Synthesis of two and antibacterial activity of one novel oxime ether derivatives of erythromycin A

Article abstract of DOI:10.1016/S0014-827X(03)00194-0

The synthesis of novel erythromycin A 9-O-(2-ethenesulfony-ethyl)-oxime and erythromycin A 9-O-(3-oxo-butyl)-oxime from erythromycin A (EA) by the Michael reaction is described and to describe the effects of transformation of ketone in position 9 of EA to an oxime ether. This transformation occurred in a single step without protecting of any functional moiety of erythromycin oxime and zero waste manner in good yield. The antibacterial screen of EA 9-O-(2-ethenesulfony-ethyl)-oxime is also reported.

Full text of DOI:10.1016/S0014-827X(03)00194-0

Il Farmaco 58 (2003) 1011ꢀ1015
/
www.elsevier.com/locate/farmac
Synthesis of two and antibacterial activity of one novel oxime ether
derivatives of erythromycin A
H.A. Dondas a,*, N. Yaktubay b
a Department of Chemistry, Faculty of Pharmacy, Yenisehir Campus, University of Mersin, 33342 Mersin, Turkey
b Department of Pharmacology, Faculty of Medicine, University of Cukurova, Adana, Turkey
Received 3 January 2003; accepted 1 July 2003
Abstract
The synthesis of novel erythromycin A 9-O-(2-ethenesulfony-ethyl)-oxime and erythromycin A 9-O-(3-oxo-butyl)-oxime from
erythromycin A (EA) by the Michael reaction is described and to describe the effects of transformation of ketone in position 9 of EA
to an oxime ether. This transformation occurred in a single step without protecting of any functional moiety of erythromycin oxime
and zero waste manner in good yield. The antibacterial screen of EA 9-O-(2-ethenesulfony-ethyl)-oxime is also reported.
´
# 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
Keywords: Erythromycinerythromycin oxime ether; Michael addition
1. Introduction
EA’s resorption variability after oral administration was
its instability in the gastric juice [3].
Erythromycin A (EA) is the most widely used and
effective macrolide antibiotic against most Gram-posi-
tive and some Gram-negative bacteria [1]. EA is an
orally administered antibiotic and the main reason for
its resorption variability and consequently the low
antibiotic level in the serum is its instability in the
gastric juice. It is well known that in acidic condition EA
gives first an internal enolic ether and secondly an
internal ketal by reaction with the ketone in position 9
and hydroxy groups in position 6 and 12. Neither
product exhibits antibiotic activity and this ketal
formation is irreversible [2] and EA oxime and some
of its derivatives are much more stable to mineral acid
[3]. The transformation of the ketone in position 9 to an
oxime is a possible way of preventing internal ketaliza-
tion. As a semisynthetic macrolide the discovery of
roxitromycin, (9-[O[(2-methoxyethoxy)methyl]oxime]-
erythromycin), is the result of a rational and scientific
process, based on the fact that at least one reason for
This instability is due to the reactivity of the ketone in
position 9 in acidic medium and one chemical approach
was to mask it by an oxime function. Several derivatives
of EA ketone had been described including the hydra-
zone [4], numerous oxime, erythromycylamine and
erythromycylamine-aldehyde condensation products
[3ꢀ6] and oxime ether derivatives [7,8] but as EA 9-O-
/
(2-ethenesulfony-ethyl)-oxime (3) and EA 9-O-(3-oxo-
butyl)-oxime (4) were practically unexplored. It has been
reported that the reaction of oximes with activated
olefins: e.g. reaction of cyclopentanone oxime with
ethenesulfonyl-benzene, has provided cyclopentanone
O-(2-benzenesulfonyl-ethyl)oxime [9] and reaction of
propan-2-one oxime with acrylic acid methyl ester
provided isopropylaminooxy-propionic acid methylester
[10]. This versatile methodology has been applied to
synthesis the novel erythromycin oxime ether deriva-
tives.
In this paper, we have considered it of interest to
study synthesis and the effects of such modifications on
the spectrum of antibacterial activity. We anticipate that
this work might lead to the erythromycin oxime ethers
analogs with a different synthetic route and different
activity profile. Both compounds have been prepared
from EA oxime and activated olefins via Michael
* Corresponding author.
