Arvind K Singh et al.

, IJSID, 2011, 1 (3), 353-361

ISSN:2249-5347

IJSID
International Journal of Science Innovations and Discoveries
Research Article
An International peer Review Journal for Science

Available online through www.ijsidonline.info

SYNTHESIS, CHARACTERIZATION AND ANTIBACTERIAL ACTIVITY OF 1, 3, 4-THIADIAZOLE DERIVATIVES Arvind k. singh*, R.Parthsarthy, Kshitiz jyoti, Geeta mishra

*kamla Nehru institute of management and technology (faculty of pharmacy), Sultanpur, Uttar Pradesh, India, *National institute of medical sciences, Jaipur, Rajasthan, India. ABSTRACT
Received: 13.09.2011 Modified: 09.10.2011 Published: 29.12.2011
*Corresponding Author

2,5-Disubstituted 1,3,4-thiadiazoles show various pharmacological activities and have been synthesized from different compounds .2,5 Disubstituted 1,3,4- thiadiazole is popularly known for its antibacterial, antinflammatory and antihypertension activities etc.It is known that the synthesis of heterocyclic compounds tend to contain various types of structure in molecules . The formation of ring system involves the condensation process Synthesis of derivatives of 1,3,4- thiadiazole from different acids . All the newly synthesized compound have been characterized by the IR, 1H NMR, MASS Spectral data by structural elucidation & all newly synthesized compounds were evaluated by the antimicrobial activity by disc diffusion methods & anti inflammatory INTRODUCTION method by carreageenan rat paw edema method.

Name: Arvind. K. Singh Place: Sultanpur Uttar Pradesh, India E-mail: adi_arv26@rediffmail.com

Key word: Thiosemicarbazide, 1, 3, 4-thiadiazole, antibacterial activity. INTRODUCTION

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INTRODUCTION Mostly five membered aromatic system having three heteroatoms at symmetrical position have been studied because of their physiological properties1-2. It is also well established that various derivatives of 1,3,4thiadiazole exhibit broad spectrum of pharmacological properties such as antibacterial and antifungal activities.3 1,3,4- thiadiazole showed antibacterial properties similar to those of well known sulfonamide drugs4. prompted by their finding and in continuation of our efforts in synthesizing various condensed bridge pharmaceutically active groups 5. During recent years there has been intense investigation of different classes of thiadiazole compounds, many of which known to possess interesting biological properties such as antimicrobial, antituberculosis, antiinflammatory, anticonvulsants, antihypertensive, antioxidant, anticancer and antifungal activity6. EXPERIMENTAL All chemicals were supplied by (Merck & S.D.fine chemicals Lucknow India). Meltingpoint(m.p) was determined by open capillary tube method. Purification of the compounds was checked by column chromatography & silica gel-G(60-120mess) & silica gel GF254 (4:1) for preparation of the TLC plates and used the solvent system 5% ethyl acetate in pet. Ether & spots were seen under iodine vapours & UV light chamber. IR spectra were obtained on a Perkin-Elmer 1720 FT-IR-Spectrometer (KBr-soln/pellets). 1HNMR spectra were noted an Brucker Ac-400 MHz spectrometer using TMS as internal standard in DMSO/CDCl3 & mass spectra (m/z %) recorded by VG ZAB-HS (FAB) instrument. General procedure for the 1,3,4-thiadiazole derivatives: Taken the molar concentration (1:1) of aromatic acid & Thionyl chloride in RBF & shake properly and kept the solution for refluxing for 3 to 4 hour at 600C with CaCl2 guard tube & then get product that was the aromatic acid chloride &. added to (0.01 mole) thiosemicarbazide was mixed properly & added the excess benzene as the solvent & shake properly chemical in Round bottom flask & refluxed for 3 to 6 hour at 600C. Then the product was washed with NaHCO3 & recrystalized with ethanol. Checked the TLC for proper finding the solvent chloroform and ethyl- acetate (8:6). Characterization of compounds (Ia-Ig:) Compound Ia:[N-(5-phenyl-[1,3,4] thiadiazole-2-yl)-benzamide] :IR Data (KBr cm-1)): 680.4 (thiadiazole),

