Evaluating spectrophotometric and antioxidant activity methods for phenolic compounds quantification in natural deep eutectic solvents

Abstract

Natural deep eutectic solvents (NaDES) are a new generation of sustainable solvents with unique physicochemical properties derived from their natural composition. This study addressed the accuracy of spectrophotometric analytical methods in quantifying phenolic compounds, namely chlorogenic acid, in NaDES, both as a standard and as part of more complex natural extracts. Comparative analyses of different quantification methods were conducted, using HPLC as the reference, revealing the influence of solvent components on the accuracy of results. Pearson’s correlation was evaluated between the colorimetric and chromatographic methods through the comparison of standard curves. The Folin-Ciocalteu reagent method was highlighted as the most correlated with the reference method in NaDES composed of lactic acid, while the Fast Blue BB method was noted for NaDES composed of glycerol, both with Pearson’s R values greater than 0.99. Variations in pH and solvent composition affected the sensitivity of the spectrophotometric methods, requiring a customized approach for each solvent. However, when spectrophotometric methods were applied to sunflower meal extracts, none, not even when antioxidant activity evaluation methods were used, was able to replicate the order of extraction efficiency revealed by the HPLC analysis, namely: AL:G > EtOH > CC:GL > U:GL > B:AL. The best results were achieved – correctly identifying the two best extraction solvents – when a solvent-directed method was used to analyze each extract, which resulted in extracts containing 3,075.78 μg CGA mL⁻¹ AL:G and 2,384.83 μg CGA mL⁻¹ EtOH, but also 2,002.67 μg CGA mL⁻¹ B:AL; 1,488.68 μg CGA mL⁻¹ U:GL and 579,14 μg CGA mL⁻¹ CC:GL. This study contributed to a more comprehensive understanding of the challenges associated with the spectrophotometric analysis of phenolic compounds in NaDES, highlighting the need to select the right analytical method for each different solvent formulation.