Utilization of 1,3-Dioxolanes in the Synthesis of α-branched Alkyl and Aryl 9-[2-(Phosphonomethoxy)Ethyl]Purines and Study of the Influence of α-branched Substitution for Potential Biological Activity

Article abstract of DOI:10.1080/15257770.2018.1506884

Syntheses of α-branched alkyl and aryl substituted 9-[2-(phosphonomethoxy)ethyl]purines from substituted 1,3-dioxolanes have been developed. Key synthetic precursors, α-substituted dialkyl [(2-hydroxyethoxy)methyl]phosphonates were prepared via Lewis acid mediated cleavage of 1,3-dioxolanes followed by reaction with dialkyl or trialkyl phosphites. The best preparative yields were achieved under conditions utilizing tin tetrachloride as Lewis acid and triisopropyl phosphite. Attachment of purine bases to dialkyl [(2-hydroxyethoxy)methyl]phosphonates was performed by Mitsunobu reaction. Final α-branched 9-[2-(phosphonomethoxy)ethyl]purines were tested for antiviral, cytostatic and antiparasitic activity, the latter one determined as inhibitory activity towards Plasmodium falciparum enzyme hypoxanthine-guanine-xanthine phosphoribosyltransfesase. In most cases biological activity was only marginal.

Full text of DOI:10.1080/15257770.2018.1506884

Nucleosides, Nucleotides and Nucleic Acids
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Utilization of 1,3-Dioxolanes in the
Synthesis of α-branched Alkyl and Aryl 9-[2-
(Phosphonomethoxy)Ethyl]Purines and Study
of the Influence of α-branched Substitution for
Potential Biological Activity
Karel Pomeisl, Radek Pohl, Robert Snoeck, Graciela Andrei & Marcela
Krečmerová
To cite this article: Karel Pomeisl, Radek Pohl, Robert Snoeck, Graciela Andrei & Marcela
Krečmerová (2018): Utilization of 1,3-Dioxolanes in the Synthesis of α-branched Alkyl and
Aryl 9-[2-(Phosphonomethoxy)Ethyl]Purines and Study of the Influence of α-branched
Substitution for Potential Biological Activity, Nucleosides, Nucleotides and Nucleic Acids, DOI:
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NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
https://doi.org/10.1080/15257770.2018.1506884
Utilization of 1,3-Dioxolanes in the Synthesis of
a-branched Alkyl and Aryl 9-[2-(Phosphonomethoxy)
Ethyl]Purines and Study of the Influence of a-branched
Substitution for Potential Biological Activity
Karel Pomeisla,b, Radek Pohla, Robert Snoeckc, Graciela Andreic, and
ꢀ
ꢁa
Marcela Krecmerova
aInstitute of Organic Chemistry and Biochemistry, Czech Academy of Sciences, Prague, Czech
b
c
Republic; Institute of Physics, Czech Academy of Sciences, Prague, Czech Republic; Rega
Institute for Medical Research, Katholieke Universiteit Leuven, Belgium
ABSTRACT
ARTICLE HISTORY
Received 1 March 2017
Accepted 26 July 2018
Syntheses of a-branched alkyl and aryl substituted 9-[2-(phos-
phonomethoxy)ethyl]purines from substituted 1,3-dioxolanes
have been developed. Key synthetic precursors, a-substituted
dialkyl [(2-hydroxyethoxy)methyl]phosphonates were prepared
via Lewis acid mediated cleavage of 1,3-dioxolanes followed
by reaction with dialkyl or trialkyl phosphites. The best prepa-
rative yields were achieved under conditions utilizing tin tetra-
chloride as Lewis acid and triisopropyl phosphite. Attachment
of purine bases to dialkyl [(2-hydroxyethoxy)methyl]phospho-
nates was performed by Mitsunobu reaction. Final a-branched
9-[2-(phosphonomethoxy)ethyl]purines were tested for antivi-
ral, cytostatic and antiparasitic activity, the latter one deter-
mined as inhibitory activity towards Plasmodium falciparum
enzyme hypoxanthine-guanine-xanthine phosphoribosyltrans-
fesase. In most cases biological activity was only marginal.
KEYWORDS
Acyclic nucleoside
phosphonates; PMEG; PEEG;
1,3-dioxolanes; dialkyl
phosphite; trialkyl
phosphite
Introduction
Acyclic nucleoside phosphonates (ANPs) are compounds with a broad
spectrum of biological activities.[1] Three of them became already marketed
antiviral drugs: cidofovir, 1-(S)-[3-hydroxy-2-(phosphonomethoxy)propyl]-
cytosine, for the treatment of cytomegalovirus retinitis[2] and off
label also for the treatment of several other DNA-viral infections[3] and
antiretrovirals adefovir, 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA)
and tenofovir, (R)-9-[2-(phosphonomethoxy)propyl]adenine (PMPA). The
last two compounds are marketed as oral prodrugs: adefovir dipivoxil
(anti-hepatitis B drug HepseraTM) and tenofovir disoproxil fumarate (TDF,
anti-hepatitis B and anti-HIV drug VireadTM).[4,5]
ꢀ
ꢁ
CONTACT Marcela Krecmerova
marcela@uochb.cas.cz
Institute of Organic Chemistry and Biochemistry,
Czech Academy of Sciences, Prague, Czech Republic.
ꢁ
ꢁ
Dedicated to the memory of Professor Antonın Holy (1936–2012).
ß 2018 Taylor & Francis Group, LLC

2
K. POMEISL ET AL.
Special attention is currently being paid to the newly developed prodrug
tenofovir alafenamide (TAF), a compound with a greater antiviral activity
and better distribution into lymphoid tissues than TDF.[6]
Purine ANPs of the (phosphonomethoxy)ethyl (PME) series are also
reported as cytostatic agents.[7] These compounds are: N-9-[2-(phosphono-
methoxy)ethyl]guanine (PMEG), N-9-[2-(phosphonomethoxy)ethyl]-2,6-
diaminopurine (PMEDAP) and its N6-alkyl derivatives, e.g. N6-cyclopropyl-
9-[2-(phosphonomethoxy)ethyl]-2,6-diaminopurine
(cPrPMEDAP).[8]
Moreover, the bis(amidate) prodrug of cPrPMEDAP (GS-9219) was investi-
gated by Gilead Sciences in clinical trials as a drug for the treatment of
non-Hodgking lymphoma. Unfortunately, the trials had to be discontinued
due to numerous side effects seen in humans but not observed previously
in animal models. Later on, the compound was licensed to VetDC com-
pany and further investigated as the veterinary anti-cancer agent VDC-
1101. Recently, the drug has been approved by FDA as a cytostatic for the
R
treatment of canine lymphoma under the trade name TanoveaV-CA1.[9]
Some guanine and hypoxanthine ANPs are currently also investigated as
potential antiparasitic agents, especially antimalarials. The target is the purine
salvage enzyme of the Plasmodium parasite hypoxanthine-guanine-xanthine
phosphoribosyltransferase (HGXPRT). In general, 6-oxopurine phosphoribo-
syltransferases (PRTases) synthesize the purine nucleoside monophosphates
required for nucleic acid production. The human enzyme can only use hypo-
xanthine or guanine as its naturally occurring purine substrates while the
Plasmodium enzyme is also able to utilize xanthine as a substrate.
Plasmodium falciparum lacks not only the de novo pathway for purine
nucleotide synthesis but also other purine salvage enzymes that provide alter-
native pathways to purine nucleotides. The catalytic activity of HGXPRT is
therefore essential for Plasmodium falciparum survival and inhibition of this
enzyme should arrest growth and division of the parasite.[10,11]
Various ANPs containing a 6-oxopurine base were found to be good
inhibitors of the human and Plasmodium falciparum 6-oxopurine phos-
phoribosyltransferases. Activity found in 9-[2-(phosphonomethoxy)ethyl]-
guanine (PMEG), (S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]guanine
(HPMPG) and 9-[2-(phosphonoethoxy)ethyl)]guanine (PEEG) encouraged
their further modifications, e.g. synthesis of hypoxanthine PEE derivative
(PEEHx),[11] various a- or b-branched PEE derivatives,[12] branched aza-
ANPs[13] or 2-hydroxy-3-phosphonomethyl (“iso-HPMP”) derivatives.[14]
The inhibitory potency and selectivity (Plasmodium enzym PfHGXPRT vs
the human HGPRT) is affected by acyclic chain structure, especially its
length and branching (Figure 1).[11–14]
In this study, we describe synthesis of novel phosphonomethoxyethyl
(PME) derivatives with modifications in the base moiety and branching in

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
3
the aliphatic side-chain (Figure 2). Compounds were designed to investigate
the potential influence of this a-branching of the (phosphonomethoxy)ethyl
chain on their ability to inhibit PfHGXPRT. In addition, general screening
of other biological activities (antiviral, cytostatic) of this class of ANP will
be performed regarding results of other side-chain branched derivatives
reported previously.[15–17] Especially, we directed our efforts to develop-
ment of preparative synthetic methods for the introduction of alkyl and
aryl substituents to the ANP phosphonomethoxy group.
Since syntheses of a-branched PMEG derivatives have never been
described in the literature, their development still remains a synthetic and
biochemical challenge.
Results and discussion
The choice of methods for the introduction of alkyl or aryl substituents to
the phosphonomethyl group of ANPs is strongly limited.[15–17] The reason
for that, in our experience, is worse reactivity of secondary phosphonoalkyl
alcohols and corresponding phosphonoalkyl bromides or tosylates utilized
as key ANP precursors (Scheme 1).
Figure 1. ANPs with inhibitory activity towards HG(X)PRT.
Figure 2. New a-branched (phosphonomethoxy)ethyl (PME) derivatives prepared in this study.

4
K. POMEISL ET AL.
Secondary phosphonoalkyl alcohols 2,3 were prepared by the published
procedure from aldehyde 1 (acetaldehyde or benzaldehyde) and diisopropyl
phosphite in the presence of sodium methoxide.[18] Their tosylation carried
[19]
out in the presence of triethylamine and 4-dimethylaminopyridine
afforded tosylates 4 and 5. The bromo derivative 7 was synthesized by
Arbuzov reaction from benzyl bromide and triisopropyl phosphite[20] fol-
lowed by bromination of the intermediate 6 with N-bromosuccinimide.[21]
While introduction of various leaving groups to form intermediates 4, 5
and 7 proceeded satisfactorily, further conversion of these compounds to
the desired a-branched (phosphonomethoxy)ethyl synthons 8 completely
failed (Scheme 1, reactions (b) and (d)).
On the other hand, syntheses using Lewis acid mediated 1,3-dioxolane
cleavage afforded the appropriate building blocks 8a-e in preparative yields
(Scheme 2, Table 1).
1,3-Dioxolanes 10a-g were obtained by the reaction of commercially
available carbonyl compounds 9a-g with 1,2-dihydroxyethane catalyzed
by p-toluenesulfonic[22] or hydrochloric acid. For the preparation of key
phosphonate intermediates 8c-f and 14a-c we utilized Lewis acids such
as iodotrimethylsilane or tin tetrachloride. The specific cleavage of a
dioxolane ring by iodotrimethylsilane is reported.[17] However, we have
found iodotrimethylsilane cleavage not suitable for all substituted dioxo-
lanes. As depicted in Scheme 1 and Table 1, the dioxolane ring is first
cleaved with iodotrimethylsilane into the hypothetical intermediate 11.
This reaction step is followed by the reaction of 11 with diisopropyl
phosphite in the presence of sodium hydride. Finally, a silyl group of
the intermediate 12 was removed with aqueous potassium fluoride.
Unfortunately, this synthesis afforded very low preparative yields with
dimethyl and phenyl substituted phosphonates 14 b and 14c while
monosubstituted benzyloxymethyl derivative 14a was obtained in accept-
able preparative yield. Methoxybenzyl and nitrophenyl substituted
Scheme 1. Critical steps in attempts to a-branched phosphonate precursors. Reagents and con-
ditions: (a) MeONa, HP(O)(OiPr)2, Et2O,rt; (b) BrCH2CH2OTr, NaH, THF, 0 ꢀC ! rt ! 70 ꢀC; (c)
TsCl, Et3N, DMAP, CH2Cl2, rt; (d) HOCH2CH2OBn, NaH, DMF; (e) (EtO)3P, 140 ꢀC; (f) NBS, Bz2O2,
CCl4, 60 ꢀC.

