Edited by: Alberto Valdés, Spanish National Research Council (CSIC), Spain
Reviewed by: Kristian Pastor, University of Novi Sad, Serbia; Marija Banožić, University of Osijek, Croatia
This article was submitted to Nutrition and Food Science Technology, a section of the journal Frontiers in Nutrition
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The increasing demand for medical cannabis urges the development of new and effective methods for the extraction of phytocannabinoids. Deep eutectic solvents (DESs) are an alternative to the use of hazardous organic solvents typically used in the industry. In this study, hydrophilic and hydrophobic DESs were developed based on terpenes, sugars, and natural organic acids as green extraction media for the extraction of cannabis bioactive compounds. The factors influencing the extraction of bioactive components, such as the type of DESs and extraction time, were investigated. Initial screening in hemp showed that the DES composed of Men: Lau (a 2:1-M ratio) had a greater extraction efficiency of cannabidiol (CBD) and cannabidiolic acid (CBDA) (11.07 ± 0.37 mg/g) of all the tested DESs and higher than ethanol. Besides having a higher or equivalent extraction yield as the organic solvents tested, DESs showed to be more selective, extracting fewer impurities, such as chlorophyll and waxes. These results, coupled with the non-toxic, biodegradable, low-cost, and environmentally friendly characteristics of DESs, provide strong evidence that DESs represent a better alternative to organic solvents.
Cannabis is one of the world’s oldest cultivated and widely distributed plants (
With the current changes in cannabis policies and new legislation for its medical use and cultivation, in the last 20 years, cannabis has been witnessing a revival, and, nowadays, domesticated forms of cannabis are spread and cultivated all over the world, exclusively for industrial purposes (
Cannabis is a psychoactive plant that contains more than 500 different chemical compounds, of which cannabinoids are the main constituents (
Biosynthesis of cannabinoids happens along with cannabis secondary metabolism and starts with the bond between geranyl pyrophosphate (GPP) and olivetolic acid, creating cannabigerolic acid (CBGA). From this precursor, specific enzymes derivate other cannabinoids acids as cannabidiolic acid (CBDA) and Δ9-tetrahydrocannabinol acid (THCA) that can be converted non-enzymatically into their decarboxylated forms, cannabidiol (CBD), and Δ9-tetrahydrocannabinol (THC), respectively, either by light or heat, while in storage or when combusted (
Traditionally, the extraction of cannabinoids is performed using organic solvents, including hydrocarbons (e.g., hexane) and alcohols (e.g., ethanol, methanol). This method of extraction is cheap, easy to operate, and does not require sophisticated equipment; however, the solvents used are flammable, toxic, and non-biodegradable, risking human health, besides having a huge environmental impact (
Other alternatives based on green chemistry have been pursued. Particularly, due to the drawbacks associated with existing processes, the demand for methods that have high extraction yields, low cost, with potential for scale-up production and environmentally friendly persists.
Deep eutectic solvents (DESs) are a new class of green solvents and have received great attention as extraction media. DESs, introduced in the beginning of the 21st century, are prepared by simply mixing at least one hydrogen bond acceptor (HBA) with one hydrogen bond donor (HBD) at an appropriate molar ratio to form a eutectic mixture (
The majority of DESs proposed so far are based on renewable resources, such as carboxylic and amino acids, sugars, amines, representing a new generation of green solvents (
To date, numerous articles detailing the use of these solvents as extraction (phenolic acids, flavonoids, and polyphenols) and separation media (dissolving lignocelluloses, separation of an azeotropic mixture, and solid-liquid extraction) are reported in the literature (
Publications have already shown the potential of NADES as extraction media to obtain phytocannabinoids (
The chemicals used for the preparation of DESs included Betaine (99%), DL-Menthol (≥ 95%), Nile Red (≥ 98%) purchased from Sigma, Lactic acid (85%), Lauric acid (98%), Myristic acid (≥ 98%) obtained from Sigma–Aldrich, L-proline (98%) from Scharlau, D-(+)-glucose anhydrous (≥ 95%), Stearic acid (98%) from Merck/Sigma, Ethanol (96%) from Valente and Ribeiro, Methanol PA from Honeywell, Folin-Ciocalteau Reagent from Panreac, L-Ascorbic acid (99%), Quercetin (HPLC grade), which were purchased from Sigma and were used as purchased.
