CN112424208A - 具有免疫增强活性的氨基酸矿物质复合物及包含其的食品、药学或饲料用组合物 - Google Patents
具有免疫增强活性的氨基酸矿物质复合物及包含其的食品、药学或饲料用组合物 Download PDFInfo
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- CN112424208A CN112424208A CN201880095680.6A CN201880095680A CN112424208A CN 112424208 A CN112424208 A CN 112424208A CN 201880095680 A CN201880095680 A CN 201880095680A CN 112424208 A CN112424208 A CN 112424208A
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Abstract
本发明涉及具有免疫增强活性的氨基酸矿物质复合物及包含其的食品、药学或饲料用组合物,更详细地,涉及将上述氨基酸矿物质复合物作为有效成分而能够增强人类或除了人类之外的动物的免疫力或者增大抗病毒疫苗的抗体产生率的食品(或食品添加剂)、药品、饲料(或饲料添加剂)组合物。
Description
技术领域
本发明涉及具有免疫增强活性的氨基酸矿物质复合物及包含其的食品、药学或饲料用组合物,更详细地,涉及将上述氨基酸矿物质复合物作为有效成分而能够增强人类或除了人类之外的动物的免疫力或者增大抗病毒疫苗的抗体产生率的食品(或食品添加剂)、药品、饲料(或饲料添加剂)组合物。
背景技术
对于矿物质营养,氨基酸矿物质复合物的优点在于,在粘膜细胞及植物细胞中通过主动运输(active transport)或其他机制容易被吸收。即,若使用氨基酸作为搬运分子来使矿物质被吸收,则具有为了吸收而回避与对活性部位的竞争相关的问题及与矿物质之间的特殊微量要素的吸收阻碍功能等的优点。
矿物质氨基酸复合物通常通过α-氨基酸与金属离子之间的反应来生成,且上述复合物为了具有环结构而需要具有2或其以上的原子价的金属离子。在这种反应中,金属离子的正电荷与α-氨基酸的氨基或羧基的负电荷反应而被中和。
对于矿物质氨基酸复合物的结构、化学式及生物体利用性的相关文献有很多种,代表性地,可以举例阿什米德等(Ashmead et al.,Chelated Mineral Nutrition,(1982),Chas.C.Thomas Publishers,Springfield,Ill.)、阿什米德等(Ashmead et al.,Intestinal Absorption of Metal Ions,(1985))、阿什米德等(Ashmead et al.,FoliarFeeding of Plants with Amino Acid Chelates,(1986))及美国专利第4,020,158号、第4,167,564号、第4,216,143号、第4,721,644号、第4,599,152号、第4,774,089号、第4,830,716号、第4,863,898号、第4,725,427号等。
现有技术文献
专利文献
美国专利第4,020,158等
美国专利第4,167,564等
非专利文献
Ashmead et al.,Chelated Mineral Nutrition,(1982),Chas.C.ThomasPublishers,Springfield,Ill.Chas.C.Thomas Publishers,Springfield,Ill.
Ashmead et al.,Intestinal Absorption of Metal Ions,(1985)
Ashmead et al.,Foliar Feeding of Plants with Amino Acid Chelates,(1986)
发明内容
技术问题
本发明用于提供具有新型化学式的氨基酸矿物质复合物。
并且,本发明用于提供上述氨基酸矿物质复合物的制备方法。
并且,本发明用于提供包含上述氨基酸矿物质复合物的免疫活性增强用组合物。
解决问题的方案
本发明涉及氨基酸矿物质复合物,其特征在于,分子式为C4H9NO7Zn,且为下述化学式1的天门冬氨酸锌水合物:
上述天门冬氨酸锌水合物可以为水溶性。上述水溶性是指1g的上述天门冬氨酸锌水合物溶解于25℃的1ml至30ml,优选1ml至15ml水中的特性。
