Six-membered ring phosphates and phosphonates as model compounds for cyclic phosphate prodrugs: Is the anomeric effect involved in the selective and spontaneous cleavage of cyclic phosphate prodrugs?

Article abstract of DOI:10.1021/jo8017473

(Chemical Equation Presented) In recent years, several six-membered ring phosph(on)ates and phosphonamides have been reported as potent prodrugs against liver diseases such as hepatitis B and C and also as antitumor agents. Apparently, the success for the

Full text of DOI:10.1021/jo8017473

Six-Membered Ring Phosphates and Phosphonates As Model
Compounds for Cyclic Phosphate Prodrugs: Is the Anomeric Effect
Involved in the Selective and Spontaneous Cleavage of Cyclic
Phosphate Prodrugs?
Silvano Cruz-Gregorio,† Vicente Rodriguez-Palacios,† Herbert Ho¨pfl,‡ Leticia Quintero,†
and Fernando Sartillo-Piscil*,†
Centro de InVestigacio´n de la Facultad de Ciencias Qu´ımicas, BUAP, 14 Sur Esq. San Claudio, San
Manuel, 72570, Puebla, Me´xico, and Centro de InVestigaciones Qu´ımicas, UniVersidad Auto´noma del
Estado de Morelos, AVenida UniVersidad 1001, 62209, CuernaVaca, Me´xico
fsarpis@siu.buap.mx
ReceiVed August 12, 2008
In recent years, several six-membered ring phosph(on)ates and phosphonamides have been reported as
potent prodrugs against liver diseases such as hepatitis B and C and also as antitumor agents. Apparently,
the success for their biological activity depends on the selective cleavage of the C4-O3 bond within the
respective P-heterocyclic ring. Empirical observations have suggested that the group attached to the C4
position (aryl or pyridyl group) is responsible for the selective cleavage. In this regard, we show in the
present work that the configuration at the P-atom, the conformation of the P-heterocyclic ring, and
particularly, the anomeric effect are involved in the spontaneous and selective cleavage of the C4-O3
bond in cyclic phosph(on)ates. We arrived at this assumption based on the conformational and
configurational study of simple model phosphates and phosphonates, where it was observed that the
spontaneous conversion of unstable six-membered ring phosphates to their most stable six-membered
ring phosphate (4d, 6d and 7d to 5d), by a selective C4-O3 bond cleavage, depends on both: the
stereochemistry of the aryl group at C4 and the electronic nature of the substituent attached to the P-atom.
Thus, we postulated that the anomeric effect weakens the C4-O3 bond within the 1,3,2-dioxaphospho-
rinane ring, favoring thus their selective cleavage and spontaneous conversion, similarly to the proposed
mechanistic mode of action of six-membered ring P-heterocyclic prodrugs.
Introduction
ring phosphates (2-oxo-1,3,2-dioxaphosphorinanes) and
derivatives.1-9 The information extracted from such studies
stimulated the Bentrude group10 to postulate an ingenious
model of interaction between the active site of the enzyme
and the dioxaphosphorinane ring of cyclic adenosine mono-
phosphate (cAMP), also known as the second messenger for
the cellular processes. On the basis of the empirical estimation
of the ∆G° for the chair h twist equilibrium of neutral model
One of the topics in physical-organic chemistry is the
conformational and configurational analysis of six-membered
* To whom correspondence should be addressed. Tel: +52 2222 2955500
ext. 7391.
† BUAP.
‡ Universidad Auto´noma del Estado de Morelos.
10.1021/jo8017473 CCC: $40.75
Published on Web 11/24/2008
2009 American Chemical Society
J. Org. Chem. 2009, 74, 197–205 197

Cruz-Gregorio et al.
FIGURE 1. Chair-twist equilibrium of cyclic adenosine monophosphate (cAMP).