Before extraction, hemp leaves and inflorescences were grinded using a commercial blender. This method allowed a reduction of the sample size and a higher contact between the DES and the plant material, resulting in an increase of the extraction efficiency, consequently, increasing the final yield. The inflorescences, leaves, and seeds were grinded in a lab mill (IKA® Tube-mill control) at 6,000 rpm for 45 s, with a particle size range between 1 mm and 180 μm. The samples were kept in an amber flask in a dark place to protect them from light, under room temperature.
Deep eutectic solvents were produced by the heating-stirring method. This method was selected since it is simple and allows the preparation of multiple DESs simultaneously, and it can be easily scaled up. DESs were obtained by mixing the HBAs and HBDs at the desired molar ratio as shown in
An overview of the tested DESs.
No. | HBA | HBD | Abbreviation | Molar ratio |
DES 1 | Betaine | L(+)-Lactic Acid | Bet:Lac | 1:2 |
DES 2 | Glucose | L(+)-Lactic Acid | Lac:Gluc | 1:5 |
DES 3 | L-Proline | L(+)-Lactic Acid | Pro:Lac | 1:1 |
DES 4 | Menthol | L(+)-Lactic Acid | Men:Lac | 2:1 |
DES 5 | Menthol | Lauric Acid | Men:Lau | 2:1 |
DES 6 | Menthol | Myristic Acid | Men:MyA | 4:1 |
DES 7 | Menthol | Stearic Acid | Men:StA | 8:1 |
Generally, the greater the intermolecular attractions, the larger the polarity. Thus, polarity is generally a solubilization property. To study NADES polarity, solvatochromic studies have been performed using Nile red. Polarity intervals can be identified by referencing against a standard.
where h is Planck’s constant, c is light speed, and λmax = wavelength of a maximum of UV absorbance.
After their synthesis, the water content of the DESs was measured using Karl Fischer (Metrohm). For each titration, ≅ 50 mg of the samples was injected. The measurements were made in triplicate.
Density, ρ, and viscosity, η, data of the prepared DESs were measured from 293.15 to 343.15 K at atmospheric pressure, with a densimeter/viscosimeter (SVM 3001 from Anton Paar). The temperature was controlled with an accuracy of ± 0.01 K.
Bioactive compounds from hemp samples were extracted by mixing the DES with the plant matrix in a solid-liquid ratio of 1:10 (W/W). After being briefly mixed, cannabinoids were extracted for 90 min in an ultrasonic bath [Model XUB5, Formatura (Type Solution)] (water temperature at 60°C; ultrasonic power, 100 W) in cycles of 15 min; a sample was collected for future kinetic analyses. Then centrifuged (Model Victor Nivo 3S, ILC) at 6,000 rpm for 15 min to separate the liquid from the solid phase. Each experiment was repeated three times for each DES, and the respective cannabinoids were quantified by HPLC.
To compare the efficiency of the DES extraction method, a Soxhlet extraction with ethanol was performed. In this method, 2 g of hemp was extracted with 70 ml of ethanol on a Soxhlet apparatus for 90 min. The resulting solutions were transferred into a weighted flask, the solvent evaporated in a rotatory evaporator and the oils obtained saved at 4°C for future studies. The extractions were carried in triplicate, and the respective cannabinoids were quantified by HPLC analyses.
The samples were prepared by diluting the obtained extracts in methanol (a 1:10 w/w ratio) and then stirred to until a homogenous solution was obtained. The solution was then filtered using a hydrophilic PTFE syringe filter with a 0.20-μm pore size (FilterLab) before analysis.
HPLC analysis of the hemp extracts was carried out using an Agilent Infinity 1100 HPLC System, and an Agilent 1100 series photodiode-array detector (DAD) for detection and recording at UV/Vis 220 nm. The cannabinoids chromatographic separations were achieved using a Kinetex C-18 column (100 mm × 4.6 mm ID and 2.6-μm particle size, 100 Å pore size). The method used for the HPLC analysis was adapted from the Cannabinoids on Raptor ARC-18 Restek LC_GN0553 methodology as described elsewhere (
As mobile phase A:0.1% Formic acid in water and B:0.1% Formic acid in acetonitrile were used, the solvent flow was kept constant at 1.5 ml/min with the following gradient profile:0.00–4.00 min 25% of solution A, 75% of solution B, 0.00–4.01 min 0% of solution A, 100% B of solution and 4.01–7.00 min 25% of solution A, 75% of solution B. The column oven was kept at 50°C during the run, and the injection volume was of 5 μL. The identification and the quantification of cannabinoids were based on CBD, CBDA external standards.
