title: Performance enhanced UV/vis spectroscopic microfluidic sensor for ascorbic acid quantification in human blood.
abstract: Quantitative analysis of antioxidants in a fast, simple and accurate manner is of great importance in the view of real-time monitoring the health of individuals. Recently, we have developed a UV/vis spectroscopic microfluidic sensor to specifically quantify ascorbic acid based on the immobilization of ascorbate oxidase, a relatively unstable enzyme. In this work, three different strategies for the immobilization of the unstable enzyme, including alumina sol-gel encapsulation, physisorption to PDMS channels with, and without alumina xerogel modification, were compared to build a microsensor. We found that the loading amount of the enzyme is not the determinative factor for the performance of the microfluidic biosensor but the retained activity of the enzyme and diffusion in the microfluidic channel. Taking into account of the two factors, the protocol of adsorbing enzymes to alumina (Al2O3) xerogel modified PDMS surface was demonstrated to be the best for preparing the microfluidic sensor among the utilized protocols. The microsensor prepared under the optimized protocol was further used to quantify ascorbic acid in human blood, where only dozens of microliters of blood (few drops) was required, demonstrating its potential application in clinical diagnosis. The developed strategy is featured with optimized enzymatic activity, simple process of microfluidic platform, low sample consumption, and straightforward spectrophotometry based detection.
authors.affiliation: International Iberian Nanotechnology Laboratory (INL), Av. Mestre José Veiga, 4715-330 Braga, Portugal. Electronic address: hongyan.bi@inl.int.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Bi
authors.affiliation: INESC Microsistemas e Nanotecnologias (INESC MN), Rua Alves Redol, 9-1, 1000-029 Lisbon, Portugal, and Physics Department, Instituto Superior Técnico, Universidade de Lisboa, 1000 Lisbon, Portugal.
authors.forename: Carla M
authors.initials: CM
authors.lastname: Duarte
authors.affiliation: International Iberian Nanotechnology Laboratory (INL), Av. Mestre José Veiga, 4715-330 Braga, Portugal.
authors.forename: Marina
authors.initials: M
authors.lastname: Brito
authors.affiliation: International Iberian Nanotechnology Laboratory (INL), Av. Mestre José Veiga, 4715-330 Braga, Portugal; UCIBIO-REQUIMTE, Laboratory of Toxicology, Biological Sciences Department, Faculty of Pharmacy, University of Porto, 4050-313 Porto, Portugal.
authors.forename: Vânia
authors.initials: V
authors.lastname: Vilas-Boas
authors.affiliation: INESC Microsistemas e Nanotecnologias (INESC MN), Rua Alves Redol, 9-1, 1000-029 Lisbon, Portugal, and Physics Department, Instituto Superior Técnico, Universidade de Lisboa, 1000 Lisbon, Portugal.
authors.forename: Susana
authors.initials: S
authors.lastname: Cardoso
authors.affiliation: International Iberian Nanotechnology Laboratory (INL), Av. Mestre José Veiga, 4715-330 Braga, Portugal.
authors.forename: Paulo
authors.initials: P
authors.lastname: Freitas
mesh_terms: D000327:Adsorption; D000537:Aluminum Oxide / Q000737:chemistry; D001204:Ascorbate Oxidase / Q000737:chemistry / Q000378:metabolism; D001205:Ascorbic Acid / Q000097:blood* / Q000378:metabolism; D015374:Biosensing Techniques / Q000379:methods*; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D057230:Limit of Detection; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D044367:Phase Transition; D013056:Spectrophotometry, Ultraviolet / Q000379:methods*
chemical_list: D004129:Dimethylpolysiloxanes; D004800:Enzymes, Immobilized; C013830:baysilon; D001204:Ascorbate Oxidase; D000537:Aluminum Oxide; D001205:Ascorbic Acid
keywords: Antioxidant analysis; Human blood; PDMS microfluidic chip; Sol-gel; UV/vis spectroscopic sensor