SCHEME 1. Proposed Prodrug Cleavage Mechanism for
Cyclic 4-Aryl-phosphorinanes
phosphates, they proposed that cAMP in a twisted conforma-
tion could be appropriate for such enzyme-cAMP interac-
tions (Figure 1).10
On the other hand, more recently, Erion introduced a new
class of six-membered ring phosphates and phosphonates for
targeting phosph(on)ate-based drugs to the liver (HepDirect
prodrugs).12-15 Apparently, this new class of prodrugs combines
properties of rapid liver cleavage with high plasma and tissue
stability to achieve increased drug levels in the liver.12 An
additional feature of these new prodrugs is that the aryl group
attached to the C4 position in a cis orientation, seems to be a
crucial element for the biological activity.12 Mechanistic studies
suggested that prodrug I undergoes oxidation to the hydroxylated
intermediate II, which subsequently eliminates unsaturated
ketone IV and releases the biologically active nucleoside
monophosphate III (Scheme 1).12-15
Similarly, cyclic aryl phosphoramides V represent another
important class of prodrugs that have been used for drug delivery
in tumor treatments.16-18 Similar to Erion’s prodrugs, this cyclic
aryl-phosphoramides incorporate a p-nitrophenyl group at the
C4 position that after bioreduction by nitroreductasa (NTR)
affords hydroxylamine VI that facilitates the C4-O3 bond
cleavage producing the anionic cytotoxic species VII (Scheme
2).17,18
On this basis, it seems evident that the aryl group attached
to the C4 position of the heterocyclic ring provides the driving
force for the prodrug mechanism action; however, this is only
a simple observation.19 Therefore, we suppose that, as for
cAMP, where the conformation of the 1,3,2-dioxaphosphorinane
ring seems to be determinant for the mechanistic action in the
cellular media,10,11 the biological activity of these new cyclic
prodrugs should depend on the conformation of the heterocyclic
ring. In this sense, this article reports our results on the
conformational and configurational study of conformationally
restricted six-membered ring phosph(on)ates having an aryl
group with different electronic demand at the C4 position and
P-atom, which may provide mechanistic information on cyclic
prodrug cleavage.
We have recently carried out the conformational and con-
figurational study of a series of conformationally restricted
4-sustituted-1,3,2-dioxaphosphorinanes.20-22 In this study we
have found a new way for monitoring with high precision the
conformational equilibria between two conformers. For those
cyclic phosphates with RP configuration that have a phenyl group
equatorially oriented at C4 position, a chair-boat equilibrium
(C1hB1) was proposed,20-22 whereas a chairhtwist equilib-
rium (C1hT) was suggested for their SP-epimers (Scheme 3).
Conformers C2 and B2 (of which B2 has been proposed in
related cyclic phosphates23-25), were not considered in study,
first because the equilibrium C1hB1 was supported by trapping
both conformers in the solid state within the same crystal
´
(1) Frank, E.; Wolfling, J. Curr. Org. Chem 2007, 11, 1610.
(2) Gallagher, M. J. In Phosphorus-31 NMR Spectroscopy in Stereochem-
ical Analysis; Verkade, J. G., Quin, L., Eds; VCH: Deerfield Beach, FL,
1987; Chapter 9.
(3) Bentrude, W. G.; Setzer, W. N. Stereospecificity in 31P-Element
Coupling: Proton-Phosphorus Coupling In Phosphorus-31 NMR Spectroscopy
in Stereochemical Analysis; Verkade, J. G., Quin, L. D., Eds; VCH: Weinheim,
1987.
(4) Bentrude, W. G. Steric and Stereoelectronic Effects in 1,3,2-Dioxaphos-
phorinanes. In Methods in Stereochemical Analysis; Juaristi, E., Ed.; VCH: New
York, 1995.
(5) Gorenstein, D. G. In Phosphorus-31 NMR: Principles and Applications;
Gorenstein, D. G., Ed.; Academic Press: New York, 1984.
(6) Maryanoff, B. E.; Hutchins, R. O.; Maryanoff, C. A. Top. Stereochem.
1979, 11, 187.
(7) Gorenstein, D. G. Chem. ReV. 1987, 87, 1047.
(8) Nifant’ev, E. E.; Zavalishina, A. I. Russ. Chem. ReV. 1982, 51, 1601.
(9) Arshinova, R. P. Russ. Chem. ReV. 1984, 53, 351.
(10) Nelson, K. A.; Bentrude, W. G.; Setzer, W. N.; Hutchinson, J. P. J. Am.
Chem. Soc. 1987, 109, 4058.
(11) See also: (a) Yu, J. H.; Arif, A. M.; Bentrude, W. G. J. Am. Chem. Soc.
1990, 112, 7451.
(12) Erion, M. D.; Reddy, K, R.; Boyer, S. H.; Matelich, M. C.; Gomez-
Galeno, J.; Lemus, R. H.; Ugarkar, B. G.; Colby, T. J.; Schanzer, J.; van Poelje,
P. D. J. Am. Chem. Soc. 2004, 126, 5154.
(13) Erion, M. D.; van Poelje, P. D.; Mackenna, D. A.; Colby, T. J.; Montag,
A. C.; Fujitaka, J. M.; Linemeyer, D. L.; Bullough, D. A. J. Pharmacol. Exp.
Ther. 2005, 312, 554.
(14) Hecker, S.; Erion, M. D. J. Med. Chem. 2008, 51, 2328.
