Convenient and efficient syntheses of N6-and N 4-substituted adenines and cytosines and their 2-deoxyribosides

Article abstract of DOI:10.1080/15257770.2012.742198

Convenient and efficient methods of the synthesis of N⁶-and N⁴-substituted derivatives of adenine and cytosine and their 2-deoxyribosides were developed. The reactions of either unprotected nucleobases (adenine, cytosine) or unprotected 2-deoxyribosides with aryl or alkyl aldehydes give corresponding Schiff bases that can be reduced to the target title compounds with high overall yields. In the case of aryl aldehydes the imine derivatives are obtained in the presence of methoxides in methanol and reduced with sodium borohydride. The corresponding reactions with alkyl aldehydes require the use of acetic acid and borane dimethyl sulfide complex instead.

Full text of DOI:10.1080/15257770.2012.742198

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Convenient and Efficient Syntheses of
N 6- and N 4- Substituted Adenines and
Cytosines and their 2′-Deoxyribosides
Ewelina Adamska a , Jan Barciszewski a & Wojciech T. Markiewicz a
a Institute of Bioorganic Chemistry, Polish Academy of Sciences,
Poznań, Poland
Published online: 05 Dec 2012.
To cite this article: Ewelina Adamska , Jan Barciszewski & Wojciech T. Markiewicz (2012):
Convenient and Efficient Syntheses of N 6- and N 4- Substituted Adenines and Cytosines and their 2′-
Deoxyribosides, Nucleosides, Nucleotides and Nucleic Acids, 31:12, 861-871
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Nucleosides, Nucleotides and Nucleic Acids, 31:861–871, 2012
Copyright ꢀC Polish Academy of Sciences
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770.2012.742198
CONVENIENT AND EFFICIENT SYNTHESES OF N6- AND N4-
SUBSTITUTED ADENINES AND CYTOSINES AND THEIR
2ꢀ-DEOXYRIBOSIDES
Ewelina Adamska, Jan Barciszewski, and Wojciech T. Markiewicz
Institute of Bioorganic Chemistry, Polish Academy of Sciences, Poznan´, Poland
Convenient and efficient methods of the synthesis of N 6- and N 4-substituted derivatives of ade-
2
nine and cytosine and their 2ꢁ-deoxyribosides were developed. The reactions of either unprotected
nucleobases (adenine, cytosine) or unprotected 2ꢁ-deoxyribosides with aryl or alkyl aldehydes give
corresponding Schiff bases that can be reduced to the target title compounds with high overall yields.
In the case of aryl aldehydes the imine derivatives are obtained in the presence of methoxides in
methanol and reduced with sodium borohydride. The corresponding reactions with alkyl aldehydes
require the use of acetic acid and borane dimethyl sulfide complex instead.
Keywords Kinetin; furfuryladenine; furfurylcytosine; 2ꢁ-deoxyribosides; nucleobase;
Schiff base
INTRODUCTION
Kinetin, N 6-furfuryladenine, previously assumed to be an unnatural and
synthetic compound, was identified in several natural sources.[1–8] Properties
of kinetin and its riboside in addition to their cytokinin properties, include
a cytotoxic effect on mouse, human, and plant tumor cells, antiproliferative
and apoptogenic effects against human cancer cells.[5–10] Modified nucleo-
sides and to some extent derivatives of nucleobases are widely investigated
as potential antiviral and antitumor agents.[11–14]
Chemical synthesis of such derivatives of nucleobases and nucleosides
usually involves rather complex transformations and requires a thorough
use of protecting groups and organic media. The first reported approach,
giving products with good to moderate yields was based on the acylation of
adenine followed by the reduction of resulting N 6-acyladenines with lithium
aluminum hydride.[15] Other methods involve preparation of either purine
Received 13 August 2012; accepted 17 October 2012.
The authors are thankful for the financial support of the Ministry of Science and Higher Education
of the Republic of Poland (Grant No. PBZ-MNiSW-07/I/2007/11).
Address correspondence to Wojciech T. Markiewicz, Institute of Bioorganic Chemistry, Polish
Academy of Sciences, Noskowskiego 12/14, PL 61-704 Poznan´, Poland. E-mail: markwt@ibch.poznan.pl
861

862
E. Adamska et al.
or pyrimidine derivatives with good leaving groups at positions either 6 or
4 that are then subjected to a nucleophilic attack with appropriate amines.
