RU2457483C2 - Method of analysing reducing substances - Google Patents

Abstract

FIELD: chemistry.

SUBSTANCE: method involves oxidation of reducing substances with copper (II) compounds in an alkaline medium while heating on a boiling water bath for 10 minutes, bringing the volume of the cooled solution to a defined value, determining optical density at wavelength 670 nm, making an adjustment to partial oxidation of non-reducing carbohydrates, which is subtracted from the optical density reading, and estimating concentration of reducing substances from a calibration curve.

EFFECT: high accuracy and reproducibility of analysis.

5 ex, 6 tbl, 4 dwg

Description

The invention relates to a method for determining reducing substances and can be used in confectionery, caramel and sugar production.

The content of reducing substances in caramel is strictly regulated and is no more than 20% for non-acidified caramel, no more than 23% for caramel with an addition of acid in excess of 0.6% and no more than 32% for products with lactose [GOST 6477-88 Caramel. General specifications.]. Excess reducing sugars can lead to adsorption of moisture from the air and dampening of the product. The lack of reducing substances causes the crystallization of sucrose inside the product, which affects its quality during prolonged storage.

By the number of reducing substances, the degree of hydrolysis of starch in sugar production is judged, this indicator mainly determines the type of product: for low-sugared molasses, the content of reducing substances is 26-35%, for caramel acid and caramel enzymatic - 36-44%, for maltose - 38 % or more, for highly sugared - 45% or more [GOST R 52060-2003 Starch syrup. General specifications].

Known methods based on the determination of the content of reducing substances, based on the titrimetric determination of copper oxide (I), released as a result of the recovery of divalent copper by reducing substances. The main disadvantage of these methods is the subjectivity in determining the end of titration, as well as the need for substitution titration, which increases the complexity of the method, and also significantly affects the measurement error [GOST 5903-89. Confectionery. Methods for the determination of sugar. - S.131-141].

An analogue of the invention is a photocolorimetric method based on the decolorization of a solution of potassium hexacyanoferrate (ferricyanide) (III) during the reaction with reducing substances. The amount of reducing substances is judged by the residue of ferricyanide after the reaction. The remainder of ferricyanide is determined by the optical density in cuvettes with a layer thickness of 10 mm and with a filter having a wavelength of 440 nm relative to a blank sample [GOST 5903-89. Confectionery. Methods for the determination of sugar. - S.144-147].

The disadvantages of this method include:

1) the use of potassium ferricyanide for the oxidation of reducing substances, which, compared with the Feling solution, significantly oxidizes sucrose;

2) when the solution is boiling, it is concentrated due to intensive evaporation of moisture, resulting in an increase in optical density, causing an unsystematic error in parallel experiments (it is impossible to achieve the same heating rate, boiling rate, cooling rate);

3) the calibration graph does not obey the Bouguer-Lambert-Beer law, therefore this method cannot guarantee a reliable result - Fig. 1.

The closest analogue (prototype) is a method based on photocolorimetry of a copper-alkaline solution (Feling solution) after reaction with reducing substances. A solution of copper sulfate, an alkaline solution of Rochelle salt, a solution of yellow blood salt and the test solution are added to the test tube. After the test tube is heated in a boiling water bath for 3 minutes, after which the mixture is photometric on a photocolorimeter at 670 nm relative to distilled water in cuvettes with a working layer thickness of 1 cm. In parallel, an empty experiment is carried out without heating. The content of reducing substances is determined by the equation for the calibration graph [RU 2366931 C1 of 05.03.2008].

The disadvantages of this method are:

1) when the solution is kept in a boiling water bath, its concentration changes due to intensive evaporation of moisture, resulting in an increase in optical density, causing a non-systematic measurement error;

2) the absence of a correction for the partial oxidation of non-reducing carbohydrates (for example, sucrose) by the Feling reagent - Fig. 2, which is extremely necessary to consider when analyzing confectionery products;

3) insufficient holding time in a boiling water bath - 3 minutes: during this time the reducing substances do not have time to completely react, the optical density changes, which also introduces an error in the determination - Fig. 3.

The technical result of the proposed method is to increase the accuracy and reproducibility of the analysis.

The technical result is achieved by the fact that the oxidation of reducing substances by copper (II) compounds in an alkaline medium when heated in a boiling water bath for 10 minutes, bringing the volume of the cooled solution to a certain value, determining the optical density at a wavelength of 670 nm, introducing a correction for partial oxidation non-reducing carbohydrates, which is subtracted from the readings of optical density, and an estimate of the concentration of reducing substances according to the calibration graph.

The essence of the proposed method is as follows.

Reagents: solution I: 34.66 g CuSO ₄ · 5H ₂ O in 1 liter of solution; solution II: 70 g of NaOH, 173 g of Rochelle salt (potassium sodium tartrate) and 4 g of yellow blood salt g in 1 liter. It is possible to prepare solution II without yellow blood salt, in which case filtration or centrifugation is necessary before removing the optical density of the test solution.

