International Journal of Biopharmaceutics DESIGN, SYNTHESIS

50
Saritha Garrepalli. et al. / International Journal of Biopharmaceutics. 2012; 3(1): 50-54.
e- ISSN 0976 - 1047
Print ISSN 2229 - 7499
International Journal of Biopharmaceutics
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IJB
DESIGN, SYNTHESIS AND BIOLOGICAL EVALUATION OF
BENZOXAZOLE DERIVATIVES AS NEW ANTI-INFLAMMATORY
AGENTS
Saritha Garrepalli1,*, Manne Pavan Kumar1, Ambati Praneeth Sai1,
Bommalla Sharanya2, Ganneboina Jyothir Mai2, Chikoti Abhinavgandhi3
1
St.Peter's College of Pharmacy, Madikonda, Kazipet, Warangal, A.P, India - 506142.
St. Peter's Institute of Pharmaceutical Sciences, Vidyanagar, Hanamkonda, Warangal, A.P, India - 506001.
3
Mallige College of Pharmacy, Silverpura, Banglore, India - 560090.
2
ABSTRACT
New series of 2-methyl-N-(substituted arylidene) benzoxazole -5-carbohydrazide derivatives were synthesized by the
reaction of Schiff bases of 2-methyl benzoxazole-5-carbohydrazide with appropriate aromatic aldehydes. The chemical
structures of the synthesized compounds were confirmed by means of IR, 1HNMR, Mass spectral analysis. Further, the
synthesized compounds (VIa-VIf) were screened for anti-inflammatory activity by using Carrageenan – induced paw edema
rat model. The results showed that, compound VI c was significantly (p<0.0001) reduced the inflammation there by showed
a promising anti-inflammatory activity; whereas the compounds i.e., VI a, VI d, VI e, VI f moderately reduced the
inflammation. Only one compound VI b showed very poor anti-inflammatory activity after one hour of administration.
Key words: Benzoxazole derivatives, IR, ¹H NMR, Mass spectroscopy and Anti-inflammatory activity.
INTRODUCTION
Recent
observations
suggest
that
substituted
benzoxazoles and related heterocycles, possess potential
activity with lower toxicities in the chemotherapeutic
approach in man (Haugwitz RD et al., 1982, Hisano T et
al., 1982). Careful literature survey revealed that targets
containing benzoxazole moiety, either isolated from
plants or accessed by total synthesis, have remarkable
biological activities (Anita hari et al., 2001). For
example, antimicrobial (Sultan Nacak et al., 1997),
antihistaminic (Kastura, Y et al., 1992), antiparasitics
(Qian et al., 2001), herbicidal (Peter Paul Wilhelm et al.,
1989), antiviral (Surendra Bahadhur et al., 1981),
anti allergic and anthelmintic activities (Gopal Krishna B
et al., 2005), antifungal (Ismail Y et al., 2000),
Cyclooxygenase Inhibiting (Shrinivasa R et al., 2003),
antitumor (Aiello S et al., 2008), antiulcer (Kastura Y et
al., 1992), anticonvulsant (Siddiqui N et al., 2008),
hypoglycemic (Arakova K et al., 1997), antiinflammatory (Sondhi S et al., 2006, Unlu S et al., 2003)
Corresponding Author
Saritha Garrepalli
E-mail: [email protected]
2-Methyl-N-(substituted arylidene) 1,3-benzoxazole-5carbohydrazide
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Saritha Garrepalli. et al. / International Journal of Biopharmaceutics. 2012; 3(1): 50-54.
and anti-tubercular activity (Klimensova V et al., 2002)
Anti-inflammatory activity of benzoxazole derivatives
were also reported in the literature. The title compounds
were synthesized by treating the 2-methyl benzoxazole-5carbohydrazide with appropriate aromatic aldehydes to
get a new series of 2-methyl-N-(substituted arylidene)
benzoxazole-5-carbohydrazide (VI a-VI f).
MATERIALS AND METHODS
All the reagents and solvents used were of laboratory
grade. The melting points of synthesized compounds
were determined by open capillary method and were
uncorrected. The purity and homogeneity of compounds
were checked using TLC technique. IR spectra
(Rajmohan K et al., 1973) of compounds were recorded
using
KBr
pellets
on
Perkin
Elmer
337
spectrophotometer. 1H-NMR spectra (Finhorn et al.,
1900) were recorded on Bruker Avance-300 MHz
Spectrophotometer using dimethyl sulfoxamide as
solvent at Aptuit laurus, hydrabad. Mass Spectra of the
synthesized compounds were recorded on Liquid
Chromatography Mass Spectrometer at Aptuit laurus,
hydrabad.