3421 (NH), 1623 (C=O).1HNMR Data( DMSO-d6,,ppm): 8.76 (s, 1H, J= 7.633), 7.77 (d,3H,7.623), 7.6 (5Hm,7.61).MASS Data m/z (%): C15H11N3SO, calculated: C=64.05, H=3.91, N=14.94, found: C=63.11, H=3.85, N=14.72%). Compound-Ib:[3-chloro-N-[5-(3-chloro-phenyl)-[1,3,4] thiadiazole-2-yl]-benzamide] :IR (KBr cm-1): 673.6

(thiadiazole), 3333.7 (NH), 1623.8 (C=O). 1HNMR (DMSO): 8.4(1H, s, J=7.73), 7.9(4Hx2, 7.709).MASS Data m/z: C15H9 Cl2N3SO, calculated: C=51.44, H=2.59, N=12.00, found: C=51.85, H=2.59, N=12.10%). Compound-Ic:[4-chloro-N-[5-(4-chloro-phenyl)-[1,3,4]thiadiazole-2-yl]-benzamide] :IR (KBr cm-1) 673.6

(thiadiazole), 333.7 (NH), 1623.8 (C=O).1HNMR (DMSO): 8.4(1H, s, J=7.73), 7.9(4Hx2, 7.709).MASS Data m/z: C15H9Cl2N3SO, calculated: C=51.44, H=2.59, N=12.00, found: C=51.85, H=2.59, N=12.10%).
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Compound-Id:[4-Nitro

-N-[5-(2-Nitro-phenyl)-[1,3,4]

thiadiazole-2-yl]-benzamide]:IR

(KBr

cm-1)

657.7

(thiadiazole), 3404.2 (NH), 1628.8 (C=O).1H NMR Data ( DMSO-d6,,ppm): 8.7(1H, s, NH), 8.1(5x2H, m,aromaticH).MASS Data m/z (%): C15H9N5SO5: Calculated: C=48.52, H=2.44, N=18.86%), found:C=48.12, H=2.40, N=18.71%). General procedure for the compounds and Characterization :(IIa-IId) Compound IIa: 2-amino-5-phenyl-1, 3, 4-thiadiazole: Benzaldehyde (3gm, 0.1M) is dissolved in warm alcohol (42.4ml) and thiosemicarbazide (3gm, 0.1M) is dissolved in warm water (49.45 ml) separately and were mixed both solutions slowly with continuous stirring. Then cool the mixture it produces the product immediately. This was filtered with suction and dried. The product was recrystallized from 50% ethanol to Found compound (I).Compound (I) Thiosemicarbazone) (2gm, 0.05M) was suspended in (67ml) warm water. FeCl3 (4.436gm, 0.15M) was dissolved in (54.7 ml) water quantitatively, and added to Compound (I) suspension slowly with constant stirring. The mixture was heated at 80-90 C for 45 min. This solution was filtered hot and then added the citric acid (4.74gm, o.11M) and Sodium citrate (3.3gm, 0.05M) with continuous stirring . The resulting mixture is neutralized with aq. Ammonia (10%). The mixture was filtered with suction, dried and recrystallized with 50% alcohol to produce the compound (IIa). . Benzene: ethyl acetate: ethanol (4:2:1) was used as mobile phase. The spots were visualized by exposure to iodine vapours.IR: cm-1 3403 (NH), 1514 (C=N), 1057 (N-N), 686 (C-S).1HNMR: (DMSO-d6, , ppm); 7.77-7.72(2H, d, NH2), 7.45-7.41(4H, m, ArH) MASS: (m/2 %) Anal. Calculated for C8H7N3S: C, 53.58; H, 3.90; N, 23.44; S, 17.86. Found C, 53.91; H, 3.93; N, 23.58; S, 17.97. %. Compound: - 2-amino-5-(4-methoxyphenyl)-1, 3, 4-thiadiazole Anisaldehyde (3gm, 0.1M) is dissolved in warm alcohol (42.4ml) and thiosemicarbazide (3gm, 0.1M) is dissolved in warm water (49.45 ml) separately and were mixed both solutions slowly with continuous stirring. Then cool the mixture it produces the product immediately. This was filtered with suction and dried. The product was recrystallized from 50% ethanol to Found compound (I).Compound (I) Thiosemicarbazone) (2gm, 0.05M) was suspended in (67ml) warm water. FeCl3 (4.436gm, 0.15M) was dissolved in (54.7 ml) water quantitatively, and added to Compound (I) suspension slowly with constant stirring. The mixture was heated at 80-90 C for 45 min. This solution was filtered hot and then added the citric acid (4.74gm, o.11M) and Sodium citrate (3.3gm, 0.05M) with continuous stirring . The resulting mixture is neutralized with aq. Ammonia (10%). The mixture was filtered with suction, dried and recrystallized with 50% alcohol to produce the compound (IIb). . Benzene: ethyl acetate: ethanol (4:2:1) was used as mobile phase. The spots were visualized by exposure to iodine vapours.IR: (KBr cm-1 )3410 (NH), 1512 (C=N), 1022 (N-N), 621 (C-S). 1HNMR: (DMSO-d6, , ppm); 8.01(2H, s, NH2), 7.79-7.77(3H, d, OCH3), 6.97-6.89(4H, m, Ar-H). MASS: (m/2 %) Anal. Calculated for C9H9N2OS: C, 51.60; H, 4.30; N, 20.08; O, 7.65; S, 15.30%. Found C, 51.90; H, 4.32; N, 20.18; O, 7.68; S, 15.37. %.

