uses of azo

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48

Saudi Pharmaceutical Journal, Vol. 15, No. 1, January 2007

SYNTHESIS AND ANTIBACTERIAL ACTIVITY OF NOVEL

PYRAZOLO[3,4-b]QUINOLINE BASED HETEROCYCLIC AZO

COMPOUNDS AND THEIR DYEING PERFORMANCE

Sanjay F. Thakor, Dinesh M. Patel, Manish P. Patel and Ranjan G. Patel

3-Amino-6-methyl-1H-pyrazolo[3,4-b]quinoline was synthesized in good yield. Monoazo com-

pounds were prepared using this intermediate as diazo component with various heterocycliccoupling components. All the azo compounds were characterized by their percentage yield,melting point, elemental analysis, UV-visible spectra, IR-spectra and dyeing performance on

nylon and polyester fibres and by their antibacterial activity against gram positive and gramnegative bacteria.

Key Words: 3-amino-6-methyl-1H-pyrazolo[3,4-b]quinoline, azo compounds, synthesis, dyeing,

fastness properties, antibacterial activity.

Introduction

The chemistry of quinoline has gained increasingattention due to its various diverse pharmacologicalactivities (1-3). Quinoline ring fused with five or sixmembered ring in linear fashion is found in naturalproducts as well as in synthetic compounds of biolo-gical interest. Dictemine and skimmianine are the

examples of two naturally occurring compoundswhich are associated with smooth muscle contra-

cting properties.Biological importance of azo compounds is well

known for their use as antineoplastics (4), antidia-betics (5), antiseptics (6) and other useful chemo-therapeutic agents. It has been found that the activityof azo linkage increases on the incorporation of

suitable heterocyclic moiety. Pyrazole derivativesare also considered as potent biologically activecompounds (7,8). With this background it has beenthought worth to synthesize linearly fused 3-amino-6-methyl-1H-pyrazolo[3,4-b]quinoline and use as a

diazo component to prepare some new azo com-pounds as possible antibacterial agents. All the azocompounds were also applied on nylon and polyesterfibres as disperse dyes and their dyeing performancehave assessed. The general structure of azo

compounds is shown in Figure 1.

NNH

N

CH3

N N R

6a-i

Figure 1. General structure

3JJ6JJH1J[b-4 3] K K

K

*Department of Chemistry, Sardar Patel University, Vallabh

Vidyanagar-388 120, Gujarat, INDIA.

*To whom correspondence should be addressed.

E-mail: [email protected]


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AZO-QUINOLINES AS ANTIBACTERIALS 49

Experimental

All melting points are uncorrected and deter-mined by an electrothermal melting point apparatus

and expressed inoC. All the coupling components

used were synthesized in laboratory except 8-

hydroxy quinoline which was purchased from Spe-ctrochem Co. The IR spectra were recorded on aNicolet Impact-400D FT-IR spectrophotometer us-ing KBr pellets technique. The 1H-NMR spectrawere recorded on a Hitachi R-1500, 60MHzinstrument using TMS as the internal standard.

Chemical shifts are given in (ppm). The mass spe-ctrum was carried out on a Jeol D-300 model. The

absorption spectra (

max) of all the azo compoundsolutions in DMF were recorded on a Shimadzu UV-240 spectrophotometer.

Preparation of 2-chloro-6-methyl-3-quinoline-car-

boxaldehyde (1):

The title compound was synthesized following asequence of reactions according to a procedure des-cribed in the literature (9,10). Yield 69%; m.p. 123-

125oC.

Preparation of 2-chloro-6-methyl-3-quinoline-car-

boxaldehyde oxime (2):

The title compound was synthesized following areaction according to a procedure described in the

literature (11). Yield 75%; m.p.193-195oC.

Preparation of 2-chloro-6-methyl-3-quinoline-

carbo-nitrile (3):

The title compound was synthesized following a

reaction according to a procedure described in theliterature (12). Yield 70%; m.p.181-183

oC.

