Pavel Kloucek

In vitro growth-inhibitory effect of plant-derived extracts and compounds against Paenibacillus larvae and their acute oral toxicity to adult honey bees.

In total, 26 natural compounds of various chemical classes (flavonoids, alkaloids, terpenoids) and 19 crude extracts from selected plants were tested in vitro for antibacterial activity against three strains of P. larvae, the causal agent of American Foulbrood Disease of honey bees (AFB) by the broth microdilution method. Among the individual substances, sanguinarine (MIC 4 microg/ml), followed by thymoquinone, capsaicin, trans-2-hexenal and nordihydroguaiaretic acid (MIC 4-32 microg/ml) possessed the strongest antibacterial effect. In case of extracts, common hop (Humulus lupulus L.) and myrtle (Myrtus communis L.) methanolic-dichloromethane extracts exhibited the highest growth-inhibitory effect with MICs ranging from 2 to 8 microg/ml. Acute oral toxicity of the most active natural products was determined on adult honey bees, showing them as non-toxic at concentrations as high as 100 microg peer bee. Our study leads to identification of highly potent natural products effective against AFB in vitro with very low MICs compared to those reported in literature, low toxicity to adult honey bees and commercial availability suggesting them as perspective, low cost and consumer-acceptable agents for control of AFB.

Evaluation of Antimicrobial and Anti-Inflammatory Activities of Seed Extracts from Six Nigella Species

Seed extracts from six species of the genus Nigella (Family Ranunculaceae)-Nigella arvensis, Nigella damascena, Nigella hispanica, Nigella nigellastrum, Nigella orientalis, and Nigella sativa-obtained by successive extraction with n-hexane, chloroform, and methanol, were tested for their antimicrobial activity against 10 strains of pathogenic bacteria and yeast using the microdilution method as well as for anti-inflammatory properties by in vitro cyclooxygenase (COX)-1 and COX-2 assay. Chemical characterization of active extracts was carried out including free and fixed fatty acid analysis. Comparison of antimicrobial activity showed that N. arvensis chloroform extract was the most potent among all species tested, inhibiting Gram-positive bacterial and yeast strains with minimum inhibitory concentration (MIC) values ranging from 0.25 to 1 mg/mL. With the exception of selective inhibitory action of n-hexane extract of N. orientalis on growth of Bacteroides fragilis (MIC = 0.5 mg/mL), we observed no antimicrobial activity for other Nigella species. Anti-inflammatory screening revealed that N. sativa, N. orientalis, N. hispanica, N. arvensis n-hexane, and N. hispanica chloroform extracts had strong inhibitory activity (more than 80%) on COX-1 and N. orientalis, N. arvensis, and N. hispanica n-hexane extracts were most effective against COX-2, when the concentration of extracts was 100 microg/mL in both COX assays. In conclusion, N. arvensis, N. orientalis, and N. hispanica seeds, for the first time examined for antimicrobial and anti-inflammatory effects, revealed their significant activity in one or both assays.

Antifungal effect of essential oil components against Aspergillus niger when loaded into silica mesoporous supports

BACKGROUND Essential oil components (EOCs) are known for their antifungal properties; however, their high volatility limits their application as antimicrobial agents. Strategies used for controlling the volatility of EOCs include encapsulation or loading into porous materials. This study evaluated the in vitro antifungal activity of selected EOCs (carvacrol, cinnamaldehyde, eugenol and thymol) against the fungus Aspergillus niger when loaded into MCM-41 and β-cyclodextrin (β-CD). RESULTS Carvacrol and thymol in Mobil Composition of Matter No. 41 (MCM-41) displayed remarkable enhanced antifungal properties in comparison to the pure or β-CD-encapsulated EOCs. In fact, carvacrol and thymol were able to maintain antifungal activity and inhibit fungal growth for 30 days, suggesting better applicability of these EOCs as natural preservatives. CONCLUSIONS The sustained antifungal effect of EOCs encapsulated into silica mesoporous supports was described.

