CN116392635B - 一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料及其制备方法与应用 - Google Patents
一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料及其制备方法与应用 Download PDFInfo
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Abstract
本发明提供了一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料及其制备方法与应用。所述复合抗菌材料的制备方法，包括步骤：将纳米压电颗粒均匀分散于有机溶剂A中，然后倒入模具中，待溶剂蒸发后，加入聚合物溶液，待溶剂蒸发后，得到表面嵌有纳米压电颗粒的聚合物薄膜；将所得聚合物薄膜置于含有金属盐水溶液和甲醇水溶液的混合溶液中，调节体系pH值，然后将混合物进行超声处理，之后经洗涤、干燥，得到。本发明的复合抗菌材料可在机械能条件下来诱导活性氧产生从而杀灭细菌对抗体内植入物感染。本发明提供了一种按需和非侵入性治疗植入物相关感染的方法，可达到预防和治疗植入物相关感染的目的。
Description
技术领域
本发明涉及一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料及其制备方法与应用，属于医疗器械植入物抗菌领域。
背景技术
植入物相关感染(IAI)是植入术后并发症的主要原因，具有严重的后果。当前正值各种功能的生物医学植入物革新时期，感染的风险也进一步增加。植入物相关感染是所有医院获得性感染中最常见的并发症之一，因为它们提供了易受致病菌粘附和免疫抵抗生物膜形成影响的基质，对人类健康构成巨大威胁也增加了医疗方面的压力。植入物相关感染具有高发病率和高死亡率的特点，同时患者要花费高昂的住院费用。少部分植入物相关感染可通过感染控制措施得以预防，然而大部分的植入物相关感染难以通过无菌技术和环境控制降低感染的发生。
目前，感染仍然是植入物失败的主要原因，经常需要进行第二次手术来移除植入物，这对长期住院的患者在身体和心理方面都有严重影响。植入物相关感染中最常见的病原体是金黄色葡萄球菌(S.aureus)，若不及时清除会形成生物膜，尤其是在惰性表面上形成，例如广泛用作矫形植入物的金属、聚合物材料等医疗器械。目前IAI的临床预防和治疗仅限于全身使用抗生素和植入物移除手术(物理移除感染组织或植入物)。然而，由于生物屏障的存在，抗生素在细菌生物膜内的局部浓度较低，使抗生素的全身给药不能有效预防和治疗IAI。
因此迫切需要一种非侵入性和有效治疗植入物相关感染的替代策略。目前，已经开发了许多基于生物医学植入物的表面抗菌方法，主要包括抗生素或金属离子的局部释放、抗菌肽的接枝、光敏剂的修饰等方法。其中，抗菌肽和金属离子是消耗性、不可再生和不可控的，可能对周围组织表现出一定的毒性。例如：中国专利文献CN108392679A公开了一种植入材料表面抗菌修饰的方法，以固体氧化锌为原料，通过简单步骤修饰在以心脏起搏器为代表的金属植入材料(钛、钽、钴、铬、钼)和高分子植入材料(硅胶、聚亚安酯、聚醚醚酮、聚乳酸)表面。
此外，利用非侵入的超声等声波机械能产生活性氧(ROS)的声动力学疗法已成为潜在的抗菌策略。压电材料是一类可以捕获机械能的材料，可以将超声波等机械能转化为电能并和周围介质发生氧化还原反应引发纳米催化效应，报道具有产生ROS抗菌的效果。因此，开发一种工艺简单的超声响应抗菌的纳米压电颗粒/聚合物复合材料具有重要的意义。目前，制备表面具有纳米压电涂层的聚合物，并在机械能(超声)条件下提供一种按需和非侵入性治疗植入物相关感染的应用尚未有报道。为此，提出本发明。
发明内容
针对现有技术的不足，本发明提供了一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料及其制备方法与应用。本发明提出的纳米压电颗粒/聚合物复合抗菌材料联合使用浇注法和压电沉积法等技术合成，可以在机械能(例如超声)条件下来诱导活性氧产生从而杀灭细菌对抗体内植入物感染。因此本发明提供了一种按需和非侵入性治疗植入物相关感染的方法，可达到预防和治疗植入物相关感染的目的。
本发明的技术方案如下：
