Chinese Journal of Animal Nutrition >

2023 , Vol. 35 >Issue 7: 4321 - 4330

DOI: https://doi.org/10.12418/CJAN2023.402

RESEARCH PAPER

Effects of Lactobacillus plantarum on Muscle Quality and Antioxidant Function of Broilers under Ammonia Stress

  • MA Yanhua , 1 ,
  • LI Kui 2 ,
  • SHEN Yuanyuan 2 ,
  • LIU Jinsong 3 ,
  • FENG Jie 4 ,
  • ZHANG Ruiqiang , 2, *
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  • 1 College of Biological and Food Engineering, Puyang Vocational and Technical College, Puyang 457000, China
  • 2 Collage of Animal Science and Technology, College of Veterinary Medicine, Zhejiang A&F University, Hangzhou 311300, China
  • 3 Zhejiang Vegamax Biotechnology Co., Ltd., Anji 313300, China
  • 4 College of Animal Science, Zhejiang University, Hangzhou 310058, China
* lecturer, E-mail:

Received date: 2022-12-20

  Online published: 2023-07-11

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Abstract

This experiment was conducted to investigate the effects of Lactobacillus plantarum on muscle quality and antioxidant function of broilers under ammonia stress. A total of 288 healthy yellow-feathered broilers of 1-day-old were randomly divided into 3 groups with 8 replicates per group and 12 broilers per replicate. Broilers in the control group and ammonia stress group were fed a basal diet, and those in ammonia stress+Lactobacillus plantarum group were fed the basal diet supplemented with 500 mg/kg Lactobacillus plantarum (live bacteria count was 3.5×108 CFU/g). The experiment lasted for 70 days. From day 63 of the experiment, ammonia stress group and ammonia stress+Lactobacillus plantarum group were treated with ammonia stress for 7 days, while the control group was not affected by ammonia. The results showed as follows: 1) compared with the control group, the pH at 45 min and the redness (a*) value at 45 min and 24 h in breast muscle of broilers after slaughter in ammonia stress group were significantly decreased (P<0.05), the brightness (L*) value at 45 min and 24 h and the cooking loss at 24 h were significantly increased (P<0.05), the contents of moisture and ash were significantly increased (P<0.05), the contents of organic matter and crude protein was significantly decreased (P<0.05), the activities of total superoxide dismutase (T-SOD), catalase (CAT) and glutathione peroxidase (GPX) as well as total antioxidant capacity (T-AOC) were significantly decreased (P<0.05), the malondialdehyde content was significantly increased (P<0.05), and the mRNA relative expression levels of nuclear factor E2-related factor 2 (Nrf2), GPX1, CAT, superoxide dismutase 1 (SOD1) and superoxide dismutase 2 (SOD2) were significantly decreased (P<0.05). 2) Compared with ammonia stress group, the pH at 45 min and 24 h and the a* value at 24 h in breast muscle of broilers after slaughter in ammonia stress+Lactobacillus plantarum group were significantly increased (P<0.05), the L* value at 24 h and the drip loss at 24 and 48 h were significantly decreased (P<0.05), the contents of moisture and ash were significantly decreased (P<0.05), the crude protein content was significantly increased (P<0.05), the activities of T-SOD, CAT and GPX as well as T-AOC were significantly increased (P<0.05), and the SOD1 mRNA relative expression level was significantly increased (P<0.05). In conclusion, dietary Lactobacillus plantarum can improve the muscle quality and muscle composition in breast muscle of broilers under ammonia stress, and enhance the antioxidant capacity in muscle by regulating Nrf2 signaling pathway, so as to alleviate oxidative stress damage caused by ammonia stress.

