A Three-Step Preparation of Dihydroxyacetone Phosphate Dimethyl Acetal

Full text of DOI:10.1021/jo9823902

J . Org. Chem. 1999, 64, 4943-4945
4943
A Th r ee-Step P r ep a r a tion of
prepared. This synthesis was later modified by Weiling
and E5 who made the diethyl acetal. Because 1 is
Dih yd r oxya ceton e P h osp h a te Dim eth yl
4
Aceta l†
converted nearly quantitatively to DHAP by acid hy-
drolysis, it is an excellent precursor for the production
of this biochemical. However, the preparation of 1 is
somewhat lengthy, requiring six steps from a com-
mercially available material. Similarly, 2 is converted in
Edward L. Ferroni,* Victoria DiTella, Nancy Ghanayem,
Rebecca J eske, Christopher J odlowski,
Matthew O’Connell, J ennifer Styrsky, Robyn Svoboda,
Ajay Venkataraman, and Beth M. Winkler
>
95% yield to DHAP by acid hydrolysis, but its synthesis
requires six steps from acetone (or five steps from 1,3-
Department of Chemistry, Benedictine University,
6
dibromoacetone). Compounds 3 and 4 are synthetically
5
700 College Road, Lisle, Illinois 60532-0900
more attractive because they are made easily and ef-
7
ficiently in only three steps from 1,3-dibromoacetone and
Received December 7, 1998
8
the dihydroxyacetone dimer, respectively. Effenberger
9
and Straub originally synthesized 4 from the dihydroxy-
In tr od u ction
acetone dimer in 1987. This preparation was later
modified by Pederson et al.3 and then by J ung et al.,
a
8
Dihydroxyacetone phosphate (DHAP) is a biochemical
that acts as a substrate for a number of enzymes,
significantly improving the overall yield. However, 4 has
one significant drawbacksit is converted to DHAP with
1
including triose phosphate isomerase, R-glycerol phos-
1
1
8
7
phate dehydrogenase, and several types of aldolases.
only a 66% yield. Similarly, 3 is converted in <73% yield
Because the aldolases can be used in the synthesis of a
variety of carbohydrates on a preparative scale, DHAP
to DHAP. It must be noted, however, that these impure
2
DHAP preparations made from 3 and 4 have been shown
7
,8
has taken on a new importance as a precursor molecule
in organic synthesis. As a result, there is interest in
developing new ways of producing this compound.
Because DHAP itself is unstable, there have been two
general strategies for its synthesis: (1) enzymatic prepa-
ration of DHAP solutions for immediate use and (2)
chemical synthesis of stable DHAP precursors (contain-
ing a protected ketone group) that may be stored indefi-
nitely. The limitations of enzymatic preparations have
been discussed elsewhere3 and will not be addressed
herein. Several stable DHAP precursors have been
reported in the literature: dihydroxyacetone phosphate
to be useful for at least some enzymatic syntheses.
The ideal precursor would combine the benefits out-
lined above. It would be efficiently converted into DHAP
(like 1 and 2), and itself be synthesized easily in just a
few steps from a cheap, commercially available compound
(like 3 and 4). Furthermore, the synthesis of the ideal
precursor molecule would not involve the use of expensive
reagents, such as Pt or Pd, metals that are used in the
synthesis of all four of the precursors shown above. In
this regard, we report herein the synthesis of 1 by a
three-step method that is inexpensive, is easily carried
out, and can be accomplished on a gram scale.
4
4,5
dimethyl acetal (or diethyl acetal ) (1), 3-acetoxy hy-
doxyacetone phosphate dimethyl acetal6 (2), 3-bromo
hydroxyacetone phosphate dimethyl acetal (3), and 2,5-
diethoxy-p-dioxane-2,5-dimethanol O-2 ,O-5 -bis-phos-
Resu lts a n d Discu ssion
7
1
1
Dihyrdoxyacetone dimethyl acetal (6) (see Scheme 1)
was made according to a slight modification of a previ-
ously published method,10 the only difference being the
phate (4).8
1
0
manner of product isolation. Cesarotti et al., who
reported a yield of 75.7%, purified 6 by column chroma-
tography. In this study, distillation was used because it
was cheaper and more convenient. Three fractions were
obtained, with fraction 2 found to be compound 6, isolated
in 41% yield. Although the data are not presented herein,
both NMR and TLC analyses of fractions 1 and 3
indicated the presence of 6, though these fractions were
impure. However, no attempt was made to isolate 6 from
these fractions and thus improve the yield of the reaction.
