Guillermo Reglero

Chemical composition and antimicrobial activity of Rosmarinus officinalis L. Essential oil obtained via supercritical fluid extraction

The chemical composition and antimicrobial activity of essential oil-rich fractions obtained by supercritical CO2 extraction from Rosmarinus officinalis L. were investigated. Gas chromatography-mass spectroscopy analysis of these fractions resulted in the identification of 33 compounds of the essential oil. The main components of these fractions were alpha-pinene, 1,8-cineole, camphor, verbenone, and borneol, constituting ca. 80% of the total oil. The antimicrobial activity was investigated by the disc diffusion and broth dilution methods against six microbial species, including gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis), gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa), a yeast (Candida albicans), and a fungus (Aspergillus niger). All of the essential oil-rich fractions obtained showed antimicrobial activity against all of the microorganisms tested, with inhibition zones and minimal bactericidal and fungicidal concentration values in the range of 17 to 33 mm and 2.25 to 0.25 mg/ml, respectively. The most active fraction was the one obtained in experiment 4 (4% ethanol as modifier; extraction pressure, 25 MPa; extraction temperature, 60 degrees C). S. aureus was found to be the most sensitive bacteria to the rosemary extracts, whereas the least susceptible was A. niger. alpha-Pinene, 1,8-cineole, camphor, verbenone, and borneol standards also showed antimicrobial activity against all the microorganisms tested, borneol being the most effective followed by camphor and verbenone. In that way, it was confirmed that essential oil from experiment 4, with the best antimicrobial activity, presented the highest quantity of camphor, borneol, and verbenone.

Liquid chromatographic-mass spectrometric analysis of supercritical-fluid extracts of rosemary plants.

A two-step supercritical fluid extraction process of rosemary leaves, on a pilot plant scale, is proposed to divide the oleoresin into two fractions with different antioxidant activities and essential oil composition. Rosemary leaves were extracted by using different conditions of pressure and temperature as well as different conditions for fractionation of the extracts. Conditions can be tuned to selectively extract one antioxidant fraction with almost no residual aroma. In the present investigation, the antioxidant fraction was exhaustively studied in terms of antioxidant activity measurements as well as of chemical composition. An LC-MS method was adapted to perform the analysis and identification of the compounds responsible for the antioxidant activity of the extracts. Different extraction and fractionation conditions were studied in order to correlate the process conditions with the antioxidant activities obtained.

Analysis of volatile fruit components by headspace solid-phase microextraction

A method based on headspace solid-phase microextraction (HS-SPME) has been developed for the analysis of volatile compounds in fruits. Conditions of sampling have been tuned. Repeatability and recoveries obtained with this method have been determined by using a mixture of typical components of fruit aroma. Several fruits, such as raspberries, strawberries, blackberries, banana and mango, have been analysed by the proposed method. The patterns obtained include compounds typically found in fresh fruits and compounds formed during processing or storage. The results showed the use of the technique for fruit characterization and its potential as a routine method for analyzing changes in key flavour compounds during different fruit processing regimes.

Isolation of essential oil from different plants and herbs by supercritical fluid extraction

Supercritical fluid extraction (SFE) is an innovative, clean and environmental friendly technology with particular interest for the extraction of essential oil from plants and herbs. Supercritical CO(2) is selective, there is no associated waste treatment of a toxic solvent, and extraction times are moderate. Further, supercritical extracts were often recognized of superior quality when compared with those produced by hydro-distillation or liquid-solid extraction. This review provides a comprehensive and updated discussion of the developments and applications of SFE in the isolation of essential oils from plant matrices. SFE is normally performed with pure CO(2) or using a cosolvent; fractionation of the extract is commonly accomplished in order to isolate the volatile oil compounds from other co-extracted substances. In this review the effect of pressure, temperature and cosolvent on the extraction and fractionation procedure is discussed. Additionally, a comparison of the extraction yield and composition of the essential oil of several plants and herbs from Lamiaceae family, namely oregano, sage, thyme, rosemary, basil, marjoram and marigold, which were produced in our supercritical pilot-plant device, is presented and discussed.

