Synthesis, characterization and physicochemical analysis of some mannofuranoside derivatives with potent antimicrobial activity

Article abstract of DOI:10.13005/ojc/330606

We have synthesized protected mannofuranose as a glycosyl donor and some heterocyclic moieties as glycosyl acceptor. Coupling of glycosyl donor and acceptor by glycosylation, results in the formation of mannofuranoside derivatives. Antimicrobial potential of synthesized compounds were tested against five different human pathogenic bacteria and two fungi by using microdilution method. Interestingly, all synthesized mannofuranoside derivatives gave antimicrobial activity. Cumulatively, inhibitory concentration (IC₅₀) against bacteria were found to be in the range of 84.28-309.43 μg/ml, while, IC₅₀ against fungus were in the range of 0.59-3.82 mg/ml. Ampicillin and Fluconazole were used as positive control. Further, physicochemical parameters of synthesized mannofuranoside derivatives were analyzed using in-silico approach. Fortunately, none of our synthesized mannofuranoside derivatives gave violation of Lipinski’s Rule of Five. Thus, we can safely state that these synthesized mannofuranoside derivatives could be considered as potential anti-microbial drug candidates.

Full text of DOI:10.13005/ojc/330606

ISSN: 0970-020 X
CODEN: OJCHEG
017, Vol. 33, No.(6):
Pg. 2731-2741
ORIENTAL JOURNAL OF CHEMISTRY
An International Open Free Access, Peer Reviewed Research Journal
2
www.orientjchem.org
Synthesis, Characterization and Physicochemical Analysis
of Some Mannofuranoside Derivatives with Potent
Antimicrobial Activity
1
1
1
2
FIROJ HASSAN , MALIK NASIBULLAH , NASEEM AHMAD , AZHAR KAMAL , SYED
3
2
1
MOHAMMAD DANISH RIZVI , MOHD SAJID KHAN and ABDUL RAHMAN KHAN *
1
Department of Chemistry, Kursi Road, Integral University, Lucknow-226026, India.
2
Department of Biosciences, Kursi Road, Integral University, Lucknow-226026, India.
3
Department of Pharmacology & Toxicology, College of Pharmacy
University of Hail, PO Box no. 2440, Saudi Arabia.
*
Corresponding author E-mail: firojhassan42@gmail.com, arahman@iul.ac.in
http://dx.doi.org/10.13005/ojc/330606
(Received: August 25, 2017; Accepted: September 20, 2017)
ABSTRACT
We have synthesized protected mannofuranose as a glycosyl donor and some heterocyclic
moieties as glycosyl acceptor. Coupling of glycosyl donor and acceptor by glycosylation, results
in the formation of mannofuranoside derivatives. Antimicrobial potential of synthesized compounds
were tested against five different human pathogenic bacteria and two fungi by using microdilution
method. Interestingly, all synthesized mannofuranoside derivatives gave antimicrobial activity.
Cumulatively, inhibitory concentration (IC ) against bacteria were found to be in the range of
50
84.28-309.43 μg/ml, while, IC50 against fungus were in the range of 0.59-3.82 mg/ml. Ampicillin and
Fluconazole were used as positive control. Further, physicochemical parameters of synthesized
mannofuranoside derivatives were analyzed using in-silico approach. Fortunately, none of our
synthesized mannofuranoside derivatives gave violation of Lipinski’s Rule of Five. Thus, we can
safely state that these synthesized mannofuranoside derivatives could be considered as potential
anti-microbial drug candidates.
Keywords: Glycosylation, Mannofuranoside, Antibacterial, Antifungal, Physicochemical parameter.
INTRODUCTION
The potential of sugars as starting compounds for
highly efficient syntheses of carbohydrate
conjugates are now well recognized with regard to
antibacterial, antiviral, antineoplastic, antiprotozoal
Microbial food contaminations have been
the cause of severe public health concern over the
1
3-9
last few decades hence antibacterial and antifungal
and antifungal activity . Heterocyclic chemistry has
agents are necessary for food preservation. Many
researchers have focused their research on finding
high potential antibacterial and antifungal agents .
wide array of biologically active compounds isolated
from natural sources or synthesized in the
2
10-12
13,14
laboratory, such as coumarins
, quinolines
,

2
732
KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
1
5
16,17
triazoles and phenolics
etc. They are also if Apparatus
Electronics India, model-934 used for the
significant activity to these molecules. From the determination of Melting points. ES mass spectra
present as a part of giant molecule, imparting
were recorded in Merck M-8000 LCMS system or
Micromass Quadro LCMS system and HR/EI mass
were done on JEOL-600H at 70eV. Proton NMR
previous studies, it is realized that heterocyclic and
aromatic rings are coupled with carbohydrate to
1
8
enhance the biological activity like some
acyl-substituted D-glucofuranose were discovered
1
3
and C-NMR spectra were recorded on Bruker
Avance DPX-300 MHz or Avance DPX-200 MHz
FT Bruker spectrometers, using deuteriated solvents
and Tetramethyl Silane as an internal standard. The
chemical shift values from downfield to upfield in
19
as more biologically active than D-glucofuranose.
furthermore, when biologically active molecule is
connected to another nuclei may possess
2
0
excellentbiological activity.
1
3
both proton NMR and C-NMR spectra. For
synthesized compounds, proton NMR data is
recorded in the order; chemical shifts values in ppm
Several glycoconjugates such as
glycolipids, glycoproteins or glycopeptides &
glycosylated natural products are frequently being
used as antimicrobial drugs and also emerging as
(
δ), multiplicities are indicated as bs (broadened
singlet), s (singlet), d (doublet), dd (double doublet),
t (triplet), q (quartet) and m (multiplet). Coupling
constant ‘J’ reported in Hz. Perkin-Elmer’s Spectrum
RX I FT-IR spectrophotometer was used for recording
IR spectra using KBr disc. Elemental analysis was
carried out on Carlo-Erba-1108 instrument.