The total phenolic content (TPC) was determined for individual extracts using the Folin–Ciocalteu method and was adopted from Waterhouse A.L, with some modifications (
The calibration curve was obtained, preparing a stock solution with concentration of 5 g/L, dissolving 0.5 g of gallic acid in 10 ml of ethanol and 90 ml of distilled water. From this solution, five standard solutions obtained, with concentrations of 50, 100, 150, 250, and 500 mg/L, The five standard solutions obtained, with concentrations of 50, 100, 150, 250, and 500 mg/L, respectively, were used to acquire the calibration standard curve. Briefly, 20 μL of extract (diluted 1:10 w/w in methanol) was mixed with 1.58 ml of distilled water and 100 μL of Folin–Ciocalteu reagent; the solution was mixed and incubated at room temperature for 7 min. Then, 300 μL of a Na2CO3 solution was subsequently added to the mixture and incubated at 40°C for 30 min. Afterward, the absorbance was measured, utilizing a microplate reader (Model Victor Nivo 3S, ILC) at 750 nm.
The flavonoid content of individual extracts was measured according to Navarro J.M. et al. (
Terpene’s identification was performed by SPME/GC-MS, using the method described by Stenerson K, with adaptations (
The removal and the quantification of waxes were carried out through a process called “winterization” of oil and were applied to the obtained extracts. The analysis was carried out in triplicate. Briefly, 1–3 g of extract was dissolved in 10–30 ml of ethanol (96%), respectively, and stirred until a homogeneous solution was obtained. The solution was then left to cool down in the freezer (-20°C) to induce wax precipitation for 24 h. The samples were then centrifugated at 6,000 rpm for 15 min to separate the waxes from the solution. The solid residue was then left to evaporate the remaining ethanol overnight and then weighted to calculate the wax concentration (% W/W). The supernatant was then transferred for a volumetric balloon to evaporate the solvent by a rotary evaporator. The recovered dewaxed extracts were kept at 4°C and in darkness until analysis.
Chlorophyll (CHL) content was determined by following the method previously described by Arnon (
Results of three measurements were expressed as μg total CHL/g extract.
The antioxidant activity of the extracts was measured based on their scavenging activity of the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical. About 150 μL of the extracts was added to 4 ml of the DPPH working solution. After incubating for 40 min at room temperature, the absorbance of the preparations was taken at 517 nm by a microplate reader (Model Victor Nivo 3S, ILC). Sample antioxidant activity was compared to standard ascorbic acid concentrations (1–500 μg/ml). Then, the % inhibition was calculated by the following equation:
From the calibration curves, determined from different concentrations of the extracts, the IC50 was obtained. IC50 value denotes the concentration of the sample required to scavenge 50% of the DPPH free radicals.
The solubility of CBD and CBDA in PBS was determined
The results of cannabinoids, wax, and chlorophyll extraction where all statistically treated with Graph Pad Prism 6. To indicate statistically significant differences between means, the mean value obtained from each DESs tested was compared to the control using one-way ANOVA and a confidence interval of 95% (
Seven DESs were successfully prepared as homogenous liquids, without any crystal precipitation at normal ambient temperature, excluding for Men:StA, which is solid at room temperature. All the systems appear as a transparent liquid except for Pro:Lac, which is a yellowish liquid.
The determination of the physico-chemical properties of DESs is extremely important since they have a significant influence on the solvent properties, affecting solvents suitability for specific applications. Thus, polarity, water content, density, and viscosity of the prepared systems were determined.
Experimental data on polarity obtained using Nile red as a probe and the water content measured by the Karl Fischer titrator are shown in
Experimental water content (%) and polarity (E
DESs | Water Content (%) | E |
Bet:Lac | 6.7 ± 0.8 | 50.16 |
Lac:Gluc | 9.0 ± 0.6 | 44.67 |
Pro:Lac | 4.5 ± 0.5 | 49.58 |
Men:Lac | 2.4 ± 0.1 | 51.80 |
Men:Lau | 0.1 ± 0.01 | 53.34 |
Men:MyA | 0.1 ± 0.02 | 53.34 |
Men:StA | 0.2 ± 0.02 | – |
Nile red is a molecule whose florescence is influenced by the polarity of where it is dissolved. The polarity evaluation of Men:StA could not be assessed since this measurement is performed at room temperature at which the solvent is in the solid state, making it impossible to read on the spectrophotometer. The higher the E
The determination of density and viscosity was carried out at atmospheric pressure from 293.15 to 338.15 K. Due to their high viscosity, it has not been possible to determine the viscosity in the whole range of temperatures for all of them. Results are shown in
Experimental densities, ρ, expressed in g/cm3, of the DESs:Betaine lactic acid Bet:Lac (1:2), lactic acid glucose Lac:Gluc (5:1), proline lactic acid Pro:Lac (1:1), menthol lactic acid Men:Lac (2:1), menthol lauric acid (2:1), menthol myristic acid Men:MyA (4:1), and menthol stearic acid (8:1).
