Due to high nutritional value of butter, extensive studies have been undertaken to improve its functional properties focusing on the reduction of cholesterol. One of the health benefits of probiotics is their ability to reduce serum cholesterol. The main aim of this study was to investigate the characteristics, Fatty Acid Profiles, rheological behavior and sensory properties of butter samples produced by adding adjunct culture with the ability of cholesterol assimilation. Lactobacillus brevis, Pediococcus pentosaceus, Neoscardovia arbecensis and Lactobacillus pentosus were used as adjunct culture in the process of industrial butter. Results shown that using all examined strains as an adjunct starter reduced the amount of saturated fatty acids in butter and increased the amount of unsaturated fatty acids. In the butter sample prepared with Lactobacillus brevis, the highest amount of unsaturated fatty acids was observed (33.2%) and was the softest butter in rheological properties. Sensory analysis showed a significant differences between all treatments (P<0.05) than control and the sample with Neoscardovia arbecensis was more acceptable than the others. It is concluded that due to the high health effect properties of the examined strains and their acceptable technological characteristics, we do encourage dairy industries involved in butter production to incorporate these strains in their product and introduce novel functional butter to the market

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