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Compound IIc: - 2-amino-5-(3-nitrophenyl)-1, 3, 4-thiadiazole 3-nitrobenzaldehyde (3gm, 0.1M) is dissolved in warm alcohol (42.4ml) and thiosemicarbazide (3gm, 0.1M) is dissolved in warm water (49.45 ml) separately and were mixed both solutions slowly with continuous stirring. Then cool the mixture it produces the product immediately. This was filtered with suction and dried. The product was recrystallized from 50% ethanol to Found compound (I).Compound (I) Thiosemicarbazone) (2gm, 0.05M) was suspended in (67ml) warm water. FeCl3 (4.436gm, 0.15M) was dissolved in (54.7 ml) water quantitatively, and added to Compound (I) suspension slowly with constant stirring. The mixture was heated at 8090 C for 45 min. This solution was filtered hot and then added the citric acid (4.74gm, o.11M) and Sodium citrate (3.3gm, 0.05M) with continuous stirring . The resulting mixture is neutralized with aq. Ammonia (10%). The mixture was filtered with suction, dried and recrystallized with 50% alcohol to produce the compound (IIc). . Benzene: ethyl acetate: ethanol (4:2:1) was used as mobile phase. The spots were visualized by exposure to iodine vapours. IR:(KBr cm-1) 3406 (NH), 1525 (C=N), 1402 (NO2), 1053 (N-N), 680 (C-S). 1HNMR: (DMSO-d6, , ppm); 8.62(2H, s, NH) 2.65-2.55(4H, m, Ar-H). MASS: (m/2 %) Anal. Calculated for C8H6N4O2S: C, 42.83; H, 2.67; N, 24.98; O, 14.27; S,14.27%. Found C, 43.03; H, 2.68; N, 25.10; O, 14.34; S, 14.34. %. Compound IId: - 2-amino-5-(6-hydroxyphenyl)-1, 3, 4-thiadiazole: Salicylaldehyde (3gm, 0.1M) is dissolved in warm alcohol (42.4ml) and thiosemicarbazide (3gm, 0.1M) is dissolved in warm water (49.45 ml) separately and were mixed both solutions slowly with continuous stirring. Then cool the mixture it produces the product immediately. This was filtered with suction and dried. The product was recrystallized from 50% ethanol to Found compound (I).Compound (I) Thiosemicarbazone) (2gm, 0.05M) was suspended in (67ml) warm water. FeCl3 (4.436gm, 0.15M) was dissolved in (54.7 ml) water quantitatively, and added to Compound (I) suspension slowly with constant stirring. The mixture was heated at 80-90 C for 45 min. This solution was filtered hot and then added the citric acid (4.74gm, o.11M) and Sodium citrate (3.3gm, 0.05M) with continuous stirring . The resulting mixture is neutralized with aq. Ammonia (10%). The mixture was filtered with suction, dried and recrystallized with 50% alcohol to produce the compound (IId) . Benzene: ethyl acetate: ethanol (4:2:1) was used as mobile phase. The spots were visualized by exposure to iodine vapours. IR:(KBr cm-1 ) 3396 (NH), 3139 (OH), 1482 (C=N), 105 (N-N), 705 (C-S). 1HNMR: (DMSO-d6, , ppm); 7.86(1H, s, OH), 7.237.34(4H, m, Ar-H), 6.98-6.86(2H, d, NH2). MASS: (m/2 %) Anal. Calculated for C8H7N3OS: C, 49.07; H, 3.60; N, 21.52; O, 8.19; S, 16.39%. Found C, 49.46; H, 3.60; N, 21.64; O, 8.24; S, 16.48. %. Antibacterial Activity:7 The synthesized compounds were evaluated for their antimicrobial activity against Staphylococcus aureus (gram+ve), B.Subtilis (gram+ve), P.aeuroginosa (gram-ve) and Escherichia coli (gram-ve) by cup plate agar diffusion method by measuring the inhibition zone in mm. Concentration (10g/disc) was used as a standard drug for antibacterial activity. The compounds were screened for antibacterial activity against E.coli, S.aureus, P. aeuroginosa, in nutrient agar medium. These sterilized agar media were poured into petri-dishes and allowed to
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solidify. On the surface of the media microbial suspensions were spread with the help of sterilized triangular loop. A stainless steel cylinder of 8 mm diameter (pre-sterilized) was used to bore cavities. All the synthesized compounds (5g/disc) were placed serially in the cavities with the help of micropipette and allowed to diffuse for 1.0 hr. DMF was used as a solvent for all the compounds, and as a control. These plates were incubated at 370C for 34 hrs for antibacterial activity. The zone of inhibition observed around the cups after incubation was measured and percent inhibition of the compounds was calculated. Table I- Spectral data of compound (Ia-Id and IIa-IId)
S.N. Code MolecularFormula IR (cm1) Analytical data
1HNMR(DMSO,d6,,ppm)