2
1-28
anti-cancer drug candidates
. Encouraged by
these results, we have synthesized some glycosides
for antibacterial and antifungal evaluation. During
the synthesis of glycosides, protection of a particular
functional group of monosaccharides is not only
necessary for the modification of the remaining
functional groups but also useful during the synthesis
RESULTS AND DISCUSSION
2
9,30
of novel derivatives of great importance
.
The glycosyl donor was synthesized from
Glycosylation of alcohol-acceptor precursor D-mannose, led to the formation of 2,3,5,6-di-O-
as heterocycles, with O-(2,3,5,6-di-O-α-D- isopropylidene-α-D-mannofuranose (C1). The later
3
6
mannofuranosyl) trichloroacetimidate , as donor was treated with trichloroacetonitrile to gave 2,3,5,6-
precursor in the presence of catalyst trimethylsilyl di-O-isopropylidene-1-O-trichloroacetamidyl-α-D-
36
trifluoromethanesulfonate (TMSOTf) to give mannofuranose (C2) in Scheme-1 .
31
glycosidic products , further antimicrobial activity
of synthesized compounds were evaluated using
various strains of bacteria& fungi to determine
O
O
OH
O
O
O
OC(NH)CCl
3
O
OH
O
OH
3 2 3
CCl CN, K CO
PTSA, Acetone
Reflux, 28 hrs
3
2-35
Inhibitory Concentration (IC )
.
HO
HO
2 2
CH Cl , RT, 24 hrs
50
OH
O
O
O
O
D-mannose
MATERIALS AND METHODS
C1
C2
Scheme 1. Synthesis of mannofuranosyl donor36
All the synthesized compounds were
routinely checked for their purity on silica gel 60
The structure of the C1 was in accordance
(
E. Merck) TLC plates and their spots were
with its spectroscopic data such as proton NMR,
visualized by charring the plates with 5% H SO in isopropylidene group (>C(CH ) ) at δ 1.46-1.33
2
4
3 2
13
MeOH. Column chromatography on Merck silica gel ppm. C NMR spectrum displayed signal of methyl
(
100-200 mesh) was used for the purification of carbons of isopropylidene group (>C(CH ) ) at δ
3
2
2
5.8-27.0 ppm. The mass spectra gave a peak at
synthesized compounds. Combined organic extract
were dehydrated using anhydrous Na SO and dried
-1
2
61 (M+1). FT-IR for –OH at 3431 cm with 87%
2
4
yield. The synthesized glycosyl donor (C2) was
solutions were evaporated under vacuum. Solvents
used in column chromatography were purchased
from E. Merck, sd fine chemicals and Qualigens.
Reagents used in the synthesis were purchased
from, Avara syntheses, Sigma Aldrich and Himedia.
established by proton NMRsignals at δ 8.59 ppm
13
for –NH group, C NMR spectrum displayed signal
of methyl carbons of isopropylidene group
(
>C(CH ) ) δ 27.1-24.9 ppm, at δ 160.8 ppm for
3 2
acetamidyl carbon. Mass spectrum gave a peak at

KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
2733
-1
O
4
6
03 (M+1), FT-IR for –NH group was 3794 cm with
6% yield.
CH3
O
O
O
O
O
NH
C
O
OH
O
O
TMSOTf
O
O
CCl
3
+
Mol sieve 4 A°
O
O
O
CH
2
Cl
2
,
RT,
3
hrs
O
O
CH
3
The aglycon 7-(2-hydroxyethoxy)-4-
O
O
C4
C3
methyl coumarin (C3) was synthesized by the
reaction of 7-hydroxy-4-methyl coumarin with
C2
Scheme 3. Catalysed glycosidation of coumarin
derivatives with glycosyl donor
2
-bromoethanol and potassium carbonate in the
37
presence of ethanol in Scheme-2.
Synthesized glycosyl donor of D-mannose
C2 was reacted with Glycosyl acceptor vanillin
in the presence of trimethylsilyl trifluoro-
CH
3
CH
3
OH
Ethanol, K
2
CO
3
+
HO
Br
9
0°C, 24 hrs
o
methanesulfonate (TMSOTf), 4 A molecular sieve
O
O
OH
O
O
O
2
-bromo ethanol
in anhydrous dichloromethane to formed 3-O-
C3
Scheme 2. Synthesis of 7-(2-hydroxyethoxy)-4-
methyl coumarin
methoxy-4'-O-α-(2,3,5,6-di-O-isopropylidene-
mannofuranosyl) benzoic acid (C5) Chemical
structure of the compound C5 was confirmed by
proton NMR, and FT-IR.Proton NMRspectra gave
signals of isopropylidene proton (>C(CH ) ) singlet
The structure of C3 was established by
1
3
proton NMR, C NMR, Mass and FT-IR. Proton
3
2
at δ 1.56-1.32 ppm, singlet at δ 3.92 ppm (-OCH )
NMRspectra gave a triplet at δ 3.69 ppm, (-CH OH),
3
C
2
13
and also a singlet at δ 11.74 ppm (-COOH).
1
3
and δ 4.33 ppm (-CH OAr). C NMR spectrum
2
NMR spectrum displayed signal of methyl carbons
displayed a signal at δ 61.3 ppm (-CH OH), at δ
2
of isopropylidene group (>C(CH ) ) at δ 26.2-23.8
3
2
6
9.9 ppm (-CH OAr) and at δ 161.9 ppm for carbonyl
2
ppm, at δ 56.2 ppm (-OCH ) and at δ 169.5 ppm for
3
carbon, Mass spectrum gave a peak at 221 (M+1),
carbonyl carbon, Mass spectrum gave a peak at
-
1
-1
FT-IR gave frequency 3452 cm (-OH), 1720cm
>C=O) for compound C3 with 58 % yield.
