CN106198661A - Electrochemical determination method of phenylethylamine - Google Patents

Abstract

The invention relates to the technical field of electroanalytical chemistry, and specifically discloses an electrochemical determination method for phenethylamine. The method specifically comprises the following steps: S11. A gold electrode modified with colloidal gold is used as a working electrode, a platinum wire electrode is used as an auxiliary electrode, and an Ag/AgCl electrode is used as a reference electrode to construct a three-electrode electrochemical test system; S12. The electrode system is placed in an alkaline solution, and the i-t curve method is used to scan when the initial potential is 0.08~0.12V. After the baseline is stable, add the solution to be tested, and then scan until it is stable, and measure the presence of phenylethylamine on the electrode. The current difference of the response; S13. According to the linear regression equation between the current difference of the phenylethylamine response on the electrode and the concentration of the phenylethylamine, the content of the phenylethylamine is calculated. The method can quickly detect the content of phenylethylamine with low detection limit, strong anti-interference and good reproducibility.

Description

A kind of electrochemical determination method of phenylethylamine

technical field

The invention relates to the technical field of electroanalytical chemistry, in particular to an electrochemical determination method of phenethylamine.

Background technique

Biogenic amine is a general term for a class of nitrogen-containing small molecular weight compounds. According to the structural characteristics of biogenic amines, they can be divided into three categories: (1) aliphatic, such as cadaverine, putrescine, spermidine, spermine, etc.; (2) aromatic, such as phenylethylamine, tyramine, etc.; (3) ) heterocyclic group, such as tryptamine, histamine, etc. Biogenic amine exists in the tissues of various animals and plants. It is a normal active ingredient in biological organisms and plays an important physiological role in animal and plant organisms. For example, spermine, spermidine, putrescine, and cadaverine are all indispensable parts of biologically active cells and play key roles in regulating the synthesis of proteins and nucleic acids and the stability of biofilms. In addition, biogenic amines also widely exist in a variety of foods, especially fermented foods and protein-rich foods. Such as: meat products, aquatic products, cheese, sausage, beer, wine, etc. The main biogenic amines in food include: histamine, tyramine, β-phenylethylamine, cadaverine, putrescine, tryptamine, spermine and spermidine and other monoamines and polyamines. In foods rich in protein such as meat products and aquatic products, the enzymes produced by related microorganisms can catalyze the removal of carboxyl groups from free amino acids to produce biogenic amines, so the content of biogenic amines is closely related to their quality.

Phenylethylamine is also called 2-phenylethylamine or β-phenylethylamine, its molecular formula is C ₈ H ₁₁ N, its chemical structure is relatively simple, and it belongs to aromatic amines. Widely present in food such as chocolate, milk, meat and wine, especially food after microbial fermentation. In the human body, phenylethylamine is an alkaloid and monoamine neurotransmitter, which can increase the level of dopamine in the extracellular fluid, and at the same time inhibit the activation of dopamine nerves, and is used for the treatment of depression. In the human brain, β-phenylethylamine acts as a neuromodulator, neurotransmitter and tracer amine. As an important pharmaceutical and dye intermediate, phenylethylamine is mainly used in the synthesis of the following drugs in medicine: stimulants, antidepressants, hallucinogens, ecstasy stimulants, appetite suppressants, bronchodilators, etc. Phenylethylamines can be used as important drugs in pharmacology. However, if phenethylamines are present in food in excess, they will cause certain harm to the body. Therefore, it is particularly important to detect the content of biogenic amines in foods such as fermented foods, animal source foods and wine.

Contents of the invention

The technical problem to be solved by the present invention is to provide an electrochemical determination method for phenethylamine in order to overcome the lack of a method capable of rapidly detecting the content of phenethylamine in the prior art. When the method is used to detect phenethylamine, the detection speed is fast and the detection limit is low.

The above-mentioned technical problems to be solved by the present invention are realized through the following technical solutions:

An electrochemical determination method of phenylethylamine adopts a gold electrode modified by colloidal gold, and when the initial potential is 0.1V, the i-t curve method is used to determine the content of phenylethylamine.

The present invention shows through a large number of experimental studies of the inventor, adopts the gold electrode modified with colloidal gold prepared by the present invention, uses the i-t curve method to measure phenylethylamine under the conditions described in the present invention, and can quickly detect phenylethylamine in the sample content, and low detection limit, strong anti-interference, good reproducibility.

Preferably, the electrochemical assay method of described phenylethylamine specifically comprises the following steps:

S11. Using colloidal gold-modified gold electrodes as working electrodes, platinum wire electrodes as auxiliary electrodes, and Ag/AgCl electrodes as reference electrodes, construct a three-electrode electrochemical test system;

S12. Place the three-electrode system in an alkaline solution, and use the i-t curve method to scan when the initial potential is 0.08~0.12V. After the baseline is stable, add the solution to be tested, and then scan until it is stable. The current difference across the electrodes responds;

S13. Calculate the content of phenylethylamine according to the linear regression equation between the current difference (-∆I, nA) and the concentration of phenethylamine (c, μmol/L) that phenethylamine responds to on the electrode.

