CN106038484B - 一种蓝莓花色苷壳聚糖纳米乳液的制备方法及应用 - Google Patents
一种蓝莓花色苷壳聚糖纳米乳液的制备方法及应用 Download PDFInfo
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Abstract
本发明提供了一种蓝莓花色苷壳聚糖纳米乳液的制备方法及应用,属于纳米技术领域,所述制备方法包括:蓝莓果渣超声提取得到蓝莓花色苷提取液;冷冻干燥;将蓝莓花色苷粗提物加入浓度为0.5~0.8mg/mL壳聚糖盐酸盐溶液中,得到混合液Ⅰ;将pH为6.0±0.1,浓度为0.5~2.0mg/mL羧甲基壳聚糖溶液逐滴加入到混合液Ⅰ中,得到蓝莓花色苷壳聚糖纳米乳液。本发明制备的蓝莓花色苷壳聚糖纳米乳液可以用于制备滴眼液和蓝莓花色苷饮料中。本发明制备方法简单,制备的蓝莓花色苷壳聚糖纳米乳液的平均粒径小于500nm,同时,本发明制备的蓝莓花色苷壳聚糖纳米乳液中的花色苷随贮存时间的延长,损失率极小,制备的滴眼液和饮料具有花色苷含量高,性质稳定等特点。
Description
技术领域
本发明涉及一种纳米乳液及其应用,特别是涉及蓝莓花色苷壳聚糖纳米乳液及其制备方法和在滴眼液及饮料中的应用。
背景技术
纳米技术自20世纪80年代被科学界提出后,正在科技发展的今天飞速崛起,已经成为当今世界各国空前发展的世界前沿技术之一。纳米技术被定义为以粒子大小均匀分布在1-500 nm 之间的物质为主体,此研究主体大小在此尺度范围内具有与宏观物质、同时也与单个孤立原子不同特性的新兴的功能技术。纳米技术涵盖领域广泛, 包括纳米材料学、纳米生物学和纳米显微学等方面, 它建立了一种崭新的思维方式, 使人类能够利用越来越小、越来越精确的物质和越来越精细的技术成品来满足更高层次的要求。目前, 由于纳米技术具有的独特优势以及人们对健康和重大疾病防治等问题的日益关注, 纳米技术已经广泛应用于生物医药领域。2003年9月,美国学者首先报道了关于将纳米技术应用到农业和食品行业的研究,这对食品行业的发展无疑具有巨大的推动作用。“纳米食品技术”的概念也被欧洲研究学者随后提出,此概念在狭义的定义上是指经过原子、分子等的修饰达到生成新型的食品分子技术的目的。而广泛的定义指纳米食品技术可以在食品生产整个过程发挥作用,如生产、加工、包装、运输等。结合纳米技术构建生物活性成分运输系统,改善功能分子的稳定性、氧化性、生物利用率等诸方面的问题,是纳米食品技术的核心和关键。这种新型的纳米运输系统诸如纳米乳液、聚合物纳米囊、脂质体、纳米微球等用于包埋营养功能成分并达到保护和靶向释放功能分子的纳米载体技术。
花青素,是黄酮类化合物,是广泛存在于植物中的天然水溶性色素,因主要以糖苷的形式存在,所以也称花色苷。科研结果表明:花青素是现今所发现的最有效的天然水溶性自由基清除剂。随着对花青素更加深入的研究,研究者们发现花青素不仅有助于老年性白内障的预防和糖尿病性白内障的发生,而且可以通过强大的抗氧化作用消除或缓解人体眼部疲劳。此外,花青素还具有其他功效:①延缓衰老、激活巨噬细胞,使血清免疫球蛋白免受自由基的侵害;②提高免疫力、降低心脑血管疾病的发病率;③降低糖尿病及其并发症的发病机率、改善退行型老年痴呆、保护基因的完整性;④降低癌症的发病率、抗发炎等诸多功能。联合国粮农组织把富含花青素的蓝莓列为“人类五大健康食品之一”,并因其拥有活化视网膜的作用,被誉为“飞行员的早餐”,说明它在改善视力有着非比寻常的效果。此外,世界卫生组织和联合国粮农组织组成的食品添加剂联合专家委员会对花色苷的毒理性进行考察,结果显示,花色苷是一类安全、无毒、多功能的天然抗氧化剂。
