JPS6185397A - Separation of saccharides - Google Patents

Abstract

PURPOSE:To separate a saccharide component from a mixture of specific saccharides, in high efficiency, by carrying out the liquid chromatography using a column packed with a specific ion exchange resin, and using water or an aqueous solution as an elutant. CONSTITUTION:A saccharide mixture containing sucrose, maltose, fructose and sorbitol is separated by the liquid chromatography using one or more columns packed with a apcking agent consisting of an SO3<->-containing ion exchange resin (e.g. sulfonated porous styrene-ethylmonovinylbenzene-divinyl benzene terpolymer) wherein the counter ion of the SO3<-> is Ag<+> and Ba<++> and/or Sr<++>, and using water or an aqueous solution (e.g. 10<-4> M/l aqueous solution of NaOH) as an elutant. The volume ratio of the resin having Ag<+> as the count er ion of SO3<-> to the resin having Ba<++> and/or Sr<++> is 9/1-1/9.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Application Field) The present invention relates to sugars and sugar alcohols, particularly sucrose and maltose, which are commonly contained in foods.

This invention relates to a method for separating fructose and sorbitol using liquid chromatography.

(Prior art) Methods for separating saccharides by liquid chromatography include: 1) partition-type separation using a filler chemically bonded with amine groups (for example, silica gel chemically bonded with aminopropylsilyl groups); Journal Chromatography (J, Cbromatogr, ) 144 volume 16
Page 9 (1977, 1.1.2) Separation by molecular sieve using cross-linked dextran gel etc. [written by S.C.Churms].

Advances in Carbohydrate Chemistry
rateChemistry) Volume 25, page 13, Academic Press
Published in 1970, USA].

3) Separation as boric acid complex ions using anion exchange resin (Analytical Chemistry (Ana ICb)
em, ) vol. 52, p. 1079 (19SO)), 4)
Separation using a compromise mode of molecular sieving and partitioning using a salt-type cation exchange resin [Journal Chromatography (
J, Chromatogr., Vol. 103, p. 229 (1975)], etc. are known. 2 of these
Methods 4) and 4) use water as an eluent and are therefore easy to operate.

(Problems to be solved by the invention) However, it is difficult to separate sugars (sucrose, maltose, fructose) and sugar alcohols (sorbitol), which are abundantly contained in foods, especially confectionery.

This is not possible with method 2) above. Further, even with the method 4) above, a satisfactory separation result has not been achieved.

Generally, when saccharides are separated using salt-type cation exchange resin particles, it is known that the separation performance depends on the type of counter ion (the type of cation that constitutes the salt).

In the separation of trisaccharides, cations other than Ag" ions, such as Lr", Na", K++ Rb", C
Alkali metal ions such as s+, Mg, Ca
, Sr, alkaline earth metal ions such as Ba, Fe
, Pe, Co.

Transition metal ions such as Ni'' and Cu2+ have no effect as counter ions, and Ag+ ions are suitable counter ions for separating the trisaccharides sucrose and maltose.However, when the counter ions are Ag4 ions.

Good separation of fructose and sorbitol is possible. Suitable counter ions for separating fructose and sorbitol are alkaline earth metal ions such as Sr'' and Ba2+, and by using one or more of these as a counter ion, fructose and sorbitol can be separated. However, good separation of multiple trisaccharides, namely sucrose and maltose, is not possible.

Therefore, the present invention solves the problems of the prior art described above,
An object of the present invention is to provide a method for suitably separating a saccharide mixture containing sucrose, maltose, fructose, and sorbitol by liquid chromatography.

(Means for Solving the Problems) The present invention provides SO3-group-containing ion exchange resin particles containing sucrose, maltose, fructose, and sorbitol from a saccharide mixture containing each of them.
A method for separating saccharides, characterized by separation by liquid chromatography using water or an aqueous solution as an eluent using one or more columns packed with a packing material having Ag+ and Ba++ and/or Sr++ as a 3-group counterion. Regarding.

