CN108514123A - 从毛蚶提取的活性多肽制备复配物方法 - Google Patents
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Abstract
本发明公开一种从毛蚶提取的活性多肽制备复配物方法,包括毛蚶活性多肽的提取,复配物的制备,冷却保存,复配物的制备步骤为:按重量份计取粉碎后的盐蒿粉末和纤维素酶,加入蒸馏水并调pH值至5.5~6.0,于45~50℃下酶解30‑35min,过滤取酶解液,加入无水乙醇和抗氧化剂搅拌20‑25min,加热干燥得浸膏,备用,将牛奶、维生素A、白糖、葡萄汁、蜂蜜、毛蚶活性多肽和柠檬汁溶解得基液,将浸膏加入2‑2.3倍体积的水加热溶解并加入浸膏质量45~48%的基液,得复配物,制备的复配物具备独特的风味,有益于口腔健康,在保存过程及开瓶后营养物质不易氧化,具有预防肿瘤和调节人体免疫的作用。
Description
技术领域
本发明属于饮料制备技术领域,具体涉及一种从毛蚶提取的活性多肽制备复配物方法。
背景技术
毛蚶( Arca subcrenata Lischke) ,软体动物门,瓣鳃纲,蚶目,蚶科,毛蚶属,主要分布于中国、朝鲜和日本沿海。毛蚶的营养与药用价值较高,其壳和肉皆有药用价值。中医认为其性温味甘,归脾、肝经,具温中健胃、滋阴补血之功效,临床上主治贫血、体虚等,毛蚶多肽提取物能够显著抑制多株肿瘤细胞的增殖,但是现有技术中对毛蚶活性多肽的提取率并不高并且其应用领域并不广泛。
饮料是人们重要的生活消费品,目前市场上,饮料数量和饮料种类丰富多彩,各具特色。然而,除包装饮用水之外,其他饮料,如碳酸饮料、果蔬汁饮料、茶饮料等,通常含糖量较高。研究表明,长期食用高含糖、尤其是高含蔗糖的食品、已成为诱发糖尿病、肥胖症等病症的病因。
现有技术提供了较多的保健饮料,例如中国发明授权专利文献,授权公告号CN103054035 B,该发明涉及一种保健饮料,公开了一种喉爽饮料及制备方法,该饮料以蒲公英提取物、菊花提取物、薄荷油、糖、糖醇、非糖甜味剂、酸味剂等原料,通过调配、热罐装、高温瞬时灭菌等加工工艺而得到。该发明可以减少饮料中糖的使用量,并且通过合理配伍,各种配料功效互补,从而协同发挥良好的润喉利咽、洁齿防龋等功能,但是该饮料在开瓶后可能会在短时间内饮料中的营养物质会氧化降低其保健功效。
发明内容
本发明的目的在于提供一种从毛蚶提取的活性多肽制备复配物方法,制备的复配物具备独特的风味,有益于口腔健康,在保存过程及开瓶后营养物质不易氧化,具有预防肿瘤和调节人体免疫的作用。
本发明为实现上述目的所采取的技术方案为:从毛蚶提取的活性多肽制备复配物方法,包括毛蚶活性多肽的提取,复配物的制备,冷却保存,复配物的制备步骤为:按重量份计取粉碎后的盐蒿粉末和纤维素酶,加入蒸馏水并调pH值至5.5~6.0,固液比为1g:10-17ml,于45~50℃下酶解30-35min,过滤取酶解液,加入无水乙醇和抗氧化剂搅拌20-25min,加热干燥得浸膏,备用,将牛奶、维生素A、白糖、葡萄汁、蜂蜜、毛蚶活性多肽和柠檬汁溶解得基液,将浸膏加入2-2.3倍体积的水加热溶解并加入浸膏质量45~48%的基液,得复配物。选用盐蒿为主原料使制备的复配物中的主要口感为盐蒿风味并且添加葡萄汁、蜂蜜等成分进一步丰富复配物的口感同时降低白糖的使用量使复配物老少皆宜,利用盐蒿粉末为主原料制备所得复配物具有很好的清热、消积、扩张血管、防止心脏病和增强人体免疫力的功效,即复配物具有很好的保健效果,在复配物制备过程中加入毛蚶活性多肽、维生素A、牛奶等成分使其协同发挥作用提高复配物的抗氧化、润喉、口腔保健的效果,并且进一步提升复配物的口感,增强其香味,同时遮蔽复配物可能存在的苦涩感,人们在长期饮用复配物的过程中,复配物可促进人体未活化的淋巴细胞增殖,起到预防肿瘤的效果。