The cyclic phosphate triester 7 was made according to
Compound 1 was synthesized by Ballou and Fischer4
in 1956, the first time a stable precursor of DHAP was
1
1
the general procedure reported by Penny and Belleau
†
Supported by the Howard Hughes Medical Institute (HHMI
#
711191-528601 and HHMI #71196-528602) and the NSF (Instrumen-
tation and Laboratory Improvement, USE-9052273).
1) Walsh, C. Enzymatic Reaction Mechanisms; W. H. Freeman: San
Francisco, CA, 1979; pp 593, 594, 745.
2) For example: (a) Henderson, I.; Sharpless, K. B.; Wong, C.-H.
(3) (a) Pederson, R. L.; Esker, J .; Wong, C.-H. Tetrahedron 1991,
47, 2643. (b) Toone, E. J .; Simon, E. S., Bednarski, M. D.; Whitesides,
G. M. Tetrahedron 1989, 45, 5365.
(4) Ballou, C. E.; Fischer, H. O. L. J . Am. Chem. Soc. 1956, 78, 1659.
(5) Weiling, X.; E, W. YoujHuaxue 1986, 3, 201.
(6) Valentin, M.-L.; Bolte, J . Bull. Soc. Chim. Fr. 1995, 132, 1167.
(7) Gefflaut, T.; Lemaire, M.; Valentin, M.-L.; Bolte, J . J . Org. Chem.
1997, 62, 5920.
(
(
J . Am. Chem. Soc. 1994, 116, 558. (b) Valentin, M.-L.; Bolte, J .
Tetrahedron Lett. 1993, 34, 8103. (c) Wong, C.-H.; Liu, K. K.-C.;
Kajimoto, T.; Chen, L.; Zhong, Z.; Dumas, D. P.; Liu, L.-C.; Ichikawa,
Y.; Shen, G.-J . Ind. J . Chem. 1993, 32B, 135. (d) Fessner, W.-D.; Badia,
J .; Eyrisch, O.; Schneider, A.; Sinerius, G. Tetrahedron Lett. 1992, 33,
(8) J ung, S. H.; J eong, J .-H.; Miller, P.; Wong, C.-H. J . Org. Chem.
1994, 59, 7182.
5
9
1
7
231. (e) Schmid, W.; Whitesides, G. M. J . Am. Chem. Soc. 1990, 112,
670. (f) Whitesides, G. M.; Wong, C.-H. Angew. Chem., Int. Ed. Engl.
985, 24, 617. (g) Lehninger, A. L.; Sice, J . J . Am. Chem. Soc. 1955,
7, 5343.
(9) Effenberger, F.; Straub, A. Tetrahedron Lett. 1987, 28, 1641.
(10) Cesarotti, E.; Antognazza, P.; Pallavicini, M.; Villa, L. Helv.
Chim. Acta 1993, 76, 2344.
(11) Penny, C. L.; Belleau, B. Can. J . Chem. 1978, 56, 2396.
1
0.1021/jo9823902 CCC: $18.00 © 1999 American Chemical Society
Published on Web 06/09/1999

4
944 J . Org. Chem., Vol. 64, No. 13, 1999
Notes
4
Sch em e 1. Syn th esis of Dih yd r oxya ceton e
procedure of Ballou and Fischer. NMR spectra were recorded
a
on a 300 MHz instrument. Chemical shifts were determined
relative to TMS in CDCl or to HOD (4.8 ppm) in D O. A
3 2
13
P h osp h a te Dim eth yl Aceta l
calibrated thermometer was used for measuring melting points,
but no stem correction was made.
2
,2-Dim eth oxy-1,3-p r op a n ed iol (6). This compound was
synthesized by a slight modification of the method of Cesarotti
et al.10 Dihydroxyacetone dimer (5) (75 g, 0.42 mol) was stirred
overnight in a solution containing 90 mL of trimethyl orthofor-
mate, 1000 mL of anhydrous methanol, and 300 mg of p-toluene
sulfonic acid monohydrate. The reaction was halted by adding
9
00 mg of anhydrous sodium carbonate and by stirring the
mixture overnight, during which time most of the salt dissolved.
The solvent plus unreacted trimethylorthoformate was removed
by evaporation under reduced pressure, and the resulting oil
was distilled in vacuo at 2 Torr. Three fractions (fraction 1, bp
105-115 °C; fraction 2, bp 115-125 °C; fraction 3, bp. 125-130
a
(
a) HC(OMe)3, MeOH, cat. TsOH; (b) PhOP(O)Cl2, C6H5N; (c)
0
.1 M Ba(OH)2, 96 °C.