一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，所述的复合抗菌材料由聚合物基底和表面负载金属纳米粒子的纳米压电颗粒组成；所述表面负载金属纳米粒子的纳米压电颗粒嵌于聚合物基底两个表面中的一个表面上；所述表面负载金属纳米粒子的纳米压电颗粒为单侧沉积金属纳米粒子的不对称纳米结构；所述聚合物基底为聚己内酯(PCL)、聚醚醚酮(PEEK)、聚二甲基硅氧烷(PDMS)、聚氨酯(PU)、聚乙烯基吡咯烷酮(PVP)、聚甲基丙烯酸甲酯(PMMA)、聚乙烯醇(PVA)、聚乳酸(PLA)或聚乙烯(PE)；所述纳米压电颗粒为钛酸钡、铌酸锂、钛酸铅或氧化锌；所述金属纳米粒子为Au、Pt、Pd、Al或Ni纳米粒子。
根据本发明优选的，所述纳米压电颗粒的粒径为100-500nm；所述金属纳米粒子的粒径为5-50nm。
根据本发明优选的，所述金属纳米粒子为Au纳米粒子。
根据本发明，上述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，包括步骤如下：
(1)将纳米压电颗粒均匀分散于有机溶剂A中，然后倒入模具中，待溶剂挥发后，向模具中加入聚合物溶液，待溶剂挥发后，得到表面嵌有纳米压电颗粒的聚合物薄膜；
(2)将步骤(1)中所得表面嵌有纳米压电颗粒的聚合物薄膜置于含有金属盐水溶液和甲醇水溶液的混合溶液中，调节体系pH值，然后将混合物进行超声处理，之后经洗涤、干燥，得到具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料。
根据本发明优选的，步骤(1)中所述有机溶剂A为无水乙醇；所述纳米压电颗粒为钛酸钡、铌酸锂、钛酸铅或氧化锌；所述有机溶剂A的体积与纳米压电颗粒的质量之比为0.1-10mL:1mg，进一步优选为0.5-5mL:1mg。
根据本发明优选的，步骤(1)中所述模具为玻璃培养皿；所述有机溶剂A的体积与模具底部表面积之比为0.03-0.15mL:1cm2，进一步优选为0.05-0.1mL:1cm2。
根据本发明优选的，步骤(1)中所述聚合物为聚己内酯(PCL)、聚醚醚酮(PEEK)、聚二甲基硅氧烷(PDMS)、聚氨酯(PU)、聚乙烯基吡咯烷酮(PVP)、聚甲基丙烯酸甲酯(PMMA)、聚乙烯醇(PVA)、聚乳酸(PLA)或聚乙烯(PE)；所述聚合物的重均分子量Mw为10000-1000000。
根据本发明优选的，步骤(1)中所述聚合物溶液的浓度为0.1-0.2g/mL；所述聚合物溶液是将聚合物加入有机溶剂B中得到的，所述有机溶剂B为甲苯、二氯甲烷、四氢呋喃或乙酸乙酯，进一步优选为二氯甲烷。
根据本发明优选的，步骤(1)中所述聚合物溶液中的聚合物与纳米压电颗粒的质量比为100-4000:1，进一步优选为300-3000:1。
根据本发明优选的，步骤(1)中溶剂挥发的温度均为20-30℃。
根据本发明，步骤(1)中纳米压电颗粒嵌于聚合物薄膜与模具接触的一面的表面。
根据本发明优选的，步骤(2)中所述金属盐为HAuCl4、K2PtCl4、Na2PdCl4、AlCl3或NiCl2，进一步优选为HAuCl4；所述金属盐水溶液的浓度为10-100mmol/L，进一步优选为10-50mmol/L，更优选为20mmol/L。
根据本发明优选的，步骤(2)中所述甲醇水溶液的体积百分比浓度为10-20％。
根据本发明优选的，步骤(2)中所述混合溶液中金属盐水溶液和甲醇水溶液体积比为1:1-2；所述混合溶液完全没过聚合物薄膜即可。
根据本发明优选的，步骤(2)中所述调节体系pH值为将体系的pH值调节至9-10；使用K2CO3调节体系的pH值。
根据本发明优选的，步骤(2)中所述超声处理的时间为30-60min，超声频率为20-40kHz，超声功率为60-90W，超声处理的温度为0-5℃。
根据本发明优选的，步骤(2)中所述洗涤为用去离子水洗涤3-5次，所述干燥为在室温下干燥至恒重。
根据本发明，上述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的应用，用于制备治疗植入物相关感染的制品。
本发明的技术特点及有益效果如下：
1、本发明制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，联合使用浇注法和压电沉积法等技术合成，该复合抗菌材料可在超声刺激下原位产生ROS从而杀灭贴壁的细菌，从而对抗体内感染。本发明制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，纳米压电颗粒具有不对称结构，能够实现植入物表面即细菌感染的一侧产生抗菌效果，同时避免了对聚合物基底的影响。
