CN111203280A - 一种具有类尿酸酶活性的铂纳米团簇及其制备方法 - Google Patents
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Abstract
本发明公开了一种具有类尿酸酶活性的铂纳米团簇的制备方法,以氯铂酸作为反应物金属前驱体,鱼精蛋白作为修饰剂稳定并诱导铂纳米颗粒形成多级团簇结构。制得的铂纳米团簇具有类尿酸酶活性,其活性可与天然尿酸酶相媲美;制备方法简单易操作、一步合成、易于批量生产;反应条件温和,仅需常温常压,安全无污染对环境友好,设备投资少成本低,重现性良好;所用蛋白安全性高、热稳定性好、具有很高的营养性和功能性;所得产物具有无毒、生物相容性好、纯化和储存简单、稳定性良好、催化活性高等特点,对人体高尿酸相关疾病(如痛风)的治疗具有重要的价值。
Description
技术领域
本发明涉及纳米生物技术领域,具体涉及一种具有类尿酸酶活性的铂纳米团簇及其制备方法。
背景技术
近年来,随着纳米材料的发展,一些纳米材料逐渐被发现具有与天然蛋白酶类似的高效催化活性,且克服了天然酶制备复杂、价格昂贵、易失活等缺点,被定义为“纳米酶”。纳米酶催化活性高、成本低、易保存、稳定性好,而且具有纳米材料的特性,在疾病诊断、食品安全、生物传感、疾病治疗等领域有着广泛的应用前景。目前已经开发了多种纳米酶可以用于多种疾病的治疗,包括肿瘤、炎症、阿尔茨海默症、帕金森综合症等,而对于痛风的研究则相对较少。痛风是由尿酸沉积引起的一种常见的炎症性关节炎,可导致严重的关节畸形和残疾。由于缺乏尿酸酶,人体内的尿酸水平比几乎其他所有哺乳动物都高。因此,开发一种更有效、更安全的痛风新疗法具有重要意义。考虑到人体内缺乏尿酸酶,我们试图寻找一种合适的催化剂代替尿酸酶来催化尿酸的氧化降解。
众所周知,纳米铂由于表面活性中心多、催化效率高、生物相容性好等优点,在工业催化、光学、电学以及纳米医学等领域都有着重要应用。目前大部分研究主要集中在阐明和改善纳米铂的催化性能,特别是合成具有高效催化性能的复杂形貌铂纳米材料,往往合成步骤复杂且催化性能单一。由于纳米酶的催化性能与其组成、结构和尺寸等因素密切相关,通过调控纳米铂的结构及表面性质可以进一步发挥其在纳米医学中的潜在作用,可以作为高效的尿酸氧化酶模拟酶催化降解尿酸,为痛风的治疗提供一种新的选择。
发明内容
有鉴于此,本发明的目的是提供一种具有类尿酸酶活性的铂纳米团簇的制备方法。本发明以鱼精蛋白为修饰剂和稳定剂制得团簇状铂纳米酶,该纳米酶具有高效的类尿酸酶活性。
实现本发明目的的技术解决方案是:一种鱼精蛋白稳定的铂纳米团簇的制备方法,以氯铂酸作为金属前驱体,硼氢化钠为还原剂,鱼精蛋白作为修饰剂稳定并诱导铂纳米颗粒形成团簇结构。具体步骤如下:
(1)将氯铂酸水溶液与鱼精蛋白水溶液混合,室温下磁力搅拌一段时间得到鱼精蛋白-氯铂酸混合前体溶液;
(2)缓慢滴加新鲜配制冰水浴的硼氢化钠溶液到鱼精蛋白-氯铂酸混合前体溶液中,保证反应体系中氯铂酸与硼氢化钠的摩尔比为1:5~1:10,在1200~1500 r/min条件下继续搅拌1~4 h,产物纯化,置于4℃储存备用。
较佳的,鱼精蛋白以鱼精蛋白水溶液的形式加入,鱼精蛋白水溶液的浓度为0.2~1.0 mg/mL。
具体的,鱼精蛋白为硫酸鲑鱼精蛋白。
较佳的,氯铂酸以氯铂酸水溶液的形式加入,氯铂酸水溶液的浓度为0.4~0.8mM。
较佳的,鱼精蛋白-氯铂酸混合前体溶液中,鱼精蛋白浓度为0.1~0.5 mg/mL,氯铂酸浓度为0.2~0.4 mM。
较佳的,步骤(1)中,室温下在1200~1500 r/min条件下磁力搅拌15~30 min。
较佳的,冰水浴的硼氢化钠溶液温度为0~4℃。
与现有技术相比,本发明具有如下优点:
(1)制备方法简单易操作、一步合成、易于批量生产;(2)反应条件温和,仅需常温常压,安全无污染对环境友好,设备投资少成本低,重现性良好;(3)所用蛋白安全性高、热稳定性好、具有很高的营养性和功能性;(4)所得产物具有无毒、生物相容性好、纯化和储存简单、稳定性良好、催化活性高等特点,对人体高尿酸相关疾病(如痛风)的治疗具有重要的意义;(5)本发明制备的铂团簇纳米酶具有高效的尿酸酶活性,催化降解尿酸符合一级反应规律。
附图说明
