日粮ω6/ω3比例对肉鸡脂肪酸组成及肉品质的影响
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摘要
本论文通过四个试验研究了日粮不同ω6/ω3多不饱和脂肪酸(polyunsaturated fatty acids, PUFA)比例(ω6/ω3)对肉鸡生产性能、屠体品质、脂肪沉积、肉品质及脂肪酸组成的影响;从脂质代谢关键基因、肉质候选基因以及脂肪酸代谢基因的mRNA表达水平上,探讨了日粮ω6/ω3对脂肪代谢的影响机制;建立了鸡原代肝实质细胞分离培养方法,研究了脂肪酸对体外肝细胞增殖的影响。
试验一研究了日粮不同ω6/ω3比例对AA鸡生产性能、屠体品质、脂肪沉积、肉品质及脂肪酸组成的影响。结果表明:①饲喂10:1的日粮显著增加6周龄AA鸡的体重和日采食量;②随着日粮ω6/ω3比值的降低,屠宰率和净膛率显著下降;低ω6/ω3比值的日粮增加腹脂率、皮下脂肪和肌间脂肪;③日粮ω6/ω3比值从18下降到2.6时,胸、腿肌滴水损失显著下降;除日粮比值从5降到2.6时胸肌b*值下降外,胸、腿肌变化规律相似,a*值和b*值变化趋势相反:a*值下降,b*值上升;④日粮ω6/ω3的降低,ω3 PUFA中ALA、EPA和DPA显著增加;⑤肝脏中ALA含量低于胸肌,而其长链衍生物C20:3、EPA和DHA的含量则是肝脏中含量较高。本试验证明,在日粮3 %的油脂添加水平下,通过调整油脂类型来降低日粮ω6/ω3,能在一定程度改善AA鸡肉品质,并有效地增加胸肌中人体必需的ω3长链PUFA沉积,获得营养丰富的鸡肉。
试验二研究了长期改变日粮脂肪酸组成对北京油鸡脂肪沉积、肉品质及脂肪酸组成的影响。结果表明:①饲喂ω6/ω3为5:1的日粮能提高北京油鸡体重和饲料报酬。②饲喂10:1的日粮,肌间脂带宽和胸、腿肌IMF显著(P < 0.001)高于饲喂其它比值的日粮;除腿肌IMF在13周龄最低外,其他屠体指标随着日龄的增加而极显著(P < 0.001)增加。③当日粮比值从27降至5时,胸、腿肌剪切力、b*、L*值以及胸肌滴水损失和a*值和IMP含量显著增加;随着日龄的增加,胸、腿肌pH值、a*值显著下降;13周龄时胸肌剪切力和IMP含量达到最大,滴水损失最小。④随着日粮ALA含量的增加,胸肌和肝脏中大多数ω3 PUFA(ALA、EPA和DPA)而极显著增加(P < 0.001),但DHA不受日粮的影响。⑤肝脏组织中LA和ALA的含量低于相应的胸肌;日龄对肝脏和胸肌脂肪酸组成的影响存在差异,随着日龄的增加,肝脏ALA、DPA和总的ω3 PUFA含量在13周龄时最高;而胸肌大多数的ω3 PUFA(EPA、DPA、DHA和总ω3 PUFA)、所有的ω6 PUFA随日龄增加显著减少(P < 0.001)。本试验进一步证明,降低日粮ω6/ω3能够显著增加ALA向其长链衍生物EPA和DPA的转化能力;10:1的日粮显著增加IMF含量,可能是由于该比值能够增强脂肪酸的内源合成能力;13周龄时胸肌ω6/ω3最低、IMP最高,为北京油鸡13周龄上市提供了新的依据。
试验三利用Q-PCR技术研究日粮ω6/ω3对不同品种、日龄肉鸡肉品质和脂类代谢相关基因表达的影响。结果表明:①10和2.6的ω6/ω3比值能显著增加6周龄北京油鸡Δ6 FAD基因的表达水平;6周龄Δ6 FAD基因的表达显著高于13和22周龄。②北京油鸡A-FABP mRNA表达水平随日龄的增加而显著增加。③随着日粮ω6/ω3的降低,腹脂LPL mRNA表达显著增加;北京油鸡LPL表达水平在6周龄时最高。④日粮ω6/ω3的改变对ALDH1A1和CAPN1影响极显著,但影响趋势相反,随着日粮比值的降低,ALDH1A1显著降低,而CAPN1却显著上升。⑤10:1的日粮显著增加AA鸡和北京油鸡Apo-AⅠ基因表达,油鸡该基因的表达要显著低于AA鸡。本研究发现,幼龄动物Δ6 FAD和LPL表达较高,A-FABP水平较低;日粮ω6/ω3对LPL、ALDH1A1、Apo-AⅠ和CAPN1影响显著;不同品种间Apo-AⅠ和H-FABP表达存在差异。上述结果为研究日粮脂肪酸组成对肉质和脂肪代谢提供了依据。
试验四研究了LA、ALA和不同LA/ALA比值对体外鸡肝实质细胞增殖的影响。结果表明:使用0.05 %的胶原酶Ⅱ进行原位二步灌流分离法,不需要大量的胶原酶,细胞分离效果好,细胞活率高。LA、ALA及不同比例的LA/ALA处理肝实质细胞72 h后,LA/ALA比值为10的处理组细胞增殖作用最强。
综上所述,动物试验表明,在较低的油脂添加水平下,降低ω6/ω3能够有效地增加鸡肉中ω3 PUFA的含量,分子试验也证明了该结果,同时发现了幼龄动物具有较强的向长链脂肪酸转化的能力。10:1的日粮显著增加北京油鸡IMF含量和Apo-AⅠ基因表达,同时体外研究发现,10:1的LA/ALA比值对鸡肝实质细胞增殖有促进作用,此结果可能与该比值对体内脂肪代谢及整个机体内环境的稳定与正常生长密切相关。
Four experiments were conducted to study the effects of dietaryω6/ω3 on growth performance, carcass traits, fat deposition, meat quality, fatty acids composition and expression of key genes related to meat quality or lipid metabolism in chicken. Method for isolation and culture of chicken hepatocytes was established, by which effect of fatty acids on proliferation of hepatocytes was examined.