(15) Hecker, S. J.; Reddy, K. R.; van Poelje, P. D.; Sun, Z.; Huang, W.;
Varkhedkar, V.; Reddy, M. V.; Fujitaki, J. M.; Olsen, D. B.; Koeplinger, K. A.;
Boyer, S. H.; Linemeyer, D. L.; MacCoss, M.; Erion, M. D. J. Med. Chem.
2007, 50, 3891.
(19) Erion suggested that the prodrug cleavage proceeds via an initial
oxidation at C4 (see ref 12), and Hu suggested that an electron-donating group
in the aromatic ring facilitates the prodrug cleavage (see ref 16), however, no
fundamental explanations for the selective prodrug cleavage were given.
(20) Cruz-Gregorio, S.; Sa´nchez, M.; Clara-Sosa, A.; Be´rnes, S.; Quintero,
L.; Sartillo-Piscil, F. J. Org. Chem. 2005, 70, 7107.
(21) Sartillo-Piscil, F.; Cruz-Gregorio, S.; Sa´nchez, M.; Quintero, L. Tetra-
hedron 2004, 60, 3001.
(22) Sartillo-Piscil, S.; Cruz-Gregorio, S.; Sa´nchez, M.; Hopfl, H.; Anaya
de Parrodi, C.; Quintero, L. Tetrahedron 2003, 59, 4077.
(16) Borch, R. F.; Millard, J. A. J. Med. Chem. 1987, 30, 427.
(17) Hu, L.; Yu, C.; Jiang, Y.; Han, J.; Li, Z.; Browne, P.; Race, P. R.;
Knox, R. J.; Searle, P. F.; Hyde, E. I. J. Med. Chem. 2003, 46, 4818.
(18) Li, Z.; Han, J.; Jiang, Y.; Browne, P.; Knox, R. J.; Hu, L. Bioorg. Med.
Chem. 2003, 11, 4171.
(23) Hermans, R. J. M.; Buck, H. M. J. Org. Chem. 1988, 53, 2077.
(24) Neeser, J.-T.; Tronchet, J. M. J.; Charollais, E. J. Can. J. Chem. 1983,
61, 13.
(25) Morr, M.; Ernts, L.; Mengel, R. Liebigs Ann. Chem. 1982, 651.
198 J. Org. Chem. Vol. 74, No. 1, 2009

Anomeric Effect in the SelectiVe CleaVage of Prodrugs
SCHEME 2. Proposed Prodrug Cleavage Mechanism for Cyclic 4-Nitro-aryl-phosphoramides
SCHEME 3. Conformations for Conformationally Restricted 4-Substituted-1,3,2-dioxaphosphorinanes
TABLE 1. Synthesis of the Diastereoisomeric 1,3-Diol Precursors
2 and 3
arising: (i) what is the role of the configuration and conformation
of the 1,3,2-dioxaphosphorinane ring in the prodrug cleavage
mechanism? (ii) Is the prodrug cleavage initiated by delocal-
ization of the oxygen lone pair into the aromatic ring facilitating
the C4-O3 bond cleavage? As previously proposed for cyclic
phosphoramide prodrugs, or (iii) is the cleavage a combination
of both contributions? To answer the above questions, we
synthesized a series of conformationally restricted 4-aryl-1,3,2-
dioxaphosphorinanes to analyze the effect that the aryl group
placed at C4 position exerts on the conformation and config-
uration of the 1,3,2-dioxaphosphorinane ring.
Results and Discussions
The syntheses of the 4-aryl-1,3,2-dioxaphosphorinanes 4-7
were achieved in two steps starting from diacetone-D-glucose
1. First, the diastereomeric 1,3-diol precursors 2 and 3 were
prepared applying our one-pot procedure of sequential hydroly-
sis-oxidation-Grignard reagent addition to the commercially
available diacetone-D-glucose (Table 1).22 Phosphorylation of
1,3-diols with phenyldichlorophosphate and triethylamine in
CH2Cl2 gave the corresponding diastereomeric 1,3,2-dioxaphos-
phorinane pairs in good yields (Scheme 4).
a Yields are for the mixture of diastereoisomers. b Information
extracted from ref 22.
The diastereoisomeric mixtures were separated by column
chromatography (except for 4d, 6d, and 7d) allowing to record
1
and assign completely the H, 13C and 31P NMR data for each
lattice22 (for R ) Ph equatorially oriented), and second, because
the C1hT equilibrium could be established based on NMR data.
Interestingly, a reverse situation was observed for cyclic
phosphate having RP configuration and when R ) Ph is axially
oriented. In this case, the equilibrium observed was C1hT
instead the expected C1hB1 equilibrium. Apparently, the
pseudoaxial force of the OPh group, which should induce that
the heterocyclic ring acquires the B1 conformation, was inhibited
by steric interactions when the phenyl group is axially
oriented.26-31 It is important to note that the relative orientation
between the phenoxy and the phenyl group was cis, and
coincidently, Erion had found that better prodrugs were those
with cis relative orientation.12 Thus, the following questions are
(26) The phenyl group placed at the C4 position has been used to lock
the spontaneous chair-chair equilibrium of 1,3,2-dioxaphosphorinane rings
into a chair conformation that orients the phenyl group equatorially. See
refs 27-31.
