CN110063961B - 一种刺五加皂苷b的新用途 - Google Patents
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Abstract
本发明涉及动物抗冷应激技术领域,具体涉及一种刺五加皂苷B的新用途,所述新用途为所述刺五加皂苷B用于动物抗冷应激。本发明提供的刺五加皂苷B的新用途,其有效成分皂苷类物质能够穿过动物的血脑屏障直接作用于抵抗冷应激的神经中枢,提高了畜禽抵抗冷应激相关激素的分泌。刺五加作为天然植物成分不仅对环境没有污染,动物也不会对其产生依赖性,是保证畜禽健康、提高经济效益的有效手段,在养殖领域具有十分重要的意义。
Description
技术领域
本发明涉及动物抗冷应激技术领域,具体涉及一种刺五加皂苷B的新用途。
背景技术
寒冷刺激是北方寒区动物最普遍的应激原,冬季畜禽的养殖,特别是早春和晚秋母猪产仔、冬季母牛产犊、早春孵化鸡雏都会由于气温低而导致动物发生冷应激,冷应激所引起的动物营养代谢紊乱、生长缓慢、抗病性差甚至死亡是制约北方畜牧业和养殖业发展的主要因素之一。
当动物体遭受寒冷时,需要摄取大量的饲料用于维持体温、加强代谢以及提高免疫力,从而使机体处于健康的动态平衡,但这将会降低饲料的转化效率,影响动物的生产性能。提高动物自身的抗冷应激能力,不仅会提高养殖效益,也为健康养殖打下基础。
为了减小冷应激对动物的影响,目前通常采用增加畜舍保温隔热层、窗口设置挡风导流板以及挂门帘的方式;或者采用煤炉,暖风炉等保温措施,但上述方式不但会增加养殖投入,而且会加大人工劳动强度;
发明内容
本发明为了解决上述技术问题,提供一种刺五加皂苷B的新用途。
本发明包括以下技术方案:一种刺五加皂苷B的新用途,所述新用途为所述刺五加皂苷B用于动物抗冷应激。
进一步地,所述刺五加皂苷B用于动物抗冷应激的时间为动物冷应激条件下12-15天。
优选地,所述刺五加皂苷B用于动物抗冷应激的时间为动物冷应激条件下第14天。
进一步地,所述刺五加皂苷B对所述动物下丘脑抗冷应激作用的影响通过测定注射有刺五加皂苷B的动物下丘脑中的Hsp70蛋白表达量来反应。
优选地,所述Hsp70蛋白表达量的测定方法中采用的实验动物为小鼠。
更进一步地,采用Hsp70蛋白表达量的测定方法之前还包括步骤:使用不同浓度的刺五加皂苷B分别在饲养1、7、14、21d之后腹腔注射小鼠,浓度为0、2、4、8mg/kg,每组6个重复,每个重复一只。
采用上述技术方案,包括以下有益效果:本发明提供了一种刺五加皂苷B的新用途,其有效成分皂苷类物质能够穿过动物的血脑屏障直接作用于抵抗冷应激的神经中枢,提高了畜禽抵抗冷应激相关激素的分泌。刺五加作为天然植物成分不仅对环境没有污染,动物也不会对其产生依赖性,是保证畜禽健康、提高经济效益的有效手段,在养殖领域具有十分重要的意义。
附图说明
图1为小鼠冷应激后第14天时不同浓度刺五加皂苷B下丘脑中Hsp70蛋白的相对表达量;
图2为冷应激0d、1d、7d、14d、21d时,注射4mg/kg刺五加皂苷B下丘脑中蛋白的相对表达。
具体实施方式
下面通过具体的实施例并结合附图对本发明做进一步的详细描述。
实施例:
本实施例提供了一种刺五加皂苷B的新用途,刺五加皂苷B作为一种天然的中草药饲料添加剂,是我国北方地区常见的中草药之一,刺五加根茎和果实中含有丰富的皂苷类物质。所述新用途为所述刺五加皂苷B用于动物抗冷应激。所述刺五加皂苷B用于动物抗冷应激的时间为动物冷应激条件下12-15天,即在动物处于冷应激条件下的第12-15天为动物注射刺五加皂苷B,本实施例中,最佳地,所述刺五加皂苷B用于动物抗冷应激的时间为动物冷应激条件下第14天。所述刺五加皂苷B用于动物抗冷应激的注射浓度为3-54mg/kg,本实施例中,最佳地,刺五加皂苷B用于动物抗冷应激的注射浓度为4mg/kg;即,根据动物的重量来计算注射浓度,本实施例所提供的注射时间和浓度能够使得动物处于最佳抗冷应激的状态,即在动物抗冷应激的第14天,以4mg/kg的注射浓度对动物进行注射,可达到最佳的效果。
本实施例中采用刺五加皂苷B,不仅有利于动物抗冷应激,而且能够提高幼龄动物免疫能力,增强动物机体抗氧化性、增强机体抵抗自由基的效果。