SYNTHESIS AND CHARACTERIZATION OF
COMPOUNDS
1) 4-Hydroxy-3-nitro-benzoic acid methyl ester (II)
In a 1 lit. Three necked round bottom flask equipped with
water condenser, mechanical stirrer and thermometer, of
p-hydroxy methyl benzoate (10 g, 0.74 mol) was placed.
A mixture of
-
and then run in the
nitrating mixture in p-hydroxy methyl benzoate with
stirring, while maintaining the temperature of the reaction
between 5 to 15°; the addition continued up to 1 h.
Poured the reaction mixture in to of crushed ice (70 g).
Filtered
off
the crude m-nitro, p-hydroxy methyl
benzoate at the pump and wash with cold water. Transfer
the solids into 500 ml flask and stirred it with ice cold
methanol in order to remove a small amount of ortho
isomer and other impurities. The mixture was filtered
with suction and washed with little methanol and dried in
the air. Then the product was recrystallised using
methanol as solvent (Rajmohan K et al., 1973).
Percentage Yield- 84%, M.P. 65-67°, Rf-0.76 (Ethyl
acetate: methanol, 1:1).
2) 3-Amino-4-hydroxy-benzoic acid methyl ester (III)
In a 500 ml three necked flat bottom flask equipped with
reflux condenser with guard tube, compound II (10 g)
was dissolved in boiling alcohol (50%, 100 ml) and
sodium dithionate was added to this boiling alcohol
solution until it becomes almost colorless. Then the
alcohol was reduced to one third of its volume by
distillation and the residual liquid was triturated with ice
cold water. The resulting colorless, shiny product was
filtered, washed with cold water, dried and recrystalise
using methanol as solvent (Winter CA et al., 1962).
Percentage Yield- 70%, M.P. 110-112°, Rf-0.67
(Saturated methanol).
3) 2-methyl benzoxazole-5-carboxylic acid methyl
ester (IV)
Compound III (0.01mol) was heated with acetic acid
(0.1mol) in excess under reflux for 2h. The reaction
mixture was cooled and poured in crushed ice (100 gm)
with stirring. The product thus separated was filtered
under suction and washed with cold water. The products
were recrystallised by using methanol as a solvent.
Percentage Yield- 60%, M.P. 70-72°C, Rf-0.7
(Saturated methanol).
4) 2-methyl benzoxazole-5-carboxylic acid hydrazide
(V)
A mixture of an appropriate 2-subtituted benzoxazole-5carboxylic acid methyl ester IV (0.001 mol) in alcohol
(25 ml) and hydrazine hydrate (99%, 0.015 mol) was
heated under reflux on water bath for 4 hours. The
alcohol was reduced to half of its volume and cooled.
The product separated was filtered and washed with
small portions of cold alcohol and then with cold water,
repeatedly and dried. The resultant product was
recrystallised using methanol as solvent.
Percentage Yield- 80%, M.P. 144-146°C, Rf-0.7
(Methanol).
5) Synthesis of 2-methyl-N-(substituted arylidene)
benzoxazole-5-carbohydrazide(VI)
The 2- methyl benzoxazol-5-carboxylic acid hydrazides
(V, 0.01mol) and appropriate aromatic aldehydes
(0.015mol) in alcohol(20ml) with 2 to 3 drops of acetic
acid, heated under reflux on a water bath for one hour.
The solvent was removed to possible extent by
distillation under reduced pressure. The product thus
obtained was filtered, washed with water dried and
purified by recrystallization from suitable solvent to
produce the compounds GH1-GH8. The physical data of
these benzoxazole derivatives were given in table1.
RESULTS AND DISCUSSION
Compound VI a
IR (KBr, cm-1): 3400(OH), 3200 (NH), 1692 (C=O),
1592 (C=N), 1259 (C-O-C), 700 ( C-H). ¹H NMR
(DMSO-d6) d: 11.5 (S, 1H, OH), 9.9(S,1H,NH), 7-8 (d,
8H, Ar-H), 2.35 (S, 3H, CH3); MS (m/z): M+ calculated
296, found 295.
Compound VI b
IR (KBr, cm-1): 3238(NH), 1693 (C=O), 1610 (C=N),
1249 (C-O-C), 690 (C-H).
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Saritha Garrepalli. et al. / International Journal of Biopharmaceutics. 2012; 3(1): 50-54.
