The energy and nitrogen availability from softwood and softwood-hardwood blend ammonium sulfite liquor (ASL) were compared to urea-sulfur and urea-sulfur-molasses by in vitro techniques. Fluid collected from rumen fistulated sheep, previously adapted to a high energy ration containing 4% softwood-hardwood blend ASL (dry matter basis) was incubated with test substrates, in triplicate, for a 12-hr period. In vitro rumen microbial metabolism and growth were evaluated by gas, protein and volatile fatty acid (VFA) production. Gas production was insufficient as a criterion to evaluate in vitro microbial metabolism of the various substrates, as there were no differences in the rates or total gas production. However, total VFA production increased with softwood, and softwood-hardwood blend ASL's at the 4, 8 and 12% levels when compared to substrates with urea-sulfur and urea-sulfur-molasses. Acetate and butyrate production was higher but propionate production was lower with the ASL's than with urea-sulfur and urea-sulfur-molasses diets. Nitrogen from low levels of ASL (4%) was not as well utilized as that from urea containing diets. High levels (20 to 100%) of ASL had an inhibitory effect on gas production and protein synthesis. Softwood ASL, at low levels, appeared to be more beneficial to microbial metabolism than the softwood-hardwood blends of ASL.