How do in vitro digestion and cell metabolism affect the biological activity and phenolic profile of grape juice and wine.

Cabernet Sauvignon grape juice and wine underwent in vitro digestion, resulting in a reduction of most phenolic compounds (10%-100% decline), notably impacting anthocyanins (82%-100% decline) due to pH variations. However, specific phenolics, including p-hydroxybenzoic, protocatechuic, vanillic, p-coumaric, gallic and syringic acids, and coumarin esculetin, increased in concentration (10%-120%). Grape juice and wine samples showed comparable polyphenolic profile during all phases of digestion. Antioxidant activity persisted, and inhibition of angiotensin-I converting enzyme was improved after the digestion process, likely because of increased concentrations of listed phenolic acids and esculetin. Digested grape juice displayed comparable or superior bioactivity to red wine, indicating it as a promising source of accessible grape polyphenols for a broader audience. Nevertheless, Caco-2 cell model metabolization experiments revealed that only 3 of 42 analyzed compounds passed to the basolateral compartment, emphasizing the significant impact of digestion on polyphenol bioactivity, suggesting potential yet unmeasurable and overlooked implications for human health.

Anthriscus sylvestris-Noxious Weed or Sustainable Source of Bioactive Lignans?

Anthriscus sylvestris (L.) Hoffm. (Apiaceae), commonly known as wild chervil, has gained scientific interest owing to its diverse phytochemical profile and potential therapeutic applications. The plant, despite being categorized as a noxious weed, is traditionally used in treating various conditions like headaches, dressing wounds, and as a tonic, antitussive, antipyretic, analgesic, and diuretic. Its pharmacological importance stems from containing diverse bioactive lignans, especially aryltetralins and dibenzylbutyrolactones. One of the main compounds of A. sylvestris, deoxypodophyllotoxin, among its wide-ranging effects, including antitumor, antiproliferative, antiplatelet aggregation, antiviral, anti-inflammatory, and insecticidal properties, serves as a pivotal precursor to epipodophyllotoxin, crucial in the semisynthesis of cytostatic agents like etoposide and teniposide. The main starting compound for these anticancer medicines was podophyllotoxin, intensively isolated from Sinopodophyllum hexandrum, now listed as an endangered species due to overexploitation. Since new species are being investigated as potential sources, A. sylvestris emerges as a highly promising candidate owing to its abundant lignan content. This review summarizes the current knowledge on A. sylvestris, investigating its biological and morphological characteristics, and pharmacological properties. Emphasizing the biological activities and structure-activity relationship, this review underscores its therapeutic potential, thus encouraging further exploration and utilization of this valuable plant resource.

Comprehensive study of phenolic profile and biochemical activity of monovarietal red and white wines from Fruska Gora region, Serbia.

This paper presents a comprehensive study of commercially available monovarietal red and white wines from Fruska Gora mountain in Serbia, conducted to characterize their chemical and biochemical properties. Namely, 72 wines of 18 different Vitis vinifera varieties, including international, domestic and autochthonous ones, were assessed. Phenolic profile of wines was determined qualitatively, using spectrophotometric methods, and quantitatively, using LC-MS/MS and HPLC-UV/VIS methods. Quantified phenolic compounds pointed out differences between red and white wines, where some compounds stood out as distinctive characteristic of analyzed varieties, while others served as chemotaxonomic markers of the Fruska Gora region. Antioxidant and inhibitory properties towards acetylcholinesterase were evaluated for all wines and their relation to phenolic composition was investigated. This study is valuable since chemical and biochemical properties of domestic and autochthonous Fruska Gora varieties are reported for the first time. Also, international varieties studied herein gave distinctive information about the terroir of Fruska Gora region, which is scarce in scientific literature.

Quantitative HPLC-UV Study of Lignans in Anthriscus sylvestris.

