JPS6040839B2 - Method for producing amorphous maltose syrup - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for producing high maltose syrup, and more specifically, the present invention relates to a method for producing high maltose syrup.
Using a culture in which the ratio of 8-amylase activity and Q-amylase activity produced by M. l.
1-84%, maltotriose content 8-15%, glucose content 0.4% or less, degree of polymerization 4 or more, 4-8%
The present invention relates to a method for producing amorphous maltose syrup having a sugar composition of

Methods for producing glucose, starch syrup, maltose, high-fructose sugar, etc. using starch as a raw material have been known for a long time, and have become a large sector in the starch saccharification industry.

Recently, products with particularly high maltose content, ie, high maltose syrup, have attracted attention.

The reasons for this include the following characteristics. '1' A recent trend is that people prefer foods with less sweetness. {21
It has a refreshing sweetness and is not bitter, and its quality of sweetness is superior to that of sugar and grape sugar.

[3] Chemical properties such as browning and decomposition during heating are more stable than other mister sugar and isomerized sugar.

■ Physical properties are similar to sucrose, so it can be used as a substitute for sucrose without changing the physical properties of foods.

Currently, when producing high maltose syrup on an industrial scale, starch treatment is carried out using 8-amylase alone or a combination of 6-amylase and Q-1,6 glucosidase.

In the case of 8-amylase alone, a product with a maltose content of 45 to 60% is obtained;
, 6-glucosidase has been used in combination to obtain products with a maltose content of 70% or more.

However, when trying to increase the maltose content to 80% or more, maltose crystallizes, which is inconvenient, and it is difficult to obtain an amorphous maltose syrup with a maltose content of 80% or more. However, although saccharified liquid with a maltose content of 80% or more has been obtained so far, this method is for separating crystalline maltose, and the maltose content is 80% or more.
% or more, in some cases 90% or more, but since the remaining sugar is mainly sugar with a degree of polymerization of 4 or more, maltose tends to crystallize, making it difficult to make maltose syrup for food additives.

The present inventors conducted research to produce maltose syrup containing 80% or more maltose, which is advantageous for food additives, and coincidentally discovered that the presence of Q-1,6 glucosidase in Bacillus polymyxa 8-amylase. Below D. Maltose content is 80 when applied to liquefied starch of E5 or less.
It was discovered that it was possible to produce high maltose syrup with a concentration of more than %.

However, this phenomenon cannot be explained theoretically, so further research revealed that when Bacillus polymyxa is cultured in an appropriate medium and produces 3-amylase, the bacterium produces Q-amylase in addition to 8-amylase. This amount of Q-amylase is moderately involved in the production of maltose and maltotriose, and sugars that cannot be broken down by 8-amylase alone are also converted to maltose or maltotriose.
It has become clear that maltose syrup can be obtained which is decomposed to sulfur and has a maltose content of 80% or more and is mainly composed of oligosaccharides with a degree of polymerization of 3 or less and is difficult to crystallize. The present invention was completed based on these findings, and D. In the presence of Q-1,6 glucosidase in liquefied starch below E5, the activity ratio of B-amylase and Q-amylase is 1:0.
．． 02 to 0.2, maltose content 81 to 84%, maltotriol content 8 to 15%.
, a method for producing an amorphous maltose syrup characterized by producing a syrup having a sugar syrup composition of 4 to 8% with a glucose content of 0.4% or less, a degree of polymerization of 4 or more, and a sugar content of 4 to 8%.

The starch that is the raw material of the present invention may be any starch such as potato, sweet potato, corn, tapioca, etc., and it can be processed by D. It is necessary to liquefy to E5 or lower.

In addition, the starch concentration at this time can be freely selected below 40%, but if the concentration is too high, the viscosity will be too strong, and if it is too low, the cost of concentrating the saccharified liquid will be increased. , preferably 20 to 35%. As the enzyme used, any of various commercially available Q-amylases may be used as Q-amylase used for liquefying starch. About 2000 u' of Q-amylase is added to starch milk and liquefied by heating at about 80-90 am.

After about 20 minutes, D. Liquefied starch of E5 or lower is obtained. This D. Liquefied starch below E5 contains Q-1,6 glucosidase and 8-amylase produced by Bacillus polymyxa and Q.
- A culture with an amylase activity ratio of 1:0.02 to 0.2, or an enzyme obtained therefrom, is added.

Q-1,6 glucosidase is a well-known Aerobacterium (A
erobacter), Pseudomonas (Pse
Enzymes produced by bacteria of the genus Momonas can be used.