-1
4
(
11 (M+1), FT-IR gave frequency at 1690 cm
(
-1
>C=O), 3298, 2925 and 2870 cm (-CH and
3
–
CH stretching), while frequency for -OH was
2
Glycosylation of C3 as an alcohol-
absent. The proton NMRsignals of -OH and –NH
was absent in C5. It was formed with 45 % yield.
acceptor, with 2,3,5,6-di-O-isopropylidene-1-O-
trichloroacetamidyl-α-D-mannofuranose, C2 as
donor in the presence of trimethylsilyl
O
OCH
3
O
OCH3
o
trifluoromethanesulfonate (TMSOTf) and 4A
NH
C
O
HO
O
O
O
O
TMSOTf
molecular sieve in anhydrous dichloromethane
gave 7-O-ethoxy-α-(2,3,5,6-di-O-isopropylidene-
mannofuranosyl)-4-methyl-coumarin (C4)
O
CCl
3
+
Mol sieve 4 A°
CHO CH
Cl
,RT,
2
2
CHO
3
hrs
O
O
O
O
Vanillin
C5
(
Scheme-3) as colorless crystalline solid. Chemical
C2
structure of the compound C4 was confirme of
Scheme 4. Synthesis of mannosylated phenolics
isopropylidene proton (>C(CH ) ) singlet at δ 1.49-
3
2
1
.32 ppm, singlet at δ 1.97 ppm (-CH ), triplet at δ
Synthesized glycosyl donor C2 was
coupled with 8-Hydroxy quinoline, 1-Hydroxy
benzotriazole and 2-(Hydroxymethyl) furan in
the presence of catalytic amount of trimethylsilyl
3
3
.69 ppm (-CH CH OAr) and also a triplet at δ 4.33
2
2
13
ppm (-CH CH OAr). C NMR spectrum displayed
2
2
signal of methyl carbons of isopropylidene group
>C(CH ) ) at δ 26.8-26.0 ppm, at δ 18.9 ppm (-CH ),
o
trifluoromethanesulfonate (TMSOTf), 4A molecular
(
3
2
3
sieve in anhydrous dichloromethane at room
temperature to gave 8-O-α-(2,3,5,6-di-O-
isopropylidene-mannofuranosyl)-quinoline (C6),
at δ 61.3 ppm (-CH CH OAr), at δ 69.8 ppm
2
2
(
-CH CH OAr) and at δ 164.1 ppm for carbonyl
2 2
carbon, Mass spectrum gave a peak at 463 (M+1),
FT-IR gave frequency at 1720 cm (>C=O), 3280,
1-O-α-(2,3,5,6-di-O-isopropylidene mannofuranosyl)-
-
1
1
H-benzo-1,2,3-triazole (C7) and2-O-methyl-α-
-
1
2
988cm (-CH3 and -CH2 stretching), while
(2,3,5,6-di-O-isopropylidene-mannofuranosyl)
furan (C8) with 56, 67, 69% yield respectively.
Structure of these compounds was established by
frequency for -OH was absent. The proton
NMRsignals of –OH and –NH was absent in C4,
formed with 59 % yield.
13
proton NMR, C NMR, Mass and FT-IR.

2
734
KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
Proton NMRspectra of C6 gave signals of NCL Pune, India were grown to mid-logarithmic
isopropylidene proton (>C(CH ) ) singlet at δ 1.36- phase and harvested by centrifugation, washed with
3
2
1
7
.25 ppm, doublet at δ 8.73 ppm (H-1), triplet at δ
.52 ppm (H-2), doublet at δ 7.58 ppm (H-3) and
1
0 mM sodium phosphate buffer (SPB) at pH 7.4,
5
and diluted to 2 x 10 colony forming units (CFU)/ ml in
SPB containing 0.03% Nutrient broth (NB).
Mannofuranoside derivatives were serially diluted
in 250 µL of nutrient broth (NB) medium in 96-well
microtitre plates to achieve the desired
concentrations (25-400µg/ml) with bacterial
inoculum (5 x 10 CFU per well).After incubation at
37 C overnight, the IC was taken of the synthesized
compound that provides a 50% reduction in growth
of bacterial strains compared to the growth in the
control well.
1
3
multiplet at δ 7.41-7.16 ppm. C NMR spectrum
displayed signal of methyl carbons of
isopropylidene group (>C(CH ) ) at δ 27.0-24.7 ppm,
3
2
carbon of aromatic ring was observed at δ 163.9,
52.3, 148.0, 138.4, 136.3, 128.7, 127.9, 122.0, 118.1
ppm, Mass spectrum gave a peak at 388 (M+1), FT-
1
4
-
1
IR gave frequency at 2988 and 2881 cm (-CH3
and –CH stretching), while frequency for -OH was
absent.
o
50
2
Proton NM Rspectra of C7 gave signals of
isopropylidene proton (>C(CH ) ) at δ 1.47-1.31 ppm
3
2
Antifungal activity
Determination of inhibitory concentration (IC ):
13
and multiplet at δ 7.69-7.38 ppm (Ar-H). C NMR
spectrum displayed signal of methyl carbons of
isopropylidene group (>C(CH ) ) at δ 26.0-24.7 ppm,
50
The utilization of microplate bioassays, or
broth microdilution tests, to measure the biological
action of synthetic compounds against fungal
pathogens has expanded in recent years. This
method has been recognized as the most
encouraging invitro bioassay for evaluating
3
2
carbons of aromatic ring was observed at δ 144.8,
44.7, 135.1, 130.8, 129.8, 129.6, 128.8, ppm, Mass
spectrum gave a peak at 378 (M+1), FT-IR gave
1
-
1
frequency at 3039 and 2917 cm (-CH and –CH
3
2
stretching), while frequency for -OH was absent.
