CN108434369A - 治疗鸡大肠杆菌病的益生菌复方中药发酵液 - Google Patents
治疗鸡大肠杆菌病的益生菌复方中药发酵液 Download PDFInfo
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Abstract
本发明公开了一种治疗鸡大肠杆菌病的益生菌复方中药发酵液,它是由下述方法制备的:1)称取黄芪、黄连、乌梅、白头翁、茯苓、黄芩、金银花、大青叶、葛根、千里光、神曲、甘草、山楂、泽泻、诃子、明矾、黄柏、麦芽,加水,加热30~50min,过滤,收集滤液,浓缩,即为水提中药液;2)保加利亚乳杆菌与植物乳杆菌活化,按1:1~6接种在MRS液体培养基,36~38℃下恒温培养20~30h,得到复方益生菌制剂;3)将步骤2)的复方益生菌制剂接种到步骤1)的水提中药液中,接种量为中药液体积分数的3~5%,发酵,得到治疗鸡大肠杆菌病的益生菌复方中药发酵液;本发明发酵液的抑菌效果好,能维护肠道微生态平衡、提高机体免疫力,并具有有抑菌消炎的效果。
Description
技术领域
本发明属于鸡大肠杆菌病的防治技术领域,具体涉及治疗鸡大肠杆菌病的益生菌复方中药发酵液。
背景技术
鸡大肠杆菌病是我国临床常见的鸡病之一,是由致病性大肠杆菌引起的。具有发病快、死亡率高、病程长的特点,其主要的病型有胚胎和幼雏的死亡、败血症、气囊炎、心包炎、输卵管炎、肠炎、腹膜炎和大肠杆菌性肉芽肿等。由于常和霉形体病合并感染,又常继发于其他传染病(如新城疫、禽流感惑、传染性支气管炎、巴氏杆菌病等),使治疗十分困难。目前本病已成为危害养业的重要传染病,常造成巨大的经济损失。
目前对于大肠杆菌病多采用抗菌药物治疗,但由于此类药物的过量使用,已导致大肠杆菌的耐药谱不断扩大,防治效果下降,同时抗菌药物在机体内的残留,易引起菌群失调,降低了肉蛋产品品质,影响了人类健康和公共卫生安全。中草药具有天然、使用安全、无毒副作用、无化学物质残留、不易产生抗药性等特点,但是存在疗效慢的特点,目前市售的用于治疗鸡大肠杆菌病的中草药制剂虽然种类很多,但实际使用效果不尽人意;中药发展至今,加工方法过于简单,产品始终难以摆脱粗、大、黑的特点,尤其是散剂,保留了中药材的有效和无效部位,且药物粒径大,粗纤维含量高,病畜尤其是家禽难以消化和吸收,致使中药材中抗菌或抗病毒的有效成分难以充分发挥疗效,同时也造成了中药材宝贵资源的大量浪费。因此开发一种含有益生菌的中药符合制剂可弥补以上药物的缺陷。
发明内容
本发明目的是提供一种取材容易、制备方便、减少药物残留及毒副作用、价格低廉疗效较高的治疗鸡大肠杆菌病的益生菌复方中药发酵液。
治疗鸡大肠杆菌病的益生菌复方中药发酵液,它是由下述方法制备的:
1)称取黄芪、黄连、乌梅、白头翁、茯苓、黄芩、金银花、大青叶、葛根、千里光、神曲、甘草、山楂、泽泻、诃子、明矾、黄柏、麦芽,加水,加热30~50min,过滤,收集滤液,浓缩,即为水提中药液;
2)保加利亚乳杆菌与植物乳杆菌活化,按1:1~6接种在MRS液体培养基,36~38℃下恒温培养20~30h,得到复方益生菌制剂;
3)将步骤2)的复方益生菌制剂接种到步骤1)的水提中药液中,接种量为中药液体积分数的3~5%,发酵,得到治疗鸡大肠杆菌病的益生菌复方中药发酵液;
步骤1)中所述的黄芪10~30份、黄连5~10份、乌梅6~15份、白头翁13~50份、茯苓13~50份、黄芩15~25份、金银花10~30份、大青叶5~15份、葛根13~50份、千里光15~25份、神曲10~20份、甘草6~30份、山楂10~20份、泽泻5~10份、诃子5~15份、明矾5~10份、黄柏5~20份、麦芽15~50份;
所述的黄芪20份、黄连10份、乌梅10份、白头翁30份、茯苓30份、黄芩20份、金银花20份、大青叶10份、葛根30份、千里光20份、神曲15份、甘草20份、山楂15份、泽泻10份、诃子10份、明矾10份、黄柏10份、麦芽25份;
