Scale-up studies of the supercritical fluid extraction of triterpenic acids from Eucalyptus globulus bark

The supercritical fluid extraction (SFE) of Eucalyptus globulus deciduous bark is investigated, under the context of biorefinery, aiming at the definition of a pathway for the production of enriched extracts in triterpenic acids (TTAs). Scale-up studies are performed, well supported by modeling and by experiments carried out at lab, intermediate and pilot scales (0.5, 5.0 and 80.0 L, respectively). By acknowledging the significant role played by cosolvent (ethanol) addition on the concentration of TTAs, extraction curves are measured at 200 bar and 40 °C, using two different ethanol contents, 2.5% and 5.0% (wt). Taking into account the results achieved by four distinct models, the intraparticle diffusion is confirmed as the prevailing mass transfer resistance in both conditions. Accordingly, the ratio between CO₂ flow rate and biomass weight is adopted as scale-up criterion. Despite the geometric differences between the three units, the measured extraction yields and TTAs concentration evidence good agreement, which validates the established scale-up rule. In the whole, our results legitimate the technical viability of the SFE of E. globulus deciduous bark for future exploitation at larger scales.     

Supercritical Carbon Dioxide and Microwave-Assisted Extraction of Functional Lipophilic Compounds from Arthrospira platensis

Arthrospira platensis biomass was used in order to obtain functional lipophilic compounds through green extraction technologies such as supercritical carbon dioxide fluid extraction (SFE) and microwave-assisted extraction (MAE). The temperature (T) factor was evaluated for MAE, while for SFE, pressure (P), temperature (T), and co-solvent (ethanol) (CS) were evaluated. The maximum extraction yield of the obtained oleoresin was (4.07% ± 0.14%) and (4.27% ± 0.10%) for SFE and MAE, respectively. Extracts were characterized by gas chromatography mass spectrometry (GC-MS) and gas chromatography flame ionization detector (GC-FID). The maximum contents of functional lipophilic compounds in the SFE and MAE extracts were: for carotenoids 283 ± 0.10 μg/g and 629 ± 0.13 μg/g, respectively; for tocopherols 5.01 ± 0.05 μg/g and 2.46 ± 0.09 μg/g, respectively; and for fatty acids 34.76 ± 0.08 mg/g and 15.88 ± 0.06 mg/g, respectively. In conclusion, the SFE process at P 450 bar, T 60 °C and CS 53.33% of CO₂ produced the highest yield of tocopherols, carotenoids and fatty acids. The MAE process at 400 W and 50 °C gives the best extracts in terms of tocopherols and carotenoids. For yield and fatty acids, the MAE process at 400 W and 70 °C produced the highest values. Both SFE and MAE showed to be suitable green extraction technologies for obtaining functional lipophilic compounds from Arthrospira platensis.     

Supercritical fluid extraction of flavonoids from Maydis stigma and its nitrite-scavenging ability

Supercritical fluid carbon dioxide (SF-CO₂) extraction (SFE) of flavonoids from Maydis stigma and its nitrite-scavenging ability were investigated. The effects of extraction time, particle size and co-solvent composition in terms of water content in ethanol were first optimized. Then, a Box-Behnken design combined with response surface methodology (RSM) was employed to study the effects of three independent variables (temperature, pressure and co-solvent amount) on the extraction yield of flavonoids. A maximal extraction yield of flavonoids of approximately 4.24 mg/g of M. stigma by SFE was obtained under optimal conditions (a temperature of 50.88 °C, a pressure of 41.80 MPa, a co-solvent amount of 2.488 mL/g and an extraction time of 120 min with 0.4-mm particle sizes and 20% aqueous ethanol as the co-solvent). Furthermore, the nitrite-scavenging ability of the flavonoid-enriched SFE extracts was assessed using the Griess reagent. The flavonoid-enriched SFE extracts exhibited the highest scavenging ability on nitrite (88.1 ± 3.04%) at the concentration of 500 μg/mL and at pH 3.0. The nitrite-scavenging ability of the extracts appeared to be concentration dependent but negatively correlated with the pH.     