Preparation of 3-amino-6-methyl-1H-pyrazolo[3,4-

b]quinoline (4):

The title compound was synthesized following areaction according to a procedure described in the

literature (12). Yield 65%; m.p.331oC.Preparation details of pyrazolo[3,4-b]quinoline

based azo compounds (6a-i)

Diazotization of 3-amino-6-methyl-1H-pyrazolo[3,4-b]quinoline (Scheme 1) was carried out withhydrochloric acid as follows:

Diazotization:

Concentrated hydrochloric acid (2 ml, 0.016mol) was added to a well stirred suspension of 3-amino -6- methyl -1H- pyrazolo [3,4-b] quinoline 4

(0.534 g 0.0027 mol) in water (20 ml) and themixture was heated up to 70

oC and maintained at

that temperature till a clear solution obtained. Aftercooling the solution to 0-5

oC in an ice bath, a solu-

tion of sodium nitrite (0.75 g, 0.0054 mol) in water(20 ml) was added drop wise over a period of 10minutes with stirring. The reaction was stirred at a

temperature below 5oC for an hour. The excess of

nitrous acid (tested for using starch iodide paper)

was removed by adding required amount ofsulphamic acid solution (10%). The clear diazoniumsalt solution 5 thus obtained was used immediatelyin the coupling reaction (Scheme 1).

3-methyl-4-((6-methyl-1H-pyrazolo[3,4-b]quinolin-

3-yl)diazenyl)-1-phenyl-1H-pyrazol-5-ol(6a):

General coupling procedure:

3-methyl - 1- phenyl 5 -pyrazolone (0.469 g,0.0027 mol) was dissolved in sodium hydroxidesolution (7 ml, 10% W/V). The solution was cooledto 0-5

oC in an ice-bath. To this well stirred solution,

the above diazonium solution 5was added dropwisekeeping temperature below 5 oC. The reaction masswas further stirred for 2 hrs at 0-5 oC maintaining thepH 8.0 by adding required amount of 10% sodium

carbonate solution. The reaction mass was thenheated up to 60

oC and diluted with water (80 ml).

The dye was filtered off, washed with hot water untilthe washings were neutral, dried and powered. Theproduct was purified by dissolving in DMF and

pouring in water. 1H-NMR(DMSO-d6): 2.48 (s,3H, CH3quinoline), 2.33 (s, 3H, CH3pyrazole), 8.60(s, 1H, NH), 7.41-7.97 (m, 9H, ArH), 11.34 (s, 1H,OH). m/z 383 (M+), 384(M+1), 406(Na adduct),276.9, 183, 142.

The following compounds were prepared in si-milar manner.

1 - (3 - chlorophenyl ) -3- methy l -4- ((6-methyl-1H-

pyrazolo [3,4-b]quinolin-3-yl)diazenyl)-1H-pyrazol-

5-ol (6b).1H-NMR (DMSO-d6): 2.50 (s, 3H, CH3

quinoline), 2.46 (s, 3H, CH3pyrazole), 8.66 (s, 1H,NH), 7.47-7.99 (m, 8H, ArH), 11.20 (s, 1H, OH).

3-amino-4-((6-methyl-1H-pyrazolo[3,4-b]quinolin-3-yl)diazenyl)-1-phenyl-1H-pyrazol-5-ol(6c):

1H-NMR (DMSO-d6): 2.55 (s, 3H, CH3

quinoline), 6.50 (s, 2H, NH2), 8.59 (s, 1H, NH),7.40-7.95 (m, 9H, ArH), 11.33 (s, 1H, OH).



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50 THAKOR ET AL

2-amino-5-((6-methyl-1H-pyrazolo[3,4-b]quinolin-

3-yl)diazenyl)benzo[d]thiazol-6-ol(6d):1H-NMR (DMSO-d6): 2.50 (s, 3H, CH3quino-

line), 7.10 (s, 2H, NH2), 8.65 (s, 1H, NH), 7.33-7.98(m, 6H, ArH), 10.12 (s, 1H, OH).

