Andreana Marino

Extraction methods and bioautography for evaluation of medicinal plant antimicrobial activity

A comparative study on the antimicrobial properties of extracts from medicinal plants obtained by two different methods was carried out. The screening of the antimicrobial activity of extracts from six plants was conducted by a disc diffusion test against Gram‐positive, ‐negative and fungal organisms. The most active extracts (inhibition diameter ≥12 mm) were assayed for the minimum inhibitory concentration and submitted to phytochemical screening by thin‐layer chromatography and bioautography. The results obtained indicate that the diethyl ether extracts were the most efficient antimicrobial compounds. The activity was more pronounced against Gram‐positive and fungal organisms than against Gram‐negative bacteria. Bioautography showed that the antimicrobial activity was probably due to flavonoids and terpenes.

Antimicrobial activity of Mitracarpus scaber extract and isolated constituents.

G. BISIGNANO, R. SANOGO, A. MARINO, R. AQUINO, V. D’ANGELO, M.P. GERMANO★,R. DE PASQUALE & C. PIZZA.2000.The antimicrobial activity of a methanol extract and isolated constituents of Mitracarpus scaber, a species used in folk medicine by West African native people, was evaluated against Staphylococcus aureus and Candida albicans strains. The mitracarpus methanol extract possesses both antibacterial and antimycotic activities (minimum inhibitory concentration—MIC 31·25 and 62·50 μg ml−, respectively). This extract was subsequently fractioned and monitored by bioassays leading to the isolation of seven compounds screened for antibacterial and antimycotic activities. Among these compounds, gallic acid and 3,4,5‐trimethoxybenzoic acid inhibited the growth of Staph. aureus (MIC 3·90 and 0·97 μg ml−). 4‐Methoxyacetophenone and 3,4,5‐trimethoxyacetophenone effectively inhibited C. albicans (MIC 1·95 μg ml−). The other compounds (kaempferol‐3‐O‐rutinoside, rutin and psoralen) which were also isolated showed low antibacterial and antimycotic activities (125–500 μg ml−).

In vitro antimycoplasmal activity of oleuropein

The activity of oleuropein, a phenolic glycoside contained in olive oil, was investigated in vitro against Mycoplasma hominis, Mycoplasma fermentans, Mycoplasma pneumoniae and Mycoplasma pirum. Oleuropein inhibited mycoplasmas at concentrations from 20 to 320 mg/l. The MICs of oleuropein to M. pneumoniae, M. pirum, M. hominis and M. fermentans were 160, 320, 20 and 20 mg/l, respectively.

In vitro activity of carvacrol against staphylococcal preformed biofilm by liquid and vapour contact.

Carvacrol is an important component of essential oils and recently has attracted much attention as a result of its biological properties, such as a wide spectrum of antimicrobial activity. The aim of this study was to evaluate the effect of carvacrol in liquid and vapour phase on preformed biofilms of Staphylococcus aureus and Staphylococcus epidermidis by determining biofilm biomass and cultivable cell numbers, and by using epifluorescence and scanning electron microscopy. Carvacrol was able to reduce biofilm biomass and cell viability more effectively when used with liquid contact rather than with vapour phase. The efficacy of treatment with carvacrol vapour was found to be dependent on exposure time. The predominance of red fluorescence using a LIVE/DEAD BacLight Viability kit (Molecular Probes) and the partially destroyed biofilm architecture as determined by microscopy in treated samples provided evidence for the efficacy of carvacrol. The findings of this investigation suggest a potential application for carvacrol in the inactivation of staphylococcal biofilms.

Comparative analysis of flavonoid profile, antioxidant and antimicrobial activity of the berries of Juniperus communis L. var. communis and Juniperus communis L. var. saxatilis Pall. from Turkey.

The present study was designed to define and compare the flavonoid composition and the biological potential of berries methanol extracts of Juniperus communis L. var. communis (Jcc) and Juniperus communis L. var. saxatilis. Pall. (Jcs) from Turkey. Total polyphenols (Folin-Ciocalteau method) were 3-fold higher in Jcc (59.17 +/- 1.65 mg GAE/g extract) than in Jcs (17.64 +/- 0.09 mg GAE/g extract). Flavonoid and biflavonoid content, evaluated by HPLC-DAD-ESI-MS analysis, was higher in Jcc (25947 +/- 0.86 and 4346 +/- 3.95 microg/g extract) than in Jcs (5387 +/- 34.88 and 1944 +/- 26.88 microg/g extract). The HPLC analysis of Jcc allowed the separation of 16 flavonoids; hypolaetin-7-pentoside and quercetin-hexoside are the main compounds. Moreover, gossypetin-hexoside-pentoside and gossypetin-hexoside were identified for the first time in Jcc berries. In Jcs eight flavonoids were identified: quercetin-hexoside and isoscutellarein-8-O-hexoside are the most abundant compounds. The in vitro antioxidant activity was determined using different methods; Jcc was found to be more active than Jcs in the DPPH test (IC(50) of 0.63 +/- 0.09 mg/mL and 1.84 +/- 0.10 mg/mL) in reducing power assay (12.82 +/- 0.10 ASE/mL and 64.14 +/- 1.20 ASE/mL), and in TBA assay (IC(50) of 4.44 +/- 0.70 microg/mL and 120.07 +/- 3.60 microg/mL). By contrast, Jcs exhibited more elevated Fe(2+) chelating ability than Jcc. The extracts were also studied for their antimicrobial potential, displaying antimicrobial capacity only against Gram-positive bacteria.

