CN107271366B - 检测硝基苯酚的比色瓶/管 - Google Patents
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Abstract
本发明提供了一种检测硝基苯酚的比色瓶/管,所述的瓶/管体的底部为平面,瓶/管体中在所盛放的硝基苯酚富集材料的顶层中心处设置有一个反光片,光源发出的光线穿过底部平面后进入硝基苯酚富集材料,然后被反光片反射后,反射光线穿过底部平面射出;硝基苯酚富集材料包括主体基材和改性材料,主体基材用于形成透明或半透明的材料主体,改性材料用于与硝基苯酚类化合物发生相互作用并进行富集,主体基材为聚丙烯酰胺类化合物或硅橡胶类化合物,改性材料为环糊精类化合物。该比色瓶/管对待测溶液中的硝基苯酚类化合物进行富集及长光程双重信号放大,屏蔽了溶液中有色物及颗粒物的干扰,并进一步提高了光学比色检测的灵敏度。
Description
技术领域
本发明提供了一种用于高灵敏检测硝基苯酚类有色物质的检测装置,用于在环境监测及微生物鉴定等领域。
背景技术
硝基苯酚类化合物包含2-硝基苯酚,3-硝基苯酚及4-硝基苯酚,溶于碱性溶液和热水,微溶于冷水,溶液会呈现无色或金黄色,且化合物在溶液中电离的越多,溶液的黄色越明显,即当pH升高,其黄色会变深。利用该性质硝基苯酚可作为酸碱指示剂使用。酚羟基可与众多官能团形成酯键,糖苷键等,利用这一性质可合成较多酶水解的底物,水解后产生的硝基苯酚可呈现黄色,从而对酶活进行定量。如:4-硝基苯磷酸二钠为碱性磷酸酶显色底物,3-硝基苯基辛酸酯为脂肪酶显色底物,2-硝基苯-β-D-半乳糖苷为半乳糖苷酶显色底物,对硝基苯基-α-D-吡喃葡萄糖苷为α-葡萄糖苷酶显色底物,邻硝基苯基-β-D-葡萄糖醛酸苷为葡萄糖醛酸苷酶显色底物。对上述水解酶酶活检测在疾病诊断,环境监测,生命科学研究等方面有着较为重要的应用。
使用现有的酶底物显色方法在复杂样品或有色样品检验时往往由于样品中其他颜色的干扰,及样品中微粒对光的散射造成检验结果不准确。另外光学比色灵敏度不够高,往往颜色底物产生至微摩尔级别时方能产生较为明显的信号差异,造成检测检出限不够低。
发明内容
为了解决现有技术中存在的问题本申请设计了一种检测硝基苯酚的比色瓶/管。该比色瓶/管对待测溶液中的硝基苯酚类化合物进行富集及长光程双重信号放大,屏蔽了溶液中有色物及颗粒物的干扰,并进一步提高了光学比色检测的灵敏度。因此在环境监测及微生物鉴定等方面有着无可比拟的优势。
实现本发明上述目的所采用的技术方案为:
一种检测硝基苯酚的比色瓶/管,至少包括光源、光电检测器以及盛放有透明或半透明的硝基苯酚富集材料的透明瓶/管体,所述的瓶/管体为以下结构:
瓶/管体的底部为平面,瓶/管体中在所盛放的硝基苯酚富集材料的顶层中心处设置有一个反光片,光源发出的光线穿过底部平面后进入硝基苯酚富集材料,然后被反光片反射后,反射光线穿过底部平面射出,被光电检测器接收;
所述的硝基苯酚富集材料包括主体基材和改性材料,其中所述主体基材用于形成透明或半透明的材料主体,改性材料用于与硝基苯酚类化合物发生相互作用并进行富集,所述主体基材为聚丙烯酰胺类化合物或硅橡胶类化合物,所述改性材料为环糊精类化合物。
所述的环糊精类化合物为α-环糊精、β-环糊精、γ-环糊精及其氨基、羟丙基衍生物,聚(二烯丙基二甲基氯化铵),N,N-二甲基环己胺,四庚基溴化铵,十六烷基三甲基溴化铵中的一种或两种以上的混合物。
所述的主体基材为聚丙烯酰胺改性凝胶,所述的改性材料与主体基材的质量比为1:20~1000。
本发明提供的比色瓶/管在使用时,光由瓶底照入,通过上述硝基苯酚富集材料,材料里的硝基苯酚吸收入射光对应波长的光。光线设计为底部往上小角度照射,照射的硝基苯酚富集材料上方贴有一反光膜,光线反射往下继续通过硝基苯酚富集材料,富集材料里的硝基苯酚继续对应波长的光吸收直至光线由瓶底射出,光线瓶底射出的位置则放置感光元件,对上述光程的吸光度进行定量。
本发明中通过比色瓶/管形状的设计,瓶内光路的设计及瓶/管底硝基苯酚富集材料的引入,对溶液中的硝基苯酚类化合物进行富集及长光程双重信号放大,屏蔽了溶液中有色物及颗粒物的干扰,并进一步提高了光学比色检测的灵敏度。该比色瓶/管配合简单的光源及感光元件,即可对硝基苯酚类有色物质进行高灵敏的检测,在环境监测及微生物鉴定等方面有着无可比拟的优势。