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
5
Scheme 2. Synthesis of a-branched PME-synthons. Reagents and conditions: (a) HOCH2CH2OH,
p-TsOH, toluene, reflux; (b) TMSI, cyclohexene, ꢂ78 ꢀC! 0 ꢀC; (c) HP(O)(OiPr)2,NaH, cyclohexene,
0 ꢀC! rt; d) 10% aq. KF, EtOH, rt; e) SnCl4, P(OR3)3, CH2Cl2, ꢂ78 ꢀC! 0 ꢀC.
Table 1. Synthesis of a-branched PME-synthons.
R1
R2
Yield of 8 (%)
Yield of 14 (%)
ꢁ
Entry
a
b
c
H
Me
H
CH2OBn
Me
Ph
–
–
46
92
50
45a (9)b
28
d
H
–
e
f
H
H
90
0
–
0
-
g
h
ꢁ
Me
H
Ph
Me2N
0
0
–
Yields related to reaction with a) tin tetrachloride b) iodotrimethylsilane.
dioxolanes 10f and 10e were not converted into the desired phospho-
nate 14f and 14e at all.
The unfavorable cleavage can be overcome when tin tetrachloride is
used. This reagent leads to higher yields of 14 b and 14c as well as to other
aromatic phosphonates: 14d and 14e. The explanation is that tin tetrachlor-
ide is a weaker Lewis acid than iodotrimethylsilane. This statement can be
supported by the considerable differences in bond dissociation energy of
Si-O (799 kJ/mol), Si-I (243 kJ/mol) and Sn-O (528 kJ/mol), Sn-Cl (350 kJ/
mol) pairs.[23] As for the use of aromatic dioxolanes 10c–e under tin tetra-
chloride treatment, high yields were obtained with compounds 8d and 8e
bearing electron-accepting aromatic groups. On the other hand, some
observed results indicate the instability of the intermediate 11 which is
probably predisposed towards secondary cleavage of the ether group. This

6
K. POMEISL ET AL.
cleavage was successfully clarified only for dioxolane derivative 10f by char-
acterization of the final product 13 by NMR spectroscopy.
Based on these observations, we assume that modification of nucleophi-
licity and affinity of the ether group to Lewis acids could be main stabiliza-
tion or destabilization factors for intermediates 11 and 15. The role of an
a-branched halogen in the cleavage is not clear but we suppose that the
higher electronegativity of the chlorine atom in 15 may contribute to a
lower nucleophilicity of the ether group. Therefore, this intermediate could
be better stabilized. In contrast, reactions with dioxolanes bearing bulky
and further functionalized groups such as compounds 10 g or 10 h seem to
be more difficult based on their marginal reactivity towards Lewis acids. As
proven by NMR spectroscopy, these reactions afford complex mixtures of
products; the method of dioxolane cleavage thus remains limited for simply
substituted alkyl and aryl dioxolanes.
To prepare the final acyclic nucleoside phosphonates, several synthetic
protocols have been studied. The first attempt consisted in heating of 2-
amino-6-chloropurine (16b) sodium salt with tosyl derivative 17 (prepared
from 14a by reaction with TsCl in the presence of triethylamine and N,N-
dimethylaminopyridine, Scheme 3). Surprisingly, this procedure was not
successful. The favored reaction did not proceed under heating to 100 ꢀC
while increase of temperature resulted in decomposition of the starting
material. In contrast, mild Mitsunobu conditions[24] were a successful
choice for condensation of purines with all starting phosphonates (Table
2). Thus obtained ANP esters were deprotected with bromotrimethylsilane
followed by acid hydrolysis of the 6-chloro substituent[25] to give free phos-
phonic acids 19a–j bearing guanine or hypoxanthine base moiety. As
shown in Table 2 and Scheme 3, reaction of 18j with Lewis acid proceeded
with the parallel removal of a benzyl group from the side chain.
Scheme 3. Synthesis of a-branched PMEG and PMEHx derivatives. Reagents and conditions: (a)
Et3N, DMAP, TsCl, 0 ꢀC! rt; (b) NaH, DMF, 100 ꢀC; (c) NaH, DMF, 120 ꢀC; (d) DIAD, Ph3P, THF, rt,
27–69%; (e) Me3SiBr, MeCN, rt; (f) HCl, H2O, reflux.

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
7
Table 2. Synthesis of a-branched PMEG and PMEHx derivatives.
Entry
R1
R2
R3
R4
Yield of 18 (%)
Yield of 19 (%)
a
b
c
d
e
Me
Me
H
H
H
Me
Me
Ph
Ph
iPr
iPr
Me
Me
Me
H
NH2
H
NH2
H
69
27
59
43
44
16
28
54
29
70
f
H
H
H
Me
Me
Me
NH2
H
27
48
52
30
54
16
g
h
NH2
i
i
j
j
H
H
H
H
CH2OBn
CH2OH
CH2OBn
CH2OH
iPr
H
iPr
H
H
NH2
NH2
38
–
33
–
–
64
–
H
76
2-Amino-6-chloropurine derivatives can be also transformed to their xan-
thine counterparts as shown for compound 18j (Scheme 4). The first step
was deprotection of the isopropyl ester groups with bromotrimethylsilane.
After that, the chlorine atom in position 6 can be replaced with an oxo
group by heating with hydrochloric acid to give guanine derivative 19j.
Regarding low solubility and worse reactivity of guanine compounds in gen-
eral, we also tried an alternative consisting in the treatment of 2-amino-6-
chloropurine intermediate 20 with acetic acid and isoamyl nitrite. However,
bromination of the C-8 position of thus formed xanthine derivative 21
occurred when transformation of 18j to 21 was performed in situ. We
assume radical initiation mechanism of this C-8 bromination[25] when bro-
motrimethylsilane could serve as a source of bromine and isoamyl nitrite as
oxidizer (Scheme 5). Considering release of hydrogen bromide during
hydrolysis of Lewis acids such as phosphorous bromide[26] we suppose its
analogous formation during hydrolysis performed with an excess of bromo-
trimethylsilane (Scheme 5a). As shown by the formation of hypothetical
intermediates in Scheme 5b, the oxidative effect of isoamyl nitrite on hydro-
gen bromide may result in significant release of bromine. Concurrently, iso-
amyl nitrite can be reduced to nitric oxide[27] and isoamyl bromide. It is
plausible that nitric oxide is formed by decomposition of unstable nitrous
acid in acidic conditions.
Although we pursued originally non-brominated xanthine derivatives,
the obtained phosphonate 20 became a good precursor for insertion of aryl
substituents at the C-8 position. An example is Suzuki coupling using phe-
nylboronic acid for the preparation of compound 22. While C-8 coupling
reactions with boronic acids have been described previously for 2,6-diami-
nopurine phosphonate esters (PMEDAP)[28] no similar procedure is
reported for free phosphonic acids. Contrary to ANP esters, these com-
pounds are soluble in water and reaction can be performed successfully in

8
K. POMEISL ET AL.
Scheme 4. Synthesis of xanthine derivatives. Reagents and conditions: (a) Me3SiBr, MeCN, rt;
(b) HCl, H2O, reflux; (c) CH(Me)2CH2CH2ONO, AcOH, rt; (d) PhB(OH)2, H2O, Pd(PPh3)4, Na2CO3.
Scheme 5. Proposed mechanism of oxidation of bromotrimethylsilane during the treatment
with isoamyl nitrite.
aqueous solutions using an appropriate Pd-catalyst in the presence of
sodium carbonate as a nucleophilic activator (Scheme 4).
6-Chloropurine derivative 18i was selected as the starting material either
for preparation of 6-alkylamino and 6-alkoxy derivatives or for preparation
of side-chain fluorinated analogs. Use of perfluorobutanesulfonyl fluoride
for nucleophilic displacement of a side-chain hydroxy group with fluorine
was planned similarly as described in the syntheses of 3-fluoro-2-phospho-
nomethoxypropyl (FPMP) derivatives.[29–31] This weakly corrosive reagent
is stable and little moisture-sensitive at room temperature and therefore
easy to handle.[29]
For this purpose, the critical step was removal of the protecting benzyl
group. Direct catalytic hydrogenation of 18i is not beneficial due to simul-
taneous reduction of the 6-chloropurine base to purine. Also conversion of
6-chloropurine base to hypoxanthine before hydrogenation doesnꢁt solve the

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
9
problem. As observed in our previous syntheses this modification prevents
fluorination of the side chain completely.[29]
Therefore, we protected the purine base with a stable alkyloxy group. A
number of alkyl modified purine derivatives have been tested previously as
substrates of various biochemical targets, e.g. purine nucleoside phosphory-
lase inhibitors or antiviral agents.[32] In addition, 6-alkoxy purine ANP
derivatives could increase their ability to transport through cell membranes
and play a role of prodrugs;[33] release of the corresponding guanine ANPs
in the cell is supposed. Since the release of the parent (guanine) compound
could depend on the electron-withdrawing effect of an alkyl group we
introduced the trifluoroethyloxy group to the position 6. This goal was realized
by reaction of 18i with trifluoroethanol in the presence of sodium t-butoxide
(Scheme 6). The product 23 was subjected to catalytic hydrogenation to give
hydroxy derivative 24. Fluorination of 24 using perfluorobutanesulfonyl fluo-
ride was performed in the presence of DBU as a mild base.[29] Unfortunately,
elimination of a hydroxy group occurred and compound 26 was formed
instead of the desired fluoro derivative 25. In comparison to fluorination of
HPMP derivatives, a different reaction mechanism is probably involved.
Besides 6-alkoxy modified purine derivatives we also developed a synthesis
of a-branched 6-amino and 6-alkylamino purine ANPs. In cells, 6-alkylamino
derivatives may undergo enzymatic deamination to the corresponding guanine
counterparts, similarly as described for cytostatic N6-cyclopropyl 9-[(2-phos-
phonomethoxy)ethyl]purine (cPrPMEDAP) and other N6-alkyl PMEDAP.[34,35]
The enzyme capable of this conversion was identified as N6-methyl-AMP/
dAMP aminohydrolase.[36] Transformation of 6-chloro derivatives 18i and18j
to 6-amino derivatives 28a–d was performed by heating with the appropriate
amines or ammonia in an autoclave (Scheme 7, Table 3).
Scheme 6. Reagents and conditions: (a) CF3CH2OH, t-BuONa, dioxane, reflux, 44%; (b) H2, Pd,/
C, MeOH, rt, 78%; (c) C4F9SO2F, DBU, 100 ꢀC, 66%; (c) Me3SiBr, MeCN, lutidine, rt, 16%.

10
K. POMEISL ET AL.
Phosphonate esters 28a–d were subsequently hydrolyzed with bromotri-
methylsilane to free phosphonic acids 29a–d and the benzyl group depro-
tected by catalytic hydrogenation (Table 3). However, in the case of 29a
deprotection of the benzyl group did not proceed even when hydrogenation
was performed over palladium sponge.
Biological activity
All newly prepared compounds have been evaluated in biological tests tar-
geted to antiviral, cytostatic and antimalarial activity. In studies targeted to
Plasmodium falciparum enzyme PfHGXPRT, compounds 19a–j, 21, 22 and
27 revealed decreased inhibitory activity in comparison with 9-[2-(phos-
phonoethoxy)ethyl)]guanine (PEEG)[11] or a- and b-branched PEE deriva-
tives.[12] Regarding the proposed affinity of PEE derivatives to the enzyme,
low kinetic values might be probably caused by the insufficient length of
Scheme 7. Transformation of 6-chloropurine ANPs to 6-amino and 6-alkylamino derivatives.
Reagents and conditions: (a) R2NH2/alcohol or dioxane (see Table 3), autoclave, 110 ꢀC, 57–60%;
(b) Me3SiBr, MeCN, rt; (c) H2, Pd/C, MeOH, H2O, rt,15–66%.
Table 3. Synthesis of a-branched PMEA and PMEDAP derivatives.
Entry
R1
NH2
H
R2
R3
CH2OPh
H
Solvent
MeOH
Yield of 28 (%)
Yield of 30 (%)
a
H
58
66
33
15
b
Dioxane
c
d
H
NH2
Me
Me
H
H
EtOH
EtOH
57
60
66
64

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
11
aliphatic chains. Thus, ANPs from this study are probably not able to fit
the enzyme through the proposed purine and phosphate binding sites.[7] In
comparison with the active compound PMEG[7] our results indicate that
substitution of the phosphonate a-position with hydrophobic groups may
strongly disrupt the flexibility of the PMEG phosphonate linker. While the
flexibility of PMEG could partially compensate for a shorter phospho-
noalkyl chain length at the binding sites of a dynamic enzyme[7] it is possi-
ble that optimization of branched phosphonate interactions with the
enzyme is avoided due to their potential steric hindrance. In contrast, the
alkyl group in a- and b-branched PEE unambiguously supported their
inhibitory activity. However, the effect is selective towards human enzyme
HGPRT only.[6] Although these compounds also possess considerable
effects on PfHGXPRT the observed values were decreased by a-substitu-
tion. Therefore, the higher inhibitory activity probably corresponds more
likely with the extended chain length of PEE derivatives.
None of the compounds presented in this study possesses significant
cytostatic activity in vitro estimated in mouse lymphocytic leukemia L1210
cells (ATCC CCL 219), CCRF-CEM T lymphoblastoid cells (human acute
lymphoblastic leukemia, ATCC CCL 119), human promyelocytic leukemia
HL-60 cells (ATCC CCL 240) and human cervix carcinoma HeLa S3 cells
(ATCC CCL 2.2) at a concentration 10 lmol lꢂ1.
Moderate antiviral activities against DNA viruses were found in several
compounds. Compound 19i exhibits antiviral activity against Herpes
simplex virus 2 (HSV-2, G strain) at 9 lM concentration which is compara-
ble with brivudine (10 lM) or cidofovir (2 lM). However, this effect is
substantially lower compared to ganciclovir (0.03 lM). Modest activity
against varicella-zoster virus (VZV) in human embryonic lung (HEL) cells
was found for compounds 19a, 19c, 19f, 19g, 19h and 21 (ꢃ15–50 lM).
The highest activity against both TKþ VZV and TKꢂ VZV strains was
found for compounds 19 h and 19 g in the series bearing a-branched
4-nitrophenyl substituents (TKþ 14.8–16.1 lM; TKꢂ 17.5–23.1 lM).
As for the enzymatic studies, the catalytic activity of compounds 30a-d
has been studied on N6-methyl-AMP aminohydrolase. We have found that
neither alkoxy nor methylamino substituted derivatives exhibited significant
effects in comparison with similar PMEDAP and FPMPDAP methylamino
substituted derivatives.
Conclusions
In summary, we developed the methods for the preparation of various
a-branched alkyl and aryl substituted ANPs as potential biological
modifiers of efficient cytostatic PMEG and antiviral PMEA and PMEDAP.

