Nano-sized clusters of a teicoplanin ψ-aglycon-fullerene conjugate. Synthesis, antibacterial activity and aggregation studies

Article abstract of DOI:10.1016/j.ejmech.2012.06.054

Glycopeptide antibiotic derivative teicoplanin ψ-aglycone has been bound covalently to a fullerenopyrrolidine derivative using azide-alkyne 1,3-dipolar cycloaddition reaction. The aggregation of the antibiotic-fullerene conjugate in aqueous solution has been studied. The conjugate exhibited antibacterial activity against enterococci resistant to teicoplanin.

Full text of DOI:10.1016/j.ejmech.2012.06.054

European Journal of Medicinal Chemistry 54 (2012) 943e948
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Nano-sized clusters of a teicoplanin
j-aglycon-fullerene conjugate. Synthesis,
antibacterial activity and aggregation studies
a
a
a
a
b
c
d
}
Szilvia Tollas , Ilona Bereczki , Attila Sipos , Erzsébet Roth , Gyula Batta , Lajos Daróczi , Sándor Kéki ,
Eszter Ostorházi e, Ferenc Rozgonyi e, Pál Herczegh a
,
*
a Department of Pharmaceutical Chemistry, University of Debrecen, Medical and Health Science Center, Egyetem tér 1, H-4032 Debrecen, Hungary
b Department of Organic Chemistry, University of Debrecen, Egyetem tér 1, H-4032 Debrecen, Hungary
c Department of Solid State Physics, University of Debrecen, Egyetem tér 1, P.O. Box 2, H-4010 Debrecen, Hungary
d Department of Applied Chemistry, University of Debrecen, Egyetem tér 1, H-4032 Debrecen, Hungary
e Microbiology Laboratory, Department of Dermatology, Venerology and Dermatooncology, Semmelweis University, Mária u. 41, H-1085 Budapest, Hungary
a r t i c l e i n f o
a b s t r a c t
Article history:
Glycopeptide antibiotic derivative teicoplanin
j-aglycone has been bound covalently to a full-
Received 24 April 2012
Received in revised form
15 June 2012
Accepted 27 June 2012
Available online 4 July 2012
erenopyrrolidine derivative using azide-alkyne 1,3-dipolar cycloaddition reaction. The aggregation of the
antibiotic-fullerene conjugate in aqueous solution has been studied. The conjugate exhibited antibac-
terial activity against enterococci resistant to teicoplanin.
Ó 2012 Published by Elsevier Masson SAS.
Keywords:
Teicoplanin
Fullerene
j-aglycone
1,3-Dipolar cycloaddition
Multivalent aggregates
1. Introduction
molecules results in derivatives with high antibacterial and in some
cases anti-influenza virus activity. We have also reported [9] that
Antimicrobial drug resistance is a serious and recurring problem
in the fight against infectious bacteria. Therefore, development of
new antibiotics is an important field of medicinal chemistry.
Glycopeptide type antibiotics vancomycin and teicoplanin are
widely used for treating dangerous Gram-positive bacterial infec-
tions that are resistant to other antibiotics.
Since glycopeptide-resistant enterococci and Staphylococcus
aureus as well as teicoplanin-resistant Staphylococcus haemolyticus
have emerged, in the past two decades an intensive research has
been directed towards the synthesis of new glycopeptide antibiotic
derivatives [1e3].
