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Antibacterial effects of curcumin: an in vitro

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Antibacterial effects of curcumin: an in vitro minimum inhibitory concentration study

Hayati Gunes, Dumrul Gulen, Reyhan Mutlu, Abdullah Gumus, Tekin Tas and Aynur Eren Topkaya
Toxicol Ind Health published online 4 October 2013
DOI: 10.1177/0748233713498458

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Article
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Antibacterial effects of curcumin: © The Author(s) 2013
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an in vitro minimum inhibitory DOI: 10.1177/0748233713498458
tih.sagepub.com
concentration study

Hayati Gunes1, Dumrul Gulen1, Reyhan Mutlu1,

Abdullah Gumus1, Tekin Tas2 and Aynur Eren Topkaya1

Abstract
Aim: To evaluate the antibacterial effect of curcumin with the minimum inhibitory concentration (MIC)
method in standard bacterial strains. Methods: The in vitro antibacterial activity of curcumin was evaluated
against methicillin-sensitive Staphylococcus aureus (MSSA) (ATCC 29213), methicillin-resistant Staphylococcus
aureus (MRSA) (ATCC 43300), Enterococcus faecalis (ATCC 29212), Bacillus subtilis (ATCC 6633), Pseudomonas
aeruginosa (ATCC 27853), Escherichia coli (ATCC 25922) and Klebsiella pneumoniae (ATCC 700603) using the
macrodilution broth susceptibility test method. After incubation in tubes, the antibacterial activity of curcumin
was detected by a lack of turbidity, which indicated the inhibition of bacterial growth. The concentration in the
tube with the highest dilution showing no turbidity was defined as the MIC. Results: The curcumin MIC values
were 175 mg/ml, 129 mg/ml, 219 mg/ml, 217 mg/ml, 163 mg/ml, 293 mg/ml and 216 mg/ml against P. aeruginosa, B.
subtilis, MSSA, MRSA, E. coli, E. faecalis and K. Pneumonia, respectively. Conclusion: This study revealed
antibacterial effects of curcumin against standard bacterial strains in high concentrations. Animal experiments
have demonstrated that curcumin applied at high doses has strong antibacterial activity. There is a need for
further in vivo studies to shed light on antibacterial effects of curcumin with high concentrations.

