Desarrollo e implementación de métodos electroquímicos para determinar la capacidad antioxidante de muestras orgánicas

  1. Alberto Palma López
Supervised by:
  1. Jesús Fernández Arteaga Director
  2. José Miguel Rodríguez Mellado Director
  3. Mercedes Ruiz Montoya Director

Defence university: Universidad de Huelva

Fecha de defensa: 15 May 2015

  1. Rafael Rodriguez Amaro Chair
  2. Inmaculada Giráldez Díaz Secretary
  3. Alfonso Salinas Castillo Committee member

Type: Thesis


Oxidative stress is one of the most relevant biochemical processes recently studied that arises from an imbalance between antioxidant and prooxidant substances. This phenomenon is involved in many chronic degenerative diseases: Alzheimer's, Parkinson's, atherosclerosis, cancer etc, The incorporation of natural products to the diet, such as fruits, vegetables, spices and condiments, teas and infusions, coffees, etc., guarantees the exogenous contribution of phenolic antioxidants required to establish a proper status of "redox homeostasis" in the body. In this sense, in addition to offering numerous benefits associated with anticancer and anti-inflammatory properties, they are an effective defense against the accumulation of free radicals in the body, where oxygen reactive species (ROS) play a leading role, being responsible for accelerating the process of Cellular Aging and the onset of these diseases. The in vitro antioxidant capacity of phenolic active substances present in spices and condiments, as well as in samples of tea and infusions, were determined through cyclic voitammetry (CV) and the DPPH* radical scavenging assay. Low values of oxidative potential (EPja) obtained from CV under previously optimized experimental conditions (CA0 = 5*10'4 M, pH = 10.5 and v = 0.1 V-s'1) are indicative of a higher antioxidant activity. Thus, high values of antiradical power (ARP) indicate the increase of the scavenging DPPH* radical activity. A good correlation between the results collected from both methodologies can be established, permitting to set a limiting value of the Ep>a, apart from which the prooxidant behavior of the analyzed compounds can be noticed. Taking into account the difficulty of the analysis of the EPta in mixtures with high content of antioxidants, the lack of precision of the value of ARP and the relation with a synthetic radical, the present investigation has opted for the development of a novel and promising electrochemical methodology based on the differential pulse voitammetry (DPV), as an alternative to the methods described and the rest of traditional methods, to determine the in vitro antioxidant capacity. It is a fast, reproducible, precise and low cost test to determine the radical scavenging activity (antioxidant activity) with a detailed description of the interaction mechanism between the antioxidant and the biological radicals of interest hydroperoxyl (HO2) and superoxide (Or). This methodology is based on the formation of a complex of Hg2+with HO2 and HO", which are generated during electrooxidation of H202 on mercury drop electrode (MDE) in basic medium (pH 10.51), establishing adimensional parameters for the quantification of the antioxidant capacity based on the value of the slope of the linear range obtained in the representation of the percentage of reduction of the peak area associated with the generated complex, Hg(H02)(H0), versus the amount or volume of individual antioxidant added; the inverse of the necessary amount of antioxidant vaiue to decrease by 10% the area of the peak, C10; and ultimately, the p10 parameter, defined as the inverse value of the volume of individual antioxidant or mixture of antioxidants necessary to reduce by 10% the area of peak, V10. Finally, the study of three-dimensional graphics of response surfaces obtained through the development and implementation of experimental designs of synthetic antioxidant mixtures allowed to glimpse, as well as to quantify, significant evidence of synergistic and antagonistic effects on the overall antioxidant response.