上述氨基酸矿物质复合物可以是免疫活性增强用复合物。
并且,本发明涉及上述氨基酸矿物质复合物的制备方法,包括:将锌前驱体及天门冬氨酸以1:0.8至2.5的摩尔(mole)比投入水中的步骤;以及在50℃至100℃的温度下加热上述步骤的锌前驱体及天门冬氨酸的混合水溶液10分钟至24小时来进行反应的步骤。
并且,本发明涉及将上述氨基酸矿物质复合物作为有效成分的免疫活性增强用组合物。
上述免疫活性增强用组合物可以为食品组合物、食品添加剂组合物、药学组合物、饲料组合物或饲料添加剂组合物。
上述免疫活性增强用组合物可以与抗病毒用疫苗一起或者依次给药。
上述抗病毒用疫苗可以为如下疫苗:
用于预防新城病、传染性支气管炎、球虫性腹泻病、禽痘(fowl pox)、禽霍乱病、呼肠孤病毒引起的腱鞘炎(病毒性关节炎)、禽喉气管炎、禽脑脊髓炎、传染性法氏囊病(IBD)、马立克氏病、沙门氏菌感染、鸡败血支原体(Mycoplasma gallisepticum)、禽鼻炎、禽疱疹及猪肺炎支原体、产蛋量下降综合征、传染性鼻炎(副鸡嗜血杆菌)、滑液囊支原体(mycoplasma synovia)或禽呼肠孤病毒的禽用疫苗;
用于预防或治疗胸膜肺炎、萎缩性鼻炎、伪狂犬病、猪丹毒、猪细小病毒、大肠杆菌肠毒素中毒、猪肺炎支原体、流感、钩端螺旋体病、大肠杆菌感染、猪繁殖和呼吸障碍综合症(PRRS)、博德特氏菌及A型及D型杆菌(multocida)感染、副猪嗜血杆菌感染、产气荚膜梭菌感染、轮状病毒感染、链球菌感染、格拉塞氏病(Glasser’s Disease)、肺炎、支气管炎博德特菌感染的哺乳类家禽用疫苗;或者
用于预防流感、甲型肝炎、乙型肝炎、丙型肝炎、单纯疱疹病毒(2型)、脊髓灰质炎、白喉、百日咳、B型流感嗜血杆菌(Hib)、麻疹、流行性腮腺炎、风疹、伤寒病、水痘(鸡痘)、登革热、爱泼斯坦-巴尔二氏病毒感染、人***瘤病毒感染、肺炎球菌感染、髓膜球菌感染、肺炎双球菌感染、病毒性髓膜炎、轮状病毒感染、蜱传脑炎、旅行者腹泻、霍乱、黄热病或结核的人类用疫苗。
发明的效果
本发明的氨基酸矿物质复合物为水溶性,从而在剂型化为粉末、颗粒、发泡锭等片剂的情况下,也可以快速地溶解于水中,在制备成液剂的情况下,也不会沉淀,而且矿物质吸收率优秀。
并且,将本发明的氨基酸矿物质复合物作为有效成分的免疫活性增强用组合物够增强人类或除了人类之外的动物的免疫力或者增大抗病毒疫苗的抗体产生率。
并且,当向除了人类之外的动物给予将本发明的氨基酸矿物质复合物作为有效成分的免疫活性增强用组合物时,可以增加饲料效率来减少用于达到相同增体量的饲料量,且可以增加相同期间的增体量。
并且,将本发明的氨基酸矿物质复合物作为有效成分的免疫活性增强用组合物显示显著降低在粪尿中产生的氨及硫化氢的产生量的效果,因此,对于人可以将其用作减轻屁味、老人味的除味用组合物,对于除了人类之外的动物,尤其,对于牛、猪等家畜,可以将其用作畜牧业粪尿味降低用组合物,从而可以对畜牧业农户的环境改善作出贡献。
附图说明
图1示出化学式1的天门冬氨酸锌水合物与相邻的天门冬氨酸锌水合物的天门冬氨酸(第二天门冬氨酸)进行配位键结合的示意图。
具体实施方式
以下,更详细地说明本发明。
本发明涉及氨基酸矿物质复合物,其特征在于,分子式为C4H9NO7Zn,且为下述化学式1的天门冬氨酸锌水合物:
上述天门冬氨酸锌水合物可以为水溶性。上述水溶性是指1g的上述天门冬氨酸锌水合物溶解于25℃的1ml至30ml,优选1ml至15ml的水中的特性。
上述氨基酸矿物质复合物可以使免疫活性增强用复合物。
上述水溶性氨基酸矿物质复合物可以通过将锌前驱体及天门冬氨酸以1:0.8至2.5的摩尔(mole)比投入水中的步骤;以及在50℃至100℃的温度下加热上述步骤的矿物质前驱体及天门冬氨酸的混合水溶液10分钟至24小时来进行反应的步骤来制备。
本发明涉及将上述氨基酸矿物质复合物作为有效成分的免疫活性增强用组合物。
上述锌前驱体为可以用作食品、药品或饲料的锌盐或锌氧化物,优选地,可以使用作为食品添加物安全性已被确认的如葡萄糖酸锌或硫酸锌等水溶性锌盐或者非水溶性的氧化锌,更优选地,增大抗病毒疫苗的抗体产生率的活性更优异的氧化锌。尤其,以往制备天门冬氨酸锌复合物的过程中,认为使用容易溶解于水中的水溶性锌盐是理所当然的,但是,经确认,当使用非水溶性的氧化锌的情形下在规定温度下加热规定时间以上时,形成效果更优异的天门冬氨酸锌复合物。
