H,
7.9; O, 32.72.
NMR (CDCl3) d 13.8, 16.4, 19.3, 28.0 (d, 1JCP=142 Hz,
3
CH2 P), 31.9, 61.7, 67.6, 67.7, 68.2, 104.3 (d, JCP=
17.4 Hz, CH acetal). Anal. calcd for C15H33O6P: C,
52.9; H, 9.70; O, 28.2. Found: C, 52.8; H, 9.75; O, 28.3.
(‹)-Diethyl (2,2-dibutoxy-1-hydroxy-ethyl)-1-phosphonate
(13d). Compound 13d was prepared from the acetal 12
(3.1 g, 22 mmol) in 77% yield (4.0 g) by following the
same procedure as described for 13c, except that diethyl-
phosphite was used in place of diethylmethylphos-
(‹)-4,4-Diethoxy-3-hydroxy-butyl-1-phosphonic acid,
dilithium salt (15b). Compound 15b was prepared from
13b (1.5 g, 5.06 mmol) in 96% yield (1.23 g) by following
the same procedure as described for 15a. 31P NMR
1
1
phonate. 31P NMR (CDCl3) d 21.3; H NMR (CDCl3)
(D2O) d 22.9; H NMR (D2O) d 1.22 (t, 6H, CH3 ace-
tal), 1.3±2.0 (m, 4H, CH2 CH2 P), 3.5±3.9 (m, 5H,
d 0.9 (t, 6H, CH3 acetal), 1.31 (t, 6H, CH3 ester), 1.3±1.5
(m, 4H, CH2), 1.5±1.7 (m, 4H, CH2), 2.9 (dd, JHH=
3
3
CH O, OCH2 acetal), 4.46 (d, JHH=4.5 Hz, 1H,
5.5 Hz, 3JHP=14.5 Hz, 1H D2O exchangeable), 3.45±3.8
O OH O); 13C NMR (D2O) d 17.1, 27.9 (d, JCP
=
1
3
(m, 4H, OCH2 acetal), 3.85±4.0 (td, JHH=5.0 Hz,
2JHP=5.0 Hz, 1H, CH O), 4.05±4.3 (m, 4H, CH2O
121 Hz, CH2 P), 29.2, 66.7, 67.2, 75.3 (d, 3JCP=14.0 Hz,
CH O) 107.3. Anal. calcd for C8H17O6PLi2: C, 37.8; H,
6.7; O, 37.8. Found: C, 37.65; H, 6.9; O, 38.1.
3
3
ester), 4.73 (dd, JHH=5.1 Hz, JHP=14.5 Hz, 1H, CH
acetal); 13C NMR (CDCl3) d 13.9, 16.8, 19.3, 31.8, 63.0,
1
63.7, 67.4, 68.4, 69.0 (d, JCP=157 Hz, CH P), 101.1
(‹)-2,2-Dibutoxy-1-hydroxy-ethyl-1-phosphonic acid,
dilithium salt (15c). Compound 15c was prepared from
13d (1.4 g, 4.6 mmol) in 92.5% yield by following the
same procedure described as for 15a. 31P NMR (D2O) d
(d, 2JCP=7.2 Hz, CH acetal). Anal. calcd for C14H31O6P:
C, 51.53; H, 9.51; O, 29.45. Found: C, 50.98; H, 9.82; O,
30.14.
1
3.97; H NMR (D2O) d 0.92 (t, 6H, CH3 acetal), 1.25±
Diethyl (3,3-diethoxy-2-oxo-propyl)-1-phosphonate (14a).
Compound 14a was prepared from 13a (0.25 g,
0.88 mmol) in 76% yield (0.190 g) by following the same
procedure as described for 5a, using 0.18 g (2.22 mmol)
of dicyclohexylcarbodiimide and 3.5 mL of dry DMSO.
1.5 (m, 4H, CH2), 1.5±1.7 (m, 4H, CH2), 3.5±3.85 (m,
4H, OCH2 acetal), 3.85±4.0 (m, 1H, CH O), 4.70 (m,
1H, O CH O); 13C NMR (D2O) d 15.9, 21.5, 33.9,
70.5, 72.8 (d, JCP=154 Hz, CH P), 102.4. Anal. calcd
for C10H21O6PLi2: C, 42.55; H, 7.44; O, 34.04. Found:
C, 42.32; H, 7.66; O, 34.47.