附图说明
图1为聚丙烯酰胺做为瓶/管底透光硝基苯酚富集材料时的比色瓶/管结构图;
图2为图1的俯视图;
图3为碱性磷酸酶底物检测金黄色葡萄球菌生长曲线图;
图4为以β-半乳糖苷酶为生长指示酶的粪大肠菌群生长曲线图;
图5为以使用检测硝基苯酚比色瓶/管进行的碱性磷酸酶检测图。
具体实施方式
下面结合具体实施例对本发明做详细具体的说明,但是本发明的保护范围并不局限于以下实施例。
实施例一比色瓶/管的设计及制作
比色瓶/管上方透光硝基苯酚富集材料由聚丙烯酰胺为基材改性而来,入射光线与瓶身轴线之间夹角使用小角度,比色瓶/管的结构采用图1和图2中进行设计,此时在硝基苯酚富集材料上方中央粘结一反光膜片,膜片覆盖部分区域,不影响溶液中硝基苯酚类化合物的富集。光线照射至膜片处发生反射与入射光呈对称反射出至光电检测探头。
实施例二瓶/管底透光硝基苯酚富集材料制备
主体基材使用使用聚丙烯酰胺改性凝胶,使用丙烯酰胺,甲基丙烯酸甲基丙烯酸十八酯质量比10:1作为聚合单体,聚合单体总质量为1g,加入0.3g甲叉双丙烯酰胺作为交联剂,再加入1g的pH8.8的1.5M的三羟甲基氨基甲烷-盐酸缓冲液,加入10%(W/W)过硫酸铵及10mg四甲基乙二胺,按质量比1/100加入α-环糊精、β-环糊精、γ-环糊精、及其氨基、羟丙基衍生物,聚(二烯丙基二甲基氯化铵),N,N-二甲基环己胺,四庚基溴化铵,十六烷基三甲基溴化铵中的一种或几种,补水至总质量为10g,混匀。取一定量混匀后的加入改性剂的聚丙烯酰胺改性凝胶加入比色瓶/管静置至固化,此处无厚度或剂量控制。凝胶固化后,凝胶表面中央加入一片反光膜片,利用亲水性胶水及膜片自重实现固定。
实施例三利用碱性磷酸酶活的微生物培养比色实验
LB培养基培养金黄色葡萄球菌至O.D.为1,使用无菌水梯度稀释,稀释后的菌液1/10(V/V)接种入含1%4-硝基苯磷酸二钠无菌过滤的2%蛋白胨水溶液的比色瓶,37℃培养,实时检测405nm光吸收,结果如附图3所示。图3中纵坐标为405nm实时吸光度值与初始吸光度值的比值,横坐标为培养检测时间。
实施例四利用半乳糖苷酶活的粪大肠菌群培养比色实验
LB培养基培养大肠埃希氏菌ATCC25922至O.D.为1,使用无菌水梯度稀释,稀释后的菌液1/10(V/V)接种入含1%2-硝基苯-β-D-半乳糖苷无菌过滤的5%蛋白胨水溶液的比色瓶,44.5℃培养,实时检测405nm透光率,结果如附图4所示。图4中纵坐标为405nm实时透光率,横坐标为培养检测时间。
实施例五碱性磷酸酶酶活检测实验
配置20mL溶液加入比色瓶,溶液含50mM三羟甲基氨基甲烷-盐酸(pH 9.3at 25C),1mM MgCl2,0.1mM ZnCl2,1mM亚精胺及10mM 4-硝基苯磷酸二钠,1/100(V/V)加入0.01U/mL碱性磷酸酶,实时检测405nm光吸收,结果如附图5所示。图5中纵坐标为405nm实时吸光度值与初始吸光度值的比值,横坐标为孵育时间。
Claims (3)
1.一种检测硝基苯酚的比色瓶/管,至少包括光源、光电检测器以及盛放有透明或半透明的硝基苯酚富集材料的透明瓶/管体,其特征在于:所述的瓶/管体为以下结构:
瓶/管体的底部为平面,瓶/管体中在所盛放的硝基苯酚富集材料的顶层中心处设置有一个反光片,光源发出的光线穿过底部平面后进入硝基苯酚富集材料,然后被反光片反射后,反射光线穿过底部平面射出,被光电检测器接收;
所述的硝基苯酚富集材料包括主体基材和改性材料,其中所述主体基材用于形成透明或半透明的材料主体,改性材料用于与硝基苯酚类化合物发生相互作用并进行富集,所述主体基材为聚丙烯酰胺类化合物或硅橡胶类化合物,所述改性材料为环糊精类化合物。
2.根据权利要求1所述的检测硝基苯酚的比色瓶/管,其特征在于:所述的环糊精类化合物为α-环糊精、β-环糊精、γ-环糊精及其氨基、羟丙基衍生物、 聚(二烯丙基二甲基氯化铵)、 N,N-二甲基环己胺、 四庚基溴化铵、 十六烷基三甲基溴化铵中的一种或两种以上的混合物。
3.根据权利要求2所述的检测硝基苯酚的比色瓶/管,其特征在于:所述的主体基材为聚丙烯酰胺改性凝胶,所述的改性材料与主体基材的质量比为1:20~1000。
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