上述锌前驱体及天门冬氨酸的摩尔比意指锌与天门冬氨酸的摩尔比,1:0.8至2.5摩尔,优选地,1:1.5至2.2摩尔是有利的。在化学式1的天门冬氨酸锌水合物中,锌与天门冬氨酸的理论摩尔比为1:2,但是,当相比于锌前驱体添加过量的天门冬氨酸时,在不溶性氧化锌被减少的情形下能够更容易地形成水溶性天门冬氨酸锌水合物。
在上述进行反应的步骤中,在50℃至100℃的温度下加热10分钟至24小时,尤其,在水溶性锌盐的情况下,仅在上述范围内相对低的温度下加热短时间,例如,在50℃至80℃的温度下加热10分钟至1小时也可以结束反应,但是,在氧化锌的情况下,在80℃至100℃的温度下加热1小时以上,优选地,加热1.5小时。另一方面,水溶性锌盐或氧化锌都在3小时以内大部分约95%,优选地,99%以上的天门冬氨酸进行反应,因此,即使加热超过3小时不溶物或未反应物不会减少得多,在上述进行反应的步骤中,为了缩短反应时间,可以利用超声波处理。
还可以包括从上述步骤的反应液中去除不溶物的步骤。就去除不溶物的步骤而言,也可以在将水溶性锌盐用作锌前驱体的情况下添加,但是,尤其在将氧化锌用作锌前驱体的情况下,可能更需要此步骤。
在去除上述不溶物的步骤中,可以使用数μm以下,例如,0.2μm的滤纸或滤布进行过滤,且在上述过滤之前,或者与上述过滤无关地,可以通过离心分离来去除不溶物,
不同于以往的天门冬氨酸锌复合物的制备方法,上述制备方法除了锌前驱体、天门冬氨酸及水之外不需要额外的添加剂且由于制备工艺简单而具有能够降低工序费用的优点。
另一方面,本说明书中,“作为有效成分”意指包含在达到本发明的矿物质氨基酸复合物的功效或活性时所需要的充分的量。在本发明的一具体例中,本发明的组合物中,食品组合物、药学组合物或饲料组合物包含例如0.001mg/kg以上,优选地,包含0.1mg/kg以上,更优选地,包含10mg/kg以上,更加优选地,包含100mg/kg以上,更加优选地,包含250mg/kg以上,最优选地,包含0.1g/kg以上的矿物质氨基酸复合物。只不过,食品添加剂组合物或饲料添加剂组合物中包含10g/kg以上的矿物质氨基酸复合物,优选地,包含50g/kg以上,更优选地,包含100g/kg以上,也可以仅纯粹使用矿物质氨基酸复合物。
本发明的药学组合物不仅包含用于人类的药学组合物,而且还包含除了人类之外的如禽类或哺乳动物等兽医学用药学组合物。药学组合物通过除了上述有效成分之外还可以使用药剂学上适当且生理学上可接受的辅助剂来制备,作为上述辅助剂,可以使用赋形剂、崩解剂、甜味剂、结合剂、包衣剂、膨胀剂、润滑剂、光滑剂或香味剂等。
上述药学组合物可以与抗病毒用疫苗一起或者依次给药。为此,本发明的药学组合物可以与抗病毒用疫苗一起被剂型化,也可以各自单独地被剂型化。
上述抗病毒用疫苗可以为用于预防新城病、传染性支气管炎、球虫性腹泻病、禽痘、禽霍乱病、呼肠孤病毒引起的腱鞘炎(病毒性关节炎)、禽喉气管炎、禽脑脊髓炎、传染性法氏囊病(IBD)、马立克氏病、沙门氏菌感染、鸡败血支原体、禽鼻炎、禽疱疹及猪肺炎支原体、产蛋量下降综合征、传染性鼻炎(副鸡嗜血杆菌)、滑液囊支原体或禽呼肠孤病毒的禽用疫苗;用于预防或治疗胸膜肺炎、萎缩性鼻炎、伪狂犬病、猪丹毒、猪细小病毒、大肠杆菌肠毒素中毒、猪肺炎支原体、流感、钩端螺旋体病、大肠杆菌感染、猪繁殖和呼吸障碍综合症(PRRS)、博德特氏菌及A型及D型杆菌感染、副猪嗜血杆菌感染、产气荚膜梭菌感染、轮状病毒感染、链球菌感染、格拉塞氏病、肺炎、支气管炎博德特菌感染的哺乳类家禽用疫苗;或者用于预防流感、甲型肝炎、乙型肝炎、丙型肝炎、单纯疱疹病毒(2型)、脊髓灰质炎、白喉、百日咳、B型流感嗜血杆菌(Hib)、麻疹、流行性腮腺炎、风疹、伤寒病、水痘(鸡痘)、登革热、爱泼斯坦-巴尔二氏病毒感染、人***瘤病毒感染、肺炎球菌感染、髓膜球菌感染、肺炎双球菌感染、病毒性髓膜炎、轮状病毒感染、蜱传脑炎、旅行者腹泻、霍乱、黄热病或结核的人类用疫苗。
为了给药上述药学组合物,除了上述记载的有效成分之外,还可以通过包含一种以上的药学上可接受的载体来优选地制剂化为药学组合物。
上述药学组合物的制剂形态可以为颗粒剂、散剂、片剂、包衣锭、胶囊剂、栓剂、液剂、糖浆、汁、悬浊剂、乳剂、滴定剂或可以注射的液剂等。例如,为了制剂化为片剂或胶囊剂的形态,有效成分可以与如乙醇、丙三醇、水等口服且无毒性的药学上可接受的非活性载体结合。并且,在希望或需要时,可以包含适合的结合剂、润滑剂、崩解剂及发色剂或它们混合物。适合的结合剂包含如淀粉、明胶、葡萄糖或β-乳糖等天然糖、如玉米甜味剂、***胶、黄蓍胶或油酸钠等天然及合成胶、硬脂酸钠、硬脂酸镁、苯甲酸钠、醋酸钠、氯化钠等,但并不限定于此。崩解剂包含淀粉、甲基纤维素、琼脂、膨润土、黄原胶等,但并不限定于此。
