Synthesis and properties of glycolaldehyde di- and triphosphate

Article abstract of DOI:10.1055/s-2002-19325

Use of photolabile acetal protecting groups enables the synthesis of glycolaldehyde di- and triphosphate 4 and 5, which undergo enolisation at significantly lower pH values than glycolaldehyde phosphate 1. At pH > 10, 5 is converted to 1 and inorganic pyrophosphate.

Full text of DOI:10.1055/s-2002-19325

170
LETTER
Synthesis and Properties of Glycolaldehyde Di- and Triphosphate
S
ynthesis and Prop
n
erties of
G
ly
t
coalde
h
hyde
D
i- and
o
T
riphospha
n
Department of Chemistry, University of Manchester, Oxford Road, Manchester M13 9PL, UK
Fax +44(161)2754939; E-mail: john.sutherland@man.ac.uk
Received 29 October 2001
matography (Dowex® 1X8-400 eluting with an aqueous
ammonium bicarbonate gradient) and then converted to
the tetramethylammonium salt (treatment with H-
Dowex® followed by titration to pH 7 with tetramethy-
lammonium hydroxide solution). Ozonolysis in methanol
at –78 °C followed by reductive work-up then provided 4
Abstract: Use of photolabile acetal protecting groups enables the
synthesis of glycolaldehyde di- and triphosphate 4 and 5, which un-
dergo enolisation at significantly lower pH values than glycolalde-
hyde phosphate 1. At pH > 10, 5 is converted to 1 and inorganic
pyrophosphate.
Key words: aldehydes, bioorganic chemistry, ozonolysis, phos-
phorylations, photochemistry
1
contaminated with ca. 10% 1 as judged by H and 31P
NMR (Scheme 2).1,5 No resonances for non-hydrated al-
dehydic protons were observed in the 1H NMR spectrum
indicating that both species were > 95% hydrated. It is not
clear how 1 is formed during this procedure although it is
possible that a carbonyl oxide or -methoxy-hydroperox-
ide attacks the -phosphorus nucleophilically resulting in
pyrophosphate cleavage.6 Fortuitously the monophos-
phate 1 served as a convenient internal standard in subse-
quent experiments. Although dimethylallyl triphosphate
was successfully prepared by alkylation of tetrakis(tetra-
n-butylammonium) hydrogen triphosphate, subsequent
ozonolysis with reductive work-up gave at least four gly-
colaldehyde derivatives by 1H NMR.
The aldolisation of glycolaldehyde phosphate 1 in the
presence of formaldehyde has been shown by Eschen-
moser et al. to constitute an efficient route to pentose-2,4-
diphosphates with ribose-2,4-diphosphate 2 predominat-
ing.1 The sequence involves reaction of glycolaldehyde
phosphate with formaldehyde to give glyceraldehyde-2-
phosphate 3 which acts as an aldol acceptor to a second
molecule of 1 (Scheme 1).
O–
O–
P
O–
P
H
O
O
P
–O
O–
P
–O
–O
O
CH2O
O
O
1
O
O
O
O–
H
H
OH
O
HO
HO
OH
i) - iv)
O
HO
O
O
O
O–
O
O
O–
P
P
P
P
(rac)-3
(rac)-2
1
– •2Me4N+•H+
– •3Na+
O
O– O
O
O
O– O O
6
4
Scheme 1
+ 1•2Na+ (9:1)
An aldolisation route to nucleic acid backbones would
gain credence as a potentially prebiotic pathway if it could
be shown to operate under conditions at which nucleic ac-
ids (in particular RNA) are stable. The phosphorylation of
glycolaldehyde using amidotriphosphate2 and the aldol re-
action of 3 with 1 in the presence of double layer metal hy-
droxide minerals3 both fulfil this criterion, taking place in
aqueous solutions at near neutral pH. The aldol reaction of
1 with formaldehyde however only proceeds in a strongly
alkaline medium,1 conditions under which RNA is hydro-
lytically labile. In connection with related work we had
cause to prepare glycolaldehyde diphosphate 4 and glyco-
laldehyde triphosphate 5 the properties of which suggest a
possible solution to this problem.
Scheme 2 Reagents and conditions: i) O3, MeOH, –78 ºC; ii) Me2S;
iii) repeated evaporation from H2O; iv) Na-Dowex®
Investigation of the sample of 4 containing 1 by 1H NMR
(in D2O) revealed that the -protons of 4 fully exchange at
20 °C, pD 8.0 after 4 days (t1/2 ca. 20 h) whereas those of
1 do not (Figure 1). The greater electron-withdrawing ca-
pacity of the diphosphate group compared to the mono-
phosphate group is probably responsible for this
phenomenon. The increase in electron-withdrawal pre-
sumably biases the hydration equilibrium of 4 in favour of
hydrate further than that of 1 but the residual aldehyde of
4 is rendered intrinsically more prone to enolisation than
the residual aldehyde of 1. Evidently the effect on intrinsic
enolisation rate outweighs the effect on the hydration
equilibrium. This extremely subtle electron-demand/hy-
dration controlled enolisation chemistry prompted us to
establish a secure synthetic route to 5 and a cleaner route
to 4.
Alkylation of tris(tetra-n-butylammonium) hydrogen py-
rophosphate with dimethylallyl bromide in dry acetoni-
trile as described by Dixit et al.4 furnished dimethylallyl
diphosphate 6 which was purified by ion-exchange chro-
Synlett 2002, No. 1, 28 12 2001. Article Identifier:
1437-2096,E;2002,0,01,0170,0172,ftx,en;D24901ST.pdf.
© Georg Thieme Verlag Stuttgart · New York
ISSN 0936-5214
Unable to use late stage ozonolysis to reveal the sensitive
aldehyde groups of 4 and 5, we developed a strategy based