Key words： Lactobacillus plantarum; ammonia stress; broilers; muscle quality; antioxidant function

Cite this article

MA Yanhua , LI Kui , SHEN Yuanyuan , LIU Jinsong , FENG Jie , ZHANG Ruiqiang . Effects of Lactobacillus plantarum on Muscle Quality and Antioxidant Function of Broilers under Ammonia Stress[J]. Chinese Journal of Animal Nutrition, 2023 , 35(7) : 4321 -4330 . DOI: 10.12418/CJAN2023.402

在对动物源性食品日益增长的需求推动下,国内畜禽生产模式由传统的自由放养稳步转向更专业化和规模化的集中饲养体系,这大幅提升了生产能力和生产效率[1]。而肉鸡作为家禽的重要组成部分,近年来市场不断发展,规模不断扩大,预计2023年全球年产量将达到1.052 6亿t[2]。但随着肉鸡养殖业规模化和集约化的提高,养殖中的问题也愈发凸显,养殖中排放的大量有害气体如硫化氢、氨气等,对肉鸡的危害非常严重,可造成肉鸡生长性能下降、疾病增多以及肉质变差等[3-4]。因此,如何保障肉鸡产品质量安全,促进肉鸡产业实现健康发展,是畜禽业发展需要解决的重要问题。
畜禽养殖场氨气浓度过高易导致畜禽总抗氧化能力降低、免疫力下降和抗病力降低,继而引发各种畜禽疾病,导致生长性能下降,死亡率大幅提高,造成惨重的经济损失[5]。过高浓度的氨气暴露还会影响畜禽采食,降低饲料转化率,导致蛋白质和能量水平的摄入减少,进而影响动物肌肉内氧化型肌纤维的比例和脂肪含量,使畜禽的肌肉品质下降[6]。
近年来,有关益生菌用于畜禽养殖的研究结果显示,许多益生菌均可增强机体免疫力和抗病力,抑制有害微生物生长,提高畜禽的生长性能[7⇓-9]。植物乳杆菌是一种革兰氏阳性菌,菌种呈短杆状,有时成对或成链状,不产芽孢,属于同型发酵乳酸菌,在发酵过程中只产生乳酸,是典型的兼性厌氧菌,有很强的发酵碳水化合物的能力,较耐盐[10]。研究显示,植物乳杆菌能够改善肠道形态结构和菌群结构,提高机体免疫能力和抗病力,促进动物生长[11⇓-13]。本课题组在此前的研究中发现,饲粮中添加植物乳杆菌具有提高肉鸡生长性能、促进消化器官发育和改善肉鸡有机物和粗蛋白质表观消化率的作用[14]。但是,有关植物乳杆菌调控肉鸡肌肉品质的作用及其发生机理的研究鲜见报道。因此,本文拟通过动物试验,研究植物乳杆菌对氨气应激下的肉鸡胸肌肌肉品质、肌肉成分、抗氧化能力和抗氧化相关基因表达的调控作用,为缓解肉鸡养殖中氨气应激导致的肉鸡肌肉品质下降和促进家禽养殖高质量发展提供理论依据。

1 材料与方法

1.1 试验材料

本试验所用植物乳杆菌由浙江某公司提供,活菌数为3.5×108 CFU/g。

1.2 试验设计

试验选择288只体重均匀、健康的1日龄黄羽肉鸡,随机分为3个组,分别是对照组(CON组)、氨气应激组(AM组)和氨气应激+植物乳杆菌组(AM+LP组),每组8个重复,每个重复12只鸡。对照组和氨气应激组饲喂基础饲粮,氨气应激+植物乳杆菌饲喂在基础饲粮中添加500 mg/kg植物乳杆菌的饲粮;各组饲粮均不添加任何抗生素。基础饲粮为粉料,参考NRC(1994)配制,其组成及营养水平见表1。
表1 基础饲粮组成及营养水平(风干基础)

