Synthesis and structure-activity relationships of C-glycosylated oxadiazoles as inhibitors of glycogen phosphorylase

DOI: 10.1016/j.bmc.2009.04.036

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Bioorganic and Medicinal Chemistry p. 4773 - 4785 (2009)

Update date:2022-08-02

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Authors:

Toth, Marietta

Kun, Sandor

Bokor, Eva

Benltifa, Mahmoud

Tallec, Gaylord

Vidal, Sebastien

Docsa, Tibor

Gergely, Pal

Somsak, Laszlo

Praly, Jean-Pierre

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Article abstract of DOI:10.1016/j.bmc.2009.04.036

A series of per-O-benzoylated 5-β-d-glucopyranosyl-2-substituted-1,3,4-oxadiazoles was prepared by acylation of the corresponding 5-(β-d-glucopyranosyl)tetrazole. As an alternative, oxidation of 2,6-anhydro-aldose benzoylhydrazones by iodobenzene I,I-diac

Full text of DOI:10.1016/j.bmc.2009.04.036

Bioorganic & Medicinal Chemistry 17 (2009) 4773–4785

Contents lists available at ScienceDirect

Bioorganic & Medicinal Chemistry

journal homepage: w ww.elsevier.com/locate/bmc

Synthesis and structure–activity relationships of C-glycosylated oxadiazoles

as inhibitors of glycogen phosphorylase

Marietta Tóth ^a, Sándor Kun ^a, Éva Bokor ^a, Mahmoud Benltifa ^b,c,d,e, Gaylord Tallec ^b,c,d,e

b,c,d,e,

Sébastien Vidal ^b,c,d,e, Tibor Docsa ^f, Pál Gergely ^g, László Somsák ^a,, Jean-Pierre Praly

^aDepartment of Organic Chemistry, University of Debrecen, POB 20, H-4010 Debrecen, Hungary

^bUniversité de Lyon, Institut de Chimie et Biochimie Moléculaires et Supramoléculaires associé au CNRS, UMR 5246, Laboratoire de Chimie Organique 2, Bâtiment Curien,

43 boulevard du 11 Novembre 1918, F-69622 Villeurbanne, France

^cUniversité Lyon 1, F-69622 Villeurbanne, France

^dCNRS, UMR5246, Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Laboratoire de Chimie Organique 2, Bâtiment Curien,

43 boulevard du 11 Novembre 1918, F-69622 Villeurbanne, France

^eCPE-Lyon, F-69616 Villeurbanne, France

^fCell Biology and Signaling Research Group of the Hungarian Academy of Sciences at the Department of Medical Chemistry, Medical and Health Science Centre,

University of Debrecen, Egyetem tér 1, H-4032 Debrecen, Hungary

^gDepartment of Medical Chemistry, Medical and Health Science Centre, University of Debrecen, Egyetem tér 1, H-4032 Debrecen, Hungary

a r t i c l e i n f o

a b s t r a c t

Article history:

A series of per-O-benzoylated 5-b-

-glucopyranosyl-2-substituted-1,3,4-oxadiazoles was prepared by

Received 8 January 2009

Revised 15 April 2009

Accepted 17 April 2009

Available online 23 April 2009

acylation of the corresponding 5-(b-

D-glucopyranosyl)tetrazole. As an alternative, oxidation of 2,6-anhy-

dro-aldose benzoylhydrazones by iodobenzene I,I-diacetate afforded the same oxadiazoles. 1,3-Dipolar

cycloaddition of nitrile oxides to per-O-benzoylated b- -glucopyranosyl cyanide gave the corresponding

5-b- -glucopyranosyl-3-substituted-1,2,4-oxadiazoles. The O-benzoyl protecting groups were removed

by base-catalyzed transesteriﬁcation. The 1,3,4-oxadiazoles were practically inefﬁcient as inhibitors of

rabbit muscle glycogen phosphorylase b while the 1,2,4-oxadiazoles displayed inhibitory activities in

the micromolar range. The best inhibitors were the 5-b-D-glucopyranosyl-3-(4-methylphenyl- and -2-

Keywords:

C-Glycosyl compounds

1,3,4-Oxadiazoles

1,2,4-Oxadiazoles

Glycogen phosphorylase

Inhibitors

naphthyl)-1,2,4-oxadiazoles (K_i= 8.8 and 11.6

M, respectively). A detailed analysis of the structure–

activity relationships is presented.

1. Introduction

tion of glycogen, has been shown to be a target for the treatment

of T2DM.^14,15Since GP appears as the rate limiting enzyme of glyco-

gen degradation, its inhibition may offer a means for regulating

blood sugar levels.

The worldwide increasing prevalence of Type 2 Diabetes Mellitus

(T2DM) has become a major health problem for most of the world’s

population.¹Several oral hypoglycemic agents^2–4(sulfonylureas,

A number of inhibitors have been discovered and designed in

recent years targeting the different binding sites identiﬁed for

GP.^20,25Among these inhibitors, a large array of glucose derivatives

binds at the catalytic site of the enzyme.^20,25Among the ﬁrst efﬁ-

cient glucose analogs were the N-acyl-glucopyranosylamines, for

example, 1a,b,h,i (Chart 1), and further inhibitor design led to

biguanides, thiazolidinediones, a

-glucosidase inhibitors⁵) are now

being used to help diabetic patients to reduce hyperglycemia. Such

symptomatic treatments are intended to reach normal physiological

blood glucose levels. However, they have several undesirable side

effects and may also cause hypoglycemia.⁶These drugs are inade-

quate for 30–40% of patients.⁷Due to the appearance and spreading

of T2DM among young adults as well as children, the coming dec-

ades must face severe economic and health service burdens.^8–10

Fuelled by all these facts, several new therapeutic possibilities are

under investigation.^11–13Among them, liver glycogen phosphory-

lase (GP), an enzyme responsible for the phosphorylative degrada-

the discovery of more potent N-acyl-N⁰-b-

D-glucopyranosyl-ureas

like 2a,b,h,i. As another class of efﬁcient molecules, we developed

several inhibitors having C-glucosyl heterocyclic structural ele-

ments such as tetrazole 3, benzimidazole 4, benzothiazole 5,

1,3,4-oxadiazole^21,226a and 1,2,4-oxadiazoles 7a,b,i and 8b,^23,24

as well as hydroquinone derivatives.²⁶Several of these compounds

displayed an inhibition against rabbit muscle glycogen phosphory-

lase b (RMGPb) in the low micromolar range.

* Corresponding authors. Tel.: +36 52 512 900; fax: +36 52 453 836 (L.S.); fax:

+33 4 78 89 89 14 (J.-P.P.).

E-mail addresses: somsak@tigris.unideb.hu, somsak@tigris.klte.hu (L. Somsák),

jean-pierre.praly@univ-lyon1.fr (J.-P. Praly).

In the 3-b-

D-glucopyranosyl-5-substituted 1,2,4-oxadiazole

series, the 2-naphthyl derivative 7i was shown to be the best

doi:10.1016/j.bmc.2009.04.036

4774

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

1a (R = Me)

1b (R = Ph)

32¹⁶

81¹⁶

2a (R = Me)

2b (R = Ph)

305¹⁹

4.6¹⁹

144¹⁷

444¹⁸

10¹⁸

2h (R = 1-naphthyl)

2i (R = 2-naphthyl)

15²⁰

0.35²⁰

1h (R = 1-naphthyl)

1i (R = 2-naphthyl)

No inhibition²¹

11²¹

9²²

229²¹

76²²

7a (R = Me) no inhibition at 625 µM^c

7b (R = Ph) 10 % inhibition at 625 µM²³

7i (R = 2-naphthyl) 38²³

64²⁴

212²¹

145²²

Chart 1. Inhibition of rabbit muscle glycogen phosphorylase^a(RMGP) b (K_i[

M]) by selected glucose-based derivatives^b.

^aBecause of a ꢀ80% homology between the liver and muscle isoforms of GP, it is a general practice to use the more readily available RMGP for kinetic studies.

^bNumbering of molecules is based on the complete set of compounds presented in Table 1.^cUnpublished results.

inhibitor.²³This ﬁnding was similar to the observations made with

the N-acyl-b-

-glucopyranosylamines¹⁸1a,b,h,i and N-acyl-N⁰-b-

ent study, we have investigated the use of acyl chlorides in pyri-

dine or toluene, and also applied a DCC-mediated acylation by

the corresponding carboxylic acid³⁰(Scheme 1). The yields for

the acylation of tetrazole 10, obtained from 9 as earlier,²¹were

slightly higher in pyridine than in toluene, and the DCC coupling

proved less satisfactory to get oxadiazoles 11.

D-

glucopyranosyl-ureas^19,202a,b,h,i. Especially in the latter series of

compounds, the presence of a hydrophobic aromatic appendage

properly oriented (comparing 2h to 2i) proved beneﬁcial for the

strong binding to GP. This result was explained by the favorable

interactions of the inhibitor in the so-called b-channel¹⁹in the

vicinity of GP’s catalytic site which is an empty space surrounded

by amino-acid residues of mixed character.^14,20

A widely used method for the preparation of unsymmetrically

2,5-disubstituted-1,3,4-oxadiazoles is the oxidation of acylhydra-

zones by using a variety of oxidizing agents.^31–36This method

was extended to anhydro-aldose benzoylhydrazone 12 which

was obtained from 9 by our recently published method³⁷and oxi-

dized by iodobenzene I,I-diacetate (PIDA) to give oxadiazole 11b in

excellent yield (Scheme 1). The two alternative methods for getting

11b are similar in the number of synthetic steps as well as techni-

cal difﬁculties. However, route e–f gave higher overall yield as

compared to route a–b or a–d (Scheme 1). To show the applicabil-

ity of this oxidative method, anhydro-aldose benzoylhydrazones

14a–c⁴¹were also converted to the corresponding oxadiazoles

15a–c in good yields (Scheme 2). Removal of the O-acyl protecting

groups from oxadiazoles 11 was achieved by the Zemplén protocol

to give glucopyranosyl oxadiazoles 6 (Scheme 1). Raney-nickel cat-

alyzed reduction of the nitro group in 6e afforded amino com-

pound 6f.

Since the preliminary results obtained with the two other

isomeric oxadiazoles (6a and 8b) were encouraging with respect

to GP inhibition, we planned a broader study of structure–activity

relationships of C-glucosylated oxadiazoles. We present herein the

synthesis of 5-(b-D-glycopyranosyl) derivatives of 2-substituted-

1,3,4- and 3-substituted-1,2,4-oxadiazoles with a diverse set of

aromatic residues as well as a detailed comparative analysis of

structure–activity relationships of the different isomeric oxadiaz-

ole derivatives.

2. Results and discussion

The nature of the aryl substituent of the oxadiazole ring is inﬂu-

encing the binding to the enzyme’s catalytic site. We therefore pre-

pared a series of molecules incorporating either basic (p-amino),

acidic (p-hydroxy) or neutral (p-methyl, p-methoxy, p-nitro) sub-

stituents on the phenyl residues as well as larger aromatic moieties

(1- and 2-naphthyl). The syntheses of the target oxadiazoles were

2.2. Synthesis of 3-aryl-5-(b-D-glucopyranosyl)-1,2,4-

oxadiazoles²⁴

achieved using per-O-benzoylated b-D-glucopyranosyl cyanide 9 as

a common starting material.

1,3-Dipolar cycloaddition of nitrile oxides to nitriles provides an

efﬁcient and short route to diversely substituted 1,2,4-oxadiazoles.

The nitrile oxides can be easily obtained from the corresponding

hydroximoyl chloride precursors 16 which will undergo a dehydro-

halogenation in the presence of a base. The addition of the base must

occur in the presence of the dipolarophile in order to avoid the

formation of nitrile oxide dimers. Hydroximoyl chlorides are there-

fore synthesized in a two-step process from the corresponding alde-

2.1. Synthesis of 2-aryl-5-(D-glycopyranosyl)-1,3,4-oxadiazoles

C-Glycosyl-1,3,4-oxadiazoles were prepared earlier in various

sugar conﬁgurations by acylation of 5-( -glycosyl)tetrazoles with

an excess of acid chlorides or anhydrides,^27–29compound 6a being

the only b-

-glucopyranosyl derivative among them.²¹In the pres-

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4775

OBz

b, c or d

BzO

OBz

10 (94)

OBz

11b Ar = Ph (78^b, 56^d, 91^f) 6b Ar = Ph (91)

11c Ar = 4-Me-Ph (39^c)

11d Ar = 4-MeO-Ph (40^c)

11e Ar = 4-O₂N-Ph (63^c)

11h Ar = 1-Naphthyl (75^b)

11i Ar = 2-Naphthyl (62^b)

11l Ar = 4-AcO-Ph (41^d)

6c Ar = 4-Me-Ph (67)

6d Ar = 4-MeO-Ph (65)

6e Ar = 4-O₂N-Ph (76)

6f Ar = 4-H₂N-Ph (51)

6g Ar = 4-HO-Ph (45)

6h Ar = 1-Naphthyl (91)

6i Ar = 2-Naphthyl (96)

OBz

BzO

OBz

12 (90)

Scheme 1. Reagents and conditions: (a) NH₄N₃, DMF; (b) ArCOCl, pyridine, 90 °C; c) ArCOCl, toluene, reﬂux; (d) ArCOOH, DCC, toluene, reﬂux; (e) ArCONHNH₂, Raney-Ni,

NaH₂PO₂, AcOH, H₂O, pyridine, 40 °C; (f) PhI(OAc)₂, CH₂Cl₂, rt; (g) NaOMe, MeOH, rt; (h) Raney-Ni, H₂, MeOH, 60 °C. Yields (%) are indicated in parentheses.

(AcO)_n

13a-c

14a (86)

14b (58)

14c (84)

15a (78)

15b (67)

15c (57)

AcO

OAc

AcO

OAc

-Ara

AcO

a β-

OAc

-Xyl

OAc

-Gal

AcO

c α-D

b β-

Scheme 2. Reagents and conditions: (a) ArCONHNH₂, Raney-Ni, NaH₂PO₂, AcOH, H₂O, pyridine, 40 °C; b) PhI(OAc)₂, CH₂Cl₂, rt. Yields (%) are indicated in parentheses.

hydes by a reaction with hydroxylamine hydrochloride to afford the

oxime intermediates which are then treated with N-chlorosuccini-

mide to afford the desired hydroximoyl chlorides 16 (Scheme 3).

The puriﬁcation of the oxime intermediates was required in order

to obtain reproducible results and also to reach high purity of the

hydroximoyl chlorides since their puriﬁcation by standard column

chromatography was usually not successful due to their poor stabil-

ity. Glucosyl cyanide 9 was then reacted with the hydroximoyl chlo-

rides 16 in the presence of Et₃N at low concentration in reﬂuxing

toluene to afford the desired 1,2,4-oxadiazoles 17. The addition of

the base was achieved at a slow rate with a syringe pump. Subse-

quent transesteriﬁcation of the benzoate esters under Zemplén con-

ditions afforded the expected 1,2,4-oxadiazoles 8 in high yields. A

small proportion (4–8%) of 1,2-endo-glycals 18b–e,i were also

isolated while their formation could not be detected for other aryl

substituents. We previously observed that a substituent at the 5-po-

were madewith other ester protectedcarbohydrate derivativessuch

as glycosyl cyanides^29,38–40C-glycosyl thiadiazole⁴¹or benzothia-

zole.²⁹Interestingly, this elimination was not observed in the

1,3,4-oxadiazole series 11. Finally, the reduction of the p-nitro-

phenyl-substituted derivative 8e to the corresponding p-aminophe-

nyl oxadiazole 8f was achieved under standard hydrogenation

conditions (Pd–C 10%, H₂, 1 atm).