Further preferably, the alkaline solution described in step S12 is a NaOH solution with a pH value of 12-13; the initial potential is 0.08-0.10V.

Further preferably, the linear regression equation described in step S13 is -∆I = 7.783c + 1.181 (R=0.997).

Preferably, the colloidal gold-modified gold electrode described in step S11 is prepared by a method comprising the following steps:

S21. dissolving chitosan in the acetic acid solution that is 0.5～2.0% by volume fraction is made into the chitosan acetic acid solution that chitosan content is 1～3mg/mL;

S22. Add 5~15 mmol/L chloroauric acid (HAuCl ₄ ) solution to chitosan acetic acid solution, stir for 20~40min, the volume ratio of chitosan acetic acid solution and chloroauric acid (HAuCl ₄ ) solution is 1.5 ~3︰1;

S23. Add 0.05 ~ 0.2mol/L aqueous solution of sodium borohydride dropwise under stirring until the solution becomes transparent wine red to obtain a colloidal gold solution;

S24. Take the gold electrode, after polishing and cleaning, drop a colloidal gold solution on the surface of the gold electrode, and dry it to obtain a gold electrode modified by colloidal gold.

Further preferably, in step S21, dissolving chitosan in an acetic acid solution with a volume fraction of 1.0-2.0% is prepared as a chitosan-acetic acid solution with a chitosan content of 2-3 mg/mL.

Most preferably, in step S21, dissolving chitosan in an acetic acid solution with a volume fraction of 1.0% is made into a chitosan acetic acid solution with a chitosan content of 2 mg/mL.

Further preferably, in step S22, add 10-15 mmol/L chloroauric acid (HAuCl ₄ ) solution to the chitosan acetic acid solution, stir for 20-30 min, the chitosan acetic acid solution and the chloroauric acid (HAuCl ₄ ) solution The volume ratio is 1.5~2:1.

Most preferably, in step S22, add 10 mmol/L chloroauric acid (HAuCl ₄ ) solution to the chitosan acetic acid solution, stir for 30 minutes, the volume ratio of chitosan acetic acid solution and chloroauric acid (HAuCl ₄ ) solution is 2 ︰1.

Further preferably, in step S23, a 0.1-0.2 mol/L sodium borohydride aqueous solution is added dropwise under stirring.

Most preferably, in step S23, a 0.1 mol/L aqueous solution of sodium borohydride is added dropwise under stirring.

Further preferably, in step S24, 2-5 μL of colloidal gold solution is dripped on the surface of the gold electrode.

Most preferably, in step S24, 3 μL of colloidal gold solution is dripped on the surface of the gold electrode.

Preferably, the electrochemical determination method of phenylethylamine is used to measure the content of phenylethylamine in fish sauce, and the test step also includes a pretreatment step, and the pretreatment step is as follows:

Take a sample of fish sauce, filter it with a filter membrane, add sodium chloride to saturate the solution, measure 8~12mL of saturated solution, adjust the pH to 12~13 with NaOH solution, and then add 4~6mL of n-butane with a volume ratio of 1:1 Alcohol-chloroform mixture, oscillate and centrifuge, take the upper organic phase, repeat 2-5 times, combine the organic phases, dry them, add NaOH solution with a pH value of 12-13 to dissolve, and obtain the solution to be tested.

Most preferably, the described pretreatment steps are as follows:

Take a fish sauce sample, filter it with a filter membrane, add sodium chloride to make the solution saturated, measure 10mL of the saturated solution, adjust the pH to 12.7 with NaOH solution, and then add 5mL of n-butanol-chloroform with a volume ratio of 1:1 to mix solution, shaken and centrifuged, take the upper organic phase, repeat twice, combine the organic phases, dry them, add NaOH solution with a pH value of 12.7 to dissolve, and obtain the solution to be tested.

Beneficial effects: (1) The present invention provides a brand-new electrochemical method for the determination of phenethylamine; (2) The method of the present invention can quickly detect the content of phenethylamine, and the detection limit is low (Example data It shows that the detection limit is 1.4×10 ^{- 8} mol/L), strong anti-interference and good reproducibility; (3) The method of the present invention is used to determine the phenylethylamine in the fish sauce sample accurately and reliably.

Description of drawings

Fig. 1 is the graph of the relationship between the response current difference of phenethylamine solution and the concentration of phenethylamine.

Figure 2 is a comparative diagram of the current difference between tryptamine and phenethylamine at the same concentration.

detailed description

The present invention is further explained below in conjunction with specific examples, but the examples do not limit the present invention in any form.

