Involvement of Deep Eutectic Solvents in Extraction by Molecularly Imprinted Polymers—A Minireview

Substantial research activity has been focused on new modes of extraction and refining processes during the last decades. In this field, coverage of the recovery of bioactive compounds and the role of green solvents such as deep eutectic solvents (DESs) also gradually increases. A specific field of DESs involvement is represented by molecularly imprinted polymers (MIPs). The current state and prospects of implementing DESs in MIPs chemistry are, based on the accumulated experimental data so far, evaluated and discussed in this minireview.


Introduction
Green chemistry and technologies related to it contribute to the improvement of the environment, and also provide a significant economic impact. Remarkable progress has been achieved, mainly in the area of seeking new methods of obtaining chemicals, in particular phytochemicals from plant materials, from natural renewable resources and from waste matter. The goal is to isolate the target compounds or substances selectively, and at the same time eliminate and remove undesirable by-products. Phytochemicals are part of a broad and diverse group of chemical compounds, classified according to their chemical structures and functional properties. As typical representatives, polyphenols, terpenes, amino acids, and proteins can be mentioned [1].
In order to extract desired substances, extraction techniques are currently used-among which the most widely utilized are Soxhlet extraction, accelerated solvent extraction, ultrasound-assisted extraction, microwave-assisted extraction and supercritical fluid extraction [2]. Target chemical compounds differ in their polarity, stability and physical properties, thus rendering a single-step extraction with one solvent for all the compounds from real plant materials generally impossible. To extract, separate and purify the desired substances, several organic solvents are commonly utilized. However, they are often volatile, toxic, flammable, explosive, and their biodegradability is low. That is the rationale behind innovative methods of extraction and separation of analytes in natural materials, which would reduce the consumption of organic solvents, and also improve the efficiency, selectivity and kinetics of extraction. Deep eutectic solvents (DESs) are such alternative solvents.
For isolating, purifying and pre-concentrating individual target substances from primary fractions of extracts, selective sorbents are suitable. Molecularly imprinted polymers (MIPs) have already proven inorganic and organic substances from waste, and materials of biological origin [10]. Results of numerous studies have shown that the usage of "green" solvents often brings about higher extraction efficiency compared to the use of conventional solvents. In recent years, the effort of scientists and technologists has been directed to application of DESs in combination with modern extraction techniques [4,5,11], such as: ultrasound-assisted extraction (UAE) [12][13][14][15], negative pressure cavitation extraction (NPC) [13], enzyme assisted extraction (EAE) [3], supercritical fluid extraction (SFE) [16], microwave-assisted extraction (MAE) [9,11], microwave hydrothermal extraction [17], subcritical water extraction [18], and percolation extraction [14]. One of the most important classes of extractable target compounds is polyphenols, exhibiting antioxidant properties, radical scavenging activity, and pharmaceutical and beneficial medical effects [19]. Plant polyphenols comprise the most numerous and widespread group of natural substances isolated from materials of plant origin. Several papers focused on DESs-based extraction of polyphenols, especially flavonoids and phenolic acids from plants, such as Dictamnus albus, Foeniculum vulgare, Origanum majorana, mint, Salvia officinalis [20], Platycladi Cacumen [15], Sophora japonica [21], and others [4,5]. Duan et al. [22] [23] investigated the extraction of protocatechuic acid, catechins, epicatechin and caffeic acid from Trachycarpus fortune using DESs as the extraction medium. In order to prepare the DESs, ChCl was mixed with ethylene glycol, glycerol, xylitol, phenol, formic acid, citric acid, oxalic acid and malonic acid. Being environmentally friendly, with low vapor pressure, non-flammability and good thermal stability, DESs proved their high potential for the extraction and purification of polyphenols. The highest extraction yield of protocatechuic acid and epicatechin was achieved using a mixture of ChCl and formic acid in a 1:1 molar ratio at the extraction temperature of 40 • C in a 6-h procedure [23]. Jeong et al. [24] tested 26 DESs, including 9 betaine-based DESs, 8 containing citric acid and 9 containing glycerols, in the process of extraction of catechin from green tea Camellia sinensis. Their results have shown that the mixture containing betaine, glycerol and glucose in molar ratio of 4:20:1 and a pH of 7.16 was the most suitable in comparison with other organic solvents. Škulcová et al. [25] applied various types of DESs to extract compounds from spruce bark (Picea abies). The overall content of polyphenols was determined using the method with Folin-Ciocalteu agent. The polyphenol content in eutectic extracts ranged from 41 to 463 mg of gallic acid equivalent to 100 g of extract. The results of extraction of particular compounds have been thoroughly described in several other publications, as well as in papers by members of the research team [4,26]. The utilization of DESs in polymer synthesis is a new and rapidly developing application area, too. DESs can be used in several phases in the processing, dissolving, extraction, synthesis, and modification of polymers. In recent years, a growing interest concerning DESs' role in the preparation of selective sorbents based on polymers with molecule-imprinted polymers, as well as the utilization of sophisticated approaches towards the molecularly imprinted polymer design, which can significantly reduce the time and cost in optimizing their production.

