【作者】 刘振春；

【导师】 吴伟；

【作者基本信息】 吉林农业大学 ， 动物营养与饲料科学， 2011， 博士

【摘要】 鹅肥肝是一种高级营养食品,富含蛋白质、脂肪、维生素、卵磷脂、甘油三酯、各种酶、核糖核酸、脱氧核糖核酸和多种微量元素,具有较高的经济价值。鹅肥肝的形成受多种因素如遗传、营养、填饲方法、温度等的影响。鹅肥肝的脂肪沉积与其体内甘油三酯、高密度脂蛋白、脂肪酸合成酶活性的动态变化密切相关。从分子水平探讨填饲过程中FAS的多寡和活性的高低等动态变化规律,对控制动物肝脏脂肪的沉积具有重要意义,进而从分子生物学的角度研究酶活性在填饲诱发脂肪沉积差异中的表达与调控,不仅能明确调节机制,还可以为人类的肝病、糖尿病、心血管疾病等肝脂质代谢异常的研究提供某些依据。本研究以朗德鹅为研究对象,通过填饲试验及FQ-PCR等分子生物学技术,系统地研究在相同营养条件下不同填饲周龄朗德鹅的产肝性能、血清生化指标、肝组织中营养成分、不同填饲周龄FAS基因表达规律,揭示脂肪沉积规律和“肥肝”形成的某些分子机制,为进一步研究朗德鹅科学填饲及分子营养提供技术参考和依据。本研究还对鹅肥肝在不同贮藏温度和生物保鲜剂的贮藏特性进行了初步研究。通过安装电子感应称,比较准确地测定每只鹅不同周龄的填饲量,进而测定脂肪酸合成酶FAS在不同周龄的表达量及不同周龄肝内外脂肪的沉积量,从三位一体角度来研究填饲对脂肪合成酶的表达影响及其对肥肝增重的作用,为今后肥肝填饲提供有价值的技术参数。研究结果如下：1不同填饲周龄朗德鹅填饲量、体重增长及脂肪沉积规律朗德鹅平均每天及每周填饲量差异显著(P<0.05)；填饲不同周龄朗德鹅肝屠比差异不显著(P>0.05),料肝比差异极显著(P<0.01)。鹅肥肝重显著增加,填饲不同周龄肥肝重分别增加了130.48%、92.98%、72.38%、23.60%,肝内脂肪沉积差异极显著(P<0.01)。填饲3周鹅肥肝即达到A级标准,平均重达805g,肝重指数增加了0.7,提高了666.67%；填饲4周鹅肥肝平均重995g,提高了847.62%。填饲第3周肝重增加迅速,且与填饲量正相关,料肝比最低,因此填饲期应选择3周为宜。填饲不同周龄朗德鹅屠宰体重、肠脂重、腹脂重及皮脂重等性能显著增加。随填饲周龄增加,肝外组织脂肪沉积也随之增加。填饲不同周龄朗德鹅体重分别增加了15.63%、34.51%、46.26%、47.46%,差异显著(P<0.05)；肠脂重分别增加了39.85%、55.91%、31.72%、23.30%,差异显著(P<0.05)；腹脂重分别增加了27.88%、36.84%、38.73%、6.53%,差异显著(P<0.05)；皮脂重分别增加了341.91%、32.28%、21.07%、11.01%,差异显著(P<0.05)。2填饲不同周龄朗德鹅血清生化指标变化规律及肝内外脂肪沉积规律总胆固醇含量填饲前与填饲1周、填饲3周与填饲4周之间差异不显著(P>0.05)；而填饲1周与填饲2周、填饲2周与填饲3周之间差异显著(P<0.05)。甘油三酯除填饲3周和4周之间差异不显著(P>0.05)外,填饲前、填饲1周、填饲2周、填饲3周之间差异显著(P<0.05)。高密度脂蛋白只有填饲1周和填饲2周之间差异显著(P<0.05),其它填饲周龄之间差异不显著(P>0.05)。低密度脂蛋白和极低密度脂蛋白填饲前与填饲1周、填饲3周与填饲4周之间差异不显著(P>0.05),填饲1周、填饲2周、填饲3周之间差异显著(P<0.05)。总蛋白填饲前与填饲1周、填饲2周与填饲3周之间差异不显著(P>0.05)；填饲1周与填饲2周、填饲3周与填饲4周之间差异显著(P<0.05)。尿素和血糖各填饲周龄之间的水平差异不显著(P>0.05)。尿酸只有填饲前和填饲1周间差异显著(P<0.05),其它周龄之间差异不显著(P>0.05)。血清中ALT的活性在填饲前、填饲1周、填饲2周、填饲3周之间差异显著(P<0.05)；而填饲3周与填饲4周差异不显著(P>0.05)。血清中AST活性在填饲1周、填饲2周、填饲3周、填饲4周之间差异不显著(P>0.05)；而填饲前与填饲1周间差异显著(P<0.05)。血清中ALK的活性在填饲1周、填饲2周、填饲3周之间差异显著(P<0.05)；而填饲前与填饲1周、填饲3周与填饲4周间差异不显著(P>0.05)。填饲后肝脏中水分和蛋白质含量逐渐降低,总脂、甘油三酯、不饱和脂肪酸含量明显升高,各阶段水平均与填饲前差异显著(P<0.05)。而饱和脂肪酸差异不显著(P>0.05)。鹅肥肝在填饲前、填饲不同周龄的卵磷脂含量分别为1.75%、2.40%、4.85%、6.85%、7.90%,差异显著(P<0.05)；从填饲前到填饲4周含量增加4倍左右。3填饲不同周龄FAS基因表达及肝内外脂肪沉积规律FAS基因在填饲不同周龄的表达量逐渐增高。肥肝组织中FAS基因表达丰度与肥肝重、体重、肠脂、腹脂和皮脂均呈显著正相关(P<0.05)。肥肝组织中FAS基因表达量与肥肝组织中的总脂、饱和脂肪酸及不饱和脂肪酸、甘油三酯均呈显著正相关(P<0.05)。肥肝组织中FASmRNA表达量与血清中ALT、AST、ALK活性无显著相关性(P>0.05)。FAS表达量直接影响肥肝的沉积效率,肝脏FAS活性越高,肥肝性能就越好。FAS基因在鹅肥肝形成过程及体脂沉积过程中具有重要调控作用,它是促进鹅肝内外脂肪沉积的调控基因。4鹅肥肝的贮藏特性鹅肥肝在-20℃条件下冻藏效果最好。正交试验并经验证,0.05%溶菌酶、0.3%乳酸链球菌素、5%山梨酸钾的复合保鲜剂为鹅肥肝保鲜剂的效果最好。更多还原

【Abstract】 Goose fatty liver is a senior nutritional food with high economic value, which is rich in protein, fat, vitamin, lecithin, TG, DNA, RNA and microelement. Formation of goose fatty liver is influenced by many factors such as heredity, nutrition, overfeeding method, temperature, etc. Fat deposition in goose fatty liver was closely correlated with dynamic changes of TG, HDL and activity of FAS in the body. Researching the dynamic changes of FAS and its activity during the force-feeding in molecular level is significant for controlling fat deposition in animal liver. It is very important to research expression and regulation of enzyme during the period of fat deposition induced by