DK147236B - PARTICULATED CELL MASS PRODUCT OF GLUCOSE ISOMERASE, AND PROCEDURE FOR ITS PREPARATION - Google Patents

Description

147236

The invention relates to a dried, particulate cell mass product of glucose isomerase, and to a process for its preparation. According to the prior art, it was previously considered necessary to activate the glucose isomerase by adding cobalt ions to the glucose syrup (food syrup).

Since cobalt compounds are considered toxic at the same time, this measure necessitated the reduction of cobalt ion concentration in the isomerized syrup (product syrup) to a value in the range of 10 ppm (parts per million), e.g. by an ion exchange process.

2 14723.6

Another solution to this problem is the use of process conditions that redundant the activation of the enzyme by the addition of cobalt ions. As an example of the use of such process conditions, reference is made to the specification of Danish Patent Application No. 1097/76.

It has recently been recognized, cf. eg. description of Danish Patent Application No. 3858/75 that iron ions are also capable of activating glucose isomerase enzymes, which has led to proposals to add small amounts of an iron salt to the food syrup. In this connection, it should be noted that the glucose syrup itself often contains small amounts of dissolved iron.

However, the addition of a dissolved iron salt to the food syrup is associated with practical difficulties.

First, adding to the food syrup involves a dosing process. Next, the addition must be continuously checked by chemical analysis of the feed syrup before it is fed to the isomerization reactor. Finally, the saturation point of the enzyme product is likely to be reached during the life of a column filling, because the capacity of the enzyme product with regard to the binding of iron ions is either very low or at least limited. Once the saturation point is reached, the iron salt will begin to seep into the product stream. The presence of iron ions in the product syrup may give rise to color formation to such an extent that it will be necessary to remove them, e.g. by ion exchange, a process that will further increase reprocessing costs. All in all, adding the iron salt to the food syrup is disadvantageous.

According to the above-mentioned Danish patent application No. 3858/75, glucose isomerase is activated in insoluble form by pretreatment with an aqueous iron salt solution.

However, it is further recommended to supplement the food syrup with an iron salt on the grounds that during pre-treatment the iron bound is dissociated and removed with the product syrup. Therefore, the pretreatment with iron does not appear to be able to remedy the disadvantages mentioned above.

The object of the present invention is to provide an enzyme product in which the iron salt is incorporated into the enzyme product. This principle will be advantageous if the iron remains so firmly bound that there is virtually no leakage of iron ions from the enzyme product during its practical life.

Glucose isomerase is an intracellular enzyme that is not isolated from the microorganism for the preparation of an active enzyme product (see, e.g., U.S. Patent Nos. 3,821,086, 3,779,869 and 3,980,521). In this specification, the terms "cell mass product" and "particulate cell mass product" are used in connection with products and particles thereof which are formed or otherwise prepared from the microorganism's cell material by treatment with organic reagents, e.g. glutaraldehyde, proteins or agglomerates, e.g. poly-electrolytes. On a weight basis, the glucose isomerase content of the cell mass product generally represents a very small portion of the product as a whole.

Surprisingly, it has now been found that cell mass products of glucose isomerase are capable of binding considerable amounts of iron and furthermore that prolonged contact between the glucose isomerase product and food and / or product syrup results in very small losses of iron. The presence of an amount of a few ppm of an iron salt in the product syrup is permissible. The amount of iron that can be incorporated into the cell mass product far exceeds the amount needed to activate the glucose isomerization.

147236 4

In particular, it has been found possible to incorporate non-toxic, water-soluble iron salts in the solid state into the cell mass product in association with this being brought into particulate form, e.g. immediately prior to an extrusion process. Under appropriate circumstances, it is also possible to incorporate the iron salts in the form of concentrated aqueous solutions.

Accordingly, the dried particulate cell mass product of glucose isomerase according to the invention is peculiar in that, based on the dry matter content of the cell mass product, at least 0.05% by weight of iron in the form of a non-toxic water-soluble iron salt is incorporated herein.

The invention also relates to a process for the preparation of said dried particulate cell mass product of glucose isomerase, characterized in that at least 0.05 wt.% Of iron in the cell mass product of its dry matter content is incorporated in the form of a non-toxic, water-soluble iron salt and to a particulate product and dried.

According to a preferred embodiment of the process of the invention, the solid salt is incorporated in the cell mass product of glucose isomerase.