壳聚糖是天然多糖甲壳素的脱乙酰基产物,是一种含有游离氨基的碱性多糖,具有多种生理功能。经降解和化学修饰后的壳聚糖,在某些方面具有比壳聚糖更好的生物活性。壳聚糖及其降解物和修饰物安全性良好,且具有可降解性和组织相容性,在医药领域具有很高的应用价值。大量的研究已经证实壳聚糖及其衍生物具有以下药理活性:①抗菌活性;②调节脂质代谢;③调节凝血功能;④抗氧化性能。
但目前稳定花青素的研究主要集中在辅色作用和微胶囊化技术上,通过纳米乳液提高花青素稳定及其制备的方法到目前为止还较未发现。纳米乳液包埋技术作为纳米科技的核心技术之一,在功能性食品组分的运输载体构建方面显示出极大的潜力。
发明内容
本发明的目的是提供蓝莓花色苷含量高且稳定性好,经过长期存放后,花色苷损失量极低的蓝莓花色苷壳聚糖纳米乳液,所述制备原料主要包括蓝莓果渣、壳聚糖盐酸盐和羧甲基壳聚糖,具体制备方法依次包括如下步骤:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣用pH=3.0±0.1、60~65%(v/v)乙醇水溶液超声提取,得到蓝莓花色苷提取液;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将蓝莓花色苷粗提物加入浓度为0.5~0.8mg/mL壳聚糖盐酸盐溶液中, 反应8~30min,,得到混合液Ⅰ;
第四步:将pH为6.0±0.1,浓度为0.5~2.0mg/mL 羧甲基壳聚糖溶液逐滴加入到混合液Ⅰ中,反应0.5~2h, 得到蓝莓花色苷壳聚糖纳米乳液。
进一步,所述超声提取的条件为:超声功率150-600W,提取温度50~70℃,超声时间15-50 min。
进一步,所述羧甲基壳聚糖溶液和混合液Ⅰ的混合比例为:1:10~1:2(v/v)
进一步,所述蓝莓花色苷粗提物添加到壳聚糖盐酸盐溶液的添加比例为:1:6~1:2(m/v)。同时,本发明还提供这种蓝莓花色苷壳聚糖纳米乳液的一种应用,即采用本发明的蓝莓花色苷壳聚糖纳米乳液用于制备滴眼液。
为了满足滴眼液舒适、安全等要求,所述滴眼液的制备原料还包括:聚乙烯醇、氯化钠、硫柳汞。
所述滴眼液的配方为:聚乙烯醇1~1.4%,氯化钠0.4~0.8%,硫柳汞0.001-0.004%,余量是蓝莓花色苷壳聚糖纳米乳液。
本发明还提供了蓝莓花色苷壳聚糖纳米乳液另外一种应用,所述的蓝莓花色苷壳聚糖纳米乳液用于制备蓝莓花色苷饮料。
进一步,所述蓝莓花色苷饮料的主要成分还包括:甜味剂和酸味剂。
具体的,所述蓝莓花色苷饮料的配方为:甜味剂3~14%,酸味剂0.08~0.32%,余量是蓝莓花色苷壳聚糖纳米乳液。
有益效果