°
C), which solidified upon cooling, were obtained and recrystal-
lized from toluene. Recrystallized fraction 2, 46.7 g (0.34 mol,
1%), was found to be pure by elemental analysis.
Anal. Calcd. for C : C, 44.11; H, 8.88. Found: C, 44.05;
H, 9.08.
Sch em e 2
4
5
12 4
H O
Compound 6 was very hygroscopic and was stored at room
temperature in a desiccator protected from moisture.
5
,5-Dim et h oxy-2-p h en oxy-2-oxo-1,3,2-d ioxa p h osp h or i-
n a n e (7). Compound 6 (15.0 g) was dissolved in 65 mL of
anhydrous pyridine in a two-neck 100 mL round bottom flask.
The solution was cooled in an ice/salt bath, and 15 mL of phenyl
dichlorophosphate was added dropwise stirring over the course
of 1 h, keeping the temperature of the reaction mixture below
1
0 °C. The mixture was stirred for an additional 20 h at room
temperature. Solid pyridinium hydrochloride was removed by
filtration of the mixture on a Buchner funnel, the solids were
washed with benzene, and the combined filtrate plus washes
were evaporated to dryness under reduced pressure (T < 40 °C).
The residue was triturated with water, and the resulting solid
was isolated by centrifugation at 9000g for 10 min. The solid
was washed consecutively with 50 mL each of water, saturated
sodium bicarbonate, and water. The crude product was dissolved
in 300 mL of warm benzene after drying the solid overnight on
a watch glass (or in a vacuum desiccator for several hours). The
resulting cloudy mixture was stirred while heating gently for
for the reaction of phenyl dichlorophosphate with diols.
As evidenced by NMR spectra and the results of elemen-
tal analyses, 7 was found to be pure, and so it was not
1
1
recrystallized. In their work, Penny and Belleau pre-
pared a number of cyclic phosphate triesters and obtained
yields ranging from 47 to 78%. The 72% yield reported
herein for the preparation of 7 falls within this range.
The hydrolysis of 7 was carried out at 96 ( 2 °C in 0.1
M barium hydroxide. When 7, which is insoluble in water,
was added to the hot barium hydroxide solution, a two-
phase mixture resulted. Within 20 min, a homogeneous
solution was observed as 7 was (presumably) converted
to 8 (Scheme 2), an anion soluble in water. As evidenced
by enzyme assays, 8 was hydrolyzed to 1 in 16 h, under
the conditions of the reaction. The conversion of 7 to 1
2
0 min, cooled for 10 min, and then dried by the addition of
anhydrous sodium sulfate. Filtration of the mixture followed by
evaporation of the solvent under reduced pressure yielded 21.9
1
g (72%) of a white, fluffy solid: mp 100-101 °C;
δ 3.27, (s, 3 H), 3.36, (s, 3 H), 4.25-4.43 (m, 4 H), 7.16-7.4 (m,
.6 H). Anal. Calcd for C11 P: C, 48.2; H, 5.51; P, 11.3.
Found: C, 48.2; H, 5.63; P, 11.4.
H NMR (CDCl3)
4
15 6
H O
Compound 7 was stored at -18 °C, but it could be kept at
room temperature for several months without significant de-
composition as evidenced by NMR.
(66% yield) took advantage of the very large difference
in the rate of basic hydrolysis of phosphate di- and
monoesters. It is known that phosphate diester monoan-
ions react with hydroxide rapidly as compared with the
2
,2-Dim eth oxy-1,3-p r op a n ed iol P h osp h a te (1). Compound
7
(2.5 g) was added in one portion to 250 mL of a stirred 0.10 M
solution of barium hydroxide that had been preheated to 94-
98 °C in an oil bath. The compound melted, and the two phases
that resulted disappeared within 20 min as the phenyl ester was
hydrolyzed. The solution, protected from the atmosphere by the
use of a soda lime tube, was heated with stirring for 16 h (the
extent of the reaction was determined enzymatically as described
below), at which time the reaction mixture was removed from
the oil bath and allowed to cool for 1.5 h. A solution of 8.1 g of
dicyclohexylammonium sulfate (27.3 mmol, a 10% excess) in 50
mL of water was added, and the resulting mixture stirred for
rate of reaction of hydroxide with phosphate monoester
dianions.12 Thus, the cleavage of 1 to 6 (Scheme 2) was
slow enough to allow 1 to be isolated from the reaction
solution.
The three-step synthesis of 1 presented herein is
straightforward and relatively inexpensive. The overall
yield of 20% was disappointing, but since the reactions
can be carried out on a multigram scale, it is a convenient
method for the preparation of an excellent precursor of
dihydroxyacetone phosphate.