Keywords
Acute toxicity, curcumin, antibacterial effect, standard strain, MIC

Introduction et al., 2012a, 2012b), burn wound healing (Kulac

Medicinal plants are used in folk medicine in different
parts of the world for the treatment of many diseases.
Products from some plants, fungi, bacteria and other
organisms continue to be used either as pure com-
pounds or as extracts (Araújo and Leon, 2001). Many
of these compounds, such as curcumin, are derived
from plants and purified. Curcumin is derived from
Curcuma longa L (Araújo and Leon, 2001). It is a
naturally occurring yellow pigment commonly known
as turmeric, which is used as flavouring in both
vegetarian and non-vegetarian food dishes. It also has
digestive properties (Govindarajan, 1980) and has been
employed for centuries in the Indian subcontinent
for the treatment of a variety of illnesses, such as the
common cold, coughs, jaundice, inflammatory bowel
conditions and arthritis (Ammon and Wahl, 1991).
Anti-inflammatory (Chandra and Gupta, 1972; Huang
et al., 1992), antioxidant (Pulla Reddy and Lokesh,
1992; Erenoglu et al., 2011), anti-apoptotic (Aktas
et al., 2013), antiprotozoal (Araújo et al., 1999; Ras-
mussen et al., 2000), nematocidal (Kiuchi et al.,
1993) antiretroviral (Mazumber et al., 1995),
antifungal (Martins et al., 2009; Sharma et al., 2010),
antimalarial (Nandakumar et al., 2006; Reddy et al.,
2005) and antibacterial (Chopra et al., 1941) effects
of curcumin have been identified. However, the anti-
bacterial effects have been identified in only a limited
1
Department of Medical Microbiology, Medical Faculty, Namık
Kemal Üniversitesi, Tekirdag, Turkey
2
Department of Medical Microbiology, Medical Faculty, Abant
Izzet Baysal Üniversitesi, Bolu, Turkey
Corresponding author:
Hayati Gunes, Department of Medical Microbiology, Medical
Faculty, Namık Kemal Üniversitesi, Tıp Fak., Tekirdag 59100,
Turkey.
Email: dr_hgunes@yahoo.com
2 Toxicology and Industrial Health
number of bacteria and in a narrow concentration range
(Bhavani Sankar and Murthy, 1979).
The aim of this study was to evaluate the antibac-
terial effect of curcumin of standard bacterial strains
using the minimum inhibitory concentration (MIC)
method.
Materials and methods
Curcumin compound was obtained from Sigma Chemi-
cals Co. (St Louis, Missouri, USA). It was stored at
20 C until extraction as a powder and allowed to warm
to room temperature before the experiment. The purity
of curcumin was accepted as 67% according to the infor-
mation given by the manufacturer. The in vitro antibac-
terial activity of curcumin was evaluated against
methicillin-sensitive Staphylococcus aureus (MSSA),
methicillin-resistant S. aureus (MRSA), Enterococcus
faecalis, Bacillus subtilis, Pseudomonas aeruginosa,
Escherichia coli and Klebsiella pneumoniae by the
macrodilution broth susceptibility test method.
All the strains were stored as frozen stocks with
15% glycerol at 80 C. Before the experiment, the
stock strains were subcultured on blood agar plates.
Determinations of the MICs of curcumin against the
strains above were determined by broth macrodilution
using twofold serial dilutions in a Mueller–Hinton
Broth medium. The test was carried out in small
sterile glass tubes according to the method reported
previously (Clinical and Laboratory Standards Insti-
tute, 2011). Briefly, the solution of curcumin was
prepared in ethanol to ensure complete solubilisation
in a concentration of 1 mg/ml. Ten sterile tubes were
prepared. A total of 500 ml of Mueller–Hinton Broth
was dispensed into each tube. Curcumin solution
(500 ml) was added to the first tube. The solution was
doubly diluted to achieve per ml: 500 mg, 250 mg,
125 mg, 62.5 mg, 31.3 mg, 15.7 mg, 7.9 mg, 4 mg,
2 mg and 1 mg concentrations. Fresh cultures of stan-
dard strains were prepared, and a standardised bacter-
ial suspension of 0.5 McFarland turbidity was diluted
with serial dilutions, and bacterial count adjusted as
106 cfu/ml. 500 ml bacterial solution was added to
each dilution to yield fold diluted suspensions
(between 250 mg/ml and 0.5 mg/ml). Thus bacterial
concentration was adjusted as 5105 cfu/ml. A suitable
solvent control (ethanol), a positive growth (strains
and ethanol in antibiotic-free medium) and a negative
control (bacteria-free medium and curcumin solution)
were added to the tubes. The tubes were incubated at
37 C for 24 h. After incubation, the antibacterial
Table 1. Standard bacterial strains and their curcumin MIC
values.
Bacteria MIC values (mg/ml)
Staphylococcus aureus (MSSA) 219
S. aureus (MRSA) 217
Enterococcus faecalis 293
Bacillus subtilis 129
Pseudomonas aeruginosa 175
Escherichia coli 163
Klebsiella pneumoniae 216
MSSA: Methicilline susceptible S. aureus; MRSA: Methicilline resis-
tance S. aureus.
activities of curcumin in the tubes were detected by
a lack of turbidity, which indicated the inhibition of
bacterial growth. The concentration in the tube with
the highest dilution showing no turbidity was defined
as the MIC. After detection of the MIC, the first con-
centrations were changed and different concentrations
were achieved. These changes were aimed at capturing
the concentration of the MIC and the previous resis-
tance value for each strain. Serial experiments were
conducted, and the exact MIC values were obtained.
Results
Curcumin showed antibacterial activities against bac-
terial strains at a MIC ranging between 129 mg/ml and
293 mg/ml. The results are given in Table 1.
Discussion
Antibiotic resistance in bacteria is a major worldwide
health problem. To solve this problem, there is an
