Phenolic and flavonoid content and antioxidants capacity of pressurized liquid extraction and perculation method from roots of Scutellaria pinnatifida A. Hamilt. subsp alpina (Bornm) Rech. f.

The extract was separated from roots of Scutellaria pinnatifida using perculation and pressurized liquid extraction (PLE). A circumscribed central composite (CCC) was used to optimize the effective extraction variables. For achieving maximum extraction yield via PLE the temperature, pressure, static time, dynamic time, and the solvent flow rate were adjusted 65.8 °C, 39.2 bar, 12.9 min, 18.9 min, and 0.76 mL/min, respectively. Ferric reducing antioxidant power (FRAP) (mmol/g) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (mg/mL) were evaluated and the highest antioxidant activity was observed from the PLE extract. The total phenolic and flavonoid content was calculated and a good correlation founded between phenolic content and antioxidant activity. The results indicated the root of this plant is a potential source of natural antioxidants and flavonoids. The PLE method is quicker and it has more extraction yield than perculation.     

Optimization of multi-stage countercurrent extraction of antioxidants from Ginkgo biloba L. leaves

Multi-stage countercurrent extraction (MCE) as a novel extraction technique was used to extract antioxidants from Ginkgo biloba leaves. Orthogonal array design (OAD) was employed to optimize the ratio of 60% ethanol to raw material (8–16 mL/g), extraction time (30–60 min) and extraction temperature (60–80 °C) to obtain a high yield of antioxidants from G. biloba leaves by MCE. The optimum conditions were a ratio of 60% ethanol to raw material of 16 mL/g and extraction time of 30 min at 80 °C. Under these conditions, the yields of flavonoids and total phenolics were 1.74% and 2.42%, respectively, and DPPH radicals scavenging activity of the extract was 89.97%. Compared with heat-reflux extraction, MCE had obvious advantages of less extraction time and lower solvent and energy consumption. It may be used as a promising technique for the extraction of bioactive compounds from plant materials.     

Characterization of wheat bran oil obtained by supercritical carbon dioxide and hexane extraction

Supercritical carbon dioxide (SC-CO₂) and soxhlet extraction using was carried out to extract oil from wheat bran oil. For SC-CO₂, the pressure and temperature were ranging from 10 to 30 MPa and 313.15–333.15 K. The extraction was performed in a semi batch process with a CO₂ flow rate of 26.81 g/min for 2 h. Wheat bran oil was characterized to investigate the quality. Acid value (AV) and peroxide value (POV) were higher in hexane extracted oil compared to SC-CO₂ extracted oil. Induction period was measured by rancimat test. The oil obtained by SC-CO₂ extraction had higher capability to delay the oxidation by surrounding environment. The DPPH radical scavenging activity was also measured. The SC-CO₂ extracted oil showed higher radical scavenging activity compared to hexane extracted oil.     

Optimization of extraction time and temperature for antioxidant activity of edible wild mushroom,Pleurotus porrigens

The extraction time and temperature of Pleurotus porrigens were optimized for the maximization of 2,2-diphenyl-1-picryhydrazyl (DPPH) radical scavenging and 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonate) (ABTS) radical cation inhibition activities, ferric reducing/antioxidant power (FRAP) and total phenolic content (TPC) using response surface methodology (RSM). A rotatable central composite design consisting of 14 experimental runs with three replicates at the central points was applied and second-order polynomial models were used to describe the experimental data regarding the responses. The experimental results adequately fitted into the second-order polynomial models with significant linear, quadratic and interaction effects of the independent variables. The optimized conditions were 372.8 min/32.0 °C (DPPH); 340.9 min/36.8 °C (ABTS); 240.0 min/38.1 °C (FRAP); and 310.1 min/43.6 °C (TPC) with corresponding yields of 32.66%; 91.21%; 7.91 mM Fe²⁺ equivalent/100 g; and 494 mg gallic acid equivalent/100 g, respectively. The experimental values were close with those predicted values, indicating suitability of the model employing RSM for optimizing the extraction time and temperature on antioxidant activity from P. porrigens.     

