Supercritical Antisolvent Fractionation Research Articles

Reply to Frański, R.; Beszterda-Buszczak, M. Comment on "Villalva et al. Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori. Antioxidants 2022, 11, 1849".

Franski and Beszterda-Buszczak [...].

Comment on Villalva et al. Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori. Antioxidants 2022, 11, 1849.

Villalva et al. evaluated the potential utility of an Achillea millefolium (yarrow) extract in the control of H. pylori infection. The agar-well diffusions bioassay was applied to determine the antimicrobial activity of yarrow extracts. The supercritical anti-solvent fractionation process of yarrow extract was made to give two different fractions with polar phenolic compounds and monoterpenes and sesquiterpenes, respectively. Phenolic compounds were identified by HPLC-ESIMS by using the accurate masses of [M-H]- ions and the characteristic product ions. However, some of the reported product ions seem to be disputable, as described below.

Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori.

The main objective of this work is to evaluate the potential utility of an Achillea millefolium extract (yarrow extract, YE) in the control of H. pylori infection. The supercritical anti-solvent fractionation (SAF) process of YE allowed the obtaining of two different fractions: yarrow’s precipitated fraction (YPF), enriched in most polar phenolic compounds (luteolin-7-O-glucoside, luteolin, and 3,5-dicaffeoylquinic acid), and yarrow’s separator fraction (YSF), enriched in monoterpenes and sesquiterpenes, mainly containing camphor, artemisia ketone, and borneol. YE was effective in reducing reactive oxygen species (ROS) production in human gastric AGS cells by 16% to 29%, depending on the H. pylori strain. YPF had the highest inhibitory activity (38–40%) for ROS production. YE modulated the inflammatory response in AGS gastric cells, decreasing IL-8 production by 53% to 64%. This IL-8 inhibition also showed a strain-dependent character. YPF and YSF exhibited similar behavior, reducing IL-8 production, suggesting that both phenolic compounds and essential oils could contribute to IL-8 inhibition. YSF showed the highest antibacterial activity against H. pylori (6.3–7.1 log CFU reduction, depending on the strain) and lower MIC (0.08 mg/mL). Results obtained have shown that YE and SAF fractions (YPF and YSF) were effective as antioxidant, anti-inflammatory, and antibacterial agents regardless of the H. pylori strain characteristics.

Supercritical antisolvent fractionation of antioxidant compounds from Lavandula luisieri (Rozeira) Riv.-Mart.

There is a renewed research interest in food industry in natural additives to improve food shelf life and provide preventive or therapeutic effects on chronic ailments. The aim of this study was to optimise the concentration of three bioactives, rosmarinic, oleanolic and ursolic acids from Lavandula luisieri ethanolic extracts using the supercritical antisolvent fractionation technique. In order to evaluate the influence of pressure and CO2 flow rate in the process, response surface methodology was employed. Actives quantification was accomplished using HPLC-Photodiode array. Rosmarinic acid, was completely retained and concentrated in the precipitation vessel, while oleanolic and ursolic acids distributed between both fractions. The supercritical antisolvent fractionation process is a useful and green technology to concentrate bioactive in a fine solid to be applied as natural preservative in food products. The optimum conditions for higher mass recovery and concentration of actives were 130 bar and CO2 feeding flow rate 30 g/min, respectively.

Supercritical fractionation of antioxidants from algerian Opuntia ficus ‐ indica (L.) Mill. seeds

The recovery of active ingredients from vegetable wastes is a very attractive research field for the development of a sustainable economy. In order to revalue the waste from the extraction of oil of Opuntia ficus-indica seeds, in this study cakes resulting from the pressing were macerated in ethanol and then a Supercritical Antisolvent Fractionation (SAF) technique was used for the recovery and enrichment of the antioxidants present in the ethanolic extract. Catechin, epicatechin, and ferulic acid, as characteristic antioxidant components, were identified and quantified by HPLC in the fractions obtained through SAF. A series of 11 SAF experiments was programed by Response Surface Methodology (RSM) to optimize the recovery of the original extract and the yield in antioxidants. Optimal SAF conditions were: 15 MPa pressure, 30 g min−1 CO2 flow rate, and 13.6 MPa, 10 g min−1 CO2 flow rate for extract mass recovery (77.96%), and epicatechin yield (3.81%), respectively. Practical applications Due to the worldwide diffusion of Opuntia ficus-indica (L.) Mill. seeds, its valorization for the recovery of bioactive compounds with potential diversified applications (food, cosmetic, pharmaceutical, and material sectors) can represent an important source of additional income for the producers allowing, at the same time, the production at reduced costs of bioactive-based products. Industrial large scale implementation of such recovery strategies requires the development of a cost-effective process. Emerging technologies based on Supercritical Antisolvent assistance has been increasingly investigated for this purpose. This work demonstrated the possibility of exploiting the SAF technique to obtain extracts remarkably enriched in valuable antioxidants from industrial wastes resulting from the obtaining of seed oils. The operation time is short and the activity of the antioxidants contained in the starting material preserved. This laboratory- model could advantageously be scaled to a acomercial plant within the framework of Green Chemistry.