对于制剂化为液态溶液的组合物,作为药学上可接受的载体为适合用于灭菌及生物体,可以使用盐水、灭菌水、林格氏液、缓冲生理盐水、白蛋白注射溶液、右旋糖溶液、麦芽糖糊精溶液、丙三醇、乙醇或通过混合这些成分中的一种以上来使用,根据需要,可以添加抗氧化剂、缓冲剂、抑菌剂等其他常规的添加剂。并且,还可以附加地添加稀释剂、分散剂、表面活性剂、结合剂及润滑剂来制剂化为如水溶液、悬浊液、乳浊液等注射用剂型、丸剂、胶囊、颗粒或片剂。
进一步的,可以利用作为该领域的适当的方法的在Remington's PharmaceuticalScience,Mack Publishing Company,Easton PA中公开的方法而根据各疾病或成分来优选地制剂化。
本发明的药学组合物可以通过口服或非口服方式给药,在非口服给药的情况下,可以通过静脉内注入、皮下注入、肌肉注入、腹腔注入、经皮给药等方式给药,优选为口服给药。
本发明的药学组合物的适当的给药量可以根据如制剂化方法、给药方式、患者或动物的年龄、体重、性别、病态、饮食、给药时间、给药途径、***速度及反应灵敏度等主要因素而多样化,普通的熟练的医生兽医可以容易决定及处方对所希望的治疗或预防具有效果的给药量。根据本发明的优选实例,本发明的药学组合物的一日给药量为0.001-10g/Kg。
就本发明的药学组合物而言,根据本发明所属技术领域的普通技术人员容易实施的方法并利用药学上可接受的载体和/或赋形剂来制剂化,从而可以制备为单位容量形态或者通过放入大容量容器内来制备。此时,剂型可以为油或水性介质中的溶液、悬浊液或乳化液形态、或者浓缩剂、粉末剂、颗粒剂、片剂或胶囊剂形态,还可以包含分散剂或稳定剂。
在这里所使用的术语“有效量”是指研究者、兽医、医生或其他临床医师所认为的在组织***、动物或人类中诱导生物学或医学性反应的有效成分或药学组合物的量,其包含诱导相应疾病或障碍的症状的缓解的量。对于普通技术人员而言,对本发明的有效成分的有效量及给药次数根据所需的效果而变化是显而易见的。因此,需要给药的最佳给药量可以由普通技术人员容易决定,可以根据包括疾病的种类、疾病的严重度、组合物中所含有的有效成分及其他成分的含量、剂型的种类、以及患者或动物的年龄、体重、一般健康状态、性别及饮食、给药时间、给药途径及组合物的分泌率、疗程、同时使用的药物在内的多种因素来调节。对于本发明的预防、治疗或改善方法,在成人或成体的情况下,当将矿物质氨基酸复合物以一日一次给药数次时,优选地,给药0.001mg/kg至10g/kg的剂量。
与上述药学组合物相同的方式制剂化根据本发明的食品组合物或食品添加剂组合物,从而可以利用为功能性食品或者添加至各种食品中。作为可以添加上述食品添加剂组合物的食品有,例如,饮料类、酒精饮料类、饼干类、减肥杂粮饼、乳制品、肉类、巧克力、披萨、拉面、其他面类、口香糖类、冰淇淋类、维生素复合物、健康辅助食品类等。
本发明的食品组合物不仅包含作为有效成分的矿物质氨基酸复合物,而且还可以包含制备食品时通常添加的成分是,例如,包含蛋白质、碳水化合物、脂肪、营养素、调味剂及香味剂。作为上述碳水化合物的例,如葡萄糖、果糖等单糖;如麦芽糖、蔗糖、低聚糖等二糖;及如糊精、环糊精等多糖等常规的糖以及如木糖醇、山梨糖醇、赤藓糖醇等糖醇。作为香味剂,可以使用天然香味剂[索马汀、甜叶菊提取物(例如,莱鲍迪甙A、甘草甜素等])及合成香味剂(糖精、阿斯巴甜等)。例如,在本发明的食品组合物制备为口服液或饮料类的情况下,除了本发明的米、米糠或粗糠提取物之外,还可以包含柠檬酸、液态果糖、白糖、葡萄糖、醋酸、苹果酸、果汁、以及各种植物提取液等。
本发明提供包含将上述矿物质氨基酸复合物作为有效成分来包含的免疫活性增强用食品组合物的健康功能食品。所谓健康功能食品是指将米糠提取物或米糠粉末添加至饮料、茶类、香料、口香糖、饼干类等食品材料中或者由胶囊化、粉末化、悬浊液等制备的食品,且在摄取其时对健康带来特定效果,但不同于一般药品,由于将食品作为原料,因此具有在长期服用时不会产生副作用等的优点。如此获得的本发明的健康功能食品能够日常摄取,因此非常有用。就这种健康功能食品中的米糠提取物或米糠粉末的添加量而言,根据作为对象的健康功能食品的种类不同而无法统一规定,但是,能够以在不影响食品原有的味道的范围内的量添加即可,且相对于对象食品,通常以0.01重量百分比至50重量百分比的范围添加,优选地,以0.1重量百分比至20重量百分比的范围添加。并且,在丸剂、颗粒剂、片剂或胶囊剂形态的健康功能食品的情况下,通常以0.1重量百分比至100重量百分比的范围添加,优选地,以0.5重量百分比至80重量百分比的范围添加。在一具体例中,本发明的健康功能食品可以为丸剂、片剂、胶囊剂或饮料的形态。