Table 1 Composition and nutrient levels of basal diets (air-dry basis) %

项目
Items		 1~35日龄
1 to 35
days of age		 36~70日龄
36 to 70
days of age
原料Ingredients
玉米Corn 54.0 54.0
豆粕Soybean meal 23.2 15.0
膨化大豆Extruded soybean 5.0 3.0
玉米酒糟Corn distiller’s grains 8.2 8.0
米糠Rice bran 9.0
玉米麸Corn gluten 2.0
大豆油Soybean oil 1.6 3.5
石粉Limestone 2.1 2.4
发酵豆粕
Fermented soybean meal		 2.6
磷酸氢钙CaHPO4 2.0 1.8
食盐NaCl 0.3 0.3
预混料Premix1) 1.0 1.0
合计Total 100.0 100.0
营养水平Nutrient levels2)
代谢能ME/(MJ/kg)2) 12.20 12.92
粗蛋白质CP 20.20 17.10
赖氨酸Lys 1.20 1.06
蛋氨酸+半胱氨酸Met+Cys 0.90 0.73
钙Ca 0.86 0.74
总磷TP 0.60 0.58

1)预混料为每千克饲粮提供 The premix provided the following per kilogram of diets:VA 10 000 IU,VD3 2 000 IU,VE 30 IU,VK 1.3 mg,VB1 2.2 mg,VB2 8 mg,VB6 4 mg,VB12 13 μg,泛酸 pantothenic acid 10 mg,烟酸 niacin 1 mg,生物素 biotin 0.04 mg,氯化胆碱 choline chloride 400 mg,Fe 80 mg,Cu 7.5 mg,Mn 110 mg,Zn 65 mg,Se 0.3 mg,I 1.1 mg,载体为沸石 the carrier was zeolite。

2)代谢能为计算值,其余为实测值。ME was a calculated value, while the others were measured values.

试验期70 d。于试验第63天起,氨气应激组和氨气应激+植物乳杆菌组分别在08:30和16:30通入高纯氨气,使氨气浓度达到(25±3) mg/m3(氨气浓度的设定参考Yi等[15]的研究),持续7 d,对照组不通氨气。

1.3 饲养管理

本试验于浙江惠嘉生物科技股份有限公司试验基地开展。黄羽肉鸡饲养在气候舱中,气候舱可以自动控制舱内的温度、湿度、光照和通风量等因素,第1周气候舱内温度保持在34~36 ℃,随后每周降低2~3 ℃,温度降至20 ℃左右后保持不变;24 h光照(白天自然光照,晚上白炽灯补光);相对湿度保持在60%~70%;调节气候舱内换气扇的通风换气量,保证在下一次通入氨气时,氨气浓度处于15 mg/m3以上。试验期间鸡只自由采食和饮水。免疫接种及消毒、疾病预防按常规进行。

1.4 样品采集

氨应激试验进行7 d后,每组各选取生长状况相近的8只试验鸡(每个重复1只)空腹12 h后屠宰,采集左侧胸肌测定屠宰后45 min和24 h的肌肉品质,同时采集右侧胸肌样品保存于-80 ℃冰箱中,用于测定肌肉成分,分析鸡肉抗氧化能力和相关基因表达。

1.5 指标检测

1.5.1 肌肉品质的测定

分别在肉鸡屠宰后45 min和24 h,用装载肌肉专用电极pH计(HI99121,Hanna Instruments,Inc)测定胸肌的pH;用色差仪(CX0857,ColorFlexEZ,HunterLab)测定胸肌的亮度(L*)、黄度(b*)和红度(a*)值。切割并准确称取(7±2) g重的肌肉样品,将各组样品标号,悬挂放置于4 ℃冰箱中,在放置24和48 h后各测1次样品的重量并记录,计算各样品24和48 h的滴水损失。切割并准确称取(7±2) g重的肌肉样品,将各组样品标号,在70 ℃的蒸箱中放置30 min,再次称重并记录,计算各样品的蒸煮损失。

