Building blocks for Polyamide Nucleic Acids: Facile synthesis using potassium fluoride doped natural phosphate as basic catalyst

Article abstract of DOI:10.1081/NCN-120019491

Potassium fluoride doped natural phosphate, inexpensive and environmentally friendly catalyst, is shown to be an efficient basic catalyst for the N1/N9 alkylation of different nucleobases as synthons for PNAs.

Full text of DOI:10.1081/NCN-120019491

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Building Blocks for Polyamide Nucleic Acids: Facile
Synthesis Using Potassium Fluoride Doped Natural
Phosphate as Basic Catalyst
A. Alahiane a , M. Taourirte a , A. Rochdi a , N. Redwane a , S. Sebti b , J. W. Engels c & H. B.
Lazrek a d
a Laboratoire de Chimie Bioorganique, Faculté des Sciences Semlalia , Universite Cadi
Ayyad , Marrakech, Maroc
b Laboratoire de Chimie Organique Appliquée, Faculté des Sciences BenM'Sik , Université
Hassan II , Sidi Othmane Casablanca, Maroc
c Institute of Organic Chemistry and Chemical Biology, Johann Wolfgang Goethe University ,
Frankfurt am Main, Germany
d Laboratoire de Chimie Bioorganique, Faculté des Sciences Semlalia , Universite Cadi
Ayyad , Marrakech, BP2390, Maroc
Published online: 24 Jun 2011.
To cite this article: A. Alahiane , M. Taourirte , A. Rochdi , N. Redwane , S. Sebti , J. W. Engels & H. B. Lazrek (2003) Building
Blocks for Polyamide Nucleic Acids: Facile Synthesis Using Potassium Fluoride Doped Natural Phosphate as Basic Catalyst,
Nucleosides, Nucleotides and Nucleic Acids, 22:2, 109-114, DOI: 10.1081/NCN-120019491
To link to this article: http://dx.doi.org/10.1081/NCN-120019491
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NUCLEOSIDES, NUCLEOTIDES & NUCLEIC ACIDS
Vol. 22, No. 2, pp. 109–114, 2003
Building Blocks for Polyamide Nucleic Acids:
Facile Synthesis Using Potassium Fluoride
Doped Natural Phosphate as
Basic Catalyst
A. Alahiane,1 M. Taourirte,1 A. Rochdi,1 N. Redwane,1 S. Sebti,2
J. W. Engels,3 and H. B. Lazrek1,
*
1
´
Laboratoire de Chimie Bioorganique, Faculte des Sciences Semlalia,
Universite Cadi Ayyad, Marrakech, Maroc
2
´
´
Laboratoire de Chimie Organique Appliquee, Faculte des Sciences BenM’Sik,
´
Universite Hassan II, Sidi Othmane Casablanca, Maroc
3Institute of Organic Chemistry and Chemical Biology,
Johann Wolfgang Goethe University,
Frankfurt am Main, Germany
ABSTRACT
Potassium fluoride doped natural phosphate, inexpensive and environmentally
friendly catalyst, is shown to be an efficient basic catalyst for the N1=N9 alkyla-
tion of different nucleobases as synthons for PNAs.
In 1991, Nielson et al.[1] developed a new class of oligonucleotide analogues
known as Polyamide (or Peptide) Nucleic Acids (PNAs) in which the entire sugar
phosphate backbone has been replaced by a peptide-like backbone. These oligomers
´
*Correspondence: H. B. Lazrek, Laboratoire de Chimie Bioorganique, Faculte des Sciences
Semlalia, Universite Cadi Ayyad, Marrakech, BP2390, Maroc; E-mail: hblazrek@caramail.com.
109
DOI: 10.1081/NCN-120019491
Copyright # 2003 by Marcel Dekker, Inc.
1525-7770 (Print); 1532-2335 (Online)
www.dekker.com

110
Alahiane et al.
Scheme 1.
of nucleobase are derived from N-(2-aminoethyl) glycine which recognize and bind
strongly to specific DNA or RNA sequences.[2] These characteristics make them
potentially and extremely useful as an antisense or antigene drug.[3]
The application of inorganic solid acids as heterogeneous catalysts for organic
synthesis is an area of intense research. Silica gel, alumina, montmorillonite, zeolite
and natural phosphate have been shown to function as effective catalyst for liquid-
phase organic transformations.[4] The advantages of these heterogeneous catalysts
over the homogeneous systems include stability, ease handling, lack of corrosion
and other environmental hazards, and ease of recovery and regeneration.
We have shown recently that natural phosphate is a new Lewis acids catalysts
for 1,3-dipolar cycloaddition[5] and for acyclonucleoside synthesis.[6] In continuation
of our program on the use of natural phosphate as catalyst and in the search of
an alternative strategy which would open the way to a combination of PNA and
oligonucleotides synthesis, we have developed a new and easy synthesis of the PNA
precursor (ethyl acetate-nucleobase) using a cheap KF doped natural phosphate as a
basic catalyst (Sch. 1, 2, 3).
The use of potassium fluoride on alumina (KF=Al2O3) as a base for functiona-
lization of amide and N-alkylation, has been described in the literature.[7] In order to
assess influence of natural phosphate doped with KF as basic catalyst on the synth-
esis of ethyl acetate-nucleobase derivatives, a number of experiments were performed
to optimize reaction conditions. Results of these studies, are summarized in Table 1.
Both KF and NP=KF (175=25) had a weak catalytic activity (entries 1 and 2).
When the amount of NP=KF was increased (350=50),the reaction yield was tripled
Scheme 2.