就根据本发明的饲料添加剂组合物而言,以猪、鸡、鸭、牛、羊、山羊、狗等家畜的饲料组合物的固体成分基准,以0.01至1重量百分比添加及混合上述饲料添加剂组合物而向各种动物给药来使用,更优选地,可以用作猪饲料的添加剂。
本发明的上述饲料添加剂组合物可以制备成发酵饲料、配合饲料、颗粒形态及青贮饲料(silage)等形态的饲料组合物,但并不限定于此,可以根据需要进行变形来制备。
发明的实施方式
以下,通过实施例等更加详细地说明本发明,但是,以下不应解释为通过实施例等缩小或限制本发明的范围和内容。并且,若基于以包括以下实施例的本发明的公开内容,则就普通技术人员可以容易实施没有具体揭示实验结果的本发明而言,是明了的,且这种变形及修改包括在所附的发明要求保护范围内也是理所当然的。
实施例1-1:制备天门冬氨酸锌复合物(使用硫酸锌)
将60g的天门冬氨酸放入500ml的水中并进行搅拌,且向其中放入硫酸锌(35重量百分比的锌)并进行搅拌,并以60℃的温度充分反应30分钟以上后,进行离心分离后冻干上清液,从而获得约70g的水溶性天门冬氨酸锌复合物。但以1:2的摩尔(mole)比的含量投入上述硫酸锌与上述天门冬氨酸。
比较例1-1:制备谷氨酸锌复合物(使用硫酸锌)
将60g的谷氨酸放入500ml的水中并进行搅拌,且向其中放入硫酸锌(35%的锌)并进行搅拌,并以50℃至100℃的温度充分反应30分钟以上后,进行离心分离后冻干上清液,由此获得约70g的水溶性谷氨酸锌复合物。但是,以1:2的摩尔(mole)比的含量投入上述硫酸锌与上述谷氨酸。
比较例1-2:制备天门冬氨酸铁复合物(使用乳酸铁)
将60g的天门冬氨酸放入500ml的水中并搅拌,且向其中放入乳酸铁(35%的铁)并进行搅拌,并以50℃至100℃的温度充分反应30分钟以上后,进行离心分离后冻干上清液,由此获得约70g的乳酸铁天门冬氨酸复合物。但是,以1:2的摩尔(mole)比的含量投入上述乳酸铁与上述天门冬氨酸。
实验例1:反应收率评价
实施例1-1、实施例1-2、比较例1-1、比较例1-2的反应收率的比较结果如下列表1所示。通过ICP分析法分析所生成的矿物质氨基酸复合物中所含有的矿物质的含量,并用其除于所投入的矿物质前驱体的理论矿物质含量(锌或铁粉)来计算反应收率。
表1
区分 | 实施例1-1 | 实施例1-2 | 比较例1-1 | 比较例1-2 |
反应收率(%) | 100% | 100% | 48% | 60% |
上述结果显示,实施例1-1及实施例1-2的反应收率为100%,说明全部参与反应而制备水溶性天门冬氨酸锌复合物。与此相反,使用谷氨酸的比较例1-1中,上述谷氨酸未完全溶解于水中,因此反应收率为约50%以下,明显减少,使用乳酸铁的比较例1-2中,反应收率为约60%,也明显减少。
实施例2-1:制备饲料添加剂组合物
向100重量份的米糠中放入20重量份的上述实施例1-1的天门冬氨酸锌复合物并进行混合后,在80℃的干燥机中进行干燥而获得水分含量为10%以内的固体,在滚碎粉碎机中进行粉碎并加工成好看的粉末,由此制备饲料添加剂组合物。
比较例2-1:制备饲料添加剂组合物
除了使用比较例1-1的谷氨酸锌复合物代替实施例1-1来制备饲料添加剂组合物之外,以与上述实施例2-1相同的方法进行。
比较例2-2:制备饲料添加剂组合物
除了使用比较例1-2的乳酸铁天门冬氨酸复合物代替实施例1-1来制备饲料添加剂组合物之外,以与上述实施例2-1相同的方法进行。
实验例2:猪饲料效率及粪内恶臭物质评价
为了调查向猪供给饲料添加剂组合物后对饲料效率和发育产生的效果,将通过上述实施例2-1及比较例2-1、比较例2-2制备的饲料添加剂组合物以0.2重量百分比与猪饲料进行混合来供给,并将结果表示在以下表2中。
[表2]
区分 | 实施例2-1 | 比较例2-1 | 比较例2-2 |
饲养头数 | 30只 | 30只 | 30只 |
饲料摄取量 | 223kg | 285kg | 281kg |