The 8-amylase and Q-amylase used for saccharification can be produced by a strain of the same origin or a strain of different origin, as long as their activity ratio is 1:0.02 to 0.2. However, Bacillus polymyxa produces both at the same time by the same strain, and B-amylase versus Q
- Particularly preferred since it yields a suitable proportion of amylase. The reaction is carried out at 60 ml for about 20 to 4 hours.

The obtained saccharified liquid is purified by activated carbon treatment, ion exchange resin treatment, etc., and concentrated by vacuum concentration to obtain a thick syrup of 70% or more. The thick syrup obtained here will not crystallize even when stored for long periods at room temperature.

Next, test examples and examples of the present invention will be shown, and the enzyme activity measurement method used here will be shown.

I Q-1,6 glucosidase activity: M
Take 0.5 μl of the substrate solution dissolved in phosphate buffer (pH 6.0) and make 100 μl into a test tube, and after preheating at 40 μl for 5 minutes, add 0.5 μl of the appropriately diluted enzyme solution. 40
℃ for 18 minutes.

After each stroke, add DNS reagent 1' and mix to stop the reaction.

Thereafter, the mixture was heated in boiling water for 10 minutes to develop color, cooled in running water, 8 cores of purified water was added, and the mixture was uniformly mixed. The absorbance was then measured at a wavelength of 54 hm. Activity (above u') is expressed as (absorbance of test - absorbance of control) x dilution factor. 2 8
-Amylase activity: 0.5 ml of soluble starch was added to M/20 phosphate buffer (pH 7.
0) Add 100 μl of substrate solution dissolved in 100 μl to 100 μl: Place in a square flask, place in a constant overflow tank at 40°C, preheat for at least 5 minutes, add 1 μl of diluted enzyme solution, and heat to 40°C. React for 3 minutes.

After 30 minutes, Fehling's alkaline solution 2 is added to stop the reaction, and then the top of Fehling's steel solution 2 is added.

Boil for another 2 minutes, then cool, and after cooling 30% (w'w)
After adding 2 parts of potassium iodide and 2 parts of 25% (w/w) sulfuric acid, immediately titrate with 0.0% sodium thiosulfate. (A') Separately, the order of adding the enzyme solution and Fehling's alkaline solution was reversed and the same procedure was used as a control (B [), P-amylase activity (u/secretion) was 1.62
Expressed as x (B-A)/10 x enzyme dilution factor. 3 Q-
Amylase activity: 1 tablet of Amylase Test "Daiichi" (sold by Daiichi Chemical Co., Ltd.) was added to M/10 acetate buffer containing 2 hM calcium chloride (p
H6.0) 6' was added to make a substrate solution, and after preheating at 40℃ for 5 minutes, 0.1 I of diluted enzyme solution was added and reacted.
0 after 5 minutes. Add 1 part of sodium hydroxide to stop the reaction.

After passing through a furnace, the absorbance at 620' was measured, and the same operation was carried out using water instead of the enzyme solution as a control. Q
-Amylase activity (u/ship) is expressed as (absorbance of sample - absorbance of symmetrical) x dilution factor. Test example 1 dextrin 10%, ammonium sulfate 0.75%, defatted soybean powder 1
%, NaCIO. The medium having a composition of 3% ammonium phosphorus and 0.1% ammonium phosphorus (pH 6.5) was placed in an 800 fermenter and sterilized under high pressure at 120°C.

Pre-soluble starch 0.5%, ammonium phosphorus 0.1%, KCI
O. 0.02% MgSO, 200.02% MgSO, 0.2% yeast extract (pH 7.3) 20 pieces of medium were placed in a 30 capacity fermenter, 120 pieces were autoclaved for 030 minutes, and Bacillus, Polymixa NO. ．． 55, FERM-P
No. The entire amount of the seed culture solution inoculated with 3952 and aerated culture (20mpm, 0.5VVm) for 2 hours at 3030 was inoculated into the above fermenter, and cultured at 370 throughout the entire culture period.
During cultivation, the pH was maintained at 6.5 and 0.5 with NaOH.
1008 Terama Aerial Bicycle (30 ratio pm, 0.4 VVm) was cultured, and after the culture was completed, bacteria were sterilized using a Draval type centrifuge, and the sterilized solution was concentrated under reduced pressure to produce 8-amylase 1050 u/', Q-amylase A concentrated crude enzyme solution of 95 u/' was obtained. Furthermore, by subjecting this concentrated crude enzyme solution to salting with ammonium sulfate, DEAE/cellulose treatment, and CM-cellulose treatment, 8
-Amylase and Q-amylase were separated and purified.

