Synthesis and biological evaluation of N-cinnamoyl and mandelate metformin analogues

Article abstract of DOI:10.14233/ajchem.2016.19633

A series of N,N-dimethyl-N1-[3-(substituted phenyl)-1-oxo-2-propenyl]biguanides were synthesized by coupling a solution of metformin in pyridine with different cinnamoyl chloride derivatives in ether for 3 h in addition to synthesis of some five molecules of metformin-mandelates. All the synthesized cinnamoyl metformins and a few metformin-mandelates were characterized by IR, NMR and Mass spectroscopic techniques. All the synthesized compounds were also evaluated for their antioxidant activity by DPPH scavenging method and nitric oxide scavenging method. All the compounds exhibited good antioxidant activity.

Full text of DOI:10.14233/ajchem.2016.19633

Asian Journal of Chemistry; Vol. 28, No. 9 (2016), 1895-1898
A
SIAN
J
OURNAL OF HEMISTRY
C
http://dx.doi.org/10.14233/ajchem.2016.19633
Synthesis and Biological Evaluation of N-Cinnamoyl and Mandelate Metformin Analogues
1
2
4
V. ANITHA KUMARI , K. BHARATHI , K. PRABHU3,* and K. PONNUDURAI
1Department of Pharmaceutical Chemistry, Nova College of Pharmacy, Jangareddygudem-534 447, India
2Department of Pharmaceutical Chemistry, Sri Padmavathi Mahila Viswa Vidyalayam, Tirupathi-517 502, India
3Department of Pharmacognosy and Phytochemistry, School of Pharmacy, Vivekananda Group of Institutions, Batasingaram-501 511, India
4Department of Pharmacology and Toxicology, School of Pharmacy, Vivekananda Group of Institutions, Batasingaram-501 511, India
*Corresponding author: E-mail: prabhu.cognosy@gmail.com
Received: 22 November 2015;
Accepted: 15 February 2016;
Published online: 1 June 2016;
AJC-17911
A series of N,N-dimethyl-N1-[3-(substituted phenyl)-1-oxo-2-propenyl]biguanides were synthesized by coupling a solution of metformin
in pyridine with different cinnamoyl chloride derivatives in ether for 3 h in addition to synthesis of some five molecules of metformin-
mandelates. All the synthesized cinnamoyl metformins and a few metformin-mandelates were characterized by IR, NMR and Mass
spectroscopic techniques. All the synthesized compounds were also evaluated for their antioxidant activity by DPPH scavenging method
and nitric oxide scavenging method. All the compounds exhibited good antioxidant activity.
Keywords: Antioxidants; Metformin-mandelates, Cinnamoyl metformins, Nitric oxide, DPPH.
INTRODUCTION
EXPERIMENTAL
A continuous demand for a search for antioxidant molecules
either by semi-synthetic or by synthetic means is inevitable.
Double bond of cinnamic acid derivatives probably participates
in stabilizing the radical by resonance. Increasing the number of
hydroxy and methoxy substituents increases the potency of
antioxidant side-chain and conjugated double bond in the side
chain could have a stabilizing effect by resonance on the phenoxy
radical. Hydroxy cinnamic acids are the most widely represented
phenolic acids in food vegetables, strengthening their potential
role as nutritional antioxidants [1]. It was discovered that
cinnamic acid, a naturally occurring aromatic fatty acid of low
toxicity, has a long history of human exposure [2]. Cinnamic
acid exhibited several pharmacological activities, including
antimicrobial and anti-inflammatory activity as well as platelet
aggregation and its quartenary ammonium salts were reported
to exhibit plant growth retardant activity [3]. Several hydroxylated
cinnamic acid derivatives were found to have inhibitory effect
on mutagenesis, in a dose dependent manner [2,4].
Melting points were determined in open capillaries on a
melting point apparatus (Tempo) & were uncorrected. FTIR
spectra were recorded using Thermo Nicolet 670 spectrometer.
1H NMR spectra of the compounds in deuteriated dimethyl
sulphoxide were recorded on GEMINI-300 MHz &AVANCE-
200 MHz. All chemicals used are of AR grade. Purity of the
compounds was checked by thin layer chromatography, using
aluminium plates, precoated with silicagel GF 254. Ethanol
was used as the eluent. The spots were visualized in the ultraviolet
light chamber.
General method for synthesis of cinnamic acids: A
mixture of substituted aldehydes (5 g, 0.33 mol), malonic acid
(7.5 g, 0.72 mol) dissolved in pyridine (15 mL) and piperidine
(0.25 mL) was heated under reflux for 2 h on a water bath.
The reaction mixture was allowed to cool and poured into the
excess of water, containing enough HCl. The solid mass, which
separated out was filtered washed with little water, dried
and recrystallized from glacial acetic acid and alcohol. By
following the same procedure, different analogues of cinnamic
acid were prepared. In present work, we have synthesized 12
substituted cinnamic acids (Fig. 1).
In present study, certain cinnamic acid and mandelic acids
were prepared and coupled with metformin, to synthesize novel
cinnamic acid and mandelic acid derivatives, could result in
different pharmacological effects like anti-inflammatory,
cardioprotective, neuroprotective, anticonvulsant and anti-
hypertensive properties which are having oxidation properties
as the causative factor.
General method for synthesis of N-[3-(substituted
phenyl)-2-propenyl-1-amido]metformin: A mixture of substi-
tuted cinnamic acid (0.137 g, 1.0 mmol) and ether (10 mL)
and thionyl chloride (10 mL) were taken in an iodine flask