40,41
antifungal activity .The antifungal strains A. flavus
MTCC 277) and T. viridens (MTCC 167) for the
isopropylidene proton (>C(CH ) ) singlet at δ 1.42- test were obtained from NCIM, NCL Pune, India.
(
Proton NMR spectra of C8 gave signals of
3
2
1
.329 ppm, singlet at δ 3.92 ppm (H-1), doublet at δ Mannofuranoside derivatives were serially diluted
6
.24 ppm (H-2), triplet at δ 6.27ppm (H-3), doublet in 250 µl of SD medium in 96-well microtitre plates
13
at δ 6.98 ppm (H-4). C NMR spectrum displayed
signal of methyl carbons of isopropylidene group
to achieve the desired concentrations (0.5-5 mg/
2
ml) with fungal inoculum (5 x 10 CFU per well).The
(>C(CH ) ) at δ 26.0-24.7 ppm, carbons of furan ring
3 2
invitro antifungal activities of the test compounds
under investigation were also done by Poisons Food
was observed at δ 152.8, 143.8, 113.7, 112.7 ppm,
Mass spectrum gave a peak at 341 (M+1), FT-IR
gave frequency at 3282 and 2974 cm (-CH and
–
absent. The proton NMR signals of –OH and –NH
was absent in C6, C7and C8.
42
technique and the technique was used with some
-
1
43
3
modification by using 0.5-5 mg of compounds per
CH stretching), while frequency for -OH was
2
ml of Sabouraud Dextrose (SD) medium. The
diameter of radial growth of the test fungi was
o
measured after 2 to 3 days of incubation at 27±1 C
and expressed as percent mycelia growth inhibition
following the formula given below:
Antibacterial activity
Determination of inhibitory concentration (IC ):
Thedisk diffusion method was used for
the preliminary antibacterial evaluation of compounds
C1, C2, C4-C8. The inhibitory concentrations (IC )
5
0
3
8
O
O
O
NH
C
O
O
OR
O
CCl
3
+
Different
TMSOTf
O
Heterocyclic
moieties
Mol sieve 4 A°
CH2Cl2 ,RT,
5
0
1
.5 hrs
O
O
of mannofuranoside derivatives showing inhibition
in the preliminary tests, were determined by the
microtitre plate technique using micro dilution
O
O
C2
N
N
N
N
O
39
method (Table-1) Briefly, Staphylococcus aureus
ATCC 6538), Bacillus pumilis (MTCC 160), Bacillus
C8
(
C6
C7
Scheme 5. Catalysed glycosidation of different
heterocycles with glycosyl donor
subtilis (MTCC 441), Bacillus cerius (MTCC1305),
Escherichia coli (ATCC 25923), obtained from NCIM

KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
2735
I = C-T/CX100
important part of drug discovery. These analyses
involve assessment of various molecular
descriptors that would help in prediction of drug-
likeliness. There are various in- silico tools available
to calculate the molecular properties of a compound
to assess its pharmacokinetic profile. In fact, these
in-silico tools have helped a lot to overcome the
Where, I = Percentage of inhibition.
C = Optical density of the fungal colony at 595 nm
in control (DMSO)
T = Optical density of the fungal colony in treatment.
Calculation of Physicochemical Properties
4
5,46
issues related to the failure of clinical trials
.
To check the physicochemical properties
of synthesized mannofuranoside derivatives,
Molinspiration property count instrument(http://
www.molinspiration.com/cgi-bin/properties) was
utilized. Different parametersas topological polar
surface area (TPSA), molecular weight, miLogP, the
number of hydrogen bond donors, the number of
hydrogen bond acceptors, number of rotatable
Molinspiration is a tool which was used in
the present study to screen the pharmacokinetic
property of mannofuranoside derivatives i.e., C1,
C2, C4-C8. According to the Lipinski’s Rule of Five
(
Lipinski et al., 2001) studied mannofuranoside
derivatives posses’ drug like property [Table-2].
Lipinski’s Rule of Five is based on the observation
that the majority of oral drugs have a molecular
weight ≥5000, Log P value ≥5, hydrogen bond
donor ≥5, hydrogen bond acceptor ≥10 and
rotatable bonds ≥10. Any compound which violates
more than one of these rules might have problem.
Interestingly, none of our synthesized manno-
furanoside derivatives gave violation of Lipinski’s
Rule of Five. Thus, we can safely state that these
compounds could be considered as potential drug
candidates.
44
bonds, and violations of Lipinski’s rule of five were
estimated using Molinspiration tool [Table-2].
