Novel acyclic amide-conjugated nucleosides and their analogues

Article abstract of DOI:10.1080/15257770902736467

An effective one-step synthesis of new amide-conjugated nucleosides and their analogues, in the presence of 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4- methylmorpholinium chloride (DMT-MM) as the condensing agent, is presented. Substrate subunits carrying carboxylic group were obtained by acidic hydrolysis of Michael-type adducts of various 5-substituted uracil derivatives to methyl acrylate. Amine substrate was synthesized by reduction of 1-(2′- cyanoethyl)thymine with sodium borohydride in the presence of nickel (II) chloride as catalyst. Other applied amine substrates were 5′-amino- 5′-deoxythymidine and 5-aminouracil. Copyright Taylor & Francis Group, LLC.

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Novel Acyclic Amide-Conjugated
Nucleosides and Their Analogues
Slawomir Boncel a & Krzysztof Walczak a
a Department of Organic Chemistry, Biochemistry and Biotechnology,
Silesian University of Technology, Gliwice, Poland
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Nucleosides, Nucleotides and Nucleic Acids, 28:103–117, 2009
Copyright ꢀC Taylor & Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770902736467
NOVEL ACYCLIC AMIDE-CONJUGATED NUCLEOSIDES AND THEIR
ANALOGUES
Slawomir Boncel and Krzysztof Walczak
Department of Organic Chemistry, Biochemistry and Biotechnology, Silesian University
of Technology, Gliwice, Poland
An effective one-step synthesis of new amide-conjugated nucleosides and their analogues, in the
2
presence of 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM) as the
condensing agent, is presented. Substrate subunits carrying carboxylic group were obtained by acidic
hydrolysis of Michael-type adducts of various 5-substituted uracil derivatives to methyl acrylate.
Amine substrate was synthesized by reduction of 1-(2ꢁ-cyanoethyl)thymine with sodium borohydride
in the presence of nickel (II) chloride as catalyst. Other applied amine substrates were 5ꢁ-amino-5ꢁ-
deoxythymidine and 5-aminouracil.
Keywords Acyclic nucleosides; aminolysis; 5-substituted uracil derivatives; DMT-MM;
amide bond formation
INTRODUCTION
Systems of amide-conjugated units of diversely functionalized acyclic
nucleosides may be considered as potential medicines, for instance
in combination anti-HIV therapy due to synergic effect, e.g. ddC-
MKC analogue conjugated nucleoside (Figure 1, I).[1] Furthermore,
5ꢁ-acetamido-5ꢁ-deoxythymidine derivatives (II) exhibit significant activ-
ity against the P388 mouse leukaemia line.[2] 5ꢁ-Amino-5-fluoro-2ꢁ,5ꢁ-
dideoxy-β-uridine derivatives (III)—with appropriate spacers containing
the amide bonds—exhibit cytotoxicity of considerable decrease compar-
ing to unsubstituted nucleosides.[3] Amide-linked heterodimer synthons
consisting of acyclic nucleoside units and 5ꢁ-amino-5ꢁ-deoxythymidine are
PNA/DNA chimeras (IV).[4] 6ꢁ-Amide-bond based aromatic and non-
Received 8 October 2008; accepted 10 December 2008.
Address correspondence to Slawomir Boncel and Krzysztof Walczak, Department of Organic
Chemistry, Biochemistry and Biotechnology, Silesian University of Technology, Krzywoustego 444–100,
Gliwice, Poland. E-mail: krzysztof.walczak@polsl.pl; slawomir.boncel@polsl.pl
103

104
S. Boncel and K. Walczak
FIGURE 1 Various amide-conjugated analogues of nucleosides.
aromatic nikkomycin conjugates (V) greatly inhibit chitin synthases.[5]
Polyamide-based minor-groove binders were designed as analogues of dis-
tamycin, netropsin, and the oligopeptide analogues.[6] Moreover, amide
bonds are also exploited in the construction of diverse anticancer ther-
mosensitive polymer-drug conjugates; a precursor bearing active moiety is
represented by structure VI.[7] Separately from medicinal application of the
title compounds, a very intriguing amide-conjugated quasi-nucleosides were
reported (VII).[8] These structures, formally being 1,ω-bolaamphiphilic,

