Method of determining triterpene saponins in vegetable raw material and medications

FIELD: medicine.

SUBSTANCE: method of determination of triterpene saponins in vegetable raw material and medications includes dissolution of saponin-containing fraction in mixture water-ammonium buffer, determination of its optic density and calculation of saponin content in terms of oleanolic acid, under specified conditions.

EFFECT: claimed method represents express method, facilitates analysis and increases degree of reliability of obtained results.

2 ex

 

The method of determination of triterpene saponins in plant raw materials and medicines.

The invention relates to methods of quantitative determination of active drug substances in plant material and obtained from him the drugs and can be used in pharmaceutical and food industries.

Known methods of determination of saponins method colorimetry using various chromogenic reagent solution of ferric chloride, a solution of phenylhydrazine, petaluridae antimony [Nagorno VA, jijina GT, Kartashov A.K. // Sugar industry. 1966. No. 8. P.39-43]. However, the coloration of the resulting compounds are unstable in time, and the analysis results are poorly reproducible.

The known method of determination of saponins in ginseng extract (Patent RF №1480568 IPC G01N 33/68 from 30.11.1994 g), which are obtained from raw materials and dosage forms of ginseng extract containing saponins, mixed with vanilla and sulfuric acid and heated to obtain a purple-red coloring, then the resulting solution spectrophotometery at λ=544 nm. As a standard sample used alcohol solution of vitamin D2.

The technical result consists in the universality and expressnet method of determination of triterpene saponins in plant raw materials and medicines is the courthouse square.

The technical result is achieved in that in the method of determination of triterpene saponins in plant raw materials and medicines, including cooking saponinosteroids fractions, determination of the optical density and the calculation of the contents of saponins of pre-constructed calibration curve, according to the invention for the preparation of fractions as solvent a mixture of water - ammonia buffer 1:1, the optical density is determined at λ=210 nm, and the content of saponins calculated in terms of oleanolic acid.

Since the determination of saponins spectrophotometric methods based on the absorption of electromagnetic radiation of the chromophore groups are not all molecules of saponin, but only its aglycone, in our case, oleanolic acid, our proposed method provides the possibility of quantitative determination of the total content of individual saponins in the analyzed object in terms of oleanolic acid is the most reactive part of the molecule that determines its pharmacological activity. The absorption maxima of saponin and oleanolic acid are the same and are stable over a long period of time (1 h)that facilitates the analysis and increases the reliability of the results.

Capodistrias FR is the Ktsia, obtained from vegetable raw materials or medicinal product, dissolved in a mixture of water - ammonia buffer [State Pharmacopoeia of the USSR: 2). General methods of analysis. Medicinal plant raw materials / the USSR Ministry of health. - 11th ed. - M.: Medicine, 1989 - 400 S.] (1:1), spectrophotometric at a wavelength of λ=210 nm, measure the optical density of a solution of saponins, expect their content in a fraction of pre-constructed calibration curve. The content of saponins calculated in terms of oleanolic acid

Example 1. Determination of saponins in plant raw materials (sugar beet roots).

1. Preparation of calibration graph.

The original solutions oleanolic acid (Patent for invention №2258709 of 28.06.2004, the Method of obtaining oleanolic acid) is prepared by dissolving accurately weighed oleanolic acid and subsequent preparation of a series of working solutions by dilution of the source to the desired concentration of ammonium buffer based on the ratio of aqueous phase buffer 1:1. The working concentration range of oleanolic acid is 0.01-0.4 mg/ml as the reference solution, a mixture of distilled water - ammonia buffer (1:1). Spectrophotometric measurements were carried out using spectrophotometer SF-46. Using the average solution concentration, relieve range Pogue is osenia in the wavelength range of 180-300 nm and set the maximum absorption of oleanolic acid (λ=210 nm). In accordance with the established maximum absorption gain calibration dependence of the optical density of the standard solutions of oleanolic acid on their concentration. The calibration equation D=S-0.0044 (- concentration, mg/ml; D - optical density).

2. Preparation of the test solution.

The sample obtained from vegetable raw materials (for example, 250 grams of sugar beet roots) capodistria fraction, weighing about 0.25 g (accurately weighed), dissolved in a volumetric flask of 100 ml capacity in a mixture of water - ammonia buffer (1:1). An aliquot of the resulting solution with a volume of 1 ml was placed in a volumetric flask with a capacity of 25 ml, bring to label the above mixture (concentration of saponins about 0.08 mg/ml).

3. Measurements and calculation of the content of saponins. The resulting solution spectrophotometery at λ=210 nm. As a comparison solution, a mixture of distilled water - ammonia buffer (1:1). The content of triterpene saponins in terms of oleanolic acid are calculated according to the formula:

If the optical density of the test solution D=0,412, the content of saponins in the solution in terms of oleanolic acid is C=0,03648 mg/ml and With the average molecular weight of saponins 1000 g/mol, the content of saponins in the solution amounted to 0.08 mg/ml, the resulting fraction is 80%.

Recip is installed, the data correlate well with data for the determination of saponins by well-known methods [Nagorno VA, Gigina GT, Kartashov, A.//Sugar industry. 1966. No. 8. P.39-43]. The relative error of determination in the case of the analysis of plant materials does not exceed ±6%.

Example 2. Determination of saponins in tablets "Caporal".

1. The construction of the calibration curve analogously to example 1.

2. Preparation of the test solution.

A portion of crushed tablets of about 0.15 g (accurately weighed) was dissolved in a mixture of ammonium buffer - water 1:1 in a volumetric flask with a capacity of 50 ml. Aliquot of the resulting solution with a volume of 1 ml was placed in a flask with a capacity of 25 ml and adjusted to the mark above mixture.

3. Measurements and calculation of the content of saponins.

The solution spectrophotometric - at λ=210 nm, the absorbance value is 0,501. The content of saponins terms of the aglycone calculated according to the formula above, in the test solution, it is 0,0427 mg/ml Using the value of their average molar mass, taking into account the dilution and average weight of the tablet, get the content of saponins - 0,039 g in one pill. The error of determination of saponins by this method is ±3%, which is specified in the ND range of contents (being 0.036-0,044 g in one pill, Rel. error ±10%).

The developed method for determination of saponins does not require the use of specific reagents, is to Express the time required on the I analysis, approximately 20 minutes. The stability of maximum absorption is necessary to conduct repeated measurements of the same sample. Proposed-translation method the content of saponins on the aglycone is rational, as it allows to estimate their number in terms of the reactive part of the molecule that determines its chemical, pharmacological and other types of activities.

The method of determination of triterpene saponins in plant raw materials and medicines, including cooking saponinosteroids fractions, determination of the optical density and the calculation of the contents of saponins of pre-constructed calibration curve, characterized in that for the preparation of fractions as solvent a mixture of water: ammonia buffer 1:1, the optical density is determined at λ=210 nm, and the content of saponins calculated in terms of oleanolic acid.



 

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