其一,本发明提出的蓝莓花色苷壳聚糖纳米乳液相比普通蓝莓花色苷提取液,具有以下优点:(1)稳定性高,可防止粒子间的聚集和储存过程中的重力分层,在不添加稳定剂的条件下存放6个月不发生分层现象;(2)本发明的蓝莓花色苷壳聚糖纳米乳液的比表面积较大,能长期保持动力学稳定性,在储存过程中无絮凝、聚集、沉淀、奥斯特瓦尔德成熟现象,可以看成是一种“近热力学稳定”体系;(3)本发明的蓝莓花色苷壳聚糖纳米乳液光学透明,散射光弱,可用于透明液态滴眼液和饮料中,对原有产品的颜色影响甚微。
其二,本发明的蓝莓花色苷壳聚糖纳米乳液在滴眼液中可以最大程度的保留蓝莓花色苷
稳定性,该滴眼液具有成色透明,稳定性好,根据申请人的加速实验,存放72小时的花色苷残留率是87.2%,而对照组只有7.1%,存在显著差异。由此证明,经过长时间存放后,纳米乳液和滴眼液里的蓝莓花色苷损失率低,本发明可以有效的将花色苷保留在滴眼液中,该滴眼液的配方均为国家规定允许添加的成分,安全可靠,同时能够有效缓解眼部疲劳,维护视觉健康、增进眼部微细血管循环、提高微血管和静脉流动、保护微血管作用、防止近视再加深、预防重度近视及视网膜剥离病变之产生,促进视网膜细胞中的视紫质再生,预防近视,增进视力等功效。
其三,本发明工艺简单,制备过程不需特殊设备,可自发形成,纳米乳粒径一般为1~500nm;本发明制备的蓝莓花色苷壳聚糖纳米乳液黏度低,制备成的滴眼液可减少眼睛的疼痛和不适;本发明的蓝莓花色苷壳聚糖纳米乳液还具有缓释和靶向作用。最后,本发明的蓝莓花色苷壳聚糖纳米乳液具有提高药物的溶解度,减少药物的分解,可形成对药物的保护作用并提高对药物的吸收,提高药物的生物利用度。大部分天然功能性色素对光、热、氧和其他加工条件较为敏感,将功能性色素包埋制成纳米乳能够起到保护作用,提高其加工应用稳定性,兼有延长贮藏期的作用。
纳米乳液是热力学不稳定动力学稳定的体系,小粒径能够增强布朗运动从而减轻重力沉降影响,能够防止在储存过程中乳液分层、沉降或絮凝现象的发生。本发明蓝莓花色苷壳聚糖纳米乳液的粒径均小于500nm,大大降低了蓝莓花色苷乳液的粒径,使蓝莓花色苷乳液达到纳米级别,并且粒径小于大部分可见光波长,对光的散射效应微弱,纳米乳液体系通常呈透明或半透明状态,用作脂溶性活性物质载体添加到饮料中且不影响产品的透明度。
附图说明
图1:本发明实施例1制备的蓝莓花色苷壳聚糖纳米乳液粒径分布图。
具体实施方式
下面结合具体实施例子进一步阐明本发明,应理解这些实施例仅用于说明本发明,而不用于限制本发明的范围,在阅读了本发明之后,本领域的技术人员对本发明各种等价形式的修改均落于本申请所附权利要求所规定的范围。
实施例1
本实施例提供了一种蓝莓花色苷壳聚糖纳米乳液,所述蓝莓花色苷壳聚糖纳米乳液由蓝莓果、壳聚糖盐酸盐和羧甲基壳聚糖制备而成,其的制备方法如下:
蓝莓花色苷壳聚糖纳米乳液的制备过程如下:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣按照料液比1:20g/mL,加入pH=3.0±0.1、60%(v/v)乙醇水溶液,并在超声功率300W,提取温度60℃的条件下,超声提取30min,得到蓝莓花色苷提取液;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将8mg蓝莓花色苷粗提物加入30mL浓度为0.6mg/mL壳聚糖盐酸盐溶液,在磁力搅拌条件下反应10min,得到混合液Ⅰ;
