Synthesis and Properties of 7‐Substituted 7‐Deazapurine (Pyrrolo[2,3‐d]pyrimidine) 2′‐Deoxyribonucleosides实验方法详情页

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Synthesis and Properties of 7‐Substituted 7‐Deazapurine (Pyrrolo[2,3‐d]pyrimidine) 2′‐Deoxyribonucleosides

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Abstract

7?Substituted 7?deazapurine (pyrrolo[2,3?d ]pyrimidine) 2??deoxyribonucleosides are synthesized by stereoselective nucleobase anion glycosylation. The introduction of a halogen at C7 is performed regioselectively either on the nucleobase or on the nucleoside. The pK a values of a series of 7?deazapurine 2??deoxyribonucleosides are provided, and fluorescence properties are also discussed.

Keywords: pyrrolo[2,3?d]pyrimidine; 7?deazapurine; 2??deoxyribonucleoside; nucleobase anion glycosylation; stereoselectivity; regioselective halogenation; pKa values; fluorescence; quenching

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Basic Protocol 1: Preparation of 7‐Substituted 7‐Deazapurine Nucleosides Related to 2′‐Deoxyadenosine and 2′‐Deoxyinosine
Basic Protocol 2: Preparation of 7‐IODO‐7‐Deazapurine Nucleosides Related to 2′‐Deoxyguanosine and 2′‐Deoxyxanthosine
Basic Protocol 3: Preparation of 7‐Bromo‐7‐Deaza‐2′‐Deoxyisoguanosine
Basic Protocol 4: Determination of pKa Values of Nucleosides
Reagents and Solutions
Commentary
Literature Cited
Figures
Tables

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Basic Protocol 1: Preparation of 7‐Substituted 7‐Deazapurine Nucleosides Related to 2′‐Deoxyadenosine and 2′‐Deoxyinosine Materials

Potassium hydroxide (KOH) powder (purity, ≥85%; Sigma)
Anhydrous acetonitrile (MeCN)
Tris[2‐(2‐methoxyethoxy)ethyl]amine (TDA‐1)
4‐Chloro‐5‐iodo‐7H ‐pyrrolo[2,3‐d ]pyrimidine ( S.7 ; see Pudlo et al., )
2‐Deoxy‐3,5‐di‐O ‐(p ‐toluoyl)‐α‐D‐erythro ‐pentofuranosyl chloride ( S.8 ; see Hoffer, ; Rolland et al., )
Anhydrous dichloromethane (CH 2 Cl 2 )
Silica gel 60 (particle size, <0.063 mm; Merck)
Ethyl acetate (EtOAc)
Petroleum ether (b.p. range, 40° to 80°C)
Isopropanol (i ‐PrOH)
Methanol (MeOH)
1,4‐Dioxane
25% (v/v) ammonium hydroxide (NH 4 OH)
Palladium catalyst Pd(PPh 3 ) 4
Copper iodide (CuI)
Argon gas
Anhydrous dimethylformamide (DMF)
Triethylamine (Et 3 N)
Cyclopentylacetylene
Sodium methoxide (NaOMe)
2 M NaOH
1 M HCl

10‐, 50‐, and 100‐mL round‐bottom flasks
3‐cm‐diameter Buchner funnel with filter paper circles
Rotary evaporator connected to a vacuum pump
3 × 50– and 4 × 50–cm chromatography columns
0.2‐mm‐thick silica gel 60 F 254 aluminum TLC plates (Merck)
254‐nm UV lamp
High‐vacuum pump (final pressure, <1 mmHg)
Steel bomb (autoclave)
Reflux condenser
Heating mantle with controller

Additional reagents and equipment for flash chromatography ( appendix 3E ) and thin‐layer chromatography (TLC; appendix 3D )

Basic Protocol 2: Preparation of 7‐IODO‐7‐Deazapurine Nucleosides Related to 2′‐Deoxyguanosine and 2′‐Deoxyxanthosine Materials