One of the successful ways of obtaining new semisynthetic
glycopeptide antibiotic derivatives, active against resistant bacteria,
is the introduction of lipophilic side chains into the antibiotic
molecules [4e6]. Recently, in a study of semisynthetic derivatiza-
tion of vancomycin, ristocetin and teicoplanin aglycons [7e9] we
have also found that attachment of lipophilic groups to those
such compounds form nanosized aggregates in water solution. The
glycopeptide-type antibiotics inhibit cell-wall biosynthesis of
Gram-positive bacteria by forming a stable hydrogen-bonded
complex with the terminal
D-alanyl-D-alanine sequence of the
muramyl pentapeptide intermediate formed during the biosyn-
thesis of mureide peptidoglycan [3]. Seven types of vancomycin
resistance in enterococci are known, VanC is an intrinsic resistance
phenotype in Enterococcus casseliflavus and Enterococcus gallina-
rum, the other six resistance mechanisms (VanA, B, D, G, L) are
acquired. The various acquired glycopeptide resistance genotypes
are associated with mobile genetic elements, which allow resis-
tance to spread not only clonally but laterally. In Europe the most
prevalent genotypes of glycopeptide resistance are vanA and vanB,
resulting resistance to vancomycin and teicoplanin, or only to
vancomycin, respectively [10,11]. The resistance results from
a change in the peptidoglycan termini from
lactate [12] thus weakening the complex formation with the anti-
biotics. Since the -Ala- -Ala is a repeating unit of the bacterial
D-Ala-D-Ala to D-Ala-D-
D
D
peptidoglycan it can be assumed that its multivalent interactions
with multimeric glycopeptide antibiotics can enhance the binding
potency and the antibacterial effect. Indeed, covalent dimers
* Corresponding author. Tel.: þ36 52 512 900 22895; fax: þ36 52 512 914.
E-mail address: herczeghp@gmail.com (P. Herczegh).
0223-5234/$ e see front matter Ó 2012 Published by Elsevier Masson SAS.
http://dx.doi.org/10.1016/j.ejmech.2012.06.054

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S. Tollas et al. / European Journal of Medicinal Chemistry 54 (2012) 943e948
[13,14], trimers [15] and polymers [16] of vancomycin exhibited
higher antibacterial activity against resistant strains. Since multi-
valency can be achieved by self-assembled clusters of molecules,
we postulated that the very potent antibacterial activity of our
recently published, aggregating teicoplanin-J-aglycon derivatives
[9] originates from a cluster effect, from a multivalent interaction of
the antibiotic aggregate with the bacterial cell-wall repeating units.
Those derivatives contained lipophilic groups such as n-decyl or
biphenyl substituents. It was obvious for us to extend this principle
to other glycopeptide derivatives, introducing larger lipophilic
substituents into them. It is well known that fullerene (C60
)
derivatives can form self-assembled supramolecular nano-
structures in water [17]. Since derivatization of fullerene molecule
is well established [18], the biomedical applications of functional-
ized fullerene nanomaterials are an intensively studied area of
research [19]. It is assumed that covalent attachment of the bulky,
very lipophilic fullerene to the teicoplanin pseudoaglycone mole-
cule will result in amphiphilic adducts forming large clusters in
water solution and hopefully exhibiting antibacterial activity
against vancomycin resistant enterococci. Here we report the
synthesis and studies of cluster formation and evaluation of anti-
bacterial activity of such an antibiotic-C60 conjugate.
Scheme 2. Synthesis of 3,4,5-trisubstituted benzaldehyde building block 7.
carrying a propargyl group, ready for the subsequent click reaction
with 9 azido-aglycon derivative. Copper(I)-catalyzed 1,3-dipolar
cycloaddition (click) reaction of 8 with compound 9 resulted in
the final product 10 (Scheme 3).