Novel Acyclic Amide-Conjugated Nucleosides
105
may form extraordinary topologies like supramolecular fibres (e.g., double-
helical ropes), microtubes or microcrystals. The nature of the observed
self-organization is based upon hydrogen bonds and π-π interactions,
similarly to multiple interactions elucidating quaternary structure of nu-
cleic acids, proteins and nucleic acid-protein complexes. Currently, a
number of methods of amide bond formation in nucleoside derivatives
is known and exploited. In a typical procedure, nucleoside containing
amine group is treated with appropriate acyl halide or other acyl donors
containing easily replaceable group under basic conditions.[9,10] Acylation
using carboxylic acids,[11] esters,[12] or acyl chlorides are common.[13]
The acylation reaction in the presence of magnesium chloride[14,15] or
9-borabicyclo[3.3.1]nonane (9-BBN) give products in excellent yields. [16]
When acids are used as acylating agents the condensation reaction have
to be supported in many cases by the presence of condensing agent,
e.g. DCC.[17] The formed as a by-product 1,3-dicyclohexylurea is poorly
soluble in most of organic solvents, but the removing of its traces may
be troublesome. The formation of the amide bond in the reaction of
appropriate carboxylic derivative with amine in the presence of 2-chloro-
1-methylpyridinium iodide and triethylamine[18] or by treatment of pen-
tachlorophenyl carboxylic esters with the corresponding amines[8] were also
reported.
In this article, we report effortless and efficient method for novel conju-
gated nucleosides synthesis by coupling of two subunits bearing carboxylic
and amino group respectively in the presence of DMT-MM as a condensing
agent.
RESULTS AND DISCUSSIONS
The units containing carboxylic group (2a–e) were synthesized by acidic
hydrolysis of Michael adducts of 5-substituted uracil derivatives to methyl
acrylate (1a–e) (Scheme 1).[19] In turn, mainly utilized subunit carrying
primary amine group, namely 1-(3ꢁ-aminopropyl)thymine trifluoroacetate
4 was synthesized by reduction of Michael adduct (3) of thymine to acry-
lonitrile with sodium borohydride, in the presence of nickel chloride (co-
catalyst) and di-tert-butyl carbonate as a “trapping” agent in methanol.[20]
In preliminary experiments for the synthesis of amide-conjugated
nucleosides the simple aminolysis approach was applied. Unfortunately,
these experiments revealed several synthetic problems. First, aminolysis
of methyl esters (instead of carboxylic acids) with primary aliphatic
amines required significant (400% and higher) excess of nucleophile
(methylamine and 2-aminoethanol). 5-Aminouracil appeared as entirely
unreactive towards ester group under investigated conditions (Scheme 2).
Moreover, prolonged time of the reaction course under basic conditions

106
S. Boncel and K. Walczak
SCHEME 1 Synthesis of subunits for amide-conjugated nucleosides.
supported the formation of products of retro-Michael reaction. When 1b
was refluxed in the presence of DBU from post reaction mixture thymine
1bc (Michael donor) was isolated. Furthermore, 5-bromo- and 5-nitrouracil
derivatives under the above-mentioned conditions undergo undesired
reactions. 5-Bromouracil derivative 1c treated with N -centered nucleophiles
under depicted conditions (Scheme 3) gave, apart from the desired
3-(5-bromo-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-N -(2-hydroxyethyl)
propanamide 1ca, 5-bromouracil 1cc (formed via retro-Michael
reaction) and product of subsequent substitution of bromine atom with
2-aminoethanol, namely N -(2-hydroxyethyl)-3-[5-(2-hydroxyethylamino)-
2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl]propanamide 1cb. Our previous
experiments also revealed that direct Michael-type addition of uracils to
acrylic acid procured work-up problems in the purification of product
and loss of selectivity. Thus, mainly due to the competitive polymerization
reaction, an additional stage of hydrolysis had to be implicated.
As the consequence, this synthetic pathway had to be relinquished as
completely ineffective in the case of the synthesis of intentional conjugated
nucleosides.
Considering the optimal conditions of amide-coupling reaction led
us to astonishingly convenient non-carbodiimide one-step condensing
agent, namely 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium
chloride—DMT-MM.[21] Its versatility of usage owes to non-toxicity, non-
irritation, working under mild conditions, capability of recycling, and prac-
tically total removal of products its dissolution from postreaction mixture:

Novel Acyclic Amide-Conjugated Nucleosides
107
SCHEME 2 Aminolysis of 3-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1-(2H)-yl) propanoate with se-
lected N -nucleophiles.
SCHEME 3 Aminolysis of 3-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1-(2H)-yl)propanoate with 2-
aminoethanol.

