CN116831224B - 一种发酵巨菌草牛饲料的制作方法 - Google Patents
一种发酵巨菌草牛饲料的制作方法 Download PDFInfo
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- A23K30/10—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder
- A23K30/15—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder using chemicals or microorganisms for ensilaging
- A23K30/18—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder using chemicals or microorganisms for ensilaging using microorganisms or enzymes
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
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- A—HUMAN NECESSITIES
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- A23K30/10—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder
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- A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
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Abstract
本发明属于巨菌草青贮技术领域，具体涉及一种发酵巨菌草牛饲料的制作方法，包括如下步骤：选择处于3个月生长期，200～300cm的巨菌草整株收割；刈割的巨菌草粉碎为2‑3厘米的细丝，晾晒至含水量为60～65％；每吨粉碎后的巨菌草，按质量比计，添加3％玉米粉、1％糖蜜、0.3％菌剂，所述菌剂由EM菌原液和蜜环菌提取液按照体积比1:1混合制备而成，然后按照常规管理方法，进行发酵，通过本发明方法发酵的巨菌草饲料适口性好，能提高采食量和饲料效率，丰富了反刍动物的饲料来源。
Description
技术领域
本发明属于巨菌草青贮技术领域，具体涉及一种发酵巨菌草牛饲料的制作方法。
背景技术
牧草是所有反刍动物饲粮的基础，因此，牧草消化率对家畜干物质采食量、瘤胃功能及生产性能具有重要意义(Raffrenato等，2017)。研究表明，提高家畜饲喂的优质青贮饲料的瘤胃发酵能力与饲喂精料一样有益，因为它可以增加挥发性脂肪酸的能量供应和微生物蛋白质合成的蛋白质供应。
巨菌草(Pennisetum giganteum)是一种优质型牧草，属多年生禾本科作物，是一种菌草，具有生长速度快、繁殖性能强、营养价值高、抗逆性强等优良特性。巨菌草在维持生态环境、畜牧业发展以及能源开发利用等方面具有巨大潜力，然而，由于高大和结实的竹状结构，使巨菌草很难被青贮，从而降低了巨菌草饲料在反刍动物中的消化和利用，目前已有部分巨菌草的青贮研究，但效果不尽如人意，故进一步的提升巨菌草的发酵品质，丰富反刍动物的饲料种类很有必要。
蜜环菌Armillaria mellea(Vahl.ex Fr.)Quel.是口蘑科Tricholomataceae的兼性寄生真菌，也是一种药食兼用真菌，又名糖蕈、栎菌等，其胞外产物中含有多种活性物质，酶如漆酶、纤维素酶、木聚糖酶；多糖类：蜜环菌多糖；萜类：原伊鲁烷倍半萜芳香酸酯类、二萜、三萜等；酚性化合物及其苷类，以及多种氨基酸合微量元素等，目前的主要研究为其活性成分的培养、提取和分离后的药用作用。
