单一和复合乳酸菌添加水平对苜蓿青贮营养品质及蛋白组分的影响
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摘要
以第1茬现蕾期刈割的紫花苜蓿(Medicago sativa)为原料(干物质含量为42.56%),结合康奈尔碳水化合物-蛋白质体系(CNCPS),研究不同浓度的单一和复合乳酸菌对苜蓿青贮品质及蛋白组分的影响。发酵40d后取样分析其青贮发酵品质和营养成分,结果表明,所有乳酸菌处理组都能显著降低苜蓿青贮饲料的氨态氮含量(P<0.05),并且均未检测到丁酸;添加量为1×10~7 cfu·g~(-1)的单一乳酸菌处理组氨态氮含量最低,乳酸含量最高;除添加量为1×10~5cfu·g~(-1)的复合菌处理组外,其它乳酸菌处理组的pH均显著低于无添加剂处理组(P<0.05);乳酸菌处理后粗蛋白含量无显著变化(P>0.05);复合菌处理组的非蛋白氮含量显著低于无添加剂处理组(P<0.05);单一和复合乳酸菌都能降低蛋白组分中的非蛋白氮(PA)含量,增加快速降解真蛋白质(PB1)和中速降解真蛋白质(PB2)含量,复合菌处理的真蛋白质(PB)含量显著高于对照的(P<0.05)。综上所述,高浓度的单一植物乳杆菌(Lactobacillus plantarum)处理的苜蓿青贮发酵品质最好,复合菌能保护更多的真蛋白不被降解。
The objective of this study was to determine the effects of applying single or multiple lactic acid bacteria(LAB)at ensiling on the fermentation quality and protein fractions of alfalfa(Medicago sativa).The protein fractions of alfalfa silage were calculated by Cornell net carbohydrate and protein system(CNCPS).The silage material was the first-cut alfalfa(425.6g DM·kg-1 fresh herbage)harvested at budding stage.Forage was packed into 21 laboratory silos and ensiled for 40 days.The results showed that all treatments with LAB had significantly lower(P<0.05)concentration of NH_3-N than untreated group,and butyric acid was not detected in these treatments.The treatment of single-strain(Lactobacillus plantarum)at 1×10~7 cfu·g~(-1) fresh herbage had the lowest concentration of NH_3-N and the highest concentration of lactic acid.Silages treated with lactic acid bacteria(other than multi-strains at 1×10~5 cfu·g~(-1) fresh herbage)characterized by lower(P<0.05)pH value.There was no(P>0.05)treatment effect on the content of crude protein.The content of nonprotein nitrogen(NPN)in silages with all additives of LAB was reduced when compared with untreated,and silages treated with compound LAB characterized by lower(P<0.05)NPN.Additives with single and multistrains of LAB both resulted in alfalfa silage with low concentration of non-protein nitrogen(PA),high concentration of immediately degraded protein(PB1)and intermediately degraded protein(PB2).The concentration of true protein(PB)in silage with multi-strains of LAB was higher(P<0.05)than untreated.Therefore,high concentration of L.plantarum was beneficial to improve the fermentation quality of alfalfa,and the proteolysis,in some extent,could be inhibited by multiple LAB at ensiling.
