CN113966838A - 一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒及制备方法 - Google Patents
一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒及制备方法 Download PDFInfo
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- CN113966838A CN113966838A CN202111197927.5A CN202111197927A CN113966838A CN 113966838 A CN113966838 A CN 113966838A CN 202111197927 A CN202111197927 A CN 202111197927A CN 113966838 A CN113966838 A CN 113966838A
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Abstract
本发明公开了一种虾青素纳米结构脂质载体‑壳聚糖凝胶微粒及制备方法,微粒包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油1‑5%、乙酰化单双甘油脂肪酸酯1‑10%、维生素E醋酸酯1‑10%、吐温类乳化剂1‑10%、聚甘油类乳化剂1‑10%、大豆卵磷脂0.5‑5%、余量为去离子水,微粒由以下制备方法制得:油相的制备;水相的制备;预乳化处理;虾青素纳米结构脂质载体的制备;虾青素NLC‑壳聚糖多层乳液的制备;添加三聚磷酸钠溶液;虾青素纳米结构脂质载体‑壳聚糖凝胶微粒的制备。本发明具有提高虾青素的稳定性和生物利用度,提升虾青素的水分散性,便于添加至功能食品或保健品中等优点。
Description
【技术领域】
本发明属于凝胶微粒的技术领域,尤其涉及一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒及制备方法。
【背景技术】
虾青素是一种酮式类的胡萝卜素,结构中含有长长的共轭不饱和双键,其末端具有不饱和酮基和羟基基团,通过吸引自由基和提供给自由基电子来清除自由基。虾青素作为天然抗炎的抗氧化剂之一,它是类胡萝卜素合成的最高级别产物。虾青素在体外和体内模型中都显示出潜在的生物活性,对动物和人类代谢有一定有益作用。它是唯一能通过血脑屏障的类胡萝卜素,可以阻止视网膜氧化,维持中枢神经系统的健康,对黄斑变性也有很好的防治作用。此外,虾青素对于免疫反应还可以进行调节,抗炎抗感染和调节肝脂代谢,预防肿瘤和心血管疾病、延缓糖尿病等慢性病的发展。关于运动方面,它能够减轻运动性缺血再灌注损伤,延缓运动性疲劳及防止继发性器官损伤,进而提高运动能力。
基于脂质的活性物传递系统以生理相容性的脂质为载体,可以用来增加难溶性活性物的吸收,提高生物利用度。纳米结构脂质载体是将固态脂质和空间上不相容的液体脂质在一定温度下混合制备得到的载体。液态油的加入破坏了晶格排列,增加了脂质微粒结构中不规则晶型的比例,使其承载活性物的空间容量增加从而提高载体的载药能力,载体固化而不结晶,呈均一的、含有“液态纳米室”的固态载体状态。纳米结构脂质载体能够明显提高难溶性活性物的溶解度,和其他胶体传递系统相比(乳液、脂质体等),具有较高的物理稳定性,同时活性成分被包裹或负载于固态脂质的内部,减少了活性氧在液态油脂之间的流动性,能显著地提高活性物的化学稳定性。
壳聚糖是目前自然界中已知的唯一带有正电荷的碱性多糖,它具有良好的生物相容性、生物可降解性、无毒性和生物粘附性,且有抗菌消炎、促进伤口愈合、抗酸抗溃疡及直接抑制肿瘤细胞的作用,与现有的抗癌药合用可增强抗癌效果。且壳聚糖来源丰富,价格低廉,使得壳聚糖可以作为理想的营养物输送材料。壳聚糖在磷酸根离子的交联下,可形成具有三维网络状结构的凝胶。壳聚糖凝胶在伤口愈合、软骨组织工程以及药物载体方面有着广泛的应用。
目前自然条件下虾青素存在水溶性差、不稳定并且易于氧化降解和褪色从而使生物活性丧失等问题,导致虾青素的生物利用度比较低,会造成资源的浪费,这给其在食品工业中的应用带来诸多不便。
【发明内容】