Those who do not indicate the specific conditions in the examples are carried out according to the conventional conditions or the conditions suggested by the manufacturer. Among them, the reagents or instruments used, those whose manufacturers are not indicated, are all commercially available conventional products.

The electrochemical determination method of embodiment 1 phenethylamine

S11. Using colloidal gold-modified gold electrodes as working electrodes, platinum wire electrodes as auxiliary electrodes, and Ag/AgCl electrodes as reference electrodes, construct a three-electrode electrochemical test system;

S12. Place the three-electrode system in an alkaline solution, and use the i-t curve method to scan when the initial potential is 0.10V. After the baseline is stable, add the solution to be tested, and then scan until it is stable, and measure the phenethylamine on the electrode. The current difference of the response;

S13. Calculate the content of phenylethylamine according to the linear regression equation between the current difference (-∆I, nA) and the concentration of phenylethylamine (c, μmol/L) that phenethylamine responds to on the electrode; the linear regression The equation is -∆I = 7.783c + 1.181 (R=0.997);

The colloidal gold-modified gold electrode described in step S11 is prepared by a method comprising the following steps:

S21. dissolving chitosan in the acetic acid solution that volume fraction is 1.0% is made into the chitosan acetic acid solution that chitosan content is 2mg/mL;

S22. Add 10 mmol/L chloroauric acid (HAuCl ₄ ) solution to the chitosan acetic acid solution, stir for 30 minutes, the volume ratio of chitosan acetic acid solution and chloroauric acid (HAuCl ₄ ) solution is 2:1;

S23. Add 0.1mol/L sodium borohydride aqueous solution dropwise under stirring until the solution becomes transparent wine red to obtain a colloidal gold solution;

S24. Take the gold electrode, after polishing and cleaning, drop 3 μL of colloidal gold solution on the surface of the gold electrode, and dry to obtain the gold electrode modified by colloidal gold.

Embodiment 2 electrochemical performance test

(1) Standard curve establishment

Using the method described in Example 1, the change of the current response of phenethylamine in the solution on the gold electrode modified by colloidal gold was investigated. As shown in Figure 1, the current difference (-∆I, nA) and the concentration of phenylethylamine (c, μmol/L) in response to phenylethylamine on the electrode range from 4.13×10 ^-8 to 4.72×10 ^-6 mol/L There is a good linear relationship within the range, and its linear regression equation is: -∆I =7.783c + 1.181 (R=0.997), based on three times the signal-to-noise ratio (S/N = 3), it is obtained that the modified gold electrode is The detection limit of ethylamine is 1.4×10 ^-8 mol/L.

(2) Anti-interference test

According to the method described in Example 1, the interference of tyramine, histamine, spermine, spermidine, putrescine, cadaverine and tryptamine to the determination of phenylethylamine at 10 times the concentration of phenylethylamine was investigated. The results showed that, except for tryptamine, the above other biogenic amines had no obvious interference to the determination of phenylethylamine.

Interference analysis of tryptamine: with 2.49×10 ^-7 mol/L, 5.8×10 ^-7 mol/L, 1.23×10 ^-6 mol/L phenethylamine solution as a control, the same concentration of tryptamine was investigated for the determination of phenylethylamine The interference of amines, the results are shown in Figure 2.

The results show that the current difference of the same concentration of tryptamine is significantly larger than that of phenylethylamine, and tryptamine has less interference, and the content of tryptamine in aquatic products and their seasonings is relatively small. Therefore, the influence of tryptamine will not be considered in the follow-up sample test.

(3) Reproducibility test

The method described in Example 1 is used to measure the phenethylamine solution whose concentration is 1.66×10 ^-7 mol/L and 3.32×10 ^-7 mol/L, and parallel measurement is performed 6 times at the same time, and the current difference of the 6 measurements is as follows: Table 1 shows.

From the data in the table, it can be seen that the relative standard deviation (RSD) of phenethylamine solutions with concentrations of 1.66×10 ^-7 mol/L and 3.32×10 ^-7 mol/L measured by the modified electrode are 4.7% and 4.0% respectively . The results show that the fabrication of the modified electrode has good reproducibility.

Determination of phenylethylamine in embodiment 3 fish sauce

Get the fish sauce condiment purchased in the supermarket in a clean small beaker, filter it in a 50mL beaker with a 0.22 μm cellulose acetate filter, add sodium chloride to make the solution saturated in the resulting filtrate. Measure 10.00mL of the saturated solution into a 50mL centrifuge tube, adjust the pH to 12 with 1.0mol/L NaOH solution, then add 5.0mL of n-butanol-chloroform (1:1) mixture, vortex for 5min, and place at 3600r/ min Centrifuge for 10 min, take the upper organic phase, repeat twice, and combine the two solutions. The mixture was evaporated to near dryness in a water bath at 60°C, and then blown to dryness with nitrogen. Finally, 1 mL of pH 12.7 NaOH solution was added to dissolve the residue, and the resulting solution was the solution to be tested. Then measure the phenethylamine content according to the method described in Example 1.