(27) Jankowski, S.; Marczak, J.; Olczak, A.; Glowka, M. L. Tetrahedron
Lett. 2006, 47, 3341.
(28) Ten Hoeve, W.; Wynberg, H. J. Org. Chem. 1985, 50, 4508.
(29) Hulst, R.; Zijlstra, R. W. J.; de Vries, N. K.; Feringa, B. L. Tetrahedron:
Asymmetry 1994, 5, 1701.
(30) Hulst, R.; Visser, J. M.; de Vries, N. K.; Zijlstra, R. W. J.; Kooijman,
H.; Smeets, W.; Spek, A. L.; Feringa, B. L. J. Am. Chem. Soc. 2000, 122, 3135.
(31) Sartillo-Piscil, F.; Meza-Leo´n, R.; Quintero, L. ReV. Soc. Quim. Me´x.
2002, 46, 330.
(32) It is important to mention that in this work, the C4 position of the 1,3,2-
dioxaphosphorinane nomenclature corresponds to the C5′ position of xylofuranose
nomenclature.
J. Org. Chem. Vol. 74, No. 1, 2009 199

Cruz-Gregorio et al.
SCHEME 4. Synthesis of the 4-Aryl-1,3,2-dioxaphosphorinanes 4-7
1
TABLE 2. Representative H and 31P NMR Chemical Shifts for
TABLE 3. Representative Coupling Constants for Cyclic
,
Cyclic Phosphatesa b
Phosphatesa b
,
phosphate
H1′
H2′
H3′
H4′
H5′
31P
phosphate
H5′-P
H3′-P
H5′-H4′
H4′-H3′
4a
5a
6a
7a
4b
5b
6b
7b
4c
5.60
6.02
6.01
6.15
5.63
6.03
6.02
6.15
5.63
6.02
6.02
6.16
5.63
6.06
5.99
6.14
4.61
4.73
4.78
4.83
4.62
4.73
4.79
4.84
4.62
4.73
4.79
4.84
4.61
4.73
4.77
4.83
5.12
5.01
4.95
4.92
5.12
5.00
4.96
4.93
5.12
5.00
4.94
4.91
5.09
4.98
4.96
4.98
4.42
4.40
4.53
4.63
4.38
4.37
4.51
4.61
4.38
4.37
4.49
4.59
4.39
4.36
4.52
4.63
5.84
5.75
5.63
5.68
5.83
5.73
5.57
5.64
5.82
5.73
5.55
5.63
5.78
5.69
5.57
5.66
-12.9
-15.2
-13.8
-13.0
-12.6
-14.9
-12.9
-12.1
-12.6
-15.0
-12.9
-12.0
-12.4
-14.8
-13.4
-12.9
4a
5a
6a
7a
4b
5b
6b
7b
4c
<1
<1
10.2
11.4
<1
<1
9.6
10.8
<1
<1
9.9
10
4.8
<1
7.2
3.6
4.8
<1
7.2
3.2
4.8
<1
6.8
3.6
4.5
<1
-
2.3
1.6
3.6
4.0
1.6a
<1
4.0
4.4
<1
<1
4.4
4.4
<1
<1
3.6
3.6
3.0
2.0
3.2
3.2
2.8
2.0
3.6
3.2
2.4
2
3.6
3.6
2.1
2.1
3.6
3.2
5c
6c
5c
6c
7c
7c
4dc
5d
6dc
7dc
4dc
5d
6dc
7dc
<1
<1
10.0
9.3
-
a Spectra recorded at 400 and 161.8 MHz in CDCl3 for 1H and 31P,
respectively. b Chemical shifts (δ) given in ppm. c NMR data measured
from the crude reaction mixture.
a Spectra recorded at 400 and 161.8 MHz in CDCl3 for 1H and 31P,
respectively. b Coupling constants (J) given in Hz. c NMR data
measured from the crude reaction mixture.
cal calculations.20 Due to the structural relationship and similar
NMR data for 4b, 4c, and 4d, the same C1hB1 can be
proposed. For their P-epimers 5a-d, in which the phenoxy
groups are axially and the aryl groups are equatorially oriented,
it can be proposed that the C1 conformer is highly populated.