本发明的目的就是解决背景技术中的问题,提出一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒及制备方法,能够提高虾青素的稳定性和生物利用度,提升虾青素的水分散性,便于添加至功能食品或保健品中。
为实现上述目的,本发明提出了一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,所述虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油1-5%、乙酰化单双甘油脂肪酸酯1-10%、维生素E醋酸酯1-10%、吐温类乳化剂1-10%、聚甘油类乳化剂1-10%、大豆卵磷脂0.5-5%、余量为去离子水。
作为优选,所述吐温类乳化剂为吐温80、吐温60和吐温40中的一种或多种。
作为优选,所述聚甘油类乳化剂为十聚甘油单油酸酯、十聚甘油单硬脂酸酯、十聚甘油单月桂酸酯、六聚甘油单油酸酯、六聚甘油单硬脂酸酯、六聚甘油单月桂酸酯、分子蒸馏单甘脂中的一种或多种。
本发明还提出了一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,包括以下几个步骤:
油相的制备:按配比称取乙酰化单双甘油脂肪酸酯、吐温类乳化剂、聚甘油类乳化剂,加热溶解成液体,之后加入虾青素油和维生素E醋酸酯,搅拌分散均匀,获得油相;
水相的制备:将大豆卵磷脂溶解分散于去离子水中,获得水相;
预乳化处理:将上述水相加入到油相中,加热搅拌,完成预乳化;
虾青素纳米结构脂质载体的制备:对预乳化后的溶液进行微射流均质处理,冷却至室温,获得虾青素纳米结构脂质载体;
虾青素NLC-壳聚糖多层乳液的制备:将壳聚糖盐酸盐按质量浓度1~4%的比例加入上述的虾青素纳米结构脂质载体中,搅拌溶解均匀,获得虾青素NLC-壳聚糖多层乳液;
添加三聚磷酸钠溶液:在高剪切条件下,将三聚磷酸钠溶液添加到上述的虾青素NLC-壳聚糖多层乳液中;
虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备:先预冻一段时间,后进行冷冻干燥,最终获得虾青素纳米结构脂质载体-壳聚糖凝胶微粒。
作为优选,所述三聚磷酸钠溶液的质量浓度为2.5~10%。
作为优选,所述微射流均质处理由微射流均质机进行,并设置微射流均质机的微射流压力为9000~15000psi,循环2~4次。
作为优选,所述预冻在冰箱中进行,冰箱温度设置为-75~-85℃。
作为优选,所述冷冻干燥在冷阱中进行,冷阱温度设置为-75~-85℃,冷阱的真空度小于0.1Torr。
作为优选,所述预乳化处理中加热方式采用水浴加热,水浴加热的温度设置在60~80℃。
作为优选,所述高剪切条件是在高剪切分散乳化机下进行,所述高剪切分散乳化机的转速设置为9000~15000rpm。
本发明的有益效果:1).本发明通过将虾青素油负载于纳米结构脂质载体的中间固态核中,减少了油脂中活性氧的流动性,解决了虾青素水溶性和稳定性差的应用瓶颈;2)维生素E醋酸酯是一种营养成分,同时也是一种抗氧化保护剂,纳米结构脂质载体中采用维生素E醋酸酯为液态油脂,可进一步保护虾青素;3)纳米结构脂质载体中使用了大豆卵磷脂为乳化剂,大豆卵磷脂可以和壳聚糖静电自组装,在纳米结构脂质载体的表面覆盖上一层壳聚糖,形成多层乳液,进一步保护虾青素,延缓其释放,提高生物利用度;4)多层乳液表面的壳聚糖,在三聚磷酸钠溶液磷酸根离子的交联下,得到凝胶微粒,形成三位网络状的囊壳,可进一步保护虾青素,延缓其释放,提高生物利用度;5)采用冷冻干燥的方法将凝胶微粒制备成固体,增加其食用便利性和多样性。
本发明的特征及优点将通过实施例结合附图进行详细说明。
【附图说明】
图1是本发明一种实施例的制备方法流程图;
图2是本发明一种实施例的虾青素光稳定性检测结果示意图;
图3是本发明一种实施例的虾青素体外释放情况结果示意图。
【具体实施方式】
本发明提出了一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油1-5%、乙酰化单双甘油脂肪酸酯1-10%、维生素E醋酸酯1-10%、吐温类乳化剂1-10%、聚甘油类乳化剂1-10%、大豆卵磷脂0.5-5%、余量为去离子水。
进一步地,吐温类乳化剂为吐温80、吐温60和吐温40中的一种或多种。
