The objective of the study was to determinate optimal functions of new silage inoculants as probiotic agents for fermentative processes of rumen microbes. The inoculants were Lactobacillus plantarum, Lactobacillus fermentum and Enterococcus faecium. Allinoculants of whole corn plant were not effective for increasing conjugated linoleic acid and trans vaccenic acid concentration in fermentation fluid and for decreasing of rumen biohydrogenation of C18 fatty acids in RUSITEC. Results suggest that silageswith different inoculants induce changes in rumen metabolism that might be related to differences in the extent of rumen biohydrogenation of polyunsaturated fatty acids in RUSITEC. It will probably be necessary to increase the concentration of polyunsaturated fatty acids using oilseeds or plant oils as supplements in corn silage substrates. Therefore, future experiments for increasing conjugated linoleic acid and trans vaccenic acid in fermentation fluid and milk will be necessary.