According to yet another preferred embodiment of a process according to the invention, a cell mass product of glucose isomerase is prepared which is made up of a concentrate of whole or partially disrupted cell material made from microorganisms of Bacillus coagulance, which concentrate has a dry matter content of from 3 to 30% by weight per volume unit, reacted with from 0.01 to 1.0 part by weight of glutaraldehyde per unit volume. part by weight of dry matter content of the concentrate to form a coherent solid product, after which water is removed and the glucose isomerase product is removed.

According to a further preferred embodiment of the process according to the invention, at least 0.5% by weight of magnesium oxide and at least 2% by weight of solid glucose are incorporated in the solid state, both calculated on the basis of the dry matter content of the cell mass product.

Thus, the present invention provides a particulate product which is readily suitable for soaking in glucose syrup prior to its use for isomerization.

As mentioned, at least 0.05% by weight, usually from 0.05% to 2.0% by weight, of iron in the form of a non-toxic, water-soluble iron salt is incorporated into the cell mass product. Incorporation of more than 2.0% by weight of iron is possible, but serves no practical purpose. The preferred iron content is in the range of 0.2 to 0.5% by weight, preferably about 0.2 to 0.25% by weight.

Only a little or none of the iron incorporated in the cell mass product in an amount of from 0.05 to 2.0% will pass into the product syrup during the practical life of the enzyme. In fact, the ferrous enzyme product is capable of further absorbing iron from an ferrous syrup. For example, a food syrup fed to the isomerization column with an iron content of 4 ppm may very well leave it as a product syrup with an iron content of less than 1 ppm.

In practice, it has been found that increased productivity and / or improved stability of the glucose isomerase product can be achieved by further admixture of other solids to the cell mass product. In particular, the initial pH drop, which occurs 1-2 days after a new column loading with fresh enzyme product is used, has caused some problems. A decrease in the column filling pH is undesirable because it causes shrinkage of the column filling, which in turn can cause channel formation. A further consequence of this may be a decrease in activity and, in more severe cases, in the stability of the enzyme product. Incorporation of from 0.5 to 3.0% by weight of magnesium oxide, based on the dry weight of the cell mass product, has been found to substantially reduce the initial pH drop and thus provide a relatively stable pH in the product syrup.

Furthermore, addition to the premix of glucose in solid form (e.g., glucose monohydrate) in amounts of from 2 to 15% by weight based on the dry mass of the cell mass product has been found to be desirable, the glucose acting mainly as a diluent in the dry mixing process.

According to a preferred embodiment, a premix of the water-soluble iron salt is prepared in solid state with magnesium oxide and glucose, after which it is mixed into the cell mass product prior to the extrusion process leading to the particulate product.

The following iron salts are preferred:

Ferrisulfate Ferrosulfate

Ferric chloride Ferrolactate

Ferric citrate Ferrocitrate

Ferriammonium citrate Ferroacetate

Ferrin nitrate Ferripyrophosphate

The invention will then be described in more detail with reference to the following examples.

In the examples, the following terminology is used:

Definition of activity

The unit of enzyme activity is defined as the amount of enzyme that catalyzes the conversion of glucose to fructose at an initial rate of 1 micromol of fructose per day. mine. under given reaction conditions.

Determination of activity

The activity is determined under the following conditions: Food syrup 40% by weight dissolved glucose pH of food syrup 8.5

Mg 0.004 M

Temperature 65 ° C

Column, diam. / Height height 2.5 / 35 cm Fluid flow downwards

The activity is expressed in IGIC units per g of enzyme product.

In long-range isomerization experiments, the curves for the activity decay are adjusted in exponential curves with the following formula: * »“ bt A = A0e where t is the time in hours after the start of the isomerization process, A and A are the activities for times t ° -1 and 0, respectively, and b is a constant with the dimension h

From this equation the half-life is defined in hours as 147236 8 Τι = ln 2% -

Productivity

Productivity is defined as the amount of glucose 1 kg per day. kg of enzyme product which in a given time is converted into a mixture containing 45% fructose.

In the examples, productivity is calculated according to the above equation after an isomerization time of 2 x Th.

Iron

The iron content is determined by the ortho-phenanthro-lin method (Nordic Methodology Committee for Foods no.

22, 1955, U.D.C. 664.7: 546.72).

Color

The color intensity is measured according to the CIRF method.

color stability

The color stability is determined after one hour of heating the syrup to 100 ° C at pH 4.2 according to the CIRF method.

