阅读说明：本技术 一种二价汞离子免仪器定量检测方法 (Instrument-free quantitative detection method for divalent mercury ions ) 是由 张云 姚茂茂 聂瑾芳 陈意静 于 2019-09-29 设计创作，主要内容包括：本发明公开了一种二价汞离子免仪器定量检测方法。利用Hg<Sup>2+</Sup>诱导纳米金探针表面保护剂脱附后团聚,以介导纸芯片中纳米多孔膜孔道“开-关”状态来选择性调控彩色试剂在纸芯片中流动长度的机制。Hg<Sup>2+</Sup>浓度与彩色试剂流动长度成反比关系。通过使用直尺量取纸芯片中彩色试剂的流动长度,就能实现纳摩尔水平Hg<Sup>2+</Sup>的便携式定量检测。本发明方法具有操作简单、成本低廉、分析快速、不需使用专业分析仪器设备、适于Hg<Sup>2+</Sup>的家庭检测和野外现场分析等突出优点。本方法能直接推广应用于医学诊断、环境监测、食品安全等诸多领域里各类溶液样品中Hg<Sup>2+</Sup>分析物的简单、经济、快速、灵敏、特异的便携式定量检测。(The invention discloses an instrument-free quantitative detection method for divalent mercury ions. Utilization of Hg 2+ Inducing the surface protective agent of the nanogold probe to agglomerate after desorption so as to mediate the on-off state of the pore canal of the nanoporous membrane in the paper chip to selectively regulate and control the flowing length of the color reagent in the paper chip. Hg is a mercury vapor 2+ The concentration is inversely related to the length of the color reagent flow. Nanomolar levels of Hg can be achieved by measuring the flow length of the color reagent in the paper chip using a ruler 2+ The portable quantitative detection. The method has the advantages of simple operation, low cost, quick analysis, no need of using professional analytical instruments and equipment, and suitability for Hg 2+ The method has the outstanding advantages of home detection, field analysis and the like. The method can be directly popularized and applied to Hg in various solution samples in various fields of medical diagnosis, environmental monitoring, food safety and the like 2+ simple, economical, rapid, sensitive, specific portable quantitative detection of analytes。)

1. An instrument-free quantitative detection method for divalent mercury ions is characterized by comprising the following specific steps:

(1) Preparing a round paper micro-area chip and a rectangular paper chip with hydrophobic substance patterning, and respectively dropwise adding a fixed nano-gold probe and a color reagent, so as to align and sequentially stack circular areas in the round paper micro-area chip, the round nano-porous membrane and the rectangular paper chip to prepare a three-dimensional paper chip;

(2) isovolumetrically mixing a divalent mercury ion sample solution and a buffer solution containing high-concentration sodium chloride, dropwise adding the divalent mercury ion sample solution and the buffer solution into the circular paper micro-area chip on the uppermost layer of the three-dimensional paper chip prepared in the step (1), then observing the flowing condition of a color reagent in the rectangular paper chip, measuring the flowing length of the color reagent in the rectangular paper chip by using a ruler after 5 ~ 30 min, wherein the flowing length is inversely related to the concentration of divalent mercury ions, and thus realizing instrument-free quantitative detection of the divalent mercury ions;

The circular paper micro-area chip is in a square shape with the side length of 1.5 ~ 2 cm, the middle part of the chip is a circular hydrophilic paper body with the diameter of 2 ~ 7 mm, and the rest part of the chip is filled with wax or hydrophobic polymer;

the rectangular paper chip is rectangular with the length of 3 ~ 5 cm and the width of 1 ~ 2 cm, the middle part is a circular hydrophilic paper body and a rectangular hydrophilic paper channel which are communicated, and the rest part is filled with wax or hydrophobic polymer, wherein the diameter of the circular hydrophilic paper body is 2 ~ 7 mm, the length of the rectangular hydrophilic paper channel is 2.5 ~ 4.5.5 cm, and the width of the rectangular hydrophilic paper channel is 1 ~ 2 mm;

The nano-gold probe is dropwise fixed in the whole round hydrophilic paper body of the round paper micro-area chip, and a small molecule protective reagent which can ensure that nano-gold is stably dispersed in a high-concentration salt solution is modified on the surface of the nano-gold probe, wherein the particle size of the nano-gold is 10 ~ 30 nm;

The color reagent is dropwise added and fixed in the whole round hydrophilic paper body of the rectangular paper chip, a color blot formed by the color reagent in the round hydrophilic paper body has good water solubility, and a color solution formed after being dissolved by a reaction solution can spontaneously flow to the rectangular hydrophilic paper channel from the round hydrophilic paper body under the action of a capillary tube;

The round nano porous membrane is one of an inorganic porous membrane and an organic porous membrane with the diameter of 2 ~ 7 mm, has good hydrophilicity, and has the average diameter of pore channels of 50 ~ 200 nm;

