Whether cleaning our homes or manufacturing edible oils, solvents are a crucial part of modern life. Yet, toxic organic solvents pose risks to both human health and the environment. The advent of green chemistry led chemical producers to rethink solvents, opting instead for natural sources. However, replacing organic solvents comes with challenges. Natural solvents are often too expensive or inefficient as a synthesis medium.
Despite the barriers, researchers in academia and industry are finding ways to replace traditional extractants, such as ethyl acetate, petroleum ether, methanol, ethanol, and chloroform. Fermentation now serves as a greener means of producing once petrochemically sourced solvents. And enzymatic synthesis has become commonplace as a means of avoiding some hazardous organic solvents. Deep eutectic solvents (DES) also show potential as organic solvent replacements.
"These solvents are better for the environment because the chemicals are tame and the solution uses less water," says Thilini Dissanayake, PhD student under the supervision of Nandika Bandara at the University of Manitoba in Winnipeg, Canada. "They do not require extreme temperatures or pH that would negatively affect a protein. In our lab, the benefits to the environment and protein function are the main reasons we are interested in this type of extraction."
DES are a combination of renewably sourced components, many composed entirely of plant metabolites (such as ammonium salts, sugars, and organic acids). They have proven successful in metal plating industries; however, early attempts by researchers to use them in oilseed extraction produced insufficient results with some oil retained in the solvent. The undesirable addition of organic solvents were ultimately required to achieve sufficient extraction yields (https://doi.org/10.1016/j.fuproc.2017.02.004).
Now, researchers are giving DES a second look to determine how these environmentally friendly solvents can be valuable in food chemistry. Reports indicate DES has a higher extraction yield for polyphenols, chitin, pectin, lipids, and proteins compared to conventional solvents (https://doi.org/10.1016/j.foodchem.2022.135079). Is there a future for large-scale commercialization of DES in food chemistry?
A NEW KIND OF SOLVENT
DES are a class of ionic liquid (IL) analogues, sharing similar characteristics and properties. While IL form from systems composed primarily of one type of discrete anion and cation. By contrast, DES are systems formed from a eutectic mixture of Lewis or Brønsted acids and bases which contain a variety of anionic/cationic species. Although the physical properties of DES mimic those of IL, their chemical properties are significantly different, conducive to a variety of applications.
In the last three decades, potential for new chemical technologies accelerated IL research. However, DES research is still in its infancy with the first paper on the subject only published in 2001. The two areas of focus for DES studies in that timeframe were metal processing and as a synthesis media. DES has become known as an environmentally benign alternative for synthesis, especially for metals that require hazardous means for plating or processing.
DES contain large, nonsymmetric ions with low lattice energy and hence low melting points. They usually form through a complex of quaternary ammonium salt with another metal salt or a hydrogen bond donor (HBD). The charge delocalization occurring through hydrogen bonding between, for example a halide ion and the hydrogen-donor moiety, is responsible for the melting point decrease of the mixture relative to the melting points of the individual components. The term DES refers to liquids close to the eutectic composition of the mixtures, where the molar ratio of the components gives the lowest melting point.
The difference in the freezing point at the eutectic composition of a binary mixture compared to that of a theoretical ideal mixture, ΔTf, is related to the magnitude of the interaction between A and B. The larger the interaction, the larger ΔTf (Figure 2).
DES are classified according to the nature of the complexing agent. Type III eutectics, formed from choline chloride and hydrogen bond donors, solvate a wide range of transition metal species making them a popular research subject. The liquids are simple to prepare, low cost, and biodegradable. The wide range of hydrogen bond donors available also means that this class of DES is particularly versatile. The liquid’s physical properties are easily tailored for specific applications. This class of DES has achieved popularity for applications in glycerol removal from biodiesel, metal oxide processing, and cellulose derivative synthesis, among other industrial uses.