12
K. POMEISL ET AL.
We newly described the various reaction courses for the cleavage of
1,3-dioxolane ring using described Lewis acids such as trimethylsilyl iodide
and tin tetrachloride. We have found that treatment of some alkyl and
aryl-substituted dioxolanes with trimethylsilyl iodide resulted in low yields
of favored phosphonate synthons. In contrast, the ether group is stabilized
using tin tetrachloride to afford acceptable preparative yields. For that
reason, we assume that unfavorable cleavage is connected with a higher
affinity of trimethylsilyl iodide and the nucleophility of ether oxygen
as a consequence of different a-branched substitution.
As for the biological activity, the shorter phosphonoalkyl chain and
a-substitution strongly decreased PfHGXPRT or human HGXPRT-
inhibitory activity of guanine and xanthine modified PME phosphonates
in comparison with PMEG and PEEG-derivatives. We assume the inability
of binding of our a-branched PMEG derivatives may be caused a) by insuf-
ficient length between the proposed purine and phosphate binding site and
b) by steric hindrance of their bulky substituents in the chains. It can be
concluded that a-substitution is not a favored tool for modification of both
purine PME and PEE inhibitors as well as compounds mimicking antiviral
phosphonates PMEA and PMEDAP.
Experimental
General methods
Solvents were evaporated at 40 ꢀC/1 kPa and compounds dried at 50 Pa.
Column chromatography was performed on silica gel 60 mm (Fluka).
Analytical TLC was performed on silica gel 60 F254 plates (Merck). KGaA,
Germany) in solvent systems: chloroform-methanol 97:3 (S1); 9:1 (S2); 15:1
(S3); 50:1 (S4) 25:1 (S5), ethyl acetate-acetone-ethanol-water 18:3:2:2 (S6)
or 16:3:3:2 (S7) and isopropanol-25% NH4OH-water 7:1:2 (S8). Preparative
TLC was carried out on 45 ꢄ 18 ꢄ 0.4 cm loose-layer silica gel plates (silica
gel containing UV indicator prepared by Service laboratories of the
Institute (S9). IR spectra were recorded on a FTIR Spectrometer Bruker IFS
1
55 (Equinox) in KBr, CCl4 and CHCl3. H and 13C NMR spectra were
measured on a Bruker Avance 600 spectrometer (1H at 600.1 MHz, 13C at
150.9 MHz); Bruker Avance 500 spectrometer (1H at 500.0 MHz, 13C at
125.7 MHz, 13 P at 202.3 MHz) or Bruker Avance 400 (1H at 400.1 MHz,
13C at 100.6 MHz, 31 P at 162.0 MHz) in CDCl3 (referenced to the residual
solvent signal in 1H (d ¼ 7.26 ppm) or solvent signal in 13C
1
(d ¼ 77.00 ppm)); DMSO (referenced to the residual solvent signal in H
(d ¼ 2.50 ppm) or solvent signal in 13C (d ¼ 39.7 ppm)); CD3OD (referenced
1
to the residual solvent signal in H (d ¼ 3.31 ppm) or solvent signal in 13C
(d ¼ 49.00 ppm)) or in D2O (þNaOD) with dioxane 1 as an internal

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
13
Figure 3. General numbering scheme for assignment of NMR signals.
1
standard (3.75 ppm for H and 69.30 ppm for 13C). Chemical shifts (d,
ppm) and coupling constants (J, Hz) were obtained by first-order analysis
of the spectra. Complete signal assignment was done using standard H,H-
COSY, H,C-HSQC and H,C-HMBC experiments. General numbering
scheme for assignment of NMR signals is depicted in Figure 3. Mass spec-
tra were measured by ESI technique on LCQ Fleet or LTQ Orbitrap XL
(Thermo Fisher Scientific) and on a ZAB-EQ (VG Analytical) spectrometer
using FAB (ionization with xenon, accelerating voltage 8 kV, glyc-
erol matrix).
Materials and chemicals
Standard chemicals and ion-exchange resins were purchased from Sigma-
Aldrich (Czech Republic). Starting dioxolanes 10a-g were prepared by reac-
tion of appropriate oxo compounds with glycol in presence of p-toluenesul-
fonic acid or hydrochloric acid according to ref. [22]. Hydroxy derivatives 2
and 3 were prepared by condensation of aldehyde 1 with diisopropyl phos-
phite.[18] Bromo derivative 7 was prepared by 2-step procedure from benzyl
[20,21]
bromide according to procedures in refs.
Tosylation of hydroxy
derivatives 2 and 3 to compounds 4 and 5 was carried out analogously as
described in ref.[19] Anhydrous solvents such as N,N-dimethylformamide,
tetrahydrofuran and acetonitrile were purchased from Sigma-Aldrich and
Penta (Czech Republic).
Synthesis of a-branched phosphonates 8c–g and 14b;
general procedure
Tin chloride (7.94 mmol) was added dropwise at ꢂ78 ꢀC under argon to a
solution of dioxolane 10b-g (4.90 mmol) in dichloromethane (50 mL) and

14
K. POMEISL ET AL.
the mixture was stirred at ꢂ78 ꢀC for 25 min. Triisopropyl phosphite or tri-
methyl phosphite (7.44 mmol) was added, the mixture stirred for 30 min
and then allowed to warm to rt during 3 h. The mixture was stirred at rt
for 16 h and treated with 50 mL ethyl acetate. Saturated aqueous solution of
NaHCO3 (10 mL) was added and the product was extracted into ethyl ace-
tate (3x 50 mL). The combined organic extract was dried over MgSO4,
evaporated and the residue chromatographed on silica gel (gradient of
chloroform-methanol 95:5 to 90:10 for 8c, 8e-g and 14 b or 15:1 for 8d).
Dimethyl [(2-hydroxyethoxy)(phenyl)methyl]phosphonate (8c)
Yield 587 mg (46%) of a yellowish oil; Rf (S1) 0.27; IR: (CCl4) mmax 3604,
3414, 3090, 3066, 3032, 2956, 1603, 1495, 1465, 1453, 1262, 1185, 1100,
1067, 1055, 1038, 921, 855, 698 cmꢂ1. ESIMS: m/z 283.0 (M þ Na)þ (99).
HRMS (ESI): m/z calcd for C11H17O5NaP (M þ Na)þ 283.0633; found
1
283.0706. H NMR (500.0 MHz, CDCl3): 3.20 (bs, 1 H, OH); 3.57 (m, 1 H,
CHaHbO); 3.69 (d, 3 H, JH,P ¼ 10.4, CH3OP); 3.71 (d, 3 H, JH,P ¼ 10.6,
CH3OP); 3.72–3.80 (m, 3 H, CHaHbO, CH2OH); 4.74 (d, 1 H, JH,P ¼ 15.8,
CHP); 7.34 (m, 1 H, H-p-Ph); 7.39 (m, 2 H, H-m-Ph); 7.44 (m, 2 H, H-o-
Ph). 13C NMR (125.7 MHz, CDCl3): 53.64 (d, JC,P ¼ 7.2, CH3OP); 53.94
(d, JC,P ¼ 6.8, CH3OP); 61.72 (CH2OH); 73.03 (d, JC,P ¼ 12.2, CH2O); 78.91
(d, JC,P ¼ 168.7, CHP); 127.68 (d, JC,P ¼ 5.8, CH-o-Ph); 128.58 (d, JC,P ¼ 2.5,
CH-m-Ph); 128.64 (d, JC,P ¼ 3.1, CH-p-Ph); 134.49 (d, JC,P ¼ 1.6, C-i-Ph).
Dimethyl [(4-chlorophenyl)(2-hydroxyethoxy)methyl]phosphonate (8d)
Yield 1.44 g (92%) of colorless oil; Rf (S1) 0.35; IR: (CCl4) mmax 3414, 2937,
1459, 1261, 1186, 1113, 1092, 1067, 1055, 842, 569. ESIMS: m/z 295.0.
(M þ H)þ (12). HRMS (ESI): m/z calcd for C11H17O5ClP (M þ H1)þ
1
295.0497; found 295.0498. H NMR (500.0 MHz, CDCl3): 3.55 (m, 1 H,
CHaHbO); 3.71, 3.73 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.5, CH3O); 3.72–3.79 (m, 3 H,
CH2OH, CHaHbO); 4.72 (d, 1 H, JC,P ¼ 15.9, CHP); 7.34–7.39 (m, 4 H, CH-
o,m-C6H4Cl). 13C NMR (125.7 MHz, CDCl3): 53.72 (d, JC,P ¼ 7.2, CH3O);
54.02 (d, JC,P ¼ 6.8, CH3O); 61.66 (CH2OH); 73.12 (d, JC,P ¼ 12.1, CH2O);
78.25 (d, JC,P ¼ 169.5, CHP); 128.81 (d, JC,P ¼ 2.5, CH-m-C6H4Cl); 128.97
(d, JC,P ¼ 5.8, CH-o-C6H4Cl); 133.08 (d, JC,P ¼ 1.7, C-i-C6H4Cl); 134.55
(d, JC,P ¼ 3.8, CH-p-C6H4Cl).
Dimethyl [(2-hydroxyethoxy)(4-nitrophenyl)methyl]phosphonate (8e)
Yield 1.35 g (90%) of a yellowish oil; Rf (S2) 0.51. IR: (CHCl3) mmax 3605,
3413, 3083, 2958, 1608, 1602, 1526, 1493, 1459, 1416, 1350, 1287, 1187,
1108, 1097, 1065, 1052, 1042, 1017, 865, 834, 694 cmꢂ1. ESIMS: m/z 328.0

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
15
(M þ Na)þ (100). HRMS (ESI): m/z calcd for C11H16O7NNaP (M þ Na)þ
1
328.0483; found 328.0557. H NMR (500.0 MHz, CDCl3): 3.63 (m, 1 H,
CHaHbO); 3.72–3.84 (m, 3 H, CH2OH, CHaHbO); 3.76, 3.78 (2 ꢄ d,
2 ꢄ 3 H, JH,P ¼ 10.6, CH3O); 4.90 (d, 1 H, JC,P ¼ 17.1, CHP); 7.63 (m, 2 H,
CH-o-C6H4NO2); 8.25 (m, 2 H, CH-m-C6H4NO2). 13C NMR (125.7 MHz,
CDCl3): 53.88 (d, JC,P ¼ 7.2, CH3O); 54.22 (d, JC,P ¼ 6.8, CH3O); 61.67
(CH2OH); 73.80 (d, JC,P ¼ 11.2, CH2O); 78.28 (d, JC,P ¼ 167.4, CHP); 123.67
(d, JC,P ¼ 2.6, CH-m-C6H4NO2); 128.25 (d, JC,P ¼ 5.3, CH-o-C6H4NO2);
142.26 (d, JC,P ¼ 1.7, C-i-C6H4NO2); 147.90 (d, JC,P ¼ 3.7, CH-p-C6H4NO2).
Diisopropyl [2-(2-hydroxyethoxy)propan-2-yl]phosphonate (14b)
Yield 596 mg (45%) of a colorless oil; Rf (S1) 0.27; IR: (CCl4) mmax 3606,
3393, 2980, 1467, 1385, 1375, 1385, 1361, 1245, 1223, 1196, 1177, 1142,
1087, 1004, 989, 885 cmꢂ1. ESIMS: m/z 291.1 (M þ Na)þ (100). HRMS
1
(ESI) m/z calcd for C11H25O5NaP (M þ Na)þ 291.1259; found 291.1332. H
NMR (500.0 MHz, CDCl3): 1.33 (d, 12 H, Jvic ¼ 6.2, (CH3)2CH); 1.42 (d,
6 H, JH,P ¼ 15.2, (CH3)2C); 3.69 (m, 2 H, CH2OH); 3.75 (m, 2 H, CH2O);
4.75 (dsep, 2 H, JH,P ¼ 7.4, Jvic ¼ 6.2, CH(CH3)2). 13C NMR (125.7 MHz,
CDCl3): 22.46 (d, JC,P ¼ 3.9, (CH3)2C); 23.89 (d, JC,P ¼ 5.1, (CH3)2CH);
24.25 (d, JC,P ¼ 3.1, (CH3)2CH); 62.30 (CH2OH); 65.17 (d, JC,P ¼ 3.8,
CH2O); 70.89 (d, JC,P ¼ 7.6, CH(CH3)2); 74.24 (d, JC,P ¼ 167.5, CP).
Synthesis of a-branched phosphonates 14a-c and 13;
general procedure
Trimethylsilyl iodide (14.6 mmol) in hexane (4 mL) was added dropwise at
ꢂ78 ꢀC under argon to a solution of dioxolane 10a-c or 10f (14.6 mmol) in
cyclohexene (8 mL) and the mixture was stirred at ꢂ78 ꢀC for 1 h. The mix-
ture was then allowed to warm to 0 ꢀC. In parallel, diisopropylphosphite
(17.0 mmol) was dissolved in cyclohexane (16 mL) and sodium hydride
(17.0 mmol) was added at 0 ꢀC. This mixture was stirred at 0 ꢀC for 1 h.
The mixture containing the intermediate 11 was then added dropwise
(using a cold syringe) and the resulting mixture was stirred at rt for 17 h.
The mixture was evaporated, the residue partitioned between chloroform
(20 mL) and water (5 mL). An organic layer was dried over MgSO4 and
evaporated. The residue was dissolved in ethanol (8 mL) and aqueous 10%
potassium fluoride and the solution was evaporated in vacuo. The residue
was dissolved in chloroform (25 mL) and washed with water (4 mL). An
organic layer was dried over MgSO4, evaporated and the residue was chro-
matographed on silica gel (chloroform-methanol 97:3).

