PSIV-1 - A Comparison of Chromatography Methods to Estimate Ruminal VFA Concentrations

The objective of this study was to describe two chromatography equipment and their methods (EM) and to evaluate their adequacy in estimating ruminal volatile fatty acid concentrations (VFA). Adequacy was assessed through precision and accuracy using three standard mixtures of known acetate, propionate, and butyrate concentrations. The standard mixtures were prepared for VFA analysis using high-performance liquid chromatography (HPLC) or gas chromatography (GC). Each mixture was injected ten times into each EM. The comparison was assessed with rumen fluid samples from four cannulated steers offered three diets at 2% BW. Diet A simulated a forage-based diet offered to cattle during the winter. Diet B simulated a grower-type diet offered to weaned calves. Diet C simulated a finisher-type diet offered to finishing cattle. Rumen fluid was collected three hours after the morning feeding for seven days for each diet and strained through 8 μm porosity fiberglass wool. Two 2-mL aliquots were stored at -20°C for HPLC analysis, while two 8-mL aliquots were diluted with 2 mL of 25% meta-phosphoric acid and stored at -20°C for GC analysis. Chromatograms without a flat baseline were removed from the analysis. For the adequacy evaluation, HPLC (R² = 0.997; Cb = 0.874) was more precise and accurate at estimating total VFA than GC (R² = 0.447; Cb = 0.763). When compared with the standards, HPLC estimated less (P < 0.001) total VFA (98.8 ± 10.3 mM) than GC (110.5 ± 17.4 mM). Concentrations for acetate, propionate, and butyrate in rumen fluid samples were estimated for each EM and analyzed using a random coefficients model. Similarly, estimates for acetate, propionate, and butyrate were less for HPLC than GC (P ≤ 0.002). VFA estimation differs depending on EM chosen. Further research should identify the source of difference in VFA estimation from each EM.