步骤1)中所述的浓缩至每毫升滤液中0.4~0.5g生药;
步骤2)中所述的接种比例为1:4;
步骤3)中所述的发酵在发酵罐中进行,转速100~300rpm、时间36~60h、温度36~38℃;
所述的转速200rpm、时间48h、温度37℃;
步骤3)中所述的接种量为4%。
发明详述:
葛根具有发表解肌,生津止渴,升阳止泻,使表解里和;白头翁具有清热解毒,凉血止痢,对肠粘膜有收敛作用,故能止泻,是治疗湿热泄泻的要药;黄连具有清热燥湿,肠黄作泻,热痢后重,尤以肠胃湿热之证为宜;黄芩具有清热燥湿,湿热泻痢;黄柏具有湿热泄泻,治疗泻痢;金银花具有清热解毒,治热毒泻痢,常与黄芩为伍治疗热毒泄泻,兼有宣散作用;神曲具有消食化积,健胃和中,治疗脾虚泄泻;茯苓具有脾虚湿困,水湿停滞,既能健脾又能利湿,能补能泻;乌梅具有涩肠止泻,用于久泻久痢;诃子具有涩肠止泻,用于久泻久痢;泽泻具有能利水渗湿,治疗泻痢不止;黄芪具有补气,适用于脾肺气虚,泄泻;大青叶具有清热凉血,胃热毒;千里光具有清热、解毒,菌痢,泄泻;甘草具有性温,补脾胃,治脾胃虚弱,还具有调和诸药的作用,许多配方配伍本品;山楂具有消食健胃,治疗食积不消,止泻;麦芽具有消食和中,治疗脾胃虚弱;明矾具有收敛止血,可用于久泻不止,刺激胃黏膜引起反射性呕吐,制止肠黏膜分泌,因而具有止泻效果。
本发明提供的一种治疗鸡大肠杆菌病中药组合物,以清热、凉血、解毒,涩肠止泻,久泻久痢,调和诸药,兼调肠胃湿热,表里兼治;本发明中诸药合用共成清热止泻之剂,药物经有机组合,辩证配伍和相互辅佐,增强了该方清热燥湿止泻的功效,从而达到治愈鸡大肠杆菌病的目的;所述的中药组合物被保加利亚乳杆菌、植物乳杆菌发酵,产生的发酵液,能维护肠道微生态平衡、提高机体免疫力,并具有有抑菌消炎的效果;在选用本发明治疗鸡的大肠杆菌病时,抓住“湿、热、止泻、久痢”四点,所谓“热”主要表现为发热、拉白色稀粪便,气味恶臭难闻,并且病鸡精神沉郁、眼半睁半闭,喜欢趴着,个别鸡卧地不起,站不起来,舌苔发黄,脉滑而数,食欲减退等,如果符合上面这些症状,则可以认为是“湿热”所致的腹泻;应用要点:拉白稀、兼湿热、舌苔发白、脉数。
本发明提供了一种治疗鸡大肠杆菌病的益生菌复方中药发酵液,它是由下述方法制备的:1)称取黄芪、黄连、乌梅、白头翁、茯苓、黄芩、金银花、大青叶、葛根、千里光、神曲、甘草、山楂、泽泻、诃子、明矾、黄柏、麦芽,加水,加热30~50min,过滤,收集滤液,浓缩,即为水提中药液;2)保加利亚乳杆菌与植物乳杆菌活化,按1:1~6接种在MRS液体培养基,36~38℃下恒温培养20~30h,得到复方益生菌制剂;3)将步骤2)的复方益生菌制剂接种到步骤1)的水提中药液中,接种量为中药液体积分数的3~5%,发酵,得到治疗鸡大肠杆菌病的益生菌复方中药发酵液;本发明发酵液的抑菌效果好,能维护肠道微生态平衡、提高机体免疫力,并具有抑菌消炎的效果。
附图说明
图1 保加利亚乳杆菌、植物乳杆菌复方制剂生长曲线。
具体实施方式
实施例1 复方益生菌制剂配比优化工艺
将保加利亚乳杆菌、植物乳杆菌分别活化:从-80℃冰箱取出保加利亚乳杆菌、植物乳杆菌菌种,用手将离心管预热将其菌种溶解,然后在超净工作台内取出溶解的两种菌种100µL,分别接种于含5mLMRS液体培养基的两个试管内,37℃培养12h,然后在培养后的试管各取出100µL菌液,分别加入含5mLMRS液体培养基的两个试管,在37℃培养12h,然后放入4℃冰箱备用。
按保加利亚乳杆菌与植物乳杆菌比例分别为1:1、1:2、2:1接种在MRS液体培养基(乳酸细菌培养基)上,37℃培养箱中恒温培养12h,培养结束后用分光光度计于600nm测定OD值,结果显示1:2时 OD值最大;将保加利亚乳杆菌与植物乳杆菌分别按1:2、1:3、1:4、1:5、1:6比例,37℃培养箱中恒温培养24h,培养结束后用分光光度计于600nm测定OD值,比较大小,结果显示1:4时OD值最大,因此将保加利亚乳杆菌与植物乳杆菌1:4比例作为复方益生菌制剂的最佳比例;保加利亚乳杆菌与植物乳杆菌1:4时培养生长曲线见图1。
