Simultaneous analysis of organic acids and inorganic anions in silage by capillary electrophoresis

Organic acids and some inorganic anions may have an effect on the nutritive quality of silage. To facilitate the simultaneous and quantitative measurement of nine organic acids (formate, acetate, propionate, butyrate, n-valerate, iso-valerate, n-hexanate, citrate and lactate) and three inorganic anions (nitrate, nitrite and chloride) in silage, we optimized an analysis method using capillary zone electrophoresis (CZE) with a photodiode-array detector (PDA). We then validated the optimized method in a collaborative study. Samples were extracted with water, purified through a cartridge column to remove interferences, diluted with water, filtered with a centrifugal filter and applied on the CZE. The separation was carried out with a fused-silica capillary column at 30 kV. A wavelength of 350 nm was selected for indirect UV detection and a wavelength of 275 nm was selected as a reference. A 30 mM 2,6-pyridinedicarboxylic acid (PDC) solution was used as a background electrolyte (BGE) at pH 12 with 0.5 mM of cetyltrimethylammonium hydroxide (CTAH).For the recovery study, the silage was spiked with 3.0 or 6.0 g/kg of each of the nine organic acids and three inorganic anions. The mean recoveries of the acids were 0.996–1.097 and the coefficient of variation for the repeatability (CV) was within 0.071. The collaborative study included five laboratories. Grass silage was spiked with 7.0 g/kg of each analyte. The mean recoveries of the acids were 0.843–1.143, the coefficients of variation for the repeatability and reproducibility (CVr and CVR) were zero to 0.058 and 0.033–0.081, respectively, and the HorRat values were 0.8–1.9. From the measurements of farm silage samples, chloride, nitrate, formate, citrate, acetate, lactate, butyrate and iso-valerate were detected, and their mean ranges were 2.7–4.4, 0–0.9, 0–2.6, 0–1.8, 0.7–5.5, 0–12.3, 0–3.4 and 0–2.4 g/kg, respectively, and the coefficients of variation for the repeatability were within 0.087, 0.079, 0.083, 0.095, 0.091, 0.070, 0.103 and 0.037, respectively. Over a long-term storage period of nine months, nitrate and citrate decreased. The acetate and lactate contents were higher in the high-moisture silage than the low-moisture silage.