进一步地,聚甘油类乳化剂为十聚甘油单油酸酯、十聚甘油单硬脂酸酯、十聚甘油单月桂酸酯、六聚甘油单油酸酯、六聚甘油单硬脂酸酯、六聚甘油单月桂酸酯、分子蒸馏单甘脂中的一种或多种。
以下通过具体实施例进一步对本发明进行说明,下述实施例仅用于说明本发明而对本发明没有限制:
实施例1
一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油1g、乙酰化单双甘油脂肪酸酯1g、维生素E醋酸酯1g、吐温801g、十聚甘油单油酸酯1g、大豆卵磷脂0.5g、去离子水94.5g。
参阅图1,该虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,包括以下几个步骤:
S01,油相的制备:按上述配比称取乙酰化单双甘油脂肪酸酯、吐温80、十聚甘油单油酸酯,于80℃水浴中加热溶解成液体,之后加入1g虾青素油和1g维生素E醋酸酯,搅拌分散均匀,获得油相;
S02,水相的制备:称取0.5g大豆卵磷脂和94.5g去离子水,将大豆卵磷脂加入去离子水中,溶解搅拌分散均匀,并于80℃水浴中保温,获得水相;
S03,预乳化处理:在500rpm搅拌条件下将上述水相加入到油相中,并于80℃水浴中加热搅拌30min,完成预乳化;
S04,虾青素纳米结构脂质载体的制备:微射流均质机预热,设置微射流均质机的微射流压力为10000psi,循环3次,对预乳化后的溶液进行微射流均质处理,均质完成后冷却至室温,获得虾青素纳米结构脂质载体;
S05,虾青素NLC-壳聚糖多层乳液的制备:将壳聚糖盐酸盐按质量浓度1%的比例加入上述的虾青素纳米结构脂质载体中,搅拌溶解均匀,获得虾青素NLC-壳聚糖多层乳液;
S06,添加三聚磷酸钠溶液:在高剪切分散乳化机的10000rpm高剪切条件下,取质量浓度2.5%的三聚磷酸钠溶液40ml逐滴添加到上述的虾青素NLC-壳聚糖多层乳液中,边加边剪切,直至添加完成;
S07,虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备:将S06得到的溶液放入冰箱中在-80℃温度下预冻24h,之后放入冷冷阱进行冷冻干燥,冷阱的温度设置为-80℃,冷阱的真空度小于0.1Torr,冷冻24h,最终获得虾青素纳米结构脂质载体-壳聚糖凝胶微粒。
实施例2
一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油2g、乙酰化单双甘油脂肪酸酯4g、维生素E醋酸酯10g、吐温605g、十聚甘油单油酸酯10g、大豆卵磷脂5g、去离子水64g。
参阅图1,该虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,包括以下几个步骤:
S01,油相的制备:按上述配比称取乙酰化单双甘油脂肪酸酯、吐温60、十聚甘油单油酸酯,于60℃水浴中加热溶解成液体,之后加入2g虾青素油和10g维生素E醋酸酯,搅拌分散均匀,获得油相;
S02,水相的制备:称取5g大豆卵磷脂和64g去离子水,将大豆卵磷脂加入去离子水中,溶解搅拌分散均匀,并于60℃水浴中保温,获得水相;
S03,预乳化处理:在500rpm搅拌条件下将上述水相加入到油相中,并于60℃水浴中加热搅拌30min,完成预乳化;
S04,虾青素纳米结构脂质载体的制备:微射流均质机预热,设置微射流均质机的微射流压力为12000psi,循环3次,对预乳化后的溶液进行微射流均质处理,均质完成后冷却至室温,获得虾青素纳米结构脂质载体;
S05,虾青素NLC-壳聚糖多层乳液的制备:将壳聚糖盐酸盐按质量浓度4%的比例加入上述的虾青素纳米结构脂质载体中,搅拌溶解均匀,获得虾青素NLC-壳聚糖多层乳液;
S06,添加三聚磷酸钠溶液:在高剪切分散乳化机的12000rpm高剪切条件下,取质量浓度10%的三聚磷酸钠溶液40ml逐滴添加到上述的虾青素NLC-壳聚糖多层乳液中,边加边剪切,直至添加完成;
S07,虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备:将S06得到的溶液放入冰箱中在-80℃温度下预冻24h,之后放入冷冷阱进行冷冻干燥,冷阱的温度设置为-80℃,冷阱的真空度小于0.1Torr,冷冻24h,最终获得虾青素纳米结构脂质载体-壳聚糖凝胶微粒。
实施例3