¹H NMR (DMSO-d6) d: 9.7 (s, 1H, NH), 7-8 (d, 8H, ArH,CH), 3.0 (s, 6H, CH3),2.35(s, 3H, CH3); MS (m/z): M+
calculated 323, found 322
Compound VI c
IR (KBr, cm-1): 3207(NH), 1669 (C=O), 1607 (C=N),
1298 (C-O-C);
¹H NMR (DMSO-d6) d: 9.9 (s, 1H, NH), 7.0-8.0 (d, 8H,
Ar-H, CH), 6.9 (d, 2H, Ar-H),3.7 (s,3H,CH3), 2.3 (s, 3H
, CH3) MS (m/z): M+ calculated 310.0, found 309.08.
Compound VI d
IR (KBr, cm-1): 3200(NH), 1709 (C=O), 1630 (C=N),
1293 (C-O-C);
¹H NMR (DMSO-d6) d: 9.7 (s, 1H, NH), 7.0-8.0 (d, 8H,
Ar-H,CH), 2.4(s, 3H, CH3)
MS (m/z): M+ calculated 314.0, found 313.10
Compound VI e
IR (KBr, cm-1): 3282(NH), 1706 (C=O), 1685 (C=O),
1352 (C-O-C); ¹H NMR (DMSO-d6) d: 9.7 (s, 1H, NH),
7.2-8.0 (d, 8H, Ar-H,CH), 2.4(s, 3H, CH3) MS (m/z): M+
calculated 297.0, found 298.
Compound VI f
IR (KBr, cm-1): 3200 (NH), 1684.26 (C=O), 1615
(C=N), 1296 (C-O-C);
¹H NMR (DMSO-d6) d: 9.7 (s, 1H, NH), 7.-8 (d, 8H, ArH,CH), 2.35 (s, 3H, CH3 )
MS (m/z): M+ calculated 325, found 324
SCHEME OF SYNTHESIS
TABLE 1. PHYSICAL DATA OF 2-METHYL-N-[SUBSTITUTED ARYLIDENE]-1,3-BENZOXAZOLE –5CARBOHYDRAZIDE (VI A-VI F )
Melting point
S.No.
Compound
Ar
Mol. Formula
(%) Yield
(oC)
1
VI a
4-Hydroxy phenyl
C16H13N303
230
90
2
VI b
4-Dimethyl amino phenyl
C18H18N402
205
86
3
VI c
4-Methoxy phenyl
C17H15N303
220
85
4
VI d
4-Chlorophenyl
C16H12N302Cl
224
80
5
VI e
4-Flourophenyl
C16H12N302F
213
75
6
VI f
2-Nitrophenyl
C16H12N404
242
81
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Saritha Garrepalli. et al. / International Journal of Biopharmaceutics. 2012; 3(1): 50-54.
TABLE 2. ANTI INFLAMMATORY
BENZOXAZOLE-5-CARBOHYDRAZIDE
S.NO.
Compound
Ar
1
2
3
4
5
6
7
8
VIa
VIb
VIc
VId
VIe
VIf
Standard
Control
4-OH C6H4
4-N(CH3)2C6H4
4-OCH3C6H4
4-ClC6H4
4-FC6H4
2-NO2C6H4
Diclofenac sod.
Sod.CMC
*P < 0.05,
*P < 0.001,
ACTIVITY
OF
2-METHYL-N-[SUBSTITUTED
Paw volume in mL (Mean ± S.D.)
2hr
3hr
0.32 ± 0.12
0.35 ± 0.10
0.40 ± 0.08*
0.42 ± 0.05
0.36 ± 0.05
0.46 ± 0.05
0.02 ± 0.00
0.32 ± 0.12
0.40 ± 0.14
0.42 ± 0.09
0.33 ± 0.05
0.46 ± 0.15
0.225 ± 0.05**
0.25 ± 0.05**
0.375 ± 0.05**
0.375 ± 0.05**
1hr
0.22 ± 0.15*
0.37 ± 0.04
0.10 ± 0.00
0.17 ± 0.05**
0.25 ± 0.1
0.20 ± 0.00
0.15 ± 0.05**
0.38 ± 0.05
ARYLIDENE]
4hr
0.25 ± 0.10*
0.05 ± 0.05
0.33 ± 0.05
0.35 ± 0.05
0.40 ± 0.08*
0.30 ± 0.10
0.25 ± 0.05***
0.375 ± 0.05**
*P < 0.0001
TABLE 3. ANTI INFLAMMATORY
BENZOXAZOLE-5-CARBOHYDRAZIDE
S.NO.
