In this study, the efficiency of using zeolite-based adsorbents in a solid phase extraction (SPE) procedure of alpha-tocopherol and beta-sitosterol isolation from Sunflower Oil Deodorizer Distillate (SuDOD) without pre-treatment was investigated. The results showed that 99.2% a-tocopherol and 97.3% beta-sitosterol were successfully isolated as pure fractions from SuDOD, when desilicated ZSM-5-type zeolite (DSiZSM-5) was used as adsorbent on a SPE. A simple and rapid HPLC method for simultaneous alpha-tocopherol and beta-sitosterol analysis was developed and validated according to AOAC guidelines. It was found that the inclusion of a DSiZSM-5 SPE step increased the precision of the alpha-tocopherol and beta-sitosterol analysis. In conclusion, DSiZSM-5 zeolite was proven to be an efficient adsorbent which can be used not only for the recovery of alpha-tocopherol and beta-sitosterol from SuDOD in industrial scale, but also in a laboratory scale clean-up method prior to the analysis of alpha-tocopherol and beta-sitosterol.