The buffer solution containing high concentration of sodium chloride has a pH value of 7, wherein the concentration of sodium chloride is 100 ~ 150 mM, and the buffer solution does not contain Hg²⁺A component that precipitates or complexes;

in the step (2), the bivalent mercury ion sample solution and the buffer solution containing high-concentration sodium chloride are mixed and then dripped into the circular paper micro-area at the uppermost layer of the three-dimensional paper chip for reaction, so that bivalent mercury ions and nanogold react to generate gold-amalgam, the probe is caused to agglomerate in the high-concentration salt solution after desorption of the small molecular protective agent on the surface of the probe, the formed large-size compound blocks the pore of the nanoporous membrane in the three-dimensional paper chip, and the volume of the reaction solution flowing to the circular area of the rectangular paper chip at the lowermost layer to dissolve the color reagent is reduced.

Technical Field

The invention belongs to the technical field of nano chemical sensing, and particularly relates to an instrument-free quantitative detection method for divalent mercury ions.

Background

Divalent mercury ion (Hg)²⁺) Is one of heavy metal ions with serious physiological toxicity. Hg is a mercury vapor²⁺Can be accumulated in animals and plants, and gradually enriched by food chain, and its concentration can be thousandsthe increase of the traditional Chinese medicine is tens of thousands or millions, and the traditional Chinese medicine finally threatens human health, for example, the nervous system, the skin mucosa, the urinary system, the reproductive system and the like of the human body can be seriously damaged, and even death can be caused. The standard national sanitary Standard for Drinking Water (GB 5749 and 2006) limits that the mercury content in drinking water must not exceed 0.001 mg/L (about 5 nM). To ensure Hg in food, water and environment²⁺Does not affect the health and life safety of human body, and trace Hg²⁺The quantitative detection of the method is significant. Existing conventional gold standard Hg²⁺The quantitative detection technology mainly comprises inductively coupled plasma mass spectrometry, atomic absorption spectrometry, atomic emission spectrometry, atomic fluorescence spectrometry and the like. However, these quantitative methods have problems of expensive analysis cost, tedious and time-consuming operation steps, and unsuitability for home use and field analysis.

Disclosure of Invention

The invention aims to provide an instrument-free quantitative detection method for divalent mercury ions aiming at the defects of the existing quantitative technology.

The idea of the invention is as follows: in the experiment, Hg is found²⁺Can react with the nano-gold to generate gold-amalgam, which leads to the desorption of the small molecule protective agent modified on the surface of the nano-probe. Then, the nano gold is agglomerated in a high-concentration salt solution, and the formed large-size compound can block nano-size pore channels of the nano porous membrane. Meanwhile, the on-off state of the pore channel of the nano porous membrane can be further used for selectively regulating and controlling the flowing length of the color reagent in the paper chip. Hg is a mercury vapor²⁺The concentration is inversely proportional to the flow length. The flow length of the color reagent in the paper chip is measured by visual observation and a ruler, and the Hg can be used for reading signals (the flow length of the color reagent) instead of expensive and portable professional analytical instruments such as an inductively coupled plasma mass spectrometer, an atomic absorption spectrometer and an atomic fluorescence spectrometer, so that Hg can be realized²⁺The method is simple, low in cost and portable and accurate in quantitative analysis.

The method comprises the following specific steps:

(1) Preparing a round paper micro-area chip and a rectangular paper chip with hydrophobic substance patterns, respectively dropwise adding a fixed nano-gold probe and a color reagent, and aligning and sequentially overlapping round areas in the round paper micro-area chip, the round nano-porous membrane and the rectangular paper chip to obtain the three-dimensional paper chip.

(2) and (2) mixing the divalent mercury ion sample solution and a buffer solution containing high-concentration sodium chloride in equal volume, dropwise adding the mixture into the circular paper micro-area chip on the uppermost layer of the three-dimensional paper chip prepared in the step (1), observing the flow condition of the color reagent in the rectangular paper chip, measuring the flow length of the color reagent in the rectangular paper chip by using a ruler after 5 ~ 30 min, wherein the flow length is inversely related to the concentration of the divalent mercury ions, and thus realizing the instrument-free quantitative detection of the divalent mercury ions.

The round paper micro-area chip is a square with the side length of 1.5 ~ 2 cm, the middle part is a round hydrophilic paper body with the diameter of 2 ~ 7 mm, and the rest part is filled with wax or hydrophobic polymer.

The rectangular paper chip is rectangular with the length of 3 ~ 5 cm and the width of 1 ~ 2 cm, the middle part is a circular hydrophilic paper body and a rectangular hydrophilic paper channel which are communicated, and the rest part is filled with wax or hydrophobic polymer, wherein the diameter of the circular hydrophilic paper body is 2 ~ 7 mm, the length of the rectangular hydrophilic paper channel is 2.5 ~ 4.5.5 cm, and the width of the rectangular hydrophilic paper channel is 1 ~ 2 mm.