Propolis extracts obtained by low pressure methods and supercritical fluid extraction

Propolis is a natural product used for centuries by human kind, due to several evidenced biological activities: antioxidant, antimicrobial, anti-inflammatory, antitumor and anti-HIV. Extracts from propolis, used in food, pharmaceutical and cosmetic industries, present quality and composition related to the extraction method applied. Natural compounds with biological activity can be obtained by conventional techniques, such as Soxhlet and Maceration, or by alternative methods such as supercritical fluid extraction (SFE). Thus, the aim of this work was to compare propolis extraction yields obtained by different procedures, for instance, SFE in one stage, with CO₂ and CO₂ plus co-solvent, and SFE in two stages, as well as Soxhlet and Maceration as low pressure extraction methods using ethanol, ethyl acetate, chloroform, n-hexane, water and mixtures of water/ethanol. The operational conditions for SFE in one stage with pure CO₂ were: 30, 40 and 50 °C and from 100 to 250 bar. The SFE with co-solvent was performed at 150 bar and 40 °C and ethanol concentrations of 2, 5 and 7% (w/w). The highest yield was obtained by chloroform Soxhlet extraction (73 ± 2%, w/w) whereas for SFE the maximum yield was 24.8 ± 0.9%, using 5% ethanol as co-solvent. For SFE in two stages, 100 and 150 bar were used in the first stage while 250 and 300 bar were applied in the second stage, at 40 °C. The yields were 8.4 ± 0.7 (150 bar) and 5.1 ± 0.7 (250 bar), for stages 1 and 2, respectively. The chemical composition of the propolis material was determined by HPLC analysis. The experimental data were correlated using four models based on differential mass balance equations: (1) the Sovová’s model; (2) the logistic model (3) the diffusion model and (4) the simple single plate model (SSP). The logistic model provided the best adjustment for propolis SFE curves.     

Investigation of Oleuropein Content in Olive Leaf Extract Obtained by Supercritical Fluid Extraction and Soxhlet Methods

The separation processes of the phenolic compounds from solid plant matrixes are of great importance. In the scope of developing more efficient methods to separate olive leaf extract, dried and ground olive tree leaves from Aegean region of Turkey were extracted by means of Soxhlet and supercritical fluid extraction (SFE) methods. In the Soxhlet method, different types of solvents (hexane, water, ethanol, methanol, and methanol/hexane (3:2, v/v) mixture) were used to determine the effect of the solvent type on the extraction performance. In the SFE method, the effect of pressure (100–300 bar), temperature (50 and 100°C), and type of co-solvent on the amount of both extract and oleuropein were investigated. Ethanol, methanol and water were selected as co-solvent in 20% (v/v) amount. Quantitative analysis was performed by using a liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) technique. The results of SFE were compared with those obtained by the Soxhlet method. Whereas the highest oleuropein yield was achieved via the Soxhlet method through methanol with the value of 37.84 mg/g dried leaf, the best oleuropein yield was achieved with the value of 14.26 mg/g dried leaf by using CO₂ modified by methanol at 300 bar and 100°C in the SFE method.     

Effects of Operating Parameters on the Cinnamaldehyde Content of Extracted Essential Oil Using Various Methods

Supercritical fluid extraction (SFE) of essential oils from commercial cinnamon bark was compared with essential oils that were obtained by hydrodistillation. Effects of operating parameters (pressure, temperature and extraction time of SFE) on the extraction yield and the composition of the extracted volatile oil were studied. Moreover, in the hydrodistillation process, the effect of the pH of the solvent on the concentration of cinnamaldehyde in the extracted volatile oil was studied. The maximum yield of extract in the SFE process is about 7.8 % at 70 °C and 240 bar. The maximum concentration of cinnamaldehyde in the SFE process was obtained at 70 °C and 160 bar, and the maximum concentration of this component in hydrodistillation was achieved at pH = 4.1.     