Application of compressed fluid-based extraction and purification procedures to obtain astaxanthin-enriched extracts from Haematococcus pluvialis and characterization by comprehensive two-dimensional liquid chromatography coupled to mass spectrometry.

The green microalga Haematococcus pluvialis has been widely studied due to its capacity to accumulate great amounts of astaxanthin, a high-value carotenoid with biological activities. In the present work, two green compressed fluid-based processes, pressurized liquid extraction (PLE) and supercritical antisolvent fractionation (SAF), are integrated to obtain an astaxanthin-enriched extract from this microalga. PLE was carried out using pressurized ethanol as solvent, for 20min, at 10MPa, and 50°C as extraction temperature. Subsequently, the obtained extract was processed by SAF to further purify the carotenoid fraction. The SAF process was optimized using a 3-level factorial experimental design and considering three experimental variables: (i) CO2 pressure (10-30MPa), (ii) percentage of water in the PLE extract (20-50%), and (iii) PLE extract/supercritical-CO2 flow rate ratio (0.0125-0.05). Total carotenoid content was evaluated in both extracts and raffinates. Best results were obtained at 30MPa, 0.05 feed/SC-CO2 mass flow rate, and 20% (v/v) of water in the feed solution, achieving values of 120.3mgg-1 carotenoids in extract (in the SAF extract fraction), which were significantly higher than those obtained in the original PLE extract. In parallel, a new fast two-dimensional comprehensive liquid chromatography (LC×LC) method was optimized to get the full carotenoid profile of these extracts in less than 25min. This is the first time that the use of a C30 column is reported in an on-line LC×LC system. Graphical abstract.

Supercritical Carbon Dioxide Antisolvent Fractionation for the Sustainable Concentration of Lavandula luisieri (Rozeira) Riv.- Mart Antimicrobial and Antioxidant Compounds and Comparison with Its Conventional Extracts.

Lavandula stoechas subsp. luisieri is a Spanish subspecies from the Lamiaceae family. Its essential oil has been traditionally used for several medical applications though little is known about other extracts. Similar to many other studies aiming to obtain traditional plant extracts to be used in different applications, this work evaluated the antioxidant and antimicrobial activities of Lavandula luisieri extracts and the correlation with their composition. Traditional hydrodistillation and ethanolic maceration were used to obtain the essential oil and the maceration extract, respectively. A green and sustainable methodology was applied to the maceration extract that was under a Supercritical Antisolvent Fractionation process to obtain a fine solid enriched in rosmarinic acid and the terpenes oleanolic and ursolic acids. Antimicrobial activities of all extracts and pure identified compounds (rosmarinic and ursolic acids) were evaluated against five bacterial strains; Listeria monocytogenes, Enterococcus faecium, Staphylococcus aureus, Salmonella Typhimurium and Escherichia coli and were compared with the pure compounds identified, rosmarinic and ursolic acids. All strains were sensitive against L. luisieri essential oil. The solid product obtained from the supercritical process was concentrated in the identified actives compared to the maceration extract, which resulted in higher antimicrobial and DPPH scavenging activities. The supercritical sustainable process provided L. luisieri compounds, with retention of their antimicrobial and antioxidant activities, in a powder exemptof organic solvents with potential application in the clinical, food or cosmetic fields.

Supercritical antisolvent fractionation as a tool for enhancing antiproliferative activity of mango seed kernel extracts against colon cancer cells

Mango seed kernel (MSK) has demonstrated to be a valuable source of bioactive phenolic compounds with antiproliferative activity against colon cancer cell lines. In this work, a phenolic MSK extract obtained by sequential pressurized liquid extraction (PLE) was submitted to supercritical antisolvent fractionation (SAF) to attain enriched fractions with improved inhibitory effect on HT-29 colon cancer cells. SAF was optimized using a Box-Behnken experimental design and considering as main factors: pressure, percentage of water in the feed extract and feed/sc−CO2 flow rate ratio. The highest inhibitory cell proliferation effect (70.51 ± 1.14%) was obtained in the extract using 50% of water v/v in the feed and 0.0625 feed/sc−CO2 ratio at 15 MPa. A targeted phytochemical profiling by LC-q-TOF-MS/MS followed by a multivariate statistical analysis of the observed bioactivity and the chemical composition evidenced the presence of characteristic compounds which might explain the enhanced antiproliferative activity of the optimal SAF extract.