Moreover, Absorption % was ascertained by the
strategy for Zhao et al. (2002) utilizing the recipe
%
of Absorption = 109 [0.345 × TPSA]
Physicochemical properties prediction
Earlier, lead drug compounds fails at
different clinical trials stages as a result of in
appropriate pharmacokinetic profiling. But, now- a-
days to cut down these failures, physicochemical
profiling of new drug candidates has become an
The growth inhibition of different strains of
bacteria (S. aureus (ATCC 6538), B. pumilis (MTCC
1
60), B. subtilis (MTCC 441), B. cerius
Table. 1: Antibacterial (IC50 µg/ml) and Antifungal activity (*IC50 mg/ml) of compounds
C1, C2, C4-C8
Compounds
S. aureus B. pumilis B. Subtilis
B. cerius
E.coli
A. flavus
T. viridens
C1
C2
NA
267.67
±0.17)
NA
NA
NA
NA
NA
NA
NA
NA
NA
1.84(±0.04) 1.86(±0.02)
1.09(±0.01) 2.30(±0.05)
(
C4
NA
157.61
±0.32)
NA
NA
NA
NA
1.18(±0.05) 1.66(±0.03)
2.21(±0.03) 1.08(±0.01)
0.75(±0.02) 0.59(±0.01)
1.23(±0.05) 2.98(±0.04)
2.91(±0.02) 3.82(±0.03)
(
C5
234.62
221.65
(±0.25)
101.12
(±0.13)
NA
309.43
(±0.19)
154.78
(±0.21)
156.67
±0.34)
NA
(
±0.16)
84.28
±0.11)
NA
C6
102.68
(±0.16)
NA
178.96
(±0.31)
NA
(
C7
(
C8
294.15
NA
NA
NA
(
±0.31)
4.14
±0.7)
-
Ampicillin
Fluconazole
14.15
(±0.06)
-
11.11
(±0.09)
-
8.51
(±0.42)
-
28.03
(±0.10)
-
-
-
(
0.10(±0.01) 0.08(±0.01)
NA, not active, (±) shows SD

2
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KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
Table. 2: Physicochemical Parameters of Compounds C1, C2, C4-C8 and standard drugs
Compounds
Physiochemical parameters
a
b
c
d
e
f
g
i
%
Abs
TPSA
MW
miLogP
HBD
HBA
RB
Lip. vio
Rule
C1
C2
C4
C5
C6
C7
C8
-
-
<500
d”5
0.70
2.85
3.30
2.52
2.97
2.34
2.17
-0.87
-0.12
<5
1
1
0
1
0
0
0
4
<10
6
7
9
9
7
9
7
7
d”10
1
4
6
5
3
3
4
4
d”1
0
0
0
0
0
0
0
0
86.09
81.65
76.27
73.83
85.43
79.28
85.35
70.10
80.82
66.40
79.26
94.85
101.94
68.30
86.12
68.54
112.73
81.66
260.29
404.67
462.50
410.42
387.43
377.40
340.37
349.41
306.88
Ampicillin
Fluconazole
1
7
5
0
a
%
Abs = % of Absorption= 109 [0.345 × Topologicl Polar Surface Area]
b
2
TPSA = Topological Polar Surface Area (Å)
c
MW = Molecular Weight
miLogP = Logarithm of compound partition coefficient between n-octanol and water
HBD = Hydrogen Bond Donors
HBA = Hydrogen Bond Acceptors
RB = Rotatable bond
Lip. vio = Lipinski’s Violation
d
e
f
g
i
1
00
100
80
C2
C5
C6
C8
9
8
7
6
5
4
3
2
1
0
0
0
0
0
0
0
0
0
0
C4
C6
6
4
2
0
0
0
0
B.subtilis (MTCC 441)
S.aureus (ATCC 6538)
0
50
100
150
200
250
300
350
400
450
Concentration (µg/ml)
0
50
100
150
200
250
300
350
400
450
Fig.1. Growth Inhibition of S. aureus at Different
Concentration of Synthesized Mannofuranoside
Derivatives (C2, C5, C6 & C8) was Recorded
After 18 hours
Concentration (µg/ml)
Fig.3. Growth Inhibition of B. subtilis at Different
Concentration of Synthesized Mannofuranoside
Derivatives (C4, & C6) was Recorded After 20 hours
90
90
80
70
60
50
40
30
20
10
0
C5
C6
C7
8
0
C5
C6
70
60
50
40
30
B. cerius (MTCC1305)
B.pumilis (MTCC160)
20
10
0
0
50
100
150
200
250
300
350
400
450
0
50
100
150
200
250
300
350
400
450
Concentration (µg/ml)
Concentration (µg/ml)
Fig.4. Growth Inhibition of B. cereus at Different
Concentration of Synthesized Mannofuranoside
derivatives (C5, C6 & C7) was Recorded
After 19 hours
Fig.2. Growth Inhibition of B. pumilis at Different
Concentration of Synthesized Mannofuranoside
Derivatives (C5 & C6) was Recorded After 20 hours
EXPERIMENTAL
(
MTCC1305), E. coli (ATCC 25923) and fungi A.
flavus (MTCC 277), T.viridens (MTCC 167) obtained Synthesis of 2,3,5,6-di-O-isopropylidene-α-D-
36
mannofuranose (C1)
from NCIM, National chemical laboratories, Pune)
at different concentration were presented in Fig. 1-7.
Yield 87%, Rf 0.74, mp = 124-126 pC;
Proton NMR (300 MHz, CDCl ): δ 4.83-4.79 (m, 1H);
3

KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
2737
1
3
80
70
60
50
40
30
20
10
0
(s, 3H); C NMR (75 MHz, CDCl ) δ: 113.4, 112.7,
3
C6
1
7
2
09.4, 109.2, 101.2, 101.0, 97.2, 85.6, 80.1, 79.7,
9.4, 79.1, 78.6, 76.8, 76.2, 73.4, 73.0, 70.1, 67.1,
7.0, 26.8, 25.9, 25.8; MS(m/z) = 261 (M+1); IR
KBr): (ν cm ): 3431, 2982, 2875; Anal. Calcd for
-1
(
C H O : Calculated C, 55.37; H, 7.74 Found C,
12
20
6
5
5.29; H, 7.71
E.coli (ATCC 25923)
Synthesis of 2, 3, 5, 6-di-O-isopropylidene-1-O-
trichloroacetamidyl-α-D-mannofuranose (C2)
36
Yield 66%, Rf 0.92, mp = 103-105 pC;
0
50
100
150
200
250
300
350
400
450
Proton NMR (300 MHz, CDCl ): δ 8.59 (s, 1H, -NH);
Concentration (µg/ml)
3
δ 6.26 (s, 1H); δ 4.94 (m, 2H); δ 4.42 (m, 1H); 4.04 (m,
3H); δ 4.05-4.17 (dd, J = 2.1 Hz, 1.2 Hz, 2H); δ 1.59,
Fig.5. Growth Inhibition of e.coli at different
concentration of synthesized mannofuranoside
derivative (C6) was recorded After 16 hours
13
δ 1.50 , δ 1.45 & δ 1.33 (s, 3H); C NMR (CDCl , 75
3
C1
C2
MHz) δ: 160.8, 113.6, 112.7, 109.6, 109.2, 104.9,
1
00
C4
C5
C6
C7
C8
101.4, 91.1, 85.6, 84.9, 83.0, 80.5, 79.8, 79.4, 79.1,
7
6.8, 73.4, 72.8, 67.2, 66.8, 27.1, 26.1, 25.3, 24.9;
8
6
4
2
0
0
0
0
0
-1
MS(m/z) = 404 (M+1); IR (KBr): (ν cm ): 3794, 3334,
2988Anal. Calcd for C H O NCl : Calculated C,
14
20
6
3
40.68; H, 4.98; N, 3.46; Found C, 41.08, H, 4.81, N
3.38.
