Recycling of microbial N and estimation of protein degradation by in vitro gas production

Rumen fluid (RF) activity has great impact on in vitro incubation measurements for feed evaluation in ruminant nutrition. When feed protein degradability is estimated from the release of ammonia (NH3), the presence of protozoa in RF and their release of NH3 from phagocytosis of rumen bacteria may influence degradation estimates. The objective of this study was to improve the in vitro gas production (GP) method, as applied to protein degradation measurements, by defaunation of RF. Soybean meal was incubated as a protein source along with graded levels of carbohydrates and buffered RF, which was either defaunated by centrifugation or untreated. Gas production and NH3 were recorded continuously during 23 h. Extrapolation of these recordings to zero GP was used as a theoretical value for NH3 release from feed protein degradation at zero microbial growth. Results showed that defaunation reduced NH3 concentration in blank incubations, which was particularly evident at longer incubation intervals compared to untreated RF. This effect was likely caused by eliminating bacterial N recycled by protozoa. Defaunation lowered NH3 release in the soybean meal incubations by 55% (P<0.002) and also the estimates of in vitro degradation of crude protein (IVDP) at different incubation times, while GP was only reduced by 13% (P<0.008). However, incubations with defaunated RF resulted in depleted NH3 concentrations in early stages of incubation, which led to unreliable estimates of IVDP for these intervals. Further improvements of the method should therefore consider an adequate addition of NH3 at the start of the incubation. These improvements are important to eventually establish the methods as an alternative to other protein evaluation methods.