Selective Sorbents Based on Molecularly Imprinted Polymers
Molecularly imprinted polymers (MIPs) are synthetic tailor-made materials with a pre-defined selectivity for a template (frequently target compound), or closely related compounds for which they were designed. These materials are obtained by polymerizing functional and cross-linking monomers around a template molecule, which lead to a highly cross-linked three-dimensional network polymer ( Figure 1). Figure 1. Scheme of MIP preparation [27].
The monomers are selected according to their ability to interact with the functional groups of the template molecule. After polymerization, the template molecules are extracted/removed from the polymeric matrix and binding sites, having their shape, size, and functionalities complementary to the target molecule established. Cavities are able to specifically recognize the target molecule in complex mixtures. The resulting polymers are stable, robust and resistant to organic solvents, high temperatures, and a wide range of pH. In the most common method of preparation, monomers form a complex with a template through covalent or non-covalent interactions. The advantages of the non-covalent approach are the easy formation of the template-monomer complex, the easy removal of the templates from the polymers, fast binding of templates to MIPs and the possibility to prepare for a wide variety of compounds. MIPs are widely applied in the separation, cleaning and pre-concentration of compounds. Conventional MIPs preparation techniques include polymerization in block, precipitation, emulsion, multistep, swelling, suspension and other types of polymerization. The obtained particle size can vary from nano-to micro-particles, from irregular to spherical particles [27,28]. Despite the many advantages of MIPs-such as selectivity, sorption properties, and robustness-they also have disadvantages. When conventional techniques are used, the high quantities of organic solvents as porogens are consumed in preparation process. Water is rarely used, because it can form strong interactions with the template and/or the monomers, and thus destabilize complex formation and also interfere in the formation of specific imprinting sites. Details on the synthesis of MIPs are given elsewhere [29][30][31]. The use of DESs is an alternative and "green" strategy in MIP preparation, which can eliminate some disadvantages of traditional techniques and solvents. The relationship between DESs and MIPs can be realized in three ways: 1) the usage of a DES in a MIP preparation with the DES acting as a medium/porogen or a reactant incorporated in the MIP; 2) the use of a DES for biomass extraction with subsequent isolation of target compound(s) from the extract by a MIP; 3) the use of a DES as solvent for the extraction of target compound(s) from MIP. While there are a number of examples that meet point 1 (see Table 1), data to meet points 2 and 3 are very rare. In MIPs preparation methodologies, the DESs can be applied as medium or porogen, functional monomer [32][33][34], MIPs modifier [35,36], or MIPs The monomers are selected according to their ability to interact with the functional groups of the template molecule. After polymerization, the template molecules are extracted/removed from the polymeric matrix and binding sites, having their shape, size, and functionalities complementary to the target molecule established. Cavities are able to specifically recognize the target molecule in complex mixtures. The resulting polymers are stable, robust and resistant to organic solvents, high temperatures, and a wide range of pH. In the most common method of preparation, monomers form a complex with a template through covalent or non-covalent interactions. The advantages of the non-covalent approach are the easy formation of the template-monomer complex, the easy removal of the templates from the polymers, fast binding of templates to MIPs and the possibility to prepare for a wide variety of compounds. MIPs are widely applied in the separation, cleaning and pre-concentration of compounds. Conventional MIPs preparation techniques include polymerization in block, precipitation, emulsion, multistep, swelling, suspension and other types of polymerization. The obtained particle size can vary from nano-to micro-particles, from irregular to spherical particles [27,28]. Despite the many advantages of MIPs-such as selectivity, sorption properties, and robustness-they also have disadvantages. When conventional techniques are used, the high quantities of organic solvents as porogens are consumed in preparation process. Water is rarely used, because it can form strong interactions with the template and/or the monomers, and thus destabilize complex formation and also interfere in the formation of specific imprinting sites. Details on the synthesis of MIPs are given elsewhere [29][30][31]. The use of DESs is an alternative and "green" strategy in MIP preparation, which can eliminate some disadvantages of traditional techniques and solvents. The relationship between DESs and MIPs can be realized in three ways: 1) the usage of a DES in a MIP preparation with the DES acting as a medium/porogen or a reactant incorporated in the MIP; 2) the use of a DES for biomass extraction with subsequent