Supercritical anti-solvent fractionation of Artemisia absinthium L. conventional extracts: tracking artemetin and casticin

In the current work the Supercritical Anti-solvent Fractionation (SAF) methodology was applied to conventional extracts from Artemisia absinthium L. (wormwood). Artemetin and casticin, two compounds found in wormwood extracts and with high structural similarity, were tracked in all the experiments. A Response Surface Methodology (RSM), based on Central Composite Design (CCD) was used for both the experimental design and the results correlation. Studied variables were pressure (from 8.0–15.0 MPa) and CO2 flow rate (from 10 to 60 g·min−1), while temperature (40 °C) and feed solution flow rate (0.45 mL·min−1) were maintained constant. Overall achieved yields were around 70%, being downstream vessel yields much higher than those in the precipitation vessel. The conditions predicted to reach an optimal overall yield and fractionation were 80 MPa and 10 g·min−1 (composite desirability = 0.7443). Casticin and artemetin were mainly obtained in the downstream vessel, being this fraction more enriched in artemetin than casticin.

Supercritical anti-solvent fractionation for improving antioxidant and anti-inflammatory activities of an Achillea millefolium L. extract

Achillea millefolium L. is a plant widely used in traditional medicine. Nowadays, there is a growing concern about the study of its bioactive properties in order to develop food and nutraceutical formulations. Supercritical anti-solvent fractionation (SAF) of an A. millefollium extract was carried out to improve its antioxidant and anti-inflammatory activities. A selective precipitation of phenolic compounds was achieved in the precipitation vessel fractions, which presented an antioxidant activity twice than original extract, especially when fractionation was carried out at 10 MPa. The main phenolic components identified in this fraction were luteolin-7-O-glucoside, 3,5-dicaffeoylquinic acid, 6-hidroxyluteolin-7-O-glucoside and apigenin-7-O-glucoside. However, separator fractions presented higher anti-inflammatory activity than precipitation vessel ones, particularly at 15 MPa. This fact could be related to separator fractions enrichment in anti-inflammatory compounds, mainly camphor, artemisia ketone and borneol.Therefore, SAF produced a concentration of antioxidant and anti-inflammatory compounds that could be used as high-added valued ingredients.

Comparative Study of Green Sub- and Supercritical Processes to Obtain Carnosic Acid and Carnosol-Enriched Rosemary Extracts with in Vitro Anti-Proliferative Activity on Colon Cancer Cells

In the present work, four green processes have been compared to evaluate their potential to obtain rosemary extracts with in vitro anti-proliferative activity against two colon cancer cell lines (HT-29 and HCT116). The processes, carried out under optimal conditions, were: (1) pressurized liquid extraction (PLE, using an hydroalcoholic mixture as solvent) at lab-scale; (2) Single-step supercritical fluid extraction (SFE) at pilot scale; (3) Intensified two-step sequential SFE at pilot scale; (4) Integrated PLE plus supercritical antisolvent fractionation (SAF) at pilot scale. Although higher extraction yields were achieved by using PLE (38.46% dry weight), this extract provided the lowest anti-proliferative activity with no observed cytotoxic effects at the assayed concentrations. On the other hand, extracts obtained using the PLE + SAF process provided the most active rosemary extracts against both colon cancer cell lines, with LC50 ranging from 11.2 to 12.4 µg/mL and from 21.8 to 31.9 µg/mL for HCT116 and HT-29, respectively. In general, active rosemary extracts were characterized by containing carnosic acid (CA) and carnosol (CS) at concentrations above 263.7 and 33.9 mg/g extract, respectively. Some distinct compounds have been identified in the SAF extracts (rosmaridiphenol and safficinolide), suggesting their possible role as additional contributors to the observed strong anti-proliferative activity of CA and CS in SAF extracts.