The p-benzyloxy-phenyl derivative 8j was initially prepared in

order to obtain the p-hydroxy-phenyl substituted oxadiazole 8g

through hydrogenolysis of the benzyl group. But, since the direct

access to 8g could be achieved using the general synthetic route

from

a-chloroarylaldoxime 16g, we therefore considered the

p-benzyloxy-phenyl derivative 8j as an additional candidate for

the inhibition of GP. The benzo[1,3]dioxol-5-yl substituted 1,2,4-

oxadiazole 8k was synthesized in order to evaluate its inhibition

towards the enzyme since the hydrophobic aromatic residue could

interact with the b-channel of GP and also since this aromatic res-

idue can be found as a pharmacophore in a series of natural prod-

ucts and synthetic drugs.

sition of the 1,2,4-oxadiazole ring possessing an acid-labile a-proton

would be susceptible of deprotonation or even b-elimination if a

leaving group is present at the b-position.²²Similar observations

OBz

BzO

OBz

17b Ar = Ph (94)

8b Ar = Ph (92)

OBz

18b Ar = Ph (8)

17c Ar = 4-Me-Ph (53)

17d Ar = 4-MeO-Ph (90)

17e Ar = 4-O₂N-Ph (99)

17g Ar = 4-HO-Ph (41)

17h Ar = 1-Naphthyl (55)

17i Ar = 2-Naphthyl (77)

17j Ar = 4-BnO-Ph (69)

8c Ar = 4-Me-Ph (92)

8d Ar = 4-MeO-Ph (75)

8e Ar = 4-O₂N-Ph (76)

8f Ar = 4-H₂N-Ph (99)

8g Ar = 4-HO-Ph (37)

18c Ar = 4-Me-Ph (8)

18d Ar = 4-MeO-Ph (5)

18e Ar = 4-O₂N-Ph (4)

18i Ar = 2-Naphthyl (8)

16b Ar = Ph (98)

16c Ar = 4-Me-Ph (76)

16d Ar = 4-MeO-Ph (78)

16e Ar = 4-O₂N-Ph (91)

16g Ar = 4-HO-Ph (39)

16h Ar = 1-Naphthyl (91)

16i Ar = 2-Naphthyl (64)

16j Ar = 4-BnO-Ph (51)

16k Ar = Benzo[1,3]dioxol-5-yl (81)

8h Ar = 1-Naphthyl (79)

8i Ar = 2-Naphthyl (76)

17k Ar = Benzo[1,3]dioxol-5-yl (74) 8j Ar = 4-BnO-Ph (56)

8k Ar = Benzo[1,3]dioxol-5-yl (60)

Scheme 3. Reagents and conditions: (a) NH₂OHꢁHCl, EtOH, NaOH, 70 °C then NCS, DMF, rt; (b) Et₃N, toluene, reﬂux; (c) NaOMe, MeOH, rt; (d) H₂, Pd–C, MeOH, rt. Yields (%)

are indicated in parentheses.

4776

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

2.3. Enzymatic evaluation of oxadiazoles as GP inhibitors

family displayed usually weak inhibition with the exception of

the acetyl derivative 1a with a K_ivalue of 32 M. The methyl

The kinetic parameters of the synthesized molecules were then

determined according to our previously described enzymatic pro-

tocol.²⁶We compared the inhibition properties of C-glucosylated

1,3,4-oxadiazoles 6 and 1,2,4-oxadiazoles 8 with the activity of

substituted oxadiazoles 6a and 7a and acetyl urea 2a showed poor

(6a, 2a) or no detectable (7a) inhibition.

The 3-C-glucosylated 1,2,4-oxadiazoles 7 with a large series of

aromatic substituents displayed the same inhibition pattern with

the 2-naphthyl derivative 7i as the best inhibitor in the series.

The phenyl substituted oxadiazole 7b was a poor inhibitor and

the p-nitrophenyl derivative 7e displayed the worst inhibition in

the series. This detrimental effect exerted by the nitro group was

not observed in the acyl urea series (compare 7e and 2e).

Nevertheless, the biological activity of these 3-C-glucosylated

1,2,4-oxadiazoles 7 was always weaker than the corresponding

regioisomeric 1,2,4-oxadiazoles 7, N-acyl-b-

ines 1 and N-acyl-N⁰-b-

-glucopyranosyl ureas 2 (Table 1).

In the N-acyl-N⁰-b-

-glucopyranosyl urea series 2, an overall

D-glucopyranosylam-

equivalent inhibition in the micromolar range was observed for

the benzoyl analogs (2b–g R = (p-substituted)phenyl). The 1-naph-

thoyl derivative 2h was a somewhat worse inhibitor of GP. A

simple change to the 2-naphthoyl moiety (2i) remarkably

improved the inhibition compared to 2h (43-fold increase) and

more moderately in comparison to the benzoyl derivative 2b

(13-fold increase). The 2-naphthoyl derivative 2i was the ﬁrst glu-

cose-based inhibitor with sub-micromolar activity against GP. In

N-acyl-N⁰-b-

D-glucopyranosyl ureas 2.

The 5-C-glucosylated 1,2,4-oxadiazoles 8 are generally better

inhibitors than the corresponding 3-C-glucosylated counterparts

7 and followed a similar trend with the 2-naphthyl derivative 8i

being among the best (displaying a value very close to the stron-

gest inhibitor 8c) and the p-nitrophenyl 8e the worst in the series.

The weak inhibition observed for the p-benzyloxy-phenyl deriva-

tive 8j might be attributed to the steric hindrance of the benzyl

the series of N-acyl-b-D-glucopyranosylamines 1, a similar obser-

vation could be made since the 2-naphthoyl derivative 1i was

the best inhibitor, found much more effective (44-fold) than 1h,

with a regioisomeric 1-naphthoyl residue. Other members of this

Table 1

Inhibition (K_ior IC₅₀

M)) of rabbit muscle glycogen phosphorylase b by N-acyl-b-D-glucopyranosylamines 1a–c,h,i, N-acyl-N⁰-b-D-glucopyranosyl ureas 2a–c,e–i, 2-(b-D-

glucopyranosyl)-5-substituted-1,3,4-oxadiazoles 6a–i, 3-(b-^D-glucopyranosyl)-5-substituted-1,2,4-oxadiazoles 7a–e,h,i and 5-(b-D-glucopyranosyl)-3-substituted-1,2,4-oxadiaz-

oles 8b–k

O N

ꢂCH₃

32¹⁶

305¹⁹

4.6¹⁹

2.3²⁰

—

212²¹

145²²

No inhibition at 625

—

81¹⁶

10% at 625

M²³

144¹⁷

4500

—

No inhibition at 625

350

8.8

20.4

550

OMe

NO₂

3.3²⁰

6.0²⁰

6.3²⁰

15²⁰

0.35²⁰

No inhibition at 625 lM

650

—

20% at 625

19.4

NH₂

—

444¹⁸

10¹⁸

—

10% at 625

No inhibition at 625

19.0

10% at 625

38²³

11.6^a

—

No inhibition at 625 lM

OBn

>625

—

A K_ivalue of 2.4 lM was measured independently by N. G. Oikonomakos and co-workers (unpublished results).

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4777

vdW

cyanide as a common starting material. Its transformation either

into the corresponding 5-(b- -glucopyranosyl)tetrazole and subse-

quent acylation or into anhydro-aldose acylhydrazones followed

by oxidation furnished 2-(b- -glucopyranosyl)-5-substituted-

1,3,4-oxadiazoles. 1,3-Dipolar cycloaddition of nitrile oxides to

the cyanide moiety afforded 5-(b- -glucopyranosyl)-3-substi-

tuted-1,2,4-oxadiazoles. The regioisomeric 3-(b- -glucopyrano-

β-channel

N⁴

₃N

H₂O

Protein

syl)-5-substituted-1,2,4-oxadiazoles were prepared previously by

ring closure of intermediate O-acyl-amidoximes.

β-channel

Enzyme kinetic evaluation of these inhibitors showed that the

nature of the oxadiazole ring attached to C-1 of glucopyranose is

strongly inﬂuencing the inhibitory activity towards RMGPb. While

the 1,3,4-oxadiazoles proved practically inactive, the 1,2,4-oxadiaz-

ole series displayed inhibition in the micromolar range with the 5-

glucosylated derivatives being superior to their 3-glucosylated

regioisomers. The nature of the p-substituent of the phenyl moiety

exhibited a similar inﬂuence on the inhibition of GP in each series.

In addition, the size and orientation of the aromatic substituent of

the oxadiazole strongly modulated the activity. The 2-naphthyl

derivatives in both 1,2,4-oxadiazole series were among the

best inhibitors. A possible explanation was proposed to understand

the differences in the inhibitory strengths of the isomeric

oxadiazoles.

₄N

H₂O

₄N

H₂O

Protein

Figure 1. Observed and probable orientations of the heterocyclic moieties of

C-glucosyl oxadiazole type inhibitors upon binding to the catalytic site of GP.

moiety in comparison to the naphthyl residue and also to the rota-

tion around the methylene benzylic group creating an entropy loss

for the binding process.

Very surprisingly, the 1,3,4-oxadiazoles 6 with a substituent

larger than methyl did not display any meaningful inhibition

against RMGPb. To explain this unexpected result, we propose to

consider the binding peculiarities of these molecules to the active

site of GP as revealed by X-ray crystallography. Because of the lack

of binding of the 1,3,4-oxadiazoles one has to go back to the struc-

ture of the enzyme–inhibitor complex obtained with the methyl-

ated analogue 6a, while an X-ray structure is available for the

enzyme-complex of the 2-naphthyl substituted 8i.²⁵

There are no direct H-bonds between the heterocycle of 6a and

the protein, however, an extensive H-bond network involving

nitrogen atoms N-3 and N-4 exists with the participation of water

molecules.²²As a result, 6a binds in the orientation shown in

Figure 1a. The compound can be accommodated at the catalytic

site with essentially no disturbance of the protein structure. The

relatively small methyl group makes 6 van der Waals interactions

while the b-channel remains unoccupied.

4. Experimental

4.1. General methods

Thin-layer chromatography (TLC) was carried out on alumi-

num sheets coated with Silica Gel 60 F₂₅₄(Merck). TLC plates

were inspected by UV light (k = 254 nm) and developed by treat-

ment with a mixture of 10% H₂SO₄in EtOH/H₂O (1/1 v/v) fol-

lowed by heating. Silica gel column chromatography was

performed with Geduran^ÒSilica Gel Si 60 (40–63

lm) purchased

from Merck (Darmstadt, Germany). Reversed-phase silica gel col-

umn chromatography was performed on a Varian Bond Elut C18

(20 mm, 15 mm diameter). ¹H and ¹³C NMR spectra were re-

corded at 23 °C using Bruker AC200, DRX300, WP 360 SY, or

DRX500 spectrometers with TMS or the residual solvent as the

internal standard. The following abbreviations are used to ex-

plain the observed multiplicities: s, singlet; d, doublet; dd, dou-

blet of doublet; ddd, doublet of doublet of doublet; t, triplet;

td, triplet of doublet; q, quadruplet; m, multiplet; br, broad; p,

pseudo. Structure elucidation was deduced from 1D and 2D

NMR spectroscopy which allowed, in most cases, complete signal

assignments based on COSY, HSQC and HMBC correlations.

Atoms in the pyranosyl rings are numbered by primed ﬁgures

and the aryl substituents with double-primed ﬁgures while those

of the oxadiazole ring have the oxa sufﬁx. NMR solvents were

purchased from Euriso-Top (Saint Aubin, France) or Sigma–Al-

drich. HRMS (LSIMS) mass spectra were recorded in the positive

mode using a Thermo Finnigan Mat 95 XL spectrometer. MS (ESI)

mass spectra were recorded in the positive mode using a Thermo

Finnigan LCQ spectrometer. Optical rotations were measured

using a Perkin Elmer polarimeter. Elemental analyses were per-

formed at the Service Central d’Analyses du CNRS (Vernaison,

France).

Similarly, no direct H-bonds could be observed between the

heterocycle of 5-glucosyl-1,2,4-oxadiazole 8i and the protein.²⁵

The nitrogen atom N-4 takes part in a H-bond network mediated

by water molecules. Due to the isomeric constitution of the het-

erocyclic part of this molecule as compared to 6a, the 3-substi-

tuent is in

a favorable position to make interactions with

residues surrounding the b-channel. The aromatic moiety makes

11 van der Waals contacts in the b-channel for the 2-naphthyl

derivative 8i. These interactions result in a strong binding with

the orientation of 8i represented by Figure 1d. For the isomeric

oxadiazoles 7 (Fig. 1c), a similar interaction pattern and orienta-

tion can be postulated. In the case of the 1,3,4-oxadiazole series

with large aromatic substituents (6), interactions in the b-chan-

nel would require a rotameric orientation of the heterocycle as

shown in Figure 1b. However, this conformation would ulti-

mately result in a loss of participation of N-3 and N-4 in the

H-bond network. In the other orientation (like in Fig. 1a), the

accommodation of large substituents in the vicinity of the cata-

lytic site would most probably signiﬁcantly disturb the protein

structure. These factors together prevent compounds 6 from

strong binding to GP.

Method A: To a solution of tetrazole 10 (700 mg, 1.08 mmol) in

abs. pyridine (10 mL) the corresponding acid chloride (5.40 mmol)

was added. The reaction mixture was stirred at 90 °C for 1 h. It was

then cooled to rt, water was added, and extracted with CH₂Cl₂

(3 ꢃ 40 mL). The combined organic phase was washed with satu-

rated aqueous NaHCO₃solution and water. The organic phase

was dried (MgSO₄), concentrated under diminished pressure and

3. Conclusion

The synthesis of three series of C-glycosylated oxadiazoles

could be achieved from per-O-benzoylated b-D-glucopyranosyl

4778

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

the crude product was puriﬁed by column chromatography (hex-

ane–EtOAc 3:1) and then crystallized from EtOH.

4.2. Synthesis of 5-aryl-2-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,3,4-oxadiazoles

Method B: Perbenzoylated glucopyranosyl-tetrazole 10 was dis-

solved in abs. toluene (100 mg/mL) then acid chloride (1.2–

2 equiv) was added in one portion. The mixture was reﬂuxed and

monitored by TLC. After disappearance of the starting material

the solvent was removed under diminished pressure, the residue

was puriﬁed by column chromatography if necessary and crystal-

lized from EtOH.

Method C: An anhydro-aldose benzoylhydrazone (12 or 14,

0.25 mmol) was dissolved in CH₂Cl₂(2.4 mL) and PIDA

(159 mg, 0.50 mmol) was added. The reaction mixture was stir-

red at rt. When the reaction was complete (TLC: EtOAc/hexane,

1:1), the solution was diluted with CH₂Cl₂(4 mL) and water

(2 mL). The aqueous layer was extracted with CH₂Cl₂

(3 ꢃ 4 mL), then the combined organic layers were washed with

cold saturated NaHCO₃solution (1 ꢃ 5 mL), dried (Na₂SO₄) and

ﬁltered. The solvent was removed by evaporation under dimin-

ished pressure and the residue puriﬁed by column chromatogra-

phy (EtOAc/hexane, 1:2).

4.2.1. 5-(b-D-Glucopyranosyl)-2-phenyl-1,3,4-oxadiazole (6b)

Prepared according to method G. Yield: 91% (white crystals).