16
K. POMEISL ET AL.
Diisopropyl [2-(benzyloxy)-1-(2-hydroxyethoxy)ethyl]phosphonate (14a)
Yield 2.62 g (50%) of a colorless oil; Rf (S1) 0.32. IR: (CCl4) mmax 3411, 1497,
1453, 1386, 1375, 1247, 1105, 1070, 1038, 1029, 1005, 989, 733, 697, 574 cmꢂ1.
FABMS: m/z 361.0 (M þ H)þ (24). HRMS (FAB): m/z calcd for C17H30O6NP
(M þ H)þ 361.1702; found 361.1775. Anal. Calcd for C17H29O6P (%): C,
1
56.66; H, 8.11; P, 8.59. Found: C, 56.23; H, 8.03; P, 8.22. H NMR (500 MHz,
CDCl3): 1.30, 1.31, 1.32 and 1.33 (4 ꢄ d, 4 ꢄ 3H, Jvic ¼ 6.2, (CH3)2CH); 3.69
(ddd, 1 H, Jgem ¼ 10.7, Jvic ¼ 9.3, JH,P ¼ 3.8, CHaHbCHP); 3.71 (m, 2 H,
CH2OH); 3.78 (ddd, 1 H, Jgem ¼ 10.7, JH,P ¼ 3.3, Jvic ¼ 2.4, CHaHbCHP); 3.84,
3.91 (2 ꢄ m, 2 ꢄ 1H, CH2O); 3.93 (ddd, 1 H, JH,P ¼ 10.7, Jvic ¼ 9.3, 2.4, CHP);
4.56, 4.61 (2 ꢄ d, 2 ꢄ 1H, Jgem ¼ 12.1, CH2Ph); 4.73, 4.79 (2 ꢄ dsep, 2 ꢄ 1H,
JH,P ¼ 7.4, Jvic ¼ 6.2, CH(CH3)2); 7.27–7.38 (m, 5 H, H-o,m,p-Ph). 13C NMR
(125.7 MHz, CDCl3): 23.88, 24.01, 24.02, 24.22 (4 ꢄ d, JC,P ¼ 4.0, (CH3)2CH);
61.78 (CH2OH); 70.27 (d, JC,P ¼ 12.6, CH2CHP); 71.35 (d, JC,P ¼ 7.3,
CH(CH3)2); 71.92 (d, JC,P ¼ 7.0, CH(CH3)2); 73.51 (d, JC,P ¼ 1.1, CH2Ph);
75.34 (d, JC,P ¼ 4.5, CH2O); 76.86 (d, JC,P ¼ 165.6, CHP); 127.68 (CH-o-Ph);
127.85 (CH-p-Ph); 128.46 (CH-m-Ph); 137.57 (C-i-Ph).
Diisopropyl [2-(2-hydroxyethoxy)propan-2-yl]phosphonate (14b)
1
Yield 352 mg (9%) of a colorless oil. H, 13C NMR and HRMS data were
identical with those obtained for 14 b prepared by the method with tin
chloride (see above).
Diisopropyl [(2-hydroxyethoxy)(phenyl)methyl]phosphonate (14c)
Yield 1.19 g (28%) of a yellowish oil. Rf (S1) 0.35. ESIMS: m/z 339.1
(M þ Na)þ (100). HRMS (ESI): m/z calcd for C15H24O5NaP (M þ Na)þ
239.1259; found 239.1101. 1H NMR (400.1 MHz, CDCl3): 1.45 (d, 6 H,
Jgem ¼ 10.5, (CH3)2CH); 2.22 (bs, 1 H, OH); 3.56 (m, 1 H, CHaHbO); 3.71–3.79
(m, 3 H, CHaHbO and CH2OH); 4.84 (m, 2 H, JHP ¼ 10.5, (CH3)2CH), 4.73 (d,
1H, JHP ¼ 15.8, CHP); 7.33 (m, 1 H, H-p-Ph), 7.38 (m, 2 H, H-m-Ph); 7.43 (m,
2H, H-o-Ph). 13C NMR (125.7 MHz, CDCl3): 23.90 (d, JC,P ¼ 3.9, (CH3)2CH),
23.96 (d, JC,P ¼ 4.0, (CH3)2CH), 53.63, (d, JC,P ¼ 7.2, CH3)2CH); 53.92
(JC,P ¼ 6.9, CH3)2CH); 61.71 (CH2OH); 73.02 (d, JC,P ¼ 12.2, CH2O); 78.90 (d,
JC,P ¼ 168.7, CHP); 127.67 (d, JC,P ¼ 5.8, CH-o-Ph); 128.57 (d, JC,P ¼ 2.5, CH-
m-Ph); 128.63 (d, JC,P ¼ 3.2, CH-p-Ph); 134.48 (d, JC,P ¼ 1.6, C-i-Ph).
Diisopropyl 1-hydroxy-1-[(4-methoxyphenyl)methyl]phosphonate (13)
Yield 2.86 g (68%) of a yellowish oil. FABMS: m/z 303.1 (M þ H)þ (100).
HRMS (FAB): m/z calcd for C14H23O5P (M þ H)þ 303.1283; found

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
17
303.1257. 1H NMR (400.1 MHz, CDCl3): 1.38 (dd, 6 H, Jgem ¼ 8.00,
CH3CHO); 3.80 (s, 3 H, CH3O-Ph); 4.62 (dsept, 2 H, Jgem ¼ 6.92, JHP ¼ 0.95,
CH3CHO); 4.89 (dd, 1 H, Jgem ¼ 9.62, JHP ¼ 3.87, P-CHOH); 4.62 (s, 2 H,
CH2Ph); 6.90 and 7.43 (2 x d, 4 H, Jgem ¼ 9.20, CH ¼ C).
2-[2-(Benzyloxy)-1-(diisopropoxyphosphoryl)ethoxy]ethyl 4-
methylbenzenesulfonate (17)
Tosyl chloride (484 mg, 2.54 mmol) and a catalytic amount of DMAP
(13 mg) were added to 0 ꢀC cold solution of 14a (702 mg, 2.11 mmol) and
triethylamine (704 mL, 5.07 mmol) in dichloromethane (10 mL). The mix-
ture was allowed to warm to rt and stirred for 10 days, then diluted with
chloroform (20 mL) and washed with water (2 mL). The organic extract
was dried over MgSO4 and evaporated and the residue purified by column
chromatography on silica gel (chloroform-methanol 50:1) to give 332 mg
1
(32%) of yellowish oil; Rf (S4) 0.50; H NMR (400.1 MHz, CDCl3): 1.21-
1.40 (m, 6 H, CH3CHO); 2.41 (s, 3 H, CH3-Ph); 3.58–4.05 (m, 6 H, CH2-
O); 4.16 (sept, 2 H, Jgem ¼5.49, CH3CHO); 4.62 (s, 2 H, CH2Ph); 7.21-7.42
(m, 9 H, CH ¼ C). ESIMS: m/z 537.2 (M þ Na)þ (100). HRMS (ESI): m/z
calcd for C24H35O8NaPS (M þ Na)þ 538.1682; found 538.1599.
Synthesis of purine phosphonates 18a–j. General procedure
A mixture of 16a or 16 b (2.37 mmol), phosphonate synthon 8c-f or 14a-b
(1.83 mmol) and triphenylphosphine (2.38 mmol) was stirred in dry tetra-
hydrofuran (20 mL), after that cooled to 0 ꢀC and DIAD (2.74 mmol) was
added dropwise. The stirring was continued for 23 h at rt, the mixture was
evaporated and the residue purified by preparative TLC chromatography
(S9) in systems described below.
Diisopropyl {2-[2-(6-chloro-9H-purin-9-yl)ethoxy]propan-2-
yl}phosphonate (18a)
Yield 477 mg (69%) of a colorless oil; chromatography in system chloro-
form-methanol (25:1) and (15:1); Rf (S5) 0.48. IR: (CCl4) mmax 2981, 1592,
1563, 1497, 1467, 1404, 1385, 1375, 1362, 1335, 1247, 1214, 1106, 1003,
987, 884, 567, 648 cmꢂ1. ESIMS: m/z 405.0 (M þ H)þ (30). HRMS (ESI):
m/z calcd for C16H27O4N4ClP (M þ H)þ 405.1380; found 405.1454. Anal.
Calcd for C16H26O4NClP (%): C, 47.47; H, 6.47; Cl, 8.76; P, 7.65. Found: C,
1
47.26; H, 6.77; P, 8.29; Cl 7.65. H NMR (500.0 MHz, CDCl3): 1.26, 1.28
(2 ꢄ d, 2 ꢄ 6 H, Jvic ¼ 6.2, (CH3)2CH); 1.32 (d, 6 H, JH,P ¼ 14.9, (CH3)2C);
4.02 (m, 2 H, H-20); 4.46 (m, 2 H, H-10); 4.65 (dsep, 2 H, JH,P ¼ 7.4,
Jvic ¼ 6.2, CH(CH3)2); 8.32 (s, 1 H, H-8); 8.74 (s, 1 H, H-2). 13C NMR

18
K. POMEISL ET AL.
(125.7 MHz, CDCl3): 22.27 (d, JC,P ¼ 4.2, (CH3)2C); 23.85 (d, JC,P ¼ 5.1,
(CH3)2CH); 24.15 (d, JC,P ¼ 3.1, (CH3)2CH); 44.61 (CH2-10); 61.85 (d,
JC,P ¼ 3.1, CH2-20); 70.85 (d, JC,P ¼ 7.6, CH(CH3)2); 74.49 (d, JC,P ¼ 166.6,
C-40); 131.37(C-5); 146.55 (CH-8); 150.78 (C-6); 151.70 (C-4); 151.75
(CH-2).
Diisopropyl {2-[2-(2-amino-6-chloro-9H-purin-9-yl)ethoxy]propan-2-
yl}phosphonate (18b)
Yield 207 mg (27%) of a white foam. Chromatography in S6; Rf (S6) 0.48.
IR: (CCl4) mmax 3538, 3506, 3428, 3316, 2981, 1609, 1596, 1565, 1514, 1465,
1408, 1385, 1375, 1363, 1309, 1246, 1214, 1178, 1146, 1106, 1004, 988, 884,
644 cmꢂ1. ESIMS: m/z 420.0 (M þ H)þ (15). HRMS (ESI): m/z calcd for
1
C16H28O4N5ClP (M þ H)þ 420.1489; found 420.1562. H NMR (500.0 MHz,
CDCl3): 1.27, 12.9 (2 ꢄ d, 2 ꢄ 6 H, Jvic ¼ 6.2, (CH3)2CH); 1.32 (d, 6 H,
JH,P ¼ 14.9, (CH3)2C); 3.96 (m, 2 H, H-20); 4.24 (m, 2 H, H-10); 4.66 (dsep,
2 H, JH,P ¼ 7.4, Jvic ¼ 6.2, CH(CH3)2); 5.07 (bs, 2 H, NH2); 7.93 (s, 1 H, H-
8). 13C NMR (125.7 MHz, CDCl3): 22.29 (d, JC,P ¼ 4.1, (CH3)2C); 23.87 (d,
JC,P ¼ 5.1, (CH3)2CH); 24.19 (d, JC,P ¼ 3.0, (CH3)2CH); 44.06 (CH2-10);
61.96 (d, JC,P ¼ 3.3, CH2-20); 70.83 (d, JC,P ¼ 7.6, CH(CH3)2); 74.43 (d,
JC,P ¼ 166.5, C-40); 125.10 (C-5); 143.72 (CH-8); 151.06 (C-6); 153.66 (C-4);
158.84 (C-2).
Dimethyl {[2-(6-chloro-9H-purin-9-
yl)ethoxy](phenyl)methyl}phosphonate (18c)
Yield 428 mg (59%) of a slightly yellowish oil; chromatography in S6; Rf
(S6) 0.48. IR: (CCl4) mmax 3066, 3032, 2955, 1592, 1563, 1497, 1454, 1427,
1405, 1336, 1305, 1262, 1227, 1216, 1185, 1105, 1058, 1039, 921, 835, 698,
569, 537, 476 cmꢂ1. ESIMS: m/z 397.1 (M þ H)þ (63). HRMS (ESI): m/z
1
calcd for C16H19O4N4ClP (M þ H)þ 397.0754; found 397.0827. H NMR
(500.0 MHz, CDCl3): 3.57, 3.65 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.6, CH3OP); 3.85
(ddd, 1 H, Jgem ¼ 10.5, J2 b,1 b ¼ 6.4, J2 b,1 a ¼ 3.2, H-20b); 3.91 (ddd, 1 H,
0
0
0
0
Jgem ¼ 10.5, J2 a,1 a ¼ 6.8, J2 a,1 b ¼ 3.1, H-20a); 4.46 (ddd, 1 H, Jgem ¼ 14.8,
0
0
0
0
J1 b,2 b ¼ 6.4, J1 b,2 a ¼ 3.1, H-10b); 4.55 (ddd, 1 H, Jgem ¼ 14.8, J1 a,2 a ¼ 6.8,
0
0
0
0
0
0
J1 a,2 b ¼ 3.2, H-10a); 4.58 (d, 1 H, JH,P ¼ 15.8, H-40); 7.22–7.31 (m, 5 H, H-
o,m,p-Ph); 8.37 (s, 1 H, H-8); 8.67 (s, 1 H, H-2). 13C NMR (125.7 MHz,
CDCl3): 44.11 (CH2-10); 53.66 (d, JC,P ¼ 7.2, CH3OP); 53.68 (d, JC,P ¼ 6.9,
CH3OP); 68.19 (d, JC,P ¼ 13.7, CH2-20); 78.88 (d, JC,P ¼ 170.2, CH-40);
127.77 (d, JC,P ¼ 6.0, CH-o-Ph); 128.64 (d, JC,P ¼ 2.2, CH-m-Ph); 129.03 (d,
JC,P ¼ 2.9, CH-p-Ph); 131.49 (C-5); 133.28 (d, JC,P ¼ 1.9, C-i-Ph); 146.34
(CH-8); 150.92 (C-6); 151.62 (C-4); 151.75 (CH-2).
0
0