Trail 1 was carried out to assess the effects of dietaryω6/ω3 on growth performance, carcass traits, fat deposition, meat quality and fatty acids composition of AA broiler. The results showed that birds fed 10: 1 diet incorporated significantly higher body weight (BW) and average daily feed intake (ADFI) than fed the other birds. Percentage of dressing (PD), eviscerated carcass (PEC) decreased significantly as dietaryω6/ω3 was reduced; percentage of abdominal fat (PAF), subcutaneous fat (SBF) and intermuscular fat (IF) were higher in birds fed lower ratio diets. Drip loss (DL) in both breast and thigh muscle increased significantly in birds fedω6/ω3 ratio from 18 to 2.6. Except b* value in breast muscle decreased as birds fed dietary ratio from 5 to 2.6, the tendency for breast and thigh muscle were almost similar; a* values progressively increased as the diets contained increasingω3 content, changes in b* were also significant but the pattern was almost the reverse for changes in a*. Decreasing the dietaryω6/ω3 clearly increased the contents of ALA, EPA and DPA, but not for DHA. Liver had lower ALA, higher C20: 3, EPA and DHA than breast muscle. The results clearly demonstrated that decreasing the dietaryω6/ω3 by supplementing 3 % oil, improves meat quality of AA broiler, and increases deposition of desirableω3 andω6 long chain PUFA in the edible tissues, thereby achieving nutritionally enriched meat.
Trail 2 was conducted to study the effects of dietaryω6/ω3 on fat deposition, meat quality and fatty acids composition of Beijing-you chicken. The results showed that birds fed 5: 1 diet gained significantly higher BW and feed intake to gain ratio (F/G) than fed the other diets. For both breast and thigh muscle, birds fed the 10:1ω6/ω3 diet incorporated significantly (P < 0.05) higher intramuscular fat and IF than did birds fed the other diets; all carcass traits increased significantly (P < 0.001) from 6 to 22 wk. Shear force (SF), L* or b* value of meat color and DL in breast muscle decreased, while inosine-5’-monophosphate (IMP) and a* value increase, as the diets contained increasingω3 content, up to the 5:1ω6/ω3 (P < 0.01). Regardless of diet, a* value of meat color and pH at 1h postmortem declined (P < 0.001) between 6 and 22 wk. Chickens fed for 13 wk had the highest (P < 0.001) SF and IMP content, and lowest DL in breast muscle. Decreasing the dietaryω6/ω3 significantly increased (P < 0.001) the contents of ALA, EPA and DPA, but not DHA in both breast muscle and liver. Both LA and ALA were at lower levels in liver tissue lipid than in muscle. The most obvious difference in the effect of time on fatty acid composition between muscle and liver was that the percentage of ALA, DPA, and totalω3 PUFA were highest at 13 wk, but the percentage of mostω3 PUFA (EPA, DPA, DHA, and totalω3 PUFA) and allω6 PUFA decreased (P < 0.001) with time. This study clearly shows that decreasing the dietaryω6/ω3, without change in total dietary lipid, increases the efficiency of ALA to its long chain derivatives, with the exception of DHA. The pronounced effects of the diets on IMF could indicate that the 10:1ω6/ω3 ratio was optimal for de novo lipogenesis. The highest IMP content and lowestω6/ω3 in breast muscle at 13 wk provided proof for marketing the Beijing-you chicken.