(Figure 1) Test Example 2 A 25% potato starch slurry was liquefied using a commercially available liquefiable bacterial amylase to obtain a liquefied liquid with a DE of 2.3. 8-Amylase 4 U per evening of starch dry matter,
and Q-1,6 glucosidase (Pulteam K2000
: ABM Chemicals Limited) was added to 7 U of starch, and the purified Q-amylase of Bacillus polymixa obtained in Test Example 1 was added with various activity ratios to 8-amylase to adjust the pH to 6.0 to 5.0. When saccharification was carried out for 60 hours and 3 hours while maintaining the amount within the range, the results shown in Table 1 were obtained.

Table 1 To measure the sugar composition, the saccharified solution was spotted on furnace paper, developed three times with butanol:pyrimidine:water = 6:4:3, the corresponding portion of each sugar was cut out, and the water-extracted solution was extracted with phenol. It was determined by the sulfuric acid method.

However, only glucose was directly quantified in the saccharified solution using the glucose oxidase method.

No. of Table 1 1~No. The saccharified solution having the sugar composition obtained in step 10 was purified by ion exchange resin treatment and activated carbon treatment, and then vacuum concentrated to a solid content of 75% and left at room temperature for one month. 2~No. No crystals were deposited in the syrup obtained from Example 7. Furthermore, as shown in Table 1, by adding Q-amylase to B-amylase, high maltose syrup with a maltose content of 80% or more can be obtained, but even if there is too much Q-amylase, Good results were not obtained, therefore, the activity ratio of 8-amylase to Q-amylase was 1:0.02-0.
2 was found to be appropriate. Example 1 Potato starch 4.0k9, calcium chloride dihydrate 7 days, Diasmen (Q-amylase: product of Daiwa Kasei Co., Ltd.), 1
Mix 500 blood with water and prepare starch milk of 6.5 stem.

Separately, heat the water to 80 qo or more and pour in the starch milk while stirring.

Further heating is performed to react the enzyme for 80 to 900 CI minutes. Furthermore, live steam was blown into the reaction system and heated at 110°C for 30 minutes, and then cooled to 6000°C. Thus DE
．． 3.65 liquefied starch was obtained, and to this was added 5 u of bruzyme K2000 (Q-1,6 glucosidase: product of ABM Chemical Limited) and Test Example 1 per starch dry matter.
A concentrated crude enzyme solution containing 8-amylase and Q-amylase (5 u of 3-amylase activity per starch mixture) obtained by the method described above was added, and saccharification was carried out at 60°C for 24 hours while maintaining the pH between 6.0 and 5.0. The bran composition of this saccharified solution is 0.24% glucose and 82.78% maltose.
, maltotriose 12.64%, other sugars 4.34%
%Met. This material is used for decolorization using normal activated carbon and ion exchange.
The mixture was purified using a sterile resin and concentrated to a solid content concentration of 75% using a vacuum concentration method to obtain a syrup that does not easily crystallize at room temperature. Example 2 Corn starch 3.5k9 was used as a starch raw material, 10 tadiasmen (Q-amylase: product of Daiwa Kasei Co., Ltd.) 2000 skin was added, and DE2 was prepared according to Example 1.
．． 84 liquefied starch was obtained, and this was added with 5 u of starch dry matter per evening.
Pullteam K2000 (Q-1,6 glucosidase:
ABM Chemical Ltd. product) and 5 u of 8-amylase and Q-amylase-containing concentrated enzyme solution obtained by the method of Test Example 1 (ruled as 8-amylase activity per starch) were added, and 6.0 Saccharification was carried out for 4 feeding hours at 60 qo while maintaining the temperature at ~5.0.

The bran composition of this saccharified solution was 0.18% glucose, 81.30% maltose, 12.25% maltotriose, and 6.27% other sugars. This saccharified liquid was decolorized using conventional activated carbon, purified using an ion exchanger resin, and concentrated to a solid content concentration of 75% using a vacuum concentration method to obtain a syrup that does not easily crystallize at room temperature.

[Brief Description of the Drawings] Figure 1 shows the production of Bacillus polymyxa in Test Example 1. CM the crude enzyme solution containing 8-amylase and Q-amylase.
- It is a figure purified by cellulose treatment. A...o-amylase, B...3-amylase, C...
...Protein, D...Salt concentration.

Claims (1)

[Claims] 1. A saccharifying enzyme having an activity ratio of β-amylase and α-amylase of 1:0.02 to 0.2 is applied to liquefied starch having a DE of 5 or less in the presence of α-1,6 glucosidase to reduce the maltose content to 81 to 0.2. 84%, maltotriose content 8-15%, glucose content 0.4 or less, degree of polymerization 4
A method for producing amorphous maltose syrup characterized by producing syrup having a sugar composition having a sugar content of 4 to 8% as described above.