Such activated precursors were obtained from nucleoside substrates (ribo-
sides and 2ꢁ-deoxyribosides of uracil, thymine, hypoxanthine, guanine) in a
reaction with: thionyl chloride, arylsulphonyl chlorides, methanesulphonyl
chloride, phosphoryl chloride, p-chlorophenyl phosphodichloridate, hex-
amethyldisilazane.[16–23] Another approach relies on the transformation of
the exo-amino group of 2ꢁ-deoxyadenosine into the 1,2,4-triazole residue in
the reaction with 1,2-bis[(dimethylamino)methylene]hydrazine.[24] The use
of N 4-arylsulphonyl derivatives was reported in the case of cytosine nucleo-
sides.[25] Yet, an unprotected 2ꢁ-deoxyadenosine was successfully reacted with
hydrogen sulfide or selenide and further transaminated with an excess of dif-
ferent primary amines.[26–28] All these methods led to nucleoside derivatives
that could be subjected of N -glycoside bond cleavage in order to provide
kinetin or its analogs as well as N 4-substituted cytosines.[29] Direct alkylation
of adenine leads to N 9-substituted derivatives[30] and is rather applicable to
adenosine derivatives.[31]
Undoubtedly, there is a need for simple and cheap procedures that
would shorten synthetic routes to compounds of pharmacological interest.
Recently, we became interested in the synthesis of cytosine deriva-
tives, as N 4-furfurylcytosine (FC) was found in natural sources, similarly
to N 6-furfuryladenine.[32] FC exhibits anti-aging effects, inhibits the activ-
ity of methyltransferase DNMT1 and probably can inhibit the expression
of methyltransferase DNMT1.[33,34] We report here a simple approach for
the synthesis of exo-amino substituted derivatives of adenine, cytosine as well
as their 2ꢁ-deoxyribosides, based on the synthesis of Schiff bases from the
corresponding aldehydes and nucleobase or nucleoside substrates.
RESULTS AND DISCUSSION
A series of adenine (3–5, 7–16) derivatives was synthesized analogously to
the case of cytosine.[32–34] It turned out that only a minor modification of the
reaction conditions was needed. The solubility of adenine (1) as well as other
nucleobases is rather low in organic solvents. However, it dissolves quite well
in methanol in the presence strong bases. Thus, the reaction of adenine (1)
with ‘aromatic’ aldehydes proceeds smoothly and almost quantitatively in
anhydrous methanol in the presence of magnesium or sodium methoxides
as ‘drying agents’[32,34] (Scheme 1) to form Schiff base derivatives. It was nec-
essary to increase the molar excess of methoxide from 6 to 10 equivalents.
However, it was not possible to achieve the complete reaction of adenine (1)
despite the use of higher excess of basic catalyst, prolonged reaction time
or an increase of temperature. All such efforts resulted in the formation
of by-products and gave unclean reaction mixtures. Then, imines were sub-
jected to reduction with sodium borohydride in the reaction media without

Syntheses of Substituted Adenines and Cytosines
863
SCHEME 1 Reagents and conditions: (i) Mg(MeO)2 (10 equiv.), aromatic aldehyde (6 equiv.), MeOH,
55◦C, 3 h; (ii) NaBH4 (8 equiv.), 0.5 h, MeOH, rt; (iii) AcOH (10 equiv.), alkyl aldehyde (4 equiv.),
MeOH, reflux, 3 h; (iv) a) BH3–DMS (2 equiv.), THF, CH2Cl2, 12 h; b) HCl, MeOH aq.
any work-up procedure (Scheme 1) leading to the desired N 6-methylaryl
adenine derivatives (3–6) in high yields (Scheme 1). This approach allowed
us to obtain many compounds in a short time without protecting adenine at
N 9 position. It is interesting that under the same reaction conditions unpro-
tected 2ꢁ-deoxyadenosine (2) can be converted to appropriate N 6-alkylaryl
derivatives as well (6) (Scheme 1).
In the case of alkyl aldehyde as a substrate, the reaction was found to
be efficiently catalyzed by acetic acid (7–16) (Scheme 1). The use of basic
conditions (methoxide) for alkyl aldehydes led to condensation products.
Again, it was necessary to increase the excess of the catalyst to drive the
Schiff base formation to completion. The derived Schiff bases were reduced
with borane dimethyl sulfide complex to corresponding products after the
removal of the catalyst. The applied work-up procedure included an ex-
traction step to avoid purification difficulties due to the formation of high
boiling alcohols from excess aldehydes (Schemes 1, 2).
Thus, the procedure developed by us opens an easy access to adenine
derivatives starting with appropriate aldehyde substrates.
Synthesis of many cytosine derivatives was recently reported.[32–34] Here
we show that our method can be applied to obtain N 4-alkylcytosine deriva-
tives from alkyl aldehydes of higher molecular weight than butyl aldehyde.