7‐(2‐Deoxy‐β‐D‐erythro ‐pentofuranosyl)‐2‐(formylamino)‐4‐methoxy‐7H ‐pyrrolo[2,3‐d ]pyrimidine ( S.12 ; see Seela and Driller, )
Anhydrous acetonitrile (MeCN)
Isobutyric anhydride
Triethylamine
Silica gel 60 (particle size, <0.063 mm; Merck)
Anhydrous dichloromethane (CH 2 Cl 2 )
Acetone
Cyclohexane
N ,N ‐Dimethylformamide (DMF)
N ‐Iodosuccinimide
5% (w/v) sodium bicarbonate (NaHCO 3 )
Anhydrous sodium sulfate (Na 2 SO 4 )
2 M NaOH
1 M HCl
Methanol (MeOH)
Sodium methoxide (NaOMe)
10% (v/v) acetic acid (AcOH) in H 2 O
Sodium nitrite (NaNO 2 )
Ethanol (EtOH)
Sodium iodide (NaI)
Chlorotrimethylsilane (Me 3 SiCl)
0.1 M NaH 2 PO 4 buffer, pH 7.0 (see recipe )

25‐, 50‐, and 250‐mL round‐bottom flasks
Rotary evaporator connected to a vacuum pump
4 × 50–cm chromatography columns
0.2‐mm‐thick silica gel 60 F 254 aluminum TLC plates (Merck)
254‐nm UV lamp
3‐cm‐diameter Buchner funnel with filter paper circles
High‐vacuum pump (final pressure, <1 mmHg)
Separatory funnel
5‐cm‐diameter funnel with folded 10‐cm‐diameter Whatman no. 1 filter
Reflux condenser
Heating mantle with controller

Additional reagents and equipment for flash chromatography ( appendix 3E ) and thin‐layer chromatography (TLC; appendix 3D )

Basic Protocol 3: Preparation of 7‐Bromo‐7‐Deaza‐2′‐Deoxyisoguanosine Materials

2‐Amino‐4‐chloro‐7H ‐pyrrolo[2,3‐d ]pyrimidine ( S.17 ; see Seela et al., )
Anhydrous pyridine
Pivaloyl chloride
Potassium hydroxide (KOH) powder (purity, ≥85%; Sigma)
Anhydrous dichloromethane (CH 2 Cl 2 )
N ‐Bromosuccinimide
Methanol (MeOH)
Anhydrous acetonitrile (MeCN)
Tris[2‐(2‐methoxyethoxy)ethyl]amine (TDA‐1)
2‐Deoxy‐3,5‐di‐O ‐(p ‐toluoyl)‐α‐D‐erythro ‐pentofuranosyl chloride ( S.8 ; see Hoffer, ; Rolland et al., )
Silica gel 60 (particle size, <0.063 mm; Merck)
Petroleum ether (b.p. range, 40° to 80°C)
1,4‐Dioxane
25% (v/v) ammonium hydroxide (NH 4 OH)
1:5 (v/v) acetic acid (AcOH)/H 2 O
Sodium nitrite (NaNO 2 )
Isopropanol (i ‐PrOH)
100‐ and 250‐mL round‐bottom flasks
Rotary evaporator connected to a vacuum pump
3‐cm‐diameter Buchner funnel with filter paper circles
High‐vacuum pump (final pressure, <1 mmHg)
5 × 50–cm chromatography columns
0.2‐mm‐thick silica gel 60 F 254 aluminum TLC plates (Merck)
254‐nm UV lamp
Steel bomb (autoclave)
5 × 20–cm Serdolit AD‐4 column (resin particle size, 0.1 to 0.2 mm; Serva)

Additional reagents and equipment for flash chromatography ( appendix 3E ) and thin‐layer chromatography (TLC; appendix 3D )

Basic Protocol 4: Determination of pKa Values of Nucleosides Materials

4‐Amino‐5‐bromo‐7‐(2‐deoxy‐β‐D‐erythro ‐pentofuranosyl)‐3,7‐dihydro‐2H ‐pyrrolo[2,3‐d ]pyrimidin‐2‐one ( S.4c ; see protocol 3 )
Phosphate buffer solution, pH 4.5 (see recipe )
1 M H 3 PO 4
3 M NaOH

