pyrazolines are nitrogen - containing heterocyclic compounds , well known for their pronounced biological activity .
it was demonstrated that the combination of pyrazole with azole ring , linked to each by one sigma bond , led to more biologically active targets ; for example , pyrazolylthiazoles showed excellent antimicrobial activities . continuing our work in this research field [ 69 ] and in an attempt to identify new and potent antimicrobial agents
, we tried here to generate new benzofuryl 2-pyrazolin-1-ylthiazoles as antimicrobial agents using simple methods .
the starting pyrazoline-1-carbothioamide 5 was prepared by treatment of 2-acetylbenzofuran 1 with equivalent of 4-flurobenzaldehyde 2 in the presence of 10% alcoholic naoh in 90% ethanol with stirring at room temperature to give chaconne 3 .
reaction of chalcone 3 with equivalent amount of thiosemicarbazide was performed in ethanol in the presence of 2.5 equivalent of sodium hydroxide to the target precursor 5 .
the resulting pyrazoline-1-carbothioamide 5 was cyclized to the corresponding 2-(3-(benzofuran-2-yl)-5-(4-fluorophenyl)-4,5-dihydro-1h - pyrazol-1-yl)-4-methyl-5-(p - subs.phenyldiazenyl)thiazole derivatives 8a  d by reaction with hydrazonoyl halides 6a  d in anhydrous ethanol and in the presence of an equivalent of triethylamine ( scheme 1 ) .
the reaction product structures were elucidated by means of nmr , ms spectroscopy , and elemental analyses ( table 1 ) .
h nmr spectra of 8a  d contained two doublet - doublet and one triplet signals due to the presence of ch2 adjacent asymmetric carbon .
the mass spectra of 8a  d showed the molecular ion peaks at m / z 481 , 499 , 515 and 561 , respectively in agreement with the calculated masses .
the reaction between pyrazoline-1-carbothioamide 5 and the equivalent amount of -haloketones , for example , phenacyl bromides derivatives , 2-bromoacetylbenzofuran , and 3-bromoacetylcoumarin , was performed in refluxing ethanol to yield pyrazol-1-ylthiazoles 1315 in good yields via the intermediates a ( scheme 2 ) .
the h nmr of compounds 1316 indicate the disappearance of nh2 signal due to blocking of nh2 with thiazole nucleus .
the mass spectra of 13a , b16 showed the molecular ion peaks at m / z 473 , 518 , 479 , 507 , and 465 , respectively in agreement with the calculated masses .
all the new synthesized compounds were screened for their antibacterial and antifungal activities at 100  g / ml concentration against four gram - positive bacteria ( staphylococcus aureus atcc 29213 ; b. subtilis atcc6633 ; b. megaterium atcc 9885 ; sarcina lutea ) , three gram - negative bacteria ( klebsiella pneumoniae atcc13883 ; pseudomonas aeruginosa atcc27953 ; e. coli atcc 25922 ) , and two yeast ( saccharomyces cerevisiae and candida albicans nrrl y-477 ) .
most of the newly synthesized compounds showed good antimicrobial activities with respect to the control drugs .
compound 8c exhibited the highest potency against all tested organisms with respect to reference drugs .
compounds 8d and 13b inhibited the growth of staphylococcus aureus atcc 29213 with inhibition zones 23 , 22  mm , respectively , while compound 5 showed excellent activities against klebsiella pneumoniae atcc13883 ; pseudomonas aeruginosa atcc27953 ; and e. coli atcc 25922 with inhibition zone about 24  mm . also , compound 8c showed the highest activity against staphylococcus aureus atcc 29213 , saccharomyces cerevisiae , and candida albicans nrrl y-477 with inhibition zone about 23  mm .
the minimum inhibitory concentration ( mic ) of the synthesized compounds against highly inhibited organisms is reported in table 3 .
compounds 5 revealed low mic ( 200  g / ml ) against staphylococcus aureus atcc 29213 , b. megaterium atcc 9885 , and candida albicans nrrl
, compound 8a exhibited high mic ( 16  g / ml ) against b. subtilis atcc6633 ( table 3 ) .
elemental analytical data were carried from the microanalytical unit , cairo university , giza , egypt .
the ir spectra were recorded in potassium bromide disks on a jasco ft / ir-6100 .
h nmr spectra were run on joel - eca 500  mhz in deuterateddimethyl sulphoxide ( dmso - d6 ) .
chemical shifts values (  ) are given in parts per million ( ppm ) .
the mass spectra were performed using mass varian mat ch-5 spectrometer at 70  ev .