下丘脑是动物体温调节的基本中心,可以在抗冷应激机制中发挥重要作用。Hsp70是用于检测冷应激的可靠且灵敏的分子标记物。因此本实施例提供通过实验检测小鼠下丘脑中Hsp70蛋白表达量来确定刺五加皂苷B的最适宜注射浓度和最佳作用时间;
所述Hsp70蛋白表达量的测定方法为:
(1)动物饲养
试验鼠购自长春市亿斯实验动物技术有限公司(中国吉林长春),按照〖实验动物饲养管理条例〗进行饲养,自由采食饮水,低温(10±1)℃;常温(21±1)℃,每天清理鼠笼,保持干净清洁,定时通风。
使用不同浓度的刺五加皂苷B分别在饲养1、7、14、21d之后腹腔注射小鼠,浓度为0、2、4、8mg/kg,每组6个重复,每个重复一只;试验结束后采集血液、下丘脑样品,
(2)取实验动物下丘脑组织样品,加入预冷的pH为7.4的磷酸缓冲液清洗,用液氮研磨后放到EP管中,低温离心20min,3000r/min,吸取上清液,分装后待检测;
(3)研磨组织的同时取出4℃冰箱中的试剂盒,并在室温下平衡30分钟;
(4)标准品的稀释:取5个EP管,向每个EP管中加入150μL标准品稀释液,标记为1、2、3、4、5;在标记5的管中加入150μL标准品,轻轻震荡混匀后,取出150μL,加入到下一管中,按照此方法依次倍比稀释;
(5)加样:每个样品做三个重复,空白孔中只加显色剂和终止液;将标准孔依次加入稀释的标准品50μL,链霉素-HRP 50Ml(由于抗体已经放入标准品中,所以不加);向样品中分别加入40μL样品、50μL链霉亲和素-HRP和10μL抗-Hsp70抗体;加样结束后,盖上封板膜,轻轻摇动使其混匀,放在37℃恒温箱中,孵育60分钟;
(6)洗涤:先用蒸馏水稀释浓缩洗涤液,备用;温育结束后,弃去残留液体;加入洗涤液,甩干,拍板,依次重复5次;
(7)显色:分别向每个孔的空间中加入50μL显色剂A和B,轻轻摇动并混合,放在37℃恒温箱中,避光显色10分钟;
(8)终止:向每个孔加入50μL终止溶液以终止反应;
(9)测定:10分钟内,用分光光度计在450nm的检测波长下测量吸光度(OD值);
(10)计算:用标准品的浓度和OD值计算出线性回归方程,然后根据测定出的样品OD值,计算其相应的样品浓度。
图1为小鼠冷应激后第14天时不同浓度刺五加皂苷B下丘脑中Hsp70蛋白的相对表达量,图2为冷应激0d、1d、7d、14d、21d时,注射4mg/kg刺五加皂苷B下丘脑中蛋白的相对表达。由图1可知,在14d小鼠下丘脑中Hsp70蛋白含量随着刺五加皂苷B浓度的增加,呈先升高后降低的趋势。4mg/kg皂苷组中Hsp70蛋白的相对表达量显著高于0mg/kg、2mg/kg和8mg/kg组(p<0.05)。图2所示,随着冷应激时间的增加,小鼠下丘脑中Hsp70蛋白含量先升高后降低,14d时,Hsp70蛋白相对表达量显著高于其他组(p<0.05),说明此时机体处于冷应激状态,因此选用4mg/kg和14d为刺五加皂苷B的最佳刺激浓度和时间。
以上所述仅为本发明的优选实施例而已,并不用于限制本发明,对于本领域的技术人员来说,本发明可以有各种更改和变化。凡在本发明的精神和原则之内,所作的任何修改、等同替换、改进等,均应包含在本发明的保护范围之内。
Claims (6)
1.一种刺五加皂苷B的新用途,其特征在于,所述新用途为所述刺五加皂苷B作为唯一活性成分用于制备动物抗冷应激药物;所述刺五加皂苷B用于动物抗冷应激的时间为动物冷应激条件下第14天。
2.根据权利要求1所述的新用途,其特征在于,所述刺五加皂苷B用于动物抗冷应激的注射浓度为3-5mg/kg。
3.根据权利要求2所述的新用途,其特征在于,所述刺五加皂苷B用于动物抗冷应激的注射浓度为4mg/kg。
4.根据权利要求1所述的新用途,其特征在于,所述刺五加皂苷B对所述动物下丘脑抗冷应激作用的影响通过测定注射有刺五加皂苷B的动物下丘脑中的Hsp70蛋白表达量来反应。
5.根据权利要求4所述的新用途,其特征在于,所述Hsp70蛋白表达量的测定方法中采用的实验动物为小鼠。
6.根据权利要求5所述的新用途,其特征在于,采用Hsp70蛋白表达量的测定方法之前还包括步骤:使用不同浓度的刺五加皂苷B分别在饲养1、7、14、21d之后腹腔注射小鼠,浓度为0、2、4、8mg/kg,每组6个重复,每个重复一只。
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