title: Solid-phase extraction and GC-MS analysis of potentially genotoxic cleavage products of β-carotene in primary cell cultures.
abstract: A validated method for the simultaneous determination of prominent volatile cleavage products (CPs) of β-carotene in cell culture media has been developed. Target CPs comprised β-ionone (β-IO), cyclocitral (CC), dihydroactinidiolide (DHA), and 1,1,6-trimethyltetraline (TMT). CPs were extracted by solid-phase extraction applying a phenyl adsorbent, eluted with 10% (v/v) tetrahydrofuran in n-hexane, and identified and quantified by gas chromatography-mass spectrometry with electron impact ionization. Method validation addressed linearity confirmation over two application ranges and homoscedasticity testing. Recoveries from culture media were between 71.7% and 95.7% at 1.0 μg/ml. Precision of recoveries determined in intra-day (N = 5) and inter-day (N = 15) assays were <2.0% and <4.8%, respectively. Limit of detection and limit of quantification of the analysis method were <18.0 and <53.0 ng/ml for β-IO, CC, and TMT, whereas 156 and 474 ng/ml were determined for DHA, respectively. Although extractions of blank matrix proved the absence of interfering peaks, statistical comparison between slopes determined for instrumental and total method linearity revealed significant differences. The method was successfully applied in selecting an appropriate solvent for the fortification of culture media with volatile CPs, including the determination of their availability over the incubation period. For the first time, quantification of volatile CPs in treatment solutions and culture media for primary cells becomes accessible by this validated method.
authors.affiliation: Department of Molecular Biology, Division of Chemistry and Bioanalytics, University of Salzburg, Salzburg, Austria.
authors.forename: G
authors.initials: G
authors.lastname: Martano
authors.forename: C
authors.initials: C
authors.lastname: Vogl
authors.forename: E
authors.initials: E
authors.lastname: Bojaxhi
authors.forename: N
authors.initials: N
authors.lastname: Bresgen
authors.forename: P
authors.initials: P
authors.lastname: Eckl
authors.forename: H
authors.initials: H
authors.lastname: Stutz
mesh_terms: D056809:Alveolar Epithelial Cells / Q000737:chemistry* / Q000166:cytology; D000818:Animals; D002478:Cells, Cultured; D008401:Gas Chromatography-Mass Spectrometry; D022781:Hepatocytes / Q000737:chemistry* / Q000166:cytology; D015394:Molecular Structure; D051381:Rats; D052616:Solid Phase Extraction; D013237:Stereoisomerism; D019207:beta Carotene / Q000032:analysis*
chemical_list: D019207:beta Carotene