E-mail addresses: yakdas25@mersin.edu.tr, yakdas25@hotmail.
com (H.A. Dondas).
´
0014-827X/03/$ - see front matter # 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
doi:10.1016/S0014-827X(03)00194-0

1012
H.A. Dondas, N. Yaktubay / Il Farmaco 58 (2003) 1011ꢀ1015
/
Scheme 1.
reaction (Schemes 1 and 2) and the antibacterial
activities of EA 9-O-(2-ethenesulfony-ethyl)-oxime (3)
compared with that of EA (Tables 2a and 2b).
Table 1
1H chemical shifts (ppm) and 1Hꢀ1
(vinylysulfone)erythromycin A 9-ethoxime (3) at 600 MHz NMR in
/
H coupling constants (Hz) for O-
CDCl3
1.1. Synthesis and structure determination (chemistry)
Position
1H
Ma
Position
1H
Ma
1
22
23
24
25
4.41
3.33
t
EA oxime (2) [12,17] with divinylsulfone (toluene,
reflux, 3d) gave O-Michael adduct (3) in 76% yield. As
shown in Scheme 1 compound (2) underwent a facile
and chemoselective O-Michael addition upon treatment
with divinylsulfone to afford the O-Michael adduct (3)
in good yield in one pot reaction without protection of
any functional moiety of corresponding EA oxime (2).
The adduct (3) was isolated and the structures of it were
2
2.89
4.11
1.98
3.52
dq
bd
m
d
t
3
6.66
m
d
4
6.45(Ha)
6.16(Hb)
4.45
5
d
6
1?
2?
3?
4?
5?
6?
7?, 8?
1ƒ
2ƒ
3ƒ
4ƒ
5ƒ
6ƒ
7ƒ
8ƒ
d
7
1.90
2.38
dd
m
3.02
2.55
m
m
m
m
d
8
9
1.6
3.98
10
11
12
13
14
15
16
17
18
19
20
21
2.69
3.62
dq
d
1
2
determined by 600 MHz H NMR (Table 1), D-COSY
studies and FAB MS technique (see Section 3) and also
from the literature data of known erythromycin deriva-
1.28
2.15
s
5.07
1.54
0.85
1.18
1.08
1.45
1.17
1.29
1.12
dd
m
t
4.88
2.35
bd
dd
tives and by a comparison with previous study [11ꢀ
/
d
d
s
3.05
4.00
1.11
1.24
3.30
dd
dq
bd
s
d
d
s
s
Ma, multiplicity of 1H resonances.
13,17]. Such structure is also consistent with the known
chemistry of the erythromycins [14,15]. Previous studies
shown that direct O-alkylation of erythromycin oxime
allowed access to E-stereoisomers which are more
Scheme 2.

H.A. Dondas, N. Yaktubay / Il Farmaco 58 (2003) 1011ꢀ
/1015
1013
interesting than the Z-ones [6ꢀ
under giving conditions and from the spectral data, we
obtain same stereoisomers.
/
12]. We assume that,
2. Experimental
2.1. Material and methods
Similarly the reaction of (2) with methylvinyl ketone
(DCM, 25 8C, 7d) gave O-(methylketone)erythromycin
A 9-ethoxime (4) in 69% yield via a facile and
chemoselective O-Michael addition under mild condi-
tion in one pot reaction (Scheme 2). The structure of
Nuclear magnetic resonance spectra and decoupling
experiments were determined at 400 and 600 MHz on a
Bruker AM spectrometer. Chemical shifts are given in
parts per million (d) downfield from tetramethylsilane
as internal standard. Spectra were determined in deuter-
1
compound (4) was determined by 400 MHz H NMR,
2D-COSY studies and FAB MS technique (see Section
1
3). The H NMR spectra of (4) in CDCl3 shown the
iochloroform. The following abbreviations are used; sꢁ
singlet, dꢁdoublet, tꢁtriplet, qꢁquartet, mꢁmulti-
plet, brꢁbroad and brsꢁbroad singlet, brdꢁbroad
doublet. Flash column chromatography was performed
using silica gel 60 (230ꢀ400 mesh). Melting points were
/
/
/
/
/
/
/
/
pattern similar to those of (3), indicating the C-9
ketoxime to O-(methylketone)erythromycin A 9-ethox-
ime. Further the mass spectra (FAB MS) of O-alkyl
ketone EA 9-ethoxime derivatives (4) exhibited the
characteristic protonated molecular ion peaks with the
increasing mass units due to the additional alkyl ketone
groups (see Section 3). This was also confirmed by a
comparison of the results which were previously re-
/
determined on a Kofler hot stage apparatus and are
uncorrected. Mass spectra were recorded at 70 eV on a
VG Autospec mass spectrometer. Specific rotations
were measured at ambient temperature with an Optical
Activity Ltd, AA-1000 polarimeter. All solvents were
purified before use. Divinylsulfone and methylvinyl
ketone were purchased from Aldrich Chemical Com-
pany and ERY were obtained from Sigma.