达到110kg的时间 | 160.7日 | 189.8日 | 181.1日 |
如上述表1所示,可知:当向猪饲料中添加实施例2-1的饲料添加剂组合物时,显示达到110kg的时间缩短约20日左右的结果,因此饲料摄取量也减少约20.7%。并且,为了分析与猪粪中的恶臭相关的氨及硫化氢的产生量,在上述中结束实验时采取新鲜的粪便且将该粪便以特殊方式制造的每个塑料容器中装入100g,随着时间的经过(3小时后、24小时后),利用数码气体测量仪(多气体监视器PGM-7800,RAE***公司,美国(Multi GasMonitor PGM-7800,RAE Systems Inc.,USA))测量在室温下产生的气体中氨及硫化氢的浓度,并将其结果表示在以下表3中。
[表3]
如上述表3所示,从结果可知,相比于比较例2-1、比较例2-2,实施例2的在粪尿中产生的恶臭气体中氨及硫化氢的浓度显著低。
实施例1-2:制备天门冬氨酸锌复合物(使用氧化锌)
将57.5g天门冬氨酸放入500ml的水中并进行搅拌,且向其中放入17.5g的氧化锌(81重量百分比的锌)并进行搅拌,且以90℃的温度充分反应120分钟以上后,利用0.2μm的滤纸过滤反应液,在65℃的温度下将滤液浓缩3小时至80%后,进行真空干燥来获得水溶性天门冬氨酸锌复合物。
实验例3:确认天门冬氨酸锌复合物的结构
利用布鲁克智能APEX II x射线检晶仪(Bruker SMART APEX II X-rayCrystallograph(XRC))及布鲁克SHELXTL结构分析程序(Bruker SHELXTL StructureAnalysis Program)确认实施例1-2的天门冬氨酸锌复合物的结构。
晶体学数据
分子式:C4H9NO7Zn
分子量:248.49
晶系:斜方晶系
空间组:P2(1)2(1)2(1)
Z4
计算的密度:1.956Mg/m3
温度:296(1)K
绝对结构参数:0.00
结晶大小:0.18×0.12×0.08mm3
F(000)504
吸收系数:2.920mm-1
表4中示出原子坐标(×104)及等效各向同性位移(equivalent isotropicdisplacement)参数U(eq)定义为正交化的(orthogonalized)Uij张量(tensor)的追踪(trace)的1/3。
[表4]
[表5]
为了生成同等原子,使用对称变换。#1-x,y-1/2,-z+3/2;#2-x,y+1/2,-z+3/2。
[表6]
[表7]
表8中示出扭转角(torsion angles)(°)。
[表8]
通过综合上述结果确认到实施例2-1的水溶性天门冬氨酸锌复合物的分子式为C4H9NO7Zn,且为下述化学式1的天门冬氨酸锌水合物。
化学式1:
就上述化学式1的天门冬氨酸锌水合物而言,由于锌与两个水分子配位键结合,与第一天门冬氨酸的两个羟基中的两个氧原子、一个氨基中的一个氮原子配位键结合,与第二天门冬氨酸的一个氧原子配位键结合,从而被确认为由具有总共六个配位键的锌与天门冬氨酸以1:1的摩尔比结合而成的新型天门冬氨酸锌水合物。
将上述化学式1的天门冬氨酸锌水合物与相邻的天门冬氨酸锌水合物的天门冬氨酸(第二天门冬氨酸)进行配位键结合的示意图表示在图1中。
比较例3:常用天门冬氨酸锌复合物
使用分子式为C8H10N2O8Zn.2H,分子量为329.58,且锌与天门冬氨酸以1:2的摩尔结合的络合物(非水溶性)来制备饲料添加剂组合物。
实验例4:猪***抗体形成能力
为了调查饲料添加剂组合物对猪的血液特性及***抗体生成能力产生的影响,将氧化锌粉末、硫酸锌粉末、上述实施例1-1的天门冬氨酸锌复合物(使用硫酸锌)、实施例1-2的天门冬氨酸锌复合物(使用氧化锌)及比较例3的天门冬氨酸锌复合物分别添加至饲料中来进行动物实验6周。开始实验时的体重为25.56±2.22kg。
饲料使用根据NRC(2012)要求量配合的以玉米-大豆粕为主的饲料,向上述饲料中添加0.1重量百分比的氧化锌粉末(CON 1)、0.3重量百分比的硫酸锌粉末(CON 2)、0.3重量百分比的实施例1-1的天门冬氨酸锌复合物(实施例1-1-0.3)、0.1重量百分比的实施例1-2的天门冬氨酸锌复合物(实施例1-2-0.1)、0.3重量百分比的实施例1-2的天门冬氨酸锌复合物(实施例1-2-0.3)以及0.3重量百分比的比较例3的天门冬氨酸锌复合物(比较例3-0.3),并将240头的三元杂交种(杜洛克约克郡长白(Duroc x Yorkshire x Landrace))生长猪处理4次,每次处理反复8次,每次反复中完全任意分配5头。上述饲料是自由摄取的,且水是以利用自动供水机能够自由摄取的方式供给的。