第四步:用蒸馏水配制1.5mg/mL 羧甲基壳聚糖溶液,并用柠檬酸将羧甲基壳聚糖的pH调整为6.0±0.1。取10mL上述羧甲基壳聚糖,逐滴加入到30mL混合液Ⅰ中, 完全融合后反应1h, 即得蓝莓花色苷壳聚糖纳米乳液。
实施例2
本实施例提供了一种蓝莓花色苷壳聚糖纳米乳液,所述蓝莓花色苷壳聚糖纳米乳液由蓝莓果、壳聚糖盐酸盐和羧甲基壳聚糖制备而成,其的制备方法如下:
蓝莓花色苷壳聚糖纳米乳液的制备过程如下:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣按照料液比1:25g/mL,加入pH=3.0±0.1、65%(v/v)乙醇水溶液,并在超声功率150W,提取温度70℃的条件下,超声提取15min,得到蓝莓花色苷提取液;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将10mg蓝莓花色苷粗提物加入30mL浓度为0.6mg/mL壳聚糖盐酸盐溶液,在磁力搅拌条件下反应20min,得到混合液Ⅰ;
第四步:用蒸馏水配制1.0mg/mL 羧甲基壳聚糖溶液,并用柠檬酸将羧甲基壳聚糖的pH调整为6.0±0.1。取15mL上述羧甲基壳聚糖,逐滴加入到30mL混合液Ⅰ中, 完全融合后反应2h, 即得蓝莓花色苷壳聚糖纳米乳液。
实施例3
本实施例提供了一种蓝莓花色苷壳聚糖纳米乳液,所述蓝莓花色苷壳聚糖纳米乳液由蓝莓果、壳聚糖盐酸盐和羧甲基壳聚糖制备而成,其的制备方法如下:
蓝莓花色苷壳聚糖纳米乳液的制备过程如下:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣按照料液比1:15g/mL,加入pH=3.0±0.1、62%(v/v)乙醇水溶液,并在超声功率600W,提取温度50℃的条件下,超声提取40min,得到蓝莓花色苷提取液;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将5mg蓝莓花色苷粗提物加入30mL浓度为0.8mg/mL壳聚糖盐酸盐溶液,在磁力搅拌条件下反应8min,得到混合液Ⅰ;
第四步:用蒸馏水配制0.5mg/mL 羧甲基壳聚糖溶液,并用柠檬酸将羧甲基壳聚糖的pH调整为6.0±0.1。取9mL上述羧甲基壳聚糖,逐滴加入到30mL混合液Ⅰ中, 完全融合后反应0.8h, 即得蓝莓花色苷壳聚糖纳米乳液。
实施例4
本实施例提供了一种蓝莓花色苷壳聚糖纳米乳液,所述蓝莓花色苷壳聚糖纳米乳液由蓝莓果、壳聚糖盐酸盐和羧甲基壳聚糖制备而成,其的制备方法如下:
蓝莓花色苷壳聚糖纳米乳液的制备过程如下:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣按照料液比1:20g/mL,加入pH=3.0±0.1、60%(v/v)乙醇水溶液,并在超声功率400W,提取温度60℃的条件下,超声提取50min,得到蓝莓花色苷提取液;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将15mg蓝莓花色苷粗提物加入30mL浓度为0.5mg/mL壳聚糖盐酸盐溶液,在磁力搅拌条件下反应30min,得到混合液Ⅰ;
