CN110419660A - 一种野樱莓酵素及其制备方法 - Google Patents
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Abstract
本发明公开了一种野樱莓酵素及其制备方法。它是以新鲜野樱莓为原料,经枯草芽孢杆菌和/或乳酸菌发酵制备而成。本发明制备的野樱莓酵素,以枯草芽孢杆菌和/或乳酸菌作为发酵菌株,发酵产物中GABA的含量大幅提升,具有显著的解酒作用,为开发解酒的天然饮品及药品奠定了基础。
Description
技术领域
本发明属于食品级酵素制备技术领域,具体涉及一种野樱莓酵素及其制备方法。
背景技术
野樱莓原产于北美洲东部,最初接触它的是美洲土著印第安人。约14世纪左右,他们将野樱莓浆果作为健康的食物长期持久的食用,并且也将它制成能帮助治疗感冒的饮用原料。野樱莓果实及其提取物对心脑血管疾病具有特殊的疗效,花青素和黄酮还有助于保持尿道健康;极高的维生素含量,具有抗衰老功能,但是,野樱莓果实口感酸涩,不适合直接食用。野樱莓被称为“花青素之王”,野樱莓每100g含有1480mg花青素、664mg原花青素、1752mg多酚类物质,总抗氧化剂超过2000mg,ORAC抗氧化指数达到160.6umol TE/g。也正是因为丰富的抗氧化物质,使得野樱莓系列的产品广受消费者,尤其是女性消费者的青睐,在欧洲被称为“超级食品”,在亚洲有“口服化妆品”的美称。
中国于上世纪九十年代开始引种,经过十多年的研究,种植育苗技术已经成熟。目前国内总种植面积约6万亩,果实年产量约4500吨。2018年10月,野樱莓在中国通过审批,意味着野樱莓产品可以在国内正式生产销售。目前国内自主研发生产的野樱莓产品非常少,尚属萌芽状态,市场一片空白。
发明内容
本发明的目的在于提供一种野樱莓酵素及其制备方法。
发明人在努力开发野樱莓加工产品的时候,发现野樱莓的乳酸菌发酵产物中GABA的含量大幅提升,而GABA与解酒相关,发明人在此基础上拓展,发现野樱莓的枯草芽孢杆菌发酵产物中GABA的含量也大幅提升,而其他发酵菌如啤酒酵母等,则不能提升发酵产物中GABA的含量,因此,选定枯草芽孢杆菌和乳酸菌进行发酵实验。
一种野樱莓酵素,其是以新鲜野樱莓为原料,经微生物发酵制备而成。
所述微生物为枯草芽孢杆菌和/或乳酸菌。
所述野樱莓酵素的制备方法,按照如下步骤进行:
(1)按照重量份数,取新鲜的野樱莓100-200份,加入水100-250份,打浆,加入葡萄糖15-25份,磷酸二氢钾5-10份,混合均匀;
(2)将步骤(1)制备的混合物料置于高压容器中,121℃灭菌30-50min,待灭菌锅降温到50-70℃时取出;
(3)接入微生物菌液,将物料放入37℃摇床中,在100-200r/min条件下摇6-12d;
(4)将发酵好的物料用脱脂棉网过滤,再用抽滤漏斗抽滤,进一步去除滤渣得到微生物酵素;将到微生物酵素置于容器中,密封后灭菌121℃,20-40min,置于4℃下保存。
所述微生物菌液为枯草芽孢杆菌菌液和/或乳酸菌菌液。
所述微生物菌液的接种量为接种物料的0.05%-0.5%,微生物菌液中活菌数为1-3X108cfu/mL。
所述野樱莓酵素在制备饮料中的应用。
所述野樱莓酵素在制备解酒饮品或药品中的应用。
本发明的有益效果:本发明制备的野樱莓酵素,以枯草芽孢杆菌和/或乳酸菌作为发酵菌株,发酵产物中GABA的含量大幅提升,具有显著的解酒作用,为开发解酒的天然饮品及药品奠定了基础。
具体实施方式
下面结合具体实施例对本发明做进一步说明。
实施例1枯草芽孢杆菌发酵野樱莓酵素的制备
一种野樱莓酵素的制备方法,按照如下步骤进行:
(1)按照重量份数,取新鲜的野樱莓16kg,加入水20kg,打浆,加入葡萄糖2.0kg,磷酸二氢钾0.6kg,混合均匀;