A.flavus (MTCC 277)
Synthesis of 7-(2-hydroxyethoxy)-4-methyl
coumarin (C3)
A mixture 7-hydroxyl-4-methyl coumarin
(5.0 gm, 22.72 mmol), 2-bromoethanol (5.0 gm,
40.65 mmol) and potassium carbonate (3.0 gm,
0
1
2
3
4
5
Concentration mg/ml
Fig. 6. Growth inhibition of A. flavus strain
at different concentration of synthesized
mannofuranoside derivatives (C1, C2, C4-C8)
21.89 mmol) are taken in flat bottom flask (25 ml)
and refluxed for 24 h. in ethanol (100 ml). After
cooling treated with ether and water (1:1) then
separated. The aqueous layer further washed with
ether (2 x 20 ml), both organic layers combined;
add sodium sulphate to dry and solvent evaporate
under reduced pressure obtained residue was
directly allow to column chromatography using
hexane/ethyl acetate (4/6) to give the desired
compound C3 with interesting yield for
crystallization ether was added and 7-(2-
hydroxyethoxy)-4-methyl coumarin obtained in the
form of white crystalline solid.
was recorded after 48 hours
C1
1
10
00
C2
C4
C5
C6
C7
C8
1
9
8
7
6
5
4
3
2
1
0
0
0
0
0
0
0
0
0
0
T.viridens (MTCC 167)
0
1
2
3
4
5
Concentration mg/ml
Yield 58%, mp = 128-130 pC; Proton NMR
Fig.7. Growth inhibition of T. viridens strain
at different concentration of synthesized
mannofuranoside derivatives (C1, C2, C4-C8)
(
300 MHz, CDCl ): δ 7.67 (d, J = 1.5 Hz, 1H); δ 6.83(d,
3
J = 2.3 Hz, 1H); δ 6.93 & δ 6.91 (dd, 1.6 & 1.2 Hz,
H); δ 6.57 (s, 1H); δ 1.97 (s, 3H); δ 2.6 (s, 1H,); δ 3.69
1
were recorded after 52 hours
13
(t, 2H); δ 4.33 (t, 2H); C NMR (75 MHz, CDCl ) δ:
3
δ 4.46-4.42 (m, 1H); δ 4.61-4.63 (d, J = 5.6 Hz, 1H,);
.18-4.21 (m, 1H); δ 4.05-4.17 (dd, J = 2.1 Hz, 1.2
Hz, 2H); δ 3.1 (s, 1H); δ 1.47, δ 1.46, δ 1.38 & δ 1.33
1
1
61.9, 161.6, 155.3, 152.8, 125.8, 114.1, 113.3,
12.7, 112.3, 111.6, 103.5, 101.8, 69.9, 61.3, 19.3;
4
-1
MS (m/z) = 221 (M+1); IR (KBr) (ν cm ): 3452; 3021,

2
738
KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
2
982, 1720 Anal. Calcd for C H O : Calculated C, δ 7.16 (d, J = 1.5 Hz, 1H); δ 5.69 (s, 1H), δ 4.93 (m,
,
11
8
4
64.07; H, 4.89; Found C, 64.01, H, 4.74.
1
H); δ 4.89 (m, 1H); δ 4.41-4.38 (m, 1H); δ 4.63-4.61
(
d, J = 7.0 Hz, 1H); δ 4.13-4 .19 (dd, J = 2.2 Hz, 1.2
Synthesis of 7-O-ethoxy-α-(2,3,5,6-di-O-
isopropylidene-mannofuranosyl)-4-methyl-
coumarin (C4)
Hz, 2H); δ 3.92 (s, 3H); δ 1.56, δ 1.53, δ 1.36 & δ 1.32
13
(s, 3H); C NMR (75 MHz, CDCl ) δ: 169.5, 151.1,
3
1
1
2
45.8, 144.7, 126.7, 124.3, 124.2, 123.8, 116.4,
15.1, 111.4, 103.6, 101.2, 85.5, 71.2, 56.2, 26.2,
To synthesized C4, the mixture of C2 (1.21
gm, 3.0 mmol) and C3 (0.61 gm, 3.0 mmol) was
stirred in dry dichloromethane CH Cl (15 ml) in the
5.2, 25.0, 23.8; MS(m/z) = 411 (M+1); IR (KBr)
-1
2
2
(ν cm ): 3298, 2925, 2870, 1690;Anal. Calcd for
C H O : Calculated C, 58.53; H, 6.39; Found C,
presence of trimethylsilyl trifluoro-methanesulfonate
20
26
9
TMSOTf (60 µl, 0.30 mmol) and molecular sieve
58.62, H, 6.35.
o
(1.0 gm, 4A ) for 1.5 h. at room temperature and
then solvent evaporated under reduced pressure
obtained residue was directly allow to column
chromatography using hexane/ethyl acetate (9/1)
to give the desired compound C4 as white crystalline
solid with interesting yield.
