Nucleic acid related compounds. 136. Synthesis of 2-amino- and 2,6-diaminopurine derivatives via inverse-electron-demand Diels-Alder reactions

Article abstract of DOI:10.1135/cccc20061029

Thermal inverse-electron-demand Diels-Alder reactions of 5-aminoimidazoles and 2,4,6-tris-(ethoxycarbonyl)-1,3,5-triazine (2) with spontaneous retro-Diels-Alder loss of ethyl cyanoformate and elimination of ammonia give 2,6-bis(ethoxycarbonyl)purines. A report that selective alkaline hydrolysis followed by acid-catalyzed decarboxylation gave 6-(ethoxycarbonyl)purine products was not in harmony with known reactions in purine chemistry. Our reinvestigation has shown that the 6-(ethoxycarbonyl) group undergoes preferential base-promoted hydrolysis, as expected, but regioselectivity for attack of hydroxide at the carbonyl group at C6 is not high (relative to hydrolysis of both C2 and C6 esters). The structure of 9-benzyl-2- (ethoxycarbonyl)purine was determined by X-ray crystallography and confirmed by Curtius rearrangement of the azidocarbonyl analogue to give 2-amino-6- benzylpurine. Acid-catalyzed decarboxylation of the 2,6-dicarboxylate formed during hydrolysis gave 9-benzylpurine, and Curtius rearrangement of 2,6-bis(azidocarbonyl)-9-benzylpurine gave 2,6-diamino-9-benzylpurine. Attempted applications of inverse-electron-demand Diels-Alder reactions of 2 with nucleoside derivatives were problematic.

Full text of DOI:10.1135/cccc20061029

Nucleic Acid Related Compounds
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NUCLEIC ACID RELATED COMPOUNDS. 136. SYNTHESIS OF
2-AMINO- AND 2,6-DIAMINOPURINE DERIVATIVES VIA
INVERSE-ELECTRON-DEMAND DIELS–ALDER REACTIONS+
1,
Xiaoyu LIN and Morris J. ROBINS *
Department of Chemistry and Biochemistry, Brigham Young University, Provo,
1
Utah 84602-5700, U.S.A.; e-mail: morris_robins@byu.edu
Received March 5, 2006
Accepted May 25, 2006
Dedicated to Professor Antonín Holý on his 70th birthday in appreciation of outstanding achievements
in the area of nucleic acid chemistry and a long and pleasant friendship.
Th erm al in verse-electron -dem an d Diels–Alder reaction s of 5-am in oim idazoles an d 2,4,6-tris-
(eth oxycarbon yl)-1,3,5-triazin e (2) with spon tan eous retro-Diels–Alder loss of eth yl
cyan oform ate an d elim in ation of am m on ia give 2,6-bis(eth oxycarbon yl)purin es. A report
th at selective alkalin e h ydrolysis followed by acid-catalyzed decarboxylation gave
6-(eth oxycarbon yl)purin e products was n ot in h arm on y with kn own reaction s in purin e
ch em istry. Our rein vestigation h as sh own th at th e 6-(eth oxycarbon yl) group un dergoes pref-
eren tial base-prom oted h ydrolysis, as expected, but regioselectivity for attack of h ydroxide at
th e carbon yl group at C6 is n ot h igh (relative to h ydrolysis of both C2 an d C6 esters). Th e
structure of 9-ben zyl-2-(eth oxycarbon yl)purin e was determ in ed by X-ray crystallograph y an d
con firm ed by Curtius rearran gem en t of th e azidocarbon yl an alogue to give 2-am in o-
6-ben zylpurin e. Acid-catalyzed decarboxylation of th e 2,6-dicarboxylate form ed durin g h y-
drolysis gave 9-ben zylpurin e, an d Curtius rearran gem en t of 2,6-bis(azidocarbon yl)-
9-ben zylpurin e gave 2,6-diam in o-9-ben zylpurin e. Attem pted application s of in verse-
electron -dem an d Diels–Alder reaction s of 2 with n ucleoside derivatives were problem atic.
Keyw ord s: Purin es; Curtius rearran gem en t; Diels–Alder in verse-electron -dem an d reaction s;
5-Am in oim idazoles; Triazin es; Nucleosides.
Holý an d coworkers2 h ave reported studies on organ om etallic cross-
couplin g reaction s of h alopurin e derivatives an d n ucleosides. Hocek an d
coworkers3 h ave m ade m ajor con tribution s in th is area, an d reviews4 are
available. We recen tly h ave developed oth er cross-couplin g m eth ods5 in -
+ Part 135 see ref.1
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© 2006 Institute of Organic Chemistry and Biochemistry
doi:10.1135/cccc20061029

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Lin, Robins:
cludin g reaction s with n on -con ven tion al leavin g groups5b,5c to produce
com poun ds with carbon -lin ked substituen ts on th e purin e rin g of
n ucleoside derivatives.