Additionally, we describe here two derivatives of 2ꢁ-deoxycytidine (18) that
could be obtained from an unprotected nucleoside. Thus, the reaction with
two aryl aldehydes, benzaldehyde and furfural, catalyzed with methoxide

864
E. Adamska et al.
SCHEME 2 Reagents and conditions: (i) Mg(MeO)2 (5 equiv.), aromatic aldehyde (6 equiv.), MeOH,
55◦C, 3 h; (ii) NaBH4 (9 equiv.), 0.5 h, MeOH, rt; (iii) AcOH (5 equiv.), alkyl aldehyde (4 equiv.), MeOH,
reflux, 3 h; (iv) a) BH3 –DMS (2 equiv.), THF, CH2Cl2, 12 h; b) HCl, MeOH aq.
gave after the reduction of Schiff derivatives the desired products with yields
of 40% and 75% respectively (Scheme 2).
EXPERIMENTAL
All reagents were commercially available. Solvents, except of methanol,
were used without further purification. Reaction progress was usually moni-
tored by TLC using Merck silica gel 60 F254 plates and UV light at 254 nm.
All chromatographic purifications were carried out on the silica gel 60
0.0063–0.200 mm (Merck) and on the silica gel 60 0.040–0.063 mm or on
1
the silanized silica gel 60 0.063–0.200 mm (Merck, No 107719). H-NMR
(300 MHz) spectra were recorded on a Varian 300 MHz spectrometer using
DMSO-d6 as a solvent. 13C-NMR spectra were recorded on a Bruker 400 MHz
spectrometer or a Varian 300 MHz spectrometer using DMSO-d6, D2O or
MeOH-d4 as solvents and internal standards. Mass spectra (EMS) were mea-
sured with ESI Micro Q-TOF (Bruker). UV spectra of all compounds in
methanol were measured using Jasco V-650 spectrophotometer and molar
extinction coefficients at 260 nm (ε260) were calculated.
General Procedure of Synthesis of Adenine and
2ꢀ-Deoxyadenosine Derivatives with Aryl Substituent at N6
Adenine (1) (500 mg, 3.7 mmol, 1 equiv.) or 2ꢁ-deoxyadenosine
(2) (929 mg, 3.7 mmol, 1 equiv.) and appropriate aromatic aldehyde
(22.2 mmol, 6 equiv.) were added to methanol (ca. 30 mL) containing mag-
nesium methylate (3.19 g, 37 mmol, 10 equiv.). The reaction was carried for
3 hours at 55◦C. Then sodium borohydride (1.12 g, 29.6 mmol, 8 equiv.)

Syntheses of Substituted Adenines and Cytosines
865
was slowly added to the mixture and the reduction reaction was continued
for 30 minutes at room temperature. After that, the pH of the mixture was
adjusted to 7 with conc. HCl and then the solvents were removed by evap-
oration under reduced pressure. Water (30 mL) was added to the residue
and the mixture was extracted with ethyl acetate (30 mL). The organic layer
was extracted with an aqueous solution of hydrochloric acid (1 M, 30 mL).
The combined aqueous extracts were neutralized with potassium hydroxide
and extracted with ethyl acetate (50 mL). The organic layer was dried with
magnesium sulfate and then evaporated under reduced pressure to give
products as a white solid. The yields range: from 40% to 64%.
N6-Benzyladenine (3) ε260 13.1 M−1 cm−1. Yield 64%. MS (ESI) m/z
1
[M+H]+ calcd 226.11; found 226.11. H-NMR (400 MHz, DMSO) δ 4.7 (s,
2H, CH2 of benzyl); 7.2 -7.3 (m,5H, aromatic); 7.2 -7.3 (s, 1H, NH-6); 8.0 (s,
1H, H-8); 8.1 (s, 1H, H-2); 12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO)
δ 42.95; 126.54; 127.17; 128.15; 129.23; 138.80; 149.25; 152.32; 154.33.
N6-(p-Methylbenzyl)adenine (4) ε260 10.6 M−1 cm−1. Yield 40%. MS (ESI)
m/z [M+H]+ calcd 240.12; found 240.12. 1H-NMR (400 MHz, DMSO) δ 2.2
(s, 3H, CH3 of p-methylbenzyl); 4.7 (s, 2H, CH2 of p-methylbenzyl); 7.2 -7.3
(m, 4H, aromatic); 7.2 -7.3 (s, 1H, NH-6); 8.0 (s, 1H, H-8); 8.1 (s, 1H, H-
2); 12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 20.61; 62.72; 126.44;
127.18; 128.54; 128.69; 129.01; 135.54; 138.77; 152.30; 171.97.