The ion exchange resin particles in the present invention preferably have +SO3- groups in an amount of 2 meq/g or more per gram of resin, and particularly preferably 3 to 5 meq/g.When the SO3- groups are less than 2 meq/g, .

Hydrophilicity tends to be insufficient. In terms of separation performance, cation exchange resins are essentially SO3H groups or SO3
It is preferred that the H group does not have any groups neutralized by inorganic cations other than those mentioned above. here.

"Substantially" means that the SO31-1 group or its neutralized group is not detected by neutralization titration or the like.

In addition, the particle size of the ion exchange resin particles in the present invention is
1 to 20 μm for liquid chromatography analysis and 20 to 20 μm for liquid chromatography fractionation.
Preferably, the thickness is 300 μm.

In addition, there is no particular limit to the degree of pores of the ion exchange resin particles in the present invention, but the exclusion limit is 400 to I
It is preferable to set it within the range of 0'.

The SO3-group-containing ion exchange resin particles are preferably sulfonated porous styrene-divinylbenzene copolymer particles. In such ion exchange resin particles,
The 503-group is bonded to a styrene-divinylbenzene copolymer, which is a copolymer of divinylbenzene and a styrene monomer and contains 2 to 60% by weight of divinylbenzene. is preferable. As the styrenic monomer.

Examples include styrene, ethyl monovinylbenzene, vinyltoluene, α-Me-styrene, etc., and may also contain other monomers such as vinyl acetate, methacrylic acid, and acrylic acid (in a range of i75 or less).

The styrene-divinylbenzene copolymer is in the form of particles having pores.

The sulfonated porous styrene-divinylbenzene copolymer particles can be produced by a conventionally known method.

For example, TTF, divinylbenzene, and styrene monomers are subjected to suspension polymerization in the presence of a water-insoluble organic solvent such as amyl alcohol or toluene, and the resulting particles are isolated and swollen with a swelling agent such as dichloroethane or trichloroethane. It can be obtained by adding concentrated sulfuric acid or chlorosulfuric acid, etc., and carrying out a sulfonation reaction at room temperature to 120°C. The above ion exchange capacity can be adjusted by appropriately adjusting the reaction conditions during the reaction with concentrated sulfuric acid or chlorosulfuric acid.1 The degree of pores of the ion exchange resin in the present invention is determined during the suspension polymerization. The particle size can be adjusted by appropriately selecting the suspension polymerization conditions, mainly by adjusting the type and amount of the water-insoluble organic solvent.

Note that it is preferable to use divinylbenzene in an amount of 5 to 60% by weight based on the total amount of the above reaction components.

In addition, as styrenic monomers, the above-mentioned ones tend to be used.
．． Other monomers such as vinyl acetate, methacrylic acid, and acrylic acid may be used in an amount of 5% by weight or less and 11% or less based on the total amount of reaction components. In addition, during RS polymerization, peroxides such as benzoyl peroxide, etc.

Commonly used polymerization initiators can be used.

I cut it. - The sulfonated porous styrene-divinylbenzene copolymer particles have an ion exchange capacity of 2 meQ/g
Those mentioned above are preferable, and those of 3 to 5 meq/g are particularly preferable.

The filler used in the present invention includes SO3- group-containing ion exchange resin particles that simultaneously contain Ag+ and Ba++ and/or Sr++ as counter ions for the 503-group, and those that contain only Ag+ as a counter ion. with Ba++ and/or Sr□+ as counterions
Only those with the following can be used. However, for those with only Ag+ as a counter ion and those with only Ba++ and/or Sr++ as counter ions, these should be mixed and packed in a column, or they should be packed in columns separately and connected. used. That is,
It is sufficient that Ag+, which is a counter ion of the 503-group, and Ba+4 and/or Sr2 are present in the column packing as a whole.