The objective of this study was to determine the effects of applying single or multiple lactic acid bacteria(LAB)at ensiling on the fermentation quality and protein fractions of alfalfa(Medicago sativa).The protein fractions of alfalfa silage were calculated by Cornell net carbohydrate and protein system(CNCPS).The silage material was the first-cut alfalfa(425.6g DM·kg-1 fresh herbage)harvested at budding stage.Forage was packed into 21 laboratory silos and ensiled for 40 days.The results showed that all treatments with LAB had significantly lower(P<0.05)concentration of NH_3-N than untreated group,and butyric acid was not detected in these treatments.The treatment of single-strain(Lactobacillus plantarum)at 1×10~7 cfu·g~(-1) fresh herbage had the lowest concentration of NH_3-N and the highest concentration of lactic acid.Silages treated with lactic acid bacteria(other than multi-strains at 1×10~5 cfu·g~(-1) fresh herbage)characterized by lower(P<0.05)pH value.There was no(P>0.05)treatment effect on the content of crude protein.The content of nonprotein nitrogen(NPN)in silages with all additives of LAB was reduced when compared with untreated,and silages treated with compound LAB characterized by lower(P<0.05)NPN.Additives with single and multistrains of LAB both resulted in alfalfa silage with low concentration of non-protein nitrogen(PA),high concentration of immediately degraded protein(PB1)and intermediately degraded protein(PB2).The concentration of true protein(PB)in silage with multi-strains of LAB was higher(P<0.05)than untreated.Therefore,high concentration of L.plantarum was beneficial to improve the fermentation quality of alfalfa,and the proteolysis,in some extent,could be inhibited by multiple LAB at ensiling.
引文
[1]玉柱,孙启忠.饲草青贮技术.北京:中国农业大学出版社,2011:8-10,62-65,121-122.Yu Z,Sun Q Z.Forage Silage Technology.Beijing:China Agriculture Press,2011:8-10,62-65,121-122.(in Chinese)
[2]万里强,李向林,何峰.添加乳酸菌和纤维素酶对苜蓿青贮品质的影响.草业科学,2011,28(7):1379-1383.Wan L Q,Li X L,He F.The effects of adding lactic acid bacteria and cellulose on alfalfa silage quality.Pratacultural Science,2011,28(7):1379-1383.(in Chinese)
[3]张庆,张万军,田吉鹏,王雨,李旭娇,玉柱.乳酸菌青贮技术研究进展.草业科学,2014,31(2):328-333.Zhang Q,Zhang W J,Tian J P,Wang Y,Li X J,Yu Z.Advances in lactic acid bacteria silage technology research.Pratacultural Science,2014,31(2):328-333.(in Chinese)
[4]刘辉,卜登攀,吕中旺,李发弟,刘士杰,张开展,王加启.凋萎和不同添加剂对紫花苜蓿青贮品质的影响.草业学报,2015,24(5):126-133.Liu H,Bu D P,Lyu Z W,Li F D,Liu S J,Zhang K Z,Wang J Q.Effects of wilting and additives on fermentation quality of alfalfa(Medicago sativa)silage.Acta Prataculturae Sinica,2015,24(5):126-133.(in Chinese)
[5]Francisco E,Contreras-Govea,Muck R E,Broderick G,Weimer P J.Lactobacillus plantarum effects on silage fermentation and in vitro microbial yield.Animal Feed Science and Technology,2013,179(1):61-68.
[6]Weinberg Z G,Muck R E.New trends and opportunities in the development and use of inoculants for silage.FEMS Microbiology Reviews,1996,19(1):53-68.
[7]张增欣,邵涛.青贮添加剂研究进展.草业科学,2006,23(9):56-63.Zhang Z X,Shao T.Research progress in silage additive.Pratacultural Science,2006,23(9):56-63.(in Chinese)
[8]张庆,王显国,玉柱.乳酸菌添加剂和含水量对紫花苜蓿青贮的影响.中国奶牛,2015(18):37-40.Zhang Q,Wang X G,Yu Z.Effects of lactic acid bacteria inoculants and moisture content on fermentation quality of alfalfa silage.China Dairy Cattle,2015(18):37-40.(in Chinese)
[9]郭旭生,周禾,刘桂霞.苜蓿青贮过程中蛋白的分解及抑制方法.草业科学,2005,22(11):50-54.Guo X S,Zhou H,Liu G X.Proteolysis in alfalfa silage and its inhibiting method.Pratacultural Science,2005,22(11):50-54.(in Chinese)
[10]Nagel S A,Broderick G A.Effect of formic acid or formaldehyde treatment of alfalfa silage on nutrient utilization by dairy cows.Journal of Dairy Science,1992,75(1):140-154.