第四步:用蒸馏水配制2.0mg/mL 羧甲基壳聚糖溶液,并用柠檬酸将羧甲基壳聚糖的pH调整为6.0±0.1。取3mL上述羧甲基壳聚糖,逐滴加入到30mL混合液Ⅰ中, 完全融合后反应0.5h, 即得蓝莓花色苷壳聚糖纳米乳液。
为了测定本发明制备的纳米乳液的性能,申请人测定了本发明纳米乳液中花色苷稳定性,并将蓝莓花色苷粗提物直接溶于等体积的蒸馏水中,制备了对照组。
将实施例1得到的蓝莓花色苷壳聚糖纳米乳液和对照组的蓝莓花色苷溶液置于37℃恒温培养箱中,间隔检测蓝莓花色苷含量。比较两种液体中花色苷的稳定性。
申请人用以下方法检测蓝莓花色苷壳聚糖纳米乳液及对照组中的蓝莓花色苷含量,具体步骤如下:
采用UHPLC 检测花色苷含量
具体方法为:分别取2mL上述液体,再用0.45 um微孔滤膜过滤后,使用UHPLC检侧,所述UHPLC的条件为:A相:0.5%甲酸水,B相:甲酸甲醇乙腈水,进样流速:0.2mL/min, 柱温:35.0 °C,使用紫外检测器,波长为510nm,并采用以下的梯度洗脱,具体见下表。并用矢车菊素3-O-葡萄糖苷标准品绘制标准曲线,测定出花色苷含量。
梯度洗脱条件
经检测,对照组在37 ℃ 恒温培养箱中储存3天后,花色苷保留率为7.1%,而在蓝莓花色苷壳聚糖纳米乳液中,花色苷保留率为87.2%。
每天测定的数据变化如下:
在37℃贮藏条件下,花色苷含量的变化
由上表可以看出,实施例1组相对于对照组,其花色苷的稳定性显著增强。
同时,申请人还用美国Beckman coulter 公司的Delsa Max Pro 粒度仪测定了实施例1~实施例4的粒径及电位,实施例1制备的蓝莓花色苷壳聚糖纳米乳液粒径分布图见图1,实施例1~4的平均粒径和电位如下表所示:
实施例1~4的粒径电位测定结果表
电位可以反映一个体系的稳定性,无论电位的正负,绝对值越大,表明体系越稳定,研究表明,当羧甲基壳聚糖溶液的浓度为0.5或2.0mg/mL时,其电位显著降低,体系的稳定性显著下降,申请人使用2.5mg/mL和0.3mg/mL羧甲基壳聚糖溶液逐滴加入到30mL混合液Ⅰ中,2.5mg/mL羧甲基壳聚糖溶液逐滴加入到30mL混合液Ⅰ中后,无法形成纳米乳液,出现大量沉淀;0.3mg/mL羧甲基壳聚糖溶液逐滴加入到30mL混合液Ⅰ中后,电位急剧降低而无法形成稳定的纳米乳液体系。
实施例5
本实施例提供了蓝莓花色苷壳聚糖纳米乳液在滴眼液中的应用:
用实施例1~4制备的蓝莓花色苷壳聚糖纳米乳液配制滴眼液,其配方如下:
配方一:聚乙烯醇1g,氯化钠0.4g,硫柳汞0.001g,蓝莓花色苷壳聚糖纳米乳液98.599g。
配方二:聚乙烯醇1.2g,氯化钠0.5g,硫柳汞0.003g,蓝莓花色苷壳聚糖纳米乳液98.297g。
配方三:聚乙烯醇1.4g,氯化钠0.8g,硫柳汞0.004g,蓝莓花色苷壳聚糖纳米乳液97.796g。
按照上述配方配制的100g滴眼液稳定性好,经过长时间存放后,滴眼液里的蓝莓花色苷含量高,有效的将花色苷融入到滴眼液中,该滴眼液的配方均为国家规定允许添加的成分,安全可靠,同时能够有效缓解眼部疲劳,维护视觉健康、增进眼部微细血管循环、提高微血管和静脉流动、保护微血管作用、防止近视再加深、预防重度近视及视网膜剥离病变之产生,促进视网膜细胞中的视紫质再生,预防近视,增进视力等功效。