1)体重变化
开始供给饲料时及结束时测量体重并将量体重变化表示在表9中。
[表9]
区分 | CON 1 | CON 2 | 实施例1-1-0.3 | 实施例1-2-0.1 | 实施例1-2-0.3 | 比较例3-0.3 |
开始时 | 22.56 | 22.56 | 22.56 | 22.56 | 22.56 | 22.56 |
结束时 | 53.62b | 53.74b | 54.43ab | 55.40a | 54.76ab | 54.55a |
在上述6周的饲养实验中,未观察到体重的显著变化,但相比于CON 1及CON 2,在实施例1-1-0.3、实施例1-2-0.1、实施例1-2-0.3及比较例3-0.3中,体重有所增加。
2)血液中锌及免疫球蛋白含量
为了测量血液中锌及免疫球蛋白含量,结束实验节点(6周),利用E3 EDTA真空管(碧迪真空***,富兰克林湖,新泽西州(Bec ton Dickinson Vacutainer systems,Granklin Lakes,NJ))从相同对象的颈静脉(Jugular vein)采取5ml的血液后,并从在4℃的温度下以3000rpm离心分离15分钟后在所获得的血清中测量锌含量及免疫球蛋白含量,表示在表10中。
[表10]
区分 | CON 1 | CON 2 | 实施例1-1-0.3 | 实施例1-2-0.1 | 实施例1-2-0.3 | 比较例3-0.3 |
锌(μg/dL) | 100c | 98c | 158b | 177ab | 205a | 180ab |
IgG(mg/dL) | 472b | 466b | 480b | 504ab | 546a | 489b |
相比于CON 1及CON 2,实施例1-1-0.3、实施例1-2-0.1、实施例1-2-0.3及比较例3-0.3的血液中锌含量显著增加。尤其,即使CON 1及CON 2的锌供给量相比于实施例1-1-0.3、实施例1-2-0.3及比较例3-0.3更多,但血液中锌含量在上述实施例中更高,在这点上示出本发明的矿物质氨基酸复合物和比较例3的天门冬氨酸锌复合物的高的生物体利用率。相比于CON 1及CON 2,实施例1-1-0.3及比较例3-0.3的血液中免疫球蛋白含量没有显著差异,但实施例1-2-0.3中显著增加。尤其,将给药量降低至1/3的实施例1-2-0.1中也没有显著差异,但呈现明显增加的倾向。
3)***抗体形成率
在上述实验例3的2)中获得的血清中分析***抗体滴度(FM DV-O型)。***抗体滴度检查是,通过SP ELISA每次执行SP抗体检查来分析测量的个别抗体滴度(percentage inhibition titer,PI数值)而考察抗体滴度持续程度、根据各疫苗接种时期的抗体滴度形成程度。利用简单线性回归分析(simple linear regression analysis)验证结果。使用PrioCHECK FMDV型O ELISA试剂盒(Prionics Lely stad B.V,荷兰)并根据由制造商提供的方法进行SP抗体检查。就E LISA结果判定而言,根据制造商的计算式{100-(corrected OD450 tes t sample/correcte OD 450max)×100}dp(100-(校正的OD450试验样本/校正的OD 450最大)×100}dp)而获得PI数值,当PI数值为50以上时,判定为阳性,当小于50时,判定为阴性。
[表11]
区分 | CON 1 | CON 2 | 实施例1-1-0.3 | 实施例1-2-0.1 | 实施例1-2-0.3 | 比较例3-0.3 |
抗体(PI) | 47.38b | 47.52b | 51.88b | 55.65b | 78.07a | 54.45b |
阳性率 | 50 | 50 | 75 | 100 | 100 | 75 |
相比于CON 1及CON 2,实施例1-1-0.3、实施例1-2-0.1及比较例3-0.3中,***抗体形成率有所增加,但并没有显著差异,而相比于CON 1及CON 2,仅在实施例1-2-0.3中显著增加。在CON 1及C ON 2中,***抗体阳性率为50%,但在实施例1-1-0.3及比较例3-0.3中,***抗体阳性率为75%,在实施例1-2-0.1及实施例1-2-0.3中,***抗体阳性率为100%。