2. Results and discussion
2.1. Chemistry
2.1.1. Preparation of teicoplanin
J-aglycone-fullerene conjugate
2.2. Studies on aggregation of 10
For the conjugation of the antibiotic molecule to fullerene we
chose the copper(I) catalyzed 1,3-dipolar cycloaddition reaction of
a terminal alkyne with an azido group known as “click” reaction
[20,21]. Since the water solubility of the aglycon is rather low we
introduced some hydrophilic tetraethylene glycol moieties on the
fullerene derivative bearing the terminal alkyne group. For the
derivatization of the fullerene molecule the versatile Prato reaction
was chosen [22], which is a 1,3-dipolar cycloaddition of an azo-
methine ylide generated by the thermal reaction of an N-alkyl
glycine with an aldehyde. For this purpose tetraethylene glycol
(TEG) derivative 1 [23] was alkylated with t-butyl bromoacetate
(2) and the acid protecting group was removed by hydrolysis to
give 3 glycine derivative. (Scheme 1). For obtaining the aldehyde
reaction partner methyl gallate (4) was alkylated with tetra-
ethylene glycol monotosylate [24] resulting in 5. The methox-
ycarbonyl group was then reduced (6) and the alcohol was
oxidized with MnO2 affording the 7 benzaldehyde derivative
(Scheme 2). Prato reaction of 3, 7 and fullerene resulted in the
pyrrolidine derivative 8 with four TEG chains and one of them
Since we expected that a possible antibacterial activity of
compound 10 against resistant bacteria could be explained by
formation of a self-assembled multivalent ligand from the amphi-
philic molecule of 10, the examination of its cluster formation
properties were obvious.
According to our dynamic light scattering studies the effective
diameter of the aggregates formed in the solution of 10 in the
concentration range 0.01e0.1 mg/ml increased considerably with
the increasing concentration. The effective diameter of aggregates at
a concentration of 0.1 mg/ml was found to be 134 nm and the particle
size increased up to 316 nm when the concentration was raised to
0.1 mg/ml (Fig. 1). These observations indicate that concentrated
solutions facilitate for the formation of larger-sized aggregates.
On transmission electron microscopy (TEM) images we have
detected aggregates with about 200 nm diameter (Fig. 2). Studying
clusterecluster aggregation phenomena of fullerene-cyclodextrin
conjugates, Samal and Geckeler [25] reported an unexpected
phenomenon: the formation of larger aggregates on dilution of
aqueous solution of the conjugates. One year later Hallwass et al.
[26] reported that their pulsed field gradient NMR studies were
inconsistent with those results obtained by Samal and Geckeler. In
water solution, in the minimal antibacterial concentration range,
our teicoplanin-pseudoaglycon fullerene conjugate 10 formed on
dilution definitely smaller aggregates than in higher concentration.
On the TEM electron micrograph, aggregates of 200 nm clusters of
10 can be detected demonstrating the complexity of the aggrega-
tion phenomenon. So we can conclude that at least for this
particular fullerene derivative we could not observe the unex-
pected solute aggregation phenomenon on dilution reported by
Samal and Geckeler.
2.3. Antibacterial activity of antibiotic-fullerene conjugate 10
Although there is an intensive research activity in the domain
of biomedical applications of functionalized fullerene-based
Scheme 1. Synthesis of tetraethylene glycol precursor 3.

S. Tollas et al. / European Journal of Medicinal Chemistry 54 (2012) 943e948
945
Scheme 3. Synthesis of the aimed fullerene derivative 10.
nanomaterials [19], our compound is the first antibiotic derivative
3. Conclusions
of fullerene. The antibacterial activity was evaluated on a panel of
various Gram-positive bacteria and is summarized in Table 1.
Compound 10 showed moderate activities against MSSA, MRSA,
lower than teicoplanin but exhibited activity against Enterococcus
faecalis 15376 VanAþ, which is completely resistant to vanco-
mycin and teicoplanin. This observation supports our hypothesis:
the aggregation of the antibiotic forms a multivalent ligand for
the repeating terminal D-Ala-D-Ala sequence of the bacterial cell
wall peptidoglycan. This multivalent interaction can form
stronger complexes. Glycopeptide resistance of enterococci and in
We suppose that in the case of compound 10 the multivalency
can overcome the known resistance mechanism towards glyco-
peptides as it has been demonstrated for E. faecalis 15376 VanAþ.