Optimization of antioxidant hydrolysate production from flying squid muscle protein using response surface methodology

The squid muscle protein, extracted from by-products of flying squid (Ommastrephes bartrami) was hydrolyzed by five proteases (pepsin, trypsin, papain, alcalase and flavourzyme). DPPH radical scavenging power was used to evaluate antioxidative activity of hydrolysates. The hydrolysate obtained by papain exhibited the most excellent potential of antioxidative activity. Furthermore, response surface methodology (RSM) was employed to optimize hydrolysis conditions, including enzyme to substrate (E/S) ratio, reaction temperature, and hydrolysis time. The optimum conditions obtained were as follows: E/S ratio of 1.74%, temperature of 51 °C and time of 46 min, under which, DPPH radical scavenging activity of 74.25% was obtained. Moreover, it was found that the optimum hydrolysate of 8 mg/mL displayed relatively stronger inhibitory effect on lipid peroxidation compared with α-Tocopherol of 0.1 mg/mL.     

Cosolvent-modified supercritical carbon dioxide extraction of phenolic compounds from bamboo leaves (Sasa palmata)

This study highlights the possibility of supercritical carbon dioxide for extracting phenolic compounds from bamboo leaves that have shown antioxidant and anticancer activities. The CO₂ extraction solvent was modified by adding ethanol–water mixture cosolvent of different concentrations to allow extraction of both polar and non-polar compounds. Conventional Soxhlet extraction was also done to investigate the advantages of supercritical extraction over the conventional extraction method. For addition of 5% (mol) of a 25:75 (mol:mol) ethanol–water mixture solvent to CO₂, the highest amount of polyphenols (7.31 ± 0.06 mg/g bamboo leaves in catechin equivalents) and radical scavenging activity (3.65 ± 0.05 mg/g bamboo leaves in BHA equivalents) at 20 MPa and 95 °C, could be obtained among the mixture cosolvents studied. For Soxhlet extraction with a 25:75 (mol:mol) ethanol–water mixture, 1.48 times the amount of phenolic compounds (10.85 ± 0.52 mg/g bamboo leaves in catechin equivalents), could be isolated compared with the supercritical extraction method, however, the radical scavenging activity (3.30 ± 0.05 mg/g bamboo leaves in BHA equivalents) was 0.90 times lower than the extract obtained from the supercritical extraction method. The seven major antioxidative compounds identified from the SC-CO₂ extraction method were: (1) dl-alanine, (2) gluconic acid, (3) phosphoric acid, (4) ß-siosterol, (5) β-amyrene, (6) α-amyrin acetate and (7) friedelin.     

Subcritical water extraction of steviol glycosides from Stevia rebaudiana leaves and characterization of the raffinate phase

The objectives of this work were to obtain steviol glycosides of S. rebaudiana leaves, possessing natural and noncaloric sweetener properties, using subcritical water extraction; to assess optimum extraction conditions; to determine biological activities of Stevia extracts and to characterize the raffinate phase. A Box–“Bhenken” statistical design was used to evaluate the effects of various values of temperature (100–150 °C), time (30–60 min) and flow rate (2–6 ml/min) at a pressure of 230 bar applying a solid/liquid ratio of 1:10 (m:v). The most effective parameter was temperature (p < 0.005). Optimum extraction conditions were elicited as 125 °C, 45 min, 4 ml/min flow rate which yielded 38.67 mg/g stevioside and 35.68 mg/g rebaudioside A. The total phenolic, flavonoid contents and DPPH free radical scavenging activity were found as 48.63 mg gallic acid/g extract, 29.81 mg quercetin/g extract and 92.50%, respectively. After extraction, total chlorophyll, carotenoid contents and dietary fibers were quantified as 31.91 mg/100 g, 5.71 mg/100 g and 4.98% in the raffinate phase. Hence, both extract and raffinate phases of S. rebaudiana leaves can be utilized as sources of natural sweeteners, fibers and coloring agents in the industry.     