1
5 min. The barium sulfate precipitate was removed by cen-
trifugation at 10000g for 10 min and washed three times with
0 mL of water. The combined supernatant and washings were
5
evaporated to dryness under reduced pressure (T < 40 °C). The
residue remaining was dissolved in boiling hot absolute ethanol,
and the mixture was vacuum filtered on a Buchner funnel using
filter paper in order to remove the excess dicyclohexylammonium
sulfate and any salts of inorganic phosphate that may have been
present. If the filtrate was still cloudy, then it was passed
Exp er im en ta l Section
Gen er a l. All reagents and solvents (including anhydrous
solvents) were purchased from either the Sigma or Aldrich
Chemical Co. and were used without further purification.
Dicyclohexylammonium sulfate was synthesized according to the
(
13) The authors wish to thank Dr. Karol Bruzik, Dr. David Rausch,
(12) Westheimer, F. H. Science 1987, 235, 1173.
and William Lambert for obtaining NMR spectra.

Notes
J . Org. Chem., Vol. 64, No. 13, 1999 4945
1
through a 0.45 µm nylon filter. The filtrate was evaporated to
dryness under reduced pressure, and the residue was dissolved
in about 30 mL of water. The mixture was filtered through a
sodium salt for NMR analysis as described above. H NMR (D
2
O)
δ 3.28 (s, 6 H), 3.63 (s, 2 H), 3.76 (d, J ) 5.5 Hz, 2 H). Anal.
Calcd for the dicyclohexylammonium salt C17 O: C,
H
39
N
2
O
7
P‚H
2
0
.45 µm nylon filter, and the filtrate was evaporated to dryness
47.2; H, 9.56; N, 6.48. Found: C, 46.9; H, 9.56; N, 6.48. The
combined yield for the two crops was 2.61 g (66%).
under reduced pressure. The solid obtained was dissolved in 8
mL of water, the pH of the solution was adjusted to 10.1 with
cyclohexylamine, and 35 mL of acetone was added. The mixture
was allowed to stand overnight at 4 °C, and the white solid,
which crystallized, was collected by filtration on a Buchner
funnel, containing a medium porosity glass frit. The solid was
washed with several portions of acetone and allowed to air-dry
on the funnel for several hours. Yield ) 1.74 g. For NMR
analysis, 30-50 mg of the solid was dissolved in 1 mL of water,
The rate of ring opening of 7 to produce 1 was monitored
enzymatically. A 0.5 mL aliquot of the barium hydroxide
hydrolysis reaction solution was removed at various times and
+
mixed with 0.5 g of Dowex-50(H ) for 60 s. The resulting mixture
was filtered, and the Dowex was washed twice with 0.5 mL of
water. The combined filtrate plus washings (pH = 2) was
incubated at 65 °C for 25 min to hydrolyze the dimethyl acetal
of dihydroxyacetone phosphate. This solution was transferred
quantitatively to a 10.00 mL volumetric flask and diluted to the
mark with a 0.05 M Tris buffer, pH 7.5. An aliquot of this
buffered solution was then assayed enzymatically for dihydroxy-
+
and the solution was stirred with about 0.5 g of Dowex-50(H )
for 1 min to remove the cyclohexylammonium groups. After
filtration to remove the Dowex polymer, solid NaHCO
added in portions until the pH was about 7. The solution was
freeze-dried, and the resulting solid was dissolved in D
O. 1
NMR (D O) δ 3.28 (s, 6 H), 3.63 (s, 2 H), 3.76 (d, J ) 5.4 Hz, 2.1
H). Anal. Calcd for the dicyclohexylammonium salt C17 P‚
O: C, 47.2; H, 9.56; N, 6.48. Found: C, 47.4; H, 9.64; N, 6.53.
The filtrate and washes from above were combined and placed
3
was
14
acetone phosphate. Note that although this method was
2
H
adequate to measure the extent of reaction, it appeared that the
hydrolysis of the acetal at 65 °C for 25 min was sensitive to pH.
Preliminary observations seemed to indicate that if the pH of
the combined filtrate plus washings was above 2, then incom-
plete hydrolysis of the acetal occurred. When the pH was lower,
then quantitative hydrolysis seemed to occur. More work must
be done to produce a reproducible acetal hydrolysis procedure
at 65 °C.
2
39 2 7
H N O
H
2
overnight at 4 °C in order to obtain a second crop. Yield ) 0.87
g. The dicyclohexylammonium salt was converted into the
(14) Bergmeyer, H. U. Methods of Enzymatic Analysis; Academic
Press: New York, 1965; p 246.
J O9823902