Similarly, a difference in the relative composition of the essential oils of different species of the Lamiaceae family by Luo et al., (2019) and species of the Apiaceae family by Ngahang Kamte et al., (2018) has been observed. The production and accumulation of metabolites in individuals with a similar species that are growing in different regions may be influenced by environmental factors, such substances appear to play a chemical interface between the plant and the environment (Gutbrodtet et al., 2012). Environmental factors (eg, average temperature, precipitation, and longitude) are likely to play a role in the selection of specialized genotypes/chemotypes for monoterpenes that lead to different chemistry (Taft et al., 2015).
In almost all studies on this plant, thymol had the highest percentage of essential oil with diffrent amount, that the highest amount of thymol by Weisany et al., (2019) from Kurdistan region of Iran (91.15%) observed. Similarly, differences in the proportion of constituent components of essential oils between samples of one plant from different geographical areas, particularly the predominant composition, have been observed by Lukas et al., (2009) for Origanum syr-iacum L., and by Sampaio and Batista (2017) for Tithonia diversifolia (Hemsl.) A.Gray species. These data from literature indicatethat the surrounding environment seems to significantly influencethe production and accumulation of volatile constituents in aerialparts of T. daenensis. Due to the high percentage of thymol, this species can be considered as the main base of this valuable compound. Numerous studies have shown that thymol is one of the phenolic compounds that have been shown to have very effective antimicrobial and antimicrobial properties in EOs (Villanueva-Bermejo et al., 2015; Gavaric et al., 2015 and Ultee et al., 1999).
As reported in Safaei Ghomi et al., (2019) and Talebi et al., (2011), oxygenated sesquiterpene (35.3% and 33.14%), and in the study of Jamzad et al. (2008), oxygenated monoterpene (49%) contained the highest percentage of EO compounds of this species, which did not correspond to the present results.
The yields and effects of medicinal plants vary depending on the locations of growth, climatic and ecological conditions, field operations, growth stages, and genetic traits (Millauskas et al., 2004). Oleic acid (9-octadecenoic acid) is a component of omega-9 fatty acids. This fatty acid is effective in treating cancer and cardiovascular disease, autoimmune diseases, Parkinson's, Alzheimer's, inflammatory diseases and high blood pressure. Its derivatives have a regulatory role on the cell membrane and are used as an anticancer drug that induces cell apoptosis and cell division (Sales-Campos et al., 2013; Choque et al., 2014 and Gonçalves et al., 2014). Linoleic acid is one of the most abundant unsaturated fatty acids in the human diet, which is one of the omega-6 fatty acids and plays an active role in the growth and general health of the human body (Sales-Campos et al., 2013).
Although some common compounds in the present study were similar to the results of Mirza et al., (2001); however, there was a significant difference between the two studies. In particular, none of the major constituents of the essential oil present, was recorded in Mirza et al., (2001). These differences are most likely due to the chemotypic differences that result from the environmental and climate conditions of the studied habitats (Yavari et al., 2010). Caryophyllene oxid inhibits abnormal accumulation of fluid in the intercellular space of body tissues and antitumor (Jaimand and Rezaei, 2006).
Palmitic acid as the predominant compound with amount 9.1% have recorded by Morteza-Semnani et al., (2006). Germacrene D and bicyclogermacrene by Morteza-Semnani et al., (2006) 8.9% and 5.1%, by Sajjadi and Somae, (2004) 16.9% and 16.6%, and by Meshkatalsadat et al., (2007) 32.9% and 7.3% was reported. Differences in the types and percentages of constituents may be due to genetic or non-genetic variations in response to environmental differences such as soil chemical composition and physiographic factors and or quantity of plant material (Zuzarte and Salgueiro, 2015) or various growth stages of the collected species (Ghasemi Pirbalouti et al., 2013), and or different methods for the extraction of EOs (Hashemi et al., 2010). Germacrene are a class of volatile organic hydrocarbons, in particular terpenes. Germacrene in some plant species possess antimicrobial and insecticidal properties (Telascrea et al., 2007).
Basically in all previous reports, 1,8-cineole with a content of 48.72% (Risaliti et al., 2019), 37.06% (Moussi et al., 2019), 32.90% (Pereira et al., 2017), 57.88% (BenAbada et al., 2020), and 26.89% (Bajalan et al., 2017) had the highest percentage of EO of this species and also Camphor with 11.72% (Risaliti et al., 2019),11.81% (Pereira et al., 2017), 18.99% (BenAbada et al., 2020 ), and 24.82% (Bajalan et al., 2017) was the second main compound in previous reports. α-pinene, which accounted for the most in the present study, in previous reports with a content of 9.86% (Risaliti et al., 2019), 10.71% (Pereira et al., 2017), 9.18% (BenAbada et al., 2020), and 20.81% (Bajalan 2017), was the third major constituent of essential oils.These results indicate that the essential oils of these five Lamiaceae species can be considered as economically viable natural sources for this various high-value compounds. Similar trend was observed by Luo et al., (2019) for six species of lamiaceae.
