ORIGINAL ARTICLE Sitosterol Thermo-oxidative Degradation Leads to the Formation of Dimers, Trimers and Oligomers: A Study Using Combined Size Exclusion Chromatography/Mass Spectrometry Magdalena Rudzinska • Roman Przybylski • Yuan Yuan Zhao • Jonathan M. Curtis Received: 30 October 2009 / Accepted: 21 April 2010 / Published online: 29 May 2010 Ó AOCS 2010 Abstract Phytosterols are recognized as functional food components with cholesterol reducing properties in humans. The formation of phytosterol oligomers as a result of the thermo-oxidative degradation of sitosterol is shown to occur. The existence of oligomers is demonstrated by size exclusion chromatography (SEC) and confirmed by combined SEC-atmospheric pressure chemical ionization mass spectrometry (SEC/APCI-MS). A speculative struc- ture for the sitosterol dimer with 3,7 0 linkage is proposed consistent with data from tandem mass spectrometry and exact mass measurements. Higher molecular weight spe- cies arising from the formation of trimers or higher olig- omers are seen in the mass spectra. Fragments of sitosterol formed by thermo-oxidative processes are also shown to oligomerize and their common structural characteristics are demonstrated by tandem mass spectrometry. The results presented provide evidence for the possible formation of oligomeric species involving sterols in addition to those known for acylglycerides in vegetable oils subjected to extreme oxidative stress such as in frying. Keywords Sitosterol Á Oxyphytosterols Á Thermo-oxidative degradation Á Oligomers Á LC/MS Á SEC Á HPLC-APCI/MSMS Abbreviations APCI Atmospheric pressure chemical ionization ELSD Evaporative light scattering detector ESI Electrospray ionization HPLC High pressure liquid chromatography LC/MS Liquid chromatography–mass spectrometry SEC Size exclusion chromatography Introduction Sterols comprise a major portion of the unsaponifiable matter of most vegetable oils and are mainly present as free, esterified with fatty and phenolic acids, and as glucosides. Phytosterols similar in chemical structure to cholesterol, are prone to oxidation particularly at elevated temperatures as used during frying [1, 2]. Oxidative deg- radation of phytosterols leads to the formation of oxidized sterol derivatives, volatile flavor components and oligo- mers [3–6]. Dutta et al. [7] studied the formation of phytosterol oxides during potato chips and French fries frying in different oils. Lampi et al. [8] and Soupas et al. [9] established that the sterol structure and oil matrix are the main factors affecting the formation of oxidative derivatives. The mechanism of sterol oxidation is similar to free radical oxidation of fatty acids [10]. The initial step involves the formation of a C-7 carbon centered radical adjacent to the double bond at C-5 and C-6 in ring B of the M. Rudzinska Faculty of Food Science and Nutrition, Poznan University of Life Sciences, Poznan, Poland Y. Y. Zhao Á J. M. Curtis Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Canada R. Przybylski (&) Department of Chemistry and Biochemistry, University of Lethbridge, Lethbridge, AB T1K 3M4, Canada e-mail: roman.przybylski@uleth.ca 123 Lipids (2010) 45:549–558 DOI 10.1007/s11745-010-3433-0