The X-ray crystallographic study for phosphate 5d confirmed
that the assignments of their conformation and configuration
are correct (Figure 3).33
cyclic phosphate, in order to establish the absolute configuration
at the phosphorus atom, and the C4 carbon (or C5′),32 as well
as to determine the conformational equilibrium for the 1,3,2-
dioxaphosphorinane rings (see Tables 2 and 3).
For phosphates 4a and 5a, the absolute configuration at the
phosphorus atom has been determined previously, taking into
account an analysis of the chemical shifts of the anomeric
hydrogen atoms (H1′).22 The remaining phosphates 4b-d and
5b-d were assigned by analogy. Thus, when the OPh group is
oriented cis to H1′, the signal of this anomeric hydrogen atom
is shifted to upper field (compared with its P-epimer) and the
absolute configuration at the phosphorus atom is RP. For this
reason, phosphates 4a, 4b, 4c, and 4d possess RP configuration,
and phosphates 5a, 5b, 5c, and 5d SP configuration (see Figure
2).
The value of the vicinal H-P coupling constants for 4a-d
and 5a-d (3JH5′-P < 1 Hz) suggests that the aryl group is located
in equatorial or pseudoequatorial position. Therefore, the
absolute configuration at C4 (or C5′) have to be R. As mentioned
above, for phosphate 4a, a C1hB1 equilibrium was proposed
previously,21,22 based on NMR, X-ray diffraction22 and theoreti-
On the other hand, the determination of the configuration at
the phosphorus atoms for the cyclic phosphates 6 and 7 turned
out to be more complicated. It is important to mention that based
on Gorenstein’s criteria (which establish that upfield shifted 31
P
NMR signals can be attributed to phosphorinanes having their
phenoxy group axially oriented34-36), the absolute configuration
at the P-atom for 6a and 7a has been previously assigned.22
(33) Crystallographic data were deposited in Cambridge Crystallographic Data
Base (CCDC-660542).
(34) Gorenstein, D. G.; Rowell, R. J. Am. Chem. Soc. 1979, 101, 4925.
(35) Gorenstein, D. G.; Rowell, R.; Findlay, J. J. Am. Chem. Soc. 1980, 102,
5077.
(36) Majoral, J. P.; Pujol, R.; Navech, J. Bull. Soc. Chim. Fr. 1972, 606.
200 J. Org. Chem. Vol. 74, No. 1, 2009

Anomeric Effect in the SelectiVe CleaVage of Prodrugs
FIGURE 2. Assignment of the absolute configuration at the phosphorus atom in phosphates 4 and 5.
FIGURE 3. Perspective view of the molecular structure of 5d showing that it possesses C1 conformation.
However, we now believe that this criteria could give misleading
assignments of the P-atom configuration for phosphorinanes that
are unable to permanently orient the phenoxy group in an axial
position. Unfortunately, our own criteria for the assignment of
the configuration at P-atom can not be applied because both:
the proposed chair C1 (with the phenoxy group axially oriented)
and the twist conformation T orient the phenoxy groups away
from H1′. However, according to the 3JH5′-P coupling constants
for phosphates 6 and 7 (3JH5′-P ) 9.0-11.5 Hz, see Table 3) a
C1hT equilibrium that is strongly directed to T could be
suggested. This was corroborated by X-ray crystallographic
study of phosphate 6c,37 showing an almost perfect T confor-
mation (Figure 4).
In the perspective view given in Figure 4, we can see that
the aryl group at C4 is located in an apparent pseudoequatorial
position, so that the phosphorinane ring acquires the T
conformation. We can also observe that the phenoxy group is
directed away from H1′, and the presence of any shielding effect
on H1′ becomes evident. The X-ray crystallographic study of
phosphate 6c allowed determining the absolute configuration
at the P-atom for the diastereomeric phosphate mixture (6c/
7c). Therefore, with the aid of their 31P NMR chemical shifts
we could assign the absolute configuration at the P-atom for
the rest of the cyclic phosphates. Thus, cyclic phosphates shifted
to higher field like 6a, 6b, and 6d (-13.8, -12.9 and -13.4
ppm, respectively) were assigned as RP, and their respective
diastereoisomeric cyclic phosphates 7a, 7b, and 7d (-13.0,
-12.1 and -12.9 ppm, respectively) were assigned as SP (Table
2). Thus, with the aid of the crystallographic study for 6c, we
realized that the absolute configurations at the P-atom for cyclic
phosphates 6a and 7a were previously misassigned (cyclic
phosphates 4b and 4b′ in reference 22).
