title: Zirconium silicate assisted removal of residual proteins after organic solvent deproteinization of human plasma, enhancing the stability of the LC-ESI-MS response for the bioanalysis of small molecules.
abstract: An efficient blood plasma clean-up method was developed, where methanol protein precipitation was applied, followed by zirconium silicate assisted exclusion of residual proteins. A strong binding of zirconium (IV) silicate to the proteins enabled the elimination of remaining proteins after solvent deproteinization through a rapid solid-phase extraction (SPE) procedure. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI/TOF MS) was used for monitoring the proteins during clean-up practice applied to human plasma samples. The proteins were quantified by colorimetric detection using the bicinchoninic acid (BCA) assay. The presented analytical strategy resulted in the depletion of >99.6% proteins from human plasma samples. Furthermore, high-performance liquid chromatography hyphenated to diode-array and electrospray ionization mass spectrometric detection (HPLC-DAD/ESI MS) was applied for qualitative and quantitative analysis of the caffeoylquinic acids (CQAs) and their metabolites in human plasma. The procedure demonstrated high recoveries for the standard compounds spiked at different concentrations. Cynarin and chlorogenic acid were recovered in the range of 81-86% and 78-83%, respectively. Caffeic acid was extracted in the excess of 89-92%, while ferulic acid and dihydroxyhydrocinnamic acid showed a recovery of 87-91% and 92-95%, respectively. The method was partially validated in accordance with FDA-Industry Guidelines for Bioanalytical Method Validation (2001). The presented scheme improves the clean-up efficacy of the methanol deproteinization, significantly reduces the matrix effects and provides a great analytical tool for the isolation of small molecules from human plasma.
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria.
authors.forename: Shah
authors.initials: S
authors.lastname: Hussain
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Pezzei
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria; ADSI-Austrian Drug Screening Institute, Innrain 66a, 6020 Innsbruck, Austria.
authors.forename: Yüksel
authors.initials: Y
authors.lastname: Güzel
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria.
authors.forename: Matthias
authors.initials: M
authors.lastname: Rainer
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria. Electronic address: Christian.W.Huck@uibk.ac.at.
authors.forename: Christian W
authors.initials: CW
authors.lastname: Huck
authors.affiliation: Institute of Analytical Chemistry and Radiochemistry, CCB-Center for Chemistry and Biomedicine, Leopold-Franzens University, Innrain 80/82, 6020 Innsbruck, Austria; ADSI-Austrian Drug Screening Institute, Innrain 66a, 6020 Innsbruck, Austria.
authors.forename: Günther K
authors.initials: GK
authors.lastname: Bonn
mesh_terms: D001798:Blood Proteins / Q000302:isolation & purification*; D011232:Chemical Precipitation; D002726:Chlorogenic Acid / Q000302:isolation & purification*; D002853:Chromatography, Liquid / Q000379:methods; D002934:Cinnamates / Q000302:isolation & purification*; D006801:Humans; D017640:Silicates / Q000737:chemistry*; D052616:Solid Phase Extraction / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D015040:Zirconium / Q000737:chemistry*
chemical_list: D001798:Blood Proteins; D002934:Cinnamates; D017640:Silicates; D002726:Chlorogenic Acid; C003784:zircon; C100257:cynarine; D015040:Zirconium
keywords: Caffeoylqinic acids; Human plasma; Protein precipitation; Solid-phase extraction; Zirconium silicate