Investigation of antifungal effect of thyme and cardamom essential oils

Countless pests and diseases threat foods around the world. Most of them are related to pathogenic fungal diseases. Worldwide, post harvest losses have been estimated at 50% because of fungal and bacterial infections. Fungal growth on fresh fruits and vegetables is responsible for food spoilage. Moulds are immanent biological agents that are able to colonize foods because of their potential to synthesize a wide variety of hydrolytic enzymes. They cause pathologic disorders in plants bringing serious economic losses for food producers. Mould growth depends on factors such as pH, water activity (aw), solute concentration, temperature, atmosphere, time, etc. Rhizopus, Aspergillus and Penicillium species cause reduction of shelf life, decrease of quality and rapid deterioration of fruits and vegetables especially after harvesting. Aspergillus species have long been known to be common contaminants of human foods and animal feeds. The occurrence of Aspergillus in foods and feeds is dependent on the substrate and environmental factors. Mycotoxins, low molecular weight secondary metabolites, are produced by various fungi especially genus of Aspergillus, Penicillium, Fusarium. The primary mycotoxins are aflatoxins, ochratoxins, zearalenone, fumonisin, and patulin. Mycotoxins are hazardous to human and animal health due to the carcinogenic, mutagenic, teratogenic, lethal effect. Their activity depends on the type of toxin and their concentration in the food. Poor storage and tropical climate are the suitable environmental conditions that support mould growth and mycotoxin production. Ochratoxin A (OTA) is a mycotoxin commonly present in grapes, cereals, species, coffee, cocoa. It is a toxic metabolite produced by several fungi of the genera Aspergillus and Penicillium, mainly produced by Penicillium verrucosum, P. nordicum, Aspergillus niger, A. ochraceus and A. carbonarius under different environmental conditions. Some of the factors that influence growth and production of Aspergillus toxins are temperature, pH, moisture, antifungal agents and type of substrate. OTA in cereals is mainly produced by Penicillium verrucosum while OTA on grapes, coffee and cocoa is formed by Aspergillus carbonarius. Ochratoxin A has been classified as possible human carcinogen by International Agency for Research on Cancer (IARC). In recent years, consumers' choices are moving towards foods that contain lower levels of chemical preservatives and exhibit more fresh-like and natural characteristics. Traditional medicine, particularly herbal medicines, has increased considerably in both developed and developing countries. Global and national markets for medicinal herbs have been growing rapidly and significant economic incomes are being realized. Investigations have turned to the study of natural antifungals such as essential oils. Essential oils are aromatic, volatile, natural oily liquids obtained by hydrodistillation from plant material and are usually mixtures of several components. They are easily decomposed, environmentally friendly and non toxic. Their natural antimicrobial activity is commonly related to the chemical structure of their components, the concentration in which they are present, and their interactions, which can affect their bioactive properties. The mechanisms of action essential oils can be explained by changes in permeability of cell membranes, granulation of the cytoplasm and cytoplasmic membrane rupture. Temperature, pH, water activity (aw), medium conditions and the microorganism involved may influence the stability of the antifungal activities of essential oils. The aim of this study was to determine the effect of thyme essential oil (0.01; 0.03; 0.06; 0.09; 0.12; 0.15 µg / mL) and cardamom essential oil (0.3; 0.6; 0.9; 1.2; 1.5; 1.8; 2.1; 2.4; 3 µg / mL) on growth and ochratoxin A production by three isolates (35-03X1, 3904-X4, 57-03D1) of Aspergillus carbonarius isolated from dried figs collected from the Aegean region. Thyme and cardamom essential oils were selected aganist to the effects of Aspergillus carbonarius The plates incubated at 30°C and 0.97 water activity. OTA production was determined after 7, 10 and 14 day of incubation at each value of essential oils and concentrations. The growth of fungi evaluated by daily measurement of the colony diameter (mm). Growth rates were calculated with primary model of Baranyi. OTA detection and quantification carried out by high performance liquid chromatography (HPLC) with fluorescence detection after extraction with methanol. Thyme and origanum essential oils demonstrated the highest inhibition zones and ginger essential oil showed lowest inhibition zones of against development of A. carbonarius 3904-X4 isolate. A. carbonarius 3904-X4 isolate has been completely inhibited by using 2.5 µL thyme and origanum essential oils. A. carbonarius 35-03X1 and 57-03D1 isolates completely inactivated 0.09 µg/mL while 3904-X4 isolate inactivated 0.12 µg/mL of thyme essential oil. Complete inactivation for A. carbonarius 35-03X1 and 57-03D1 isolates was provided as 0.6 µg/mL concentration of cardamom essential oil. A. carbonarius 3904-X4 isolate inactivated at 0.9 µg/mL concentration of cardamom essential oil. Thyme essential oil demonstrated higher antifungal effect than cardamom essential oil. The lowest minimum inhibitory concentration (MIC) was obtained from 0.09 µg/mL thyme ve 0.6 µg/mL cardamom essential oils. The highest MIC values were determined for bothof them against the A. carbonarius 3904-X4 isolates. The highest MIC values for thyme and cardamom essential oils were 0.12 µg/mL and 3.0 µg/mL, respectively. In this study, the minimum fungicidal concentration (MFC) values were determined as equal to MIC values for all oils and isolates. According to control, A. carbonarius 3904-X4 isolate showed rapid development by using 0.01 µg/mL thyme essential oil. This isolate was followed by A. carbonarius 35-03X1 and 57-03D1 isolates respectively. The results of statistical analysis, 0.01 µg/mL concentration thyme essential oil had same effect with control, different effect with 35-03X1 isolate. The effect of thyme essential oil is different from the control at 0.03 µg/mL and higher concentrations for the A. carbonarius 3904-X4 and 57-03D1 isolates. Inhibitory effect of cardamom essential oil against isolates of Aspergillus carbonarius is lower than thyme essential oil. The lowest growth rate for cardamom essential oil was determined 1.05 mm/day 1.2 µg/mL concentration oil against 57-03D1 isolate. A. carbonarius 57-03D1 isolate was resistant than other isolates against the thyme and cardamom essential oils of the OTA production. Thyme essential oil was completely prevented OTA production in the studied concentrations. In cases where the presence of toxin, OTA production decreased between 26.5- 99.83 % compared to the control. In this study 3904-X4 isolate was showed better development than other isolates in the thyme and cardamom essential oils analogously to literature findings.