An electrochemical detection method for vitamin a detection

Abstract

The present application relates to the technical field of VA detection, in particular to an electrochemical detection method for vitamin A detection, comprising the following steps: preparing an electro-oxidation modified glassy carbon electrode; treating VA retinol with perchloric acid to acidify retinol into retinoic acid; using the method of SWV to optimize the buffer solution formula, and finally realizing high sensitivity detection of VA; beneficial effects are that: the electrochemical detection method for vitamin A detection proposed in the present application oxidizes retinol into retinoic acid by using the oxidant perchloric acid, the pretreatment process is simple, and the detection sensitivity can be greatly improved; the optimized reagent formula has low cost and good detection effect; meanwhile, the present research places the oxidant such as perchloric acid in the buffer solution, and reacts with VA in the buffer solution, greatly saving the pretreatment step; the electrochemical detection method using SWV optimizes parameters and improves detection sensitivity.

Description

Technical Field

[0001] This invention relates to the field of vitamin A detection technology, specifically to an electrochemical detection method for vitamin A detection. Background Technology

[0002] Natural vitamin A, also known as retinol, is an unsaturated monohydric alcohol with a lipid ring. It is a fat-soluble vitamin that can prevent night blindness and vision loss, and maintain the health of the surface of tissues or organs. Vitamin A plays a very important physiological role in maintaining vision, immune function, embryonic development, and inhibiting tumors.

[0003] In the existing technology, it is crucial to strengthen the detection of vitamin A in the human body. Currently, there are various methods for detecting vitamin A, including fluorescence assay, gas chromatography, liquid chromatography, liquid chromatography-tandem mass spectrometry, and electrochemical analysis. Among them, electrochemical methods are widely used due to their simple operation and convenient equipment.

[0004] However, existing electrochemical methods for detecting VA have drawbacks such as high detection limits and high CV values, as well as problems such as complex electrode preparation and high detection limits. Summary of the Invention

[0005] The purpose of this invention is to provide an electrochemical detection method for vitamin A detection, in order to solve the problems mentioned in the background art.

[0006] To achieve the above objectives, the present invention provides the following technical solution: an electrochemical detection method for vitamin A detection, the electrochemical detection method comprising the following steps: Preparation of electro-oxidation modified glassy carbon electrodes; VA retinol was treated with perchloric acid to acidify it into retinoic acid; The buffer solution formulation was optimized using the SWV method, ultimately achieving highly sensitive detection of VA.

[0007] Preferably, when preparing the electro-oxidative modified glassy carbon electrode, the glassy carbon electrode is polished with 0.05 μm-1 μm alumina powder for 1-3 minutes in sequence, and then ultrasonically cleaned with pure water, anhydrous ethanol, and pure water in sequence for 1-3 minutes.

[0008] Preferably, the glassy carbon electrode, after ultrasonic treatment, is thoroughly cleaned. A platinum wire electrode is used as the counter electrode, and a saturated calomel electrode or a silver-silver chloride electrode is used as the reference electrode. Cyclic voltammetry is performed in 0.01 mol / L-1 mol / L potassium ferricyanide, 0.01 mol / L-1 mol / L potassium chloride, and 0.01 mol / L-1 mol / L PBS solutions until the electrode stabilizes. The initial electrode potential is -1 to 0.2 V, and the final electrode potential is 0.8 to 1.2 V. The scan rate is 20-200 mV / s, and the number of scan cycles is 10-60. Cyclic voltammetry is performed until the electrode stabilizes, thus successfully preparing the electro-oxidation modified electrode.

[0009] Preferably, perchloric acid is used to treat VA retinol. When acidifying retinol into retinoic acid, perchloric acid oxidant is used to oxidize retinol into retinoic acid. At the same time, the perchloric acid oxidant is placed in a buffer solution and reacts with retinol in the buffer solution to produce retinoic acid.

[0010] Preferably, the perchloric acid oxidant comprises the following components: 950 μL phosphate solution + 950 μL organic reagent solution + 100 μL 0.01 mol / L-10 mol / L strong oxidant. The phosphate solution can be replaced by citrate buffer solution.