图1是实施例1中氯铂酸浓度为0.3 mM,鱼精蛋白浓度为0.2 mg/mL,硼氢化钠浓度为1.5 mM制得的铂纳米团簇的TEM图。
图2是实施例1中所制备的铂纳米团簇的粒径分布直方图。
图3是铂纳米团簇催化尿酸氧化降解的时间依赖性紫外-可见光谱图。
图4是铂纳米团簇催化降解尿酸氧化降解百分比与时间之间的关系。
具体实施方式
实施例1
制备鱼精蛋白修饰的铂纳米团簇
第一步:在温度为25℃,磁力搅拌的条件下,将4.25 mL 0.4 mg/mL鱼精蛋白水溶液加入到4.25 mL 0.6 mM 氯铂酸水溶液中;第二步:所得混合液预混搅拌15 min,搅拌速度为1500 r/min,得到鱼精蛋白-氯铂酸混合前体溶液;第三步:将1.5 mL 10 mM新鲜配制冰水浴的硼氢化钠溶液缓慢滴加到鱼精蛋白-氯铂酸混合前体溶液中,持续搅拌反应2 h。第四步:产物纯化,4℃储存备用,得到团簇状铂纳米酶,其微观结构和尺寸分布分别如图1和图2所示。
实施例2
制备鱼精蛋白修饰的铂纳米团簇
第一步:在温度为25℃,磁力搅拌的条件下,将4.25 mL 0.4 mg/mL鱼精蛋白水溶液加入到4.25 mL 0.6 mM 氯铂酸水溶液中;第二步:所得混合液预混搅拌30 min,搅拌速度为1250 r/min,得到鱼精蛋白-氯铂酸混合前体溶液;第三步:将1.5 mL 10 mM新鲜配制冰水浴的硼氢化钠溶液缓慢滴加到鱼精蛋白-氯铂酸混合前体溶液中,持续搅拌反应4 h。第四步:产物纯化,4℃储存备用,得到团簇状铂纳米酶。
实施例3
制备鱼精蛋白修饰的铂纳米团簇
第一步:在温度为25℃,磁力搅拌的条件下,将4.25 mL 1.0 mg/mL鱼精蛋白水溶液加入到4.25 mL 0.8 mM 氯铂酸水溶液中;第二步:所得混合液预混搅拌15 min,搅拌速度为1500 r/min,得到鱼精蛋白-氯铂酸混合前体溶液;第三步:将1.5 mL 10 mM新鲜配制冰水浴的硼氢化钠溶液缓慢滴加到鱼精蛋白-氯铂酸混合前体溶液中,持续搅拌反应2 h。第四步:产物纯化,4℃储存备用,得到团簇状铂纳米酶。
实施例4
鱼精蛋白修饰的铂纳米团簇的类尿酸酶活性的测定
在1300μL超纯水中加入1500 μL 0.2 mM尿酸溶液,然后加入200 μL以上实施例中所制备的团簇状铂纳米酶使其浓度为0.02 mM,最终反应溶液体积为3 mL,在37℃条件下反应30min,以2 min时间间隔测定体系的紫外-可见光谱。如图3所示,谱图中尿酸在290 nm处特征吸收峰随时间逐渐降低表明其具有良好的尿酸酶活性。图4表明,尿酸的催化降解符合一级反应。
Claims (10)
1.一种鱼精蛋白稳定的铂纳米团簇的制备方法,其特征在于,以氯铂酸作为金属前驱体,硼氢化钠为还原剂,鱼精蛋白作为修饰剂稳定并诱导铂纳米颗粒形成团簇结构,具体步骤如下:
(1)将氯铂酸水溶液与鱼精蛋白水溶液混合,搅拌得到鱼精蛋白-氯铂酸混合前体溶液;
(2)缓慢滴加冰水浴的硼氢化钠溶液到鱼精蛋白-氯铂酸混合前体溶液中,保证反应体系中氯铂酸与硼氢化钠的摩尔比为1:5~1:10,在1200~1500 r/min条件下继续搅拌1~4h进行还原反应,纯化。
2. 如权利要求1所述的方法,其特征在于,鱼精蛋白水溶液的浓度为0.2~1.0 mg/mL。
3.如权利要求1或2所述的方法,其特征在于,鱼精蛋白为硫酸鲑鱼精蛋白。
4. 如权利要求1所述的方法,其特征在于,氯铂酸水溶液的浓度为0.4~0.8 mM。
5. 如权利要求1所述的方法,其特征在于,步骤(1)中,室温下在1200~1500 r/min条件下磁力搅拌15~30 min。
6. 如权利要求1所述的方法,其特征在于,鱼精蛋白-氯铂酸混合前体溶液中,鱼精蛋白浓度为0.1~0.5 mg/mL,氯铂酸浓度为0.2~0.4 mM。
7.如权利要求1所述的方法,其特征在于,冰水浴的硼氢化钠溶液温度为0~4℃。
8. 如权利要求1所述的方法,其特征在于,在1200~1500 r/min条件下继续搅拌1~4h进行还原反应。
9.如权利要求1-8任一项所述的方法制备的鱼精蛋白稳定的铂纳米团簇。
10.如权利要求1-8任一项所述的方法制备的鱼精蛋白稳定的铂纳米团簇在催化尿酸的氧化降解中的用途。
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