LETTER
Synthesis and Properties of Glycoaldehyde Di- and Triphosphate
171
OCH2Ar
OBz
H
i) - iii)
iv)
ArCH2O
O
HO
OH
OBz
4 (CH)
1 (CH)
4 (CH2)
8
7
v), vi)
1 (CH2)
vii) (n=1) or
viii) (n=2),
ix), x)
OCH2Ar
O
OCH2Ar
OSO2CF3
ArCH2O
ArCH2O
O–
O
P
P
9
O–
O
O–
n
O
10 (n=1, •3Na+); 11 (n=2, •4Na+)
xi)
NO2
Ar =
OH
HO
O–
O
O
n
P
P
O
O–
O
O–
4 (n=1, •3Na+); 5 (n=2, •4Na+)
Scheme 3 Reagents and conditions: i) BzCl, pyridine; ii) O3,
CH2Cl2, –78 ºC; iii) Me2S; iv) o-NO2-C6H4CH2OSiMe3,
Me3SiOSO2CF3, –78 ºC to r.t. (over 3 h); v) LiOH (1 M), H2O/1,4-
dioxan (1:1); vi) (CF3SO2)2O, pyridine/CH2Cl2, 0 ºC, 2 min (HCl(aq.)
(1 M) quench); vii) (Bun N)3HP2O7, CH3CN; viii) (Bun N)4HP3O10,
Figure 1 1H NMR spectra (500 MHz, D2O, pD 8.0 with HOD sup-
pression) of a mixed sample containing 1 and 4 (1:9) maintained at
20 °C; a) t = 0, b) t = 4 days
4
4
CH3CN; ix) HPLC (C-18: Et3NH HCO3 (50 mM), CH3CN gradient);
upon the use of photolabile protecting groups. Benzoyl x) Na-Dowex®; xi) h , EtOAc:dil. HCl(aq.) (pH 4.5)
glycolaldehyde 7 was prepared conveniently by quantita-
tive dibenzoylation of Z-but-2-ene-1,4-diol followed by
ozonolysis with reductive work-up in 86% yield. The
the gem-diol of 5 is likely to have a pKa near 10 it is prob-
bis(o-nitrobenzyl) acetal 8 was obtained in 70% yield us-
ing Noyori’s procedure;7,8 other methods were unsuccess-
ful (Scheme 3). Saponification of the ester function of 8 in
75% yield followed by careful triflylation (quantitative)
gave the key intermediate 9. Displacement of the triflate
with tris(tetra-n-butylammonium) hydrogen pyrophos-
phate followed by reverse-phase HPLC gave the acetal 10
in 16% yield (compromised for purity). Deprotection at
pH 4.5 by irradiation with a standard slide projector in a
biphasic solvent system9 proceeded smoothly to give 4 in
quantitative yield and a high state of purity as judged by
1H and 31P NMR. All data were consistent with those ob-
tained from the route outlined in Scheme 2.5 Displace-
ment of the triflate with tetrakis(tetra-n-butylammonium)
hydrogen triphosphate was successful and, after similar
HPLC purification, 11 was obtained in 33% yield (again
compromised for purity) from 9. Photolytic deprotection
of 11 proceeded smoothly and the triphosphate 5 was ob-
tained in quantitative yield, in a high state of purity as
judged by 1H and 31P NMR.10
able that the hydrolysis mechanism involves attack of the
hydrate alkoxide on the -phosphorus atom resulting in
the displacement of inorganic pyrophosphate and the for-
mation of the cyclic intermediate 12 which can then ring-
open to 1 (Scheme 4).11
O
O
H
O–
HO
P
O–
O–
O–
O
pH > 10
H2O
O
O
O
O
O
O
1
- PPi
-H+
P
P
P
P
OH
O
HO
O
O–
O–
O–
5
O
O–
12
O
O
O–
P
O–
O
Scheme 4 Proposed mechanism for the conversion of 5 to 1
The ease with which 5 enolises coupled with its propensi-
ty for hydrolysis suggests that it might be possible at low-
er pH values, where RNA is stable, for 5 to aldolise with
With pure samples of 4 and 5 in hand, an investigation of
their solution behaviour was undertaken. The two com-
pounds were found to undergo comparable H/D exchange
(t1/2 ca. 20 h) in D2O at 20 °C, pD 8.0; conditions under
which 1 remains unchanged. Further studies were carried
out with 5 at higher pH values. At pH 10 and above, 5 was
observed to hydrolyse to 1 and inorganic pyrophosphate
(1H, 31P NMR) with a half-life of ca. 1 hour at pH 10.5,
20 °C. Since the hydrolysis is not seen below pH 10 and
formaldehyde
giving glyceraldehyde-2-triphosphate
which could then hydrolyse to 3. Current work is aimed at
addressing this possibility and at investigating possible
prebiotic routes to 5.
Acknowledgement
This work was supported by the BBSRC and the EPSRC.
Synlett 2002, No. 1, 170–172 ISSN 0936-5214 © Thieme Stuttgart · New York