(2)将步骤(1)制备的混合物料置于高压容器中,121℃灭菌45min,待灭菌锅降温到60℃时取出;
(3)接入枯草芽孢杆菌菌液,将物料放入37℃摇床中,在160r/min条件下摇8d;所述枯草芽孢杆菌菌液的接种量为接种物料的0.2%,枯草芽孢杆菌菌液中活菌数为2X108cfu/mL;
(4)将发酵好的物料用脱脂棉网过滤,再用抽滤漏斗抽滤,进一步去除滤渣得到微生物酵素;将到微生物酵素置于容器中,密封后灭菌121℃,30min,置于4℃下保存。
实施例2乳酸菌发酵野樱莓酵素的制备
一种野樱莓酵素的制备方法,按照如下步骤进行:
(1)按照重量份数,取新鲜的野樱莓16kg,加入水20kg,打浆,加入葡萄糖2.0kg,磷酸二氢钾0.6kg,混合均匀;
(2)将步骤(1)制备的混合物料置于高压容器中,121℃灭菌45min,待灭菌锅降温到60℃时取出;
(3)接入乳酸菌菌液,将物料放入37℃摇床中,在160r/min条件下摇8d;所述乳酸菌菌液的接种量为接种物料的0.2%,乳酸菌菌液中活菌数为 2X108cfu/mL;
(4)将发酵好的物料用脱脂棉网过滤,再用抽滤漏斗抽滤,进一步去除滤渣得到微生物酵素;将到微生物酵素置于容器中,密封后灭菌121℃,30min,置于4℃下保存。
实施例3混合发酵野樱莓酵素的制备
一种野樱莓酵素的制备方法,按照如下步骤进行:
(1)按照重量份数,取新鲜的野樱莓16kg,加入水20kg,打浆,加入葡萄糖2.0kg,磷酸二氢钾0.6kg,混合均匀;
(2)将步骤(1)制备的混合物料置于高压容器中,121℃灭菌45min,待灭菌锅降温到60℃时取出;
(3)接入枯草芽孢杆菌菌液和乳酸菌菌液,将物料放入37℃摇床中,在 150r/min条件下摇8d;所述枯草芽孢杆菌菌液的接种量为接种物料的0.1%,枯草芽孢杆菌菌液中活菌数为1X108cfu/mL;所述乳酸菌菌液的接种量为接种物料的0.1%,乳酸菌菌液中活菌数为1X108cfu/mL;
(4)将发酵好的物料用脱脂棉网过滤,再用抽滤漏斗抽滤,进一步去除滤渣得到微生物酵素;将到微生物酵素置于容器中,密封后灭菌121℃,30min,置于4℃下保存。
实验例1:样品中GABA(γ-氨基丁酸)的测定
采用高效液相色谱法,对照实验样品为野樱莓打浆后的过滤液;测试步骤如下:
(1)衍生试剂的配制:精确称取OPA 0.1g、β-巯基乙醇130μL溶解在1mL乙腈和0.5mL甲醇中,然后加入用0.5M硼酸溶液定容至20mL;
(2)硼酸缓冲液的配制:取硼酸0.62g和KOH 0.52g溶于20mL蒸馏水中;
(3)流动相A(乙酸钠缓冲液)的配制:取无水乙酸钠8.0g溶于1000m L双蒸水,加入三乙胺220μL,用5%冰醋酸调pH为7.2,再加入5mL的四氢呋喃,混匀后过0.22um过滤膜;
(4)流动相B的配制:取8.0g乙酸钠,用双蒸水定容至1000mL,然后用2%的乙酸溶液调节pH为7.2,再按乙酸钠溶液:乙腈:甲醇=1:2:2(体积比) 混合后过0.22um针孔过滤膜;
(5)样品的衍生:吸取衍生试剂和预处理后的发酵液按照1:1的比例混合,室温反应5min,取20μL进样;
(6)色谱分析:采用C18色谱柱(4.6mm×250mm),A、B混合流动相流速为1.0mL/min,柱温30℃,检测波长338nm,梯度洗脱;
(7)标准曲线的绘制与样品分析:精确称量GABA标准品10mg,用去离子水定容至100mL,然后精确量取5、10、20、30、40、50μL的标准品溶液,分别加入495、490、480、470、450mL的双蒸水,衍生测定,然后按照峰面积和GABA浓度绘制标准曲线;
GABA浓度(x)在0-5.0g/L的范围内与峰面积(y)的线性方程为: y=519667.23x-18125.35,R2=0.9975。