[0011] Preferably, the phosphate buffer solution formulation is as follows: a 0.001 mol / L to 1 mol / L potassium dihydrogen phosphate solution and a 0.001 mol / L to 1 mol / L dipotassium hydrogen phosphate solution are mixed in a ratio of 1:9 to 9:1 to form a phosphate solution. The dihydrogen phosphate is a mixture of potassium dihydrogen phosphate, sodium dihydrogen phosphate, and potassium dihydrogen phosphate in any molar ratio; the dihydrogen phosphate is a mixture of dipotassium hydrogen phosphate, disodium hydrogen phosphate, and dipotassium hydrogen phosphate in any molar ratio. The citrate buffer solution formulation is as follows: a 0.001 mol / L to 0.9 mol / L citric acid solution and a 0.001 mol / L to 0.9 mol / L sodium citrate solution are mixed in any molar ratio to form a citrate buffer solution.

[0012] Preferably, the organic reagent dimethylformamide is ① aromatic hydrocarbons: benzene, toluene, xylene; ② aliphatic hydrocarbons: pentane, hexane, octane; ③ alicyclic hydrocarbons: cyclohexane, cyclohexanone, toluenecyclohexanone; ④ halogenated hydrocarbons: chlorobenzene, dichlorobenzene, dichloromethane; ⑤ alcohols: methanol, ethanol, isopropanol; ⑥ ethers: diethyl ether, propylene oxide; ⑦ esters: methyl acetate, ethyl acetate, propyl acetate; ⑧ ketones: acetone, methyl butyl ketone, methyl isobutyl ketone; ⑨ glycol derivatives: ethylene glycol monomethyl ether, ethylene glycol monoethyl ether, ethylene glycol monobutyl ether; ⑩ others: acetonitrile, pyridine, phenol.

[0013] Preferably, the strong oxidant is perchloric acid or perchlorate. The perchlorate is composed of one or more of sodium perchlorate, potassium perchlorate, ammonium perchlorate, lithium perchlorate, magnesium perchlorate, and silver perchlorate. Alternatively, it can be one or a mixture of strong oxidants of the same concentration, such as KMnO4, KClO3, H2SO4, HCl, HNO3, MnO2, and FeCl3.

[0014] Preferably, the perchloric acid oxidant oxidizes retinol to retinoic acid in 90-150 seconds. After adding the sample to be tested to the buffer solution, stir for 30-45 seconds, let stand for 60-105 seconds, and then perform electrochemical detection.

[0015] Preferably, when optimizing the buffer solution formulation using the SWV method, the electrochemical detection method of SWV is adopted, and the electrochemical parameters are as follows: initial electrode potential of the instrument: -0.1-0.3V; termination electrode potential: 1.1-1.3V; settling time: 2-20s; potential increment: 0.001-0.01V; amplitude: 0.005-0.1V; frequency: 50-150Hz; enrichment voltage: 0.1-0.6V; enrichment time: 80-220s.

[0016] Compared with the prior art, the beneficial effects of the present invention are: The electrochemical detection method for vitamin A proposed in this invention oxidizes retinol to retinoic acid using perchloric acid as an oxidant. This simplifies the pretreatment process and significantly improves detection sensitivity. The optimized reagent formulation is cost-effective and provides good detection results. Furthermore, this study places oxidants such as perchloric acid in a buffer solution, allowing them to react with vitamin A in the buffer solution, greatly reducing pretreatment steps. The electrochemical detection method using SWV optimizes parameters and enhances detection sensitivity. Attached Figure Description

[0017] Figure 1 A schematic diagram illustrating the linear relationship between VA concentration and current in this invention. Detailed Implementation