100‐mL volumetric flask
100‐mL beaker
pH meter
UV‐Vis spectrophotometer

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Figure 1.10.1 The structures of selected 7‐deazapurine 2′‐deoxyribonucleosides. Purine and systematic numbering systems are shown in the first two structures. Compounds: S.1 , 7‐deaza‐2′‐deoxyadenosine; S.2 , 7‐deaza‐2′‐deoxyguanosine; S.3 , 2‐amino‐7‐deaza‐2′‐deoxyadenosine; S.4 , 7‐deaza‐2′‐deoxyisoguanosine; S.5 , 7‐deaza‐2′‐deoxyinosine; S.6 , 7‐deaza‐2′‐deoxyxanthosine. References: S.1a (Seela and Kehne, ); S.1b ‐e (Seela and Thomas, ; Seela and Zulauf, ); S.1f (Seela et al., ); S.2a (Winkeler and Seela, ); S.2b‐e (Ramzaeva and Seela, ); S.3a (Seela et al., ); S.3b‐d (Seela and Peng, ); 4a (Seela and Wei, ); S.4b,c (Seela and Peng, ); S.5a (Seela and Menkhoff, ); S.5b ‐d (Ramzaeva et al., ); S.6a (Seela et al., ); and S.6b ‐d (Seela and Shaikh, ).

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Figure 1.10.2 Preparation of 7‐deaza‐7‐iodo‐2′‐deoxyadenosine (S.1d ) and 7‐deaza‐7‐iodo‐2′‐deoxyinosine (S.5d ) is carried out by glycosylation of an iodinated nucleobase. S1.d can then be used in a palladium‐catalyzed Sonogashira cross‐coupling reaction to yield the 7‐alkynyl derivative S.10 (Seela et al., ). TDA‐1, tris[2‐(2‐methoxyethoxy)ethyl]amine; Tol, p ‐toluoyl.

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Figure 1.10.3 Preparation of 7‐iodo‐7‐deaza‐2′‐deoxyguanosine (S.2d ) and 7‐iodo‐7‐deaza‐2′‐deoxyxanthosine (S.6d ) is carried out by iodination of a precursor nucleoside. i ‐Bu2 O, isobutyric anhydride; DMF, N,N ‐dimethylformamide; NIS, N ‐iodosuccinimide.

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Figure 1.10.4 Preparation of 7‐bromo‐7‐deaza‐2′‐deoxyisoguanosine (S.4c ) is carried out by glycosylation of a brominated nucleobase followed by amination at C4 and selective deamination at C2′. NBS, N ‐bromosuccinimide; Piv, pivaloyl; TDA‐1, tris[2‐(2‐methoxyethoxy)ethyl]amine; Tol, p ‐toluoyl. For structure of S.8 , see Figure .

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Figure 1.10.5 Structures of 7‐deazapurine 2′‐deoxyribonucleosides with interesting fluorescence properties.

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Literature Cited

Literature Cited
	Albert, A. and Serjeant, E.P. 1971. Determination of ionization constants by spectrometry. In The Determination of Ionization Constants, pp. 44‐64. Chapman & Hall, London.
	Balow, G., Mohan, V., Lesnik, E.A., Johnston, J.F., Monia, B.P., and Acevedo, O.L. 1998. Biophysical and antisense properties of oligodeoxynucleotides containing 7‐propynyl‐ , 7‐iodo‐ and 7‐cyano‐7‐deaza‐2‐amino‐2′‐deoxyadenosines. Nucl. Acids Res. 26:3350‐3357.
	Hoffer, M. 1960. α‐Thymidin. Chem. Ber. 93:2777‐2781.
	Latimer, L.J. and Lee, J.S. 1991. Ethidium bromide does not fluoresce when intercalated adjacent to 7‐deazaguanine in duplex DNA. J. Biol. Chem. 266:13849‐13851.
	LePecq, J.B. and Paoletti, C. 1967. A fluorescent complex between ethidium bromide and nucleic acids. Physical‐chemical characterization. J. Mol. Biol. 27:87‐106.
	Kasai, H., Ohashi, Z., Harada, F., Nishimura, S., Oppenheimer, N

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