The beverage tea is produced from the plant Camellia sinenesis. A recent investigation
reported that the antibacterial activity obtained using methanoic extract of black tea diluted in
ethanol is enhanced in combination with Chloramphenicol, Levofloxacin and Gentamycin
when applied against S. mutans (Mughal et al., 2010). However, information regarding
possible synergy between green tea extracts and antibiotics is scant. This study investigated
the modification of the antibacterial activity of aqueous green tea extract in conjunction with
Penicillin G against the gram positive bacterium Bacillis subtilis.
MATERIAL AND METHODS
Dried green tea leaves of mass 10g were packed into small metal thimbles and immersed in
boiling distilled water (100cm3) for 10 minutes. The contents were drained into a sterilised
beaker to form the stock solution. Samples of green tea extract with relative concentrations of
0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, and 1 were prepared; the sample of relative concentration of 1 was obtained by diluting the stock solution with distilled water in the ratio
1:4. In addition, distilled water was used as a control solution.
Bacterial samples of Bacillus subtilis were cultured on nutrient agar using Petri dishes, with
three antibiotic discs (Penicillin G, 6mg) on each surface. Agar samples had been prepared by
agar dilution (Wiegand et al., 2008) using the green tea extract samples of the respective
concentrations (agar: tea extract 3:1); there were three Petri dishes per concentration. The
dishes were sealed and incubated at 30°C for 24 hours.
Following incubation, the area of each inhibition zone was estimated. Inhibition area was
plotted against green tea concentration. Strength of association was measured using Pearson’s
correlation. A linear regression line was fitted to the data, and a statistical test based on the tdistribution
(Bland, 1995) was applied to determine whether the gradient observed was
A positive association (Pearson’s correlation r = 0.755) was found between the inhibition
area and the relative concentration of green tea extract. The regression line, shown in the
Figure, indicates a relationship of strong statistical significance (p<0.001).
These findings suggest synergism between green tea extract and antibiotics, in like manner to
that reported by Mughal et al. (2010) for black tea extract. Mughal et al. investigated
synergism for Chloramphenicol, Tetracycline, Levofloxacin and Gentamycin using black tea
extract concentrations between 0.1 mg/ml and 1.0 mg/ml. The minimum inhibitory
concentration (MIC) values found were 0.1 mg/ml for Chloramphenicol, Levofloxacin,and
Gentamycin. Tetracycline, with an MIC value of 0.5 mg/ml, was found not to enhance
antibacterial activity. It was not possible to estimate the MIC value for Penicillin G with
green tea extract as synergy was observed even at the lowest extract concentration.
It is difficult to make a direct comparison between the green tea extract and black tea extract
studies. Firstly, different ranges of tea extract concentrations were used; based on the method
of preparation for the stock solution the concentrations of green tea extract in the agar were
much greater, being in the range 0.5 mg/ml to 5.0 mg/ml. Secondly, this study had an
aqueous solvent whereas the black tea extract study used methanol, and methanolic extract
diluted in ethanol. Jonathan and Fasidi (2003) reported that compared to water, methanol is a
more effective solvent for extracting antimicrobial substances. Thirdly, this investigation
involved agar dilution whereas the well diffusion method was used by Mughal et al. Finally,
different antibiotics were tested in the two studies. In the light of these differences in study
design, a comparison of green tea extract and black tea extract under the same experimental
conditions is required.
This investigation has shown that aqueous green tea extract solution significantly enhances
the antibacterial effect of Penicillin G against the Bacillus subtilis bacterium. These findings
highlight the potential that green tea extract may offer in addressing the growing problem of
bacterial resistance to antibiotics. However, more research is merited, to determine whether
similar effects occur with other types of solvent, antibiotic and bacteria.
This research was conducted by the author as a student at Mid-Kent College. Statistical
advice was obtained from Mr. N. Smeeton, King’s College London. The author is grateful for
the helpful comments of a referee.
1. Bland M. An Introduction to Medical Statistics (Second Edition). Oxford: Oxford
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2. Jonathan SG, Fasidi IO. Antimicrobial activities of two Nigerian edible macro-fungi -
Lycoperdon Pusilum and Lycoperdon Giganteum. Afr J Biomedical Research 2003;
3. Mughal T, Tahir A, Qureshi S, Nazir T, Rasheed M. Antibacterial activity of black tea
against Streptococcus mutans and its synergism with antibiotics. J App Pharm 2010;
Wiegand I, Hilpert K, Hancock REW. Agar and broth dilution methods to determine
the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat Protoc
2008; 3(2): 163-175.