Norsesquiterpene hydrocarbon, chemical composition and antimicrobial activity of Rhaponticum carthamoides root essential oil

A detailed analysis of Rhaponticum carthamoides (Willd.) Iljin root essential oil was carried out by GC, GC-MS and GC-FTIR techniques. In total, 30 components were identified, accounting for 98.0% of total volatiles. A norsesquiterpene 13-norcypera-1(5),11(12)-diene (22.6%), followed by aplotaxene (21.2%) and cyperene (17.9%), were isolated and their structures confirmed by 1D and 2D-NMR spectra (COSY, ROESY, HSQC, HMBC and INADEQUATE). Selinene type sesquiterpenes and aliphatic hydrocarbons were among minor constituents of the essential oil. The oil exhibited antimicrobial activity against 5 of 9 strains of bacteria and yeast, when tested using broth micro-dilution method. Minimum inhibitory concentrations ranged between 32 and 256 microg/ml.

Thymoquinone vapor significantly affects the results of Staphylococcus aureus sensitivity tests using the standard broth microdilution method

The broth microdilution (BMD) method is widely used for the determination of minimum inhibitory concentrations of antimicrobial agents, including volatile oils and their components. In this series of various experiments, we have demonstrated the influence of thymoquinone (TQ) vapor on the results of the BMD test performed with Staphylococcus aureus as a model organism. The spread of vapor from the TQ containing wells (32-512 μg/mL) caused the complete inhibition of staphylococcal growth in adjoining wells initially containing bacterium-inoculated pure Mueller-Hinton broth only and thus produced false positive results of the test. The ability of TQ to pass into the adjoined wells was subsequently confirmed by gas chromatography-mass spectrometry, whereas TQ at concentrations up to 84 μg/mL was detected in these wells after five hours. Based on these results, we suppose that vapors of TQ as well as of other naturally occurring volatile compounds and their mixtures (for example essential oils and plant extracts) can significantly influence results of the standard BMD assay. These observations, therefore, call for development of new appropriate BMD method suitable for assessment of antimicrobial activity of volatile substances.

Identification of potential sources of thymoquinone and related compounds in Asteraceae, Cupressaceae, Lamiaceae, and Ranunculaceae families

AbstractIn this study, forty-seven plant species belonging to seven families were analysed by GC and GC-MS for the contents of pharmacologically effective quinones: dithymoquinone (DTQ), thymohydroquinone (THQ), and thymoquinone (TQ). The results showed that detectable amounts (≥1 mg kg−1) of at least one of these compounds have been found in three species of both Monarda (M. didyma, M. media, and M. menthifolia) and Thymus (T. pulegioides, T. serpyllum, and T. vulgaris) genera, two Satureja (S. hortensis and S. montana) species, and in single representatives of Eupatorium (E. cannabinum), Juniperus (J. communis), and Nigella (N. sativa) genera. The maximum contents of THQ and TQ were found in M. media aerial parts and M. didyma inflorescences (2674 and 3564 mg kg−1 of dried weight, respectively) in amounts significantly exceeding their maximum contents in N. sativa seeds (THQ = 530 mg kg−1 and TQ = 1881 mg kg−1), which are generally considered as the main natural source of both of these compounds. As a conclusion, M. didyma (bergamot) and M. media (purple bergamot) can be recommended as new prospective natural sources of THQ and TQ for pharmaceutical or food industries.

