The aim of this study was to investigate the effect of diets containing lipid sources with different fatty acid profiles on ruminal pH, NH₃-N, volatile fatty acids (VFA) and microbial protein synthesis. Ten Nellore steers with initial body weight of 268 ± 27 kg, ruminally cannulated, were used in a double 5 × 5 Latin square design (20 d of each period). Dietary treatments were: without fat (WF), palm oil (PO), linseed oil (LO), protected fat (PF; Lactoplus), and whole soybeans (WS). The roughage feed was corn silage (600 g/kg on a DM basis) plus concentrate (400 g/kg on a DM basis). To evaluate rumen fermentation parameters, rumen fluid samples (approximately 80 mL) were collected manually on the last day of the experimental period (20 d), both before supplying the diet (time zero) and 1, 2, 4, 6, 8, 10, 12, and 14 h after feeding. Microbial protein synthesis was calculated via urinary total excretion of purine derivatives (allantoin + uric acid). The diet with PF and WF increased the concentration of NH3-N (P < 0.001); however, the diet did not change in VFAs (P > 0.05), such as the molar percentage of acetate (ACE), propionate (PROP), butyrate and the ACE:PROP ratio. There was a tendency (P = 0.06) to change ruminal pH values and total VFA (P = 0.092) with lipid diets. The higher NH3-N and tendency to reduce pH values in the WF and PF diets could be linked to the higher DMI (nitrogen and organic matter), and consequently higher ruminal fermentation. The higher production of microbial N (P = 0.030) was in the animals fed WF, LO and WS, whereas animals of the PO diet had the lowest production. The higher microbial protein synthesis was found in animals on the diet with LO and WS (P = 0.040). Diets with some type of protection (PF and WS) caused less disturbance on ruminal fermentation.