CN109125744A - 一种具有mri与ct双模态成像功能的钆掺杂氧化铪纳米颗粒的制备方法 - Google Patents
一种具有mri与ct双模态成像功能的钆掺杂氧化铪纳米颗粒的制备方法 Download PDFInfo
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Abstract
本发明公开了一种具有核磁(MRI)与电子计算机断层扫描(CT)双模态成像功能的钆掺杂的氧化铪(HfO2:Gd)纳米颗粒的制备方法。此方法是利用微波水热技术,其基本步骤为:含铪与钆的前驱体溶液制备、微波水热、离心洗涤、冷冻干燥。通过微波水热制备得到的HfO2:Gd纳米颗粒具有良好的分散性且大小为~65nm。研究表明HfO2:Gd颗粒具有良好的生物相容性以及双模态成像功能(MRI与CT成像)。本发明制备过程简单,制备所需时间短,原料成本低廉,整个制备过程在空气气氛中进行,无需特殊装置。
Description
技术领域
本发明涉及双模态影像纳米材料,具体是指一种具有MRI和CT造影功能的钆掺杂的氧化铪纳米颗粒的制备方法。
背景技术
医学影像技术为肿瘤的早期诊断与治疗评价提供了可靠依据,成为备受重视的研究热点。目前医学影像技术主要包括磁共振成像(MRI),电子计算机X线断层扫描技术(CT),荧光成像,正电子发射断层摄影术(PET)等。近年来,这些医学成像技术被广泛用于各种疾病的诊断。单一的成像技术虽然有自身的优势,然而同时也存在其局限性,因而不能达到精确诊断的目的。例如,CT是目前最常用的非侵入性医学成像手段,可以对高电子密度的材料形成优质图像,但是软组织对X射线的吸收效果相似,所以用CT很难分辨软组织间的不同。因此在使用CT时往往需注射CT增强造影剂使软组织的成像更清晰。MRI虽然空间分辨率高和软组织对比度佳,但是它没有光学成像灵敏。因此,单一的成像手段不能够精确获取全部信息,而用多种成像手段获得的信息相叠加,可以解决单一成像手段存在的灵敏度和分辨率的问题。CT软组织分辨率低,但成本低、分辨率相对高,是目前临床上最广泛使用的成像手段;而MRI虽然灵敏度较低,但软组织和空间分辨率高,因此将两者结合起来可以有效地进行功能互补。所以对CT/MRI造影剂的开发很有必要。
铪(Hf)因为原子序数较大(Z=72),具有较高的X射线吸收系数大,因而可以产生较好的CT增强成像性能。而临床上最常用的CT造影剂是小分子的碘的化合物,尽管它们可以提供安全可靠的成像,但是易被肾脏排出,成像窗口短,非特异性分布等缺点导致CT图像不清晰。同时,对于磁共振造影剂,含礼的对比剂具有较高的灵敏度和生物相容性。是含礼对比剂在临床的广泛应用,
纳米技术的迅速发展为实现有效的肿瘤的早期诊断和有效治疗提供了契机。纳米材料本身的纳米尺寸就赋予了它生物医学应用的极大优势。稳定性良好的纳米颗粒(尺寸为10~500nm)注射进入生物活体内后,显现长的血液循环时间。通过增强渗透滞留效应(EPR)或者癌细胞的主动靶向,纳米颗粒可以聚集到肿瘤部位,而在正常组织中浓度较低,降低对正常组织的毒副作用,因此可将应用领域拓展到靶向显像或治疗。与此相反,通过肾脏清除和网状内皮系统(RES)的吞噬可分别将小分子或者微米尺寸的颗粒从血液中迅速清除出去此外,由于多种不同功能可以整合在单个颗粒上,所以纳米颗粒是设计多模态造影纳米探针的理想平台。因此,纳米造影剂在基础研究和临床应用领域都备受关注,制备具有CT/MRI双模态的氧化铪基造影剂在肿瘤诊疗一体化应用中具有广泛的前景。
发明内容
本发明的目的是针对目前纳米颗粒合成的复杂性,以及分散性问题,提出一种具有MRI和CT造影功能的钆掺杂的氧化铪纳米颗粒的制备方法。
本发明采用如下技术方案:一种具有MRI与CT双模态成像功能的钆掺杂的氧化铪(HfO2:Gd)纳米颗粒的制备方法,包括以下步骤:
(1)将1.200g四氯化铪和0.464g氯化钆六水合物溶解100mL超纯水溶液中,在60℃恒温水域中磁力搅拌2h得到溶液A;
(2)将(1)中所得的A溶液分别取10mL分放在三个烧杯中,分别缓慢的滴加0.05mol/L氢氧化钠溶液10mL,在常温下继续搅拌1h得到透明的微波水热前驱体溶液;
