CN101505611B - 用于动物饲料的酶团粒 - Google Patents
用于动物饲料的酶团粒 Download PDFInfo
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- CN101505611B CN101505611B CN2007800292648A CN200780029264A CN101505611B CN 101505611 B CN101505611 B CN 101505611B CN 2007800292648 A CN2007800292648 A CN 2007800292648A CN 200780029264 A CN200780029264 A CN 200780029264A CN 101505611 B CN101505611 B CN 101505611B
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- enzyme
- zinc
- enzyme granules
- granule
- granules
- Prior art date
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- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/20—Inorganic substances, e.g. oligoelements
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/189—Enzymes
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/20—Inorganic substances, e.g. oligoelements
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K40/00—Shaping or working-up of animal feeding-stuffs
- A23K40/20—Shaping or working-up of animal feeding-stuffs by moulding, e.g. making cakes or briquettes
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K40/00—Shaping or working-up of animal feeding-stuffs
- A23K40/25—Shaping or working-up of animal feeding-stuffs by extrusion
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K40/00—Shaping or working-up of animal feeding-stuffs
- A23K40/30—Shaping or working-up of animal feeding-stuffs by encapsulating; by coating
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Polymers & Plastics (AREA)
- Zoology (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Animal Husbandry (AREA)
- Inorganic Chemistry (AREA)
- Fodder In General (AREA)
- Enzymes And Modification Thereof (AREA)
Abstract
本发明涉及包含饲料酶和有机酸锌盐的饲料团粒。
Description
发明领域
本发明涉及酶团粒(enzyme granules),尤其是用于动物饲料的酶团粒。本发明还涉及所述团粒的制造。
发明背景
在动物饲料的相关技术领域中,一个公知的事实是饲料的制粒(pelleting)是必需的,因为将饲料制粒能增加尤其是饲料的淀粉部分的可消化性。此外,已知将动物饲料制粒能减少粉尘问题。
在制备饲料丸粒(pellets)的过程中,人们认为有必要对碎饲料(mash feed)进行蒸汽处理以杀灭可能存在的致病微生物并部分地糊化淀粉以改善饲料的物理性质,由此70-120℃左右的蒸汽处理是适当的。饲料中存在的活性化合物(如酶)在高的温度和湿度下不是非常稳定,因此,必须使用大大过量的酶,或者将不含酶的饲料组分制粒并加以蒸汽处理,然后将含酶的浆液或溶液包覆到经蒸汽处理的团粒上。然而,这种包覆过程繁琐费事(cumbersome),而且往往不适用于饲料粉碎机(feed mill)设备。
WO 92/12645中记载了一项获得改进的饲料用酶团粒的尝试。WO92/12645描述了T团粒(T-granules),其包覆有脂肪(fat)或蜡(wax)。将所述T团粒与饲料组分混合,蒸汽处理,随后制粒。通过这项发明,有可能对包含酶的颗粒进行热处理,而避免在热处理后进行费事的酶包覆。蜡包覆的T颗粒的应用是本领域的一项显著进步,因为有可能在蒸汽制粒(steampelleting)过程中维持可接受的酶活性。WO 2006034710描述了另一项改善酶团粒的制粒稳定性的尝试,其中发现用盐包层(salt coating)包覆酶团粒能改善制粒稳定性。
某些饲料粉碎机是在非常激烈的条件下运行的,这些条件对于经过改进的酶颗粒而言非常严苛,因此仍然需要改善制粒稳定性。
本发明为这个问题提供了一种解决方案,其是通过将有机酸锌盐与酶一起掺入。
利用锌和镁的无机盐使用于动物饲料的酶团粒稳定化是本领域已知的,参见WO 97/05245。已经令人惊讶地发现,使用有机酸锌盐不但能增加造粒后的酶稳定性,还能增加蒸汽制粒过程中的酶稳定性。
发明概述
本发明的一个目的是提供一种具有良好的制粒稳定性的酶团粒。本发明的另一个目的是提供一种用于获得具有良好制粒稳定性的酶团粒的简单方法。
已经令人惊讶地发现,有机酸锌盐如果与酶一起配制,对制粒过程中的酶稳定性以及本身的稳定性(per se stability)有非常积极的作用。
因此,本发明在第一个方面中提供一种适合用于动物饲料组合物的团粒,其包含饲料酶和有机酸锌盐。
本发明还提供包含本发明的酶团粒的饲料组合物。
发明详述
介绍
我们令人惊讶地发现,通过添加有机酸锌盐,有可能增加团粒中所含的酶在蒸汽制粒过程中的稳定性。
定义
术语“溶液”(solution)意指两种或更多种物质的均匀的混合物。
术语“悬液”(suspension)意指悬浮于液体中的微细颗粒。
术语“粒度”(particle size)意指团粒的质量平均直径。
术语“酶核心”(enzyme core)意指包含酶的粗团粒(raw granule),例如(thus)施加任何包层之前的团粒。
除非另有定义,本申请中使用的所有技术和科学术语的含义与本发明相关领域的普通技术人员通常理解的含义相同。说明书和权利要求书中使用的单数“一个”、“一种”和“该”包含复数的指称,除非上下文明显地另有所指。例如,术语“团粒”可能包括多个团粒。
“有机锌盐”(organic zinc salt)是指有机酸的锌盐。
团粒
当述及本发明的团粒时,它可以是单个团粒或数个团粒。
本发明的团粒可具有基质结构(matrix structure),其中组分被均匀地混合,或者可为分层的(layered)团粒,其包含核心和围绕核心的一个或多个层。
特别良好地适于蒸汽制粒,并且作为蒸汽处理的制粒饲料组合物的一部分的本发明的团粒,包括含酶区域(enzyme comprising region),其中存在有机酸锌盐以使酶稳定化。在本发明的具体实施方案中,所述酶和有机酸锌盐存在于均匀的基质中。包含酶和有机酸锌盐的所述基质可以包含其它辅助组分。
发现本发明的团粒的合适粒度为0-2000μm,更具体为50-1000μm。更进一步为75-700μm。本发明的团粒在一个具体的实施方案中可具有700μm以下的粒度。在本发明的另一个具体的实施方案中,成品团粒的粒度是100-800μm。在本发明的更具体的实施方案中,成品团粒的粒度是300-600μm。在本发明的最具体的实施方案中,成品团粒的粒度是450-550μm。在本发明的另一个具体的实施方案中,成品团粒的粒度在400μm以下。在本发明的另一个最具体的实施方案中,本发明的团粒的粒度在250μm以上,350μm以下。
在本发明的具体实施方案中,本发明的团粒的粒度为210-390μm。
含酶基质(enzyme comprising matrix)
所述团粒的含酶基质或区域可以是团粒的核心,围绕核心的层,或者,如果整个团粒的结构是均匀的话,可以是团粒本身。酶层和核心之间可以存在有层,或者酶层可以紧邻着核心。核心可以是惰性的颗粒(particle)。
含酶区域可以是包含酶和有机酸锌盐的均匀掺混物(blend)。所述掺混物可以构成核心,或者其可以构成围绕惰性核心颗粒的基质层(matrixlayer)。酶基质可以包含其它造粒剂。
由包含酶和有机酸锌盐的均匀掺混物构成,或者由其上施加有包含酶有机酸锌盐的层的惰性核心构成的核心颗粒,在一个具体实施方案中具有20-800μm的粒度。在本发明的更具体的实施方案中,核心粒度为50-500μm。在本发明的一个进而更具体的实施方案中,核心粒度为100-300μm。在本发明的最具体的实施方案中,核心粒度为150-250μm。在本发明的另一个具体的实施方案中,核心粒度为400-500μm。
所述含酶核心颗粒在一个具体实施方案中具有20-800μm的粒度。在本发明的更具体的实施方案中,核心粒度为50-500μm。在本发明的一个进而更具体的实施方案中,核心粒度为100-300μm。在本发明的一个最具体的实施方案中核心粒度为150-250μm。在本发明的另一个具体的实施方案中,核心粒度为400-500μm。
惰性核心颗粒(inert core particles):