T/K | Bet:Lac (1:2) | Lac:Gluc (5:1) | Pro:Lac (1:1) | Men:Lac (2:1) | Men:Lau (2:1) | Men:MyA (4:1) | Men:StA (8:1) |
293.15 | 1.20 | 1.28 | 1.27 | 0.95 | 0.90 | 0.90 | |
298.15 | 1.20 | 1.26 | 0.89 | ||||
303.15 | 1.19 | 1.28 | 1.26 | 0.95 | 0.89 | 0.89 | |
308.15 | 1.19 | 1.26 | 0.89 | ||||
313.15 | 1.19 | 1.27 | 1.25 | 0.94 | 0.88 | 0.88 | 0.88 |
318.15 | 1.19 | 1.24 | 0.88 | 0.88 | |||
323.15 | 1.18 | 1.26 | 1.24 | 0.93 | 0.87 | 0.87 | 0.87 |
328.15 | 1.18 | 1.24 | 0.87 | 0.87 | |||
333.15 | 1.18 | 1.25 | 1.24 | 0.92 | 0.87 | 0.87 | 0.87 |
338.15 | 1.17 | 1.23 | 0.86 | ||||
343.15 | 1.17 | 1.23 | 0.86 | 0.86 |
Experimental viscosities, η, expressed in mPA/s, of the DESs:Betaine lactic acid Bet:Lac (1:2), lactic acid glucose Lac:Gluc (5:1), proline lactic acid Pro:Lac (1:1), menthol lactic acid Men:Lac (2:1), menthol lauric acid (2:1), menthol myristic acid Men:MyA (4:1).
T/K | Bet:Lac (1:2) | Lac:Gluc (5:1) | Pro:Lac (1:1) | Men:Lac (2:1) | Men:Lau (2:1) | Men:MyA (4:1) | Men:StA (8:1) |
293.15 | 818.73 | 85.25 | 46.32 | ||||
298.15 | 1266 | 32,03 | |||||
303.15 | 819 | 330.73 | 38.77 | 18.46 | 24.25 | ||
308.15 | 545 | 14.39 | |||||
313.15 | 375 | 152.88 | 20.30 | 11.42 | 13.96 | 16.61 | |
318.00 | 261 | 9.21 | 12.62 | ||||
323.15 | 190 | 79.30 | 997.4 | 11.85 | 7.54 | 8.71 | 9.81 |
328.15 | 142 | 675.1 | 6.25 | 7.77 | |||
333.15 | 108 | 45.18 | 472.4 | 7.56 | 5.25 | 5.82 | 6.27 |
338.15 | 84 | 338.9 | 4.46 | 5.14 | |||
343.15 | 66 | 246.1 | 3.83 | 4.27 |
The high viscosity of DESs can make them difficult to handle in industrial processes during processing and filtration, even though it sharply decreases when temperature increases (
The choice of the solvent, in this case the DES, is one of the most important things in solid-liquid extraction. Therefore, seven different DESs were chosen as potential candidates. The screening was carried out, fixing a 1:10 solid-liquid ratio for the extraction of bioactive compounds from
Extraction of CBD + CBDA from the hemp using different DESs. The extraction conditions were as follows: 60°C, 90 min, and a 1:10 solid-liquid ratio (w/w). The symbol * means that there are statistical differences between Men:Lau and EtOH.
As it can be seen, the extraction solvent used has a significant effect on the extraction efficiency. The extraction yield decreased with the use of hydrophilic DESs (Bet:Lac, Lac:Gluc, and Pro:Lac). Menthol-based hydrophobic system produced the highest yields of all DESs that were tested (Men:Lac, Men:Lau, Men:MyA, and Men:StA). In summary, the best result for the hydrophobic DES was obtained for Men:lau (11.07 ± 0.4 mg/g), and, for the hydrophilic ones, Pro:lac (6.1 ± 0.3 mg/g) was the best solvent. These results indicate that the interactions between DESs and target compounds affect the extraction ability of DESs. This effect could be accounted for two reasons: among other characteristics, polarity is one of the most important properties, and it is a key indicator of the DESs dissolving capability (
In this work, a soxhlet extraction with ethanol was performed both to characterize the plant and compare the extraction efficacy of this solvent with the tested DESs under the same extraction conditions. The extraction efficiency of Men-Lau (11.07 ± 0.4 mg/g) has demonstrated to exceed the performance of ethanol (8.19 ± 1.7 mg/g).