It is preferable that Ag+ and Ba" and/or Sr++, which are counter ions of SO3- groups present in the column packing material, exist in an equivalent ratio of the former/latter in a range of 9/1 to 1/1. This ratio If it is too large, the separation of sucrose and maltose will be good, but the separation of fructose and sorbitol will be poor; if it is too small, the separation of fructose and sorbitol will be good, but the separation of sucrose and maltose will be poor. - The above equivalent ratio is preferably 4/1 to 2/1.

Preferably, the filler is prepared and used as follows to adjust the ratio of the counterions.

That is, SO3- has only Ag+ as a counter ion.
Group-containing ion-exchange resin particles and 503-group-containing ion-exchange resin particles having only alkali metal ions and/or alkaline earth metal ions as counterions are separately prepared, and each of them has a 503-group-containing ion-exchange resin particle containing only an alkali metal ion and/or an alkaline earth metal ion as a counterion. Measure the equivalent weight of the counter ions in advance, use this as a guideline, decide the weight ratio or volume ratio to be used for each, and either mix them and fill the column, or fill them in separate columns and then connect them. However, it is easy and preferable to use. Therefore, for K, two types of 5o3-group-containing ion exchange resin particles with almost the same characteristics such as particle size, ion exchange capacity, and porosity are prepared, and the same equivalent amount of the niger loop ion is present in each as a counter ion. It is particularly preferable to do so. In this case, 1 and both are A g4/' r(a '+
And/or Sr++ may be used in an equivalent ratio of 9/1 to 1/1 by weight or volume of 9/1 to 1/1, and is easy to handle.

In the ion exchange resin particles in which the counter ion of the 503- group is Ag, the counter ion of the SO1 group is Na4. S which is H+ etc.
It is obtained by treating ion exchange resin particles containing 03~ groups with an aqueous solution of a water-soluble silver salt such as silver nitrate.

In addition, the counter ion of the 503- group is Ba++ and/or Sr
The SO3- group-containing ion exchange resin particles that are ++ are S
SOa in which the counter ion of the O3- group is Na", II"
It can be obtained by treating - group-containing ion exchange resin particles with an aqueous solution of varius and/or strontium metal salts, hydroxides, and the like.

Furthermore, simultaneous KAg'' and B as counterions of the SO3- group
The SO3- group-containing ion exchange resin particles having a++ and/or Sr''1 are prepared by adding SO3-group-containing ion exchange resin particles whose counter ion to the SO3 group is Ag'' to an appropriate amount of Ba''.
1 and/or Sr"k, or
l'18 (the counter ion of the h- group is Ba++ and/or Sr
++ SO3- group-containing ion exchange resin particles are treated with an aqueous solution containing an appropriate amount of Ag'. Or, the counter ion of the 503- group is Na+.

It can be obtained by treating ion exchange resin particles containing SO3- groups such as H+ with an aqueous solution containing an appropriate amount of Ag+ and an appropriate amount of Ba++ and/or Sr++.

In the present invention, an aqueous swimsuit solution is used as an eluent, and the aqueous solution is methanol.

Organic solvents such as ethanol, propatool, acetonitrile, and tetrahydrofuran, salts, acids, or alkaline substances diluted to such an extent that the counter ion of the SO3 group in the SO3 group-containing ion exchange resin particles are not replaced, dissolved in water. be.

Eluent and 1. In this case, it is particularly preferable to use an aqueous solution of an alkaline substance. This is because kneading extends the life of the column packing material more than other eluents.

Examples of the alkaline substances used include hydroxides, hydrogen carbonates, and carbonates of alkali metals and alkaline earth metals, which exhibit alkalinity when made into an aqueous solution, and specifically include I, ioH, NaOH.

Xo)T, Sr(OH)2, Ba(OH)
z, NaHCOs, KHCOs.