Apparently, the conformational and configurational behavior
of all new 4-aryl-1,3,2-dioxaphosphorinanes resulted as ex-
pected, according to the conformational equilibria proposed in
Scheme 3. Furthermore, a very interesting result was observed
when we tried to phosphorylate the 1,3-diol 2d under standard
conditions (see Scheme 4). Analyzing the crude mixture
reaction, an equimolar amount of cyclic phosphates 4d and 5d
was observed; however, when we tried to separate them by
column chromatography, we obtained a fraction that contained
only 5d in high yield (65%) plus a fraction which was a mixture
of 4d and 5d in a ratio of 2:1 (15%). An even more interesting
observation was made for 1,3-diol 3d. Under the same reaction
conditions for the phosphorylation, diol 3d was not converted
to the expected cyclic phosphates 6d and 7d, instead 5d was
obtained exclusively38 (Scheme 5). This suggests that under this
reaction conditions, the cyclic phosphates 6d and 7d are rapidly
and spontaneously converted to the apparently more stable cyclic
(37) Crystallographic data were deposited in Cambridge Crystallographic Data
Base (CCDC-660543).
(38) Observedby1Hand31P-NMRafterpurificationbycolumnchromatography.
J. Org. Chem. Vol. 74, No. 1, 2009 201

Cruz-Gregorio et al.
FIGURE 4. Perspective view of the molecular structure of 6c showing that it possesses T conformation.
SCHEME 5. Unexpected Phosphorylation of 1,3-Diols, 2d and 3d
phosphate 5d. To prove this, diol 3d was treated with a stronger
base (t-BuOK) at -78 °C and indeed, initially an almost
therefore we found that t85 (time to convert 85% of 6d and 7d
to 5d) was >45 h for 4d, 7.5 h for 7d and 6.5 h for 6d (Figure
6 and Table 4). The observation of a linear behavior in the
spontaneous conversion suggests nonequilibration processes of
4d, 6d, and 7d during the conversion to 5d, so that a common
intermediary F it might be proposed, which is followed by a
very fast recombination. This mechanism would be similar to
the prodrug cleavage mechanism proposed for cyclic aryl-
phosphoramides.17
These results indicate that phosphate 5d is the most stable
cyclic phosphate among the four possible diastereoisomers (4d,
5d, 6d, and 7d, see Scheme 7). This is because, first of all, the
axial orientation of the phenoxy group at the phosphorus atom
provides stability to 5d by the anomeric effect,40,41 and
additionally, the equatorial orientation of the aryl group at the
1
equimolar amount of 6d and 7d was observed by H and 31P
NMR, which within a period of 8 h at room temperature in
CDCl3, were spontaneously converted to 5d (Scheme 5).
Apparently, the cyclic phosphates 4d, 6d, and 7d prefer to
spontaneously convert to 5d rather than hydrolyze the 1,3,2-
dioxaphosphorinane ring.39 By monitoring the phosphorylation
reactions of diols 2d and 3d, we could determine the time
required for the spontaneous conversions of 4d, 6d, and 7d to
5d. The cyclic phosphates 6d and 7d are converted to 5d in
approximately 10 h in CDCl3, meanwhile phosphate 4d required
more than 50 h in CDCl3 to convert into 5d (Scheme 6).
It is important to mention that the conversions of 4d, 6d,
and 7d into 5d follow a linear behavior until 85% of conversion,
(39) It is well documented that cyclic phosphates undergo hydrolysis in protic
media or basic conditions (see refs 7 and 34), however, the cyclic phosphates
4d, 6d, and 7d showed to be resistant to hydrolysis in methanol.
(40) Juaristi, E.; Cuevas, G. The Anomeric Effect; CRC Press, Inc.: Boca
Raton, FL, 1995.
(41) Juaristi, E.; Cuevas, G. Tetrahedron 1992, 48, 5019.
202 J. Org. Chem. Vol. 74, No. 1, 2009

Anomeric Effect in the SelectiVe CleaVage of Prodrugs
SCHEME 6. Proposed Mechanism for the C4-O3 Bond Cleavage of Phosphates 4d, 6d, and 7d and Proposed Putative
Intermediate F
SCHEME 7. Proposed Route for the Conversion of 4d, 6d,
and 7d to 5d
SCHEME 8. Anomeric Effect in 1,3,2-Dioxaphosphorinanes
(eq 1) That Might Weaken the C4-O3 Bonds (eq 2)
SCHEME 9. Postulated Mesomers H, G, and F Involved in
the Conversion of 4d, 6d, and 7d to 5d
C4 position reduces steric interactions considerably (see X-ray
structure 5d in Figure 3). In the case of 4d, the nonaxial
orientation of the phenoxy group does not provide stabilization
by the anomeric effect; meanwhile strong steric repulsions are
present in 7d due to the axial orientation of the aryl group at
C4. The absence of an anomeric effect (nonaxially orientation
of phenoxy group), and strong steric repulsions (axial orientation
of the aryl group at C4), make phosphate 6d the most unstable
cyclic phosphate and, therefore, it converts more rapidly into
the most stable phosphate 5d (see Scheme 7).