( e)-1-(benzofuran-2-yl)-3-(4-fluorophenyl)prop-2-en-1-one 3 ; hydrazonoyl halides ; 1-(benzofuran-2-yl)-2-bromoethanone 9 ; 3-(2-bromoacetyl)-2h - chromen-2-one 10 ; and 2,3-dichloroquinoxaline   were prepared according to the literature . to a suspension of chalcone 3
( 10  mmol , 2.66  g ) and sodium hydroxide ( 25  mmol , 1.0  g ) in ethanol ( 50  ml ) , thiosemicarbazide ( 12  mmol , 1.1  g ) was added .
the mixture was refluxed for 12  h , then left to cool ; the solid product was filtered off , washed with ethanol , and dried . yield 52% ; m.p .
) max / cm 3460 , 3335 ( nh2 ) ; h nmr ( dmso - d6 )  3.09 , 3.14 ( dd , 1h , ch , j = 3.05  hz , j = 3.05  hz ) , 3.94 , 4.06 ( dd , 1h , ch , j = 11.45  hz , j = 11.5  hz ) , 5.88 ( t , 1h , ch , j = 3.05  hz , j = 7.65  hz ) , 7.137.43 ( m , 9h , ar - h ) , 9.44 ( s , 2h , nh2 , d2o - exchangeable ) ; ms m / z ( % ) : 339 ( m , 75 ) , 60 ( 100 ) . to a suspension of compound 5 ( 1  mmol , 0.34  g ) in ethanol ( 20  ml ) , the 1  mmol of appropriate reagent { ( appropriate hydrozonoyl chlorides 6 + et3n ) ; ( appropriate phenacyl bromides 9 ) ; ( 2-bromoacetylbenzofuran 10 ) ; ( 3-bromoacetylcoumarin 11 ) ; or ( 3,4-dichloroquinoxaline 12 ) } was added and heated under reflux for 4  h. after cooling , the precipitate was collected by suction filtration . yield 58% ; m.p.180 - 2c ; h nmr ( dmso - d6 )  2.54 ( s , 3h , ch3 ) , 4.09 , 4.11 ( 2dd , 2h , ch , j = 10.7  hz , j = 9.95  hz ) , 5.87 ( t , 1h , ch , j = 10.7  hz , j = 9.95  hz ) , 7.187.73 ( m , 14h , ar - h ) ; ms m / z ( % ) : 481 ( m , 75 ) , 95 ( 100 ) . yield 66% ; m.p . 201 - 3c ; h nmr ( dmso - d6 )  2.54 ( s , 3h , ch3 ) , 4.07 , 4.12 ( 2dd , 2h , ch , j = 10.7  hz , j = 9.95  hz ) , 5.89 ( t , 1h , ch , j = 10.7  hz , j = 9.95  hz ) , 7.197.73 ( m , 13h , ar - h ) ; ms m / z ( % ) : 499 ( m , 80 ) , 95 ( 100 ) . yield 72% ; m.p . 206 - 8c ; h nmr ( dmso - d6 )  2.53 ( s , 3h , ch3 ) , 4.10 , 4.34 ( 2dd , 2h , ch , j = 10.7  hz , j = 9.95  hz ) , 5.89 ( t , 1h , ch , j = 10.7  hz , j = 9.95  hz ) , 7.197.73 ( m , 13h , ar - h ) ; ms m / z ( % ) : 515 ( m , 70 ) , 95 ( 100 ) .
128 - 30c ; h nmr ( dmso - d6 )  2.55 ( s , 3h , ch3 ) , 4.10 , 4.34 ( 2dd , 2h , ch , j = 10.7  hz , j = 9.95  hz ) , 5.87 ( t , 1h , ch , j = 10.7  hz , j = 9.95  hz ) , 7.197.72 ( m , 13h , ar - h ) ; ms m / z ( % ) : 561 ( m , 62 ) , 95 ( 100 ) . yield 49% ; m.p .
