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CN-122016687-A - Silver metallized microbubbles in Hg2+Detection and application of alloy microbubble nano enzyme prepared by detection in detection of reducing drugs

CN122016687ACN 122016687 ACN122016687 ACN 122016687ACN-122016687-A

Abstract

The invention discloses an application of silver metallized microbubbles in Hg 2+ detection and an alloy microbubble nano enzyme prepared by the silver metallized microbubbles in detection of a reducing drug, belonging to the technical field of functional material preparation and nano enzyme sensing. Aims at solving the problems of high background signal, poor reproducibility and poor stability of the existing carrier nano enzyme. The method comprises the steps of mixing silver metallized microbubbles with TMB solution, adding a sample to be detected, standing until alloy microbubble nano enzyme floats to the top end of the solution after reaction, measuring absorbance by adopting an enzyme-labeled instrument, or shooting a photo of the reaction solution by using handheld electronic equipment and extracting RGB values, and calculating to obtain Hg 2+ concentration in the sample to be detected according to a pre-established concentration standard curve. The method utilizes the buoyancy of the micro-bubbles to realize spontaneous separation, and can complete quantitative analysis by combining an enzyme-labeled instrument or handheld electronic equipment, and is mainly used for on-site rapid detection of Hg 2+ .

Inventors

  • QIN XIAOJIE
  • YANG FAN
  • Liao Xiyue
  • ZHANG YUYUAN
  • Meng Biyu

Assignees

  • 广西医科大学

Dates

Publication Date
20260512
Application Date
20260415

Claims (10)

