Application of response surface methodology and central composite design for the optimisation of supercritical fluid extraction of essential oils from Myrtus communis L. leaves

Essential oils of Myrtus communis L. leaves were obtained using supercritical fluid extraction (SFE) and hydrodistillation methods. The experimental parameters of SFE such as pressure, temperature, modifier volume, static and dynamic extraction time were optimised using a central composite design after a 2ⁿ⁻¹ fractional factorial design. The chemical compositions of the SFE extract were identified by GC–MS and determined by GC–FID. The major components of essential oils obtained by hydrodistillation were α-pinene (31.8%), 1,8-cineole (24.6%), limonene (14.8%), linalool (8.3%) and α-terpinolene (4.8%). However, by using the supercritical carbon dioxide in optimum conditions, only three components represented more than 85% of the extract. Therefore, by using the proper SFE conditions, the supercritical extraction is more selective than the conventional hydrodistillation methods. The oil yields based on the hydrodistillation was 0.47% (v/w). Extraction yields based on the SFE varied in the range of 0.5–6.3% (w/w) under different conditions.     

Comparison of essential oil compositions of Salvia mirzayanii obtained by supercritical carbon dioxide extraction and hydrodistillation methods

Essential oil of Salvia mirzayanii cultivated in Iran was obtained by hydrodistillation and supercritical (carbon dioxide) extraction methods. The oil was analysed by capillary gas chromatography using flame ionization and mass spectrometric detections. The compounds were identified according to their retention indices and mass spectra (EI, 70 eV). The effects of different parameters such as pressure, temperature, modifier volume and extraction times (dynamic and static) on the supercritical fluid extraction (SFE) of S. mirzayanii oil were investigated. The results showed that, under a pressure of 35.5 MPa, temperature of 35 °C, 6% methanol, dynamic extraction time of 50 min and static extraction time of 30 min, extraction was more selective for the linalyl acetate. Thirty four compounds were identified in the hydrodistilled oil. The major components of S. mirzayanii were linalyl acetate (7.6%), 1,8-cineole (8.0%), linalool (9.0%) and 8-acetoxy linalool (11.0%). However, by using supercritical carbon dioxide in optimum conditions, only three components contain more than 63% of the oil. The yield of the obtained oil based on hydrodistillation was 2.20% (v/w). Extraction yield based on the SFE varied in the range of 1.50–9.67% (w/w) under different conditions. The results revealed that, in Iranian S. mirzayanii oil, linalyl acetate is a major component.     

Volatile compounds of leaves and fruits of Mangifera indica var. coquinho (Anacardiaceae) obtained using solid phase microextraction and hydrodistillation

Volatile compounds present on fruits and leaves of Mangifera indica var. coquinho were investigated by headspace solid phase microextraction (HS-SPME) and hydrodistillation (HD). Conventional techniques, such as hydrodistillation, may impart chemical changes to the original oil composition being also time-consuming. On the other hand, HS-SPME provides solvent-less extractions, shorter extraction times and may supply complementary information about the composition of the compounds. The HS-SPME technique was previously evaluated by the comparative study among the fibres: commercial PDMS, NiTi-ZrO₂ and NiTi-ZrO₂-PDMS. The fibre NiTi-ZrO₂-PDMS showed better sensitivity and precision and was used on the extraction of components. The influence of several parameters like the time and temperature of extraction and desorption time were examined to obtain better efficiency. Fruits and leaves were analysed in mature and immature stages. The profiles corresponding to the volatile compounds detected by both techniques are discussed.     