Mp = 165–167 °C. ½a _D²⁰

ꢄ

¼ þ33 (c 0.2, MeOH). ¹H NMR (360 MHz,

CD₃OD): d = 8.04-7.52 (m, 5H, H-ar) 4.61 (d, 1H, J = 10.6 Hz, H-

1⁰), 3.87 (pdd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a), 3.78 (t, 1H,

J = 9.3 Hz, H-2⁰or H-3⁰or H-4⁰), 3.68 (dd, 1H, J = 5.3 Hz,

J = 11.9 Hz, H-6⁰b), 3.53-3.40 (m, 3H, H-2⁰or H-3⁰or H-4⁰and H-

5⁰). ¹³C NMR (90 MHz, CD₃OD): d = 166.9, 165.6 (C-2oxa, C-5oxa),

133.5, 130.5, 130.5, 128.1, 128.1, 124.7, 82.9 (C-1⁰), 79.1, 74.7,

73.5, 71.3 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.7 (C-6⁰). Anal. Calcd for

C₁₄H₁₆N₂O₆(308.29): C, 54.54; H, 5.23; N, 9.09. Found: C, 54.27;

H, 5.30; N, 8.94.

4.2.2. 5-(b-D-Glucopyranosyl)-2-(4-methylphenyl)-1,3,4-

oxadiazole (6c)

Prepared according to method G. Yield: 83 mg, 67% (white crys-

tals). Mp = 185–188 °C.

a ²_D⁰

ꢄ

¼ þ19 (c 0.14, H₂O). ¹H NMR

Method D: A solution of arylaldehyde (40 mmol), hydroxyl-

amine hydrochloride (80 mmol, 2 equiv) and sodium hydroxide

(80 mmol, 2 equiv) in EtOH (50 mL) was stirred at 78 °C for 2 h.

The suspension was ﬁltered and the solid washed with EtOH

(2 ꢃ 20 mL). The ﬁltrate was evaporated and the residue dissolved

in EtOAc (150 mL). The organic layer was washed with water

(3 ꢃ 70 mL), dried (Na₂SO₄), ﬁltered and evaporated to dryness.

A portion of the crude arylaldoxime (7 mmol) was then dissolved

in DMF (10 mL) and NCS (7 mmol, 1 equiv) was added in 8–10

portions. The reaction was slightly exothermic and NCS portions

were added slowly in order to maintain the temperature at 35–

40 °C. If no heat was generated after the ﬁrst two additions of

NCS, a stream of HCl (generated ex situ from NaCl and H₂SO₄)

was bubbled through the solution in order to start the reaction

then stopped when the temperature reached 35–40 °C. The reac-

tion was then stirred for 3 h at rt and poured into EtOAc

(100 mL). The organic layer was washed with water (3 ꢃ 50 mL),

dried (Na₂SO₄), ﬁltered and evaporated to dryness. The hydroxi-

moyl chlorides 16b–e,g–k were used for cycloadditions without

further puriﬁcation.

Method E: A solution of glucosyl cyanide 9 (0.5 mmol) and a

hydroximoyl chloride (2.5 mmol, 5 equiv) in toluene (5 mL) was

stirred at 110 °C under argon. Triethylamine (3.75 mmol,

7.5 equiv) was dissolved in toluene (5 mL) and slowly added in

12 h with a syringe pump. The reaction was stirred at 110 °C

for an additional 12 h then the solvent was evaporated under

diminished pressure. The residue was puriﬁed by ﬂash silica

gel column chromatography to afford the desired 1,2,4-oxadiaz-

oles 17.

(360 MHz, D₂O): d = 7.77 (d, 2H, J = 7.9 Hz, H-ar), 7.31 (d, 2H,

J = 7.9 Hz, H-ar), 4.72 (d, 1H, J = 10.6 Hz, H-1⁰), 3.92 (pdd, 1H,

J < 1 Hz, J = 11.9, H-6⁰a), 3.86-3.55 (m, 5H, H-2⁰, H-3⁰, H-4⁰, H-5⁰,

H-6⁰b), 2.34 (s, 3H, PhCH₃). ¹³C NMR (90 MHz, D₂O): d = 166.8,

164.0 (C-2oxa, C-5oxa), 144.7, 130.5, 127.5, 119.9, 81.2 (C-1⁰),

77.3, 73.2, 72.2, 69.9 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 61.3 (C-6⁰), 21.4

(PhCH₃). Anal. Calcd for C₁₅H₁₈N₂O₆(322.32): C, 55.90; H, 5.63;

N, 8.69. Found: C, 55.82; H, 5.55; N, 8.61.

4.2.3. 5-(b-D-Glucopyranosyl)-2-(4-methoxyphenyl)-1,3,4-

oxadiazole (6d)

Prepared according to method G. Yield: 65% (yellowish amor-

phous solid). ½a _D²⁰

ꢄ

¼ þ15 (c 0.16, H₂O). ¹H NMR (360 MHz, D₂O):

d = 7.68 (d, 2H, J = 7.9 Hz, H-ar), 6.89 (d, 2H, J = 7.9 Hz, H-ar), 4.71

(d, 1H, J = 9.2 Hz, H-1⁰), 3.95 (pdd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a),

3.86-3.81 (m, 5H, H-2⁰, H-6⁰b, OMe), 3.69-3.64 (m, 2H, H-3⁰, H-

5⁰), 3.57 (t, 1H, J = 9.2 Hz, H-4⁰). ¹³C NMR (90 MHz, D₂O):

d = 166.3, 163.8 (C-2oxa, C-5oxa), 129.4-115.1 (CH-ar, C^IV-ar),

81.2 (C-1⁰), 77.5, 73.2, 72.3, 69.9 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 61.4 (C-

6⁰), 56.1 (OMe). Anal. Calcd for C₁₅H₁₈N₂O₇(338.32): C, 53.25; H,

5.36; N, 8.28. Found: C, 53.19; H, 5.30; N, 8.35.

4.2.4. 5-(b-D-Glucopyranosyl)-2-(4-nitrophenyl)-1,3,4-

oxadiazole (6e)

Prepared according to method G. Puriﬁed by column chroma-

tography (CHCl₃/MeOH, 9:1) Yield: 76% (yellow crystals).

Mp = 124–126 °C. ½a _D²⁰

ꢄ

¼ þ22 (c 0.52, DMSO). ¹H NMR (360 MHz,

CD₃SOCD₃): d = 8.44 (d, 2H, J = 9.3 Hz, H-ar), 8.30 (d, 2H,

J = 9.3 Hz, H-ar), 5.44 (d, 1H, J = 5.3 Hz, OH), 5.24 (d, 1H,

J = 4.0 Hz, OH), 5.14 (d, 1H, J = 5.3 Hz, OH), 4.63–4.58 (m, 2H, H-

1⁰, OH), 3.74–3.37 (m, 6H, H-2⁰, H-3⁰, H-4⁰, H-5⁰, H-6⁰a, H-6⁰b). ¹³C

NMR (90 MHz, CD₃SOCD₃): d = 164.8, 163.1 (C-2oxa, C-5oxa),

149.3, 128.6, 128.1, 124.7, 81.9 (C-1⁰), 77.2, 72.7, 71.8, 69.9 (C-2⁰,

C-3⁰, C-4⁰, C-5⁰), 61.0 (C-6⁰). Anal. Calcd for C₁₄H₁₅N₃O₈(353.29):

C, 47.60; H, 4.28; N, 11.89. Found: C, 47.69; H, 4.34; N, 11.80.

Method F: A solution of the acyl-protected carbohydrate deriva-

tives 17 (0.15 mmol) and NaOMe (50 lL, 1 M in MeOH) in MeOH

(3 mL) was stirred at rt for 3 h. The solution was then neutralized

to pH 5 with a cation exchange resin (DOWEX 50WX2, H⁺form).

The resin was ﬁltered off and washed with MeOH (3 ꢃ 10 mL) then

the ﬁltrate was evaporated under diminished pressure. The residue

was puriﬁed by ﬂash silica gel column chromatography to afford

the desired carbohydrate derivatives 8.

4.2.5. 2-(4-Aminophenyl)-5-(b-D-glucopyranosyl)-1,3,4-

Method G: The benzoylated compounds 11 were dissolved in a

mixture of abs. MeOH and abs. CHCl₃and 1 M methanolic sodium

methoxide solution was added to the solutions in catalytic amount.

The reaction mixture was kept at room temperature for a given

time and then neutralized with a cation exchange resin Amberlyst

15 (H⁺form). After ﬁltration and removal of the solvent under

diminished pressure, the crude product was puriﬁed by crystalliza-

tion from Et₂O or by column chromatography to afford the depro-

tected carbohydrate derivatives 6.

oxadiazole (6f)

Compound 6e (60 mg, 0.17 mmol) was dissolved in a mixture of

abs. EtOAc (2 mL) and abs. MeOH (2 mL), and reduced by H₂

(1 atm) using Raney-Ni as catalyst at 60 °C for 8 h. After ﬁltration

and evaporation of the solvents the residue was puriﬁed by col-

umn chromatography (CHCl₃/MeOH, 4:1) Yield: 28 mg, 51% (yel-

lowish crystals). Mp = 215–217 °C. ½a _D²⁰

ꢄ

¼ þ16 (c 0.12, DMSO). ¹H

NMR (360 MHz, CD₃SOCD₃): d = 7.68 (d, 2H, J = 7.9 Hz, H-ar), 6.70

(d, 2H, J = 7.9 Hz, H-ar), 5.98 (s, 2H, NH₂), 5.39 (d, 1H, J = 5.3 Hz,

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4779

OH), 5.21 (d, 1H, J = 5.3 Hz, OH), 5.13 (d, 1H, J = 5.3 Hz, OH), 4.64

(dd, 1H, J = 6.6 Hz, J = 5.3 Hz, OH), 4.47 (d, 1H, J = 9.3 Hz, H-1⁰),

3.74-2.20 (m, 6H, H-2⁰, H-3⁰, H-4⁰, H-5⁰, H-6⁰a, H-6⁰b). ¹H NMR

(360 MHz, CD₃OD): d = 7.74 (d, 2H, J = 9.3 Hz, H-ar), 6.74 (d, 2H,

J = 9.3 Hz, H-ar), 4.57 (d, 1H, J = 9.3 Hz, H-1⁰), 3.89 (pdd, 1H,

J < 1 Hz, J = 10.6 Hz, H-6⁰a), 3.79 (t, 1H, J = 9.3 Hz, H-2⁰), 3.70 (dd,

1H, J = 4.0 Hz, J = 11.9 Hz, H-6⁰b), 3.54-3.34 (m, 3H, H-3⁰, H-4⁰,

H-5⁰). ¹³C NMR (90 MHz, CD₃OD): d = 167.8, 164.3 (C-2oxa, C-

5oxa), 154.1, 129.7, 115.2, 111.7, 82.9 (C-1⁰), 79.2, 74.7, 73.4,

71.3 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.7 (C-6⁰). Anal. Calcd for C₁₄H₁₇N₃O₆

(323.31): C, 52.01; H, 5.30; N, 13.00. Found: C, 50.94; H, 5.22; N,

13.10.

(LSIMS, glycerol) m/z = 309 [M+H]⁺. HRMS (LSIMS, glycerol) m/

z = C₁₄H₁₇N₂O₆[M+H]⁺calcd 309.1087, found 309.1088. Analytical

data for 18b: R_f= 0.51 (EtOAc/MeOH, 9:1). ¹H NMR (300 MHz,

CD₃OD): d = 8.10–7.96 (m, 5H, H-ar), 6.13 (d, 1H, J = 2.8 Hz, H-2⁰),

4.34 (dd, 1H, J = 2.8 Hz, J = 7.2 Hz, H-3⁰), 4.06 (ddd, 1H, J = 2.3 Hz,

J = 5.2 Hz, J = 9.5 Hz, H-5⁰), 4.01 (dd, 1H, J = 2.3 Hz, J = 12.6 Hz, H-

6⁰a), 3.93 (dd, 1H, J = 5.2 Hz, J = 12.6 Hz, H-6⁰b), 3.75 (dd,

J = 7.2 Hz, J = 9.5 Hz, H-4⁰). ¹³C NMR (75 MHz, CD₃OD): d = 172.9

(C-5oxa), 169.8 (C-3oxa), 141.1 (C-1⁰), 132.7, 130.7, 128.4, 127.7

(C^IV-ar), 112.9 (C-2⁰), 82.3 (C-5⁰), 70.2 (C-3⁰), 69.7 (C-4⁰), 61.9 (C-

6⁰). MS (LSIMS, glycerol) m/z = 291 [M+H]⁺. HRMS (LSIMS, glycerol)

m/z = C₁₄H₁₅N₂O₅[M+H]⁺calcd 291.0981, found 291.0981.

4.2.6. 5-(b-

-Glucopyranosyl)-2-(4-hydroxyphenyl)-1,3,4-

4.3.2. 5-(b-

D-Glucopyranosyl)-3-(4-methylphenyl)-1,2,4-oxadi-

oxadiazole (6g)

azole (8c) and 5-(2-deoxy-^D–D

-arabino-hex-1-enopyranosyl)-3-

Prepared according to method G. Yield: 45% (yellowish amor-

(4-methylphenyl)-1,2,4-oxadiazole (18c)

phous solid). ½a _D²⁰

ꢄ

¼ þ20 (c 0.2, MeOH). ¹H NMR (360 MHz, D₂O):

A solution of 17c (250 mg, 0.38 mmol) was treated according to

method F. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 3:2 then EtOAc then EtOAc/MeOH 9:1) to

afford 8c (103 mg, 92%) as a colorless oil and 18c (11 mg, 8%) as

a colorless oil. Analytical data for 8c: R_f= 0.38 (EtOAc/MeOH,

d = 7.84 (d, 2H, J = 7.9 Hz, H-ar), 6.97 (d, 2H, J = 7.9 Hz, H-ar), 3.98

(pdd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a), 3.91-3.79 (m, 3H, H-1⁰, H-2⁰,

H-6⁰b), 3.73-3.68 (m, 2H, H-3⁰, H-5⁰), 3.60 (t, 1H, J = 9.2 Hz, H-4⁰).

¹³C NMR (90 MHz, D₂O): d = 163.7, 160.4 (C-2oxa, C-5oxa), 129.9-

114.9 (CH-ar, C^IV-ar), 81.1 (C-1⁰), 77.3, 73.2, 72.2, 69.9 (C-2⁰, C-3⁰,

C-4⁰, C-5⁰), 61.4 (C-6⁰). Anal. Calcd for C₁₄H₁₆N₂O₇(324.29): C,

51.85; H, 4.97; N, 8.64. Found: C, 51.79; H, 4.91; N, 8.71.