  1. 1. A colorimetric detection method of Hg 2+ , comprising the steps of: Mixing silver metallized microbubbles with TMB solution containing H 2 O 2 , adding a sample to be tested containing Hg 2+ , and reacting at room temperature for 10-15 min; Standing until the alloy microbubble nano enzyme floats to the top end of the solution after the reaction; measuring absorbance at 652 nm by using an enzyme-labeled instrument, or taking a picture of the reaction solution by using a handheld electronic device and extracting RGB values; And calculating to obtain the Hg 2+ concentration in the sample to be detected according to a pre-established absorbance-Hg 2+ concentration standard curve or an R/B value-Hg 2+ concentration standard curve.
  2. 2. The colorimetric detection method of Hg 2+ according to claim 1, wherein the preparation of silver metallized microbubbles comprises the steps of: S1, dispersing hollow glass microbubbles in a piranha solution, stirring at room temperature for reaction of 0.5-2 h, washing with ultrapure water until washing solution is neutral, and vacuum drying to obtain hollow glass microbubbles with activated surfaces; s2, adding the hollow glass microbubbles with the surface activated into a polyethyleneimine solution with the concentration of 1-8 mg/mL, carrying out rotary mixing reaction at room temperature for 0.5-12 h, standing until the hollow glass microbubbles float to the top end of the solution after the reaction, sucking the lower layer solution, washing by using ultrapure water, and carrying out vacuum drying to obtain the hollow glass microbubbles wrapped by polyethyleneimine; s3, adding hollow glass microbubbles wrapped by polyethyleneimine into a buffer solution containing 1-1000 pmol single-stranded DNA rich in cytosine bases, rotating and mixing at room temperature for reaction of 0.5-4 h, standing until the compound floats to the top end of the solution after the reaction, sucking the lower-layer solution, and washing by using ultrapure water to obtain an intermediate product loaded with the single-stranded DNA; S4, mixing the intermediate product loaded with single-stranded DNA with silver nitrate solution with the concentration of 0.1-50 mM, carrying out rotary mixing reaction at room temperature for 10-200 min, standing until the compound floats to the top end of the solution after the reaction, absorbing the lower layer solution and washing by using ultrapure water to obtain hollow glass microspheres loaded with silver ions; And S5, mixing the hollow glass microspheres loaded with silver ions with sodium borohydride solution with the concentration of 0.1-50 mM, carrying out vortex reaction for 10-120: 120 min, standing until the product floats to the top end of the solution after the reaction, sucking the lower solution, and washing with ultrapure water to obtain the silver metallized microbubbles.
  3. 3. The colorimetric detection method of Hg 2+ according to claim 1, wherein the step of taking a photograph of the reaction solution and extracting RGB values using a handheld electronic device comprises: Placing the reacted porous plate on a shooting bracket, wherein a standard colorimetric strip is preset in a non-detection area of the porous plate, and the standard colorimetric strip comprises at least one color block with a known RGB value; using a handheld electronic device to shoot images containing all the reaction holes and the standard colorimetric strips simultaneously in the same view field; extracting R, G, B values of the central area of each reaction hole in the image, and extracting R, G, B values of the standard colorimetric strips; Performing color correction on the image by using the corresponding relation between the known RGB value and the extracted value of the standard colorimetric strip to obtain a corrected RGB value of the reaction hole; And calculating an R/B ratio based on the corrected RGB value of the reaction hole, and calculating Hg 2+ concentration in the sample to be detected according to a pre-established R/B value-Hg 2+ concentration standard curve.
  4. 4. The colorimetric detection method of Hg 2+ according to claim 3, wherein the non-detection region of the multi-well plate is provided with a plurality of micro-scale color patches, the micro-scale color patches being in one-to-one correspondence with and disposed adjacent to the reaction well edges of the multi-well plate; when the handheld electronic equipment is used for shooting images, the image area of each reaction hole and the image area of the corresponding miniature standard color block are captured at the same time; Extracting R, G, B values of the central area of each reaction hole, and simultaneously extracting R, G, B values of the miniature standard color block corresponding to the reaction hole; And calculating R/B ratio based on the RGB values corrected by each reaction hole, and calculating Hg 2+ concentration in the sample to be detected according to a pre-established R/B value-Hg 2+ concentration standard curve.
  5. 5. The colorimetric detection method of Hg 2+ according to claim 4, wherein the non-detection region of the multi-well plate is further provided with an information encoding region in which accurate color value data of each micro-scale standard color patch is stored; Before using the handheld electronic equipment to shoot an image, firstly reading an information coding area through the handheld electronic equipment to acquire an accurate color value of each miniature standard color block; after the RGB values of the micro standard color blocks corresponding to the reaction holes are extracted, the RGB values of the corresponding reaction holes are respectively and independently corrected by utilizing the corresponding relation between the accurate color values and the extracted values of the micro standard color blocks, so as to obtain R, G, B values after correction of each reaction hole; And calculating an R/B ratio based on the RGB value corrected by each reaction hole, and calculating the Hg 2+ concentration in the sample to be detected according to a pre-established R/B value-Hg 2+ concentration standard curve.
  6. 6. A colorimetric detection method of a reducing drug, comprising the steps of: Mixing silver metallized micro-bubbles with TMB solution containing H 2 O 2 and Hg 2+ solution, and reacting at room temperature for 10-15 min so as to combine Hg 2+ with the silver metallized micro-bubbles to generate alloy micro-bubble nano-enzyme with peroxidase-like activity; Adding a sample to be detected containing a reducing drug, and reacting at room temperature for 10-15 min; Standing until the alloy microbubble nano enzyme floats to the top end of the solution after the reaction; measuring absorbance at 652 nm by using an enzyme-labeled instrument, or taking a picture of the reaction solution by using a handheld electronic device and extracting RGB values; And calculating according to a pre-established absorbance-drug concentration standard curve or an R/B value-drug concentration standard curve to obtain the concentration of the reducing drug in the sample to be detected.
  7. 7. The colorimetric detection method of Hg 2+ according to claim 1, wherein the detection of Hg 2+ is achieved by a portable colorimetric sensor comprising a signal reporting unit, a chromogenic substrate, a reaction vessel and a signal reading unit; The signal reporting unit is silver metallized micro bubbles; the chromogenic substrate is a mixed solution containing TMB and H 2 O 2 ; the reaction container is a porous plate for containing silver metallized microbubbles, a chromogenic substrate and a sample to be tested; The signal reading unit is a handheld electronic device and is used for shooting an image of the reaction solution and extracting RGB values for quantitative analysis.
  8. 8. An alloy microbubble nano enzyme is characterized by taking silver metallized microbubbles prepared by the method in claim 2 as a raw material, and is prepared by mixing silver metallized microbubbles with TMB solution containing H 2 O 2 and Hg 2+ solution with the concentration of 10-100 mu M, reacting at room temperature for 10-15-min, and washing with ultrapure water after the reaction to obtain the alloy microbubble nano enzyme.
  9. 9. Use of silver metallized micro-bubbles according to claim 2 in Hg 2+ detection.
  10. 10. The use of alloy microbubble nano-enzyme according to claim 8 in detection of a reducing drug, wherein the reducing drug is one or more of ascorbic acid, N-acetylcysteine, D-penicillamine, dimercaptosuccinic acid, vitamin K, edaravone, resveratrol and curcumin.