Extraction of Cuminum cyminum essential oil by combination technology of organic solvent with low boiling point and steam distillation

Extraction of essential oil from Cuminum cyminum seeds using a combination of organic solvent with low boiling point and steam distillation was explored. The effect of different parameters, such as particle size (40, 60, 80 mesh), temperature (°C) 10, 15, 20 and extraction time (3, 5, 8 h), on the extraction yield was investigated using three-level orthogonal array design. The experimental results showed that the temperature had the largest effect on the yield of the extract (oleoresin), followed by extraction time and particle size. The optimum parameters, such as temperature, particle size, and extraction time were in turn 20 °C, 80 mesh, and 8 h. Essential oil of C. cyminum seeds obtained by supercritical fluid extraction (SFE), hydrodistillation (HD), combination technology of organic solvent with low boiling point and steam distillation (OS-SD) were further analysed by gas chromatographic/mass spectrometric detection to compare the extraction methods. Forty-five compounds in the C. cyminum essential oil were identified, showing that the composition of the extraction by different methods was mostly similar, whereas relative concentration of the identified compounds was apparently different. General characteristics of the C. cyminum essential oil obtained by different methods were further compared, and OS-SD was considered as the optimum process among the three processes to obtain C. cyminum essential oil for high quality, simple technology and low cost.     

Supercriticial fluid extraction of flavors and fragrances from Hyssopus officinalis L. cultivated in Iran

Hyssopus officinalis L. (hyssop) as a food ingredient has its own importance in flavor industry and also in sauce formulations. Supercritical fluid extraction (SFE) of hyssop, cultivated in Iran, was performed at various pressures, temperatures, extraction (dynamic and static) times and modifier (methanol) concentrations using an orthogonal array design with an OA₂₅(5⁵) matrix conditions. Pressure, temperature and modifier in the SFE system influenced the extraction yield. Also, the composition of the extracted oils was greatly impacted by the operating conditions. Main components of the extracts under different SFE conditions were sabinene (4.2–17.1%, w/w), iso-pinocamphene (0.9–16.5%) and pinocamphene (0.7–13.6%). The extraction of sabinene, for example, was favored at 100 atm, 55 °C, 1.5% (v/v) methanol, 30 min dynamic time and 35 min static time. Use of SFE under different conditions can allow targeting the extraction of different constituents.     

Optimization of subcritical fluid extraction of seed oil from Nitraria tangutorum using response surface methodology

Subcritical fluid extraction (SFE) technology was used to extract oil from Nitraria tangutorum seed. The best possible combination of extraction parameters was found using response surface methodology (RSM) in a three-variable, three-level Box-Behnken experimental design (BBD). The optimum extraction parameters were an extraction time of 40 min, an extraction pressure of 0.60 MPa, an extraction temperature of 44 °C and a raw material particle size of 0.45 mm. Conventional solvent extraction and supercritical CO₂ fluid extraction were comparatively used. The yield of seed oil obtained using SFE was 12.92%, which was similar to or higher than the other methods. The chemical compositions of the seed oil, determined by GC–MS, indicate that its unsaturated fatty acids content was 97%. SFE proved to be an effective technique for extracting oil from N. tangutorum.     

Essential oil composition and antimicrobial activity of Origanum majorana L. extracts obtained with ethyl alcohol and supercritical carbon dioxide

Volatile components of marjoram (Origanum majorana L.) essential oil obtained by hydrodistillation and extracts obtained by solvent extraction with ethyl alcohol and supercritical fluid extraction (SFE) were investigated. The compositions of volatile compounds in essential oil, ethanolic and SFE extracts were determined by GC and GC–MS. The antimicrobial properties of marjoram solvent extracts were investigated with microbiological tests against food borne fungi and bacteria strains. Extracts obtained by SFE at high pressure and temperature showed significantly stronger antimicrobial properties in comparison to the slight inhibitory effects of the ethanolic extract. The results support the notion that extracts obtained by SFE might have a role as flavourings and natural colourants as well as use as preservatives in food and cosmetic systems.     

In vitro antioxidant activities of Rosmarinus officinalis extracts treated with supercritical carbon dioxide

The leaves of Rosmarinus officinalis (rosemary) were subjected to supercritical CO₂ extraction (SFE). Different sources of variability, including location (Izmir, Canakkale and Mersin) and harvesting time (December, March, June and September), were considered. Among active constituents of rosemary, carnosic acid, carnosol and rosmarinic acid were analyzed by HPLC. Variability of the amounts of active constituents appears to be due to different geographical locations of growth and seasonal variations. The levels of the constituents were higher in the months of December 2003 and September 2004. In addition to this, 12 SFE extracts were screened for their radical-scavenging capacities and antioxidant activities by various in vitro assays, namely total phenol assay, DPPH radical-scavenging activity and trolox equivalent antioxidant capacity (TEAC).     

