First synthesis of 2'-deoxyfluoropuromycin analogues: Experimental insight into the mechanism of the staudinger reaction

DOI: 10.1002/chem.200700058

First Synthesis of 2’-Deoxyfluoropuromycin Analogues: Experimental Insight

into the Mechanism of the Staudinger Reaction

Adib Charafeddine, Wissam Dayoub, Hubert Chapuis, and Peter Strazewski*^[a]

Abstract: The N⁶,N⁶-dedimethyl-2’-de-

yl azide and an active ester of the N-

protected amino acid using highly effi-

cient solution conditions for the Stau-

dinger–Vilarrasa coupling, to obtain

the corresponding carboxamide directly

from the in situ formed iminophos-

phorane. This coupling reaction fur-

nished sterically quite demanding

amides in 94% isolated yields under

very mild conditions and should there-

fore be of a more general value. Under

certain reaction conditions we isolated

(amino)acyltriazene derivatives from

which dinitrogen was not eliminated.

These secondary products are trapped

and stabilized witnesses of the first in-

termediate of the Staudinger reaction,

the phosphatriazenes (phosphazides,

triazaphosphadienes) which usually

eliminate dinitrogen in situ and rapidly

rearrange into iminophosphoranes,

unless they are derived from conjugat-

ed or sterically bulky azides and phos-

phines. The acyltriazenes could either

be thermally decomposed or converted

to the corresponding N-alkyl carboxa-

mides through proton-assisted elimina-

tion of dinitrogen. All compounds were

carefully characterized through MS

oxyfluoro analogue of puromycin

(=

methyltyrosylamido]adenosine),

2’,3’-regioisomer and a 3’-cytidyl-5’-(2’-

deoxyfluoro)puromycyl dinucleotide

3’-deoxy-N⁶,N⁶-dimethyl-3’-[O-

its

analogue were synthesized following an

approach involving i) the diastereospe-

cific nitrite-assisted formation of a lyxo

nucleosidic 2’,3’-epoxide from an ade-

nosine-2’,3’-ditriflate derivative in a bi-

phasic solvent mixture; ii) the regio-

and stereoselective epoxide ring open-

ing with sodium azide under mildly

acidic aqueous conditions, iii) the ste-

reospecific introduction of the fluor

atom using DAST and iv) the reaction

between the nucleosidyl or dinucleotid-

Keywords: antibiotics · antitumor

agents

· azides · nucleotides ·

1

spectrometry, H, ¹⁹F, ³¹P and ¹³C NMR

Staudinger Vilarrasa · triazenes

spectroscopy.

Introduction

ysis of the ester bond linking the peptide to the 3’-terminal

oxygen atom of the P-site bound tRNA by the a-amino

group of the A-site bound aminoacyl-tRNA (the a-amino

nucleophile attacks one face of the planar ester group). This

attack was thought to yield an oxyanion containing a tetra-

hedral carbon intermediate which would resolve to the

products,^[1]a deacylated P-site bound tRNA and an A-site

bound peptidyl-tRNA with a peptide chain elongated by

one amino acid. After translocation of the elongated peptid-

yl-tRNA into the P-site with the help of a translocation fac-

tor·GDP complex, new aminoacyl-tRNAs—each in conjunc-

tion with an elongation factor·GTP complex—can test the

new codon presented at the decoding site. The cognate ami-

noacyl-tRNA is detached from the elongation factor

through GTP-to-GDP hydrolysis and processed, thus, pro-

tein synthesis continues.

The ribosome is the macromolecular machine responsible

for protein synthesis in all cells. The peptidyl transferase

active site of the ribosome has two substrate binding sites

and utilizes two tRNAs as the reaction substrates: an amino-

acyl-tRNA bound to the A-site and a peptidyl-tRNA bound

to the P-site. The peptidyl transfer reaction involves aminol-

[a] A. Charafeddine, Dr. W. Dayoub, Dr. H. Chapuis,

Prof. Dr. P. Strazewski

Laboratoire de Synthse des BiomolØcules

Bâtiment Eugne Chevreul (5ime Øtage)

UMR UCBL-CNRS 5246, ICBMS

UniversitØ Claude Bernard Lyon 1

Domaine Scientifique de la Doua, 43bvd du 11 novembre 1918

69622 Villeurbanne Cedex (France)

Fax : (+33)472-431-323

The tRNA substrates are aligned in the active site by base

pairing between the CCA sequence at the 3’-end of each

tRNA and complementary sequences within the A and P-

sites of the rRNA that make up the peptidyl transferase

center.^[2]The ribosome can also catalyze amide-bond forma-

E-mail: strazewski@univ-lyon1.fr

Supporting information for this article is available on the WWW

under http://www.chemeurj.org/ or from the corresponding authorꢀs

website: All NMR spectra including correlation spectra and signal as-

signments

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Chem. Eur. J. 2007, 13, 5566 – 5584

FULL PAPER

tion using the minimal A-site

substrate puromycin, a nucleo-

side antibiotic comprised of

3’-amino-3’-deoxy-N⁶,N⁶-dimeth-

yladenosine coupled to the

amino acid O-methyl-l-tyro-

sine (3’-deoxy-N⁶,N⁶-dimethyl-

3’-[O-methyltyrosylamido]ade-

nosine, 1).^[3]The antitumor an-

tibiotic puromycin inhibits pro-

tein synthesis as a consequence

of its striking resemblance to

the aminoacyladenyl 3’-termi-

nus of aminoacyl-tRNA. It has

Figure 1. Natural puromycin and its 2’-deoxyfluorinated analogues.

been demonstrated that puro-

mycin competes with aminoacyl-tRNA at the A site and

subsequently interacts with the peptidyl-tRNA at the P site

causing premature release of the polypeptide chains from

the ribosome.^[4]For this reason, puromycin has long since

been used in the investigation of the peptidyl transfer reac-

tion.^[5]Today puromycin is broadly used as a tool for molec-

ular biologists.

synthesis, and kinetic parameters for the interaction of these

analogues with procaryotic P-site substrates will be reported

in the future.

In the puromycin analogues 2 and 3 the 2’-hydroxyl group

of both natural puromycin and the peptide accepting 3’-ter-

minal A76 residue of aminoacyl-tRNA, a function known to

serve as a hydrogen acceptor of the imino hydrogen atom of

the ribosomal A-site uridylate residue U2620,^[10]is replaced

by a close to isosteric and perhaps weakly hydrogen bonding

fluor atom.^[11]The 2’-fluorine atom allows for a qualitatively

undisturbed peptidyl transfer when present at the 3’-termi-

nal adenylate of an A site-bound aminoacyl-tRNA.^[12]Its

high electronegativity may have a favorable impact on the

ribofuranose pucker, since 2’-F is likely to force, through the

stereoelectronic gauche effect stronger than the one of 2’-

OH, a more defined orientation of the vicinal 3’-substituent

bearing the reactive nucleophile (Figure 1), the more so, as

its a-amino group accumulates a partial positive charge

during peptidyl transfer before it deprotonates, which may

to some extent deplete electron density on N3’ in the first

transition state and thus further reinforce the stereoelec-

tronic gauche effect.^[13]

The catalytic mechanism of the peptidyl transfer reaction

is an area of ongoing research interest both for its medicinal

and for its evolutionary relevance. Evolutionarily, RNA cat-

alysis of peptide-bond formation is the reaction required to

bridge between a world dominated by RNA catalysts and

the one of modern cells dominated by protein catalysts.^[6]

Medicinally, the peptidyl transferase center with its sur-

rounding substrate and product channels is the binding site

for several naturally and synthetically derived antibiotics.^[7,8]

Microbial resistance to antibiotics is growing and spread-

ing rapidly. Thus, new approaches for the development of

novel antibiotics are clearly needed and, since about one

half of the currently used antibiotics target the ribosome—

mostly the large subunit—structure-based drug design using

the large subunit structure should be useful. The use of the

structural information has greatly increased the speed with

which new potential drug candidates can be developed. Fur-

thermore, structural variation of puromycin to obtain more

active analogues may be extremely useful in elucidating var-

ious aspects of protein synthesis. For this purpose, one of

our priorities is to synthesize new and powerful antibiotics

that will be capable of supplementing or replacing those an-

tibiotics that become ineffective due to bacterial resistance

mutations.

Numerous antibiotics have been shown to bind to bacteri-

al and/or eucaryotic ribosomes and inhibit protein synthesis

but only puromycin is a pure A-site binder that inhibits pro-

tein synthesis irreversibly and produces truncated, dysfunc-

tional bacterial C-terminal puromycyl peptides which are

likely to boost an immune response in the infected host or-

ganism. Therefore a comparison of puromycin analogues 2

and 3 (Figure 1) and other analogues^[9]with natural puromy-

cin in assays on bacterial ribosomes will be carried out. Spe-

cifically, the ability of the puromycin analogues to serve as

peptide acceptors during peptidyl transfer, to inhibit protein

Puromycin is known to carry out a much faster peptidyl

transfer reaction when it is present as a dinucleotide where

the 5’-terminus is a cytidylate that mimics C75 of a transfer

RNA.^[14]The cytidine residue in 3 is expected to stabilize

the analogueꢀs interaction with the ribosome through base

pairing with the complementary A site residue G2588, as is

the case for C75 of aminoacyl-tRNA.^[10]

The new fluorinated puromycin analogues

2 and 3

(Figure 1) are synthesized in seven and thirteen steps, re-

spectively, from adenosine (4). The synthetic route for the

preparation of 2 and 3 is outlined in Schemes 1 and 3.

Results

The synthetic pathway (Scheme 1) is evident and conceptu-

ally similar to published procedures^[15a,b]but each step was

re-evaluated and changed so as to render the synthesis more

efficient and practical. It begins with the synthesis of 5, a

protected form of adenosine (4).

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5567

P. Strazewski et al.

The tritylation of 4 furnished at most 34% of the desired

ditritylated compound 5.^[15c]The first challenge was to effi-

ciently obtain diastereoisomerically pure lyxo-epoxide 7

(Scheme 1) from an activated form of diol 5,^[15a]best

through an efficient synthesis of ditriflate 6a.^[15d]Our first

attempts with the highly reactive triflic chloride and 4-dime-

thylaminopyridine (DMAP)—found to be indispensa-

ble^[15e]—produced in significant yields chlorinated (6b),

eliminated (6c) and monotriflated (6d) side products

1

(Scheme 2) which were identified by H and ¹⁹F NMR spec-

tra (cf. Supporting Information),^[16]as well as through ESI

MS spectra (not shown). Triflic anhydride and DMAP pro-

duced 6a in 83.7% yield along with 15% isolated 6d that

were triflated to 6a in a subsequent batch. Both 6a and 6d

proved quite stable and could be readily worked up and pu-

rified by chromatography over silica gel without detectable

elimination or hydrolysis.

In our hands, rather than from a 2’,3’-O,O-dimesylate and

sodium hydroxide in ethanol,^[15a,b,17]lyxo-epoxide 7 was

more reliably and cleanly obtained from ditriflate 6a using

tetrabutylammonium nitrite in a biphasic water/toluene mix-

ture.^[15d]The initially formed mononitrite–monotriflic diester

intermediate, where the nitrite bearing carbon configuration

had been inversed (major regioisomer: probably 3’-O-ni-

trite-2’-O-triflate),^[16]hydrolyzed to the corresponding

Scheme 2. Reactivity difference between TfCl and Tf₂O in the presence

of DMAP.

served.^[15a]Higher regioselectivities were obtained with

mono

A

sodium azide in refluxing acetone, but only after a much

prolonged reaction time (20 h).^[17]We obtained the desired

regioisomer 8a in higher yields after only 60–90 minutes re-

action time using soluble sodium azide in aqueous DMF in

the presence of ammonium chloride.^[18]The combined isolat-

ed yield for 8a and 8b was 93–96% and the ring opening se-

lectivity better than 4:1 in favor of the desired regioisomer.

The isomers could be readily

reacting to the doubly inverted vicinal dinitrite—and subse-

quently ring-closed in situ to 7.

Epoxides are most often opened by nucleophiles such as

the azide anion under anhydrous neutral or slightly basic

conditions using well soluble lithium azide in anhydrous

DMF, but rather weak regioselectivities have been ob-

separated through chromatog-

raphy over silica gel and con-

verted using DAST (Et₂NSF₃)

to the corresponding deoxy-

fluoro derivates 9a and 9b, re-

spectively, under inversion of

the configuration of the alco-

hol function.^[19]The best isolat-

ed yields were obtained when

the formation of the 2’-O-

SF₂NEt₂intermediate at ambi-

ent temperature clearly pre-

ceded the nucleophilic substi-

tution by F^ꢀat 808C.

We attached the amino acid

to the nucleoside by means of

a direct coupling of azide 9a

to an OBt active ester of N-

Boc-O-methyltyrosine in 94%

yield under the recently pub-

Scheme 1. Synthesis of 2’,3’-dideoxy-2’-fluoro-3’-[O-methyltyrosylamido]adenosine (2). a) Tr-Cl, DMAP, Py,

808C/4 h 40 min; b) Tf₂O, Py, DMAP, 08C/30 min, then RT/3 h; c) Bu₄N⁺NO₂^ꢀ, MePh/H₂O, RT/40 h; d) NaN₃,

NH₄Cl, DMF/H₂O, reflux (1008C)/1 h 30 min; silica gel chromatography; e) DAST, Py, Tol, RT/30 min, then

lished

Staudinger–Vilarrasa

808C/45 min; f) Boc-l-Tyr(Me)-OH, HOBt, DIC, P(nBu)₃, 08C/30 min, then RT/overnight; g) CF₃COOH,

A

coupling conditions as devel-

oped for the synthesis of non-

fluorinated puromycin ana-

CH₂ClCH₂Cl, RT/4 h. * This combined yield was obtained when monotriflate 6d (see text, Experimental Sec-

tion and Supporting Information) was reused for the synthesis of 6a. ** Yield with respect to consumed 8a

(=applied ꢀ recovered).

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2’-Deoxyfluoropuromycin Analogues

logues.^[9]Compound 10a was

FULL PAPER

deprotected

with

TFA

(CF₃COOH)^[15c]in dichloro-

ethane to give 2 as the TFA

salt in 10.5% overall yield

from adenosine (4).

The isolation of the pure

and highly water soluble target

compound 2·TFA necessitates

the following procedure. The

deprotection reaction must be

quenched with an excess meth-

anol to avoid partial retrityla-

tion. The resulting mixture is

concentrated under reduced

pressure to a volume that can

be directly applied on a silica

gel column. Chromatography

yields the target compound as

a mixture of a-amine and a-

ammonium salt as determined

1

by H-decoupled ¹⁹F NMR (in-

tegral ratio between the signals

at ꢀ76.9 ppm for TFA^ꢀand

ꢀ196.5 ppm for 2’-F of com-

pound 2 ꢁ1.34:1). To enhance

the solubility in water and to

Scheme 3. Synthesis

a) CF₃COOH, CH₂ClCH₂Cl, RT/2 h; b) N,N-di

ethylthiotetrazole,

N⁴-acetyl-5’-O-dimethoxytrityl-2’-O-triisopropylsilyloxymethylcytid-3’-yl-(2-cyanoethyl)-

N,N-diisopropylphosphoramidite, dry CH₃CN, RT/20 min, 2) 0.2m I₂/THF/pyridine/H₂O, RT; d) Fmoc-l-

Tyr(Me)-OH, HOBt, DIC, P(nBu)₃, 08C/30 min, then RT/overnight; e) 1) 33% CH₃NH₂/EtOH, RT/35 min, 2)

TBAF, THF, RT/40 min; f) 80% AcOH/H₂O, RT/20 min.

of

cytid-3’-yl-5’-[2’,3’-dideoxy-2’-fluoro-3’-(O-methyltyrosyl)amido]adenylate

(3).