[0018] To make the objectives, technical solutions, and advantages of the present invention clear and complete, the embodiments of the present invention will be further described in detail below with reference to the accompanying drawings. It should be understood that the specific embodiments described herein are only some, not all, embodiments of the present invention, and are merely illustrative of the embodiments of the present invention. They are not intended to limit the embodiments of the present invention. All other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0019] Example 1 This invention provides a technical solution: an electrochemical detection method for vitamin A detection, the electrochemical detection method comprising the following steps: Prepare an electro-oxidative modified glassy carbon electrode. Grind the glassy carbon electrode sequentially with 0.05 μm-1 μm alumina powder for 1-3 minutes, then ultrasonically clean it sequentially with pure water, anhydrous ethanol, and pure water for 1-3 minutes. After cleaning the ultrasonically treated glassy carbon electrode, use a platinum wire electrode as the counter electrode and a saturated calomel electrode or a silver-silver chloride electrode as the reference electrode. Cyclic voltammetry is performed in 0.01 mol / L-1 mol / L potassium ferricyanide, 0.01 mol / L-1 mol / L potassium chloride, and 0.01 mol / L-1 mol / L PBS solutions until the electrode stabilizes. The initial electrode potential is -1 to 0.2 V, and the final electrode potential is 0.8 to 1.2 V. The scan rate is 20-200 mV / s, and the number of scan cycles is 10-60. Cyclic voltammetry is performed until the electrode stabilizes, indicating successful preparation of the electro-oxidative modified electrode. Retinol (VA) is treated with perchloric acid to acidify it into retinoic acid. Then, the perchloric acid oxidant is used to oxidize the retinol to retinoic acid. Simultaneously, the perchloric acid oxidant is placed in a buffer solution, where it reacts with the retinol to produce retinoic acid. The perchloric acid oxidant comprises: 950 μL phosphate solution + 950 μL organic reagent solution + 100 μL 0.01 mol / L-10 mol / L strong oxidant. The phosphate buffer solution consists of: 0.001 mol / L-1 mol / L potassium dihydrogen phosphate solution and 0.001 mol / L-1... A mol / L dipotassium hydrogen phosphate solution is mixed in a ratio of 1:9 to 9:1 to form a phosphate solution. The dihydrogen phosphate is a mixture of potassium dihydrogen phosphate, sodium dihydrogen phosphate, or any molar ratio of potassium dihydrogen phosphate and sodium dihydrogen phosphate. The dihydrogen phosphate is a mixture of dipotassium hydrogen phosphate, disodium hydrogen phosphate, or any molar ratio of dipotassium hydrogen phosphate and disodium hydrogen phosphate. The organic reagent dimethylformamide is selected from: ① aromatic hydrocarbons: benzene, toluene, xylene; ② aliphatic hydrocarbons: pentane, hexane, octane; ③ alicyclic hydrocarbons: cyclohexane, cyclohexanone, toluenecyclohexanone; ④ halogenated hydrocarbons: chlorobenzene, dichlorobenzene, dichloromethane; ⑤ alcohols: methyl... Alcohols, ethanol, isopropanol; ⑥ Ethers: diethyl ether, propylene oxide; ⑦ Esters: methyl acetate, ethyl acetate, propyl acetate; ⑧ Ketones: acetone, methyl butyl ketone, methyl isobutyl ketone; ⑨ Glycol derivatives: ethylene glycol monomethyl ether, ethylene glycol monoethyl ether, ethylene glycol monobutyl ether; ⑩ Others: acetonitrile, pyridine, phenol; Strong oxidizing agents are perchloric acid and perchlorate. Perchlorate is composed of one or more of sodium perchlorate, potassium perchlorate, ammonium perchlorate, lithium perchlorate, magnesium perchlorate, and silver perchlorate, or it can be KMnO4, KClO3, H2SO4, HCl, HNO3, Mn One or more of the strong oxidants, such as O2 and FeCl3, are used; the perchloric acid oxidant oxidizes retinol to retinoic acid in 90-150 seconds. After adding the sample to be tested to the buffer solution, stir for 30-45 seconds, let stand for 60-105 seconds, and then perform electrochemical detection. The phosphate solution can be replaced with a citrate buffer solution. The citrate buffer solution formula is: 0.001 mol / L-0.9 mol / L citric acid solution and 0.001 mol / L-0.9 mol / L sodium citrate solution are mixed in any molar ratio to form the citrate buffer solution. The buffer solution formulation was optimized using the SWV method to achieve highly sensitive detection of VA. The electrochemical detection method using SWV was employed, with the following electrochemical parameters: initial electrode potential -0.1-0.3V, termination electrode potential 1.1-1.3V, settling time 2-20s, potential increment 0.001-0.01V, amplitude 0.005-0.1V, frequency 50-150Hz, enrichment voltage 0.1-0.6V, and enrichment time 80-220s.