[11]Wu R D,Guo X S,Kazuo A.Characterization of peptides in ensiled alfalfa treated with different chemical additives.Animal Science Journal,2013,84(12):774-781.
[12]赵广永.用净碳水化合物-蛋白质体系评定反刍动物饲料营养价值.中国农业大学学报,1999,4(增刊):71-76.Zhao G Y.The Use of cornell net carbohydrate and protein system(CNCPS)for the evaluation of some Chinese feeds of ruminants.Journal of China Agricultural University,1999,4(S):71-76.(in Chinese)
[13]许庆方.影响苜蓿青贮品质的主要因素及苜蓿青贮在奶牛日粮中应用效果的研究.北京:中国农业大学博士学位论文,2005:24-26.Xu Q F.Studies of the factors of affecting alfalfa silage quality and the utilization of alfalfa silage in dairy cows diet.PhD Thesis.Beijing:China Agricultural University,2005:24-26.(in Chinese)
[14]Broderick G A,Kang J H.Automated simultaneous determination of ammonia and amino acids in ruminal fluids and in vitro media.Journal of Dairy Science,1980,63(1):64-75.
[15]张丽英.饲料分析及饲料质量检测技术.北京:中国农业大学出版社,2007:53-56,67-78.Zhang L Y.Feed Analysis and Forage Quality Detection Technology.Beijing:China Agriculture Press,2007:53-56,67-78.(in Chinese)
[16]Licitra G,Hernandez T M,Van Soest P J.Standardization of procedures for nitrogen fractionation of ruminant feeds.Animal Feed Science Technology,1996,57(4):347-358.
[17]Sniffen C J,O’Connor J D,Van Soest P J,Fox D G,Russell J B.A net carbohydrate and protein system for evaluating cattle diets:II.Carbohydrate and protein availability.Journal of Animal Science,1992,70(11):3562-3577.
[18]张金霞,乔红霞,刘雨田.水分和添加剂对紫花苜蓿青贮品质的影响.草业科学,2014,31(4):766-770.Zhang J X,Qiao H X,Liu Y T.Effects of moisture and additives on feed quality of alfalfa silage.Pratacultural Science,2014,31(4):766-770.(in Chinese)
[19]徐炜,师尚礼,张文渝,王兴中,王永福,倪兴铎,李小平,王新强,王得文,王皎.不同添加剂对低水分紫花苜蓿青贮品质的影响.草原与草坪,2014,34(1):49-54.Xu W,Shi S L,Zhang W X,Wang X Z,Wang Y F,Ni X D,Li X P,Wang X Q,Wang D W,Wang J.Effect of different additives on quality of low moisture alfalfa silage.Grassland and Turf,2014,34(1):49-54.(in Chinese)
[20]Yuan X,Guo G,Wen A.The effect of different additives on the fermentation quality,in vitro digestibility and aerobic stability of a total mixed ration silage.Animal Feed Science and Technology.2015,207:41-50.
[21]Zhang Q,Li X,Zhao M.Lactic acid bacteria strains for enhancing the fermentation quality and aerobic stability of Leymus chinensis silage.Grass and Forage Science,2015:DOI:10.1111/gfs.12190.
[22]Heron S J E,Edwards R A,Phillips P.Effect of pH on the activity of Lolium multiflorum protease.Journal of Science Food Agriculture,1989,46(3):267-277.
[23]Rooke J A,Armstrong D G.The importance of the form of nitrogen on microbial protein synthesis in the rumen of cattle receiving grass silage and intrarumen infusions of sucrose.British Journal of Nutrition,1989,61(1):113-121.
[24]Winters A L,Cockburn J E,Dhanoa M S,Merry R J.Effects of lactic acid bacteria in inoculants on changes in amino acid composition during ensilage of sterile and non-sterile ryegrass.Journal of Applied Microbiology,2000,89(3):442-451.