惰性核心颗粒,诸如安慰剂颗粒(placebo particles)、载体颗粒(carrierparticles)、无活性核(inactive nuclei)、无活性颗粒(inactive particles)、丸核心用糖丸颗粒(non-pareil particles)、非活性颗粒或种子(non active particles orseeds),是不含酶或者仅含少量酶的颗粒,在其上可以用含有所述酶的包覆混合物形成层。它们可以与有机或无机材料如无机盐、糖、糖醇、小有机分子如有机酸或盐、淀粉、粉(flour)、经处理的粉、纤维素、多糖、矿物质如粘土或硅酸盐、或上述两种或更多种物质的组合一起配制。
在本发明的具体实施方案中,要包覆的颗粒是无活性颗粒。在本发明的更具体的实施方案中,核心颗粒的材料选自下组:无机盐、糖醇、小有机分子、淀粉、粉、纤维素和矿物质。
惰性颗粒可以用多种造粒(granulation)工艺来制备,其包括:结晶、沉淀、锅包衣(pan-coating)、流化床包衣(fluid bed coating)、流化床团聚(fluid bedagglomeration)、旋转雾化(rotary atomization)、挤出(extrusion)、喷射造粒(prilling)、滚圆(spheronization)、粉碎方法(size reduction methods)、转鼓造粒(drum granulation)、和/或高剪切造粒(high shear granulation)。
酶
本发明的语境中的饲料酶可以是任何酶或者不同酶的组合,其适于给予动物,意思是食用该酶将以某种方式对动物带来营养上的好处。因此,当提及“酶”时,一般应理解其包括一种饲料酶或多种饲料酶的组合。在具体的实施方案中,其不被解释为包括具有医学意义上的治疗功能的酶。
饲料酶应当为饲料/食品级,从而意味着它们不得对动物有害,并且饲料/食品级意味着它应该符合食品级酶的推荐纯度规格。在具体的实施方案中,这意味着所述酶符合联合国粮农组织/世界卫生组织食品添加剂联合专家委员会(Joint FAO/WHO Expert Committee on Food Additives,JECFA)和美国食品化学法典(Food Chemical Codex,FCC)给出的食品级酶的推荐纯度规格。
在一个具体的实施方案中,所述酶应包含少于每克30个大肠杆菌类细菌(coliform bacteria),并且包含少于50000/g的活菌数(viable count)。
本发明的团粒以干重为基础包含约0.0005至约50%的团粒酶组分。例如,在本发明的实施方案中酶的重量百分比在团粒中占至少0.0005到约25%、至少0.001到约15%、至少0.01到约10%、至少0.1到约10%、至少1.0到约10%、至少1.0到约8%、至少1.0到约5%、以及至少2.0到至少5%。每吨饲料25到400克稳定酶团粒的典型剂量将递送约0.0001至约80g的活性酶蛋白每吨饲料,酶团粒的剂量可高达每吨5000克每吨饲料。
应该理解,酶变体(例如,通过重组技术制备的)包含在术语“酶”的意义中。这样的酶变体的例子在例如EP 251,446(Genencor)、WO 91/00345(NovoNordisk)、EP 525,610(Solvay)和WO 94/02618(Gist-Brocades NV)中公开。
可以根据NC-IUBMB,1992的酶命名法手册(handbook EnzymeNomenclature)对酶加以分类,另参见互联网上的ENZYME站点http://www.expasy.ch/enzyme/。ENZYME是一个酶命名相关信息库。它主要基于国际生物化学与分子生物学联合会(IUB-MB)的推荐命名,AcademicPress,Inc.,1992,它还对每一类已被表征并已获得EC(酶学委员会)编号的酶进行了描述(Bairoch A.The ENZYME database,2000,Nucleic Acids Res28:304-305)。这种IUB-MB酶命名法基于它们的底物特异性,有时基于它们的分子机制;这样的分类并不反映这些酶的结构特征。
几年前,已提出另一种基于氨基酸序列相似性将某些糖苷水解酶,例如内切葡聚糖酶、木聚糖酶、半乳聚糖酶、甘露聚糖酶、葡聚糖酶和α-半乳糖苷酶分为家族的分类法。它们目前分为90个不同的家族:参见CAZy(ModO)因特网站点(Coutinho,P.M.&Henrissat,B.(1999)Carbohydrate-Active Enzymes服务器,在URL:http://afmb.cnrs-mrs.fr/~cazy/CAZY/index.html(相应的文章:Coutinho,P.M.&Henrissat,B.(1999)Carbohydrate-active enzymes:an integrated databaseapproach.在“Recent Advances in Carbohydrate Bioengineering”中,H.J.Gilbert,G.Davies,B.Henrissat和B.Svensson编,The Royal Society of Chemistry,Cambridge,pp.3-12;Coutinho,P.M.&Henrissat,B.(1999)The modularstructure of cellulases and other carbohydrate-active enzymes:an integrateddatabase approach.在“Genetics,Biochemistry and Ecology of CelluloseDegradation”中.,K.Ohmiya,K.Hayashi,K.Sakka,Y.Kobayashi,S.Karita和T.Kimura编,Uni Publishers Co.,Tokyo,pp.15-23)。
可掺入本发明的团粒中的酶类包括氧化还原酶(EC 1.-.-.-)、转移酶(EC2.-.-.-)、水解酶(EC 3.-.-.-)、裂合酶(EC 4.-.-.-)、异构酶(EC 5.-.-.-)和连接酶(EC6.-.-.-)。
本发明上下文中优选的氧化还原酶是过氧化物酶(EC 1.11.1)、漆酶(EC1.10.3.2)和葡糖氧化酶(EC 1.1.3.4)]。商业上可得到的氧化还原酶(EC 1.-.-.-)的实例是GluzymeTM(该酶可从Novozyme A/S获得)。更多的氧化还原酶可从其它供应商获得。优选的转移酶是以下任何亚类的转移酶:
a转移一碳基团的转移酶(EC 2.1);
b转移醛或酮残基的转移酶(EC 2.2);酰基转移酶(EC 2.3);
c糖基转移酶(EC 2.4);
d转移脂族烃基(alkyl)或芳基,不包括甲基的转移酶(EC 2.5);和
e转移含氮基团的转移酶(EC 2.6)。
本发明上下文中最优选的转移酶类型是转谷氨酰胺酶(蛋白质-谷氨酰胺γ-谷氨酰基转移酶;EC 2.3.2.13)。
合适的谷氨酰胺转移酶的其它例子在WO96/06931(Novo Nordisk A/S)中描述。
本发明上下文中优选的水解酶是:羧酸酯水解酶(EC 3.1.1.-)如脂肪酶(EC 3.1.1.3);肌醇六磷酸酶(phytase)(EC 3.1.3.-),例如3-肌醇六磷酸酶(EC3.1.3.8)和6-肌醇六磷酸酶(EC 3.1.3.26);糖苷酶(EC 3.2,它落入本文“糖酶”表示的组中),如α-淀粉酶(EC 3.2.1.1);肽酶(EC 3.4,也称作蛋白酶);以及其它羰基水解酶。商业上可得到的肌醇六磷酸酶的例子包括Bio-FeedTMPhytase(Novozymes)、RonozymeTM产品系列(DSM Nutritional Products)、NatuphosTM(BASF)、FinaseTM(AB Enzymes)以及PhyzymeTM产品系列(Danisco)。其它优选的肌醇六磷酸酶包括WO 98/28408、WO 00/43503和WO 03/066847中描述的那些。
在本上下文中,术语“糖酶”不仅用来表示能够打断糖链(例如淀粉或纤维素),特别是五元或六元环结构的糖链的酶(即糖苷酶,EC 3.2),还表示能够使糖异构化的酶,例如将六元环结构如D-葡萄糖异构化为五元环结构如D-果糖。
相关的糖酶包括下列物质(圆括号内为EC号):
α-淀粉酶(EC 3.2.1.1)、β-淀粉酶(EC 3.2.1.2)、葡聚糖1,4-α-糖苷酶(EC3.2.1.3)、内-1,4-β-葡聚糖酶(纤维素酶,EC 3.2.1.4)、内-1,3(4)-β-葡聚糖酶(EC3.2.1.6)、内-1,4-[β]-木聚糖酶(EC 3.2.1.8)、葡聚糖酶(EC 3.2.1.11)、几丁质酶(EC 3.2.1.14)、多聚半乳糖醛酸酶(EC 3.2.1.15)、溶菌酶(EC 3.2.1.17)、β-糖苷酶(EC 3.2.1.21)、α-半乳糖苷酶(EC 3.2.1.22)、β-半乳糖苷酶(EC 3.2.1.23)、淀粉-1,6-糖苷酶(EC 3.2.1.33)、木聚糖1,4-β-木糖苷酶(EC 3.2.1.37)、葡聚糖内-1,3-β-D-糖苷酶(EC 3.2.1.39)、α-糊精内-1,6-α-糖苷酶(EC3.2.1.41)、蔗糖α-糖苷酶(EC 3.2.1.48)、葡聚糖内-1,3-α-糖苷酶(EC 3.2.1.59)、葡聚糖1,4-β-糖苷酶(EC 3.2.1.74)、葡聚糖内-1,6-β-糖苷酶(EC 3.2.1.75)、半乳聚糖酶(EC3.2.1.89)、阿拉伯聚糖内-1,5-α-L-阿拉伯糖苷酶(EC 3.2.1.99)、乳糖酶(EC3.2.1.108)、壳聚糖酶(chitosanases)(EC 3.2.1.132)和木糖异构酶(EC 5.3.1.5)。
在本上下文中肌醇六磷酸酶是催化肌醇六磷酸盐(myo-inositolhexakisphosphate(肌醇六磷酸盐))的水解成(1)肌醇(myo-inositol)和/或(2)它的单、二、三、四和/或五磷酸盐以及(3)无机磷酸。
已知有三种不同类型的肌醇六磷酸酶:所谓的3-肌醇六磷酸酶(别名1-肌醇六磷酸酶;肌醇六磷酸盐3-磷酸水解酶,EC 3.1.3.8),所谓的4-肌醇六磷酸酶(别名6-肌醇六磷酸酶,基于1L编号系统而不是1D编号的名称,EC3.1.3.26),以及所谓的5-肌醇六磷酸酶(EC 3.1.3.72)。就本发明而言,将全部三种包括在肌醇六磷酸酶的定义中。
就本发明而言,肌醇六磷酸酶的活性可以且优选以FYT为单位来确定,一个FYT是在下列条件下每分钟释放1微摩尔无机正磷酸的酶量:pH 5.5;温度37℃;底物:浓度为0.0050mol/l的肌醇六磷酸钠(C6H6O24P6Na12)。合适的肌醇六磷酸酶测定法在WO 00/20569的实施例1中描述。FTU用于测定饲料和预混合物中的肌醇六磷酸酶活性。