ported for erythromycin oxime ethers [7ꢀ10,17].
/
It is also interesting to note that in both cases the
reactions proceeds chemoselectively in a single step
without protection any functional moiety of correspon
ding erythromycin oxime (2) and underwent a facile and
chemoselective O-Michael addition upon giving reac-
tion conditions (Schemes 1 and 2).
2.2. EA oxime (2) were prepared according to literature
procedure [12,17]
The product was obtained as colourless solid in 75%
The antibacterial activity of O-(vinylsulfone)erythro-
mycin A 9-ethoxime (3) against a range of bacteria has
been described. The compound tested has been found
active in some cases against Gram-positive bacteria
tested (Tables 2a and 2b).
yield. M.p. 170 8C, [a]Dꢁ
/
ꢂ70 (c, 1 g/100 ml, EtOH) m/
/
z (%) (FAB): 749 (Mꢃ1, 4), 591(2), 174(7), 158(100),
/
116 (49), 98(29) and 72(23). H and 13C chemical shifts
1
1
(ppm) and Hꢀ
/
1H coupling constants (Hz) have been
reported by McGill et al. [12].
In conclusion, the synthesis of novel O-(vinylsulfo-
ne)erythromycin A 9-ethoxime (3) and O-(vinylketo-
ne)erythromycin A 9-ethoxime (4) were achieved by O-
Michael addition reaction. This approach also opens up
the possibility of constructing other oxime ether deriva-
tives at the C-9 position of the macrolide core of
erythromycin. Since the main reason for low antibiotic
level of EA in the serum is instantly in the gastric juice
and its oxime ether derivatives are much more stable to
mineral acids, the result obtained can be considered
interesting enough for further investigations.
2.3. O-(Vinylysulfone)erythromycin A 9-ethoxime (3)
A solution of EA oxime (2) (0.3 g, 0.40 mmol) and
divinylsulfone (0.047 g, 0.4 mmol) in dry toluene (40 ml)
was stirred and boiled under reflux for 3 days. After
cooling the solvent was removed in vacuo and the
residue chromatographed on silica eluting with 1:9:90 v/
v NH3/MeOH/CH2Cl2 to afford the product (3) (0.264
g, 76%) as a pale yellow solid that crystallised from
dichloromethaneꢀ
118ꢀ120 8C. [a]Dꢀ
HRMS: 866.4809, C41H74N2O15S: 866.4798, m/z
(FAB): 867 (Mꢃ1, 5), 709(3), 174(9), 158(100),
/
hexane as colourless prisms, m.p.
/
/
ꢂ
/
64.8 (c 1 g/100 ml, CHCl3).
/
Table 2a
Antibacterial activity of EA and the compound (3)
116(38), 98(10), 72(13) and 59(3).