AHCTREUNG

isolate

a

homogenous salt

AHCTREUNG

form of the target compound,

this solution (about 10 mm)

was carefully acidified with

more TFA until ¹⁹F NMR showed complete formation of the

2·TFA salt (¹⁹F NMR signal integral ratio TFA^ꢀ/2’-F 3:1)

which consistently resulted in a pH of 3.4. Such a solution

can be safely lyophilized to furnish 2·TFA as a well soluble

white powder.

The synthesis of dinucleotide analogue 3 (Scheme 3)

starts from intermediate 9a which was de- and reprotec-

ted,^[15c,20]to give first 11a then 12 (9b was deprotected to

give 11b, cf. Experimental Section and Supporting Informa-

tion). Compound 12 was coupled to a commercial cytidine

phosphoramidite precursor according to standard proce-

dures.^[21]Azidodinucleotide 13 was an equally efficient cou-

pling partner under the Staudinger–Vilarrasa conditions

(94% yield). Complete deprotection of 14a in three steps

furnished the target compound 3 with 5.3% overall yield

from 4.

During the optimization of the Staudinger–Vilarrasa cou-

pling reaction conditions a slight modification of the proto-

col—holding the reaction temperature at 08C all times—was

sufficient to allow for the appearance of a secondary prod-

uct of very similar polarity (similar R_fvalues on TLC plates)

and showing reasonably similar ¹H but distinct ¹⁹F NMR

spectra from the ones of the corresponding and desired

amides and a molecular mass of +28 Da higher than the

target compounds. Apparently, trapping the initially formed

phosphatriazene with the activated amino acid had become

possible before it eliminated dinitrogen (Scheme 4). We de-

tected the acyltriazene derivatives in varied but substantial

amounts, as determined by ESI mass (Table 1), ¹H and

¹⁹F NMR spectra (Figures 2 and 3, respectively) after chro-

matographic purification of the reaction mixtures but no

separation of the acyltriazene from the amide. When tribu-

tylphosphine was first mixed with 9b separately at 08C fol-

lowed by the addition to a solution containing the activated

amino acid at 08C, we obtained after 48 h reaction time (in-

stead of the usual 16 h) even more of the acyltriazene 10d

(Table 1, entry 3). We obtained the highest acyltriazene-to-

amide ratio, approximately 1:1, with the coupling at 08C on

dinucleotide 13 (Table 1, entry 4).

1

The H and ¹⁹F NMR spectra of the mixtures 10a + 10c

(Table 1, entry 1), 10b + 10d (Table 1, entry 3) and 14a +

14b (Table 1, entry 4) confirmed the assumption of compa-

rable volatility of the molecular ions of the amides and acyl-

triazenes—although some acyltriazene contents may appear

slightly underestimated in the ESI mass spectra—and re-

vealed useful additional informations about the nature of

the acyltriazenes (Figures 2 and 3).

In all three mixtures the presence of the acyltriazene in

deuterochloroform gave rise to several more or less broad-

1

ened H resonances (Figure 2) at low (9.5–10.5 ppm for 10c,

10d and 14b) and very low fields (13–14 ppm for 14b). The

corresponding hydrogen atoms exchanged rapidly against

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5569

P. Strazewski et al.

The ¹⁹F resonances of the same amide/acyltriazene mix-

tures consistently showed a population of several isomeric

forms of the latter usually at higher fields than the corre-

sponding amide (Figure 3). The pure azides 9a, 9b, 11a, 11b

and 12 and the pure amides 10a, 10b, 2, 15 and 3 appeared

1

as single isomers showing single peaks in the H-decoupled

¹⁹F NMR spectra (Supporting Information), not so the dia-

stereoisomeric and possibly aggregated (hydrogen bonded)

forms of 14a (Figure 3, upper spectrum) or 13 (Supporting

Information).^[22–32]

Since the chromatographic separation of the acyltriazenes

from the corresponding amides was too difficult on a

column or impossible, we resorted to chemical means

(Table 2). The mixture 14a + 14b was first heated in reflux-

ing 1,4-dioxane which caused the decomposition of all prod-

ucts (entry 1). We replaced dioxane with DMF and found

for the mixtures 10a + 10c and 14a + 14b that at 1108C

the amides remained stable whereas the acyltriazenes de-

composed (entry 2). After successfully isolating acyltriazene

10d through preparative TLC, we attempted to convert it to

amide 10b as described by Inazu and collegues^[33]and suc-

ceeded quite unexpectedly (Table 2, entries 3and 4).

Discussion

In this synthesis three chemical steps are of particular im-

portance: the stereospecific formation of a nucleosidic lyxo

2’,3’-epoxide from a cis-vicinal ditriflate, the practical and

regioselective ring-opening with inorganic azide and the

very high yielding in situ coupling of the organoazide with

an active ester of an amino acid. This Staudinger–Vilarrasa

coupling, a particularly efficient variant of the so-called

modified Staudinger reaction^[34]of a which an attractive

water-compatible version became known as the Staudinger

ligation,^[35]is the most important key step, since it bears a

great potential in more general synthetic contexts.

Scheme 4. Formation of amides 10a, 10b and 14a and acyltriazenes 10c,

10d and 14b: a) Boc-l-Tyr(Me)-OH, HOBt, DIC, P(nBu)₃, 08C over-

night; b) Fmoc-l-Tyr(Me)-OH, HOBt, DIC, P(nBu)₃, 08C overnight.

AHCTREUNG

Table 1. Ratio of amides 10a, 10b and 14a versus acyltriazenes 10c, 10d

and 14b, respectively, under conditions that favor the enrichment of acyl-

triazenes.

Entry Compounds Reagents

Ratio^[b]amide/

acyltriazene

1

2

3

9a

Boc-l-Tyr(Me)-OH, HOBt, DIC,

(nBu)₃, 08C, overnight^[a]

Boc-l-Tyr(Me)-OH, HOBt, DIC,

(nBu)₃, 08C, overnight^[a]

Boc-l-Tyr(Me)-OH, HOBt, DIC,

08C, overnight

91:9 10a/10c

P

ACHTREUNG

The Staudinger reaction^[36]is a redox reaction occurring be-

tween a phosphine as the reducing agent and an organoa-

zide as the oxidizing agent, to initially produce a phospha-

triazene (triazaphosphadiene, phosphazide) by nucleophilic

attack of the phosphorus atom of the phosphine at the ter-

minal nitrogen atom (Na) of the organoazide.^[37]In the fol-

lowing step, the intermediate phosphatriazene undergoes an

intramolecular rearrangement via a four-membered P-N-N-

N-ring transition state to yield a second intermediate, the

iminophosphorane, with concomitant loss of N₂. The result-

ing iminophosphorane is hydrolyzed in the most common

version to the amine and phosphorane oxide. The imino-

phosphoraneꢀs highly nucleophilic nitrogen atom, however,

can react with almost any kind of electrophilic reagent (the

modified versions), thus resulting in many reactions of sig-

nificant synthetic importance.^[34]

9b

87–90:13–10^[c]

10b/10d

78:22 10b/10d

P

ACHTREUNG

9b +

P(nBu)₃

A

(08C)

4

13

Fmoc-l-Tyr(Me)-OH, HOBt,

53:47 14a/14b

DIC, P

A

[a] P

ACHTREUNG

dure as described in the Experimental Section. [b] The ratios were deter-

mined through the ratio of the ESI⁺mass peak intensities of the corre-

sponding amide and acyltriazene (=amide + 28) molecular ions assum-

ing a comparable volatility of both molecular ions (for quantification by

NMR see further on in the text). [c] Result from two independent experi-

ments.

deuterium from added D₂O (example 10b + 10d + D₂O).

All resonances of acyltriazene 14b, the “exchangeable” as

well as the “non-exchangeable” ones (see SI), vanished

completely upon prolongued contact with added D₂O (com-

pare pure 14a with 14a + 14b + D₂O in Figure 2).

Phosphatriazenes: Horner and Gross^[38]were the first to in-

vestigate the mechanism of the Staudinger reaction. They

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Chem. Eur. J. 2007, 13, 5566 – 5584

2’-Deoxyfluoropuromycin Analogues

FULL PAPER

showed that in some cases,

phosphatriazenes are quite

stable under the usual condi-

tions of the Staudinger reac-

tion in organic solvents at 0–

28C, but evolve dinitrogen

upon heating to 50–1508C.

They noticed the formation of

phosphatriazene

following

second-order kinetics. Later

Leffler and Temple^[39]rein-

vestigated this result, con-

firmed it and refined the

theory. They postulated a free

energy profile of the Stauding-

er reaction in which the phos-

phatriazene complex is rather

stable, more stable than the re-

agent pair (PPh₃

+ PhN₃),

thus, saw the phosphatriazene

as a kinetically trapped, mostly

enthalpically stabilized reaction

intermediate. It was also found

that the fragment PN₃R in the

isolated phosphatriazenes was

acyclic, that is, the azide bound

to the P^IIIsite with Na, its ter-

minal nitrogen atom. In the

isolated compounds the chains

PN₃C were nearly planar and

had the trans (E) configuration

with respect to the central

ꢀ

Na Nb bond which exhibits,

partially, double-bond charac-

teristics (references in [37b]).

pK_ameasurements have shown

that phosphatriazenes are less

basic than the corresponding

iminophosphoranes.^[40]As the

parameters 1 in the Hammett

correlations of pK_avalues for

phosphatriazenes are much

smaller than for the corre-

sponding iminophosphoranes,

it was concluded that, for the

Figure 2. Comparison of the down-

1

field region of the H NMR spectra in

CDCl₃at 298 K of amides and mix-

tures of amides and acyltriazenes

(marked with an asterisk, unidentified

resonances marked with a dot). The

amide-acyltriazene ratios according to

the integration of sufficiently well re-

solved ¹H resonances are: 10a/10c

ꢁ91:9 (H1’ and NH amide); 10b/10d

ꢁ74:26 (H2 + H8), 71.5:28.5 (H4’)

and 70.5:29.5 (OCH₃, not shown here,

see Supporting Information).

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5571

P. Strazewski et al.

strated that the elimination of

the two nitrogen atoms from

ꢀ

the phosphatriazene were Na

Nb and the corresponding

product was therefore the imi-

nophosphorane containing Ng.

This demonstration confirmed

through experiment that the

decomposition of phosphatria-

zenes is an intramolecular

mechanism via

a four-mem-

bered ring transition state.

In the groups of Molina,^[41a]

Rzepa,^[45]Grützmacher^[46]and

Tian and Wang^[47]the Stau-

dinger reaction profile was in-

vestigated by means of ab

initio calculations using several

methods and two kinds of basis

sets: restricted Hartree–Fock

(RHF/6-31G*),^[41,45]

coupled

cluster calculations on single

and double substitutions from

the Hartree–Fock determinant

(including non-iteratively cal-

culated triple excitations) using

geometries from fully correlat-

Figure 3. Comparison of the ¹H-decoupled ¹⁹F NMR spectra in CDCl₃at 298 K of mixtures of amides (#) and

acyltriazenes (*). The amide-acyltriazene ratios according to the integration of the ¹⁹F resonances are: 14a/14b

ꢁ59–55:41–45 (F2’); 10a/10c ꢁ90:10 (F2’); 10b/10d ꢁ69.5:30.5 (F3’). The signal marked with a dot could not

be found in the fully coupled ¹⁹F NMR spectrum (Supporting Information).

ed second order Møller–Plesset perturbation calculations

Table 2. Heating of product mixtures (acyltriazene + amide) under vari-

ous conditions.

(CCSD(T)/6-31G**//MP2 popular

A

a

hybrid Hartree-Fock/density functional theory method

(B3LYP/6-31G*^[45,47]or B3LYP/6-311G**)^[45]that included

zero-point vibrational energy corrections and stationary

point Gibbs free energies on the reaction profiles and a

number of different substituents on phosphorous and Ng, as

well as bulk (continuum) solvent effects.^[47]

Entry Starting

compounds^[a]

Reaction

conditions

Results^[a]

1

2

3

4

53:47

14a/14b

91:9 10a/10c

53:47 14a/14b

<5:>95

dioxane reflux (1008C), 14a and 14b

60 min

DMF 1108C, 40 min

decompose

10a, 14a are stable

10c, 14b decompose

58:42 10b/10d

DOWEX-H⁺toluene

reflux (1308C), 60 min

These studies reconstructed the Staudinger reaction—very

briefly—-as one that proceeds through two major steps:

i) the system moves fastest through an initial P-Na-Nb-NgC

cis-transition state, DDG^°_{298 K}(TS1ꢀreactands)=21.5–

24.9 kcalmol^ꢀ1for the alkyl-substituted reactands,^[47]—im-

probably slower through a regioisomeric P-Ng(C)-Nb-Na

and usually slower still through a P-Na-Nb-NgC trans-transi-

tion state—to form the cis intermediate being usually (for

most alkyl-substituted reactands) slightly more stable than

10b/10d

58:42 10b/10d 10% AcOH toluene

reflux (1308C), 30 min

100:0 10b/10d

[a] Identified through TLC and quantified by ESI⁺MS of the crude reac-

tion mixtures.

phosphatriazenes, protonation occurs at Na, the N atom ad-

jacent to phosphorus. Later a few cis (Z) configured phos-

phatriazenes have been isolated where steric bulkiness and

structural constraints of the carbon backbone seemed to ex-

plain the unusual preference to a large part, although a fa-

vorable cis P···Ng overlap (weak attractive interaction) may

also have contributed.^[41]

the trans isomer, DDG_{298 K}(trans–cis)=2.2–4.0 kcalmol^{ꢀ1 [47]}

;

ii) the cis intermediate can either rapidly isomerise to the

trans intermediate, DDG^°_{298 K}(TS1ꢀcis)=10.8–12.7 kcal

mol^ꢀ1for most alkyl-substituted reactands,^[47]or it forms a

four-membered ring in passing through at least one (or

more) transition states, DDG^°_{298 K}(TS2ꢀcis)=17.4–26.2 kcal

mol^ꢀ1for most alkyl-substituted reactands,^[47]with retention

of the original configuration at phosphorus, whereafter N₂

dissociates over a very small or no barrier.

Interestingly, the free energy barriers of cis–trans isomeri-

sation and even reversion, DDG^°_{298 K}(cisTS–cis)=7.5–

12.7 kcalmol^ꢀ1for the alkyl-substituted P-Na-Nb-NgC cis-

phosphatriazenes,^[47]are lower than the ones for cyclization

Phosphatriazenes to iminophosphoranes: The thermal con-

version into iminophosphoranes through the mechanism

previously studied was experimentally investigated.^[36,37a,42]

In general the reaction seemed to proceed without either

free radicals or nitrene participation^[39,43a]and possibly with

retention of configuration at the phosphorus atom.^[43b]By

using ¹⁵N labeling techniques Bock and Schnçller^[44]demon-

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2’-Deoxyfluoropuromycin Analogues

FULL PAPER

and dissociation of N₂which makes the formation of this in-

termediate fully reversible (thermodynamic control). As a

result of the flat potential energy surface in the second for-

ward step, that is, the quasi isoenthalpic and isoergonic

P···Ng overlap transition state, P–Ng ring closure, Nb···Ng

loosening, followed by the cleavage of NaꢂNb (N₂), the

Staudinger reaction essentially takes one rate-limiting barri-

er to form the cis-phosphatriazene and another similarly

rate-limiting one to rearrange into the iminophosphorane

and dissociate N₂. It seems that, unlike Leffler and Templeꢀs

presupposition of a kinetically trapped enthalpically favored

phosphatriazene (deep energy well, high barriers), this inter-

mediate is relatively labile but may be nevertheless quite

abundant in the steady state and thus efficiently trapped for

entropic reasons. Despite an established P–Ng contact the

system may move back and forth over a number of flat tran-

sition states between several kinetically labile cyclic inter-

mediate structures along the reaction pathway, it may very

easily reopen to the P-Na-Nb-NgC phosphatriazene (cis or

trans), even reverse to phosphine and azide, before an ap-

propriate cyclic intermediate eventually dissociates N₂.