肌醇六磷酸酶的优选实例是微生物肌醇六磷酸酶,如真菌或细菌肌醇六磷酸酶,例如源自下列的肌醇六磷酸酶:
i.子囊菌纲(Ascomycetes),如EP 684313或US 6139902公开的那些子囊菌;泡盛曲霉(Aspergillus awamori)PHYA(SWISSPROT P34753,Gene133:55-62(1993));黑曲霉(Aspergillus niger)(无花果曲霉(Aspergillus ficuum))PHYA(SWISSPROT P34752,Gene 127:87-94(1993),EP 420358);泡盛曲霉PHYB(SWISSPROT P34755,Gene 133:55-62(1993));黑曲霉PHYB(SWISSPROT P34754,Biochem.Biophys.Res.Commun.195:53-57(1993));构巢翘孢霉(Emericella nidulans)PHYB(SWISSPROT O00093,Biochim.Biophys.Acta 1353:217-223(1997));
ii.嗜热霉属(Thermomyces)或腐质霉属(Humicola),如公开于WO97/35017的细毛嗜热霉(Thermomyces lanuginosus)肌醇六磷酸酶;
iii.担子菌纲(Basidiomycetes),例如隔孢伏革菌属(Peniophora)(WO98/28408和WO 98/28409);
iv.其它真菌肌醇六磷酸酶,如公开于JP 11000164(青霉属(Penicillium)肌醇六磷酸酶)或WO98/13480(亚克红曲霉(Monascus anka)肌醇六磷酸酶)中的那些;
v.芽孢杆菌属(Bacillus),例如枯草芽孢杆菌(Bacillus subtilis)PHYC(SWISSPROT O31097,Appl.Environ.Microbiol.64:2079-2085(1998));芽孢杆菌属菌种(Bacillus sp).PHYT(SWISSPROT O66037,FEMS Microbiol.Lett.162:185-191(1998);枯草芽孢杆菌PHYT_(SWISSPROT P42094,J.Bacteriol.177:6263-6275(1995));AU 724094或WO 97/33976中公开的肌醇六磷酸酶;
vi.大肠杆菌(Escherichia coli)(US 6110719);
vii.柠檬酸杆菌属(Citrobacter),如弗氏柠檬酸杆菌(Citrobacter.freundii)(公开于WO 2006/038062、WO 2006/038128,或具有UniProt Q676V7的序列)、布氏柠檬酸杆菌(Citrobacter braakii)(公开于WO 2004/085638(GeneseqpADU50737)和WO 2006/037328)、和无丙二酸柠檬酸杆菌(Citrobacteramalonaticus)或Citrobacter gillenii(公开于WO 2006/037327);
viii.其它细菌肌醇六磷酸酶,如来自布丘氏菌属(Buttiauxella)的肌醇六磷酸酶(公开于WO 2006/043178);
ix.酵母肌醇六磷酸酶,例如来自西方许旺酵母(Schwanniomycesoccidentalis)(例如US 5830732中公开的);以及
x.具有与(i)-(ix)的任一肌醇六磷酸酶的成熟氨基酸序列有至少75%同一性的氨基酸序列的肌醇六磷酸酶;
xi.包含一个或多个氨基酸取代、缺失和/或插入的(i)-(ix)的肌醇六磷酸酶的变体;
xii.(i)-(ix)的等位变体;
xiii.(i)-(ix)的保留肌醇六磷酸酶活性的片段;或
xiv.基于(i)-(ix)设计的并具有肌醇六磷酸酶活性的合成多肽。
优选的肌醇六磷酸酶变体的例子公开于例如WO 99/49022、WO99/48380、WO 00/43503、EP 0897010、EP 0897985、WO 2003/66847,以及上文提及的WO 2006/038063、WO 2006/038128和WO 2006/43178中)。
商业上可得到的蛋白酶(肽酶)的例子包括KannaseTM、EverlaseTM、EsperaseTM、AlcalaseTM、NeutraseTM、DurazymTM、SavinaseTM、OvozymeTM、PyraseTM、Pancreatic Trypsin NOVO(PTN)、Bio-FeedTM Pro和Clear-LensTMPro(均可获自Novozymes A/S,Bagsvaerd,Denmark)。其它优选的蛋白酶包括WO 01/58275和WO 01/58276中描述的那些。
其它商业上可得到的蛋白酶包括RonozymeTM Pro、MaxataseTM、MaxacalTM、MaxapemTM、OpticleanTM、PropeaseTM、PurafectTM和Purafect OxTM(可由Genencor International Inc.、Gist-Brocades、BASF或DSM NutritionalProducts得到)。
商业上可得到的脂肪酶的例子包括LipexTM、LipoprimeTM、LipopanTM、LipolaseTM、LipolaseTM Ultra、LipozymeTM、PalataseTM、ResinaseTM、NovozymTM435和LecitaseTM(均可由Novozymes A/S得到)。
其它商业上可得到的脂肪酶包括LumafastTM(来自GenencorInternational Inc.的门多萨假单胞菌(Pseudomonas mendocina)脂肪酶);LipomaxTM(来自Gist-Brocades/Genencor Int.Inc.的类产碱假单胞菌(Ps.Pseudoalcaligenes)脂肪酶);和来自Solvay enzymes的芽孢杆菌属菌种脂肪酶。从其它供应商可获得更多的脂肪酶。
商业上可得到的糖酶的例子包括Alpha-GalTM、Bio-FeedTM Alpha、Bio-FeedTM Beta、Bio-FeedTM Plus、Bio-FeedTM Wheat、Bio-FeedTM Z、NovozymeTM 188,CarezymeTM、CelluclastTM、CellusoftTM、CelluzymeTM、CeremylTM、CitrozymTM、DenimaxTM、DezymeTM、DextrozymeTM、DuramylTM、EhergexTM、FinizymTM、FungamylTM、GamanaseTM、GlucanexTM、LactozymTM、LiquezymeTM、MaltogenaseTM、NatalaseTM、PentopanTM、PectihexTM、PromozymeTM、PulpzymeTM、NovamylTM、TermamylTM、AMGTM(Amyloglucosidase Novo)、MaltogenaseTM、SweetzymeTM和AquazymTM(均可由Novozymes A/S得到)。更多的糖酶可获得自其它供应商,如RoxazymeTM和RonozymeTM产品系列(DSM Nutritional Products),AvizymeTM、PorzymeTM和GrindazymeTM产品系列(Danisco,Finnfeeds),以及NatugrainTM(BASF)、PurastarTM和PurastarTM OxAm(Genencor)。
其它商业上可得到的酶类包括MannawayTM、PectawayTM、StainzymeTM和RenozymeTM。
在本发明的具体实施方案中,所述饲料酶选自下组:内切葡聚糖酶、内-1,3(4)-β-葡聚糖酶、蛋白酶、肌醇六磷酸酶、半乳聚糖酶、甘露聚糖酶、葡聚糖酶和α-半乳糖苷酶,参考WO 2003/062409,该文献在此通过引用并入。
特别合适的饲料酶类包括:淀粉酶,磷酸酶,如肌醇六磷酸酶,和/或酸性磷酸酶;糖酶,如淀粉分解酶和/或植物细胞壁降解酶,包括纤维素酶,如β-葡聚糖酶和/或半纤维素酶,如木聚糖酶或半乳聚糖酶;蛋白酶或肽酶,例如溶菌酶;半乳糖苷酶、果胶酶、酯酶,脂肪酶,特别是磷脂酶如哺乳动物胰磷脂酶A2和葡萄糖氧化酶。特别地,饲料酶具有中性和/或酸性最适pH。在本发明的具体实施方案中,所述饲料酶选自下组:淀粉酶、磷酸酶、肌醇六磷酸酶、纤维素酶、β-葡聚糖酶、半纤维素酶、蛋白酶、肽酶、半乳糖苷酶、果胶酶、酯酶、脂肪酶和葡萄糖氧化酶。
在本发明的具体实施方案中,所述酶选自下组:淀粉酶、蛋白酶、β-葡聚糖酶、肌醇六磷酸酶、木聚糖酶、磷脂酶和葡糖氧化酶。
有机酸锌盐
有机酸锌盐在一个具体的实施方案中是水溶性的。
当与用于动物的饲料一起使用时,重要的是团粒中所用的材料具有一定的纯度,因此在本发明的一个实施方案中,所述有机酸锌盐是食品级的。所述有机酸锌盐可以选自但不限于下组锌盐:柠檬酸盐、苹果酸盐、马来酸盐、丙二酸盐、甲硫氨酸盐(methionate)、琥珀酸盐、乳酸盐、甲酸盐、乙酸盐、丁酸盐、丙酸盐、苯甲酸盐、酒石酸盐、抗坏血酸盐、葡糖酸盐,氨基酸水合物的锌螯合物(zinc chelates of amino acids hydrates)和它们的组合。
在本发明的具体实施方案中,有机酸锌盐选自下组锌盐:柠檬酸盐、苹果酸盐、马来酸盐、丙二酸盐、甲硫氨酸盐、琥珀酸盐、乳酸盐、甲酸盐、乙酸盐,以及氨基酸水合物的锌螯合物。有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌(zinc methionate)、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌、甲硫氨酸锌(zincmethionine)和它们的组合。
有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌和它们的组合。
有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌、甲硫氨酸锌和它们的组合。