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
19
Dimethyl {[2-(2-amino-6-chloro-9H-purin-9-
yl)ethoxy](phenyl)methyl}phosphonate (18d)
Yield 332 mg (43%) of a white foam; chromatography in S6; Rf (S6) 0.62.
IR: (CHCl3) mmax 3533, 3424, 3089, 3065, 2957, 1633, 1610, 1602, 1567,
1515, 1495, 1456, 1408, 1310, 1283, 1258, 1189, 1105, 1056, 1041, 1030,
836, 699, 644, 577, 541 cmꢂ1. ESIMS: m/z 434.2 (M þ Na)þ (26). HRMS
(ESI): m/z calcd for C16H19O4N5ClNaP (M þ Na)þ 434.0763; found
434.0755. 1H NMR (500.0 MHz, CDCl3): 3.57, 3.64 (2 ꢄ d, 2 ꢄ 3 H,
0
0
JH,P ¼ 10.6, CH3OP); 3.77, 3.83 (2 ꢄ ddd, 2 ꢄ 1 H, Jgem ¼ 10.5, J2 ,1 ¼ 6.5,
0
3.2, H-20); 4.23, 4.32 (2 ꢄ ddd, 2 ꢄ 1 H, Jgem ¼ 14.7, J1 ,2 ¼ 6.5, 3.2, H-1 b);
4.59 (d, 1 H, JH,P ¼ 15.4, H-40); 7.26 (m, 2 H, H-o-Ph); 7.28–7.32 (m, 3 H,
H-m,p-Ph); 7.97 (s, 1 H, H-8). 13C NMR (125.7 MHz, CDCl3): 43.47 (CH2-
10); 53.66 (d, JC,P ¼ 6.9, CH3OP); 53.69 (d, JC,P ¼ 7.2, CH3OP); 68.28 (d,
JC,P ¼ 13.9, CH2-20); 78.85 (d, JC,P ¼ 169.9, CH-40); 125.10 (C-5); 127.79 (d,
JC,P ¼ 5.9, CH-o-Ph); 128.62 (d, JC,P ¼ 2.2, CH-m-Ph); 128.93 (d, JC,P ¼ 2.9,
CH-p-Ph); 133.41 (d, JC,P ¼ 2.0, C-i-Ph); 143.38 (CH-8); 151.13 (C-6);
153.59 (C-4); 158.85 (C-2).
0
0
Dimethyl {[2-(6-chloro-9H-purin-9-yl)ethoxy](4-
chlorophenyl)methyl}phosphonate (18e)
Yield 347 mg (44%) of a yellowish oil; chromatography in S6; Rf (S6) 0.41. IR:
(CCl4) mmax 2955, 1592, 1564, 1499, 1491, 1466, 1435, 1405, 1336, 1294, 1277,
1262, 1216, 1197, 1185, 1148, 1110, 1093, 1057, 1038, 1017, 934, 713, 648,
638, 570 cmꢂ1. ESIMS: m/z 431.0 (M þ H)þ (26). HRMS (ESI): m/z calcd for
1
C16H18O4N4Cl2P (M þ H)þ 431.0364; found 431.0437. H NMR (500.0 MHz,
CDCl3): 3.59, 3.67 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.6, CH3OP); 3.84, 3.90 (2 ꢄ ddd,
0
0
0
2 ꢄ 1 H, Jgem ¼ 10.5, J2 ,1 ¼ 6.6, 3.1, H-2 ); 4.47 (ddd, 1 H, Jgem ¼ 14.8,
J1 b,2 ¼ 6.6, 3.1, H-1 b); 4.56 (d, 1 H, JH,P ¼ 15.5, H-40); 4.57 (ddd, 1 H,
0
0
0
0
0
0
Jgem ¼ 14.8, J1 a,2 ¼ 6.6, 3.1, H-1 a); 7.20 (m, 2 H, H-o-C6H4Cl); 7.28 (m, 2 H,
H-m-C6H4Cl); 8.38 (s, 1 H, H-8); 8.70 (s, 1 H, H-2). 13C NMR (125.7 MHz,
CDCl3): 44.00 (CH2-10); 53.74 (d, JC,P ¼ 7.2, CH3OP); 53.76 (d, JC,P ¼ 6.9,
CH3OP); 68.34 (d, JC,P ¼ 13.5, CH2-20); 78.12 (d, JC,P ¼ 170.9, CH-40); 128.92
(d, JC,P ¼ 2.3, CH-m-C6H4Cl); 129.01 (d, JC,P ¼ 5.9, CH-o-C6H4Cl); 131.39 (C-
5); 131.80 (d, JC,P ¼ 2.0, C-i-C6H4Cl); 134.99 (d, JC,P ¼ 3.6, C-p-C6H4Cl);
146.28 (CH-8); 150.94 (C-6); 151.50 (C-4); 151.79 (CH-2).
Dimethyl {[2-(2-amino-6-chloro-9H-purin-9-yl)ethoxy](4-
chlorophenyl)methyl}phosphonate (18f)
Yield 220 mg (27%) of a white foam; chromatography in S6; Rf (S6) 0.29.
IR: (CHCl3) mmax 3533, 3423, 2958, 1632, 1610, 1602, 1567, 1515, 1491,

20
K. POMEISL ET AL.
1408, 1310, 1457, 1295, 1281, 1256, 1189, 1149, 1111, 1092, 1057, 1042,
1016, 711, 641, 568 cmꢂ1. ESIMS: m/z 446.0 (M þ H)þ (16). HRMS (ESI):
1
m/z calcd for C16H19O4N5Cl2P (M þ H)þ 446.0473; found 446.0546. H
NMR (500.0 MHz, CDCl3): 3.54, 3.57 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.6, CH3OP);
3.72 (ddd, 1 H, Jgem ¼ 10.5, J2 b,1 b ¼ 6.3, J2 b,1 a ¼ 3.4, H-20b); 3.76 (ddd, 1 H,
0
0
0
0
Jgem ¼ 10.5, J2 a,1 a ¼ 6.5, J2 a,1 b ¼ 3.3, H-20a); 4.17 (ddd, 1 H, Jgem ¼ 14.7,
0
0
0
0
J1 b,2 b ¼ 6.3, J1 b,2 a ¼ 3.3, H-10b); 4.26 (ddd, 1 H, Jgem ¼ 14.7, J1 a,2 a ¼ 6.5,
0
0
0
0
0
0
J1 a,2 b ¼ 3.4, H-10a); 4.50 (d, 1 H, JH,P ¼ 15.4, H-40); 7.14 (m, 2 H, H-o-
C6H4Cl); 7.22 (m, 2 H, H-m-C6H4Cl); 7.88 (s, 1 H, H-8). 13C NMR
(125.7 MHz, CDCl3): 43.45 (CH2-10); 53.69, 53.74 (2 ꢄ d, JC,P ¼ 6.6,
CH3OP); 68.43 (d, JC,P ¼ 13.3, CH2-20); 78.24 (d, JC,P ¼ 170.4, CH-40);
125.11 (C-5); 128.87 (d, JC,P ¼ 2.1, CH-m-C6H4Cl); 129.07 (d, JC,P ¼ 5.9,
CH-o-C6H4Cl); 132.11 (d, JC,P ¼ 1.9, C-i-C6H4Cl); 134.89 (d, JC,P ¼ 3.5, C-
p-C6H4Cl); 143.24 (CH-8); 151.24 (C-6); 153.56 (C-4); 158.88 (C-2).
0
0
Dimethyl {[2-(6-chloro-9H-purin-9-yl)ethoxy](4-
nitrophenyl)methyl}phosphonate (18g)
Yield 388 mg (48%) of a yellowish foam; chromatography on silica gel (gra-
dient ethyl acetate-chloroform-methanol 21:20:1 to 21:20:3); Rf (S6) 0.64.
IR: (CHCl3) mmax 3082, 3064, 2958, 1607, 1594, 1566, 1527, 1499, 1466,
1428, 1405, 1350, 1338, 1290, 1259, 1197, 1185, 1148, 1116, 1106, 1055,
1041, 1015, 936, 865, 835, 694, 648, 569, 522 cmꢂ1. ESIMS: m/z 464.1
(M þ Na)þ (20). HRMS (ESI): m/z calcd for C16H17O6N5ClNaP (M þ Na)þ
464.0504; found 464.0497. Anal. calcd for C16H17O6N5ClP (%): C, 43.50; H,
1
3.88; P, 7.01; Cl, 8.03. Found: C, 43.25; H, 4.18; P, 6.80; Cl 7.96. H NMR
(500.0 MHz, CDCl3): 3.63, 3.66 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.7, CH3OP); 3.96
0
0
0
0
0
(t, 2 H, J2 ,1 ¼ 4.9, H-2 ); 4.52, 4.61 (2 ꢄ dt, 2 ꢄ 1 H, Jgem ¼ 14.9, J1 ,2 ¼ 4.9,
H-10); 4.70 (d, 1 H, JH,P ¼ 16.1, H-40); 7.47 (m, 2 H, H-o-C6H4NO2); 8.18
(m, 2 H, H-m-C6H4NO2); 8.37 (s, 1 H, H-8); 8.71 (s, 1 H, H-2). 13C NMR
(125.7 MHz, CDCl3): 44.00 (CH2-10); 53.82 (d, JC,P ¼ 5.3, CH3OP); 53.88 (d,
JC,P ¼ 5.5, CH3OP); 68.25 (d, JC,P ¼ 11.8, CH2-20); 78.29 (d, JC,P ¼ 168.0,
CH-40); 123.77 (d, JC,P ¼ 2.3, CH-m-C6H4NO2); 128.32 (d, JC,P ¼ 5.4, CH-o-
C6H4NO2); 131.48 (C-5); 141.02 (d, JC,P ¼ 1.9, C-i-C6H4NO2); 146.08 (CH-
8); 148.11 (d, JC,P ¼ 3.4, C-p-C6H4NO2); 151.14 (C-6); 151.61 (C-4); 151.92
(CH-2).
Dimethyl {[2-(2-amino-6-chloro-9H-purin-9-yl)ethoxy](4-
nitrophenyl)methyl}phosphonate (18h)
Yield 434 mg (52%) of a yellowish foam; chromatography in S6; Rf (S6)
0.49. IR: (KBr) mmax 3325, 2955, 1630, 1614, 1562, 1521, 1408, 1349, 1309,

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
21
1291, 1255, 1188, 1105, 1050, 1036, 866, 832, 786, 736, 694, 640, 561 cmꢂ1.
ESIMS: m/z 457.1 (M þ H)þ (23). HRMS (ESI): m/z calcd for
1
C16H19O6N6ClP (M þ H)þ 457.0714; found 457.0787. H NMR (600.1 MHz,
CDCl3): 3.65, 3.67 (2 ꢄ d, 2 ꢄ 3 H, JH,P ¼ 10.6, CH3OP); 3.88, 3.90 (2 ꢄ ddd,
0
0
0
2 ꢄ 1 H, Jgem ¼10.2, J2 ,1 ¼ 6.0, 3.7, H-2 ); 4.30, 4.37 (2 ꢄ ddd, 2 ꢄ 1 H,
Jgem ¼ 14.9, J1 ,2 ¼ 6.0, 3.7, H-1 ); 4.70 (d, 1 H, JH,P ¼ 16.2, H-40); 5.05 (bs,
0
0
0
2 H, NH2); 7.46 (m, 2 H, H-o-C6H4NO2); 7.95 (s, 1 H, H-8); 8.18 (m, 2 H,
13
H-m-C6H4NO2). C NMR (150.9 MHz, CDCl3): 43.47 (CH2-10); 53.83 (d,
JC,P ¼ 6.9, CH3OP); 53.92 (d, JC,P ¼ 7.2, CH3OP); 69.21 (d, JC,P ¼ 12.0, CH2-
20); 78.31 (d, JC,P ¼ 167.9, CH-40); 123.73 (d, JC,P ¼ 2.4, CH-m-C6H4NO2);
125.17 (C-5); 128.37 (d, JC,P ¼ 5.4, CH-o-C6H4NO2); 141.22 (d, JC,P ¼ 2.0,
C-i-C6H4NO2); 143.10 (CH-8); 148.06 (d, JC,P ¼ 3.4, C-p-C6H4NO2); 151.43
(C-6); 153.56 (C-4); 158.89 (C-2).
Diisopropyl {2-(benzyloxy)-1-[2-(6-chloro-9H-purin-9-
yl)ethoxy]ethyl}phosphonate (18i)
Yield 345 mg (38%) of a colorless oil; chromatography in S6; Rf (S6) 0.67.
ESIMS: m/z 497.1 (M þ H)þ (30). HRMS (ESI): m/z calcd for
1
C22H30O5N4ClP (M þ H)þ 497.1698; found 497.1720. H NMR (500.0 MHz,
CDCl3): 1.22, 1.26, 1.27, 1.29 (4 ꢄ d, 4 ꢄ 3 H, Jvic ¼ 6.2, (CH3)2CH); 3.63
(ddd, 1 H, Jgem ¼ 10.8, Jvic ¼ 8.9, JH,P ¼ 3.1, CHaHbO); 3.78 (ddd, 1 H,
Jgem ¼ 10.8, JH,P ¼ 4.4, Jvic ¼ 2.1, CHaHbO); 3.83 (ddd, 1 H, JH,P ¼ 10.8,
0
Jvic ¼ 8.9, 2.1, H-40); 4.09 (dddd, 1 H, Jgem ¼ 10.9, J2 b,1 ¼ 6.3, 3.4, JH,P ¼ 0.6,
0 0
H-20b); 4.21 (ddd, 1 H, Jgem ¼ 10.9, J2 a,1 ¼ 6.3, 3.4, H-2 a); 4.44, 4.46 (2 ꢄ d,
0
0
2 ꢄ 1 H, Jgem ¼ 11.9, CH2Ph); 4.46, 4.51 (2 ꢄ ddd, 2 ꢄ 1 H, Jgem ¼ 14.7,
0
0
0
J1 ,2 ¼ 6.3, 3.4, H-1 ); 4.67, 4.68 (2 ꢄ dsep, 2 ꢄ 1 H, JH,P ¼ 7.5, Jvic ¼ 6.2,
CH(CH3)2); 7.22 (m, 2 H, H-o-Ph); 7.28 (m, 1 H, H-p-Ph); 7.31 (m, 2 H, H-
m-Ph); 8.71 (s, 1 H, H-8); 8.87 (s, 1 H, H-2). 13C NMR (125.7 MHz,
CDCl3): 23.87, 23.96 (2 ꢄ d, JC,P ¼ 4.8, (CH3)2CH); 24.02, 24.06 (2 ꢄ d,
JC,P ¼ 3.7, (CH3)2CH); 44.15 (CH2-10); 70.05 (d, JC,P ¼ 11.1, CH2O); 70.55
(d, JC,P ¼ 5.6, CH2-20); 71.41, 71.65 (2 ꢄ d, JC,P ¼ 7.1, CH(CH3)2); 73.42
(CH2Ph); 77.26 (d, JC,P ¼ 165.4, CH-40); 127.53 (CH-o-Ph); 127.84 (CH-p-
Ph); 128.42 (CH-m-Ph); 131.41 (C-5); 137.40 (C-i-Ph); 146.64 (CH-8);
150.72 (C-6); 151.69 (CH-2); 151.74 (C-4).
Diisopropyl {1-[2-(2-amino-6-chloro-9H-purin-9-yl)ethoxy]-2-
(benzyloxy)ethyl}phosphonate (18j)
Yield 309 mg (33%) of a yellowish foam; chromatography in S6; Rf (S6)
0.48. IR: (CHCl3) mmax 3533, 3423, 1641, 1610, 1604, 1567, 1548, 1515,
1497, 1455, 1408, 1388, 1376, 1309, 1245, 1180, 1077, 1103, 1007, 994, 698,

