Location: Agroecosystems Management Research2012 Annual Report
1a. Objectives (from AD-416):
The contribution of dietary oxidized fats to total energy intake has markedly increased with the higher inclusion rates of corn co-products and various other supplemental fat sources, which may cause suboptimal pig performance and negatively affect intestinal health. Only recently, have data been published on the impact of fat rancidity (oxidation) on fat digestibility and pig growth performance. However, no data are available on the impact of dietary oxidized fat on digestible and metabolizable energy (DE and ME, respectively) content, nitrogen retention, or its impact on oxidative stress and intestinal barrier function in growing pigs. Thus, there is a critical need to understand the impact of fat type and oxidation level on dietary energy (DE and ME) and nitrogen utilization, as well as its intestinal and physiological effects. There are limited data suggesting that feeding oxidized fat to swine increases oxidative stress, but the impact of dietary oxidized fat on intestinal barrier function and mucosal immunity is unknown. Therefore, the objectives of the proposed research are to: 1) Determine the variation in DE and ME content of 4 dietary fat sources ranging from saturated animal fat (tallow and poultry fat) and 2 unsaturated vegetable oils containing either low or high amounts of linoleic acid (canola and corn oil, respectively); 2) Determine the impact of lipid oxidation on DE and ME content and nitrogen retention; and 3) Determine the impact of dietary fat source and lipid oxidation on indicators of oxidative stress and intestinal barrier function.
1b. Approach (from AD-416):
Obj. 1: Four different lipid sources (2 animal fats and 2 vegetable oils) will be selected based on degree of saturation. The nutrient balance experiment consists of utilizing 72 barrows weighing approximately 20 kg, which will be randomly assigned to 1 of 9 dietary treatments. The basal diet will be formulated to satisfy the nutrient requirements as suggested by the NRC (1998). Additional diets will consist of supplementing 10% of each lipid on top of the basal diet. Test diets will consist of both normal and oxidized samples of lipid, consisting of: poultry fat (an unsaturated animal fat), tallow (a saturated animal fat), corn oil (an unsaturated vegetable oil with a high content of linoleic acid), and canola oil (an unsaturated vegetable oil with a low content of linoleic acid). Obj. 2: Oxidation of lipid sources will be achieved by heating an aliquot of each oil at 190°C for 6 hours with compressed air at room temperature (22-24°C) bubbled into the sample at a flow rate of 97 cm 3/min. Pigs will be fed their respective experimental diets twice daily at an amount equivalent to approximately 4% of their body weight and will be provided ad libitum access to water. Pigs will be housed individually in metabolism crates designed for total, but separate collection of feces and urine. Pigs will be allowed to adapt to the experimental diets for 28 days followed by a 4-day total collection period. At the end of the collection period, all pigs will be fasted for 24 hours and urine will be collected to determine the concentration of polar and non-polar secondary oxidation products to determine in vivo production of secondary oxidation products. Urine 4-HNE concentrations normalized to urine creatinine concentrations and serum concentrations of TBARS will be measured as indicators of oxidative stress. Energy and nitrogen concentrations will be determined by adiabatic bomb calorimetry and thermo combustion, respectively, and nutrient balance and DE and ME determinations will be calculated. Obj. 3: On the final day of the experiment for Objective 2, the impact of dietary fat source and fat oxidation on intestinal barrier function will be determined by administering an oral dose of a cocktail containing 10 grams of lactulose and 2 grams of mannitol to all pigs. Urine will be collected for a period of 6 hours into a container with chlorhexidine to prevent microbial contamination. Urinary concentrations of lactulose and mannitol will be determined by HPLC, and the lactulose:mannitol ratio will be calculated as an indicator of small intestinal permeability. Blood samples will be collected from each pig, and serum concentrations of endotoxin will be determined by the fluorescent PyroGene® rFC assay as another indicator of intestinal barrier function. Blood samples will be collected after an overnight fast and at 3 hours after feeding for determination of endotoxin in both the fasted and fed state. Serum a-1- acid glycoprotein and haptoglobin concentrations will be measured as indicators of systemic inflammation. Fecal concentrations of IgA will be determined on freshly collected and frozen fecal samples as a marker of mucosal immunity.
3. Progress Report:
Four lipids were obtained and peroxidized by a slow or rapid method and, subsequently, characterized by various methods. Analysis showed that a high peroxide value accurately indicated the high degree of lipid peroxidation, but a moderate or low peroxide value may be misleading due to the unstable characteristics of hydroperoxides as indicated by the unchanged peroxide value of rapidly oxidized corn and canola oil compared to their original, unoxidized state. Additional tests which measured secondary peroxidation products (p-anisidine, thiobarbituric acid reactive substances, hexanal, 4-hydroxynonenal, and 2, 4-decadienal) were suggested to provide a better indication of lipid peroxidation than peroxide value for lipids subjected to a high degree of peroxidation. Similar to peroxide value analysis, it was suggested that these tests may also not provide irrefutable information regarding the extent of peroxidation due to the volatile characteristics of secondary peroxidation products and the ever changing stage of lipid peroxidation. For the predictive tests, active oxygen method stability accurately reflected the increased lipid peroxidation caused by the slow and rapid oxidation treatments as indicated by the increased active oxygen method stability value in corn and canola oil, but not in poultry fat and tallow, which indicates a potential disadvantage of the active oxygen method stability test. The oxidative stability index assay successfully showed the increased lipid peroxidation caused by slow or rapid oxidation treatments in all lipids, but it too may have disadvantages similar to anisidine, thiobarbituric acid reactive substances, hexanal, 4-hydroxynonenal, and 2, 4-decadienal, because the oxidative stability index assay directly depends on quantification of the volatile secondary peroxidation products. Overall, the data suggests that to accurately analyze the peroxidation damage in lipids, measurements should be determined at appropriate time intervals by more than one test and include different types of peroxidation products simultaneously.