4.014.05 ( dd , 2h , ch2 , j = 11.45  hz , j = 10.33  hz ) , 5.73 ( t , 1h , ch , j = 5.35  hz , j = 6.1  hz ) , 7.177.92 ( m , 14h , ar - h ) ; ms m / z ( % ) : 473 ( m , 80 ) , 91 ( 100 ) . yield 58% ; m.p . 208 - 10c ; h nmr ( dmso - d6 )  4.014.08 ( dd , 2h , ch2 , j = 11.45  hz , j = 10.33  hz ) , 5.71 ( t , 1h , ch , j = 5.35  hz , j = 6.1  hz ) , 7.177.92 ( m , 14h , ar - h ) ; ms m / z ( % ) : 518 ( m , 49 ) , 91 ( 100 ) . yield 63% ; m.p . 252 - 4c ; h nmr ( dmso - d6 )  4.044.07 ( dd , 2h , ch2 , j = 11.45  hz , j = 10.33  hz ) , 5.73 ( t , 1h , ch , j = 5.35  hz , j = 6.1  hz ) , 6.88 ( s , 2h , benzofuryl - h ) , 7.197.70 ( m , 13h , ar - h ) ; ms m / z ( % ) : 479 ( m , 100 ) .
233 - 4c ; h nmr ( dmso - d6 )  4.05 , 4.08 ( dd , 2h , ch2 , j = 11.45  hz , j = 10.33  hz ) , 5.72 ( t , 1h , ch , j = 5.35  hz , j = 6.1  hz ) , 7.227.72 ( m , 14h , ar - h ) , 8.23 ( s , 1h , coumarinyl - h ) ; ms m / z ( % ) : 507 ( m , 100 ) .
; h nmr ( dmso - d6 )  4.02 , 4.06 ( dd , 2h , ch2 , j = 11.45  hz , j = 10.33  hz ) , 5.70 ( t , 1h , ch , j = 5.35  hz , j = 6.1  hz ) , 7.227.72 ( m , 13h , ar - h ) ; ms m / z ( % ) : 465 ( m , 100 ) .
chemical compounds were individually tested against a panel of gram - positive and gram - negative bacterial pathogens and yeast .
antimicrobial tests were carried out by the agar well diffusion method   using 100  l of suspension containing 1  10  cfu / ml of pathological tested bacteria and 1  10  cfu / ml of yeast spread on nutrient agar ( na ) and sabourund dextrose agar ( sda ) , respectively . after the media had cooled and solidified , wells ( 10  mm in diameter )
were made in the solidified agar and loaded with 100  l of tested compound solution prepared by dissolving 100  mg of the chemical compound in one ml of dimethyl sulfoxide ( dmso ) .
the inculcated plates were then incubated for 24  h at 37c for bacteria and at 28c for yeast .
ciprofloxacin ( 50  g / ml ) and ketoconazole ( 50  g / ml ) were used as standard for antibacterial and antifungal activities respectively .
after incubation time , antimicrobial activity was evaluated by measuring the zone of inhibition against the test organisms and compared with that of the standard .
the experiment was carried out in triplicate , and the average zone of inhibition was calculated .
the bacteriostatic activity of the active compounds ( having inhibition zones ( iz )  18  mm ) was then evaluated using the twofold serial dilution technique .
twofold serial dilutions of the tested compounds solutions were prepared using the proper nutrient broth .
the final concentration of the solutions was 200 , 100 , 50 , and 25  g / ml .
the tubes were then inoculated with the test organisms , grown in their suitable broth at 37c for 24 hours for bacteria ( about 1  10  cfu / ml and 1  10  cfu / ml of yeast ) , and each 5 ml received 0.1  ml of the above inoculum and incubated at 37c for 24 hours . the lowest concentration showing no growth was taken as the minimum inhibitory concentration ( mic ) .
this precursor reacted with thiosemicarbazide in strong basic medium to afford the n - pyrazoline thioamide 5 .
the new compounds were tested for their antimicrobial activities and significant activities due to presence of three nucleuses : benzofuran , pyrazole , and thiazole .
also , some substituent increases the antimicrobial activities such as chloro substituent in compounds 8c and 13a .
 a new series of 2-pyrazolin-1-ylthiazoles 8a  d and 1316 was synthesized by cyclization of n - thiocarboxamide-2-pyrazoline with different haloketones and 2,3-dichloroquinoxaline . the structures of the new compounds were confirmed by elemental analyses as well as nmr , ir , and mass spectral data . 
 the newly synthesized compounds were evaluated for their antimicrobial activities , and also their minimum inhibitory concentration ( mic ) against most of test organisms was performed . amongst the tested ones , 
 compound 8c displayed excellent antimicrobial activity . 