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Ampere-type biosensor for detecting benzopyrene as well as preparation method and application thereof

A biosensor, benzopyrene technology, applied in the direction of material electrochemical variables, etc., can solve problems such as difficulties in electron transfer, and achieve the effects of easier control of operating parameters, good reproducibility and stability, and simple operation

Inactive Publication Date: 2010-10-27
SOUTH CENTRAL UNIVERSITY FOR NATIONALITIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, on the unmodified electrode, the direct electron transfer of P450 enzyme is very difficult, because the heme is embedded in the protein and it is difficult to interact with the electrode surface. Various electrode modification techniques are used to realize the direct electrochemistry of P450.

Method used

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  • Ampere-type biosensor for detecting benzopyrene as well as preparation method and application thereof
  • Ampere-type biosensor for detecting benzopyrene as well as preparation method and application thereof
  • Ampere-type biosensor for detecting benzopyrene as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Example 1: Preparation of an amperometric biosensor for detecting benzopyrene: add 10 mg of nano-montmorillonite to 1 mL of distilled water and continue to stir for 12 hours. After standing for 5 hours, centrifuge in a centrifuge at 3500 rpm for 30 minutes and take the supernatant to obtain a nano Montmorillonite suspension, 0.5 mg of dihexadecyl phosphoric acid was added to 0.5 ml of nano-montmorillonite suspension, and ultrasonically dispersed in a 700w ultrasonic instrument for 16 hours to obtain a dispersion of nano-montmorillonite and dihexadecyl phosphoric acid ; The cytochrome p4501A1 of 1mg is directly dissolved in the phosphate buffer solution of 1mL pH7.0 to obtain the cytochrome p4501A1 liquid; Crack the graphite electrode and dry it at room temperature to obtain an amperometric biosensor for detecting benzopyrene.

[0017] Put the resulting cytochrome p4501A1 / nano-montmorillonite-dicetyl phosphate-modified pyrolytic graphite electrode in a pH7.0 phosphate bu...

Embodiment 2

[0019] Example 2: Preparation of an amperometric biosensor for detecting benzopyrene: Add 10 mg of nano-montmorillonite to 1 mL of distilled water and continue to stir for 12 hours. After standing for 3 hours, centrifuge in a centrifuge at 3500 rpm for 30 minutes and take the supernatant to obtain nano Montmorillonite suspension, 2 mg of dihexadecyl phosphoric acid was added to 1 ml of nano-montmorillonite suspension, and ultrasonically dispersed in a 700W ultrasonic instrument for 12 hours to obtain a dispersion of nano-montmorillonite and dihexadecyl phosphoric acid ; Dissolve 1 mg of cytochrome p4501A1 directly in 1 mL of Tris-HCl buffer solution of pH 7.0 to obtain cytochrome p4501A1 solution; mix the dispersion with equal volumes of cytochrome p4501A1 solution, take 5 microliters of the mixed solution and apply it on a glass carbon electrode, and dried at room temperature to obtain an amperometric biosensor for detecting benzopyrene.

Embodiment 3

[0020] Example 3: Preparation of amperometric biosensor for detecting benzopyrene: Add 15 mg of nano-montmorillonite to 1 mL of distilled water and continue to stir for 10 hours. After standing for 5 hours, centrifuge in a centrifuge at 5000 rpm for 20 minutes and take the supernatant to obtain nano Montmorillonite suspension, 1.0 mg of dihexadecyl phosphoric acid was added to 0.5 ml of nano-montmorillonite suspension, and ultrasonically dispersed in a 700W ultrasonic instrument for 15 hours to obtain a dispersion of nano-montmorillonite and dihexadecyl phosphoric acid ; Dissolve 1mg of cytochrome p4501A1 directly in 1mL of pH 5.0 phosphate buffer to obtain cytochrome p4501A1 solution; mix the dispersion with equal volumes of cytochrome p4501A1, take 5 microliters of the mixed solution and apply it on the ordinary spectrum graphite The electrode is dried at room temperature to obtain an amperometric biosensor for detecting benzopyrene.

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Abstract

The invention relates to an ampere-type biosensor for detecting benzopyrene as well as a preparation method and application thereof. The preparation method of the ampere-type biosensor for detecting the benzopyrene comprises the following steps of: adding 10-20mg of nano montmorillonoid in 1-2mL of distilled water, continuously stirring for 10-15h, standing for 3-5h, and then centrifuging for 20-30min in a centrifuge with 3000-5000 revolutions per minute, taking supernatant to obtain a nano montmorillonoid suspension solution, adding 0.5-2mg of dihexadecyl hydrogen phosphate into 0.5-1ml of the nano montmorillonoid suspension solution and ultrasonically dispersing for 12-16h in a 700W ultrasonic instrument to obtain a nano montmorillonoid and dihexadecyl hydrogen phosphate dispersing solution; directly dissolving 1mg of cytochrome p4501A1 in 1mL of buffer solution with the pH value of 5.0-8.0 to obtain a cytochrome p4501A1 solution; and mixing the dispersing solution with the cytochrome p4501A1 solution in the same volume, taking 5mL of mixing solution to be coated on a substrate, and airing at room temperature to obtain the finished product. The prepared biosensor can not only realize direct electron transfer of the cytochrome p4501A1 and ensure that the bioactivity, the stability and the reproducibility are better, but also has simple operation and easily-controlled conditions.

Description

technical field [0001] The invention relates to an ampere-type benzopyrene biosensor, a preparation method and an application, which are suitable for direct electrochemical research of cytochrome p4501A1 and actual detection of benzopyrene. Background technique [0002] Cytochrome P450 is a heme-containing enzyme that can metabolize many phase I enzyme reactions. Cytochrome P4501A1 (CYP1A1) is the most important metabolic enzyme in the activation process of benzopyrene. For a long time, cytochrome P4501A1 has been used to detect benzopyrene in vitro, but cytochrome P450 reductase is required to provide the electrons required for the redox reaction of cytochrome P4501A1. Now people are interested in the research of the third generation of biosensors, that is, the direct electrochemical research of enzymes. The electrodes can directly provide electrons for P450 to make P450 play a catalytic role without other electron donors. However, direct electron transfer of P450 enzymes ...

Claims

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Application Information

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IPC IPC(8): G01N27/26
Inventor 吴云华
Owner SOUTH CENTRAL UNIVERSITY FOR NATIONALITIES
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