|Reynal, S - UW-MADISON|
Submitted to: Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: December 1, 2004
Publication Date: December 15, 2004
Citation: Broderick, G.A., Reynal, S.M. 2004. Methods for quantifying microbial protein flow from the rumen of lactating dairy cows using 15n-ammonia, total purines and urinary excretion of purine derivatives. Workshop sobre Adaptação do Modelo Cornell Net Carbohydrate and Protein System-CNCPS às condições tropicais: treinamento e seminário. Technical Abstract: The high-quality microbial protein synthesized in the rumen of dairy cows contributes the majority of total AA flowing to the small intestine. Poor utilization of dietary crude protein (CP) may result from inefficient ruminal conversion of degraded protein to microbial protein, contributing to substantial losses of urinary N. Optimization of microbial protein synthesis in the rumen should increase feed efficiency while reducing the most polluting form of excretory N. Studies on the factors affecting microbial yield and efficiency rely on accurate measurement of microbial protein synthesis. However, errors associated with unrepresentative microbial sampling and imprecise microbial markers contribute to variation when determining microbial protein synthesis. Among several markers, total purines (TP) and 15N-ammonium salts have been used most extensively to estimate microbial protein yield. Theoretically, the external marker 15N offers several advantages over TP. The 15N should be evenly distributed throughout the microbial cell; thus, microbial lysis during isolation would not affect 15N enrichment but could result in loss of purines from cell cytoplasm, lowering of the purine:N ratio and overestimating microbial flow. Moreover, unless contamination occurs, 15N-labeled protein leaving the rumen that was enriched in excess of natural abundance can only be of microbial origin, while some portion of the purines leaving the rumen may be of dietary origin or result from intra-ruminal lysis of microbial cells. Although low recoveries of TP have been reported when using the spectrophotometric procedure of Zinn and Owens, recent modifications of this method have substantially improved purine recovery. Direct quantitation with virtually complete recovery of adenine (A) and guanine (G) using an HPLC assay resulted in microbial protein flows that were highly correlated (r = 0.94) to those determined with the modified Zinn and Owens method. However, results from comparisons between TP and 15N reported in the literature have been inconsistent. Use of TP resulted in higher, lower, and similar values for microbial yield compared to 15N. Alternatively, microbial protein flow from the rumen can be estimated from the urinary excretion of purine derivatives (PD) using published equations relating PD excretion to total purine flow from the rumen and microbial NAN:purine ratios. This non-invasive approach is based on the principle that most of the urinary PD are derived principally from microbial nucleic acids. However, the extent of endogenous contribution to urinary PD and the proportions of total PD excreted through non-renal routes may vary depending on the nutritional and physiological status of the animal. Moreover, the use of equations developed under experimental conditions that differ substantially from those of the study where they are applied may also result in biased estimates of microbial flow. The paper describes our methods for using TP or 15N as microbial markers for quantifying microbial protein flows from the rumen, determined using sampling at the omasal canal, or using urinary PD excretion in lactating dairy cows.