To envisage trapping reactions that scavenge the phospha-

triazenes into stabilized derivatives that prevent the cis in-

termediate from dissociating N₂, the calculated natural

charges on Na and Ng in the P-Na-Nb-NgC cis-phosphatria-

zenes are of particular interest (Table 3in ref. [47]). Upon

formation of an alkyl-substituted P-Na-Nb-NgC cis-phos-

phatriazene it is Na that faces the strongest rise in electron

density—some 0.16 to 0.24 electron charge surplus with re-

spect to the azide, depending on the nature of the substitu-

ents. However, the highest absolute negative charge density

is always on Ng, be it in the azide or the P-Na-Nb-NgC cis-

phosphatriazene: ꢀ0.31 to ꢀ0.61 on Ng versus ꢀ0.14 to

ꢀ0.29 on Na of the P-substituted phosphatriazenes (versus

ꢀ0.04 to ꢀ0.07 on Na of the azides). This questions the for-

merly predicted preferred protonation site and insinuates

comparable nucleophilicities of Na and Ng depending on

the steric accessibilty of each.

usually simple, derived from acetate or succinate or carbonic

acid derivatives (carbamates and ureas), the role of which

was to slow down the too rapid formation of the DNA dam-

aging alkyldiazonium salts and concomitantly amines (for

instance, DNA damaging b-chloroethylamine) in unacylated

triazenes; the diacyls also served as linkers between triazene

and signal peptides. Many of these studies were devoted to

the hydrolytic decomposition—most prominently, enzymatic

or basic acyl hydrolysis versus acidic cleavage of the

RNN···NC(O)R’ bond to give the amide and the diazonium

salt—metabolisation, nucleobase and DNA damaging mech-

anisms of acyltriazenes which were compared with the ones

of the related triazenes.^[51]The aqueous acid-induced frag-

mentation of 3-acyl-1,3-dialkyl^[49d]and 3-acyl-3-alkyl-1-aryl-

triazenes^[50a]into the respective diazonium salts and amides

was experimentally observed. RHF/3-21G calculations in

vacuo proposed conformational preferences, confirmed a

generally reduced basicity with respect to the unacylated tri-

azenes and suggested the major proton affinities to be at N1

(Ng) and the carbonyl oxygen atom, not N3(N a) directly

linked to the carbonyl group.^[49e]Consequently, the forma-

+

tion of RN₂

and R’NHC(O)R’’ from RNg=Nb-

Na(R’)C(O)R’’ was proposed to be preceded by the proto-

nation of the carbonyl oxygen atom,^[49d]but the fragmenta-

tion behavior of certain 3-acyl-3-alkyl-1-aryltriazenes ques-

tioned the generality of this assumption.^[50a]

Some a-aminoacyltriazenes were studied as perhaps

better prodrugs with respect to their solubility in water.^[50d]

Another strategy was followed by Vaughan and collegues,

who investigated 1-aryl-3,3-dialkyltriazenes, bistriazenes, tri-

azines and triazinones.^[52]Triazinones can be considered as

cyclic acyl- or carbonyltriazenes. Interestingly, they found

their way into the clinics as antitumor drugs against, for in-

stance, brain tumors and metastatic melanoma.^[53]None of

the above pharmacologically highly interesting and useful

compounds were synthesized from organoazides using the

Staudinger route.

Inazu and collegues reported, in their attempts to synthe-

size N-b-glucopyranosyl Ng-asparagines and Nd-glutamines,

that is, carboxamides from glucopyranosyl azides and unacti-

vated carboxylic acids derived from aspartate^[54]and gluta-

mate, thus, following the original Staudinger–Vilarrasa con-

ditions,^[34a]on the formation of an acyltriazene (a protected

derivative of Ng-glucopyranosylazo-l-asparagine) in varying

amounts at low temperatures and using simple trialkylphos-

phines, not phosphites.^[33]The authors did not specify the

configuration of the acyltriazeneꢀs N=N double bond but de-

scribed the conversion of the acyltriazene to the N-glucopyr-

anosyl amide using non-aqueous acid and heat (AcOH or

acidic cation exchange resin in refluxing toluene) without

proposing a mechanism that would explain this intramolecu-

lar way of eliminating N₂and concomitant 1,3-shift of the

acyl group. This reaction clearly contrasts the aqueous acid-

induced fragmentation of acyltriazenes into diazonium salts

and not rearranged amides or deacylation followed by elimi-

nation of N₂from the resulting triazenes to give amines and

alcohols, as described earlier.

Experimental and theoretical studies on the natural

charges in iminophosphoranes showed varied but consistent-

ly high charge separations, that is, high negative natural

charges on nitrogen and high positive natural

charges on phosphorus rationalizing the higher nucleophilic-

ity of iminophosphoranes with respect to phosphatriazenes

and suggesting that, in spite of the commonly used double

bond ylide-like representation, they are best described as

non-hypervalent partly zwitterionic compounds with quite

ꢀ

+

ꢀ

short N

P bond lengths owing to electrostatic contraction

and possibly n–s* hyperconjugation.^[48]

Acyltriazenes: This class of compounds is well known since

1983, especially since its stabilizing prodrug effect was being

explored by Michejda, Smith Jr., Kroeger Smith and colle-

gues,^[49]as well as by Rosa and collegues^[50]for a number of

1,3-dialkyl- and 1-alkyl-3-aryltriazenes known to be muta-

genic due to their DNA alkylation and double-strand cross-

linking features. The studied acyl (carbonyl) groups were

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5573

P. Strazewski et al.

All N-acyl-N-alkyltriazenes described in the literature are

assumed to bear a linear acyl-NNN-C constitution. The only

structural proof of it stems from a crystal structure of a ben-

zoyltriazene that was obtained from the benzoylation of a

quite stable monoalkyltriazene that was isolated from the

incomplete catalytic hydrogenation in ethyl acetate of the

corresponding azide into the desired amine by Gaoni

(Figure 4),^[55]thus, following the classical Staudinger

route.^[36]The isolated monalkyltriazenes were exceptionally

stable.

tained the X-ray crystal structures of both Z and E isomers

of the same compound, 1-benzyl-3-(1,3-dimesitylimidiazol-2-

ylidene)triazene.^[57b]A crystal that contained the Z isomer

was obtained only once. In the crystal structures of both Z

and E the molecules occupied two crystallographically

unique positions in the asymmetric unit. In the crystal con-

taining the Z isomer this isomer was ordered in one crystal-

lographically unique position while in the other position it

was found partly disordered around the triazene moiety. In

the E crystal both crystallographically unique positions were

ordered and E configured. In solution, the isomerisation be-

Cmoch identified in a [D₆]DMSO solution of selectively

labeled [1-¹⁵N]-1-carbamoyl-3-(tetrazolo

A

tween the E and Z forms could not be resolved by H NMR

1

yl)triazene one single isomer, the N1H tautomer of uniden-

tified stereochemistry around N2=N3, thus, a strong prefer-

ence for the H₂N-CO-NH-N=N-R over the H₂N-CO-N=N-

NH-R tautomer, R being an aromatic N-heterocycle. The

compound was obtained from acid hydrolysis of the corre-

sponding 1-cyanotriazene using 6n HCl in acetone at 608C,

proved therefore exceptionally stable as well.^[56]

at ꢀ808C in [D₈]toluene. Most interestingly, the authors

managed to extrude N₂from the compound to obtain the

N-benzyl-1,3-dimesityl-2-iminoimidazoline in >95% yield

through heating of the triazene in DMSO above 1208C. To

the best of our knowledge, this is the only other example of

an intramolecular N₂elimination and concomitant 1,3-alkyl-

or -acyl shift involving one carbon and three nitrogen atoms,

apart from the one reported by Inazu and collegues^[33]and

our preliminary results reported here.^[59–61]

Triazenes: Tautomers and cis–trans (E–Z) isomers of unacy-

lated triazenes were studied experimentally^[31b,51b,57]and the-

oretically.^[31b,58]In particular, Khramov and Bielawski ob-

Although Khramov and Bielawskiꢀs crystalline triazene

differs somewhat from the ones discussed above, since it

is a 1-alkyl-3-arylidenetriazene

rather than a 1-alkyl-3-aryl- or

U

a 1,3-dialkyltriazene, or 1-acyl-

3-aryl- or -alkyltriazene for

that matter, hence contains

two formal double bonds (R=

ꢀ

N N=N R’) rather than one,

it strongly suggests through ex-

periment that, most probably,

triazenes and perhaps also

acyltriazenes, unlike possibly

many phosphatriazenes in the

non-solid state,^[47]prefer the E

(trans) configuration around

the (partial) N=N double

bond, although the Z (cis) con-

figuration may be in some

cases almost equally well ac-

cessible and, more importantly,

that the E–Z isomerization is

always a fast process at ambi-

ent temperature.

This was experimentally

confirmed by Limbach and

collegues^[31b]who showed in a

series of impressingly minute

and precise NMR experiments

an estimated preference of the

E over the Z isomer of 1,3-di-

(p-fluorophenyl)triazene

of

some 300:1 through the inte-

gration of ¹⁹F resonances that

could be resolved at ꢀ358C

but coalesced at 258C. Their

Figure 4. Top: Crystal structure TUGGOR (from CCDC) showing the linear s-trans arrangement of the ben-

zoyltriazene. Bottom: Reaction scheme taken from ref. [55].

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Chem. Eur. J. 2007, 13, 5566 – 5584

2’-Deoxyfluoropuromycin Analogues

FULL PAPER

main objective was to study, using small amounts of dimeth-

yl- and trimethylamine, the base-catalyzed tautomerisation

mechanism of the NH tautomers of FPh-NH-N=N-PhF, the

¹H NMR resonance of which appeared at d 11.4 or around

12 ppm (compare to Figure 2). It could only be “frozen out

of exchange” (out of line broadening and coalescence)

below some ꢀ1258C. The authors elucidated an experimen-

tal 5.5 kcalmol^ꢀ1tautomerisation free energy barrier and

concluded from preliminary hybrid HF-DFT calculations

(B3LYP/6-31+G**), taking into account the polarized con-

tinuum solvent method and the self-consistent reaction field

for chloroform and tetrahydrofuran, that the fastest passage

for the proton happened with the help of a base-assisted hy-

drogen-bond switching through a transition state without

dissociation of the triazene-base contact ion pair. The exper-

imental barrier is in agreement with theoretical studies on

water-assisted 1,3-hydrogen shifts in triazene proper,^[58b]

1

while earlier H NMR measurements suggested tautomerisa-

tion free energy barriers at 295 K of various 1,3-dialkyltria-

zenes in methanol of, according to the transition state

theory, 12.8–16.0 kcalmol^ꢀ1.^[51b]

Trapping phosphatriazenes before and without the loss of

N₂: Taken together all above studies, we can describe the

various possible reactivities in Scheme 5. The upper half of

the scheme, that is, the reactions before the production of

phosphorane oxide, is much more certain than the reactions

in the lower half of the scheme. However, it is the lower

half that describes the access to many potentially valuable

products. In the context of the synthesis presented here we

have to explain two observed results: A) Why do we see in

the NMR spectra of the solutions that contain acyltriazenes,

at ambient temperature, more than one proton resonance at

low fields and more than one fluorine resonance at higher

fields than for those of the amides (Figures 2 and 3)?

Scheme 5. The Staudinger reaction and variants. E and Z refer to the

configuration between Nb and Na (N2 and N1) bearing significant

double-bond character. The semiquantitative rate descriptions (related to

the calculated free energies at 298 K and 1 atm) and structures before

trapping with water or R’’COX (X = leaving group) were taken and

summarized from Tian and Wang,^[47]so were the natural charges on P

and N atoms of the reactants and the cis-phosphatriazene that refer to

the calculated system Me₃P + N₃Me , Me₃PN₃Me. The Mulliken charg-

es on the iminophosphorane were taken from (first line) H₃PNH,^[48b]

(second line) Me₃PNH,^[48a](third line) H₃PNH,^[48d]and (fourth line for N)

Me₃PNH.^[48f]Not specified partial charges were not replaced by formal

ones. The semiquantitative rate descriptions after trapping refer to NMR

experiments with carbamoyl and unacylated triazenes, the corresponding

structures are related to the ensemble of previous studies (see text). The

structures of the transitions states ° after trapping with R’’COX and

before fragmentation are mere suggestions. The positive charges in the

transition states were placed in anticipation of the products formed (di-

azonium salt and Na-amide not shown).

If all studies on triazenes and acyltriazenes point to very

ꢀ

rapid E–Z isomerizations and N1 N3prototropies, we are

forced to conclude that we may have obtained constitutional

isomers that could not be in thermodynamic equilibrium.

Alternatively, we did observe several E–Z isomers and/or

ꢀ

N1 N3tautomers indeed but their interconversion was sig-

nificantly slowed down by aggregation through stable hydro-

gen bonding. Our earlier experience with hydrogen-bonded

systems in chloroform^[62]contradicts this latter possibility

but we cannot exclude it completely. B) Trapping an E or Z

configured phosphatriazene with an acyl group is expected

to result in a thermodynamically controlled mixture of rap-

idly interconverting E and Z Na-acyltriazenes where the

NaH tautomers should prevail. Nevertheless, how can we be

certain that only Na of a P-Na-Nb-NgC phosphatriazene

will react with the electrophile? Thus far the formation of

the regioisomeric Ng-acyltriazene (lowest central compound

in Scheme 5) from the Na-phosphatriazene has not been de-

scribed despite the theoretically obvious nucleophilicity of

Ng.

the Na···Nb bond catalyzed by hydronium ions and leading

to Na-carboxamides and highly electrophilic and mutagenic

Nb-Ng-diazonium compounds that either alkylate or hydro-

lyze or eliminate to more stable follow-up products. We

seem to have observed this reaction after adding D₂O to a

chloroform solution containing amide 14a and triazene 14b.

After a few days, only 14a remained (lowest spectrum in

Figure 2). Like the formerly studied 3-acyl-1,3-dialkyl^[49d]

and 3-acyl-3-alkyl-1-aryltriazenes,^[50a]14b was not stable in

contact with water over a long period of time. The proposed

protonation of the oxygen atom of acyltriazenes^[49d]in aque-

The best studied degradation pathway of acyltriazenes

under neutral or slightly acidic conditions is the fission of

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5575

P. Strazewski et al.

ous solutions seems indeed to be a reasonable assumption

and is depicted in Scheme 5.

Conclusion

The formation of the rearranged Ng-carboxamides under

acidic anhydrous conditions, as observed here and by Inazu

and collegues,^[33]is more difficult to understand. It necessi-

tates the protonation of Ng bearing a relatively high calcu-

lated proton affinity,^[49e]to loosen the carbonyl···Na, as well

as the Nb···Ng bonds and to attract the carbonyl group to-

wards Ng. Such a rearrangement needs to pass through a

four-membered ring transition state, as depicted in

Scheme 5, a reaction pathway similar, in a sense, to the rear-

rangement of N-alkyl-N-nitrosoamides into N-alkyl-diazo-

esters that subsequently eliminate dinitrogen to form O-

alkyl esters.^[60,61]If, however, no rearrangement is needed to

form the experimentally observed Ng-carboxamides because

the phosphatriazene reacted directly to the regioisomeric

Ng-acyltriazene in the first place, then at least two reasona-

ble Ng-protonated transition states would lead to the elimi-

nation of dinitrogen, as shown in the lowest part of

Scheme 5.

