Direct Method of Sucrose and Raffinose Determination Using 5-Diazouracil† ,1 Boguslaw Samotus, Henryk Koloczek, 2 Elz ˙bieta Do ¨rre, Malgorzata Stefaniuk, Andrzej S ´ cigalski, and Adam S ´ widerski Department of Biochemistry, University of Agriculture, Al. 29 Listopada 54, 31-425 Krako ´w, Poland Received June 1, 1998; accepted July 7, 1998 5-Diazouracil can serve for direct sucrose and raffinose determination in pure solutions obeying Lambert–Beer law. Fructose, glucose, melezitose, maltose, and their mixtures interfere in sucrose (and raffinose) determination but their influence can be eliminated. The method was applied to natural honeys and honey substitutes. © 1998 Academic Press INTRODUCTION The role of sucrose in plant metabolism is very well recognized today. In some parts of plants it accumulates in large quantities; in other parts, in trace amounts only. This disaccharide is also an important additive in different products of the food industry and sometimes it can be an indicator of adulteration (e.g., honey). In many plants and natural products sucrose can be found with some other soluble sugars, e.g., glucose and fructose. As a rule sucrose is indirectly determined using any reducing method and two deter- minations are performed: before and after short inversion. The value of sucrose is obtained by subtracting these two determinations and multiplying the result by 0.95. If the relative concentration of sucrose compared with other reducing sugars is high, the precision of the determination is good. On the other hand, a low level of sucrose makes the determination uncertain. In that case, direct sucrose determination should follow. In 1933, while studying the chemical properties of 5-diazouracil [5-diazo-2,4-dihy- droxypyrimidine (DAU)], Raybin (1) discovered a new color reaction of sucrose with this reagent. He examined a set of compounds for the reaction with DAU and found that raffinose was the only sugar to give a similar but fainter color. Four years later Raybin (2) stated that some other sugars with glucose–fructose linkage similar to that of sucrose (1,2-glycosidic bond) showed the blue-green color typical of sucrose and raffinose. Then, other authors applied the “Raybin test” to identify sucrose in some plants and food products. French et al. (3) tested planteose, raffinose, melezitose, and sucrose. They stated that sucrose and raffinose gave a definite bluish precipitate, but planteose and melezitose (which have a 1,2-glycosidic linkage) yielded only a reddish-brown precipitate. However, Cerbulis (4), who examined different sugars in cacao beans taken from the production line † In memory of Professor Boguslaw Samotus. This was the last work supervised by the Professor. 1 This work was partially sponsored by Grant 6P04G03311 from Committee of Scientific Research (KBN, Poland). 2 To whom correspondence should be addressed. MICROCHEMICAL JOURNAL 60, 153–166 (1998) ARTICLE NO. MJ981652 0026-265X/98 $25.00 Copyright © 1998 by Academic Press All rights of reproduction in any form reserved. 153