CN109142288B - 利用荧光光谱法测定囊泡生成的方法 - Google Patents
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Abstract
本发明公开了一种利用荧光光谱法测定囊泡生成的方法,取比色管分别加入表面活性剂,再分别加入不同体积梯度的对甲苯胺溶液,然后置于超声仪中,边超声边滴加尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后倒入比色皿中,然后在相同的测试条件下进行荧光测量,随着对甲苯胺加入量的增加,混合体系的荧光强度最大值先升高后降低则说明有囊泡生成,混合体系的荧光强度最大值持续降低则说明无囊泡生成。本发明的测定方法操作简单且成本低廉,避免了因普通透射电镜抽真空导致囊泡破裂,也避免了用冷冻电镜扫描而造成的高额成本。
Description
技术领域
本发明属于囊泡生成测定技术领域,具体涉及一种利用荧光光谱法测定囊泡生成的方法。
背景技术
荧光试剂尼罗红是一种脂溶性噁嗪类染料,在亚微米胶束浓度区域表现出相当大的吸收。当分散在含水表面活性剂或胶束溶液中时,尼罗红的分子倾向于通过π-π堆积相互作用形成非吸收性二聚体或H型聚集体。由于尼罗红对局部环境的敏感性,有利于用于研究各种非均相体系的局部极性,包括胶束和预胶束。
对甲苯胺(P-methyaniline)是一种无色、有毒的固体,微溶于水,在工业上对甲苯胺是重要的染料和农药中间体。同样它也是污染水环境的主要污染物之一,它的去除和资源化对人们的生活具有重大的意义。对甲苯胺与阴离子表面活性剂具有非共价的作用,从而诱导表面活性剂聚集并导致CMC向低浓度转变,静电吸引力是对甲苯胺与表面活性剂相互作用的主要驱动力。
表面活性剂是两亲分子,它们可以通过静电作用、氢键、范德华力作用等相互作用自组装形成液晶、胶束、囊泡等不同形态的聚集体,因此而被人们广泛研究。囊泡与人体内细胞中的细胞膜很相似,通常利用囊泡来模拟细胞膜在不同环境中的状态。由于pH、浓度、电解质和温度等因素都会影响囊泡的形成,所以亟需找到有效测定囊泡生成的方法。
发明内容
本发明解决的技术问题是提供了一种简单快速且灵敏度较高的利用荧光光谱法测定囊泡生成的方法,该方法可以简单快速地测定出有无囊泡生成,易于操作且成本低廉。
本发明为解决上述技术问题采用如下技术方案,利用荧光光谱法测定囊泡生成的方法,其特征在于具体步骤为:取规格一样的25mL比色管11个,分别加入1-2.5mL之间相同体积的摩尔浓度为0.1mol/L的表面活性剂,再分别加入0-5mL之间不同体积梯度的摩尔浓度为5×10-2mol/L的对甲苯胺溶液,然后置于超声仪中,边超声边滴加0.25mL摩尔浓度为2.5×10-4mol/L的尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后倒入比色皿中,然后在相同的测试条件下进行荧光测量,随着对甲苯胺加入量的增加,混合体系的荧光强度最大值先升高后降低则说明有囊泡生成,混合体系的荧光强度最大值持续降低则说明无囊泡生成,所述表面活性剂为SDSN、SDS或SDBS,测试条件为:入射和出射狭缝宽度均为5nm,扫描速率为1200nm/min,最大激发波长为λex=549nm,测量范围为550-750nm。
本发明的测定方法操作简单且成本低廉,利用荧光强度的变化即可测定有无囊泡生成,避免了因普通透射电镜抽真空导致囊泡破裂,也避免了用冷冻电镜扫描而造成的高额成本。
附图说明
图1是表面活性剂加入量为1mL时混合体系荧光强度最大值随对甲苯胺加入量的变化曲线;
图2是表面活性剂加入量为1.5mL时混合体系荧光强度最大值随对甲苯胺加入量的变化曲线;
图3是表面活性剂加入量为2mL时混合体系荧光强度最大值随对甲苯胺加入量的变化曲线;
图4是表面活性剂加入量为2.5mL时混合体系荧光强度最大值随对甲苯胺加入量的变化曲线;
图5是表面活性剂为SDBS时的透射电镜图;
图6是表面活性剂为SDS时的透射电镜图。
具体实施方式
以下通过实施例对本发明的上述内容做进一步详细说明,但不应该将此理解为本发明上述主题的范围仅限于以下的实施例,凡基于本发明上述内容实现的技术均属于本发明的范围。
实施例1