In view of the fact that we observed in the NMR spectra

of our acyltriazenes at ambient temperature several fluorine

resonances (Figure 3) and several distinct water-exchangea-

ble low-field proton resonances, two of which between d 13

and 14 ppm (Figure 2) where usually only hydrogen-bonded

systems resonate (such as adenosine-paired uridine imino

protons), we are inclined to take those regioisomeric Ng-

acyltriazenes into consideration, at least in a mixture con-

taining Na- and Ng-acyltriazenes. Whereas R-Ng=Nb-NaH-

Two new fluorinated puromycin analogues were synthesized

with the purpose of testing their protein synthesis inhibiting

activity in vitro and antibiotic activity in procaryotes. An op-

timized seven and thirteen steps procedure is presented for,

respectively, 2 and 3 from adenosine which includes a very

efficient Staudinger–Vilarrasa coupling reaction between an

organoazide and the 1-oxybenzotriazole ester of an amino

acid derivative. The coupling reaction leads to carboxamides

in 94% isolated yield under very mild reaction conditions at

ambient temperature and is therefore likely to be of more

general value than presented here, for instance, as an alter-

native for obtaining sterically demanding peptides from

azides not amines.^[34t]

The same reaction at lower temperatures generates mix-

tures of carboxamides and acyltriazenes of a priori unknown

tauto-, stereo- and regioisomery. A careful analysis of the

literature on the Staudinger reaction and acyltriazenes re-

vealed that this medicinally highly valuable class of com-

pounds—small water-soluble cyclic carbonyl triazene com-

pounds are among the rare clinical cancerostatics that are

able to pass the blood–brain barrier and that also cure meta-

static melanoma—never has been approached by through

the Staudinger reaction. The assumed mechanism of action

of acyltriazenes at physiological or slightly acidic pH is the

ꢀ

proton-assisted cleavage of a nitrogen nitrogen bond such

that a carboxamide and an alkyl or aryl diazonium salt are

generated in situ alkylating nucleic acids of which the ulti-

mately crosslinked DNA double strands become lethal to

the affected cell.

COR’ possibly “looks more stable”, R-Ng(COR’)-Nb=Na-H

A

seems to be ideally suited to form various hydrogen-bonded

aggregates in chloroform, with themselves, with Na-acyltria-

zenes, as well as with Ng-carboxamides, perhaps just stably

enough to be observed as distinct objects on the NMR time

scale.

We tested here different, anhydrous proton-assisted acyl-

triazene cleavage conditions and obtained, after the elimina-

tion of dinitrogen, different carboxamides that formally but

not necessarily resulted from a N1,N3shift of the aminoacyl

group. Since the chemical reaction mechanism is hardly

known, we publish, interpret and discuss all spectroscopic

data on our compounds and compare them to what was

known before, in order to gain a maximum insight into a

synthetically attractive variant of the Staudinger reaction

and into an exciting class of compounds, the acyltriazenes.

Last but not least, we wish to enrich the literature, using all

our fluorinated compounds, with fully interpreted^[22]

In the COSY of the 78:22 10b + 10d mixture (Table 1,

entry 3) we found a clear correlation (Supporting Informa-

tion, page 67) between a H2’ resonance at d 5.5 ppm (show-

ing a large vicinal H–F coupling) and a resonance at

6.6 ppm (both marked with asterisks in Figure 2) that ap-

peared smaller in the acyltriazene-depleted 9:1 10b + 10d

mixture (Supporting Information, page 72) and was water-

exchangeable, hence, rapidly vanished upon contact with

D₂O (Supporting Information, page 70). We therefore as-

signed the water-exchangeable part of the 6.6 ppm reso-

nance to an amide hydrogen atom of the major acyltriazene.

Whereas in 1-carbamoyl-3-aryltriazenes there seems to be a

strong preference for the H₂NCO-NH-N=N-Ar over the

H₂NCO-N=N-NH-Ar tautomer,^[56]we might have been ob-

serving, rather than R’CO-NH-N=N-R, tautomeric R’CO-

N=N-NH-R where R contains H2’ vicinal to R’CONNNH.

Of course, only a combination of NMR experiments on spe-

cifically ¹⁵N-labeled acyltriazenes and high-level ab initio

calculations will be able to fully answer these open ques-

tions.

1

¹⁹F NMR spectra (besides H, ¹³C and ³¹P), a research tool

still in its youth despite the rapidly growing number of me-

dicinally valuable fluorinated organic compounds. The

target compounds will be subjected to biological assays for

antibiotic activity.

Experimental Section

General: Pyridine was dried over CaH₂and freshly distilled, CH₂Cl₂and

THF were dried over molecular sieves 4 . Other reagents were used as

purchased. ¹H, ¹³C, ¹⁹F, and ³¹P NMR spectra were recorded in CDCl₃,

[D₆]DMSO, CD₃OD, and H₂O/D₂O (5%), at 300.1, 75.5, 282.4, and

121.5 MHz, respectively. Chemical shifts are given in ppm relative to re-

sidual CHCl₃(d7.26) or CH₂Cl₂(d5.29) or CHD₂OD (d3.31) or CH₃OH

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2’-Deoxyfluoropuromycin Analogues

FULL PAPER

3

ACHTREUNG

3

(d3.44) or CHD₃SOCD₃(d2.50) for ¹H, CDCl₃(d77.23) or CD₃OD

(d49.00) for ¹³C as internal references, and CFCl₃(d0) for ¹⁹F and H₃PO₄

(d0) for ³¹P as external references. Signals were attributed based on H–D

exchange (¹H), COSY, DEPT (¹³C) and HSQC spectra. Signal shapes

and multiplicities: br=broad, s=singlet, d=doublet, t=triplet, q=quar-

tet, quint=quintuplet, sext=sextuplet, m=multiplet. Scalar coupling

constants J are given in Hertz (Hz). Mass spectra (MS and HRMS) were

obtained using electron ionisation (EI) and chemical ionisation (CI), fast

atomic bombardment (FAB from CH₂Cl₂or H₂O/MeOH 9:1) and elec-

trospray ionization (ESI, from CH₂Cl₂or H₂O/MeOH 9:1), in part with

time-of-flight detection (TOF). Infrared spectra were obtained on a

Bruker IFS 66, Perkin-Elmer 681. Flash chromatography was performed

on silica gel 60 (0.04–0.063mm). Thin-layer chromatography (TLC plate,

Merck, silica gel on Aluminium, 20X) was performed on a pre-coated

silica gel F254 plates with fluorescent indicator. The detection of com-

pounds was carried out with UV light (254 nm). Nucleosides were visual-

ized on TLC plates by subsequent spraying with concentrated H₂SO₄and

2% naphtoresorcinol solutions in ethanol, followed by heating. UV/Vis

spectra were recorded on a Perkin-Elmer Lambda Bio 40 spectropho-

tometer equipped with a deuterium and tungsten-halogen lamp. The

buffer solutions were prepared with water purified through the Nanopure

Ultrapure D4742 water system of Barnstead. The salts (biochemical qual-

ity) were obtained from Fluka. The melting points (m.p.) and, for amor-

phous solids, melting ranges (m.r.) were determined in capillary tubes

heated electrically in a silicon oil bath (Büchi apparatus), are given in

degree Centigrade and are uncorrected. Analytical RP-HPLC purifica-

tion: 2508 mm Eurospher 100/5 RP₁₈column (Knauer), flow rate:

1.3mLmin ^ꢀ1; UV detection at 260 nm. Eluants for HPLC were prepared

with water purified through the Milli-Q system. CH₃CN was HPLC

grade: A) H₂O, TFA 0.05m, CH₃CN 1%; B) CH₃CN/H₂O 9:1.

1H, J

3

2

(dd, 1H, J=3.9, J=7.4 Hz, H-4’), 3.68 (dd, 1H, J=4.2, J=11.2 Hz, H-

5’), 3.36 ppm (dd, 1H, J=3.9, J=11.2 Hz, H-5’’); ¹⁹F NMR (CDCl₃): d=

ꢀ74.55, ꢀ74.69 (2s, 2CF₃); ¹³C NMR (CDCl₃): d=154.6, 152.9, 148.7,

145.0, 143.1, 139.0, 129.2, 128.7, 128.2, 128.1, 127.7, 127.2, 121.7, 118.5 (q,

¹J=320.0 Hz), 118.3 (q, ¹J=320.0 Hz), 88.1, 85.4, 81.8, 81.7, 80.4, 71.8,

61.7 ppm; HRMS (FAB⁺): m/z: calcd for C₅₀H₃₉F₆N₅O₈S₂: 1015.2144;

found: 1015.2142 [M]⁺.

3

2

9-(2’,3’-Anhydro-6-N,5’-O-ditrityl-b-d-lyxofuranosyl)adenine (7): Com-

pound 6a (2.6 g, 2.56 mmol) was dissolved in toluene (55 mL) containing

ꢀ

Bu₄N⁺NO₂(5.9 g, 20.48 mmol) and water (7 mL). After vigorously stir-

ring the reaction mixture for 40 h, the reaction mixture was extracted

with tBuOCH₃(90 mL) and water (2130 mL), dried with anhydrous

MgSO₄, filtered and concentrated in vacuo under reduced pressure. The

residue was purified by chromatography on a column of silica gel (MePh/

EtOAc 9:1 ! MePh/EtOAc 8:2) to give 6c (227 mg, 10%) and 7

(1.190 g, 64%) as a white amorphous solid. 7: M.r. 105–1158C; R_f=0.51

(MePh/EtOAc 8:2); ¹H NMR (CDCl₃): d=8.04, 8.02 (2s, 2H, H-2, H-8),

7.46–7.15 (m, 30H, 6C₆H₅), 6.95 (s, 1H, N⁶-H), 6.28 (s, 1H, H-1’), 4.22

(t, 1H, J(4’,5’/5’’)=6.3Hz, H-4 ’), 4.02, 4.01 (2d, 2H, J=5.8 Hz, H-2’, H-

3’), 3.49 (dd, 1H, ³J=6.2, ²J=9.3Hz, H-5 ’), 3.37 ppm (dd, 1H, ³J=6.5,

²J=9.4 Hz, H-5’’); ¹³C NMR (CDCl₃): d=154.3, 152.6, 148.9, 145.1, 143.8,

139.1, 129.2, 128.8, 128.1, 128.1, 127.4, 127.1, 120.5, 87.3, 81.0, 81.0, 71.6,

62.4, 57.4, 56.7 ppm; HRMS (FAB⁺): m/z: calcd for C₄₈H₃₉N₅O₃:

733.3053; found: 733.3048 [M]⁺.

3

9-(3’-Azido-3’-deoxy-6-N,5’-O-ditrityl-b-d-arabinofuranosyl)adenine (8a)

and 9-(2’-azido-2’-deoxy-6-N,5’-O-ditrityl-b-d-arabinofuranosyl)adenine

(8b): A mixture of 7 (925 mg, 1.26 mmol), NH₄Cl (135 mg, 2.52 mmol),

NaN₃(492 mg, 7.56 mmol), DMF (4 mL), and H₂O (600 mL) was heated

under reflux at 1008C for 1 h 30 min. The reaction mixture was extracted

with CH₂Cl₂(35 mL) and water (35 mL). The organic layer was washed

with water (335 mL), dried with anhydrous MgSO ₄and concentrated in

vacuo under reduced pressure. The residue was purified by chromatogra-

phy on a column of silica gel (MePh/EtOAc 8:2 ! MePh/EtOAc 7:3) to

give 8a (770 mg, 78.6%) and 8b (174 mg, 17.8%) as white amorphous

solids.

6-N,5’-O-Ditrityladenosine (5):

A mixture of adenosine (4) (10 g,

37.42 mmol) dried by coevaporation with pyridine, DMAP (3.84 g,

31.43 mmol), and TrCl (35 g, 125.35 mmol) in pyridine (500 mL) was

heated at 808C. The progress of the reaction was followed by TLC

(EtOAc/MeOH 9:1). After 4 h 40 min the reaction was cooled down to

ambient temperature and quenched with EtOH (150 mL). The reaction

mixture was concentrated in vacuo and coevaporated with toluene (2

250 mL). The residue was suspended in toluene (250 mL), well shaken,

filtered and the precipitate rinsed several times with toluene. The filtrate

was concentrated in vacuo, the residue was dissolved in a minimal

amount of CH₂Cl₂, a mixture of toluene and EtOAc (85:15, 75 mL) was

added, then concentrated under reduced pressure until opaqueness oc-

curred, and the solution was kept at room temperature to recrystallize 5.

Compound 5 (7.2 g) was filtered after 3.5 h and the filtrate was applied

on a column of silica gel (MePh/EtOH 4:1:0.2) to give addditional

amounts of 5 (2.2 g). Total yield: 34%; white solid. M.p. 213–2168C

(214–2178C);^[15b]R_f=0.37 (MePh/EtOAc/EtOH 4:1:0.2); ¹H NMR

Compound 8a: M.r. 114–1188C; R_f=0.38 (MePh/EtOAc 8:2); ¹H NMR

(CDCl₃): d=8.00, 7.98 (2s, 2H, H-2, H-8), 7.35–7.15 (m, 30H, 6C₆H₅),

7.05 (s, 1H, N⁶-H), 6.04 (d, 1H, J

3

ACHTREUNG

3

A

ACHTREUNG

AHCTREUNG

(dd, 1H, ³J=3.5, ²J=10.7 Hz, H-5’), 3.27 ppm (dd, 1H, ³J=4.2, ²J=

10.7 Hz, H-5’’); ¹³C NMR (CDCl₃): d=154.7, 152.1, 148.0, 144.9, 143.5,

140.8, 129.2, 128.8, 128.1, 127.5, 127.2, 121.4, 87.5, 85.4, 80.1, 77.1, 71.8,

65.8, 63.1 ppm; IR (CH₂Cl₂): n=2100 cm^ꢀ1(N₃st); HRMS (FAB⁺): m/z:

calcd for C₄₈H₄₀N₈O₃: 776.3223; found: 776.3225 [M]⁺.

Compound 8b: M.r. 119–1228C; R_f=0.35 (MePh/EtOAc 9:1); ¹H NMR

([D₆]DMSO): d=8.36 (s, 1H, H-2), 7.82 (s, 1H, H-8), 7.46 (s, 1H, N⁶-H),

(CDCl₃): d=7.88, 7.83(2s, 2H, H-2, H-8), 7.48, 7.44 (d, 1H, ²J

ACHTREUNG

3

7.36–7.18 (m, 30H, 6C₆H₅), 5.91 (d, 1H, J

A

1H, ³J=5.7 Hz, OH-3’/2’), 5.21 (d, 1H, ³J=5.7 Hz, OH-2’/3’), 4.73(q,

1H, ³J=5.2 Hz, H-2’), 4.29 (q, 1H, ³J=5.3Hz, H-3 ’), 4.06 (q, 1H, ³J=

4.7 Hz, H-4’), 3.20 ppm (brd, 2H, J=4.5 Hz, H-5’’ + H-5’); ¹³C NMR

(CDCl₃): d=154.4, 151.7, 148.1, 145.0, 143.5, 138.3, 129.2, 128.7, 128.1,

128.1, 127.4, 127.2, 121.4, 91.2, 87.3, 86.6, 76.3, 73.2, 71.7, 63.9 ppm;

HRMS (FAB⁺): m/z: calcd for C₄₈H₄₁N₅O₄: 751.3159; found: 751.3160

[M]⁺.

AHCTREUNG

A

ACHTREUNG

4’), 3.54 ppm (d, 1H, ³J=5.6 Hz, H5’ + H5’’); ¹³C NMR (CDCl₃): d=

154.6, 151.6, 146.7, 144.7, 143.9, 139.9, 129.1, 128.8, 128.1, 127.9, 127.2,

127.1, 122.0, 90.1, 87.3, 82.7, 75.7, 72.4, 71.7, 62.1 ppm; IR (CH₂Cl₂): n˜ =

2114.2 cm^ꢀ1(N₃st); HRMS (FAB⁺): m/z: calcd for C₄₈H₄₀N₈O₃:

776.3223; found: 776.3219 [M]⁺.