9:1).

a ²_D⁰

ꢄ

¼ þ14 (c 1, MeOH). ¹H NMR (300 MHz, CD₃OD):

d = 7.74 (d, 2H, J = 8.1 Hz, H-ar), 7.34 (d, 2H, J = 8.1 Hz, H-ar), 4.64

(d, 1H, J = 9.8 Hz, H-1⁰), 3.90 (pdd, 1H, J < 1 Hz, J = 12.3 Hz, H-6⁰a),

3.78 (dd, 1H, J = 8.6 Hz, J = 9.8 Hz, H-2⁰), 3.71 (dd, 1H, J = 5.3 Hz,

J = 12.3 Hz, H-6⁰b), 3.55-3.45 (m, 3H, H-3⁰, H-4⁰, H-5⁰), 2.41 (s, 3H,

CH₃Ph). ¹³C NMR (75 MHz, CD₃OD): d = 178.1 (C-5oxa), 169.6 (C-

3oxa), 143.3 (C-1⁰⁰), 130.8 (2C, C-2⁰⁰, C-6⁰⁰), 128.4 (2C, C-3⁰⁰, C-5⁰⁰),

125.0 (C-4⁰⁰), 83.0 (C-5⁰), 79.2 (C-3⁰), 75.2 (C-1⁰), 74.0 (C-2⁰), 71.3

(C-4⁰), 62.8 (C-6⁰), 21.6 (CH₃Ph). MS (ESI) m/z = 323.0 [M+H]⁺,

345.1 [M+Na]⁺, 666.9 [2 M+Na]⁺. HRMS (ESI) m/z = C₁₅H₁₉N₂O₆

[M+H]⁺calcd 323.1243, found 323.1245. Analytical data for 18c:

R_f= 0.38 (EtOAc/MeOH, 9:1). ¹H NMR (300 MHz, CD₃OD): d = 7.94

(d, 2H, J = 8.1 Hz, H-ar), 7.34 (d, 2H, J = 8.1 Hz, H-ar), 6.12 (d, 1H,

J = 2.8 Hz, H-2⁰), 4.33 (dd, 1H, J = 2.8 Hz, J = 7.2 Hz, H-3⁰), 4.06

(ddd, 1H, J = 2.6 Hz, J = 5.4 Hz, J = 9.5 Hz, H-5⁰), 4.01 (m, 1H, H-

6⁰a), 3.94 (dd, 1H, J = 5.4 Hz, J = 12.6 Hz, H-6⁰b), 3.75 (dd,

J = 7.2 Hz, J = 9.5 Hz, H-4⁰), 2.41 (s, 3H, CH₃Ph). ¹³C NMR (75 MHz,

CD₃OD): d = 172.8 (C-5oxa), 169.9 (C-3oxa), 143.4 (C-1⁰), 141.2,

130.8 (2C), 128.4 (2C), 124.9 (C^IV-ar), 112.8 (C-2⁰), 82.3 (C-5⁰),

70.3 (C-3⁰), 69.8 (C-4⁰), 62.0 (C-6⁰), 21.6 (CH₃Ph). MS (ESI) m/

z = 304.8 [M+H]⁺, 327.0 [M+Na]⁺, 630.8 [2M+Na]⁺. HRMS (ESI) m/

z = C₁₅H₁₇N₂O₅[M+H]⁺calcd 305.1137, found 305.1136.

4.2.7. 5-(b-D-Glucopyranosyl)-2-(1-naphthyl)-1,3,4-oxadiazole (6h)

Prepared according to method G. Yield: 91% (colorless syrup).

a ²_D⁰

ꢄ

¼ þ35 (c 0.2, MeOH). ¹H NMR (360 MHz, CD₃OD): d = 8.96–

7.50 (m, 7H, H-ar), 4.69 (d, 1H, J = 10.6 Hz, H-1⁰), 3.91–3.82 (m,

2H, H-2⁰or H-3⁰or H-4⁰and H-6⁰a), 3.70 (dd, 1H, J = 5.3 Hz,

J = 11.9 Hz, H-6⁰b), 3.57–3.44 (m, 3H, H-2⁰or H-3⁰or H-4⁰and H-

5⁰). ¹³C NMR (90 MHz, CD₃OD): d = 166.7, 165.3 (C-2oxa, C-5oxa),

135.3, 134.1, 131.2, 130.0, 130.0, 129.2, 127.9, 126.6, 126.1,

121.1, 82.9 (C-1⁰), 79.1, 74.7, 73.6, 71.3 (C-2⁰, C-3⁰, C-4⁰, C-5⁰),

62.7 (C-6⁰). Anal. Calcd for C₁₈H₁₈N₂O₆(358.35): C, 60.33; H,

5.06; N, 7.82. Found: C, 60.22; H, 5.30; N, 7.98.

4.2.8. 5-(b-D-Glucopyranosyl)-2-(2-naphthyl)-1,3,4-oxadiazole (6i)

Prepared according to method G. Yield: 96% (white crystals).

Mp = 219–221 °C. ½a _D²⁰

ꢄ

¼ ꢂ5 (c 0.1, DMSO). ¹H NMR (360 MHz,

CD₃SOCD₃): d = 8.66–7.66 (m, 7H, H-ar), 5.49 (d, 1H, J = 9.2 Hz, H-

1⁰), 3.72–3.16 (m, 6H, H-2⁰, H-3⁰, H-4⁰, H-5⁰, H-6⁰a, H-6⁰b). ¹³C

NMR (90 MHz, CD₃SOCD₃): d = 164.6, 164.0 (C-2oxa, C-5oxa),

134.2, 132.4, 129.3, 128.9, 128.3, 127.9, 127.3, 127.1, 122.8,

120.4, 81.9 (C-1⁰), 77.3, 72.7, 71.7, 69.9 (C-2⁰, C-3⁰, C-4⁰, C-5⁰),

61.0 (C-6⁰). Anal. Calcd for C₁₈H₁₈N₂O₆(358.35): C, 60.33; H,

5.06; N, 7.82. Found: C, 60.23; H, 5.18; N, 7.65.

4.3.3. 5-(b-D-Glucopyranosyl)-3-(4-methoxyphenyl)-1,2,4-oxadi-

azole (8d) and 5-(2-deoxy-

-arabino-hex-1-enopyranosyl)-3-(4-

methoxyphenyl)-1,2,4-oxadiazole (18d)

A solution of 17d (187 mg, 0.25 mmol) was treated according to

method F. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 3:2 then EtOAc then EtOAc/MeOH 9:1) to

afford 8d (62 mg, 75%) as a white solid and 18d (3 mg, 5%) as a pale

yellow solid. Analytical data for 8d: R_f= 0.08 (EtOAc). Mp = 152–

4.3. Synthesis of 3-aryl-5-(b-D-glucopyranosyl)-1,3,4-oxadiazoles

4.3.1. 5-(b-

and 5-(2-deoxy-

oxadiazole (18b)

-Glucopyranosyl)-3-phenyl-1,2,4-oxadiazole (8b)

D-arabino-hex-1-enopyranosyl)-3-phenyl-1,2,4-

153 °C (MeOH/hexane). ½a _D²⁰

ꢄ

¼ þ16 (c 0.75, MeOH). ¹H NMR

A solution of 17b (155 mg, 0.21 mmol) was treated according to

method F. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 3:2 then EtOAc then EtOAc/MeOH 9:1) to

afford 8b (67 mg, 92%) as a yellow solid and 18b (5 mg, 8%) as a

yellow solid. Analytical data for 8b: R_f= 0.41 (EtOAc /MeOH, 9:1).

(300 MHz, CD₃OD): d = 8.00 (d, 2H, J = 8.9 Hz, H-ar), 7.05 (d, 2H,

J = 8.9 Hz, H-ar), 4.63 (d, 1H, J = 9.6 Hz, H-1⁰), 3.91 (pdd, 1H,

J < 1 Hz, J = 12.3 Hz, H-6⁰a), 3.86 (s, 3H, OCH₃), 3.79 (dd, 1H,

J = 8.8 Hz, J = 9.6 Hz, H-2⁰), 3.72 (dd, 1H, J = 4.8 Hz, J = 12.3 Hz, H-

6⁰b), 3.50-3.43 (m, 3H, H-3⁰, H-4⁰, H-5⁰). ¹³C NMR (75 MHz, CD₃OD):

d = 177.9 (C-5oxa), 169.3 (C-3oxa), 163.8 (C-4⁰⁰), 130.1 (2C), 120.0

(C^IV-ar), 115.2 (2C), 83.0 (C-5), 79.2 (C-3⁰), 75.2 (C-1⁰), 74.0 (C-2⁰),

71.2 (C-4⁰), 62.8 (C-6⁰), 56.0 (OCH₃). MS (LSIMS, glycerol) m/z = 339

[M+H]⁺. HRMS (LSIMS, glycerol) m/z = C₁₅H₁₉N₂O₇[M+H]⁺calcd

339.1192, found 339.1191. Analytical data for 18d: R_f= 0.17

(EtOAc). ¹H NMR (500 MHz, CD₃OD): d = 8.00 (d, 2H, J = 8.9 Hz, H-

ar), 7.07 (d, 2H, J = 8.9 Hz, H-ar), 6.11 (d, 1H, 1H, J = 2.8 Hz, H-2⁰),

4.34 (dd, 1H, J = 2.8 Hz, J = 7.3 Hz, H-3⁰), 4.06 (ddd, 1H, J = 2.3 Hz,

Mp = 129–130 °C (MeOH/hexane). ½a _D²⁰

ꢄ

¼ þ16 (c 1, MeOH). ¹H

NMR (300 MHz, CD₃OD): d = 8.10–8.06 (m, 2H, H-ar), 7.55–7.50

(m, 3H, H-ar), 4.66 (d, 1H, J = 9.8 Hz, H-1⁰), 3.91 (pdd, 1H, J < 1 Hz,

J = 12.2 Hz, H-6⁰a), 3.80 (dd, 1H, J = 8.5 Hz, J = 9.8 Hz, H-2⁰), 3.72

(dd, 1H, J = 4.5 Hz, J = 12.2 Hz, H-6⁰b), 3.50–3.40 (m, 3H, H-3⁰, H-

4⁰, H-5⁰). ¹³C NMR (75 MHz, CD₃OD): d = 178.2 (C-5oxa), 169.6 (C-

3oxa), 132.6, 130.1 (2C), 128.4 (2C), 127.8 (C^IV-ar), 83.0 (C-5⁰),

79.2 (C-3⁰), 75.1 (C-1⁰), 74.0 (C-2⁰), 71.2 (C-4⁰), 62.7 (C-6⁰). MS

4780

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

J = 5.4 Hz, J = 9.5 Hz, H-5⁰), 4.03 (dd, 1H, J = 2.3 Hz, J = 12.7 Hz, H-6⁰a),

3.93 (dd, 1H, J = 5.4 Hz, J = 12.7 Hz, H-6⁰b), 3.87 (s, 3H, OCH₃), 3.74

(dd, J = 7.3 Hz, J = 9.5 Hz, H-4⁰). ¹³C NMR (125 MHz, CD₃OD):

d = 171.6 (C-5oxa), 168.5 (C-3oxa), 162.9 (C-4⁰⁰), 140.1 (C-1⁰), 129.0

(2C), 118.8 (C^IV-ar), 114.5 (2C), 111.7 (C-2⁰), 81.2 (C-5⁰), 69.2 (C-3⁰),

68.7 (C-4⁰), 60.9 (C-6⁰), 54.9 (OCH₃). MS (LSIMS, glycerol) m/z = 321

[M+H]⁺. HRMS (LSIMS, glycerol) m/z = C₁₅H₁₇N₂O₆[M+H]⁺calcd

321.1087, found 321.1087.

6.88 (d, 2H, J = 8.7 Hz, H-3⁰⁰, H-5⁰⁰), 4.60 (d, 1H, J = 9.9 Hz, H-1⁰),

3.88 (pdd, 1H, J < 1 Hz, J = 11.7 Hz, H-6⁰a), 3.77 (dd, 1H, J = 8.7 Hz,

J = 9.9 Hz, H-2⁰), 3.70 (m, 1H, H-6⁰b), 3.54–3.44 (m, 3H, H-3⁰, H-4⁰,

H-5⁰). ¹³C NMR (75 MHz, CD₃OD): d = 176.7 (C-5oxa), 168.4 (C-

3oxa), 160.8 (C-4⁰⁰), 129.1 (C-2⁰⁰, C-6⁰⁰), 117.7 (C-1⁰⁰), 115.8 (C-3⁰⁰,

C-5⁰⁰), 74.1 (C-1⁰), 72.9 (C-2⁰), 81.9, 78.1, 70.2 (C-3⁰, C-4⁰, C-5⁰),

61.7 (C-6⁰). MS (ESI) m/z = 347.0 [M+Na]⁺. HRMS (ESI) m/z =

C₁₄H₁₆N₂O₇[M+Na]⁺calcd 347.0855, found 347.0857.

4.3.4. 5-(b-D-Glucopyranosyl)-3-(4-nitrophenyl)-1,2,4-oxadiazole

4.3.7. 5-(b-D-Glucopyranosyl)-3-(1-naphthyl)-1,2,4-oxadiazole (8h)

(8e) and 5-(2-deoxy-

phenyl)-1,2,4-oxadiazole (18e)

-arabino-hex-1-enopyranosyl)-3-(4-nitro-

A solution of 17h (388 mg, 0.43 mmol) and NaOMe (5 mg) in

MeOH/CH₂Cl₂(10 mL, 9:1) was stirred at rt for 24 h. The reaction

was neutralized to pH 5–6 with Amberlite IR-120 resin (H⁺form)

and the resin washed with MeOH (3 ꢃ 10 mL). The ﬁltrate was

evaporated under diminished pressure and the residue was puri-

ﬁed by ﬂash silica gel column chromatography (PE/EtOAc, 1:1 then

EtOAc then EtOAc/MeOH 95:5) to afford 8h (125 mg, 79%) as a

white solid. R_f= 0.45 (EtOAc/MeOH, 9:1). Mp = 182–183 °C.

A solution of 17e (258 mg, 0.34 mmol) was treated according to

method F. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 3:2 then EtOAc then EtOAc/MeOH 9:1) to

afford 8e (89 mg, 76%) as a yellow solid and 18e (4 mg, 4%) as a yel-

low solid. Analytical data for 8e: R_f= 0.08 (EtOAc). Mp = 141–

142 °C (MeOH/hexane).

a ²_D⁰

ꢄ

¼ þ22 (c 0.5, MeOH). ¹H NMR

(300 MHz, CD₃OD): d = 8.43–8.36 (m, 2H, H-ar), 8.34–8.30 (m,

2H, H-ar), 4.70 (d, 1H, J = 9.8 Hz, H-1⁰), 3.92 (dd, 1H, J = 1.7 Hz,

J = 12.0 Hz, H-6⁰a), 3.81 (dd, 1H, J = 9.8 Hz, J = 8.8 Hz, H-2⁰), 3.72

(dd, 1H, J = 5.4 Hz, J = 12.0 Hz, H-6⁰b), 3.54 (dd, 1H, J = 8.5 Hz,

J = 8.8 Hz, H-3⁰), 3.49-3.42 (m, 2H, H-4⁰, H-5⁰). ¹³C NMR (75 MHz,

CD₃OD): d = 178.9 (C-5oxa), 168.3 (C-3oxa), 151.1 (C^IV-ar), 133.6

(C^IV-ar), 129.6 (2C), 125.3 (2C), 83.1 (C-5⁰), 79.1 (C-3⁰), 75.1 (C-1⁰),

73.9 (C-2⁰), 71.2 (C-4⁰), 62.8 (C-6⁰). MS (LSIMS, glycerol) m/z = 354

[M+H]⁺. HRMS (LSIMS, glycerol) m/z = C₁₄H₁₆N₃O₈[M+H]⁺calcd

354.0937, found 354.0940. Analytical data for 18e: R_f= 0.17

(EtOAc). ¹H NMR (500 MHz, CD₃OD): d = 8.43–8.37 (m, 2H, H-ar),

8.34–8.30 (m, 2H, H-ar), 6.18 (d, 1H, J = 2.8 Hz, H-2⁰), 4.35 (dd,

1H, J = 2.8 Hz, J = 7.3 Hz, H-3⁰), 4.07 (ddd, 1H, J = 1.9 Hz, J = 4.7 Hz,

J = 9.5 Hz, H-5⁰), 4.03 (dd, 1H, J = 1.9 Hz, J = 12.3 Hz, H-6⁰a), 3.95

(dd, 1H, J = 4.7 Hz, J = 12.3 Hz, H-6⁰b), 3.77 (dd,1H, J = 7.3 Hz,

J = 9.5 Hz, H-4⁰). ¹³C NMR (125 MHz, CD₃OD): d = 173.6 (C-5oxa),

168.5 (C-3oxa), 151.2 (C^IV-ar), 141.1 (C-1⁰), 133.6 (C^IV-ar), 129.6,

125.3 (2C), 113.4 (C-2⁰), 82.3 (C-5⁰), 70.2 (C-3⁰), 69.6 (C-4⁰), 61.8

a ²_D⁰

ꢄ

¼ þ14:0 (c 1, MeOH). ¹H NMR (500 MHz, CD₃OD): d = 8.83

(d, 1H, J = 8.4 Hz, H-8⁰⁰), 8.24 (d, 1H, J = 7.1 Hz, H-2⁰⁰), 8.09 (d, 1H,

J = 8.2 Hz, H-4⁰⁰), 7.99 (d, 1H, J = 7.9 Hz, H-5⁰⁰), 7.65–7.58 (m, 3H,

H-3⁰⁰, H-6⁰⁰, H-7⁰⁰), 4.73 (d, 1H, J = 9.8 Hz, H-1⁰), 3.92 (dd, 1H,

J < 1 Hz, J = 11.7 Hz, H-6⁰a), 3.85 (dd, 1H, J = 9.2 Hz, J = 9.8 Hz, H-

2⁰), 3.74 (dd, 1H, J = 5.4 Hz, J = 11.7 Hz, H-6⁰b), 3.57–3.47 (m, 3H,

H-3⁰, H-4⁰, H-5⁰). ¹³C NMR (125 MHz, CD₃OD): d = 176.4 (C-5oxa),

168.9 (C-3oxa), 134.4 (C-1⁰⁰), 132.1 (C-4⁰⁰), 130.8 (C-8a⁰⁰), 129.4 (C-

2⁰⁰), 128.8 (C-5⁰⁰), 127.6 (C-6⁰⁰), 126.5 (C-7⁰⁰), 126.0 (C-8⁰⁰), 125.1

(C-3⁰⁰), 123.7 (C-4a⁰⁰), 82.0, 78.2 (C-3⁰, C-5⁰), 74.2 (C-1⁰), 73.0 (C-

2⁰), 70.2 (C-4⁰), 61.7 (C-6⁰). MS (ESI) m/z = 381.2 [M+Na]⁺. HRMS

(ESI) m/z = C₁₈H₁₈N₂O₆[M+Na]⁺calcd 381.1062, found 381.1062.