Extraction of pollen lipids by SFE‐CO2 and determination of free fatty acids by HPLC

Rape bee pollen lipids obtained by petrol ether extraction (PEE) or supercritical fluid (carbon dioxide) extraction (SFE) were compared with regard to their free fatty acid (FFA) components. Optimal SFE conditions were selected by carrying out the Taguchi method with an OA9 (3³) matrix design, and are as follows: extraction pressure at 35 MPa, temperature at 45 °C, and dynamic extraction time at 90 min. The lipid yield based on PEE was 7.42 wt‐% and the extracts of the desired analytes based on SFE varied in the range of 3.23–5.58 wt‐% under different conditions. With the optimized procedure, the lipid yield was 6.09 wt‐%. The FFA in the lipids were separated with a pre‐column derivation method and 1‐[2‐(p‐toluenesulfonate) ethyl]‐2‐phenylimidazole [4,5‐f]9,10‐phenanthrene as labeling regent, followed by high‐pressure liquid chromatography (HPLC) with fluorescence detection. HPLC analysis shows that the lipids contain abundant unsaturated fatty acids (UFA) in high to low concentrations as follows: linolenic acid (18:3), oleic acid (18:1), linoleic acid (18:2), nervonic acid (24:1), and lignoceric acid (20:4). The UFA contents in the SFE extracts were higher than those after PEE. The results indicated that SFE under suitable conditions is more selective than conventional PEE with regard to lipid extraction and preservation of their quality.     

Bioactive extracts of orange (Citrus sinensis L. Osbeck) pomace obtained by SFE and low pressure techniques: Mathematical modeling and extract composition

The objective of this study was to obtain orange (Citrus sinensis L. Osbeck) pomace extract using supercritical fluid extraction (SFE) with CO₂ and with CO₂ and co-solvent. In order to evaluate the high pressure method in terms of process yield, extract composition and biological activity, low pressure methods were also applied to obtain orange extracts, such as ultrasound (UE) and soxhlet (SOX), with different organic solvents, and hydrodistillation (HD). The SFE conditions were temperatures of 313.15 K and 323.15 K and pressures from 100 to 300 bar. The SFE kinetics and mathematical modeling of the overall extraction curves (OEC) were also investigated. The antioxidant potential of the extracts was evaluated by the DPPH method, by the Folin-Ciocalteau method and by the β-carotene/linoleic acid bleaching method. The antimicrobial activity of the extracts was also studied. The main compounds identified were l-limonene, palmitic and oleic acids, n-butyl benzenesulfonamide and β-sitosterol.     

Extraction of phenolic fraction from guava seeds (Psidium guajava L.) using supercritical carbon dioxide and co-solvents

In this work the supercritical fluid extraction (SFE) with carbon dioxide (CO₂) and with ethyl acetate (EtAc) and ethanol (EtOH) as co-solvents was applied to obtain the phenolic fraction from guava seeds (Psidium guajava L.). The extraction was explored at various operating conditions, using 10, 20 and 30 MPa and 40, 50 and 60 °C. The use of EtAc and EtOH as co-solvents in SFE was also studied. The supercritical process was compared with traditional techniques such as Soxhlet extraction using EtAc and EtOH as solvents. The quality of the different extracts, obtained using SFE and Soxhlet methods and different solvents, was evaluated through the antioxidant activity, obtained by the collection methods of scavenging DPPH and bleaching of β-carotene, and also through the total phenolic content (TPC) of the samples, by the Folin-Ciocalteu method. The antioxidant potential indicates the use of ethanol as co-solvent as the best modifier in SFE, used in concentration of 10% (w/w) at 50 °C and 30 MPa. The quality of the extracts obtained by SFE with EtOH varied with the operating conditions of temperature and pressure, with higher values obtained at 10 and 20 MPa for TPC results and also antioxidant methods. The process yield of the phenolic fraction was also evaluated for all the extraction procedures studied (SFE and Soxhlet), with results varying from 0.380 to 1.738% (w/w).     

Optimization of the supercritical fluid extraction of triterpenic acids from Eucalyptus globulus bark using experimental design

The supercritical CO₂ extraction of E. globulus deciduous bark was carried out at different temperatures (40–60 °C), pressures (100–200 bar), and ethanol contents (0.0–5.0 wt. %) to study triterpenic acids (TTAs) recovery. A factorial design of experiments and response surface methodology were implemented to analyze the influence of these variables upon extraction and perform its optimization. The best conditions were 200 bar, 40 °C and 5% ethanol, for which the statistically validated regression models provided: extraction yield of 1.2% (wt.), TTAs concentration of 50%, which corresponds to TTAs yield of 5.1 g/kg of bark and a recovery of 79.2% in comparison to the Soxhlet value. The trends of the free and acetylated TTAs were very different, due to their distinct CO₂-philic character caused by dissimilar polarities: the acetyl derivatives approached a plateau near 200 bar and 5% ethanol, while the free TTAs extraction always increased in the range of conditions studied.     