Perspectives on the Application of Supercritical Antisolvent Fractionation Process for the Purification of Plant Extracts: Effects of Operating Parameters and Patent Survey

Background: A recently developed process entitled Supercritical Antisolvent Fractionation (SAF) was recently described for the fractionation of plant extract solutions using supercritical fluids to give two or more fractions containing bioactive compounds with widely differing polarities. An updated overview of SAF of natural products is presented in this article. Objective: The goal of this work is to show some perspectives about the use of supercritical CO2 (SC-CO2) applied to the fractionation of plant extract compounds, such as phenolic compounds, lipids, and carotenoids; among other components, those extracted from seeds, leaves, stems, roots and other parts of the plants are described. The main features and mechanisms of antisolvent techniques that contribute to the understanding of the fundamentals of the supercritical antisolvent fractionation (SAF) process are also described. Method: The influence of various operating parameters, such as, CO2 temperature and pressure, CO2 flow rate, among others, during SAF process on the purification efficiency and particle morphology and the main differences about the patented SAF processes are reviewed. Results: The various experimental works intend to optimize their processes through adaptations or modifications of equipments that constitute the SAF unit, as the study of process parameters. Conclusion: SAF process is a technique to produce particles with high yield and purity of bioactive compounds. On the other hand, patents developed using this type of process are very scarce, less attention being given be given to the potential of this technique to purify plant extracts with a very complex content of bioactive compounds.

Supercritical antisolvent fractionation of rosemary extracts obtained by pressurized liquid extraction to enhance their antiproliferative activity

Rosemary bioactivity is correlated with the presence of phenolic diterpenes (carnosic acid, carnosol) and phenolic acids (rosmarinic acid). In the present work, two processes, pressurized liquid extraction (PLE) and supercritical antisolvent fractionation (SAF) were integrated to separate and enrich these families of compounds. The optimization of the SAF process was carried out using a response surface methodology (RSM) with three factors: (i) CO2 pressure, (ii) percentage of water in the PLE extract and (iii) PLE extract/SC-CO2 flow ratio. The selected responses were the relative amount of each compound (rosmarinic acid (RA) and carnosic acid+carnosol (CA+CS)), total phenolic content, antioxidant activity and antiproliferative activity against HT-29 colon cancer cells. The higher CA+CS content was obtained at 100bar, 50% (v/v) of water in the feed solution and 0.025 feed/SC-CO2 mass flow ratio, achieving values of 478.1mg/g extract and 17% of cell survival after 24h of treatment.

Optimization of rutin isolation from Amaranthus paniculatus leaves by high pressure extraction and fractionation techniques

A procedure based on the application of pressurized liquid extraction (PLE) was employed to recover the major Amaranthus spp. flavonoid glycoside rutin from dried ground plant leaves. A central composite design (CCD) was used to optimize rutin extraction; three factors were considered: extraction solvent (water/ethanol ratio, from 0/100 to 100/0), extraction time (from 5 to 20min) and extraction temperature (from 50 to 200°C). Optimization of the three extraction parameters revealed that water/ethanol ratio in the solvent mixture was the most-significant factor influencing rutin yield, followed by extraction temperature. The total rutin recovery varied from 6.95 to 14.03g/kg dry matter (DM) depending on the experimental conditions tested. The optimum extraction conditions for the highest rutin content were 70/30 (v/v) water/ethanol ratio, at 188°C and 20min as static extraction time. In this case rutin recovery from the leaves (14.30g/kg DM) was 4-times higher compared to that attainable using conventional ultrasound assisted extraction (3.62g/kg DM) with methanol:ethanol mixture (90:10) as solvent. Rutin recovery from defatted and non-defatted seeds of amaranth under optimum PLE conditions was 35.3 and 41.1mg/kg DM, respectively, whereas using ultrasound assisted extraction 19.7 and 34.6mg/kg DM were obtained, respectively.Extracts obtained under optimum conditions were further fractionated by using supercritical antisolvent fractionation (SAF). Extracts of amaranth leaves containing 70% water were mixed with supercritical CO2 and separated into two fractions according to the molecular mass and solubility of the contained compounds. A central composite experimental design was applied for the determination of the best fractionation conditions to maximize rutin enrichment. Maximum rutin concentration in raffinate (22.57g/kg) was recovered at 15MPa using a feed mixture flow rate of 0.3mL/min.

Supercritical antisolvent fractionation of lignans from the ethanol extract of flaxseed

Supercritical antisolvent fractionation (SAF) was evaluated in the fractionation and concentration of a family of beneficial plant compounds called lignans from the ethanol extract of flaxseed. The amount of lignans obtained in different fractions was studied under varying pressures (10–30MPa), CO2 flow rates (5–15kgh−1) and times of treatment (60–180min) in a three-stage fractionation column with constant temperature fixed at 313, 323 and 333K. The determination of lignan content was performed by HPLC coupled with a coulometric array detector. The effects of each individual variable as well as their interactions were investigated using a full factorial design with three factors and two levels and the optimal conditions were calculated through response surface methodology. A statistically significant increase in lignan content was obtained after the SAF process; from an average initial lignan content of 1.66±0.13gL−1 it was possible to obtain a total lignan content ranging from 3.42 to 12.96gL−1. We conclude that SAF is an appropriate technique for the isolation of lignans from flaxseed.

