## 5 Using DES to Extract Bioactive Compounds from Other Natural Sources
Nevertheless, their combination with microextraction techniques has been also carried out in this case [45; 47; 48]. Remarkable is the work developed by [PERSON] et al. [47] in which a vortex assisted-dispersive liquid-liquid microextraction (VA-DLLME) using trioctylmethylammonium chloride (TOMAC):isoamyl alcohol in a molar ratio 1:4 was used as extractant for the evaluation of folic acid in wheat  
Figure 2: Citrus essential oil deterepation approach intensified by in situ formation of a DES between a quaternary salt and terpenoids. Reprinted from [PERSON]; [PERSON]; [PERSON]; [PERSON]; [PERSON]; [PERSON] Deep Deterpenation of Citrus Essential Oils Intensified by in Situ Formation of a Deep Eutectic Solvent in Associative Extraction. Industrial and Engineering Chemistry Research, 2020, 59 (19), 9223–9232. with permission from American Chemical Society [42]. Copyright 2020.  
flour. With this aim, 5 g of sample were initially submitted to a SLE by ammonium buffer. After that, the aqueous supernatant (1 mL) and together with 9 mL of ammonium buffer were located in a centrifuge tube and 150 mL of NADESs dissolved in 250 mL were rapidly injected. Afterwards, the mixture was vortexed during 1 min, centrifuged and the NADESs drop enriched with folic acid was diluted and injected in the HPLC-UV system for its analysis. The different parameters affecting the extraction performance such as pH, salt amount and DES volume were carefully evaluated, as well as the type of DESs. In this case, three different DESs, benzyltriethylammonium chloride (BTEAC):thymol, BTEAC:octanoic acid and TOMAC:isoamyl alcohol were tested in various molar ratios. Results showed that the ion-paring (electro-static interaction) between TOMAC and folic acid carboxylic groups were more efficient than the interactions established for the other DESs increasing the extraction efficiency. Additionally, as it was indicated by the authors, this hydrophobic DESs removes emulsifiers and reduces solvent consumption compared to hydrophilic DESs, leads to the extent of DESs efficiency in water samples without the use of emulsifier solvents and, additionally, it can eliminate ion-pairing agents needed for folic acid extraction. The combination of VA-DLLME with NADESs resulted to be a suitable approach for the evaluation of folic acid in wheat flour with LOQs in the range 1-3 mg/kg and recovery higher than 90%. Finally, it should be highlighted that, as described in previous sections, the application of NADESs for the evaluation of the discussed matrices involves the re-extraction with organic solvents on many occasions. This fact is related to the limitation of DESs to be volatilized and their incompatibility with chromatographic and detection systems which complicate the possibilities of preconcentration and the reliable evaluation of the compounds of interest. In addition, the necessity of introducing a re-extraction process reduces the green nature of the procedures using toxic organic solvents and increase the complexity of the developed approaches. In this sense, further research is essential to develop new strategies that allow solving such limitations.