Extraction of low-molar-mass phenolics and lipophilic compounds from Pinus pinaster wood with compressed CO2

Near-supercritical and supercritical CO₂ was used to extract low-molar-mass phenolics and lipophilic compounds from Pinus pinaster wood. Extraction of samples containing sapwood and knotwood was carried out at 10⿿25 MPa and 30⿿50 °C to assess the influence of the operational conditions on the yields of total extracts and phenolics, as well as on the radical scavenging capacity of extracts. The use of ethanol as a co-solvent increased both the extraction yields and the concentration of phenolics in extracts. Operating under selected conditions (25 MPa, 50 °C, 10% ethanol), the extraction yield accounted for 4.1 wt% of the oven-dry wood. The extracts contained up to 7.6 g of phenolic compounds (measured as gallic acid equivalents) per 100 g extract, and showed one third of the radical scavenging capacity of Trolox. Native resin acids accounted for about 24 g per 100 g extracts, whereas flavonoids, lignans, stilbenes and juvabiones were found at lower proportions.     

Optimization of microwave-assisted enzymatic extraction of polyphenols from waste peanut shells and evaluation of its antioxidant and antibacterial activities in vitro

A microwave-assisted enzymatic extraction (MAEE) method was developed and optimized to enhance the polyphenols extraction yield from waste peanut shells. The optimum conditions were as follows: irradiation time 2.6 min, amount of cellulase 0.81 wt.%, a pH of 5.5, and incubation at 66 °C for 2.0 h. Under these conditions, the extraction yield of total polyphenols could reach 1.75 ± 0.06%, which was higher than other extraction methods including heat-refluxing extraction, ultrasonic-assisted extraction and enzyme-assisted extraction. The structural changes of the plant material after different extractions observed by scanning electron microscopy provided visual evidence of the disruption effect. Moreover, the crude extract was then purified by NKA-9 resin, the polyphenols content in the purified extract increased to 62.73%. The antioxidant activities of the crude and purified polyphenols extract were evaluated by DPPH and hydroxyl radicals, reducing power and β-carotene bleaching test. The antibacterial activities of purified extract were also tested using Oxford cup method. The results indicated that the MAEE method was efficient and environment-friendly, and the polyphenols have significant antioxidant and antibacterial activities, which can be used as a source of potential antioxidant and preservative.     

Extraction of phenolic compounds from pitanga (Eugenia uniflora L.) leaves by sequential extraction in fixed bed extractor using supercritical CO2, ethanol and water as solvents

With the goal of maximizing the extraction yield of phenolic compounds from pitanga leaves (Eugenia uniflora L.), a sequential extraction in fixed bed was carried out in three steps at 60 °C and 400 bar, using supercritical CO₂ (non-polar) as solvent in a first step, followed by ethanol (polarity: 5.2) and water (polarity: 9.0) in a second and third steps, respectively. All extracts were evaluated for global extraction yield, concentration and yield of both polyphenols and total flavonoids and antioxidant activity by DPPH method (in terms of EC₅₀). The nature of the solvent significantly influenced the process, since the extraction yield increased with solvent polarity. The aqueous extracts presented higher global extraction yield (22%), followed by ethanolic (16%) and supercritical extracts (5%). The study pointed out that the sequential extraction process is the most effective in terms of global extraction yield and yield of polyphenols and total flavonoids, because it produced the more concentrated extracts on phenolic compounds, since the supercritical ethanolic extract presented the highest phenolics content (240.5 mg GAE/g extract) and antioxidant capacity (EC₅₀ = 9.15 μg/mL). The most volatile fraction from the supercritical extract, which is similar to the essential oils obtained by steam distillation or hydrodistillation, presented as major compounds the germacrenos D and B + bicyclogermacrene (40.75%), selina-1,3,7(11)-trien-8-one + selina-1,3,7(11)-trien-8-one epoxide (27.7%) and trans-caryophyllene (14.18%).     