Several years ago we were in trigued by a com m un ication 6 th at described
th e preparation of 2- an d 6-(eth oxycarbon yl)-substituted purin e bases an d
n ucleosides by in verse-electron -dem an d Diels–Alder (DA) reaction s7. Th at
approach 6 em ployed cycloaddition of 2,4,6-tris(eth oxycarbon yl)-
1,3,5-triazin e (2) (Fig. 1) with electron -rich am in oim idazole derivatives
such as 5-am in o-1-ben zylim idazole (4) (gen erated in situ by decarboxyl-
ation of 5-am in o-1-ben zylim idazole-4-carboxylic acid). Th e in itially form ed
bicyclic adducts un dergo spon tan eous retro-DA loss of eth yl cyan oacetate
an d arom atization by loss of am m on ia7a,7b to give 2,6-bis(eth oxy-
carbon yl)purin es. Th e sam e process with 5-am in o-1-(β-D-ribofuran osyl)-
im idazole-4-carboxam ide was reported6 to give th e correspon din g purin e
n ucleoside, an d we en vision ed use of th is m eth odology for th e syn th esis of
sugar-m odified an alogues th at could be con jugated with am in o acids an d
peptides at C2 an d/or C6.
FIG. 1
Th e (in verse-electron -dem an d Diels–Alder cycloaddition , retro-Diels–Alder fragm en tation , loss
of am m on ia) reaction cascade
It was stated: “Selective fun ction alization of th e 2- or 6-eth oxycarbon yl
group can be readily ach ieved because of th e large differen ce in reactivity
between th e ester groups. For exam ple, treatm en t of 5 with on e equivalen t
of sodium h ydroxide followed by decarboxylation gave 9-ben zyl-6-eth oxy-
carbon ylpurin e (65%) with n o detectable 9-ben zyl-2-eth oxycarbon ylpurin e
or 9-ben zylpurin e”6. Th at structure assign m en t was based on an alogy with
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Nucleic Acid Related Compounds
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selective reduction of th e 2-(eth oxycarbon yl) group on th e m odel
6-am in o-2,4-bis(eth oxycarbon yl)-5-(m eth ylth io)pyrim idin e (A) (Fig. 2) with
sodium boroh ydride7a. Steric bias in opposition to reduction of th e
6-(eth oxycarbon yl) group adjacen t to th e 5-substituen t was in voked to ra-
tion alize selective reduction of th e 2-(eth oxycarbon yl) group. However,
steric effects of an en docyclic n itrogen (N7) in th e purin e rin g (a fused im i-
dazo[4,5-d]pyrim idin e) of 5 are n ot equivalen t to th ose of a 5-m eth yl-
sulfan yl group on th e m on ocyclic pyrim idin e rin g of A. Also th e
m eth ylsulfan yl substituen t m igh t exert sign ifican tly differen t electron ic
effects, an d reduction with NaBH4 is substan tially differen t th an h ydrolysis
with NaOH.
FIG. 2
6-Am in o-2,4-bis(eth oxycarbon yl)-5-m eth ylsulfan ylpyrim idin e (A);
2,6-Bis(acetam ido)-9-(2,3,5-tri-O-acetyl-β-D-ribofuran osyl)purin e (B);
2-(Acetam ido)-6-am in o-9-(β-D-ribofuran osyl)purin e (C)
Sch aeffer an d Th om as8 h ad sh own th at treatm en t of a 2,6-dich loropurin e
n ucleoside derivative with m eth an olic am m on ia gave 2-ch loroaden osin e by
displacem en t of ch loride from C6, an d th at treatm en t of a 2-ch loro-
6-m eth oxypurin e an alogue with am m on ia in m eth an ol at elevated tem per-
ature resulted in displacem en t of m eth oxide from C6 rath er th an ch loride
from C2. Num erous exam ples of regiospecific displacem en t of th e
6-substituen t from 2,6-disubstituted purin e derivatives9 (especially with two
iden tical groups) are kn own , an d treatm en t of 2,6-bis(acetam ido)-
9-(2,3,5-tri-O-acetyl-β-D-ribofuran osyl)purin e (B) (Fig. 2) with m eth an olic
am m on ia at 0 °C resulted in selective am m on olysis of th e acetam ido group
at C6 to give 2-acetam ido-6-am in o-9-(β-D-ribofuran osyl)purin e10 (C). Th us,
C6 is m ore reactive th an C2 with n ucleoph iles, an d th is effect also is ob-
served at th e carbon yl group of acetam ido substituen ts. We con sidered it
m uch m ore likely th at treatm en t of 9-ben zyl-2,6-bis(eth oxycarbon yl)purin e
(5) with on e equivalen t of h ydroxide un der con trolled con dition s would
produce 9-ben zyl-2-(eth oxycarbon yl)purin e-6-carboxylate rath er th an th e
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Lin, Robins:
9-ben zyl-6-(eth oxycarbon yl)purin e-2-carboxylate6 regioisom er. Th erefore,
th e m odel reaction s were rein vestigated prior to th e pursuit of an alogous
ch em istry with n ucleosides.