[25]Mckersie B D.Effect of pH on proteolysisin ensiled legume forage.Agronomy Journal,1985,77(1):81-86.
[26]Jones B A,Satter L D,Muck R E.Influence of bacterial inoculants and substrate addition to lucerne ensiled at different dry matter contents.Grass and Forage Science,1992,47(1):19-27.
[27]Broberg A,Jacobsson K,Strm K,Schnürer J.Metabolite profiles of lactic acid bacteria in grass silage.Applied and Environmental Microbiology,2007,73(17):5547-5552.
[28]Pichard G,van Soest P J.Protein solubility of ruminant feeds.In:Ithaca N Y.(ed).Proc Cornell Nutrition Conf.Department of Animal Science,Cornell University,1977:91-98.
[29]Rooke J A,Armstrong D G.The importance of the form of nitrogen on microbial protein synthesis in the rumen of cattle receiving grass silage and continuous intrarumen infusions of sucrose.British Journal of Nutrition,1989,61(1):113-121.
[30]Krishnamoorthy U C,Sniffen C J,Stem M D,van Soest P J.Evaluation of a mathematical model of rumen digestion and an in vitro simulation of rumen proteolysis to estimate the rumen-undegraded nitrogen content of feedstuffs.British Journal of Nutrition,1983,50(3):555-568.
[31]王燕,杨方,陈常栋,王海威,张微微,张永根.康奈尔净碳水化合物-蛋白质体系预测小肠可消化粗蛋白质含量.动物营养学报,2012,24(7):1274-1282.Wang Y,Yang F,Chen C D,Wang H W,Zhang W W,Zhang Y G.Predicting utilizable crude protein content in small intestine by cornell net carbohydrate and protein system.Acta Zoonutrimenta Sinica,2012,24(7):1274-1282.(in Chinese)
[32]Krishnamoorthy U C,Muscato T V,Sniffen C J,van Soest P J.Nitrogen fractions in selected feedstuffs.Journal of Dairy Science,1982,65(2):217-225.
[33]Francisco E C G,Richard E M,David R M,Paul J W.Microbial inoculant effects on silage and in vitro ruminal fermentation,and microbial biomass estimation for alfalfa,bmr corn,and corn silages.Animal Feed Science and Technology,2011,163(1):2-10.
[34]Nsereko V L,Smiley B K,Rutherford W M,Spielbauer A,Forrester K J,Hettinger G H,Harman E K,Harman B R.Influence of inoculating forage with lactic acid bacteria strains that produce ferulate esterase on ensilage and ruminal degradation of fiber.Animal Feed Science and Technology,2008,145(1-4):122-135.
[2]万里强,李向林,何峰.添加乳酸菌和纤维素酶对苜蓿青贮品质的影响.草业科学,2011,28(7):1379-1383.Wan L Q,Li X L,He F.The effects of adding lactic acid bacteria and cellulose on alfalfa silage quality.Pratacultural Science,2011,28(7):1379-1383.(in Chinese)
[3]张庆,张万军,田吉鹏,王雨,李旭娇,玉柱.乳酸菌青贮技术研究进展.草业科学,2014,31(2):328-333.Zhang Q,Zhang W J,Tian J P,Wang Y,Li X J,Yu Z.Advances in lactic acid bacteria silage technology research.Pratacultural Science,2014,31(2):328-333.(in Chinese)
[4]刘辉,卜登攀,吕中旺,李发弟,刘士杰,张开展,王加启.凋萎和不同添加剂对紫花苜蓿青贮品质的影响.草业学报,2015,24(5):126-133.Liu H,Bu D P,Lyu Z W,Li F D,Liu S J,Zhang K Z,Wang J Q.Effects of wilting and additives on fermentation quality of alfalfa(Medicago sativa)silage.Acta Prataculturae Sinica,2015,24(5):126-133.(in Chinese)
[5]Francisco E,Contreras-Govea,Muck R E,Broderick G,Weimer P J.Lactobacillus plantarum effects on silage fermentation and in vitro microbial yield.Animal Feed Science and Technology,2013,179(1):61-68.
