This study investigates the ascorbic acid (AA) assay, that is a cell-free method commonly used for measuring the oxidative potential (OP) of atmospheric aerosol, as a biologically relevant metric for assessing PM adverse effects to human health mediated by oxidative stress. This assay uses AA, the most abundant physiological antioxidant, as a simplified model of the synthetic respiratory tract lining fluid [1]. The conventional procedure is based on spectrophotometric measures of the depletion rate of ascorbate oxidized by redox-active species to produce reactive oxygen species (ROS). Different assay experimental conditions are investigated to more closely in vitro reproduce the complexity of particle−lung interactions, in particular the composition of antioxidant solution was varied to better represent surrogate lung fluid (SLF). In addition, an alternative method is proposed, that measures the generation rate of hydrogen peroxide as a reaction product. It uses 4-Nitrophenyl Boronic Acid (4-NPBA) as a selective probe for H2O2, since it is stechiometrically converted into 4-nitrophenol, which can be spectrophotometrically quantified by measuring light absorption at 405 nm [2]. A comparative investigation of the performances of the two assays was performed with standard solutions of the individual redox-active species commonly found in ambient PM, that were found the most efficient species in oxidizing AA, such as copper and 1,2- naphthoquinone. A good agreement was found with complementary data obtained by following AA depletion and H2O2 production: the sum of the two species is approximately the same (~80%) as the initial concentration of AA added in the UV cuvette (100 μmol). Finally, the obtained results were associated with responses of biomolecular tests for investigating oxidative cellular damage promoted by specific PM redox-active components on skin tissue cells, as the human organ with the greatest surface and the major route by which organisms enter in contact with air pollutants [3]. Preliminary results of cellular tests show that the treatment with standard solutions of the individual redox-active species modifies the gene expression of the inflammatory markers, such as Interleukin 6 and Interleukin 6. In addition, after 24 hours of treatment, the expression of specific genes involved in the redox response was dysregulated, such as superoxide dismutase 1, Glutathione peroxidase and glutathione reductase.

Ascorbic acid as molecular model for measuring oxidative potential of atmospheric aerosol in a-cellular assays

Maria Chiara Pietrogrande
Primo
Writing – Original Draft Preparation
;
BERTOLI, ILARIA
Writing – Original Draft Preparation
;
Franco Cervellati
Writing – Original Draft Preparation
;
RUSSO, Mara
Writing – Original Draft Preparation
;
Francesco Manarini
Writing – Original Draft Preparation
;
Giuseppe Valacchi
Ultimo
Writing – Original Draft Preparation
2018

Abstract

This study investigates the ascorbic acid (AA) assay, that is a cell-free method commonly used for measuring the oxidative potential (OP) of atmospheric aerosol, as a biologically relevant metric for assessing PM adverse effects to human health mediated by oxidative stress. This assay uses AA, the most abundant physiological antioxidant, as a simplified model of the synthetic respiratory tract lining fluid [1]. The conventional procedure is based on spectrophotometric measures of the depletion rate of ascorbate oxidized by redox-active species to produce reactive oxygen species (ROS). Different assay experimental conditions are investigated to more closely in vitro reproduce the complexity of particle−lung interactions, in particular the composition of antioxidant solution was varied to better represent surrogate lung fluid (SLF). In addition, an alternative method is proposed, that measures the generation rate of hydrogen peroxide as a reaction product. It uses 4-Nitrophenyl Boronic Acid (4-NPBA) as a selective probe for H2O2, since it is stechiometrically converted into 4-nitrophenol, which can be spectrophotometrically quantified by measuring light absorption at 405 nm [2]. A comparative investigation of the performances of the two assays was performed with standard solutions of the individual redox-active species commonly found in ambient PM, that were found the most efficient species in oxidizing AA, such as copper and 1,2- naphthoquinone. A good agreement was found with complementary data obtained by following AA depletion and H2O2 production: the sum of the two species is approximately the same (~80%) as the initial concentration of AA added in the UV cuvette (100 μmol). Finally, the obtained results were associated with responses of biomolecular tests for investigating oxidative cellular damage promoted by specific PM redox-active components on skin tissue cells, as the human organ with the greatest surface and the major route by which organisms enter in contact with air pollutants [3]. Preliminary results of cellular tests show that the treatment with standard solutions of the individual redox-active species modifies the gene expression of the inflammatory markers, such as Interleukin 6 and Interleukin 6. In addition, after 24 hours of treatment, the expression of specific genes involved in the redox response was dysregulated, such as superoxide dismutase 1, Glutathione peroxidase and glutathione reductase.
atmospheric aerosol, oxidative potential, a-cellular assays, ascorbic acid
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11392/2398324
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