The magnesium oxide used in the examples is a heavy quality ER / B type from Pharmelko, Milan,

Italy.

The process according to the invention is illustrated in more detail in the following examples. Those with labeled test conditions are comparative trials.

Example 1 9 147236

Addition of ferric citrate, ferrolactate and ferric sulfate in connection with magnesium oxide and glucose. Addition of ferric oxide.

The starting material is a cell mass product of glucose isomerase made from a concentrate of whole or partially disrupted cell material of the microorganism Bacillus coagulans. The concentrate having a dry matter content of from 3 to 30% by weight per unit weight. by volume, from 0.01 to 1.0 parts by weight of glutaraldehyde per unit volume. part by weight of dry matter content of the concentrate. Thereby, a coherent solid product is formed from which the water is removed by filtration on a filter press (cf. Example 5 of U.S. Patent No. 3,980,521).

The filter cake is granulated by means of an oscillating granulation apparatus with a screen of 1 cm holes.

The coarse granule contains approx. 76% water (determined by drying at 105 ° C). It is divided into 6 portions.

+ ^ A. 8.5 kg of the coarse granulated filter cake is extruded by means of an axial extrusion apparatus which is provided with a filter with holes having a diameter of 0.8 mm. The extrudate is dried in a fluidized state with air at a temperature of 60 - 65 ° C to a water content of approx.

10%.

B. To 8.5 kg of the coarse granulated filter cake is added a mixture of 20 g of magnesium oxide, 85 g of glucose monohydrate and 40 g of ferric citrate with an iron content of 16%.

The mixture is extruded after thorough mixing and dried in an analogous manner as described under point A.

· / 10 ounce C. 8.5 kg of the coarse granulate is mixed with a mixture of 20 g of magnesium oxide, 85 g of glucose monohydrate and 40 g of ferrolactate having an iron content of about 19%. The mixture is extruded after thorough mixing and dried in an analogous manner as described under point A.

D. 8.5 kg of the coarse granulate is mixed with a mixture of 20 g of magnesium oxide, 85 g of glucose monohydrate and 30 g of ferric sulphate having an iron content of approx. 20%. The mixture is extruded after thorough mixing and dried in an analogous manner as described under point A.

+ ^ E. 8.5 kg of the coarse granulate is thoroughly mixed with a mixture of 20 g of magnesium oxide and 85 g of glucose monohydrate. The mixture is extruded and dried in an analogous manner as described under point A.

+ ^ F. 8.5 kg of coarse granulate is thoroughly mixed with 25 g of ferric oxide containing approx. 58% iron. The mixture is extruded and dried in an analogous manner as described under point A.

The products are screened to obtain a product with a grain size of between 0.35 mm and 1.0 mm and the products analyzed.

In the running syrup flow, the pH is measured in samples taken after 20 and 43 hours. Before the pH determination, the samples are cooled to 25 ° C.

Table I

11 147236 | Product J_Activity_ | _ | pH of external syrup | | | Found | Corrected | % [ after_[

I I IGIC / q I IGIC / ct I increase I 20 hours I 43 hours I

III III I

| + k I 246 | 246 I 0 I 6.68 | 7.62 | | B | 307 | 326 | 33 | 7.99 | 8.20 | | C | 296 | 315 | 28 | 7.60 | 7.98 | | D | 308 | 328 | 33 | 7.90 | 8.14 | | +) E | 254 | 267 | 8 | 7.99 | 8.20 | | +) F | 257 | 260 | 6 | 6.86 | 7.65 |

I I _! _! _! _! _ L

As shown in Table I, an activity increase of significance is obtained only by the addition of soluble iron compounds. By adding ferric oxide, only an increase of approx. 6% compared to approx. 30% for the soluble salts.

Example 2

Addition of magnesium oxide + glucose and magnesium oxide + glucose + iron salt.

A filter cake is prepared in an analogous manner as described in Example 1. The filter cake is granulated by means of an oscillating granulating apparatus, which is provided with a screen of 1 cm holes.

The coarse granule contains approx. 79% water. It is divided into 5 portions of 8.5 kg.

147236 12 + ^ A. 8.5 kg is extruded and dried in an analogous manner as described in Example 1A without the addition of additives.

+ ^ B. To 8.5 kg granulated filter cake is added 25 g of magnesium oxide. Extruded after thorough mixing and dried in an analogous manner as described under point A.