7-((6-methyl - 1H-pyrazolo [3,4-b] quinolin -3- yl)

diazenyl) quinolin-8-ol(6e):1H-NMR (DMSO-d6): 2.42 (s, 3H, CH3quinoline),

8.68 (s, 1H, NH), 7.10-8.85 (m, 9H, ArH), 10.26 (s,

1H, OH).

2-mercapto-6-methyl-5-((6-methyl-1H-pyrazolo[3,4-

b]quinolin-3-yl)diazenyl)pyrimidin-4-ol(6f):1

H-NMR (DMSO-d6): 2.42 (s, 3H, CH3quinoline), 8.60 (s, 1H, NH), 2.58 (s, 3H, CH3

pyrimidine), 7.34-7.95 (m, 4H, ArH), 10.23 (s, 1H,OH), 12.35 (s, 1H, SH).

7-hydroxy-4-methyl-8-((6-methyl-1H-pyrazolo[3,4-

b]quinolin-3-yl)diazenyl)-2H-chromen-2-one.(6g):.1H-NMR (DMSO-d6): 2.33 (s, 3H, CH3

quinoline), 8.66 (s, 1H, NH), 2.55 (s, 3H, CH3coumarin), 7.50-7.98 (m, 7H, ArH), 9.90(s, 1H,OH).

5-hydroxy-1,4-dimethyl-6-((6-methyl-1H-pyrazolo

[3,4-b]quinolin-3-yl)diazenyl)Pyrimidin-2 (1H)-one.(6h):1H-NMR (DMSO-d6): 2.40 (s, 3H, CH3

quinoline), 8.60 (s, 1H, NH), 1.2 (s, 3H, CH3), 2.79(s, 3H, CH3-N),7.45-7.96 (m, 4H, ArH), 11.45(s, 1H,OH).

3-(4-hydroxyphenyl)-1-(4-methoxyphenyl)-2-thioxo-

3- ( ( 6- methyl - 1H - pyrazolo [3,4-b] quinolin-3-yl)

diazenyl)dihydropyrimidin-4,6(1H,5H)-dione.(6i):1H-NMR (DMSO-d6): 2.48 (s, 3H, CH3

quinoline), 8.55 (s, 1H, NH), 3.80 (s, 3H, OCH3),3.20 (s, 2H, CH2), 6.97-7.98 (m, 10H, ArH), 10.08(s,

1H, OH).

Dyeing method:

Dyeing of nylon and polyester has been carriedout for 2% shade following standard procedures(13).

Fastness properties:

The fastness to light, sublimation and perspire-ation were assessed in accordance with AATCC

/15/1985. The rubbing fastness test was carried outwith a crockmeter (Atlas) in accordance with

AATCC/88/1988 and the wash fastness test inaccordance with IS: 765-1979. By observing thealternation of dyed pattern light fastness rating wasgiven by grey-scale (1 to 8). Where, 1-very poor, 2-poor, 3-fair, 4-fairly good, 5-good, 6-very good, 7-excellent, 8-outstanding. Rating for sublimation,

perspiration and wash fastness is given by grey-scale(1 to 5). Where, 1-poor, 2-fair, 3-good, 4-very goodand 5-excellent. Data of fastness properties are givenin Table 3.

Antibacterial activity:

Antibacterial activities of synthesized compou-nds were examined in vitroby known agar diffusion

cup method (14-17). All the compounds were testedfor activity against gram-positive bacteria like Baci-llus cereus ATCC 10987, Staphylococcus aureusATCC 6538 and Bacillus subtilis ATCC 6633 andgram-negative bacteria Escherichia coli ATCC10536. The culture medium was nutrient agar. All

the compounds were dissolved in DMF (500ppmconcentration) and DMF used as control. Norfloxa-cin was employed as the standard drug. The resultsare summarized in Table 4.