Wild chervil (Anthriscus sylvestris) is a wild-growing plant from the Apiaceae family, used as a food in Europe and eastern Asia. Due to its high content of lignans known to possess anti-inflammatory, antiproliferative, antiviral and other activities, it represents a potential functional food. However, quantitative data on lignans are still scarce and limited to deoxypodophyllotoxin, nemerosin and yatein. In this paper, a newly developed and validated reverse-phase HPLC-UV method was used to evaluate the content of 14 lignans in both aerial parts and roots of A. sylvestris. The most abundant root components were found to be deoxypodophyllotoxin (2.0-42.8 mg/g), nemerosin (2.0-23.4 mg/g), yatein (1.1-18.5 mg/g), podophyllotoxone (0.7-20.5 mg/g), guaiadequiol (0.8-8.3 mg/g) and dimethylmatairesinol (0.1-5.2 mg/g). Despite the high intra-population variability, a general trend of an increased lignan content during plant development could be observed in the root samples, whereas an opposite trend was observed in the herb samples. A validation study indicated that some of the investigated compounds-7-oxoaryltetralins and dibenzylbutyrolactones-have low stability and require cold storage in the dark. Furthermore, dibenzylbutyrolactones were confirmed to undergo a fast cis-trans isomerization; therefore, only the total content of these isomers should be reported.

Roots of Apium graveolens and Petroselinum crispum-Insight into Phenolic Status against Toxicity Level of Trace Elements.

Celery (Appium graveolens L.) and parsley (Petroselinum crispum (Mill.) Fuss) are herbs utilized in the everyday diet as spices and culinary flavorings, often used in the chemical and medicinal industries. Despite the knowing benefits of different plants from the Apiaceae family, their chemical composition is closely associated with various extrinsic factors. Environmental loading with trace elements (TEs) can modify a plant's metabolic pathways, change bioactive compounds production, cause plant pollution, and consequently provoke human health issues. Therefore, we established this research aiming to unravel the linkage between TEs accumulation and phenolic status in celery and parsley. Higher As, Cd, and Ni levels were observed in celery, which was followed by greater DPPH∙ radical scavenging activity and higher coumarins content. Contrary, parsley accumulated chromium to a greater extent, was richer in flavonoids, apigenin, and its glucosides. No significant difference between species was found in total phenolic contents, where ferulic and chlorogenic acid dominated in both species. A direct relationship between TEs and selected secondary metabolites was proven by the standardized major axis model. Besides abundant bioactive compounds, analyzed plant species showed a moderate hazard index in the children population, since the hazard index was higher than 1. Therefore, future perspectives should be turned towards the production of genotypes with a lower potential for toxic elements accumulation, so the health benefits of plant food will be more prominent.

Comprehensive study of Anthriscus sylvestris lignans.

Wild chervil (Anthriscus sylvestris) is a widespread, wild-growing herbaceous plant from Apiaceae family, known for high content of lignans related to podophyllotoxin, and thus representing a promising new source for their industrial isolation. The data on detailed chemical profile of A. sylvestris lignans are still lacking. By combining fractionation with non-targeted LC-DAD-ESI-MS/MS metabolite profiling, we have identified, fully or tentatively, 46 lignans, 12 of which were never reported in A. sylvestris and 19 in any biological source. The dominant lignans were found to be nemerosin, yatein, deoxypodophyllotoxin, podophyllotoxin, podophyllotoxone and guayadequiol. In addition to well-known dibenzylbutyrolactones, aryltetralins and 7-oxygenated aryltetralins, we found several oxygenated lignan classes previously undescribed in A. sylvestris - 7-hydroxy, 7-oxo and 8-hydroxydibenzylbutyrolactones, a 7'-oxotetrahydrofuran and a 7-hydroxyarylnaphthalene. To facilitate future rapid classification and identification of lignans in raw extracts, UV, MS and NMR spectral features of different lignan classes are described.

Health assessment of medicinal herbs, celery and parsley related to cadmium soil pollution-potentially toxic elements (PTEs) accumulation, tolerance capacity and antioxidative response.