RESULTS AND DISCUSSION
Trim erization of eth yl cyan oform ate (1) (Sch em e 1) to give 2,4,6-tris-
(eth oxycarbon yl)-1,3,5-triazin e11 (2) h as been reported several tim es. We
foun d th e gen eral m eth od of Bozh kova an d Heim gartn er11b (with a rough ly
m easured addition of dry HCl gas) to be m ost reliable. Th e un stable
5-am in o-1-ben zylim idazole-5-carboxylic acid (3) was prepared by h ydroly-
sis12 of its eth yl ester (prepared by a two-step in situ rin g closure of eth yl
SCHEME 1
Syn th esis of substituted purin e derivatives
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Nucleic Acid Related Compounds
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2-am in o-2-cyan oacetate13 with trieth yl orth oform ate an d ben zylam in e14
)
followed by acidification . In situ decarboxylation of 3 in DMF at 80 °C pro-
d u ced th e u n stable 5-am in o-1-ben zylim id azole (4), an electronrich
dien eoph ile for in verse-electron -dem an d Diels–Alder cycloaddition 7 with
electron -deficien t dien e 2. Th e 9-ben zyl-2,6-bis(eth oxycarbon yl)purin e (5)
product was obtain ed in 76% yield wh en th e reported6 2:1 ratio of 3/2 was
used. Th e yield was dim in ish ed (67%) with a 1:1 ratio of th ese reactan ts,
but 2 was m ore readily accessible th an 3.
Careful treatm en t of 5 with on e equivalen t of sodium h ydroxide un der
th e reported6 con dition s (0 °C for 1 h an d 25 °C for 2 h ) gave a m ixture of
recovered startin g m aterial 5 (22%) an d th ree h ydrolysis products. Sodium
9-ben zyl-2-(eth oxycarbon yl)purin e-6-carboxylate (6a) was th e m ajor prod-
uct an d sodium 9-ben zyl-6-(eth oxycarbon yl)purin e-2-carboxylate (6b) was
th e decidedly m in or com pon en t. Disodium 9-ben zylpurin e-2,6-dicarb-
oxylate (6c) was produced in sign ifican t am oun ts (typical 1H NMR ratios
were 67:8:25 for 6a/6b/6c, respectively). Treatm en t of 5 un der m ore vigor-
ous con dition s (2 equivalen ts of NaOH, 25 °C for 3 h ) gave 6a/6b/6c ratios
of 20:4:76 (1H NMR). Un der a variety of con dition s, h ydrolysis of 5 was n ei-
th er regiospecific n or was 6b ever th e m ajor product. Mon oester 6a was th e
predom in an t isom er produced, as expected8–10 from oth er reaction s of
iden tically 2,6-disubstituted purin e com poun ds with n ucleoph ilic reagen ts,
but th e <3:1 selectivity for base-prom oted h ydrolysis of th e m ore suscepti-
ble C6 ester relative to h ydrolysis of both C2 an d C6 esters was low.
Also in con trast with th e earlier report6, th erm al decarboxylation of th e
m ixture from th e m ore selective h ydrolysis gave two isolated m aterials. Th e
m in or m aterial was 9-ben zylpurin e (10) an d th e m ajor (68% based on re-
covered startin g m aterial) was a m ixture of two com poun ds (1H NMR ratio
of ~10:1). Th e predom in an t isom er was 9-ben zyl-2-(eth oxycarbon yl)purin e
(11), wh ich was con firm ed by X-ray crystallograph y (Fig. 3) an d con version
in to 2-am in o-9-ben zylpurin e15 (14). Decarboxylation of th e disodium salt
6c gave 9-ben zylpurin e16 (10).
Treatm en t of purified 11 with h ydrazin e m on oh ydrate gave 9-ben zyl-
2-(h ydrazin ocarbon yl)purin e (12), wh ich un derwen t diazotization to give
9-ben zyl-2-(azidocarbon yl)purin e (13). A toluen e solution of 13 was h eated
at 110 °C to effect Curtius rearran gem en t, an d 2-am in o-9-ben zylpurin e15
(14) was obtain ed in 74% overall yield (two-stage con version from 12). We
th en probed th e poten tial for selective h ydrazin olysis of th e esters as a pos-
sible route to purin e derivatives with am in o an d carboxyl groups at C2
an d/or C6. However, solution s of 5 in DMF treated with various m olar ra-
tios of h ydrazin e m on oh ydrate at 0 °C sh owed slow an d com plex ch an ges,
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Lin, Robins:
wh ereas h eatin g such solution s resulted in disappearan ce of startin g 5 with -
out form ation of defin ed products. We foun d th at treatm en t of 5 with
N2H4·H2O/EtOH (3:10) at 22 °C for 20 m in gave a sin gle product (TLC). Be-
cause con cen tration of such solution s resulted in form ation of addition al
UV-active m aterials, H2O was added an d partially evaporated several tim es.