[6]Weinberg Z G,Muck R E.New trends and opportunities in the development and use of inoculants for silage.FEMS Microbiology Reviews,1996,19(1):53-68.
[7]张增欣,邵涛.青贮添加剂研究进展.草业科学,2006,23(9):56-63.Zhang Z X,Shao T.Research progress in silage additive.Pratacultural Science,2006,23(9):56-63.(in Chinese)
[8]张庆,王显国,玉柱.乳酸菌添加剂和含水量对紫花苜蓿青贮的影响.中国奶牛,2015(18):37-40.Zhang Q,Wang X G,Yu Z.Effects of lactic acid bacteria inoculants and moisture content on fermentation quality of alfalfa silage.China Dairy Cattle,2015(18):37-40.(in Chinese)
[9]郭旭生,周禾,刘桂霞.苜蓿青贮过程中蛋白的分解及抑制方法.草业科学,2005,22(11):50-54.Guo X S,Zhou H,Liu G X.Proteolysis in alfalfa silage and its inhibiting method.Pratacultural Science,2005,22(11):50-54.(in Chinese)
[10]Nagel S A,Broderick G A.Effect of formic acid or formaldehyde treatment of alfalfa silage on nutrient utilization by dairy cows.Journal of Dairy Science,1992,75(1):140-154.
[11]Wu R D,Guo X S,Kazuo A.Characterization of peptides in ensiled alfalfa treated with different chemical additives.Animal Science Journal,2013,84(12):774-781.
[12]赵广永.用净碳水化合物-蛋白质体系评定反刍动物饲料营养价值.中国农业大学学报,1999,4(增刊):71-76.Zhao G Y.The Use of cornell net carbohydrate and protein system(CNCPS)for the evaluation of some Chinese feeds of ruminants.Journal of China Agricultural University,1999,4(S):71-76.(in Chinese)
[13]许庆方.影响苜蓿青贮品质的主要因素及苜蓿青贮在奶牛日粮中应用效果的研究.北京:中国农业大学博士学位论文,2005:24-26.Xu Q F.Studies of the factors of affecting alfalfa silage quality and the utilization of alfalfa silage in dairy cows diet.PhD Thesis.Beijing:China Agricultural University,2005:24-26.(in Chinese)
[14]Broderick G A,Kang J H.Automated simultaneous determination of ammonia and amino acids in ruminal fluids and in vitro media.Journal of Dairy Science,1980,63(1):64-75.
[15]张丽英.饲料分析及饲料质量检测技术.北京:中国农业大学出版社,2007:53-56,67-78.Zhang L Y.Feed Analysis and Forage Quality Detection Technology.Beijing:China Agriculture Press,2007:53-56,67-78.(in Chinese)
[16]Licitra G,Hernandez T M,Van Soest P J.Standardization of procedures for nitrogen fractionation of ruminant feeds.Animal Feed Science Technology,1996,57(4):347-358.
[17]Sniffen C J,O’Connor J D,Van Soest P J,Fox D G,Russell J B.A net carbohydrate and protein system for evaluating cattle diets:II.Carbohydrate and protein availability.Journal of Animal Science,1992,70(11):3562-3577.
[18]张金霞,乔红霞,刘雨田.水分和添加剂对紫花苜蓿青贮品质的影响.草业科学,2014,31(4):766-770.Zhang J X,Qiao H X,Liu Y T.Effects of moisture and additives on feed quality of alfalfa silage.Pratacultural Science,2014,31(4):766-770.(in Chinese)
[19]徐炜,师尚礼,张文渝,王兴中,王永福,倪兴铎,李小平,王新强,王得文,王皎.不同添加剂对低水分紫花苜蓿青贮品质的影响.草原与草坪,2014,34(1):49-54.Xu W,Shi S L,Zhang W X,Wang X Z,Wang Y F,Ni X D,Li X P,Wang X Q,Wang D W,Wang J.Effect of different additives on quality of low moisture alfalfa silage.Grassland and Turf,2014,34(1):49-54.(in Chinese)
[20]Yuan X,Guo G,Wen A.The effect of different additives on the fermentation quality,in vitro digestibility and aerobic stability of a total mixed ration silage.Animal Feed Science and Technology.2015,207:41-50.