Celery and parsley are recognized as medicinal herbs and nutraceutical vegetables due to their valuable pharmacological properties and numerous health benefits. However, in recent years, soil loadings with various PTEs have become a serious concern across the world, leading to plants pollution, which can consequently diminish their quality and safety for human consumption. Therefore, we attempted to quantify quality and safety of celery and parsley grown in Cd polluted soil. We examined the presence of PTEs: As, Cu, Fe, Mn, Ni, Cu and Cd in soil and selected herbs, as well as their physiological responses to different Cd exposures (control-without Cd addition, 3 and 6 µg/g Cd of dry soil). Following elevation of Cd in plants, both species showed increasing trend of As, Pb and Cu in plants, which overcome safe limits, with exception for Cu. Further, celery showed strong phytoextraction ability (99.9 µg/g Cd of dry weight) with high potential to tolerate Cd due to the efficient antioxidative machinery. Besides that herbs pollution was evident on the basis of target hazard quotients (HQ), hazard index (HI) and cancerogenic risk (CR), revealing that chronic consumption of contaminated herbs can consequently endanger human health. HI was greater than 1, while CR exceeded safe limits in treated plants, with exception for As. In the point of view of toxicology and food safety, growing of medicinal plants should be strictly regulated and distinguished based on the purpose of growing, and further herbs usage.

Anti-Acinetobacter baumannii activity of selected phytochemicals alone, in binary combinations and in combinations with conventional antibiotics.

Acinetobacter baumannii is one of the emerging multidrug- and pandrug-resistant pathogens. The aim of the study was to determine anti-A. baumannii activity of selected terpenes, terpenoids and phenylpropanoids alone, in binary combinations, and in combinations with conventional antibiotics using microdilution-checkerboard and time-kill curve method. The most effective were terpenoids carvacrol (7.0-28.0 µg mL-1) and thymol (22.0-76.0 µg mL-1), as well as phenylpropanoid compound eugenol (90.5-304.0 µg mL-1), with the active concentrations were comparable to antibiotics. The binary combinations showed additive or indifferent effects. The combination gentamicin-carvacrol was synergistic only against reference strain (FICI = 0.50), while other combinations were additive. The best bacteriostatic activity showed carvacrol, thymol or eugenol in combination with ciprofloxacin (FICI range 0.11-0.50) against both the reference and multidrug-resistant strains. The synergistic effect was further confirmed by time kill curve method and obtained after only 15 h. The results indicate a new possible therapeutic strategy against multiple resistant A. baumannii.

The Effects of Trifluoromethylated Derivatives on Prostaglandin E2 and Thromboxane A2 Production in Human Leukemic U937 Macrophages.

BACKGROUND: A convenient approach to modulation of the inflammation has an influence on the production of inflammatory mediators - icosanoids, generated in arachidonic acid (AA) metabolism. The common therapeutic activity of non-steroidal anti-inflammatory drugs (NSAID), such as aspirin, includes inhibition of two crucial enzymes of AA metabolism - cyclooxygenase- 1 and -2 (COX-1/2), with certain risk for gastrointestinal and renal intolerance. Ever since the enrolment of COX-2, particularly overabundance of its main products prostaglandin E2 (PGE2) and thromboxane A2 (TXA2) in numerous pathological processes was recognized, it became a significant therapeutic target. OBJECTIVE: The aim of this study was to examine the effects of synthesized organo-fluorine compounds on PGE2 and TXA2 production in the inflammation process. METHODS: Trifluoromethyl compounds were synthesized from N-benzyl trifluoromethyl aldimine, commercially available 2-methyl or 2-phenyl α-bromo esters (ß-lactams trans-1 and trans-2 and trifluoromethyl ß-amino ester, respectively) and methyl 2-isocyanoacetate (2-imidazoline trans-4). The reactions proceeded with high geometric selectivity, furnishing the desired products in good yields. The influence of newly synthesized compounds on PGE2 and TXA2 production in human leukemic U937 macrophages on both enzyme activity and gene expression levels was observed. RESULTS: Among the tested trifluoromethyl compounds, methyl trans-1-benzyl-5-(trifluoromethyl)- 4,5-dihydro-1H-imidazole-4-carboxylate (trans-4) can be distinguished as the most powerful antiinflammatory agent, probably due to its trifluoromethyl-imidazoline moiety. CONCLUSION: Some further structural modifications in tested compounds and particularly in the synthesis of different trifluoromethyl imidazolines could contribute to the development of new COX-2 inhibitors and potent anti-inflammatory agents.