Supercritical fluid extraction and hydrodistillation for the recovery of bioactive compounds from Lavandula viridis L’Hér

The chemical profiles of bioactive essential oil and extracts obtained by hydrodistillation (HD) and supercritical fluid extraction (SFE), respectively, from Lavandula viridis were compared. The SFE was performed at 40 °C and at extraction pressures of 12 or 18 MPa in two different separators. Evaluation of the essential oil and SFE extracts by GC–FID and GC–IT–MS revealed that oxygen-containing monoterpenes were the major constituents in both cases, but there were important differences between the chemical profiles produced by the different extraction techniques. More compounds were isolated by HD but higher yields were achieved by SFE. Camphor was the main component identified in the essential oil (31.59 ± 1.32%), and in extracts from the first (1.61 ± 0.34%) and second SFE separators (22.48 ± 1.49%) at 12 MPa. In contrast, the first separator SFE extract at 18 MPa (heavy compounds) was dominated by myrtenol (5.38 ± 2.04%) and camphor (4.81 ± 1.93%), whereas the second separator SFE extract (volatiles) was dominated by verbenone (13.97 ± 5.27%). The essential oil and heavy compound extracts from the first separator possessed antioxidant and anti-cholinesterase activities. Our data show that phytochemicals from the aerial parts of L. viridis could be developed as natural antioxidant and anti-cholinesterase drugs, with particular applications in the symptomatic treatment of Alzheimer’s disease.     

Antioxidant properties of Agaricus blazei, Agrocybe cylindracea, and Boletus edulis

Three mushrooms are currently available in Taiwan, including Agaricus blazei, Agrocybe cylindracea, and Boletus edulis. Their ethanolic and hot water extracts were prepared and antioxidant properties studied. Ethanolic extracts from three mushrooms were more effective than hot water extracts in antioxidant activity using the conjugated diene method and scavenging ability on 1,1-diphenyl-2-picrylhydrazyl radicals whereas hot water extracts were more effective in reducing power, scavenging ability on hydroxyl radials and chelating ability on ferrous ions as evidenced by their lower EC₅₀ values. Overall, for both extracts, B. edulis was more effective among antioxidant properties assayed. Naturally occurring antioxidant components including total tocopherols (3.18-6.18 mg/g) and total phenols (5.67-5.81 mg/g) were found in the extracts and their contents were associated (r=0.636-0.907) with EC₅₀ value of antioxidant properties. Based on the results obtained, both extracts from these three mushrooms were effective in antioxidant properties.     

Comparative chemical composition,antioxidant and hypoglycaemic activities of Juniperus oxycedrus ssp. oxycedrus L. berry and wood oils from Lebanon

Juniper (Juniperus oxycedrus) is used in European cuisine for its distinguishing flavour. J. oxycedrus ssp. oxycedrus berry and wood essential oils were tentatively identified by GC and GC/MS. Fifty compounds were identified in the berry oil and 23 compounds were identified in the wood oil. The J. oxycedrus ssp. oxycedrus berry oil was characterised by high contents of α-pinene (27.4%) and β-myrcene (18.9%). Other important compounds were α-phellandrene (7.1%), limonene (6.7%), epi-bicyclosesquiphellandrene (2.3%) and δ-cadinene (2.2%) while, in the wood oil, δ-cadinene (14.5%) is a major main component, together with cis-thujopsene (9.2%) and α-muurolene (4.9%). In vitro evaluation of antioxidant activity by the DPPH method showed a significant activity for both oils with IC₅₀ values of 1.45 μl/ml for wood and 7.42 μl/ml for berries. Hypoglycaemic activity was investigated through the inhibition of α-amylase. The results revealed that oil obtained by hydrodistillation from J. oxycedrus ssp. oxycedrus wood exhibits an interesting activity with IC₅₀ of 3.49 μl/ml.     

