In vitro, in vivo and in silico evaluation of the anti-inflammatory potential of Hyssopus officinalis L. subsp. aristatus (Godr.) Nyman (Lamiaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal properties of hyssop have been used in traditional medicine since ancient times, inter alia, in diseases/conditions with an inherent inflammatory process. AIM OF THE STUDY: Accordingly, the aim of this study was to investigate the anti-inflammatory properties of hyssop herb preparations (essential oil and methanol extracts) in vivo, in vitro and in silico. MATERIALS AND METHODS: For in vitro testing of essential oils and extracts of hyssop herb, the cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) enzyme assays were used. In vivo anti-inflammatory potential of the extracts (at doses of 50, 100 and 200 mg/kg) was assessed using the carrageenan-induced rat paw edema test. Molecular docking and dynamics were used for in silico testing of the inhibitory activity of chlorogenic (CA) and rosmarinic (RA) acids, as the dominant compounds in the tested methanol extracts against COX-1 and COX-2 enzymes. RESULTS: Significant inhibitory activity was shown in the COX-2 test regarding extracts (essential oils did not exhibit any significant activity). Namely, all analyzed extracts, at a concentration of 20 µg/mL, showed a percentage of inhibition of COX-2 enzyme (54.04-63.04%), which did not indicate a statistically significant difference from the positive control of celecoxib (61.60%) at a concentration of 8.8 µM. In vivo testing showed that all methanol extracts of hyssop herb, at the highest test dose of 200 mg/kg in the third and fourth hours, after carrageenan administration, exhibited a statistically significant (p < 0.05) inhibitory effect on the increase in rat paw edema in relation to control. This activity is comparable or higher in relation to the reference substance, indomethacin, at a concentration of 8 mg/kg. The preliminary in silico results suggest that investigated compounds (RA and CA) showed better inhibitory activity against COX-1 and COX-2 than standard non-steroidal anti-inflammatory drug (NSAID), ibuprofen, as evident from the free binding energy (ΔGbind in kJ mol-1). The binding energies of the docked compounds to COX-1 and -2 were found to be in the range between -47.4 and -49.2 kJ mol-1. Ibuprofen, as the one NSAID, for the same receptors targets, showed remarkably higher binding energy (ΔGbind = -31.3 kJ mol-1 to COX-1, and ΔGbind = -30.9 kJ mol-1 to COX-2). CONCLUSION: The results obtained not only support the traditional use of hyssop herb in inflammatory conditions in folk medicine, but also open the door to and the need for further in vivo testing of extracts in order to examine the molecular mechanism of anti-inflammatory activity in living systems and possibly develop a new anti-inflammatory drug or supplement.

Antioxidant, Antigenotoxic and Cytotoxic Activity of Essential Oils and Methanol Extracts of Hyssopus officinalis L. Subsp. aristatus (Godr.) Nyman (Lamiaceae).

Hyssopus officinalis L. is a well-known aromatic plant used in traditional medicine and the food and cosmetics industry. The aim of this study is to assess the antioxidant, genotoxic, antigenotoxic and cytotoxic properties of characterized hyssop essential oils and methanol extracts. Chemical composition was analyzed by gas chromatography - mass spectrometry (GC-MS) and liquid chromatography with diode array detection and mass spectrometry (LC-DAD-MS), respectively. Antioxidant activity was examined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing/antioxidant power (FRAP) tests; genotoxic and antigenotoxic activity were examined by the comet assay, while cytotoxicity was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide dye (MTT) test against tumor cell lines (SW480, MDA-MB 231, HeLa) and non-transformed human lung fibroblast cell lines (MRC-5). The essential oils were rich in monoterpene hydrocarbons (e.g., limonene; 7.99-23.81%), oxygenated monoterpenes (1,8-cineole; 38.19-67.1%) and phenylpropanoids (methyl eugenol; 0.00-28.33%). In methanol extracts, the most abundant phenolics were chlorogenic and rosmarinic acid (23.35-33.46 and 3.53-17.98 mg/g, respectively). Methanol extracts expressed moderate to weak antioxidant activity (DPPH IC50 = 56.04-199.89 µg/mL, FRAP = 0.667-0.959 mmol Fe2+/g). Hyssop preparations significantly reduced DNA damage in human whole blood cells, induced by pretreatment with hydrogen peroxide. Methanol extracts exhibited selective and potent dose- and time-dependent activity against the HeLa cell line. Results of the current study demonstrated notable H. officinalis medicinal potential, which calls for further investigation.