NaNO2 an d HCl/H2O were th en added to th e solution of in term ediate 7,
an d a toluen e solution of th e resultin g diazotization product 8 was h eated.
Th e Curtius rearran gem en t product, 2,6-diam in o-9-ben zylpurin e17 (9), was
isolated in 74% yield (overall for th ree steps from 5).
FIG. 3
X-ray crystal structure of 9-ben zyl-2-(eth oxycarbon yl)purin e (11) (CCDC 608646)
Attem pts to duplicate th e results an d yields reported6 for in verse-
electron -dem an d DA reaction s of 2 with 5-am in oim idazole riboside (de-
rived from base-prom oted h ydrolysis of 5-am in oim idazole-4-carboxam ide
riboside12 (15) (Sch em e 2) followed by th erm al decarboxylation un der
acidic con dition s) were problem atic. Both th e sodium an d potassium salts
of 5-am in oim idazole-4-carboxylate riboside (16) were prepared as described
by Robin s an d coworkers12. Subjection of eith er salt to variation s of th e re-
ported gen eral con dition s6 gave ≤30% yields of purified product 17 (con -
tam in ated with m in or am oun ts of an un kn own UV-active im purity after
ch rom atograph y).
We h ad prepared 5-am in o-3′,5-an h ydro-1-(β-D-xylofuran osyl)im idazole-
4-carboxam ide18 (20) by h eatin g aqueous solution s of 2′,3′-an h ydro-
aden osin e (18) (form ation of th e ch arged cyclon ucleoside an d rin g
open in g) followed by base-prom oted h ydrolysis of th e resultin g 5-carbox-
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Nucleic Acid Related Compounds
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SCHEME 2
Attem pted application s of th e reaction cascade with 2 an d 5-am in oim idazole-4-carboxylic acid
n ucleoside derivatives
12
am idin e 19. Vigorous h eatin g of 20 with KOH/H2O
gave th e potassium
carboxylate 21. Heatin g solution s of catalytically acidified 21 an d triazin e 2
un der various con dition s failed to produce 9-(3-am in o-3-deoxy-
β-D-xylofuran osyl)-2,6-bis(eth oxycarbon yl)purin e (22), wh ich m igh t h ave
been expected to result from an in situ reaction cascade (decarboxyl-
ation -in verse-electron -dem an d DA–retro-DA-elim in ation of am in o sugar)
an alogous to th e process with th e m odel am in oim idazole 4. It appears th at
in situ reaction cascades with 2 an d 5-am in oim idazole n ucleoside deriva-
tives are problem atic.
In con clusion , th e Boger reaction cascade7a,7b with 2,4,6-tris(eth oxy-
carbon yl)-1,3,5-triazin e (2) an d 5-am in o-1-ben zylim idazole (4) proceeds as
described6. However, th e con trolled, base-prom oted h ydrolysis of th e
resultin g 9-ben zyl-2,6-bis(eth oxycarbon yl)purin e (5) is <3:1 selective for
h ydrolysis of on e versus both ester groups, an d th e selectivity for
m on o-h ydrolysis of th e ester fun ction s at C6:C2 is ≥5:1. Th e m ajor product
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Lin, Robins:
of con trolled h ydrolysis followed by decarboxylation is 9-ben zyl-
2-(eth oxycarbon yl)purin e (11), an d n ot th e reported6 6-(eth oxycarbon yl)
regioisom er (th e derived carboxam ide6 m ust also be th e C2 isom er). On th e
basis of our in vestigation s, a m ore prom isin g route is required for th e prep-
aration of n ovel n ucleoside con jugates derived from rich ly fun ction alized
platform s such as structure 22. Oxidation of 2- an d/or 6-(h ydroxym eth yl)
substituen ts on purin e an d n ucleoside derivatives (readily accessible by th e
m eth odology of Hocek an d coworkers3b) to th e carboxylate level m igh t be a
plausible n ew approach .
EXPERIMENTAL
Gen eral
Un corrected m eltin g poin ts were determ in ed with a capillary tube apparatus. UV spectra (λ,
n m ; ε) were determ in ed with solution s in MeOH un less oth erwise in dicated. 1H NMR (300
MHz) an d 13C NMR (75 MHz) spectra (δ, ppm ; J, Hz) were determ in ed with solution s in
DMSO-d6 (with in tern al Me4Si) un less oth erwise in dicated. “Observed” 1H NMR m ultiplici-
ties are with in quotation m arks if m ore com plex splittin g pattern s are expected.
High -resolution m ass spectra (MS) were determ in ed un der FAB con dition s (with a glycerol or
th ioglycerol m atrix) un less oth erwise in dicated; m eth an e (CH4) was used for ch em ical ion -
ization (CI) m ass spectral determ in ation s. Reagen t grade ch em icals were used, an d solven ts
were distilled before use. CH2Cl2, CH3CN, an d toluen e were dried over an d distilled from
CaH2. Volatile m aterials were flash evaporated at <35 °C un der h ouse vacuum (~10 m m Hg)
or at m ech an ical oil-pum p vacuum (<0.1 m m Hg). “Diffusion crystallization ” was perform ed
as described19 with th e solven t com bin ation s in dicated.