N6-(5-Hydroxymethylfurfuryl)adenine (5) Yield 40%–65%. MS (ESI)
m/z [M+H]+ calcd 246.09, found 246.09, [M+Na]+ calcd 268.08; found
268.08. 1H-NMR (400 MHz, DMSO) δ 4.3 (s, 2H, 5-CH2 of hydroxymethyl);
5.1 (s, 2H, CH2 of furfuryl); 6.1 (s, 2H, H-3 and H-4 of furfuryl); 7.1 (s, 1H,
H-2); 8.1 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 55.68; 69.25; 107.22;
107.35; 130.01; 130.02; 152.37; 154.63.
2ꢁ-Deoxy-N6-furfuryladenosine (6) ε260 13.1 M−1 cm−1. Yield 45%. MS
(ESI) m/z [M+H]+ calcd 332.13 [M+H]+; found 332.13 [M+Na]+ calcd
354.12; found 354.12. 1H-NMR (400 MHz, DMSO) δ 2.2 (m, 1H, H-2ꢁ); 2.7
(m, 1H, H-2ꢁꢁ); 3.5 (m, 1H, H-5ꢁ); 3.6 (m, 2H, H-5ꢁꢁ); 3.9 (q, JA = 4.0 Hz. JB
= 6.8 Hz, 1H, H-3ꢁ); 4.4 (m, 1H, H-4ꢁ); 4.6 (s, 2H, CH2 of furfuryl); 5.1 (q, JA
= 4.9 Hz, JB = 6.3Hz, 1H, OH-3ꢁ); 5.2 (d, J = 3,9 Hz, 1H, OH-5ꢁ); 6.2 (d, J =
4.4 Hz, 1H, H-3 of furfuryl); 6.3 (m, 1H, H-4 of furfuryl); 6,3 (m, 1H, H-1ꢁ);
7.5 (m, 1H, H-5 of furfuryl); 8.2 (s, 1H, H-2); 8.2 (s, 1H, HN-6); 8.3 (s, 1H,
H-8). 13C-NMR (100 MHz, DMSO) δ 36.46; 59.73; 61.85; 70.93; 83.92; 87.99;
106.61; 110.40; 128.29; 139.62; 141.78; 152.21; 152.88; 154.31.
General Procedure of Synthesis of 2ꢀ-Deoxycytidine Derivatives
with Aryl Substituent at N4
2ꢁ-Deoxycytidine (18) (613 mg, 2.70 mmol, 1 equiv.) and appropriate
aromatic aldehyde (16.20 mmol, 6 equiv) were added to methanol (ca.
25 mL) containing magnesium methylate (1.2 g, 13.50 mmol, 5 equiv.).
The reaction was continued for 3 hours at 55◦C. Then sodium borohydride

866
E. Adamska et al.
(923 mg, 24.30 mmol, 9 equiv.) was slowly added to the mixture and the
reduction reaction was continued for 30 minutes at room temperature. After
that, the pH of the mixture was adjusted to 7 with conc. HCl and then
the solvents were removed by evaporation under reduced pressure. Water
(25 mL) was added to the residue and the mixture was extracted with ethyl
acetate (25 mL). The organic layer was extracted with an aqueous solution
of hydrochloric acid (0.05 M, 25 mL). The combined aqueous extracts were
neutralized with aq. potassium hydroxide (1 M, 1.3 mL) and extracted with
ethyl acetate (25 mL). The organic layer was dried with magnesium sulfate
and then evaporated under reduced pressure to give a product as a white
solid. The yields range: about 75%.
N4-Benzyl-2ꢁ-deoxycytidine (19) MS (ESI) m/z [M+H]+ calcd 318.14;
found 318.14, [M+Na]+ calcd 340.13; found 340.13. Yield 75%. 1H-NMR
(400 MHz, DMSO) δ 1.9 (m, 1H, H-2ꢁ); 2.1 (m, 1H, H-2ꢁꢁ); 3.5 (q, JA =
20.8 Hz, JB = 9.2 Hz, 2H, H-5ꢁ, H-5ꢁꢁ); 3.7 (q, 1H, JA = 9.2 Hz, JB = 4.8 Hz,
H-3ꢁ); 4.1 (m, 1H, H-4ꢁ); 4.5 (d, J = 7.6 Hz, 2H, CH2 of benzyl); 4.9 (t, J =
7.2 Hz, 1H, -OH); 5.1 (d, J = 5.6 Hz, 1H, -OH); 5.8 (d, J = 10 Hz, 1H, H-5);
6.1 (t, J = 8 Hz, 1H, H-1ꢁ); 7,3 (m, 5H, aromatic); 7.7 (d, J = 10.4 Hz, 1H, H-
6); 8.1 (t, J = 8 Hz, 1H, NH-4). 13C-NMR (100 MHz, DMSO) δ 42.24; 47.16;
64.19; 73.44; 89.21; 89.68; 100.02; 129.89; 130.41; 131.28; 131.68; 135.01;
139.60; 142.92; 159.60; 165.95.