overfeeding from the aspect of molecular biology since it not only identifies the regulation mechanism but also provides reference for the study of disease of liver, cardiovascular and diabetes for humane abnormal fat metabolize.Landes goose was researched as object by means of biology technology of FQ-PCR and force-feeding experiment. Properties of liver, biochemical index of serum, nutrients in liver tissue under the same nutritional conditions was systematically studied, including expression regulations of FAS gene under different week-old of force-feeding. Fat deposition law and some molecular mechanism of formation of fatty liver" was revealed. And it can provide some consultancy and reference for further study of molecular nutrition and Landes goose scientifical force-feeding. Storage characteristics of storage temperature and biological preservative on goose fatty liver were preliminarily conducted in the paper. Amount of force-feeding of each goose in different week-old was accurately measured by electronic sensor installed. Then expression amount of FAS and deposition in and out of the livers at different ages were measured. From the perspective of the trinity, effects of force-feeding on the expression and role of weight gaining of fatty liver were investigated, which provided valuable technical parameters for future research of fatty liver fore-feeding. Results obtained were shown as follows:1. Regulation of the number of feedings weight and deposition of fat in Landes goose liver with different feeding weeksThere were significant discrepants in the daily and weekly weight of feed in average (P＜0.05) and the ratio of the liver to slaughtering weight (P>0.05) during feeding weeks. Simultaneously, the ratio of material to liver showed extreme significant (P<0.01).The weight of liver increased obviously by 130.48%.92.98%、72.38%、23.60%respectively from one to four feeding weeks. In addition, the livers which were 805g in average came to A standard in the third week and the index of liver weight increased 0.7 by 666.67%. The livers with 995g in average in the fourth week were increased by 847.62%. The weight of livers which had a positive correlation with the number of feeding increased obviously and the ratio of material to liver came to the least.Force-feeding three weeks was appropriate.The weight of slaughter, intestinal fat, abdominal fat and leather fat increased significantly after each feeding week. What’s more, the deposition of extra-hepatic adipose tissue also increased significantly with time extended. The weight of goose with different feeding weeks raised by 15.63%、34.51%、46.26%、47.46%respectively(P＜0.05). In addition, the weight of intestinal fat was increased obviously by 39.85%、55.91%、31.72%、23.30% respectively(P＜0.05), the weight of abdominal fat also raised significantly by 27.88%、36.84%、38.73%、6.53%respectively (P＜0.05),and the weight of leather fat also increased significantly by 341.91%、32.28%、21.07%、11.01%,respectively (P＜0.05).2 The law of blood lipid metabolism and the fat