The nano-gold probe is dropwise fixed in the whole round hydrophilic paper body of the round paper micro-area chip, and a small molecule protective reagent which can enable nano-gold to be stably dispersed in a high-concentration salt solution is modified on the surface of the nano-gold probe, wherein the particle size of the nano-gold is 10 ~ 30 nm.

The color reagent is dripped and fixed in the whole round hydrophilic paper body of the rectangular paper chip, the color blot formed by the color reagent in the round hydrophilic paper body has good water solubility, and the color solution formed after being dissolved by the reaction solution can spontaneously flow from the round hydrophilic paper body to the rectangular hydrophilic paper channel under the action of capillary.

The round nano porous membrane is one of an inorganic porous membrane and an organic porous membrane with the diameter of 2 ~ 7 mm, has good hydrophilicity, and has the average diameter of a pore channel of 50 ~ 200 nm.

Said containsThe pH of the buffer solution with high concentration of sodium chloride is 7, wherein the concentration of sodium chloride is 100 ~ 150 mM, and the buffer solution does not contain Hg²⁺A component in which precipitation or complexation occurs.

In the step (2), a bivalent mercury ion sample solution and a buffer solution containing high-concentration sodium chloride are mixed in an isometric mode and then are dripped into the circular paper micro-area chip on the uppermost layer of the three-dimensional paper chip to react, bivalent mercury ions and nanogold react to generate gold-amalgam, so that the probe is agglomerated in a high-concentration salt solution after being desorbed on the surface of the probe, and the formed large-size compound is used for reducing the volume of a reaction solution which flows to the circular area of the rectangular paper chip on the lowermost layer to dissolve a color reagent and flows in the rectangular channel by blocking the pore channel of the nano porous membrane on the middle layer of the three-dimensional paper chip.

Compared with the prior conventional gold standard Hg²⁺Compared with the quantitative detection method, the invention has the outstanding advantages that:

(1) the cost of porous paper base materials such as filter paper, chromatographic paper, nitrocellulose membrane and the like is low, the paper chips can be prepared in batches, and the size of a single chip is small and exquisite (centimeter level, convenient to store, carry and use).

(2) detection of Hg with paper chip²⁺The operation process is very simple, and only two steps are involved, namely the dropwise addition of the sample solution and the measuring of the flowing length of the color reagent.

(3) Quantitative signal reading can be carried out only by measuring the flowing length of the color reagent in the paper chip by the ruler, so that Hg can be realized while the analysis cost is greatly reduced²⁺Home testing and field analysis.

(4) The method can be directly popularized and applied to Hg in various solution samples in the fields of medical diagnosis, environmental monitoring, food safety and the like²⁺Simple, economical, rapid, sensitive and specific portable quantitative detection of analytes.

drawings

FIG. 1 is a schematic diagram of the instrument-free quantitative detection method of divalent mercury ions according to the present invention.

The labels in the figure are: 1-circular paper micro-area chip (1)-1-round hydrophilic paper, 1-2-paraffin or hydrophobic polymer); 2-small molecule protection reagent modified nano gold probe (2-1-nano gold, 2-2-small molecule protection reagent); 3-circular nanoporous membrane; 4-rectangular paper chips (4-1-round hydrophilic paper body, 4-2-rectangular hydrophilic paper channel, 4-3-paraffin or hydrophobic polymer); 5-a color agent; 6-a three-dimensional paper chip (top view) assembled by overlapping the round paper micro-area chip fixed with the reagent, the round nano porous membrane and the rectangular paper chip fixed with the reagent; 7-equal volume of mixed solution of sample solution and buffer solution; 8-Hg²⁺(ii) a 9-nanogold-amalgam agglomerate complex; 10-flow length of color reagent solution in the rectangular channel of the rectangular paper chip (three-dimensional paper chip cross-sectional view).

FIG. 2 shows Hg used in example 1 of the present invention²⁺The instrument-free quantitative detection method is used for respectively detecting a blank water sample (blank) without analyte ions and 40 nM Hg²⁺Water sample, other 15 metal ion water samples (ion concentration is 10 mu M), and metal ions and 40 nM Hg²⁺comparison of the signal values (length, flow length of the color reagent red ink) obtained for the mixed water samples. Error bars in the figures are the standard deviation of the results of three replicates. The time for measuring the flowing length of the red ink is 25 min after the water sample is dripped.

FIG. 3 shows Hg used in example 2 of the present invention²⁺Instrumentally-free quantitative detection method for analyzing a series of Hg with concentration range of 2.5 ~ 640 nM²⁺Length of water sample and Hg²⁺Log value of concentration (Log [ Hg)²⁺]) The operating curve in between. Error bars in the figures are the standard deviation of the results of three replicates. The time for measuring the flowing length of the red ink is 25 min after the water sample is dripped.

Detailed Description

The following examples further illustrate the invention but are not intended to limit the invention thereto.