Hydrolysis of pumpkin oil cake protein isolate and free radical scavenging activity of hydrolysates: Influence of temperature,enzyme/substrate ratio and time

The effect of hydrolysis parameters (temperature, initial enzyme/substrate ratio and time) on the hydrolysis of pumpkin oil cake protein isolate (PuOC PI) with acid protease from Aspergillus niger and the antioxidant potency of the obtained hydrolysates were studied by response surface methodology (RSM). The hydrolysis progress, measured by the degree of hydrolysis (DH), was described by a second-order polynomial model (R² = 0.77) and the conditions for optimum DH (42.94%) were found at temperature of 40 °C, enzyme/substrate ratio (E/S) 4.38 HUT/mg of substrate proteins and 85 min. The antiradical activity (AA) of the PuOC PI hydrolysates was examined by the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) assay; all hydrolysates showed a concentration dependent scavenging activity against DPPH radicals. The AA of hydrolysates was influenced by process parameters and was presented also by a second-order polynomial model (R² = 0.7). The conditions to achieve the highest DH did not result hydrolysates with the optimum AA; the highest AA ranged from 34% to 40% and were found in hydrolysates obtained at 50 °C.     

Optimization of subcritical water extraction of antioxidants from Coriandrum sativum seeds by response surface methodology

Subcritical water extraction (SWE) of antioxidants from Coriandrum sativum seeds (CSS) was optimized by simultaneous maximization of the total phenolics (TP) and total flavonoids (TF) yield and antioxidant activity, using IC₅₀ value. Box–Behnken experimental design (BBD) on three levels and three variables was used for optimization together with response surface methodology (RSM). Influence of temperature (100–200 °C), pressure (30–90 bar) and extraction time (10–30 min) on each response was investigated. Experimentally obtained values were fitted to a second-order polynomial model and multiple regression. Analysis of variance (ANOVA) was used to evaluate model fitness and determine optimal conditions. Moreover, three-dimensional surface plots were generated from employed mathematical model. The optimal SWE conditions obtained in simultaneous optimization were temperature of 200 °C, pressure of 30 bar and extraction time of 28.3 min, while obtained values of TP and TF yields and IC₅₀ value at this experimental point would be 2.5452 g GAE/100 g CSS, 0.6311 g CE/100 g CSS and 0.01372 mg/ml, respectively. Moreover, good and moderate linear correlation was observed between antioxidant activity (IC₅₀ value) and total phenolics content (R² = 0.965), and total flavonoids content (R² = 0.709) which indicated that these groups of compounds are responsible for antioxidant activity of C. sativum extracts.     

Sequential extraction of bioactive compounds from Melia azedarach L. in fixed bed extractor using CO2, ethanol and water

Melia azedarach L. is a plant with wide use in folk medicine since it contains many bioactive compounds of interest. The present study aimed to extract bioactive compounds from M. azedarach fruits by a sequential process in fixed bed using various solvent mixtures. Extractions were performed at 50 °C and 300 bar in four sequential steps using supercritical CO₂ (scCO₂), scCO₂/ethanol, pure ethanol, and ethanol/water mixture as solvents, respectively. The efficacy of the extraction process was evaluated by extraction yield and kinetics, and analysis of extracts by: (1) thin layer chromatography (TLC), (2) phenolics content, (3) reduction of surface tension of water, (4) gas chromatography (GC–MS), (5) electrospray ionization mass spectrometry (ESI–MS) and (6) antiviral activity. The overall extraction yield reached 45% and TLC analysis showed extracts with different composition. extract obtained from CO₂/ethanol mixture (SCEE) exhibited the greatest ability to reduce surface tension of water from 72.4 mN m⁻¹ [1] of pure water to 26.9 mN m⁻¹ of an aqueous solution of 40 g L⁻¹. The highest phenolics contents were observed in both the hydroalcoholic extract and scCO₂/ethanolic extract. Volatile oils were not detected in the supercritical extracts by GC–MS. MS analyses identified the fatty acids: linoleic, palmitic and myristic acid in the supercritical extract (SCE), and the phenolics: caffeic acid and malic acid in the other extracts. In addition, SCE and SCEE extracts showed significant inhibition percentage against Herpes Simplex Virus Type 1. The extraction process proposed in the present study produced extracts with significant potential for application in food and pharmaceutical industries.     

