290
under alkaline conditions, with sodium carbonate. The natural deep eutectic solvent
extracts are simply diluted in water and/or methanol (1–10%) and sometimes centrifuged prior to dilution. The resulting blue coloration is spectrophotometrically
detected in the wavelength range 675–765 nm (Fig. 7.11), and the corresponding
absorbance is directly proportional to the concentration of “phenolics” (Georgantzi
et al. 2017; Chanioti and Tzia 2018; Wan Mahmood et al. 2019; Obluchinskaya
et al. 2019).
The quantification remains relative because it is expressed as standard equivalents, mostly gallic acid, but could be expressed as catechin or even phloroglucinol
equivalents (Obluchinskaya et al. 2019). Consequently, Folin-Ciocalteu assay could
not be considered as a direct titration of polyphenols in the extracts, even if obtained
results have been shown to correlate very often with quantification based on chromatographic methods in conventional media (Singleton et al. 1999). Secondly,
Folin-Ciocalteu assay is not specific toward polyphenols. It also reacts with monophenolics (which are not true polyphenols as explained in Sect. 7.2), and many other
reductive species may interfere in the assay (Singleton et al. 1999). The interference
could be of several natures: inhibiting, additive, or enhancing (Singleton et al.
1999). Especially, the components of natural deep eutectic solvent such as sugars,
citric acid, diols, etc., may be potent interferents, sometimes inducing a “positive
false.” However, only a very few works mentioned blank experiments that were carried out with “pure” natural deep eutectic solvent (Jeong et al. 2018; Chanioti and
Tzia 2018; Panić et al. 2019b).
Total flavonoids’ content in natural deep eutectic solvent extracts was also
reported utilizing AlCl 3 spectrophotometric assay, although less often than the
Folin-Ciocalteu test (Manousaki et al. 2016; Bakirtzi et al. 2016; Patsea et al. 2017,
Georgantzi et al. 2017; Athanasiadis et al. 2018a, b; Jeong et al. 2018; Yoo et al.
2018). The colorimetric assays are very often coupled to chromatographic analyses,
while some works are only based on these latter analyses.
Fortunately, (natural) deep eutectic solvent extracts are compatible with the most
popular chromatographic separation techniques (Fernández et al. 2018b). After
extraction, the deep eutectic solvent phases containing the target polyphenol analytes could be directly introduced into HPLC (Yang et al. 2016; Duan et al. 2016;
Chanioti and Tzia 2018; Wan Mahmood et al. 2019; Panić et al. 2019b; Rajha et al.
2019; El Kantar et al. 2019) or LC systems (Bajkacz and Adamek 2017; Athanasiadis
et al. 2018a, b, c). In some works, the recovery of polyphenols from (natural) deep
eutectic solvent phases has been performed through SPE, SLE, or addition of antisolvent prior to chromatography (García et al. 2016; Fu et al. 2017c; Wang et al.
2017; Cao et al. 2018c; Mamilla et al. 2019; Pal and Jadeja 2019; Tian et al. 2019).
Only the chromatographic analyses could be considered as true titration of polyphenols in extracts, allowing a clean identification along with quantification of specific
polyphenols.
In works using combination of Folin-Ciocalteu and chromatography, some discrepancies could be noted, although rough trends remain acceptable to a certain
point. For instance, the extraction of polyphenols in olive pomace was achieved in
a series of deep eutectic solvents made from mixture of choline chloride and either
L. Percevault et al.
under alkaline conditions, with sodium carbonate. The natural deep eutectic solvent
extracts are simply diluted in water and/or methanol (1–10%) and sometimes centrifuged prior to dilution. The resulting blue coloration is spectrophotometrically
detected in the wavelength range 675–765 nm (Fig. 7.11), and the corresponding
absorbance is directly proportional to the concentration of “phenolics” (Georgantzi
et al. 2017; Chanioti and Tzia 2018; Wan Mahmood et al. 2019; Obluchinskaya
et al. 2019).
The quantification remains relative because it is expressed as standard equivalents, mostly gallic acid, but could be expressed as catechin or even phloroglucinol
equivalents (Obluchinskaya et al. 2019). Consequently, Folin-Ciocalteu assay could
not be considered as a direct titration of polyphenols in the extracts, even if obtained
results have been shown to correlate very often with quantification based on chromatographic methods in conventional media (Singleton et al. 1999). Secondly,
Folin-Ciocalteu assay is not specific toward polyphenols. It also reacts with monophenolics (which are not true polyphenols as explained in Sect. 7.2), and many other
reductive species may interfere in the assay (Singleton et al. 1999). The interference
could be of several natures: inhibiting, additive, or enhancing (Singleton et al.
1999). Especially, the components of natural deep eutectic solvent such as sugars,
citric acid, diols, etc., may be potent interferents, sometimes inducing a “positive
false.” However, only a very few works mentioned blank experiments that were carried out with “pure” natural deep eutectic solvent (Jeong et al. 2018; Chanioti and
Tzia 2018; Panić et al. 2019b).
Total flavonoids’ content in natural deep eutectic solvent extracts was also
reported utilizing AlCl 3 spectrophotometric assay, although less often than the
Folin-Ciocalteu test (Manousaki et al. 2016; Bakirtzi et al. 2016; Patsea et al. 2017,
Georgantzi et al. 2017; Athanasiadis et al. 2018a, b; Jeong et al. 2018; Yoo et al.
2018). The colorimetric assays are very often coupled to chromatographic analyses,
while some works are only based on these latter analyses.
Fortunately, (natural) deep eutectic solvent extracts are compatible with the most
popular chromatographic separation techniques (Fernández et al. 2018b). After
extraction, the deep eutectic solvent phases containing the target polyphenol analytes could be directly introduced into HPLC (Yang et al. 2016; Duan et al. 2016;
Chanioti and Tzia 2018; Wan Mahmood et al. 2019; Panić et al. 2019b; Rajha et al.
2019; El Kantar et al. 2019) or LC systems (Bajkacz and Adamek 2017; Athanasiadis
et al. 2018a, b, c). In some works, the recovery of polyphenols from (natural) deep
eutectic solvent phases has been performed through SPE, SLE, or addition of antisolvent prior to chromatography (García et al. 2016; Fu et al. 2017c; Wang et al.
2017; Cao et al. 2018c; Mamilla et al. 2019; Pal and Jadeja 2019; Tian et al. 2019).
Only the chromatographic analyses could be considered as true titration of polyphenols in extracts, allowing a clean identification along with quantification of specific
polyphenols.
In works using combination of Folin-Ciocalteu and chromatography, some discrepancies could be noted, although rough trends remain acceptable to a certain
point. For instance, the extraction of polyphenols in olive pomace was achieved in
a series of deep eutectic solvents made from mixture of choline chloride and either
L. Percevault et al.