Techniques of forage evaluation based on in-vitro rumen gas production have the potential for monitoring rumen fermentation kinetics of different energy yielding compounds. Observations in our laboratory led to identify proteins as a source of error in gas pressure measurements and the underlying hypothesis was that dissolved C02 is up taken by basic compounds and prevented from escaping to the bottle head-space. Bases would be generated during hydrolysis of proteins, and the higher the protein content the higher the potential for C02 uptake by ammonia. The experimental methodology was based on in-vitro batch rumen incubation system, a pressure transducer, manual gas measurement and frequent gas release. In a purely chemical assay a protein-free rumen-like buffer incubation media was dosed with six levels of ammonium hydroxide (NH40H) as an alkaline agent and gas pressure recorded at hourly intervals. Gas production was depleted at a rate of 0.24 psi/mgN/bottlc of I JO mL as NH40H increased from O to 25 mg/bottle. In a rumen in-vitro incubation experiment six levels of soy peptone or casein hydrolysate containing O to 160 mg protein/bottle were dosed to a basal substrate of 300 mg purified cell walls of Lolium multiflorum Lam. Results showed an inverse relationship between gas production and level of protein supplementation, being more severe at longer incubation time. Another experiment compared soy peptone and ammonium sulphate as sources of organic and inorganic nitrogen supplemented to same basal substrate at 15 mgN/bottle. A consistent depiction in apparent gas production was observed as the proportion of organic N was increased. When ammonium sulphate was substituted by 100% organic N, gas production at end point was underestimated by 28%. In another experiment soybean meal and leafy fractions of lucerne, red clover and orchardgrass were dosed to supplement the basal cell wall substrate with l O or 25 mg protein/bottle. Opposite effects in digestibility and gas production were observed by increasing supplementation. In-vitro dry matter digestibility increased from an average 52.7 to 60.8%. However, gas production diminished from 410, 421, 418 and 409 rnL·g-1 to 392, 393, 377, and 347 mL·g-1 fermented dry matter in all four forages respectively. The results confirmed that ammonia resulting from ruminal hydrolysis of nitrogenous compounds performs as sink for dissolved C02 thus interfering with the measurement of gas actually produced in the in-vitro system. This effect was large enough to offset the ammonia untaken by rumen microbes and the C02 released from buffer for fatty acids neutralization. Modelling was sensitive to this artifact and kinetic parameters were altered, particularly in pool sizes.