Recovery of betulinic acid from plane tree (Platanus acerifolia L.)

Betulinic acid (3β, hydroxy-lup-20(29)-en-28-oic acid) is a bioactive triterpenic acid which was identified in various botanical sources and in considerable amounts in the bark of plane tree (Platanus acerifolia L.). In this work, the recovery of betulinic acid from plane tree bark was studied using different liquid solvent based extraction methods, namely solid–liquid extraction (SLE), ultrasound assisted extraction (UAE) and pressurized liquid extraction (PLE). Furthermore, preliminary studies of the supercritical fluid extraction (SFE) of plane tree bark are also reported.The liquid solvent based extraction techniques (SLE, UAE and PLE) were carried out using ethanol and ethyl acetate, and produced a recovery of betulinic acid in the range 10–15 mg/g of bark, with concentrations around 25–35% mass. A betulinic acid enrichment in the ethanolic extracts was possible by means of a simple precipitation step adding water. The precipitate contained 42–46% mass of betulinic acid and high recovery (>95%). Increasing the extraction temperature, by means of the PLE assays, has not resulted in an improvement of betulinic acid recovery.The preliminary SFE assays produced lower recoveries of betulinic acid (0.5–8 mg/g) with respect to liquid extraction. The addition of ethanol as cosolvent produced a significant improvement of both betulinic acid recovery and concentration in the SFE extract.     

Study on Oleuropein Extraction from Olive Tree (Olea europaea) Leaves by means of SFE: Comparison of Water and Ethanol as Co-Solvent

The present study focuses on developing methods for olive leaf extraction and deals with obtaining extract, rich in oleuropein, which is the most abundant phenolic compound in olive leaves. Supercritical fluid extraction (SFE) was applied to the dried and ground olive leaves by using CO₂ as supercritical (SC) fluid in the presence of water and ethanol as co-solvent. The influences of operating parameters by means of co-solvent content (0-1 mL/min), temperature (50 and 100°C) and pressure (100-300 bar) on both extract and oleuropein yields were investigated. Quantitative analysis was performed by using a liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) technique. The experimental results obtained by using SC-CO₂ alone were not satisfactory, and it was seen that addition of a polar modifier is necessary in order to improve yield and selectivity of the process. It was observed that CO₂ modified by water and ethanol showed nearly the same extract performance, where CO₂ modified by water is better for high oleuropein yield.     

Supercritical CO2 extraction of glycosides from Stevia rebaudiana leaves: Identification and optimization

The aim of this work was to optimize the glycoside composition of Stevia rebaudiana leaves using supercritical fluid extraction (SFE). A Box-Behnken statistical design was used to evaluate the effect of various values of pressure (150–350 bar), temperature (40–80 °C) and concentration of ethanol-water mixture (70:30) as co-solvent (0–20%) by CO₂ flow rate of 15 g min⁻¹ for 60 min. The most effective variables were co-solvent concentration (P < 0.005) and temperature (P ≤ 0.005). Evaluative criteria for both dependent variables (stevioside and rebaudioside A yields) in the model was assigned maximum. Optimum extraction conditions were elicited as 211 bar, 80 °C and 17.4% which yielded 36.66 mg/g stevioside and 17.79 mg/g rebaudioside A. Total glycosides composition were close to those obtained using conventional water extraction (64.49 mg/g) and a little higher than ethanol extraction (48.60 mg/g) demonstrating challenges for industrial scale application of SFE.     

