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Molecules 2021, 26, 1516 8 of 13 extracting oil from Osmanthus fragans flower using MAE with deep eutectic solvents and reflux extraction with ethanol [34]. The antioxidant activity results from a synergy among the different phenolic com- pounds. Because the Curcuma longa L. antioxidant activity results from different mecha- nisms, in the present study, three methods were employed (DPPH, FRAP and ABTS) to compare the two extraction methods. Among the three techniques, DPPH showed the lowest antioxidant activity. Similar results were observed previously, in the extraction of Curcuma longa oil by Soxhlet vs. supercritical fluid CO2 and ethanol [29]. As previously observed in oil extracted from seeds and flowers [29,32], with the FRAP assay a considerable difference was detected in the antioxidant activity between the samples extracted by MAE (255.66 ± 0.24 mg GAE/g) and Soxhlet (73.37 ± 0.18 mg GAE/g) (Table 5). Finally, the ABTS method is considered excellent for the evaluation of the antioxidant activity of several substances and can be applied to both liposoluble and hydrosoluble compounds. The results obtained for the ABTS test with MAE and Soxhlet, even if quite similar, showed a significant difference p < 0.05 between both techniques. Other authors showed lower values of ABTS in the extraction of Curcuma longa using supercritical fluid CO2 and ethanol [29] compared with our data. The differences in the values between the antioxidant assays could be due to the different mechanisms involved. The DPPH assay measures the ability of a substance to donate a hydrogen to the DPPH* free radical, whereas the FRAP method is based on the measurement of the reduction of the ferric ion-TPTZ complex. In the ABTS assay, the activity is based on the capacity of the Curcuma longa L. oil to decrease the amount of ABTS•+ cation radical preformed in the solution. This reducing capacity of Curcuma longa L. oil is an important indicator of their antioxidant capacity [34]. Taking into account the obtained data, it can be concluded that the MAE technique provides samples with higher TPC and this phenolic extract exhibited higher antioxidant properties compared with traditional methods. 3. Materials and Methods 3.1. Raw Materials and Chemicals Curcuma root (Curcuma longa L.) from India was purchased in a local market in Bizkaia, Spain. The root was washed in water to remove impurities and dried at 50 ± 0.05 ◦C. Then, the root was milled with a cutting mill (Retsch SM 2000, Haan, Germany) and was sieved with a 0.5 × 0.5 mm mesh. The powder obtained from the curcuma root was stored in the dark at room temperature. Ethanol (EtOH, analytical standard), ethyl acetate (C4H8O2, HPLC grade), methanol (MeOH, HPLC grade), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 6-hydroxy-2,5,7,8- tetramethylchromane-2-carboxylic acid (Trolox), bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 2,4,6-tris(2-pyridyl)-s-triazine (TPTZ) were purchased from Sigma- Aldrich (Madrid, Spain). Glacial acetic acid (CHCOOH, technical grade), sodium chlo- ride (NaCl), sodium acetate (CH3COONa), sodium hydrogen phosphate (Na2HPO4), potassium dihydrogen phosphate (KH2PO4), potassium chloride (KCl), potassium per- oxodisulphate (K2S2O8) and hydrochloric acid (HCl, 37%) were purchased from Panreac AppliChem (Barcelona, Spain). Sodium carbonate anhydrous (NaCO, general-purpose grade) was supplied by Fisher (Madrid, Spain) and Iron (III) chloride hexahydrate (FeCl3 ̇6H2O) was obtained from Acros Organics (Madrid, Spain). Gallic acid mono- hydrate (C7H6O5 ̇H2O, extra pure) and Folin-Ciocalteu reagent was obtained from Scharlau (Barcelona, Spain).PDF Image | Microwave-Assisted Extraction of Curcuma
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