Bial’s reagent

Bial s reagent (for pentoses) dissolve 1 g of orcinol in 500 mL of 30% HCl to which 30 drops of a 10% ferric chloride solution have been added. 

By careful choice of both the reaction conditions and the phenolic compound used, it may be possible to produce a colour that is characteristic of a particular carbohydrate or related group, so giving some degree of specificity to the method. Thus, Seliwanoff s test uses hydrochloric acid and either resorcinol or 3-indolylacetic acid to measure fructose with minimal interference from glucose. The colour produced by pentoses with orcinol (Bial s reagent) or p-bromoaniline is sufficiently different from that produced by hexoses to permit their quantitation in the presence of hexoses. However, none of the methods based on the formation of furfural or its derivatives can be considered to be entirely specific. 

Staining solution (Bial s reagent), prepared freshly 50 mg orcine monohydrate is dissolved in 12.5 ml distilled water, 20 ml HC1 37%, and 0.5 ml 1% ferric chloride... 

Qualitatively, the presence of the nonulosaminic acids is best indicated by the brilliant-red coloration formed on addition of an acidic solution of p-dimethylaminobenzaldehyde (the so-called direct Ehrlich reaction ), and by the bright-purple coloration which develops upon boiling with Bial s reagent for several minutes at 100°. In addition, the nonulosaminic acids give a blue-violet coloration with Dische s diphenylamine reagent for deoxypentoses and a positive reaction in the tryptophan-perchloric acid test. Since no single one of these color reactions is absolutely specific for a nonulosaminic acid, it is advisable to carry out at least two of these for a qualitative analysis. For quantitative determinations, all four reactions have been employed, using either A/ -acetylneuraminic acid (m. p., 183-185° [a] —32.0°) or methoxyneuraminic acid [m. p., 200° (dec.) [a]o —55.0°] as colorimetric standards. 

A comparison between certain chemical characteristics of the siaUc, iV-acetylneuraminic, and methoxyneuraminic acids, especially their functional groups and the color reactions afforded with Ehrlich s and Bial s reagents, suggested a close relationship between these nonulosaminic acids and indicated that methoxyneuraminic acid might be the methyl glycoside of a completely deacylated sialic acid. This concept was substantiated with the conversion of A-acetylneuraminic acid into methoxyneuraminic... 

Fig. 141. Thin-layer chromatogram of a mixture of gangliosides (Mix.) and individual gangliosides (A, B, C and B) from human brain [96]. Layer silica gel G solvent n-butanol-pyridine-water (3+2 + 1) time of run 2.5—3h amounts ca 50 p.g of each spray reagent Bial s reagent...

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