在本发明的具体实施方案中,有机阴离子选自羧酸根(carboxylates)。在一个具体实施方案中,锌盐选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌和它们的组合。
在本发明的具体实施方案中,有机酸锌盐选自下组简单有机酸(少于10个碳原子)的锌盐。在本发明的具体实施方案中,有机酸锌盐包括氨基酸的锌盐,如锌赖氨酸螯合物或锌甲硫氨酸螯合物(zinc methionine chelate)。在一个具体的实施方案中,有机酸锌盐不包括氨基酸。
在本发明的更具体的实施方案中,有机酸锌盐选自下组:柠檬酸锌、乙酸锌、甲硫氨酸锌、锌甲硫氨酸螯合物(zinc methionine chelate),和它们的组合。
在本发明的具体实施方案中,有机酸锌盐以酶核心为基础的添加量为0.01-15%w/w。在本发明的更具体的实施方案中,有机酸锌盐以酶核心为基础的添加量为0.05-10%w/w。在本发明的还更具体的实施方案中,有机酸锌盐的添加量为0.1-7%w/w。在本发明的最具体的实施方案中,有机酸锌盐的添加量为酶核心的0.5-3.5%w/w。所述量是对无水锌盐计算的。“无水”是指盐中不存在结晶水。
在本发明的具体实施方案中,有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌和它们的组合,且锌化合物以酶核心为基础的添加量为0.01-15%w/w。
有机酸锌盐可以以溶液、分散体、乳液的形式或以固体形式添加。在本发明的具体实施方案中,有机酸锌盐以溶液形式添加。在一个具体实施方案中,所述溶液是水溶液。在一个具体实施方案中,所述有机酸锌盐作为液体添加。在另一个实施方案中,所述有机酸锌盐作为粉末添加。
在本发明的具体实施方案中,有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌和它们的组合,且锌化合物以酶核心为基础的添加量为0.01-15%w/w,且有机酸锌盐作为粉末添加。
在一个具体实施方案中,所述有机酸锌盐与酶一起作为混合物存在于团粒中。在本发明的另一个具体实施方案中,所述有机酸锌盐存在于围绕酶层的层中。
在本发明的具体实施方案中,有机酸锌盐可以选自下组:柠檬酸锌、苹果酸锌、马来酸锌、丙二酸锌、甲硫氨酸锌、琥珀酸锌、乳酸锌、甲酸锌、乙酸锌、丁酸锌、丙酸锌、苯甲酸锌、酒石酸锌、抗坏血酸锌、葡糖酸锌、甲硫氨酸锌、赖氨酸锌和它们的组合,且锌化合物以酶核心为基础的添加量为0.01-15%w/w,且所述有机酸锌盐与酶一起作为混合物存在于团粒中。
附加的造粒剂
所述颗粒可包含附加的材料,如粘合剂、填充剂、纤维材料、稳定剂、增溶剂、悬浮剂、粘度调节剂、轻球体(light spheres)、增塑剂、盐、润滑剂和香料。
本发明的粘合剂可以是合成聚合物、包括脂肪的蜡、发酵液、糖、盐或多肽。
合成聚合物
合成聚合物意指通过合成手段聚合形成骨架的聚合物。
本发明的合适的合成聚合物具体地包括聚乙烯吡咯烷酮(PVP)、聚乙烯醇(PVA)、聚乙酸乙烯酯、聚丙烯酸酯、聚甲基丙烯酸酯、聚丙烯酰胺、聚磺酸酯、聚羧酸酯,以及它们的共聚物,尤其是水溶性聚合物或共聚物。
在本发明的具体实施方案中,合成聚合物是乙烯基聚合物。
蜡
在本发明上下文中“蜡”应当理解为具有25-150℃、特别是30-100℃、更特别是35-85℃、最特别是40-75℃的熔点的聚合材料。在室温25℃,蜡优选以固态存在。优选的是,所述下限在蜡开始熔解的温度与团粒或包含团粒的组合物的通常贮藏温度20-30℃之间设定合理的距离。
对于某些团粒,所述蜡的优选特征在于其应当是水溶性或水可分散的,所述蜡应当分解和/或溶解,使掺入颗粒中的活性成分的快速释放和溶解到水溶液中。水溶性蜡的例子是聚乙二醇(PEG’s)。在水溶性蜡中,在水溶液中可分散的为甘油三酯和油类。对于某些团粒,优选所述蜡是不溶性的。
在本发明的具体实施方案中,所述蜡组合物是亲水组合物。在一个具体的实施方案中,包含在蜡组合物中的成分的至少25%w/w,优选至少50%w/w、优选至少75%w/w、优选至少85%w/w、优选至少95%w/w、优选至少99%w/w是可溶于水的。
在另一个实施方案中,所述蜡组合物是亲水的,且可分散于水溶液中。
在一个具体的实施方案中,所述蜡组合物包含少于75%w/w,优选少于50%w/w、优选少于25%w/w、优选少于15%w/w、优选少于5%w/w、优选少于1%w/w的疏水成分。
在一个具体的实施方案中,所述蜡组合物包含少于75%w/w,优选少于50%w/w、优选少于25%w/w、优选少于15%w/w、优选少于5%w/w、优选少于1%w/w的水不溶性成分。
合适的蜡是具有一种或多种上述性质的有机化合物或有机化合物的盐。
本发明的蜡组合物可包含任何化学合成的蜡。其也同样可包含分离自天然来源的蜡或其衍生物。因此,本发明的蜡组合物可包含选自下列非限制性的蜡的列表的蜡。
-聚乙二醇,PEG。可从市场上获得具有不同分子量的不同PEG蜡,其中具有低分子量的PEG还具有低熔点。合适的PEG的例子是PEG 1500、PEG 2000、PEG 3000、PEG 4000、PEG 6000、PEG 8000、PEG 9000等,例如来自BASF(Pluriol E系列)或来自Clariant或Ineos。还可以使用聚乙二醇的衍生物。
-聚丙烯(例如来自BASF的聚丙二醇Pluriol P系列)或聚乙烯或它们的混合物。还可以使用聚丙烯和聚乙烯的衍生物。
-环氧乙烷、环氧丙烷的聚合物,或它们的共聚物是有用的,如用于嵌段聚合物(block polymers)中,所述嵌段聚合物例如来自BASF的PluronicPE 6800。乙氧基化脂肪醇的衍生物。
-分离自天然来源的蜡,如巴西棕榈蜡(Carnauba wax)(熔点为80-88℃)、小烛树蜡(Candelilla wax)(熔点为68-70℃)和蜂蜡。其它天然蜡或其衍生物是来自动物或植物的蜡,例如海洋来源的。氢化的植物油或动物油脂。这样的蜡的例子是氢化牛脂、氢化棕榈油、氢化棉籽和/或氢化大豆油,其中在本申请中使用的术语“氢化”应理解为不饱和的碳水化合物链的饱和,例如在甘油三酯中,其中碳=碳双键转化为碳-碳单键。氢化棕榈油是商业上可得到的,例如来自Hobum Oele und Fette GmbH-Germany或Deutche Cargill GmbH-Germany。
-脂肪酸醇,如来自Condea Chemie GMBH-Germany的线性长链脂肪酸醇NAFOL 1822(C18,20,22),具有55-60℃的熔点。脂肪酸醇的衍生物。
-单酸甘油酯和/或甘油二酯,如硬脂酸甘油酯,其中硬脂酸是十八酸和十六酸的混合物,是有用的蜡。其例子是来自Danisco Ingredients,Denmark的Dimodan PM。
-脂肪酸,例如氢化的线性长链脂肪酸和脂肪酸的衍生物。
-石蜡(Paraffines),即固体烃。
-微晶蜡。
在进一步的实施方案中,在本发明中有用的蜡可见于C.M.McTaggart等,Int.J.Pharm.19,139(1984)或Flanders等,Drug Dev.Ind.Pharm.13,1001(1987)中,两者均通过引用并入本文。
在本发明的具体实施方案中,本发明的蜡是两种或更多种不同的蜡的混合物。
在本发明的具体实施方案中,所述蜡选自由PEG、脂肪酸、脂肪酸醇和甘油酯组成的组。
在本发明另一个具体的实施方案中,所述蜡从合成蜡中选择。在本发明的更具体的实施方案中,本发明的蜡是PEG。在本发明的最具体的实施方案中,所述蜡选自下组:牛脂、PEG和棕榈油。
发酵液(fermentation broth)
根据本发明的发酵液包含微生物细胞和/或其细胞碎片(生物质)。
在优选的实施方案中,所述发酵液包含至少10%、更优选至少50%、还更优选至少75%,最优选至少90%或至少95%的来源于发酵的生物质。在另一个优选的实施方案中,所述发酵液含有0-31%w/w的干物质、优选0-20%w/w、更优选0-15%w/w,如10-15%w/w的干物质,0%的干物质被排除在所述范围之外。所述生物质可占到干物质的多至90%w/w、优选多至75%w/w、更优选多至50%w/w,同时酶可占到干物质的多至50%w/w、优选多至25%w/w、更优选多至10%w/w。
多糖
本发明的多糖可以是未修饰的天然存在的多糖或经修饰的天然存在的多糖。
合适的多糖包括纤维素、果胶、糊精和淀粉。所述淀粉可以是可溶或不溶于水的。
在本发明的具体实施方案中,所述多糖是淀粉。在本发明的具体实施方案中,所述多糖是不溶性淀粉。
在本发明上下文中,来自多种植物来源的天然存在的淀粉是合适的(作为淀粉本身,或作为改性淀粉的起点),有关的淀粉包括来自下述的淀粉:稻、玉米(corn)、小麦、马铃薯、燕麦、木薯(cassava)、西谷椰子(sago-palm)、木薯(yuca)、大麦、甘薯(sweet potato)、高粱、薯蓣(yams)、黑麦、粟/黍(millet)、荞麦(buckwheat)、竹芋(arrowroot)、芋头(taro)、芋类(tannia),并可例如以粉(flour)的形式存在。
在本发明的上下文中,木薯淀粉是优选的淀粉之一;在此处要提及的是木薯和木薯淀粉有多种同义词,包括木薯(tapioca)、树薯(manioc)、树薯(mandioca)和木薯(manihot)。
如在本发明上下文中所使用的,术语“改性淀粉”表示天然存在的淀粉,其已经历某种至少部分的化学修饰、酶修饰和/或物理或物理化学修饰,并且相对于“亲本”淀粉通常显示改变的特性。
在本发明的具体实施方案中,所述团粒包含多糖。
盐
核心可包含附加的盐。所述盐可以是无机盐,例如硫酸盐、亚硫酸盐、磷酸盐、膦酸盐、硝酸盐、氯化物或碳酸盐、或简单有机酸(小于10个碳原子,例如6个或更少的碳原子)的盐,如柠檬酸盐、丙二酸盐或乙酸盐。在这些盐中,阳离子的例子是碱或碱土金属离子,尽管铵离子或第一过渡系(transition series)的金属离子,如钠、钾、镁、钙、锌或铝。阴离子的例子包括氯、碘、硫酸根、亚硫酸根、亚硫酸氢根、硫代硫酸根、磷酸根、磷酸二氢根、磷酸氢根、次磷酸根、焦磷酸二氢根、碳酸根、碳酸氢根、硅酸根、柠檬酸根、苹果酸根、马来酸根、丙二酸根、琥珀酸根、乳酸根、甲酸根、乙酸根、丁酸根、丙酸根、苯甲酸根、酒石酸根、抗坏血酸根或葡糖酸根。特别是,可以使用硫酸、亚硫酸、磷酸、膦酸、硝酸、氯或碳酸的碱或碱土金属盐,或简单无机酸的盐例如柠檬酸盐、丙二酸盐或乙酸盐。具体的例子包括NaH2PO4、Na2HPO4、Na3PO4、(NH4)H2PO4、K2HPO4、KH2PO4、Na2SO4、K2SO4、KHSO4、ZnSO4、MgSO4、CuSO4、Mg(NO3)2、(NH4)2SO4、硼酸钠、乙酸镁和柠檬酸钠。