Supercritical fluid extraction of α- and β-acids from hops compared to cyclically pressurized solid–liquid extraction

In this paper, two solid–liquid extraction techniques, supercritical fluid extraction (SFE) with and without modifiers and cyclically pressurized solid–liquid extraction with a Naviglio Extractor, were compared on the basis of extraction of acidic compounds contained in hops flowers. The hops extracts were analyzed by electro-kinetic capillary chromatography (MECK). The results showed that the technique using supercritical carbon dioxide was more effective for the isolation of β acids; the use of ethanol as a co-solvent, as reported in the literature, produced a heterogeneous extract, while cyclically pressurized solid–liquid extraction showed a greater extraction capacity for α acids. Consequently, both techniques are valid for the extraction of α and β acids from hops. By suitably varying the parameters of the two extractive procedures, it will be possible to obtain extracts for use in the production of beer and dietary supplements and drugs. Furthermore, based on the SFE CO₂ extraction process, a mathematical model was applied to the examined process, and a numerical simulation was performed, leading to a model that provides direction for the optimization of further experiments.     

Antioxidant and antimicrobial potential of cajazeira leaves (Spondias mombin) extracts

Cajazeira leaves (Spondias mombin) have their highlighted use as antioxidant and natural antimicrobial, which justifies the objective of this work to evaluate the biological activities of different extracts. In order to evaluate the antioxidant and antimicrobial potentials of the cajazeira leaves, extractions at low pressure and high pressure were performed. The low pressure extractions (PLE) were carried out using Soxhlet (SOX) and tip ultrasound, using different solvents. Supercritical fluid extraction (SFE) was evaluated at temperature of 40?60°C and pressure of 150?300 bar besides extraction with cosolvent. Higher yields were obtained with the use of more polar solvents at LPE. The extracts obtained by SOX with ethanol and others polar solvents presented the best TPC values and antioxidant activity. The extracts at LPE with hexane and ethyl acetate and SFE presented better antimicrobial activity. Through liquid chromatography of high efficiency, it was possible to identify compounds with recognized biological activity, like ellagic acid, gallic acid and catechin.     

Radical-scavenging activity of extracts from Cordia verbenacea DC obtained by different methods

The possibility of using the leaves of Cordia verbenacea as a new source of natural antioxidant compounds was investigated. In the present work, extracts from C. verbenacea were obtained using different extraction methods: supercritical fluid extraction (SFE), Soxhlet (SE), hydrodistillation and maceration, with the objective to evaluate the methods in terms of yield and antioxidant potential. The high-pressure technique was applied using pure CO₂ and CO₂ with co-solvent at different temperatures and pressures (30, 40 and 50 °C and 100, 200, and 300 bar). Organic solvents with different polarities were used to obtain extracts by low-pressure extraction processes. The extracts were evaluated according to their antioxidant activity using total phenolic content, scavenging abilities on DPPH radical, total antioxidant activities (ABTS^•+), superoxide anion radical-scavenging (O₂^•) and protection against lipid peroxidation in vitro (LPO). Ethyl acetate fraction obtained by maceration and extract isolated by SE using 25% aqueous mixture of ethanol possessed the highest scavenger activity against DPPH radical (IC₅₀ = 9.2 ± 0.4 μg/ml, IC₅₀ = 27.4 ± 0.1 μg/ml, respectively). The SFE with 8% ethanol as a co-solvent produced extracts with distinguished increase in the antioxidant activity. The Soxhlet extract with ethyl acetate exhibited a strong reduction of lipid peroxidation (IC₅₀ = 209 ± 3 μg/ml) value comparable to the standard rutin (IC₅₀ = 203 ± 2 μg/ml). The results indicate that extracts of C. verbenacea have important potential as a source of bioactive compounds with antioxidant activity.     

Ultrasonic extraction of ferulic acid from Ligusticum chuanxiong

The extraction of ferulic acid, a pharmacologically active ingredient from the root of Ligusticum chuanxiong, with ultrasonic extraction was investigated. Percolation and supercritical fluid extraction (SFE) were employed to make comparisons with ultrasonic extraction. Three variables, which included the concentration of solvent, the ratio of solvent volume/sample (mL/g), and extraction time, were found to have a great influence on ultrasonic extraction. The optimum extraction was with pure ethanol with a solvent volume/sample ratio 8:1 (mL/g) and extraction time of 30 min. Under the optimum extraction conditions, the extraction yield could reach 8.8% which was higher than that using SFE with ethanol as co-solvent and a content of ferulic acid of 0.7%; both the yield and the content were higher than those obtained by percolation. Ultrasonic extraction significantly shortened the time required for the extraction process. Overall, ultrasonic extraction was shown to be highly efficient in the extraction of ferulic acid from Ligusticum chuanxiong.     