2,4,6-Tris(eth oxycarbon yl)-1,3,5-triazin e (2)
Dry h ydrogen ch loride gas was bubbled (15–30 bubbles/s) for 20 m in in to eth yl cyan o-
form ate (0.5 m l, 0.50 g, 5 m m ol) at 0 °C, an d th e solution was stored at am bien t tem pera-
ture for 6 days. Th e resultin g crystals were filtered, wash ed with Et2O, an d recrystallized
from EtOH to give 2 (438.5 m g, 88%) as a white solid; m .p. 170–172 °C (lit.11a m .p. 170–171 °C).
1H NMR (300 MHz, CDCl3): 4.56 (q, J = 7.2, 2 H, CH2); 1.46 (t, J = 7.2, 3 H, CH3). 13C NMR
(75 MHz, CDCl3): 167.1 (C=O), 161.4 (arom atic), 64.2 (CH2), 14.2 (CH3). MS (CI): m/z
298.1044 (MH+ [C12H16N3O6] = 298.1039).
5-Am in o-1-ben zylim idazole-4-carboxylic Acid (3)
Saturated NaHCO3/H2O (30 m l) was added with vigorous stirrin g to a suspen sion of eth yl
cyan oglyoxylate oxim e (10 g, 0.07 m ol) in water (60 m l) un der a n itrogen atm osph ere. Th e
resultin g clear yellow solution was treated with 2-g portion s of solid sodium dith ion ite (34 g,
0.21 m ol) at a rate th at gave con trolled effervescen ce, an d stirrin g was con tin ued at am bien t
tem perature for an addition al 30 m in . Th e solution was th en extracted (CHCl3, 3 × 50 m l)
an d dried (Na2SO4). Volatiles were evaporated to give eth yl 2-am in o-2-cyan oacetate13 (6.13 g,
68%) as an oil, wh ich m ust be used im m ediately in th e n ext step.
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Nucleic Acid Related Compounds
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Trieth yl orth oform ate (1 m l, 0.89 g, 6.0 m m ol) was added dropwise to a stirred solution
of eth yl 2-am in o-2-cyan oacetate (426 m g, 3.33 m m ol) in CH3CN (4 m l, fresh ly distilled),
an d th e stirred solution was h eated at reflux for 50 m in an d th en cooled to am bien t tem pera-
ture. Ben zylam in e (0.50 m l, 0.49 g, 4.58 m m ol) was added, an d th e red solution was stored
at am bien t tem perature for 16 h . Volatiles were evaporated, an d th e residue was
ch rom atograph ed (0.5 to 0.8% MeOH/CH2Cl2) to give eth yl 5-am in o-1-ben zyl-
im idazole-4-carboxylate14 (364 m g, 45%) as a yellow solid. Recrystallization of th is m aterial
(EtOH) gave lon g n eedles; m .p. 167–170 °C (lit.14 m .p. 156 °C). UV: λm ax 269 (13 800), λm in
225 (6140). 1H NMR (300 MHz, CDCl3): 7.40–7.28 (m , 3 H, Ph ): 7.23–7.13 (m , 3 H, Ph an d
H2); 5.05 (br s, 4 H, CH2Ph an d NH2); 4.31 (q, J = 7.2, 2 H, CH2); 1.35 (t, J = 7.2, 3 H, CH3).
13C NMR (75 MHz, CDCl3): 164.4 (CO), 145.2 (C4), 134.3, 129.5, 128.8, 127.2 (Ph ), 131.7
(C2), 111.8 (C5), 60.1 (OCH2), 45.0 (CH2Ph), 14.8 (CH3). MS: m/z 246.1251 (MH+ [C13H16N3O2] =
246.1243).
A suspen sion of eth yl 5-am in o-1-ben zylim idazole-4-carboxylate (478 m g, 1.95 m m ol) in
EtOH (20 m l) an d 0.5 M NaOH/H2O (20 m l) was refluxed for 5 h . Volatiles were evaporated,
H2O (5 m l) was added, an d 5% HCl/H2O was added to pH ~5. Th e off-wh ite solid th at pre-
cipitated was filtered an d wash ed with on e portion of H2O. Th is m aterial was dried un der
vacuum at am bien t tem perature for 30 m in to give 3 (quan titative), wh ich was used im m e-
diately in th e n ext step.
9-Ben zyl-2,6-bis(eth oxycarbon yl)purin e (5)
A m ixture of 2 (438 m g, 1.47 m m ol) an d 3 (664 m g, 3.05 m m ol) in DMF (7.9 m l) was
h eated at 80 °C for 2 h un der a n itrogen atm osph ere, an d th en cooled to am bien t tem pera-
ture. Volatiles were evaporated, CH2Cl2 (10 m l) was added, an d th e suspen sion was filtered.
