Abstract

It is well established that promoting the balance of nutrients and plant secondary metabolites (PSM) by feeding diverse forage physiologically improves ruminant production. However, the underlying mechanism remains unclear. To investigate the physiological mechanism related to the improvement of physiological stress tolerance, ruminants were fed diverse forage. Oxidative stress markers were quantified, and serum metabolomics was performed. Six crossbred Shiba wethers (32.8±9.2kg BW) were arranged in a replicated 3×3 Latin square design. The treatments were feeding only Sudan grass hay (100% SDN); feeding a mixture of Sudan grass and alfalfa hay (70:30, SDN-ALF); and feeding a mixture of Sudan grass, timothy grass, and alfalfa hay (35:35:30; SDN-TMT-ALF). Each diet group was fed its specific diet for 21days with a 14-day adaptation period. Feed intake and digestibility, blood biochemistry, total antioxidant capacity (TAC), and superoxide dismutase (SOD) were analysed. In addition, blood serum metabolites were assessed by liquid chromatography-tandem mass spectrometry. The DM intake and DM, organic matter, and CP digestibility were higher (P<0.05) in the SDN-TMT-ALF group than in the SDN group. The TAC was higher (P<0.01) in the SDN-TMT-ALF and SDN-ALF groups (809.51 and 813.7µM, respectively) than the SDN group (720.69µM), while the SOD level was unchanged (P=0.06) among the treatments. Total serum cholesterol and NH3 levels were higher (P<0.05) in the SDN-TMT-ALF group (89.17mg/dL and 242.42µg/dL, respectively) than in the SDN group (71.00mg/dL and 89.17µg/dL). Additionally, the levels of nine metabolites in serum differed among the treatments (P<0.05). Linoleic acid (LA) and cortisone, which are related to LA metabolism and the steroid biosynthesis pathway, were upregulated by the SDN-ALF and SDN-TMT-ALF diets compared to the SDN diet, suggesting the contribution of ALF to altering the metabolites. The levels of hippuric acid, which is a metabolite of phenolic compounds, were higher (P<0.001) in the animals fed SDN, which contained higher phenolic and luteolin concentrations than the other diets. Pathway analysis suggested that the higher cortisone levels were derived from cholesterol due to upregulated glycolysis metabolism, which was positively related to increased ingestion, digestibility, and serum LA levels in animals given mixed forage. In conclusion, physiological stress tolerance in the animals was regulated by upregulation of LA and steroid hormone metabolism, which was associated with an increase in TAC rather than the ability of the animal to regulate its PSM intake.

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