甲烷为温室气体，排放量虽不及二氧化碳，但其温室效能是等量二氧化碳的25倍，能够增强辐射及温室效应，每年畜牧业排放的甲烷占据全球排放量的16％，是人类活动中最大的甲烷排放源。其中，反刍动物甲烷排放量约占畜牧业排放量的73％，甲烷调控减排是反刍动物营养研究的一个重要领域。发酵处理可以改善饲料的营养价值，降低饲料中不好消化的纤维和一些抗营养因子、毒素等。日粮中碳水化合物类型，是影响反刍动物甲烷排放的重要因素，其可通过影响瘤胃pH或瘤胃微生物的种类、数量和产物来影响甲烷排放。因此，改善饲料的营结构，养利用营养调控技术有效地控制并减少甲烷气体产生，不仅能提高饲料利用率，节约生产成本，还能减轻甲烷对大气层的破坏，获得环境和经济的双重收益。
发明内容
本发明的一个目的是提供一种发酵巨菌草牛饲料的制作方法，以提升巨菌草草饲料的品质，丰富动物的饲料来源，本发明的另一个目的是提供一种肉牛饲喂日粮，通过营养调控，从而减少肉牛的甲烷排放量，增加饲料利用率。
为实现以上目的，本发明提供以下技术方案：
一种发酵巨菌草牛饲料的制作方法，包括如下步骤：
(1)选择处于3个月生长期，200～300cm的巨菌草整株收割；
(2)刈割的巨菌草粉碎为2-3厘米的细丝，晾晒至含水量为60～65％；
(3)每吨粉碎后的巨菌草，按质量比计，添加3％玉米粉、1％糖蜜、0.3％
菌剂，所述菌剂由EM菌原液和蜜环菌A9提取液按照体积比1:1混合制备而成，然后按照常规管理方法，进行青贮发酵。
进一步的，所述玉米粉粉碎后过50目筛。
进一步的，蜜环菌A9提取液的制备方法为：将蜜环菌A9，接种至液体发酵培养基中，在25℃、180r/min摇床中培养10d,取出发酵液，经5000r/min,离心20min后，去除沉淀，获得蜜环菌A9提取液。
进一步的，所述液体发酵培养基为：马铃薯粉3％、葡萄糖1％、蛋白胨1％、酵母膏0.1％、KH2PO40.15％、MgSO40.075％、CaCl20.01％、维生素B10.001％，余量为去离子水，pH6.0。
本发明获得的巨菌草饲料主要应用在反刍动物饲养中，进一步的，所述反刍动物为牛，进一步的，所述反刍动物为肉牛，优选的，饲喂肉牛时所述巨菌草饲料和精料的质量比为6:4。
本发明达到的技术效果为：
本发明提供的一种发酵巨菌草牛饲料的制作方法制备获得巨菌草饲料，感官评分为优等，适口性较好，显著提升了巨菌草饲料的碳水化合物含量，通过肉牛饲喂实验，使用本发明的巨菌草饲料饲喂肉牛，显著降低甲烷的日平均排放量，提高饲料利用率，节约生产成本，对环境友好。
具体实施方式
下面结合具体实施例对本发明作进一步说明，实施例中未注明具体条件者，按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者，均为可以通过市售购买获得的常规产品。
实施例1
1、青贮原料及添加剂
选择处于3个月生长期、200～300cm的巨菌草整株收割，并用一个牧草铡草机将刈割的巨菌草切成长2-3厘米的细丝，晾晒至含水量为60～65％备用，玉米粉粉碎后过50目筛备用，EM菌原液:有效微生物含量300亿CFU/ml,购于徐州森田生物科技有限公司；蜜环菌A9购于华中农业大学菌种实验中心；糖蜜:含糖量47％,购自山东金畅鑫化工科技有限公司。
蜜环菌A9提取液制备：
将蜜环菌斜面菌种接种到固体PDA(马铃薯200克，葡萄糖20克，琼脂15克，蒸馏水1000毫升自然PH，121℃灭菌20min)平板上25℃，黑暗培养活化14d。在超净台下，用打孔器取长势一致的菌丝，接种于液体PDA培养基(马铃薯200克，葡萄糖20克，蒸馏水1000毫升,自然PH，121℃灭菌20min)中，25℃、180r/min摇瓶培养7d，收集菌丝体悬液作为液体菌种，以接种量10％，接种至液体发酵培养基中，在25℃、180r/min摇床中培养10d,取出发酵液，经5000r/min,离心20min后，去除沉淀，获得蜜环菌A9提取液。