Building Blocks for Polyamide Nucleic Acids
111
Scheme 3.
Table 1. Catalyst influence on the N-alkylation of uracil.
Entry
Catalyst
Weight ratio (mg=mg)a
Time (h)
Yield (%)b
1
2
3
KF
NP=KF
NP=KF
50
175=25
350=50
10
2
1.5
28
15
50
aThe amount of catalyst used in reactions with 100 mg of uracil.
bPurification by silica gel chromatography.
(entry 3). The reaction was monitored by thin layer chromatography, and it was
stopped when the N1,N3-bisalkylated product appeared. This procedure appears
to be regioselective and gives only the N1 isomer for uracil (Sch. 1).
To expand the scope and the synthetic utility of this reaction using NP=KF
(350=50), we next examined the N-alkylation of other nucleobases under similar
conditions (Sch. 2, 3 and Table 2).
The poor solubility of unprotected nucleobases (cytosine, adenine and guanine)
excluded their use in most reactions. Introduction of a protecting group was neces-
sary to increase the solubility of these bases.[8,9]
Table 2. Alkylation of different heterocyclic bases.
Entry
Heterocyclic base
Time (h)
Yield (%)a
1
2
3
4
5
6
Uracil
Thymine
4-N-Benzoylcytosine
4-N-Benzoyladenine
2-N-Acetylguanine
2-N-acetyl-6-O-(N,N-diphenylcarbamoyl)guanine
1a
1b
1c
1d
1e
1f
1.5
2
1
4
4
50
55
60
50
43b
70c
3
aPurification by silica gel chromatography.
bN9=N7: 60=40 yield ratio.
cN9=N7: 95=5 yield ratio.

112
Alahiane et al.
These protected nucleobases were alkylated to give the desired N1-alkylated
pyrimidines (entries 1–3) and N9-alkylated purines derivatives along or with other
regioisomers (entries 4–6). It was reported that 6-O-(N,N-diphenylcarbamoyl) pro-
tected guanine 1f has been reported to undergo alkylation with high regioselectivity
to give in some cases 99=1 ratio in favour of the N9 regioisomer.[9]
Interesting, we found that the 2-N-acetyl 6-O-(N,N-diphenylcarbamoyl)guanine
1f (Sch. 3) when reacted with ethyl bromoacetate in the presence of NP=KF afforded
a nearly 95=5 ratio of N9=N7 isomers in 70% yield (entry 6, Table 2).
All compounds were characterized fully by spectroscopic and elemental
analyses, which were found to be in accordance with the proposed structures.[8]
In conclusion, we showed that our method, in general, provides the desired ethyl
acetate-nucleobases in yields comparable to those reported in the literature using
basic conditions.
On the other hand the use of KF doped natural phosphate as basic catalyst
provides a significant new and effective method for the environmentally compatible
and practical synthesis of these derivatives.
EXPERIMENTAL
The 1H NMR spectra were recorded using a Brucker AC 250 MHz spectrometer
DMSO-d6 was used as a solvent and internal reference. Mass spectra (MS) were
obtained with JEOL JMS DX 300 instrument using fast atomic bombardment
(FABþ). Thin layer chromatography was performed on plates of kieselgel 60 F254
(Merck) and short-wave ultraviolet light (254 nm) was used to detect the UV-absorb-
ing spots. Column chromatography separation was carried out on silica gel (0.063–
0.2 mm Merck).
For the preparation of natural phosphate see Ref.[4c]
Preparation of Doped Natural Phosphate
350 mg of natural phosphate and 50 mg of KF were mixed in 10 mL of water and
evaporated to dryness and dried for 6 h at 150ꢀC. The obtained solid residue was
used as basic catalyst in alkylation reactions.
General Procedure
A typical experimental procedure is described for uracil. To a mixture of uracil
(100mg, 0.892 mmol) and ethyl bromoacetate (2 eq) in dry acetonitrile (12 mL) was
added NP=KF (350 mg=50mg). After stirring for 1.5 h at reflux, the mixture was filtered
and the filtrate was concentrated in vaccuo. Purification of the residue by flash column
chromatography gave N-1 (ethyl acetate) uracil (80 mg, 50%) as a white precipitate.
1-(Ethoxycarbonylmethyl)uracil 2a. Yield: 50%, Rf ¼ 0.62 (CHC13=MeOH)
1
(90=10, v=v), H NMR d: 11.4 (s, 1H, NH-3); 7.65 (d, 1H, H-6, J ¼ 7.8 Hz); 5.65
(d, 1H, H-5, J ¼ 7.8 Hz); 4.55 (s, 2H, NCH2); 4.17 (q, 2H, COOCH2CH3); 1.23
(t, 3H, COOCH2CH3). MS (FABþ, GT) m=z 199 [M þ H]þ.