Nat CO3, K2CO3"4. These alkaline substances are used at a concentration of 5 x 10-5 to 10-3 M/l. If the concentration is less than 5 x 10-5 Nj//, the life of the column packing material will be shortened. If it exceeds 10-3 M/l.

Ion exchange with the counter ion of the column packing material can no longer be ignored, and the separation performance changes.

In the method of the present invention, the sample is injected from the sample injection port,
High-performance liquid chromatography analysis in which the above eluent is passed through a column packed with double column packing materials, and the passing material is detected by a detector to draw a chromatogram. It can be applied to preparative separation, etc., in which a column internal pressure sample is passed through and collected every 9 fractions. In such a separation method, the column packing material used in the present invention does not function as an ion exchange resin.

Also, to fill the column with column packing material.

A conventionally known method can be used, and it is preferable to press fit the slurry into a 9% slurry.

9 In the present invention, sucrose, maltose,
The form of the saccharide mixture containing fructose and sorbitol is arbitrary. However, when applying liquid chromatography, it is made into a suitable form 1, for example, an aqueous solution in which at least the above liquid and a sugar alcohol are dissolved.

(Function) In the present invention, since the counter ion of the SO1 group in the column packing material is Ag+, it is useful for separating sucrose and maltose. On the other hand, the counter ion of SO3- group is B
a''/Sr++ helps in the separation of fructose and sorbitol.The separation of these sugars and sugar alcohols is not inhibited by each other by using the above two types of counterions, and therefore, it is synergistic. becomes possible.

(Example) Next, an example of the present invention and a comparative example will be shown.

Example 1 Particle size 12±2 μnl, ion exchange capacity 4.3 meQ
Glass Filter J [Put it on. A 50% silver nitrate aqueous solution 11 is poured into this, and the counter ion of the 503-group is completely KAg'', and the water is swollen with the drained water washed with water to form a slurry.
shall be.

Particle size 10 μm, ion exchange capacity 4.3 meq/g
and 10 g of sulfonated porous styrene-dihinylbenzene copolymer particles with a degree of crosslinking of 10% were swollen with 150 g of water to form a slurry, and Ba(OH)2.
An aqueous solution prepared by dissolving 6.8 g of 81-T2O in water and binding it to 500 m/m was mixed and stirred for 1 hour. Then filter,
Wash with water, swell with water and pressurize to form a slurry. This is referred to as slurry (B).

Mix slurry (A) and slurry (Bl well, 107
A stainless steel column of φ×30011II11 was filled to obtain a column fcl.

This was set in a high-performance liquid chromatography analyzer, and a sample containing sucrose, maltose, glucose, maltitol, fructose, and sorbitol (hereinafter referred to as sample I) was analyzed. The analysis conditions were a flow rate of 1
．． 0+nj? /min, eluent is 10””M/l NaOH
The aqueous solution and column temperature were 60° C., and a differential refractometer was used as a detector to obtain a chromatogram. The obtained chromatogram is shown in FIG.

In Figure 1, beak 1 is sucrose.

Beak 2 is maltose and beak 3 is glucose.

Beak 4 is maltitol, Beak 5 is fructose, and Beak 6 is sorbitol.

Example 2 In Example 1, Ba (OH)2 ・8 HzO6,
8g instead of Sr (OH)2.81-ho 5.
A column packing material was obtained in the same manner as in Example 1 except that 7 g was used.

After this, it was packed into a column in the same manner as in Example 1.

A chromatogram of sample 1 was obtained by high performance liquid chromatography. The obtained chromatogram is shown in FIG.

In FIG. 2, each peak name is the same as in Example 1.

Example 3 In Example 1, Ba (OH)z ・8 H2O
Ba (OH)2.8 H2O3 instead of 6.8g,
A column packing material was obtained in the same manner as in Example 1 except that 4g and 5r(OH)z.81hO2, 9g were used.

Thereafter, a column was filled in the same manner as in Example 1, and a chromatogram of sample (2) was obtained by high performance liquid chromatography (r). The obtained taromadogram is shown in FIG.