Anti-Acinetobacter baumannii activity of Rumex crispus L. and Rumex sanguineus L. extracts

Objective: To examine the effect of Rumex crispus (R. crispus) and Rumex sanguineus (R. sanguineus) plant extracts against isolates of Acinetobacter baumannii (A. baumannii) from wounds, including multidrug-resistant strains.Methods: Six prepared Rumex extracts were subjected to liquid chromatography-tandem mass spectrometry. Antimicrobial activity of extracts and pure compounds (catechin, quercetin, isoquercitrin, emodin, and gallic acid) was examined by a microtiter plate method, while for determination of compound binary combinations activity a checkerboard method was applied. Active fractions of extracts were detected by agar-overlay high-performance thin-layer chromatography-bioautography assay followed by liquid chromatography - diode array detection - mass spectrometry analysis. Results: A total of 28 compounds were detected in two extracts of R. crispus and 26 compounds in four different R. sanguineus extracts, with catechin as a dominant component. Anti-A. baumannii activity was confirmed for all six R. sanguineus and R. crispus extracts at the concentration range from 1 to 4 mg/mL. Neither examined single compounds nor their binary combinations exhibited an anti-A. baumannii activity (MIC>256 μg/mL). The bioautography showed that fractions with the most prominent anti-A. baumannii activity tended to contain more polar compounds, predominantly flavonol (quercetin and kaempherol) glycosides; but also fractions containing flavanone (eriodictyol) glycosides and anthraquinone (emodin) glycosides; and less polar eriodictyol aglycone. Conclusions: The results justify and elucidate the traditional application of R. sanguineus and R. crispus extracts for wound healing, indicating the necessity for their further examination in combat against multidrug-resistant A. baumannii isolates from wounds.

Investigation of the chemical composition and biological activity of edible grapevine (Vitis vinifera L.) leaf varieties.

Chemical composition and bioactivity of eight edible Vitis vinifera L. leaf varieties originating from Fruska Gora, Serbia, including two autochthonous varieties, were evaluated. The LC-MS/MS analysis confirmed the presence of 28 natural products in leaf samples, among which most abundant were glycosylated flavonoids, ellagic and chlorogenic acids. The highest amounts of most of the tested compounds were detected in leaves of autochthonous variety Sila. Bioactivity was assessed by a set of in vitro antioxidant, neuroprotective and cytotoxic assays. Leaves of all examined varieties exhibited average bioactivity, with leaves of autochthonous variety Zupljanka standing out as the most promising sample. To conclude, investigated grapevine leaves proved to be a rich source of nutraceuticals, primarily antioxidants, and their further employment as medicinal food in human diet and beneficial raw material for food, pharmaceutical and cosmetic industries should be supported.

Alchemilla vulgaris agg. (Lady's mantle) from central Balkan: antioxidant, anticancer and enzyme inhibition properties.

The current study was designed to evaluate the phytochemical profile and biological properties (antioxidant, enzyme inhibitory and cytotoxic activity) of methanolic, ethanolic, ethyl-acetate and water extracts of Alchemilla vulgaris from Southeast Serbia (central Balkan) which was traditionally used to alleviate and treat many diseases. Bioactive compounds were characterized by LC-MS/MS technique and biological properties were evaluated using antioxidant, enzyme inhibitory and cytotoxic assays. Twenty-six phenolic compounds were quantified in methanolic, ethanolic, ethyl acetate and water extracts, whereas the highest yields were found in ethyl-acetate (EA) extract (gallic acid, caffeic acid, catchin, quercetin). This extract has also shown the greatest antioxidant, anticancer and inhibitory enzyme activities which were demonstrated for the first time in this study. The obtained results indicated that Alchemilla vulgaris from South Serbia possesses high potential for pharmaceutical applications.

Heavy metal accumulation in vegetable species and health risk assessment in Serbia.