A question that still has to be addressed is: how does the
anomeric effect affect the spontaneous cleavage of the C4-O3
bond? Thus, if it is assumed that the anomeric effect consists
in the interaction of two orbitals in the ground state
(nOTσ*P-OR),7,40,41 giving thus a double-bond-no-bond reso-
nance structure B (Scheme 8, eq 1), then a congruent explanation
for the C4-O3 bond cleavage is that the contribution of
mesomer B weaken the C4-O3 bond strength, thus favoring
the formation of mesomer C (Scheme 8, eq 2).
Now, it is evident that the spontaneous conversion of 4d, 6d
and 7d to 5d is not exclusively caused by the delocalization of
the oxygen lone pair through the aromatic ring, as previously
considered17,18 (based on the analogy with phosphoramides
prodrugs, see Schemes 2 and 6), but that the configuration and
the conformation of the 1,3,2-dioxaphosphorinane rings are also
determinant, and therefore, we postulate that: in the spontaneous
conversion of 4d, 6d and 7d to 5d, the anomeric effect plays a
major role in the above-mentioned, prodrug cleavage mechanism.
J. Org. Chem. Vol. 74, No. 1, 2009 203

Cruz-Gregorio et al.
SCHEME 10. Synthesis of 4-Alkyl-1,3,2-dioxaphosphonates 8-11
SCHEME 11. Route Proposed for the Conversion of 11d to the More Stable Cyclic Phosphonate 8d
SCHEME 12. Orbital Interactions (nOT π*PdO) in
1,3,2-Dioxaphosphonates (Anomeric Effect)
Therefore, the previous assumption that: “the cleavage of the
C4-O3 bond is caused by a simple delocalization of the oxygen
lone pair through the aromatic ring affording intermediate F”
(see Scheme 6) seems to be insufficient to explain our results.
Therefore, mesomers G and H should be proposed as additional
electronic structures involved in the spontaneous cleavage of
the C4-O3 bond in 1,3,2-dioxaphosphorinanes (Scheme 9).
With this new assumption related to the intervention of the
anomeric effect in the spontaneous cleavage of the C4-O3
bond, we should expect a strong manifestation of above
mesomers when an efficient electron-donating group is attached
to C4 and when the antibonding σ*P-X orbital is a good electron
acceptor (e. g., P-OPh).42,43 In this context, it is reasonable to
expect a lower spontaneous conversion of cyclic phosphates, if
the phenoxy group is replaced by a less electronegative group,
such as an alkyl group. In order to prove this statement, we
proceeded to synthesize and examine a series of 4-alkyl-1,3,2-
dioxaphosphonates, which have been prepared from 1,3-diols
(2b-d and 3b-d) through phosphorylation reactions using
propyl dichlorophosphonate (Scheme 10).
FIGURE 5. Perspective view of the molecular structure of 8d showing
that it possesses C1 conformation.
that phosphonate 11d was the only phosphonate that converted
to the more stable cyclic phosphonate 8d in 60% yield by NMR
over 30 days (Scheme 11).
Now, phosphonate 8d is the most stable cyclic phosphonate,
because the aryl group is oriented equatorially and the phos-
phoryl group is oriented axially. The latter provides stabilization
by the anomeric effect, although in this case through different
orbital interactions (nOTπ*P ) O) (Scheme 12).45,46
The molecular structure and conformation (in solid state) of
phosphonate 8d has been confirmed by X-ray diffraction (Figure
5).47
As we anticipated, the cyclic 4-aryl-phosphonates 8d, 9d, 10d,
and 11d have been shown to be very stable in solution and are
stable to chromatography.44 However, it is important to mention
(44) Fully spectroscopic characterization is provided in the Supporting
Information.
(45) Herna´ndez, J.; Ramos, R.; Sastre, N.; Meza, R.; Hommer, H.; Salas,
M.; Gordillo, B. Tetrahedron 2004, 60, 10927.
(46) Verkade, J. G. Phosphorus Sulfur 1976, 2, 251.
(47) Crystallographic data were deposited in Cambridge Crystallographic Data
Base (CCDC-687297).
(42) Deslongchamps, P. Kinetic Anomeric Effect. In Stereoelectronic effects
in Organic Chemistry; Pergamon: Oxford, 1983.
(43) Kirby, A. J. The Anomeric and Related Stereoelectronic Effects at
Oxygen; Springer: Berlin, 1983.
204 J. Org. Chem. Vol. 74, No. 1, 2009

Anomeric Effect in the SelectiVe CleaVage of Prodrugs
heterocyclic systems and will function analogously albeit on
different time scales. In this regard, related experimental efforts
and theoretical calculations are in progress and will be published
in due course.