Trail 3 investigated effects of dietaryω6/ω3 on gene expression related to meat quality and lipid metabolism in different breeds and age by Q-PCR. Results showed that the expression ofΔ6 FAD in abdominal fat was highest in Beijng-you chickens of 6 wk fed dietary ratio of 10 and 2.6. A-FABP expression increased significantly (P < 0.001) from 6 to 22 wk. Decreasing the dietaryω6/ω3 significantly increased (P < 0.001) the expression of LPL in abdominal fat, and the highest expression level for LPL occurred at 6 wk for Beijing-you chicken. The expression of CAPN1 progressively increased as the diets contained increasingω3 content, the changes in ALDH1A1 were also significant but the pattern was almost the reverse of changes in CAPN1. Apo-AⅠgene expression in Beijing-you chicken was lower than that in AA broiler, and fed 10: 1 diet was higher than other diets. In conclusion, the expression ofΔ6 FAD and LPL was higher in younger birds, LPL, ALDH1A1, Apo-AⅠand CAPN1 expression was significantly influenced by varying dietaryω6/ω3, there were significantly different of Apo-AⅠand H-FABP expression between two breeds.
Trail 4 examined effect of LA, ALA and different LA/ALA ratio on proliferation of chicken hepatocytes. The results showed that high purity and viability of hepatocytes could be obtained by two steps in situ circulatory perfusion method with little 0.05 % collagenaseП. MTT results showed that ratio of 10 for LA/ALA promoted proliferation of hepatocytes after culture for 72 h.
In conclusion, this study from in vivo suggests that, the efficiency of the chicken to convert ALA to EPA and DPA can be improved by feeding a diet of lowerω6/ω3 ratio with a relatively low amount of total added fat. The following molecular test demonstrated this result further, and found the best ability of conversion to long chain fatty acids occurred in younger birds. Furthermore, the results from in vitro that the 10: 1 ratio of LA/ALA can promote the proliferation of hepatocytes, along with the highest IMF content and Apo-AⅠexpression in Beijing-you chicken fed 10: 1 diet in vivo, may be closely associated with the influence of this ratio on lipid metabolism, stability and growth of organism.
试验一研究了日粮不同ω6/ω3比例对AA鸡生产性能、屠体品质、脂肪沉积、肉品质及脂肪酸组成的影响。结果表明:①饲喂10:1的日粮显著增加6周龄AA鸡的体重和日采食量;②随着日粮ω6/ω3比值的降低,屠宰率和净膛率显著下降;低ω6/ω3比值的日粮增加腹脂率、皮下脂肪和肌间脂肪;③日粮ω6/ω3比值从18下降到2.6时,胸、腿肌滴水损失显著下降;除日粮比值从5降到2.6时胸肌b*值下降外,胸、腿肌变化规律相似,a*值和b*值变化趋势相反:a*值下降,b*值上升;④日粮ω6/ω3的降低,ω3 PUFA中ALA、EPA和DPA显著增加;⑤肝脏中ALA含量低于胸肌,而其长链衍生物C20:3、EPA和DHA的含量则是肝脏中含量较高。本试验证明,在日粮3 %的油脂添加水平下,通过调整油脂类型来降低日粮ω6/ω3,能在一定程度改善AA鸡肉品质,并有效地增加胸肌中人体必需的ω3长链PUFA沉积,获得营养丰富的鸡肉。
试验二研究了长期改变日粮脂肪酸组成对北京油鸡脂肪沉积、肉品质及脂肪酸组成的影响。结果表明:①饲喂ω6/ω3为5:1的日粮能提高北京油鸡体重和饲料报酬。②饲喂10:1的日粮,肌间脂带宽和胸、腿肌IMF显著(P < 0.001)高于饲喂其它比值的日粮;除腿肌IMF在13周龄最低外,其他屠体指标随着日龄的增加而极显著(P < 0.001)增加。③当日粮比值从27降至5时,胸、腿肌剪切力、b*、L*值以及胸肌滴水损失和a*值和IMP含量显著增加;随着日龄的增加,胸、腿肌pH值、a*值显著下降;13周龄时胸肌剪切力和IMP含量达到最大,滴水损失最小。④随着日粮ALA含量的增加,胸肌和肝脏中大多数ω3 PUFA(ALA、EPA和DPA)而极显著增加(P < 0.001),但DHA不受日粮的影响。⑤肝脏组织中LA和ALA的含量低于相应的胸肌;日龄对肝脏和胸肌脂肪酸组成的影响存在差异,随着日龄的增加,肝脏ALA、DPA和总的ω3 PUFA含量在13周龄时最高;而胸肌大多数的ω3 PUFA(EPA、DPA、DHA和总ω3 PUFA)、所有的ω6 PUFA随日龄增加显著减少(P < 0.001)。本试验进一步证明,降低日粮ω6/ω3能够显著增加ALA向其长链衍生物EPA和DPA的转化能力;10:1的日粮显著增加IMF含量,可能是由于该比值能够增强脂肪酸的内源合成能力;13周龄时胸肌ω6/ω3最低、IMP最高,为北京油鸡13周龄上市提供了新的依据。