108

109

110
S. Boncel and K. Walczak
SCHEME 4 Synthesis of amide-conjugated nucleosides containing short linkage or a sugar constituent.
coupling by-product—4,6-dimethoxy-1H-1,3,5-triazin-2-one (DMT-OH) and
product of DMT-MM decomposition 2,4-dimethoxy-6-morpholin-4-yl-1,3,5-
triazine (DMTM). Hence, DMT-MM and its analogues are at present of high
synthetic importance in the formation of both amide[21] (e.g., synthesis of
peptides) and ester bonds.[22] The target compounds were synthesized via
condensation of carboxylic and amine groups deriving from correspond-
ing stricte- or quasi-nucleoside units (Scheme 4). Under typical applied
conditions, the coupling reactions were performed at ambient temperature
in THF or 1,2-dimethoxyethane with usage of 10% molar excess of amine
substrate salt or pure amine and 10% molar excess of DMT-MM in relation
to acidic substrate (Scheme 4). Total decay of limiting reagents was achieved
in 1.5–24 hours. In a prevailing number of cases, products were precipitating
during the course of the reactions and required only single crystallization.
In the two cases (5b, 5e), isolation on chromatographic column was applied.
The products, conjugated nucleosides (5a–5e) were isolated in moderate to
very good yields, except 5-nitrouracil derivatives. This fact may be colligated
with the known susceptibility of the 5-nitrouracil ring to nucleophilic attack
initiated by amino group, and in consequence, ring opening reactions.[23]
To extend application of the explored condensation method, 5ꢁ-amino-
5ꢁ-deoxythymidine 6 was applied as an amino substrate. The latter was
synthesized in Mitsunobu reaction of thymidine with phthalimide, followed

Novel Acyclic Amide-Conjugated Nucleosides
111
by cleavage of phthalimide moiety under basic conditions.[24] The expected
condensation product (7) was obtained in satisfactory yield. It should
be mentioned that we did not observe products of competitive acylation
neither on 3ꢁ-hydroxyl group nor on nitrogen N-3 of thymidine. On the
other hand, the illustration of significant effectiveness of DMT-MM as the
condensing agent was reaction between 2b and 5-aminouracil 8. Although
all of the substrates were practically insoluble in most of commonly used
solvents, product of coupling was isolated, after total time of reaction
of 120 hours, in 52% yield. Spectroscopic data of the amide-conjugated
nucleosides built from the above mentioned subunits are collected in Tables
1 (1H NMR) and 2 (13C NMR).
CONCLUSION
In summary, we have synthesized novel amide-conjugated nucleosides
and their acyclic analogues with usage of convenient condensing agent
DMT-MM. It was demonstrated that application of DMT-MM enables direct
amide formation on various compounds bearing amine groups, e.g. in 5-
aminouracil or 5ꢁ-amino-5ꢁ-deoxythymidine.
EXPERIMENTAL
General
NMR spectra were recorded at 300 MHz for 1H NMR and 75.5 MHz for
13C NMR on a Varian Inova 300 MHz in DMSO-d6 solution; δ values are
in parts per million relative to tetramethylsilane as an internal standard.
Elemental analyses were obtained using a Perkin-Elmer 240C (Poland)
apparatus. All used reagents were purchased from Aldrich (Germany). TLC
60F254 plates and silica gel 60 (0.040–0.063 mm) were purchased from
Merck (Poland). Melting points were measured at Betius apparatus and are
uncorrected.
5-Substituted 3- (3,4-dihydro- 2,4-dioxopyrimidin-1 (2H)-yl) propanoic
acids (2a–e). General procedure. Michael-type adduct of uracil to methyl
acrylate (1a–e) (1 mmol) was refluxed in 5% HClaq (8 ml). During first 5–15
minutes of refluxing solid phase disappeared. During the next 15 minutes of
boiling neo-crystalline material appeared. The mixture was then cooled to
room temperature and kept for 16 hours in refrigerator. Afterwards crystals
were filtered off, rinsed with ice water to neutral pH and dried on air.
3-(3,4-Dihydro-2,4-dioxopyrimidin-1(2H)-yl)propanoic acid (2a). Color-
less needles; yield 62%; m.p. 176–179◦C, lit.[25] 183–185◦C; 1H NMR: δ: 2.62
(t, 2H, CH2, J = 6.3 Hz); 3.86 (t, 2H, >NCH2, J = 6.3 Hz); 5.53 (d, 1H, H5,
J = 7.5 Hz); 7.63 (d, 1H, H6, J = 7.5 Hz); 11.25 (br s, 1H, H3); 12.42 (br s,
1H, COOH); 13C NMR: δ: 32.8; 44.22; 100.6; 146.2; 150.9; 163.8; 172.4.