Description

Application of silver metallized microbubble in Hg 2+ detection and alloy microbubble nano enzyme prepared by silver metallized microbubble in detection of reducing medicine Technical Field The invention belongs to the technical field of functional material preparation, the technical field of nano enzyme sensing and the technical field of portable rapid detection, and particularly relates to application of silver metallized microbubbles in Hg 2+ detection and alloy microbubble nano enzyme prepared by the silver metallized microbubbles in detection of a reducing drug. Background Mercury ions are widely used as high-toxicity heavy metal pollutants in environmental water and food matrixes, and the residual levels of the mercury ions directly relate to ecological safety and human health, so that a convenient and accurate detection method needs to be established. At present, conventional detection means for mercury ions are mostly dependent on large-scale precise instruments such as inductively coupled plasma mass spectrometry and atomic absorption spectrometry, and the like, and the detection precision is high, but the problems of high equipment acquisition cost, complex operation flow, long detection period, dependence on professional technicians and the like exist, so that the detection means are difficult to adapt to actual requirements of on-site rapid screening and high-throughput detection of large-scale samples, and are particularly inconvenient to popularize and apply in basic scenes with limited resources. Colorimetric sensing technology based on nano materials is gradually applied to mercury ion detection research in recent years, and is attracting attention because of the advantages of visual response and relatively simple operation. The nanoenzyme colorimetric sensing system becomes a research hotspot in the field by virtue of the characteristics of good material stability and relatively low preparation cost. However, the existing nano enzyme sensing system still has obvious defects in practical application that most nano enzyme materials are in a uniform dispersion state in a detection reaction system, and after the reaction is finished, the nano enzyme and the reaction system are separated through additional steps such as centrifugation and filtration, so that stable signal reading can be performed, the complexity of detection operation is increased, material loss or change of the state of the reaction system can be caused in the separation process, and the accuracy of a detection result and the reproducibility among batches are further influenced. The hollow glass microbubbles are used as a low-density microcarrier, have natural floating characteristics, can realize spontaneous separation after reaction theoretically through buoyancy, and are hopeful to simplify the separation step in the detection process. However, the hollow glass microbubbles have smooth surfaces and strong chemical inertness, and it is difficult to directly realize uniform and stable loading of the functional identification component and the signal response component. The existing surface functionalization modification technology for hollow glass microbubbles, in particular to a surface metallization modification method, often faces the problems of uneven metal component loading, weak binding force with a carrier, easy falling and agglomeration and the like, so that the modified microbubble materials have larger performance difference among batches, insufficient signal response stability, are difficult to meet the requirement of mercury ion conventional detection on the consistency of the material performance, and restrict the practical application of the materials in the field of sensing detection. Disclosure of Invention It is an object of the present invention to solve at least the above problems and to provide at least the advantages to be described later. The invention also aims to provide a colorimetric detection method of Hg 2+, which can realize selective identification of a target object based on the specific amalgamation reaction of silver and Hg 2+, and can complete on-site rapid quantitative detection of Hg 2+ under the condition of no need of a large instrument by combining absorbance measurement or RGB analysis of handheld electronic equipment (such as a smart phone or a tablet personal computer and the like), thereby simplifying the detection flow, reducing the operation threshold and adapting to the actual application requirements of scenes such as water quality monitoring, environment screening and the like. To achieve these objects and other advantages and in accordance with the purpose of the invention, there is provided a colorimetric detection method of Hg 2+, comprising the steps of: Mixing silver metallized microbubbles with TMB solution containing H 2O2, adding a sample to be tested containing Hg 2+, and reacting at room temperature for 10-15 min; Standing until the alloy microbubble nano enzyme floats