Evaluation of antibacterial effect of sesame oil, olive oil and their synergism on Staphylococcus aureus in vitro

INTRODUCTION

Nowadays, infectious diseases are the second cause of death in the world.¹ These infections are followed by exorbitant expenses of treatment and increase in mortality and the outbreak of the infection. According to statistics, nosocomial infections are the main cause of the death of 88 thousand people in the world and the reason of 4.5 billion economic losses in a year.² One of the common bacteria involved in nosocomial infections, which hospitalized patients infected continually, is Staphylococcus aureus.³

S. aureus is a gram-positive Cocci bacteria which could produce pestiferous toxic factors, it is considered in pathogens that interface between humans and animals. S. aureus can survive on dry surfaces for months and infects its host from skin, mouth and nose. Its major hosts are humans and the percentage of its incidence in human is about 30%. According to a report from world health organization (WHO) this bacteria is the most common cause of the post-operative wound infections.^4-8 Researches show that this bacteria is the cause of many skin infections and diseases such as pneumonia, meningitis, osteomyelitis, endocarditis and bacteremia.⁹

Resistance to antibiotics could be seen in many bacteria such as S. aureus, For example some strains of this organism are resistant to aminoglycoside antibiotics, methicillin and primary antibiotic like penicillin. This antibiotic resistance is growing continually, and this problem is related to the patient's duration of hospitalization.^3,10,11

In recent years, due to the side effects and drug resistance of bacteria, herbal medicines are used as a substitutes.¹² Although about 25 to 50 percent of drugs are gained from plants and the usage of medicinal plants has been increased, it still is needed to pay more attention to these ways of struggling against bacteria.¹³

One of these herbs is Sesame (Sesamum indicum L.) that is a flowering plant in the genus Sesamum and the family of Pedaliaceae. Sesame is an annual plant and native to tropical areas. 50% of a sesame seed consist of oil, containing Sesamol, Sesamolin, Sezamin, Sesaminol, γ-tocopherol and polyphenols, Excellent stability of this oil against oxidation is the reason of its usage as a solvent in pharmaceutical industry.^14-17 This oil is a rich source of non-saturated fatty acids like oleic acid and linoleic acid with percentage of 39.09% and 40.39%, respectively.¹⁸ An important feature of this oil is its anti-bacterial and anti-inflammatory properties. So, it has many usages, like treating superficial burns and also it has an important role in traditional medicine.¹⁹

Olive tree (Olea europea L.) is another herb, it grows in tropical and subtropical regions and it has been used for medical purposes since many years ago.^20,21 High consumption of olive oil as a rich source of lipids enhanced people's health in the Mediterranean regions.^22,23

This oil contains phenolic and polyphenolic compounds such as oleuropein and hydroxytyrosol that eliminate free radicals. Olive oil contains fat-soluble vitamins, especially vitamin E, which is one of the most important protective factors against free radicals. It has antimicrobial,
anti-inflammatory and antioxidant activity and is used topically to treat skin diseases.^24-27

Synergism of vegetable oils with no changing in chemical and biological processes, could alter their physicochemical properties and increases their antioxidant activity.²⁸

Since S. Arrears' is resistant to a variety of antibiotics, Using plant sources such as olive oil and sesame oil with their antimicrobial properties is important. So, this study was aimed to determine in vitro antibacterial effect of olive oil, sesame oil and their synergism on S.aureus.

METHODS

In this experimental study, at first, the standard strains of lyophilized S. Areas ATCC 25923 were provided by the Iranian Research Organization for Science and Technology. After culture, some biochemical tests (coagulase, catalase, Dnase) were done to ensure purity of strains. Then, bacterial suspensions based on standard 0.5 McFarland (1.5×10⁸ CFU/ml) was prepared.²⁹

For preparation of olive oil and sesame oil, cold pressing method was used. It should be noted that sesame oil was crude because crude oil, which is obtained by cold pressing method is more stable than the refined oil.

In the next step, antibacterial properties of oils were evaluated by micro-broth dilution method. This means that olive oil, sesame oil and their synergism conducted separately in 96-well sterile plates on
S. aureus, based on the Clinical and Laboratory Standards Institute in triplicate for each sample.³⁰

To do bacteriological tests, sesame and olive oils at concentrations of 2 mg/ml,
4 mg/ml, 8 mg/ml, 16 mg/ml, 32 mg/ml,
64 mg/ml, 128 mg/ml, 256 mg/ml,
512 mg/ml, 1024 mg/ml; synergism of them with equal proportions of sesame oil and olive oil at similar concentrations were prepared. Then, 100 µL of Mueller Hinton broth was introduced into each well. After that, concentration of each oil added to Mueller Hinton broth. Afterwards, 100 µL of bacterial suspension, equal to 0.5 Mc Farland added to the wells. The first well contains only bacterial suspension and Mueller Hinton Broth (positive control) and second well containing Mueller Hinton broth and oil (negative control). After that the samples were incubated at 37 °C for
24 h. Then, optical densities of the samples were read at 650 nm wavelength with an ELISA reader (State Fax 2100, USA).

To determine the minimum inhibitory concentration (MIC), concentrations of the last wells that did not have any turbidity (the lowest concentration) was considered MIC. To determine the minimum bactericidal concentration (MBC), all wells without turbidity were cultured on Mueller Hinton agar and then incubated for 24 h at 37 °C. The lowest concentration of oil that bacteria did not grow in it was considered as MBC.³⁰

RESULTS

Determination of MIC and MBC of the oils against S. aureus was measured with micro-broth dilution technique, and indicated the amount of MIC for sesame oil, olive oil, and their synergism; 32 mg/ml,
32 mg/ml and 8 mg/ml; and also their MBC; 128 mg/ml, 128 mg/ml and 32 mg/ml, respectively (Table 1), (Figure 1).

Table 1: Pattern of S. aureus Sensitivity

(-): indicates bacterial growth, (+): indicates inhibition of growth, (++): indicates the bacteria has been killed.

According to the results MIC and MBC of olive oil and sesame oil are similar, but their synergism has more antibacterial activity than olive oil and sesame oil individually. These results indicate that the mixture of these oils has a positive synergy.

Figure 1: MIC and MBC of olive oil, sesame oil and their synergism.

DISCUSSION