Comparison of HS-SPME with hydrodistillation and SFE for the analysis of the volatile compounds of Zisu and Baisu,two varietal species of Perilla frutescens of Chinese origin

Content and composition of the volatiles of Baisu and Zisu, two varieties of Perilla frutescens, obtained by SFE, HS-SPME, and hydrodistillation, were compared. A total of 64 compounds were identified by GC/MS. Perillaldehyde (21.9–58.1%) and perilla ketone (54.5–83.5%) were the main volatile components of Zisu and Baisu, respectively. They might be partly responsible for the aroma and taste of perilla. The abundance of perilla ketone indicates that it is not safe to use Baisu as vegetable, and Baisu should not be used as substitute for Zisu. SFE extracted oil with high yield, and extracted some high molecular-weight compounds that did not contribute to the aroma. The HS-SPME was simple and sensitive for the analysis of volatile compounds of fresh perilla. The results showed that the drying process influenced the volatile components of perilla.     

Central composite design for the optimization of supercritical carbon dioxide fluid extraction of fatty acids from Borago officinalis L. flower

Abstract: In the present study, fatty acids and essential oils of the flower of borage (Borago officinalis L.) were obtained by supercritical carbon dioxide fluid extraction under different conditions. The extracts obtained were compared to oils of borage flower oil isolated by hydrodistillation. The obtained oils were analyzed by gas chromatography mass spectrometry. The compounds were identified according to their retention indices and mass spectra. The experimental parameters of supercritical fluid extraction (SFE) were optimized using a central composite design after a full factorial experimental design. Extraction yields based on SFE varied in the range of 0.02% to 1.96% (w/w), and the oil yield based on the hydrodistillation was 0.05% (v/w). The optimum conditions of SFE were obtained at a pressure of 350 atm, a temperature of 65 °C, a methanol modifier volume of 100 μL, and static and dynamic extraction time of 10 min. Main components of the extracts under optimum SFE conditions were palmitic acid, linoleic acid, γ‐linolenic acid, and oleic acid. The results indicated that by using the suitable extraction conditions, SFE is more effective than the conventional hydrodistillation method in the extraction of fatty acids and the preservation of its quality. Practical Application: SFE is a good technique for the extraction of oils from plants. The extraction yields by SFE are more than the conventional method. SFE is used on a large scale for production of essential oils and pharmaceutical products from plants.     

Supercritical carbon dioxide fluid extraction of Hibiscus cannabinus L. seed oil: A potential solvent-free and high antioxidative edible oil

The supercritical fluid extraction (SFE) trends and antioxidant activities of Hibiscus cannabinus seed oils were studied. SFE results indicate that extraction pressure is the major factor determining the oil yield. In comparison, classic Soxhlet extraction (SOX/L) yielded higher oil content than SFE (P < 0.05). However, no significant differences in oil content were observed in SFE at 600 bars/80 °C, rapid Soxhlet extraction (SOX/S) and conventional ultra-sonic assisted solvent extraction (SONIC) (P > 0.05). Antioxidant activities of H. cannabinus seed oils were compared with 7 types of commercial edible oils. DPPH scavenging activity test indicated that H. cannabinus seed oil extracted by SFE at 200 bars/80 °C possessed the highest antiradical activity whereas beta-carotene bleaching (BCB) assay revealed that all H. cannabinus seed oils (except for SFE at 400 bars/80 °C and 600 bars/80 °C) exhibited higher antioxidant activity than all commercial edible oils (P < 0.05). Thus, SFE – H. cannabinus seed oil may serve as an excellent source of solvent-free edible oil with high antioxidant properties.     