Phytochemical Properties of Satureja kitaibelii, Potential Natural Antioxidants: a New Insight.

Satureja kitaibelii Wierzb. ex Heuff. has a great importance in Serbian ethnopharmacology/herbal traditional medicine, as well as a flavoring food additive. Ethanol extract of aerial parts of Satureja kitaibelii analyzed by liquid chromatography-mass spectrometry revealed the presence of 18 compounds among which the most abundant were phenolic acids, flavonoids, jasmonic acid derivatives and rosmanol. The extracts were rich in total phenolics and flavonoid contents, while rosmarinic acid was the dominant compound (18.30-29.52 mg/g). As assessments of antioxidant properties of natural extracts are important because of their growing use in medicine and food industry, antioxidant activity of ethanol extracts of Satureja kitaibelii was analyzed by several assays. The half maximal scavenging capacity (SC50) of 2,2'-diphenyl-1-picrylhydrazyl ranging from 71.20 to 125.65 µg/mL, the total antioxidant capacity from 272.37 to 714.12 mg ascorbic acid/g, and ferric ion reducing antioxidant power ranging from 0.74 to 1.94 µmol Fe/mg, clearly imply a significant antioxidant potential of Satureja kitaibelii. The extracts inhibit growth of Micrococcus luteus and Pseudomonas aeruginosa with inhibition zones 20-30 and 16-26 mm, respectively. Antioxidant and antibacterial activity of compounds identified in extracts suggest a great potential for Satureja kitaibelii application as valuable food ingredient.

Antioxidant, anti-inflammatory and gastroprotective activity of Filipendula ulmaria (L.) Maxim. and Filipendula vulgaris Moench.

ETHNOPHARMACOLOGICAL RELEVANCE: Meadowsweet (Filipendula ulmaria (L.) Maxim.) and dropwort (Filipendula vulgaris Moench) are herbaceous perennials employed in folk medicine for their antirheumatic, antipyretic and anti-ulcer properties. AIM OF THE STUDY: To assess ethnomedicinal claims through investigation of antioxidant, anti-inflammatory and gastroprotective effects of F. ulmaria and F. vulgaris lyophilized flower infusions (LFIs) as well as the F. vulgaris isolated flavonoids spiraeoside, kaempferol 4'-O-glucoside, astragalin 2'-O-gallate, mixture of hyperoside 2'-O-gallate and isoquercitrin 2'-O-gallate, and a tannin tellimagrandin II. MATERIALS AND METHODS: Free radical scavenging activity of the tested samples was determined by examining their ability to neutralize DPPH and OH radicals in vitro, whereas reducing properties were assessed in Ferric Reducing Antioxidant Power (FRAP) assay. Anti-inflammatory activity was studied ex vivo in human platelets by monitoring the effect on eicosanoid biosynthesis. Gastroprotective action was estimated in animal model of acute gastric injury induced by ethanol. RESULTS: LFIs and spiraeoside exerted activities comparable to those of positive control in DPPH-radical scavenging and FRAP antioxidant assays, whereas notable hydroxyl radical scavenging ability was demonstrated only for spiraeoside (IC50 = 5.1µg/mL). Among tested samples, astragalin 2″-O-gallate (IC50 = 141.1µg/mL) and spiraeoside (IC50 = 4.69µg/mL) the most markedly inhibited production of pro-inflammatory prostaglandin E2 and 12(S)-hydroxy-(5Z,8Z,10E,14Z)-eicosatetraenoic acid in human platelets, respectively. Examination of LFIs (100-300mg/kg, p.o.) gastroprotective action in rats revealed their capacity to preserve mucosal integrity. In addition, spiraeoside (50mg/kg, p.o.) and tellimagrandin II (40mg/kg, p.o.) showed ulcer preventive ability. CONCLUSION: Current study supports documented traditional use of investigated herbs and indicates that flavonoid and tannin components are partially responsible for the demonstrated pharmacological activities.