Conclusions
We can summarize the following findings: the 1,2-O-
isopropylidene-xylofuranose moiety functions efficiently as
conformationally restricted template for the conformational and
configurational study of cyclic phosph(on)ates, providing thus
a convenient way for monitoring conformational equilibria and
assigning the relative configuration at the P-atom. Experimental
evidence that support that the anomeric effect can weaken the
C4-O3 bond in six-membered ring phosph(on)ates, causing a
spontaneous conversion of conformationally unstable cyclic
phosph(on)ates to their corresponding cyclic more stable phos-
ph(on)ates has been presented. Finally, this spontaneous conver-
sion of phosphates 6d and 7d to 5d represents a novel two step
way for the inversion of the configuration at the C5-atom in
some 5-aryl-xylofuranose derivatives (e.g., 3d f 2d).48
FIGURE 6. Conversion of 6b and 7b versus formation of 5d.
TABLE 4. Conversion of 6d and 7d versus Formation of 5d
Monitored by 31P NMR
time (min) phoshate 7d (%) phosphate 6d (%) phosphate 5d (%)
0
5
10
15
20
25
30
45
22.3
22.2
21.7
21.0
21.0
20.9
20.8
20.3
19.5
18.4
15.6
15.0
14.4
13.2
13.1
12.5
12.6
11.4
7.3
43.8
43.3
42.5
42.0
41.1
40.6
39.4
37.7
35.3
32.7
30.5
27.8
25.6
24.0
22.7
21.1
19.2
18.3
13.7
12.7
11.6
10.4
9.3
33.9
34.5
35.8
37.0
37.9
38.5
39.8
42.0
45.2
48.9
53.9
57.3
60.1
62.8
64.2
66.4
68.1
70.3
79.0
80.4
82.0
83.6
85.1
85.0
Experimental Section
General. The reagents were obtained from commercial sources
and used without purification. The solvents were used as technical
grade, and freshly distilled prior use. NMR studies were carried
out with 400 and 300 MHz spectrometers. Internal reference (TMS)
1
for H and 13C. Chemical shifts are stated in parts per million.
COSY, HSQC, and NOESY experiments have been carried out in
order to assign the 1H and 13C spectra completely. High resolution
Mass spectra (FAB+ ion mode).
60
75
90
105
120
135
140
150
165
180
220
235
250
265
325
340
Protocol for the Synthesis of Unstable Cyclic Phosphates
6d and 7d and Determination of t85. To a solution of diol 3d
(0.2 g, 0.67 mmol) and potassium t-butoxide (1.69 mL, 1.7 mmol)
in dry Et2O (30 mL) at -78 °C, was added dropwise phenyl
dichlorophosphate dissolved in 2 mL of dry Et2O (0.12 mL, 0.8
mmol). The reaction mixture was allowed to stir for 10 min before
to add 1 mL of H2O at -30 °C. The organic phase was separated
from the aqueous phase and dried with Na2SO4. The organic phase
was evaporated under reduced pressure at -20 °C. NMR data for
unstable phosphates 6d and 7d were obtained from the crude
reaction mixture (Table 4).
6.9
6.3
6.0
5.6
5.0
8.0
Acknowledgment. We gratefully acknowledge the financial
support from CONACyT (grant number: 62203). We dedicate
this work to Professor Dr. Hermann Roth for the unconditional
support to this research group.
These results demonstrate that aryls group having an electron-
donating group placed at C4 position are not sufficient to
promote the C4-O3 bond cleavage in 1,3,2-dioxaphosphorinane
compounds, but that stereoelectronic interactions are even more
important and determinant factors.
It is noteworthy to mention that, in this work have been
presented experimental evidence that suggest that both cyclic
prodrug cleavage mechanism (for the cyclic phosph(on)ate and
phosphoramide prodrugs) might function similarly. However,
it is also important to understand that the results presented here
do not attempt to show a close relationship between instability
and biologically activity of the cyclic prodrugs, however, we
do try to expose, by affecting the reactivity or stability of the
cyclic phosphates, that stereoelectronic interactions like the
anomeric effect are involved in the cyclic prodrugs cleavage.
Additionally, although the presented study was carried out with
cyclic phosph(on)ates, we anticipated similar behavior with other
Supporting Information Available: Experimental section
including spectroscopic, crystallographic data, and Cif files for
compounds 5d, 6c, and 8d. This material is available free of
charge via the Internet at http://pubs.acs.org.
JO8017473
(48) First, phosphorylation, then spontaneous conversion (in situ), and finally
removal of the phosphate group with lithium aluminium hydride (LAH):
J. Org. Chem. Vol. 74, No. 1, 2009 205