Oregano and Thyme Essential Oils Encapsulated in Chitosan Nanoparticles as Effective Antimicrobial Agents against Foodborne Pathogens

The use of natural compounds with biocidal activity to fight the growth of bacteria responsible for foodborne illness is one of the main research challenges in the food sector. This study reports the preparation and physicochemical characterization of chitosan nanoparticles loaded with Thymus capitatus (Th-CNPs) and Origanum vulgare (Or-CNPs) essential oils. The nanosystems were obtained by ionotropic gelation technique with high encapsulation efficiency (80–83%) and loading capacity (26–27%). Nanoparticles showed a spherical shape, bimodal particle size distribution, and good stability (zeta potential values > 40 mV). The treatment of the nanosuspensions at different temperatures (4 and 40 °C) and storage times (7, 15, 21, and 30 days) did not affect their physicochemical parameters and highlights their reservoir ability for essential oils also under stressful conditions. Both Or-CNPs and Th-CNPs exhibited an enhanced bactericidal activity against foodborne pathogens (S. aureus, E. coli, L. monocytogenes) than pure essential oils. These ecofriendly nanosystems could represent a valid alternative to synthetic preservatives and be of interest for health and food safety.     

Determination of Seven Active Ingredients in Three Plant Essential Oils by Using Micellar Electrokinetic Chromatography

A simple micellar electrokinetic chromatography (MEKC) method was developed for determination of citronellal, citral (Z; E), α-pinene, limonene, linalool, and eugenol in plant essential oils (EOs). A buffer consisting of 20 mM Na₂B₄O₇, 50 mM SDS, 20% (v:v) methanol adjusted to pH 9.5 was found to provide a very efficient and stable electrophoretic system for the analysis. The validation of the method included linearity, LODs, LOQs, precision (intra - and inter - day variation of migration time and peak area), and recovery. Seven terpenoids presented good linearity (R ² > 0.9960) within the test ranges; LODs (S/N = 3) and LOQs (S/N = 10) were 0.2–1.8 µg/mL and 0.8–5.9 µg/mL, respectively. The precision and accuracy were satisfactory, with the overall intra- and inter-day variation (for migration time and peak area, RSD%) being less than 7.0%, and recoveries of this method were greater than 91% at spiked levels. The proposed method was successfully applied to the determination of seven terpenoids in clove oil, litsea cubeba oil, and citronella oil, respectively.     

Essential Oils of Seven Lamiaceae Plants and Their Antioxidant Capacity

Oxidative stress has been reported as a cause of many diseases like Parkinson′s, Alzheimer′s, cardiovascular disease, and diabetes. Oxidative stress can also lead to cancer formation by promoting tumor development and progression. Antioxidants derived from Lamiaceae plants play an important role in natural medicine, pharmacology, cosmetology, and aromatherapy. Herein, we examine the antioxidative capacity of essential oils from seven aromatic Lamiaceae plants against the synthetic radicals DPPH and ABTS. Among the essential oils analyzed, the most robust scavenging capacities were found in mixtures of volatile compounds from thyme and savory. The scavenging activity of tested EOs against the ABTS radical was clearly higher than activity towards DPPH. Analysis of essential oils with weaker antioxidant activity has shown that volatile compounds from marjoram, sage, and hyssop were more active than EOs from lavender and mint. It can be suggested that the potent antioxidant capacity of thyme (Thymus vulgaris) and savory (Satyreja hortensis) are related to a high level of phenolic constituents, such as thymol and carvacrol. On the other hand, the elevated antioxidative power of marjoram, sage, and hyssop essential oils may also be due to their terpinene, o-cymene, terpinolene, and terpinen-4-ol constituents. Although non-phenolic components are less active than thymol or carvacrol, they may affect antioxidant capacity synergistically.     