isolation of target compound(s) from the extract by a MIP; 3) the use of a DES as solvent for the extraction of target compound(s) from MIP. While there are a number of examples that meet point 1 (see Table 1), data to meet points 2 and 3 are very rare. In MIPs preparation methodologies, the DESs can be applied as medium or porogen, functional monomer [32][33][34], MIPs modifier [35,36], or MIPs template [33,37]. Such systems will be abbreviated as DES-MIPs. Some authors postulated that the interaction of a DES with the functional monomer, and/or the surface of a MIP improved the affinity, selectivity and adsorption of target compounds. Such systems will be abbreviated as DES-MIPs. Many publications showed that produced DES-MIPs were suitable for the specific and selective recognition of target compounds in real samples and were characterized by stability, reusability, a high imprinting factor, fast binding kinetics, and high adsorption capacity [38][39][40]. Some authors also reported the advantages of the DES-MIPs in comparison with MIPs from conventional monomers. The advantages of DESs as monomer compared with conventional monomers are due to their high content of functional groups, allowing unique interactions with template molecules, which result in the higher affinity and selectivity of DES-MIPs. A further advantage is the higher rigidity of DES-MIPs, which can prevent their shrinkage or swelling. Moreover, the liquid phase of DES is advantageous in including the monomer in the bulk of DES or by substituting the media or solvent [36,38,41].  Chloromycetin, thiamphenicol [51] The actual researches are focused on new innovative approaches in DES-MIPs preparations. DES-MIPs were prepared on the surface of carrier material (magnetite), using ChCl and acrylic acid (1:2) as a functional monomer. This experimental approach avoids the immersion of the template during polymerization and facilitates its removal [33]. Extraction, including solid phase extraction, is a very complex process. For a better understanding of the adsorption of the adsorbate, investigation of the adsorption kinetics is useful. Moreover, the kinetic parameters are useful for designing and modeling the adsorption process, since they can provide information on the number of adsorbed molecules during the adsorption process. To investigate the kinetics of the adsorption process, the pseudo-first order, pseudo-second order models, and intraparticle diffusion model are used [38,39]. The latest results published in the last five years and evaluating the state-of-the-art of methods and technologies applied in the field of DES-MIPs utilization document a wide potential range of these systems in obtaining and/or purifying value-added substances [32,[35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51] (Table 1). Extraction methods using DES-MIPs show excellent adsorption ability and selectivity for the selection of templates or target compounds in case studies. In these studies, ChCl acts as HBA in binary or ternary ChCl-based DESs. Research groups of Li and Row [36,43] applied the ternary system containing betaine (HBA), ethylene glycol and water (1:2:1) for extraction of levofloxacin, tetracycline from millet extract [43] and levofloxacin from green bean [36]. Levofloxacin as the target compound was isolated from green been with the recovery reaching 95.2% [36]. From millet, levofloxacin (94.5%), and tetracycline (93.3%), were extracted [43]. Li and Row [42] also studied the application of ternary systems and DES containing caffeic acid:ChCl formic acid in a different molar ratio, and this system as a functional monomer in MIP synthesis. Polymeric sorbent was applied for the purification of levofloxacin. Recovery of levofloxacin for different DES-MIPs (molar ratio for DES: 1:1:0.5; 1:2:1; 1:3:1.5) ranged from 83.2% to 91.3%. Hybrid monomer γ-aminopropyltriethoxysilane-methacrylic acid (KH-550-MAA) was modified by DESs composed of ChCl and ethylene glycol, glycerol or 1,4-butanediol acting as HBD with template (target compound) rutin, scoparone, quercetin were evaluated as more effective from the viewpoint of recoveries of the target compounds compared with hybrid molecular imprinted polymers (HMIPs) modified with ionic liquids [35]. Modified by DES and ionic liquids, the HMIPs were developed for high recognition towards rutin, scoparone, and quercetin in Herba Artemisiae Scopariae. The best extraction recoveries were found for the system ChCl:glycerol (1:3)-HMIPs (rutin 92.27%; scoparone 87.51%; quercetin 80.02%). The attention of the authors of papers [50,51] was focused on milk analysis with DES-MIPs. A molar ratio of 1:1 of a mixture of ChCl:ethylene glycol, ChCl:glycerol, or ChCl:propylene glycol [50] or a mixture of ChCl:glycerol (1:2, n/n) [51], with template chloramphenicol, were used in DES-MIPs preparation for milk analysis. These sorbents were applied for extraction of chloromycetin (CHL) and thiamphenicol (THI), which are still used illegally in some animals intended for food production all over the world. DES-MIPs in dispersive liquid-liquid microextraction or in solid-phase extraction show higher recoveries for both analytes/templates (87.23% for CHL; 83.17% for THI /91.23% for CHL; 87.02% for THI, respectively) than MIPs prepared without DES.