The higher activity of 10 compared to teicoplanin in case of various
S. epidermidis strains is also noteworthy. In summary, we can
conclude that searching for new glycopeptide antibiotic analogs
capable to form aggregates in solution is a promising way of
obtaining new compounds for fighting against resistant bacteria.
some cases of S. aureus is based on the replacement of the acyl-
D
-
4. Experimental section
Ala- -Ala terminus with an acyl- -Ala- -lactate terminus. In this
D
D
D
way one of the hydrogen bonds between the bacterial peptido-
glycan and the antibiotic is abolished, and the interaction
decreases by 3 orders of magnitude. We suppose that in the
case of compound 10 the multivalency can overcome this resis-
tance mechanism as it has been demonstrated for E. faecalis
15376 vanAþ. The higher activity of 10 compared to teicoplanin in
case of various Staphylococcus epidermidis strains is also
remarkable.
4.1. Materials
Teicoplanin pseudoaglycon azido derivative was prepared as
described in our previous work [9]. Fullerene was purchased from
SES Research, Houston, Texas. Azido derivative of teicoplanin
pseudoaglycon was prepared as described in ref 9. Tetraethylene
glycol, toluenesulfonyl chloride, sodium hydride, propargyl
bromide, trifluoroacetic acid were purchased from Aldrich. All

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S. Tollas et al. / European Journal of Medicinal Chemistry 54 (2012) 943e948
Table 1
350
300
250
200
150
100
50
Antibacterial activity of compound 10.
Bacteria
Teicoplanin
MIC ( g/ml)
Compound 10
m
Bacillus subtilis ATCC 6633
0.5
0.5
0.5
4
16
8
4
Staphylococcus aureus MSSA ATCC 29213
Staphylococcus aureus MRSA ATCC 33591
Staphylococcus epidermidis
2
ATCC 35984 biofilm
Staphylococcus epidermidis mecA
Enterococcus faecalis ATCC 29212
Enterococcus faecalis 15376 VanA
Enterococcus faecalis ATCC 51299 VanB
16
1
256
0.5
1
6
16
16
MIC: Minimum Inhibition Concentration, ATCC: American Typed Culture Collection,
MRSA: Methicillin Resistant Staphylococcus aureus, vanA: vanA gene positive, vanB:
vanB gene positive.
performed on a Brookhaven Light Scattering instrument equipped
with a BI-9000 digital dynamic correlator. The laser light source
was a solid state, vertically polarized laser operating at 533 nm. DLS
0
0
0.02
0.04
0.06
0.08
0.1
measurements were performed at 25 ꢀC at scattering angle
q
¼ 90ꢀ.
The autocorrelation function of the scattered light intensity was
acquired in the homodyne mode. For the determination of particle
sizes and particle size distributions the NNLS (Non-Negative
Constraint Least Squares) calculation combined with multiple pass
analysis was applied.
c (mg/mL)
Fig. 1. Dynamic light scattering results for compound 10.
solvents and reagents were purchased from commercial sources
and were used as received.
4.2.2. Transmission electron microscopy image analysis
4.2. Instrumental methods
TEM measurements were performed using a Jeol 2000FX-II
equipment operating at 200 kV accelerating voltage. Sample
preparation: the sample solutions were dried on 400 mesh copper
microgrids covered by amorphous carbon substrate layer.
NMR spectra were recorded with Bruker DRXII-500 and DRX-400
spectrometers in CDCl3 or DMSO-d6 solutions. Chemical shifts have
been reported in parts per million (ppm) downfield from tetrame-
thylsilane. The assignment of the final product was performed on the
basis of previously reported full assignation of teichoplanin aglycon
using 1D and 2D NMR techniques (HSQC) [7]. Matrix-assisted laser
desorption/ionization time of flight mass spectrometry (MALDI-TOF
MS) analysis of products was carried out using Bruker BIFLEX III
mass spectrometer (Bruker Daltonik GmbH, Germany).
4.3. Antibacterial activity
The efficacy of the prepared compounds was determined with
the broth micro dilution method according to the CLSI guideline.