N4-Furfuryl-2ꢁ-deoxycytidine (20) ε260 7.4 M−1 cm−1. Yield 78%. MS (ESI)
m/z [M+H]+ calcd 308.12; found 308.12, [M+Na]+ calcd 330.11; found
330.11. 1H-NMR (400 MHz, DMSO) δ 2.0 (m, 1H, H-2ꢁ); 2.1 (m, 1H, H-2ꢁꢁ);
3.4 (d, J = 1.5 Hz, 1H, H-3ꢁ); 3.5 (m, 2H, H-5ꢁ); 3.9 (m, 1H, H-4ꢁ); 4.5 (d, J =
5.1 Hz, 2H, CH2 of furfuryl); 5.8 (m, 1H, H-1ꢁ); 6.2 (m, 1H, H-2 of furfuryl);
6.3 (m, 1H, H-5); 6.4 (m, 1H, H-3 of furfuryl); 7.5 (m, 1H, H-4 of furfuryl); 7.6
(d, J = 6.2 Hz, 1H, H-6); 8.2 (m, 1H, NH-4). 13C-NMR (100 MHz, DMSO) δ
45.64; 60.32; 69.39; 74.08; 84.08; 89.09; 94.45; 107.21; 110.45; 142.20; 151.81;
155.31; 160.94; 163.23.
General Procedure of Synthesis of Adenine Derivatives with Alkyl
Substituent at N6
Adenine (1) (500 mg, 3.7 mmol, 1 equiv.) and an alkyl aldehyde
(14.8 mmol, 4 equiv.) were added to methanol (ca. 30 mL) containing acetic
acid (2.1 mL, 37 mmoles, 10 equiv.). The mixture was refluxed for 3 hours.
Then, the solvents were removed by evaporation under reduced pressure. Af-
ter that, the imine was isolated by partitioning between ethyl acetate (50 mL)
and water (50 mL). Then the organic lawyer was evaporated and the reaction
mixture was partitioned between hexane (50 mL) and methanol (50 mL).
The methanolic solution was evaporated to dry residue. The Schiff base was
dissolved in methylene chloride (30 mL) and then reduced with 2M borane

Syntheses of Substituted Adenines and Cytosines
867
dimethyl sulfide complex in THF (3.7 mL, 7.4 mmol, 2 equiv.). After that,
water (3 mL) and acetone (10 mL) were added to the reaction flask. Then all
solvents were evaporated and then conc. HCl (1 mL) in methanol (10 mL)
and water (10 mL) were added to the reaction mixture. The mixture was
left for 12 hours. Then the solvents were evaporated and the product was
separated by silanized silica gel column chromatography with a mixture of
water and acetone or silica gel ethyl acetate and methanol.
N6-n-Ethyladenine (7) ε260 10.5 M−1 cm−1. Yield 57%. MS (ESI)
m/z [M+H]+ calcd 164.09; found 164.09, [M+Na]+ calcd 186.08; found
186.07. 1H-NMR (400 MHz, DMSO) δ 1.1 (t, J = 7.3 Hz, 3H, CH3 of ethyl);
3.5 (t, J = 6.3 Hz, 2H, H-1 of ethyl); 7.5 (t, J = 6.3 Hz, 1H, NH-6); 8.0 (s, 1H,
H-8), 8.1 (s, 1H, H-2). 13C-NMR (100 MHz, DMSO) δ 15.62; 35.35; 118.61;
141.15; 152.15; 152.61; 154.29.
N6-n-Propyladenine (8) ε260 12.5 M−1 cm−1. Yield 23%. MS (ESI) m/z
[M+H]+ calcd 178.11 [M+H]+; found 178.11, calcd [M+Na]+ 200.09;
found 200.09. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 7.2 Hz, 3H, CH3
of propyl); 1.6 (m, 2H, H-2 of propyl); 3.5 (s, 2H, H-1 of propyl); 7.5 (s,
1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR
(100 MHz, DMSO) δ 15.62; 23.57; 42.85; 140.01; 154.29.