1896 Kumari et al.
Asian J. Chem.
R
following the above procedure better yields were obtained by
using thionyl chloride (Fig. 2).
COOH
COOH
+
H2C
CHO
OH
Substituted benzaldehydes
Malonic acid
R
C O
Pyridine
OH
reflux, 2 h
Piperidine
R
with stirring
Thionyl chloride, ether reflux,
60 °C, 2 h
CH CH.COOH
OH
CO Cl
Cinnamic acids
Substituted
with stirring
R
Thionyl chloride, ether reflux,
60 °C, 2 h
NH NH
H2NCNHCN(CH3)2.HCl in pyridine, ether
R
Reflux, 3 h
CH CH.CO.Cl
Cinnamoylchloride
NH NH
OH
NH
NH
R
H
Reflux, 3 h
CO NH C N C N(CH3)2
H2NC NHCN(CH3)2.HCl in pyridine, ether
NH
NH
Fig. 2. Scheme of synthesis
CH CH.CO NH C NH CN (CH3)2
R
Antioxidant screening of cinnamoyl and mandelate-
metformins
Fig. 1. Synthesis of substituted cinnamic acids
and heated under reflux at 60 °C for 2 h, with stirring. Then
the reaction mixture was allowed to be concentrated under
reduced pressure. To the distillate, add ether (20 mL) and a
solution of metformin (330 mg, 2 mmol) in pyridine (5 mL).
The reaction mixture in the flask was heated under reflux for
3 h.Again the reaction mixture was concentrated under reduced
pressure. To the distillate, add 20 mL of ethanol and 100 mL
distilled water. The byproduct was precipitated out and filtered.
The product obtained was recrystallized by a mixture of
ethanol:water (1:5) and later was confirmed by TLC. Better
yields were obtained by using thionyl chloride. In present work,
about 12 cinnamic acids were condensed analogues with
metformin synthesized by following the above procedure.
General method for the synthesis of mandelic acid
derivatives: A mixture of substituted mandelic acid (0.175 g,
1 mmol) and ether (10 mL) and thionyl chloride (10 mL) were
taken in an iodine flask and heated under reflux at 60 °C for 2
h, with constant stirring. Then the reaction mixture was allowed
to be concentrated under reduced pressure. To the distillate,
add ether (20 mL) and a solution of metformin (330 mg,
2 mmol) in pyridine (5 mL). The reaction mixture in the flask
was heated under reflux for 3 h. Again the reaction mixture
was concentrated under reduced pressure. To the distillate, add
20 mL of ethanol and 100 mL distilled water. The byproduct
was precipitated out and filtered. The product obtained was
recrystallized from a mixture of ethanol:water (1:5), later was
confirmed by TLC. In present work, about 5 mandelic acid
condensed derivatives were synthesized with metformin, by
Diphenyl, 2-picryl hydrazyl (DPPH) radical scavenging
activity:0.1 mM solution of DPPH was prepared in ethanol.
To this solution, 3 mL of the test solution was added at different
concentration (25-800 µg/mL). Equal amount of distilled water
was added to the control. The mixture was shaken well
and incubated at room temperature (23 2 °C) for 0.5 h. The
absorbance was read at 517 nm using a spectrophotometer.
Nitric oxide (NO) scavenging: Nitric oxide scavenging
activity was measured by using a spectrophotometer. Sodium
nitroprusside (5 mM) in phosphate buffered saline (0.25 g of
sodium dihydrogen phosphate, 0.253 g of disodium hydrogen
phosphate and 0.82 g of sodium chloride were dissolved in
sufficient quantity of water to produce 100 mL) was mixed
with different concentrations of the ethanolic extracts (25-800
µg/mL) dissolved in distilled water and incubated at 25 °C for
0.5 h. A control without test compound but with equivalent
amount of distilled water was taken. After 0.5 h, 1.5 mL of the
incubation solution was removed and diluted with 1.5 mL of
Griess reagent (1 % w/v sulphanilamide, 2 % v/v phosphoric
acid and 0.1 % w/v naphthyl ethylene diamine dihydro-
chloride).
The absorbance of the chromophore formed during
diazotization of the nitrite with sulphanilamide and subsequent
coupling with naphthylethylene diamine was measured at 546 nm.
Determination of lipid peroxidation: Malondialdehyde
were measured as an index of lipid peroxidation. Peroxidation
was induced with Fe2+ (0.02 mM), ascorbate (1 mM) and H2O2
(0.5 mM) in a final volume of 0.5 mL. Test compounds were