2、本发明制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，可在体内发生再感染的情况下非侵入性地进行治疗，不依赖抗生素的使用，最终达到预防和治疗植入物相关感染的目的。
3、本发明的提出可以为控制种植物表面细菌感染提供新的表面处理技术和研究思路，有助于为植入物表面修饰技术和表面抗菌理论提供有益的指导。
附图说明
图1为本发明的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备流程示意图。
图2为实施例3制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的FIB-SEM图和选中区域的相应X射线能谱分析(EDS)图谱。
图3为对比例1制备的聚合物薄膜及实施例1-3制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的SEM图和相应EDS图谱。
图4为实施例3制备的聚合物表面纳米压电涂层剥离出来单个金属负载纳米压电颗粒的TEM图。
图5为试验例1中对比例1制备的聚合物薄膜PCL和实施例1-3制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料在超声刺激下的ROS产量曲线图。
图6为试验例2中具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料在超声作用下杀灭金黄色葡萄球菌后的菌落生长情况图。
图7为试验例2中具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料在超声作用下细菌活/死染色的荧光图(上)和SEM图(下)。
具体实施方式
下面通过具体实施例和附图对本发明作进一步的说明。本发明的实施例是为了使本领域的技术人员更好的理解本发明，并不是对本发明做任何的限制。
同时下述实施例中所述实验方法，如无特殊说明，均为常规方法；所述试剂和材料，如无特殊说明，均可从商业途径获得。
实施例中所用钛酸钡纳米颗粒的粒径为100-300nm；实施例中所用聚己内酯(PCL)重均分子量Mw为80000。
实施例1
一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，包括步骤如下：
(1)将1mg钛酸钡纳米颗粒(BTO NPs)均匀分散于5mL无水乙醇中，然后倒入直径为10cm的玻璃培养皿中，25℃下，溶剂挥发后，钛酸钡纳米颗粒均匀地沉积在容器底的表面上；
(2)将聚己内酯(PCL)溶解在二氯甲烷(CH2Cl2)中，得到浓度为0.15g/mL的聚合物溶液，然后将20mL上述溶液倒入步骤(1)的玻璃培养皿中；25℃下，溶剂挥发后，剥离容器底部，得到表面嵌有纳米压电颗粒的聚合物薄膜，纳米压电颗粒嵌于聚合物薄膜与模具接触的一面的表面；
(3)将步骤(2)中所得薄膜置于含有10mL HAuCl4(20mmol/L)水溶液和20mL体积百分比浓度为10％甲醇水溶液的混合溶液中，使用K2CO3调节体系pH值至9.7，然后将混合物放入超声清洁器(40kHz，80W)中，在4℃下进行超声处理60分钟。之后用去离子水洗涤3次、在室温下干燥至恒重，得到具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，记为piezoPCL(1)。
实施例2
一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法如实施例1所述，所不同的是：步骤(1)中钛酸钡纳米颗粒(BTO NP)的添加量为5mg，所得产物记为piezoPCL(5)。
实施例3
一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法如实施例1所述，所不同的是：步骤(1)中钛酸钡纳米颗粒(BTO NP)的添加量为10mg，所得产物记为piezoPCL(10)。
对比例1
一种聚合物薄膜的制备方法，包括步骤如下：
将聚己内酯(PCL)溶解在二氯甲烷(CH2Cl2)中以获得浓度为0.15g/mL的聚合物溶液，然后将上述20mL溶液倒入直径为10cm的玻璃培养皿中，25℃下，溶剂挥发后，剥离容器底部薄膜，即可获得聚己内酯聚合物薄膜，记为PCL。