N6-n-Butyladenine (9) ε260 11.6 M−1 cm−1. Yield 83%. MS (ESI) m/z
[M+H]+ calcd 192.12; found 192.12. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 7.2 Hz, 3H, CH3 of butyl); 1.2 (m, 2H, H-3 of butyl); 1.4 (s, 2H, H-2 of
butyl); 3.5 (m, 2H, H-1 of butyl); 7.5 (s, 1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s,
1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 14.11; 20.70;
32.70; 42.85; 119.62; 131.37; 139.09; 153.69; 155.86.
N6-n-Pentyladenine (10) ε260 12.7 M−1 cm−1. Yield 25%. MS (ESI) m/z
[M+H]+ calcd 206.14, found 206.14.1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 6.8 Hz, 3H, CH3 of pentyl); 1.2 (m, 4H, H-4, H-3 of pentyl); 1.4 (m, 2H,
H-2 of pentyl); 3.5 (m, 2H, H-1 of pentyl); 7.5 (s, 1H, NH-6); 8.0 (s, 1H, H-8),
8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 13.62;
21.90; 28.82; 38.67; 40.34; 119.00; 138.82; 152.26; 161.00.
N6-n-Hexyladenine (11) ε260 13.3 M−1 cm−1. Yield 45%. MS (ESI) m/z
[M+H]+ calcd 220.16; found 220.16. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 7.2 Hz, 3H, H-1 of hexyl); 1.2 (m, 6H, H-2, H-3, H-4 of hexyl); 1.4 (s, 2H,
H-5 of hexyl); 3.5 (m, 2H, H-6 of hexyl); 7.5 (s, 1H, NH-6); 8.0 (s, 1H, H-8),
8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 14.25;
23.27; 27.32; 32.34; 40.94; 139.23; 153.71.
N6-n-Heptyladenine (12) ε260 12.7 M−1 cm−1. Yield 70%. MS (ESI) m/z
[M+H]+ calcd 234.17; found 234.17. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 6.7 Hz, 3H, CH3 of heptyl); 1.2 (m, 8H, H-6, H-5, H-4, H-3 of hep-
tyl); 1.4 (m, 2H, H-2 of heptyl); 3.5 (m, 2H, H-1 of heptyl); 7.5 (m, 1H,
NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR
(100 MHz, DMSO) δ 14.27; 23.26; 27.61; 32.57; 41.05; 119.24; 139.42; 153.69;
155.61.

868
E. Adamska et al.
N6-n-Octyladenine (13) ε260 13.6 M−1 cm−1. Yield 50%. MS (ESI)
m/z [M+H]+ calcd 248.19; found 248.19, [M+Na]+ calcd 270.17; found
270.17. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.4 Hz, 3H, CH3 of octyl);
1.2 (m, 10H, H-7, H-6, H-5, H-4, H-3 of octyl); 1.4 (m, 2H, H-2 of octyl); 3.5
(m, 2H, H-1 of octyl); 7.5 (m, 1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2),
12.8 (s, 1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 13.30; 22.35; 26.69; 28.05;
29.59; 31.62; 40.14; 138.66; 152.73.
N6-n-Nonyladenine (14) ε260 13.8 M−1 cm−1. Yield 20%. MS (ESI) m/z
[M+H]+ calcd 262.20; found 262.20. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 6.7 Hz, 3H, H-1 of nonyl); 1.2 (m, 12H, H-2, H-3, H-4, H-5, H-6, H-7
of nonyl); 1.4 (m, 2H, H-8 of nonyl); 3.4 (m, 2H, H-9 of nonyl); 7.6 (m,
1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR
(100 MHz, DMSO) δ 14.28; 23.33; 27.64; 29.22; 29.59; 32.62; 41.04; 139.49;
153.71
N6-n-Decyladenine (15) ε260 11.6 M−1 cm−1. Yield 20%. MS (ESI) m/z
[M+H]+ calcd 276.22; found 276.22. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 7.0 Hz, 3H, CH3 of decyl); 1.2 (m, 14H, H-9, H-8, H-7, H-6, H-5, H-4,
H-3 of decyl); 1.4 (m, 2H, H-2 of decyl); 3.5 (m, 2H, H-1 of decyl); 7.5 (m,
1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2), 12.8 (s, 1H, NH-9). 13C-NMR
(100 MHz, DMSO) δ 14.26; 23.32; 26.62; 27.62; 29.19; 32.63; 41.10; 139.63;
152.65.