The purification of hawthorn extract was achieved by solid-phase extraction process, and SPE recoveries of chlorogenic acid were 72.56%, 64.79%, 69.34% and 60.08% by DES-MIPs, DES-NIPs, MIPs and NIPs, respectively [32]. Non imprinted polymers (NIPs) are synthesized and treated under the same conditions but without the addition of the template. The results showed that strategy of modification of different systems (MIPs) by DES led to improving the properties of polymers due to the controlled morphology and homogeneity of the binding sites. In addition to the possibilities offered by MIPs and DES-MIPs, invention of a new class of MIPs-magnetic MIP (further on MMIPs)-has opened and expanded the possibilities of extraction, isolation and analysis of the desired compounds from materials of biological origin as well as separation of MIPs from reaction systems. DES-MMIPs can contain various magnetic parts as carrier, such as Fe 3 O 4 @3-aminopropyltriethoxysilane [37], Fe 3 O 4 @SiO 2 [38]; Fe 3 O 4 @MoS 2 [39]; Fe 3 O 4 @acrylic acid [40]; Fe 3 O 4 [48]; Fe 3 O 4 @SiO 2 [49]. MIPs are imprinted on the surface of magnetic parts and have usually a core-shell structure, of which the magnetic phase is the core and the polymeric phase acts as the shell [31]. One of the advantages of MMIPs lies in the fact that after the extraction or elution, particles can be easily separated using an external magnetic field rather than centrifugation or filtration. Fu et al. [39] have renewed and reinforced the interest in the recovery of proteins such as β-lactoglobulin, albumin, conalbumin from milk. The resulting magnetic polymer poly(ChCl-acrylic acid)Fe 3 O 4 @MoS 2 was in the form of nanospheres. It was characterized by good thermal stability at room temperature, and good adsorption capacity and selectivity for β-lactoglobulin. The strong antibacterial activity of this material was confirmed vs. S. aureus, E. coil and B. subtilis. In the case study [38], MMIPs containing Fe 3 O 4 @SiO 2 were used. The MIP layer was produced using methacrylic acid (MAA) as a monomer, with the following DESs as porogens:ChCl:urea (DES1); ChCl:ethylene glycol (DES2); ChCl:1,4-butanediol (DES3) and ChCl: glycerol (DES4). Ethylene glycol dimethacrylate (EGDMA) acted as the crosslinker, and 2,2-azobisisobutyronitrile (AIBN) as the initiator. The template role was performed by template I: tanshinone I, tanshinone IIA, or cryptotanshinone; template II: glycitein, genistein, or daidzein; template III: epicatechin, epigallocatechin gallate, or epicatechin gallate. In this study, systems for the extraction of substances from different substrates (Salvia miltiorrhiza bunge, Glycine max (Linn.) Merr and green tea) by non-DES-MNIP, non-DES-MMIP, DES1-MMIP, DES2-MMIP, DES3-MMIP, DES4-MMIP were compared. The system DES4-MMIP showed the best extraction ability for various substances and substrate. Extraction recoveries reached 85.57% for tanshinone I, 80.58% for tanshinone IIA, 92.12% for cryptotanshinone, 81.65% for glycitein, 87.72% for genistein, 92.24% for daidzein, 86.43% for epicatechin, 80.92% for epigallocatechin gallate, and 93.64% for epicatechin gallate. Furthermore, it was observed that DES-containing polymers showed higher selectivity for nine targets than that of systems without the DESs (for both MMIP and MNIP). It was documented that the MMIPs modified by DES are an innovative approach for the extraction of target substances with a higher selectivity and efficiency in the extraction of target compounds. These results of selective recognition and higher recoveries of polysaccharides were also confirmed in another study using seaweed as a substrate [37]. Taking Fe 3 O 4 @3-aminopropyltriethoxysilane as a carrier; fucodain and alginic acid as templates; MMA as a monomer; EGDMA as a crosslinker, and AIBN as an initiator, the MMIPs were modified by the DESs-ChCl:ehylene glycol; ChCl:glycerol; ChCl:1,4-butanediol; ChCl:urea; ChCl:formic