所述盐还可以是水合盐,即带有结晶的结合水的结晶盐水合物,如WO99/32595中所描述的。水合盐的例子包括七水合硫酸镁(MgSO4(7H2O))、七水合硫酸锌(ZnSO4(7H2O))、七水合磷酸氢二钠(Na2HPO4(7H2O))、六水合硝酸镁(Mg(NO3)2(6H2O))、十水合硼酸钠、二水合柠檬酸钠和四水合乙酸镁。
在本发明的具体实施方案中,所述粘合剂是多肽。所述多肽可选自明胶、胶原、酪蛋白、壳聚糖(chitosan)、聚天冬氨酸和聚谷氨酸。在另一个具体实施方案中,所述粘合剂是纤维素衍生物,如羟丙基纤维素、甲基纤维素或CMC。合适的粘合剂是碳水化合物粘合剂如糊精,例如Glucidex 21D或Avedex W80。
填充剂(fillers)
合适的填充剂是水溶性和/或不溶性无机盐,如磨细的碱金属硫酸盐、碱金属碳酸盐和/或碱金属氯化物,粘土如高岭土(如SPESWHITETM,英语称China Clay)、膨润土、滑石、沸石、白垩、碳酸钙和/或硅酸盐。
代表性的填充剂是硫酸二钠和木质素磺酸钙(calcium-lignosulphonate)。其它的填充剂是硅石、石膏、高岭土、滑石、硅酸铝镁和纤维素纤维。
纤维材料(fibre materials)
纯或不纯的纤维素以纤维状形式存在,如锯屑、纯纤维状纤维素、棉花或其它类型的纯或不纯的纤维状纤维素。同样地,可以使用以纤维状纤维素为基础的助滤剂。市场上有几种品牌的纤维状形式的纤维素,例如CEPOTM和ARBOCELLTM。纤维状纤维素助滤剂的有关例子是ARBOCELLBFC 200TM和ARBOCELL BC 200TM。也可使用合成纤维,如EP 304331B1中所述。
稳定剂
稳定剂或保护剂如在造粒领域所通常使用的。稳定剂或保护剂可分为几类:碱性或中性材料、还原剂、抗氧化剂和/或第一过渡系金属离子的盐。这些的每一种都可与其它相同或不同类别的保护剂一起使用。碱性保护剂的例子是碱金属硅酸盐、碳酸盐或碳酸氢盐。还原保护剂的例子是亚硫酸盐、硫代亚硫酸盐、硫代硫酸盐或MnSO4,而抗氧化剂的例子是甲硫氨酸、丁基化羟基甲苯(BHT)或丁基化羟基苯甲醚(butylated hydroxyanisol)(BHA)。特别是,稳定剂可以是硫代硫酸盐,例如硫代硫酸钠或甲硫氨酸。有用的稳定剂的其它例子是明胶、尿素、山梨糖醇、甘油、酪蛋白、聚乙烯吡咯烷酮(PVP)、羟丙基甲基纤维素(HPMC)、羧甲基纤维素(CMC)、羟乙基纤维素(HEC)、脱脂乳粉和/或食用油,如豆油或油菜油(canola oil)。在饲料团粒中具体的稳定剂是乳酸源或淀粉。在本发明的具体实施方案中,所述团粒包含根据专利申请号EP 1,117,771的乳酸源,其在此并入作为参考。优选的乳酸源是玉米浆(corn steep liquor)。本领域还公知酶底物如淀粉、脂质、蛋白质等可作为酶的稳定剂。
增溶剂
本领域技术人员已知,许多试剂,通过不同方法,能起到增加制剂的溶解性的作用,本领域已知的代表性的试剂可见于国家药典(NationalPharmacopeia)。
轻球体:
轻球体是具有低的真密度(true density)的小颗粒。通常,它们是内含空气或气体的中空球状颗粒。这样的材料通常通过使固体材料发泡(expanding)来制备。这些轻球体本质上可以是无机的或有机的。多糖如淀粉或其衍生物是优选的。是由纤维素(造纸废料)制成的非中空轻量材料的例子,它可由GranTek Inc得到。这些材料可单独或与不同的轻材料作为混合物包括在本发明的团粒中。
悬浮剂:
可以掺入悬浮剂、介质(mediator)和/或溶剂。
粘度调节剂:
可以存在粘度调节剂。
增塑剂:
本发明的增塑剂包括,例如:多元醇,如糖、糖醇、甘油、甘油三羟甲基丙烷、新戊二醇、三乙醇胺、单-、二-和三甘醇或分子量小于1000的聚乙二醇(PEG);尿素和水。
润滑剂
如本发明上下文中所使用的,术语“润滑剂”指任何减少表面磨擦、润滑颗粒表面、减少静电形成倾向和/或减少团粒脆性的试剂。润滑剂可作为抗附聚剂和润湿剂。合适的润滑剂的例子是低级聚乙二醇(PEG)和矿物油。具体地,润滑剂是矿物油或非离子型表面活性剂,更具体地,润滑剂不与其它材料混溶。
包层(coatings)
本发明的团粒可包含一个、二个或更多个附加包层。可将包层施用于所述团粒以提供额外的特征或性质。因此,例如附加包层可实现以下一种或多种效果:
(i)减少团粒的粉尘形成倾向;
(ii)保护团粒中的酶对抗环境中的不利化合物(hostile compounds);
(iii)一旦将颗粒引入液体介质(如酸性介质)中,就以期望的速率溶解;
(iv)提供更好的团粒物理强度。
本发明的包层通常作为围绕核心的一个或多个层来施加。实施方案包括一个、两个、三个或四个保护性包层。合适的包层材料是聚合物、碳水化合物、蛋白质、脂质、油脂、脂肪酸、无机盐、胶质(gums)和它们的混合物。
包层包括湿分隔离包层(moisture barrier coatings)和湿分水化包层(moisture hydrating coatings)。湿分隔离包层通过排除湿分来发挥功能,例如通过形成壳层(shell layer),所述壳层通常不吸收湿分并阻止湿分移动到团粒中或延缓湿分移动到团粒中的速率。团粒上的湿分水化包层吸收或结合游离水或水化水形式的湿分,从而起到阻止或延缓外部湿分向团粒内转运的程度或速率的作用。湿分水化包层通常占团粒的至少约35%w/w。包层中的湿分水化材料使酶热绝缘,并且将吸收一定量的湿分并将其保持在水化材料中而不会容许它透到团粒的含酶部分中。对于在蒸汽处理前含水量不明显的稳定团粒上的湿分水化包层而言,这样的包层可占到团粒的约25%w/w。湿分隔离包层通常包含疏水性材料,如疏水性聚合物,例如PVA、HPMC、酸稀化的羟丙基淀粉(acid-thinned hydroxypropyl starches)和氧化淀粉(oxidized starch);蛋白质,例如乳清和乳清蛋白浓缩物;脂质,如卵磷脂;油脂,脂肪酸,胶乳和胶质,例如阿拉伯胶。某些湿分隔离包层,如PVA和阿拉伯胶,不易被氧化,特别适用于将本发明的团粒保存于未制粒或未压片的混合物中(例如在含氯化胆碱的预混物中)时提供化学稳定性。湿分水化包层材料通常是亲水性材料,如碳水化合物和无机盐,包括水合盐。湿分水化材料的例子有:硫酸镁;硫酸钠;麦芽糖糊精;硫酸铵;糖,例如蔗糖;和天然玉米淀粉。用于保护性包层的聚合物有聚乙烯醇(PVA),聚乙二醇,聚乙烯吡咯烷酮,聚丙烯酸酯,聚氧化乙烯(PEO),聚乳酸,聚氯乙烯,聚乙酸乙烯酯,聚乙烯吡咯烷酮(PVP),纤维素醚,藻酸盐(或酯)(alginates),明胶(gelatin),改性淀粉(modified starches)和它们的取代衍生物、水解物和共聚物,如酸稀化的羟丙基淀粉,如Pure coteTM羟丙基甲基纤维素(HPMC)、甲基纤维素(MC)、羧甲基纤维素(CMC)和乙基纤维素。最优选的用于保护性包层的聚合物是PVA、改性PVA,如美国专利6,872,696描述的,和改性纤维素,如甲基纤维素和羟丙基甲基纤维素,如PCT公开号WO 99/51210中描述的,上述两篇文献通过引用并入本文。用于保护性包层的碳水化合物有麦芽糖糊精羟甲基纤维素;由玉米、高粱、竹芋、稻、小麦、黑麦、大麦、燕麦、马铃薯、薯蓣、木薯(tapioca)、木薯(cassava)、西米制成的改性或天然淀粉;以及糖类,包括蔗糖、玉米糖浆固形物、糖蜜、葡萄糖、果糖和乳糖。用于保护性包层的蛋白质有乳清粉、乳清蛋白浓缩物、乳清蛋白分离物、酪蛋白酸盐(caseinates)、大豆蛋白浓缩物和分离物、玉米醇溶蛋白(zein)、白蛋白和明胶。
可用于保护性包层的简单、复合和衍生的(simple,compound and derived)脂质有蜡(例如植物蜡、矿物蜡和合成蜡,如巴西棕榈蜡、小烛树蜡、蜂蜡、耵聍(cerumen)、carnuba、虫胶(shellac)、石蜡和微晶蜡);卵磷脂(例如单-和二甘油酯);脂肪酸(例如硬脂酸、棕榈酸、亚油酸、油酸、丁酸和花生四烯酸脂肪酸)和它们的钠、钾、钙和锌盐);以及油脂(例如氢化或部分氢化的油脂,如大豆油、玉米油、棉籽油、动物脂油、油菜油和亚麻子油)。用于保护性包层的优选脂质是卵磷脂。
用于保护性包层的无机盐包括钠、铵、钾、钙、镁和锌的硫酸盐、柠檬酸盐、氯化物盐、碳酸盐、亚硫酸盐、磷酸盐、膦酸盐和碳酸氢盐。优选的盐是镁、钠和铵的硫酸盐。
可用于保护性包层的胶质包括阿拉伯胶、瓜尔胶、琼脂、黄蓍胶、karya胶、槐豆胶(locust bean gum)、鹿角菜胶(carageenan)、黄原胶(xanthan gum)、和藻酸盐(alginates)。
本发明的保护性包层还可以包括增塑剂、润滑剂、色素和粉末,如滑石、膨润土、高岭土、玉米淀粉、硅酸镁、碳酸钙和壳聚糖。
本发明的某些实施方案通常具有单层湿分水化材料,其约为团粒的至少55%w/w。由于湿分水化包层摄取和封锁水分的能力有限,所以施加的单层包层的水平相对较高。或者,可在两层中施加湿分水化材料。本发明的其它实施方案具有利用湿分水化材料和湿分隔离材料两者的保护性包层。在这些实施方案中,湿分水化材料的量可以较低,为团粒的至少约25%w/w,而湿分隔离材料为团粒的约2%到25%w/w。同时使用湿分水化材料和湿分隔离材料将二者的保护机制组合,并通常降低了成本,尤其是湿分隔离材料的成本。
湿分隔离材料,尤其是成膜材料,可能受到机械损伤,如果单独用这些材料作为薄包层,可导致丧失对酶的保护。上述组合使两种材料的用量都可以比它们单独使用时所需用量少。考虑到湿分水化材料的存在,所述组合容许对湿分隔离层的一定损伤。
可以应用任何具有期望性质的常规包层,常规包层材料和包层方法的例子在特别是下列文献中有描述:US 4,106,991、EP 170360、EP 304332、EP 304331、EP 458849、EP 458845、WO 97/39116、WO 92/12645、WO89/08695、WO 89/08694、WO 87/07292、WO 91/06638、WO 92/13030、WO93/07260、WO 93/07263、WO 96/38527、WO 96/16151、WO 97/23606、US5,324,649、US 4,689,297、EP 206417、EP 193829、DE 4344215、DE 4322229A、DD 263790、JP 61162185A、JP 58179492或PCT/DK/01/00628。