Synthesis of 8-O-α-(2,3,5,6-di-O-isopropylidene-
mannofuranosyl)-quinoline (C6)
To synthesized C6, the mixture of C2 (1.21
gm, 3.0 mmol) and 8-Hydroxy quinoline (0.87 gm,
6
.0 mmol) was stirred in dry dichloromethane
CH Cl (15 ml) in the presence of trimethylsilyl
2
2
Yield 49%, Rf 0.62, mp = 116-118 pC;
trifluoromethanesulfonate TMSOTf (60 µl, 0.30
Proton NMR (300 MHz CDCl ): δ 7.67 (d, J = 1.5 Hz,
o
3
mmol) and molecular sieve (1.0 gm, 4A ) for 1.5 h.
1
1
1
H); δ 6.83 (d, J = 2.3 Hz, 1H); δ 6.91-6.89 (dd, 1.6 &
.5 Hz, 1H); δ 6.57 (s, 1H); δ 5.62 (s, 1H), δ 4.87 (m,
H); δ 4.91 (m, 1H) δ 4.42-4.39 (m, 1H); δ 4.62-4.60
at room temperature and then solvent evaporated
under reduced pressure obtained residue was
directly allow to column chromatography using
hexane/ethyl acetate (9/1) to give the desired
compound C6 as white crystalline solid with
sufficient yield.
(
d, J = 7.5 Hz, 1H); δ 4.16-4 .18 (dd, J = 2.0 Hz, 1.5
Hz, 2H); δ 1.97 (s, 3H); δ 3.69 (t, 2H); δ 4.33 (t, 2H); δ
13
1.49, δ 1.43, δ 1.35 & δ 1.32 (s, 3H); C NMR (75
MHz, CDCl ) δ: 164.1, 161.9, 160.2, 126.1, 125.9,
3
1
6
=
13.3, 112.8, 112.2, 111.5, 103.5, 101.7, 91.9, 69.8,
Yield 44%, Rf 0.65, mp = 110-113 pC;
1.3, 29.8, 27.0, 26.0, 26.3, 24.7, 18.9; MS(m/z)
Proton NMR (300 MHz CDCl ): δ 8.73 (d, J = 7.5 Hz,
3
-
1
463 (M+1); IR (KBr) (ν cm ): 3282, 3021, 2974,
720 ;Anal. Calcd for C H O : Calculated C, 62.33;
1
H); δ 7.52 (t, 1.5 Hz, 1H); δ 7.40 (d, J = 1.2 Hz, 1H);
δ 7.16 (d, J = 1.5 Hz, 1H); δ 7.41 (t, 1.2 Hz, 1H); δ 7.24
d, J = 1.5 Hz, 1H); δ 5.71 (s, 1H), δ 4.98 (m, 1H); δ
1
24
30
9
H, 6.54; Found C, 64.46, H, 6.62.
(
4
7
2
.91 (m, 1H) δ 4.42-4.39 (m, 1H); δ 4.62-4.60 (d, J =
Synthesis of 3-O-methoxy-4'-O-α-(2,3,5,6-di-O-
isopropylidene- mannofuranosyl) benzoic
acid (C5)
To synthesized C5, the mixture of C2 (1.21
gm, 3.0 mmol) and vanillin (0.50 gm, 3.0 mmol) was
.5 Hz, 1H); δ 4.16-4.18 (dd, J = 2.0 Hz & 1.5 Hz,
13
H); δ 1.36, δ 1.34, δ 1.27 & δ 1.25 (s, 3H); C NMR
(
75 MHz, CDCl ) δ: 163.9, 152.3, 148.0, 138.4,
3
1
1
36.3, 128.7, 127.9, 122.0, 118.1, 113.0, 110.2,
09.5, 109.2, 101.7, 101.5, 92.0, 85.6, 85.2, 81.1,
stirred in dry dichloromethane CH Cl (15 ml) in the
presence of trimethylsilyl trifluoro-methanesulfonate
2
2
80.5, 79.8, 73.4, 73.1, 67.1, 66.7, 29.8, 27.0, 26.0,
-1
25.3, 24.7; MS(m/z) = 388 (M+1); IR (KBr) (ν cm ):
TMSOTf (60 µl, 0.30 mmol) and molecular sieve
3
382, 2988, 2881;Anal. Calcd for C H O N:
2
1
25
6
o
(
1.0 gm, 4A ) for 2.5 h. at room temperature and
Calculated C, 65.10; H, 6.50, N, 3.62; Found C,
5.15, H, 6.59, N, 3.71.
then solvent evaporated under reduced pressure
obtained residue was directly allow to column
chromatography using hexane/ethyl acetate (9/1)
to give the desired compound C5 as light yellow
crystalline solid with sufficient yield.
6
Synthesis of 1-O-α-(2,3,5,6-di-O-isopropylidene-
mannofuranosyl)-1H-benzo-1,2,3-triazole (C7)
To synthesized C7, the mixture of C2 (1.21
gm, 3.0 mmol) and 1-Hydroxy benzotriazole (0.86
Yield 45%, Rf 0.42, mp = 97-99 pC; Proton gm, 6.4 mmol) was stirred in dry dichloromethane
NMR (300 MHz CDCl ): δ 11.74 (s, 1H); δ 7.81 (d, J CH Cl (15 ml) in the presence of trimethylsilyl
3
2
2
=
1.5 Hz, 1H); δ 7.41-7.38 (dd, J = 1.5 Hz & 1.2 Hz); trifluoromethanesulfonate TMSOTf (60 µl, 0.30

KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
2739
ο
mmol) and molecular sieve (1.0 gm, 4A ) for 1.5 h.
at room temperature and then solvent evaporated
under reduced pressure obtained residue was
directly allow to column chromatography using
hexane/ethyl acetate (9/1) to give the desired
compound C7 as colourless oil with sufficient yield.
were due to substituent’s which attached to parent
skeleton. The mannofuranoside derivative, 8-O-α-
(2,3,5,6-di-O-isopropylidene-mannofuranosyl)-
quinoline (C6) have IC50 84.28 µg/ml against S.