2’,3’-Di-O-trifluoromethylsulfonyl-6-N,5’-O-ditrityladenosine (6a): Com-

pound 5 (2.30 g, 3.06 mmol) and DMAP (0.935 g, 7.65 mmol) were dis-

solved in pyridine (70 mL). This solution was cooled to 08C and a solu-

tion of triflic anhydride (4 mL, 24.48 mmol) was added in a dropwise

manner. The reaction mixture was held at 08C for 30 min and then al-

lowed to warm to room temperature over a period of 3h. The reaction

mixture was then added to CH₂Cl₂(200 mL) extracted with water

(200 mL at 08C). The solvents were removed under reduced pressure and

the residue purified by chromatography on a column of silica gel (MePh/

EtOAc 9:1) to give 6d (0.40 g, 15%) and 6a (2.60 g, 83.7%) as a white

amorphous solid. 6a: M.r. 92–1028C; R_f=0.58 (MePh/EtOAc 9:1);

3’-Azido-2’,3’-dideoxy-2’-fluoro-6-N,5’-O-ditrityl-adenosine (9a): Diethyl-

amino sulfur trifluoride (DAST, 0.9 mL, 6.87 mmol) was added in a drop-

A

wise manner to a solution of 8a (835 mg, 1.075 mmol) in toluene (14 mL)

and pyridine (1.5 mL), and stirred at room temperature for 30 min before

heating the reaction mixture to 808C. After 45 min EtOAc (70 mL) was

added and the organic layer washed successively with 7% NaHCO₃

(60 mL) and water (60 mL), dried with anhydrous MgSO₄, filtered, and

concentrated in vacuo under reduced pressure. The residue was purified

on a silica gel column (MePh/EtOAc 8:2) to give recovered 8a (44 mg,

5.3%) and 9a (626 mg, 75%) as a yellowish amorphous solid. 9a: M.r.

106–1128C; R_f=0.76 (MePh/EtOAc 8:2); ¹H NMR (CDCl₃): d=8.07,

7.97 (2s, 2H, H-2, H-8), 7.46–7.26 (m, 30H, 6C₆H₅), 7.09 (s, 1H, N⁶-H),

¹H NMR (CDCl₃): d=7.88 (s, 1H, H-2), 7.83(s, 1H, H-8), 7.38–7.16 (m,

3

30H, 6C H₅), 6.98 (s, 1H, N⁶-H), 6.45 (t, 1H, J ꢁ5.4 Hz, H-2’), 6.21 (d,

6

Chem. Eur. J. 2007, 13, 5566 – 5584

ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemeurj.org

5577

P. Strazewski et al.

6.18 (dd, 1H, ³J

(2’,1’)=1.2, ³J(2’,3’)=4.6, ³J

(3’,2’)=4.7, ³J(3’,4’)=8.7, ³J(3’,F)=22.7 Hz, H-3’), 4.34 (td, 1H, ³J(4’,5’/

5’’)=3.5, ³J(4’,3’)=8.7 Hz, H-4’), 3.66 (dd, 1H, ³J(5’,4’)=3.1, ²J

(5’,5’’)=

(5’’,4’)=4.0, ²J

(5’’,5’)=11.0 Hz, H-

(F,3’)=22.5, ³J

(F,1’)=

C

G

³J

G

N

ACHTREUNG

G

N

U

ACHTREUNG

N

G

U

ACHTREUNG

G

(dt, ³J

ACHTREUNG

E

ACHTREUNG

19.8, ²J

U

ACHTREUNG

145.0, 143.5, 139.3, 129.1, 128.7, 128.1, 128.1, 127.4, 127.1, 121.6, 94.1 (d,

¹J

³J

A

N

³J

A

N

A

G

¹⁹F NMR (CDCl₃): d=ꢀ191.73(“quint” =dt, ²J

C

(F,2’)=

IR (CH₂Cl₂): n˜ =2112.7 cm^ꢀ1(N₃st); HRMS (ESI⁺TOF): m/z: calcd for

C₄₈H₄₀F₁N₈O₂: 779.3258; found: 779.3272 [M+H]⁺.

³J

N

ACHTREUNG

3

A

N

ESI⁺MS: m/z: calcd for C₆₃H₆₁FN₇O₆(10b): 1030.5; found: 1030.3; calcd

for C₆₃H₆₁FN₉O₆(10d): 1058.5; found: 1058.2 [M]⁺.

2’-Azido-2’,3’-dideoxy-3’-fluoro-6-N,5’-O-ditrityladenosine

(9b):

The

same procedure as for 8a was applied to 8b (35 mg, 0.045 mmol) to give

after colmun chromatography 9b (29 mg, 83%) as a yellowish amor-

phous solid. M.r. 108–1148C; R_f=0.51 (MePh/EtOAc 9:1); ¹H NMR

2’,3’-Dideoxy-2’-fluoro-3’-(O-methyltyrosyl)aminoadenosine (2): Com-

pound 10a (220 mg, 0.21 mmol) was dissolved in a mixture of CF₃COOH

and C₂H₄Cl₂(8.5:1.5, 3.2 mL) and kept at room temperature for 4 h.

MeOH (10 mL) was added to the reaction mixture and concentrated in

vacuo under reduced pression to a volume of ꢁ1 mL. After chromatog-

raphy over silica gel (EtOAc 100% ! EtOAc/MeOH 9:1 ! EtOAc/

MeOH/H₂O 9:1:0.2 ! EtOAc/MeOH/H₂O 8:2:0.3) the fractions were

combined, evaporated in vacuo then lyophilized to give 2 (91.3mg) as a

white amorphous solid (¹⁹F NMR signal integral ratio 1:1.34). Compound

2 (91.3mg) was dissolved in H ₂O nanopure (13.5 mL) to obtain a concen-

tration of ꢁ10 mm and acidified to pH 3.4 (100% salt), then lyophilized

to obtain 2·TFA (102.8 mg, 86%) as a white amorphous solid. M.r. 134–

1468C; R_f=0.37 (EtOAc/MeOH/H₂O 8:2:0.3); ¹H NMR (CD₃OD): d=

8.47, 8.23(2s, 2H, H-2, H-8), 7.21–6.95 (2d, 4H, C ₆H₄Tyr), 6.35 (d, 1H,

(CDCl₃): d=7.88, 7.84 (2s, 2H, H-2, H-8), 7.40–7.21 (m, 30H, 6C₆H₅),

3

6.95 (s, 1H, N⁶-H), 6.00 (d, 1H, J

A

³J

A

G

<

1.3, ²J

(2’,F)=23.5 Hz, H-2’), 4.45 (td, 1H,

G

A

N

ACHTREUNG

10.7 Hz, H-5’), 3.39 ppm (dd, 1H, ³J=4.3Hz, ²J=10.7 Hz, H-5’’);

¹⁹F NMR (CDCl₃): d=ꢀ195.15 ppm (ddd, ³J

²J(F,3’)=53.5 Hz, F-3’); IR (CH₂Cl₂): n˜ =2117.7 cm^ꢀ1(N₃st); HRMS

G

(ESI⁺TOF): m/z: calcd for C₄₈H₄₀F₁N₈O₂: 779.3258; found: 779.3249

[M+H]⁺.

3’-[(2-N-tert-Butoxycarbonyl-O-methyl)-l-tyrosyl]amido-2’,3’-dideoxy-2’-

fluoro-6-N,5’-O-ditrityladenosine (10a): A mixture of Boc-l-Tyr(Me)-OH

(106 mg, 0.36 mmol) and HOBt (48 mg, 0.36 mmol) was coevaporated

from anhydrous THF. This solution (6 mL) was cooled down to 08C

under N₂for 10 min then DIC (50 mL, 0.31 mmol) was added. After

10 min a solution of 9a (200 mg, 0.257 mmol) and (nBu)₃P (146 mL,

0.59 mmol) in THF (5.4 mL) was added. The reaction mixture was stirred

at RT overnight, then coevaporated with CH₂Cl₂(40 mL), EtOAc

(80 mL) were added. The organic phase was extracted with satd NaHCO₃

(40 mL) and H₂O (50 mL), dried with anhydrous MgSO₄, filtered, and

concentrated in vacuo under reduced pressure. The residue was purified

on a silica gel column (MePh/EtOAc 7:3) to give 10a (249 mg, 94%) as a

white amorphous solid. M.r. 115–1248C; R_f=0.51 (MePh/EtOAc 7:3);

¹H NMR (CDCl₃): d=7.99, 7.95 (2s, 2H, H-2, H-8), 7.50–7.16 (m, 30H,

6C₆H₅), 6.96 (s, 1H, N⁶-H), 6.89, 6.73(2d, J=8.6 Hz, 4H, C₆H₄Tyr),

3

ACHTREUNG

³J

2’), 5.02 (ddd, 1H, ³J

A

N

(2’,F)=52.4 Hz, H-

A

N

ACHTREUNG

3

4.09 (t, 1H, ³J

A

G

U

ꢁ3.2, ³J

ACHTREUNG

OCH₃), 3.46 (dd, 1H, ³J

T

ACHTREUNG

3.08 ppm (2dd, 2H, ²J

(b₁,b₂) =13.8, ³J

ACHTREUNG

¹⁹F NMR (CD₃OD): d=ꢀ196.48 (ddd, ³J

A

²J(F,2’)=52.5 ppm, F-2’); ¹³C NMR (CD₃OD): d=170.5, 160.8, 156.7,

A

153.0, 149.8, 141.2, 131.6, 127.2, 120.4, 115.5, 95.1 (d, ¹J

186.1 Hz), 89.4 (d, ²J(F,C1’)=34.4 Hz), 83.3 (d, ³J

(F,C4’) < 1 Hz), 60.8,

(F,C2’)=

N

ACHTREUNG

55.8, 55.7, 50.9 (d, ²J(F,C3’)=16.8 Hz), 37.9 ppm; HRMS (CI⁺): m/z:

G

calcd for C₂₀H₂₅F₁N₇O₄: 446.1952; found: 446.1953[ M+H]⁺.

3’-Azido-2’,3’-dideoxy-2’-fluoroadenosine (11a): Compound 9a (531 mg,

0.68 mmol) was treated with a mixture of CF₃COOH/C₂H₄Cl₂(8.5:1.5,

8.8 mL). After 3h the reaction mixture was worked up as described

above and purified by chromatography over silica gel (EtOAc/MePh 7:3

! EtOAc/MeOH 9:1 ! EtOAc/MePh/MeOH 9:0.5:0.5) to give 11a

(151 mg, 75%) as a white solid. M.p. 182–1838C; R_f=0.33 (MePh/EtOAc

1:9); ¹H NMR ([D₆]DMSO): d=8.35, 8.16 (2s, 2H, H-2, H-8), 7.39 (brs,

6.17 (d, 1H, ³J

5.36 (dd, 1H, ³J

³J(3’,2’)=4.4, ³J(3’,4’)=9.9, ³J

ACHTREUNG

A

ACHTREUNG

G

ACHTREUNG

NH carbamate), 4.29–4.15 (brd, 1H, J=6.8 Hz, CH-a), 3.91–3.82 (m,

1H, H-4’), 3.75 (s, 3H, OCH₃), 3.46–3.37 (m, 2H, H-5’, H-5’’), 2.95 (dd,

1H, ³J

²J

C

G

A

3

ACHTREUNG

G

2H, NH₂), 6.31 (dd, 1H, J

1H, ³J(2’,1’)=2.1, ³J(2’,3’)=4.5, ³J

³J(OH,5’/5’’)=5.6 Hz, OH-5’), 4.72 (ddd, 1H, ³J

³J(3’,F)=21.4 Hz, H-3’), 4.12 (dt, 1H, ³J(4’,3’)=7.8, ³J(4’,5’/5’’)=3.1 Hz,

H-4’), 3.77 (ddd, 1H, ³J(5’,OH)=5.6, ³J(5’,4’)=3.2, ²J

(5’,5’’)=12.5 Hz,

H-5’), 3.63 ppm (ddd, 1H, ³J(5’’,OH)=6.0, ³J(5’’,4’)=3.6, ²J

(5’’,5’)=

12.5 Hz, H-5’’); ¹H NMR (CDCl₃): d=8.33, 7.86 (2s, 2H, H-2 and H-8);

6.50 (dd, 1H, ³J(OH,5’)=2.4, ³J

(OH,5’’)=11.8, OH-5’); 6.09–5.89 (m,

2H, H-1’, H2’), 5.68 (brs, 2H, NH₂), 4.62–4.60 (m, 1H, H-3’), 4.24 (brt,

1H, H-4’), 3.96 (brdd, 1H, ³J(5’,OH)=1.7, ³J

(5’,5’’)=13.3 Hz, H-5’),

(5’’,5’)=12.5, ³J

(5’’,4’)=1.4 Hz, H-5’’);

(F,3’)=21.2, ³J

(F,1’)=18.7,

A

(1’,F)=18.3Hz, H-1 ’), 5.81 (ddd,

(F,3’)=27.5, ³J

G

(F,2’)=

A

N

ACHTREUNG

52.6 Hz, F-2’); ¹³C NMR (CDCl₃): d=171.5, 158.8, 155.7, 154.3, 152.7,

A

ACHTREUNG

148.2, 145.1, 143.6, 138.2, 130.3, 129.2, 129.0, 128.5, 128.1, 127.4, 127.1,

A

ACHTREUNG

121.6, 114.4, 94.2 (d, ¹J

A

G

A

N

ACHTREUNG

87.3, 81.5 (d, ³J

A

N

ACHTREUNG

²J(F,C3’)=16.2 Hz), 37.7, 28.5 ppm; HRMS (FAB⁺): m/z: calcd for

G

C₆₃H₆₀F₁N₇O₆: 1029.4589; found: 1029.4584 [M]⁺.

A

ACHTREUNG

2’-[(2-N-tert-Butoxycarbonyl-O-methyl)-l-tyrosyl]amido-2’,3’-dideoxy-3’-

fluoro-6-N,5’-O-ditrityladenosine (10b) and 2’-[(2-N-tert-butoxycarbonyl-

O-methyl)-l-tyrosyl]triazeno-2’,3’-dideoxy-3’-fluoro-6-N,5’-O-ditritylade-

nosine (10d): Same procedure as for 10a except that the reaction temper-

ature remained at 08C overnight. The extracted reaction mixture was pu-

rified on a silica gel column (MePh/EtOAc 8:2) to give the mixture of

10b and 10d as a white amorphous solid, ratio 78:22 according to ESI

MS. R_f=0.38 for 10b and 0.41 for 10d (MePh/EtOAc 8:2); ¹H NMR

(CDCl₃): d=10.34 (br, NH amide 10d), 8.20, 8.07 (2s, 2H, H-2, H-8

10b), 7.91 (brs, H-2 10d), 7.88 (s, H-8 10d), 7.47–7.20 (m, 30H, 6C₆H₅),

7.05 (s, 1H, N⁶-H), 6.90, 6.60 (2d, J=8.6 Hz, 4H, C₆H₄Tyr 10b), 6.82–

A

ACHTREUNG

3.71 ppm (brtd, 1H, ³J

C

G

ACHTREUNG

G

ACHTREUNG

²J

N

ACHTREUNG

<

calcd for C₁₀H₁₁F₁N₈O₂: 294.0989; found: 294.0986 [M]⁺.