液体发酵培养基：马铃薯粉3％、葡萄糖1％、蛋白胨1％、酵母膏0.1％、KH2PO40.15％、MgSO40.075％、CaCl20.01％、维生素B10.001％，余量为去离子水，用氢氧化钠或柠檬酸调整培养液的pH至6.0，121℃灭菌20min后备用。
2、实验设计
将每吨备用巨菌草按质量比添加3％玉米粉、1％糖蜜、0.3％菌剂，具体设置如表1，利用裹包机将巨菌草裹包发酵。糖蜜和菌剂使用时，按照每吨菌草兑水10L，将糖蜜和菌剂与水混合均匀后喷洒在待发酵菌草上，对照组用等量水处理后，装入聚乙烯塑料袋，裹包，密封；每个处理组制作5个裹包，每个裹包100kg，于室温下参照DB22/T 2914-2018管理方法，存储发酵45天。
表1试验设计
| 处理 | |
| CK | 等量水 |
| T1 | 3％玉米粉、1％糖蜜 |
| T2 | 3％玉米粉、1％糖蜜、0.3％菌剂(EM菌原液) |
| T3 | 3％玉米粉、1％糖蜜、0.3％菌剂(EM菌原液：蜜环菌A9提取液＝1：1) |
3、测定指标
3.1发酵结束后，对饲料样品进行感官评定参照德国农业协会制定的青贮饲料感官评定标准的《青贮饲料质量评定标准》，观察饲料的气味、颜色、质地根据稻秸青贮饲料的气味(分)、色泽(分)和质地(分)感官指标进行感官评定，其中，优等为30～40分，良好为20～30分，一般为10～20分，差为分10以下。
3.2营养成分测定指标及方法
按照孟庆翔等《全株玉贮制作与质量评价》，张崇玉等的《饲料中性洗涤纤维(NDF)和酸性洗涤纤维(ADF)快速测定方法在生产中的应用》，进行常规营养物质的测定，包括：干物质(DM)、粗蛋白(CP)、中性洗涤纤维(NDF)、酸性洗涤纤维(ADF)、可溶性碳水化合物(WSC)等，粗蛋白测定采用凯氏定氮法。
3.3发酵品质测定
取青贮样品20.00g，加入180mL的蒸馏水，或青贮饲料与水的质量体积比为1：9。经高速粉碎机(5000r/min)粉碎匀浆1min，先后通过四层纱布和定性滤纸过滤、分装。取滤液，按照标准DB15/T 1458-2018的方法测定滤液中有机酸(乳酸、乙酸、丙酸、丁酸)含量、采用比色法测定青贮饲料中氨态氮含量的方法，pH值在浸提液制备后及时测定，有机酸和氨态氮在浸提液制备后24h内测定完毕。
4、结论
4.1巨菌草中营养成分分析
由表2可知巨菌草DM含量在常规青贮原料范围内。
表2处理前巨菌草营养成分分析(干物质基础)
4.2不同处理方式对巨菌草发酵饲料的感官评分的分析
由表3可知，不同处理下，与对照组相比，处理组的感官总平均分均显著高于对照组，T1与对照组相比，总评分显著提高，说明添加玉米粉和糖蜜改善了巨菌草青贮的适口性，T2、T3与对照组相比，再添加菌剂后，感官总评分极显著提高，表明菌群进一步的提升了巨菌草青贮饲料的风味、质地和适口性。
表3不同处理方式对巨菌草发酵饲料的感官评分的影响
| 项目 | 色泽 | 气味 | 质地 | 总评分 | 等级 |
| CK | 9.37±0.52Cc | 6.62±0.29Cc | 5.61±0.22Cc | 21.61±0.34Dd | 良 |
| T1 | 9.31±0.48Cc | 8.57±0.58Bb | 6.51±0.51Bb | 24.39±0.52Cc | 良 |
| T2 | 10.52±0.40Bb | 11.64±0.55Aa | 7.01±0.57ABb | 29.17±0.51Bb | 良 |
| T3 | 11.83±0.19Aa | 12.00±1.00Aa | 7.85±0.27Aa | 31.68±1.16Aa | 优 |