在本发明的具体实施方案中,附加包层是根据US 4,106,991或EP0,569,468(在此通过引用并入)的蜡包层。有关合适的蜡参见上文“蜡”一节。在本发明的具体实施方案中,附加包层可包含PVA、PEG、牛脂和/或棕榈油。任选地,可以用包层混合物包覆团粒。
在本发明的具体实施方案中,用盐包层包覆团粒。
所述盐可以从“盐”一节选择。
附加包层材料
所述包层可包含附加包层材料,如在上述“附加的造粒剂”的节中提及的粘合剂、填充剂、纤维材料、酶稳定剂、盐、增溶剂、悬浮剂、粘度调节剂、轻球体、增塑剂、盐、润滑剂和香料。另外的包层成分可以是色素。
色素
合适的色素包括,但不限于,精细磨碎的增白剂(finely dividedwhiteners),如二氧化钛或高岭土,有色色素、水溶性着色剂、以及一种或多种色素和水溶性着色剂的组合。
酶核心的制备
酶核心包含酶和有机酸锌盐。
用于制备酶核心的方法可见于Handbook of Powder Technology;Particlesize enlargement,作者为C.E.Capes;Volume 1;1980;Elsevier。制备方法包括已知的饲料和团粒配制技术,即:
a)喷雾干燥产物,其中在喷雾干燥塔中雾化液体含酶溶液以形成小液滴,在它们在沿干燥塔下降的过程中干燥形成含有酶的颗粒状物质。该方法可产生非常小的颗粒(Michael S.Showell(编);Powdered detergents;Surfactant Science Series;1998;vol.71;page 140-142;Marcel Dekker)。
b)成层(layered)产物,其中酶作为层涂覆在预成型的惰性核心颗粒周围,其中将含有酶的溶液雾化,通常在其中预成型的核心颗粒被流化的流化床装置中,含有酶的溶液附着于核心颗粒并干燥,在核心颗粒的表面上留下干酶的层。如果能找到期望尺寸的有用的核心颗粒,则该方法能获得具有期望尺寸的颗粒。这种类型的产品描述于例如WO 97/23606中。
c)吸收的(absorbed)核心颗粒,其中不是将酶涂覆为核心周围的层,而是使酶被吸收到核心的表面上和/或表面中。这样的方法描述于WO97/39116中。
d)挤出或制粒的产品,其中含有酶的糊剂被压成丸粒(pellets)或在压力下通过小的开口挤出并切割为颗粒,随后颗粒被干燥。这样的颗粒通常具有相当大的尺寸,因为制有挤出开口的材料(通常是具有钻孔的板)对整个挤出口的容许压降设定了限制。此外,当使用小的开口时,非常高的挤出压力可增加酶糊剂的产热,这对酶是有害的(Michael S.Showell(编辑);Powdered detergents;Surfactant Science Series;1998;vol.71;page 140-142;Marcel Dekker)。
e)喷射造粒产物,其中将酶粉末悬浮于熔化的蜡中,并将悬浮液喷射(例如通过转盘喷雾器)到冷却室中,在此冷却室中液滴快速地固化(Michael S.Showell(编辑);Powdered detergents;Surfactant Science Series;1998;vol.71;page 140-142;Marcel Dekker)。所获产物中酶均匀分布于惰性材料中而不是集中在其表面上。同样地,US 4,016,040和US 4,713,245是涉及此技术的文献。
f)混合器造粒产物,其中将酶液体添加于常规造粒组分的干燥粉末组合物。将液体和粉末以合适的比例混合,液体的水分为干燥粉末所吸收,干燥粉末的组分开始粘着并团聚,构成颗粒,形成包含酶的团粒(granulates)。这样的方法描述于US 4,106,991(NOVO NORDISK)和有关的文献EP170360B1(NOVO NORDISK)、EP 304332B1(NOVO NORDISK)、EP 304331(NOVO NORDISK)、WO 90/09440(NOVO NORDISK)和WO 90/09428(NOVO NORDISK)中。在该方法的一种具体产品中,其中不同的高剪切混合器可用作造粒机,将由酶、填充剂和粘合剂等组成的团粒与纤维素纤维混合以增强颗粒得到了所谓的T-团粒(T-granulate)。增强的团粒更加坚固,释放较少的酶粉尘。
g)粉碎(size reduction),其中通过磨碎或压碎含有酶的较大的团粒、丸粒、片体(tablet)、坯块(briquettes)等产生核心。通过将磨碎或压碎的产物过筛来获得所需要的核心颗粒级分。尺寸过大和尺寸过小的团粒可重复利用。粉碎描述于(Martin Rhodes(编辑);Principles of Powder Technology;1990;Chapter 10;John Wiley&Sons)中。
h)流化床造粒。流化床造粒包括将颗粒悬浮于空气流中并经喷嘴喷射液体到流态化的颗粒上。被喷射的液滴击中的颗粒湿润并发粘。粘性的颗粒与其它颗粒碰撞并粘附于其上而形成团粒。
i)可对核心进行干燥,如在流化床干燥器中干燥。本领域技术人员可以使用饲料或酶工业中其它已知的用于干燥团粒的方法。所述干燥优选在25-90℃的产品温度进行。对于某些酶,重要的是包含酶的核心在用盐涂覆之前含有少量的水。如果在除去过量水之前用盐涂覆水敏感性酶,则水分会被截留在核心中并可能给酶的活性造成不利的影响。在干燥之后,所述核心优选含有0.1-10%w/w的水。
团粒的包层
可合适地使用本领域已知的常用包层和方法,如描述于丹麦PA 200200473、WO 89/08694、WO 89/08695、270608B1和/或WO 00/01793中的包层。常用包层物质的其它例子可见于US 4,106,991、EP 170360、EP 304332、EP 304331、EP 458849、EP 458845、WO 97/39116、WO 92/12645A、WO89/08695、WO 89/08694、WO 87/07292、WO 91/06638、WO 92/13030、WO93/07260、WO 93/07263、WO 96/38527、WO 96/16151、WO 97/23606、WO01/25412、WO 02/20746、WO 02/28369、US 5879920、US 5,324,649、US4,689,297、US 6,348,442、EP 206417、EP 193829、DE 4344215、DE 4322229A、DE 263790、JP 61162185A和/或JP 58179492。
可通过与上述“酶核心的制备”一节中相同的方法制备所述包层。
所获得的团粒可进行滚圆(如球状化),如在MarumeriserTM中,或压实。
所述团粒可加以干燥,例如在流化床干燥器中。本领域技术人员可以使用饲料或酶工业中用于干燥团粒的其它已知的方法。所述干燥优选在25-90℃的产品温度进行。
饲料丸粒的制造
在饲料丸粒的制造中,优选在制粒之前引入蒸汽处理,该工序称作调理(conditioning)。在接下来的制粒步骤中,施力使饲料通过模具,将所得的条状物(strand)切成不同长度的合适丸粒。在所述调理步骤中,加工温度可上升到60-100℃。
通过将包含饲料酶的团粒与期望的饲料组分混合来制备饲料混合物(碎饲料(mash feed))。将混合物导入调理器(conditioner),例如带有蒸汽喷射的阶式混合器(cascade mixer)。在调理器中通过喷射蒸汽将饲料加热到指定的温度60-100℃,例如60℃、70℃、80℃、90℃或100℃,所述温度在调理器的出口测量。停留时间可从数秒变化至数分钟,甚至数小时。如5秒、10秒、15秒、30秒、1分钟、2分钟、5分钟、10分钟、15分钟、30分钟和1小时。在本发明的具体实施方案中,温度是100℃,停留时间是60秒。
在本发明的具体实施方案中,蒸汽处理过程中的加工温度是至少60℃。在本发明的更具体的实施方案中,蒸汽处理过程中的加工温度是至少70℃。在本发明的还更具体的实施方案中,蒸汽处理过程中的加工温度是至少80℃。在本发明的最具体的实施方案中,蒸汽处理过程中的加工温度是至少90℃。
将饲料从调理器引入压力机例如Simon Heesen压力机,并挤压成具有不同长度例如15mm的丸粒。在挤压之后,将丸粒置于空气冷却器中并冷却指定的时间,例如15分钟。
本发明一个具体的实施方案是一种制造饲料组合物的方法,包括步骤:
i.将饲料组分与本发明的团粒混合,
ii.蒸汽处理所述组合物(i),和
iii.将所述组合物(ii)制粒。
在本发明的具体实施方案中,对本发明的团粒进行蒸汽处理和/或制粒。
动物饲料
本发明的团粒适用于动物饲料组合物。将团粒与饲料组分混合。这样的饲料组合物称为碎饲料(mash feed)。团粒的特性使其可以用作完全适合作为动物饲料的组合物的组分,所述组合物是被蒸汽处理且随后被制粒的。本发明的一个具体实施方案是一种经过蒸汽处理的制粒的包含团粒的饲料组合物,所述团粒包含饲料酶和有机酸锌盐。
术语“动物”包括所有的动物。动物的例子是非反刍和反刍动物,如牛、绵羊和马。在一个具体的实施方案中,所述动物是非反刍动物。非反刍动物包括单胃动物,如猪(pigs)或猪(swines)(包括但不限于小猪、饲育猪(growingpigs)和母猪(sows));家禽例如火鸡和家鸡(包括但不限于肉用仔鸡(broilerchickens)、蛋鸡(layers));小牛(young calves),和鱼类(包括但不限于鲑鱼)。
本发明的饲料可包含植物蛋白质。本申请中使用的术语“植物蛋白质”指如下所述的任何化合物、组合物、制备物或混合物:其包括至少一种来自或源于植物的蛋白质,包括修饰的蛋白质和蛋白质衍生物。在具体的实施方案中,所述植物蛋白质的蛋白含量是至少10、20、30、40、50或60%(w/w)。
植物蛋白质可来自植物蛋白质源,如豆类(legumes)和谷类(cereals),例如来自豆科(Fabaceae)(豆科(Leguminosae))、十字花科(Cruciferaceae)、藜科(chenopodiaceae)和禾本科(Poaceae)植物的物质,如大豆粉(soy bean meal)、羽扇豆粉(lupin meal)和油菜籽粉(rapeseed meal)。