1.5.2 肌肉成分的测定

样品预处理:将屠宰后采集的胸肌样品适当切碎,称重后置于洁净的已知重量的培养皿中,在65 ℃烘箱中烘干48 h后,取出回潮24 h,称重并记录,粉碎后装于自封袋备用,后续测定参考《饲料分析及饲料质量检测技术》[16]。
水分和粗灰分含量的测定:取洁净坩埚烘干至恒重,于干燥器中冷却,称重,称取2 g左右的肉粉置于坩埚中,在105 ℃烘箱中烘干至恒重,于干燥器中冷却,称重,结合样品预处理水分含量结果计算胸肌实际水分含量。随后将坩埚放在电炉上炭化至无烟,移入陶瓷纤维马弗炉(SX2-4-10NP,广州越特科学仪器有限公司)中于505 ℃下灼烧5 h,等待炉温降下后取出,于干燥冷却,称重,结合样品预处理水分含量结果计算胸肌粗灰分实际含量。
粗蛋白质含量的测定:称取0.2 g左右胸肌肉粉试样,在消化管中消化,冷却消煮液后用常量蒸馏法吸收氨气,用盐酸标准滴定溶液滴定,记录滴定体积,结合样品预处理水分含量结果计算胸肌粗蛋白质含量。
粗脂肪含量的测定:称取胸肌肉粉试样2 g左右,放入滤纸中折成密封小包,放入105 ℃烘箱中烘干2 h,在干燥器中冷却后称重。将滤纸包放入预干燥处理的索氏抽提管中,在干燥的抽提瓶中加入足量石油醚,浸泡过夜后,进行抽提,控制石油醚回流速度适当。取出试样在通风橱中晾干滤纸包,在105 ℃烘箱中烘干至恒重,于干燥器中冷却,称重并记录,结合样品预处理水分含量结果计算胸肌粗脂肪含量。
有机物含量的测定:通过胸肌水分和粗灰分含量直接计算出胸肌有机物含量。

1.5.3 抗氧化指标的测定

取0.18 g左右胸肌肉试样放入2 mL匀浆管,加入9倍体积预冷处理的生理盐水,再加入适量的灭菌匀浆珠,经匀浆仪充分匀浆后,在4 000 r/min、4 ℃离心机中离心10 min,取上清液作为各组肉鸡的胸肌组织匀浆液,保存在-80 ℃冰箱中。根据南京奥青生物技术有限公司二喹啉甲酸(BCA)蛋白浓度测定试剂盒说明书测定蛋白浓度。根据南京建成生物工程研究所提供的试剂盒进行抗氧化指标的测定:采用亚铁离子(Fe2+)还原法测定组织中总抗氧化能力(T-AOC),采用酶促比色法测定谷胱甘肽过氧化物酶(GPX)活性,采用钼酸铵法测定过氧化氢酶(CAT)活性,采用黄嘌呤氧化酶法测定总超氧化物歧化酶(T-SOD)活性,采用硫代巴比妥酸法测定丙二醛(MDA)含量。

1.5.4 抗氧化相关基因表达的测定

根据TRIzol总RNA提取试剂及说明书,从肉鸡胸肌中提取总RNA。使用Nano-300分光光度仪测定总RNA的浓度。将RNA反转录为cDNA并将其储存在-20 ℃冰箱中,为随后实时荧光定量PCR(RT-qPCR)做准备。使用SYBR试剂盒进行RT-qPCR扩增反应。PCR培养程序如下:95 ℃预变性5 min后进入循环;95 ℃变性30 s;58~60 ℃退火30 s;72 ℃延长30 s,共39次循环;熔解曲线分析,95 ℃ 5 s(温度变化率4.4 ℃/s),60 ℃ 1 min(温度变化率2.2 ℃/s),95 ℃(温度变化率0.11 ℃/s),收集方法为连续收集,每1 ℃ 5次。
使用参照基因β-肌动蛋白(β-actin,F:TGCTGTGTTCCCATCTATCG;R:TTGGTGACAATACCGTGTTCA)运行和归一化样品。最终,记录循环时间,并通过2-ΔΔCt法计算肉鸡胸肌抗氧化相关基因核因子E2相关因子2(Nrf2)、Kelch样环氧氯丙烷相关蛋白1(Keap1)、血红素加氧酶1(HO1)、CAT、超氧化物歧化酶1(SOD1)、超氧化物歧化酶2(SOD2)和GPX1 mRNA相对表达量,目的基因的引物序列见表2。
表2 引物序列

