*2.3. Mineral Analysis*

Trace mineral concentrations in the feed components, plasma, and tissue samples were analyzed using a double-beam atomic absorption spectrophotometer (AA-7000 Series, Shimadzu Co., Kyoto, Japan) with a graphite furnace (GFA-7000, Shimadzu Co., Kyoto, Japan). The microwave-assisted digestion method using closed pressure vessels (MWS 4 Speedwave, Berghof Co., Eningen, Germany) was used for decomposition of all samples in two replicates. All samples, except plasma, were wet acid-digested with a mixture of concentrated nitric acid and hydrogen peroxide. Mineral concentration of Mn in muscles was determined using a graphite furnace atomic absorption spectrophotometer with deuterium background correction and pyrolytic-coated graphite tubes [23]. The certificate reference materials of bovine liver BCR-185, bovine muscle ERM-BB184, and pig kidney ERM-BB186 from the Institute for Reference Materials and Measurements (IRMM, Geel, Belgium) and ClinCheck control of lyophilized human blood (Recipe, Munich, Germany) were included in each analysis to verify instrument accuracy. Mineral concentrations in all samples except plasma were expressed as mg/kg of DM and in plasma as mg/L.