1
1
31P NMR (CDCl3) d 20.2; H NMR (CDCl3) d 1.22 (t,
6H, CH3 acetal), 1.31 (t, 6H, CH3 ester), 3.25 (d,
2JHP=21.8 Hz, 2H, CH2 P), 3.5±3.7 (m, 4H, OCH2
acetal), 4.0±4.2 (m, 4H, CH2O ester), 4.71 (s, 1H,
(‹)-2-Hydroxy-3-oxo-propyl-1-phosphonic acid, di-
sodium salt (16a). Compound 16a was prepared from
15a (0.050 g, 0.17 mmol) in 96% yield by following the
same procedure described as for 7a, except that the pH
was adjusted to 7.4 with 1 N NaOH, then freeze-dried.
O CH O); 13C NMR (CDCl3) d 13.6, 16.3, 35.7 (d,
3
1JCP=132 Hz, CH2 P), 62.5, 63.6, 101.7 (d, JCP
=
9.8 Hz, CH acetal), 201.6. Anal. calcd for C11H23O6P:
C, 46.8; H, 8.15; O, 34.0. Found: C, 46.5; H, 8.27; O, 34.2.
1
31P NMR (D2O) d 20.6; H NMR (D2O) d 1.6±1.95 (m,
2H, CH2 P), 3.8±3.95 (m, 1H, CH O), 4.88 (d,
3JHH=4.4 Hz, 1H, HCˆO, hydrated form); 13C NMR
Diethyl (4,4-diethoxy-3-oxo-butyl)-1-phosphonate (14b).
Compound 14b was prepared from 13b (0.35 g,
1
(D2O) d 32.3 (d, JCP=136 Hz, CH2 P), 72.4, 94.4 (d,

P. Page et al. / Bioorg. Med. Chem. 7 (1999) 1403±1412
1411
3JCP=16.4 Hz, CˆO, hydrated form). Anal. calcd for
C3H5O5PNa2, H2O: C, 16.67; H, 3.24; O, 44.43. Found:
C, 16.99; H, 3.45; O, 44.82.
2,3-Dioxo-propyl-1-phosphonic acid (18a). Compound
18a was prepared from acetal 17a (0.110 g, 0.30 mmol)
by following the same procedure described for 9a,
except that the incubation was carried out for 24 h at
room temperature. 31P NMR (D2O+H2O) d 14.6; 13C
(‹)-3-Hydroxy-4-oxo-butyl-1-phosphonic acid, disodium
salt (16b). Compound 16b was prepared from 15b
(0.060 g, 0.235 mmol) in 96% yield (1.20 g) by following
the same procedure described as for 16a. 31P NMR
(D2O) d 23.7; 1H NMR (D2O) d 1.4±1.9 (m, 4H,
CH2 CH2 P), 3.75±3.85 (m, 1H, CH O), 4.92 (d,
3JHH=5.1 Hz, 1H, HC O, hydrated form); 13C NMR
1
NMR (D2O+H2O) d 34.9 (d, JCP=127 Hz, CH2 P),
3
92.9 (d, JCP=14.9 Hz, HCˆO, hydrated form), 200.5
(CˆO ketone).
3,5-Dioxo-butyl-1-phosphonic acid (18b). Compound
18b was prepared from acetal 17b (0.280 g, 0.75 mmol)
by following the same procedure as described for 18a.
31P NMR (D2O+H2O) d 25.2; 13C NMR (D2O+H2O)
1
(D2O) d 25.75 (d, JCP=135 Hz, CH2 P), 27.65, 76.5
3
(d, JCP=16.0 Hz, CH O) 94.2 (CˆO, hydrated form.
1
Anal. calcd for C4H7O5PNa2, H2O: C, 20.87; H, 3.91;
O, 41.74. Found: C, 21.62; H, 4.09; O, 42.26.
d 18.9 (d, JCP=125 Hz, CH2 P), 33.6, 92.3 (HCˆO,
3
hydrated form), 201.2 (d, JCP=13.6 Hz, CˆO ketone).
(‹)-1-Hydroxy-2-oxo-ethyl-1-phosphonic acid, dilithium
salt (16c). Compound 16c was prepared from 15c
(0.100 g, 0.35 mmol) in 91% yield (0.055 g) by following
the same procedure described as for 16a except that 1 M
LiOH was used to adjust the pH to 7.4. 31P NMR
(D2O) d 6.6; 1H NMR (D2O) d 3.8±4.0 (m, 1H, CH O),
HCˆO (hydrated form) mixed up with H2O; 13C NMR
Diethyl (2-oxo-3-hydroxy-propyl)-1-phosphonate (19a).
