Improved Benzidine Sugar Reagent for Paper Chromatograms

Abstract

BENZIDINE has been found to be a useful reagent for sugar detection on paper chromatograms^1,3 and for quantitative determination in test-tubes⁴. Horrocks³ first found that benzidine–acetic acid detected reducing aldoses on paper chromatograms, but ketoses were detected only at high concentration. Bacon and Edelman¹ added trichloracetic acid to the benzidine–acetic acid reagent to increase its acid strength and claimed that it detected less than one microgram of glucose in free or combined form. Its sensitivity to fructose was considerably less. An acid is required for the formation of furfuraldehyde from sugars. Furfuraldehyde, or a degradation product of it, then reacts with an aromatic amine or phenol (benzidine in this case) to form a coloured product. It is important that the acid should he strong enough to cause the reaction to proceed, but, in the case of paper chromatography, it should not cause partial destruction of the paper or produce a dark background colour. For this purpose, organic acids (acetic, phthalic, or trichloracetic acid) have been used. However, when mineral acids at low concentrations are used, an improved reagent is obtained without paper destruction or serious background interference.