urgent need to develop new antibacterials. S. aureus
is found in humans as a commensal microorganism.
It is carried, persistently or intermittently, in the ante-
rior nares, the throat, the axilla and the perineum by
approximately half of the general population (Price
et al., 2013; Wertheim et al., 2005). MSSA, which can
be part of the normal flora, can acquire methicillin
resistance. MRSA is the cause of severe hospital infec-
tions. Health-care providers play a significant role in
transferring bacteria from one patient to another and
in contributing to the development of invasive disease.
S. aureus is responsible for toxic shock syndrome,
postoperative wound infection and food poisoning. In
a previous study, the MIC value of curcumin was
187.5–500 mg/ml against S. aureus (Sivasothy et al.,
2013; Tajbakhsh et al., 2008). Similar results were
obtained in the present study. Moreover detected MIC
Gunes et al.
value were quite similar for MSSA and MRSA (219
mg/ml for MSSA and 217 mg/ml for MRSA)
E. faecalis is also responsible for hospital infections.
To the best of our knowledge, there has been no study
on the MIC value of curcumin against enterococci. The
present study detected an MIC value of 293 mg/ml
against E. faecalis. A previous study reported that
curcumin-loaded foam inactivated E. faecalis com-
pletely (Heggel et al., 2010).
B. subtilis does not generally cause human infection,
although flora bacteria can cause severe infections,
especially in immunocompromised patients. In a recent
report, B. subtilis was identified as the cause of bacter-
emia and mediastinitis following oesophageal perfora-
tion (Jeon et al., 2012). A previous study reported that
the MIC value of curcumin against B. subtilis was
125 mg/ml (Wang et al., 2009). The present study found
a similar value (129 mg/ml).
P. aeruginosa causes several infections, such as
septicemia, urinary tract infections and severe pneu-
monia, especially in patients with immune deficiency
and burn patients with primary wound infection
(Ekrami and Kalantar, 2007; Pandey et al., 2011). A
previous study showed that curcumin significantly
decreased the density of P. aeruginosa in some food
products (Hosny et al., 2011). The present study
demonstrated that curcumin inhibited P. aeruginosa
grown in a 175 mg/ml concentration. Therefore, the
result is consistent with the previous finding.
E. coli is present in the human intestine and causes
lower urinary tract infections, diarrhoea and septicae-
mia (Crichton, 2008). In a previous study, the curcu-
min MIC value was between 93.8 mg/ml and 250 mg/ml
against E. coli (Tajbakhsh et al., 2008; Wang et al.,
2009). In the present study, it was 163 mg/ml, consis-
tent with the values reported earlier.
K. pneumoniae is an opportunistic pathogen, which
causes community-acquired and nosocomial infec-
tions, especially in immunocompromised individuals.
It is an important source of various infections, includ-
ing mild urinary infections, severe bacteraemia and
pneumonia, and can be associated with a high rate
of mortality and morbidity (Struve and Krogfelt,
2004). To the best of our knowledge, there has been
no report as yet in the literature of the MIC value of
K. pneumoniae. The present study detected an MIC
value of 216 mg/ml against K. pneumoniae.
In this study, the inhibitory activity of curcumin against
the above-mentioned bacteria was achieved at much
higher concentrations than usual drug concentrations. For
example, the highest concentration of susceptibility for S.
3
aureus was 32 mg/ml, and 8 mg/ml and 16 mg/ml for enter-
ococci and enterobacteriaceae, respectively (Clinical and
Laboratory Standards Institute, 2011). Some studies have
reported that curcumin can be used at higher than normal
doses of drugs without any toxic effects. In a phase II trial,
curcumin was given to patients with advanced pancreatic
cancer in an 8-g daily dose. None of the patients showed
toxic effects (Dhillon et al., 2008). In another study, cur-
cumin was administered to rats in a 2-g/kg dose, and toxic
effects were not observed (Dudhatra et al., 2012). Further
studies are needed to determine the limit of toxic effects in
humans. If curcumin does not produce toxic effects in
humans at the same dose as in rats, a person with a weight
of 80 kg could tolerate a dose of curcumin up to 160 g. In
this case, the doses administered in the present study
remain within the limits of sensitivity. Curcumin has also
been used in local therapy. In a burn wound study, curcu-
min was applied topically as a powder, and wound healing
was more rapid than in a control group (Kulac et al.,
2013). In another study, the application of curcumin as
a foam to wounds infected with E. faecalis inactivated
E. faecalis infection completely (Heggel et al., 2010).
In conclusion, the present study detected antibacter-
ial effects of curcumin against standard bacterial strains
but at higher doses than that of normal drugs. Although
animal experiments showed curcumin can be used in
high doses, new studies necessary to determine the limit
of toxic dose. There is a need for further in vivo studies
to shed light on antibacterial effects of curcumin with
high concentrations. In addition to these, the effective-
ness of curcumin to different infections should be tested.
Acknowledgements
The authors thank Mehmet Başli for helping in detection of
MIC values.
Funding
This research received no specific grant from any funding
agency in the public, commercial, or not-for-profit sectors.
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