4.3.8. 5-(b-

D-Glucopyranosyl)-3-(2-naphthyl)-1,2,4-oxadiazole

(8i) and 5-(2-deoxy-D-arabino-hex-1-enopyranosyl)-3-(2-

naphthyl)-1,2,4-oxadiazole (18i)

A solution of 17i (340 mg, 0.44 mmol) was treated according to

method F. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 3:2 then EtOAc then EtOAc/MeOH 9:1) to af-

ford 8i (119 mg, 76%) as a yellow foam and 18i (11 mg, 8%) as a white

(C-6⁰). MS (ESI) m/z = 692.7 [2 M+Na]⁺. HRMS (ESI) m/z = C₁₄H₁₄

N₃O₇[M+H]⁺calcd 336.0832, found 336.0836.

solid. Analyticaldatafor 8i: R_f= 0.51 (EtOAc/MeOH, 9:1). ½a _D²⁰

¼ þ14

ꢄ

4.3.5. 3-(4-Aminophenyl)-5-(b-

oxadiazole (8f)

-glucopyranosyl)-1,2,4-

(c 1, MeOH). ¹H NMR (300 MHz, CD₃OD): d = 8.62 (s, 1H, H-ar), 8.99

(d, 1H, J = 8.6 Hz, H-ar), 7.98–7.92 (m, 3H, H-ar), 7.59–7-55 (m, 2H,

H-ar), 4.70 (d, 1H, J = 9.8 Hz, H-1⁰), 3.93 (pdd, 1H, J < 1 Hz,

J = 12.2 Hz, H-6⁰a), 3.84 (dd, 1H, J = 8.9 Hz, J = 9.8 Hz, H-2⁰), 3.74

(dd, 1H, J = 4.5 Hz, J = 12.2 Hz, H-6⁰b), 3.59–3.49 (m, 3H, H-3⁰, H-4⁰,

H-5⁰). ¹³C NMR (75 MHz, CD₃OD): d = 178.3 (C-5oxa), 169.7 (C-

3oxa), 136.2 (C^IV-ar), 134.5 (C^IV-ar), 130.0, 129.9, 129.01, 128.98,

128.9, 128.1, 125.1 (C^IV-ar), 124.6, 83.0 (C-5⁰), 79.2 (C-3⁰), 75.2 (C-

1⁰), 74.0 (C-2⁰), 71.3 (C-4⁰), 62.8 (C-6⁰). MS (ESI) m/z = 359.0 [M+H]⁺,

381.0 [M+Na]⁺, 738.9 [2M+Na]⁺. HRMS (ESI) m/z = C₁₈H₁₉N₂O₆

[M+H]⁺calcd 359.1243, found 359.1244. Analytical data for 18i:

R_f= 0.61 (EtOAc/MeOH, 9:1). ¹H NMR (300 MHz, CD₃OD): d = 8.61

(s, 1H, H-ar), 8.10 (dd, 1H, J = 1.6 Hz, J = 8.6 Hz, H-ar), 8.03–7.97

(m, 3H H-ar), 7.61–7.57 (m, 2H, H-ar), 6.16 (d, 1H, J = 2.8 Hz, H-2⁰),

4.36 (dd, 1H, J = 2.8 Hz, J = 7.2 Hz, H-3⁰), 4.09–4.05 (m, 2H, H-5⁰, H-

6⁰a), 3.95 (dd, 1H, J = 5.3 Hz, J = 12.5 Hz, H-6⁰b), 3.77 (dd, 1H,

J = 7.2 Hz, J = 9.5 Hz, H-4⁰). ¹³C NMR (75 MHz, CD₃OD): d = 173.0

(C-5oxa), 169.9 (C-3oxa), 141.2 (C-1⁰), 136.3 (C^IV-ar), 134.5 (C^IV-ar),

130.1, 129.9, 129.02, 128.96, 128.2, 125.0 (C^IV-ar), 124.6, 113.0 (C-

2⁰), 82.3 (C-5⁰), 70.3 (C-3⁰), 69.8 (C-4⁰), 62.0 (C-6⁰). MS (ESI) m/z =

341.0 [M+H]⁺, 363.0 [M+Na]⁺, 702.9 [2M+Na]⁺. HRMS (ESI) m/z =

C₁₈H₁₇N₂O₅[M+H]⁺calcd 341.1137, found 341.1136.

A solution of 8e (30 mg, 8.2

mol) and Pd–C 10% (5 mg) in

MeOH (5 mL) was stirred at rt under H₂(1 atm.). After 24 h, the

solution was ﬁltered through a pad of Celite, washed with MeOH

(3 ꢃ 10 mL). The ﬁltrate was evaporated under diminished pres-

sure and the residue was puriﬁed by reversed-phase silica gel col-

umn chromatography (water) to afford 8f (30 mg, 99%) as a pale

brown foam. ½a ²_D⁰

ꢄ

¼ ꢂ7:8 (c 0.95, MeOH). ¹H NMR (300 MHz,

CD₃OD): d = 7.57 (d, 2H, J = 9.2 Hz, H-2⁰⁰, H-6⁰⁰), 6.73 (d, 2H,

J = 9.2 Hz, H-3⁰⁰, H-5⁰⁰), 3.85 (m, 1H, H-6⁰a), 3.63 (m, 1H, H-6⁰b),

3.56 (m, 1H, H-1⁰), 3.20–3.42 (m, 4H, H-2⁰, H-3⁰, H-4⁰, H-5⁰). ¹³C

NMR (75 MHz, CD₃OD): d = 176.8 (C-5oxa), 166.1 (C-3oxa), 155.1

(C-4⁰⁰), 129.8 (C-2⁰⁰, C-6⁰⁰), 113.7 (C-3⁰⁰, C-5⁰⁰), 113.5 (C-1⁰⁰), 77.7 (C-

1⁰), 80.0, 78.4, 72.7, 70.8 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.0 (C-6⁰).

4.3.6. 5-(b-D-Glucopyranosyl)-3-(4-hydroxyphenyl)-1,2,4-

oxadiazole (8g)

A solution of 17g (173 mg, 0.27 mmol) and NaOMe (5 mg) in

MeOH/CH₂Cl₂(10 mL, 9:1) was stirred at rt for 24 h. The reaction

was neutralized to pH 5–6 with Amberlite IR-120 resin (H⁺form)

and the resin washed with MeOH (3 ꢃ 10 mL). The ﬁltrate was

evaporated under diminished pressure and the residue was puri-

ﬁed by ﬂash silica gel column chromatography (PE/EtOAc, 1:1 then

EtOAc then EtOAc/MeOH 95:5) to afford 8g (32 mg, 37%) as a white

4.3.9. 3-(4-Benzyloxyphenyl)-5-(b-D-glucopyranosyl)-1,2,4-

oxadiazole (8j)

A solution of 17j (205 mg, 0.25 mmol) and NaOMe (5 mg) in

MeOH/CH₂Cl₂(5 mL, 9:1) was stirred at rt for 24 h. The reaction

gum. R_f= 0.39 (EtOAc/MeOH, 85:15). ½a _D²⁰

ꢄ

¼ þ10:4 (c 1, MeOH). ¹H

NMR (300 MHz, CD₃OD): d = 7.88 (d, 2H, J = 8.7 Hz, H-2⁰⁰, H-6⁰⁰),

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4781

was neutralized to pH 5–6 with Amberlite IR-120 resin (H⁺form)

and the resin washed with MeOH (3 ꢃ 10 mL). The ﬁltrate was

evaporated under diminished pressure and the residue was puri-

ﬁed by ﬂash silica gel column chromatography (PE/EtOAc, 1:1 then

EtOAc then EtOAc/MeOH 95:5) to afford 8j (57 mg, 56%) as a white

The reaction mixture was reﬂuxed for 3 h then evaporated in vac-

uum. Crystallisation from EtOH gave 11c (379 mg, 70%) as white

crystals. Mp = 170–172 °C. ½a _D²⁰

ꢄ

¼ ꢂ200 (c 0.19, CHCl₃). ¹H NMR

(360 MHz, CDCl₃): d = 8.05–7.80 (m, 10H, H-ar), 7.57–7.29 (m,

14H, H-ar), 6.10 (t, 1H, J = 9.3 Hz, H-2⁰), 6.02 (t, 1H, J = 9.3 Hz, H-

3⁰), 5.87 (dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-4⁰), 5.27 (d, 1H,

J = 9.3 Hz, H-1⁰), 4.70 (dd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a), 4.54

(dd, 1H, J = 5.3 Hz, J = 11.9 Hz, H-6⁰b), 4.38 (ddd, 1H, J < 1 Hz,

J = 5.3 Hz, J = 10.6 Hz, H-5⁰), 2.41 (s, 3H, PhCH₃). ¹³C NMR

(90 MHz, CDCl₃): d = 166.1, 165.7, 165.2, 164.9 (C@O), 160.7,

142.7 (C-2oxa, C-5oxa), 133.6–120.6 (CH-ar, C^IV-ar), 77.0 (C-1⁰),

73.7, 71.9, 70.2, 69.1 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 63.0 (C-6⁰), 21.6

(PhCH₃). Anal. Calcd for C₄₃H₃₄N₂O₁₀(738.76): C, 69.91; H, 4.64;

N, 3.79. Found: C, 69.99; H, 4.57; N, 3.72.

foam. R_f= 0.31 (EtOAc/MeOH, 9:1). ½a _D²⁰

¼ þ10:5 (c 1.13, MeOH).

ꢄ

¹H NMR (300 MHz, CD₃OD): d = 8.00 (d, 2H, J = 9.0 Hz, H-2⁰⁰, H-

6⁰⁰), 7.50–7.25 (m, 5H, CH₂Ph), 7.13 (d, 2H, J = 9.0 Hz, H-3⁰⁰, H-5⁰⁰),

5.15 (s, 2H, CH₂Ph), 4.63 (d, 1H, J = 9.6 Hz, H-1⁰), 3.91 (pdd, 1H,

J < 1 Hz, J = 11.4 Hz, H-6⁰a), 3.80 (dd, 1H, J = 8.7 Hz, J = 9.6 Hz, H-

2⁰), 3.72 (m, 1H, H-6⁰b), 3.47 (m, 3H, H-3⁰, H-4⁰, H-5⁰). ¹³C NMR

(75 MHz, CD₃OD): d = 177.9 (C-5oxa), 169.3 (C-3oxa), 162.8 (C-

4⁰⁰), 138.2 (C^IV-ar), 130.0 (C-2⁰⁰, C-6⁰⁰), 129.6, 129.0, 128.6 (CH₂Ph),

120.3 (C-1⁰⁰), 116.4 (C-3⁰⁰, C-5⁰⁰), 75.1 (C-1⁰), 73.9 (C-2⁰), 83.0, 79.2,

71.2 (C-3⁰, C-4⁰, C-5⁰), 71.1 (CH₂Ph), 62.7 (C-6⁰). MS (ESI) m/z =

437.2 [M+Na]⁺. HRMS (ESI) m/z = C₂₁H₂₂N₂O₇[M+Na]⁺calcd

437.1325, found 437.1325.

4.4.3. 2-(4-Methoxyphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,3,4-oxadiazole (11d)

To a solution of tetrazole 10 (500 mg, 0.77 mmol) in abs. tolu-

ene (5 mL) was added 4-methoxy-benzoyl chloride (1.54 mmol).

The reaction mixture was reﬂuxed for 1 h. Then it was cooled to

rt, washed with saturated aqueous NaHCO₃solution and water.

The organic phase was dried (MgSO₄), concentrated under dimin-

ished pressure and puriﬁed by column chromatography (hexane/

4.3.10. 3-(1,3-Benzodioxol-5-yl)-5-(b-D-glucopyranosyl)-1,2,4-

oxadiazole (8k)

A solution of 17k (280 mg, 0.36 mmol) and NaOMe (5 mg) in

MeOH/CH₂Cl₂(15 mL, 2:1) was stirred at rt for 2 h. The reaction

was neutralized to pH 5–6 with Amberlite IR-120 resin (H⁺form)

and the resin washed with MeOH (2 ꢃ 10 mL). The ﬁltrate was

evaporated under diminished pressure and the residue was puri-

ﬁed by ﬂash silica gel column chromatography (PE/EtOAc, 1:1 then

EtOAc then EtOAc/MeOH 85:15) to afford 8k (74 mg, 60%) as a pale

EtOAc, 1:1) to give 11d (235 mg, 40%) as a syrup. ½a _D²⁰

¼ ꢂ261 (c

ꢄ

0.19, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.05–7.80 (m, 10H,

H-ar), 7.36–7.27 (m, 12H, H-ar), 6.97–6.95 (d, 2H, J = 9.3 Hz, H-

ar), 6.12 (dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-2⁰), 6.04 (dd, 1H,

J = 9.3 Hz, J = 10.6 Hz, H-3⁰), 5.89 (t, 1H, J = 9.3 Hz, H-4⁰), 5.28 (d,

1H, J = 9.3 Hz, H-1⁰), 4.71 (pdd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a),

4.55 (dd, 1H, J = 5.3 Hz, J = 11.9 Hz, H-6⁰b), 4.40 (ddd, 1H,

J = 2.6 Hz, J = 5.3 Hz, J = 9.3 Hz, H-5⁰), 3.80 (s, 3H, OMe). ¹³C NMR

(90 MHz, CDCl₃): d = 166.0, 165.8, 165.6, 165.1 (C@O), 162.5,

160.4 (C-2oxa, C-5oxa), 133.5–114.3 (CH-ar, C^IV-ar), 77.0 (C-1⁰),

73.6, 71.8, 70.2, 69.0 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.9 (C-6⁰), 55.3

(OMe). Anal. Calcd for C₄₃H₃₄N₂O₁₁(754.76): C, 68.43; H, 4.54; N,

3.71. Found: C, 68.34; H, 4.61; N, 3.63.

yellow foam. R_f= 0.41 (EtOAc/MeOH, 85:15). ½a _D²⁰

¼ þ18:0 (c 1,

ꢄ

DMSO). ¹H NMR (300 MHz, CD₃OD): d = 7.64 (dd, 1H, J = 1.6 Hz,

J = 8.1 Hz, H-5⁰⁰), 7.48 (d, 1H, J = 1.6 Hz, H-3⁰⁰), 6.95 (d, 1H,

J = 8.1 Hz, H-6⁰⁰), 6.05 (s, 2H, OCH₂O), 4.62 (d, 1H, J = 9.8 Hz, H-1⁰),

3.91 (dd, 1H, J = 1.6 Hz, J = 12.2 Hz, H-6⁰a), 3.78 (t, 1H, J = 9.8 Hz,

H-2⁰), 3.71 (dd, 1H, J = 5.2 Hz, J = 12.2 Hz, H-6⁰b), 3.56–3.40 (m,

3H, H-3⁰, H-4⁰, H-5⁰). ¹³C NMR (75 MHz, CD₃OD): d = 178.0 (C-

5oxa), 169.3 (C-3oxa), 152.0, 149.8 (C-1⁰⁰, C-2⁰⁰), 123.4 (C-5⁰⁰),

121.5 (C^IV-ar), 109.7 (C-6⁰⁰), 108.7 (C-3⁰⁰), 103.3 (OCH₂O), 75.2 (C-

1⁰), 74.0 (C-2⁰), 83.0, 79.2, 71.2 (C-3⁰, C-4⁰, C-5⁰), 62.8 (C-6⁰). MS

(ESI) m/z = 353 [M+H]⁺, 375 [M+Na]⁺, 727 [2 M+Na]⁺. HRMS (ESI)

m/z = C₁₅H₁₆N₂NaO₈[M+Na]⁺calcd 375.0804, found 375.0802.