Bacterial strains were grown on 5% bovine blood agar plates at
35 ꢀC overnight. Appropriate numbers of colonies were suspended
in physiological saline in order to reach the density of 0.5 McFar-
land for inoculation.
Stock solutions containing different concentrations of the
substances were prepared in either distilled water or H2O and
methanol (1:1) or H2O and DMSO (1:1), respectively, depending
on the solubility of the given preparation. These were two-fold
4.2.1. Size distribution using dynamic light scattering
The solutions of compound 10 in water in a concentration range
of 0.01e0.1 mg/ml were studied by dynamic light scattering (DLS)
serially diluted from 256 to 0.5
ler-Hinton broth, and then 100
m
m
g/ml in cation-adjusted Muel-
l of each dilution was trans-
ferred into microplate holes. Inoculation was carried out
with 10
ml of each bacterial suspension. Incubation was per-
formed at 35 ꢀC for 18 h and determination of the minimal
inhibitory concentration (MIC) was made with the naked eyes on
a mirror.
4.4. Synthesis of the teicoplanin-j-aglycon-fullerene conjugate
4.4.1. Synthesis of 2
To the solution of 1 (763 mg, 3.3 mmol) in dry dichloromethane
(30 ml) triethylamine (555 l, 4.0 mmol) was added, and the
mixture was cooled to 0 ꢀC and stirred under N2 atmosphere.
t-Butyl bromoacetate (536 l, 3.63 mmol) was added dropwise to
m
m
the solution and it was stirred for overnight at rt. Then the mixture
was extracted with brine, the organic layer was dried (Na2SO4) and
the solvent was evaporated in vacuum. The residue was purified by
flash chromatography (CH2Cl2/CH3OH 95:5) to yield compound 2
(46%). dH (400 MHz; CDCl3; TMS) 1.41 (9H, s, Ce(CH3)3), 3.11 (2H, m,
Fig. 2. Transmission electron microscopy image of the cluster of compound 10.

S. Tollas et al. / European Journal of Medicinal Chemistry 54 (2012) 943e948
947
NHeCH2), 3.28 (2H, m, CH2eC^), 3.47e3.70 (17H, m, CH2CH2,
4.4.6. Synthesis of 8
C^CH, CH2CO); dC (100 MHz; CDCl3; TMS) 28.4, 59.8, 61.6, 70.0,
70.1, 70.2, 70.4, 70.5, 72.6, 80.3, 173.1;
Compound 3 (58 mg, 0.2 mmol) and compound 7 (136 mg,
0.2 mmol) were added to the solution of fullerene (72 mg,
0.1 mmol) in dry toluene (30 ml), and the mixture was stirred under
reflux for 6 h. Then the solvent was evaporated in vacuum, and the
residue was purified by flash chromatography (CH2Cl2/CH3OH 9:1
and 0.1% triethylamine) to yield compound 8 (20%).
MS (MALDI-TOF) m/z calcd for C17H31NO6Naþ [M þ Na]þ: 368.21,
found: 368.25.
4.4.2. Synthesis of 3
Compound 2 (528 mg, 1.53 mmol) was dissolved in dry
dichloromethane (30 ml) and stirred with 10 ml of 80% trifluoro-
acetic acid overnight, then the mixture was concentrated under
vacuum to yield compound 3 (87%).
dH (400 MHz; CDCl3; TMS) 2.47 (1H, m, NH), 3.21 (2H, m,
CH2eCH2eNH), 3.30 (1H, t, C^CH), 3.47e3.94 (16H, m, CH2CH2),
4.18 (2H, m, CCH2), 4.40e4.72 (2H, m, CH2NH), 8.90 (1H, br s,
COOH); dC (100 MHz; CDCl3; TMS) 47.3, 58.2, 60.2, 68.7, 69.6, 70.0,
70.2, 70.6, 72.4, 75.0, 169.5.