The Inhibition of Non-albicans Candida Species and Uncommon Yeast Pathogens by Selected Essential Oils and Their Major Compounds

The epidemiology of yeast infections and resistance to available antifungal drugs are rapidly increasing, and non-albicans Candida species and rare yeast species are increasingly emerging as major opportunistic pathogens. In order to identify new strategies to counter the threat of antimicrobial resistant microorganisms, essential oils (EOs) have become an important potential in the treatment of fungal infections. EOs and their bioactive pure compounds have been found to exhibit a wide range of remarkable biological activities. We investigated the in vitro antifungal activity of nine commercial EOs such as Thymus vulgaris (thyme red), Origanum vulgare (oregano), Lavandula vera (lavender), Pinus sylvestris (pine), Foeniculum vulgare (fennel), Melissa officinalis (lemon balm), Salvia officinalis (sage), Eugenia caryophyllata (clove) and Pelargonium asperum (geranium), and some of their main components (α-pinene, carvacrol, citronellal, eugenol, γ-terpinene, linalool, linalylacetate, terpinen-4-ol, thymol) against non-albicans Candida strains and uncommon yeasts. The EOs were analyzed by GC-MS, and their antifungal properties were evaluated by minimum inhibitory concentration and minimum fungicidal concentration parameters, in accordance with CLSI guidelines, with some modifications for EOs. Pine exhibited strong antifungal activity against the selected non-albicans Candida isolates and uncommon yeasts. In addition, lemon balm EOs and α-pinene exhibited strong antifungal activity against the selected non-albicans Candida yeasts. Thymol inhibited the growth of all uncommon yeasts. These data showed a promising potential application of EOs as natural adjuvant for management of infections by emerging non-albicans Candida species and uncommon pathogenic yeasts.     

Chemical Composition and Insecticidal Activities of Essential Oils against the Pulse Beetle

Pulse beetles, Callosobruchus chinensis and Callosobruchus maculatus, are essential pests of cowpea, gram, soybean and pulses. Application of synthetic insecticides against the pulse beetle has led to insect resistance; insecticide residues on grains affect human health and the environment. Essential oils (EOs) are the best alternatives to synthetics due to their safety to the environment and health. The main objective of the investigation was to study the chemical composition and insecticidal activities of EOs, their combinations and compounds against the pulse beetle under laboratory. Neo-isomenthol, carvone and β-ocimene are the significant components of tested oils using GC-MS. Mentha spicata showed promising fumigant toxicity against C. chinensis (LC₅₀ = 0.94 µL/mL) and was followed by M. piperita (LC₅₀ = 0.98 µL/mL), whereas M. piperita (LC₅₀ = 0.92 µL/mL) against C. maculatus. A combination of Tagetes minuta + M. piperita showed more toxicity against C. chinensis after 48 h (LC₅₀ = 0.87 µL/mL) than T. minuta + M. spicata (LC₅₀ = 1.07 µL/mL). L-Carvone showed fumigant toxicity against C. chinensis after 48 h (LC₅₀ = 1.19 µL/mL). Binary mixtures of T. minuta +M. piperita and M. spicata showed promising toxicity and synergistic activity. EOs also exhibited repellence and ovipositional inhibition. The application of M. piperita can be recommended for the control of the pulse beetle.     

Anti-Biofilms’ Activity of Garlic and Thyme Essential Oils against Salmonella typhimurium

Biofilm control by essential oil (EO) application has recently increased to preclude biofilm production on foods and environmental surfaces. In this work, the anti-biofilm effects of garlic and thyme essential oils using the minimum inhibitory concentration (MIC) method against Salmonella typhimurium recovered from different abattoir samples were investigated along with the virulence genes (InvA, SdiA and Stn genes), and the antimicrobial susceptibility profile of S. typhimurium as well. The obtained results revealed that S. typhimurium contaminated abattoir samples to varying degrees. The InvA gene was investigated in all isolates, whereas the SdiA and Stn genes were observed in four and three isolates, respectively. Utilizing the disc diffusion method, S. typhimurium isolates demonstrated substantial resistance to most of the examined antibiotics with a high multiple antibiotic resistance index. S. typhimurium isolates demonstrated biofilm formation abilities to various degrees at varied temperatures levels (4 °C and 37 °C). In conclusion, the obtained samples from the research area are regarded as a potential S. typhimurium contamination source. Furthermore, garlic essential oil (GEO) has more potential to inhibit S. typhimurium biofilm at different sub-minimum inhibitory concentrations as compared to thyme essential oil (TEO). Therefore, these EOs are considered as potential natural antibacterial options that could be applied in food industry.     