Thymol is a highly effective phenolic and antimicrobial compound in TDEO. Thymol, as the most important phenol present in plant essential oils, is an inexpensive antibacterial source and can be used in pathogenic systems (Nejad Ebrahimi et al., 2008). The antimicrobial effect of thiol is due to permeability of cell membranes, which can be clotted with membrane surface cations and disrupt vital activities (Ultee et al., 1999). Similar results have been obtained by Wetwitayaklung et al. (2009) for Aquilaria crassna L. extract against S. aureus due to the presence of some compounds. The antimicrobial effect of essential oils is not limited to their major ingredients. In fact, it is possible that compounds with a lower percentage are likely to have a synergistic effect with otheractive compounds (Khajehie et al., 2017 and Naeini et al., 2009).
TDEO contained high Thymol, and extensive research has shown that Thymol exhibits good antifungal activity against a wide range of plant pathogenic fungi and food contaminants (Villanueva-Bermejo et al., 2015 and Gavaric et al., 2015). The mechanism of action of these compounds has not been fully elucidated, but it is believed that can interfere with or damage the cell wall of the fungi, or cause their cell wall to decay (Isman and Machial, 2006). Which is inconsistent with findings Hosseini Behbahani et al., (2013) for effect of TDEO against A. sniger and C. albicans (∼53.7 and ∼37.7 mm) and Dadashpour et al., (2011) for effect of TDEO against C. albicans (∼63 mm). While no one of the EOs of the other four species did not produced any inhibition zone (ND) on A. brasiliensis, A. sniger, and C. albicans (Table 5).
But Hosseini Behbahani et al., (2013) reported a remarkable effect of TDEO (43 mm) against B. subtilis, which contradicts the present study. This effect may be related to monoterpene and phenolic compounds that are able to break down the outer membrane of Gram-negative strains (Callo et al., 2015).
In previous reports, the effect of TDEO against E. coli was ∼7 mm (GhasemiPirbalouti et al., 2010), ∼7.1 mm (Golkar et al. 2020), ∼27.7 mm (Hosseini Behbahani et al., 2013), and ∼43.67mm (Dadashpour et al., 2011). Inhibition zone TDEO was recorded against Gram-positive S. epidermidis with 9.00 ± 0.001. However, there is no reported effect of this EO on S. paratyphi-A and S. epidermidis. The difference of antimicrobial activities between species from the same genus might be due to the difference of their major compounds and also to the complexity of EOs chemical profiles. In the present study EOs against P. aeruginosa did not produce any inhibition zone. The reverse process has been developed by Luo et al., (2019) for six species of lamiaceae against P. aeruginosa, so that ROEO has created the highest inhibition zone against this bacterium. This difference in antibacterial activity can be due to the difference in the composition of the essential oil of this species in different regions. The biological activities of the essential oils depend on chemical constituents (Popović-Djordjević et al., 2019).
These results are inconsistent with the findings of Ezzatzadeh et al., (2014) for Nepeta asterotricha. Since the major constituents of NSEO were, oleic acid, stearic acid, and linoleic acid, one might argue that the microbial effect is due to these fatty acids. Fatty acids have antifungal and antibacterial activities against many microorganisms whose spectrum of activity varies based on the degree of saturation, the length of the carbon chain and the orientation of the double bond (Mattanna et al., 2014). Studies have shown that Lauric, palmitic and linolenic have potential antibacterial activity against S. aureus (McGaw et al., 2002 and Agoramoorthy et al., 2007). There are no reports of antimicrobial activity of this plant so far and these results are important.
It may be explained by different composition and percentage of active compounds in EOs, variation in subtypes of genotypes, drying and extraction methods that appear to have a significant effect on the capacity to slow down or stop the growth of bacteria or kill them. (Mohsenipour and Hassanshahaian, 2015). ROEO did not create a diameter against other microorganisms, which is in some cases consistent with the results of Risaliti et al., (2019).