MASS (Calculated) (found) H=3.91, C=63.11, H=3.85, N=14.72%)

Ia

C15H11N3SO

680.4(thiadiazole),3421 (NH),1623(C=O)

8.76 (s, 1H, 7.633),7.77(d,3H,7.623),7.6 (5Hm,7.61)

J=

C=64.05, N=14.94,

Ib

C15H9 Cl2N3SO,

673.6(thiadiazole),3333 .7(NH),1623.8(C=O)

8.4(1H,s,J=7.73),7.9(4Hx2, 7.709)

C=51.44, H=2.59, N=12.00

C=51.85, H=2.59, N=12.10%)

Ic

C15H9 Cl2N3SO

3333.7(NH), 1623.8(C=O),673.6(thia diazole)

8.4(1H,S,J=7.73),7.9(4Hx2, 7.709)

C=51.44, H=2.59, N=12.00

C=51.85,H=2.59, N=12.10%)

Id

C15H9N5SO5

657.7 (thiadiazole),3404.2(NH ),1628.8(C=O)

8.7(1H,S,NH), 8.1(5x2H,m, aromatic H)

C=48.52, H=2.44, N=18.86%),

C=48.12, H=2.40, N=18.71%)

IIa

C8H7N3S

3403(NH),1514(C=N),1 057(N-N),686(C-S) 3410(NH),1512(C=N),1 022(N-N),621(C-S)

7.77-7.72(2H,d,NH2),7.457.41(4H,m,Ar-H) 8.01(2H,s,NH2),7.797.77(3H,d,OCH3 ),6.97-6.89(4H,m,Ar-H)