Antihyperalgesic activity of Filipendula ulmaria (L.) Maxim. and Filipendula vulgaris Moench in a rat model of inflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: Meadowsweet (Filipendula ulmaria (L.) Maxim.), and dropwort (Filipendula vulgaris Moench) flowers are traditionally used to treat various ailments, including inflammatory conditions. The aim of the present study was to validate the aforementioned ethnomedicinal claim by assessing antihyperalgesic and antiedematous activities and toxicity of orally administered lyophilized flower infusions (LFIs) of F. ulmaria and F. vulgaris in experimental animals. MATERIALS AND METHODS: The phytochemical analysis of LFIs was performed by HPLC-DAD. Antihyperalgesic and antiedematous activities were estimated in a rat model of inflammation induced by intraplantar injection of carrageenan using Von Frey anesthesiometer and plethysmometer, respectively. Moreover, acute oral toxicity of LFIs in mice was evaluated by observing changes in animal behavior and mortality for a period of 14 days following the treatment. RESULTS: HPLC-DAD analysis revealed the presence of phenolic acids and flavonoids in LFIs, among which spiraeoside was identified as the principal component (56.27±1.03 and 55.67±1.82mg/g of LFI in F. ulmaria and F. vulgaris, respectively). The LFIs of F. ulmaria and F. vulgaris (100-300mg/kg; p.o.) produced significant and dose-dependent antihyperalgesic effects: ED50±SEM values were 164.8±15.4mg/kg (110.3-246.3mg/kg) and 172.2±6.2mg/kg (147.4-201.3mg/kg) for F. ulmaria and F. vulgaris, respectively. On the other hand, LFIs of both species (100-300mg/kg; p.o.) did not significantly reduce edema. Good safety profiles were evidenced in the toxicological study. The median lethal dose (LD50) of the tested extracts is likely to be greater than 2000mg/kg. CONCLUSION: The results of the present study support the use of F. ulmaria and F. vulgaris flowers in folk medicine for relieving pain in diseases with an inflammatory component.

Antibacterial activity of Veronica montana L. extract and of protocatechuic acid incorporated in a food system.

This study was designed to evaluate the antibacterial activity of the Veronica montana L. water extract and its main phenolic compound, protocatechuic acid. The antibacterial activity was determined by microdilution assay against six strains of Gram-positive and Gram-negative bacteria. Listeria monocytogenes was the most sensitive of the tested bacterial species. Antibacterial preserving properties of protocatechuic acid were also evaluated after its incorporation in cream cheese, using L. monocytogenes as commonly cheese contaminant. The compound successfully inhibited L. monocytogenes development in cream cheese, at room temperature and in refrigerator (25°C and 4°C, respectively), after 3days of inoculation. Sensory evaluation was carried out in order to validate the mentioned food system. A possible mode of action of the tested compound towards bacterial cells was assessed and appears to be direct lysis of pathogenic cytoplasmic membrane. Prediction of pharmacokinetic properties was also performed using computational analyzes. The obtained results can serve as an important platform for the development of effective natural preservatives.

Hepatoprotective effect of Filipendula hexapetala Gilib. (Rosaceae) in carbon tetrachloride-induced hepatotoxicity in rats.

The influence of methanol extract produced from the flowers of Filipendula hexapetala Gilib. (Rosaceae) on some liver biochemical parameters in rats intoxicated with carbon tetrachloride (CTC) was evaluated in this study. The results Obtained revealed that pretreatment with the extract investigated inhibited CTC-induced liver injury by decreasing lipid peroxidation and increasing the content of reduced glutathione in a dosage dependent manner, bringing the levels of all antioxidant enzymes close to control values. The administration of CTC diminished hepatic antioxidant defense mechanisms by significant reduction of peroxidase and catalase activities. The catalase activity was significantly recovered in groups treated with the extract investigated and intoxicated with a single CTC dose. A similar impact on hepatic peroxidase activity has also been observed, indicating a partial detoxication of hydrogen peroxide by both catalase and peroxidase.

Antioxidant activity of yellow dock (Rumex crispus L., Polygonaceae) fruit extract.