As a result, the concentration of phenylethylamine in the fish sauce test sample was found to be 1.85×10 ^-7 mol/L, and the content of phenylethylamine in the sample was converted to 1.11×10 ^-5 mol/L. In order to verify the accuracy of this method, a spike recovery experiment was carried out on fish sauce samples (Table 2). The recoveries of phenethylamine at different concentrations (1.66×10 ^-7 mol/L, 3.31×10 ^-7 mol/L) in fish sauce samples were 90.0%~110.8%, and the RSDs were 4.8% and 6.2%, respectively. It shows that the method is accurate and reliable.

Claims (10)

1. an electrochemical determination method of phenylethylamine is characterized in that, the gold electrode that adopts colloidal gold modification, when initial potential is 0.1V, utilizes the i-t curve method to measure the content of phenylethylamine. 2. the electrochemical assay method of phenylethylamine according to claim 1, is characterized in that, specifically comprises the steps: S11. Using colloidal gold-modified gold electrodes as working electrodes, platinum wire electrodes as auxiliary electrodes, and Ag/AgCl electrodes as reference electrodes, construct a three-electrode electrochemical test system; S12. Place the three-electrode system in an alkaline solution, and use the i-t curve method to scan when the initial potential is 0.08~0.12V. After the baseline is stable, add the solution to be tested, and then scan until it is stable. The current difference across the electrodes responds; S13. Calculate the content of phenethylamine according to the linear regression equation between the current difference of phenethylamine response on the electrode and the concentration of phenethylamine. 3. the electrochemical determination method of phenylethylamine according to claim 2, is characterized in that, the alkaline solution described in step S12 is the NaOH solution that pH value is 12～13; Described initial potential is 0.08～ 0.10V. 4. the electrochemical determination method of phenylethylamine according to claim 2, is characterized in that, the linear regression equation described in step S13 is-∆I=7.783c+1.181. 5. the electrochemical determination method of phenylethylamine according to claim 2, is characterized in that, the gold electrode of the colloidal gold modification described in step S11 is prepared by the method comprising the following steps: S21. dissolving chitosan in the acetic acid solution that is 0.5～2.0% by volume fraction is made into the chitosan acetic acid solution that chitosan content is 1～3mg/mL; S22. in chitosan acetic acid solution, add the chloroauric acid solution of 5 ~ 15 mmol/L, stir 20 ~ 40min, the volume ratio of chitosan acetic acid solution and chloroauric acid solution is 1.5 ~ 3: 1; S23. Add 0.05 ~ 0.2mol/L aqueous solution of sodium borohydride dropwise under stirring until the solution becomes transparent wine red to obtain a colloidal gold solution; S24. Take the gold electrode, after polishing and cleaning, drop a colloidal gold solution on the surface of the gold electrode, and dry it to obtain a gold electrode modified by colloidal gold. 6. the electrochemical determination method of phenylethylamine according to claim 5 is characterized in that, in step S21, dissolving chitosan in the acetic acid solution that volume fraction is 1.0～2.0% is made into chitosan content and is 2 ~3 mg/mL chitosan in acetic acid solution. 7. the electrochemical determination method of phenylethylamine according to claim 5, is characterized in that, in step S22, adds the chloroauric acid solution of 10～15 mmol/L in chitosan acetic acid solution, stirs 20～30min, The volume ratio of chitosan acetic acid solution and chloroauric acid solution is 1.5~2:1. 8. the electrochemical determination method of phenethylamine according to claim 5, is characterized in that, in step S23, add the aqueous solution of 0.1～0.2mol/L sodium borohydride dropwise under stirring. 9. the electrochemical determination method of phenylethylamine according to claim 5, is characterized in that, in step S24, the colloidal gold solution of 2～5 μ L is dripped on the gold electrode surface. 10. the electrochemical determination method of phenylethylamine according to claim 2, is characterized in that, is used for measuring the content of phenylethylamine in fish sauce, also comprises pretreatment step in its test step, described pretreatment step as follows: Take a sample of fish sauce, filter it with a filter membrane, add sodium chloride to saturate the solution, measure 8~12mL of saturated solution, adjust the pH to 12~13 with NaOH solution, and then add 4~6mL of n-butane with a volume ratio of 1:1 Alcohol-chloroform mixture, oscillate and centrifuge, take the upper organic phase, repeat 2-5 times, combine the organic phases, dry them, add NaOH solution with a pH value of 12-13 to dissolve, and obtain the solution to be tested.