Results and Discussion

The fused pyrazole intermediate, 3-amino-6-methyl-1H-pyrazolo[3,4-b]quinoline 4was prepared

by dehydration of 2-chloro-6-methyl-3-quinoline-carboxaldehyde oxime with thionyl chloride follow-ed by cyclization with hydrazine hydrate in ethanol.(Scheme 1). The fused pyrazole inter-mediate 4wasdiazotized satisfactorily at 0-5

oC by usual procedure

using hydrochloric acid and sodium nitrite. The

diazonium salt was used immediately since it deco-mposed on standing, even when cold. Subsequent

coupling reactions took place readily on adding thediazonium salt continuously to the solution of coup-ling component in alkaline medium at 0-5

oC main-

taining the pH 8.0 by adding required amount of10% sodium carbonate solution.

Spectral properties of the azo compounds:

The absorption maxima (max) and logarithm of

the molar extinction coefficient (log) of the dyes6a-i are given Table 1. The absorption maxima of

6a-iwere recorded in DMF solution. The absorptionmaxima were in the range of 383 nm to 523 nm. The

value of logarithm of molar extinction coefficient

(log) of the dyes 6a-i were in the range 4.15-4.68.



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CH3

NH2

CH3

NH O

CH3 DMF/POCl3

RefluxN

CH3

Cl

CHO

NH2OH EtOH

N

CH3

Cl

CH

N

CH3

Cl

CN NOH

SOCl2

NH2NH

2

N

CH3

NNH

NH2

Ac2O/AcOH

EtOH

N

CH3

NNH

HCl /NaNO2

N2Cl

Coupling with R

NNH

N

CH3

N N R

1

234

0-5oC

+

65

Wherein, R= various heterocyclic coupling components with their coupling position.

ON

N

CH3

ON

N

CH3

Cl

N

OH N

N

SHCH3

OH

OO

CH3

OHO

N

N

CH3

OH

CH3

N

S

NH2

OH

ON

N

NH2

O O

S

N NOMeOH

a. b. d

e.f.

c.

g. h.

i.

.

Scheme 1:Synthetic route of azo compounds 6a-i.



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52 THAKOR ET AL

Table 1: Absorption maxima, logarithm of molarextinction coefficient and dyebath exhaustion of azocompounds 6a-ion nylon and polyester.

Infrared spectra of azo compounds:

All the azo compounds showed a characteristicband at 1575-1594 cm-1 for the azo (-N=N-) group.The band at 2873-2920 cm

-1is due to C-H stretching

of methyl groups. The band at 1312-1318 cm-1 canbe attributed to bending of aromatic methyl group.The characteristic band at 3120-3178 cm-1is due to

N-H stretching of secondary amine group. The bandappears at 3032-3057 cm

-1 corresponding to C-H

stretching of aromatic rings. The band at 742 cm-1

isdue to a C-Cl stretching.

Dyeing properties of azo compounds

The azo compounds 6a-i were applied at 2%depth on polyester and nylon as disperse dyes. The

properties are given in Table 1. These dyes mostlygave various brown, orange, reddish orange and ye-llow shades with good depth on the fibres. The va-riation of the shades of the dyed fibres results fromthe different coupling components.

Table 2:Physicochemical properties of azo compounds 6a-i.

Comp.

no.

Molecular

Weightg/mol.

Melting

Point (oC)(d)

Yield

(%)

C% H% N%Molecular

formula

Calcd. Found Calcd. Found Calcd. Found

6a C21H17N7O 383 240-245 70 65.98 65.75 4.58 4.43 25.34 25.58

6b C21H16N7OCl 417 290-296 77 60.14 60.23 3.76 3.83 23.88 23.60

6c C20H16N8O 384 276-280 73 62.78 62.88 4.56 4.37 29.45 29.26

6d C18H13N7SO 375 234-237 71 57.87 57.60 4.16 4.36 26.45 26.23

6e C20H14N6O 354 225-228 75 67.34 67.59 3.58 3.44 20.14 20.33

6f C16H13N7SO 351 193-195 67 54.55 54.70 3.95 3.70 27.67 27.92

6g C21H15N5O3 385 201-204 74 65.22 65.45 3.60 3.85 18.56 18.38

6h C17H15N7O2 349 248-255 69 58.21 58.42 4.67 4.49 28.34 28.18

6i C28H21N7SO4 551 259-264 70 60.77 60.98 3.69 3.81 17.92 17.78

Absorption in

DMF

% ExhaustionComp.