The statistical analysis of the extraction performance showed statistically significant differences between Men:Lau and ethanol (one-way ANOVA,
Vági E. et al. (
To represent the kinetic curves of the process, samples were collected along the extraction every 15 min, and the results are illustrated in
Effect of time in CBD + CBDA extraction. Extraction conditions were 60°C, a 1:10 solid-liquid ratio.
The characterization of the extracts included not only the evaluation of the content in CBD and CBDA but also the quantification of other bioactive compounds, such as the TPC, total flavonoid content, and terpene content. The presence of subproducts of extraction was also evaluated, such as wax and total chlorophyll, in order to evaluate the specificity of the extraction solvents.
Phenolic compounds are important plant constituents with redox properties responsible for antioxidant activity, and their extraction requires compatible solvents with high commercial interest. The results obtained with the Folin-Ciocalteu method allowed to quantify the TPC using the gallic acid standard as equivalent. The extraction efficiency is herein assessed through the quantification of TPC in the extract as mg GAE/g hemp. The values are derived from the calibration curve and are illustrated in
Total phenolic content related to the different solvents tested and expressed in mg GAE/g hemp (extraction conditions: 60°C, 90 min, and a 1:10 S/L ratio).
The results also demonstrate that we can have different selectivities for TPC according to the composition of the DESs. Lac:Gluc extraction conditions could also be optimized for a higher extraction yield.
The total flavonoids content (TFC) was determined following the aluminum colorimetric method, which allowed to quantify these compounds using the quercetin standard as equivalent. The results were derived from the calibration curve of quercetin and expressed in mg QE/g hemp and are illustrated in
Total flavonoid content related to the different solvents tested and expressed in mg QE/g hemp (extraction conditions: 60°C, 90 min, and a 1:10 S/L ratio).
Cannabis is composed of a wide variety of terpenes that provide therapeutic benefits and properties (
Identification of the terpenes present in DESs extracts by GC-MS/SPME.
DES | α -pinene | β -pinene | β -myrcene | β -limonene | α -cariophyllene | Caryophyllene oxide | β -Selinene | |
Bet:Lac | ✓ | – | ✓ | ✓ | ✓ | ✓ | – | ✓ |
Lac:Gluc | ✓ | – | ✓ | ✓ | ✓ | ✓ | – | ✓ |
Pro:Lac | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ |
Hemp | ✓ | ✓ | ✓ | ✓ | ✓ | ✓ | – | ✓ |
From the data presented, it was possible to observe that all the tested DESs can extract the main terpenes present in hemp. It was also visible that the main fraction of terpenes present belongs to the monoterpenes (α-Pinene, β-Pinene, β-Myrcene, and β-Limonene) and to the sesquiterpenes group (
During the extraction process, some compounds are extracted alongside the targeted compounds, such as waxes, as a result of unselective extraction media. The cannabis wax layer is easily soluble in many solvents that are used for extraction, such as supercritical CO2 and organic solvents, such as ethanol. While waxes are useful for plants, they are often an undesirable by-product of extraction, since they decrease the purity of the extracts and increase the overall cost of extraction with the addition of purification steps (
Results of wax quantification (%) of hemp extracts related to the different extraction solvents.
The results showed once again a significant difference between hydrophobic and hydrophilic DESs. Hydrophilic DESs extracted more waxes, being the highest value obtained for Pro:Lac (12.46% ± 0.2), while hydrophobic extracted the lowest, being Men:Lac the DES, which extracted the least (1.19% ± 0.05). Ethanol showed a behavior very similar to the hydrophilic DESs, having the second highest extraction value (7.75% ± 2.2). These results can strongly be correlated with the polarity of the solvents, where a higher polarity can lead to a higher interaction of the solvent with the waxes, increasing the solubility and leading to a higher extraction. Menthol-based DESs showed to be more specific as extraction media, being a better choice than ethanol, for purer extracts.