Test organism
MIC (mg/ml)
EA Comp. 3
2.4. O-(Methylketone)erythromycin A 9-ethoxime (4)
A solution of EA oxime (2) (0.1 g, 0.134 mmol), and
methyl vinyl ketone (56 mg, 0.268 mmol) in dry
dichloromethane (20 ml) was stirred under N2 at room
temperature for 7 days. The solvent was removed in
vacuo and the residue subjected to column chromato-
graphy eluting with 9:1 v/v chloroform/Et3N to afford
S. aureous ATCC 29213
S. epidermidis ATCC 12228
S. faecalis ATCC 29212
C. diphtheriae G 12/6
0.39
12.50
3.12
50
0.19
1.56
0.04
B
/
0.09
E. coli ATCC 25922
100
ꢀ200
/

1014
H.A. Dondas, N. Yaktubay / Il Farmaco 58 (2003) 1011ꢀ1015
/
Table 2b
The antibacterial screen of EA-vinylsulfonyl ethoxime (3)
Test organism
Comp. (3) a
EA
Test organism
Comp. (3) a
EA
B. fragilis B70
B. fragilis BC1
4
8
1
Pr. vulgaris H
ꢀ
/
64
64
64
64
64
64
8
ꢀ
/
64
64
64
16
64
64
0.5
2
P. aeruginosa 55528
P. aeruginosa Badia
P. aeruginosa K799 61
P. aeruginosa 799 wt
S. marscescens S6
B. cereus BRL 1243
B. subtilis ATCC 6633
E. feacalis I
ꢀ
/
ꢀ
/
B. fragilis NCTC10581
E. coli 10418
8
64
ꢀ/
ꢀ/
4
E. coli DCO
E. coli DC2
ꢀ
/
64
64
64
4
64
4
ꢀ
/
ꢀ/
ꢀ/
E. coli DCOTEM-1
E. coli ESS
ꢀ/
32
0.125
0.125
0.06
4
B
/
E. coli JT425
E. coli K12/TEM-5
E. cloaae N1
ꢀ
/
64
64
64
64
64
64
64
64
16
64
64
4
4
0.125
0.125
0.125
0.125
0.125
ꢀ
/
S. aureus carter 37
S. aureus F89
8
ꢀ
/
ꢀ
/
8
E. cloaae P99
ꢀ/
S. aureus oxford
8
H. influenzae Q1
H. influenzae WM493
K. pneumoniae E70
Morax. Catarhalis 1502
Mora. CatarhalisRavasio
Morg. Morganii T361
P. mirabilis C889
S. aureus Russell
8
4
64
S. aureus V573
ꢀ/
64
8
ꢀ/64
ꢀ/
ꢀ
/
S. epidermidisPHLN20
S. agalactiae Hester
S. pneumoniae 1761
S. pneumoniae ERY2
S. pneumoniae PU 7
S. ppyogenes CN10
0.125
0.06
0.06
4
0.125
B
B
/0.06
B
/
0.06
0.125
B
B
/
1
64
64
/0.06
/
ꢀ
/
64
64
16
0.125
0.125
ꢀ/
ꢀ/
B
B
/0.06
/0.06
a
Values are MICs (minimum inhibitory concentrations in microgram/millilitre).
the O-Michael adduct (4) (0.075 g, 69%), m.p. 154ꢀ
/
volume of ethanol and adding Mueller-Hinton Broth
(MHB, Oxoid). Then the stock solutions were diluted
with MHB to obtain final drug concentrations ranging
from 400 to 0.08 mg/ml. Bacterial suspensions were
prepared from fresh cultures according to 0.5 McFar-
land Standards and were diluted with MHB to reach a
157 8C, [a]ꢁ
/
ꢂ57.5 (1 g/100 ml, CHCl3), m/z (%)
/
(FAB): 819 (Mꢃ, 37), 158(44), 109(31), 83(56) and
55(100). HRMS: 818.5119, C41H74N2O14: 818.5140, d
(400 MHz, partial data): 2.61(s, 3H, MeCꢁ
/
O), 4.43(m,
2H, H22) and 3.66(m, 2H, H24).
The 1H NMR spectra of (4) in CDCl3 shown the
pattern similar to those of (3), indicating the C-9
ketoxime to O-(methylketone)erythromycin A 9-etho-
xime. Further the mass spectra (FAB MS) of O-alkyl
ketone EA 9-ethoxime derivatives (4) exhibited the
characteristic protonated molecular ion peaks with the
increasing mass units due to the additional alkyl ketone
groups.
concentration of 1ꢄ
series were inoculated with 1 ml of bacterial suspensions
to obtain a final concentration of approximately 5ꢄ
105
CFU/ml. The inoculated tubes were incubated for 16ꢀ
/
106 CFU/ml. Antibiotic solutions
/
/
20 h at 35 8C. C. diphtheriae were reincubated for an
additional 24 h and were reread at 48 h. Minimum
inhibitory concentrations (MIC) were read as the lowest
concentration at which there was no visible growth. (*)
NLCLLS Method for Dilution Antibacterial Suscep-
tibility Test for Bacteria that Grow Aerobically*Third
/
Edition; Approved Standard NCCLS document M7-A3
NCCLS, 771 East Lancester Avenue, Villanova, PA,
1985.
