Study On The Application Of Low-gossypol Cottonseed Meal In Laying Hen Diets

Soybean meal（SBM） is currently the most commonly used plant protein ingredient in poultry feed and comprises up to 50% of the poultry diet. However, a shortage of SBM supply and the increase of SBM price prevents higher use in the poultry diet. Replacement of SBM with less expensive plant protein sources would be beneficial for reducing feed costs and saving protein resources in the animal feed industry. Cottonseed meal（CSM）, a byproduct of the process of oil extraction from cotton seeds, is an attractive alternative protein ingredient for poultry diets. However, CSM has a consistently lower feeding value, attributable to free gossypol（FG, a main anti-nutritional factor of CSM）, with lower yields of energy and lysine compared to SBM. Recognition of these undesirable factors coupled with the progress in processing techniques has made the method of oil extraction improved, which has resulted in low-gossypol CSM（LCSM）（Sterling et al., 2002）. The more refined technology without high-temperature heating greatly reduces husk and maintains nutrient density to the most extent during oil extraction, meanwhile integrated de-gossypolization of solvent processes after oil extraction highly decreases FG levels in LCSM. The purpose of this study is to investigate the application of LCSM,evaluating the effect of LCSM on performance and egg quality of laying hens. Further, i TRAQ proteomic combined with biochemical analysis was performed to investigate the impact of this substitution at the molecular level on egg whites. And then the effects of LCSM on egg quality during storage were studiedExperiment 1. Effect of LCSM on laying performance and egg quality under different dietary protein levels This experiment was designed to evaluate the effects of of LCSM on laying performance and egg quality during different dietary protein level. Hy-line W36 hens（n=432, 40 weeks old） were randomly allocated to one of six dietary treatments. Each treatment had six replicates of 12 hens each. All birds dietary protein soruce were SBM or LCSM with three crude protein level, 14.5%,16%,17.5%,respectively（LCSM14.5, LCSM16,LCSM17.5; SBM14.5,SBM16,SBM17.5）. The experiment lasted for 12 weeks. The result showed that laying performance was significantly affected by LCSM（P<0.05）.Laying performace and egg quality were not affected by LCSM or different dietary protein level during40-45 WK. However, there is significance decreace on laying performance and egg quality during46-51 WK. Compared with SBM14.5 group, feed intake was significantly reduced but egg weight was not affected in the LCSM14.5 group. However, egg weight was significantly reduced in the LCSM16 group compared with SBM16.Albumin height and Haugh unit were significantly reduced in the LCSM14.5group compared with SBM14.5 group.However, there is no difference on lbumin height and HU in the LCSM16 group compared with SBM16.FCR was lowest in the LCSM14.5,LCSM16 and SBM16 group,since significant depression was observed on egg quality in the LCSM14.5 group, thus, dietary protein level 16% was applied for inclusion of LCSM in hens’ diet.Experiment 2. Effect of different dosages of LCSM on laying performance, egg quality and histopathological assessment in laying hensThis experiment was designed to evaluate the effects of LCSM on laying performance,egg quality and histopathological assessment in inclusion rate, and to investigate relationship between FG and depressed performance in laying hens. A total of 432 40-WK-old Hy-line W36 laying hens were allocated to one of the six dietary treatments with 6 replicates of 12 birds each. The control group was corn-soybean meal basal diet, and four experimental diets was a basal diet with 50, 98.3, 144.2, or 189g/kg LCSM, respectively（correspondingly 25%, 50%, 75%, ₁00% dietary protein provided by soybean meal were replaced by LCSM）. The sixth group was a basal diet with supplemented free gossypol（FG group）. The feeding trial lasted for 12 weeks. The results showed that no significant difference was observed on egg production or FCR（feed/egg, g/g） among all groups, but feed intake and egg weight were significantly decreased in the 189 g/kg LCSM group in WK 46-51（P<0.05）. A significant decrease was not observed on shell strength, shell thickness and yolk color in all periods, but Haugh unit, albumen height and egg white protein content was reduced in 189 g/kg LCSM group in WK 46-51（P<0.05）. No significant differences were found between the control and FG group. There was no obvious difference on plasma levels of total protein, blood urea nitrogen, uric acid, and activities of albumen, alanine aminotransferase, aspartate aminotransferase and histopathology of liver, kidney and oviduct in all groups. In conclusion, decreasing of feed intake, egg weight and egg quality were observed in the 189 g/kg LCSM group. However, the adverse effect was not relevant to the presence of FG in LCSM. Dietary LCSM supplementation of 98.3 g/kg was recommended with optimum FCR and without adverse effect on egg production, quality and health of layers.Experiment 3. Proteomic comparison by i TRAQ combined with mass spectrometry of egg white proteins in laying hens（Gallus gallus） fed with soybean meal and cottonseed mealThis experiment was designed to investigate the molecular consequences of replacement of SBM using LCSM. A total of 216 40-WK-old Hy-line W36 laying hens were given 3 dietary treatments with six replicates of 12 birds each. The control group was fed a corn and soybean meal basal diet, and the other two experimental diets were with 50%（LCSM50） and ₁00%（LCSM₁00） of the dietary protein content provided by SBM replaced by LCSM. The feeding trial lasted for 12 weeks. To investigate the impact of LCSM on albumen quality at the molecular level, i TRAQ combined with biochemical analysis was performed in Hy-line W36 hens supplemented with a mixed diet of LCSM and SBM. Egg weight, albumen height, and Haugh unit were significantly reduced in the LCSM₁00 group（₁00% crude protein of SBM replaced by LCSM） compared with the SBM group（P<0.05）. A total of 15 proteins,accounting for 75% of egg white proteins with various biological functions of egg whites, were found to be reduced. This finding may relate to the reduction of albumen quality in the LCSM₁00 group. Oviduct magnum morphology and hormone analysis indicated that a reduced level of plasma progesterone caused reduced growth of the tubular gland and epithelial cells in the magnum, further decreasing egg white protein synthesis in the magnum. These findings help demonstrate the mechanisms of a LCSM diet that cause adverse effects on albumen quality, while also showing at the molecular level that SBM should not be totally replaced with LCSM in a hen diet.Experiment 4. Effect of LCSM on egg quality during storage using 2-DE followed by MALDI-TOF/TOF MS analysis in laying hensThis experiment was designed to investigate the effects of LCSM on egg quality during storage in laying hens. After 12 WK of diet feeding,a total of 540 eggs as collected for egg quality determination during storagement and two-dimensional electrophoresis（2-DE） followed by matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometer（MALDI-TOF/TOF MS） analysis. Egg from three treatments（SBM, LCSM50, LCSM₁00） were used and were stored at 4℃ at 0 d,4 WK,8WK.The result showed that the egg Haugh units（HU） in the LCSM₁00 group was significantly lower than those in the SBM and LCSM50 groups during storage to 4 WK at 4℃. Albumen p H was not changed but decreased significantly in the LCSM₁00 group during storage to 4 and 8 WK at 4 ℃. Yolk index was significantly decreased in the LCSM₁00 group after storage of 8 WK. 2-DE analysis showed that there are 18 decreased proteins in abundance in the SBM group after 8 WK storage, however, there is 36 decreased proteins in abundance in the LCSM₁00 group after 8 WK storage. Therefore, this is consistent with the result of Experiment 3 that albumen and yolk quality was more easily afftected in the eggs of hens fed with LCSM₁00 diets.