Differences among Spanish and Latin-American banana cultivars: Morphological, chemical and sensory characteristics

Physical (weight, size, shape, texture and colour), physicochemical (pH, titratable acidity, soluble solids, moisture content, total solids), chemical (soluble sugars, vitamin C, starch, pectic substances, volatile compounds) and biochemical (polyphenol oxidase and peroxidase activities, soluble proteins) characteristics and sensory attributes (appearance, flavour, odour, colour, firmness, acceptability) of banana (Musa cavendishii L.) fruits were studied in order to assess possible differences between nutritional properties and consumer acceptability of the local (Canarian) cultivars Enana and Gran Enana and the Latin-American (Colombian) Enana cultivar. Significant differences (P ≤ 0·05) were found between size and length of fruit, and between other objective measurements (lightness, yellowness, acidity, moisture content, starch, peroxidase and polyphenol oxidase activities, soluble sugars—sucrose, fructose, glucose). Also there were significant differences in vitamin C and protein content which established the higher nutritional value of the Spanish banana cultivars. The main compositional differences between the banana cultivars in terms of flavour were quantified. Purge and trap (head-space) analysis of the Spanish Enana cultivar showed it was the richest in the characteristic banana volatile aroma compounds. Sensory descriptive analysis discriminated between banana cultivars in terms of flesh colour and flesh sweetness; although panellists liked all cultivars, they preferred the Spanish Enana fruits (overall acceptability test).

Recent trends in the advanced analysis of bioactive fatty acids.

The consumption of dietary fats have been long associated to chronic diseases such as obesity, diabetes, cancer, arthritis, asthma, and cardiovascular disease; although some controversy still exists in the role of dietary fats in human health, certain fats have demonstrated their positive effect in the modulation of abnormal fatty acid and eicosanoid metabolism, both of them associated to chronic diseases. Among the different fats, some fatty acids can be used as functional ingredients such as alpha-linolenic acid (ALA), arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), gamma-linolenic acid (GLA), stearidonic acid (STA) and conjugated linoleic acid (CLA), among others. The present review is focused on recent developments in FAs analysis, covering sample preparation methods such as extraction, fractionation and derivatization as well as new advances in chromatographic methods such as GC and HPLC. Special attention is paid to trans fatty acids due its increasing interest for the food industry.

Supercritical fluid extraction of oregano (Origanum vulgare) essentials oils: Anti-inflammatory properties based on cytokine response on THP-1 macrophages

Two fractions (S1 and S2) of an oregano (Origanum vulgare) extract obtained by supercritical fluid extraction have been used to test anti-inflammatory effects on activated human THP-1 cells. The main compounds present in the supercritical extract fractions of oregano were trans-sabinene hydrate, thymol and carvacrol. Fractions toxicity was assessed using the mitochondrial-respiration-dependent 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) reduction method for several concentrations during 24 and 48 h of incubation. Concentrations higher than 30 microg/mL of both supercritical S1 and S2 oregano fractions caused a reduction in cell viability in a dose-dependent manner. Oxidized-LDLs (oxLDLs) activated THP-1 macrophages were used as cellular model of atherogenesis and the release/secretion of cytokines (TNT-alpha, IL-1beta, IL-6 and IL-10) and their respective mRNA expressions were quantified both in presence or absence of supercritical oregano extracts. The results showed a decrease in pro-inflammatory TNF-alpha, IL-1beta and IL-6 cytokines synthesis, as well as an increase in the production of anti-inflammatory cytokine IL-10. These results may suggest an anti-inflammatory effect of oregano extracts and their compounds in a cellular model of atherosclerosis.

