CS260286B1 - A method for determining vitamin A acetate - Google Patents

Abstract

The solution is intended for the determination of all-trans vitamin acetate of formula I and formula II in bulk materials by the method of liquid chromatography on silica gel with chemically bound octadecyl groups, the sample being prepared by leaching vitamin A from the bulk material with a mixture of water and a polar solvent miscible with water, with isocratic elution acetonitrile-water (83 : : 17) and detection of the eluted components by measuring the absorption of ultraviolet radiation at a wavelength of 325 nm. The solution is advantageously used for the determination of vitamin A during its production at the stage of inter-operational control, also for the determination of the vitamin A content in bulk materials manufactured for pharmaceutical purposes.

Description

The invention relates to the determination of vitamin A of formula I and formula II in bulk materials by liquid chromatography on silica gel with chemically bonded octadecyl groups.

The method of the present invention, which is novel, not yet described, is an efficient method of controlling and controlling the production process of free-flowing forms of vitamin A acetate of formulas I and II.

JU i.

^CH 3 (i)

The method used to determine vitamin A acetate in bulk products to date is a spectrophotometric method whose low selectivity and the presence of ballasts absorbing at 320 to 330 nm interfere with the accuracy and accuracy of the results (Knobloch E., Physicochemical Methods for Vitamin Determination, 1956). The separation of these ballast substances is partially solved by AO no. 246 254 by sorption on the Al2O3 column.

Vitamin A gas chromatography has been discussed by a number of authors (Vecchi M, Vetter W, Walther V, Jermstad SJ, Schutt G W. Helv. Chim. Acta 50, 5, 1967, 126; Swecley CC, Bentley R., Mehite M., Wells W., J. Amer. Chem. Soc., 85, 1963, 2,497).

Application of this physicochemical method is only possible after previous vitamin A silicification, which, however, as found by the authors, does not proceed quantitatively, the column undergoes partial decomposition and partial isomerization of vitamin A. All these deficiencies are eliminated by the method of vitamin A acetate determination by high performance liquid chromatography. . The authors (Steuerle H., Journal Chrom. 206, 1981, 2, 319-326) used iso-octane-diisopropyl ether absorption chromatography for the determination of vitamin A acetate isomers.

All-trans and 13-cis vitamin A isomers were analyzed in natural materials (Erberg DC, Hereff JC, Petter RH, J. Agric. Food. Chem., Vol. 25, no. 5, 1977) by high performance liquid chromatography on anchored cartridges. reverse phase octodecylsilane type by isocratic elution as well as adsorption chromatography after previous saponification.

The disadvantage of these processes is the complex treatment of the sample (isolation, saponification) and none of them is concerned with the determination of vitamin A acetate in bulk products.

These deficiencies of the spectrophotometric assay in bulk forms of vitamin A acetate have been overcome by elaborating a vitamin A acetate assay by liquid chromatography on chemically bonded octadecyl groups by decomposing a small amount of bulk material with a small amount of water and adding a polar solvent miscible with water. The mixture is shaken vigorously for 5 minutes and after subsequent filtration quantified by an external standard method, the appropriate portion of the prepared solution is fed to a column packed with a sorbent of silica gel coated with chemically bound octadecyl groups under isocratic elution with acetonitrile: water (83:17). of the substances is made by measuring the absorption of ultraviolet radiation at a wavelength of 325 nm.

The invention is advantageously used in the determination of vitamin A acetate in its production in the step of in-process control, also in determining the content of vitamin A in bulk materials produced for feed purposes.

The process of the invention is illustrated by way of example without limiting the scope of the invention.

Example 1

Weigh 0.150 g of bulk material into a 100 ml graduated flask. 2 ml of distilled water, 50 to 55 ° C, are added and the mixture is stirred in a water bath at 55 to 60 ° C for 5 minutes. After cooling to room temperature, isopropyl alcohol is added to the mixture and the volumetric flask is filled up to the mark. The contents were shaken for 5 minutes, then filtered through Whatman filter paper. From the filtrate, 260 µL is fed to 10 µl in a liquid chromatograph.

