Paper Chromatographic Separation and Determination of Some Water-Soluble Vitamins

1952 ◽  
Vol 24 (12) ◽  
pp. 1952-1956 ◽  
Author(s):  
J. A. Brown ◽  
M. M. Marsh
1969 ◽  
Vol 111 (4) ◽  
pp. 401-406 ◽  
Author(s):  
S. Usha Lakshmi ◽  
L. K. Ramachandran

The characteristic purple colour formed by N-formyl-N′-2,4-dinitrophenyl-hydrazine in the presence of piperidine and acetone was made the basis of a new quantitative method for the determination of formyl groups. Samples containing N-formyl groups (up to 0·4μmole) are hydrazinolysed at 97–98° for 1hr. and are dinitrophenylated after the removal of excess of hydrazine. Interference from 2,4-dinitrophenylhydrazine is eliminated by subjecting the dinitrophenylated samples to chromatography on an alumina column. Interference arising from the formation of N-acetyl-N′-2,4-dinitrophenylhydrazine, when determining formyl groups in samples containing acetyl, can be avoided by a paper-chromatographic separation before analysis. A standard procedure is described. The method gives satisfactory results when applied to N-formyl-amino acids. Gramicidin, when analysed by this method, was found to contain 0·89 mole of formyl group/mole for a molecular weight of 1880. The method indicated the absence of formyl groups from lysozyme, a protein known not to contain such groups. Generally, the analytical values obtained by the method are within 100±4% of theory.


Author(s):  
R. D. Ostapiv ◽  
V. І. Tkachenko

Vitamins are widely used in veterinary medicine, in particular, as components of premixes, feeds and feed additives to increase the productivity of farm animals. Most of these feed additives are a complex amino acid and vitamin mixtures (of ten active substances or more) based on the matrix of plant or animal origin, which can be enriched with protein. When the analyst is faced with the difficult task of analyzing such a mixture, it is usually necessary to use several variants of the HPLC method (high-performance liquid chromatography), which increases the time and cost of analysis of the premix or feed additive. The aim of this work was to optimize the validated method of analysis of riboflavin in premixes and feed additives available in the laboratory for simultaneous (during one chromatographic separation) determination of the content of the maximum amount of water-soluble vitamins. A Waters liquid chromatograph equipped with an Alliance 2690 separation module with a PAD 996 diode array detector was used for separation. Water-soluble vitamins were separated on a Luna Omega Polar C18 250 × 4.6 mm, 5 μm chromatographic column. As solvents were used 0.05 M phosphate buffer pH 3.0 and 0.1 M sodium hydroxide solution (for folic acid). The article describes the stages of liquid chromatographic separation parameters optimization of twelve vitamins and vitamin-like substances (thiamine hydrochloride, riboflavin, flavin mononucleotide phosphate, nicotinic acid, nicotinamide, calcuim pantothenate, phosidinobacino acidorinolobin, hydrocyanide). Chromatograms of standard samples solutions are given, which show the specificity of the technique. Isocratic and gradient elution schemes are investigated. Some parameters of chromatographic peaks under conditions of gradient separation of vitamins are estimated. The parameters of the chromatographic peaks do not exceed the limits recommended in the State Pharmacopoeia of Ukraine (SPU 2.0), that will allow to further carry out a more complete validation of the method for quantitative determination of water-soluble vitamins in various complex matrices: injection and oral solutions, feeds and premixes.


1964 ◽  
Vol 36 (3) ◽  
pp. 502-505 ◽  
Author(s):  
Herbert. Siegel ◽  
A. B. Bullock ◽  
G. B. Carter

1984 ◽  
Vol 67 (5) ◽  
pp. 994-998 ◽  
Author(s):  
Thomas S Hudson ◽  
Shyamala Subramanian ◽  
Rebecca J Allen

Abstract Until recently, liquid chromatographic (LC) methodology for pantothenic acid, biotin, and B12 (cyanocobalamin) has been only marginally successful. These vitamins are difficult to determine by conventional LC techniques and UV detection at 254 or 280 nm, because either the chromophore is inadequate for detection or interference from co-eluting vitamins is overwhelming. Biotin and B12 are usually present in pharmaceutical products at concentrations 100-1000 times lower than other commonly occurring water-soluble vitamins. Co-extraction of all water-soluble vitamins results in gross interferences, especially in LC when the interfering vitamins co-elute with biotin or B12. In addition, pantothenic acid and biotin are colorless in solution and do not exhibit strong UV absorption above 240 nm. As a result, they must be quantitated either by using a low UV wavelength for detection or by derivatizing the vitamin to obtain an adequate chromophore. A description of procedures for LC determination of pantothenic acid, panthenol, cyanocobalamin, and biotin in pharmaceutical products is presented. Pantothenic acid has been measured by using both a derivatization technique and low UV wavelength detection. Biotin has been quantitated by using low UV wavelength detection. The limitations of these techniques are also discussed. Chromatographic separation of cyanocobalamin is complicated by co-eluting vitamins such as riboflavin. It is detected by using the 546 nm wavelength where riboflavin does not interfere.


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