Specification
ENZYMATIC PREPARATION OF GLUCOSE SYRUP FROM STARCH
FIELD OF THE INVENTION
The present invention relates to a process for the preparation of starch-hydrolysate syrups having characteristics which render them particularly attractive for a variety of industrial applications, notably in the food industry. The invention makes it possible using one enzyme, for the first time, to obtain syrups of the above-mentioned kind which closely match syrups whose preparation previously only feasible using acid hydrolysis (i.e., non-enzymatic hydrolysis) of starch.
BACKGROUND OF THE INVENTION
Glucose syrups with a DE (Dextrose Equivalent) around 42 is widely used in industry as an ingredient in products such as hard boiled candy, toffees, fudge, fondant and the like.
Traditionally 42 DE glucose syrups are produced by standard acid conversion. A starch slurry is initially acidified to pH 2, and is then pumped into a continuous reactor which operates at elevated temperature and pressure. After a period of time the liquor is returned to atmospheric conditions, neutralised, clarified, decolourised and concentrated to the final syrup. Such acid converted glucose syrup profile shown in Figure 1 reduce the tendency of sucrose to crystallise, they slow down the tendency to shell-graining and they contribute to the characteristic "mouth-feel".
Today also enzymatic conversion of starch into glucose syrup has been suggested. However, such glucose syrups typically have a sugar spectrum which is quite different frorr the traditionally used 42 DE acid converted glucose syrup.
SUMMARY OF THE INVENTION
The present invention is based on the finding that a glucose syrup with a DE in the range from 35 to 45 having a sugar spectrum close to that of the traditionally acid converted 42 DE glucose syrup can be obtained by treating starch with a 54W substituted variant of Termamyl® (which is a
commercially available Bacillus licheniforwis a-amylase).
In the first aspect the invention relates to a process for the preparation of a glucose syrup wherein starch is treated
with a Termamyl-like a-amylase comprising a substitution in Val54 shown in SEQ ID NO: 2 or in the corresponding position in
another Termamyl-like a-amylase.
The invention also relates to a glucose syrup obtainable by the process of the invention. Further, an aspect the invention also relates to the use of said glucose syrup obtainable by the process of the invention as ingredient in food products such as hard boiled candy, toffees, fudge, fondant and the like.
Another object of the invention is to provide for the use
of a Termamyl-like a-amylase with a substitution in position Val54 using SEQ ID NO: 2 as the backbone (i.e., parent enzyme) or a corresponding position in another Termamyl-like a-amylase for preparing glucose syrup.
BRIEF DESCRIPTION OF THE DRAWINGS
Figure 1 shown the sugar spectrum of a 42 DE acid converted glucose syrup.
Figure 2 shows the sugar spectrum of a Termamyl® (i.e.,
Bacillus licheniformis a-amylase from Novo Nordisk shown in SEQ ID NO: 2) converted glucose syrup.
Figure 3 shows the sugar spectrum of a V54W substituted Bacillus licheniformis α-amylase variant converted glucose syrup of the invention. Figure 4 is an alignment of the amino acid sequences of six
parent Termamyl-like a-amylases. The numbers on the Extreme left
designate the respective amino acid sequences as follows:
1: Bacillus sp. a-amylase,
2 : Kaoamyl a-amylase),
3: Bacillus sp. a-amylase,
4: B. amyloliquefaciens a-amylase (BAN) (SEQ ID NO: 3),
5: Bacillus licheniformis a-amylase (SEQ ID NO: 2),
6: a-amylase disclosed in Tsukamoto et al., Biochemical and Biophysical Research Communications, 151 (1988), pp. 25-31.
DETAILED DESCRIPTION OF THE INVENTION
The present invention is based on the finding that a novel glucose syrup with a DE in the range from 3 5 to 4 5 having a sugar spectrum and properties close to that of the traditionally acid converted glucose syrup, often referred to as "42 DE glucose syrup" is obtained by treating starch with a Val54Trp (V54W) substituted variants of the commercially
available Bacillus licheniformis a-amylase, sold under the trade name Termamyl® (Novo Nordisk) . The DNA and protein sequence of Termamyl® is displayed in SEQ ID NO: 1 and 2, respectively.
Substitution in the Val54 position of Termamyl-like a-amylases, including the B. licheniformis a-amylase, is known from WO 97/41213 (Novo Nordisk). However, it is surprising that a Val54 substituted variant can be used to preparing a syrup--from starch having a sugar spectrum which is close to that o-f • an acid converted 42 DE glucose syrup as a glucose syrup
prepared from starch treated with parent B. licheniformis a-amylase (SEQ ID NO: 2) has a sugar spectrum quite different therefrom.
In the first aspect the invention relates to a glucose syrup (or speciality syrup) prepared by treating starch with
the Bacillus licheniformis a-amylase shown in SEQ ID NO: 2 comprising a substitution in position Val54 or a Termamyl-like a-amylase (as defined below) substituted in a position corresponding to Val54 of SEQ ID NO: 2.
The glucose syrup of the invention has properties close to that of the traditional acid converted 42 DE syrups with regard to its sugar spectrum, i.e., composition of dextrose (DPI), maltose (DP2), maltotriose (DP3), maltotetraose (DP4), maltopentaose (DP5) and a number of higher sugars such as DP10 etc. The rheological properties, such as the viscosity, resembles the traditional acid converted DE 42 syrup much closer than a corresponding syrup prepared under the same
conditions by treatment with parent B. licheniformis a-amylase (i.e., SEQ ID NO: 2).
As can been seen clearly by comparing Figure 1 to 3 a glucose syrup prepared by treating starch with the Val54Trp substituted Termamyl variant (Figure 3) has a sugar spectrum closer to that of the acid converted 42 DE syrup (Figure 1) than that of the glucose syrup prepared using parent B."
licheniformis a-amylase (Figure 2) .
By using the Val54 substituted Bacillus licheniformis a-amylase variant for preparing a glucose syrup of the invention it can be seen that especially the DPI and DP4 sugar content has been increased to a level closer to that of the traditional 42 DE acid converted glucose syrup and the DP5 sugar content has been decreased to a level closer to that of the 42 DE glucose syrup in comparison to the corresponding glucose syrup
prepared using parent B, licheniformis a-amylase. Further, the content of the higher sugars, as can be seen by comparing the peak(s) on the left side of Figures 1 to 3, are also increased to a level closer to that of the acid converted 42 DE glucose syrup in comparison to corresponding parent B. licheniformis
a-amylase converted starch glucose syrup.
According to the invention only one enzyme need to be used for producing the glucose syrup of the invention, i.e. Val54
substituted Termamyl-like a-amylase.
The glucose syrup of the invention may be prepared by treating starch with a Val54 substituted Termamyl-like
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