Experimental technique

In a 25 ml volumetric flask, pipette 5 ml of solutions I and II and the test solution to 10 ml (standard solution of reducing sugar), hold for 10 min in a boiling water bath, cool to room temperature and bring to the mark with distilled water, mix (centrifuged / filter) and read the absorbance at 670 nm relative to a blank sample (5 ml of solutions I and II, volume brought up to 25 ml) using a 10 mm wide cuvette. The optical density value is taken modulo.

If the test solution contains non-reducing carbohydrates, for example sucrose (conventionally it can be calculated as the difference between the total sugar and reducing substances) in significant quantities, the table is corrected.

Correction for non-reducing carbohydrates in determining the content of reducing substances

The mass of non-reducing carbohydrates introduced with the analyzed solution, mg Correction (subtract from the absorbance reading) 40-50 0.005 50-65 0.010 65-100 0,020 more than 110 0,035

The content of reducing substances in terms of the mass of glucose is found according to the calibration graph, previously built for a standard glucose solution with a concentration of 2 g / l - Fig. 4, or directly according to the proposed equation:

where m is the content of reducing substances in terms of the mass of glucose, mg; And - the value of optical density, taking into account the amendment.

The percentage of reducing substances in the product is found by the formula:

where M is the mass of the sample, g; V is the volume of the volumetric flask in which the sample is dissolved, ml; ν is the volume of the solution taken for analysis, ml.

Example 1

1 g of caramel (sample 1) is dissolved in a 100 ml volumetric flask. Reagents and test solution are added to 25 ml volumetric flasks: 4, 6 and 8 ml, which corresponds to 40, 60 and 80 mg of the product. Taking the moisture content of caramel 3% and the content of non-reducing carbohydrates 80% of dry matter, it is necessary to make an adjustment according to table. on the reading of the optical density in the 2nd case by 0.005 and in the third by 0.010.

The results are tabulated.

v BUT A (as amended) m ω _red Mean ± standard deviation four 0.342 0.342 9,948 24.87 25.27 ± 0.36 6 0.512 0.507 15,221 25.37 8 0.669 0.659 20,458 25.57

Example 2

1 g of caramel (sample 2) is dissolved in a 100 ml volumetric flask. Further, as Example 1.

v BUT A (as amended) m ω _red Mean ± standard deviation four 0.244 0.244 7,019 17.55 17.43 ± 0.16 6 0.360 0.355 10,350 17.25 8 0.480 0.470 14,002 17.50

Example 3

1 g of caramel, prepared in laboratory conditions by boiling sugar syrup with molasses 1: 1 to a temperature of 140 ° C, is dissolved in a 100 ml volumetric flask. Reagents and test solution are introduced into 25 ml volumetric flasks: 2, 4, 6, 8, and 10 ml, which corresponds to 20, 40, 60, 80, and 100 mg of the product. For 3 definitions, we accept the amendment of 0.005, for 4 - 0.010, for 5 - 0.020.

v BUT A (as amended) m ω _red Mean ± standard deviation 2 0.125 0.125 3.67 18.32 18.26 ± 0.05 four 0.254 0.254 7.31 18.28 6 0.379 0.374 10.94 18.23 8 0.497 0.487 14.56 18,20 10 0.607 0.597 18.28 18.28

Example 4

1 g of molasses is dissolved in a 100 ml volumetric flask. Reagents and test solution are added to 25 ml volumetric flasks: 2, 4, 5, and 6 ml, which corresponds to 20, 40, 50, and 60 mg of the product. Taking the molasses moisture of 22% and the content of non-reducing carbohydrates 70% of dry matter, no correction is necessary in any case (60 * 0.78 * 0.7 <40).

v BUT A (as amended) m ω _red Mean ± standard deviation 2 0.226 0.226 6.50 32,49 32.87 ± 0.53 four 0.439 0.439 13.00 32,49 5 0.543 0.543 16,43 32.86 6 0.651 0.651 20.17 33.62

Example 5

Analysis of bread for total sugar. After acid hydrolysis and neutralization of the aqueous extract, 6 g of a sample of bread is adjusted to 100 ml. Reagents and test solution are added to 25 ml volumetric flasks: 2, 4, 6, and 8 ml, which corresponds to 120, 240, 360, and 480 mg of the product. Amendment is not necessary.

v BUT A (as amended) m ω _red Mean ± standard deviation 2 0,079 0,079 2.43 2.02 2.03 ± 0.03 four 0.165 0.165 4.77 1.99 6 0.255 0.255 7.34 2.04 8 0.339 0.339 9.86 2.05

The proposed method has a very high reproducibility and accuracy compared to analogues.

Claims (1)

A method for the analysis of reducing substances in sugar-containing media, including the oxidation of reducing substances with copper (II) compounds in an alkaline medium when heated in a boiling water bath for 10 minutes, bringing the volume of the cooled solution to a certain value, determining the optical density at a wavelength of 670 nm, introducing an amendment for partial oxidation of non-reducing carbohydrates, which is subtracted from the readings of optical density, and estimation of the concentration of reducing substances according to the calibration graph.

Images