Sulphur containing volatile molecules belong to the most aroma active components in food with reported concentrations below 1 µg/L. Various extraction methods have been developed in the past to enrich thiols and improve detectability [1-4]. Here, we describe the development of a new extraction protocol using 2-thiopyridyl disulphide activated agarose gel (known as thiopropyl-sepharose 6B, TPS) commonly used for protein analysis [5]. Primary, secondary and tertiary thiols namely 1-hexanethiol, 2-furfurylthiol, 2-phenylethanethiol, pentane-2-thiol, 3-mercaptohexyl acetate, 2-methyltetrahydrofuran-3-thiol, 4-methoxy-2-methyl-2-butanethiol, 4-mercapto-4-methyl-2-pentanone, 1-p-menthene-8-thiol and p-mentha-8-thiol-3-one were used to study the binding affinity of the stationary phase. The optimization of the overall extraction parameters was performed by a design of experiment (DOE) approach. A selective enrichment and concentration was achieved by factor up to 70 for primary thiols on TPS and by an additional factor up to 5000 for stir bar sorptive extraction (SBSE) of the ethanolic extract obtained after elution of bound thiols with tris(2-carboxyethyl)phosphine. Limit of detection levels less than 1 µg/L could be achieved by combining TPS based extraction with SBSE-gas chromatography-mass spectrometry (GC-MS).