Antifungal Properties of Essential Oils and Their Compounds for Application in Skin Fungal Infections: Conventional and Nonconventional Approaches

Essential oils (EOs) are known to have varying degrees of antimicrobial properties that are mainly due to the presence of bioactive compounds. These include antiviral, nematicidal, antifungal, insecticidal and antioxidant properties. This review highlights the potential of EOs and their compounds for application as antifungal agents for the treatment of skin diseases via conventional and nonconventional approaches. A search was conducted using three databases (Scopus, Web of Science, Google Scholar), and all relevant articles from the period of 2010–2020 that are freely available in English were extracted. In our findings, EOs with a high percentage of monoterpenes showed strong ability as potential antifungal agents. Lavandula sp., Salvia sp., Thymus sp., Citrus sp., and Cymbopogon sp. were among the various species found to show excellent antifungal properties against various skin diseases. Some researchers developed advanced formulations such as gel, semi-solid, and ointment bases to further evaluate the effectiveness of EOs as antifungal agents. To date, most studies on the application of EOs as antifungal agents were performed using in vitro techniques, and only a limited number pursued in vivo and intervention-based research.     

Evaluation of Antibacterial and Antifungal Effects of Calcium Hydroxide Mixed with Two Different Essential Oils

Background: Calcium hydroxide is a routinely used material for root canal disinfection during root canal treatment. Natural products have great potential in terms of their antibacterial effects. This study aimed to establish an effective alternative intracanal medicament using Origanum dubium (O. dubium) and Mentha spicata (M. spicata) essential oils. Materials and Methods: O. dubium and M. spicata, collected from Lefke, Cyprus, were separately subjected to hydrodistillation. The obtained essential oil compositions were analysed simultaneously by gas chromatography (GC) and gas chromatography/mass spectrometry (GC-MS). The compositions were then divided into groups and mixed with calcium hydroxide at a 1:1 concentration; after that, the pastes were tested on Enterococcus faecalis (E. faecalis) and Candida albicans (C. albicans), which are the most common resistant pathogenic microorganisms in the root canal. The antibacterial activity of the pastes was measured using a disk diffusion assay. Results: The GC and GC-MS analyses revealed that O. dubium and M. spicata had major compositions of carvacrol (75.8%) and carvone (71.3%), respectively. Antimicrobial activity was found to be significantly higher when study groups with O. dubium essential oil were applied to both E. faecalis and C. albicans. The results also show that M. spicata, together with calcium hydroxide, demonstrated a significant antifungal effect on C. albicans when incubated for 72 h. Conclusions: M. spicata was found to be an effective antimicrobial agent on C. albicans, whereas O. dubium was found to be very effective on both E. faecalis and C. albicans. These data demonstrate that these natural essential oils may be promising candidates for alternative intracanal medicament in future routine clinical applications.     

Combination of GC-MS Molecular Networking and Larvicidal Effect against Aedes aegypti for the Discovery of Bioactive Substances in Commercial Essential Oils

Dengue is a neglected disease, present mainly in tropical countries, with more than 5.2 million cases reported in 2019. Vector control remains the most effective protective measure against dengue and other arboviruses. Synthetic insecticides based on organophosphates, pyrethroids, carbamates, neonicotinoids and oxadiazines are unattractive due to their high degree of toxicity to humans, animals and the environment. Conversely, natural-product-based larvicides/insecticides, such as essential oils, present high efficiency, low environmental toxicity and can be easily scaled up for industrial processes. However, essential oils are highly complex and require modern analytical and computational approaches to streamline the identification of bioactive substances. This study combined the GC-MS spectral similarity network approach with larvicidal assays as a new strategy for the discovery of potential bioactive substances in complex biological samples, enabling the systematic and simultaneous annotation of substances in 20 essential oils through LC₅₀ larvicidal assays. This strategy allowed rapid intuitive discovery of distribution patterns between families and metabolic classes in clusters, and the prediction of larvicidal properties of acyclic monoterpene derivatives, including citral, neral, citronellal and citronellol, and their acetate forms (LC₅₀ < 50 µg/mL).     

