Lactic acidosis is a major welfare issue affecting animal health and production systems such as dairy and feedlot beef. We used two bioassays to identify bioactive plants of Australia with the potential to prevent acidosis in ruminants. In the first bioassay, a potentially acidotic environment was induced by adding glucose to rumen fluid and pH and gas production were used to estimate the effect on acid production and microbial fermentation after 5-h incubation. Australian plants (n = 104) were screened for their ability to prevent a decline in the pH without inhibiting normal gas production, and five plants namely Eremophila glabra, Kennedia eximia, Acacia saligna, Acacia decurrens and Kennedia prorepens with such properties were identified. We investigated further the two top ranking plants, E. glabra and K. prorepens, in the second bioassay to determine the extent of their effect in vitro, by extending the incubation to 24 h and measuring d-lactate, and volatile fatty acids (VFA) in addition to pH and gas production. These were measured at 0, 5, 10, 16 and 24 h after inoculation. Eremophilaglabra maintained pH values that were higher and d-lactate concentrations that were lower than the control (P < 0.001), and comparable to the antibiotic-protected environment (AB; 12 μg of virginiamycin/ml). Eremophilaglabra and AB treatments did not restrict fermentation, as judged by gas production and VFA. Kennedia prorepens slowed the decline in pH and reduced the accumulation of lactate but inhibited gas production. We concluded that, in vitro, E. glabra was effective at controlling events that can lead to acidosis and the effect was comparable to that of virginiamycin, while K. prorepens was less effective than E. glabra and also inhibited fermentation.