To a 100-mL round-bottomed ¯ask containing 13c
(1.5 g, 5.03 mmol) in 10 mL of demineralized water were
added 0.100 mL of concentrated HCl (35%). The mix-
ture was warmed to 45 ꢀC for 2 h and monitored by
TLC (CH2Cl2:MeOH, 9:1); the solution was freeze-
dried. The resulting residue was puri®ed by ¯ash chro-
matography (CH2Cl2:MeOH, 9:1) to yield 19a as a col-
ourless oil (0.502 mg, 54%). 31P NMR (CDCl3) d 19.4;
1H NMR (CDCl3) d 1.28 (t, 3H, CH3), 3.14 (d,
2JHP=26.9 Hz, CH2 P), 3.51 (br, 1H, D2O exchange-
able), 4.10 (m, 4H, CH2O) 4.27 (s, 2H, CH2O); 13C
1
(D2O) d 71.5 (d, JCP=146 Hz, CH P), 93.4 (CˆO,
hydrated form). Anal. calcd for C2H3O5PLi2, H2O: C,
14.12; H, 2.94; O, 56.47. Found: C, 14.58; H, 3.26; O,
57.2.
1
3,3-Diethoxy-2-oxo-propyl-1-phosphonic acid, barium salt
(17a). Freshly distilled trimethylsilyl bromide (1.4 mL,
10.6 mmol) was added slowly with stirring to the acetal
14a (0.110 g, 0.39 mmol) at 20 ꢀC under nitrogen
atmosphere. The reaction mixture was stirred for 24 h at
room temperature. Excess trimethylsilyl bromide was
removed under reduced pressure and water (5 mL)
added to the remaining residue at 30 ꢀC. The aqueous
solution was washed with ether, the pH adjusted to 7.6
with saturated Ba(OH)2 then freeze-dried. The remain-
ing residue was dissolved in a minimum volume of water
and the barium salt was precipitated by addition of
three volumes of absolute ethanol. The resulting mix-
ture was kept at 0 ꢀC for 3 h, the salt collected by cen-
trifugation, washed twice with ethanol (80%, then
absolute), ethyl ether and dried in vacuo to yield 18a as
NMR (CDCl3) d 18.4, 38.7 (d, JCP=127 Hz, CH2 P),
2
62.1, 63.0, 69.0, 202.6 (d, JCP=6.5 Hz, CˆO). Anal.
calcd for C7H15O5P: C, 40.0; H, 7.14; O, 38.1. Found:
C, 40.06; H, 7.21; O, 38.24.
Diethyl (3-oxo-4-hydroxy-butyl)-1-phosphonate (19b).
Compound 19b was prepared from 13b (1.5 g,
4.41 mmol) in 70% yield by following the same proce-
dure as described for 19a. 31P NMR (CDCl3) d 30.7; 1H
NMR (CDCl3) d 1.26 (t, 3H, CH3), 1.8±2.2 (m, 2H,
CH2 P), 2.6±2.8 (m, 2H, CH2 CH2CˆO), 4.04 (m,
4H, CH2O P), 4.2 (s, 2H, CH2O); 13C NMR (CDCl3) d
1
16.4, 19.1 (d, JCP=145 Hz, CH2 P), 31.3, 61.8, 66.0,
3
208.1 (d, JCP=13.4 Hz, CˆO). Anal. calcd for C8H17
O5P: C, 42.8; H, 7.6; O, 35.7. Found: C, 43.1; H, 7.7; O,
35.4.
a white powder (0.130 g, 92%): 31P NMR (D2O) d 11.4;
2
1H NMR (D2O) d 1.20 (t, 6H, CH3), 3.10 (d, JHP
=
19.5 Hz, 2H, CH2 P), 3.5±3.65 (m, 4H, CH2 O), 5.25
Acknowledgements
(s, 1H, O CH O); 13C NMR (D2O) d 15.45, 32.2 (d,
This work received the ®nancial support of `Comite
d'Interface Chimie-Biologie du CNRS' and DRET
(GDR 1077) which are fully acknowledged.
3
1JCP=131 Hz, CH2 P), 63.1, 101.3 (d, JCP=8.9 Hz,
CH acetal), 200.9. Anal. calcd for C7H13O6PBa: C,
23.27; H, 3.6; O, 26.6. Found: C, 23.7; H, 3.97; O, 27.28.
4,4-Diethoxy-3-oxo-butyl-1-phosphonic acid, barium salt
(17b). Compound 17b was prepared from acetal 14b
(0.280 g, 0.945 mmol) in 93% yield (0.330 g) by follow-
ing the same procedure as described for 18a. 31P NMR
(D2O) d 22.2; 1H NMR (D2O) d 1.22 (t, 6H, CH3), 1.7±2.1
(m, 2H, CH2 P), 2.8±2.95 (m, 2H, CH2CˆO), 3.45±3.65
(m, 4H, OCH2), 5.17 (s, 1H, O CH O); 13C NMR
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