Continuous monitoring of heavy metal content in vegetables is of high priority for population nutrition control, as well as risk assessment for human health. The chemical composition of plants is a reliable indicator of their contamination by hazardous substances accumulated in the environment as a consequence of inadequately applied agro-technology. The main goal of this study was to examine the quality of vegetables that reach consumer markets as a function of growth location. Samples of 11 of the most common vegetable species used in the human diet were collected during a 4-year survey. Vegetables originated from local farm producers who cultivated them at different locations in Vojvodina Province, Serbia. Many vegetable samples contained disturbingly high levels of the investigated metals: cadmium, lead, nickel, and chromium. The plant species with the highest Cd accumulation was spinach, where Cd leaves exceeded the maximum permissible concentrations (MPCs) in more than half of the analyzed samples from different localities (54%). Pb concentrations in spinach were also higher than MPC values (according to Serbian law 3.0 µg/g) in 46% of all analyzed samples. Results showed that Cr levels in all tested vegetable species were below MPC values recommended by the FAO/WHO organization. The largest chromium accumulator was spinach, with average values of 2.3 µg/g, followed by beetroot and parsnips with an average concentration of 1.4 µg/g. The highest average content of Ni in all analyzed vegetable species was also recorded in spinach leaves, with an average value of 2.2 µg/g, followed by broccoli (1.7 µg/g) and tomatoes (1.5 µg/g).

A colorimetric broth microdilution method for assessment of Helicobacter pylori sensitivity to antimicrobial agents.

Helicobacter pylori is a major infective etiological agent of the upper gastrointestinal tract diseases. The bacterium exhibits resistance to various conventional antibiotics, being usually challenging for eradication. Since there is an urge to consider alternative therapeutic strategies, the aim of the study was to examine selected essential oils of plants belonging to families Cupressaceae (Juniperus communis) and Lamiaceae (Hyssopus officinalis, Salvia officinalis, Melissa officinalis, Lavandula angustifolia, Ocimum basilicum and Thymus serpyllum) against H. pylori, using an improved microdilution broth method. The oils were examined in concentration range from 0.03 to 4 µL/mL. The method comprises Brain-heart infusion broth supplemented with yeast extract, horse serum and IsoVitaleX. After 3 day incubation, an equal volume of double strengthen Christensen's urea was added into each well and incubated for additional 4 h. In wells with present H. pylori, the medium changed color from yellow to purple, allowing MIC determination even without a microtitre plate reader. The microtitre format method is convenient as it is less expensive, easier to perform and requires less amount of an anti-H. pylori agent. The improved method enhances specificity to H. pylori, as fast urease activity is almost an exclusive property of this bacterium. The application of the second step incubation with Christensen's urea decreases the possibility of false positive/negative results due to contaminant growth or commonly poor H. pylori growth. Among the examined oils, J. communis, H. officinalis and O. basilicum were not active with the highest applied concentrations, while the most active was T. serpyllum, with MIC 2.0-4.0 µL/mL. This is the first report on essential oils activity of T. serpyllum and H. officinalis against H. pylori.

Phytochemical composition and in vitro functional properties of three wild rose hips and their traditional preserves.

The aim of the present study was investigation of the phenolic profile, ascorbic acid content, antioxidant, anti-acetylcholinesterase, anti-inflammatory and cytotoxic activity of rose hips and the preserves (purée and jam) of three insufficiently examined Rosa species: Rosa dumalis Bechst., R. dumetorum Thuill. and R. sempervirens L. The liquid chromatography-tandem mass spectrometry analysis resulted in quantification of 14 of the 45 phenolic compounds examined, with ellagic acid as the most dominant. Notable antioxidant activity of all three species was confirmed through several assays. Moderate inhibition of acetylcholinesterase by extracts of all investigated Rosa species was observed. Several extracts of examined Rosa species demonstrated inhibition potency towards production of some monitored eicosanoids in cyclooxygenase-1 and 12-lipoxygenase pathways. Two R. sempervirens extracts exerted cytotoxic activity against HeLa and HT-29 cell lines, but were inactive towards MRC-5 and MCF7. The results support the potential of these rose hips as food with health-promoting properties.