3. In vitro antibacterial activity studies
3.1. Materials and methods
The following organisms were selected for use in the
study [16] Staphylococcus aureus, ATCC 29213, Staphy-
lococcus epidemidis ATCC 12228 Streptococcus faecalis,
ATCC 29212, Corynebacterium diphtheriae G 12/6 and
Escherichia coli ATCC 25922.
In vitro antibacterial activities of EA and the com-
pound were investigated in duplicate (two separate
studies) by macrodilution broth method. This was
done according to the National Committee for Clinical
Laboratory Standards (NLCLLS) M7-A3 guidelines (*).
For this purpose: Stock solutions of the drugs were
prepared by dissolving 10-mg drug in appropriate
4. Result and discussion
The in vitro antibacterial activity of O-(vinylsulfo-
ne)erythromycin A 9-ethoxime (3) against S. aureus, S.
epidemidis, S. faecalis, C. diphtheriae, E. coli has been
described previously. The compound tested has been
found active in some cases against Gram-positive
bacteria tested (Table 2a). In general, the transforma-
tion of the ketone in position 9 of EA to an oxime
resulted in a decrease of in vitro antibacterial activity
against the organism tested, but the drop in activity was

H.A. Dondas, N. Yaktubay / Il Farmaco 58 (2003) 1011ꢀ
/1015
1015
Erythromycin A 11, 12-methylene acetal, J. Antibiot. 40 (1988)
1644ꢀ1648.
[6] R.S. Egan, L.A. Freigberg, W.H. Washburn, Configuration of 9-
imino derivatives of erythromycin, J. Org. Chem. 39 (1974) 2492ꢀ
not so dramatic except for S. faecalis and E. coli.
Therefore, some more biological assays has also been
done and reported in Table 2b [17].
/
/
2494.
[7] (a) E.G. Brain, A.K. Forrest, E. Hunt, C. Shillingford, J.M.
Wilson, Erythromycin A oxime 11, 12-carbonate and its oxime
Acknowledgements
ethers, J. Antibiot. 41 (1989) 1817ꢀ1822;
/
(b) A. Nishimoto, K. Narita, S. Ohimoto, Y. Takahashi, S.
Yoshizumi, T. Yoshida, N. Kado, E. Okezaki, H. Kato, Studies
on macrolide antibiotics I. Synthesis and antibacterial activity of
erythromycin A 9-O-substituted oxime ether vderivatives against
Mycobacterium avium complex, Chem. Pharm. Bull. 49 (2001)
We are indebted to Prof. R. Grigg (Leeds University)
for helpful discussions. We thank Mersin University and
Cukurova University for financial support and Leeds
University for 600 MHz and mass spectral determina-
tion.
1120ꢀ
[8] J.C. Gasc, S.G. D’Ambrieres, A. Lutz, J.F. Chantot, New ether
oxime derivatives of erythromycin A a structureꢀactivity relation-
ship study, J. Antibiot. 44 (1991) 313ꢀ325.
[9] (a) P. Armstrong, R. Grigg, W.J. Taylor, Cycloaddition reactions
of oximes; powerful new carbonꢀcarbon bond forming metho-
dology, J. Chem. Soc. Chem. Commun. (1987) 1325ꢀ1327;
/1127.
/
/
References
/
/
[1] (a) J.A. Washington, W.R. Wilson, Erythromycin A microbial
and clinical perspective after 30 year of clinical use, Mayo Clin.
Proc. 60 (1985) 189;
(b) P. Armstrong, R. Grigg, S. Surendrakumar, W.J. Warnock,
Tandem intramolecular Michael addition and 1,3-dipolar cy-
cloaddition reactions of oximes; versatile new carbonꢀ
bond forming methodology, J. Chem. Soc. Chem. Commun.
(1987) 1327ꢀ1328.