112
S. Boncel and K. Walczak
3- (3,4- Dihydro-5- methyl-2,4- dioxopyrimidin-1 (2H)- yl) propano◦ic acid
1
(2b). Colorless needles; yield 82%; m.p. 172–174◦C, lit.[26] 174–175 C; H
NMR: δ: 1.73 (s, 3H, Me); 2.59 (t, 2H, CH2, J = 6.9 Hz); 3.81 (t, 2H, >NCH2,
J = 6.9 Hz); 7.49 (d, 1H, H6, J = 0.9 Hz); 11.23 (br s, 1H, H3); 12.41 (br s,
1H, COOH); 13C NMR: δ: 12.0; 32.9; 44.0; 108.2; 141.9; 150.8; 164.4; 172.4.
3- (3,4-Dihydro- 5-bromo- 2,4-dioxopyrimidin-1 (2H) -yl) propanoic acid
(2c). Colorless needles; yield 98%; m.p. 248–249◦C, lit.[27] 250◦C; 1H NMR:
δ: 2.63 (t, 2H, CH2, J = 6.9 Hz); 3.87 (t, 2H, >NCH2, J = 6.9 Hz); 8.17 (s,
1H, H6); 11.77 (br s, 1H, H3); 12.46 (br s, 1H, COOH); 13C NMR: δ: 32.6;
44.4; 94.2; 145.9; 150.2; 159.7; 172.3.
3-(3,4-Dihydro-5-nitro-2,4-dioxopyrimidin-1(2H)-yl)propanoic acid (2d).
Colorless needles; yield 96%; m.p. 247–249◦C, lit.[28] 261–262 ◦C; 1H NMR:
δ: 2.69 (t, 2H, CH2, J = 6.6 Hz); 4.04 (t, 2H, >NCH2, J = 6.6 Hz); 9.25 (s,
1H, H6); 12.03 (br s, 1H, H3); 12.52 (br s, 1H, COOH); 13C NMR: δ: 32.3;
45.5; 124.5; 149.3; 151.4; 155.1; 172.3.
3- (3,4-Dihydro- 5-(2-methyl- 4-nitroimidazol-1-yl)-2,4-dioxopyrimidin- 1
(2H) -yl)propanoic acid (2e). Light yellow needles; yield 65%; m.p.
1
240–241◦C; H NMR: δ: 2.27 (s, 3H, Me); 2.71 (t, 2H, CH2, J = 6.6 Hz);
3.93 (t, 2H, >NCH2, J = 6.6 Hz); 8.28 (s, 1H, H-imidazole); 8.36 (s, 1H,
H6); 11.99 (br s, 1H, H3); 12.44 (br s, 1H, COOH); 13C NMR: δ: 12.6; 32.4;
44.8; 110.0; 124.0; 145.8; 145.9; 146.6; 150.9; 159.8; 172.4. Analysis calcd.
for C11H15N5O6 (309.24): C42.72, H3.59, N22.65. Found: C42.61. H3.53,
N22.55. ESI MS: 2M+Na = 641.11; Calcd. 2M+Na = 641.47.
1-(3-Amino-propyl)-5-methyl-1H-pyrimidine-2,4-dione trifluoroacetate
(4). A. Preparation of t-Butyl 3-(3,4-dihydro-5-methyl-2,4-dioxopyrimidin-
1(2H)-yl)propylcarbamate (4-Boc-protected)
To a solution of uracil derivative 3 (10 mmol) in methanol (50 ml),
.
cooled in an ice bath, nickel (II) chloride hydrate (NiCl2 6H2O) (0.50
mmol) and di-tert-butyl carbonate (10 mmol) were added subsequently.
After 10 minutes of vigorous stirring the suspension turned light green
and NaBH4 (70 mmol) was added in small portions while intensive stirring
(0.5 hours). After addition of first portion, the mixture turned dark black
and evolution of hydrogen appeared. The internal pressure was regulated
by release of the stopper. After addition of NaBH4, the ice bath was
allowed to warm to ambient temperature (2.5 hours) and the reaction
mixture turned transparent and light gray. TLC analysis (MeOH/CHCl3 1:9
vol./vol.) indicated total consumption of nitrile (3). The excess of NaBH4
was removed by addition of ethyl acetate. The volatiles were removed under
reduced pressure and product was isolated by column chromatography
using a mixture MeOH/CHCl3 (1:9 vol./vol.).
◦
1
White crystals; 96%; m.p. 96–97 C; H NMR δ: 1.38 (s, 9H, C(CH3)3);
1.67 (t, 2H, CH2CH2CH2, J = 6.9 Hz); 1.75 (d, 3H, CH3 thymine, J =
0.6 Hz); 2.92 (q, 2H, CH2NH, J = 6 Hz); 3.61 (t, 2H, >NCH2, J = 7.2
Hz); 6.87 (br. t, 1H, NH-Boc, J = 5.4 Hz); 7.53 (d, 1H, H6, J = 1.2 Hz);

