INTERCONVERSION OF SUGAR FROM ALDOSE TO KETOSE.

A process for oxyanions or mixed complex oxyanions of germanium or tin which form stronger complexes converting aldoses or derivatives thereof into ketoses or derivatives thereof which takes place in the presence of with the ketoses or derivatives thereof than with the aldoses or derivatives thereof. Using the process the equilibrium position in the aldose-ketose reaction is shifted towards higher proportions of ketose. The process is very applicable to the conversion of glucose to fructose particularly when this conversion takes place enzymatically in the presence of glucose isomerase.

1. A process for the conversion of an aldose or aldose derivative to a ketose or ketose derivative, comprising: forming an aqueous solution containing an aldose or aldose derivative and chemically or enzymatically isomerizing said aldose or aldose derivative to a corresponding ketose or ketose derivative in the presence of a complexing reagent which is an oxyanion or mixed complex oxyanion of germanium or tin which forms a stronger complex with the ketose or ketose derivative than with the aldose or aldose derivative, maintaining said aqueous solution under aldose to ketose isomerization conditions, and recovering a product enriched in said ketose or ketose derivative.

2. A process for the conversion of an aldose or aldose derivative to a ketose or ketose derivative, comprising: forming an aqueous solution containing an aldose or aldose derivative and chemically or enzymatically isomerizing said aldose or aldose derivative to a corresponding ketose or ketose derivative in the presence to a complexing reagent which is an oxyanion or mixed complex oxyanion of germanium which forms a stronger complex with the ketose or ketose derivative than with the aldose or aldose derivative, maintaining said solution under aldose to ketose isomerization conditions, and

recovering a product enriched in the ketose or ketose derivative. This invention relates to a process for the conversion of an aldose or aldose derivative to a ketose or ketose derivative in the presence of an oxyanion and in particular to the conversion of glucose to fructose, mannose to fructose, glucose 6-phosphate to fructose 6-phosphate, maltose to maltulose, galactose to tagatose, and lactose to lactulose and to other analogous reactions such as the conversion of xylose to xylulose. The process can be carried out in the presence or absence of an enzyme catalysing the conversion. When the conversion of glucose to fructose is conducted in the presence of an enzyme, glucose isomerase, equilibrium is very often reached when 50 to 55% of the glucose in the reaction medium has been converted to fructose. Hitherto it has not been possible to convert glucose to fructose by a nonenzymic reaction without the production of by-products. There are numerous publications, for example U.S. Pat. Nos. 2,487,121, 3,432,345, 3,558,355 and 3,514,327 and German Pat. No. 1,163,307, relating to non-enzymic conversion of glucose to fructose all of which describe processes in which the fructose produced is accompanied by alkaline degradation and other products arising from the purely chemical and non-enzymic reaction. Because of this any large scale production of fructose up to the present has been based on an enzyme catalysed reaction. In the enzymic conversion process as the proportion of fructose in the enzymic reaction medium increases, the rate of the reaction whereby fructose is produced decreases. Therefore in a commercial process for converting glucose to fructose the fructose yield is optimised by balancing a loss in fructose production against the decreasing reaction rate whereby the proportion of fructose in the reaction medium can be increased. A commercially operated process may be optimised to produce a syrup in which typically 40% of glucose has been converted to fructose. In most chemical non-enzymic conversions of glucose to fructose, the amount of fructose present drops after reaching a maximum. Clearly it is advantageous to increase the proportion of glucose which can economically be converted to fructose during the reaction. This can be achieved by the effective removal of fructose from the reaction medium by the inclusion in the medium of a reagent which forms a stronger complex with fructose than with glucose although the complexing agent may have additional properties that increase or decrease the reaction rate. Reagents which have been proposed for this purpose are borate compounds -- see Y Takasaki, Agr Biol Chem 1971, 35(9), 1371-5 and USP 3,689,362