在一个具体的实施方案中,所述植物蛋白质源是来自一种或多种豆科植物如大豆、羽扇豆、豌豆(pea)或菜豆(bean)的材料。
在另一个具体的实施方案中,所述植物蛋白质源是来自一种或多种藜科植物如甜菜(beet)、糖用甜菜(sugar beet)、菠菜(spinach)或昆诺阿藜(quinoa)的材料。
植物蛋白质源的其它例子是油菜籽和甘蓝(cabbage)。
大豆是优选的植物蛋白质源。
植物蛋白质源的其它例子是谷类如大麦、小麦、黑麦、燕麦、玉米(玉蜀黍)、稻和高粱。
合适的动物饲料添加剂是酶抑制剂、脂溶性维生素、水溶性维生素、痕量矿质和常量矿质(macro minerals)。
此外,任选的饲料添加剂组分是着色剂、芳香族化合物、稳定剂、抗微生物肽、和/或至少一种选自下组的其它酶:肌醇六磷酸酶EC 3.1.3.8或3.1.3.26;木聚糖酶EC 3.2.1.8;半乳聚糖酶EC 3.2.1.89;和/或β-葡聚糖酶EC 3.2.1.4。
抗微生物肽(AMP)的例子是CAP18、LeucocinA、Tritrpticin、内源性抗微生物多肽(Protegrin-1)、Thanatin、防卫素、Ovispirin如Novispirin(RobertLehrer,2000)、及其保留抗微生物活性的变体或片段。
抗真菌多肽(AFP)的例子是巨大曲霉(Aspergillus giganteus)和黑曲霉肽类,及其保留抗真菌活性的变体和片段,如WO 94/01459和PCT/DK02/00289中所公开的。
通常,脂溶性和水溶性维生素以及痕量矿质形成意欲添加于饲料中的所谓预混合料的部分,而常量矿质则通常单独添加到饲料中。
以下是这些组分的例子的非排他性列表:
脂溶性维生素的例子是维生素A、维生素D3、维生素E和维生素K,例如维生素K3。
水溶性维生素的例子是维生素B12、生物素和胆碱、维生素B1、维生素B2、维生素B6、烟酸、叶酸和泛酸(盐)(panthothenate),如Ca-D-泛酸。
痕量矿质的例子是锰、锌、铁、铜、碘、硒和钴。
常量矿质的例子是钙、磷和钠。
在更进一步的具体实施方案中,本发明的动物饲料组合物含有0-80%玉米;和/或0-80%高粱;和/或0-70%小麦;和/或0-70%大麦;和/或0-30%燕麦;和/或0-40%大豆粉;和/或0-10%鱼粉;和/或0-20%乳清。
通过以下实施例更进一步地描述本发明,所述实施例不应视为对本发明范围的限制。
材料和方法
蒸汽测试
开发了实验室规模的蒸汽处理测试系统来模拟大规模蒸汽制粒后发现的残余活性。
实验室规模蒸汽处理测试系统包括配有夹套的5升混合器(mixer)。该混合器还在底部具有开口,可用于热电偶或用来通过管道导入蒸汽。通过使蒸汽以1.5巴通过2.0mm孔来控制用于处理团粒(granulates)的蒸汽流。通过混合器夹套用蒸汽将混合器预热以达到100℃的温度。为了防止蒸汽接头(steam connections)和待插入混合器的管道中发生冷凝,在导入团粒之前进行蒸汽冲洗(steam flush)。测试条件包括一系列精确控制时序的动作,以保证良好的重现性。在零时间,在全速旋转混合器具的同时将1kg酶团粒添加到混合器中。之后马上将蒸汽管插入混合器中使团粒达到100℃,精确经过30秒后关闭蒸汽。将混合物在100℃再维持30秒,然后将大约200克倾倒于75微米筛网上。为了立即冷却热团粒,在样品网上部置放一网,利用连接于通风装置的筛盖(lid)在60秒时间内将样品冷却。在蒸汽处理之前和之后取参考样品,并送至FYT活性测量以计算残余活性。
制粒稳定性的测量
实施例5、8和11的实验设置:
将大约50g酶团粒和10kg饲料在小型水平混合器中预混合10分钟。将该预混物在较大的水平混合器中与90kg饲料混合10分钟。将饲料从混合器中以大约300kg/小时的速率导入调理器(带有蒸汽喷射的阶式混合器)。调理器通过喷射蒸汽将饲料加热到100℃(在出口测得)。在调理器内的停留时间为60-70秒。将饲料从调理器导入到装有3.0×35mm水平模具的SimonHeesen压力机(Simon Heesen press)中并压制成长度大约15mm的丸粒。压制后将丸粒置于空气冷却器中并冷却15分钟。
饲料配方:
74.0% 粉碎玉米(Grind corn)
20.7% 烤制大豆粗磨粉(Toasted soy grits)
5.0% 大豆油(soy oil)
0.3% 矿物质和维生素的Solivit Mikro 106预混料
12% 水含量
肌醇六磷酸酶活性分析
方法:肌醇六磷酸酶将肌醇六磷酸分解为磷酸盐,释放的磷酸盐与钒和钼(molydenium)氧化物反应,生成有色(黄色)复合物。在415nm处测定吸光度。
单位:1FTU=在标准条件下(如下所给出的),每分钟释放相当于1μM磷酸盐的磷酸盐(phosphate)的酶量。
缓冲液:
提取缓冲液:0.01%Tween 20(聚氧乙烯去水山梨糖醇单月桂酸酯(polyoxyethylene sorbitan monolaurate))
底物:5mM肌醇六磷酸、0.22M乙酸盐(乙酸钠/乙酸),pH 5.5。
试剂:5mM钒酸铵、20mM四水合七钼酸铵(ammonium heptamolybdatetetrahydrate)、40mM氨水、2.4M硝酸。
步骤:
饲料提取:在500ml提取缓冲液中将50g饲料提取1小时。如果活性高于2.5FTU/g饲料,则最后进一步在提取缓冲液中稀释。(检测水平是0.1FTU/g饲料)。将样品离心(在4000rpm 15分钟)。将300μl上清液与3ml底物混合并在37℃反应60分钟。添加2ml试剂。将样品离心(在4000rpm 10分钟)。在415nm处测定吸光度。相对于使用KH2PO4制备的标准曲线确定活性。
参考WO 2003/66847。
实施例
实施例1
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
11.362 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
1.7 kg肌醇六磷酸酶浓缩物
1.275 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11.5%棕榈油和22%碳酸钙将产物包覆。
实施例2
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
11.287 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.075 kgZn-乙酸盐x 2H2O
1.7 kg肌醇六磷酸酶浓缩物
0.215 kg 10%H2SO4
0.975 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11%棕榈油和22%碳酸钙将产物包覆。
实施例3
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
11.122 kg磨细的硫酸钠
在混合器FM 50中使用如下组成的造粒液造粒:
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.240 kg Zn-乙酸盐x 2H2O
1.7 kg肌醇六磷酸酶浓缩物
0.419 kg10%H2SO4
0.75 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11.0%棕榈油和22%碳酸钙将产物包覆。
实施例4
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
10.957 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.405 kg Zn-乙酸盐x 2H2O
1.7 kg肌醇六磷酸酶浓缩物
0.438 kg 2M H2SO4
0.75 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11%棕榈油和22%碳酸钙将产物包覆。
实施例5
在100℃,调理器的出口在制粒试验中测试实施例1至实施例4中生产的样品。使用分析法EB-SM 0559.02版本01(可从Novozymes索取得到)在制粒前和制粒后饲料丸粒中测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 以[%]计的肌醇六磷酸酶残余活性 |
实施例1 | 68.5 |
实施例2 | 84.5 |
实施例3 | 84.1 |
实施例4 | 84.8 |
结论是,与没有添加有机酸锌盐的参考实施例1相比,Zn-乙酸盐x 2H2O显著改进制粒稳定性。
实施例6
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
11.267 kg磨细的硫酸钠
0.45 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.375 kg Zn-柠檬酸盐x 2H2O
1.8 kg肌醇六磷酸酶浓缩物
0.033 kg 1N H2SO4
1.025 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用10%棕榈油和14%高岭土将产物包覆。
实施例7
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂,Avedex W80
11.342 kg磨细的硫酸钠
0.6 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.150 kgZn-柠檬酸盐x 2H2O
1.8 kg肌醇六磷酸酶浓缩物
0.027 kg 1N H2SO4
1.025 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用10.0%棕榈油和14%碳酸钙将产物包覆。
实施例8
在100℃,调理器的出口在制粒试验中测试实施例6和实施例7中生产的样品。使用分析法EB-SM 0559.02版本01(可从Novozymes索取得到)在制粒前和制粒后的饲料丸粒中测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 以[%]计的肌醇六磷酸酶残余活性 |