Chemical Composition and Immuno-Modulatory Effects of Urtica dioica L. (Stinging Nettle) Extracts.

The purpose of this work was to determine the chemical profile of stinging nettle and to provide an insight into the mechanisms by which it ameliorates the immune response. Qualitative and quantitative liquid chromatography tandem mass spectrometry analyses indicated that phenolic acids (5-O-caffeoylquinic acid as dominant) and flavonol glycosides (rutin, isoquercitrin, and kaempferol 3-O-glucoside) are present in the aerial parts, while lignans (secoisolariciresinol, 9,9'-bisacetyl-neo-olivil and their glucosides) were detected in the root. Herb and root extracts expressed selective inhibition toward cyclooxygenase and lipoxygenase branches in human platelets: root extracts were better at inhibiting thromboxane production, while herb extracts were more specific toward inhibition of 12-lipoxygenase pathway. Stinging nettle extracts mildly increased monocyte chemoattractant protein-1 and growth-related oncogene release from nonstimulated intestinal epithelial cells, stimulating MyD88/NF-κB/p38 signaling, hence preserving the epithelial integrity and enhancing intestinal steady-state defense. Additionally, root extract reduced lipopolysaccharide-induced monocyte chemoattractant protein-1/growth-related oncogene secretion and cyclooxygenase-2 expression in intestinal epithelial cells, thus showing the potential protective effect against tissue damage caused by inflammation processes. These observations suggest that stinging nettle is an interesting candidate for the development of phytopharmaceuticals or dietary supplements for cotreatment of various inflammatory diseases, particularly inflammatory bowel diseases. Copyright © 2017 John Wiley & Sons, Ltd.

Binary mixture of Satureja hortensis and Origanum vulgare subsp. hirtum essential oils: in vivo therapeutic efficiency against Helicobacter pylori infection.

BACKGROUND: Helicobacter pylori can cause many gastrointestinal and also extra-gastrointestinal disorders and is a major risk factor for gastric carcinoma and MALT lymphoma. Currently, numerous antibiotic-based therapies are available; however, these therapies have numerous drawbacks, mainly due to increasing prevalence of antibiotic resistant strains. Thus, there is an urgent need to develop novel therapeutic agents against H. pylori infections. MATERIALS AND METHODS: In this study, the anti-H. pylori activity of 2:1 mixture of Satureja hortensis and Origanum vulgare subsp. hirtum essential oils (2MIX) was investigated in vivo. After screening in vitro cytotoxicity of 2MIX on mammalian cell lines, the therapeutic efficiency was studied in a mouse model, where changes in H. pylori colonization were detected by PCR and histology of gastric samples. The immune reaction of mice was tested based on cytokine and chemokine production, and the in vivo toxicity of 2MIX was also investigated by measuring ALT and AST enzyme activities and Cyp3a11 and HO-1 mRNA levels in livers of mice. RESULTS: 2MIX had not shown in vitro cytotoxicity against cell lines, only the highest concentration caused significant decrease in their survival rates. In the in vivo experiments, 2MIX successfully eradicated the pathogen in 70% of the mice. We could not detect toxicity or altered cytokine and chemokine balance after in vivo treatments in mice. CONCLUSIONS: These results show that 2MIX is effective in reducing H. pylori colonization suggesting that this essential oil mixture has great potential as a new, effective, and safe therapeutic agent against H. pylori.

Comparative study of biological activities and phytochemical composition of two rose hips and their preserves: Rosa canina L. and Rosa arvensis Huds.

