Monosaccharide is a generic term for a hydrocarbon having a carbon chain of 3 or more carbon atoms, and in nature there are a number of monosaccharides as much as 40 or more species in a variety of forms including free forms or derivatives of metabolites or biosynthetic products as well as complex carbohydrates, oligo-saccharides or polysaccharides. Industrially most important monosaccharide is D-glucose, of which the process for production has been established on an industrial scale using a method of hydrolysis of starch with an enzyme such as amylase, and has been industrially most widely used in the fields of foods, cosmetics and pharmaceuticals in large quantities. Further, D-fructose which is produced by conversion of D-glucose with isomerase is an industrially important sugar. Other industrially applied process for production of some monosaccharides includes production of L-sorbose from sorbitol by an acetic acid bacterium, production of D-galactose from lactose by β-galactosidase, and production of D-xylose from xylan by acid hydrolysis. A large number of other monosaccharides are difficult to produce in industrially large quantities because of industrial and economical limitation in raw materials, generative reaction and yield, and they almost have not yet been utilized industrially.
The present inventors have paid attention to such expensive and rarely available monosaccharides and gave a name of rare sugars to these monosaccharides; and they have continued to assiduously study the generative reaction of rare sugars for a long period of years. The present inventors have conducted a study for development on monomeric rare sugars including both D-psicose and D-allose. They, as mentioned in the non-patent document 1, have developed a process for producing D-psicose directly from D-glucose in combination of an industrially used D-xylose isomerase with D-ketohexose 3-epimerase discovered by the inventors, and thus opened a new route to a process for the continuous production of D-psicose.
D-Psicose which is generally known to exist in molasses of sweet potatoes in small quantities, is one of industrially expected sugars, particularly in food industry, since it has a sweet taste of good quality similar to D-fructose and is best suited as hardly fermentative and low calorie sugar. That is, D-psicose draws attention as a sugar which does not promote the synthesis of fat and does not accumulate body fat, particularly intraperitoneal fat in comparison with monosaccharides such as D-glucose or D-fructose (non-patent document 2). Further, it has been reported that the available energy value of D-psicose is approximately zero (non-patent document 3).
Thus, based on their success in producing D-psicose, they have succeeded in large scale production of a rare sugar D-allose by isomerization of D-psicose using L-rhamnose isomerase. In this connection, the present inventors have discovered L-rhamnose isomerase (also referred to as “L-RhI”) produced by Pseudomonas stutzeri LL-172 strain, and further elucidated that this enzyme works to catalyze the isomerization reaction of L-rhamnose to L-rhamnurose as well as L-rhamnurose to L-rhamnose. L-Rhamnose isomerase produced by Pseudomonas stutzeri LL-172 (IPOD FERM BP-08593) also acts on isomerization between D-allose and D-psicose, and thus this enzyme is capable of producing D-allose from D-psicose. In this situation, it is necessary to use an enzyme of the Pseudomonas stutzeri LL-172 origin in order to produce D-allose from D-psicose (patent document 1). Because of discovery of this enzyme, practical use of the reaction for production of rare sugars aimed by the present inventors took a great step forward; particularly, this was remarkable in production of D-allose from the substrate D-psicose on a large scale. Since it became apparent that this D-allose has various physiological activities, the research is rapidly developing. At present, it has become apparent that D-allose has an effect such as cancer cell inhibition or anti-oxidant effect important in a medical field (patent document 2).
The production of D-allose, however, includes the most expensive bottleneck step for separating D-allose from D-psicose during the course of production. In order to produce D-allose as a single product, the product contained in a mixture of sugar solutions formed due to the property of the enzyme has to be separated from the raw materials. In the reaction with L-rhamnose isomerase, D-psicose and D-allose reach to an equilibrium state of 7:3, from which mixture of sugar solutions D-allose is separated by column chromatography. Though this operation for separation is naturally required as an essential step for purification of a monomeric sugar, it is also true that the time and labor are required as the production scale of rare sugar becomes huge.
On the other hand, “sugar solution of grape sugar/fruit sugar” containing D-glucose and D-fructose has practically been used as a mixture of sugars at present. The sugar solution of grape sugar/fruit sugar has industrially produced as alternative sugar having a compositional ratio similar to ordinary sugar by converting approximately a half of D-glucose to D-fructose using xylose isomerase, and is commercialized as gum syrup for use in coffee or tea. The mixed sugar solution of D-glucose and D-fructose is difficult to crystallize and utilized in a liquid form not as crystals. This is considered because of very high solubility of D-fructose in water, requiring much time and labor and producing a disadvantage in production cost; thus, it has been utilized as such without crystallization. The name of isomerized sugar usually indicates a mixed sugar solution of D-glucose and D-fructose. This is owing to the reason that at present only the sugar solution produced by isomerization reaction of D-glucose as raw material with xylose isomerase has been industrially made fit for practical use.    Non-patent document 1: Journal of Fermentation and Bioengineering, 80, p 101, 1995    Non-patent document 2: Asia Pacific J. Clin. Nutr. 10, 233-237,    Non-patent document 3: J. Nutr. Sci. Vitaminol 48, 77-80, 2002    Patent document 1: PCT WO2004/063369 A1)    Patent document 2: PCT WO03/097820 A1