Novel Acyclic Amide-Conjugated Nucleosides
113
11.23 (br. s, 1H, N3-H). 13C NMR δ: 11.96; 28.25; 28.90; 37.10; 45.25; 77.63;
108.41; 141.58; 150.89; 155.57; 164.32. MS m/z (relative intensity) 227 (17),
210 (18), 183 (25), 167 (32), 166 (M+– t-BuOCONH, 100), 154 (83), 153
(58), 140 (67), 127 (24), 126 (60), 110 (51), 96 (36), 79 (25). Analysis calcd
for C13H21N3O4 (283.33): C55.11, H7.47, N14.83. Found: C54.73, H7.20,
N14.53.
B. Deprotection
Total amount of pure N -Boc (9.6 mmol) protected amine was refluxed
in CH2Cl2 (15 ml) for 6 hours in the presence TFA (38.4 mmol). Solvent
was removed under diminished pressure and colorless crystals of amine
trifluoroacetate were dried in a vacuum desiccator, yield 98%. Colorless
crystals; 1H NMR: δ: 1.77 (s, 1H, Me); 1.89 (quin., 2H, CH2CH2CH2, J = 7.5
Hz); 2.82 (tt, 2H, CH2NH3+, J = 6.6 Hz, J = 7.5 Hz); 3.72 (t, 2H, >NCH2,
J = 6.6 Hz); 7.55 (s, 1H, H6); 7.89 (br s, 3H, NH3+); 11.30 (s, 1H, H3); 13
C
NMR δ: 12.0; 26.9; 36.4; 44.6; 109.0; 116.0 (q, CF3, JC-F = 292 Hz); 141.3;
151.2; 158.7 (q, CF3-CO, JC-F = 35 Hz); 164.4.
Aminolysis of methyl 5-substituted 3-(3,4-dihydro-2,4-dioxopyrimidin-
1(2H)-yl)propionates. Michael-type adducts of uracils to methyl acrylate
(1b, 1c) (1 mmol) were dissolved in methanol (or ethanol in the case of
1ba) (8 ml). After dissolution, 2-aminoethanol (5 mmol) was added. The
progress of reaction was monitored by TLC. For the precise conditions see
Scheme 2. Products were isolated by column chromatography using 10%
MeOH/CHCl3 (v/v).
N-methyl-3- (5-methyl-2,4-dioxo -3,4-dihydropyrimidin-
1
(2H)-yl)
propanamide (1ba). White crystals, yield 74%; m.p. 172–173◦C; H NMR:
δ: 1.73 (s, 3H, Me thymine); 2.42 (t, 2H, CH2CO, J = 6.6 Hz); 2.56 (d, 3H,
NH-Me, J = 4.5 Hz); 3.81 (t, 2H, >CH2, J = 6.6 Hz); 7.41 (s, 1H, H6); 7.89
(d, 1H, CONHMe, J = 4.5 Hz); 11.21 (br s, 1H, H3); 13C NMR δ: 12.0; 25.4;
34.2; 44.5; 108.0; 141.9; 150.7; 164.3; 169.9. Analysis calcd. for C9H13N3O4
(211.22): C51.18, H6.20, N19.89. Found: C50.95, H6.01, N19.65.
3-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-N-(2-
1
hydroxyethyl)-propanamide (1bb). White crystals, yield 91%; m.p.
187–189◦C; 1H NMR: δ: 1.73 (s, 3H, Me thymine); 2.44 (t, 2H, CH2CONH,
J = 6.6 Hz); 3.10 (q, 2H, CH2OH, J = 6.0 Hz); 3.36 (q, 2H, NHCH2CH2, J
= 6.0 Hz); 3.81 (t, 2H, >NCH2, J = 6.6 Hz); 4.65 (t, 1H, OH, J = 5.4 Hz);
7.41 (s, 1H, H6); 7.99 (t, 1H, CONH, J = 5.4 Hz); 11.21 (br s, 1H, H3);
13C NMR: δ: 11.9; 34.3; 41.6; 44.5; 59.8; 108.0; 141.9; 150.8; 164.3; 169.7.
Analysis calcd. for C10H15N3O4 (241.25): C49.79, H6.27, N17.42. Found:
C49.55, H6.07, N17.25.
3-(5-bromo-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-N-(2-
hydroxyethyl)-propanamide
(1ca).
White
crystals;
yield
30%;
m.p.194–195◦C; 1H NMR: δ: 2.47 (t, 2H, CH2CO, J = 6.6 Hz); 3.10
(q, 2H, CH2OH, J = 6.0 Hz); 3.36 (q, 2H, CH2CH2OH, J = 6.0 Hz); 3.88