Optimum Extraction of Osthole and Imperatorin from the Fruits of Cnidium monnieri (L.) Cusson by Supercritical Fluid

This work describes an optimized supercritical fluid extraction (SFE) process of osthole and imperatorin from the fruits of Cnidium monnieri (L.) Cusson. Response surface methodology (RSM) was employed to evaluate the effects of independent variables on the yields of osthole and imperatorin. The independent variables were extraction pressure (100 to 300 bar), temperature (50 to 70°C), and ethanol flow rate (0 to 0.4 mL/min). Results indicated that the data could be well fitted to two second-order polynomial models in the selected experimental domain. The effects of independent variables of pressure, temperature, ethanol rate flow, as well as the interactions between pressure and ethanol flow rate, and between temperature and ethanol flow rate on the extraction yield were significant for osthole or imperatorin extraction (p < 0.05). The models predicted that the optimized conditions were 243 bar, 64°C, ethanol flow rate of 0.28 mL/min, giving maximum yields of 1.29% and 0.53% for osthole and imperatorin, respectively, that were in good agreement with the experimental values. The extraction yields obtained with the optimized SFE and Soxhlet extraction were compared.     

Brazilian Ginseng extraction via LPSE and SFE: Global yields, extraction kinetics, chemical composition and antioxidant activity

Brazilian Ginseng extracts of two species, Pfaffia paniculata and Pfaffia glomerata, were obtained by supercritical fluid extraction (SFE) with CO₂ and by low-pressure solvent extraction (LPSE) with methanol, hexane and ethanol. The SFE assays were conducted at pressures of 100, 200 and 300 bar, and temperatures of 30 and 50 °C. The qualitative chemical compositions of the extracts were determined by thin layer chromatography (TLC). One of the active principles of interest from P. glomerata extract, β-ecdysone, was identified and quantified by HPLC. The antioxidant activities of Brazilian Ginseng extracts were determined by the coupled reaction of linolenic acid and β-carotene. For P. paniculata, the highest SFE yield was obtained at 200 bar/50 °C (0.22%, dry basis—d.b.), while the best extraction condition for P. glomerata was obtained at 200 bar/30 °C (0.18%, d.b.). The higher extract yields obtained by LPSE were 2.0% and 5.8% (w/w, d.b.) for P. paniculata and P. glomerata, respectively, both obtained with methanol as extraction solvent. From the overall extraction curve of P. glomerata, it was possible to obtain the kinetic parameters of extraction; the duration of the CER (constant extraction rate) period was determined as 134 min. The TLC plates showed the possible presence of flavonoids in the ethanolic extract for both Pfaffia species. The antioxidant activity analysis detected that LPSE extracts had higher activity than SFE extracts.     

不同亚临界溶剂从微拟球藻湿藻泥中提取油脂

以微拟球藻(Nannochlorsis sp.)湿藻泥为原料,研究了亚临界乙醇、亚临界乙醇-正己烷共溶剂及硫酸辅助亚临界乙醇-正己烷共溶剂3种萃取体系对微藻油脂提取的影响. 结果表明,亚临界乙醇-正己烷比亚临界乙醇对湿藻细胞有更高的油脂萃取率和低的溶剂用量,加入少量硫酸可进一步提高油脂的提取率、降低溶剂用量. 微拟球藻湿藻泥(含水约70%)优化提取条件为,正己烷/乙醇体积比3:1,液固比(溶剂/藻细胞干重)7 mL/g,加入藻细胞干重6%的硫酸,1.5 MPa下90℃萃取30 min,在此条件下油脂提取率可达90%以上. 3种萃取体系获得的微藻油脂均以甘油三酯为主,甘油三酯的脂肪酸主要为C16:0, C18:1和C16:1,其中硫酸辅助亚临界共溶剂萃取的微藻油脂中甘油三酯含量最高,约占总脂的86%以上.