The objective of the in vivo study was to evaluate whether feeding graded levels of boron affect rumen microbial fermentation including pH, ammonia, volatile fatty acids and protozoa counts. In the experiment 4 Merino rams in a 4 × 4 Latin square design fed forage and concentrate with boric acid (0, 200, 300 and 400 mg · kg⁻¹ in control, B1, B2 and B3 group, respectively) were used. Each experimental period lasted 14 days, with 12 first days of diet adaptation. In comparison with control diet, boron supplementation did not increased the total volatile fatty acid concentration before and 3 h after feeding. However in both time points, acetate content was higher in B1 and B2 than in control and B3 groups, whereas propionate content was lower in all boron-supplemented groups. The iso-butyrate, n-butyrate and iso-valerate levels were influenced only 3 h after feeding. The n-valerate content was lower in B1 and B2 than in control and B3 groups. Protozoan abundance in the rumen fluid was significantly higher in animals fed B3 and control diets both before and after feeding. The boron content in rumen fluid was increased in boron-supplemented groups to average value 7.32 ppm, but dose effect was not observed. The obtained results showed that dietary boron supplementation had a dose-dependent influence on rumen microbial fermentation and protozoan abundance in yearling rams. However, the boron concentration in rumen fluid did not increased simultaneously with increased dose in a diet. Further studies are needed to estimate the most recommended dose of boron in the ruminant diets and to better understand the boron role in the processes occurring in the rumen.