取规格一样的25mL比色管11个,分别加入1mL摩尔浓度为0.1mol/L的表面活性剂SDS,再分别加入0、0.5mL、1mL、1.5mL、2mL、2.5mL、3mL、3.5mL、4mL、4.5mL和5mL摩尔浓度为5×10-2mol/L的对甲苯胺溶液,然后置于超声仪中,边超声边滴加0.25mL摩尔浓度为2.5×10-4mol/L的尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后按顺序倒入比色皿中,在相同的测试环境下:入射与出射狭缝宽度均为5nm,扫速速率为1200nm/min,最大激发波长为λex=549nm,测量范围为550-750nm,作图观察其荧光发射光谱。用同样的方法改变表面活性剂SDS加入量依次为1.5mL、2mL和2.5mL,测其荧光强度。
实施例2
取规格一样的25mL比色管11个,分别加入1mL摩尔浓度为0.1mol/L的表面活性剂SDSN,再分别加入0、0.5mL、1mL、1.5mL、2mL、2.5mL、3mL、3.5mL、4mL、4.5mL和5mL摩尔浓度为5×10-2mol/L的对甲苯胺溶液,然后置于超声仪中,边超声边滴加0.25mL摩尔浓度为2.5×10-4mol/L的尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后按顺序倒入比色皿中,在相同的测试环境下:入射与出射狭缝宽度均为5nm,扫速速率为1200nm/min,最大激发波长为λex=549nm,测量范围为550-750nm,作图观察其荧光发射光谱。用同样的方法改变表面活性剂SDSN加入量依次为1.5mL、2mL和2.5mL,测其荧光强度。
实施例3
取规格一样的25mL比色管11个,分别加入1mL摩尔浓度为0.1mol/L的表面活性剂SDBS,再分别加入0、0.5mL、1mL、1.5mL、2mL、2.5mL、3mL、3.5mL、4mL、4.5mL和5mL摩尔浓度为5×10-2mol/L的对甲苯胺溶液,然后置于超声仪中,边超声边滴加0.25mL摩尔浓度为2.5×10-4mol/L的尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后按顺序倒入比色皿中,在相同的测试环境下:入射与出射狭缝宽度均为5nm,扫速速率为1200nm/min,最大激发波长为λex=549nm,测量范围为550-750nm,作图观察其荧光发射光谱。用同样的方法改变表面活性剂SDBS加入量依次为1.5mL、2mL和2.5mL,测其荧光强度。
由图1-4可知,随着对甲苯胺溶液加入量的递增,无论表面活性剂浓度多大,表面活性剂SDS或SDSN的混合体系的荧光强度最大值都是递减的。这是由于对甲苯胺和这两种表面活性剂可以发生自组装,对甲苯胺在此过程中只作猝灭剂,使体系的聚集和组装遭到破坏,稳定性减弱,所以荧光强度递减。但在表面活性剂SDBS中,随着对甲苯胺溶液加入量的增大,混合体系的荧光强度最大值都是先升高后降低。荧光强度最大值升高是由于对甲苯胺与表面活性剂SDBS相互作用,促进了SDBS-尼罗红体系从胶束阶段转变到了囊泡阶段。尼罗红在胶束阶段时荧光吸收不太好,但尼罗红进入囊泡中,荧光强度开始增强,所以囊泡出现越多,荧光强度越大,所以开始是荧光逐步增强的;但之后体系的荧光强度开始降低,这可能是因为随着对甲苯胺溶液加入量的增大,发生了能量转移,导致荧光猝灭。这两个作用对荧光强度都有影响,荧光强度最后的增减取决于两个作用的综合结果。所以可以根据荧光强度先增后减这一现象,从而可以判定表面活性剂SDBS与对甲苯胺之间有囊泡产生。图5-6进一步验证了本发明的测定结果。
以上实施例描述了本发明的基本原理、主要特征及优点,本行业的技术人员应该了解,本发明不受上述实施例的限制,上述实施例和说明书中描述的只是说明本发明的原理,在不脱离本发明原理的范围下,本发明还会有各种变化和改进,这些变化和改进均落入本发明保护的范围内。
Claims (1)
1.利用荧光光谱法测定囊泡生成的方法,其特征在于具体步骤为:取规格一样的25mL比色管11个,分别加入1-2.5mL之间相同体积的摩尔浓度为0.1mol/L的表面活性剂,再分别加入0-5mL之间不同体积梯度的摩尔浓度为5×10-2mol/L的对甲苯胺溶液,然后置于超声仪中,边超声边滴加0.25mL摩尔浓度为2.5×10-4mol/L的尼罗红的四氢呋喃溶液,再加入二次蒸馏水定容,摇匀,放入超声仪中超声15min,将定容后的混合体系在室温环境下静置4h,再振荡摇匀后倒入比色皿中,然后在相同的测试条件下进行荧光测量,随着对甲苯胺加入量的增加,混合体系的荧光强度最大值先升高后降低则说明有囊泡生成,混合体系的荧光强度最大值持续降低则说明无囊泡生成,所述表面活性剂为SDSN、SDS或SDBS,测试条件为:入射和出射狭缝宽度均为5nm,扫描速率为1200nm/min,最大激发波长为λex=549nm,测量范围为550-750nm。
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