Th e filtrate was con cen trated an d subjected to flash ch rom atograph y (EtOAc/h exan es, 1:1)
to give 5 (398 m g, 76%) as an off-wh ite solid. Diffusion crystallization 19 (EtOAc/h exan es)
gave m aterial with m .p. 145–146.5 °C (lit.6 143–144 °C). UV: λm ax 287 (8660), λm in 253
(4060). 1H NMR (300 MHz, CDCl3): 8.43 (s, 1 H, H8); 7.42–7.30 (m , 5 H, Ph ); 5.60 (s, 2 H,
CH2Ph ); 4.66–4.52 (m , 4 H, 2 × CH2); 1.53–1.46 (m , 6 H, 2 × CH3). 13C NMR (75 MHz,
CDCl3): 163.5 (6-CO), 163.2 (2-CO), 154.5 (C6), 150.8 (C4), 149.6 (C2), 146.0 (C8), 134.2
(Ph ), 132.7 (C5), 129.5, 129.2, 128.5 (Ph ), 63.2 (OCH2), 48.2 (CH2Ph ), 14.4 (CH3). MS: m/z
377.1235 (MNa+ [C18H18N4O4Na] = 377.1225).
9-Ben zyl-2-(eth oxycarbon yl)purin e (11)
A solution of 5 (198 m g, 0.559 m m ol) in THF/EtOH/H2O (3:2:1, 7.6 m l) was cooled to 0 °C,
treated with 1 M NaOH/H2O (0.56 m l, 0.56 m m ol) un der a n itrogen atm osph ere, an d stirred
at 0 °C for 1 h an d th en at 25 °C for 2 h (as described6). Volatiles were evaporated, an d th e
residue was partition ed (EtOAc/H2O). Th e aqueous layer was extracted with EtOAc un til n o
furth er UV active m aterial was rem oved. Evaporation of th e com bin ed organ ic ph ase gave
recovered 5 (43.3 m g, 22%). Evaporation of volatiles from th e aqueous layer gave a m ixture
of sodium 9-ben zyl-2-(eth oxycarbon yl)purin e-6-carboxylate (6a), sodium 9-ben zyl-6-(eth oxy-
carbon yl)purin e-2-carboxylate (6b ), an d disodium 9-ben zylpurin e-2,6-dicarboxylate (6c).
Th is m ixtu re was treated with Ac2O (2.3 m l, 2.5 g, 24 m m ol) an d AcOH (2.3 m l, 2.4 g,
40 m m ol) an d h eated at 130 °C for 20 h . Volatiles were evaporated, th e residue was parti-
tion ed between saturated NaHCO3/H2O (25 m l) an d CH2Cl2 (30 m l), an d th e aqueous layer
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Lin, Robins:
was extracted with CH2Cl2 (3 × 25 m l). Th e com bin ed organ ic ph ase was dried (Na2SO4),
an d volatiles were evaporated. Th e residue was ch rom atograph ed (1% EtOH/CH2Cl2) to give
con tam in ated 11 (84 m g, 68% based on recovered startin g m aterial) an d 9-ben zylpurin e
(10; 13 m g, 14% based on recovered m aterial) as wh ite solids. A sam ple of 11 was purified
by diffusion crystallization 19 (EtOAc/h exan es) to give crystals; m .p. 89–91 °C (lit.6 m .p.
89–90 °C). UV: λm ax 269 (9850), λm in 241 (3820). 1H NMR (300 MHz, CDCl3): 9.36 (s, 1 H,
H6); 8.60 (s, 1 H, H8); 7.39 (s, 5 H, Ph ); 5.62 (s, 2 H, CH2Ph ); 4.59 (q, J = 7.2, 2 H, CH2);
1.51 (t, J = 7.2, 3 H, CH3). 13C NMR (75 MHz, CDCl3): 163.5 (CO), 151.7, 151.1, 148.4,
147.6, 134.2, 133.1, 129.6, 129.3, 128.7, 63.2, 48.2. MS: m/z 283.1185 (MH+ [C15H14N4O2] =
283.1195). Th e NMR spectra of 10 were iden tical to th ose of th e sam ple prepared in th e fol-
lowin g experim en t.
9-Ben zylpurin e (10)
A solution of 5 (202 m g, 0.570 m m ol) in THF/EtOH/H2O (3:2:1, 5 m l) was treated with 0.1
M
NaOH/H2O (12 m l, 1.2 m m ol) un der an argon atm osph ere at 26 °C for 3 h . Volatiles were
evaporated, an d th e residue was treated with Ac2O (7.5 m l, 8.12 g, 80 m m ol) an d AcOH
(7.5 m l, 7.87 g, 0.131 m m ol) at 130 °C for 4 days un der an argon atm osph ere. Th e reaction
m ixture was th en cooled, volatiles were evaporated, an d th e residue was partition ed between
CH2Cl2 an d saturated NaHCO3/H2O. Th e aqueous ph ase was extracted (CH2Cl2, 3×), an d th e
com bin ed organ ic ph ase was dried (Na2SO4). Volatiles were evaporated, an d th e residue was
ch rom atograph ed (1% MeOH/CH2Cl2) to give 10 (81 m g, 68%) as an off-wh ite solid.
