SU1755137A1 - Method of pyrocathechol determination - Google Patents

Abstract

SUMMARY OF THE INVENTION: A sample is treated with resorcinol and hydrogen peroxide in the presence of peroxidase at pH 7.5-8.0 and measurement of the optical density is measured in the kinetic mode. 2 tab.

Description

The invention relates to analytical chemistry or applied biology, and specifically to methods for determining pyrocatechin in the mixture, and can be used in the analysis of wastewater, in determining the content of pyrocatechin in resorcinol.

A known method for the determination of trace amounts of phenols in water is based on the condensation of 4 -aminoantipyrine with phenols.

The disadvantage of this method is that it is impossible to determine separately pyrocatechol if it is mixed with other phenols, since equally colored condensation products are formed with different phenols.

The closest to the invention to the technical essence and the achieved result is a method for determining pyrocatechin, based on the processing of the sample being analyzed with a chemical peareWF volume containing phenols followed by photometry of the resulting solution.

The disadvantage of this method is low selectivity, since the method

intended for the determination of individual phenols and various phenols give the same colored condensation products.

The purpose of the invention is to increase the selectivity of the determination.

The goal is achieved by the fact that in the method based on the processing of an analyzed sample with a chemical reagent and photometric analysis, the analyzed sample is treated with resorcinol, hydrogen peroxide in the presence of peroxidase at pH 7.5-8.0 and the change in optical density is measured in the kinetic mode.

The method is carried out as follows.

The concentration of pyrocatechol is determined. Phosphate buffer, aqueous solutions of resorcinol, peroxidase, and hydrogen peroxide are added to the sample to be analyzed. The mixture is rapidly mixed and its optical density (A) is measured at certain time intervals T for 1-2 minutes. Based on the calculated ratio of the optical density (D A) over time (DT), the concentration of resorcinol in the sample is judged.

Xi

listened to

from h

Example 1

1.6 ml of phosphate buffer pH 7.75 are added to the cuvette; 1.0 ml of resorcinol 20 mM; 1.0 ml of pyrocatechin containing 1 μg of the substance; 0.2 ml of peroxidase with a concentration of 5 µg / ml, 0.2 ml of 140 mmol / l of hydrogen peroxide solution, quickly mix the mixture and measure the change in optical density with time at L 460 nm. Ti 15c, T2 45c, DT 45 s-15 with 30 with Ai 0.014, Aa 0.053 D A 0.053 - 0.014 - 0.039

YES

st

100 7.8

Example 2. The concentration of pyrocatechol is determined. 1.6 ml of phosphate buffer pH 7.75 are added to the cuvette; 1.0 ml of resorcinol 20 mmol / l; 1.0 ml of pyrocatechin containing 1 μg of the substance; 0.2 ml peroxidase with a concentration of 5 μg / ml; 0.2 ml 140 mmol / l hydrogen peroxide solution; mix the mixture rapidly and measure the change in optical density with time at A = 460 nm.

Ti-15c, T2 45c,

LT 45 with - 15 with 30 with 0.5 min

And 1 0,014, A2 0,053,

A A 0.053-0.014 0.039

YES 5T

100 7.8

Example 3. The determination is carried out analogously to example 1, except that pH 8.0, hydrogen peroxide concentration 180 mM / l, resorcinol 50 mM / l and kinetics are removed at I 470 nm, Ti 15 s, T2 45 s, DT 30 s 0.5 min Ai 0.012, A2 0.050, YES 0.038

YES ZT

100 7.6

Example 4. The determination is carried out analogously to Example 1, except that the pH is 7.5, the hydrogen peroxide concentration is 100 mM / L and the resorcinol is 15 mM / L, measurements are made at A = 450 nm. Ti 15s, T2 45s DT 0.5min Ai 0.012, A2 0.047 YES 0.035

100 7.0

DT in minutes, YES - in units optical density (D A / DT) 100 - in arbitrary units.

The sequence of operations when making the components of the reaction mixture sample solutions, resorcinol, buffer does not matter. Peroxidase and hydrogen peroxide are added last. Hydrogen peroxide

triggers the reaction, and from the moment it is introduced, the time begins.

Example 5. All values as in Example 1, but the content of pyrocatechin 6

mcg The sequence of the components of the reaction mixture: solutions of buffer, resorcinol, pyrocatechin samples, peroxidase, hydrogen peroxide.

Ti 15s, T2 75s, DT 60s 1 min

Ai 0,026, A2 0,178 D A A2 - AI

 0.178-0.026 0.152 (YES / DT) -, 2 Example 6. All values are as in Example 4, but the sequence of the components of the mixture is different; solutions of sample, resorcinol, buffer, peroxidase, hydrogen peroxide.

Ti 15 s, T2 75 s.D T 60 s 1 min Ai 0.026, A2 0.179, D A 0.179 - 0.026

0.153

{DA / DT) - 100 15.3 In this method, the oxidation reaction of a mixture of phenols is catalyzed by a peroxidase system (peroxidase plus peroxide

hydrogen). The results presented in the tables and examples, obtained in experiments where one. from the components of the reaction mixture, 0.2 ml of peroxidase solution with a concentration of 5 μg / ml.

For the method, this is the optimal peroxidase concentration. At lower concentrations, the sensitivity of the method decreases, at higher concentrations, the linear portion of the kinetic curve of the color increase decreases. With an increase in peroxidase concentration of 5 µg / ml, the linear portion of the kinetic curve became less than 25 seconds (for pyrocatechol with a concentration of 1-2 µg / ml) and deteriorated

reproducibility of experiments.

The proposed method allows the direct determination of trace amounts of pyrocatechol in the presence of high concentrations of resorcinol (at a ratio of

Claims (2)

pyrocatechin: resorcinol to 1: 1000). Data on the advantage of the proposed method — the high selectivity of the determination of pyrocatechol — is presented in Table. 2. The formula of the invention 0 A method for the determination of pyrocatechol, including the treatment of an analyzed sample with a chemical reagent and photometric measurement, characterized in that, in order to increase the selectivity of the determination, the analyzed sample is treated with resorcinol and hydrogen peroxide in the presence of peroxidase at pH 7.5-8.0 and measured the change in optical density in the kinetic mode. Table1 The dependence of the rate of the indicator reaction on the concentration of pyrocatechin table 2