实施例2 治疗鸡大肠杆菌病的中药液的制备
称量黄芪20g、黄连10g、乌梅10g、白头翁30g、茯苓30g、黄芩20g、金银花20g、大青叶10g、葛根30g、千里光20g、神曲15g、甘草20g、山楂15g、泽泻10g、诃子10g、明矾10g、黄柏10g、麦芽25g,共315g,装入1000mL烧杯,浸泡30min,加4倍量即1260mL水(4倍是指中药组合物重量的倍数),倒入煎药壶中,加热40min,2层纱布过滤,收集滤液倒入指定的瓶中;药渣再加入2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入刚才的滤液瓶中;剩下的药渣再加2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入上两次的瓶中,中药液共计700mL,相当于每毫升滤液中生药量为0.45g。
实施例3治疗鸡大肠杆菌病的中药液的制备
称量黄芪10g、黄连10g、乌梅15g、白头翁20g、茯苓50g、黄芩15g、金银花10g、大青叶15g、葛根20g、千里光20g、神曲20g、甘草10g、山楂10g、泽泻10g、诃子5g、明矾5g、黄柏20g、麦芽40g,共315g,装入1000mL烧杯,浸泡30min,加4倍量即1260mL水(4倍是指中药组合物重量的倍数),倒入煎药壶中,加热40min,2层纱布过滤,收集滤液倒入指定的瓶中;药渣再加入2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入刚才的滤液瓶中;剩下的药渣再加2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入上两次的瓶中,中药液共计700mL,相当于每毫升滤液中生药量为0.45g。
实施例4治疗鸡大肠杆菌病的中药液的制备
称量黄芪30g、黄连5g、乌梅6g、白头翁50g、茯苓20g、黄芩25g、金银花30g、大青叶5g、葛根50g、千里光20g、神曲10g、甘草6g、山楂20g、泽泻5g、诃子8g、明矾5g、黄柏5g、麦芽15g,共315g,装入1000mL烧杯,浸泡30min,加4倍量即1260mL水(4倍是指中药组合物重量的倍数),倒入煎药壶中,加热40min,2层纱布过滤,收集滤液倒入指定的瓶中;药渣再加入2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入刚才的滤液瓶中;剩下的药渣再加2倍量蒸馏水,在煎药壶中加热30min,2层纱布过滤,滤液倒入上两次的瓶中,中药液共计700mL,相当于每毫升滤液中生药量为0.45g。
实施例5 治疗鸡大肠杆菌病的益生菌复方中药发酵液的制备
复方益生菌制剂制备方法按实施例1制备,水提中药液按实施例2制备。
本实验采取韩国生产的2.5L发酵罐进行益生菌发酵,参数有:温度、转速、复方益生菌接种量、中药量、发酵时间,还有pH、通气量不在设定范围之内。通过发酵罐进行发酵,参数设定为:转速200rpm、时间48h、温度37℃、复方益生菌制剂接种量20mL、水提中药液480mL,进行发酵,得到治疗鸡大肠杆菌病的益生菌复方中药发酵液。
实施例6 治疗鸡大肠杆菌病的益生菌复方中药发酵液的效果验证
鸡大肠杆菌病,病变在肠,其病性湿热证,其临床特征表现排白色新粪便,治疗的关键要清热燥湿,恢复小肠的正常传导功能。
