Synthesis and Antiviral Studies of Novel N-Sulphonamidomethyl piperazinyl Fluoroquinolones

A series of novel N-Sulphonamidomethyl piperzinyl fluoroquinolones were synthesized and screened antiviral activity. Eight compounds were synthesized through modifying the N4-hydrogen of piperazine in fluoroquinolones with formaldehyde and sulphanomides by Mannich reactions. The structures of the synthesized compounds were characterized by means of their IR and 1H-NMR spectral data. Synthesized compounds were screened for antiviral activity against influenza A (H1N1, H3N2, H5N1) and influenza B viruses in MDCK cell culture. The antiHIV activities of the new compounds were screened for antiviral activity against replication of HIV-1(IIIB) in MT-4 cells. Cytotoxicity of the synthesized compounds was also tested in mock-infected MDCK and MT-4 cells. Compound CF-SD and CF-SDM inhibits the influenza A (H1N1) and compound GF-SDM inhibit the replication of influenza A (H5N1) and B in MDCK cells. All compounds displayed cytostatic propertity in MT-4 cells. Among the compounds tested, GF-SDM (CC50=39.44 μM) most toxic compound in this series.

Quinolone is a versatile lead molecule for designing potential bioactive agents and its derivatives were reported to possess board spectrum antimicrobial activity.A new fluoroquinolone K12, bearing o-methoxyphenylpiperazinyl group and a difluoromethoxyl group at positions 7 and 8, respectively, was reported to have strong and selective antiHIV-1 activity [1] .The antiviral activity seemed to be related to an inhibitory effect at the RNA transcriptional level.Two K12 analogues bearing a phenyl dehydropiperidinyl moiety at position 7 were effective at inhibiting HIV-1 long terminal repeat (LTR)-driven gene expression, as well as suppressing tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) production in blood mononuclear cells, suggesting a mechanism of action mediated by inhibition of Tat functions [2] .Recently, newer synthesized aryl piperazinyl fluoroquinolones were studied for anti-HIV activity [3][4][5][6] and some of these compounds showed profound activity.A large number of fluoroquinolone have been synthesized and studied for wide range of antiviral activity [7] , but its activity against influenza virus has not been well explored.
Based on these findings, some fluoroquinolones were synthesized and screened for antiviral activity against both HIV and influenza viruses.

Melting points were determined in open capillary
tubes on a Thomas-Hoover melting point apparatus and are uncorrected.IR spectra were recorded using KBr pellets on a Jasco-410 infrared spectrophotometer, 1 H-NMR spectra were determined on Bruker AMX 400 MH Z with tetramethylsilane as an internal standard.The sample was dissolved in DMSO-d 6 and the value was measured in δ ppm.
Compounds were tested for their inhibitory effects against replication of HIV-1 (III B ) in MT-4 cells [10,11] .The MT-4 cells were grown and maintained in RPMI 1640 DM medium supplemented with 10% (v/v) heatinactivated fetal calf serum (FCS), 2 mM-glutamine, 0.1% sodium bicarbonate and 20 µg/ml gentamicin (culture medium).Inhibitory effect of test compounds on HIV-1 replications was monitored by inhibition of virus-induced cytopathic effect in MT-4 cells and was estimated by MTT assay.Briefly, 50 µl of HIV-1 (100-300 CCID 50 ) were added to a flat-bottomed microtiter tray with 50 µl of medium containing various concentrations of compounds.MT-4 cells were added at a final concentration of 6×10 5 cells/ml.After 5 d of incubation at 37°, the number of viable cells were determined by the 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT) method.Cytotoxicity of test compounds against mock-infected MT-4 cells was also assessed by the MTT method.AntiHIV activity and cytotoxicity of standard AZT were also performed by a similar method in MT-4 cells.
The substitutions take place at N 4 -hydrogen of piperazine moiety of flouoroquinolone, which represents a site amenable to significant modification.We report a study of replacing the N 4 -hydrogen of piperazine in flouroquinolone with different substitutions of sulphonamide moiety via mannich reactions to form N-sulphonamido-methyl flouroquinolone derivatives Compound CF-SD and CF-SDM inhibited the replication of influenza A (H1N1).Their inhibitory concentration (EC 50 ) was 16 and 18 µg/ml, respectively, whereas the cytotoxic concentration (CC 50 ) was found to be more than 100 µg/ml (Table 1).The compound GF-SDM inhibited the avian flu (H5N1) with the EC 50 value of 19 µg/ml and CC 50 value of >100 µg/ml.These compound also inhibited significant activity against influenza B with a EC 50 value of 11 µg/ml and CC 50 >100 µg/ml.Other compounds investigated exhibited mild inhibitory activity against influenza viruses.All the compounds exhibited more than 100 µg/ml in uninfected MDCK cells.The standard ribavirin inhibits the replication of influenza A and B viruses in the concentration of 3.7-5.9µg/ml and their cytotoxic concentration was found to be >100 µg/ml The synthesized compounds were evaluated for antiHIV activity against HIV-1 replication in acutely infected MT-4 cells (Table 2).The 50% effective concentration (EC 50 ) values of the compounds against the replication of HIV-1 were higher than the cytotoxic concentrations (CC 50 ).Whereas the standard AZT (EC 50 ) had 0.0064 µM against HIV-1 in MT-4 cells.Cytotoxic concentrations of test compounds were found to be 39-75 µM, whereas the standard AZT showed 65.9 µM in mock infected MT-4 cells Compound GF-SDM (CC 50 =39.44µM)displayed marked cytostatic activity in the MT-4 cells.Presence of 2-pyrimidinyl/4,6-dimethyl-2-pyrimidinyl group in sulphonamide (SO 2 NH 2 ) of N-sulphonamidomethyl piperzinyl flouroquinolone lead molecule is essential for antiviral activity against influenza viruses.Free SO 2 NH 2 of N-sulphonamidomethyl piperizinyl flouroquinolone substitution will reduce the antiviral activity (Table 1).These lead molecules are suitable for designing newer derivatives against influenza viruses based upon promising antiviral activity seen.

Mishra, et al.: Antibacterial Activity of Andrographis paniculata
In the present study the ethanol extract of the aerial part of Andrographis paniculata was prepared and evaluated for antimicrobial activity against eleven bacterial strains by determining minimum inhibitory concentration and zone of inhibition.Minimum inhibitory concentration values were compared with control and zone of inhibition values were compared with standard ciprofloxacin in concentration 100 and 200 µg/ml.The results revealed that, the ethanol extract is potent in inhibiting bacterial growth of both Gram-negative and Gram positive bacteria.

Key Words: Antibacterial activity, Andrographis paniculata
Andrographis paniculata (Acanthaceae) is an annual herb.It is found in Sri Lanka and throughout the plains of India specially Maharashtra, Karnataka, Uttar Pradesh, Tamilnadu, Orissa.Various medicinal properties like choleretic, antidiarrhoeal, immunostimulant and antiinflammatory have been attributed to this plant in the traditional system of Indian medicine [1][2][3][4][5] .Further reported activities are hepatoprotective, antimalarial, antihypertensive, antipyretic, antithrombotic and antidote for snake bites.The present investigation was undertaken to find out the antibacterial potentiality of the ethanol extract of the aerial part against some Gram positive and Gram negative bacteria.