Antioxidant properties of Achyranthis radix extract in rats

This study was performed to investigate the antioxidant activities of various fractions of Achyranthis radix extract using various solvents in vitro and antioxidant enzyme activities in vivo. Among the various fractions, when n-butanol was used as the extract solvent the DPPH radical scavenge rate was highest at 31.09 μg/ml. The thiobarbituric acid reaction value using n-butanol fraction was 0.33 at 532 nm after 6 days, representing a decrease of about 42% compared to control group. When the administered concentration of A. radix extract was 400 mg/kg/body weight/day, the xanthine oxidase activity in the liver was 19.5 mU/mg protein, representing a decrease of about 46% compared to the control group. The superoxide dismutase activity in the AR-E2 group was 89.4 Umol/(min mg protein), representing a decrease of about 40% compared to the control group. Catalase and glutathione peroxidase were decreased by about 24% and 58%, respectively, compared to the control group. When n-butanol fraction was used, the nitrite scavenging activity in various fractions at pH 1.2 was highest at 90.2%, an increase of about 90% compared to the control group. The results support the antioxidant potential of A. radix extract for various health supplements, pharmaceuticals, and mutraceuticals.     

Influence of pre-treatments on yield,chemical composition and antioxidant activity of Satureja montana extracts obtained by supercritical carbon dioxide

Main objective of this work was to investigate the influence of pre-treatments of Satureja montana herbal material on supercritical extraction process, in terms: influence on extraction yield, composition and antioxidant activity of extracts. Four different pretreatments were investigated: water pre-treatment, ethanol pre-treatment, ultrasound and high pressure pre-treatment. Extraction yields were in the range from 1.68 to 2.35 g/100 g. Pre-treatments with water and ethanol increase the yield of extraction for 25% and 17%, respectively. According to GC/MS results the main compound of obtained extracts was carvacrol. Analyses confirmed that carvacrol content in extracts can be significantly increased by application of pre-treatments. Ultrasound pre-treatment and high pressure pre-treatment yielded highest concentration of carvacrol in extracts (around 66%). Using these two pre-treatments of herbal material, concentration of carvacrol, in extracts obtained by supercritical carbon dioxide extraction, could be increased for around 25%. Antioxidant activity of all extract was very high and in the range of well-known antioxidants.     

Supercritical extraction of carob kibbles (Ceratonia siliqua L.)

Carob pulp kibbles, a by-product of carob been gum production, was studied as a source of bioactive agents. Firstly, the carob kibbles were submitted to an aqueous extraction to extract sugars, and supercritical fluid extraction (SFE) was applied to the solid residue of that aqueous extraction, by using compressed carbon dioxide (SC-CO₂) as the solvent and a mixture of ethanol and water (80:20, v/v) as a co-solvent. Pressure and temperature were studied in the ranges 15–22 MPa, and 40–70 °C. Particle diameter, and co-solvent percentage in ranges of 0.27–1.07 mm, and 0–12.4%, respectively, were also studied, as well as the flow rate of SC-CO₂ between 0.28 and 0.85 kg h⁻¹, corresponding, respectively, to 0.0062 and 0.0210 cm s⁻¹ of superficial velocity. The extracts were characterised in terms of antioxidant capacity by DPPH method, and total phenolics content by the Folin–Ciocalteu method. The central composite non-factorial design was used to optimise the extraction conditions, using the Statistica, version 6 software (Statsoft). The best results, in terms of yield and antioxidant capacity, were found at 22 MPa, 40 °C, 0.27 mm particle size, about 12.4% of co-solvent and a flow rate of 0.29 kg h⁻¹ of SC-CO₂. The phenolics profile of the extracts obtained at these conditions was qualitatively evaluated by HPLC-DAD. The solid residue of the supercritical extraction was also studied showing to be a dietary fiber, which can be compared to Caromax™, a carob fiber commercialised by Nutrinova Inc.     

Supercritical fluid extraction of Prunus persica leaves and utilization possibilities as a source of phenolic compounds

Mediterranean countries contribute highly on world peach production and tonnes of waste leaves are released due to pruning. The aim of this study was to investigate the utilization possibilities of the leaves by supercritical fluid extraction. A statistical design was used to evaluate the effect of temperature (40–80 °C), pressure (150–300 bar) and concentration of ethanol as co-solvent (6–20%) at a flow rate of 15 g/min and for a duration of 60 min. The most effective variables were found as pressure and co-solvent ratio (p < 0.005). Optimum extraction conditions were elicited as 60 °C, 150 bar and 6% co-solvent yielding a total phenol content of 79.92 mg GAE/g extract, EC₅₀ value of 232.20 μg/ml and a radical scavenging activity of 53.25% which was higher than the value obtained by conventional solvent extraction method (32.23%). Consequently, Prunus persica L. leaves were found as a potential phenolic source for industrial applications.     