Essential oils of different specie due to the large number of chemical compounds, which work simultaneously, have unparalleled antibacterial potentials and prevent bacterial resistance mechanisms. In addition, synergistic interactions between EOs can enhance their natural antimicrobial effect. Therefore, antimicrobial potential cannot be associated with a single component or mechanism of action (Candy et al., 2018). Meanwhile, antibiotic abuse exacerbates the harm of C. albicans (Delcour, 2009). So TDEO as a natural alternative against this fungus has significant and promising potential. The presence of phenolic monoterpenes, and especially p-cymene and Thymol, is a major cause of the strong antifungal activity of TDEO. Phenolic compounds have high antimicrobial properties and in fact, phenolic compounds in plant essential oils have the most effect on the development of the antimicrobial activity (Lemos et al., 2017; Mahboubi et al., 2017). These compounds both penetrate the cell membrane and can contribute to the clotting of cell contents (Burl & Coote 1999 and Dorman & Dean 2000). Also, although α-pinene is present in small amounts in TDEO (∼1.09%), it can be another factor influencing this antifungal activity. Research shows the antifungal effect of this compound (Dorman, 2000). In general, the synergistic effects of the diversity between the main and minor components of essential oils in their biological and antimicrobial activity should be considered (Raut and Karuppayil, 2014). The lowest MBC value of TDEO has belonged to C. albicans, that was consistent with results of Dadashpour et al., (2011).
Similar findings was observed by Morteza- Semnani et al. (2010) for H. elegans and Ahmadi et al. (2010) for H. longiflorus is consistent with. But MBC value of HIEO in other microorganisms was always equal to MIC value of HIEO. This results demonstrates the remarkable inhibitory and lethal potency of this EO against most bacteria. The higher amount of oxygenated sesquiterpenes compounds such as (-)-Caryophyllene oxide (∼24.81%) and α-Cadinol (∼7.66%) in HIEO compared to other essential oils can be one of the factors affecting this antimicrobial activity. similarly, Ali et al., (2017) observed that essnetial oil of Teucrium spicastrum Hedge & A.G.Mill. (Lamiaceae) characterized by the abundance of γ-selinene presented important antimicrobial activities against several micro-organisms.
In the study of Ebrahimabadi et l., (2009), SIEO had no inhibitory effect on any of the microorganisms, so the plant sample of this study is valuable. Differences in habitat conditions are among the most important causes of these biological traits in the plant, which has led to the dominance of acidic compounds and Sesquiterpenes, followed by the ability of this essential oil to control some bacteria. The HIEO MBC value against various bacteria (except S. epidermidis and S. aureus) was always higher than its MIC value, indicating that the lethal potency of this essential oil was lower than its inhibitory potency. Another report showed that aqueous extract from Aquilaria crassna (Wetwitayaklung et al., 2009) rich of the some sesquiterpene compound was active against some microorganisms.
Some differences in the amount of antimicrobial effects observed in this study and similar studies could be due to differences in the growth locations of the EO producing plants, the use of different methods for extraction, harvesting time and even the concentration of EOs (Has-Szymanczuk et al., 2011; Lopez et al., 2005 and Toyoshi et al., 2006). Differences in antimicrobial effects indicated differences in essential oil composition. The ROEO also showed a good inhibitory effect (31.25 µg/mL) against Sh. dysenteriae, E. coli and K. pneumonia, which was weaker than control antibiotics. The lowest MIC value of ROEO was recorded against fungi. The MBC value against the associated microorganisms was also higher than the MIC value, indicating a lower lethal force than the inhibitory power, which is consistent with the results of Messaoudi Moussi et al. (2019). The major compounds that have been attributed to the antimicrobial properties of ROEO include α-pinene, 1,8-cineole, verbenone, and linalool (Okoh, 2010 and Oluwatuyi et al., 2004), among these compounds, three conditions may occur: Synergetic, Antagonist, and Additive. Occurrence of any of these conditions affects the antimicrobial properties of essential oil (Marzouk et al., 2006). There is ample evidence that the essential compounds of essential oils also play an important role in antimicrobial properties by creating a synergistic effect between the main compounds and their easier transfer into the bacterial cell (Burt, 2004 and Tavassoli et al., 2011).
Essential oils (EOs) produced by plants as secondary metabolites, usually composed of different compounds with different ratios. The present study was conducted for the first time on five important species of the Lamiaceae family in Iran which after analysis by chromatography included a variety of compounds such as thymol, oleic acid, (-)-caryophyllene oxide, α-pinene, 1,8-cineole, palmitic acid, (+)spathulenol, germacrene D, bicyclogermacrene, phytol, camphor, and borneol. 1,8-cineole and oleic acid were found in the EOs of five plant species. The diversity of compounds of different species resulted in a variety of antibacterial and antifungal activity. TDEO had the highest inhibitory zone against various microorganisms compared to other essential oils, which had a particularly significant effect against S. aureus and A. brasiliensis. Based on the MIC value results, essentail oils of five species of Lamiaceae had a relatively good effect against Sh. dysenteriae, P. aeruginosa, E. coli, K. pneumonia, C. albicans. Overall, the MIC value of ROEO against all bacteria was significantly more effective than other EOs. These essential oils from five species of Lamiaceae can have potential applications in the food and pharmaceutical industries as natural food preservatives, natural fungicides and a potential natural alternative to some antibiotics. In future research, it is important to identify and separate the main components or general compounds of these essential oils to confirm the usefulness of these compounds.