(3)将(2)中得到的前驱体溶液,分别装入3个微波水热釜中,随后放入微波水热设备中,三个水热釜构成正三角形布置,正三角形中心与微波水热设备中心重合;设置参数为:10min上升120℃,并在120℃保温5min,继续在5min内上升到160℃,最后在160℃保温120min。微波水热工艺过程完成后,自然冷却至室温。
(4)步骤(3)中微波水热得到后的混合液放入离心管中,以6000转/分钟离心洗涤,洗涤后的产物冷冻干燥得到HfO2:Gd纳米粒子。
(5)称取100mg的HfO2:Gd纳米粒子分散在100mL的超纯水溶液中超声分散0.5h,随后加入聚丙烯酸(PAA)的水溶液(分子量为1800,浓度为2mg/mL)50mL,搅拌4h后离心并用超纯水洗涤多次去除多余的PAA分子。然后,再次将PAA改性的HfO2:Gd纳米颗粒分散在100mL的超纯水中,缓慢滴加10mL的PEG-NH2的水溶液(10mg/mL,分子量为5000kDa)继续搅拌1h。称量100mg的N-(3-二甲基氨基丙基)-N'-乙基碳二亚胺盐酸盐(EDC)加入到上述溶液中,继续搅拌12h后,离心,洗涤,并放在37℃真空干燥箱中干燥得到PEG分子改性的HfO2:Gd纳米粒子。
本发明的有益效果在于:本发明利用铪与钆的氯化前驱体水溶液,在碱性条件下生成均匀混合的铪和钆的氢氧化物,实现了钆离子的均匀掺杂。在微波水热后,生成了水相的HfO2:Gd纳米粒子且实现了Gd元素在氧化铪基相中的均匀掺杂,整个过程在水相中反应,保证了亲水性能,且制备所需时间短,无需特殊改性过程。HfO2:Gd纳米颗粒中的Hf与Gd元素分别具有CT与MRI造影功能,实现了一个纳米载体双模态成像的功能。
附图说明
图1是产物HfO2:Gd纳米粒子的TEM照片(a),以及EDX能谱分析(b);
图2是产物HfO2:Gd纳米粒子的CT影像图;其中,图2a为不同浓度梯度的HfO2:Gd纳米粒子的T1模式下的MRI图;图2b为纳米颗粒的不同浓度梯度下的弛豫时间曲线图;图2c为给裸鼠尾静脉注射200μL,20mg/mLHfO2:Gd纳米粒子后,不同时间点,裸鼠肿瘤部位的MRI成像图;
图3是产物HfO2:Gd纳米粒子的MRI影像图。其中,3a、b为不同浓度梯度的材料(HfO2:Gd纳米粒子与临床用CT造影剂碘海醇对比)的CT图,图3c为给裸鼠尾静脉注射200μL,20mg/mLHfO2:Gd纳米粒子后,不同时间点(12,24h),裸鼠肿瘤部位的CT成像图。
具体实施方式
下面结合实施例和附图对发明作进一步说明;本实施例中,以体外以及HfO2:Gd纳米粒子打入裸鼠体内的CT/MRI影像分析来说明双模态成像效果。
一种具有MRI和CT造影功能的钆掺杂的氧化铪纳米颗粒的制备方法,该方法包括以下步骤:
(1)将1.200g四氯化铪和0.464g氯化钆六水合物溶解100mL超纯水溶液中,在60℃恒温水域中磁力搅拌2h得到溶液A;
(2)将(1)中所得的A溶液分别取10mL分放在三个烧杯中,分别缓慢的滴加0.05mol/L氢氧化钠溶液10mL,在常温下继续搅拌1h得到透明的微波水热前驱体溶液;
(3)将(2)中得到的前驱体溶液,分别装入3个微波水热釜中,随后放入微波水热设备中,三个水热釜构成正三角形布置,正三角形中心与微波水热设备中心重合;设置参数为:10min上升120℃,并在120℃保温5min,继续在5min内上升到160℃,最后在160℃保温120min。微波水热工艺过程完成后,合并三个反应釜中的溶液,自然冷却至室温。该步骤中,正三角形布置是核心,其余的对称布置方法均无法得到本申请的产物。
(4)步骤(3)中微波水热得到后的混合液放入离心管中,以6000转/分钟离心洗涤,洗涤后的产物冷冻干燥得到HfO2:Gd纳米粒子。
(5)称取100mg的HfO2:Gd纳米粒子分散在100mL的超纯水溶液中超声分散0.5h,随后加入聚丙烯酸(PAA)的水溶液(分子量为1800,浓度为2mg/mL)50mL,搅拌4h后离心并用超纯水洗涤多次去除多余的PAA分子。然后,再次将PAA改性的HfO2:Gd纳米颗粒分散在100mL的超纯水中,缓慢滴加10mL的PEG-NH2的水溶液(10mg/mL,分子量为5000kDa)继续搅拌1h。称量100mg的N-(3-二甲基氨基丙基)-N'-乙基碳二亚胺盐酸盐(EDC)加入到上述溶液中,继续搅拌12h后,离心,洗涤,并放在37℃真空干燥箱中干燥得到PEG分子改性的HfO2:Gd纳米粒子。