Ethanol modified supercritical carbon dioxide extraction of flavonoids from Momordica charantia L. and its antioxidant activity

Ethanol modified supercritical carbon dioxide (SC-CO₂) extraction of flavonoids from Momordica charantia L. fruits and its antioxidant activity were performed. The influences of parameters such as temperature, extraction time and pressure on the yield of flavonoids were investigated. The antioxidant activities of flavonoids were assessed by means of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging assay and β-carotene bleaching test. The experimental data obtained indicated that pressure, temperature and time had significant effect on the extraction yield. The optimum extraction conditions, determined by the 3D response surface and contour plots derived from the mathematical models, were as follows: extraction temperature 46 °C, pressure 33.4 MPa, and extraction time 53.2 min. Under these conditions, the experimental value was 15.47 mg/g, which was well matched with value predicted by the model. The antioxidant activity of flavonoids obtained by ethanol modified SC-CO₂ extraction method had higher antioxidant activity than the flavonoids extracted by conventional solvent extraction (CSE) method. The DPPH radical-scavenging ability of flavonoids obtained by ethanol modified SC-CO₂ extraction method reached to 96.14 ± 1.02%, equivalent to the clearance rate of ascorbic acid at 1.2 mg/mL. Results indicated that ethanol modified SC-CO₂ extraction was a suitable approach for the selective extraction of flavonoids from M. charantia L.     

Response surface optimization of Smyrnium cordifolium Boiss (SCB) oil extraction via supercritical carbon dioxide

In this study, the essential oil of aerial parts of a species of a plant called Smyrnium cordifolium Boiss (SCB) was extracted by supercritical CO₂. The essence was analyzed by the method of GC/MS. Design of experiments was carried out with response surface methodology by Minitab 16 software to optimize four operating variables of supercritical carbon dioxide (SC-CO₂) extraction (pressure, temperature, CO₂ flow rate and extraction dynamic time). This is the first report announcing optimization of the operation of supercritical extraction of SCB in laboratorial conditions. Optimizing process was done to achieve maximum yield extraction. Independent variables were dynamic time (t_d), pressure (P), temperature (T) and flow rate of SC-CO₂ (Q) in the range of 30–150 min, 10–30 MPa, 40–60 °C and 0.5–1.7 ml/min, respectively. The experimental optimal recovery of essential oil (0.8431, w/w%) was obtained at 13.43 MPa, 40 °C, 150 min (dynamic) and 1.7 ml/min (CO₂ flow rate).     

Enrichment of Nigella damascena extract with volatile compounds using supercritical fluid extraction

β-Elemene, germacrene A and damascenine were extracted from lady-in-a-mist (Nigella damascena L.) seeds with supercritical carbon dioxide at 10–30 MPa and 40–60 °C. The influence of supercritical fluid extraction (SFE) conditions on the yield and concentration of volatiles in the extract and the extraction kinetics were studied. The extraction yields and the apparent solubility of volatile compounds increased with increasing density of CO₂. The highest total yield was obtained at 30 MPa and 40 °C but the selectivity for volatiles was low under these conditions. With respect to both yield of volatiles and their concentration in extract, the best results were at 12 MPa and 40 °C, either with one separator or with additional separator maintained at 5 MPa and 25 °C. The yields of β-elemene, germacrene A and damascenine reached 0.72, 3.31 and 3.65 mg g⁻¹ and their concentration in the extract was 2.62, 12.04 and 13.28 wt.%, respectively. Though the yields of germacrene A and damascenine were by about 20% higher using Soxhlet extraction with hexane than using SFE, their concentration in the extract where fatty oil prevailed was only 1.19 and 1.20 wt.%, respectively. Under the conditions of hydrodistillation, partial conversion of germacrene A to β-elemene occurred and its yield was higher than using the other methods but the composition of volatiles in the SFE extracts better corresponds to the original raw material.