acid; ChCl acetic acid; and ChCl:propionic acid-were prepared [37]. The selective recognition and separation of proteins by DES-MMIPs was evaluated by Liu et al. [40]. The DES-MMIPs were produced using carrier Fe 3 O 4 @acrylic acid, ChCl and methacrylic acid as a monomer, N,N-methylenebidacrylamide as crosslinker, ammonium persulfate, N,N,N',N´-tetramethylenediamine as polymerization initiator, and bovine hemoglobin as a template. The adsorption capacity of the DES-MMIPs and DES-MNIPs with a different amount of monomer were compared, and the results suggested that DES-MIPs had approximately three times higher absorption capacity than DES-MNIPs. The extraction method using DES-MMIPs for the determination of target compounds from green tea was used with carriers Fe 3 O 4 @MIPs [48] and Fe 3 O 4 @SiO 2 [49]. In the case of Fe 3 O 4 @MIPs [48], MIPs modified by binary DESs such as ChCl:ehylene glycol; ChCl:glycerol; ChCl: 1,4-butanediol, ChCl:urea, ChCl:formic acid, ChCl:acetic acid, ChCl:propionic acid were used. The authors of the paper [49] described magnetic polymers modified by ternary DESs, namely ChCl:oxalic acid:ethylene glycol; ChCl:oxalic acid:glycerol; ChCl:oxalic acid:propylene glycol; ChCl:caffeic acid:ethylene glycol. The resulting modified polymers showed excellent adsorption ability and selectivity. The best system for the extraction of the target substances theophylline, theobromine, (+)-catechin hydrate, and caffeic acid from green tea [49] was Fe 3 O 4 -ChCl:oxalic acid:polypropylene glycol-MMIPs (5.82; 4:32; 18.36 and 3.96 mg/g, respectively). For the binary system [48], ChCl: urea, and Fe 3 O 4 @MMIPs [48], the extraction amounts of theobromine and theophylline reached 4.87 mg/g and 5.07 mg/g green tea, respectively.

Conclusions
This minireview is focused on the application of deep eutectic solvents (DESs) for the preparation of molecularly imprinted polymers (MIPs). DESs have been designed as an environmentally friendly option for the preparation of MIPs and MMIPs, since these solvents can improve the affinity and selectivity of polymers to a target substance. The actual research showed some innovative approaches in DES-MIPs, or DES-MMIPs preparations and utilization. The applications of DES in the production of MIPs are either as a medium or solvent, as functional monomers, as MIPs modifiers, or as MIPs templates in the processes of extraction, separation or purification technologies. In the production of MIPs, the employment of DESs is based on their use as functional monomers, MIPs modifiers, and MIPs templates, as well as in extraction, separation or purification procedures.
Even though the role of DESs in molecularly imprinted technology is still largely unexplored, a rapid increase in research and the implementation of results can be expected. In particular, the use of these solvents in the production of MIPs can-to a considerable extent-expand their use as a new breakthrough technology in greener separation and analytical techniques.  APVV-15-0052, APVV-19-0185, APVV-19-0174 and VEGA 1/0403/19, VEGA 1/0412/20. This article was realized also thanks to the support for infrastructure equipment provided by the Operation Program Research and Development for the project "National Center for Research and Application of renewable energy sources" (ITMS 26240120016, ITMS 26240120028), for the project "Competence center for new materials, advanced technologies and energy" (ITMS 26240220073), and for the project "University science park STU Bratislava" (ITMS 26240220084), co-financed by the European Regional Development Fund.

Acknowledgments:
The authors would like to acknowledge the financial support of the Slovak Research and Development Agency and support for infrastructure equipment provided by the Operation Program Research and Development.

Conflicts of Interest:
The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.