Antimicrobial potential of Coriandrum sativum L. against different Candida species in vitro

The aim of this study was to evaluate the chemical and antifungal activity of the essential oil from Coriandrum sativum L. (Apiaceae) against different Candida species. The essential oil (EO) was obtained by hydrodistillation and submitted to dry-column chromatography, resulting in six fractions, which were then submitted to TLC and GC–MS analysis. The main compounds identified were alcohols: 1-decanol (24.20%); 2E-decenol (18.00%); 2Z-dodecenol (17.60%); and aldehydes (89%). Antibacterial activity of the EO and its fractions was tested against five species of Candida albicans. The EO showed antimicrobial activity against all the species of Candida tested, except for Candida tropicalis CBS 94. Fractions 4 and 6 had a greater antibiotic spectrum, probably due to the presence of such alcohols as 3-hexenol, 1-decanol, 2E-decenol and 2Z-dodecenol. In conclusion, the EO and its fractions could be used as potential antimicrobial agents to treat or prevent Candida yeast infections.     

Chemical properties of the cultivated Sideritis raeseri Boiss. & Heldr. subsp. raeseri

Phytochemical analyses of the cultivated Sideritis raeseri subsp. raeseri in four different stages of flower development were performed. Traditionally used infusion and decoction were also prepared from aerial parts in full flowering stage, and analyses of active compounds and radical scavenging capacity were performed. The highest yield of the essential oil, obtained by hydrodistillation, was noticed in the full flowering phase (0.11%), with sesquiterpene bicyclogermacrene as the main constituent (42.5%). All examined extracts contained phenolic compounds and their amounts varied from 15.3 to 34.1 mg GAE/g DW. The amounts of total phenolics in infusion and decoction were similar (46.5 and 43.9 mg GAE/100 ml, respectively). LC–ESI-MS analyses of all samples allowed the characterisation of 22 phenolic compounds. Two dominant flavone glycosides, 4′-O-methylhypolaetin-7-O-[6?-O-acetyl-β-d-allopyranosyl (1 → 2)-β-d-glucopyranoside (17) and 4′-O-methylisoscutellarein-7-O-[6?-O-acetyl-β-d-allopyranosyl-(1 → 2)]-β-d-glucopyranoside (19) were quantified using HPLC. Moreover, the mineral content and the percent of transportation were investigated.     

The inhibitory effect of Plectranthus barbatus and Plectranthus ecklonii leaves on the viability,glucosyltransferase activity and biofilm formation of Streptococcus sobrinus and Streptococcus mutans

Aqueous extracts of Plectranthus barbatus and Plectranthus ecklonii are traditionally used as anti-inflammatory and anti-fungal agents. The effect of these extracts and of its main component, rosmarinic acid, on the viability of the cariogenic bacteria, Streptococcus sobrinus and Streptococcus mutans, was determined by MIC and MBC. The influence of these extracts on the biofilm formation as well as on the inhibition of glucosyltransferase enzyme, produced by these species, was also analysed. Aqueous extracts of P. barbatus and P. ecklonii were stronger inhibitors than rosmarinic acid. MIC values of 3.8 and 4.7 mg/ml for S. sobrinus and 2.9 and 5.0 for S. mutans were obtained, while rosmarinic acid presented MIC values of 8.4 and 7.3 mg/ml. P. barbartus, P. ecklonii and rosmarinic acid presented MBC values of 9.5, 9.0 and 12.0 mg/ml for S. sobrinus, and 9.5, 10.0 and 12.5 mg/ml for S. mutans. The inhibition of biofilm formation by P. barbatus, P. ecklonii and rosmarinic acid presented IC₅₀ values of, respectively, 0.6, 1.0 and 3.1 mg/ml for S. sobrinus and 1.4 and 2.7 and 1.3 mg/ml for S. mutans. The glucosyltransferase inhibition activity by theses extracts and rosmarinic acid was calculated and IC₅₀ values presented were, respectively, 1.1, ca 1.2 and 2.1 mg/ml for S. sobrinus and 3.1, 1.6 and 3.9 mg/ml for S. mutans were obtained. These extracts may be useful in the prevention of dental carie.     