[0020] Example 2 Based on Example 1, the electrochemical detection method for vitamin A detection is implemented as follows: (1) Electrode treatment: The glassy carbon electrode was polished with 0.05 μm alumina powder for 3 minutes, and then ultrasonically cleaned with pure water, anhydrous ethanol, and pure water for 2 minutes in sequence. After ultrasonic treatment, the glassy carbon electrode was cleaned. A platinum wire electrode was used as the counter electrode and a saturated calomel electrode was used as the reference electrode. Cyclic voltammetry was performed in 0.1 mol / L potassium ferricyanide and 0.2 mol / L potassium chloride solutions until the electrode stabilized. The initial electrode potential was -0.2 V, the final electrode potential was 0.8 V, the scanning speed was 100 mV / s, and the number of scans was 50. The electro-oxidation modified electrode was successfully prepared when the electrode stabilized.

[0021] (2) Different reagent formulations: 950ul phosphate solution + 950ul dimethylformamide solution + 100ul 0.7mol / L perchlorate phosphate buffer solution formulation: 0.1mol / L potassium dihydrogen phosphate solution and 0.15mol / L dipotassium hydrogen phosphate solution are mixed in the proportions shown in the table below to form a phosphate mixed solution; Table 1. Phosphate Mixture Proportioning Table The results of testing VA at 10 μmol / L are shown in the table below: Table 2 Test Results (3) Different SWV parameters: The initial electrode potential of the instrument is 0.3V, the final electrode potential is 1.1V, and the rest time is 2s. The different SWV parameters are shown in the table below: Table 3 SWV Parameter Table The results of testing VA at 10 μmol / L are shown in the table below: Table 4. Test results of VA at 10 μmol / L (4) The following buffer solution formula is used: 950ul phosphate solution + 950ul dimethylformamide solution + 100ul 1mol / L perchloric acid. The phosphate buffer solution formula is: 0.1mol / L potassium dihydrogen phosphate solution and 0.15mol / L dipotassium hydrogen phosphate solution are mixed in a ratio of 3:7 to form a phosphate solution.

[0022] The oxidation time for perchloric acid to oxidize retinol to retinoic acid is 90s (stirring for 30s and letting stand for 60s). That is, after adding the sample to be tested to the buffer solution, stir for 30s and let stand for 60s before electrochemical detection.

[0023] SWV parameters: initial electrode potential 0.3V, termination electrode potential 1.1V, rest time 2s, enrichment voltage: 0.6V, enrichment time: 120s, potential increment: 0.008V, amplitude: 0.02V, frequency: 50Hz.

[0024] The following table shows the results of testing different concentrations of vitamin A: Table 5. Table of VA at different concentrations Retinol is oxidized to retinoic acid using oxidants such as perchloric acid. Simultaneously, the oxidants are placed in a buffer solution, where they react with retinol to produce retinoic acid. The reagent formulation was optimized to achieve highly sensitive and specific detection of vitamin A. The oxidation time and operational steps for the oxidation of retinol to retinoic acid using perchloric acid were also innovated. The electrochemical detection method using SWV was employed, with optimized parameters to improve detection sensitivity.