C,53.58% H,3.90% N,23.44%S,17.86% C,51.60% H,4.30% N,20.08%O,7.65%S,1 5.30%

C,53.91% H,3.93% N,23.58% S,17.97% C,51.90% H,4.32% N,20.18%O,7.68% S,15.37%

IIb

C9H9N3OS

IIc

C8H6N4O2S

3406(NH),1525(C=N),1 402(NO2),1053(NN),680(C-S) 3396(NH),3139(OH),14 82(C=N),105(NN),705(C-S)

8.62(2H,s,NH),2.65-2.55(4H,m,ArH)

C,42.83% H,2.67% N,24.98% O,14.27% S,14.27% C,49.07% H,3.60% N,21.52%O,8.19%S,1 6.39%

C,43.03% H,2.68% N,25.10%O,14.34% S,14.34% C,49.46% H,3.60% N,21.64%O,8.24% S,16.48%

IId

C8H7N3OS

7.86(1H,s,OH),7.23-7.34(4H,m,ArH),6.98-6.86(2H,d,NH2)

RESULT & DISCUSSION Chemistry: All compounds were structurally characterized by using IR, H1-NMR and MASS analysis. The IR absorption band around 686cm-1, 3403cm-1, 1514 cm-1, 1057 cm-1, could be attributed to the C-S-, NH, C=N, N-N,Functional group. The 1HNMR spectra 7.77-7.72(2H, d, NH2), 7.45-7.41(4H, M, Ar-H) showed the presence of singlet, doublet,
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and multiplets peaks which represented the presence of aromatic, amine, and hydroxyl protons respectively. Mass spectra (m/z %) Anal. Calcd for C8H7N3S: C,53.58; H, 3.90; N, 23.44; S, 17.86. Found C, 53.91; H, 3.93; N, 23.58; S, 17.97. % showed the calculated and founded value of the compound. Antibacterial activity: Synthesis and screening of 2-amine-5-phenyl-1,3,4-thiadiazole(IIa), 2-amine-5-(4-nitrophenyl)-1,3,4thiadiazole(IIb), 2-amine-5-(4-methoxyphenyl)-1,3,4-thiadiazole(IIc) and 2-amino-5-(5-hydroxy phenyl)-1,3,4thiadiazole(IId) were tested for the antibacterial activity against following bacteria. a) i) S. aureus, ii) B.subtilis (gram+ve); b) iii) E. coli, iv) pseudomonas (gram ve). The test compounds IIa, IIb, IIc, IId showed good

antibacterial activity against Bacillus subtilis (gram +ve) and P.aeuroginosa at lower and higher concentrations and compounds IIa, IIb showed promising antibacterial activity against Staphylococci(gram+ve) and E. Coli (gram ve) at higher and lower concentrations. Only compound IIc has shown moderate antibacterial activity against P.aeroginosa (gram -ve) where as the test compounds IIb, and IId showed moderate antibacterial activity compared to Ciprofloxacine against pseudomonas (gramve). Table: II -Physical properties of compounds (Ia-Id and IIa-IId)
Compounds 1a 1b 1c 1d IIa IIb IIc IId M. formula C15H11N3SO C15H9N3SOCl
2

M.P. 83 87 87 114 225 220 257 110

Yield 70.43 67.31 72.40 66.24 64 67.3 77.7 81.8

Solvent system Chloroform ethyl- acetate (8:6), Chloroform ethyl- acetate (8:6), Chloroform ethyl- acetate (8:6), Chloroform ethyl- acetate (8:6), Benzene: ethyl acetate: ethanol(4:2:1) Benzene: ethyl acetate:ethanol(4:2:1) Benzene: ethyl acetate:ethanol(4:2:1) Benzene: ethyl acetate:ethanol(4:2:1)
3-4 hr R- COCl +

R.f value 0.804 0.694 0.706 0.634 0.63 0.58 0.71 0.54

M.wt. 281 349 349 371 177.22 207.25 222.22 193.22

Colour Pale yellow Lemon yellow Pale yellow Brick red Slight pink Greenish brown Light Yellow Black

C15H9N3SOCl
2

C15H9N5SO3 C8H7N3S C9H9N3OS C8H6N4O2S C8H7N3OS

Scheme: 1
Reflux R - COOH + SOCl2 1st step

NH2NHCSNH2

Reflux 3 - 6 hr

2nd step

R-CONH

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Scheme: 2
O S R

H 2N

NH

NH2

H OH S R N NH (I) NH2

S te p - 1

Fe

2+

S te p - 2

N R S (II)