114
S. Boncel and K. Walczak
(t, 2H, >NCH2, J = 6.6 Hz); 4.66 (t, 1H, OH, J = 5.4 Hz); 8.00 (t, 1H,
CONH, J = 5.4 Hz); 8.07 (s, 1H, H6); 11.75 (s, 1H, H3); 13C NMR: δ:
34.0; 41.4; 45.1; 59.8; 94.0; 145.9; 150.2; 159.7; 169.6. Analysis calcd. for
C9H12BrN3O4 (306.12): C35.31, H3.95, N13.73. Found: C34.96, H3.67,
N13.45.
N-(2-hydroxyethyl)-3-[5-(2-hydroxyethylamino)-2,4-dioxo-3,4-dihydro
(2H)pyrimidin-1-yl]propanamide (1cb). White crystals; yield 35%; semi-solid
1
compound; H NMR: δ: 2.43 (t, 2H, CH2CO, J = 6.9 Hz); 2.84 (br q, 2H,
U-NHCH2CH2OH, J = 5.4 Hz); 3.10 (q, 2H, CH2OH, J = 6.0 Hz); 3.36
(q, 2H, CONHCH2, J = 6.0 Hz); 3.56 (q, 2H, U-NHCH2, J = 5.4 Hz); 3.81
(t, 2H, >NCH2, J = 6.9 Hz); 4.30 (br t, 1H, U-NHCH2, J = 5.4 Hz); 4.67
(t, 1H, OH, J = 5.4 Hz); 4.78 (t, 1H, OH, J = 5.4 Hz); 6.56 (s, 1H, H6);
8.00 (t, 1H, CONHCH2, J = 6.0 Hz); 11.34 (s, 1H, H3); 13C NMR: δ: 34.5;
41.5; 44.4; 46.0; 58.9; 59.8; 117.0; 124.3; 148.5; 160.8; 169.7. Analysis calcd.
for C11H18N4O5 (286.29): C46.15, H6.34, N19.57. Found: C46.06, H6.17,
N19.25.
Conjugated Nucleosides: 3-(3,4-Dihydro-2,4-dioxopyrimidin-1
(2H)-yl)-N-(3-(3,4-dihydro-5-methyl-2,4-dioxopyrimidin-1(2H)-
yl)propyl)propanamides (5a–e). General Procedure
To
a
suspension of 3-(3,4-dihydro-2,4-dioxopyrimidin-1(2H)-yl)
propanoic acid 2 (1 mmol) in THF (5 ml) was added dropwise N -
methylmorpholine (1 mmol), then 1-(3-aminopropyl)-5-methylpyrimidine-
2,4(1H,3H)-dione trifluoroacetate 4 (1.1 mmol) and after 10 minutes of
stirring DMT-MM (1.1 mmol) were added. The progress of reaction was
monitored by TLC analysis (20% vol. MeOH/CHCl3). After 1.5–24 hours
the white solid was filtered off and washed with cold methanol. The solvent
from the filtrate was evaporated under diminished pressure and the residue
was purified on column chromatography (20% vol. MeOH/CHCl3).
Fractions containing product after evaporation were combined with
previously obtained solid and recrystallized from ethanol/water.
3-(3,4-Dihydro-2,4-dioxopyrimidin-1(2H)-yl)-N-(3-(3,4-dihydro-5-methyl-
2,4-dioxopyrimidin-1(2H)-yl)propyl)propanamide (5a). White crystals; yield
85%; m.p. 223–225◦C. Analalysis calcd. for C15H19N5O5 (349.34): C51.57,
H5.48, N20.05. Found: C51.31, H5.18. N19.69.
3-(3,4-Dihydro-5-methyl-2,4-dioxopyrimidin-1(2H)-yl)-N-(3-(3,4-dihydro-
5-methyl-2,4-dioxopyrimidin-1(2H)-yl)propyl)propanamide (5b). White
crystals; yield 82%; m.p. 196–198◦C. Analysis calcd. for C16H21N5O5
(363.37): C52.89, H5.83, N19.27. Found: C53.01, H5.80, N19.02.
3-(5-Bromo-3,4-dihydro-2,4-dioxopyrimidin-1(2H)-yl)-N-(3-(3,4-dihydro-
5-methyl-2,4-dioxopyrimidin-1(2H)-yl)propyl)propanamide (5c). White
crystals; yield 71%; m.p. 273–275◦C. Analalysis calcd. for C15H18BrN5O5