Chlorophylls are natural-occurring pigments present in all plants. During cannabinoids extraction, these subproducts are also extracted from the plant matrix, and there is the need to be removed through purification processes, increasing the overall cost of extraction. Even though some publications have reported the therapeutic benefits of CHL, in an extract that is wanted to be as pure as possible, their presence may be seen as unfavorable (
In this study, we evaluated the total CHL present in the extracts before and after winterization; the results are expressed in CHL mg/g extract and represented in
Quantification of the total chlorophyll (μg/g extract) present in the obtained hemp extracts before and after winterization.
In this work, besides the evaluation of the potential of DESs as extractants, we also studied if their presence in the final product could potentiate the bioactivity of the extracts since many articles have claimed the use of DESs for the solubilization of poorly water-soluble drugs and transdermal drug delivery. In that way, antioxidant activity and solubility were evaluated.
Antioxidants are extremely important substances that possess the ability to protect the body from damage caused by free radical-induced oxidative stress (
The IC50 values of DPPH scavenging effect of hemp extracts (mg/L).
IC50 mg/L |
||
Before winterization | After winterization | |
Bet:Lac | 337.3 ± 37.2 | 337.3 ± 37.8 |
Lac:Gluc | 226.7 ± 16.4 | 337 ± 4.5 |
Pro:Lac | 120.2 ± 10.1 | 112 ± 9.8 |
Men:Lac | 626.8 ± 19.9 | 696.7 ± 140.4 |
Men:Lau | 670.6 ± 37.1 | 851.1 ± 344 |
Men:MyA | 773. ± 122.3 | 792.3 ± 30.1 |
Men:StA | 1689.6 ± 167.1 | – |
Ascorbic Acid | 104.5 ± 4.9 |
All the hemp extracts before and after winterization showed concentration-dependent increase in radical scavenging capacity. Among all the samples before winterization, the greatest DPPH radical scavenging potency was recorded for Pro:Lac, followed by the systems Lac:Glu and Bet:Lac. The free-radical scavenging activity of Men:Lac, Men:Lau, Men:MyA, and Men:StA was very weak and did not increase much as the concentration increased. Comparing the extracts with ascorbic acid, which was used as control, the DES Pro:Lac has shown similar IC50 values. The analysis of DPPH scavenging activity results indicated that sugar and amino acid-based DES were the most effective DPPH radical scavengers, explained by the presence of bioactive compounds, such as phenolic compounds, in the extracts, which have a significant antioxidant activity. Even though Pro:Lac did not have the highest TPC and TFC content, they can be richer in other bioactives with antioxidant activity, such as terpenes, which could explain their low IC50; in this work, the quantitative analysis of these compounds was not performed (
The results of radical scavenging activity of the pure DESs showed that DESs have very little effect on the DPPH assay results, providing evidence that the effects are due to the bioactive compounds extracted.
Poor aqueous solubility of the terpenophenolic compound cannabidiol (CBD) is a major issue in the widespread use of this therapeutic molecule (
Solubility of CBD + CBDA (mg/L) in PBS at 37°C.
The statistical analysis of the solubility values showed statistically significant differences between the different solvents (one-way ANOVA,
In this study, seven DESs were developed as greener solution media for the UAE of cannabinoids and other main bioactive compounds from hemp. Moreover, DESs were evaluated not only as solvents but also as agents to improve the bioactivity of the target compounds.
The first extraction screening in hemp showed that menthol-based hydrophobic DESs showed to be more efficient in the extraction of cannabinoids. Hydrophilic systems presented low cannabinoid extractability; however, they were able to extract important antioxidant compounds, such as phenolic compounds, flavonoids, and terpenes. Lac:Gluc had the highest TPC values (7.76 ± 1.1 mg/g) and TFC among all DESs. When comparing the extraction results with ethanol, Men:Lau was the one who presented the highest yield of CBD and CBDA (11.07 ± 0.4 mg/g). These systems also proved to be more selective, reducing the extraction of undesirable compounds, such as chlorophyll and waxes.
Bioactivity assays showed that Lac:Gluc and Pro:Lac also improve the solubility of CBD and CBDA in aqueous media. Therefore, the results of this study prove that DESs are selective green solvents, with huge potential for use in industrial applications, involving the extraction of bioactive compounds, and can further enhance the bioavailability of the active components.
The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.
FT was responsible for all the laboratory work. AP, AM, and AD were responsible for supervising and review the results. All authors contributed to the article and approved the submitted version.
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.
This work has received funding from the European Union’s Horizon 2020 -European Research Council (ERC) -under grant agreement No. ERC-2016-CoG 725034. This work was also supported by the Associate Laboratory for Green Chemistry (LAQV), which is financed by national funds from FCT/MCTES (UIDB/50006/2020).