The methanol extract of ripe Rumex crispus L. fruits was evaluated for its antioxidant potential by assays for ferric-reducing antioxidant power (FRAP), DPPH-free radical scavenging activity (DPPH) and the influence on lipid peroxidation in liposomes (LP). Considerable activity was observed in all test systems (FRAP: 9.9 mmol Fe(2+) /g; DPPH IC(50) : 3.7 µg/mL; LP IC(50) : 4.9 µg/mL), comparable to that of BHT (FRAP: 8.0 µg/mL; DPPH IC(50) : 19.4 µg/mL; LP IC(50) : 3.5 µg/mL), but lower than the activity of ascorbic acid, rutin and quercetin, used as positive control substances. The in vivo effects were evaluated in several hepatic antioxidant systems (activities of LPx, GSH-Px, Px, CAT and XOD, as well as GSH content), after treatment with the studied yellow dock extract in different doses, or in combination with carbon tetrachloride (CCl(4) ). Pretreatment with the R. crispus extract inhibited CCl(4) -induced oxidative stress by decreasing LPx and increasing GSH content in a dose dependent manner, bringing the levels of antioxidant enzymes to near control values.

Composition and antimicrobial activity of Marrubium incanum Desr. (Lamiaceae) essential oil.

The essential oil from the aerial parts of Marrubium incanum Desr. (Lamiaceae), obtained by hydrodistillation, was analyzed by GC and GC-MS. Forty-six compounds were identified, representing 96.3% of the total oil. The main components of the oil were (E)-caryophyllene (27.0%), germacrene D (26.2%) and bicyclogermacrene (11.5%). The microbial growth inhibitory properties of the isolated essential oil were determined using the agar diffusion and broth microdilution method against seven bacterial species (Staphylococcus aureus ATCC 25923, S. epidermidis ATCC 12228, Micrococcus flavus ATCC 10240, Enterococcus faecalis ATCC 29212, Escherichia coli ATCC 25922, Klebsiella pneumoniae NCIMB 9111, and Pseudomonas aeruginosa ATCC 27853), and two strains of the yeast Candida albicans (ATCC 10259 and ATCC24433). The essential oil showed activity against all the microorganisms tested, but differences in microbial susceptibility were registered.

Polyphenol contents and antioxidant activity of soybean seed extracts.

The antioxidant activity and contents of various polyphenol classes in the seeds of 20 soybean hybrids were evaluated. Total polyphenols, tannins and proanthocyanidins were determined after extraction of plant material with 70% aqueous acetone. In addition, flavonoid content was determined. Antioxidant activity of seed extracts was evaluated by DPPH free radical scavenging activity assay. A positive linear correlation between antioxidant activity and contents of total phenols, tannins and proanthocyanidins was established. The highest antioxidant activity was observed in the extracts of hybrids which have higher levels of all polyphenol classes examined. The most of the single-cross hybrids were poor in tannins which recommend them as good source for ensiled livestock feed. Results suggested that polyphenol content should be considered as an important feature of the soybean seed.

Antioxidant activity of Filipendula hexapetala flowers.

The antioxidant activity of the methanol extract of Filipendula hexapetala flowers was assessed by the assay for ferric-reducing antioxidant power (FRAP), the assay for DPPH free radical scavenging ability (DPPH) and the assay for the influence of lipid peroxidation in liposomes, induced by Fe(2+)/ascorbate system and measured by the TBA test (LP). The activity of the investigated extract in all test-systems was found to be significant. The principal constituent responsible for the observed effects was isolated and identified as spiraeoside.

Polyphenol contents and antioxidant activity of Maydis stigma extracts.

The antioxidant activity and contents of various polyphenol classes in the silks of fifteen maize hybrids with economic importance in Serbia were evaluated. Total polyphenols, tannins and proanthocyanidins were determined spectrophotometrically, after extraction of plant material with 70% aqueous acetone under sonication at room temperature. In addition, flavonoid content was determined. Antioxidant activity of aqueous acetone extracts was evaluated by FRAP assay. A positive linear correlation between antioxidant activity and contents of all investigated polyphenol classes was established. The highest antioxidant activity was observed in the extract of NS 640 hybrid, which had high levels of all polyphenol classes examined. Results suggested strongly that polyphenol content should be considered as an important feature of the herbal drug Maydis stigma. For that reason, the biological source of this herbal drug needs to be more precisely defined, as observed activities and polyphenol contents were greatly dependent on plant material source.

Antimicrobial activity of Chenopodium botrys essential oil.

Essential oil, isolated by hydrodistillation from Chenopodium botrys aerial parts, expressed significant bactericidal and fungicidal activity against selected strains of microorganisms.