In the last 5 years, more researchers have investigated DES in food chemistry applications. The solvents improve micro- and macromolecule extraction, including phenolic compounds, as well as proteins, carbohydrates, and lipids (https://doi.org/10.1016/j.foodchem.2022.135079). In addition, studies have shown food contamination from pesticides and packaging are easier to detect using DES.
EXTRACTING PROTEINS
Dissanayake earned a master’s degree studying alternatives to plastic packaging (https://news.umanitoba.ca/the-end-ofplastic-pollution-starts-in-a-field/). She used protein extracted from canola meal as the basis of her new packaging material. For her PhD proposal, she plans to use the proteins for encapsulation and requires a protein source whose functional properties are not compromised by extraction.
"I can tell from past experiments done with mung bean and hemp protein that properties like gelation are really good with this extraction method compared to other solvent extraction methods," says Dissanayake.
Other researcher groups have reported favorable extraction yields with DES, in addition to gaining the benefit of an extracted protein with better physical properties. A team of researchers in China reported a 10 percent increase in extraction yield over traditional methods when recovering soy protein using DES. The extracted protein exhibited higher hydrophobicity and heat resistance compared to natural soy (https://doi.org/10.1021/acssuschemeng.1c01848). A research team at Dalhousie University in Halifax, Nova Scotia, Canada used DES to extract protein from bamboo shoots and biorefinery processing waste. They reported an extraction yield 60 percent higher than the alkaline method and demonstrated the use of raw biomasses as a renewable protein source (https://doi.org/10.1007/s13399-020-00614-3).
The list of articles about protein extraction using DES in the scientific literature continues to grow. DES extract studies generally report mild conditions and a successful recovery method for renewable sources of proteins. However, more research is needed to understand how to systematize their use.
Nandika Bandara, assistant professor, Canada Research Chair in the Department of Food and Human Nutritional Sciences, and Dissanayake’s advisor sees potential for using DES as a novel, sustainable extraction technology for developing functionally superior protein ingredients. His lab is conducting experiments to build the knowledge to identify which solvent parameters perform best for a given protein.
"There are many factors that impact this extraction method. The molar ratio between the two solvents and the percentage of water is important," says Dissanayake. "We need more research to determine what conditions provide effective, efficient protein extractions."
EXTENDING SELF-LIFE
Another area where DES has shown potential is lipid extraction. Currently, solvent mixtures of methanol with chloroform or hexane are used to extract lipophilic metabolites. These are hazardous solvents that are unsustainable for the environment, and scientists have spent the last decade searching for alternatives. Studies evaluating DES’s effectiveness in the area have uncovered an unexpected, intriguing result.
When separate research teams performed carotenoid ultrasonic- and microwave-assisted extraction in DES from sources like apricot and pumpkin pulp they discovered they could retrieve more of the compounds. Moreover, when the carotenoids were kept in the dark in the DES, the compounds degraded by only 7.3 percent during 6 months of storage (https://doi.org/10.1016/j.ultsonch.2021.105638).
In a different paper, astaxanthin extracted with thymol and oleic acid had exceptional stability, with 40 percent of the initial astaxanthin quantity remaining intact after 13.5 hours, compared to all other samples that exhibited entire astaxanthin degradation (10.1016/j.foodchem.2022.132156). This outcome could be explained by the fact that thymol has more antioxidant activity than geraniol and menthol.
Researchers have proposed that the stability of bioactive compounds in DES relates to the hydrogen bonding interacin tions between solutes and solvent molecules. In addition, the neutral pH of DES may affect the stability of bioactive molecules. This is an aspect of DES that deserves to be explored.
ELIMINATING CONTAMINANTS
Ensuring food safety starts with detecting food contamination. Scientists are realizing the advantages of using DES for the separation and preconcentration of organic and inorganic species from water and food samples through microextraction processes.
However, toxic organic solvents, like halogenated hydrocarbons, have typically been used for solid-phase and liquid–liquid microextraction. Due to their better dissolving capacity, DES could replace common solvents in microextraction.