Table 2 Primer sequences

目的基因
Target genes		 登录号
Accession No.		 上游引物
Forward primers (5'—3')		 下游引物
Reverse primers (5'—3')
核因子E2相关因子2
Nrf2		 NM_205117.1 GATGTCACCCTGCCCTTAG CTGCCACCATGTTATTCC
Kelch样环氧氯
丙烷相关蛋白1
Keap1		 KU_321503.1 GTACCAGATCGACAGCGTGG GGCAGTGGGACAGGTTGAAG
血红素加氧酶1
HO1		 NM_205344 GGTCCCGAATGAATGCCCTTG ACCGTTCTCCTGGCTCTTGG
谷胱甘肽过氧化物酶1
GPX1		 NM_001277853.1 CGCTACAGCCGCCACTT TTCGGAGAATCCCACAACG
过氧化氢酶
CAT		 NM_001031215.1 CGTTGGCGGTAGGAGTC CCAGTGGTCAAGGCATCT
超氧化物歧化酶1
SOD1		 NM_205064.1 TTGTCTGATGGAGATCATGGCTTC TGCTTGCCTTCAGGATTAAAGTGAG
超氧化物歧化酶2
SOD2		 NM_204211.1 CAGATAGCAGCCTGTGCAAATCA GCATGTTCCCATACATCGATTCC

1.6 数据统计分析

采用Excel 2021对试验数据进行初步整理,随后使用SPSS 21.0软件进行统计分析,采用单因素方差(one-way ANOVA)进行差异显著性检验,并采用Tukey’s HSD法进行多重比较检验,试验结果以平均值和均值标准误表示,以P<0.05作为差异显著的判断标准。

2 结果

2.1 植物乳杆菌对氨气应激肉鸡肌肉品质的影响

由表3可知,与对照组相比,氨气应激组肉鸡屠宰后胸肌45 min的pH显著降低(P<0.05),屠宰后胸肌45 min和24 h的L*值显著提高(P<0.05),屠宰后胸肌45 min和24 h的a*值显著降低(P<0.05),屠宰后胸肌24 h的蒸煮损失显著提高(P<0.05)。与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡屠宰后胸肌45 min和24 h的pH显著提高(P<0.05),屠宰后胸肌24 h的L*值显著降低(P<0.05),屠宰后胸肌24 h的a*值显著提高(P<0.05),屠宰后胸肌24和48 h的滴水损失显著降低(P<0.05)。此外,氨气应激+植物乳杆菌组肉鸡肌肉品质指标与对照组相比差异均不显著(P>0.05)。
表3 植物乳杆菌对氨气应激肉鸡肌肉品质的影响

Table 3 Effects of Lactobacillus plantarum on muscle quality of broilers under ammonia stress

项目
Items		 对照组
CON group		 氨气应激组
AM group		 氨气应激+
植物乳杆菌组
AM+LP group		 均值标准误
SEM		 P值
P-value
45 min pH 6.23a 5.88b 6.20a 0.052 0.004
24 h pH 6.47ab 6.19b 6.60a 0.058 0.007
45 min亮度45 min L* 29.30b 32.07a 30.84ab 0.413 0.016
45 min红度45 min a* 27.96a 25.78b 27.42ab 0.327 0.011
45 min黄度45 min b* 47.36 47.81 47.82 0.289 0.291
24 h亮度24 h L* 46.14b 51.70a 47.75b 0.717 0.002
24 h红度24 h a* 15.53a 13.33b 14.95a 0.273 0.001
24 h黄度24 h b* 27.89 28.81 27.86 0.338 0.113
24 h滴水损失24 h drip loss/% 2.74ab 3.40a 2.16b 0.161 0.003
48 h滴水损失48 h drip loss/% 7.13ab 9.47a 6.54b 0.486 0.026
24 h蒸煮损失24 h cooking loss/% 21.10b 25.48a 22.57ab 0.614 0.007

同行数据肩标不同字母表示差异显著(P<0.05),肩标相同字母或无字母表示差异不显著(P>0.05)。下表同。

In the same row, values with different letter superscripts mean significant difference (P<0.05), while with the same letter or no letter superscripts mean no significant difference (P<0.05). The same as below.