Compound
Ar
1
2
3
4
5
6
7
VIa
VIb
VIc
VId
VIe
VIf
Standard
4-OH C6H4
4-N(CH3)2C6H4
4-OCH3C6H4
4-ClC6H4
4-FC6H4
2-NO2C6H4
Diclofenac sod
ACTIVITY
1hr
40.37
0.37
73.70
53.70
33.70
47.09
60.37
ANTI - INFLAMMATORY ACTIVITY
Carrageenan-induced rat paw edema method (Aiello s et
al., 2008) was employed for evaluating the anti
inflammatory activity of the synthesized compounds
(VIa-VIf). Wister Albino rats of either sex weighing
approx 200-350 gm were housed in clean polypropylene
cages and kept under room temperature (25±2OC), and
relative humidity 40-50% in a 12 h light-dark cycle. Food
was withdrawn 12 h before and during experimental
hours. In this study, the animals were divided into groups
as shown in the Table-2. Acute inflammation was
produced by sub plantar injection of 0.1ml of 1%
suspension of Carrageenan with 2% gum acacia in
normal saline, in the right hind paw of the rats. After oral
administration of the test compounds, the paw volume
was measured Plethysmometrically at 1, 2, 3, and 4 h
intervals. Diclofenac sodium 10mg/ml of 2% gum acacia
in normal saline was used as standard drug.
The target compounds were synthesized
according to the Scheme-1. In the current research work,
the title compounds N`[substituted sulfonyl]-1,3benzoxazole-5-carbohydrazide, were synthesized by
electrophilic aromatic substitution on p-hydroxy methyl
benzoate (I) with concentrated nitric acid and
concentrated sulfuric acid under reflux condition.
Compound (II) on reduction with sodium dithionate with
alcohol gives 3-amino-4-hydroxy-benzoic acid methyl
OF
2-METHYL-N-[SUBSTITUTED
% Inhibition of paw oedema
2hr
3hr
35.50
30.50
20.50
15.50
27.16
7.16
60.50
35.50
20.50
15.50
33.03
7.16
55.50
50.50
ARYLIDENE]
4hr
47.84
16.26
30.29
26.70
16.26
37.30
47.84
ester (III). Reaction of compound (III) with two
appropriate aliphatic acids (formic acid and acetic acid)
produced corresponding 2-subtituted benzoxazole-5carboxylic acid methyl esters. The reaction of compounds
(IV) with hydrazine hydrate in ethanol on refluxing
yielded the corresponding 2- substituted benzoxazole-5carboxylic acid hydrazides. On further reaction of
compounds (V) with the different aromatic aldehydes
derivatives afforded the corresponding six 2-methyl-N[substituted
arylidene]-1,3-benzoxazole-5carbohydrazide. The identification and characterization
of the synthesized compounds were carried out by
melting point,Thin Layer Chromatography, FT-IR, NMR
and Mass data to ascertain that all synthesized
compounds were of different chemical nature than the
respective parent compound. The yields, melting points
and physical data of newly synthesized compounds are
summarized in Table-1. All the synthesized new
benzoxazole derivatives were evaluated for their anti
inflammatory activity by using the standard as diclofenac
sodium for the period of four hours with one hour
interval.
4-methoxy phenyl group was found to be the most
potent one with an inhibition of paw volume of 73.70
percent after one hour of administration. Compounds
VId, VIf, VIa and VIe were next in the order of inhibition
of paw volume after an hour. Compounds VIb is devoid
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Saritha Garrepalli. et al. / International Journal of Biopharmaceutics. 2012; 3(1): 50-54.
of any anti inflammatory activity. However, the activity
of the test compounds is comparable with the activity of
the standard employed. The data of percent inhibition of
rat paw volume after four hours indicates that the
compound VI a with 4-hydroxy phenyl gp showed more
inhibition (47.84%) followed by the compounds VIf and
VIc with the percent inhibition of 37.31% where rest of
the compounds are found to show moderate anti
inflammatory activity.
CONCLUSION
Benzoxazole derivatives showed promising anti
inflammatory activity. Compound VIc (methoxy phenyl)
is found to be the most potent compound with an
inhibition of paw volume of 73.70 percent after an hour
of administration.
ACKNOWLEDGEMENTS
The authors are thankful to the Director and Principal,
St.Peter's College of Pharmacy, Madikonda, Kazipet,
Warangal, A.P and St. Peter's Institute of Pharmaceutical
Sciences, Vidyanagar, Hanamkonda, Warangal, A.P, for
providing laboratory facilities and financial support.
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