no.

max log Nylon Polyester6a 485 4.26 74 70

6b 481 4.15 72 68

6c 468 4.45 73 72

6d 383 4.68 64 62

6e 523 4.34 73 68

6f 394 4.17 71 706g 400 4.28 66 65

6h 477 4.56 70 68

6i 441 4.48 68 63

(d) Decomposition temperature



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Table 3:Fastness properties of azo compounds 6a-i.

Rubbing

Fastness

Perspiration

Fastness

Sublimation

Fastness

Light

Fastness

Wash

Fastness

Dry Wet Acidic Alkaline

Comp.

no.

N P N P N P N P N P N P N P

6a 4 4 4-5 5 4 5 5 4-5 5 5 5 5 5 5

6b 4 4-5 5 5 4 5 4 4 5 5 5 5 5 5

6c 4 4-5 5 4-5 4 4 4 4 5 5 5 5 5 5

6d 3-4 4 5 5 4 4-5 5 4 4-5 4 4 4-5 5 5

6e 4 3-4 5 5 5 4 4 4-5 5 4-5 5 4-5 4-5 5

6f 3-4 4 5 4-5 5 4 4 4 5 5 5 5 5 5

6g 3-4 3 5 5 4 4 4 4-5 4-5 4 5 4-5 5 4-5

6h 4 4-5 5 5 4 4 4 4 5 5 5 5 5 5

6i 3 3-4 4-5 4-5 4 4-5 4 4 4 4-5 4 4 4-5 4

N-nylon and P-polyester

Table 4: Antibacterial activity data of compounds6a-i.

The light fastness of all the disperse dyes on boththe fibres is found fair to fairly good to good. The

obtained results of washing fastness of the dyes forboth the fibres showed that they are very good toexcellent. Fastness to rubbing of dyed patterns wasvery good to excellent for all the dyes on both the

fibres. This is attributed to good penetration andaffinity of dyes to the fibres. The perspiration andsublimation fastness are very good to excellent.

Zone of inhibition (mm)

Compd. E. coli B. subtilis B. cereus S. aureus

6a 13 -- 10 --

6b 15 12 8 10

6c 18 15 8 9

6d 9 -- -- --

6e 10 8 12 9

6f 20 17 13 11

6g 8 -- -- --

6h 10 8 6 5

6i 13 10 7 9

Norfloxacin 40 30 25 37

DMF -- -- -- --

Antibacterial Assay:

The antibacterial assay indicated that none of thetested compounds showed significant activitytowards selected gram positive and gram negative

bacteria. The compound 6fexhibited higher activity

than other against all selected microorganisms. It isobserved that azo compounds 6a, 6band 6cshowedmoderate activity against E.coliand B.subtilis. Theremaining compounds displayed weak activityagainst all microbes under investigation. A closeexamination of the structure of the active compoundrevealed that the best activity was confined with the



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54 THAKOR ET AL

compound having pyrimidine as coupling compo-nent. Moreover pyrazolone as coupling componentin compounds 6a, 6b and 6c showed moderateactivity. In conclusion, the compounds havingpyrimidine and pyrazolone as coupling componentscould be useful for derivatization to develop moreselective anti-ba-cterial agents.

Acknowledgement

The authors are thankful to Dr. K C Patel and

Mr. Kishor H Chauhan, Department of Biosciences,Sardar Patel University for the antibacterial scree-

ning of the compounds. We are also thankful to CD-

RI, Lucknow for recording the mass spectral data.

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