注：不同小写字母表示差异显著(P<0.05)，不同大写字母表示差异极显著(P<0.01)。
4.3不同处理方式对巨菌草发酵饲料营养成分和发酵品质的分析
由表4、表5可知，与CK相对比，在基础底料的基础上，T2组、T3组的NDF均降低，其中T3组降低更为显著，T3组的ADF显著降低，T2组、T3组CP含量显著增加，T3组的干物质损失量最少，T1组、T2组、T3组的乳酸/乙酸值均显著增加，此外T2组、T3组的丙酸值和氨态氮含量均显著减少。
表4不同处理方式对巨菌草饲料营养成分的影响(干物质基础)单位：％
| 项目 | DM | CP | NDF | ADF | WSC |
| CK | 27.78±1.48ABb | 6.27±0.10Dd | 73.66±0.24Aa | 50.01±1.20Aa | 1.95±0.05ABb |
| T1 | 28.71±0.13Bb | 7.03±0.23Cc | 72.59±0.40Ab | 49.82±0.57Aa | 1.84±0.10Bb |
| T2 | 29.08±1.21ABab | 9.44±0.29Bb | 71.04±0.10Cc | 50.72±0.72Aa | 2.20±0.40ABab |
| T3 | 30.02±0.42Aa | 10.75±0.14Aa | 66.18±0.72Dd | 44.49±0.84Bb | 2.24±0.22Aa |
注：不同小写字母表示差异显著(P<0.05)，不同大写字母表示差异极显著(P<0.01)。
表5不同处理方式对巨菌草饲料发酵品质的影响
注：不同小写字母表示差异显著(P<0.05)，不同大写字母表示差异极显著(P<0.01)。
青贮饲料的pH值与其发酵品质好坏的密切相关，添加菌剂后对青贮品质的影响主要体现在，一是快速产酸降低pH值，加快发酵速度；二是改变发酵终产物的含量，从而更加有效的保存青贮饲料的营养物质。处理组T3较高浓度的乳酸可能是由于一方面蜜环菌胞外酶发挥了作用，该酶水解木质素、纤维素等，为附着乳酸菌的发酵提供了更多可利用的底物，另一方面，玉米粉和糖蜜也提供了不同成分元素的发酵底物，在添加EM菌液后，乳酸浓度更是显著的增加，EM菌液中含有多种有益菌种，包括乳酸菌群、酵母菌群等，充足的底物浓度和多种成分的组合才能为菌种增殖提供更充分的物质保障，促进菌群生长繁殖，加快菌体蛋白的形成，从而提高青贮中CP的含量。
在仅添加EM菌液的情况下，ADF含量变化不大，NDF含量与对照相比显著下降，但在添加蜜环菌提取液后，ADF和NDF均显著下降，中性洗涤纤维和酸性洗涤纤维是饲料中不容易消化的部分，其含量的高低影响饲料的品质，添加EM菌液下，对纤维的分解有一定的作用，可一定程度提升饲料品质，但对提升饲料整体的品质，提升饲料在动物瘤胃中的转化率，需要更多的探索。
实施例2
试验于养牛场进行，试验选择体重为(300±5)kg，的西门塔尔牛20头，随机分为2组，每个组10个重复，每个重复为1头，A组饲喂T2处理组巨菌草青贮，B组饲喂T3处理组巨菌草青贮，预试验10d，试验期60d。每个处理组的试验育肥牛单独饲养，自由采食与饮水。按照规模化养牛场的饲养管理的方式进行饲养与管理，定期免疫、驱虫。
日粮配比基础日粮参照NRC(2012)推荐肉牛的营养水平进行配制参考(表6)，精料制成颗粒。饲料采用先精后粗的饲喂方式进行饲喂。
表6日粮的组成与营养水平
精料由70.8％玉米、9％玉米干酒糟及其可溶物、8％豆粕、6.2％麸皮、1％石粉、2％小苏打、3％预混料组成。预混料：维生素A 150000IU，维生素D 20000IU，维生素E3000IU，Fe 3200mg，Mn 1500mg，Zn 2000mg，Cu 650mg，I 35mg，Se 10mg，Co 10mg，Ca 130g，P 30g。