The aim of this study was to compare phenolic profile, vitamin C content, antioxidant, anti-inflammatory and cytotoxic activity of rose hips and the preserves (purée and jam) of two Rosa species: renowned Rosa canina L. and unexplored Rosa arvensis Huds. The liquid chromatography-tandem mass spectrometry analysis of 45 phenolics resulted in quantification of 14 compounds, with quercitrin, gallic and protocatechuic acids as the most dominant. High antioxidant potential of R. canina and a moderate activity of R. arvensis extracts were determined through several assays. Purée of both species and methanol extract of air-dried R. canina hips showed some anti-inflammatory (cyclooxygenase-1 and 12-lipooxygense inhibition potency) activity. Purée of R. canina exerted cytotoxic activity only against the HeLa cell line among several others (HeLa, MCF7, HT-29 and MRC-5). The presented results support traditional use of rose hips and their fruit preserves as food with health and nutritional benefits.

Antimicrobial activity of Eucalyptus camaldulensis essential oils and their interactions with conventional antimicrobial agents against multi-drug resistant Acinetobacter baumannii.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional herbal medicine has become an important issue on the global scale during the past decade. Among drugs of natural origin, special place belongs to essential oils, known as strong antimicrobial agents that can be used to combat antibiotic-resistant bacteria. Eucalyptus camaldulensis leaves are traditional herbal remedy used for various purposes, including treatment of infections. The aim of this study was to determine antimicrobial potential of two E. camaldulensis essential oils against multi-drug resistant (MDR) Acinetobacter baumannii wound isolates and to examine possible interactions of essential oils with conventional antimicrobial agents. MATERIALS AND METHODS: Chemical composition of essential oils was determined by gas chromatography-mass spectrometry analysis (GC-MS). MIC values of essential oils against A. baumannii strains were estimated by modified broth microdilution method. The components responsible for antimicrobial activity were detected by bioautographic analysis. The potential synergy between the essential oils and antibiotics (ciprofloxacin, gentamicin and polymyxin B) was examined by checkerboard method and time kill curve. RESULTS: The dominant components of both essential oils were spatulenol, cryptone, p-cimene, 1,8-cineole, terpinen-4-ol and ß-pinene. The detected MICs for the E. camaldulensis essential oils were in range from 0.5 to 2 µl mL(-1). The bioautographic assay confirmed antibacterial activity of polar terpene compounds. In combination with conventional antibiotics (ciprofloxacin, gentamicin and polymyxin B), the examined essential oils showed synergistic antibacterial effect in most of the cases, while in some even re-sensitized MDR A. baumannii strains. The synergistic interaction was confirmed by time-kill curves for E. camaldulensis essential oil and polymyxin B combination which reduced bacterial count under detection limit very fast, i.e. after 6h of incubation. CONCLUSIONS: The detected anti-A. baumannii activity of E. camaldulensis essential oils justifies traditional use of this plant. The proven E. camaldulensis essential oil synergistic interactions with conventional antibiotics could lead to the development of new treatment strategies of infections caused by MDR A. baumannii strains in the term of antibiotic dose reduction.

Binary and Tertiary Mixtures of Satureja hortensis and Origanum vulgare Essential Oils as Potent Antimicrobial Agents Against Helicobacter pylori.

Essential oils possess strong antimicrobial activity, even against multiresistant Helicobacter pylori. Available therapies against H. pylori infection have multiple disadvantages, indicating a great need for a development of new therapeutics. The purpose of this study was to develop a potent natural product based anti-H. pylori formulation. First, anti-H. pylori activity of nine essential oils was determined, after which the most active oils were mixed in various ratios for further testing. Satureja hortensis, Origanum vulgare subsp. vulgare and O. vulgare subsp. hirtum essential oils expressed the highest activity (MIC = 2 µL mL(-1)). Their binary and ternary mixtures exhibited notably higher antimicrobial activity (MIC ≤ 2 µL mL(-1)). The most active was the mixture of S. hortensis and O. vulgare subsp. hirtum oils in volume ratio 2:1, which expressed 4 times higher activity than individual oils (MIC = 0.5 µL mL(-1)). According to GC-MS, both oils in the mixture were characterized by high content of phenols (48-73%), with carvacrol as the main carrier of antimicrobial activity. Presented in vitro study pointed out binary mixture of S. hortensis and O. vulgare subsp. hirtum essential oils in volume ratio 2:1 as promising candidate for further in vivo studies targeting H. pylori infection.