Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach
The ABTS and DPPH methods are among the most popular assays of antioxidant activity determination. Attempts to adapt them to different analytes and the search for the highest values of antioxidant activity has resulted in a large variety of assay conditions to be presented in the literature, includi...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-12-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/27/1/50 |
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| author | Rafał Wołosiak Beata Drużyńska Dorota Derewiaka Małgorzata Piecyk Ewa Majewska Marta Ciecierska Elwira Worobiej Paulina Pakosz |
| author_facet | Rafał Wołosiak Beata Drużyńska Dorota Derewiaka Małgorzata Piecyk Ewa Majewska Marta Ciecierska Elwira Worobiej Paulina Pakosz |
| author_sort | Rafał Wołosiak |
| collection | DOAJ |
| description | The ABTS and DPPH methods are among the most popular assays of antioxidant activity determination. Attempts to adapt them to different analytes and the search for the highest values of antioxidant activity has resulted in a large variety of assay conditions to be presented in the literature, including the way the measurement is made. This makes it difficult to relate the results to real oxidation systems, and often makes it impossible to compare them. Such a comparison is limited in advance by the use of stable radicals that do not exist in nature and that react differently from those generated in food or in vivo. Therefore, it is important to introduce measures aimed at standardizing the conditions of the activity assay, including reaction time and several reaction environments suitable for testing different groups of compounds. In this study, we used natural antioxidants of various structures: phenolic acids, flavonoids, peptides and corresponding amino acids, ascorbic acid and α-tocopherol, and also synthetic analogues of selected compounds. The curves of dependence of the measured absorbance on the concentration of antioxidants were described, the ranges of linearity were determined, and the value of the error made when reading in various ranges of dependencies was estimated. We also determined and compared the activity values using two popular methods (IC<sub>50</sub> and TEAC), taking into account different environments and reaction times. Based on the collected data, recommendations were formulated regarding the reaction conditions adapted to the studies of individual groups of antioxidants, and unified reaction times were proposed. Taking into account the state before reaching the equilibrium of antioxidants reacting in a complex manner, this approach may introduce a simplified reference to the competing reaction that occurs in reality. |
| first_indexed | 2024-03-10T03:30:49Z |
| format | Article |
| id | doaj.art-6142cd7523f94589a1874f85e412f205 |
| institution | Directory Open Access Journal |
| issn | 1420-3049 |
| language | English |
| last_indexed | 2024-03-10T03:30:49Z |
| publishDate | 2021-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Molecules |
| spelling | doaj.art-6142cd7523f94589a1874f85e412f2052023-11-23T11:55:49ZengMDPI AGMolecules1420-30492021-12-012715010.3390/molecules27010050Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical ApproachRafał Wołosiak0Beata Drużyńska1Dorota Derewiaka2Małgorzata Piecyk3Ewa Majewska4Marta Ciecierska5Elwira Worobiej6Paulina Pakosz7Department of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandDepartment of Food Technology and Assessment, Division of Food Quality Assessment, Institute of Food Sciences, Warsaw University of Life Sciences (WULS-SGGW), 159 Nowoursynowska Street, 02-776 Warsaw, PolandThe ABTS and DPPH methods are among the most popular assays of antioxidant activity determination. Attempts to adapt them to different analytes and the search for the highest values of antioxidant activity has resulted in a large variety of assay conditions to be presented in the literature, including the way the measurement is made. This makes it difficult to relate the results to real oxidation systems, and often makes it impossible to compare them. Such a comparison is limited in advance by the use of stable radicals that do not exist in nature and that react differently from those generated in food or in vivo. Therefore, it is important to introduce measures aimed at standardizing the conditions of the activity assay, including reaction time and several reaction environments suitable for testing different groups of compounds. In this study, we used natural antioxidants of various structures: phenolic acids, flavonoids, peptides and corresponding amino acids, ascorbic acid and α-tocopherol, and also synthetic analogues of selected compounds. The curves of dependence of the measured absorbance on the concentration of antioxidants were described, the ranges of linearity were determined, and the value of the error made when reading in various ranges of dependencies was estimated. We also determined and compared the activity values using two popular methods (IC<sub>50</sub> and TEAC), taking into account different environments and reaction times. Based on the collected data, recommendations were formulated regarding the reaction conditions adapted to the studies of individual groups of antioxidants, and unified reaction times were proposed. Taking into account the state before reaching the equilibrium of antioxidants reacting in a complex manner, this approach may introduce a simplified reference to the competing reaction that occurs in reality.https://www.mdpi.com/1420-3049/27/1/50reaction solventreaction timeassay comparisonIC<sub>50</sub>TEACphenolic |
| spellingShingle | Rafał Wołosiak Beata Drużyńska Dorota Derewiaka Małgorzata Piecyk Ewa Majewska Marta Ciecierska Elwira Worobiej Paulina Pakosz Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach Molecules reaction solvent reaction time assay comparison IC<sub>50</sub> TEAC phenolic |
| title | Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach |
| title_full | Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach |
| title_fullStr | Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach |
| title_full_unstemmed | Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach |
| title_short | Verification of the Conditions for Determination of Antioxidant Activity by ABTS and DPPH Assays—A Practical Approach |
| title_sort | verification of the conditions for determination of antioxidant activity by abts and dpph assays a practical approach |
| topic | reaction solvent reaction time assay comparison IC<sub>50</sub> TEAC phenolic |
| url | https://www.mdpi.com/1420-3049/27/1/50 |
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