4.4.4. 2-(4-Nitrophenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,3,4-oxadiazole (11e)

To a solution of tetrazole 10 (850 mg, 1.31 mmol) in abs. toluene

(9 mL) was added 4-nitro-benzoyl chloride (1.57 mmol). The reac-

tion mixture was reﬂuxed for 4.5 h then evaporated under dimin-

ished pressure. Crystallisation from EtOH gave 11e (635 mg, 63%)

4.4. Zemplén deacylation of 1,3,4-oxadiazoles

4.4.1. 2-Phenyl-5-(2,3,4,6-tetra-O-benzoyl-b-D-glucopyranosyl)-

1,3,4-oxadiazole (11b)

as yellowish crystals. Mp = 158–161 °C. ½a _D²⁰

¼> ꢂ297 (c 0.17,

ꢄ

Prepared according to method A. Yield: 78% (white solid).

Alternative preparation: To a solution of benzoic acid (19 mg,

0.15 mmol) and DCC (32 mg, 0.15 mmol) in abs. toluene was

added tetrazole 10 (100 mg, 0.15 mmol), and reﬂuxed for 4 h.

After cooling to rt the mixture was ﬁltered and evaporated.

The product 11b was crystallized from EtOH (62 mg, 56%) and

CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.36–7.29 (m, 24H, H-ar),

6.13 (t, 1H, J = 9.3 Hz, H-2⁰), 5.95 (dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-

3⁰), 5.89 (dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-4⁰), 5.29 (d, 1H,

J = 9.3 Hz, H-1⁰), 4.70 (dd, 1H, J < 1 Hz, J = 11.9 Hz, H-6⁰a), 4.53 (dd,

1H, J = 3.9 Hz, J = 11.9 Hz, H-6⁰b), 4.41 (ddd, 1H, J < 1 Hz, J = 3.9 Hz,

J = 9.3 Hz, H-5⁰). ¹³C NMR (90 MHz, CDCl₃): d = 166.0, 165.6, 165.1,

165.0 (C@O), 164.1, 162.0 (C-2oxa, C-5oxa), 149.7 (CH-ar, C^IV-ar)

133.7–128.2 (CH-ar, C^IV-ar), 77.2 (C-1⁰), 73.3, 71.8, 70.4, 68.9

(C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.8 (C-6⁰). Anal. Calcd for C₄₂H₃₁N₃O₁₂

(769.73): C, 65.54; H, 4.06; N, 5.46. Found: C, 65.48; H, 4.12;

N, 5.52.

obtained as white crystals. Mp = 186–187 °C.

a ²_D⁰

ꢄ

¼ ꢂ195 (c

0.21, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.06–7.26 (m, 25H,

H-ar), 6.11, 6.01, 5.88 (3 ꢃ pt, 3H, J ꢀ 9.3 Hz, H-2⁰, H-3⁰, H-4⁰),

5.29 (d, 1H, J = 9.3 Hz, H-1⁰), 4.71 (dd, 1H, J = 2.6 Hz, J = 11.9 Hz,

H-6⁰a), 4.54 (dd, 1H, J = 5.3 Hz, J = 11.9 Hz, H-6⁰b), 4.40 (ddd,

1H, J = 2.6 Hz, J = 5.3 Hz, J = 9.3 Hz, H-5⁰). ¹³C NMR (90 MHz,

CDCl₃): d = 166.0, 165.9 165.6, 165.1, 164.8 (C@O, C-5oxa),

162.0 (C-2oxa), 133.5–123.3 (CH-ar, C^IV-ar), 77.1 (C-1⁰), 73.6,

71.9, 70.3, 69.0 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 63.0 (C-6⁰). Anal. Calcd

for C₄₂H₃₂N₂O₁₀(724.73): C, 69.61; H, 4.45; N, 3.87. Found: C,

69.52; H, 4.11; N, 3.99.

4.4.5. 2-(1-Naphthyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-glucopyr-

anosyl)-1,3,4-oxadiazole (11h)

Prepared according to method A. Yield: 75% (white solid).

Mp = 172–174 °C. ½a _D²⁰

ꢄ

¼ ꢂ162 (c 0.21, CHCl₃). ¹H NMR (360 MHz,

CDCl₃): d = 8.22–7.28 (m, 27H, H-ar), 6.14, 6.10, 5.90 (3 ꢃ pt, 3H,

J ꢀ 9.3 Hz, H-2⁰, H-3⁰, H-4⁰), 5.34 (d, 1H, J = 9.3 Hz, H-1⁰), 4.71 (dd,

1H, J = 2.6 Hz, J = 11.9 Hz, H-6⁰a), 4.55 (dd, 1H, J = 5.3 Hz,

J = 11.9 Hz, H-6⁰b), 4.42 (ddd, 1H, J = 2.6 Hz, J = 5.3 Hz, J = 9.3 Hz,

H-5⁰). ¹³C NMR (90 MHz, CDCl₃): d = 166.1, 165.9 165.6, 165.2,

164.9 (C@O, C-5oxa), 160.7 (C-2oxa), 133.2–120.0 (CH-ar, C^IV-ar),

4.4.2. 2-(4-Methylphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,3,4-oxadiazole (11c)

To a solution of tetrazole 10 (500 mg, 0.77 mmol) in abs. tolu-

ene (5 mL) was added 4-methyl-benzoyl chloride (0.95 mmol).

4782

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

77.1 (C-1⁰), 73.6, 71.9, 70.4, 69.1 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 63.0 (C-6⁰).

Anal. Calcd for C₄₆H₃₄N₂O₁₀(774.79): C, 71.31; H, 4.42; N, 3.62.

Found: C, 71.52; H, 4.20; N, 3.64.

C₁₉H₂₀N₂O₈(404.37): C, 56.43; H, 4.99; N, 6.93. Found: C, 56.25;

H, 5.10; N, 6.80.

4.4.10. 2-Phenyl-5-(2,3,4-tri-O-acetyl-a-D-arabinopyranosyl)-

4.4.6. 2-(2-Naphthyl)-5-(2,3,4,6-tetra-O-benzoyl-b-

-glucopyr-

1,3,4-oxadiazole (15c)

anosyl)-1,3,4-oxadiazole (11i)

Prepared according to method C. Yield: 57% (colourless syrup).

Prepared according to method A. Yield: 62% (white solid).

a ²_D⁰

ꢄ

¼ þ48 (c 1.188, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.11–

Mp = 164–166 °C.

a ²_D⁰

ꢄ

¼ ꢂ318 (c 0.21, CHCl₃). ¹H NMR

8.01 (m, 2H, H-ar), 7.56–7.49 (m, 3H, H-ar), 5.66 (pt, 1H,

J = 10.0 Hz, H-2⁰), 5.46–5.44 (m, 1H, H-4⁰), 5.24 (dd, 1H, J = 3.3 Hz,

J = 10.0 Hz, H-3⁰), 4.80 (d, 1H, J = 9.5 Hz, H-1⁰), 4.21 (dd, 1H,

J = 2.3 Hz, J = 13.2 Hz, H-5⁰_ax), 3.90 (dd, 1H, J = 1.2 Hz, J = 13.2 Hz,

H-5⁰_eq), 2.24, 2.04, 1.97 (3s, 9H, CH₃). ¹³C NMR (90 MHz, CDCl₃):

d = 170.3 169.9, 169.3, (C@O), 165.7, 161.4 (C-2oxa, C-5oxa),

132.0, 129.0, 127.1 (CH-ar), 123.3 (C^IV-ar), 72.2, 70.8, 68.0, 67.0

(C-1⁰, C-2⁰, C-3⁰, C-4⁰), 68.5 (C-5⁰), 20.9, 20.6, 20.4 (CH₃). Anal. Calcd

for C₁₉H₂₀N₂O₈(404.37): C, 56.43; H, 4.99; N, 6.93. Found: C, 56.38;

H, 4.91; N, 6.70.

(360 MHz, CDCl₃): d = 8.56–7.30 (m, 27H, H-ar), 6.12, 6.06, 5.90

(3 ꢃ pt, 3H, J ꢀ 9.3 Hz, H-2⁰, H-3⁰, H-4⁰), 5.30 (d, 1H, J = 9.3 Hz, H-

1⁰), 4.72 (dd, 1H, J = 2.6 Hz, J = 11.9 Hz, H-6⁰a), 4.56 (dd, 1H,

J = 5.3 Hz, J = 11.9 Hz, H-6⁰b), 4.40 (ddd, 1H, J = 2.6 Hz, J = 5.3 Hz,

J = 9.3 Hz, H-5⁰). ¹³C NMR (90 MHz, CDCl₃): d = 166.5, 166.1 165.7,

165.2, 164.9 (C@O, C-5oxa), 163.2 (C-2oxa), 133.4–120.6 (CH-ar,

C^IV-ar), 77.1 (C-1⁰), 73.6, 71.0, 70.3, 69.1 (C-2⁰, C-3⁰, C-4⁰,C-5⁰),

63.1 (C-6⁰). Anal. Calcd for C₄₆H₃₄N₂O₁₀(774.79): C, 71.31; H,

4.42; N, 3.62. Found: C, 71.55; H, 4.08; N, 3.54.

4.4.7. 2-(4-Acetoxyphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-

4.5. Synthesis of hydroximoyl chlorides

glucopyranosyl)-1,3,4-oxadiazole (11l)

To a solution of 4-acetoxybenzoic acid (139 mg, 0.77 mmol) and

DCC (159 mg, 0.77 mmol) in abs. toluene (5 mL) was added tetra-

zole 10 (500 mg, 0.77 mmol). The reaction was heated to reﬂux

for 7 h. After cooling to rt the mixture was ﬁltered, evaporated

and puriﬁed by column chromatography (hexane/EtOAc, 2:1) to

4.5.1. N-Hydroxy-benzenecarboximidoyl chloride (16b)⁴⁵

Prepared according to method D from benzaldehyde (1.76 g,

16.6 mmol) to afford 16b (2.51 g, 98%) as a pale yellow amorphous

solid. R_f= 0.69 (PE/EtOAc, 3/1).

give 11l (247 mg, 41%) as

white amorphous product.

4.5.2. N-Hydroxy-4-methyl-benzenecarboximidoyl chloride (16c)⁴⁵

Prepared according to method D from 4-methyl-benzaldehyde

(3.6 g, 30 mmol) to afford 16c (3.85 g, 76%) as a yellow oil.

R_f= 0.37 (CH₂Cl₂). ¹H NMR (300 MHz, CDCl₃): d = 9.15 (s, 1H, OH),

7.70 (d, 2H, J = 9.0 Hz, H-ar), 7.18 (d, 2H, J = 9.0 Hz, H-ar), 2.36 (s,

3H, PhMe). ¹³C NMR (75 MHz, CDCl₃): d = 144.1, 140.3, 131.8,

129.1, 127.0, 21.2.

a ²_D⁰

ꢄ

¼ ꢂ174 (c 0.17, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.09–

7.81 (m, 10H, H-ar), 7.56–7.23 (m, 14H, H-ar) 6.10 (t, 1H,

J = 9.3 Hz, H-2⁰), 5.99 (dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-3⁰), 5.87

(dd, 1H, J = 9.3 Hz, J = 10.6 Hz, H-4⁰), 5.26 (d, 1H, J = 9.3 Hz, H-1⁰),

4.71 (dd, 1H, J = 2.6 Hz,

=11.9 Hz, H-6⁰a), 4.53 (dd, 1H,

J = 11.9 Hz, J = 5.3 Hz, H-6⁰b), 4.38 (ddd, 1H, J = 2.6 Hz, J = 5.3 Hz,

J = 9.3 Hz, H-5⁰), 2.33 (s, 3H, OAc). ¹³C NMR (90 MHz, CDCl₃):

d = 168.8 (COCH₃), 166.1, 165.7, 165.2, 165.1 (C@O), 164.8, 161.0

(C-2oxa, C-5oxa), 153.4, 133.6–120.8 (CH-ar, C^IV-ar), 77.0 (C-1⁰),

73.5, 71.8, 70.2, 69.0 (C-2⁰, C-3⁰, C-4⁰, C-5⁰), 62.9 (C-6⁰), 21.1

(COCH₃). Anal. Calcd for C₄₄H₃₄N₂O₁₂(782.77): C, 67.52; H, 4.38;

N, 3.58. Found: C, 67.58; H, 4.30; N, 3.64.

4.5.3. N-Hydroxy-4-methoxy-benzenecarboximidoyl chloride

(16d)⁴⁶

Prepared according to method D from 4-methoxy-benzalde-

hyde (2 g, 14.7 mmol) to afford 16d (2.12 g, 78%) as a pale yellow

amorphous solid.

4.4.8. 2-Phenyl-5-(2,3,4,6-tetra-O-acetyl-b-

1,3,4-oxadiazole (15a)

-galactopyranosyl)-

4.5.4. N-Hydroxy-4-nitro-benzenecarboximidoyl chloride (16e)

Prepared according to method D from 4-nitro-benzaldehyde

(2 g, 14.1 mmol) to afford 16e (1.61 g, 91%) as a yellow solid.

R_f= 0.75 (CH₂Cl₂). Mp = 101–103 °C. ¹H NMR (300 MHz, CDCl₃):

d = 8.05 (d, 2H, J = 9.0 Hz, H-3, H-5), 8.27 (d, 2H, J = 9.0 Hz, H-2,

H-6), 8.63 (s, 1H, OH). ¹³C NMR (75 MHz, CDCl₃): d = 123.6 (C-3,

C-5), 127.9 (C-2, C-6), 137.9 (C-1), 138.1 (ClCNOH), 148.8 (C-4).