Building Blocks for Polyamide Nucleic Acids
113
1-(Ethoxycarbonylmethyl)thymine 2b. Yield: 55%, Rf ¼ 0.70 (CHC13=MeOH)
1
(90=10, v=v), H NMR d: 7.21 (s, 1H, H-6); 4.40 (s, 2H, NCH2); 4.20 (q, 2H,
COOCH2CH3); 1.90 (s, 3H, CH3); 1.25 (t, 3H, COOCH2CH3), MS (FABþ, GT)
m=z 213 [M þ H]þ.
4-N-benzoyl-1-(Ethoxycarbonylmethyl)cytosine 2c. Yield: 60%, Rf ¼ 0.81
1
(CHC13=MeOH)(90=10, v=v), H NMR d: 11.25 (s, 1H, NHBz); 8.12 (d, 2H, o-H
benzoyl); 8.0–7.34 (m, 5H, H-5 and H-6; m,p-H Benzoyl); 4.64 (s, 2H, NCH2);
4.15 (q, 2H, COOCH2CH3); 1.20 (t, 3H, COOCH2CH3), MS (FABþ, GT) Mass
spectrum FABþ (GT) m=z 304 [M þ H]þ.
6-N-(Benzoyl)-9-(Ethoxycarbonylmethyl)adenine 2d. Yield: 47%, Rf ¼ 0.51
1
(CHC13=MeOH)(90=10, v=v), H NMR d: 11.24 (s; 1H, NHBz); 8.80 (s, 1H, H-2);
8.50 (s, 1H, H-8); 8.10 (d, 2H, o-H Benzoyl); 7.70–7.58 (m, 3H, m,p-H Benzoyl);
5.28 (s, 2H, NCH2); 4.24 (q, 2H, COOCH2CH3); 1.27 (t, 3H, COOCH2CH3); MS
(FABþ, GT) m=z 326 [M þ H]þ.
2-N-(Acetyl)-9-(Ethoxycarbonylmethyl)guanine 2e. Yield: 25%, Rf ¼ 0.34
(CHCl3=MeOH)(90=10, v=v), 1H NMR d: 12.12 (s, 1H, NHAc); 11.61 (s, 1H,
NH-3); 8.15 (s, 1H, H-8); 5.22 (s, 1H, NCH2); 4.18 (q, 2H, COOCH2CH3); 2.51
(s, 3H, CH3CO); 1.22 (t, 3H, COOCH2CH3); MS (FABþ, GT) m=z 280 [M þ H]þ.
2-N-(Acetyl)-7-(Ethoxycarbonylmethyl)guanine 2e0. Yield: 18%, Rf ¼ 0.52
(CHC13=MeOH)(90=10, v=v), 1H NMR d: 12.12 (s, 1H, NHAc); 11.62 (s, 1H,
NH-3); 8.15 (s, 1H, H-8); 5.21 (s, 1H, NCH2); 4.17 (q, 2H, COOCH2CH3); 2.20
(s, 3H, CH3CO); 22 (t, 3H, COOCH2CH3); MS (FABþ, GT) m=z 280 [M þ H]þ.
2-N-(Acetyl)-9-(Ethoxycarbonylmethyl)guanine 2g. Yield: 67%, Rf ¼ 0.80
(CHCl3=MeOH)(90=10, v=v),1H NMR d: 10.80 (s, 1H, NH-3); 8.70 (s, 1H, H-8);
7.35–760 (m, 10H, phenyl); 5.26 (s, 1H, NCH2); 4.20 (q, 2H, COOCH2CH3); 2.58
(s, 3H, CH3CO); 1.26 (t, 3H, COOCH2CH3); MS (FABþ, GT) m=z 555 [M þ H]þ.
ACKNOWLEDGMENTS
Financial support by the Ministry of National Education (PARS: Chim
053=1998) and by the cooperation CNRST (Morocco)=DFG (Germany) are grate-
fully acknowledged. We thank Professor J. L. Abbud (CSIC, IQFR, Madrid, Spain)
for his interest in this work, and for reviewing this manuscript. We would also like to
thank the referees for their comments.
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Received June 12, 2002
Accepted December 16, 2002