Producing butter from yoghurt is known as a traditional way practiced in Iran and elsewhere, particularly in rural areas. Lactic acid bacteria (LAB) with probiotic and cholesterol-lowering properties were isolated from traditional butter collected in different regions of Iran. The isolates were then identified and applied as adjunct culture in industrial butter production. Ten samples of traditional Iranian butter were collected from local farms. Fifty-four isolates were considered LAB due to biochemical examinations. Molecular techniques then identified ten strains showing high cholesterol reduction ability and tolerated bile and acid. The sequence analysis revealed that 4 isolates belonged to Enterococcus durans, 4 isolates to Lactobacillus, 1 isolate to Pediococcus, and 1 isolate to Neoscardovia. Afterwards, Lactobacillus brevis IBRC-M 11044, Pediococcus pentosaceus IBRC-M 11045, Neoscardovia arbecensis IBRC-M 4391 4378, and Lactobacillus pentosus IBRC-M 11043 were selected and applied as adjunct culture in producing four treatments of industrial butter. All examined strain treatments showed significant changes in cholesterol level of butter samples. Furthermore in all samples, cholesterol content was significantly lower than control (P< 0.5). The highest level of cholesterol reduction was achieved in butter sample prepared by Lactobacillus pentosus IBRC-M 11045. Sensory analysis showed that butter sample with Neoscardovia arbecensis IBRC-M 4391 4378 was more acceptable than other butter samples. Due to our finding, it is valuable to incorporate these strains in butter production and introduce novel functional butter to the market.

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