Prepared according to method C. Yield: 78% (colourless syrup).

a ²_D⁰

ꢄ

¼ ꢂ30 (c 1.06, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d 8.11–

8.09 (m, 2H, H-ar), 7.61–7.45 (m, 3H, H-ar), 5.64 (t, 1H,

J = 9.9 Hz, H-2⁰), 5.44 (d, 1H, J = 3.0 Hz, H-4⁰), 5.23 (dd, 1H,

J = 3.0 Hz, J = 9.9 Hz, H-3⁰), 4.86 (d, 1H, J = 10.0 Hz, H-1⁰), 4.28–

4.11 (m, 3H, H-5⁰, H-6⁰a, H-6⁰b), 2.24, 2.06, 2.02, 1.95 (4s, 12H,

CH₃). ¹³C NMR (90 MHz, CDCl₃): d = 170.5, 170.3, 170.0, 169.4

(C@O), 166.0 161.3 (C-2oxa, C-5oxa), 132.2, 129.1,127.4 (CH-ar),

123.5 (C^IV-ar), 75.5, 72.1, 71.4, 67.4, 66.9 (C-1⁰, C-2⁰, C-3⁰, C-4⁰,

C-5⁰), 61.6 (C-6), 20.7, 20.6, 20.5 (CH₃). Anal. Calcd for

C₂₂H₂₄N₂O₁₀(476.43): C, 55.46; H, 5.08; N, 5.88. Found: C,

55.28; H, 5.21; N, 5.67.

4.5.5. N-Hydroxy-4-hydroxy-benzenecarboximidoyl chloride

(16g)

Prepared according to method D from 4-hydroxy-benzaldehyde

(1.71 g, 14.0 mmol) to afford 16g (933 mg, 39%) as a pale orange

oil. R_f= 0.73 (PE/EtOAc, 3/2). ¹H NMR (300 MHz, CD₃SOCD₃):

d = 6.83 (d, 2H, J = 8.8 Hz, H-2, H-6), 7.61 (d, 2H, J = 8.8 Hz, H-3,

H-5), 7.95 (s, 1H, PhOH), 12.01 (s, 1H, NOH). ¹³C NMR (75 MHz,

CD₃SOCD₃): d = 116.3 (C-3, C-5), 123.5 (C-1), 133.7 (C-2, C-6),

147.3 (ClCNOH), 158.0 (C-4). MS (CI, isobutene) m/z = 172.0

[M+H]⁺. HRMS (CI, isobutene) m/z = C₇H₆ClNO [M+Na]⁺calcd

172.0165, found 172.0169.

4.4.9. 2-Phenyl-5-(2,3,4-tri-O-acetyl-b-D-xylopyranosyl)-1,3,4-

oxadiazole (15b)

Prepared according to method C. Yield: 67% (colourless syrup).

a ²_D⁰

ꢄ

¼ ꢂ87 (c 1.05, CHCl₃). ¹H NMR (360 MHz, CDCl₃): d = 8.09–

8.06 (m, 2H, H-ar), 7.56–7.49 (m, 3H, H-ar), 5.43 (pt, 1H,

J = 9.2 Hz, H-2 or H-3), 5.39 (t, 1H, J = 9.2 Hz, H-2 or H-3), 5.22–

5.13 (m, 1H, H-4⁰), 4.81 (d, 1H, J = 9.5 Hz, H-1⁰), 4.31 (dd, 1H,

J = 5.6 Hz, J = 11.5 Hz, H-5⁰_eq), 3.53 (dd, 1H, J = 10.7 Hz, J = 11.5 Hz,

H-5⁰_ax), 2.08, 2.06, 1.95 (3s, 9H, CH₃). ¹³C NMR (90 MHz, CDCl₃):

d = 170.0, 169.7 169.2, (C@O), 165.7, 161.2 (C-2oxa, C-5oxa),

132.1, 129.0, 127.1 (CH-ar), 123.2 (C^IV-ar), 72.5, 71.9, 69.5, 68.5

(C-1⁰, C-2⁰, C-3⁰, C-4⁰), 67.1 (C-5⁰), 20.3, 20.6 (CH₃). Anal. Calcd for

4.5.6. N-Hydroxy-1-naphthalenecarboximidoyl chloride (16h)⁴⁵

Prepared according to method D from 1-naphthaldehyde (2 g,

12.8 mmol) to afford 16h (1.56 g, 91%) as a brown oil. R_f= 0.84

(CH₂Cl₂). ¹H NMR (300 MHz, CD₃SOCD₃): d = 7.48–7.55 (m, 3H, H-

3, H-6, H-7), 7.66 (dd, 1H, J = 5.1 Hz, J = 7.2 Hz, H-8), 7.91–7.99

(m, 1H, H-4, H-5), 8,07 (m, 1H, H-2). ¹³C NMR (75 MHz, CD₃SOCD₃):

d = 124.0, 124.7, 126.0, 126.8, 127.6, 128.0, 129.4, 130.1, 130,4,

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4783

132.7, 132.8. MS (CI, isobutene) m/z = 206.0 [M+H]⁺. HRMS (CI, iso-

butene) m/z = C₁₁H₈ClNO [M+Na]⁺calcd 206.0373, found 206.0370.

¹H NMR (300 MHz, CDCl₃): d = 8.04 (m, 2H, H-ar), 7.94 (m, 2H, H-

ar), 7.88–7.81 (m, 6H, H-ar), 7.55–7.24 (m, 12H, H-ar), 7.18 (d,

2H, J = 8.0 Hz, H-ar), 6.08 (m, 2H, H-2⁰, H-3⁰), 5.88 (t, 1H,

J = 9.7 Hz, H-4⁰), 5.26 (d, 1H, J = 9.7 Hz, H-1⁰), 4.70 (dd, 1H,

J = 2.9 Hz, J = 12.5 Hz, H-6⁰a), 4.56 (dd, 1H, J = 5.0 Hz, J = 12.5 Hz,

H-6⁰b), 4.40 (ddd, 1H, J = 9.7 Hz, J = 5.0 Hz, J = 2.9 Hz, H-5⁰), 2.34

(s, 3H, CH₃Ph). ¹³C NMR (75 MHz, CDCl₃): d = 173.1 (C-5oxa),

168.4 (C-3oxa), 166.0, 165.7, 165.0, 164.7 (C@O), 141.7 (C-1⁰⁰),

133.5, 133.4, 133.3, 133.1, 129.8, 129.7 (2C), 129.4, 129.3 (C^IV-ar),

128.5 (C^IV-ar), 128.4, 128.3 (2C), 127.3, 123.2 (C-4⁰⁰), 77.1 (C-5⁰),

73.7 (C-3⁰), 72.4 (C-1⁰), 70.6 (C-2⁰), 69.0 (C-4⁰), 62.9 (C-6⁰), 21.5

(CH₃Ph). MS (ESI) m/z = 739.0 [M+H]⁺, 740.0 [M+Na]⁺, 1476.6

[2M+H]⁺, 1498.8 [2M+Na]⁺. HRMS (ESI) m/z = C₄₃H₃₅N₂O₁₀

[M+H]⁺calcd 739.2292, found 739.2293.

4.5.7. N-Hydroxy-2-naphthalenecarboximidoyl chloride

(16i)^45,47

Prepared according to method D from 2-naphthaldehyde (3.4 g,

21.6 mmol) to afford 42 (2.84 g, 64%) as a pale orange amorphous

solid. R_f= 0.66 (CH₂Cl₂). ¹H NMR (300 MHz, CDCl₃): d = 8.81 (s, 1H,

OH), 8.30 (s, 1H, H-ar), 7.2–7.93 (m, 4H, H-ar), 7.43–7.58 (m, 2H, H-

ar). ¹³C NMR (75 MHz, CDCl₃): d = 140.5, 134.2, 132.6, 129.5, 128.8,

128.2, 128.1, 127.6, 127.5, 126.8, 123.2.

4.5.8. 4-Benzyloxy-N-hydroxy-benzenecarboximidoyl chloride

(16j)

Prepared according to method D from 4-benzyloxy-benzalde-

hyde (3 g, 14.1 mmol) to afford 16j (1.87 g, 51%) as a pale orange

solid. R_f= 0.88 (PE/EtOAc, 9/1). Mp = 83–84 °C. ¹H NMR

(300 MHz, CD₃SOCD₃): d = 5.07 (s, 2H, OCH₂Ph), 7.01 (d, 2H,

J = 9.0 Hz, H-3, H-5), 7.31–7.36 (m, 5H, OCH₂Ph), 7.63 (d, 2H,

J = 9.0 Hz, H-2, H-6), 12.09 (s, 1H, OH). ¹³C NMR (75 MHz,

CD₃SOCD₃): d = 68.7 (OCH₂Ph), 114.3 (C-3, C-5), 124.5 (C-1),

126.1, 126.9, 127.0, 127.3, 127.9, 127.5 (C-2, C-6), 135.9, 159.3

(ClCNOH), 178.7 (C-4). MS (CI, isobutene) m/z = 262.0 [M+H]⁺.

HRMS (CI, isobutene) m/z = C₁₄H₁₂ClNO₂[M+Na]⁺calcd 262.0635,

found 262.0637.

4.6.3. 3-(4-Methoxyphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-

glucopyranosyl)-1,2,4-oxadiazole (17d)

D-

A solution composed of 9 (300 mg, 0.5 mmol), 16d (460 mg,

2.48 mmol) and Et₃N (520 L, 3.71 mmol) was treated according

to method E. The residue was puriﬁed by ﬂash silica gel column

chromatography (PE/EtOAc, 4:1) to afford 17d (336 mg, 90%) as a

white solid. R_f= 0.17 (PE/EtOAc, 4:1). Mp = 75–76 °C (CH₂Cl₂/PE).

a ²_D⁰

ꢄ

¼ ꢂ9 (c 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): d = 8.05–7.85

(m, 10H, H-ar), 7.55–7.28 (m, 12H, H-ar), 6.89 (d, 2H, J = 8.7 Hz,

H-ar), 6.06 (m, 2H, H-2⁰, H-3⁰), 5.86 (m, 1H, H-4⁰), 5.24 (m, 1H, H-

1⁰), 4.70 (dd, 1H, J = 1.9 Hz, J = 12.3 Hz, H-6⁰a), 4.56 (dd, 1H,

J = 4.8 Hz, J = 12.3 Hz, H-6⁰b), 4.38 (m, 1H, H-5⁰), 3.81 (s, 3H,

OCH₃). ¹³C NMR (75 MHz, CDCl₃): d = 173.5 (C-5oxa), 168.6 (C-

3oxa), 166.6, 166.2, 165.6 (C=O), 165.2 (C-4⁰⁰), 162.5, 134.0, 133.9,

133.8, 133.6, 130.3, 130.2, 129.9 (C^IV-ar), 129.6, 129.1 (C^IV-ar),

129.0 (C^IV-ar), 128.9, 128.8, 128.8, 119.0 (C-1⁰⁰), 114.6 (C-3⁰⁰), 77.6

(C-5⁰), 74.2 (C-2⁰or C-3⁰), 72.9 (C-1⁰), 71.1 (C-2⁰or C-3⁰), 69.5 (C-

4⁰), 63.4 (C-6⁰), 55.8 (OCH₃). MS (LSIMS, NBA) m/z = 755 [M+H]⁺.

HRMS (LSIMS, NBA) m/z = C₄₃H₃₅N₂O₁₁[M+H]⁺calcd 755.2241,

found 755.2242.

4.5.9. N-Hydroxy-benzo[1,3]dioxole-5-carboximidoyl chloride

(16k)

Prepared according to method D from benzo[1,3]dioxole-5-

carbaldehyde (4.5 g, 30 mmol) to afford 16k (4.87 g, 81%) as a pale

brown amorphous solid. R_f= 0.76 (PE/CH₂Cl₂, 3/7). ¹H NMR

(300 MHz, CD₃SOCD₃): d = 12.2 (s, 1H, OH), 7.32 (dd, 1H,

J = 8.2 Hz, J = 1.8 Hz, H-4), 7.27 (d, 1H, J = 1.8 Hz, H-6), 7.00 (d,

1H, J = 8.2 Hz, H-3), 6.10 (s, 2H, OCH₂O). ¹³C NMR (75 MHz,

CD₃SOCD₃): d = 149.1, 147.6, 135.0, 126.5, 121.5 (C-4), 108.2 (C-

3), 106.1 (C-6), 101.8 (OCH₂O).

4.6.4. 3-(4-Nitrophenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,2,4-oxadiazole (17e)

4.6. [3+2] Cycloaddition reactions

A solution composed of 9 (378 mg, 0.62 mmol), 16e (630 mg,

3.12 mmol) and Et₃N (610 L, 4.86 mmol) was treated according

4.6.1. 3-Phenyl-5-(2,3,4,6-tetra-O-benzoyl-b-D-glucopyranosyl)-

1,2,4-oxadiazole (17b)

to method E. The residue was puriﬁed by ﬂash silica gel column

chromatography (PE/EtOAc, 4:1) to afford 17e (480 mg, 99%) as

an orange solid. R_f= 0.31 (PE/EtOAc, 4:1). Mp = 78–79 °C (CH₂Cl₂/

A solution composed of 9 (350 mg, 0.58 mmol), 16b (450 mg,

2.89 mmol) and Et₃N (600 L, 4.33 mmol) was treated according

to method E. The residue was puriﬁed by ﬂash silica gel column

chromatography (PE/EtOAc, 4:1) to afford 17b (400 mg, 94%) as a

yellow solid. R_f= 0.34 (PE/EtOAc, 4:1). Mp = 71–72 °C (CH₂Cl₂/PE).

PE). ½a ²_D⁰

ꢄ

¼ ꢂ9 (c 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): d = 8.21–

7.86 (m, 14H, H-ar), 7.53–7.25 (m, 10H, H-ar), 6.21 (t, 1H,

J = 9.4 Hz, H-3⁰), 6.09 (t, 1H, J = 9.4 Hz, H-2⁰), 5.96 (t, 1H,

J = 9.4 Hz, H-4⁰), 5.37 (d, 1H, J = 9.4 Hz, H-1⁰), 4.77 (dd, 1H,

J < 1 Hz, J = 12.3 Hz, H-6⁰a), 4.62 (dd, 1H, J = 4.4 Hz, J = 12.3 Hz, H-

6⁰b), 4.51 (m, 1H, H-5⁰). ¹³C NMR (75 MHz, CDCl₃): d = 174.8 (C-

5oxa), 167.3 (C-3oxa), 166.5, 166.2, 165.6, 165.4 (C@O), 149.9 (C-

1⁰⁰), 134.1, 133.9, 133.7, 132.3 (C-4⁰⁰), 130.3, 130.2 (3C), 129.8

(C^IV-ar), 129.0 (C^IV-ar), 128.9 (3C), 128.8 (2C), 124.4, 77.7 (C-5⁰),

74.0 (C-3⁰), 72.9 (C-1⁰), 71.2 (C-2⁰), 69.5 (C-4⁰), 63.4 (C-6⁰). MS

(LSIMS, NBA) m/z = 770 [M+H]⁺. HRMS (LSIMS, NBA) m/

z = C₄₂H₃₂N₃O₁₂[M+H]⁺calcd 770.1986, found 770.1983.

a ²_D⁰

ꢄ

¼ þ5 (c (c 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): d = 8.05–

7.84 (m, 10H, H-ar), 7.53–7.24 (m, 15H, H-ar), 6.12 (m, 2H, H-2⁰,

H-3⁰), 5.92 (t, 1H, J = 9.5 Hz, H-4⁰), 5.30 (d, 1H, J = 9.5 Hz, H-1⁰),

4.73 (dd, 1H, J = 2.2 Hz, J = 12.4 Hz, H-6⁰a), 4.59 (dd, 1H, J = 4.9 Hz,

J = 12.4 Hz, H-6⁰b), 4.43 (m, 1H, H-5⁰). ¹³C NMR (75 MHz, CDCl₃):

d = 173.3 (C-5oxa), 168.3 (C-3oxa), 166.0, 165.7, 165.0, 164.7

(C@O), 133.4, 133.3, 133.2, 133.0, 131.2, 129.7 (2C), 129.6 (3C),

129.3 (C^IV-ar), 128.6 (2C), 128.5 (2C^IV-ar), 128.4 (C^IV-ar), 128.3

(2C), 128.2 (3C), 127.4 (2C), 125.9 (C-1⁰⁰), 77.2 (C-5⁰), 73.6 (C-3⁰),

72.4 (C-1⁰), 70.6 (C-2⁰), 69.0 (C-4⁰), 62.9 (C-6⁰). MS (LSIMS, NBA)

m/z = 725 [M+H]⁺. HRMS (LSIMS, NBA) m/z = C₄₂H₃₃N₂O₁₀[M+H]⁺

calcd 725.2135, found 725.2132.