Novel Acyclic Amide-Conjugated Nucleosides
115
(428.24): C42,07, H4.24, N16,35. Found: C41.67, H4.11, N15.98. ESI MS
(m/z): M+Na = 450.05, 452.04, calcd. M+Na 451.23.
3-(3,4-Dihydro-5-nitro-2,4-dioxopyrimidin-1(2H)-yl)-N-(3-(3,4-dihydro-
5-methyl-2,4-dioxopyrimidin-1(2H)-yl)propyl)propanamide (5d). Light
yellow crystals; yield 30%; m.p. 240–241◦C; Analysis calcd. for C15H18N6O7
(394.34): C45.69, H4.60, N21.31. Found: C45.52, H4.23, N21.22.
3-(3,4-Dihydro-5-(2-methyl-4-nitroimidazol-1-yl)-2,4-dioxopyrimidin-
1(2H)-yl)-N-(3-(3,4-dihydro-5-methyl-2,4-dioxopyrimidin-1(2H)-
yl)propyl)propanamide (5e). White crystals; yield 66%; Analysis calcd.
for C19H22N8O7 (474.43): C48.10, H4.67, N23.62. Found: C48.22, H4.79,
N23.29.
3-(2,4-Dioxo-3,4-dihydro-pyrimidin-1(2H)-yl)-N-[(2R,3S,5R)-3-hydroxy-
5-(5-methyl-2,4-dioxo-3,4-dihydro-pyrimidin-1(2H)-yl)-tetrahydro-furan-
2-ylmethyl]-propanamide (7). To a suspension of 3-(3,4-dihydro-2,4-
dioxopyrimidin-1(2H)-yl)propanoic acid 2a (1 mmol) in THF (5 ml)
5ꢁ-amino-5ꢁ-deoxythymidine 6 (1.1 mmol) and after 10 minutes of stirring
DMT-MM (1.1 mmol) were added. The progress of reaction was monitored
by TLC analysis (20% vol. MeOH/CHCl3). After 24 hours the white solid
was filtered off and washed with cold ethanol. The solvent from the filtrate
was evaporated under diminished pressure and the residue was purified on
column chromatography (20% vol. MeOH/CHCl3). Fractions containing
product after evaporation were combined with previously obtained solid
◦
and recrystallized from water. White crystals; yield 64%; m.p. 194–196 C;
1H NMR δ: 1.81 (s, 1H, Me); 1.91–2.16 (m, 2H, Hꢁ2); 2.49 (t, 2H, CH2,
J = 6.3 Hz); 3.18–3.41 (m, 2H, Hꢁ5); 3.71 (br s, 1H, Hꢁ4); 3.86 (t, 2H,