Dunaliella salina microalga pressurized liquid extracts as potential antimicrobials

In the present work, the antimicrobial activity of different pressurized liquid extracts obtained from Dunaliella salina microalga was tested against several microorganisms of importance for the food industry (Escherichia coli, Staphylococcus aureus, Candida albicans, and Aspergillus niger). Different solvents (hexane, petroleum ether, hexane, and water) and extraction conditions (40, 100, and 160 degrees C) were tested. Results showed that the best antimicrobial activity was obtained for each solvent at the highest extraction temperature (160 degrees C). Likewise, the extraction yield followed the same trend, i.e., increasing with extraction temperature and was at a maximum when ethanol was used as an extraction solvent. Water extracts had the lowest extraction yields. In general, the best results in terms of antimicrobial activity were obtained using petroleum ether and hexane, although ethanolic extracts also showed good antimicrobial activity. Because the main antimicrobial activity of the extracts was against bacteria, the extracts can be considered to be specifically antibacterial. The extracts were analyzed by gas chromatography-mass spectrometry in order to identify the compounds responsible for activity. Fifteen different volatile compounds as well as several fatty acids (mainly palmitic, alpha-linolenic, and oleic acids) that could have been responsible for the antimicrobial activity were identified in the extracts. beta-Cyclocitral, alpha- and beta-ionone, neophytadiene, and phytol were identified among other volatile compounds; all of these compounds have previously been described as antimicrobial agents.

Antitumor effect of 5-fluorouracil is enhanced by rosemary extract in both drug sensitive and resistant colon cancer cells.

5-Fluorouracil (5-FU) is the most used chemotherapeutic agent in colorectal cancer. However, resistance to this drug is relatively frequent, and new strategies to overcome it are urgently needed. The aim of this work was to determine the antitumor properties of a supercritical fluid rosemary extract (SFRE), alone and in combination with 5-FU, as a potential adjuvant therapy useful for colon cancer patients. This extract has been recognized as a healthy component by the European Food Safety Authority (EFSA). The effects of SFRE both alone and in combination with 5-FU were evaluated in different human colon cancer cells in terms of cell viability, cytotoxicity, and cell transformation. Additionally, colon cancer cells resistant to 5-FU were used to assay the effects of SFRE on drug resistance. Finally, qRT-PCR was performed to ascertain the mechanism by which SFRE potentiates the effect of 5-FU. Our results show that SFRE displays dose-dependent antitumor activities and exerts a synergistic effect in combination with 5-FU on colon cancer cells. Furthermore, SFRE sensitizes 5-FU-resistant cells to the therapeutic activity of this drug, constituting a beneficial agent against both 5-FU sensitive and resistant tumor cells. Gene expression analysis indicates that the enhancement of the effect of 5-FU by SFRE might be explained by the downregulation of TYMS and TK1, enzymes related to 5-FU resistance.

Supercritical Carbon Dioxide Extraction of Compounds with Antimicrobial Activity from Origanum vulgare L.: Determination of Optimal Extraction Parameters

Oregano leaves were extracted using a pilot-scale supercritical fluid extraction plant under a wide range of extraction conditions, with the goal of determining the extraction and fractionation conditions to obtain extracts with optimal antimicrobial activity. In this investigation, the essential oil-rich fractions were selectively precipitated in the second separator, and their chemical composition and antimicrobial activity were investigated. Gas chromatography-mass spectrometry analysis of the various fractions resulted in the identification of 27 compounds of the essential oil. The main components of these fractions were carvacrol, trans-sabinene hydrate, cis-piperitol, borneol, terpinen-4-ol, and linalool. Antimicrobial activity was investigated by the disk diffusion and broth dilution methods against six different microbial species, including two gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis), two gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa), a yeast (Candida albicans), and a fungus (Aspergillus niger). All of the supercritical fluid extraction fractions obtained showed antimicrobial activity against all of the microorganisms tested, although the most active fraction was the one obtained in experiment 5 (fraction was obtained with 7% ethanol at 150 bar and 40 degrees C). C. albicans was the most sensitive microorganism to the oregano extracts, whereas the least susceptible was A. niger. Carvacrol, sabinene hydrate, borneol, and linalool standards also showed antimicrobial activity against all of the microorganisms tested, with carvacrol being the most effective. Consequently, it was confirmed that essential oil from experiment 5, with the best antimicrobial activity, also presented the highest quantity of carvacrol.