dH (500 MHz; CDCl3; TMS) 2.04e2.39 (4H, m, OH), 2.44 (2H, s,
NeCH2eCH2), 3.41e3.49 (2H, m, NeCH2eC60), 3.50e3.78 (42H, m,
CH2CH2), 3.91e4.08 (2H, m, PhOCH2), 4.09e4.57 (6H, m, PhOCH2,
NeCH2eCH2), 5.03 (1H, s, C^CH), 5.19 (1H, s, NeCHePh), 5.22 (1H,
s, N-CH-Ph), 7.18 (1H, s, aromatic H), 7.33 (1H, s, aromatic H); dC
(100 MHz; CDCl3; TMS) 52.7, 58.4, 63.8, 67.2, 68.7, 68.8, 69.1, 70.0,
70.1, 70.2, 72.2, 72.3, 74.3, 75.9, 76.4, 76.7, 82.0, 108.7, 108.8, 128.5,
132.3, 135.3, 135.5, 136.0, 136.2, 138.0, 139.2, 139.4, 139.7, 139.8,
141.2, 141.3, 141.4, 141.5, 141.7, 141.8, 141.9, 142.2, 142.3, 142.6, 142.8,
144.0, 144.1, 144.3, 144.4, 144.8, 144.9, 145.1, 145.4, 145.5, 145.6,
145.8, 145.9, 146.1, 146.5, 147.0, 153.0, 153.4, 153.7, 155.9; MS
(MALDI-TOF) m/z calcd for C103H75NO19Naþ [M þ Na]þ: 1652.55,
found: 1652.48.
4.4.3. Synthesis of 5
Compound 4 (920 mg, 5 mmol) was dissolved in dry dime-
thylformamide (35 ml) in the presence of K2CO3 (2.76 g, 20 mmol),
KI (830 mg, 5 mmol) and tetraethylene glycole monotosylate (7.0 g,
20 mmol). The reaction mixture was stirred for 24 h at 80 ꢀC. After
the solvent was removed under vacuum, dichloromethane was
added to the residue and it was extracted with brine (2 ꢁ 60 ml),
and then washed with water (60 ml). The organic layer was dried
(Na2SO4), then the mixture was concentrated under vacuum. The
residue was purified by flash chromatography (CH2Cl2/CH3OH
95:5) to yield compound 5 (48%).
dH (500 MHz; CDCl3; TMS) 2.70 (3H, br s, OH), 3.55e3.75 (45H,
m, CH2CH2, OCH3), 3.80e3.90 (2H, 2t, PhOCH2), 4.20e4.30 (4H, 2t,
PhOCH2), 7.30 (2H, s, C6H2); dC (100 MHz; CDCl3; TMS) 52.7, 62.2,
69.4, 70.2, 70.9, 71.0, 71.2, 71.4, 73.1, 73.2, 109.5, 125.6, 152.8, 165.6;
MS (MALDI-TOF) m/z calcd for C32H56O17Naþ [M þ Na]þ: 735.34,
found: 735.30.
4.4.7. Synthesis of 10
Compound 8 (32.6 mg, 0.02 mmol) was dissolved in the mixture
of dry dimethylformamide (4 ml) and dry toluene (1 ml) and stirred
under Ar atmosphere. Compound 9 (28.4 mg, 0.02 mmol), trie-
thylamine (6 ml, 0.04 mmol) and Cu(I)I (5 mg, 0.026 mmol) were
added to the solution, and it was stirred overnight. It was filtrated,
the solvent was evaporated in vacuum and the residue was purified
by flash chromatography (first with toluene/methanol 6:4, then
with toluene/dimethylformamide 1:1 and finally with
dimethylformamide).