172
A. J. Lawrence, J. D. Sutherland
LETTER
+ 2 H+ – H2O]–); MS (ES-, CV -35 V): m/z (%) = 219(100),
[M3– + 2 H+ – H2O]–).
(6) For related chemistry involving acylation of -methoxy-
hydroperoxides by anhydrides see: Schreiber, S. L.; Liew,
W.-F. Tetrahedron Lett. 1983, 24, 2363.
(7) Tsunoda, T.; Suzuki, M.; Noyori, R. Tetrahedron Lett. 1980,
21, 1357.
(8) Gravel, D.; Murray, S.; Ladouceur, G. J. Chem. Soc., Chem.
Commun. 1985, 1828.
(9) Pitsch, S. Helv. Chim. Acta 1997, 80, 2286.
(10) Compound 5: 1H NMR (300 MHz, D2O, pD 8.5): = 3.78
(2 H, dd, J = 4.5, J = 7.5, CH2), 5.04 (1 H, t, J = 4.5, CH);
13C NMR (75 MHz, D2O, pD 8.5): = 68.1 (d, J = 5.5),
88.3(brs); 31P NMR (83 MHz, 1 mM EDTA in D2O, pD 9.0):
= –20.85 (t, J = 19.5), –9.70 (d, J = 19.5), –4.95 (d, J =
19.5); MS (ES-, CV -35V): m/z (%) = 321(20), [M4– + 2 H+
+ Na+ – H2O]–), 299(5) [M4– + 3 H+ – H2O]–).
References
(1) Müller, D.; Pitsch, S.; Kittaka, A.; Wagner, E.; Wintner, C.
E.; Eschenmoser, A. Helv. Chim. Acta 1990, 73, 1410.
(2) Krishnamurthy, R.; Arrhenius, G.; Eschenmoser, A. Origins
of Life and Evolution of the Biosphere 1999, 29, 333.
(3) Krishnamurthy, R.; Pitsch, S.; Arrhenius, G. Origins of Life
and Evolution of the Biosphere 1999, 29, 139.
(4) Dixit, V. M.; Laskovics, F. M.; Noall, W. I.; Poulter, C. D.
J. Org. Chem. 1981, 46, 1969.
(5) Compound 4: 1H NMR (500 MHz, D2O, pD 8.0): = 3.79
(2 H, dd, J = 4.5, J = 6.5, CH2), 5.04 (1 H, t, J = 4.5, CH);
13C NMR (125 MHz, D2O, pD 8.0): = 67.9 (d, J = 5.0), 88.5
(d, J = 9.0); 31P NMR (83 MHz, 10 mM EDTA in D2O, pD
8.0): = –9.25 (d, J = 22.0), –5.06 (d, J = 22.0); MS (ES-,
CV -15 V): m/z (%) = 237(100), [M3– + 2 H+]–; MS (ES-, CV
-25 V): m/z (%) = 237(100), [M3– + 2 H+]–), 219(100), [M3–
(11) Taylor, S. D.; Kluger, R. J. Am. Chem. Soc. 1993, 115, 867.
Synlett 2002, No. 1, 170–172 ISSN 0936-5214 © Thieme Stuttgart · New York