Implementation of an Analytical Method for Spectrophotometric Evaluation of Total Phenolic Content in Essential Oils

Over the past decade, there has been growing interest in polyphenols’ research since these compounds, as antioxidants, have several health benefits, such as preventing neurodegenerative diseases, inflammation, cancer, cardiovascular diseases, and type 2 diabetes. This study implements an analytical method to assess the total phenolic content (TPC) in essential oils using Folin–Ciocalteu’s phenol reagent and quantifies the individual phenolic compounds by liquid chromatography. Thus, the research design and methodology included: (1) extraction of essential oil from dried thyme leaves by hydrodistillation; (2) spectrophotometric measurement of TPC by Folin–Ciocalteu method; and (3) identification and quantification of individual phenolic compounds by high-performance liquid chromatography-diode array detection/electrospray ionization mass spectrometry (HPLC-DAD-ESI-MS). Results revealed a TPC of 22.62 ± 0.482 mg GAE/100 µL and a polyphenolic profile characterized by phenolic acids (52.1%), flavonoids (16.1%), and other polyphenols (31.8%). Thymol, salvianolic acid A, and rosmarinic acid were the major compounds of thyme essential oil. The proposed analytical procedure has an acceptable level of repeatability, reproducibility, linearity, LOD (limit of detection), and LOQ (limit of quantification).     

GC-MS Analysis and Microbiological Evaluation of Caraway Essential Oil as a Virulence Attenuating Agent against Pseudomonas aeruginosa

The emergence of resistant microbes threatens public health on our planet, and the emergence of resistant bacteria against the most commonly used antibiotics necessitates urgent alternative therapeutic options. One way to fight resistant microbes is to design new antimicrobial agents, however, this approach takes decades of research. An alternative or parallel approach is to target the virulence of bacteria with natural or synthetic agents. Active constituents from medicinal plants represent a wide library to screen for natural anti-virulence agents. Caraway is used as a traditional spice and in some medicinal applications such as carminative, antispasmodic, appetizer, and expectorant. Caraway essential oil is rich in terpenes that were previously reported to have antimicrobial activities. In our study, we tested the caraway essential oil in sub-inhibitory concentration as a virulence agent against the Gram-negative bacteria Pseudomonas aeruginosa. Caraway essential oil in sub-inhibitory concentration dramatically blocked protease activity, pyocyanin production, biofilm formation, and quorum sensing activity of P. aeruginosa. The gas chromatography–mass spectroscopy (GC-MS) profile of caraway fruit oil identified 13 compounds representing 85.4% of the total oil components with carvone and sylvestrene as the main constituents. In conclusion, caraway essential oil is a promising virulence-attenuating agent that can be used against topical infections caused by P. aeruginosa.     

In Vitro Activity of Essential Oils Distilled from Colombian Plants against Candida auris and Other Candida Species with Different Antifungal Susceptibility Profiles

Multi-drug resistant species such as Candida auris are a global health threat. This scenario has highlighted the need to search for antifungal alternatives. Essential oils (EOs), or some of their major compounds, could be a source of new antifungal molecules. The aim of this study was to evaluate the in vitro activity of EOs and some terpenes against C. auris and other Candida spp. The eleven EOs evaluated were obtained by hydro-distillation from different Colombian plants and the terpenes were purchased. EO chemical compositions were obtained by gas chromatography/mass spectrometry (GC/MS). Antifungal activity was evaluated following the CLSI standard M27, 4th Edition. Cytotoxicity was tested on the HaCaT cell line and fungal growth kinetics were tested by time–kill assays. Candida spp. showed different susceptibility to antifungals and the activity of EOs and terpenes was strain-dependent. The Lippia origanoides (thymol + p-cymene) chemotype EO, thymol, carvacrol, and limonene were the most active, mainly against drug-resistant strains. The most active EOs and terpenes were also slightly cytotoxic on the HaCaT cells. The findings of this study suggest that some EOs and commercial terpenes can be a source for the development of new anti-Candida products and aid the identification of new antifungal targets or action mechanisms.     

