S1
Synthesis and Antimicrobial Activities of Some 6-Methyl-3-Thioxo-2,3-Dihydro1,2,4-Triazine Derivatives
Ahmed A. El-Barbary,[a] Ashraf A. El-Shehawy,[b] and Nabiha I. Abdo[a]
[a] Chemistry Department, Faculty of Science, Tanta University, Tanta 31527, Egypt
[b] Chemistry Department, Faculty of Science, Kafr El-Sheikh University, Kafr El-Sheikh 33516,
Egypt
Email: elshehawy2@yahoo.com
Supplemental Materials
Biological Activities
TESTED ORGANISMS
The microorganisms used in this study included Gram-negative bacteria (Escherichia coli,
Pseudomonas aeruginosa, Aeromonas sp and Klebsiella sp), Gram-positive bacteria (Bacillus
subtilis and Staphylococcus aureus) in addition to the non- filamentous fungus (Candida
albicans). The strains under study were obtained from the culture collection of Bacteriology
laboratory (Microbiology Unit, Faculty of Science, Tanta University, Tanta, Egypt). Bacteria
were maintained on nutrient agar and the fungus was maintained on sabaroud agar slopes.
ANTIMICROBIAL SCREENING
Antibacterial and antifungal activities of the synthesized compounds were tested in vitro
against six different types of bacteria and one fungi strain by the cut plug method. The assay
plates were seeded with the test bacteria or fungus and inoculated with 100 μL containing the
diluted inoculum (107 CFU/mL) of each tested organism that were spread on the corresponding
media. After solidification, the wells were made and 1 mg of the synthesized chemicals were
dissolved in DMSO and inserted in the wells. The plates were incubated at 30οC for 24 h, after
which the diameters of inhibition zones were evaluated. The compounds that produced
promising inhibition zones were taken for further antimicrobial assay in liquid broth media using
different concentrations in order to determine their minimum inhibitory concentrations (MIC).
S2
The growth of each tested organism on a slant was placed in 50 mL of the corresponding
broth and incubated overnight at 30οC. The growth of the test bacteria and fungus was adjusted
to 107 CFU/mL. The synthesized compounds were dissolved in DMSO to make different
concentrations. The chemicals were pipetted and added to flaks containing broth media to give
the final concentrations (0.2-16 mg/mL). The flaks containing the different concentrations of the
chemicals were each inoculated with the test organism at 107 CFU/mL. The flaks were incubated
at 30οC for 24 h or 48 h, then decimal dilutions were prepared and the resulted surviving bacteria
or fungi after 24 h or 48 h were enumerated by the spread-plate method. The counts were carried
out in triplicate for each sample. The number of colonies for the media containing the
concentration of the chemicals (M) and for the media without the chemical compounds or their
concentrations (C) were taken on the surviving cell number (CFU) and the ratio between M and
C (M/C) was taken as indication of surviving cells and with this value, the antimicrobial activity
of different chemicals and their concentrations were evaluated (Figures S 1, S 2 and Table S 1)
Figure S 1: Inhibition growth of different concentrations of compound 1 against E. coli.
S3
Figure S 2: Inhibition growth of different concentrations of compound 1 against C. albicans.
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Table S 1: Diameter of the inhibition zone (mm) produced by 1 mg of compounds 4a,b, 5a, 11a,
12, 14 and 16.
Organism
Candida
albicans
Aeromonas
sp.
Escherichia
coli
Pseudomonas
aeruginosa
Staphylococcus
aureus
Bacillus
subtilis
Klebsiella
sp.
Compound
27± 2.9
ND
ND
ND
ND
28± 3.3
21±1.3
4a
ND
ND
20± 1.7
ND
ND
23± 2.6
ND
4b
25± 0.8
ND
ND
ND
ND
24± 1.5
ND
5a
ND
13± 2.4
ND
ND
31± 1.0
ND
ND
11a
ND
18± 2.4
ND
ND
26± 4.3
ND
ND
12
33± 2.8
ND
18± 2.4
ND
26± 2.4
16± 2.4
30± 1.0
14
ND
11± 1
30± 3.7
ND
ND
ND
16± 1.5
16
ND: Not detected under the experimental condition