2’-Azido-2’,3’-dideoxy-3’-fluoroadenosine (11b): The same procedure as

for 9a was applied to 9b (90 mg, 0.12 mmol) in CF₃COOH/C₂H₄Cl₂

(8.5:1.5, 1.5 mL) to give after colmun chromatography 11b (24 mg, 70%)

as a white solid. M.p. 172–1748C; R_f=0.27 (MePh/EtOAc 1:9); ¹H NMR

6.67 (m, 1.7H, C₆H₄Tyr 10d + NH amide 10d), 6.08 (d, 1H, ³J

9.3Hz, H-1 ’), 5.70–5.39 (m, NH amide 10b 10d), 5.43(ddd, 1H,

A

+

5578

www.chemeurj.org

ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Chem. Eur. J. 2007, 13, 5566 – 5584

2’-Deoxyfluoropuromycin Analogues

FULL PAPER

(CDCl₃): d=8.32, 7.84 (2s, 2H, H-2, H-8), 6.81 (dd, 1H, ³J

A

residue was purified on a silica gel column (EtOAc/MePh 8:2 ! EtOAc/

MePh 9:1 ! EtOAc/MeOH 9.5:0.5) to give 13 (253mg, 92%) as a white

amorphous solid. M.r. 97–1058C; R_f=0.36 (CH₂Cl₂/CH₃OH 95:5);

¹H NMR (CDCl₃): d=9.02–8.85 (brd, 1H, N⁴H-Ac, diast), 8.96 (s, 1H,

³J

A

N

2

(brs, 2H, NH₂), 5.21 (dd, 1H, ³J

(ddd, 1H, ³J(2’,3’)=4.2, ³J(2’,1’)=8.7, ³J

(4’,F’)=28.1, ³J

(4’,5’) ꢁ1.3Hz, H-4 ’), 3.97 (ddd, 1H,

(5’,4’)=1.5, ²J

(5’,5’’)=13.2 Hz, H-5’), 3.81 ppm (brt, 1H,

A

(3’,F)=54.2 Hz, H-3’), 4.96

U

G

ACHTREUNG

N⁶=CH), 8.49 (s, 1H, H-2), 8.35, 8.28 (2d, 1H, ³J

ACHTREUNG

(d“tW, 1H, ³J

U

ACHTREUNG

³J

(5’,F)=3.2, ³J

N

ACHTREUNG

DMT), 7.07, 7.02 (2d, 1H, ³J

ACHTREUNG

A

N

ACHTREUNG

³J

A

<

A

<

3

d=ꢀ194.27 ppm (dtd, ⁴J

A

(F,4’) ꢁ J

G

54.5 Hz, F-3’); ¹³C NMR (CDCl₃): d=156.4, 152.9, 148.6, 140.7, 121.6,

A

ACHTREUNG

93.9 (d, ¹J

²J

(F,C3’)=184.8 Hz), 89.0, 86.7 (d, ²J

(F,C4’)=21.6 Hz), 63.5 (d,

N

A

ACHTREUNG

²J=10.0 Hz, CH₂OSi), 5.16 (t, 1H, ³J=4.4 Hz, CH₂OSi), 5.06–4.93(m,

1H, H-3’ Cyt), 4.79–4.66 (m, 1H, H-3’ Ade), 4.64–4.59 (m, 1H, H-2’ Cyt),

4.55–4.49 (m, 1H, H-5’ Ade), 4.38–4.32 (m, 2H, H-4’ Cyt + H-5’’ Ade),

4.26–4.17 (m, 2H, H-4’ Ade + CH₂-O cyanoethyl), 3.97–3.84 (m, 1H,

CH₂-O cyanoethyl), 3.80–3.78 (4s, 6H, 2O-CH₃DMT), 3.71–3.63 (m,

3H, H-5’ Cyt + N(CH₂CH₂CH₂CH₃)), 3.46–3.35 (m, 3H, H-5’’ Cyt +

N(CH₂CH₂CH₂CH₃)), 2.67 (dt, 1H, CH₂CN), 2.47 (dt, 1H, CH₂CN),

2.15, 2.10 (2s, 3H, CH₃-Ac), 1.67–1.60 (m, 4H, N(CH₂CH₂CH₂CH₃)₂),

1.41–1.31 (m, 4H, N(CH₂CH₂CH₂CH₃)₂), 1.02–0.82 ppm (m, 30H,

3

G

N

calcd for C₁₀H₁₁F₁N₈O₂: 294.0989; found: 294.0983[ M]⁺.

N,N-Di-n-butylformamide dimethylacetal: Di-n-butyl formamide (50 mL)

and fresh dimethyl sulfate (26 mL) were mixed under an inert atmos-

phere and heated to reflux (1008C) during 4 h, then cooled to ambient

temperature and stirred over night. The mixture was worked up with ice-

cold absolute MeOH (150 mL) into which sodium (8 g) had been dis-

solved before. After the temperature returned to ambient, the solvent

was evaporated under reduced pressure, diethyl ether was added under

vigorous stirring and the precipitate was filtered off and rinsed with more

ether. The filtrate was evaporated under reduced pressure and the oily

residue distilled in vacuo at up to 1808C to give a clear colorless oil that

could be safely stored under an inert atmosphere in the cold.

N(CH₂CH₂CH₂CH₃)₂

+

Si(CH

G

+

Si(CH

U

(CDCl₃): d=ꢀ196.93(dt, ³J

A

ACHTREUNG

diast), ꢀ197.3–ꢀ198.3ppm (brm, F-2 ’, diast); ³¹P NMR (¹H decoupled,

CDCl₃): d=ꢀ1.39, ꢀ1.51 ppm (2s, diast); ¹³C NMR (CDCl₃): d=171.12,

171.07 (diast); 163.11, 163.07 (diast); 160.29, 160.23 (diast); 158.79, 158.76

(diast); 158.4, 154.87, 154.75 (diast); 152.9; 150.8; 150.6; 144.53, 144.43

(diast); 143.86, 143.82 (diast); 140.43, 140.39 (diast); 135.15, 135.08,

135.03, 134.99 (diast); 130.17, 130.10 (diast); 128.32, 128.25 (diast); 128.1;

127.31, 127.22 (diast); 126.33, 126.24 (diast); 116.73, 116.17 (diast); 113.4;

3’-Azido-6-N-(di-n-butylamino)methylene-2’,3’-dideoxy-2’-fluoroadeno-

sine (12): Compound 11a (150 mg, 0.51 mmol) was dissolved in anhy-

drous MeOH (2.6 mL) under N₂and N,N-di-n-butylformamide dimethy-

lacetal (414 mg, 2.04 mmol) was added. The solution was stirred at RT

for 30 min followed by evaporation. The residue was purified by chroma-

tography on silica gel (EtOAc/MePh 3:7 ! EtOAc/MePh 5:5 ! EtOAc/

MePh 7:3 ! EtOAc/MePh 8:2) to give 12 (206 mg, 93%) as a white

amorphous solid. M.r. 99–1118C; R_f=0.54 (EtOAc/MePh 9:1); ¹H NMR

([D₆]DMSO): d=8.94 (s, 1H, N⁶=CH), 8.47, 8.45 (2s, 2H, H-2, H8), 6.37

96.75, 96.49 (diast); 93.84, 93.67 (d diast, ¹J

90.00 (diast); 88.67, 87.42 (diast); 88.51, 87.64 (d diast, ²J

33.1 Hz); 81.10, 80.91 (diast); 79.52, 79.41 (diast); 79.26, 79.15 (diast);

78.32, 78.23 (diast, ³J

(F,C4’) < 1 Hz); 73.67, 73.19 (diast); 66.65, 66.26

(diast); 62.87, 62.30 (diast); 60.80, 60.28 (diast); 59.35, 59.16 (d diast,

²J

(F,C3’)=14.9 Hz); 55.28, 55.24 (diast); 52.0; 45.3; 30.9; 29.2; 24.84,

ACHTREUNG

AHCTREUNG

(dd, 1H, ³J

³J

(2’,3’) ꢁ4.0, ³J

OH-5’), 4.73(ddd, 1H, ³J

A

<

1.5, ³J

ACHTREUNG

AHCTREUNG

A

G

ACHTREUNG

24.80 (diast); 20.2; 19.8; 19.36, 19.26 (d diast); 17.81, 17.77 (diast); 13.9;

13.7); 11.89, 11.89 ppm (diast); IR (CH₂Cl₂): n˜ =2114.1 cm^ꢀ1(N₃st);

HRMS (ESI⁺TOF): m/z: calcd for C₆₄H₈₆F₁N₁₃O₁₃P₁Si₁: 1322.5959;

found: 1322.5908 [M+H]⁺.

A

E

ACHTREUNG

3’), 4.18–4.12 (m, 1H, H-4’), 3.83–3.58 (m, 4H, H-5’, H-5’’,

N(CH₂CH₂CH₂CH₃)), 3.46 (t, 2H, ³J=6.8 Hz, N(CH₂CH₂CH₂CH₃)),

1.67–1.54 (m, 4H, N(CH₂CH₂CH₂CH₃)₂), 1.37–1.27 (m, 4H,

N(CH₂CH₂CH₂CH₃)₂), 0.96–0.90 ppm (m, 6H, N(CH₂CH₂CH₂CH₃)₂);

¹H NMR (CDCl₃): d=8.99 (s, 1H, N⁶=CH), 8.49 (s, 1H, H-2), 7.92 (s,

4-N-Acetyl-5’-O-dimethoxytrityl-2’-O-triisopropylsilyloxymethylcytid-3’-

yl-{2-cyanoethyl}-5’-[2’,3’-dideoxy-3’-(N-9-fluorenylmethoxycarbonyl-O-

methyl-l-tyrosyl)amino-2’-fluoro-6-N-(di-nbutylamino)methylene]adeny-

late (14a): A mixture of Fmoc-l-Tyr(Me)-OH (49 mg, 0.12 mmol) and

HOBt (16 mg, 0.12 mmol) was coevaporated twice from anhydrous THF.

A THF solution (2 mL) thereof was cooled down to 08C under N₂for

10 min then DIC (16 mL, 0.10 mmol) was added. After 10 min a solution

of 13 (110 mg, 0.08 mmol) and (nBu)₃P (47 mL, 0.19 mmol) in THF

(2 mL), kept at RT, was added and the reaction mixture was stirred at

RT overnight. The reaction mixture was coevaporated with CH₂Cl₂

(10 mL) and then added to EtOAc (28 mL), extracted with satur.

NaHCO₃(14 mL) and H₂O (210 mL), dried with anhydrous MgSO₄, fil-

tered, and concentrated in vacuo under reduced pressure. Cyclohexane

1H, H-8), 6.59 (brd, 1H, ³J

H-1’, H-2’), 4.64–4.60 (m, 1H, H-3’), 4.26–4.22 (m, 1H, H-4’), 3.98 (brd,

A

1H,

³J

A

H-5’),

3.77–3.65

(m,

N(CH₂CH₂CH₂CH₃)), 3.41 (t, 2H, ³J=7.3Hz, N(C H₂CH₂CH₂CH₃)),

1.70–1.60 (m, 4H, N(CH₂CH₂CH₂CH₃)₂), 1.44–1.31 (m, 4H,

N(CH₂CH₂CH₂CH₃)₂), 0.99–0.9 ppm (m, 6H, N(CH₂CH₂CH₂CH₃)₂);

¹⁹F NMR ([D₆]DMSO): d=ꢀ198.93ppm (ddd, ³J

C

18.6, ²J

A

(m, F-2’); ¹³C NMR (CDCl₃): d=160.7, 158.4, 152.2, 150.0, 141.2, 127.2,

92.1 (d, ¹J(F,C2’)=196.3Hz), 88.0 (d, ²J

(F,C1’)=31.0 Hz), 84.7 (d,

³J

(F,C4’)=2.0 Hz), 62.4, 61.0 (d, J(F,C3’)=14.2 Hz), 52.0, 45.3, 30.9, 29.2,

A

ACHTREUNG

2

ACHTREUNG

A

20.1, 19.7, 13.8, 13.6 ppm; HRMS (CI⁺): m/z: calcd for C₁₉H₂₉F₁N₉O₂:

434.2428; found: 434.2421 [M+H]⁺.

cyclohexane. The residue (a white powder now essentially free of

nBu₃PO) was redissolved in a minimum amount of EtOAc/MeOH 95:5

and purified on a silica gel column (EtOAc/MePh 95:5 ! EtOAc !

EtOAc/MeOH 95:5) to give 14a (131 mg, 94%) as a white amorphous

solid. M.r. 106–1108C; R_f=0.39 (CH₂Cl₂/CH₃OH 95:5); ¹H NMR

(CDCl₃): d=8.97 (s, 1H, N⁶=CH), 8.62 (br, 1H, NH-acetyl), 8.56 (s, 1H,

H-2), 7.79–7.39 (m, 8H, CH arom Fmoc), 7.71 (s, 1H, H-8), 7.66–7.61

(2d, 1H, H-C6 Cyt, diast), 7.34–6.79 (m, 14H, CH arom DMT + NH

amide), 7.20–7.17 (2d, 1H, H-C5 Cyt, diast), 6.96–6.69 (2d, 4H, C₆H₄

Tyr), 6.24–5.83(m, H-1 ’ Cyt + H-1’ Ade), 5.68–5.34 (m, H-2’ Ade + H-

3’ Ade + NH carbamate), 5.22–5.17 (m, 2H, CH₂OSi), 4.93–4.91 (m, 1H,

H-3’ Cyt), 4.63–4.55 (m, 1H, H-2’ Cyt), 4.49–4.41 (m, 1H, H-5’ Ade),

4.32–4.20 (m, 4H, CH₂O Fmoc + H-4’ Cyt + H5’’ Ade), 4.06–3.93 (m,

2H, H-4’ Ade + CH-a), 3.81–3.78 (m, 2H, CH₂O cyanoethyl), 3.74–3.71

4-N-Acetyl-2’-O-triisopropylsilyloxymethyl-5’-O-dimethoxytritylcytidyl-

{3’-O^P-2-cyanoethyl}-5’-[3’-azido-2’,3’-dideoxy-2’-fluoro-6-N-(di-n-butyl-

amino)methylene]adenylate (13): Compound 12 (90 mg, 0.208 mmol) and

ethylthiotetrazole (40 mg, 0.312 mmol) were co-evaporated with anhy-

drous CH₃CN (35 mL) under reduced pressure and redissolved in anhy-

drous CH₃CN (0.7 mL). After addition of commercial N⁴-acetyl-5’-O-di-

methoxytrityl-2’-O-triisopropylsilyloxymethylcytid-3’-yl-(2-cyanoethyl)-

N,N-diisopropylphosphoramidite (425 mg, 0.437 mmol) in anhydrous

CH₃CN (0.8 mL), the solution was stirred at RT for 15 min followed by

addition of a solution of 0.2m I₂/THF/pyridine/H₂O (1.15 mL, 0.229 mmol

I₂), faint yellow color persisted at the end. The reaction mixture was con-

centrated in vacuo under reduced pressure to one half the volume, taken

up in EtOAc (36 mL), and extracted with 0.2m NaHSO₃(28 mL) and

saturated NaCl (5 mL). The organic phase was dried with anhydrous

MgSO₄, filtered, and concentrated in vacuo under reduced pressure. The

(m, 10H, H-5’ Cyt

+

H-9’’ Fmoc

+

2O-CH₃DMT

+

N(CH₂CH₂CH₂CH₃)), 3.49 (s, 3H, OCH₃Tyr), 3.40 (t, 2H,

N(CH₂CH₂CH₂CH₃)₂), 3.35–3.29 (m, 1H, H5’’ Cyt), 3.00–2.87 (m, 2H,

Chem. Eur. J. 2007, 13, 5566 – 5584

ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemeurj.org

5579

P. Strazewski et al.

CH₂-b₁,b₂), 2.59–2.05 (dt, 2H, CH₂CN, diast), 2.03(s, 3H, CH ₃-Ac), 1.72–

1.56 (m, 4H, N(CH₂CH₂CH₂CH₃)₂), 1.41–1.29 (m, 4H,

N(CH₂CH₂CH₂CH₃)₂), 1.03–0.88 ppm (m, 30H, N(CH₂CH₂CH₂CH₃)₂

Si(CH(CH₃)₂)₃Si(CH

³J(F,3’)=³J

(F,1’)=21.5,

²J

N

U

ACHTREUNG

+

³J

G

+

(CH₃)₂)₃); ¹⁹F NMR (CDCl₃): d=ꢀ194.7 (dt,

d=0.00; ¹³C NMR (D₂O/trace CD₃OD): d=174.4, 166.6, 159.1, 157.9,

U

G

²J

U

F-2’,

diast);

ꢀ194.8–

156.4, 153.9, 148.8, 141.6, 139.8, 131.5, 128.7, 119.6, 115.3, 96.5, 94.7 (d,

ꢀ196.0 ppm (2brm, F-2’, diast/aggreg); ³¹P NMR (¹H decoupled, CDCl₃):

d=ꢀ1.45, ꢀ1.50 ppm (2s, diast); ¹³C NMR (CDCl₃): d=172.7; 171.4;

162.87, 162.81 (diast); 160.4; 158.9, 158.7; 158.6, 158.4; 156.22, 156.17

(diast); 153.21, 153.16 (diast); 152.8; 150.92, 150.83 (diast); 149.51, 149.45

(diast); 144.1; 143.8; 143.7; 141.4; 141.4; 135.7, 135.2; 130.7, 130.6; 130.3,

130.2; 128.3, 128.2; 128.0, 127.9; 127.4; 127.3, 127.2; 126.76, 126.49

(diast); 125.11, 125.07; 120.19, 120.10; 116.66, 116.31 (diast); 114.0, 113.7;

¹J

(F,C4’)

N

16.6 Hz), 38.9 ppm; HRMS (ESI⁺

C₂₉H₃₇F₁N₁₀O₁₁P₁: 751.2365; found: 751.2370 [M+H]⁺.