One food contaminant of relevance to the oilseed industry is 3-Monochloropropanediol esters (3-MCPDE). They belong to the chloropropanol group and contain one or two fatty acids at the sn-1 and sn-2 positions of the glycerol backbone. They are produced during the refinery and deodorization of edible oils by both inorganic and organic chlorines that are naturally present in the edible oils.
The majority of 3-MCPDE are enzymatically hydrolyzed in the intestinal tract, releasing 3-MCPD that are then absorbed into the body. The German Federal Institute for Risk Assessment concluded that the current exposure levels in infant formula and margarine may be hazardous to human health. Infants in particular ingest 20–30 times higher amounts of 3-MCPDE than the recommended total daily intake. This is a concern since in vivo studies found 3-MCPD initiate a range of toxicological effects in the brain, heart, kidney and reproductive system. According to one study, the testis and kidneys are the primary organs affected by 3-MCPD in animal subjects.
Researchers have been working on mitigation strategies for 3-MCPDE since 2009. Current tactics include optimizing oil degumming, bleaching clays, deodorization conditions and other post-refining steps. However, these approaches are time consuming and require expensive chemicals. Recently, a team of researchers lead by Adeeb Hayyan at the University of Malaya, in Malaysia, evaluated a cost-effective approach using enzymes and DES (https://doi.org/10.1016/j.bej.2023.108958).
Instead of expensive chemicals, triacylglycerol ester bonds can be hydrolyzed by enzymes that catalyze a variety of esterification and transesterification reactions. For example, Candida rugosa lipase (CRL) for the conversion of 3-MCPDE into free 3-MCPD. Hayyan’s team used DES as a support for the enzyme to remove free 3-MCPD from extra virgin olive oil and found that it enhanced the enzymatic activity.
The team reported that their proposed enzymatic method for removing 3-MCPD in edible oils was high efficiency with acceptable levels of FFA, peroxide, acid, saponification and moisture. After optimizing the reaction temperature, reaction time and solvent amount further improved the efficiency of the method. FTIR and XPS spectra demonstrated 3-MCPD adsorption onto the surface bound enzyme. They conclude that their study provides a reliable and cost-effective method for the removal of 3-MCPD in edible oils without compromising product quality.
To date, DES are used in pesticide detection, food additives and colorant extraction, as well as for determining if packaging compounds leach into food. The published literature indicates that using DES for detecting food contaminants is simple, speedy, precise, environmentally friendly and cost-effective.
LOOKING AHEAD
Dissanayake says deep eutectic solvents have a future in the food industry, but first more fundamental research needs to take place. One exciting feature of this protein extraction method is the odorless, colorless protein it produces. The versatility that DES provides in terms of tuning the protein extraction to specific protein properties has piqued the interest of food ingredient companies.
The technique does have some limitations, though. There is no universal approach for choosing DES depending on the intended use, and not all DES are appropriate for protein extraction. As a result, it takes a lot of time and effort to search for optimal DES that make extracting proteins from a particular biomass easier. Temperature impacts the conductivity and viscosity of DES. Overall, these factors may pose scaling and practicality obstacles that deter industrial application.
"I would say the biggest advantage of this method is that the preparation is simple," says Dissanayake. However, there is currently insufficient research data to prove this method is useful beyond the benchtop. Solution parameters will be different for each type of protein undergoing extraction. The proper DES composition must also be determined before extracting from a different protein source, like raw beans versus meal.
Dissanayake says that with more optimization studies food scientists will build the foundational knowledge needed to speed up the DES composition selection process. She says, that once this baseline is established it will give the industry the confidence to invest in these sustainable solvents.
Some of the background information for this article contains excerpts from a paper published in Chemical Reviews (https://doi.org/10.1021/cr300162p) and is republished here in accordance with the Creative Common (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
About the Author
Rebecca Guenard is the editor-in-chief of INFORM magazine. She can be contacted at rebecca.guenard@aocs.org