+) C. To 8.5 kg of granulated filter cake is added 25 g of magnesium oxide and 200 g of glucose monohydrate. Extruded after mixing and dried in an analogous manner as described under point A.

+ D ^. To 8.5 kg granulated filter cake is added a mixture of 25 g magnesium oxide and 300 g glucose. It is extruded after mixing and dried.

E. 8.5 kg filter cake is mixed with a mixture of 25 g of magnesium oxide, 200 g of glucose monohydrate and 40 g of ferric sulfate containing approx. 20% iron. It is then extruded and dried.

The dried products are screened to obtain a product having a grain size of between 0.35 mm and 1.0 mm and the products analyzed.

In the running syrup, the pH is measured in samples taken after 20 and 43 hours and cooled to 25 ° C.

Table II

13 U7236

I i i i i i i

| Product | % added J_Activity_ [% | pH value in outlet | | | material | Found | Corrected for | for- | bake syrup after [| | | IGIC / g | added inactive | increase | 20 hours | 43 hours |

II 11 material IGIC / q 111 I

J II II I i I A lo | 220 | 220 | 0 I 6.85 | 7.40 | | +) B I 1 I 222 I 224 | 2 | 8.18 | 8.23 | | +) C | 10 | 216 | 240 | 9 | 8.14 | 8.22 | I + b I 14 | 216 [251 | 14 | 8.15 | 1.21 | | E | 12 | 272 | 309 | 40 | 8.15 | 8.27 |

J _! _ 1 'I_I I I_L

As can be seen from Table II, a significant increase in activity is obtained only by the addition of an iron salt.

Example 3

Addition of ferric citrate, ferric pyrophosphate, ferric aromoneium citrate and ferrous sulfate.

A coarse granulated filter cake with approx. 76% water as in Example 1 is divided into 6 portions of 8.5 kg each.

+ ^ A. 8.5 kg of granulated filter cake is extruded and dried in an analogous manner as described in Example 1 to give a comparison product.

To 8.5 kg of the coarse granulate is added a mixture of 25 g magnesium oxide, 25 g ferric citrate with approx.

16% iron and 250 g glucose monohydrate. The granulate is extruded after thorough mixing and dried in an analogous manner as described under point A.

C. 8.5 kg of the coarse granulate is extruded and dried in an analogous manner as described under point A after the addition of 25 g of magnesium oxide, 50 g of ferric citrate and 250 g of glucose monohydrate.

D. 8.5 kg of the coarse granulate is treated in an analogous manner as described under point C, except that the 50 g of ferric citrate is replaced by 30 g of ferric pyrophosphate with an iron content of about 12%.

E. To 8.5 kg of the coarse granulate are added 25 g of magnesium oxide, 250 g of glucose monohydrate and 30 g of ferric ammonium citrate, with an iron content of approx. 15%. The granulate is extruded after thorough mixing and dried in an analogous manner as described under point A.

F. To the last portion of 8.5 kg is added 25 g of magnesium oxide, 250 g of glucose monohydrate and 30 g of ferrous sulphate with an iron content of approx. 30%. The granulate is extruded after thorough mixing and dried in an analogous manner as described under point A.

The dried products are screened to obtain a product having a grain size of between 0.35 mm and 1.0 mm and the products obtained are analyzed. The pH of the expiring syrup is determined after 20 and 43 hours.

Table III

1C 147236 15

II il I

| Product J_Activity_ [% In pH of expired syrup | | | Found (Corrected | Increase | After_ [

j_ I IGIC / σ I IGIC / q 1 1 20 hours I 43 hours I

I I I I I I I I

| A | 229 | 229 I 0 | 6.64 | 7.25 | | B | 261 | 293 | 28 | 7.90 | 8.24 | | C | 273 | 306 | 34 | 7.84 | 8.22 | | D | 266 | 299 | 31 | 7.79 | 8.19 | | E | 268 | 301 | 31 | 7.70 | 8.14 | | . F | 263 | 295 | 29 | 7.87 | 8.03 |

I I _! _ L_I _! _ I

No significant difference is observed with regard to the activating effect of the iron salts used.

Example 4

Isomeriseringsforsøg.

A coarse granulated filter cake, prepared in an analogous manner as described in Example 1, is used for the following products. The filter cake contains approx. 77% water.