一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油5g、乙酰化单双甘油脂肪酸酯10g、维生素E醋酸酯4g、吐温8010g、分子蒸馏单甘脂4g、大豆卵磷脂2g、去离子水65g。
参阅图1,该虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,包括以下几个步骤:
S01,油相的制备:按上述配比称取乙酰化单双甘油脂肪酸酯、吐温80、分子蒸馏单甘脂,于60℃水浴中加热溶解成液体,之后加入5g虾青素油和4g维生素E醋酸酯,搅拌分散均匀,获得油相;
S02,水相的制备:称取2g大豆卵磷脂和65g去离子水,将大豆卵磷脂加入去离子水中,溶解搅拌分散均匀,并于60℃水浴中保温,获得水相;
S03,预乳化处理:在500rpm搅拌条件下将上述水相加入到油相中,并于60℃水浴中加热搅拌30min,完成预乳化;
S04,虾青素纳米结构脂质载体的制备:微射流均质机预热,设置微射流均质机的微射流压力为15000psi,循环3次,对预乳化后的溶液进行微射流均质处理,均质完成后冷却至室温,获得虾青素纳米结构脂质载体;
S05,虾青素NLC-壳聚糖多层乳液的制备:将壳聚糖盐酸盐按质量浓度3%的比例加入上述的虾青素纳米结构脂质载体中,搅拌溶解均匀,获得虾青素NLC-壳聚糖多层乳液;
S06,添加三聚磷酸钠溶液:在高剪切分散乳化机的15000rpm高剪切条件下,取质量浓度7.5%的三聚磷酸钠溶液40ml逐滴添加到上述的虾青素NLC-壳聚糖多层乳液中,边加边剪切,直至添加完成;
S07,虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备:将S06得到的溶液放入冰箱中在-80℃温度下预冻24h,之后放入冷冷阱进行冷冻干燥,冷阱的温度设置为-80℃,冷阱的真空度小于0.1Torr,冷冻24h,最终获得虾青素纳米结构脂质载体-壳聚糖凝胶微粒。
对制备的虾青素纳米结构脂质载体-壳聚糖凝胶微粒性能检测:
一、稳定性检测:
按如下方法进行虾青素的光稳定性测试:
将样品置于室温自然光照条件下四周,每周取适量样品并检测虾青素的含量,计算虾青素的保留率。样品设置实验组和对照组,实验组为按实施例1制备方法制得的虾青素纳米结构脂质载体-壳聚糖凝胶微粒,对照组包括虾青素NLC-壳聚糖多层乳液、虾青素乳液和虾青素NLC,虾青素乳液采用辛癸酸甘油三酯为油相,乳化剂的比例和组成同实施例1,虾青素NLC采用不含维生素E醋酸酯(VEA)的虾青素纳米结构脂质载体,以乙酰化单双甘油脂肪酸酯为固态脂质,辛癸酸甘油三酯为液态脂质,乳化剂的比例及组成同实施例1,虾青素NLC-壳聚糖多层乳液为实施例1制备方法的中间产品,光稳定试验结果如图2所示。
由图2可知,虾青素乳液的光稳定性较差,储存4周后,虾青素几乎全部被破坏;使用固态脂质乙酰化单双甘油脂肪酸酯部分替代乳液中的液态脂质后,由于固态脂质的固化作用,虾青素被束缚在固态核内,活性氧在其中的流动性降低,氧化稳定性得到了提高;进一步,在纳米结构脂质载体中使用维生素E醋酸酯为液态脂质后,由于VEA的保护作用,虾青素的稳定性得到了增强;在NLC的外层包裹上壳聚糖,由于二次包裹作用,虾青素的稳定性进一步提高;最后,将多层乳液的外层壳聚糖凝胶化后,虾青素纳米结构脂质载体-壳聚糖凝胶微粒的光稳定性最高。可见,通过以上固态脂质、维生素E醋酸酯、壳聚糖包裹和凝胶化作用可起到改善虾青素稳定性的作用。
二、体外释放
采用透析袋法来考察虾青素纳米结构脂质载体-壳聚糖凝胶微粒的释放特点。本实验分为三组,实验组为采用本发明实施例3制备方法获得的最终成品虾青素纳米结构脂质载体-壳聚糖凝胶微粒,对照组1和2分别是采用本发明实施例3制备方法获得的中间产品虾青素纳米结构脂质载体和虾青素NLC-壳聚糖多层乳液。称取一定量的样品于透析袋中,用两只透析袋夹将其封口,控制透析袋中无气泡出现,且确保封口处无漏液,然后将透析袋放入200mL释放介质中,为保证漏槽条件,本实验释放介质使用磷酸盐缓冲液(pH=7.4)与无水乙醇的混合溶液(7:3,v/v),实验时保持释放介质温度为37℃,磁力搅拌转速为150rpm。实验开始后,按照预先设定的时间点用移液枪进行取样,每次取样量为3mL,取样完成后再加入3mL新鲜释放介质,以保证释放介质总量保持不变,然后利用紫外分光光度计检测取得样品中虾青素的含量。样品体外释放情况如图3所示。
