A complete randomized block design trial with 4 in vitro incubation sets were prepared to evaluate the effect of a commercial blend of functional oils (FO) containing cashew nut shell liquid and castor oil as active ingredients (Essential, Oligo Basics Agroind. Ltda., Cascavel, Brazil) on rumen fermentation, methane production and methanogenic archaea. Bottles of 120 mL were filled with 600 mg of concentrate (same as given to 4 rumen liquid donor steers), and 80 mL of an incubation solution including rumen inoculum, mineral, buffer and reducing solutions under a CO₂ stream. Sealed bottles were incubated at 39 ± 1°C for 24 h and either dosed with 500 µg of FO/g DM of basal diet or not (control). The headspace pressure was measured and sampled (0.1 mL) at 2 h intervals, up to 12 h and then 24 h post incubation to determine gas and then methane concentration using GC. The pattern of cumulative gas/methane production (y) was fitted to the model: y = a(1−e^−b(t−c)), being a the potential cumulative gas/methane production (mL); b the production rate (mL/h) and c the lag time (h). After 12 and 24 h, 2 bottles per treatment per set were sampled for NH₃-N, volatile fatty acid (VFA) concentration and molecular analyses. The DNA was extracted using a QIAamp Kit. Specific primers were used to determine absolute abundance (Log₁₀ gene copy number/ g fresh sample) of total bacteria and hydrogenotrophic methanogenic archaea (HMA) and the relative abundance (2^(-ΔCt)) of HMA in relation to total archaea using qPCR (CFX96 Touch). Bottles supplemented with FO showed a tendency (14.26 vs. 13.50 SEM 0. 284; P = 0.06) to decrease methane production (mL/g DM substrate) and reduced the methane ratio (CH₄/gas; v/v; 0.093 vs. 0.089 SEM 0.0011; P = 0.049), where the discrete lag time for gas production increased (0.04 vs.0.12 SEM 0.024; P = 0.047). Addition of FO improved rumen fermentation, increasing molar proportion of propionate (33.5 vs. 34.3 SEM 0.24; P = 0.024) and decreasing NH₃-N concentration (305.3 vs. 284.4 SEM 5.95; P = 0.022) and relative abundance of HMA (17.8 vs. 14.7 SEM 0.96; P = 0.032).