(enzymic reaction) and J F Mendicino, J Amer Chem Soc 82, 1960, 4975 (chemical reaction). Also arene boronates have been proposed by S A Barker, P J Somers and B W Hatt in UK Specification No 1,369,185 for both chemical and enzymic reactions. Disadvantages of these prior proposals are, in the case of borate compounds, that these are toxic and could constitute a health hazard in a product intended for use as a sweetener in foodstuffs for human consumption and, in the case of benzene boronate it has a limiting solubility, cannot form complexes with a 2:1 ratio of sugar: boronate and high concentrations of fructose in the product cannot be attained at high sugar concentrations ( S A Barker, B W Hatt and P J Somers, Carbohydrates Res, 26 (1973) 41-53). Thus these complexing reactions do not lend themselves readily to use in commercially operated conversion processes. In order to develop a commercial process in which the proportion of fructose in the syrup produced is increased it is necessary to find a complexing reagent which does not have such disadvantages associated with its use. According to the present invention we provide a process for the conversion of an aldose or an aldose derivative to a ketose or ketose derivative wherein the conversion takes place in the presence of a complexing reagent which is an oxyanion or mixed complex oxyanion of germanium or tin which forms a stronger complex with the ketose or ketose derivative than with the aldose or aldose derivative. Also according to the present invention we provide a process for the conversion of an aldose or aldose derivative to a ketose or ketose derivative wherein the conversion takes place in the presence of a complexing agent which is an oxyanion or mixed complex oxyanion of germanium which forms a stronger complex with the ketose or ketose derivative than with the aldose or aldose derivative. Very suitably the aldose derivative is an aldose phosphate or a glycosyl derivative of an aldose. Whilst the invention is applicable to a wide range of conversions and in particular to those conversions specified hereinabove it is most usefully employed in the conversion of glucose to mannose to fructose. During a conversion an enzyme catalyst may be present where this enables milder reaction conditions to be employed or has other advantages such as the selective action of the enzyme employed on only one isomer (D or L) of the aldose or aldose derivative. When an enzyme-catalysed conversion is carried out the enzyme may be present in solution or in an immobilised form on a solid matrix that may be a living cell, an inactivated cell or any other suitable support. The enzyme may also be in soluble form. The isomerasetype enzyme suitable for conversions of the type (examples listed in Table

A) to which the invention is applicable may constitute one or more of a series of enzymes engaged in sequential reactions.

The present article contains a study of certain changes produced when dilute aqueous solutions of various sugars are boiled. Interest in the subject was aroused because it was observed that in a solution of glucose so treated fructose was apparently produced.1 Although the interconversion of glucose, fructose, and mannose in alkaline solution has been repeatedly studied since Lobry de Bruyn and Van Eckenstein first observed the phenomenon,2 results of quantitative studies similar to those reported do not seem to be available. The technical procedure was as follows: Pure glucose from the Bureau of Standards was dried over calcium chloride and dissolved in distilled water. Seventy-five cc. of a solution so prepared were boiled on an electric hot plate under a reflux condenser. At intervals the flask containing the solution was rapidly cooled in ice water and a sample removed for analysis. The solution was then reheated rapidly and the boiling continued. The changes in the concentration of reducing substances were determined by the copper reduction method of Benedict3 and variations in the intensity of the resorcinol reaction by the technique described by Roe4 for the determination of fructose in blood. The purest glucose and fructose obtainable were used as standards. The reaction values were estimated colorimetrically at room temperature. Pure commercial mannose and galactose prepared for the bacteriological laboratory were used in some experiments. Water solutions of glucose were studied first. It soon became evident that several conditions affected the composition of the boiled mixture. One of these was the concentration of glucose initially present. When the solution was very dilute the maximum concentration of fructose was generally reached after boiling for 4 hours, but in stronger ones such a value was not reached in very much longer periods. Results of analyses made after 6 hours are given in Table I. It can be seen that the concentration of fructose present at this time varied with the concentration of glucose initially present, but was not parallel to it. Since there was very little difference between the percentage composition of the products obtained by boiling 0.02 and 0.1 % glucose in a neutral buffer mixture (Table) it seems possible that variations in acidity may have been one of the causes of this lack of parallelism.
This invention relates to a process for effecting aldose to ketose conversion and in particular to a process for converting glucose to fructose or a mixture containing glucose and fructose or to a process for increasing the proportion of fructose in a mixture containing glucose and fructose.

Fructose is sweeter than its naturally-occurring isomer glucose and in consequence high-fructose syrups have advantages for use as sweeteners in products such as confectionary and cakes. During the past twenty years there has been considerable research into processes for producing high fructose syrups for such uses. Processes available for converting glucose to fructose or fructose-containing syrups can be enzymatic, using the enzyme glucose isomerase, or chemical. Enzymatic processes are usually preferred since chemical processes tend to produce complex mixtures of sugars comprising mannose and there is a tendency for fructose produced during chemical processes to revert to glucose or mannose under the conditions employed in the process. Chemical isomerisations of aldoses or ketoses under alkaline conditions, using for example calcium hydroxide have been extensively studied. In this and similar isomerization reactions an equilibrium is established between the ketose and the related C-2 epimeric aldoses, for example between fructose and glucose and mannose. It is a particular feature of these isomerisations that they are reversible so that for example the fructose produced by isomerisation of glucose and/or mannose can be converted back to glucose and mannose. CaCl2 (0.1M) has been used in an alkaline isomerisation of glucose and/or mannose to fructose with 0.25M KOH (G. De Wit, A. P. G. Kieboom and H. Van Bekkum, Carbohyd. Res., 74 (1979) 157). In this work CaCl2 was shown to enhance the formation of a UV absorbing material attributed to an enediol-anion species. According to the present invention we provide a process for effecting conversion of an aldose, or a substituted aldose, to a ketose, or a substituted ketose, which comprises a step wherein a reaction mixture is formed comprising an aldose or substituted aldose, water and a halide of a metal from Group II of the Periodic Table, preferably a chloride, the metal halide being present in the reaction mixture in a concentration within the range 0.5 molar to saturation. The metal halide may be introduced into the reaction mixture either as the halide per se or as a precursor which forms the halide under the conditions prevailing in the reaction. It may be introduced in the form of a complex with the aldose or substituted aldose. A feature of the process of the invention is its specificity. By this process an aldose is essentially converted solely to the corresponding ketose with essentially no concomitant production of the C-2 epimer. Additionally there is essentially no tendency for the produced ketose to revert to the aldose from which it is derived. The process of the invention is generally applicable to the conversion of aldoses into ketoses or of substituted aldoses into substituted ketoses. It may be applied to the conversion of an aldose into a ketose or into an aldose-ketose mixture. Alternatively it may be used to convert an aldose-containing mixture of sugars into a mixture containing a ketose or an aldose and a ketose. It is also very suitable for converting a sugar mixture comprising an aldose and a ketose into a mixture containing an increased proportion of a ketose. Preferably the aldose is D-glucose or L-glucose and the corresponding ketose is D-fructose or L-fructose respectively. The process of the invention is very usefully applied to effect conversion of a mixture comprising glucose