Recrystallization (Et2O/h exan es) gave 10; m .p. 97–99 °C (lit.14 m .p. 100–101 °C). UV
(MeOH): λm ax 264 (9900). 1H NMR (300 MHz, CDCl3): 9.19 (s, 1 H, H6); 9.07 (s, 1 H, H2);
8.16 (s, 1 H, H8); 7.42–7.20 (m , 5 H, Ph ); 5.49 (s, 2 H, CH2). 13C NMR (75 MHz, CDCl3):
152.0 (C4), 147.5 (C2), 146.1 (C8), 134.8 (C6), 134.0 (C5), 129.5, 129.0, 128.2 (Ph ), 47.6 (CH2).
MS: m/z 211.0989 (MH+ [C12H11N14] = 211.0983).
9-Ben zyl-2-(h ydrazin ocarbon yl)purin e (12)
A yellow solution of 11 (13.7 m g, 0.05 m m ol) in h ydrazin e m on oh ydrate (0.30 m l, 0.41 g,
8.2 m m ol) was stirred at 30 °C for 20 m in . Volatiles were evaporated, an d th e residue was
adsorbed on a sm all am oun t (~1 m l) of silica gel th at was added to a sm all colum n of silica
gel. Ch rom atograph y (2 to 4% MeOH/CH2Cl2) gave 12 (11.4 m g, 88%) as an off-wh ite solid;
m .p. 143–146 °C. UV: λm ax 273 (10 700), λm in 242 (6080). 1H NMR: 10.11 (br s, 1 H, NHCO);
9.24 (s, 1 H, H6); 8.87 (s, 1 H, H8); 7.42–7.22 (m , 5 H, Ph ); 5.59 (s, 2 H, CH2); 4.64 (br s, 2 H,
NH2). 13C NMR: 161.7 (CO), 152.0 (C2), 151.4 (C4), 148.7 (C6), 147.6 (C8), 136.4 (C5),
133.9, 128.8, 128.0, 127.7 (Ph ), 46.2 (CH2). MS (EI): m/z 268.1056 (MH+ [C13H12N6O] =
268.1073).
2-Azidocarbon yl-9-ben zylpurin e (13)
Et2O (6 m l) was layered over a solution of 12 (56.9 m g, 0.21 m m ol) in 5 M HCl/H2O (5.7 m l),
an d th e biph asic m ixture was stirred an d cooled at 0–2 °C. A solution of NaNO2 (136 m g,
1.97 m m ol) in H2O (0.6 m l) was added dropwise to th e cold m ixture un der a n itrogen atm o-
sph ere. Th e eth er layer was rem oved after 20 m in , an d Et2O/CH2Cl2 (1:1, 6 m l) was added.
Th e organ ic layer was rem oved every 20 m in an d fresh Et2O/CH2Cl2 was added to extract
th e product from th e acidic aqueous solution . After 2 h , an oth er portion of NaNO2/H2O (50 m g
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Nucleic Acid Related Compounds
1039
in 0.3 m l) was added to th e m ixture, an d stirrin g was con tin ued un til n o addition al
UV-active product was extracted in to th e organ ic layer. Th e com bin ed organ ic ph ase was
con cen trated, wash ed on ce with H2O, dried (MgSO4) for 20 h , an d filtered. Th e filtrate was
con cen trated, toluen e was added an d evaporated (2 × 10 m l) to ~10 m l, an d th is solution
was used im m ediately for th e n ext step.
2-Am in o-9-ben zylpurin e (14)
Triflu oroacetic acid (0.30 m l, 0.44 g, 3.9 m m ol) was added to a stirred solu tion of 13
(~59 m g) in toluen e (~10 m l), an d stirrin g was con tin ued at 110 °C (oil bath tem perature)
for 48 h un der a n itrogen atm osph ere. Volatiles were evaporated, an d th e residue was
ch rom atograph ed (3% MeOH/CH2Cl2) to give 14 (35.5 m g, 74%, based on startin g 12) as an
off-wh ite solid. Th is m aterial was h eated gen tly in EtOAc/MeOH (10:1) an d filtered to give
14; m .p. 179–180 °C (lit.15 180–182 °C). UV: λm ax 246, 310 (5100, 7400), λm in 238, 266
(4700, 840). 1H NMR: 8.60 (s, 1 H, H6); 8.16 (s, 1 H, H8); 7.40–7.20 (m , 5 H, Ph ); 6.54 (br s,
2 H, NH2); 5.29 (s, 2 H, CH2). 13C NMR: 160.7 (C2), 153.0 (C4), 149.2 (C6), 142.7 (C8),
137.0, 128.7, 127.7, 127.1 (Ph ), 126.7 (C5), 45.4 (CH2). MS: m/z 226.1082 (MH+ [C12H12N5]
= 226.1092).