113.5; 113.4; 97.81, 97.65 (diast); 93.6 (d, ¹J

89.16 (diast); 88.6 (d, ²J

(F,C1’)=35.6 Hz); 87.6; 82.55, 82.43 (diast); 78.6;

78.0; 77.8 (³J

(F,4’) < 1 Hz); 77.4; 67.4; 67.36, 67.24 (diast); 63.82, 63.78

(diast); 62.71, 62.66 (diast); 55.8 (d, ²J

(F,3’)=16.8 Hz); 55.4, 55.3; 55.21,

A

ACHTREUNG

We thank Sergej Gryaznov, Menlo Park, California (USA), Igor Mikhai-

lopulo, University of Kuopio (Finland) and Thierry Billard, SERCOF,

UniversitØ Claude Bernard Lyon 1 (France), for helpful comments.

AHCTREUNG

55.14; 52.0; 47.1; 45.3; 45.3; 31.1; 29.4; 24.8; 20.3; 19.9; 19.41, 19.34

(diast); 17.8; 14.1; 13.8; 11.9 ppm; HRMS (ESI⁺TOF): m/z: calcd for

C₈₉H₁₀₉F₁N₁₂O₁₇P₁Si₁: 1695.7525; found: 1695.7469 [M+H]⁺.

5’-O-Dimethoxytritylcytidylyl-{3’-O^P}-5’-[2’,3’-dideoxy-2’-fluoro-3’-(O-

methyl-l-tyrosyl)amino]adenylate (15): Compound 14a (71 mg,

0.04 mmol) was dissolved in 33% CH₃NH₂/EtOH (10 mL). The solution

was stirred at RT in a closed vessel for 35 min, then evaporated in vacuo

under reduced pressure and co-evaported with THF (25 mL), followed

by addition of 1m TBAF/THF (252 mL, 0.252 mmol) and THF (0.5 mL).

After 40 min the reaction mixture was concentrated in vacuo under re-

duced pressure. The chromatography column was conditioned with

EtOAc/MeOH/H₂O 9:2:1 and the mixture was purified by chromatogra-

phy (EtOAc/MeOH/H₂O 9:2:1 ! EtOAc/MeOH/H₂O 8:3:1 ! EtOAc/

MeOH/H₂O 8:4:1) to give 15·TBA (45 mg, 83%) as a white amorphous

solid (as the mono-tetrabutylammonium salt, as determined by ¹³C NMR

and quantified by ¹H NMR). M.r. 174–1808C; R_f=0.35 (EtOAc/MeOH/

H₂O 8:4:1); ¹H NMR (CD₃OD): d=8.24, 8.15 (2s, 2H, H-2, H-8), 7.91

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[2] a) R. R. Samaha, R. Green, H. F. Noller, Nature 1995, 377, 309–314;

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Fucini, D. Wilson, M. Peretz, M. Kessler, A. Yonath, Biopolymers

2003, 70, 19–41.

(d, 1H, ³J

DMT + 4H, C₆H₄Tyr), 6.22 (d, 1H, ³J

(d, 1H, ³J

(1’,2’)=4.1 Hz, H-1’ Cyt), 5.49–5.28 (m, 2H, H-C5 Cyt + H-2’

Ade), 5.04–4.91 (ddd + m, 2H, H-3’ Ade + H-3’ Cyt), 4.43(t, 1H,

³J

(2’,1’)=4.5 Hz, H-2’ Cyt), 4.33–3.99 (m, 4H, H-4’ Ade + H-4’ Cyt +

H5’,H5’’ Cyt), 3.78–3.68 (m, 10H, 2OCH₃DMT + OCH₃Tyr + CH-

a), 3.51–3.35 (m, 2H, H5’ H5’’ Ade), 3.26–3.21 (m, 8H,

(CH₃CH₂CH₂CH₂)₄N⁺), 3.01 (dd, 1H, ³J(b₁,a)=6.0, ²J

(b₁,b₂)=13.8 Hz,

(b₁,b₂)=13.8 Hz, H-b₂), 1.71–1.61 (m,

1.42 (sext, J=7.5 Hz, 8H,

ACHTREUNG

AHCTREUNG

[9] H. Chapuis, P. Strazewski, Tetrahedron 2006, 62, 12108–12115.

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289, 920–930.

ACHTREUNG

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H.-H. Limbach, Magn. Reson. Chem. 2001, 39, S91–S99; e) J. L.

Alonso, S. Antolínez, S. Blanco, A. Lesarri, J. C. López, W. Cami-

nati, J. Am. Chem. Soc. 2004, 126, 3244–3249; f) J. D. Dunitz, Chem-

BioChem 2004, 5, 614–621.

+

A

ACHTREUNG

3

2

ACHTREUNG

H-b₁), 2.80 (dd, 1H, J

8H,

(b₂,a)=7.7, J

(CH₃CH₂CH₂CH₂)₄N⁺),

(CH₃CH₂CH₂CH₂)₄N⁺), 1.02 ppm (t, 12H, (CH₃CH₂CH₂CH₂)₄N⁺);

¹⁹F NMR (CD₃OD): d=ꢀ197.57 ppm (ddd, ³J

U

N

²J(F,2’)=52.5 Hz, F-2’); ³¹P NMR (CD₃OD): d=0.00 ppm (q,

A

[12] J. S. Weinger, K. M. Parnell, S. Dorner, R. Green, S. A. Strobel, Nat.

Struct. Mol. Biol. 2004, 11, 1101–1106.

³J=6.5 Hz); ¹³C NMR (D₂O/trace CD₃OD): d=175.6, 170.3, 167.4, 160.2,

158.4, 157.3, 154.0, 150.0, 146.0, 142.6, 140.9, 136.8, 136.5, 131.5, 131.5,

[13] a) D. OꢀHagan, C. Bilton, J. A. K. Howard, L. Knight, D. J. Tozer, J.

Chem. Soc. Perkin Trans. 2 2000, 605–607; b) C. R. S. Briggs, D.

OꢀHagan, H. S. Rzepa, A. M. Z. Slawin, J. Fluorine Chem. 2004,

125, 19–25; c) C. R. S. Briggs, M. J. Allen, D. OꢀHagan, D. J. Tozer,

A. M. Z. Slawin, A. E. Goeta, J. A. K. Howard, Org. Biomol. Chem.

2004, 2, 732–740; d) N. E. J. Gooseman, D. OꢀHagan, A. M. Z.

Slawin, A. M. Teale, D. J. Tozer, R. J. Young, Chem. Commun. 2006,

3190–3192.

129.7, 129.5, 128.9, 128.0, 120.5, 115.2, 114.2, 96.0, 95.0 (d, ¹J

ACHTREUNG

186.3Hz), 90.8, 88.3, 89.4 (d, ²J

N

ACHTREUNG

AHCTREUNG

16.0 Hz), 40.6, 24.8, 20.7, 14.0 ppm; HRMS (ESI^ꢀTOF): m/z: calcd for

C₅₀H₅₃F₁N₁₀O₁₃P₁: ꢀ1051.3515; found: ꢀ1051.3625 [MꢀH]^ꢀ

.

Cytidyl-{3’-O^P}-5’-[2’,3’dideoxy-2’-fluoro-3’-(O-methyl-l-tyrosyl)amino]-

A

AcOH/H₂O 8:2 (15 mL). The solution was stirred at RT for 20 min and

then lyophilised. The residue was purified by chromatography over a

silica gel column which was conditioned and eluted with EtOAc/MeOH/

H₂O 7:3:2 to give, after lyophilisation from H₂O, 3 (19 mg, 82%) as a

white amorphous solid. M.r. 192–1978C; R_f=0.31 (EtOAc/MeOH/H₂O

7:3:2); ¹H NMR (D₂O): d=8.39, 8.26 (2s, 2H, H-2, H-8), 7.72 (d, 1H,

[14] a) S. R. Starck, R. W. Roberts, RNA 2002, 8, 890–903; b) M. D. Er-

lacher, K. Lang, N. Shankaran, B. Wotzel, A. Hüttenhofer, R.

Micura, A. S. Mankin, N. Polacek, Nucleic Acids Res. 2005, 33,

1618–1627; c) B. Zhang, L. Zhang, L. Sun, Z. Cui, Org. Lett. 2002,

4, 3615–3618; d) unpublished results.

[15] a) J.-T. Huang, L.-C. Chen, L. Wang, M.-H. Kim, J. A. Warshaw, D.

Armstrong, Q.-Y. Zhu, T.-C. Chou, K. A. Watanabe, J. Matulic-

Adamic, T.-L. Su, J. J. Fox, B. Polsky, P. A. Baron, J. W. M. Gold,

W. D. Hardy, E. Zuckerman, J. Med. Chem. 1991, 34, 1640–1646;

b) R. G. Schultz, S. M. Gryaznov, Nucleic Acids Res. 1996, 24, 2966–

2973; c) K. W. Pankiewicz, J. Krzeminski, L. A. Ciszewski, W.-Y.

Ren, K.A. Watanabe, J. Org. Chem. 1992, 57, 553–559; d) R. W.

Binkley, J. Carbohydr. Chem. 1994, 13, 111–123; e) E. Kattnig, M.

Albert, Org. Lett. 2004, 6, 945–948.

³J

ACHTREUNG

A

ACHTREUNG

5.65 (d, 1H, ³J

A

ACHTREUNG

³J

A

N

ACHTREUNG

4.2 Hz, H-3’ Ade), 4.41–4.37 (m, 1H, H-3’ Cyt), 4.32–4.30 (m, 1H, H-2’

Cyt), 4.23–4.20 (m, 2H, H-4’ Cyt + H5’ Cyt), 4.11–4.09 (m, 1H, H-4’

Ade), 3.93–3.88 (m, 2H, CH-a + H5’ Ade), 3.85 (s, 3H, OCH₃Tyr),

3.82–3.77 (m, 2H, H5’’ Ade + H5’’ Cyt), 3.08 (dd, 1H, ³J

(b₁,a)=6.1,

A

5580

www.chemeurj.org

ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Chem. Eur. J. 2007, 13, 5566 – 5584

2’-Deoxyfluoropuromycin Analogues

FULL PAPER

[16] During the optimization of the synthesis of ditriflate 6a we could

isolate a singly isomeric monotriflate from the reaction mixture. It

was identified by ¹H NMR to be the 2’-O triflate 6d (cf. Supporting

Information). We conclude that this regioisomer forms more readily

and seems less prone to nucleophilic substitution than the 3’-O tri-

flate, although it is more prone to b-elimination owing to the appar-

ent enhanced acidity of H1’ when compared with the H4’ or H3’ and

despite the synclinal or synperiplanar orientation of H1’-C1’-C2’-

OTf2’ (Æ358).

anomeric nucleoside derivatives). close inspection of

A

³J

G

pucker preference for the 3’-azido-3’-deoxyarabinofuranosyl deriva-

tive 8a in CDCl₃(³J(H1’a, H2’a)=5.3Hz), a ꢁ77% preference for

the North-type pucker in 2’-azido-2’-deoxyxylofuranosyl derivative

8b in CDCl₃(³J(H1’a,H2’b)=2.1 Hz), consistently and expectedly

high North preferences for the 3’-azido-2’,3’-dideoxy-2’-fluororibo-

furanosyl derivatives 9a (³J(H1’a,H2’b)=1.3Hz or ꢁ86% North in

CDCl₃), 11a (³J(H1’a,H2’b)=2.1 Hz or ꢁ78% North in

[17] T. R. Webb, H. Mitsuya, S. Brodert, J. Med. Chem. 1988, 31, 1475–

1479.

[D₆]DMSO), 12 (³J(H1’a,H2’b)

<

1.3Hz or

> 86% North in

[D₆]DMSO) and 13 (³J(H1’a,H2’b) < 1.5 Hz or >84% North in

[18] D. Bouzard, P. Di Cesare, M. Essiz, J. P. Jacquet, J. R. Kiechel, P. Re-

muzon, A. Weber, T. Oki, M. Masuyoshi, R. E. Kessler, J. Fung-

Tomc, J. Desiderio, J. Med. Chem. 1990, 33, 1344–1352.

[19] P. Van Rompaey, K. Nauwelaerts, V. Vanheusden, J. Rozenski, H.

Munier-Lehmann, P. Herdewijn, S. Van Calenbergh, Eur. J. Org.

Chem. 2003, 2911–2918.

[20] a) L. J. McBride, R. Kierzek, S. L. Beaucage, M. H. Caruthers, J.

Am. Chem. Soc. 1986, 108, 2040–2048; b) B. C. Froehler, M. D. Mat-

teucci, Nucleic Acids Res. 1983, 11, 8031–8036; c) N. Q. Nguyen-

Trung, O. Botta, S. Terenzi, P. Strazewski, J. Org. Chem. 2003, 68,

2038–2041.

CDCl₃), consistently high South preferences for the regioisomeric

2’-azido-2’,3’-dideoxy-3’-fluororibofuranosyl

derivatives

9b

(³J(H1’a,H2’b)=7.9 Hz or ꢁ83% South in CDCl₃) and 11b

(³J(H1’a,H2’b)=8.7 Hz or ꢁ92% South in CDCl₃, in agreement

with ref. [23]), a quasi absolute South preference for the regioiso-

meric puromycine analogue 10b (³J(H1’a, H2’b)=9.3Hz or ꢁ98%

South in CDCl₃, the largest ³J(H1’,H2’) measured^[23]), and of course

N

an absolute North preference (no measurable H1’a-H2’b coupling)

for all 2’-deoxy-2’-fluoropuromycine analogues (3’-carboxamido de-

rivatives) 10a, 2, 14a, 15 and 3, irrespective of the solvent (CDCl₃,

CD₃OD or D₂O at pD 7). The apparent pucker preferences of all

above furanosyl derivatives appear in agreement with Altona and

Sundaralingamꢀs^[24]and Chattopadhyayaꢀs,^[25]as well as Brunck and

Weinholdꢀs^[26]models of the dependence on the electron density de-

pleting potency (quantifyable through Mullayꢀs^[27]or other empirical

group electronegativities, see refs. [14] in [28]) of the 2’ and 3’ sub-

stituents through a subtle balance between stereoelectronic gauche

versus anomeric effects^[25]or the antiperiplanar effect^[26]that govern

the pucker equilibria of nucleosides (see discussion in ref. [23]). Ac-

cording to our measurements, the azido group seems, perhaps aston-

ishingly, slightly more electronegative (more effective in electron

[21] X.-F. Zhu, A. I. Scott, Nucleosides Nucleotides Nucleic Acids 2001,

20, 197–211.