and fructose into one containing an increased proportion of fructose. The aldose or aldose-containing mixture may have been produced and be supplied directly from another process, for example a process for the solubilisation and/or hydrolysis of cellulose such as that described in our published European Patent Specification No. 44622. Other conversions of aldoses to ketoses or of substituted aldoses to substituted ketoses which may usefully be effected by the process of the present invention include xylose to xylulose, lactose to lactulose and the following: D-mannoseD-fructose L-mannoseL-fructose D-galactoseD-tagatose D-taloseD-tagatose 6-Deoxy-L-mannose6-deoxy-L-arabino-hexulose 6-Deoxy-D-glucose6-deoxy-D-arabino-hexulose (otherwise 6-deoxy-D-fructose) For a variety of reasons including general convenience the preferred halide for use in the process is calcium chloride. Other suitable but less preferred metal halides are the chlorides or bromide of magnesium and zinc and calcium bromide. The preferred halide calcium chloride is available in anhydrous form or in a number of hydrated forms including the monohydrate CaCl2 :H2 O; the dihydrate CaCl2 :2H2 O, the tetrahydrate CaCl2 :4H2 O and the hexahydrate CaCl2 :6H2 O. The manner in which it is included in the reaction mixture depends upon which form is used. Suitably the conversion of the aldose or substituted aldose to the corresponding ketose or substituted ketose is effected in a reaction mixture the pH of which is on the acid side of neutrality. Preferably the pH of the reaction mixture when calcium or magnesium halides are used is within the range 2 to 7 and especially within the range 3 to 6.5. When zinc halides are used lower pHs are preferred. The pH of the reaction mixture may be controlled by any suitable means. Suitable means include addition of calcium carbonate, calcium hydroxide or other suitable bases either initially or during the course of the conversion. The pH may also be controlled by additions of the anhydrous chloride of a Group II metal, particularly calcium chloride, during the reaction and by suitable variations of the source of the calcium or other Group II metal chloride and of the sugar content of the reaction mixture. A suitable temperature for the contacting step is in the range 20 to 100 C., temperatures in the range 30 to 85 C. being preferred. Suitable periods are dependent on other reaction parameters such as temperature, concentrations of reagents, and pH.

Lower temperatures for instance require longer reaction times. Preferably the reaction should be performed with the exclusion of oxygen. After the reaction, the metal and/or halide ions in the resulting product may be separated by any suitable method and separated ions may be returned to the reaction step by suitable means as part of a continuous process. The ketose-containing syrup produced by the process may, after separation of metal and/or halide ions, be used as it is even if it contains a mixture of sugars or, if it is a mixture, it may be treated to separate individual sugar components. Where an aldose-ketose mixture is produced and the components are separated, the aldose component may be returned to the reaction step for further treatment. It is an advantage of the process of the invention that, when it is used to convert glucose into fructose or into high fructose syrups, significant amounts of mannose are not obtained. This is most unusual for a chemical process for converting glucose to fructose. It is also possible by the process of the invention to achieve simultaneous conversion of two C-2 epimerically related aldoses to the same ketose. Thus glucose and mannose in admixture can be converted to fructose. An example of this effect would be the exploitation of the known occurrence of a -glucan and a mannan in yeast cells (Saccharamyces cerevisiaie) initially by using CaCl2 /HCl for selective extraction/hydrolysis and after removing HCl, the calcium chloride used alone to produce fructose. It has also been shown that two other C-2 epimerically related aldoses, galactose and talose can be converted into the same ketose tagatose.