deposition with different feeding weeks in Landes gooseAlthough the content of TC was not significant between pre-overfeeding and first feeding week, the third feeding week and the forth (P>0.05), it was significant between first feeding week and second feeding week, the second week and the third (P＜0.05). The differences of TG level were significant in different feeding weeks except the difference between the third week and the forth (P＜0.05). The level of HDL was only significantly different between the first week and the second (P＜0.05). The levels of LDL and VLDL were obviously different among the first, second and the third feeding week (P＜0.05). The contents of total protein were significant between the first and the second feeding week, the third and the forth week (P＜0.05). The urea and glucose in blood were insignificant among different weeks (P>0.05). The level of UA was significant between the pre-overfeeding and the first week (P＜0.05). The activity of ALT in blood serum was significantly different among the different feeding weeks from the first to the forth (P＜0.05). In addition, the activity of AST in blood serum was not significant among different feeding weeks from the first to the forth week (P>0.05). What’s more, the activity of ALK in blood serum was obviously different among the first, the second and the third feeding week (P＜0.05). What’s more, the contents of water and protein gradually decreased. The contents of total fat、TC、USFA increased significantly(P<0.05), but SFA not obviously(P>0.05). The content of lecithin in fatty livers were 1.75%、2.40%、4.85%、6.85%、7.90%with the pre-overfeeding and the different feeding weeks (P＜0.05). The content increased four times from pre-overfeeding to the forth week.3Correlation research between expression of FAS gene and fat deposition in and out of liver under different feeding weeks The expression of FAS increased regularly among the different feeding weeks. There was a positive correlation between the expression of FAS gene and the weight of fatty liver、weight、intestinal fat、abdominal fat and leather fat(P＜0.05). What’s more, there also was a positive genetic correlation between FAS mRNA expression in fatty liver tissue and total fat, SFA and USFA, TC (P＜0.05). In addition, there was no significant correlation between FAS mRNA expression in fatty liver and ALT, AST activity in serum (P＞0.05). In one word, the expression of FAS gene directly influenced the rate of deposition.The higher activity expression of FAS genes, the more perfect the fatty liver was. The result showed that FAS gene had a important regulation function in the forming of fatty liver in goose and the deposition of fat, thus it is the regulatory gene to promote the fat deposits inside goose liver.4 Storage characteristics of goose fatty liverGoose liver frozen under the condition of-20℃is best. Orthogonal test verified that compound preservative including 0.05% lysozyme,0.3%lactic acid streptocin,5%potassic sorbic acid were best compound preservative for goose fatty liver.更多还原