选50只体型、体重相当的健康鸡,随机分成5组,其中一组作为对照组;其余4组作为实验组,通过人工复制鸡大肠杆菌病,在实验动物出现大肠杆菌病症状后,分别喂饲中药(实施例2制备的治疗鸡大肠杆菌病的中药液)、西药(环丙沙星)、实施例3制备的益生菌复方中药发酵液、蒸馏水,作为中药组、西药组、发酵液组、感染组;治疗7天后,分别测定5组实验动物的盲肠SIgA(结果见表1)、十二指肠、盲肠病理组织切片、血清抗氧化指标(丙二醛、谷胱甘肽过氧化物酶、超氧化物歧化酶);盲肠SIgA结果见表1,丙二醛、谷胱甘肽过氧化物酶、超氧化物歧化酶结果分别见表2、3、4;表1~4中,同列肩标小写字母相同者差异不显著(P>0.05)、不同者差异显著(P<0.05),大写字母不同者差异极显著(P<0.01);
结果显示,发酵液的抑菌效果高于复方中药的抑菌效果,但是略低于西药组治疗效果;综合药物毒副作用等因素,益生菌复方中药发酵液更优。
所述的发酵液的服用方法:
1)药量:发酵液和水放在一起,自由饮水服用;
2)服药时间:每天上午一次,中午一次,晚上一次。
3)药温控制:温服;
4)禁忌事项:发酵液治疗期间,加强对鸡的饲养管理,保证充足的饮水,定时定点饲喂颗粒料。
在一个月的人工复制和治疗过程中,治疗1周,治愈8只,好转1只,治愈率84.3%,有效率100%;
疗效判定标准:主要依据鸡腹泻症状是否消失,服药7天为一疗程;
具体疗效分为三级:治愈-鸡大肠杆菌病拉白稀症状完全消失;好转-鸡大肠杆菌病拉白稀症状部分消失;无效-鸡的大肠杆菌病拉白稀症状毫无消失。
Claims (7)
1.治疗鸡大肠杆菌病的益生菌复方中药发酵液,它是由下述方法制备的:
1)称取黄芪、黄连、乌梅、白头翁、茯苓、黄芩、金银花、大青叶、葛根、千里光、神曲、甘草、山楂、泽泻、诃子、明矾、黄柏、麦芽,加水,加热30~50min,过滤,收集滤液,浓缩,即为水提中药液;
2)活化的保加利亚乳杆菌和植物乳杆菌按1:1~6接种在MRS液体培养基,36~38℃下恒温培养20~30h,得到复方益生菌制剂;
3)将步骤2)的复方益生菌制剂接种到步骤1)的水提中药液中,接种量为中药液体积分数的3~5%,发酵,得到治疗鸡大肠杆菌病的益生菌复方中药发酵液;
步骤1)中所述的黄芪10~30份、黄连5~10份、乌梅6~15份、白头翁13~50份、茯苓13~50份、黄芩15~25份、金银花10~30份、大青叶5~15份、葛根13~50份、千里光15~25份、神曲10~20份、甘草6~30份、山楂10~20份、泽泻5~10份、诃子5~15份、明矾5~10份、黄柏5~20份、麦芽15~50份。
2.根据权利要求2所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:所述的黄芪20份、黄连10份、乌梅10份、白头翁30份、茯苓30份、黄芩20份、金银花20份、大青叶10份、葛根30份、千里光20份、神曲15份、甘草20份、山楂15份、泽泻10份、诃子10份、明矾10份、黄柏10份、麦芽25份。
3.根据权利要求1、或2所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:步骤1)中所述的浓缩至每毫升滤液中0.4~0.5g生药。
4.根据权利要求3所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:步骤2)中所述的接种比例为1:4。
5.根据权利要求4所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:步骤3)中所述的发酵在发酵罐中进行,转速100~300rpm、时间36~60h、温度36~38℃。
6.根据权利要求5所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:所述的转速200rpm、时间48h、温度37℃。
7.根据权利要求6所述的治疗鸡大肠杆菌病的益生菌复方中药发酵液,其特征在于:步骤3)中所述的接种量为4%。
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