生产性能的测定，屠宰性能的测定,参照标准NY/T2660-2014,在试验期间空腹称重，记录试验开始和试验结束的体重及在试验期间的采料量，参照标准NY/T2660-2014,测定平均日增重(ADG)、平均日采食量(ADFI)和料重比(F/G)等生长性能指标。
甲烷排放的测定参考刘志豪等(泌乳牛不同胎次干奶期瘤胃发酵指标与甲烷产量的特征，畜牧兽医学报2022,53(12):4296-4305)，采用GreenFeed测定***进行每个实验组的甲烷测试。
表7巨菌草青贮饲喂对牛生长性能的影响
| 组别 | ADG/(kg/d) | ADFI/g | F/G |
| A组 | 0.79±0.10b | 6.72±0.11 | 6.54±0.12a |
| B组 | 0.92±0.17a | 6.84±0.36 | 6.15±0.11b |
注：同列数据肩标不同字母表示差异显著(P<0.05)，无字母表示差异不显著(P>0.05)；下表同。
表8巨菌草青贮对牛甲烷排放量的影响
| 组别 | 甲烷排放量(g/d) |
| A组 | 240.59±6.62a |
| B组 | 167.14±7.27b |
由表7、8可以看出，经过改良发酵后的巨菌草青贮提高了牛的平均日增重和日采食量，降低料重比，提高其生长性能，说明改良后的巨菌草青贮品质好，适口性好，特别的添加蜜环菌提取液后的青贮显著降低了甲烷排放量。
甲烷是养殖环境中主要的有害气体之一，对环境和动物的危害较大，反刍动物甲烷排放量减少主要通过减少氢的产生量或者降低产甲烷菌的数量，改善瘤胃发酵性能等，有效降低反刍动物甲烷排放量，同时也能减少反刍动物饲料能量的浪费，关于降低甲烷排放的相关研究有多种，例如通过添加植物多酚以抑制甲烷菌的代谢，本发明B组的甲烷排放量与A组相比降低了30.5％的甲烷的排放量，效果显著，木质素是纤维消化的主要障碍，因为它降低了酶对纤维素和木聚糖的可及性，并可逆或不可逆地对酶进行吸附，导致结构性多糖的水解率较低。一般情况下，木质素在瘤胃厌氧条件下无法被水解，它主要通过与细胞壁碳水化合物的相互作用降低纤维的消化率，而蜜环菌的主要寄生植物是阔叶林类乔木，其胞外酶中含有漆酶、木聚糖酶、纤维素酶等多种活性物质对木质素、纤维素有降解作用，另外，其胞外多糖、蜜环菌酸、倍半萜具有不同方向的药用作用和抗菌效果，故本研究中，甲烷排放量的降低，可能是改良发酵后的巨菌草青贮改善了牛瘤胃发酵的性能，促进丙酸的合成，从而降低了甲烷生成量，也可能是蜜环菌提取液中的多糖、漆酶等活性成分抑制了甲烷菌的数量，需要进一步的分析研究。
尽管已经示出和描述了本发明的较佳实施例，本领域的普通技术人员应当理解不脱离本发明技术原理的前提下，还可以做出若干改进和润饰，这些改进和润饰仍然视为本发明的保护范围。
Claims (2)
1.一种发酵巨菌草牛饲料的制作方法，其特征在于，包括如下步骤：
(1)选择处于3个月生长期，200～300cm的巨菌草整株收割；
(2)刈割的巨菌草粉碎为2-3厘米的细丝，晾晒至含水量为60～65％；
(3)每吨粉碎后的巨菌草，按质量比计，添加3％玉米粉、1％糖蜜、0.3％菌剂，所述菌剂由EM菌原液和蜜环菌A9提取液按照体积比1:1混合制备而成，然后按照常规管理方法，进行青贮发酵；
所述玉米粉粉碎后过50目筛；
蜜环菌A9提取液的制备方法为：将蜜环菌A9，接种至液体发酵培养基中，在25℃、180r/min摇床中培养10d,取出发酵液，经5000r/min,离心20min后，去除沉淀，获得蜜环菌A9提取液；
所述液体发酵培养基为：马铃薯粉3％、葡萄糖1％、蛋白胨1％、酵母膏0.1％、KH 2 PO4 0.15％、MgSO 4 0.075％、CaCl 2 0.01％、维生素B 1 0.001％，余量为去离子水，pH6.0；
饲喂牛时所述巨菌草饲料和精料的质量比为6:4。
2.根据权利要求1所述的制备方法，其特征在于，所述牛为肉牛。
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