按照标准曲线方程根据样品的峰面积计算样品中的GABA浓度,测试结果见表1:
表1
注:*代表与实施例1组比较P<0.05;**代表P<0.01。
由表1可以看出,实施例1-2制备的野樱莓酵素中GABA含量大于未发酵的原料产品(对照组,P<0.05),实施例3采用枯草芽孢杆菌和乳酸菌联合发酵, GABA含量显著增加。
实验例2:小鼠解酒实验
实验用衡水老白干(67度):购自永辉超市;酒无罪药物,购自幸福人药房,其主要成分为葛根;ICR小鼠:6-8周龄,雄性标重,购自凯学生物科技 (上海)有限公司。
随机选取50只雄性小鼠,于当日20:00禁食,次日8:00停止,然后每组分别按照0.1、0.15、0.2、0.25、0.3mL/10g的剂量对选取的50只雄性小鼠灌胃饮用酒,同时观察灌酒小鼠的醉酒数量、醉酒时间及死亡数量。小鼠是否醉酒以小鼠的翻正反射消失作为判断指标。
饮用酒量的选择试验结果显示以0.1、0.15、0.2、0.25、0.3mL/10g分别给小鼠灌胃饮用白酒,0.2mL/10g及其以上剂量组即有小鼠发生急性酒精中毒死亡,而0.15mL/10g剂量既有翻正反射消失,使小鼠呈现醉酒状态(翻正反射消失)而又无小鼠死亡,所以本试验选用0.15mL/10g饮用酒量。
先对小鼠灌胃饮用酒,经过5min后再对小鼠灌胃实施例1,实施例2及对比例1的苦荞麦酵素。取60只雄性小鼠,随机分为5组,每组12只,5 组分别命名为:空白对照组、阳性对照组、实施例1组、实施例2组、对比例 1组。在试验前先对小鼠禁食12h,然后按照0.15mL/10g的剂量分别给空白对照组、阳性对照组、实施例1组、实施例2组、对比例1组灌胃饮用酒。5组试验组要求均在灌胃饮用酒5min后按照0.15mL/10g剂量灌胃样品和药物,然后记录5组小鼠翻正反射消失时间(醉酒潜伏时间min)和翻正反射恢复时间(醒酒时间min)。测试结果见表2:
表2
注:*代表与实施例1组比较P<0.05。
由表2可以看出,实施例1组的小鼠相对于空白对照组醉酒潜伏时间明显延长,醒酒时间显著缩短,接近阳性对照组,实施例2组的小鼠与实施例1组醉酒潜伏时间相当,实施例3组小鼠相对于实施例1组,醉酒潜伏时间明显延长,醒酒时间显著缩短,证明联合发酵的产品相对于单一发酵有明显的协同增效作用。
Claims (7)
1.一种野樱莓酵素,其特征在于,其是以新鲜野樱莓为原料,经微生物发酵制备而成。
2.根据权利要求1所述野樱莓酵素,其特征在于,所述微生物为枯草芽孢杆菌和/或乳酸菌。
3.权利要求1所述野樱莓酵素的制备方法,其特征在于,按照如下步骤进行:
(1)按照重量份数,取新鲜的野樱莓100-200份,加入水100-250份,打浆,加入葡萄糖15-25份,磷酸二氢钾5-10份,混合均匀;
(2)将步骤(1)制备的混合物料置于高压容器中,121℃灭菌30-50min,待灭菌锅降温到50-70℃时取出;
(3)接入微生物菌液,将物料放入37℃摇床中,在100-200r/min条件下摇6-12d;
(4)将发酵好的物料用脱脂棉网过滤,再用抽滤漏斗抽滤,进一步去除滤渣得到微生物酵素;将到微生物酵素置于容器中,密封后灭菌121℃,20-40min,置于4℃下保存。
4.根据权利要求3所述野樱莓酵素的制备方法,其特征在于,所述微生物菌液为枯草芽孢杆菌菌液和/或乳酸菌菌液。
5.根据权利要求3所述野樱莓酵素的制备方法,其特征在于,所述微生物菌液的接种量为接种物料的0.05%-0.5%,微生物菌液中活菌数为1-3X108cfu/mL。
6.权利要求1所述野樱莓酵素在制备饮料中的应用。
7.权利要求1所述野樱莓酵素在制备解酒饮品或药品中的应用。
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