Characterisation of esterolytic activity from two wild mushroom species, Amanita vaginata var. vaginata and Tricholoma terreum

Characterisation of esterase activities from the edible mushroom species, Amanita vaginata var. vaginata and Tricholoma terreum, were investigated. Native electrophoresis of the crude extracts prepared from both mushroom samples showed the presence of esterolytic activities. The extracts had the greatest activity in the presence of p-nitrophenyl butyrate (pNPB) as a substrate. pH and temperature optima were found to be 8.0 and 30 °C for both enzymes, respectively. V_max and K_m values were determined as 14.2 U/l and 71 μM for A. vaginata var. vaginata and 34.6 U/l and 9.6 μM for T. terreum, respectively. The pH-stability profile showed a stationary line between 3.0 and 10.0 for both enzymes. The esterolytic activities from the extracts were maintained between 10 and 40 °C for 4 h and started to decrease at 50 °C. The effects of EDTA, NaN₃, DTT and PMSF on the enzyme activity were also investigated.     

Green processes based on the extraction with pressurized fluids to obtain potent antimicrobials from Haematococcus pluvialis microalgae

In the present work, the antimicrobial activity of different pressurized liquid extracts obtained from Haematococcus pluvialis microalga was tested against several microorganisms of importance for the food industry (Escherichia coli, Staphylococcus aureus, Candida albicans and Aspergillus niger). Extractions were performed with hexane and ethanol at four different temperatures (50, 100, 150 and 200 °C) for 20 min. The results showed that extracts obtained with both solvents (hexane and ethanol) from the green motile cells of the microalgae (green phase) presented a low antimicrobial activity against all the microorganisms tested. However, the antimicrobial activity of the extracts obtained from the red hematocysts without flagella (red phase) was totally different depending on the solvent used for the extraction. Hexane extracts showed an antimicrobial activity quite similar to that obtained with the green microalgae, while the antimicrobial activity of ethanol extracts was much higher. This fact seems to indicate that compounds related to antimicrobial activity of this microalga are found in higher quantities in the red phase of the microalgae and could be relatively polar compounds. Moreover, ethanol extracts from the red phase obtained at 100 °C presented the highest antimicrobial activity. In order to identify the compounds responsible for the antimicrobial activity, a GC-MS characterization of the extracts obtained with both hexane and ethanol at 100 °C, for Haematococcus pluvialis in the green and red phases was also performed. Therefore, the highest antimicrobial activity of the ethanol extract corresponding to red Haematococcus can be associated with the presence in this extract of short-chain fatty acids, which have been previously described to possess antimicrobial activity.     

Antioxidant, antiacetylcholinesterase and antimicrobial activities of Cymbopogon schoenanthus L. Spreng (lemon grass) from Tunisia

Aqueous extract, proanthocyanidin rich extract, and organic extracts of Cymbopogon schoenanthus L. Spreng (lemon grass) shoots from three different locations in South Tunisia were screened for their antioxidant, acetylcholinesterase and antimicrobial activities. In addition to the evaluation of these activities, the contents of flavonoids and total phenolic compounds were determined.Antioxidant activity measured by DPPH assay showed that the proanthocyanidin extract exhibited higher antioxidant activity than the aqueous extract. Extract concentration providing 50% inhibition (IC₅₀) ranged from 16.4 ± 6.8 μg/mL to 26.4 ± 6.8 μg/mL. The antioxidant activity was also determined using the β-carotene/linoleic acid bleaching test. The best results (IC₅₀ = 0.11 ± 0.10 mg/mL) were obtained with the proanthocyanidin extract of the plants collected from the desert region (Dhibat).The greatest acetylcholinesterase inhibitory activity (IC₅₀ = 0.23 ± 0.04 mg/mL) was exhibited by the ethyl acetate and methanol extracts of the plants collected from the mountainous region. It seems that extracts obtained with more polar solvents gave better results.The proanthocyanidin extracts showed a good antimicrobial activity against Streptococcus sobrinus at low concentration (MIC = 4 mg/mL). Therefore, these extracts could be used to prevent carious lesions by inhibiting S. sobrinus growth.