Implementation of plant hormone quantification in Arabidopsis using UHPLC-MS/MS
José Manuel Herrero-MartÃÂnez, MarÃÂa Vergara-Barberán, MarÃÂa Jesús Lerma-GarcÃÂa and Ernesto Simó-Alfonso
University of Valencia, Spain
: J Diagnos Tech Biomed Anal
Abstract
A methodology to synthesize functionalized monoliths with Gold Nanoparticles (AuNPs) and its employ as solid-phase extraction sorbent for pre-concentration of proteins has been developed. For this purpose, glycidyl methacrylate-co-ethylene dimethacrylate polymer was used as powder material and subsequently modified with ammonia and posterior attachment of AuNPs. The resulting material was placed in an SPE cartridge. The influence of several variables (sorbent amount, eluent composition, among others) on SPE extraction was investigated. To perform this study, BSA and Cytochrome C with different pI values (4.7 and 10, respectively), were chosen as probes solutes. Protein recovery was estimated using Bradford assay. The mean recoveries were between 80 and 99.7%. The loading capacity of sorbent and its reusability were also established. Thus, the results showed that the cartridge can be reused for ca. 20 extraction with negligible variations on the sorbent capacity. The practical applicability of this sorbent was demonstrated by processing of vegetal samples for isolation of proteins and satisfactory results were obtained by assay with SDS-PAGE. Then, the present methodology represents a fast and simple pretreatment of complex sample with small amounts of proteins.
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