Determination conditions: 250 mm millimeter, 4 mm cross-section, packed with a spherical sorbent of octadecyl-coated silica gel (LiChrosorb RP-18) of average size

Vitamin A all-trans acetate content in n. j./g =

13-cis vitamin A acetate content in m. j./g = -P 15

7 µm particles, mobile phase of acetohitrile: water (83: 17), flow rate 1.5 cm ³ min ^-1 . A variable wavelength detector set at 325 nm is used for detection.

Quantification is performed by the external standard method.

f = 1.089 - conversion factor to 13 cis isomer, taken from literature (Steuerle, H., Journal Chrom., 206, 1981, 2, 319-326). Analysis result:

13-cis-vitamin A acetate is 67,000 m. U / g of all-trans-vitamin A acetate 360,000 m. j. per gram total: 427 000 m. j./g

The result of the spectral analysis according to PNY is 380,000 m. j./g.

The result of the analysis according to PV agrees with the overall material balance.

P VZ U Th

Ξ-— ---. f. 2 907 000 '

P št n _vz . 2 907 000 n _vz

Example 2

A model sample was prepared with a vitamin A acetate content of 400,000 m. j./g. It was analyzed according to the procedure of Example 1. A content of 396,000 m was determined. j./g. Example 3

A sample of bulk vitamin A acetate from imports with a declared content of 325,000 m was analyzed according to the procedure of Example 1. j./g. The content of 326 400 m was determined. j./g.

Claims (3)

260286 for 10 μL in liquid chromatography. Assay conditions: 250 mm column, 4 mm cross-section, filled with a spherical sorbent of silica gel-coated octadecyl groups (Li-Chrosorb RP-18) of average size All-trans vitamin A acetate content in nj / g = 13-cis vitamin A acetate content in mj / g = - P 15 particles of 7 µm, mobile phase of the aceto-halo: water (83: 17), 1.5 cm 3 min -1 flow rate. at 325 nm. Quantification is performed by the external standard method. f = 1.089 - conversion factor to 13 cisisomer, taken from the literature (Steuerle, H., Journal Chrom., 206, 1981, 2, 319-326). The analysis of: 13-cis-V'itamin A acetate is 67,000 IU. / all-trans-vitamin A gacetate 360,000 IU gram: 427,000 IU / g spectral analysis PNY is 380,000 IU / g. According to PV, the result of the analysis agrees with the overall material balance. Pvz n št Ξ-— ---. f. 2 907 000 Pst. 2,907,000 nv Example 2 A model sample of vitamin A acetate content of 400,000 m / g was prepared. The analysis was carried out according to the procedure described in Example 1. The content of 396,000 IU / g was determined. Example 3 According to the procedure described in Example 1, a sample of bulk acetonitrile A was analyzed with a declared content of 325,000 IU / g. A content of 326 400 m / s was determined. SUBJECT MATTER A method for the determination of vitamin Avzorec I acetate and formula II -Η-, CH: CH, CH 2. Cíh CH? X '' ··. CH 3, CH 3 COCOCH 3 (II) in bulk materials by liquid chromatography on silica gel with chemically bonded octadecyl groups, characterized in that the bulk material is mixed with water, a polar solvent miscible with with water, the mixture is shaken vigorously for 5 minutes and after filtration the filtrate is injected into a liquid chromatograph. 2. Process according to claim 1, characterized in that alcohol C1 to C6 is used as the polar solvent. 3. A process according to claim 2, wherein the polar solvent is acetonitrile, dimethyl sulfoxide or a carboxylic acid amide of the formula OZR-C1-R1-N2 wherein R is H or C1 to C3 alkyl, Whose; R1, R2 are H and C1 to C3 alkyl.