试验三利用Q-PCR技术研究日粮ω6/ω3对不同品种、日龄肉鸡肉品质和脂类代谢相关基因表达的影响。结果表明:①10和2.6的ω6/ω3比值能显著增加6周龄北京油鸡Δ6 FAD基因的表达水平;6周龄Δ6 FAD基因的表达显著高于13和22周龄。②北京油鸡A-FABP mRNA表达水平随日龄的增加而显著增加。③随着日粮ω6/ω3的降低,腹脂LPL mRNA表达显著增加;北京油鸡LPL表达水平在6周龄时最高。④日粮ω6/ω3的改变对ALDH1A1和CAPN1影响极显著,但影响趋势相反,随着日粮比值的降低,ALDH1A1显著降低,而CAPN1却显著上升。⑤10:1的日粮显著增加AA鸡和北京油鸡Apo-AⅠ基因表达,油鸡该基因的表达要显著低于AA鸡。本研究发现,幼龄动物Δ6 FAD和LPL表达较高,A-FABP水平较低;日粮ω6/ω3对LPL、ALDH1A1、Apo-AⅠ和CAPN1影响显著;不同品种间Apo-AⅠ和H-FABP表达存在差异。上述结果为研究日粮脂肪酸组成对肉质和脂肪代谢提供了依据。
试验四研究了LA、ALA和不同LA/ALA比值对体外鸡肝实质细胞增殖的影响。结果表明:使用0.05 %的胶原酶Ⅱ进行原位二步灌流分离法,不需要大量的胶原酶,细胞分离效果好,细胞活率高。LA、ALA及不同比例的LA/ALA处理肝实质细胞72 h后,LA/ALA比值为10的处理组细胞增殖作用最强。
综上所述,动物试验表明,在较低的油脂添加水平下,降低ω6/ω3能够有效地增加鸡肉中ω3 PUFA的含量,分子试验也证明了该结果,同时发现了幼龄动物具有较强的向长链脂肪酸转化的能力。10:1的日粮显著增加北京油鸡IMF含量和Apo-AⅠ基因表达,同时体外研究发现,10:1的LA/ALA比值对鸡肝实质细胞增殖有促进作用,此结果可能与该比值对体内脂肪代谢及整个机体内环境的稳定与正常生长密切相关。
Four experiments were conducted to study the effects of dietaryω6/ω3 on growth performance, carcass traits, fat deposition, meat quality, fatty acids composition and expression of key genes related to meat quality or lipid metabolism in chicken. Method for isolation and culture of chicken hepatocytes was established, by which effect of fatty acids on proliferation of hepatocytes was examined.
Trail 1 was carried out to assess the effects of dietaryω6/ω3 on growth performance, carcass traits, fat deposition, meat quality and fatty acids composition of AA broiler. The results showed that birds fed 10: 1 diet incorporated significantly higher body weight (BW) and average daily feed intake (ADFI) than fed the other birds. Percentage of dressing (PD), eviscerated carcass (PEC) decreased significantly as dietaryω6/ω3 was reduced; percentage of abdominal fat (PAF), subcutaneous fat (SBF) and intermuscular fat (IF) were higher in birds fed lower ratio diets. Drip loss (DL) in both breast and thigh muscle increased significantly in birds fedω6/ω3 ratio from 18 to 2.6. Except b* value in breast muscle decreased as birds fed dietary ratio from 5 to 2.6, the tendency for breast and thigh muscle were almost similar; a* values progressively increased as the diets contained increasingω3 content, changes in b* were also significant but the pattern was almost the reverse for changes in a*. Decreasing the dietaryω6/ω3 clearly increased the contents of ALA, EPA and DPA, but not for DHA. Liver had lower ALA, higher C20: 3, EPA and DHA than breast muscle. The results clearly demonstrated that decreasing the dietaryω6/ω3 by supplementing 3 % oil, improves meat quality of AA broiler, and increases deposition of desirableω3 andω6 long chain PUFA in the edible tissues, thereby achieving nutritionally enriched meat.