dH (500 MHz; DMSO-d6) 2.87 (2H, s, NeCH2eCH2), 3.14e4.22
(58H, m, CH2CH2, PhOCH2), 4.28 (1H, s, x5), 4.36 (1H, s, c1),
4.41e4.43 (2H, m, NeCH2eCH2), 4.81 (1H, s, x2), 5.05 (1H, m, 4f),
5.24 (2H, m, NeCHePh), 5.39 (1H,s, x3), 5.54 (1H, m, 4b), 5.61 (1H,s,
x4), 6.30 (1H, m, 7f), 6.32 (1H, m, 3b), 6.34 (1H, m, 3d), 6.41 (1H, m,
3f), 6.48 (1H, m, 7d), 6.62 (1H, m, 5e), 6.63 (1H, m, 1b), 6.65 (1H, m,
5f), 7.03 (1H, m, 1e), 7.11 (1H, m, 5b), 7.16 (1H, m, 1f), 7.18 (1H, m,
2b), 7.19 (1H, m, 6e), 7.20 (1H, s, aromatic H), 7.21 (1H, s, aromatic
H), 7.31 (1H, m, 2e), 7.46 (1H, m, 6f), 7.68 (1H, m, 2f), 7.71 (1H, m,
6b); MS (MALDI-TOF) m/z calcd for C169H131Cl2N11O42Naþ
[M þ Na]þ: 3079.88, found: 3079.53.
4.4.4. Synthesis of 6
Compound 5 (977 mg, 1.37 mmol) and NaBH4 (993 mg,
21 mmol) were dissolved in abs. ethanol and stirred for 48 h at rt.
Then the precipitate formed after the addition of 0.1 M HCl was
filtered, and the filtrate was concentrated in vacuum. The residue
was dissolved in water (60 ml), and was stirred with Serdolit blue
anion exchange resin for 15 min, then with Serdolit red cation
exchange resin for another 15 min. After filtration the solvent was
evaporated in vacuum, and the residue was purified by flash
chromatography (CH2Cl2/CH3OH 85:15) to a yield compound 6
(79%).
dH (500 MHz; CDCl3; TMS) 2.60e2.90 (4H, m, OH), 3.55e3.75
(42H, m, CH2CH2), 3.75e3.85 (2H, 2t, PhOCH2), 4.10e4.20 (4H, 2t,
PhOCH2), 4.55 (2H, s, PhCH2), 6.60 (2H, s, C6H2); dC (100 MHz;
CDCl3; TMS) 68.7, 69.5, 70.2, 70.3, 70.5, 71.7, 71.4, 72.5, 72.6, 108.8,
124.9, 152.1; MS (MALDI-TOF) m/z calcd for C31H56O16Naþ
[M þ Na]þ: 707.35, found: 707.27.
Acknowledgment
The work is supported by the TÁMOP 4.2.1/B-09/1/KONV-2010-
0007 project. The project is co-financed by the European Union and
the European Social Fund. Financial support of the Hungarian
Research Fund (K 79126, K 101850) is also acknowledged.
Appendix A. Supplementary material
4.4.5. Synthesis of 7
Supplementary material associated with this article can be
found, in the online version, at http://dx.doi.org/10.1016/j.ejmech.
2012.06.054.
To compound
6 (700 mg, 1.02 mmol) dissolved in dry
dichloromethane (70 ml), MnO2 (3.5 g, 0.04 mmol) was added and
the reaction mixture was stirred vigorously for 1 h. Then the
mixture was filtered and the solvent was evaporated in vacuum to
yield compound 7 (92%).
References
dH (500 MHz; CDCl3; TMS) 2.18e2.27 (4H, br s, OH), 3.51e3.74
(42H, m, CH2CH2), 3.80e3.90 (2H, 2t, PhOCH2), 4.19e4.31 (4H, 2t,
PhOCH2), 7.15 (2H, s, C6H2), 9.83 (1H, s, CHO); dC (100 MHz; CDCl3;
TMS) 61.3, 68.5, 69.2, 69.9, 70.0, 70.1, 70.2, 70.4, 72.2, 108.5, 131.2,
152.6, 190.6; MS (MALDI-TOF) m/z calcd for C31H54O16Naþ
[M þ Na]þ: 705.33, found: 705.30.
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