[0025] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. An electrochemical detection method for vitamin A, characterized in that: The electrochemical detection method includes the following steps: Preparation of electro-oxidation modified glassy carbon electrodes; The treatment of retinol (VA) involves acidifying it to retinoic acid. This acidification is achieved using perchloric acid as an oxidant, which reacts with the retinol in a buffer solution to produce retinoic acid. The reagents used for VA retinol treatment consist of: 950 μL phosphate solution + 950 μL organic reagent solution + 100 μL 0.01 mol / L-10 mol / L strong oxidant. The strong oxidant is perchloric acid or perchlorate, which is one or more of sodium perchlorate, potassium perchlorate, ammonium perchlorate, lithium perchlorate, magnesium perchlorate, and silver perchlorate. The phosphate buffer solution is formulated as follows: a 0.001 mol / L-1 mol / L dihydrogen phosphate solution and a 0.001 mol / L-1 mol / L dihydrogen phosphate solution, mixed in a ratio of 1:9 to 9:

1. Prepare a phosphate solution. The dihydrogen phosphate is a mixture of potassium dihydrogen phosphate, sodium dihydrogen phosphate, and potassium dihydrogen phosphate in any molar ratio. The dihydrogen phosphate is a mixture of dipotassium hydrogen phosphate, disodium hydrogen phosphate, and disodium hydrogen phosphate in any molar ratio. The perchloric acid oxidant oxidizes retinol to retinoic acid in 90-150 seconds. After adding the sample to the buffer solution, stir for 30-45 seconds, let stand for 60-105 seconds, and then perform electrochemical detection. The mixing ratio of dihydrogen phosphate and dihydrogen phosphate in the buffer solution was optimized using the SWV method. When optimizing this ratio, the electrochemical detection method of SWV was employed, with the following electrochemical parameters: initial electrode potential -0.1-0.3V, termination electrode potential 1.1-1.3V, settling time 2-20s, potential increment 0.001-0.01V, amplitude 0.005-0.1V, frequency 50-150Hz, enrichment voltage 0.1-0.6V, and enrichment time 80-220s. Finally, the VA was detected using the SWV electrochemical detection method.

2. The electrochemical detection method for vitamin A detection according to claim 1, characterized in that: When preparing the electro-oxidative modified glassy carbon electrode, the glassy carbon electrode is polished with 0.05 μm-1 μm alumina powder for 1-3 minutes in sequence, and then ultrasonically cleaned with pure water, anhydrous ethanol and pure water in sequence for 1-3 minutes.

3. The electrochemical detection method for vitamin A detection according to claim 2, characterized in that: The ultrasonically treated glassy carbon electrode was cleaned. A platinum wire electrode was used as the counter electrode, and a saturated calomel electrode or a silver-silver chloride electrode was used as the reference electrode. Cyclic voltammetry was performed in 0.01 mol / L-1 mol / L potassium ferricyanide, 0.01 mol / L-1 mol / L potassium chloride, and 0.01 mol / L-1 mol / L PBS solutions until the electrode stabilized. The initial electrode potential was -1 to 0.2 V, and the final electrode potential was 0.8 to 1.2 V. The scan rate was 20-200 mV / s, and the number of scan cycles was 10-60. Cyclic voltammetry was performed until the electrode stabilized, thus successfully preparing the electro-oxidation modified electrode.

4. The electrochemical detection method for vitamin A detection according to claim 1, characterized in that: Organic reagents include: ① Aromatic hydrocarbons: benzene, toluene, xylene; ② Aliphatic hydrocarbons: pentane, hexane, octane; ③ Alicyclic hydrocarbons: cyclohexane, cyclohexanone, toluenecyclohexanone; ④ Halogenated hydrocarbons: chlorobenzene, dichlorobenzene, dichloromethane; ⑤ Alcohols: methanol, ethanol, isopropanol; ⑥ Ethers: diethyl ether, propylene oxide; ⑦ Esters: methyl acetate, ethyl acetate, propyl acetate; ⑧ Ketones: acetone, methyl butyl ketone, methyl isobutyl ketone; ⑨ Glycol derivatives: ethylene glycol monomethyl ether, ethylene glycol monoethyl ether, ethylene glycol monobutyl ether; ⑩ Others: acetonitrile, pyridine, phenol.

Images