>NCH2, J = 6.3 Hz); 4.11 (br s, 1H, Hꢁ3); 5.30 (d, 1H, Cꢁ3-OH, J = 4.2
Hz); 5.47 (d, 1H, H5-uracil, 7.5 Hz); 6.13 (t, 1H, Hꢁ1, J = 6.9 Hz); 7.47 (s,
1H, H6-thymine); 7.52 (d, 1H, H6-uracil, 7.5 Hz); 8.18 (br s, 1H, CONH);
11.23 (br s, 1H, H3-thymine); 11.30 (br s, 1H, H3-uracil); 13C NMR δ: 1.81
12.1; 34.1; 38.4; 41.0; 44.7; 71.2; 83.9; 84.9; 100.5; 109.7; 136.2; 146.1; 150.5;
150.8; 163.7; 169.9. Analysis calcd. for C17H21N5O7 (407.38): C50.12, H5.20,
N17.19. Found: C50.30, H5.02, N6.92.
N-(2,4-Dioxo-1,2,3,4-tetrahydro-pyrimidin-5-yl)-3-(5-methyl-2,4-dioxo-
3,4-dihydro-pyrimidin-1(2H)-yl)-propionamide (9). To a suspension of
3-(3,4-dihydro-5-methyl-2,4-dioxopyrimidin-1(2H)-yl)propanoic acid 2b
(0.5 mmol) in 1,2-dimethoxyethane (10 ml) was added 5-aminouracil
8 (0.75 mmol) and after 10 minutes of stirring DMT-MM (0.75 mmol).
TLC analysis after 120 hours revealed only traces of the limiting substrate.
Subsequently the solvent was removed under diminished pressure. Yellow
residue was boiling in 10 minutes in 7 ml of water. Suspension was filtered
off and precipitate collected from the filter was recrystallized from aqueous
◦
1
DMF. Yellowish crystals; yield 52%; m.p. 322–324 C; H NMR δ: 1.73 (s,
3H, Me); 2.71 (t, 2H, CH2, J = 6.6 Hz); 3.84 (t, 2H, J = 6.6 Hz); 7.47 (s,
1H, H6-thymine); 8.01 (d, 1H, H6-uracil, J = 6.0 Hz); 9.31 (s, 1H, CONH);

116
S. Boncel and K. Walczak
10.67 (d, 1H, H1-uracil, J = 5.1 Hz); 11.24 (br s, 1H, H3-thymine); 11.44
(br s, 1H, H3-uracil); 13C NMR δ: 12.0; 34.7; 44.3; 108.1; 113.0; 130.0; 141.8;
149.7; 150.8; 160.7; 164.3; 169.3. Analysis calcd. for C12H13N5O5 (307.26):
C46.91, H 4.26, N22.79. Found: C47.28, H4.11, N22.36.
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