2.2 植物乳杆菌对氨气应激肉鸡肌肉成分的影响

由表4可知,与对照组相比,氨气应激组肉鸡胸肌水分和粗灰分含量显著提高(P<0.05),有机物和粗蛋白质含量显著降低(P<0.05)。与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌水分和粗灰分含量显著降低(P<0.05),粗蛋白质含量显著提高(P<0.05)。各组之间肉鸡胸肌粗脂肪含量无显著差异(P>0.05)。
表4 植物乳杆菌对氨气应激肉鸡肌肉成分的影响

Table 4 Effects of Lactobacillus plantarum on muscle composition of broilers under ammonia stress %

项目
Items		 对照组
CON group		 氨气应激组
AM group		 氨气应激+
植物乳杆菌组
AM+LP group		 均值标准误
SEM		 P值
P-value
水分Moisture 64.64b 67.91a 66.45b 0.853 0.025
有机物Organic matter 31.60a 27.29b 29.77ab 0.854 0.028
粗蛋白质Crude protein 17.27a 13.75b 16.73a 2.465 0.034
粗脂肪Ether extract 0.90 0.54 0.74 0.378 0.424
粗灰分Crude ash 3.77b 4.79a 3.79b 0.947 <0.001

2.3 植物乳杆菌对氨气应激肉鸡抗氧化能力的影响

由表5可知,与对照相比,氨气应激组肉鸡胸肌T-SOD、CAT和GPX活性以及T-AOC显著降低(P<0.05),而胸肌MDA含量显著提高(P<0.05)。与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌T-SOD、CAT和GPX活性及T-AOC显著提高(P<0.05)。此外,氨气应激+植物乳杆菌组肉鸡胸肌抗氧化指标与对照组相比差异均不显著(P>0.05)。
表5 植物乳杆菌对氨气应激肉鸡抗氧化能力的影响

Table 5 Effects of Lactobacillus plantarum on antioxidant capacity of broilers under ammonia stress

项目
Items		 对照组
CON group		 氨气应激组
AM group		 氨气应激+
植物乳杆菌组
AM+LP group		 均值标准误
SEM		 P值
P-value
总抗氧化能力T-AOC/(U/mg prot) 0.16a 0.07b 0.18a 0.013 <0.001
总超氧化物歧化酶T-SOD/(U/mg prot) 0.22a 0.11b 0.23a 0.807 0.001
过氧化氢酶CAT/(U/mg prot) 5.00a 3.02b 4.69a 0.155 0.001
谷胱甘肽过氧化物酶GPX/(U/mg prot) 17.20a 12.57b 18.91a 0.317 0.015
丙二醛MDA/(nmol/mg prot) 0.74b 1.23a 1.01ab 0.064 0.003

2.4 植物乳杆菌对氨气应激肉鸡抗氧化相关基因表达的影响

由表6可知,与对照组相比,氨气应激组肉鸡胸肌Nrf2、GPX1、CAT、SOD1和SOD2 mRNA相对表达量显著降低(P<0.05),胸肌HO1 mRNA相对表达量显著提高(P<0.05)。与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌SOD1 mRNA相对表达量显著提高(P<0.05),胸肌HO1 mRNA相对表达量显著降低(P<0.05)。此外,氨气应激+植物乳杆菌组肉鸡胸肌抗氧化相关基因mRNA相对表达量与对照组相比差异均不显著(P>0.05)。
表6 植物乳杆菌对氨气应激肉鸡抗氧化相关基因表达的影响