对比例2
一种纳米压电颗粒/聚合物复合抗菌材料的制备方法如实施例1步骤(1)、(2)所述，其中步骤(1)中钛酸钡纳米颗粒(BTO NP)的添加量为10mg，所得产物记为BTO/PCL。
试验例1
实施例3制备得到的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的FIB-SEM图和选择区域的相应EDS图谱如图2所示，图中2中选区部分中颗粒状的为BTO，BTO上面白色一层为拍摄时的保护剂，BTO下面黑色部分为基底，从图2中可以看出，BTO NP成功地嵌入到聚合物基质PCL中。
对比例1制备的聚合物薄膜PCL和实施例1-3制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的SEM图和相应EDS图谱如图3所示，从图3可以看出，BTO NP均匀分布在PCL表面，且Au纳米粒子沉积成功，表明聚合物表面纳米压电涂层的成功制备。
实施例3制备得到的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料剥离出来单个金属负载纳米压电颗粒的TEM图如图4所示，从图4可以看出，Au纳米颗粒仅沉积在单个BTO NP的一侧，嵌入PCL的一侧并未沉积。
将对比例1制备的聚合物薄膜PCL和实施例1-3制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料进行ROS产量测试，具体步骤如下：用DCFH-DA检测每个样品在超声下产生的ROS总量，具体步骤：将每个样品切成直径为2cm的圆圈，然后加入10mL DCFH-DA溶液(10μM)，接下来在恒定温度下进行超声(25℃，40kHz，80W)，每隔5分钟取样，使用荧光光谱仪在激发波长为488nm、吸收波长为525nm的条件下测量荧光强度，其结果如图5所示。由图5可以看出，随着负载量的增加，ROS产量增高，说明压电性能随负载纳米压电颗粒的增加而增高。
试验例2
具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料杀灭金黄色葡萄球菌(S.aureus)实验，具体步骤如下：
(1)将对比例1制备的PCL、对比例2制备的BTO/PCL以及实施例3制备的piezoPCL(10)切成直径为1cm的圆形样品，并在接种细菌之前用氧等离子体处理以增加膜的亲水性。
(2)将对数期生长的单个金黄色葡萄球菌菌落用质量分数为0.85％ NaCl溶液梯度稀释至1×106CFU mL-1。
(3)将样品置于48孔板的中心，然后向每个孔中加入200μL细菌悬浮液。将孔板在37℃下孵育1小时，使细菌在材料上沉淀后，将每个样品用超声治疗仪在功率为1W/cm2的条件下作用5分钟，然后收集所有细菌溶液，用质量分数为0.85％ NaCl溶液冲洗至总体积为1mL。之后将其稀释100倍，然后将100μL添加到琼脂板中，用涂层棒均匀地将其铺开。将平板在37℃下孵育18小时，观察并拍照记录菌落生长情况，在显微镜下观察并记录细菌活/死染色的荧光图像，并通过SEM观察细菌的形态变化。
分别加入PCL、BTO/PCL和piezoPCL(10)组的菌落生长情况如图6所示，由图6可以看出，超声后，加入piezoPCL(10)复合抗菌材料组的菌落数大大减少，而加入PCL抗菌材料组的菌落在超声前后变化不大，而BTO/PCL杀菌效果与PCL组相差不大，说明本发明制备的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料在超声条件下具有很好的灭菌效果，并且负载金属纳米粒子能提高其压电性能。
图7为具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的荧光图像和细菌SEM图，从图7中可以看出PCL组和不施加超声的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料组中细菌没有出现明显的死亡、变形和破裂，而施加超声的具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料组有明显的变形、破裂，从而导致死亡。
Claims (10)