[21]Zhang Q,Li X,Zhao M.Lactic acid bacteria strains for enhancing the fermentation quality and aerobic stability of Leymus chinensis silage.Grass and Forage Science,2015:DOI:10.1111/gfs.12190.
[22]Heron S J E,Edwards R A,Phillips P.Effect of pH on the activity of Lolium multiflorum protease.Journal of Science Food Agriculture,1989,46(3):267-277.
[23]Rooke J A,Armstrong D G.The importance of the form of nitrogen on microbial protein synthesis in the rumen of cattle receiving grass silage and intrarumen infusions of sucrose.British Journal of Nutrition,1989,61(1):113-121.
[24]Winters A L,Cockburn J E,Dhanoa M S,Merry R J.Effects of lactic acid bacteria in inoculants on changes in amino acid composition during ensilage of sterile and non-sterile ryegrass.Journal of Applied Microbiology,2000,89(3):442-451.
[25]Mckersie B D.Effect of pH on proteolysisin ensiled legume forage.Agronomy Journal,1985,77(1):81-86.
[26]Jones B A,Satter L D,Muck R E.Influence of bacterial inoculants and substrate addition to lucerne ensiled at different dry matter contents.Grass and Forage Science,1992,47(1):19-27.
[27]Broberg A,Jacobsson K,Strm K,Schnürer J.Metabolite profiles of lactic acid bacteria in grass silage.Applied and Environmental Microbiology,2007,73(17):5547-5552.
[28]Pichard G,van Soest P J.Protein solubility of ruminant feeds.In:Ithaca N Y.(ed).Proc Cornell Nutrition Conf.Department of Animal Science,Cornell University,1977:91-98.
[29]Rooke J A,Armstrong D G.The importance of the form of nitrogen on microbial protein synthesis in the rumen of cattle receiving grass silage and continuous intrarumen infusions of sucrose.British Journal of Nutrition,1989,61(1):113-121.
[30]Krishnamoorthy U C,Sniffen C J,Stem M D,van Soest P J.Evaluation of a mathematical model of rumen digestion and an in vitro simulation of rumen proteolysis to estimate the rumen-undegraded nitrogen content of feedstuffs.British Journal of Nutrition,1983,50(3):555-568.
[31]王燕,杨方,陈常栋,王海威,张微微,张永根.康奈尔净碳水化合物-蛋白质体系预测小肠可消化粗蛋白质含量.动物营养学报,2012,24(7):1274-1282.Wang Y,Yang F,Chen C D,Wang H W,Zhang W W,Zhang Y G.Predicting utilizable crude protein content in small intestine by cornell net carbohydrate and protein system.Acta Zoonutrimenta Sinica,2012,24(7):1274-1282.(in Chinese)
[32]Krishnamoorthy U C,Muscato T V,Sniffen C J,van Soest P J.Nitrogen fractions in selected feedstuffs.Journal of Dairy Science,1982,65(2):217-225.
[33]Francisco E C G,Richard E M,David R M,Paul J W.Microbial inoculant effects on silage and in vitro ruminal fermentation,and microbial biomass estimation for alfalfa,bmr corn,and corn silages.Animal Feed Science and Technology,2011,163(1):2-10.
[34]Nsereko V L,Smiley B K,Rutherford W M,Spielbauer A,Forrester K J,Hettinger G H,Harman E K,Harman B R.Influence of inoculating forage with lactic acid bacteria strains that produce ferulate esterase on ensilage and ruminal degradation of fiber.Animal Feed Science and Technology,2008,145(1-4):122-135.