4.6.5. 3-(4-Hydroxyphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,2,4-oxadiazole (17g)

A solution composed of 9 (500 mg, 0.82 mmol), 16g (700 mg,

4.13 mmol) and Et₃N (861 L, 5.29 mmol) was treated according

4.6.2. 3-(4-Methylphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,2,4-oxadiazole (17c)

to method E. The residue was puriﬁed by ﬂash silica gel column

A solution composed of 9 (385 mg, 0.64 mmol), 16c (540 mg,

3.18 mmol) and Et₃N (665 L, 4.77 mmol) was treated according

to method E. The residue was puriﬁed by ﬂash silica gel column

chromatography (PE then PE/EtOAc, 7:3) to afford 17 g (250 mg,

41%) as a white gum. R_f= 0.73 (PE/EtOAc, 1:1). ½a _D²⁰

¼ ꢂ11:7 (c

ꢄ

1.05, CH₂Cl₂). ¹H NMR (500 MHz, CDCl₃): d = 8.07–7.82 (m, 10H,

H-ar), 7.57–7.28 (m, 12H, H-ar), 6.85 (d, 2H, J = 8.7 Hz, H-3⁰⁰, H-

chromatography (PE/EtOAc, 7:3) to afford 17c (250 mg, 53%) as a

pale yellow foam. R_f= 0.40 (PE/EtOAc, 7:3). ½a _D²⁰

¼ ꢂ3 (c 1, CH₂Cl₂).

ꢄ

4784

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

5⁰⁰), 6.09–6.00 (m, 2H, H-2⁰, H-3⁰), 5.86 (t, 1H, J = 9.9 Hz, H-4⁰), 5.24

(d, 1H, J = 8.8 Hz, H-1⁰), 4.72 (pdd, 1H, J < 1 Hz, J = 12.2 Hz, H-6⁰a),

4.57 (dd, J = 5.1 Hz, J = 12.2 Hz, H-6⁰b), 4.43–4.36 (m, 1H, H-5⁰).

¹³C NMR (125 MHz, CDCl₃): d = 174.8 (C-5oxa), 173.0 (C-3oxa),

168.1, 166.2, 165.8, 165.1 (C@O), 164.8, 158.3 (C-4⁰⁰), 133.6 (C^IV-

ar), 133.8, 133.5, 133.2, 129.9, 129.8 (C-2⁰⁰, C-6⁰⁰), 129.4, 128.6,

128.56, 128.46, 128.38, 118.7 (C-1⁰⁰), 115.7 (C-3⁰⁰, C-5⁰⁰), 77.2 (C-

5⁰), 73.7 (C-3⁰), 72.5 (C-1⁰), 70.6 (C-2⁰), 69.0 (C-4⁰), 63.0 (C-6⁰). MS

(ESI) m/z = 741.0 [M+H]⁺, 763.0 [M+Na]⁺. HRMS (ESI) m/

z = C₄₂H₃₂N₂O₁₁[M+H]⁺calcd 741.2084, found 741.2082.

J = 17.5 Hz, H-6⁰a), 4.58 (dd, 1H, J = 5.0 Hz, J = 17.5 Hz, H-6⁰b),

4.40 (ddd, 1H, J = 3.0 Hz, J = 5.0 Hz, J = 10.0 Hz, H-5⁰). ¹³C NMR

(125 MHz, CDCl₃): d = 173.0 (C-5oxa), 168.1 (C-3oxa), 166.1,

165.8, 165.1, 164.7 (C@O), 161.2 (C-4⁰⁰), 136.3 (C^IV-ar), 133.6,

133.4, 133.3, 133.2, 129.9, 129.8 (C^IV-ar), 129.4, 129.2, 128.6 (C^IV-

ar), 128.4, 128.3 (OCH₂Ph), 128.1, 127.5, 118.8 (C-1⁰), 115.0 (C-3⁰⁰,

C-5⁰⁰), 77.2 (C-5⁰), 73.7 (C-2⁰), 72.5 (C-1⁰), 70.6 (C-3⁰), 70.0 (OCH₂Ph),

69.0 (C-4⁰), 63.0 (C-6⁰). MS (ESI) m/z = 831.1 [M+H]⁺, 853.2

[M+Na]⁺. HRMS (ESI) m/z = C₄₉H₃₈N₂O₁₁[M+Na]⁺calcd 853.2373,

found 853.2375.

4.6.6. 3-(1-Naphthyl)-5-(2,3,4,6-tetra-O-benzoyl-b-

4.6.9. 3-(1,3-Benzodioxol-5-yl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,2,4-oxadiazole (17h)

glucopyranosyl)-1,2,4-oxadiazole (17k)

A solution composed of 9 (600 mg, 0.99 mmol), 16h (1.02 g,

4.96 mmol) and Et₃N (1.03 mL, 7.43 mmol) was treated according

to method E. The residue was puriﬁed by ﬂash silica gel column

chromatography (PE then PE/EtOAc, 7:3) to afford 17h (424 mg,

A solution composed of 9 (300 mg, 0.49 mmol), 16k (494 mg,

2.48 mmol) and Et₃N (518 L, 3.72 mmol) was treated according to

method E. The residue was puriﬁed by ﬂash silica gel column chro-

matography (PE/EtOAc, 4:1) to afford 17k (282 mg, 74%) as a white

55%) as a dark red foam. R_f= 0.65 (PE/EtOAc, 7:3). ½a _D²⁰

ꢄ

¼ þ3:0 (c

foam. R_f= 0.18 (PE/EtOAc, 4:1). ½a _D²⁰

ꢄ

¼ ꢂ11:0 (c 1, CH₂Cl₂). ¹H NMR

1, CH₂Cl₂). ¹H NMR (300 MHz, CDCl₃): d = 8.73 (dd, J = 1.4 Hz,

J = 15.8 Hz, H-8⁰⁰), 8.06–7.85 (m, 10H, H-ar), 7.54–7.25 (m, 16H,

H-ar), 6.10 (m, 2H, H-2⁰, H-3⁰), 5.88 (t, 1H, J = 9.9 Hz, H-4⁰), 5.30

(d, 1H, J = 9.0 Hz, H-1⁰), 4.73 (dd, 1H, J = 2.9 Hz, J = 12.5 Hz, H-6⁰a),

4.58 (dd, 1H, J = 5.2 Hz, J = 12.5 Hz, H-6⁰b), 4.42 (ddd, 1H,

J = 2.9 Hz, J = 5.2 Hz, J = 9.9 Hz, H-5⁰). ¹³C NMR (75 MHz, CDCl₃):

d = 172.4 (C-5oxa), 168.8 (C-3oxa), 166.1, 165.7, 165.1, 164.8

(C@O), 133.7 (C^IV-ar), 133.6, 133.5, 133.3, 131.1, 130.4 (C^IV-ar),

129.8, 129.7, 129.5, 129.4 (C^IV-ar), 128.6, 128.53 (C^IV-ar), 128.5,

128.4, 128.3, 127.5, 126.2, 126.0, 124.9, 123.0 (C^IV-ar), 77.2 (C-5⁰),

73.6 (C-3⁰), 72.5 (C-1⁰), 70.7 (C-2⁰), 69.0 (C-4⁰), 62.9 (C-6⁰). MS

(ESI) m/z = 775.1 [M+H]⁺, 797.1 [M+Na]⁺. HRMS (ESI) m/

z = C₄₆H₃₄N₂O₁₀[M+Na]⁺calcd 797.2111, found 770.2114.

(300 MHz, CDCl₃): d = 8.06–7.82 (m, 8H, H-ar), 7.57–7.23 (m, 14H,

H-ar), 6.79 (d, 1H, J = 8.1 Hz, H-ar), 6.08 (t, 1H, J = 9.5 Hz, H-2⁰),

6.03 (t, 1H, J = 9.5 Hz, H-3⁰), 5.97 (s, 2H, OCH₂O), 5.86 (t, 1H,

J = 9.5 Hz, H-4⁰), 5.22 (d, 1H, J = 9.5 Hz, H-1⁰), 4.70 (dd, 1H,

J = 2.9 Hz, J = 12.4 Hz, H-6⁰a), 4.55 (dd, 1H, J = 5.2 Hz, J = 12.4 Hz, H-

6⁰b),4.38 (ddd, 1H, J = 2.9 Hz, J = 5.2 Hz, J = 9.5 Hz, H-5⁰). ¹³C NMR

(75 MHz, CDCl₃): d = 173.1 (C-5oxa), 168.1 (C-3oxa), 166.1, 165.7,

165.0, 164.7 (C@O), 150.2 (C^IV-ar), 148.0 (C^IV-ar), 133.5, 133.4,

133.3, 133.1, 129.8, 129.7, 129.4 (C^IV-ar), 128.5 (C^IV-ar), 128.4,

128.3, 122.5, 119.8 (C^IV-ar), 108.5, 107.4, 101.5 (OCH₂O), 77.1 (C-

5⁰), 73.6 (C-2⁰), 72.4 (C-1⁰), 70.6 (C-3⁰), 69.0 (C-4⁰), 62.9 (C-6⁰). MS

(ESI) m/z = 769 [M+H]⁺, 1537 [2 M+H]⁺, 1559 [2 M+Na]⁺. HRMS

(ESI) m/z = C₄₃H₃₃N₂O₁₂[M+H]⁺calcd 769.2033, found 769.2035.

4.6.7. 3-(2-Naphthyl)-5-(2,3,4,6-tetra-O-benzoyl-b-

-glucopyr-

4.7. Enzymology

anosyl)-1,2,4-oxadiazole (17i)

A solution composed of 9 (345 mg, 0.57 mmol), 16i (586 mg,

2.85 mmol) and Et₃N (595 L, 4.86 mmol) was treated according

Glycogen phosphorylase b was prepared from rabbit skeletal

muscle according to the method of Fischer and Krebs,⁴²using dithi-

to method E. The residue was puriﬁed by ﬂash silica gel column

othreitol instead of L-cysteine, and recrystallized at least three

chromatography (PE/EtOAc, 4:1) to afford 17i (342 mg, 77%) as a

times before use. Kinetic experiments were performed in the direc-

tion of glycogen synthesis as described previously.⁴³Kinetic data

for the inhibition of rabbit skeletal muscle glycogen phosphorylase

pale yellow foam. R_f= 0.24 (PE/EtOAc, 4:1). ½a _D²⁰

¼ ꢂ10 (c 1,

ꢄ

CH₂Cl₂). ¹H NMR (300 MHz, CDCl₃): d = 8.42 (s, 1H, H-ar), 8.00–

7.90 (m, 12H, H-ar), 7.25–7.15 (m, 14H, H-ar), 6.10–6.00 (m, 2H,

H-2⁰, H-3⁰), 5.88 (t, 1H, J = 9.8 Hz, H-4⁰), 5.28 (d, 1H, J = 9.4 Hz, H-

1⁰), 4.73 (dd, 1H, J = 2.8 Hz, J = 12.5 Hz, H-6⁰a), 4.58 (dd, 1H,

J = 5.0 Hz, J = 12.5 Hz, H-6⁰b), 4.41 (ddd, 1H, J = 2.8 Hz, J = 5.0 Hz,

J = 9.8 Hz, H-5⁰). ¹³C NMR (75 MHz, CDCl₃): d = 173.4 (C-5oxa),

168.5 (C-3oxa), 166.1, 165.8, 165.1, 164.8 (C@O), 134.6 (C^IV-ar),

133.6, 133.5, 133.4, 133.2, 132.8 (C^IV-ar), 129.8, 129.7, 129.4,

128.8, 128.6, 128.5 (C^IV-ar), 128.4, 128.3, 128.2, 127.8, 127.5,

126.7, 123.7, 123.4 (C^IV-ar), 77.2 (C-5⁰), 73.6 (C-3⁰), 72.6 (C-1⁰),

70.6 (C-2⁰), 69.0 (C-4⁰), 63.0 (C-6⁰). MS (ESI) m/z = 775.0 [M+H]⁺,

1549.7 [2M+H]⁺, 1570.8 [2M+Na]⁺. HRMS (ESI) m/z = C₄₆H₃₅N₂O₁₀

[M+H]⁺calcd 775.2292, found 775.2293.

were collected using different concentrations of

a-D-glucose-1-

phosphate (2–20 mM), constant concentrations of glycogen (1%

w/v) and AMP (1 mM), and various concentrations of inhibitors.

Inhibitors were dissolved in dimethyl sulfoxide (DMSO) and di-

luted in the assay buffer (50 mM triethanolamine, 1 mM EDTA

and 1 mM dithiothreitol) so that the DMSO concentration in the as-

say should be lower than 1%. The enzymatic activities were pre-

sented in the form of double-reciprocal plots (Lineweaver–Burk)

applying a nonlinear data analysis program. The inhibitor con-

stants (K_i) were determined by Dixon plots, by replotting the

slopes from the Lineweaver–Burk plots against the inhibitor con-

centrations.^17,44The means of standard errors for all calculated ki-

netic parameters averaged to less than 10%.

4.6.8. 3-(4-Benzyloxyphenyl)-5-(2,3,4,6-tetra-O-benzoyl-b-D-

glucopyranosyl)-1,2,4-oxadiazole (17j)

Acknowledgments

A solution composed of 9 (222 mg, 0.37 mmol), 16j (482 mg,

1.83 mmol) and Et₃N (383 L, 2.75 mmol) was treated according

to method E. The residue was puriﬁed by ﬂash silica gel column

Financial support was received from the Hungarian Scientiﬁc

Research Fund (Grants: OTKA 45927, 60620 and 61336) and the

Hungarian National Ofﬁce for Research and Technology (Öveges

József Programme: HEF_06_3-T2DMDEBR). The authors thank the

Hungarian Academy of Sciences and CNRS for supporting their

cooperation in the frame of a Projet International de Coopération

Scientiﬁque (PICS 4576). Authors are also grateful to University

Lyon 1 and the Région Rhône-Alpes for ﬁnancial support (MIRA

Recherche 2006) and a PhD stipend to MB.

chromatography (PE then PE/EtOAc, 7:3) to afford 17j (211 mg,

69%) as a deep red foam. R_f= 0.40 (PE/EtOAc, 7:3). ½a _D²⁰

¼ ꢂ14:0

ꢄ

(c 1, CH₂Cl₂). ¹H NMR (500 MHz, CDCl₃): d = 8.09–8.03 (m, 2H, H-

ar), 7.95 (d, 2H, J = 7.0 Hz, H-2⁰⁰, H-6⁰⁰), 7.92–7.87 (m, 6H, H-ar),

7.58–7.31 (m, 17H, H-ar), 7.00 (d, 2H, J = 8.5 Hz, H-3⁰⁰, H-5⁰⁰), 6.07

(m, 2H, H-3⁰, H-4⁰), 5.87 (t, 1H, J = 9.5 Hz, H-2⁰), 5.23 (d, 1H,

J = 9.5 Hz, H-1⁰), 5.12 (s, 2H, OCH₂Ph), 4.72 (dd, 1H, J = 3.0 Hz,

M. Tóth et al. / Bioorg. Med. Chem. 17 (2009) 4773–4785

4785

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