22
K. POMEISL ET AL.
644, 572 cmꢂ1. FABMS: m/z 512.1 (M þ H)þ (29). HRMS (FAB): m/z calcd
for C22H32O5N5ClP (M þ 1)þ 512.1830; found 512.1832. 1H NMR
(600.1 MHz, DMSO-d6): 1.10, 1.12, 1.15, 1.16 (4 ꢄ d, 4 ꢄ 3 H, Jvic ¼ 6.2,
(CH3)2CH); 3.50 (ddd, 1 H, Jgem ¼ 11.2, Jvic ¼ 8.3, JH,P ¼ 3.8, CHaHbO); 3.63
(ddd, 1 H, Jgem ¼ 11.2, JH,P ¼ 6.7, Jvic ¼ 2.3, CHaHbO); 3.94 (ddd, 1 H,
JH,P ¼ 10.9, Jvic ¼ 8.3, 2.3, H-40); 3.98 (ddd, 1 H, Jgem ¼ 10.3, J2 b,1 ¼ 5.9, 4.0,
0
0
0
H-20b); 4.04 (ddd, 1 H, Jgem ¼ 10.3, J2 a,1 ¼ 6.4, 4.0, H-2 a); 4.21 (ddd, 1 H,
0
0
0
0
0
0
0
Jgem ¼ 14.7, J1 b,2 ¼ 5.9, 4.0, H-1 b); 4.24 (ddd, 1 H, Jgem ¼ 14.7, J1 a,2 ¼ 6.4,
4.0, H-10a); 4.40, 4.44 (2 ꢄ d, 2 ꢄ 1 H, Jgem ¼ 12.1, CH2Ph); 4.45–4.53 (m,
2 H, CH(CH3)2); 6.90 (bs, 2 H, NH2); 7.23 (m, 2 H, H-o-Ph); 7.27 (m, 1 H,
H-p-Ph); 7.32 (m, 2 H, H-m-Ph); 8.07 (s, 1 H, H-8). 13C NMR (125.7 MHz,
DMSO-d6): 23.70 (d, JC,P ¼ 4.7, (CH3)2CH); 23.82 (d, JC,P ¼ 4.6,
(CH3)2CH); 23.95 (d, JC,P ¼ 3.5, (CH3)2CH); 24.00 (d, JC,P ¼ 3.2,
(CH3)2CH); 43.40 (CH2-10); 69.56 (d, JC,P ¼ 10.3, CH2O); 69.81 (d,
JC,P ¼ 5.8, CH2-20); 70.72 (2 ꢄ d, JC,P ¼ 6.7, CH(CH3)2); 72.32 (CH2Ph);
75.94 (d, JC,P ¼ 162.0, CH-40); 123.47 (C-5); 127.70 (CH-p-Ph); 127.71 (CH-
o-Ph); 128.44 (CH-m-Ph); 138.14 (C-i-Ph); 143.75 (CH-8); 149.42 (C-6);
154.30 (C-2); 159.94 (C-2).
Deprotection of purine phosphonates 18a–j. General procedure
Bromotrimethylsilane (1.46 mL; 10.77 mmol) was added dropwise to 0 ꢀC
cold solution of 18a-j (1.06 mmol) in acetonitrile (20 mL). The mixture was
stirred overnight at room temperature, evaporated, and the residue codis-
tilled with water (2x2 mL). The residue was heated with 1 M HCl (5 mL) at
120 ꢀC for 5 h until conversion was complete. The mixture was concen-
trated and subsequently deionized on activated charcoal. Relevant fractions
were combined and evaporated in vacuo. The residue was chromatographed
on a Dowex 1 (AcO- form) in a gradient 0-1 M acetic acid, followed by
1 M formic acid. UV absorbing fractions containing 19a-j were evaporated
to dryness and the residue lyophilized from water.
{2-[2-(6-Oxo-1,6-dihydro-9H-purin-9-yl)ethoxy]propan-2-yl}phosphonic
acid (19a)
Yield 57 mg (16%) of a white amorphous solid; Rf (S8) 0.18; m.p.
155–160 ꢀC. IR: (KBr) mmax 3100–2200, 3127, 3066, 2700, 2600, 1716, 1568,
1520, 1469, 1424, 1380, 1356, 1380, 1363, 1170, 1160, 1098, 1006, 987, 957,
934, 925, 783, 685 cmꢂ1. ESIMS: m/z 301.1 (M-H)- (100). HRMS (ESI): m/z
calcd for C10H14O6N4P (M-H)- 301.0707; found 301.0705. Anal. Calcd for
C10H15O5N4P.2H2O (%): C, 35.23; H, 5.27; N, 16.41 P, 9.60. Found: C,
1
35.51; H, 5.66; N, 16.56; P, 9.15. H NMR (500.0 MHz, D2O): 1.24 (d, 6 H,

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
23
JH,P ¼ 13.1, (CH3)2C); 3.99 (m, 2 H, H-20); 4.54 (m, 2 H, H-10); 8.33 (s, 1 H,
H-2); 8.93 (s, 1 H, H-8). 13C NMR (125.7 MHz, D2O): 23.78 (d, JC,P ¼ 3.9,
(CH3)2C); 48.63 (CH2-10); 63.31 (d, JC,P ¼ 5.9, CH2-20); 77.47 (d,
JC,P ¼ 162.5, C-40); 120.77 (C-5); 144.03 (CH-8); 150.51 (C-4); 150.63 (CH-
2); 158.55 (C-6). 31P{1H} NMR (202.3 MHz, D2O): 23.95.
{2-[2-(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)ethoxy]propan-2-
yl}phosphonic acid (19b)
Yield 94 mg (28%) of a white amorphous solid; Rf (S8) 0.11; m.p. >195 ꢀC.
ESIMS: m/z 316.1 (MH)- (100). HRMS (ESI): m/z calcd for C10H15O5N5P
1
(M-H)- 316.0810; found 316.0816. H NMR (600.1 MHz, D2O þ NaOD):
1.06 (d, 6 H, JH,P ¼ 11.9, (CH3)2C); 3.93 (m, 2 H, H-20); 4.14 (m, 2 H, H-10);
7.77 (s, 1 H, H-8). 13C NMR (125.7 MHz, D2O þ NaOD): 25.69 (d,
JC,P ¼ 4.4, (CH3)2C); 47.02 (CH2-10); 64.51 (d, JC,P ¼ 2.8, CH2-20); 79.03 (d,
JC,P ¼ 151.6, C-40); 120.10 (C-5); 142.10 (CH-8); 154.10 (C-4); 163.74 (C-2);
170.98 (C-6). 31P{1H} NMR (202.3 MHz, D2O þ NaOD): 19.54.
{[2-(6-Oxo-1,6-dihydro-9H-purin-9-yl)ethoxy](phenyl)methyl}phosphonic
acid (19c)
Yield 230 mg (54%) of a white amorphous solid; Rf (S8) 0.14; m.p.
>158 ꢀC. IR: (KBr) mmax 3200-2000, 3062, 3031, 2872, 2635, 1714, 1605,
1605, 1570, 1520, 1494, 1453, 1423, 1377, 1353, 1307, 1266, 1200, 1170,
1107, 1075, 1026, 997, 938, 783, 702, 620, 606 cmꢂ1. ESIMS: m/z 349.1 (M-
H)- (100). HRMS (ESI): m/z calcd for C14H14O5N4P (M-H)- 349.0707;
found 349.0708. Anal. Calcd for C14H15O5N4P.3H2O (%): C, 41.59; H, 5.23;
1
N, 13.86 P, 7.65. Found: C, 41.32; H, 4.71; N, 13.90; P, 8.17. H NMR
0
0
0
0
(500.0 MHz, D2O): 3.88 (ddd, 1 H, Jgem ¼ 11.6, J2 b,1 a ¼ 9.6, J2 b,1 b ¼ 2.9, H-
20b); 4.14 (ddd, 1 H, Jgem ¼ 11.6, J2 a,1 b ¼ 3.8, J2 a,1 a ¼ 3.4, H-20a); 4.48 (ddd,
0
0
0
0
1 H, Jgem ¼ 14.9, J1 b,2 a ¼ 3.8, J1 b,2 b ¼ 2.9, H-10b); 4.49 (d, 1 H, JH,P ¼ 15.4,
0
0
0
0
H-40); 4.57 (ddd, 1 H, Jgem ¼ 14.9, J1 a,2 b ¼ 9.6, J1 a,2 a ¼ 3.4, H-10a); 7.06 (m,
0
0
0
0
2 H, H-o-Ph); 7.17 (m, 2 H, H-m-Ph); 7.22 (m, 1 H, H-p-Ph); 8.15 (s, 1 H,
13
H-2); 8.97 (s, 1 H, H-8). C NMR (125.7 MHz, D2O): 48.84 (CH2-10);
70.10 (d, JC,P ¼ 13.7, CH2-20); 82.74 (d, JC,P ¼ 160.6, CH-40); 120.27 (C-5);
130.49 (d, JC,P ¼ 6.0, CH-o-Ph); 130.85 (d, JC,P ¼ 1.5, CH-m-Ph); 131.06 (d,
JC,P ¼ 2.3, CH-p-Ph); 138.27 (C-i-Ph); 143.70 (CH-8); 149.94 (C-4); 150.63
(CH-2); 158.00 (C-6). 31P{1H} NMR (202.3 MHz, D2O): 16.37.
{[2-(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)ethoxy](phenyl)methyl}phosphonic acid (19d)
Yield 112 mg (29%) of a white amorphous solid; Rf (S8) 0.11; m.p.
>245 ꢀC. IR: (KBr) mmax 3500-2000, 3457, 3332, 3139, 1693, 1623, 1601,