Table 6 Effects of Lactobacillus plantarum on expression of antioxidant-related genes of broilers under ammonia stress

项目
Items		 对照组
CON group		 氨气应激组
AM group		 氨气应激+
植物乳杆菌组
AM+LP group		 均值标准误
SEM		 P值
P-value
核因子E2相关因子2 Nrf2 1.13a 0.50b 1.03ab 0.102 0.018
Kelch样环氧氯丙烷相关蛋白1 Keap1 2.36 1.50 2.11 0.191 0.168
血红素加氧酶1 HO1 0.83b 1.65a 0.98b 0.110 0.002
谷胱甘肽过氧化物酶1 GPX1 1.54a 0.68b 1.17ab 0.124 0.011
过氧化氢酶CAT 1.38a 0.85b 1.19ab 0.077 0.009
超氧化物歧化酶1 SOD1 1.91a 1.09b 1.63a 0.107 0.002
超氧化物歧化酶2 SOD2 1.95a 1.17b 1.74ab 0.115 0.009

3 讨论

3.1 植物乳杆菌对氨气应激肉鸡肌肉品质的影响

pH、肉色、滴水损失和蒸煮损失是用来衡量畜禽肉品质的常规指标,这些指标之间密切相关[17-18]。pH对畜禽肌肉的颜色、风味、嫩度、系水力和保质期均有影响,是衡量肉品质性状的重要指标[19]。一般在测定肌肉色泽时,L*值越小,说明肌肉的颜色越浅,肌肉不发白,肉的品质越好;而a*值越高,则表示肌肉颜色越好[20]。肌肉系水力指肌肉的保水能力,通常用滴水损失和蒸煮损失作为指标来评估肌肉的系水力,滴水损失和蒸煮损失越低,则系水力越高[21]。本研究中,与对照组相比,氨气应激组肉鸡屠宰后胸肌45 min的pH以及45 min和24 h的a*值显著降低,45 min和24 h的L*值及24 h的蒸煮损失显著提高。与本研究一致,魏凤仙等[22]研究亦指出,氨气应激提高了肉鸡胸肌L*值,降低了胸肌a*值。同时,与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡屠宰后胸肌45 min和24 h的pH显著提高,24和48 h的滴水损失以及24 h的蒸煮损失显著降低,24 h的L*值显著降低,24 h的a*值显著提高,这提示植物乳杆菌可调节肉鸡胸肌肌肉保水性能,有效改善肉鸡肌肉品质。

3.2 植物乳杆菌对氨气应激肉鸡肌肉成分的影响

鸡肉主要是由水分、有机物、粗蛋白质、粗脂肪和粗灰分等五大营养成分组成,且代表了鸡肉的营养价值[23]。本研究中,与对照组相比,氨气应激组肉鸡胸肌水分和粗灰分含量显著提高,有机物和粗蛋白质含量显著降低,说明氨气应激会降低肉鸡肌肉的营养价值。同时,与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌水分和粗灰分含量显著降低,粗蛋白质含量显著提高。本实验室前期研究表明,饲粮中添加植物乳杆菌可提高肉鸡消化酶活性,改善有机物和粗蛋白质的表观消化率,提高肉鸡消化器官指数,并提高肉鸡末重和平均日增重[14],这与本试验肌肉成分的结果一致。Wang等[24]报道,植物乳杆菌处理显著上调了21日龄肉鸡回肠黏膜中脂肪酸结合蛋白1(FABP1)和钠依赖性葡萄糖转运蛋白-1(SGLT-1)的mRNA表达,显著提高了产短链脂肪酸(SCFA)细菌的相对丰度。这提示植物乳杆菌可能通过调控肠道营养转运蛋白的表达,从而改善肉鸡对饲粮中营养物质的转运和吸收利用。也有研究指出,植物乳杆菌可以增加肠道微生物的碳水化合物代谢相关途径,降低肠道微生物的核苷酸生物合成相关途径,最终促进饲粮消化成可吸收的营养物质,使肉鸡的消化和吸收效率提高,从而改善肌肉营养物质成分的沉积[25]。