aureus due to the presence of quinoline as
heterocyclic substituent. Secondary nitrogen of the
quinoline having a nucleophilic centre has been
found essential for activity. The mannofuranoside
derivative, Synthesis of 3-O-methoxy-4'-O-α-
(2,3,5,6-di-O-isopropylidene- mannofuranosyl)
benzoic acid (C5) have maximum IC50 309.43 µg/
ml against B. cerius because they have ethoxy group
as substituent. ethoxy group destabilize the ring
and reduced activity. The mannofuranoside
derivative, 8-O-α-(2,3,5,6-di-O-isopropylidene-
mannofuranosyl)-quinoline (C6) have IC50 0.59 mg/
ml against T. viridens due to the presence of
quinoline as heterocyclic substituent. Secondary
nitrogen of the quinoline having a nucleophilic
centre has been found essential for activity. The
mannofuranoside derivative, 2-O-methyl-α-
Yield 48%, Rf 0.65, mp = 118-120 pC;
Proton NMR(300 MHz CDCl ):δ 7.69 (m, 2H); δ 7.38-
.56 (m, 2 H, Ar-H); δ 5.81 (s, 1H), δ 4.94 (m, 1H); δ
.92 (m, 1H) δ 4.42-4.58 (m, 1H); δ 4.81-4.83 (d, J =
.0 Hz, 1H); δ 4.04-4.12 (dd, J = 2.0 Hz & 1.5 Hz,
H); δ 1.47, δ 1.40, δ 1.35 & δ 1.31 (s, 3H); C NMR
75 MHz, CDCl ) δ: 144.8, 144.7, 135.1, 130.8,
29.8, 129.6, 128.8, 128.1, 114.3, 114.2, 113.8,
13.2, 96.8, 96.7, 79.5, 63.2, 27.0, 26.0, 25.3, 24.8,
3
7
4
7
2
13
(
3
1
1
2
2
5
-1
4.72; MS(m/z) = 378 (M+1); IR (KBr) (ν cm ): 3039,
917;Anal. Calcd for C H O N : Calculated C,
7.29; H, 6.14, N, 11.13; Found C, 57.23, H, 6.04,
1
8
23
6
3
N, 11.28.
Synthesis of 2-O-methyl-α-(2,3,5,6-di-O-
isopropylidene-mannofuranosyl) furan (C8)
To synthesized C8, the mixture of C2 (1.21
gm, 3.0 mmol) and 2-(Hydroxymethyl) furan (0.61
ml, 6.0 mmol) was stirred in dry dichloromethane
CH Cl (15 ml) in the presence of trimethylsilyl
(
2,3,5,6-di-O-isopropylidene-mannofuranosyl)
furan (C8) have IC50 3.82 mg/ml against T. viridens
having methyl furanyl as heterocyclic substituent.
Methyl group destabilize the ring and reduced activity.
2
2
Eight mannofuranoside derivatives were
synthesized and IC50 was determined for all
derivatives. From the IC50 value depicted in table-1
it can be concluded that mannofuranoside with
quinoline as heterocyclic substituent (C6) is more
potent than coumarin analogue (C4), Phenolic (C5),
triazole (C7)and methyl furanyl analogue (C8).
trifluoromethanesulfonate TMSOTf (60 µl, 0.30
o
mmol) and molecular sieve (1.0 gm, 4A ) for 2.5 h.
at room temperature and then solvent evaporated
under reduced pressure obtained residue was
directly allow to column chromatography using
hexane/ethyl acetate (9/1) to give the desired
compound C8 as crystalline solid.
In this study, synthesized manno-
furanoside derivatives, were found to be as strong
antifungal agents. In addition, these compounds
inhibited few pathologically important gram positive
and gram negative bacteria also. As they gave
significant antifungal potential, these compounds
could be further improved to form broad spectrum
antifungal agents. In addition, none of these
mannofuranoside derivatives gave violation of
Lipinski’s Rule of Five. Further studies would be
needed to decipher the exact mechanism of
anti-microbial/anti-fungal action of these
derivatives. However, we can safely state that the
present study has provided some potent anti-
microbial drug candidates in the form of
mannofuranoside derivatives.
Yield 38%, Rf 0.59, mp = 112-114 pC; Proton
NMR (300 MHz CDCl ): δ 3.92 (s, 1H); δ 6.24 (d, d = 7.5
3
Hz, 1H); δ 6.27 (t, 1H); δ 6.98 (d, 1.2 Hz, 1H); δ 5.53 (s,
H), δ 4.83 (m, 1H); δ 4.979 (m, 1H) δ4.38-4.47 (m, 1H);
1
δ4.74-4.79 (d, J = 7.0 Hz, 1H); δ4.03-4.11 (dd, J = 2.0 Hz
13
&
1.5 Hz, 2H);δ1.42, δ1.35, δ1.31 &δ1.29; C NMR (75
MHz, CDCl ) δ: 152.8, 143.8, 113.7, 112.7, 109.2, 107.9,
3
1
01.2, 85.6, 80.1, 79.7, 73.0, 64.1, 26.0, 25.8, 24.9, 24.8;
-1
MS(m/z) = 341 (M+1); IR (KBr) (ν cm ): 3282, 2974;
Anal. Calcd for C H O : Calculated C, 59.99; H, 7.11,
17
24
7
N; Found C, 59.90, H, 7.16.
Structure activity relationship (SAR):
The synthesized compounds selected for
the antimicrobial activity had the same parental
skeleton. The most important things for the activity

2
740
KHAN et al., Orient. J. Chem., Vol. 33(6), 2731-2741 (2017)
ACKNOWLEDGEMENT
MCN00021) for the manuscript and authors are also
thankful to Mr. Wajid Ali, IIT, Guwahati, Ms. Deepti
Saxena, Tahseen Akhtar, Javed, CDRI, Lucknow,
India for their help in spectroscopic data analysis.
The authors are grateful to thank the R&D
wing of Integral University, Lucknow, for support
and provide communication number (IU/R&D/2017-
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