24
K. POMEISL ET AL.
1568, 1493, 1475, 1452, 1409, 1368, 1303, 1175, 1153, 1175, 1118, 1096,
1077, 1004, 1031, 1000, 937, 772, 700, 675 cmꢂ1. ESIMS: m/z 364.1 (M-H)-
(98). HRMS (ESI): m/z calcd for C14H15O5N5P (M-H)- 364.0816; found
364.0815. 1H NMR (500.0 MHz, D2O): 3.90 (ddd, 1 H, Jgem ¼ 11.6,
J2 b,1 a ¼ 9.9, J2 b,1 b ¼ 2.9, H-20b); 4.10 (ddd, 1 H, Jgem ¼ 11.6, J2 a,1 b ¼ 3.6,
0
0
0
0
0
0
J2 a,1 a ¼ 3.3, H-20a); 4.30 (ddd, 1 H, Jgem ¼ 14.8, J1 b,2 a ¼ 3.6, J1 b,2 b ¼ 2.9, H-
0
0
0
0
0
0
10b); 4.41 (ddd, 1 H, Jgem ¼ 14.8, J1 a,2 b ¼ 9.9, J1 a,2 a ¼ 3.3, H-10a); 4.47 (d,
1 H, JH,P ¼ 15.4, H-40); 7.12 (m, 2 H, H-o-Ph); 7.21 (m, 2 H, H-m-Ph); 7.24
(m, 1 H, H-p-Ph); 8.88 (s, 1 H, H-8). 13C NMR (125.7 MHz, D2O): 48.67
(CH2-10); 69.54 (d, JC,P ¼ 13.5, CH2-20); 83.02 (d, JC,P ¼ 160.0, CH-40);
110.40 (C-5); 130.62 (d, JC,P ¼ 6.1, CH-o-Ph); 130.82 (d, JC,P ¼ 1.2, CH-m-
Ph); 131.02 (d, JC,P ¼ 2.2, CH-p-Ph); 138.63 (C-i-Ph); 140.81 (CH-8);
152.19 (C-4); 157.72, 157.75 (C-2,6). 31P{1H} NMR (202.3 MHz,
D2O): 16.03.
0
0
0
0
{(4-Chlorophenyl)[2-(6-oxo-1,6-dihydro-9H-purin-9-
yl)ethoxy]methyl}phosphonic acid (19e)
Yield 287 mg (70%) of a white amorphous solid; Rf (S8) 0.19;
m.p. > 140 ꢀC. IR: (KBr) mmax 3200–2000, 2600, 1715, 1606, 1569, 1520,
1489, 1422, 1400, 1353, 1270, 1200, 1090, 1013, 998, 935, 903, 782, 712,
606 cmꢂ1. ESIMS: m/z 383.2 (M-H)- (99). HRMS (ESI): m/z calcd for
1
C14H13O5N4ClP (M-H)- 383.0318; found 383.0314. H NMR (600.1 MHz,
0
0
0
0
D2O þ NaOD): 3.68 (ddd, 1 H, Jgem ¼ 11.2, J2 b,1 a ¼ 10.5, J2 b,1 b ¼ 2.8, H-
20b); 3.99 (dt, 1 H, Jgem ¼ 11.2, J2 a,1 b ¼ J2 a,1 a ¼ 3.2, H-20a); 4.16 (ddd, 1 H,
0
0
0
0
Jgem ¼ 15.0, J1 b,2 a ¼ 3.2, J1 b,2 b ¼ 2.8, H-10b); 4.17 (d, 1 H, JH,P ¼ 15.1, H-40);
0
0
0
0
4.29 (ddd, 1 H, Jgem ¼ 15.0, J1 a,2 b ¼ 10.5, J1 a,2 a ¼ 3.2, H-10a); 6.81 (m, 2 H,
H-o-C6H4Cl); 6.88 (m, 2 H, H-m-C6H4Cl); 7.83 (s, 1 H, H-2); 7.99 (s, 1 H,
0
0
0
0
13
H-8). C NMR (150.9 MHz, D2O þ NaOD): 47.29 (CH2-10); 69.70 (d,
JC,P ¼ 12.5, CH2-20); 84.64 (d, JC,P ¼ 149.4, CH-40); 126.00 (C-5); 129.86
(CH-m-C6H4Cl); 131.78 (d, JC,P ¼ 5.1, CH-o-C6H4Cl); 134.34 (d, JC,P ¼ 2.5,
C-i-C6H4Cl); 140.64 (C-p-C6H4Cl); 144.00 (CH-8); 151.95 (C-4); 155.35
(CH-2); 170.06 (C-6). 31P{1H} NMR (202.3 MHz, D2O þ NaOD): 13.16.
{[2-(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)ethoxy](4-
chlorophenyl)methyl}phosphonic acid (19f)
Yield 126 mg (30%) of a white amorphous solid; Rf (S8) 0.15;
m.p. > 202 ꢀC. IR: (KBr) mmax 3200–2000, 3384, 3144, 1714, 1683, 1635,
1604, 1559, 1489, 1474, 1402, 1397, 1372, 1295, 1175, 1153, 1115, 1089,
1014, 994, 926, 775, 711 cmꢂ1. ESIMS: m/z 398.1 (M-H)- (100). HRMS
1
(ESI): m/z calcd for C14H15O5N5ClP (M-H)- 398.0427; found 398.0422. H

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
25
0
0
NMR (600.1 MHz, D2O þ NaOD): 3.65 (ddd, 1 H, Jgem ¼ 11.0, J2 b,1 a ¼ 10.2,
J2 b,1 b ¼ 2.8, H-20b); 3.91 (dt, 1 H, Jgem ¼ 11.0, J2 a,1 b ¼ J2 a,1 a ¼ 3.2, H-20a);
0
0
0
0
0
0
4.03 (ddd, 1 H, Jgem ¼ 14.9, J1 b,2 a ¼ 3.2, J1 b,2 b ¼ 2.8, H-10b); 4.17 (ddd, 1 H,
0
0
0
0
Jgem ¼ 14.9, J1 a,2 b ¼ 10.2, J1 a,2 a ¼ 3.2, H-10a); 4.20 (d, 1 H, JH,P ¼ 15.0, H-
0
0
0
0
40); 6.87 (m, 2 H, H-o-C6H4Cl); 6.98 (m, 2 H, H-m-C6H4Cl); 7.72 (s, 1 H,
13
H-8). C NMR (150.9 MHz, D2O þ NaOD): 46.78 (CH2-10); 69.85 (d,
JC,P ¼ 12.0, CH2-20); 84.61 (d, JC,P ¼ 149.1, CH-40); 120.52 (C-5); 129.98
(CH-m-C6H4Cl); 131.88 (d, JC,P ¼ 4.9, CH-o-C6H4Cl); 134.33 (d, JC,P ¼ 2.4,
C-i-C6H4Cl); 140.79 (C-p-C6H4Cl); 141.92 (CH-8); 153.62 (C-4); 163.18 (C-
2); 170.65 (C-6). 31P{1H} NMR (202.3 MHz, D2O þ NaOD): 13.23.
{(4-Nitrophenyl)[2-(6-oxo-1,6-dihydro-9H-purin-9-
yl)ethoxy]methyl}phosphonic acid (19g)
Yield 225 mg (54%) of yellow amorphous solid; Rf (S8) 0.11; m.p. 193-
195 ꢀC. IR: (KBr) mmax 3200–2000, 3072, 2870, 2640, 1714, 1606, 1570,
1519, 1490, 1404, 1377, 1349, 1317, 1267, 1204, 1168, 1119, 1106, 1011,
1001, 934, 862, 834, 783, 739, 694, 619, 605 cmꢂ1. ESIMS: m/z 394.1 (M-
H)- (100). HRMS (ESI): m/z calcd for C14H13O7N5P (M-H)- 394.0558;
found 394.0561. 1H NMR (500.0 MHz, D2O þ NaOD): 3.78 (ddd, 1 H,
Jgem ¼ 11.5, J2 b,1 a ¼ 10.4, J2 b,1 b ¼ 2.8, H-20b); 4.19 (dt, 1 H, Jgem ¼ 11.5,
0
0
0
0
J2 a,1 b ¼ J2 a,1 a ¼ 3.4, H-20a); 4.27 (ddd, 1 H, Jgem ¼ 14.9, J1 b,2 a ¼ 3.4,
0
0
0
0
0
0
J1 b,2 b ¼ 2.8, H-10b); 4.41 (ddd, 1 H, Jgem ¼ 14.9, J1 a,2 b ¼ 10.4, J1 a,2 a ¼ 3.4,
H-10a); 4.42 (d, 1 H, JH,P ¼ 15.7, H-40); 7.13 (m, 2 H, H-o-C6H4NO2); 7.85
(m, 2 H, H-m-C6H4NO2); 7.90 (s, 1 H, H-2); 8.15 (s, 1 H, H-8). 13C NMR
(125.7 MHz, D2O þ NaOD): 47.60 (CH2-10); 71.35 (d, JC,P ¼ 11.8, CH2-20);
83.89 (d, JC,P ¼ 150.8, CH-40); 125.40 (d, JC,P ¼ 1.3, CH-m-C6H4NO2);
125.92 (C-5); 131.08 (d, JC,P ¼ 5.1, CH-o-C6H4NO2); 145.54 (CH-8); 147.80
(CH-2); 148.93 (d, JC,P ¼ 2.6, C-i-C6H4NO2); 149.16 (d, JC,P ¼ 2.5, C-p-
C6H4NO2); 151.12 (C-4); 160.86 (C-6). 31P{1H} NMR (202.3 MHz,
D2O þ NaOD): 12.93.
0
0
0
0
0
0
{[2-(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)ethoxy](4-
nitrophenyl)methyl}phosphonic acid (19h)
Yield 69 mg (16%) of yellow amorphous solid; Rf (S8) 0.17; m.p.
250–253 ꢀC. IR: (KBr) mmax 3500–2000, 3314, 3109, 1706, 1687, 1634, 1605,
1562, 1519, 1491, 1349, 1175, 1107, 1012, 1086, 997, 962, 862, 835, 774,
739, 695, 678 cmꢂ1. ESIMS: m/z 409.1 (M-H)- (100). HRMS (ESI): m/z
1
calcd for C14H14O7N6P (M - 1)- 409.0667; found 409.0669. H NMR
0
0
0
0
(500.0 MHz, D2O): 3.96 (ddd, 1 H, Jgem ¼ 11.7, J2 b,1 a ¼ 9.4, J2 b,1 b ¼ 3.1, H-
20b); 4.14 (ddd, 1 H, Jgem ¼ 11.5, J2 a,1 b ¼ 4.2, J2 a,1 a ¼ 3.3, H-20a); 4.33 (ddd,
0
0
0
0

26
K. POMEISL ET AL.
1 H, Jgem ¼ 14.9, J1 b,2 a ¼ 4.2, J1 b,2 b ¼ 3.1, H-10b); 4.44 (ddd, 1 H,
0
0
0
0
Jgem ¼ 14.9, J1 a,2 b ¼ 9.4, J1 a,2 a ¼ 3.3, H-10a); 4.63 (d, 1 H, JH,P ¼ 16.1, H-40);
0
0
0
0
7.35 (m, 2 H, H-o-C6H4NO2); 8.06 (m, 2 H, H-m-C6H4NO2); 8.95 (s, 1 H,
13
H-8). C NMR (125.7 MHz, D2O): 48.56 (CH2-10); 69.92 (d, JC,P ¼ 12.7,
CH2-20); 82.54 (d, JC,P ¼ 155.8, CH-40); 110.25 (C-5); 125.85 (CH-m-
C6H4NO2); 131.43 (d, JC,P ¼ 5.4, CH-o-C6H4NO2); 140.94 (CH-8); 147.53
(C-i-C6H4NO2); 149.79 (d, JC,P ¼ 2.7, C-p-C6H4NO2); 152.35 (C-4); 157.50,
157.62 (C-2,6). 31P{1H} NMR (202.3 MHz, D2O): 13.78.
{2-Hydroxy-1-[2-(6-oxo-1,6-dihydro-9H-purin-9-yl)ethoxy]ethyl}phosphonic
acid (19i)
Yield 205 mg (64%) of a white amorphous solid; Rf (S8) 0.12; m.p. > 116 ꢀC.
IR: (KBr) mmax 3400-2200, 3127, 2600, 2400, 1710, 1571, 1521, 1457, 1424,
1378, 1352, 1269, 1205, 1170, 1117, 1064, 992, 934, 783, 548, 455 cmꢂ1.
ESIMS: m/z 305.2 (M þ H)þ (100). HRMS (ESI): m/z calcd for C9H14O6N4P
1
(M þ H)þ 305.0651; found 305.0666. H NMR (500.0 MHz, D2O): 3.59–3.70
(m, 2 H, H-40, CHaHbOH); 3.87 (m, 1 H, CHaHbOH); 4.11, 4.20 (2ꢄ dt,
0
0
0
0
0
0
2 ꢄ 1 H, Jgem ¼ 11.2, J2 ,1 ¼ 4.9, H-2 ); 4.60 (t, 2 H, J1 ,2 ¼ 4.9, H-1 ); 8.33 (s,
13
1 H, H-2); 9.05 (s, 1 H, H-8). C NMR (125.7MHz, D2O): 48.40 (CH2-10);
64.27 (d, JC,P ¼ 10.7; CH2OH); 72.07 (d, JC,P ¼ 4.1, CH2-20); 82.08 (d,
JC,P ¼ 152.9, CH-40); 120.48 (C-5); 143.97 (CH-8); 150.51 (C-4); 150.71 (CH-
2); 158.40 (C-6). 31P{1H} NMR (202.3 MHz, D2O): 15.88.
{1-[2-(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)ethoxy]-2-
hydroxyethyl}phosphonic acid (19j)
Yield 255 mg (76%) of a white amorphous solid; Rf (S8) 0; IR: (KBr) mmax
3400-2200, 3160, 1715, 1700, 1643, 1609, 1542, 1474, 1409, 1165, 1121,
1067, 1037, 1010, 933, 771, 680, 553, 475 cmꢂ1. FABMS: m/z 320.1
(M þ H)þ (32). HRMS (FAB): m/z calcd for C9H15O6N5P (M þ H)þ
1
320.0760; found 320.0765. H NMR (500.0 MHz, D2O): 3.61–3.71 (m, 2 H,
H-40, CHaHbOH); 3.89 (m, 1 H, CHaHbOH); 4.09, 4.16 (2 ꢄ dt, 2 ꢄ 1 H,
0
0
0
0
0
0
Jgem ¼ 11.7, J2 ,1 ¼ 4.9, H-2 ); 4.45 (t, 2 H, J1 ,2 ¼ 4.9, H-1 ); 9.03 (s, 1 H, H-
13
8). C NMR (125.7 MHz, D2O): 48.08 (CH2-10); 64.26 (d, JC,P ¼ 10.8;
CH2OH); 71.71 (d, JC,P ¼ 4.3, CH2-20); 82.05 (d, JC,P ¼ 153.0, CH-40); 110.29
(C-5); 141.01 (CH-8); 152.64 (C-4); 157.95, 158.08 (C-2,6).
{1-[2-(8-Bromo-2,6-dioxo-2,3-dihydro-1H-purin-9(6H)-yl)ethoxy]-2-
hydroxyethyl}phosphonic acid (21)
A solution of 18j (426 mg, 0.83 mmol) in acetonitrile (15 mL) was cooled to
0 ꢀC. Bromotrimethylsilane (1.1 mL, 8.32 mmol) was added dropwise and