优选的,复配物由以下成分及重量份组成:盐蒿粉末60-180份、纤维素酶1-3份、蒸馏水500-600份、无水乙醇20-24份、抗氧化剂0.06-0.12份、牛奶2-7份、维生素A0.01-0.03份、白糖15-30份、葡萄汁27-38份、蜂蜜45-50份、毛蚶活性多肽3-7份、柠檬汁27-30份。选用盐蒿为主原料使制备的复配物中的主要口感为盐蒿风味并且添加葡萄汁、蜂蜜等成分进一步丰富复配物的口感同时降低白糖的使用量使复配物老少皆宜,利用盐蒿粉末为主原料制备所得复配物具有很好的清热、消积、扩张血管、防止心脏病和增强人体免疫力的功效,在复配物中加入抗氧化剂可有效延长复配物中营养成分的有效时间,延长复配物的保存时间,当复配物被开瓶时,由于抗氧化剂的加入可有效降低复配物的氧化速度,保持复配物的效果。
优选的,抗氧化剂由3-5重量份的维生素C和0.1-0.6重量份的混合物组成,混合物为L-亮氨酰甘氨酸和D-亮氨酰甘氨酸,两者质量比为47~53:0.5-0.8。抗氧化剂可与O2 -·、HOO·及·OH快速反应,生成半脱氢抗坏血酸,清除单线态氧,还原硫自由基,还可使氧化的VE恢复成还原型,起到间接抗氧化的作用,并且通过加入的混合物对维生素C起到增效作用,混合物可与金属离子络合,钝化催化氧化作用,同时产生的氢离子可使抗氧化剂再生,实现永久性抗氧化的效果。
优选的,毛蚶活性多肽的提取步骤为:取新鲜毛蚶去壳,加入蒸馏水,匀浆,除杂,过滤,得滤液A,然后在滤液A中加入乙酸溶液搅拌提取,离心,取上清液,上清液中加入NaCl至溶液最终浓度为2.5mol/L,同时析出絮状沉淀,离心,得毛蚶活性多肽,通过上述方法所得的毛蚶活性多肽的纯度高,纯度在92%以上,提取率在78%~82.5%之间,提取的多肽分子量在3000~6000Da之间,体外抗肿瘤实验表明抑瘤率达到57~59%,对人体免疫力调节作用强,特别是在淋巴细胞增殖反应效果方面显著,可促进未活化的淋巴细胞增殖,还可以促进ConA活化T或B 淋巴细胞增殖。
优选的,蒸馏水中含有0.08-0.1wt%的D-苦杏仁酸和L-苦杏仁酸的混合体,D-苦杏仁酸和L-苦杏仁酸重量比为52~57:0.3~0.9,蒸馏水加入D-苦杏仁酸和L-苦杏仁酸的混合体可提高毛蚶细胞组织的亲水性,使其溶于蒸馏水中并在细胞膜下形成小孔,促使细胞内的物质泄漏,实现提高毛蚶活性多肽的得率提高。
优选的,除杂操作为:将匀浆液分别用0.15-0.25mol/L的NaOH和0.1-0.15mol/L的EDTA-2Na溶液浸泡,过滤,得滤液A,实现脱除杂蛋白、色素和矿物质等成分,避免上述成分对毛蚶活性多肽提取的影响。
优选的,复配物冷却保存温度为2-5℃,通过低温保存有效保证毛蚶活性多肽的活性,利于活性多肽及复配物其余成分在人体肠胃中的吸收效果。
与现有技术相比,本发明的有益效果为:选用盐蒿为主原料使制备的复配物中的主要口感为盐蒿风味并且添加葡萄汁、蜂蜜等成分进一步丰富复配物的口感同时降低白糖的使用量使复配物老少皆宜,利用盐蒿粉末为主原料制备所得复配物具有很好的清热、消积、扩张血管、防止心脏病和增强人体免疫力的功效,即复配物具有很好的保健效果,在复配物制备过程中加入提取率很高的毛蚶活性多肽、维生素A、牛奶等成分使其协同发挥作用提高复配物的抗氧化、润喉、口腔保健的效果,并且进一步提升复配物的口感,增强其香味,同时遮蔽复配物可能存在的苦涩感,人们在长期饮用复配物的过程中,复配物可促进人体未活化的淋巴细胞增殖,起到预防肿瘤的效果。
具体实施方式
以下结合实施例对本发明做进一步详细描述:
实施例1:
从毛蚶提取的活性多肽制备复配物方法,包括毛蚶活性多肽的提取,复配物的制备,冷却保存,具体优选操作步骤如下:
毛蚶活性多肽的提取步骤为:取新鲜毛蚶去壳,加入蒸馏水,匀浆,匀浆固液比为1:1.3,蒸馏水中含有0.08wt%的D-苦杏仁酸和L-苦杏仁酸的混合体,D-苦杏仁酸和L-苦杏仁酸重量比为55:0.6,分别用0.17mol/L的NaOH和0.1mol/L的EDTA-2Na溶液浸泡,过滤,得滤液A,然后在滤液A中按固液比1:8加入0.12mol/L的乙酸溶液搅拌提取,搅拌时间为6h,搅拌速度为50r/min,于5000r/min条件下离心10min,取上清液,上清液中加入NaCl至溶液最终浓度为2.5mol/L,同时析出絮状沉淀,于3℃条件下9000r/min离心12min,得毛蚶活性多肽;
复配物的制备步骤为:按重量份计取粉碎后的盐蒿粉末120份和纤维素酶1份,加入蒸馏水540份并调pH值至5.8,固液比为1g:14ml,于48℃下酶解32min,过滤取酶解液,加入无水乙醇22份和抗氧化剂0.08份搅拌24min,加热干燥得浸膏,备用,将牛奶5份、维生素A0.02份、白糖16份、葡萄汁28份、蜂蜜47份、毛蚶活性多肽4份和柠檬汁28份溶解得基液,将浸膏加入2.2倍体积的水加热溶解并加入浸膏质量46%的基液,得复配物,抗氧化剂由4重量份的维生素C和0.4重量份的混合物组成,混合物为L-亮氨酰甘氨酸和D-亮氨酰甘氨酸,两者质量比为48:0.7;
高温瞬时灭菌:将复配物后装瓶于103℃条件下持续10s高温瞬时灭菌;
冷却及检验:检查瓶体有无破裂等异常现象,贴上标签后将复配物于4℃低温保存。
实施例2:
从毛蚶提取的活性多肽制备复配物方法,包括毛蚶活性多肽的提取,复配物的制备,冷却保存,具体操作步骤如下:
毛蚶活性多肽的提取步骤为:取新鲜毛蚶去壳,加入蒸馏水,匀浆,匀浆固液比为1:1.5,蒸馏水中含有0.09wt%的D-苦杏仁酸和L-苦杏仁酸的混合体,D-苦杏仁酸和L-苦杏仁酸重量比为52:0.7,分别用0.2mol/L的NaOH和0.13mol/L的EDTA-2Na溶液浸泡,过滤,得滤液A,然后在滤液A中按固液比1:10加入0.13mol/L的乙酸溶液搅拌提取,搅拌时间为7h,搅拌速度为45r/min,于5000r/min条件下离心11min,取上清液,上清液中加入NaCl至溶液最终浓度为2.5mol/L,同时析出絮状沉淀,于2℃条件下9000r/min离心12min,得毛蚶活性多肽;
复配物的制备步骤为:按重量份计取粉碎后的盐蒿粉末75份和纤维素酶1份,加入蒸馏水540份并调pH值至5.8,固液比为1g:14ml,于48℃下酶解32min,过滤取酶解液,加入无水乙醇22份和抗氧化剂0.09份搅拌22min,加热干燥得浸膏,备用,将牛奶7份、维生素A0.01份、白糖20份、葡萄汁30份、蜂蜜48份、毛蚶活性多肽4份和柠檬汁27份溶解得基液,将浸膏加入2倍体积的水加热溶解并加入浸膏质量46%的基液,得复配物,抗氧化剂由4重量份的维生素C和0.4重量份的混合物组成,混合物为L-亮氨酰甘氨酸和D-亮氨酰甘氨酸,两者质量比为53:0.5。
高温瞬时灭菌:将复配物后装瓶于103℃条件下持续15s高温瞬时灭菌;
冷却及检验:检查瓶体有无破裂等异常现象,贴上标签后将复配物于4℃低温保存。
实施例3:
选用实施例1提取的毛蚶活性多肽,从中山大学实验动物中心子细胞库购买小鼠淋巴瘤细胞株 YAC-1,由浙江省实验动物中心,购买SPF 级昆明种小鼠, 雄性,体重18-20 g,普通洁净环境下饲养, 温度17-20℃ ,动物可以自由摄食和饮水。
取小鼠淋巴瘤细胞, 调整细胞密度为5 × 107个/ mL,加入20孔细胞培养板,每孔1 mL,每孔分别加入含不同浓度毛蚶活性多肽液1 mL,使毛蚶活性多肽液终浓度分别为20,100,500 μg/mL,并设阳性和阴性( 终浓度5 μg /mL Con A和等体积细胞培养液) 对照孔,每孔溶液终体积为2 mL,每组均设3复孔,置 37 ℃、5% CO2培养箱中培养18 h,2000 r/min 离心10 min,弃去培养液,65%预冷乙醇重悬,混匀,置于2℃固定过夜。固定后的细2000r /min离心10 min,弃去液体,各管加PI 染液( PI 2. 5mg,RNase 5 mg, Triton X-100 15 μL 溶于 PBS 40 mL中,过滤,2 ℃避光保存) 250 μL,混匀后避光放置 18min,经340目尼龙网过滤,流式细胞仪检测细胞周期,计算得小鼠淋巴瘤细胞进入S期的细胞百分率为42.8%。