Vol. 28, No. 9 (2016)
Synthesis and Biological Evaluation of N-Cinnamoyl and Mandelate Metformin Analogues 1897
added into the reaction mixtures at indicated concentrations just
before the addition of lipid substrates. Malondialdehyde was
measured by its reaction with thiobarbituric acid at 532 nm [5].
PDMAB (10a) 45.68 % derivatives showed significant activity.
Substituted with α-naphthol (17a) 43.99 %, 3,4-di(OCH3)
(6a) 41.0 %, unsubstituted 41.0 %, 2-chloro (13a) 39.44 %,
o-cresol (14a) 39.82 %, p-CH3 38.14 %, p-isopropyl (5a) 35.28 %,
showed moderate activity and substitution with 2-nitro (16),
p-diethylamino- (12a), α-phenol (15a) and vanillinyl (4a)
derivatives showed less activity (Table-1).
Reduction of DPPH by test compounds: Among the
seventeen compounds all the compounds were tested for the
reduction of DPPH. Among the 17 compounds, the unsubsti-
tuted derivative (1a) showed 38.1 % activity. 2-Chloro derivative
showed highest activity 54.02 %.
Substitution with p-chloro (7a) 46.33 %, α-naphthol (17a)
45.19 %, p-fluoro (9a) 45.0 %, 3,4,5-tri(OCH3) (8a) 43.81 %
and para-isopropyl (5a) 42.73 % showed significant activity.
Substitution with o-cresol (14a) 40.0 %, p-dimethylamino- (10a)
38.6 %, unsubstituted (1a) 38.1 %, vanillinyl (4a) 35.06 % and
2-nitro (16a) 33.25 % derivative showed moderate activity.
Substitution with p-CH3 (2a), p-OCH3 (3a), 3,4-di(OCH3) (6a),
α-phenol (15a) derivatives showed less activity (Table-1).
Scavenging of NO free radical: Out of 17 compounds,
all compounds were tested for their ability to scavenge NO at
100 µm concentrations. Unsubstituted derivative (1a) showed
an activity of 64.33 %. Test compounds containing different
substituents on the phenyl ring were also evaluated for NO
scavenging activity.
RESULTS AND DISCUSSION
An attempt was made to synthesize certain novel cinnamic
acid and mandelic acid derivatives. Substituted cinnamic acids
were prepared by reacting a mixture of malonic acid, pyridine
and piperidine with the corresponding substituted aldehydes,
heated under reflux for 2 h. Metformin hydrochloride was
allowed to condense with the above prepared substituted cinnamic
acids (1-12), in the presence of thionyl chloride, ether and pyridine
to obtain the desired new series of compounds 1a-12a.
Twelve compounds were synthesized, with the yields
generally 50-90 %. p-Methyl, p-methoxy, vanillinyl and p-
isopropyl derivatives were obtained at highest yields (80-90 %)
and remained analogs were produced at lowest yields (50-70 %).
Substituted mandelic acids 13-17 (Table-1) were prepared
by reacting a mixture of KOH solution with the corresponding
substituted benzene and glyoxalic acid monohydrate under