Supercritical extraction and supercritical antisolvent fractionation of natural products from plant material: comparative results on Persea indica

Supercritical fluids (SCFs) are alternative solvents in the field of Green Chemistry that are being developed as advanced separation techniques due to their combined properties such as the penetrability of a gas and the solvent power of a liquid. These characteristics are used in the supercritical fluid extraction (SFE), where compounds of interest can be extracted after mixing a SCF as a solvent with a matrix in a pressurized vessel. Supercritical antisolvent fractionation (SAF) uses the SCF as antisolvent, allowing for the precipitation of insoluble compounds in the SCF-organic solution mixture. An updated overview of SFE and SAF of natural products is presented in this article. Additionally we compare the results of SFE, SAF and organic soxhlet extraction (OSE) of Persea indica, a Macaronesian paleoendemism with strong insecticidal components. The composition of the extracts was analyzed by HPLC-MS, showing in the SFE extracts as major components ryanodol, cinnzeylanol and alkyl-γ-lactones, with their abundance varying with the extraction conditions. On the other hand, the SAF allowed for the fractionation of a liquid solution to give a ryanodol enriched extract (31 % more ryanodol than the initial ethanolic solution). The antifeedant and postingestive effects of these extracts on Spodoptera littoralis showed that the SAF extracts were the most active followed by SFE and OSE and their different effects can be partially explained by their composition.

Supercritical methodologies applied to the production of biopesticides: a review

Supercritical technologies are new, environmentally friendly, advanced separation techniques that have attracted the attention of both industry and academy in their aspirations of producing safer products with cleaner processes. In the field of biopesticides, supercritical fluids are being used in different stages, from the extraction of active ingredients from natural matrices to the encapsulation of blends during the formulation of the final commercial pesticide. This review summarizes different supercritical processes that arise in literature comprising supercritical fluid extraction, supercritical antisolvent fractionation or extraction, supercritical assisted atomization, particle from gas saturated solutions and supercritical solvent impregnation among others. The aim of this work is to give a general view of supercritical fluids in the field of biopesticides production, optimization and formulation, emphasizing in the extraction, fractionation and encapsulation and highlighting their importance when green, solvent free processes have to be designed.

Supercritical Extraction and Separation of Antioxidants from Residues of the Wine Industry

The main objective of this research is to evaluate the possibility to use residues of the wine industry to produce antioxidant concentrates, more specifically, concentrates of gallic acid, catechin, epicatechin and resveratrol. For this purpose, Supercritical Antisolvent Fractionation (SAF) has been used. This technology allows to concentrate ina dry form the biologically active principles of interest for their direct application, providing simultaneously a uniform and suitable size of the product. The most abundant compounds recovered are cathechin (48.5 mg/Kg, epicathechin (36.3 mg/kg), gallic acid (108 mg/Kg) and resveratrol (170 mg/Kg).

Supercritical antisolvent fractionation of polyethylene simulated with multistage algorithm and SAFT equation of state: staging leads to high selectivity enhancements for light fractions

A multistage algorithm coupled with the SAFT equation of state is demonstrated on supercritical antisolvent (SAS) fractionation of a model polyethylene with ethylene and 1-hexene. Staging is found to improve the overall solvent selectivities, especially for lights removal. The degree of improvement increases with decreasing size of lights: the smaller the lights the greater the sensitivity to staging. As for the single-stage fractionation, conditions that favor high solvent densities, e.g., high pressure and low ethylene concentration, usually favor high capacities but low selectivities. The temperature effect, however, is not monotonic

Phase equilibria in polymer solutions. Block-algebra, simultaneous flash algorithm coupled with SAFT equation of state, applied to single-stage supercritical antisolvent fractionation of polyethylene

A robust but rigorous flash algorithm allows for simulating multicomponent two-phase equilibria in supercritical asymmetric solutions that contain very large and very small components, such as polymers and supercritical solvents or monomers. The approach is to solve simultaneously all the equilibrium and material balance equations using a Newton iterative procedure in the framework of block algebra. The SAFT equation of state is used to estimate the required thermodynamic properties. This approach is illustrated with an example of single-stage fractionation of polyethylene with supercritical ethylene and 1-hethylene. The calculated solvent capacities and selectivities are related to ethylene concentration, pressure, and temperature