Method for lipoic acid measurement in biologically active additives by cathode voltammetry

FIELD: medicine.

SUBSTANCE: invention describes a method for lipoic acid measurement in biologically active additives by cathode voltammetry involving transferring a substance from a sample into a solution and taking voltammetric measurement; the cathode voltammetry is performed on a mercury-film electrode at potential -0.373 V of a relatively saturated silver-chloride electrode with borate buffer solution pH 9.18 at continuously current potential trace at 0.06 V/s with the determined lipoic acid content range from 4.5·106 to 1.1·10-3 mole/l.

EFFECT: improving sensitivity and expressivity of the method for measuring lipoic acid in tabletted BAAs by cathode voltammetry.

1 tbl, 1 ex, 3 dwg

 

The invention relates to the field of quantification of lipoic acid with antioxidant properties in biologically active food supplements. Method for determining cathodic voltammetry.

Lipoic acid (α-lipoic acid, 3-(4-carboxybutyl)-1,2-dithiolan, thioctic acid or Lipatova acid) is a coenzyme involved in the catalytic transfer reactions of hydrogen atoms and acyl groups, plays a major role in the metabolism of man and animals, is a powerful antioxidant.

Antioxidant properties of lipoic acid is due to the presence of two thiol groups in the molecule (Fig.1), as well as the ability to bind molecules free radicals and tissue iron, preventing its participation in the peroxide oxidation of lipids. It is known that lipoic acid not only has the independent antioxidant capacity, but also provides powerful support for the work of other antioxidant mechanisms in the body. In this respect, its protective effect is closely linked to the homeostasis in the system of glutathione and ubiquinone.

It is known that there is a complex of antioxidants, which can interact lipoic acid and maintain both lipid and aqueous antioxidant status, which is important for the treatment or prevention of various diseases, to�which notes the imbalance of redox cellular status, such as diabetes mellitus, liver dysfunction, etc.

On the basis of lipoic acid created a number of synthetic antioxidants to protect cells from oxidative stress in the form of biologically active food additives (BAA).

Effective and rapid method for determination of lipoic acid in pharmaceutical preparations is considered capillary electrophoresis with spectrophotometric method of detection (Sitton A., Schmid M. G., Gubitz G., Hassan Y. Aboul-Enein. Determination of lipoic acid in dietary supplement preparations by capillary electrophoresis // J. of Biochem. And Biophysic. Methods. - 2004. - V. 61, No. 2. - P. 119-124). In this method lipoic acid is determined in the UV region at 208 nm. The analysis time does not exceed 9 minutes. This method is used for determination of lipoic acid in the range from 0.8 to 2.5 mg/ml. the Method was developed for the determination of lipoic acid in dietary supplements diet food. The disadvantage can be considered low sensitivity of the determination of lipoic acid in dietary SUPPLEMENTS.

To ensure quality control of SUPPLEMENTS containing lipoic acid, it is necessary to develop a method for quantitative determination in her BAD. Currently, methods for the quantitative determination of lipoic acid in different objects are widely used electrochemical methods.

The closest to the subject of the invention is the voltammetric determination Lipaev�th acid in model solutions on glass-carbon electrode (G. K. Ziyatdinova, G. K. Budnikov, V. I. Pogoreltsev // Electrochemical determination of lipoic acid. Journal of analytical chemistry. 2004. T. 59. No. 3. Pp. 324-326). When the voltammetric determination of α-lipoic acid, the authors used stationary glass-carbon electrode on the background of 0.05 mol/l H2SO4while a distinct signal was observed at a potential of 0.72 V. the Range defined voltammetric contents lipoic acid ranged from 1.15·1-5up to 1.73·10-4mol/l on glass-carbon electrode.

The use of the terms in the prototype method does not provide the sensitivity of the determination of lipoic acid in real objects BAD that due to the introduction of error in the summed signal of the determination of lipoic acid through the influence of related components of the dietary Supplement.

New technical problem - increasing sensitivity and quick testing were method of determination of lipoic acid in tablet form SUPPLEMENTS the cathodic voltammetry. The task is achieved by the fact that lipoic acid is transferred from the pill form in the solution and carry out voltammetric determination using cathodic voltammetry. To develop methods for the determination of lipoic acid used the analytical signal recovery lipoic acid at a potential In -0.373 in borate �vernom solution pH 9.18 at a mercury-film electrode (RPE). Cyclic voltamperometry lipoic acid on the RPE is shown in Fig.2. The dependence of the gain current limit recovery lipoic acid from increasing its concentration in model solution linear. This area is located between the concentrations of 4.5·10-6mol/l to 1.1·10-3mol/l (Fig.3). The rate of potential sweep amounted to 0.06 In/s.

The limit of detection of lipoic acid 9.3·10-6mol/l are sufficient for use in assessing its quantitative content in the BUD.

Values of the limits of repeatability, reproducibility and critical range of concentration of lipoic acid at a confidence probability P=0.95 is shown in table.1.

Example 1. Determination of lipoic acid in tablet BAD "Lipoic acid".

One tablet triturated in a mortar to obtain a powder. The powder is weighed and transferred into a 50 ml flask, add 10-15 ml of borate buffer solution pH 9.18 and heated in a water bath at a temperature of 45-50°C for 10 minutes. After that, the solution was filtered through a paper filter, which was washed with buffer twice for 5 ml. Then in a quartz Cup with a capacity of 20 ml make 10.0 ml of the supporting electrolyte borate buffer solution pH 9.18, placed in the electrochemical cell for voltammetric analyzer (TA-2, Tomsk). Immersed in solution electrodes: and�dichotomy - mercury-film electrode, the auxiliary and the reference electrode - saturated chloride-silver. Registration background lines constantly spend in the current shooting mode with linear potential sweep rate 60 mV/s after the removal of oxygen from elektrokhimicheskoi cell with inert nitrogen gas for 15 minutes, in the range of potentials from 0 to -0.8 V. the Absence of peaks on voltamperometry and reproducible curves testified to the purity of the background. After obtaining satisfactory curves background contribute aliquot of filtrate lipoic acid a volume of 0.1 ml. Stirred solution of 10 with nitrogen gas, soothe with 20 and remove voltamperometry under the same conditions. The cathodic peak is recorded at a potential -0.373 V. the Concentration of lipoic acid was estimated by the height of the cathodic peak by the method of additives certified mixtures of lipoic acid by the standard technique.

The proposed method for the quantitative determination of lipoic acid is simple, does not require a lot of effort, a significant amount of chemicals and has a high sensitivity and rapidity.

The proposed method can be used for the quantitative determination of lipoic acid in dietary SUPPLEMENTS and dosage forms.

Table 1
Measurement range, mol/lThe limit of repeatability (relative value allowable difference between two measurement results obtained in one laboratory under conditions of repeatability), r, %Critical range (relative value allowable difference between the highest and lowest of the three measurement results obtained in one laboratory under conditions of repeatability), CROf 0.95(3),%The limit of Reproducibility (relative value allowable difference between two results
measurements obtained in different laboratories), R, %
4.5·10-6to 1.1·10-3incl.161925

The method of determination of lipoic acid in dietary supplements by cathodic voltammetry, including the transfer of substances from the sample into the solution and voltammetric determination, wherein the conducting cathodic voltammetry at a mercury-film electrode at a potential -0.373 In a relatively saturated chloride-silver electrode on the background of borate buffer solution pH 9,18 when permanently current form of the potential sweep at a speed of 0.06 In/with the region defined�holding lipoic acid 4.5·10 6to 1.1·10-3mol/L.



 

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