N NH2

Compounds No 1a

Compounds No IIa

1b

Cl

IIb

1c

IIc

Cl
1d IId

NO2

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Table: III Antibacterial activity of compounds (Ia-Id to IIa-IId) Zone of inhibition in mm. B.subtilis E.coli P.aeruginosa % (zone of % (zone of % (zone of inhibi inhibition in inhibiti inhibition in inhibition inhibition in tion mm) on mm) mm) 48 20 76.84 14 49.98 18 60 18 69.12 23 53.55 12 64 15 57.6 18 64.26 16 72 21 80.64 16 57.12 21 44 17 65.38 15 53.57 16 48 18 69.23 14 50.01 17 60 20 76.92 15 53.57 18 56 17 65.38 16 57.14 17 100 26 100 28 100 25 CONCLUSION In conclusion, simple and convenient methods for synthesis of the new substituted 1, 3, 4-thiadiazole derivatives, and reported the antibacterial and anti-inflammatory studies of these derivatives. The total of eight derivatives were synthesized and screened for their antibacterial activity against S. aureus, B. subtilis, (gram+ve) and E. coli, P. aeuroginosa (gram-ve) bacteria. The MIC of all compounds was determined by observing the zone of inhibition formed around the cup after 24 hr of incubation for antibacterial activity. Compounds were found to have moderate antibacterial activity. These substituted thiadiazole derivatives can be selected for further studies since it contains Chloro, methoxy, hydroxy, nitro groups substitution, it showed significant anti-inflammatory activity compared to Ibuprofen respectively. According to this study, we could point out that substituted 1, 3, 4thiadiazole derivatives played a very important role as anti-inflammatory and antibacterial agents. These observations may provide some predictions in order to design further promising thiadiazole compounds. ACKNOWLEDGEMENT I am thankful to my sincere project guide Dr R.Parthasarthy (Director of KNIMT faculty of pharmacy) for provide proper guidance towards our project work & thankful to CDRI, Lucknow for spectral analysis of compounds. REFERENCE 1. SAAD H (1996). Design synthesis, characterization and antibacterial activity of 1,3,4-Oxadiazole derivatives. Indian Journal.of Chemistry., 35B: 980. 2. Nanjunda S, Swamy S, Basppa, Priya Bs, Prabhuswamy B, Doreswamy BH (2006). Crystal Structure of Novel 2-butyl-4-chloro-1H-Imidazolyl-5-Carboxaldehyde. European Journal. of Medicinal Chemistry, 41: 531-538. 3. HUI XP, CHU CH, ZANGZU, WANG Q and ZHANG Q (2002). Synthesis and antibacterial activity of 1,3,4Thiadiazole derivatives of 5-amino,2-hydroxybenzoic acid. Indian Journal of. Chemistry., 41B: 2176.

Compounds

Ia Ib Ic Id IIa IIb IIc IId Ciprofloxacin

S.aures (zone of inhibitio n in mm) 12 15 16 18 11 12 15 14 25

% inhibition 72 48 64 56 64 68 72 68 100

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4. Golgolab h. Lalezari, I, hossini l, gohari (1973). Synthesis of pharmaceutically important condensed hetrocyclic 4, 6-di-substituted 1, 2, 4-Triazole-1, 3, 4-Thiadiazole derivatives as antimicrobial activity. Jurnal of advaced. Hetrocyclic. Chemistry, 10: 387-390. 5. SANDSTORM H (1968). Synthesis, characterization and pharmacological activity of 1,3,4- thiadiazole. Journal of advanced . Heterocyclic Chemistry, 9: 165. 6. KAR ASHUTOSH l(2001), A text book of medicinal chemistry, 2nd edition, new age international (p) limited, publishers, page no-372 to 396. 7. BARRY AL (1976). The antimicrobial susceptibility test: Principal and practice. Leu and Febiyer (editors), (Philadelphia, PA, USA, 7: 180.

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