就上述实验例3的所有资料而言,为了利用SAS(2013)的一般线性模型程序(General Linear Model procedure)来调查根据添加水平的功效,通过线性、二次、三次(linear、quadratic、cubic)分析并用邓肯的多范围测试(Duncan's multiple rangetest)进行处理,从而将平均之间差异在P〈0.05验证显著性。
实验例4:流感H1N1抑制效果
将29±1g的12周龄雌性(ICR)小鼠(东方生物公司,首尔,韩国(Orient Bio Inc.,Seoul,Korea))使用于实验中。实验之前,对小鼠进行7日的驯化过程后用于实验,在驯化过程中,将小鼠在调节成22±1℃的温度及55±10%湿度的饲养室自由摄取食物来饲养,将明暗周期以12小时的周期进行调整。
将上述小鼠分成每组10只,向病毒对照组、CON 1、实施例1-1、实施例1-2及比较例3给药组投H1N1病毒后,将各自的试料稀释在生理盐水中并口服给药20μg/kg/d,给药5日。本实验中所使用的阴性对照组为生理盐水,病毒对照组(virus control)为虽然被病毒感染但未给药试剂的组。
1)体重变化
开始供给饲料时及结束时测量体重并将量体重变化表示在表12中。
[表12]
区分 | 阴性对照组 | 病毒对照组 | CON 2 | 实施例1-1 | 实施例1-2 | 比较例3 |
开始时(0日) | 100.0 | 100.0 | 100.0 | 100.0 | 100.0 | 100.0 |
结束时(5日) | 106.0b | 99.1a | 99.9a | 104.8ab | 106.8b | 103.7ab |
投病毒后,相比于阴性对照组,病毒对照组及CON2中的体重减少,但是,实施例1-1,实施例1-2及比较例3中对于体重增加方面,与阴性对照组没有显著差异。
2)肺组织中H1N1病毒含量
弄死小鼠后,用滴定的方式测量肺组织中所存留的病毒的含量并表示在表13中。
[表13]
经确认,仅在实施例1-2中,肺组织中病毒含量显著减少。
Claims (8)
2.根据权利要求1所述的氨基酸矿物质复合物,其特征在于,所述天门冬氨酸锌水合物为水溶性的。
3.根据权利要求2所述的氨基酸矿物质复合物,其特征在于,用于增强免疫活性。
4.一种权利要求1至3中任一项所述的氨基酸矿物质复合物的制备方法,其中,包括:
将锌前驱体及天门冬氨酸以1:0.8至2.5的摩尔比投入水中的步骤;以及
在50℃至100℃的温度下加热所述步骤的锌前驱体及天门冬氨酸的混合水溶液10分钟至24小时而进行反应的步骤。
5.一种免疫活性增强用组合物,其特征在于,将权利要求1至3中任一项所述的氨基酸矿物质复合物作为活性成分。
6.根据权利要求5所述的免疫活性增强用组合物,其特征在于,所述组合物为食品组合物、食品添加剂组合物、药学组合物、饲料组合物或饲料添加剂组合物。
7.根据权利要求5所述的免疫活性增强用组合物,其特征在于,将所述免疫活性增强用组合物与抗病毒用疫苗一起或者依次给药。
8.根据权利要求7所述的免疫活性增强用组合物,其特征在于,
所述抗病毒用疫苗为如下疫苗:
用于预防新城病、传染性支气管炎、球虫性腹泻病、禽痘、禽霍乱病、呼肠孤病毒引起的腱鞘炎(病毒性关节炎)、禽喉气管炎、禽脑脊髓炎、传染性法氏囊病、马立克氏病、沙门氏菌感染、鸡败血支原体、禽鼻炎、禽疱疹及猪肺炎支原体、产蛋量下降综合征、传染性鼻炎(副鸡嗜血杆菌)、滑液囊支原体或禽呼肠孤病毒的禽用疫苗;
用于预防或治疗胸膜肺炎、萎缩性鼻炎、伪狂犬病、猪丹毒、猪细小病毒、大肠杆菌肠毒素中毒、猪肺炎支原体、流感、钩端螺旋体病、大肠杆菌感染、猪繁殖和呼吸障碍综合症、博德特氏菌及A型及D型杆菌感染、副猪嗜血杆菌感染、产气荚膜梭菌感染、轮状病毒感染、链球菌感染、格拉塞氏病、肺炎、支气管炎博德特菌感染的哺乳类家禽用疫苗;或者
用于预防流感、甲型肝炎、乙型肝炎、丙型肝炎、单纯疱疹病毒(2型)、脊髓灰质炎、白喉、百日咳、B型流感嗜血杆菌(Hib)、麻疹、流行性腮腺炎、风疹、伤寒病、水痘(鸡痘)、登革热、爱泼斯坦-巴尔二氏病毒感染、人***瘤病毒感染、肺炎球菌感染、髓膜球菌感染、肺炎双球菌感染、病毒性髓膜炎、轮状病毒感染、蜱传脑炎、旅行者腹泻、霍乱、黄热病或结核的人类用疫苗。
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