3.3 植物乳杆菌对氨气应激肉鸡抗氧化能力的影响

氧化应激是机体中常见的过程,它可以在体内产生多种活性氧(ROS),过量的ROS会损害蛋白质、核酸等生物大分子,发生脂质氧化,产生大量的MDA,导致组织损伤,从而助长疾病的发展[26]。抗氧化酶体系物质如CAT、GPX、谷胱甘肽(GSH)和超氧化物歧化酶(SOD)对于清除过量的ROS至关重要,从而影响肉鸡肌肉的氧化还原状态平衡[27]。本研究中,与对照组相比,氨气应激组肉鸡胸肌T-AOC及T-SOD、CAT和GPX活性显著降低,而胸肌MDA含量显著提高。赵天等[28]研究发现,氨气可引起肉羊应激,导致肉羊血清SOD、CAT活性显著降低,血清MDA含量提高;魏凤仙等[22]在探究高浓度氨气暴露对肉仔鸡机体抗氧化性能的影响时发现,与低浓度氨气对照组相比,高浓度氨气降低了肉仔鸡肌肉中T-AOC及SOD和GPX活性,这与本研究结果类似。这说明当机体在氨气应激下,体内的氧化体系和抗氧化体系失衡,体内ROS自由基可能发生改变。此外,与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌T-AOC及T-SOD、CAT和GPX活性显著提高。由此推测,植物乳杆菌可能是通过激活机体内的抗氧化酶表达抑制自由基的产生,从而提高机体抗氧化能力,减少细胞损伤[29]。

3.4 植物乳杆菌对氨气应激肉鸡抗氧化相关基因表达的影响

Nrf2作为一种含碱性亮氨酸拉链结构的转录因子,广泛表达于机体的各个组织器官中,是细胞解毒反应和抗氧化系统的主要调控因子[30]。机体中高表达的Nrf2能够减少过氧化氢诱导产生ROS和MDA[31],Keap1-Nrf2信号通路能够感知机体氧化和抗氧化系统失衡,调控细胞的氧化还原状态,是细胞对抗氧化应激的主要防御机制,同时能够促进SOD、GPX等抗氧化相关因子的表达[32]。本研究中,与对照组相比,氨气应激组肉鸡胸肌Nrf2、GPX1、CAT、SOD1和SOD2 mRNA相对表达量显著降低。Zhang等[33]研究发现,热应激会导致肉鸡肝脏Nrf2、GPX和SOD1 mRNA表达量显著降低。由此推测,氨应激与热应激对机体抗氧化功能的影响机制类似,都是通过抑制Nrf2信号通路的激活,进而抑制SOD、GPX等多种抗氧化酶的表达,造成肉鸡的氧化应激损伤,破坏机体氧化和抗氧化系统的平衡,这也与氨气应激导致肉鸡胸肌T-AOC及T-SOD、CAT和GPX活性降低的结果对应。此外,本研究发现,氨气应激提高了肉鸡胸肌HO1 mRNA相对表达量,提示机体抗氧化状态可能受其他信号的调控,其具体机制尚需进一步研究。与氨气应激组相比,氨气应激+植物乳杆菌组肉鸡胸肌SOD1 mRNA相对表达量显著提高,推测植物乳杆菌可能通过诱导Nrf2基因的表达,提高抗氧化酶的活性,减少肉鸡胸肌肌肉细胞中ROS的含量,使氧化和抗氧化系统恢复平衡,减轻氧化应激的损伤,达到保护胸肌细胞发育的目的[26]。

4 结论

饲粮中添加植物乳杆菌能够提高氨气应激肉鸡胸肌肌肉品质,改善肌肉成分,并通过调控Nrf2信号通路增强肌肉抗氧化能力,缓解氨气应激造成的氧化应激损伤。

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