Insecticidal,Repellent and Antifeedant Activity of Essential Oils from Blepharocalyx cruckshanksii (Hook. & Arn.) Nied. Leaves and Pilgerodendron uviferum (D. Don) Florin Heartwood against Horn Flies,Haematobia irritans (Diptera: Muscidae)

Haematobia irritans is a cosmopolitan obligate blood-feeding ectoparasite of cattle and is the major global pest of livestock production. Currently, H. irritans management is largely dependent on broad-spectrum pesticides, which has led to the development of insecticide resistance. Thus, alternative control methods are needed. Essential oils have been studied as an alternative due to their wide spectrum of biological activities against insects. Thus, the main aim of this study was to evaluate the insecticidal, repellent and antifeedant activity of the essential oils from Blepharocalyx cruckshanksii leaves and Pilgerodendron uviferum heartwood against horn flies in laboratory conditions. The composition of the essential oils was analyzed using gas chromatography coupled to mass spectrometry. Accordingly, α-pinene (36.50%) and limonene (20.50%) were the principal components of the B. cruckchanksii essential oil, and δ-cadinol (24.16%), cubenol (22.64%), 15-copaenol (15.46%) and δ-cadinene (10.81%) were the most abundant compounds in the P. uviferum essential oil. Mortality of flies and feeding behavior were evaluated by non-choice tests, and olfactory response was evaluated using a Y-tube olfactometer. Both essential oils were toxic to horn flies, with LC₅₀ values for B. cruckchanksii essential oil of 3.58 µL L⁻¹ air at 4 h, and for P. uviferum essential oil of 9.41 µL L⁻¹ air and 1.02 µL L⁻¹ air at 1 and 4 h, respectively. Moreover, the essential oils exhibited spatial repellency in the olfactometer using only 10 µg of each oil, and these significantly reduced the horn fly feeding at all doses evaluated. Although further laboratory and field studies related to the insectistatic and insecticide properties of these essential oils against H. irritans are necessary, B. cruckshanksii leaves and P. uviferum heartwood essential oils are promising candidates for horn fly management.     

Characterization of Primary Action Mode of Eight Essential Oils and Evaluation of Their Antibacterial Effect against Extended-Spectrum β-Lactamase (ESBL)-Producing Escherichia coli Inoculated in Turkey Meat

The current study aims to evaluate the antimicrobial activity of eight essential oils (EOs) against multidrug-resistant Escherichia coli strains, producing extended-spectrum β-lactamase (ESBL) enzymes and isolated from foods. Disc-diffusion assay showed that the inhibition diameters generated by EOs varied significantly among the tested EOs and strains. In fact, EOs extracted from Thymus capitaus, Eucalyptus camaldulensis, Trachyspermum ammi and Mentha pulegium exerted an important antimicrobial effect against tested strains, with the diameters of inhibition zones varied between 20 and 27 mm. Moreover, minimal inhibition and bactericidal concentration (MIC and MBC) values demonstrated that T. capitatus EOs generate the most important inhibitory effect against E. coli strains, with MIC values ranging from 0.02 to 0.78%. Concerning the mode of action of T. capitatus EO, the obtained data showed that treatment with this EO at its MIC reduced the viability of E. coli strains, their tolerance to NaCl and promoted the loss of 260-nm-absorbing material. In addition, in the presence of T. capitatus EO, cells became disproportionately sensitive to subsequent autolysis. Moreover, the inhibitory effect of T. capitatus was evaluated against two E. coli strains, experimentally inoculated (10⁵ CFU/g) in minced turkey meat, in the presence of two different concentrations of EO (MIC and 2 × MIC), and stored for 15 days. In both samples, EO exerted a bacteriostatic effect in the presence of concentrations equal to MIC. Interestingly, at 2 × CMI concentration, the bactericidal activity was pronounced after 15 days of storage. Our results highlighted that the use of essential oils, specially of T. capitatus, to inhibit or prevent the growth of extended-spectrum β-lactamase (ESBL)-producing E. coli in food, may be a promising alternative to chemicals.