cooling (0-5 °C). Metformin hydrochloride was allowed to
condense with the above prepared substituted mandelic acids,
in the presence of thionyl chloride, ether and pyridine to obtain
the desired new series of compounds 13a-17a. Five compounds
were prepared with the yields generally 60-90 %. α-Nitro, α-
phenol and 2-chloro derivatives were obtained at high yields
(80-90 %) and remained analogs were produced at 60-70 %.
Substitution with Cl– derivative at para position (7a)
resulted in increased activity (76.12 %) which is the highest
activity. Substitution with p-OCH3 (3a) 71.52 %, 3,4-di(OCH3)
(6a) 72.06 %, p-F (9a) 66.46 %, p-dimethylamino- (10a) 66.01
%, 2-chloro (13a) 65.90 %, o-cresol (14a) 68.48 % and α-
naphthol (17a) 65.70 % derivatives showed significant activity.
Substitution with p-CH3 62.30 %, vanillinyl (4a) 56.12 %, p-
isopropyl (5a) 58.43 %, α-phenol (15a) 59.27 %, 2-nitro (16a)
63.26 %, p-OH (11a) 56.52 %, showed moderate activity and
substitution with p-diethylamino- (12a) showed less activity
(Table-1).
Antioxidant studies
Effect of compounds 1a-17a on ferrous induced lipid
peroxidation in rat brain homogenate: Out of seventeen
compounds, all the compounds were tested for the inhibitory
effect on iron induced lipid peroxidation. Among these deri-
vatives, the unsubstituted (1a) showed 41.0 % activity. Highest
activity, showed with p-fluoro derivative (9a) (55.10 %). Substi-
tution with p-chloro (7a) 54.0 %, p-hydroxy (11a) 53.08 %,
3,4,5-trimethoxy (8a) 50.49 %, p-methoxy (3a) 49.02 % and
TABLE-1
ANTIOXIDANT ACTIVITY DATA OF N-CINNAMOYL AND MANDELATE METFORMIN ANALOGUES
Inhibition (%) at 100 µm
Reaction
time (h)
Yield
(%)
Compd.
R
m.f.
Lipid peroxidation
41.00
DPPH radical
38.1
NO scavenging
64.33
62.3
H
4-CH3
C13H17N5O
C14H19N5O
C14H19N5O2
C14H19N5O3
C16H21N5O
C15H21N5O3
C13H19N5OCl
5
5
5
5
5
6
5
5
5
5
6
6
5
6
5
5
6
72.00
80.00
81.90
92.70
82.80
63.90
58.60
55.80
66.60
60.30
56.70
63.09
87.50
62.50
76.33
89.20
92.40
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
38.14
49.02
33.72
35.28
41.00
54.00
50.49
55.10
45.68
53.08
31.06
39.44
39.82
32.49
31.78
43.99
32.0
32.7
4-OCH3
71.52
56.12
58.43
72.06
76.12
64.90
66.46
66.01
56.52
37.92
65.90
68.48
59.27
63.26
65.70
Vanillinyl
Isopropyl
3,4-di-(OCH3)
4-Cl
35.06
42.73
30.03
46.33
43.81
45.0
3,4,5-tri(OCH3) C16H23N5O4
4-F
C13H16N5OF
C15H22N6O
C13H17N5O2
C17H26N6O
C12H16N5O2
C17H21N5O3
C16H19N5O3
C12H15N6O4
C16H17N5O3
4-N(CH3)2
4-OH
4-N(C2H5)2
2-Cl
o-Cresol
α-Phenol
2-Nitro
α-Naphthol
38.6
56.52
37.92
54.02
40.0
30.63
33.25
45.19

1898 Kumari et al.
Asian J. Chem.
3. M.L. Sharma and S. Kaur, J. Indian Chem. Soc., 84, 612 (2007).
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