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
27
the mixture was stirred overnight at room temperature. The mixture was
evaporated in vacuo and then codistilled with water (2x2 mL). The residue
was heated in 2.8 M HCl (15 mL) at 120 ꢀC for 5 h until conversion was
complete. The mixture was concentrated in vacuo to a minimum volume,
diluted with 80% acetic acid (10 mL) and isoamyl nitrite (5.72 mmol) was
subsequently added. The mixture was stirred at rt for 48 h, evaporated to
dryness and deionized on a Dowex 50X8 (Hþ) in water. The product was
eluted with 2.5% aqueous ammonia, the relevant fractions were combined
and evaporated in vacuo. The residue was chromatographed on a Dowex 1
(AcO- form) in a gradient 0-1 M acetic acid, followed by 1 M formic acid.
Appropriate UV absorbing fractions containing the product 21 were evapo-
rated and the product lyophilized from water. Yield: 167 mg (50%) of a
white amorphous solid. Rf (S8) 0.12; IR: (KBr) mmax 3495, 3418, 3260, 3159,
3033, 1713, 1691, 1624, 1577, 1466, 1438, 1280, 1125, 1109, 1076, 1005,
760, 546 cmꢂ1. ESIMS: m/z 398.9 (M þ H)þ (90). HRMS (ESI): m/z calcd
for C9H13O7N4BrP (M þ H)þ 398.9700; found 398.9709. Anal. Calcd for
C9H13O7N4BrP.1.2 H2O (%): C, 25.76; H, 3.70; N, 13.35 P, 7.38. Found: C,
1
25.84; H, 3.54; N, 12.89; P, 7.80. H NMR (500.0 MHz, D2O): 3.61–3.69 (m,
2 H, H-40, CHaHbOH); 3.88 (dt, 1 H, Jgem ¼ 11.7, JH,P ¼ Jvic ¼ 6.9,
CHaHbOH); 4.00, 4.10 (2 ꢄ m, 2 ꢄ 1 H, H-20); 4.28–4.38 (m, 2 H, H-10). 13C
NMR (125.7 MHz, D2O): 49.14 (CH2-10); 63.97 (d, JC,P ¼ 10.6; CH2OH);
72.88 (d, JC,P ¼ 3.8, CH2-20); 82.02 (d, JC,P ¼ 154.1, CH-40); 118.14 (C-5);
125.90 (C-8); 145.69 (C-4); 154.50 (C-2); 161.49 (C-6). 31P{1H} NMR
(202.3 MHz, D2O): 17.14.
{1-[2-(2,6-Dioxo-8-phenyl-2,3-dihydro-1H-purin-9(6H)-yl)ethoxy]-2-
hydroxyethyl}phosphonic acid (22)
A mixture of 21 (101 mg, 0.25 mmol), phenylboronic acid (100 mg,
0.82 mmol), Pd(PPh3)4 (76 mg, 0.07 mmol) and sodium carbonate (88 mg,
0.83 mmol) in degassed H2O (40 mL) was heated (ꢃ 130 ꢀC, oil bath) under
argon for 23 h until conversion of the starting material was complete. The
mixture was cooled to rt, filtered through a Celite pad and concentrated in
vacuo to a minimum volume. The residue was chromatographed on a
Dowex 1 (AcO- form) in gradient 0–1 M acetic acid, followed by 1 M for-
mic acid. The appropriate UV absorbing fraction containing 22 was evapo-
rated and the residue lyophilized from water to give 79 mg (79%) of yellow
amorphous solid. Rf (S8) 0.17; IR: (KBr) mmax 3277, 3233, 3200, 3051, 2798,
1730, 1717, 1700, 1687, 1490, 1450, 1398, 1216, 1193, 1064, 912, 773, 705,
693 cmꢂ1. ESIMS: m/z 395.1 (M-H)- (100). HRMS (ESI): m/z calcd for
1
C15H16O7N4P (M-H)- 395.0751; found 395.0761. H NMR (500.0 MHz,
D2O): 3.57–3.66 (m, 2 H, H-40, CHaHbOH); 3.86 (m, 1 H, CHaHbOH);

28
K. POMEISL ET AL.
0
3.91, 4.07 (2 ꢄ m, 2 ꢄ 1 H, H-20); 4.44 (t, 2 H, J1 ,2 ¼ 4.9, H-1 ); 7.63–7.75
0
0
13
(m, 5 H, H-o,m,p-Ph). C NMR (125.7 MHz, D2O): 48.78 (CH2-10); 64.09
(d, JC,P ¼ 10.7; CH2OH); 72.76 (d, JC,P ¼ 3.6, CH2-20); 82.12 (d, JC,P ¼ 153.2,
CH-40); 113.47 (C-5); 126.80 (C-i-Ph); 132.12 (CH-m-Ph); 132.41 (CH-o-
Ph); 134.87 (CH-p-Ph); 145.03 (C-4); 151.24 (C-8); 154.45 (C-2); 160.11
(C-6). 31P{1H} NMR (202.3 MHz, D2O): 16.48.
Diisopropyl (2-benzyloxy-1-{2-[6-(2,2,2-trifluoroethoxy)-9H-purin-9-
yl]ethoxy}ethyl)phosphonate (23)
Trifluoroethanol (208 lL, 2.89 mmol) was dissolved in dry dioxane (35 mL).
Sodium t-butoxide (2.76 mmol) was added at 0 ꢀC and the mixture was
allowed to warm to 60 ꢀC for 20 min. Phosphonate 18j (1.3 g, 2.62 mmol)
in dioxane (40 mL) was added and the mixture was heated (120 ꢀC, oil
bath) for 6 h until conversion of 23 was almost complete (TLC in S6). The
mixture was concentrated in vacuo to a minimum volume and the residue
chromatographed on silica gel (S6). Yield 652 mg (44%) of 23 as a colorless
oil. Rf (S6) 0.54; IR: (CCl4) mmax 3063, 3034, 1606, 1577, 1499, 1483, 1449,
1415, 1387, 1376, 1347, 1258, 1203, 1166, 1120, 1105, 1089, 1005, 988, 721,
696, 647, 572, 542 cmꢂ1. ESIMS: m/z 561.1 (M þ 1)þ (34). HRMS (ESI): m/
1
z calcd for C24H33O6N4F3P (M þ H)þ 561.2084; found 561.2091. H NMR
(500.0 MHz, CDCl3): 1.21, 1.255, 1.260, 1.28 (4 ꢄ d, 4 ꢄ 3 H, Jvic ¼ 6.2,
(CH3)2CH); 3.63 (m, 1 H, CHaHbO); 3.75 - 3.84 (m, 2 H, H-40, CHaHbO);
4.08 (dddd, 1 H, Jgem ¼ 13.3, J2 b,1 ¼ 6.5, 3.6, JH,P ¼ 0.6, H-20b); 4.20 (dt,
0
0
0
0
0
0
1 H, Jgem ¼ 13.3, J2 a,1 ¼ 3.3, H-2 a); 4.41–4.51 (m, 4 H, H-1 , CH2Ph); 4.68
(dsep, 2 H, JH,P ¼ 7.5, Jvic ¼ 6.2, CH(CH3)2); 4.98–5.06 (m, 2 H, CH2CF3);
7.21 (m, 2 H, H-o-Ph); 7.24–7.31 (m, 3 H, H-m,p-Ph); 8.26 (s, 1 H, H-8);
8.50 (s, 1 H, H-2). 13C NMR (125.7 MHz, CDCl3): 23.86, 23.95 (2 ꢄ d,
JC,P ¼ 4.8, (CH3)2CH); 24.02, 24.06 (2 ꢄ d, JC,P ¼ 3.8, (CH3)2CH); 44.01
(CH2-10); 62.23 (q, JC,F ¼ 36.8, CH2CF3); 70.11 (d, JC,P ¼ 11.4, CH2O); 70.75
(d, JC,P ¼ 5.6, CH2-20); 71.34, 71.59 (d, JC,P ¼ 7.1, CH(CH3)2); 73.36
(CH2Ph); 77.22 (d, JC,P ¼ 165.2, CH-4ꢁ); 120.82 (C-5); 123.15 (d,
JC,F ¼ 277.8, CF3); 127.48 (CH-o-Ph); 127.73 (CH-p-Ph); 128.37 (CH-m-
Ph); 137.49 (C-i-Ph); 144.47 (CH-8); 151.17 (CH-2); 152.77 (C-4); 158.52
(C-6). 19 F NMR (470.3 MHz, CDCl3): ꢂ73.87 (t, JF,H ¼ 8.4).
Diisopropyl (2-hydroxy-1-{2-[6-(2,2,2-trifluoroethoxy)-9H-purin-9-
yl]ethoxy}ethyl)phosphonate (24)
A solution of phosphonate 23 (620 mg, 1.11 mmol) in methanol (10 mL)
was hydrogenated over 10% palladium on charcoal (730 mg) at rt for 53 h
until conversion was complete (TLC in S6). The mixture was filtered
through a Celite pad and the filtrate evaporated in vacuo to give 403 g (78%)

NUCLEOSIDES, NUCLEOTIDES AND NUCLEIC ACIDS
29
of 24 as a colorless oil. ESIMS: m/z 493.1 (M þ Na)þ (100). HRMS (ESI): m/
z calcd for C17H26O6N4F3NaP (Mþ Na)þ 493.1434; found 493.1427.
Diisopropyl (1-{2-[6-(2,2,2-trifluoroethoxy)-9H-purin-9-
yl]ethoxy}vinyl)phosphonate (26)
A mixture of DBU (1.18 mmol) and 92% perfluorobutane-1-sulfonyl fluo-
ride (309 mg, 0.94 mmol) was stirred in dry toluene (5 mL) at rt for 20 min.
A solution of 24 (369 mg, 0.786 mmol) in toluene (10 mL) was added and
the resulting mixture was heated at 100 ꢀC for 2 h until conversion was
complete (TLC in S6). The mixture was concentrated to a minimum vol-
ume in vacuo and the residue was chromatographed on silica gel (S6) and
preparative TLC (S9, S6). Yield 232 mg (65%) of 26 as a white amorphous
solid. Rf (S6) 0.54; IR: (CCl4) mmax 2981, 1605, 1578, 1504, 1465, 1417,
1387, 1376, 1321, 1269, 1239, 1228, 1050, 1007, 990, 888, 647 cmꢂ1. ESIMS:
m/z 453.0 (M þ H)þ (21). HRMS (ESI): m/z) calcd for C17H25O5N4F3P
1
(M þ H)þ 453.1509; found 453.1508. H NMR (500.0 MHz, CDCl3): 1.22,
1.34 (2 ꢄ d, 2 ꢄ 6 H, Jvic ¼ 6.2, (CH3)2CH); 4.12 (m, 2 H, H-20); 4.64 (m,
2 H, H-10); 4.67 (dsep, 2 H, JH,P ¼ 7.8, Jvic ¼ 6.2, CH(CH3)2); 4.82 (dd, 1 H,
JH,P ¼ 37.3, Jgem ¼ 3.1, ¼CHaHb); 5.07 (q, 2 H, JH,F ¼ 8.4, CH2CF3); 5.23
(dd, 1 H, JH,P ¼ 13.0, Jgem ¼ 3.1, ¼CHaHb); 8.17 (s, 1 H, H-8); 8.54 (s, 1 H,
H-2). 13C NMR (125.7 MHz, CDCl3): 23.64 (d, JC,P ¼ 4.6, (CH3)2CH); 23.98
(d, JC,P ¼ 4.0, (CH3)2CH); 42.86 (CH2-10); 62.28 (q, JC,F ¼ 36.9, CH2CF3);
66.07 (d, JC,P ¼ 10.2, CH2-20); 71.64 (d, JC,P ¼ 5.8, CH(CH3)2); 99.19 (d,
JC,P ¼ 24.4, ¼CH2); 120.97 (C-5); 123.13 (d, JC,F ¼ 277.7, CF3); 143.98 (CH-
8); 151.46 (CH-2); 152.74 (C-4); 154.32 (d, JC,P ¼ 227.0, C-40); 158.78 (C-6).
19 F NMR (470.3 MHz, CDCl3): ꢂ73.88 (t, JF,H ¼ 8.4).
Dilithium salt of (1-{2-[6-(2,2,2-trifluoroethoxy)-9H-purin-9-
yl]ethoxy}vinyl)phosphonic acid (27)
A solution of phosphonate 26 (161 mg, 0.36 mmol) and lutidine (39 mg,
0.36 mmol) in acetonitrile (8 mL) was cooled to 0 ꢀC. Bromotrimethylsilane
(3.56 mmol) was added dropwise and the mixture was stirred at rt for 7.5 h.
The mixture was evaporated, neutralized with diluted aqueous ammonia at
-30 ꢀC and evaporated again to dryness. The residue was chromatographed
on DEAE A-25 Sephadex (Cl– form, activated with 0.02 M TEAB) in gra-
dient 0-0.4 M TEAB. Subsequent deionization on activated charcoal at 0 ꢀC
was carried out. The product was then dissolved in water (3 mL), applied
onto a column of Dowex 50X8 (Liþ form) and the column was washed
with water. UV absorbing fraction containing 27 was evaporated and the
residue was lyophilized from water. Yield 21 mg (16%) of 27 as a white
amorphous solid. Rf (S8) 0.26; m.p. > 300 ꢀC; IR: (KBr) mmax 1633, 1610,