9-Ben zyl-2,6-bis(h ydrazin ocarbon yl)purin e (7)
Hydrazin e m on oh ydrate (0.40 m l, 0.412 g, 8.0 m m ol) was added dropwise to a stirred sus-
pen sion of 5 (86.2 m g, 0.24 m m ol) in EtOH (4.5 m l) at am bien t tem perature. A clear yellow
solution form ed, an d stirrin g was con tin ued for an addition al 20 m in (TLC sh owed com plete
reaction of 5). H2O was added an d th e solution was con cen trated (2 × 10 m l) to ~20 m l. Th is
solution was used im m ediately for th e n ext step.
2,6-Bis(azidocarbon yl)-9-ben zylpurin e (8)
Et2O (20 m l) an d HCl/H2O (37%, 10 m l) were added dropwise to a stirred aqueous solution
of 7 (~79 m g, 0.24 m m ol) in H2O (20 m l) at 0 °C. A solution of NaNO2 (563 m g, 8.10
m m ol) in H2O (2 m l) was carefully added to th e cooled biph asic m ixture at a rate th at kept
effervescen ce un der con trol, an d stirrin g at 0 °C was con tin ued. Th e Et2O layer was rem oved
after 20 m in , an d Et2O/CH2Cl2 (1:1, 20 m l) was added. Con tin uous extraction of th e prod-
uct form ed in th e aqueous ph ase was effected by rem oval of th e organ ic layer an d addition
of fresh solven ts every 20 m in . Solution s of NaNO2 (500 m g) in H2O (2 m l) were added after
two an d four h ours, an d stirrin g was con tin ued un til n o furth er UV-active product was de-
tected (TLC) in th e aqueous layer. Th e com bin ed organ ic ph ase was con cen trated, wash ed
on ce with H2O/saturated aqueous NaHCO3 (10:1), dried (MgSO4), an d filtered. Toluen e (10 m l)
was added to th e filtrate, an d th e solution was con cen trated. Toluen e (10 m l) was again
added, an d th e solution of 8 was con cen trated an d used im m ediately in th e n ext step.
2,6-Diam in o-9-ben zylpurin e (9)
Trifluoroacetic acid (0.20 m l, 0.30 g, 2.6 m m ol) was added to a solution of 8 (~85 m g, 0.24
m m ol) in toluen e, an d th e stirred solution was h eated at 100–110 °C (oil bath tem perature)
for 24 h . Volatiles were evaporated, an d th e residue was adsorbed on a sm all am oun t of silica
gel an d added to a sm all colum n of silica gel. Ch rom atograph y (2 to 3% MeOH/CH2Cl2)
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1040
Lin, Robins:
gave 9 (43.1 m g, 74%) as an off-wh ite solid th at was recrystallized (H2O) to give colorless
n eedles; m .p. 178–180 °C (lit.17 m .p. 180–181 °C). UV: λm ax 256, 281 (8300, 10 400), λm in
237, 265 (5090, 6940). 1H NMR: 7.81 (s, 1 H, H8); 7.38–7.18 (m , 5 H, Ph ); 6.84 (br s, 2 H,
6-NH2); 5.91 (br s, 2 H, 2-NH2); 5.19 (s, 2 H, CH2). 13C NMR: 157.8 (C2), 154.2 (C6), 151.7
(C4), 138.8 (C8), 137.3, 128.7, 127.6, 127.1 (Ph ), 112.4 (C5), 45.7 (CH2). MS (EI): m/z
240.1130 (MH+ [C12H12N6] = 240.1124).
X-ray Crystal Data for 11
Colorless crystals; form ula: C15H14N4O2 (FW: 282.30); m on oclin ic; space group: P2(1)/n; un it
cell dim en sion s: a = 9.7068(6) Å, b = 9.8131(6) Å, c = 15.7115(9) Å, α = 90°, β
106.9370(10)°, γ = 90°; Z = 4; R = 0.0735; GOF = 1.040.
=
CCDC 608646 con tain s th e supplem en tary crystallograph ic data for th is paper. Th ese data
can be obtain ed free of ch arge via www.ccdc.cam .ac.uk/con ts/retrievin g.h tm l (or from th e
Cam bridge Crystallograph ic Data Cen tre, 12, Un ion Road, Cam bridge, CB2 1EZ, UK; fax:
+44 1223 336033; or deposit@ccdc.cam .ac.uk).
We thank Professors N. K. Dalley and J. F. Cannon for determination of the X-ray crystal structure
of 11, and gratefully acknowledge unrestricted gift funds from pharmaceutical companies (M. J. Robins)
and a R. K. Robins Graduate Research Fellowship (X. Lin) from Brigham Young University.
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