[22] Only very recently we learned that in the laboratories of Mikhailo-

pulo and his collegues another synthetic route was pursued to syn-

thesize, among a number of similar fluorinated compounds, azide

11a (Scheme 3) and puromycine analogue 2 (Figure 1), in order to

study the influence of various substituents that replace the 2’- and/or

3’-hydroxy groups of nucleosides on the equilibria between two fura-

nose puckers, and structural informations thereof, using a maximum

number of measured vicinal coupling constants as an input for the

program PSEUROT v6.3. The regioisomer 11b was already synthe-

sized and analyzed by the authors (their compound 20)^[23]along with

other 3’-deoxyfluororibonucleosides. Here we wish to report in this

context on the approximate positions of the rapidly interconverting

North–South equilibrium of b-hydroxyazides 8a and 8b, b-fluoro-

density depletion) than

a carboxamide function—albeit much

weaker than hydroxyl—irrespective of a positive charge present or

absent in the a-position of the amido substituent. In addition, we

observed in the ¹⁹F NMR spectra of the 3’-azido-2’,3’-dideoxy-2’-

fluoro analogues 11a and 12—both bearing free hydroxyls in the 5’

position—a marked solvent dependence of the furanose pucker

preference depending on whether the spectrum was measured in

[D₆]DMSO (ꢁ78 and >86% North, respectively) or in CDCl₃. In

dimethyl sulfoxide we observed the usual ddd signal (as in all the

other 3’-azido-2’,3’-dideoxy-2’-fluoro derivatives) at d ꢀ199.2 and

ꢀ198.9 ppm, respectively, which originate from three distinct scalar

azides 9a, 9b, 11a, 11b, 12, and 13, as well as for the puromycine

analogues (carboxamido derivatives) 10a, 10b, 2, 14a, 15 and 3, as

revealed by their scalar vicinal proton–proton coupling constants be-

tween H1’ and H2’, as well as vicinal fluor–proton coupling con-

stants. Instead of analyzing our NMR data through PSEUROT, we

should like to apply a slightly modified version of the well known

and quite simple but robust model, the “10 Hertz rule-of-thumb”, to

estimate the pucker preference of our compounds in a given solvent.

According to this rule, the ³J(H1’a,H2’b) values—Ha for C5’-trans,

Hb for C5’-cis configured hydrogen atoms—indicate an approximate

tendency for either a North- or South-type pucker preference of

some pentafuranoses. Pure North-type ribo- or xylofuranose puckers

(2’-pseudoaxial substituent) give rise to a zero coupling between

H1’a and H2’b, pure South-type ribo- or xylofuranose puckers (2’-

pseudoequatorial substituent) result in close to 10 Hertz coupling

constants between H1’a and H2’b. Thanks to the above-mentioned

study by Mikhailopulo et al.^[23]we could correlate thirteen of their

³J(H1’a,H2’b) values of eleven ribofuranose derivatives with their

corresponding South preferences as determined through PSEUROT

(their compounds 2, 4, 5, 10b in DMSO, 10b in methanol, 11b in

DMSO, 11b in methanol, 12b, 17, 20, 23, 24 and 25). The linear cor-

relation coefficient was r=0.9938 and the correlation showed that,

although not overwhelmingly convincing due to the lack of very-

high-North compounds, 0% South indeed corresponds to not very

much above 0 Hz for ³J(H1’a,H2’b) but the upper limit value for

100% South is more likely to correspond to 9.5 rather than 10 Hz

(cf. Supporting Information), with an error margin of probably

Æ5%. The way we calculate from ³J(H1’a,H2’b) values the pucker

preference is thus more of a “9.5 Hertz rule-of-thumb”: we thus

multiply the ³J(H1’a,H2’b) value in Hertz with 10.53to obtain %

South (=100 ꢀ %North). Of course this rule will not apply to

F,H coupling constants, one geminal of ²J

and two different vicinals of ³J

³J(F,H)=21–22 Hz for F2’–H3’ (in the ¹H NMR spectra 52.4, 18.3

and 21.4 Hz, respectively). Were the puckers of 11a and 12 in

DMSO 100% North, these ³J

(F,H) values would represent torsional

ACHTREUNG

AHCTREUNG

angles, according to crystal structure coordinates of several nucleos-

(t)ide North puckers, of between ꢀ14 and ꢀ338 (not too far from

syn) for H1’a-C1’-C2’-F2’a and 162–1638 (close to anti) for H3’b-

C3’-C2’-F2’a. In chloroform the multiplicity of the ¹⁹F NMR signals

of 11a and 12 collapsed each to a kind of dd signal—with more

minor badly or unresolved peaks within—and shifted to around

ꢀ207.7 ppm (which folded in to +58.5 ppm when the usual limit of

¹⁹F high field detection of ꢀ206.5 ppm was applied, cf. Supporting

Information). These pseudo-dd signals did not show first-order mul-

tiplicity: the “roof effect” (order of peak intensities) seemed in-

versed—as if the downfield half of the resonance were folded in

from an even higher field—and the measured peak distances did not

reveal the true F–H coupling constants, since the apparent peak

splittings (in Hz) could not be found in the corresponding ¹H NMR

spectra in CDCl₃(cf. Supporting Information). In the latter, howev-

er, the absence of a vicinal F2’–H3’ coupling in the H3’ signals and a

1.2 ppm downfield shift of the 5’-OH signals with respect to the

¹H NMR spectra in [D₆]DMSO became evident, along with other

differences. Unfortunately, the H1’–H2’ coupling constants could not

be elucidated with certainty owing to the close proximity of those

resonances in CDCl₃. We interpret the unusual ¹⁹F NMR signals and

3

³J(H1’a, H2’a) values (arabinofuranoses) or J(H1’b, H2’) values (a-

Chem. Eur. J. 2007, 13, 5566 – 5584

ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.chemeurj.org

5581

P. Strazewski et al.

3

the strong 5’-OH downfield shift in the ¹H NMR spectra of 11a and

12 in CDCl₃with a stable intramolecular 5’-OH···N3hydrogen

bridge that forces the base into the usually less favored syn confor-

mation and the furanose pucker into one belonging to the South

type (despite F2’a) where the torsional angle H3’b-C3’-C2’-F2’a is

assumed to be close to perpendicular (ꢁ868, as for H3’b-C3’-C2’-

O2’a in the crystal structure ACADOS of 3’-O-acetyladenosine).^[29]

Apparently, the forced North-to-South rearrangement, thus, the re-

positioning of F2’a from the usually preferred pseudoaxial into the

pseudoequatorial orientation through an intramolecular hydrogen

bond, caused enhanced shielding, an upfield shift of the correspond-

ing ¹⁹F resonance of 8.6 to 8.7 ppm. Not unexpectedly, the intramo-

cinal proton–proton coupling constants J(H1’a,H2’b) that are larger

for the 2’-fluoro-3’-azides than for the corresponding 2’-fluoro-3’-

amides and smaller for the 3’-fluoro-2’-azide than for the 3’-fluoro-

2’-amide, therefore, suggesting a weaker preference for the pseu-

doaxial orientation of fluorine in vicinal cis-fluoroazides than in vici-

nal cis-fluoroamides due to a higher electronegativity of the azido

with respect to the carboxamido function, is very evidently sensed in

the fluorine resonances. The higher the population of pseudoaxially

oriented fluorine atoms, the stronger its time-averaged chemical de-

shielding, the more downfield the ¹⁹F resonance appears. The com-

parison of ¹⁹F resonances between azide-amide pairs, in which all

other atoms remain the same, reveal downfield shifts of Dd_F

(amide ꢀ azide)=1.10 ppm for 10a/9a (CDCl₃), 2.51 ppm for 13/

14a (CDCl₃), 2.72 ppm for 2 (CD₃OD)/11a (DMSO) and 3.40 ppm

for 10b/9b (CDCl₃). Recall that we observed the strongest down-

field shift for the pseudoequatorial-to-pseudoaxial rearrangement of

a fluorine (due to the formation of a stable hydrogen bridge) as we

passed from CDCl₃to DMSO of solutions of 2’-fluoro-3’-azides 11a

and 12: Dd_F=8.6–8.7 ppm. In addition, useful information on equi-

librium torsional angles can be extracted from F,H scalar coupling

constants even without a corresponding Karplus–Conroy function,

such as the ones that were very impressively and carefully worked

out by J. Chattopadhyaya and co-workers.^[28]In the compounds mea-

lecular hydrogen bond seems to be preferred in chloroform but dis-

ꢀ1

favored in dimethyl sulfoxide. An estimated 3to 5 kcalmol

free

energy drop due to the formation of 5’-OH···N3gives us an upper

limit for the compensated for rise in free energy due to the pseudo-

A

with recent measurements and calculations of its stereoelectronic

gauche effect (in general 1–2 kcalmol^ꢀ1in favor of gauche versus

anti F-C-C-N).^[13]We refrain, however, from quantifying the energet-

ics of gauche (maximal at Æ608) or anomeric effects (optimal at 0

and 1808) in geometries where torsional angles between heteroa-

toms of 908 (O4’-C1’-C2’-O2’a), 438 (O3’a-C3’-C2’-O2’a) and ꢀ1518

(N9b-C1’-C2’-O2’a) dominate the scene of North puckers (taken

from CUPYUH),^[30]while in typical South puckers the respective

torsional angles of 161.5, ꢀ36 and 808 prevail (ACADOS).^[29]We ob-

served similar but less marked differences between the ¹⁹F NMR

spectra of the regioisomeric 3’-deoxyfluoro analogues 9b (5’-trityl

ether, ꢁ83% South), 10b (5’-trityl ether, ꢁ98% South) and 11b

(5’-OH, ꢁ92% South) all in CDCl₃(cf. Supporting Information).

Azide 9b showed quite clearly a fairly resolved ddd multiplicity of

the F3’-signal at ꢀ195.1 ppm, which originates from three distinct

sured here we found vicinal F,H coupling constants ³J

29 Hz being up to three times higher than the highest vicinal H,H

coupling constant measured for 10b: ³J

(H,H)=9.3Hz (in other sys-

tems up to 15, max. 18 Hz). In other fluorinated compounds J(F,H)

N

AHCTREUNG

limiting values of between 29.5 and 47 Hz for close to anti orienta-

tions have been found.^[28]We are curious about a PSEUROT+J_H,F

analysis of our compounds, as well as that for arabinofuranose-con-

figured 2’-deoxyfluoronucleoside derivatives such as Clofarabine.^[32]

Furthermore, ¹⁹F resonances are far better sensors of diastereoiso-

meric forms than any other nucleus even though the fluorine atom

might be at quite a distance from the symmetry breaking element,

the phosphorous atom in compounds 13 and 14a in our case. Last

but not least, ¹⁹F resonances sense the spacial proximity of hydrogen

atoms, carbon atoms (through sometimes substantial long-range cou-

plings) and also nitrogen atoms. The quadrupole of the latter nu-

cleus (¹⁴N) may find its imprint in the very evident broadening of

the ¹⁹F resonance making it difficult or impossible to determine the

multiplicity in the fully coupled ¹⁹F NMR signal, as can be readily

seen for one of the two diastereoisomers of 13 and 14a each and in

all isomeric forms of the acyltriazenes 10c and 10d, but not so in

14b (see Supporting Information). A close spacial proximity be-

tween F2’ and NNN3’ or between F3’ and NNN2’ giving rise to line

broading of the ¹⁹F resonance is not surprising. We were surprised,

however, to see a line broading in only one of the diastereoisomers

of 13 and 14a and not the other, although the stereogenic centre is

phosphorous six covalent bonds away from F2’. We were somewhat

disappointed to see that the vicinal fluorine–carbon coupling con-

stants in almost all of our compounds were too small to be useful,

scalar F,H coupling constants, one geminal of ²J

F3’–H3’ and two different vicinals of ³J

(F,H)=23.5 Hz for F3’-H2’

A

AHCTREUNG

and 25.5 Hz for F3’-H4’. Amide 10b showed, like all amides, a

quintuplet of the F3’ signal at ꢀ191.7 ppm, which originates from

two distinct scalar F,H coupling constants, one geminal of twice the

value of one and the same vicinal of ³J

ACHTREUNG

AHCTREUNG

A

N

ACHTREUNG

values represent torsional angles, according to crystal structure coor-

dinates of several nucleos(t)ide South puckers, of between ꢀ154 and

ꢀ1808 (close to anti) for H2’b-C2’-C3’-F3’a and 13–348 (close to syn)

for H4’a-C4’-C3’-F3’a. The ¹⁹F NMR resonance of 11b, in contrast,

showed up as a dtd signal at ꢀ194.2 ppm which originates from

three distinct scalar F,H-coupling constants, one geminal of

²J(F,H)=54.6 Hz for F3’-H3’ (in the ¹H NMR spectrum 54.2 Hz),

T

one and the same vicinal coupling constant of ³J

F3’–H2’ and F3’–H4’ (in the H NMR spectrum 26.3and 28.1 Hz, re-

A

1

spectively), as well as an exceptional long range coupling of

⁴J

(F,H)=3.0 Hz for F3’-H5’ (not F’-H5’’, as revealed by H–D ex-

G

change in the ¹H NMR spectrum, cf. Supporting Information)! The

¹H resonance of 5’-OH shifted even more to lower fields and ap-

peared as a clearly resolved dd signal at 6.8 ppm (6.6 ppm for 5’-OH

of 11a and 12 in CDCl₃). Again, we interpret the additional F3’–H5’

long range coupling in the ¹⁹F and ¹H resonances (the only ob-

served) and the strong 5’-OH downfield shift in the ¹H NMR spec-

trum of 11b in CDCl₃with a stable 5’-OH···N3hydrogen bridge im-

possible for 9b or 10b. Since 9b, 10b and 11b all favor the pseu-

doaxial orientation of F3’a, thus, the South pucker, no large upfield

shift of the ¹⁹F resonance upon formation of the intramolecular hy-

drogen bond is expected. On the contrary, the ¹⁹F resonance of

azide 11b shifted downfield by 1.1 ppm with respect to azide 9b be-

cause its South pucker, thus, the pseudoaxial orientation of F3’a is

even higher populated than in 9b, as confirmed by a larger H1’–H2’

coupling constant in 11b (8.7 Hz) than in 9b (7.9 Hz). In conclusion,

not only are chemical shifts of ¹⁹F resonances extremely sensitive to

hydrogen bonding elsewhere in the molecule^[31]and to conforma-

tional changes. For instance, our observation from the analysis of vi-

³J

pling constants:

²J

(F,C)=14–36 Hz, j J

South-puckered 11b in CDCl₃the rigid conformation of which not

only produced long range fluorine–proton coupling of

⁴J vicinal F,C coupling of ³J-

(F3’a,H5’)=3.0 Hz but also

(F3’a,C5’)=11.6 Hz (in agreement with compound 20 in the Sup-

porting Information in ref. [23]), to the best of our knowledge, per-

haps the highest measured vicinal fluorine–carbon coupling con-

stant. According to the crystal structure coordinates ACADOS of

3’-O-acetyladenosine^[29]this value should correspond to a time-aver-

G

AHCTREUNG

aged torsional angle of 142.58. Apparently, South-puckered 3’-deox-

3

yfluororibofuranose derivatives produce J

A

9.1 and 11.6 Hz,^[23]while some 2’-deoxyfluoroarabino nucleosides

show

³J

(F2’b,C4’)=6 and 10 Hz, and one reveals a close spacial proximity

G

4

through a long-range J

A

tial ³J

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ꢁ 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Chem. Eur. J. 2007, 13, 5566 – 5584

2’-Deoxyfluoropuromycin Analogues

FULL PAPER

only other high-South compound of which we measured a ¹³C NMR

spectrum (not shown).

[23] I. A. Mikhailopulo, T. I. Pricota, G. G. Sivets, C. Altona, J. Org.

Chem. 2003, 68, 5897–5908.

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ACHTREUNG

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Todayꢀs credo seems to be (for some of us) that four-membered ring

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Received: January 14, 2007

Published online: April 23, 2007

5584

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Chem. Eur. J. 2007, 13, 5566 – 5584

Article Doi

First synthesis of 2'-deoxyfluoropuromycin analogues: Experimental insight into the mechanism of the staudinger reaction

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