Control of biofilm formation by poly-ethylene-co-vinyl acetate films incorporating nisin

The aim of this study was to evaluate the effect of poly-ethylene-co-vinyl acetate (EVA) films incorporating different concentrations (0.1%, 0.5% and 1%) of nisin on the biofilm-forming ability of Listeria monocytogenes ATCC 7644, Staphylococcus aureus 815 and Staphylococcus epidermidis ATCC 35984. Nisin was incorporated into two grades of EVA (EVA14 and EVA28) in the melt during a common film-blowing operation. The efficacy of EVA/nisin films was evaluated by biofilm biomass measurements and Live/Dead staining in combination with fluorescence microscopy. In order to evaluate whether the nisin incorporation could modify the film surface properties, contact angle measurements and scanning electron microscopy were performed. The results revealed the efficacy of EVA14/nisin films in reducing biofilm formation on their surfaces with more evident effect for S. epidermidis than L. monocytogenes and S. aureus strains. In contrast, EVA28/nisin films showed unsatisfactory activity. Fluorescence microscopy confirmed poor biofilm formation on EVA14/nisin films, also characterised by the presence of dead cells. The data presented in this study offer new potential applications for developing strategies aimed to improve the effect of antimicrobial agents.

Preservative properties of Calamintha officinalis essential oil with and without EDTA.

Aims: This study was focused on the preserving properties of Calamintha officinalis essential oil, a plant known for its diaphoretic, expectorant and aromatic properties.

Effect of alkaline pH on staphylococcal biofilm formation

Biofilms are a serious problem, cause of severe inconvenience in the biomedical, food and industrial environment. Staphylococcus aureus and S. epidermidis are important pathogenic bacteria able to form thick and resistant biofilms on various surfaces. Therefore, strategies aimed at preventing or at least interfering with the initial adhesion and subsequent biofilm formation are a considerable achievement. The aim of this study was to evaluate the effect of alkaline pH on bacterial adhesion and further biofilm formation of S. aureus and S. epidermidis strains by biofilm biomass, cell‐surface hydrophobicity, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) analysis. The results demonstrated that the amount of biofilm biomass formed and the surface hydrophobicity were significantly less than what were observed at higher levels of pH. SEM and CLSM images revealed a poorly structured and very thin biofilm (2.5–3 times thinner than that of the controls). The inhibiting effect of the alkaline pH on the bacterial attachment impaired the normal development of biofilm that arrested at the microcolony stage. Alkaline formulations could be promising towards the control of bacterial colonization and therefore the reduction of the biofilm‐related hazard. In the clinical setting, alkaline solutions or cleaners could be promising to prevent the bacterial colonization, by treating surfaces such as catheters or indwelling medical devices, reducing the risk of biofilm related infections.

Phenolic composition and biological activities of Juniperus drupacea Labill. berries from Turkey

The present study was designed to define the phenolic profile and the biological potential of berries methanol extract of Juniperus drupacea Labill. from Turkey. The total phenolic content (Folin-Ciocalteau assay) was 48.06±0.99mgGAE/g extract. The HPLC-DAD-ESI-MS analysis allowed the determination of the complete phenolic profile of J. drupacea berries. Phenolic acids represented more than 60% of the total phenolics, and tyrosol was the major one (1324±0.64μg/g extract); within the flavonoids amentoflavone was detected as the main constituent (927±0.35μg/g extract). The extract exhibited good antioxidant properties, as determined by different in vitro models: DPPH test (IC(50) 0.38±0.02mg/mL), reducing power (12.63±0.14ASE/mL), Fe(2+) chelating ability (IC(50) 2.26±0.06mg/mL), and TBA test (IC(50) 2.47±1.13μg/mL). Cytotoxicity against Artemia salina was highlighted (LC(50) 489.47±27.8μg/mL), and a significant decrease (p⩽0.05; p⩽0.01) in HepG2 cells viability was observed at the higher concentrations (5-10μg/mL). The extract displayed good antibacterial activity towards Gram-positive bacteria and in particular Staphylococcus aureus was the most susceptible strain (MIC 78.12μg/mL).

Ochratoxin A production by Aspergillus westerdijkiae in orange fruit and juice.

To evaluate the incidence of fungi producing ochratoxin A in orange fruit and juice a survey was carried out by challenge test with ochratoxin A producing strains of Aspergillus westerdijkiae used as the model system. A. westerdijkiae grew into experimental lesions caused on orange surfaces, in orange natural medium and in orange juice and produced OTA in all environments. The higher temperature of 26 degrees C had a greater influence than temperatures of 20 degrees C and 4 degrees C on the growth of the mould, and OTA production. The environmental temperature of 20 degrees C, slows down but does not prevent toxin production. At the lowest temperature of 4 degrees C no growth was observed.