N6-n-Dodecyladenine (16) ε260 12.2 M−1 cm−1. Yield 20%. MS (ESI) m/z
[M+H]+ calcd 304.25; found 204.25. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 7.0 Hz, 3H, CH3 of dodecyl); 1.2 (m, 18H, H-11, H-10, H-9, H-8, H-7,
H-6, H-5, H-4, H-3 of dodecyl); 1.4 (m, 2H, H-2 of dodecyl); 3.5 (m, 2H, H-1
of dodecyl); 7.5 (m, 1H, NH-6); 8.0 (s, 1H, H-8), 8.1 (s, 1H, H-2), 12.8 (s,
1H, NH-9). 13C-NMR (100 MHz, DMSO) δ 14.19; 23.04; 27.03; 29.71; 29.76;
29.98; 30.02; 32.29; 41.04; 138.45; 149.03; 153.10; 155.19.
General Procedure of Synthesis of Cytosine Derivatives with Alkyl
Substituent at N6
Cytosine (17) (400 mg, 3.60 mmol, 1 equiv.) and alkyl aldehyde
(14.40 mmol, 4 equiv.) were added to methanol (ca. 35 mL) containing
acetic acid (1 mL, 18.00 mmol, 5 equiv.). The mixture was refluxed for
3 hours. Then, the solvents were removed by evaporation under reduced
pressure. After that, the imine was isolated by partitioning between ethyl
acetate (20 mL) and water (20 mL). Then the organic lawyer was evapo-
rated and the reaction mixture was partitioned between hexane (20 mL)
and methanol (20 mL). The methanolic solution was evaporated to dry
residue. Then, products were isolated using reverse chromatography with
a mixture of water and acetone. The Schiff bases (2 mmol) were dissolved
in methylene chloride (5 mL) and then reduced with 2M borane dimethyl

Syntheses of Substituted Adenines and Cytosines
869
sulfide complex in THF (2 mL, 4 mmol, 2 equiv.) for 12 hours at ambient
temperature. After that water (3 mL) and acetone (10 mL) were added to
the reaction flask. Then all solvents were evaporated and conc. HCl (1 mL)
in methanol (10 mL) and water (10 mL) were added to the reaction mixture.
The mixture was left for 12 hours. Then the solvents were evaporated and
the product was separated by silanized silica gel column chromatography
with a mixture of water and acetone.
N4-n-Pentylcytosine (21) ε260 7.1 M−1 cm−1. Yield 52%. MS (ESI)
m/z [M+H]+ calcd 182.13; found 182.13, [M+Na]+ calcd 204.11; found
204.11. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.7 Hz, 3H, CH3 of pentyl);
1.2 (m, 4H, H-4, H-3 of pentyl); 1.4 (m, 2H, H-2 of pentyl); 3.2 (q, JA =
12.7 Hz, JB = 6.8 Hz, 2H, H-1 of pentyl); 5.5 (d, J = 6.7 Hz, 1H, H-5); 7.2 (d,
J = 6.7 Hz, 1H, H-6); 7.5 (t, J = 3.8 Hz, 1H, NH-4); 10.2 (s, 1H, NH-1). 13C-
NMR (100 MHz, DMSO) δ 13.88; 21.86; 28.27; 28.86; 93.27; 141.14; 156.79;
164.45.
N4-n-Hexylcytosine (22) ε260 6.2 M−1 cm−1. Yield 37%. MS (ESI)
m/z [M+H]+ calcd 196.14; found 196.15, [M+Na]+ calcd 218.13; found
218.13. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.8 Hz, 3H, CH3
of hexyl), 1.2 (m, 6H, H-5, H-4, H-3 of hexyl); 1.4 (m, 2H, H-2 of
hexyl); 3.2 (q, JA = 12.6 Hz, JB = 6.8 Hz, 2H, H-1 of hexyl); 5.5 (d,
J = 7.2 Hz, 1H, H-5); 7.2 (d, J = 7.2 Hz, 1H, H-6); 7.6 (t, J = 5.3 Hz,
2H, NH-4); 10.2 (s, 1H, NH1). 13C-NMR (100 MHz, DMSO) δ 13.90; 22.08;
26.20; 28.59; 31.05; 93.36; 141.23; 157.08; 164.52.
N4-n-Heptylcytosine (23) ε260 6.5 M−1 cm−1. Yield 37%. MS (ESI)
m/z [M+H]+ calcd 210.16; found 210.16, [2M+H]+ calcd 419.31; found
419.32. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.8 Hz, 3H, CH3 of heptyl);
1.2 (m, 8H, H-6, H-5, H-4, H-3 of heptyl); 1.4 (m, 2H, H-2 of heptyl); 3.2 (q,
JA = 12.8 Hz, JB = 6.8 Hz, 2H, H-1 of heptyl); 5.5 (d, J = 7.3 Hz, 1H, H-5);
7.2 (d, J = 7.3 Hz, 1h, H-6); 7.5 (s, 1H, NH-4); 10.2 (s, 1H, NH-1). 13C-NMR
(100 MHz, DMSO) δ 13.09; 21.95; 26.46; 28.45; 28.58; 31.24; 93.29; 141.19;
156.66; 164.35.