实施例6
本实施例提供了蓝莓花色苷壳聚糖纳米乳液在蓝莓花色苷饮料中的应用:
用实施例1~4制备的蓝莓花色苷壳聚糖纳米乳液配制蓝莓花色苷饮料,其配方如下:
配方1:蔗糖3g,柠檬酸0.08g,蓝莓花色苷壳聚糖纳米乳液96.92g。
配方2:蔗糖6g,柠檬酸0.12g,蓝莓花色苷壳聚糖纳米乳液93.88g。
配方3:蔗糖10g,柠檬酸0.23g,蓝莓花色苷壳聚糖纳米乳液89.77g。
配方4:蔗糖14g,柠檬酸0.32g,蓝莓花色苷壳聚糖纳米乳液85.68g。
配方5:蔗糖4g,木糖醇4g,酒石酸0.11g,蓝莓花色苷壳聚糖纳米乳液91.89g。
配方6:蔗糖5g,木糖醇5g,苹果酸0.20g,蓝莓花色苷壳聚糖纳米乳液89.80g。
配方7:蔗糖7g,木糖醇7g,柠檬酸0.28g,蓝莓花色苷壳聚糖纳米乳液85.72g。
按照上述配方制备的蓝莓饮料酸甜适口,存放6个月以上不发生分层现象,且在贮藏中能够保留原有的蓝莓花色苷,有效的抑制了蓝莓花色苷的分解。
Claims (7)
1.一种蓝莓花色苷壳聚糖纳米乳液的制备方法:其特征在于,所述蓝莓花色苷壳聚糖纳米乳液的制备方法依次包括如下步骤:
第一步:蓝莓果渣超声提取:将蓝莓果取汁后的果渣用pH=3.0±0.1、体积分数为60~65%乙醇水溶液超声提取,得到蓝莓花色苷提取液;其中,所述蓝莓果渣超声提取的条件为:超声功率150-600W,提取温度50~70℃,超声时间15-50 min;
第二步:冷冻干燥:将蓝莓花色苷提取液冷冻干燥,得到蓝莓花色苷粗提物;
第三步:将蓝莓花色苷粗提物加入浓度为0.5~0.8mg/mL壳聚糖盐酸盐溶液中,反应8~30min,得到混合液Ⅰ;
第四步:将pH为6.0±0.1,浓度为0.5~2.0mg/mL 羧甲基壳聚糖溶液逐滴加入到混合液Ⅰ中,反应0.5~2h, 得到蓝莓花色苷壳聚糖纳米乳液,其中,所述羧甲基壳聚糖溶液和混合液Ⅰ的混合体积比为:1:10~1:2,所述蓝莓花色苷粗提物添加到壳聚糖盐酸盐溶液的质量体积比为:1:6~1:2。
2.权利要求1所述的蓝莓花色苷壳聚糖纳米乳液在制备滴眼液中的应用,其特征在于:所述的蓝莓花色苷壳聚糖纳米乳液用于制备滴眼液。
3.根据权利要求2所述的蓝莓花色苷壳聚糖纳米乳液的应用, 其特征在于:所述滴眼液的主要成分还包括:聚乙烯醇、氯化钠、硫柳汞。
4.根据权利要求3所述的蓝莓花色苷壳聚糖纳米乳液的应用, 其特征在于:所述滴眼液的配方为:聚乙烯醇1~1.4%,氯化钠0.4~0.8%,硫柳汞0.001-0.004%,余量是蓝莓花色苷壳聚糖纳米乳液。
5.权利要求1所述的蓝莓花色苷壳聚糖纳米乳液在制备饮料的应用, 其特征在于:所述的蓝莓花色苷壳聚糖纳米乳液用于制备蓝莓花色苷饮料。
6. 根据权利要求5所述的蓝莓花色苷壳聚糖纳米乳液的应用, 其特征在于:所述蓝莓花色苷饮料的主要成分还包括:甜味剂和酸味剂。
7. 根据权利要求6所述的蓝莓花色苷壳聚糖纳米乳液的应用, 其特征在于:所述蓝莓花色苷饮料的配方为:甜味剂3~14%,酸味剂0.08~0.32%,余量是蓝莓花色苷壳聚糖纳米乳液。
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