实施例4:
从毛蚶提取的活性多肽制备的复配物的感官评定:
请10位产品质量检验所有一定感官评审经验的化验员对实施例1、实施例2和市售保健饮料对照组进行感官评定,评分标准见表1,每一指标均采用10分制,满分为50分,计算各指标得分后,乘以加权值后相加,即得感官评定分值。感官评定结果为:实施1为48.1、实施例2为48.8、对照组为44.1,由此可知,实施例1和实施例2的从毛蚶提取的活性多肽制备的复配物的感官评定得分高于对照组,更易于被消费者接受。
表1从毛蚶提取的活性多肽制备的复配物的感官评分标准
上述实施例中的常规操作为本领域技术人员所熟知,在此不进行赘述。
以上所述的实施例对本发明的技术方案进行了详细说明,应理解的是以上所述仅为本发明的具体实施例,并不用于限制本发明,凡在本发明的原则范围内所做的任何修改、补充或类似方式替代等,均应包含在本发明的保护范围之内。
Claims (7)
1.从毛蚶提取的活性多肽制备复配物方法,包括毛蚶活性多肽的提取,复配物的制备,冷却保存,其特征在于:所述复配物的制备步骤为:按重量份计取粉碎后的盐蒿粉末和纤维素酶,加入蒸馏水并调pH值至5.5~6.0,固液比为1g:10-17ml,于45~50℃下酶解30-35min,过滤取酶解液,加入无水乙醇和抗氧化剂搅拌20-25min,加热干燥得浸膏,备用,将牛奶、维生素A、白糖、葡萄汁、蜂蜜、毛蚶活性多肽和柠檬汁溶解得基液,将浸膏加入2-2.3倍体积的水加热溶解并加入浸膏质量45~48%的基液,得复配物。
2.根据权利要求1所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:所述复配物由以下成分及重量份组成:盐蒿粉末60-180份、纤维素酶1-3份、蒸馏水500-600份、无水乙醇20-24份、抗氧化剂0.06-0.12份、牛奶2-7份、维生素A0.01-0.03份、白糖15-30份、葡萄汁27-38份、蜂蜜45-50份、毛蚶活性多肽3-7份、柠檬汁27-30份。
3.根据权利要求1所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:所述抗氧化剂由3-5重量份的维生素C和0.1-0.6重量份的混合物组成,所述混合物为L-亮氨酰甘氨酸和D-亮氨酰甘氨酸,两者质量比为47~53:0.5-0.8。
4.根据权利要求1所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:所述毛蚶活性多肽的提取步骤为:取新鲜毛蚶去壳,加入蒸馏水,匀浆,除杂,过滤,得滤液A,然后在滤液A中加入乙酸溶液搅拌提取,离心,取上清液,上清液中加入NaCl至溶液最终浓度为2.5mol/L,同时析出絮状沉淀,离心,得毛蚶活性多肽。
5.根据权利要求4所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:蒸馏水中含有0.08-0.1wt%的D-苦杏仁酸和L-苦杏仁酸的混合体,D-苦杏仁酸和L-苦杏仁酸重量比为52~57:0.3~0.9。
6.根据权利要求4所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:除杂操作为:将匀浆液分别用0.15-0.25mol/L的NaOH和0.1-0.15mol/L的EDTA-2Na溶液浸泡,过滤,得滤液A。
7.根据权利要求1所述的从毛蚶提取的活性多肽制备复配物方法,其特征在于:所述复配物冷却保存温度为2-5℃。
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