In FIG. 3, each peak name is the same as in Example 1.

Comparative Example 1 A column (C) was obtained in the same manner as in Example 1, except that only the slurry (Al) was packed into the column.

A chromatogram of sample r was obtained by high performance liquid chromatography. The obtained chromatogram is shown in FIG.

In Figure 4, beak 1 sucrose.

Beak 2 is maltose, Beak 3 is H glucose.

Beak 4 is maltitol, Beak 8 is a combined beak of fructose and sorbitol.

Comparative Example 2 A column (C) was obtained in the same manner as in Example 3, except that only the slurry fil was packed.

A chromatogram of sample (2) was obtained by high performance liquid chromatography. The obtained chromatogram is shown in FIG.

In FIG. 5, each peak name is the same as in Example 1.

As is clear from the chromatogram, it can be seen that the separation of shootase and maltose is insufficient.

A schematic diagram of the apparatus used for the analyzes above is shown in FIG. 6. In this apparatus, the eluent is stored in a container 10 and is drained by a pump 11 through a pipe 12, through a column 13 and a detector 14' into a container 15. A sample is introduced through an injection port 16 (for example, a three-way cock), and a chromatograno recorder 17 is connected to the detector 14 .

(Effect of the invention) According to the invention, sucrose, maltose.

Samples containing fructose and sorbitol can be separated very well.

[Brief explanation of drawings]

FIG. 1 is a chromatogram showing the results of Example 1. FIG. 2 is a chromatogram showing the results of Example 2. FIG. 3 is a chromatogram showing the results of Example 3. FIG. 4 is a chromatogram showing the results of Comparative Example 1. FIG. 5 is a chromatogram showing the results of Comparative Example 2. FIG. 6 shows a schematic diagram of the analyzer used in Examples and Comparative Examples. Explanation of symbols 1... Sucrose peak 2... Maltose peak 3... Glucose peak 4... Maltitol peak 5... Fructose peak 6... Sorbitol peak 7... Joint peak proposal for fructose and sorbitol
3 mouths

Claims (1)

[Claims] 1. From a saccharide mixture containing sucrose, maltose, fructose and sorbitol, each is
_3^- group-containing ion exchange resin particles and the SO_3
Liquid chromatography using one or more columns packed with a column packing material having Ag^+, Ba^+^+ and/or Sr^+^+ as counter ions of the ^- group, and using water or an aqueous solution as the eluent. A method for separating saccharides, characterized by separating them by. 2. SO_ where the counter ion of SO_3^- group is Ag+^
S in which the counter ions of the 3^- group-containing ion exchange resin particles and the SO_3^- group are Ba^+^+ and/or Sr^+^+
2. The method for separating saccharides according to claim 1, wherein the O_3^- group-containing ion exchange resin particles are packed in the same column or in separate columns and connected. 3. SO_3^- group ion exchange resin particles are SO_3^-
3. The method for separating saccharides according to claim 1 or 2, wherein the group-containing porous styrene-divinylbenzene copolymer particles are used. 4. In the column packing material, the counter ion of the SO_3^- group, Ag^+, and Ba^+^+ and/or Sr^+^+ are in an equivalent ratio of the former/latter of 9/1 to 1/1. A method for separating saccharides according to any one of claims 1 to 3, wherein the saccharides are present in a certain proportion. 5.SO_ where the counter ion of SO_3^- group is Ag^+
The volume ratio of 3^- group-containing cation exchange resin particles and SO_3^- group-containing cation exchange resin particles in which the counter ion of the SO_3^- group is Ba^+^+ and/or Sr^+^+ is 9.
The method for separating saccharides according to any one of claims 1 to 3, using a column packed with a ratio of 1/1 to 1/1. 6. The method for separating saccharides according to any one of claims 1 to 4, wherein the alkali metal ion is Na^+.