(6)将PEG分子改性的HfO2:Gd纳米粒子分散在PBS溶液中,通过尾静脉注射方式打入到裸鼠体内,观察在裸鼠体内的CT与MRI影像功能。
1.形貌以及元素表征
图1为HfO2:Gd纳米粒子的TEM和EDX图片,纳米粒子的尺寸在~65nm,可以看出,Hf,O,Gd元素均存在纳米粒子基相中,证明HfO2:Gd纳米粒子的成功制备。
2.HfO2:Gd纳米粒子的MRI(T1)影像分析
图2为产物PEG分子改性的HfO2:Gd纳米粒子的T1模式下的MRI影像分析,其中图2a为不同浓度梯度的HfO2:Gd纳米粒子的T1模式下的MRI图,可以看出随着浓度增加,信号强度增加;图2b为纳米颗粒的不同浓度梯度下的弛豫时间曲线图,可以看出弛豫时间倒数与材料浓度梯度有较好的线性关系;图2c为给裸鼠尾静脉注射200μL,20mg/mLHfO2:Gd纳米粒子后,不同时间点,裸鼠肿瘤部位的MRI成像图,可以看出在24h内,随着时间的增加,小鼠部位MRI信号强度增加,证明纳米粒子在肿瘤部位的累积。
3.HfO2:Gd纳米粒子的CT影像分析
图3为产物PEG分子改性的HfO2:Gd纳米粒子的CT影像分析,其中图3a&b为不同浓度梯度的材料(HfO2:Gd纳米粒子与临床用CT造影剂碘海醇对比)的CT图,可以看出随着浓度增加,两种材料的信号强度增加,且HfO2:Gd纳米粒子信号强度增加较快,证明了HfO2:Gd纳米粒子较好的CT造影效果;图3c为给裸鼠尾静脉注射200μL,20mg/mLHfO2:Gd纳米粒子后,不同时间点(12,24h),裸鼠肿瘤部位的CT成像图,可以看出在24h点肿瘤部位的CT信号强度高于12h点,证明纳米粒子在肿瘤部位有累积作用与图2结果一致。
Claims (5)
1.一种具有MRI与CT双模态成像功能的钆掺杂氧化铪纳米颗粒的制备方法,其特征在于,包括以下步骤:
(1)将1.200g四氯化铪和0.464g氯化钆六水合物溶解100mL超纯水溶液中,在60℃恒温水域中磁力搅拌2h得到溶液A;
(2)将(1)中所得的A溶液分别取10mL分放在三个烧杯中,分别滴加0.05mol/L氢氧化钠溶液10mL,在常温下继续搅拌1h得到透明的微波水热前驱体溶液;
(3)将(2)中得到的前驱体溶液,分别装入3个微波水热釜中,随后放入微波水热设备中,三个水热釜构成正三角形布置,正三角形中心与微波水热设备中心重合;设置参数为:10min上升120℃,并在120℃保温5min,继续在5min内上升到160℃,最后在160℃保温120min。微波水热工艺过程完成后,自然冷却至室温。
(4)步骤(3)中微波水热得到后的混合液放入离心管中,离心洗涤,洗涤后的产物冷冻干燥得到HfO2:Gd纳米粒子。
(5)称取100mg的HfO2:Gd纳米粒子分散在100mL的超纯水溶液中超声分散0.5h,随后加入聚丙烯酸(PAA)的水溶液50mL,搅拌4h后离心并用超纯水洗涤多次去除多余的PAA分子。
(6)将PAA改性的HfO2:Gd纳米颗粒分散在100mL的超纯水中,滴加10mL的PEG-NH2的水溶液继续搅拌1h。称量100mg的N-(3-二甲基氨基丙基)-N'-乙基碳二亚胺盐酸盐(EDC)加入到上述溶液中,继续搅拌12h后,离心,洗涤,并放在37℃真空干燥箱中干燥。
2.根据权利要求1所述的方法,其特征在于,步骤4中离心洗涤的离心速度为6000转/分钟。
3.根据权利要求1所述的方法,其特征在于,所述步骤5中,聚丙烯酸(PAA)的水溶液浓度为2mg/mL。
4.根据权利要求1所述的方法,其特征在于,所述步骤5中,PAA分子量为1800。
5.根据权利要求1所述的方法,其特征在于,所述步骤6中,PEG-NH2的水溶液浓度为10mg/mL,PEG-NH2分子量为5000kDa。
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