/carbon
(b) P.B. Fernandes, H.C. Neu, The macrolide revival: thirty-five
years after erythromycin, The Antimicrobic Newsletter 4 (1987)
/
27ꢀ
[2] (a) J.C.H. Mao, M. Putterman, The intermolecular complex of
erythromycin and ribozome, J. Mol. Biol. 44 (1969) 347ꢀ352;
/
35.
[10] E.L. Schumann, L.A. Paquette, R.V. Heinzelmann, D.P. Wallach,
J.P. DaVanzo, M.E. Greig, The synthesis g-aminobutyric acid
transaminase inhibition of aminooxy acids and related com-
/
(b) S. Burger, in: M.E. Wolf (Ed.), Medicinal Chemistry and Drug
Discovery, vol. 2, Wiley, USA, 1995, p. 481.
pound, J. Med. Pharm. Chem. (1962) 464ꢀ477.
/
[11] S. Morimoto, Y. Misawa, T. Adachi, T. Nagate, Y. Watanabe, S.
Omura, Chemical modification of erythromycins. Synthesis and
antibacterial activity of O-alkyl derivatives of erythromycin A, J.
[3] (a) J.F. Chantot, J.C. Gasc, S.G. D’Ambrieres and A. Lutz, New
ether oxime derivatives of erythromycin A: preparation and
antibacterial activities, Program and Abstracts of the 23rd
Interscience Conference on Antimicrobial Agents Chemotherapy,
Antibiot. XLIII (1990) 286ꢀ294.
/
[12] J.M. McGill, R. Johnson, Structrural and conformational analy-
No. 447, P 165, Las Vegas, October 24ꢀ26, 1983.;
/
sis of (E)-erythromycin A oxime, Magnet. Reson. Chem. 31
(b) J.F. Chantot, A. Bryskier, J.C. Gasc, Antibacterial activities of
roxithromycin: a laboratory evaluation, J. Antibiotics 39 (1986)
(1993) 273ꢀ
[13] J.R. Everett, J.W. Tyler, The conformational analysis of erythro-
mycin A, J. Chem. Soc. Perkin Trans. 2 (1987) 1659ꢀ1667.
[14] J. Barber, J.I. Gyi, L. Lian, G.A. Morris, D.A. Pye, J.K.
Sutherland, The structure of erythromycin A in [2H6] DMSO
and buffered D2O: full assignments of the 1H and 13C NMR
/
277.
660ꢀ
/
668.
[4] E. Wildsmith, The reaction of erythromycin hydrazone with
/
nitrous acid a new route to erythromycylamine, Tetrahedron Lett.
1 (1972) 29ꢀ30.
/
[5] (a) E.H. Massey, B.S Kitchel, L.D. Martin, K. Gerzon, Anti-
bacterial activity of 9(S)-erythromycylamine-aldehyde condensa-
spectra, J. Chem. Soc. Perkin Trans. 2 (1991) 1489ꢀ1494.
[15] J.R. Everett, E. Hunt, J.W. Tyler, Ketone-hemiacetal tautomer-
/
tion product, J. Med. Chem. 17 (1974) 105ꢀ
(b) E.H. Massey, B.S Kitchel, L.D. Martin, K. Gerzon, H.W.
Murphy, Erythromycylamine, Tetrahedron Lett. 2 (1970) 157ꢀ
/
107;
ism in erythromycin A in non-aqueous solutions. An NMR
spectroscopic study, J. Chem. Soc. Perkin Trans. 2 (1991) 1481ꢀ
/
/
1487.
160;
[16] Some parts of this work were presented at the Second Interna-
tional Meeting on Pharmacy and Pharmaceutical Sciences,
(c) P.A. Lartey, S.L. Deninno, R. Faghih, D.J. Hardy, J.J.
Clement, J.J. Plattner, Synthesis and activity of C-21 alkylamino
derivatives of (9R)-erythromycylamine, J. Antibiot. 45 (1992)
Istanbul, Turkey, 6ꢀ9 September, 1998.
/
[17] H.A. Dondas, Biochemical Application of Oxime Nitrone Cy-
cloaddition Cascades, Ph.D. thesis Leeds University, Leeds, UK,
1997.
380ꢀ385;
(d) E. Hunt, D.J.C. Knowles, C. Shillingford, I.I. Zomaya,
/