In Vitro Antimicrobial Activity of Some Libyan Medicinal Plant Extracts

Abstract In vitro. antimicrobial activity of 32 extracts (ethanol, hexane, chloroform, and methanol) from eight different Libyan medicinal plants, namely Artemisia herba-alba. Asso (Compositae), Globularia alypum. L. (Globulariaceae), Helichrysum stoechas., DC. (Compositae), Peganum harmala. L. (Zygophyllaceae), Polygonum equisetiforme. Sibth. & Sm. (Polygonaceae), Pulicaria crispa. (Forssk.) Oliv. (Compositae), Rosmarinus officinalis. L. (Labiatae), and Thymus capitatus. Hoffmanns & Link. (Labiatae), was determined against seven bacteria and one yeast strain using the broth microdilution method. The results show that all plants tested possessed antimicrobial activity against at least one of the examined strains at a concentration ≤8 mg/ml. The extracts from H. stoechas., P. equisetiforme., P. crispa., R. officinalis., and T. capitatus. exhibited strongest activity against Gram-positive bacteria with minimum inhibitory concentrations (MICs) ranging from 0.5 to 8 mg/ml. Only a few extracts showed inhibitory activity against Gram-negative strains in this test, whereas those of the H. stoechas. and P. harmala. were among the strongest ones (MICs range 4–8 mg/ml). High anticandidal activity was observed for P. harmala., P. crispa., and T. capitatus., with MICs ranging from 0.25 to 1 mg/ml.

The effect of oregano essential oil on microbial load and sensory atributes of dried meat.

BACKGROUND Microbial load can be controlled using either synthetic or natural preservatives. Particular interest has been focused on the potential application of plant essential oils as safer additives for meat. However, there is no published research on the use of essential oils during the meat drying process. This study was focused on enhancing the meat drying process by using oregano essential oil (OEO) to inhibit the growth of bacteria and thus obtain a value-added dried meat product. The sensory response from assessors is presented. RESULTS It was found that the application of OEO in meat was effective in inhibiting Salmonella enteritidis and Escherichia coli. After 6 h of drying at 55 °C, 2 mL (0.038 mL L-1 air) and 1.5 mL (0.028 mL L-1 air) of OEO were considered as the minimal inhibitory concentrations (MICs) against S. enteritidis and E. coli respectively. Samples treated with 0.75 mL of OEO were more attractive for consumption compared with the control; at a higher concentration of OEO, the sensory quality of the food was affected. CONCLUSION A value-added dried meat product obtained by using OEO to enhance food safety received an acceptable sensory response from consumers.

Essential Oils in the Ranunculaceae Family: Chemical Composition of Hydrodistilled Oils from Consolida regalis, Delphinium elatum, Nigella hispanica, and N. nigellastrum Seeds

In this study, we analyzed the chemical composition of volatile oils hydrodistilled from seeds of Consolida regalis, Delphinium elatum, Nigella hispanica, and N. nigellastrum using GC and GC/MS. In C. regalis, octadecenoic (77.79%) and hexadecanoic acid (8.34%) were the main constituents. Similarly, the oils from D. elatum and N. hispanica seeds consisted chiefly of octadecadienoic (42.83 and 35.58%, resp.), hexadecanoic (23.87 and 28.59%, resp.), and octadecenoic acid (21.67 and 19.76%, resp.). Contrastingly, the monoterpene hydrocarbons α‐pinene (34.67%) and β‐pinene (36.42%) were the main components of N. nigellastrum essential oil. Our results confirm the presence of essential oils in the family Ranunculaceae and suggest chemotaxonomical relationships within the representatives of the genera Consolida, Delphinium, and Nigella. In addition, the presence of various bioactive constituents such as linoleic acid, (−)‐β‐pinene, squalene, or carotol in seeds of D. elatum, N. hispanica, and N. nigellastrum indicates a possible industrial use of these plants.

Susceptibility of some clinical isolates of Staphylococcus aureus to fractions from the aerial parts of Leuzea carthamoides

The antimicrobial activity of the dichloromethane extract from aerial parts of Leuzea carthamoides DC. was tested in vitro against 19 Staphylococcus aureus strains (ATCC 25923, CNCTC Mau 43/60, clinical isolates). The extract was fractionated by column chromatography on silica gel into six fractions (petroleum ether, toluene, dichloromethane, ethyl acetate, methanol and water). The minimum inhibitory concentrations (MICs) of the fractions ranged from 64 to 1024 µg/mL. An ethyl acetate fraction (EA 1) with the widest range of activity inhibited all of the strains with MIC in the range 128–512 µg/mL. This fraction exhibited potent activity against strains which showed associated resistance to oxacillin, ciprofloxacin and erythromycin.