1.一种具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，其特征在于，所述的复合抗菌材料由聚合物基底和表面负载金属纳米粒子的纳米压电颗粒组成；所述表面负载金属纳米粒子的纳米压电颗粒嵌于聚合物基底两个表面中的一个表面上；所述表面负载金属纳米粒子的纳米压电颗粒为单侧沉积金属纳米粒子的不对称纳米结构；所述聚合物基底为聚己内酯、聚醚醚酮、聚二甲基硅氧烷、聚氨酯、聚乙烯基吡咯烷酮、聚甲基丙烯酸甲酯、聚乙烯醇、聚乳酸或聚乙烯；所述纳米压电颗粒为钛酸钡、铌酸锂、钛酸铅或氧化锌；所述金属纳米粒子为Au、Pt、Pd、Al或Ni纳米粒子。
2.根据权利要求1所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料，其特征在于，所述纳米压电颗粒的粒径为100-500nm；所述金属纳米粒子的粒径为5-50nm；所述金属纳米粒子为Au纳米粒子。
3.权利要求1或2所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，包括步骤如下：
（1）将纳米压电颗粒均匀分散于有机溶剂A中，然后倒入模具中，待溶剂挥发后，向模具中加入聚合物溶液，待溶剂挥发后，得到表面嵌有纳米压电颗粒的聚合物薄膜；所述有机溶剂A为无水乙醇；所述有机溶剂A的体积与纳米压电颗粒的质量之比为0.1-10mL:1mg；所述有机溶剂A的体积与模具底部表面积之比为0.03-0.15mL:1cm2；所述聚合物溶液的浓度为0.1-0.2g/mL；所述聚合物溶液是将聚合物加入有机溶剂B中得到的，所述有机溶剂B为甲苯、二氯甲烷、四氢呋喃或乙酸乙酯；所述聚合物溶液中的聚合物与纳米压电颗粒的质量比为100-4000:1；
（2）将步骤（1）中所得表面嵌有纳米压电颗粒的聚合物薄膜置于含有金属盐水溶液和甲醇水溶液的混合溶液中，调节体系pH值，然后将混合物进行超声处理，之后经洗涤、干燥，得到具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料；所述金属盐为HAuCl4、K2PtCl4、Na2PdCl4、AlCl3或NiCl2；所述金属盐水溶液的浓度为10-100mmol/L；所述甲醇水溶液的体积百分比浓度为10-20%；所述混合溶液中金属盐水溶液和甲醇水溶液体积比为1:1-2；所述调节体系pH值为将体系的pH值调节至9-10；所述超声处理的时间为30-60min，超声频率为20-40kHz，超声功率为60-90W，超声处理的温度为0-5℃。
4.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（1）中所述纳米压电颗粒为钛酸钡、铌酸锂、钛酸铅或氧化锌；所述有机溶剂A的体积与纳米压电颗粒的质量之比为0.5-5mL:1mg；
所述模具为玻璃培养皿；所述有机溶剂A的体积与模具底部表面积之比为0.05-0.1mL:1cm2。
5.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（1）中所述聚合物为聚己内酯、聚醚醚酮、聚二甲基硅氧烷、聚氨酯、聚乙烯基吡咯烷酮、聚甲基丙烯酸甲酯、聚乙烯醇、聚乳酸或聚乙烯；所述聚合物的重均分子量Mw为10000-1000000。
6.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（1）中所述聚合物溶液中的聚合物与纳米压电颗粒的质量比为300-3000:1；所述溶剂挥发的温度均为20-30℃。
7.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（2）中所述金属盐为HAuCl4；所述金属盐水溶液的浓度为10-50mmol/L。
8.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（2）中所述金属盐水溶液的浓度为20mmol/L。
9.根据权利要求3所述所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的制备方法，其特征在于，步骤（2）中使用K2CO3调节体系的pH值；
所述洗涤为用去离子水洗涤3-5次，所述干燥为在室温下干燥至恒重。
10.权利要求1所述具有超声响应性的纳米压电颗粒/聚合物复合抗菌材料的应用，用于制备治疗植入物相关感染的制品。
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