由图3可知,纳米结构脂质载体中虾青素释放较快,2小时几乎完全释放。在纳米结构脂质载体的表面覆盖上壳聚糖后,由于壳聚糖膜的阻碍作用,虾青素的释放速度明显变缓,4小时才几乎释放完全。进一步,将纳米结构脂质载体外层的壳聚糖凝胶化后,由于壳聚糖凝胶微粒的三维网络状结构,虾青素的释放变得更加缓慢。由此可知,通过壳聚糖的覆盖和凝胶化,可以实现对纳米结构脂质载体中虾青素的控制释放,有望提高虾青素的生物利用度。
本发明的各种原料均可从市场购得,制备方法中采用设备均采用现有设备,上述实施例是对本发明的说明,不是对本发明的限定,任何对本发明简单变换后的方案均属于本发明的保护范围。
Claims (10)
1.一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒,其特征在于:包括虾青素纳米结构脂质载体、覆盖在虾青素纳米结构脂质载体表面的壳聚糖,所述虾青素纳米结构脂质载体由以下百分比质量的组分组成:虾青素油1-5%、乙酰化单双甘油脂肪酸酯1-10%、维生素E醋酸酯1-10%、吐温类乳化剂1-10%、聚甘油类乳化剂1-10%、大豆卵磷脂0.5-5%、余量为去离子水。
2.如权利要求1所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒,其特征在于:所述吐温类乳化剂为吐温80、吐温60和吐温40中的一种或多种。
3.如权利要求1所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒,其特征在于:所述聚甘油类乳化剂为十聚甘油单油酸酯、十聚甘油单硬脂酸酯、十聚甘油单月桂酸酯、六聚甘油单油酸酯、六聚甘油单硬脂酸酯、六聚甘油单月桂酸酯、分子蒸馏单甘脂中的一种或多种。
4.一种虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:包括以下几个步骤:
油相的制备:按配比称取乙酰化单双甘油脂肪酸酯、吐温类乳化剂、聚甘油类乳化剂,加热溶解成液体,之后加入虾青素油和维生素E醋酸酯,搅拌分散均匀,获得油相;
水相的制备:将大豆卵磷脂溶解分散于去离子水中,获得水相;
预乳化处理:将上述水相加入到油相中,加热搅拌,完成预乳化;
虾青素纳米结构脂质载体的制备:对预乳化后的溶液进行微射流均质处理,冷却至室温,获得虾青素纳米结构脂质载体;
虾青素NLC-壳聚糖多层乳液的制备:将壳聚糖盐酸盐按质量浓度1~4%的比例加入上述的虾青素纳米结构脂质载体中,搅拌溶解均匀,获得虾青素NLC-壳聚糖多层乳液;
添加三聚磷酸钠溶液:在高剪切条件下,将三聚磷酸钠溶液添加到上述的虾青素NLC-壳聚糖多层乳液中;
虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备:先预冻一段时间,后进行冷冻干燥,最终获得虾青素纳米结构脂质载体-壳聚糖凝胶微粒。
5.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述三聚磷酸钠溶液的质量浓度为2.5~10%。
6.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述微射流均质处理由微射流均质机进行,并设置微射流均质机的微射流压力为9000~15000psi,循环2~4次。
7.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述预冻在冰箱中进行,冰箱温度设置为-75~-85℃。
8.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述冷冻干燥在冷阱中进行,冷阱温度设置为-75~-85℃,冷阱的真空度小于0.1Torr。
9.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述预乳化处理中加热方式采用水浴加热,水浴加热的温度设置在60~80℃。
10.如权利要求4所述的虾青素纳米结构脂质载体-壳聚糖凝胶微粒的制备方法,其特征在于:所述高剪切条件是在高剪切分散乳化机下进行,所述高剪切分散乳化机的转速设置为9000~15000rpm。
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