Trail 2 was conducted to study the effects of dietaryω6/ω3 on fat deposition, meat quality and fatty acids composition of Beijing-you chicken. The results showed that birds fed 5: 1 diet gained significantly higher BW and feed intake to gain ratio (F/G) than fed the other diets. For both breast and thigh muscle, birds fed the 10:1ω6/ω3 diet incorporated significantly (P < 0.05) higher intramuscular fat and IF than did birds fed the other diets; all carcass traits increased significantly (P < 0.001) from 6 to 22 wk. Shear force (SF), L* or b* value of meat color and DL in breast muscle decreased, while inosine-5’-monophosphate (IMP) and a* value increase, as the diets contained increasingω3 content, up to the 5:1ω6/ω3 (P < 0.01). Regardless of diet, a* value of meat color and pH at 1h postmortem declined (P < 0.001) between 6 and 22 wk. Chickens fed for 13 wk had the highest (P < 0.001) SF and IMP content, and lowest DL in breast muscle. Decreasing the dietaryω6/ω3 significantly increased (P < 0.001) the contents of ALA, EPA and DPA, but not DHA in both breast muscle and liver. Both LA and ALA were at lower levels in liver tissue lipid than in muscle. The most obvious difference in the effect of time on fatty acid composition between muscle and liver was that the percentage of ALA, DPA, and totalω3 PUFA were highest at 13 wk, but the percentage of mostω3 PUFA (EPA, DPA, DHA, and totalω3 PUFA) and allω6 PUFA decreased (P < 0.001) with time. This study clearly shows that decreasing the dietaryω6/ω3, without change in total dietary lipid, increases the efficiency of ALA to its long chain derivatives, with the exception of DHA. The pronounced effects of the diets on IMF could indicate that the 10:1ω6/ω3 ratio was optimal for de novo lipogenesis. The highest IMP content and lowestω6/ω3 in breast muscle at 13 wk provided proof for marketing the Beijing-you chicken.
Trail 3 investigated effects of dietaryω6/ω3 on gene expression related to meat quality and lipid metabolism in different breeds and age by Q-PCR. Results showed that the expression ofΔ6 FAD in abdominal fat was highest in Beijng-you chickens of 6 wk fed dietary ratio of 10 and 2.6. A-FABP expression increased significantly (P < 0.001) from 6 to 22 wk. Decreasing the dietaryω6/ω3 significantly increased (P < 0.001) the expression of LPL in abdominal fat, and the highest expression level for LPL occurred at 6 wk for Beijing-you chicken. The expression of CAPN1 progressively increased as the diets contained increasingω3 content, the changes in ALDH1A1 were also significant but the pattern was almost the reverse of changes in CAPN1. Apo-AⅠgene expression in Beijing-you chicken was lower than that in AA broiler, and fed 10: 1 diet was higher than other diets. In conclusion, the expression ofΔ6 FAD and LPL was higher in younger birds, LPL, ALDH1A1, Apo-AⅠand CAPN1 expression was significantly influenced by varying dietaryω6/ω3, there were significantly different of Apo-AⅠand H-FABP expression between two breeds.
Trail 4 examined effect of LA, ALA and different LA/ALA ratio on proliferation of chicken hepatocytes. The results showed that high purity and viability of hepatocytes could be obtained by two steps in situ circulatory perfusion method with little 0.05 % collagenaseП. MTT results showed that ratio of 10 for LA/ALA promoted proliferation of hepatocytes after culture for 72 h.
In conclusion, this study from in vivo suggests that, the efficiency of the chicken to convert ALA to EPA and DPA can be improved by feeding a diet of lowerω6/ω3 ratio with a relatively low amount of total added fat. The following molecular test demonstrated this result further, and found the best ability of conversion to long chain fatty acids occurred in younger birds. Furthermore, the results from in vitro that the 10: 1 ratio of LA/ALA can promote the proliferation of hepatocytes, along with the highest IMF content and Apo-AⅠexpression in Beijing-you chicken fed 10: 1 diet in vivo, may be closely associated with the influence of this ratio on lipid metabolism, stability and growth of organism.
引文
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