N4-n-Octylcytosine (24) ε260 6.2 M−1 cm−1. Yield 40%. MS (ESI) m/z
[M+H]+ calcd 224.18; found 224.18. 1H-NMR (400 MHz, DMSO) δ 0.8 (t,
J = 6.8 Hz, 3H, CH3 of octyl); 1.2 (m, 10H, H-7, H-6, H-5, H-4, H-3 of octyl);
1.4 (m, 2H, H-2 of octyl); 3.2 (q, JA = 12.2 Hz, JB = 6.3 Hz, 2H, H-1 of octyl);
5.5 (d, J = 6.8 Hz, 1H, H-5); 7.2 (d, J = 6.8 Hz, 1H, H-6); 7.5 (t, J = 5.3 Hz,
1H, NH-4); 10.2 (s, 1H, NH-1). 13C-NMR (100 MHz, DMSO) δ 13.92; 22.06;
26.51; 27.38; 28.67; 28.60; 28.76; 31.25; 91.19; 156.82; 164.46.
N4-n-Nonylcytosine (25) ε260 3.2 M−1 cm−1. Yield 40%. MS (ESI) m/z
[M+H]+ calcd 238.19; found 238.19, [M+Na]+ calcd 260.17; found 260.17.
1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.8 Hz, 3H, CH3 of nonyl); 1.2 (m,
12H, H-8, H-7, H-6, H-5, H-4, H-3 of nonyl); 1.4 (m, 2H, H-2 of nonyl); 3.2
(m, 2H, H-1 of nonyl); 5.5 (d, J = 7.3 Hz, 1H, H-5); 7.2 (d, J = 6.8 Hz, 1H,
H-6); 7.5 (t, J = 4.8 Hz, 1H, NH-4); 10.2 (s, 1H, NH-1). 13C-NMR (100 MHz,

870
E. Adamska et al.
DMSO) δ 13.93; 22.07; 24.49; 25.54; 26.50; 28.64; 28.80; 28.97; 31.26; 93.25;
141.20; 156.83; 164.46.
N4-n-Decylcytosine (26) ε260 2.8 M−1 cm−1. Yield 53%. MS (ESI) m/z
[M+H]+ calcd 252.20; found 252.20, [M+Na]+ calcd 274.19; found 274.19.
1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.8 Hz, 3H, CH3 of decyl); 1.2 (m,
14H, H-9, H-8, H-7, H-6, H-5, H-4, H-3 of decyl); 1.4 (m, 2H, H-2 of decyl);
3.2 (q, JA = 12.2 Hz, JB = 6.3 Hz, 2H, H-1 of decyl); 5.5 (d, J = 6.8 Hz, 1H,
H-5); 7.2 (d, J = 6.8 Hz, 1H, H-6); 7.5 (t, J = 5.3 Hz, 1H, NH-4); 10.2 (s,
1H, NH-1). 13C-NMR (100 MHz, DMSO) δ 13.92; 22.06; 25.24; 25.48; 26.52;
28.59; 28.67; 28.94; 29.00; 31.26; 93.23; 141.16; 156.80; 164.46.
N4-n-Dodecylcytosine (27) ε260 3.3 M−1 cm−1. Yield 25%. MS (ESI)
m/z [M+H]+ calcd 280.20; found 280.20; [M+Na]+ calcd 302.19; found
302.21. 1H-NMR (400 MHz, DMSO) δ 0.8 (t, J = 6.8 Hz, 3H, CH3 of dode-
cyl); 1.2 (m, 18H, H-11, H-10, H-9, H-8, H-7, H-6, H-5, H-4, H-3 of dodecyl);
1.4 (m, 2H, H-2 of dodecyl); 3.2 (m, 2H, H-1 of dodecyl); 5.5 (d, J = 6.8 Hz,
1H, H-5); 7.2 (d, J = 6.8 Hz, 1H, H-6); 7.5 (t, J = 5.3 Hz, 1H, NH-4); 10.2 (s,
1H, NH-1). 13C-NMR (100 MHz, DMSO) δ 13.93; 19.14; 22.07; 25.48; 26.50;
28.68; 28.79; 28.99; 29.00; 31.27; 93.27; 141.17; 156.80; 164.46.
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