实施例1 | 68.5 |
实施例6 | 82.4 |
实施例7 | 84.5 |
结论是,与参考实施例1相比,Zn-柠檬酸盐x 2H2O显著改进制粒稳定性。
实施例9
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
11.212 kg磨细的硫酸钠
在混合器FM 50中使用如下组成的造粒液造粒:
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.150 kg Zn甲硫氨酸螯合物
1.7 kg肌醇六磷酸酶浓缩物
1.125 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11%棕榈油和22%碳酸钙将产物包覆。
实施例10
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
11.062 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.3 kg Zn甲硫氨酸螯合物
1.7 kg肌醇六磷酸酶浓缩物
1.2 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到850μm的产物。最后,按照如美国专利号4,106,991实施例22中描述的方式用11.0%棕榈油和22%碳酸钙将产物包覆。
实施例11
在100℃,调理器的出口在制粒试验中测试实施例9和实施例10中生产的样品。使用分析法EB-SM 0559.02版本01(可从Novozymes索取得到)在制粒前和制粒后的饲料丸粒中测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 以[%]计的肌醇六磷酸酶残余活性 |
实施例1 | 68.5 |
实施例9 | 85.4 |
实施例10 | 77.2 |
结论是,与参考实施例1相比,Zn甲硫氨酸螯合物显著改进制粒稳定性。
实施例12
测试了本身稳定性(per se stability)。以[%]计的肌醇六磷酸酶残余活性:
批次 | 40℃4周后 | 50℃4周后 | 40℃/60%RH 4周后 |
实施例1 | 88.5 | 81.9 | 51.9 |
实施例2 | 94.1 | 84.7 | 64.6 |
实施例3 | 96.5 | 90.0 | 63.0 |
实施例4 | 97.7 | 88.6 | 52.6 |
实施例6 | 94.5 | 83.5 | 77.5 |
实施例7 | 92.0 | 79.0 | 77.2 |
实施例9 | 95.9 | 84.0 | 80.4 |
实施例10 | 96.3 | 86.8 | 81.3 |
显示出特别是柠檬酸锌和甲硫氨酸锌具有很好的本身稳定性。
实施例13
制备了两种团粒:一种包含乙酸锌的团粒,和一种包含硫酸锌的团粒。
团粒1:
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
11.042 kg磨细的硫酸钠
在混合器FM 50中使用如下组成的造粒液造粒:
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.36 kg乙酸锌x 2H2O
1.475 kg肌醇六磷酸酶浓缩物
1.10 kg水
团粒2:
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
10.868 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.534 kg硫酸锌x 7H2O
1.475 kg肌醇六磷酸酶浓缩物
1.20 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到700μm的产物。
计算了与肌醇六磷酸酶单位的添加量相比,造粒和干燥后制得的样品的残余活性。使用分析法EB-SM 0511(可从Novozymes索取得到)测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 残余活性[%] |
团粒1 | 99 |
团粒2 | 93 |
结论是,与七水合硫酸锌相比,二水合乙酸锌在造粒过程中提供更好的肌醇六磷酸酶稳定化。
实施例14
制备了两种团粒:一种包含乙酸锌的团粒,和一种包含硫酸锌的团粒。
团粒1:
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
10.592 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.45 kg“玉米浆”粉
0.36 kg乙酸锌x 2H2O
1.475 kg肌醇六磷酸酶浓缩物
0.850 kg水
团粒2:
将由下列组成的粉末:
1.5 kg纤维状纤维素,Arbocel BC200
0.75 kg碳水化合物粘合剂Avedex W80
10.418 kg磨细的硫酸钠
0.75 kg碳水化合物粘合剂,Avedex W80
0.3 kg小麦淀粉
0.45 kg“玉米浆”粉
0.534 kg硫酸锌x 7H2O
1.475 kg肌醇六磷酸酶浓缩物
1.20 kg水
按照如美国专利号4,106,991实施例1中描述的方式进行造粒。
将所得的团粒在流化床中干燥到含水量低于1%并过筛,得到粒度范围为250μm到700μm的产物。
计算了与肌醇六磷酸酶单位的添加量相比,造粒和干燥后制得的样品的残余活性。使用分析法EB-SM 0511(可从Novozymes索取得到)测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 造粒后的残余活性[%] |
团粒1 | 97 |
团粒2 | 90 |
结论是,与七水合硫酸锌相比,二水合乙酸锌在造粒过程中提供更好的肌醇六磷酸酶稳定化。
按照如美国专利号4,106,991实施例22中描述的方式用9-10%棕榈油和22%碳酸钙包覆团粒1和团粒2。
然后对经包覆的团粒进行实验室蒸汽测试,其中将团粒(granulates)暴露于30秒的蒸汽处理,再在100℃继续再进行30秒,然后立即空气冷却。这些条件与大规模制粒条件相似。使用分析法EB-SM 0511(可从Novozymes索取得到)测量肌醇六磷酸酶含量。发现的肌醇六磷酸酶残余活性如下:
批次 | 以[%]计的肌醇六磷酸酶残余活性 |
团粒1 | 89 |
团粒2 | 84 |
结论是,与七水合硫酸锌相比,二水合乙酸锌在高温和高湿度下显示更好的酶稳定化。
Claims (23)
1.一种适用于饲料的酶团粒,其包含核心和围绕核心的层,其中该酶团粒包含的酶和有机酸锌盐作为混合物存在于所述酶团粒的核心中,所述有机酸锌盐选自下组:乙酸锌、柠檬酸锌和甲硫氨酸锌,所述酶是肌醇六磷酸酶,添加的有机酸锌盐的量为基于酶核心的0.05-10%。
2.权利要求1的酶团粒,其中所述酶团粒是经蒸汽处理的。
3.权利要求1或2的酶团粒,其中所述酶是食品级的。
4.权利要求1或2的酶团粒,其中所述有机酸锌盐是食品级的。
5.权利要求1或2的酶团粒,其中所述酶团粒的粒度在700μm以下。
6.权利要求1或2的酶团粒,其中所述酶团粒的粒度为210-390μm。
7.权利要求1或2的酶团粒,其中所述酶是热不稳定的。
8.权利要求1或2的酶团粒,其中所述酶团粒包含盐包层。
9.权利要求1或2的酶团粒,其中所述酶团粒包含聚合物包层。
10.权利要求1或2的酶团粒,其中所述酶团粒包含蜡包层。
11.权利要求1或2的酶团粒,其中所述酶团粒包含乳酸源。
12.一种制备权利要求1-11中任一项的酶团粒的方法,包括下列步骤:
a)制备包含有机酸锌盐和酶的核心
b)用包层材料包覆所述核心。
13.权利要求12的方法,其中所述酶团粒是在混合器、流化床、流化喷雾干燥器、喷雾流化器、喷雾干燥器或挤出机中制备的。
14.权利要求12或13的方法,其中所述有机酸锌盐作为液体添加到过程中。
15.权利要求12或13的方法,其中所述有机酸锌盐作为粉末添加到过程中。
16.权利要求12或13的方法,其中所述核心包含惰性颗粒。
17.一种饲料组合物,包含饲料组分和权利要求1-11中任一项的酶团粒。
18.权利要求17的饲料组合物,其中所述饲料组分选自下组:植物蛋白、脂溶性维生素、水溶性维生素、痕量矿物、常量矿物和它们的组合。
19.一种制粒饲料组合物,其包含权利要求1-11任一项的酶团粒。
20.一种经蒸汽处理的饲料组合物,其包含权利要求1-11任一项的酶团粒。
21.一种用于饲喂动物的方法,其包括对动物施用权利要求17-20中任一项的饲料组合物。
22.一种用于制造饲料组合物的方法,其包括下列步骤:
i.将饲料组分与权利要求1-11中任一项的酶团粒混合,
ii.对所述组合物(i)进行蒸汽处理,和
iii对所述组合物(ii)进行制粒。
23.权利要求1-11中任一项的酶团粒用于经蒸汽处理的制粒饲料组合物的用途。
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US20180242615A1 (en) | 2018-08-30 |
ES2577430T3 (es) | 2016-07-14 |
US11229223B2 (en) | 2022-01-25 |
US20080031998A1 (en) | 2008-02-07 |
CN103168928A (zh) | 2013-06-26 |
ES2712466T3 (es) | 2019-05-13 |
US20220110343A1 (en) | 2022-04-14 |
EP2051591B1 (en) | 2016-04-20 |
CN101505611A (zh) | 2009-08-12 |
EP3072399B1 (en) | 2018-12-19 |
EP2051591A1 (en) | 2009-04-29 |
WO2008017659A1 (en) | 2008-02-14 |
CN103109982A (zh) | 2013-05-22 |
EP3072399A1 (en) | 2016-09-28 |
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