147236 16 + ^ 410 / A: No addition.

+ 410 / B: Approx. 10 parts by weight of mixture 1 is added to approx. 90 parts by weight, based on dry matter, of the filter cake.

Mixture 1 consists of 100 parts glucose and 8 parts magnesium oxide.

410 / C: Approx. 2 parts by weight of mixture 2 is added to approx. 98 parts by weight of the filter cake, based on dry matter.

Mixture 2 consists of 100 parts glucose, 10 parts magnesium oxide and 12 parts ferric sulfate.

410 / D: Approx. 7 parts by weight of mixture 2 is added to approx. 93 parts by weight of the filter cake, calculated on dry matter.

+ V410 / E: No addition.

Mixtures 410 / A - 410 / E are then extruded through a 0.8 mm hole screen and then dried in a fluidized state to a water content of approx. 10%.

The iron content of the five final products is determined: 410 / A: 0.04% 410 / B: 0.03% 410 / C: 0.08% 410 / D: 0.18% 410 / E: 0.04%

Isomerizations are performed with material from products 410 / A, 410 / B, 410 / D and 410 / E under the following designations: 17 147236

Syrup: 45% by weight of redissolved glucose

Inlet pH: 8.4-0.1

Mg ++: 0.0016 M

Temperature: 62 ° C

Column size: height: 40 cm diameter: 5.8 cm volume: 1 liter Enzyme weight: 260 g.

The enzyme is soaked for 2 hours at room temperature in the above syrup, however, using a pH of 8.0 and then packed in the column.

The following results are obtained:

Table IV (a) iiiiiii "| Product | Maximum Measured | Overall | Expiry pH | Halving | Producti [| | Activity | Experimental 1 after hours_ [time T ^ | vite ef- | | | | time , || || hours | ter 2 | | | Itimer | 21 | 48 | 92 | | hours |

I I I 1 I I I I I

| 410 / A | 158 I 1293 | 6.9 | 6.8 | 7.2 | 842 | 1880 | | 410 / B | 155 | 936 | 7.4 | 7.7 | 8.0 | 818 | 1790 | | 410 / D | 202 | 1316 | 7.3 | 7.5 | 7.7 | 843 | 2295 | | 410 / E | 151 | 1147 | 6.9 | 6.9 | 7.7 | 828 | 1755 |

J _! _ I 1111_I_L

The concentration of iron in the syrup flowing from these columns is determined.

Table IV (b) 18 147236 I I Ϊ | Product J_Fe (ppm) in export syrup |

II I I I

| For 2.5 hours after | 21 hours after | 27 hours after | I I start __L_j £ grt start _ [_

II I I I

| 410 / A | <i | * 1 | <i I

I 410 / B I <1 | * 1 | <1 I

| 410 / D For approx. <1 | <1 | <1 | | 410 / E | <1 | <1 | * 1 |

J_I _! _! _ In

Further isomerization experiments are performed with material from products 410 / C, 410 / D and 410 / E using the following conditions:

Syrup: 45% by weight of redissolved glucose

Inlet pH: 8.4-0.1

Mg ++: 0.0016 M

Temperature: 6 5 ° C

Column size: height: 20 cm diameter: 2.5 cm volume: 100 ml Enzyme weight: 20 g.

The enzyme is soaked for 1 hour at room temperature in the above syrup and then packed in the column.

19 147236

The following results are obtained:

Table IV (c) T i i i i i i \ | Product | Max. measured | Total | Expiry pH | Halving | Producti | | | activity [experimental [after hours_ [time | wit ef- [I I | time, | 1 [| hours | ter 2xT ^ | | | | hours | 17 | 45 | 200 | | hours | I I 1 ..... 1 111 I 1

II I I I I I I I I

| 410 / C I 210 I 900 | 7.0 | 7.8 | 8.1 | 512 | 1510 | | 410 / D I 250 I 900 | 7.5 | 8.0 J8.2 | 484 | 1725 | | 410 / E | 190 | 900 | 6.9 | 7.4 {8.2 | 485 | 1340 |

I I I I I I I _! _ L

The concentration of iron in the syrup flowing from these columns is determined.

Table IV (d) "i j 'i" | Product J_Fe (ppm) in expiring syrup_ [| | 0 hours | 24 hours | 72 hours | 140 hours | 850 hours |

I I (bleeding) I after start | after start After__start [after start I

| 410 / C | 0.8 I <0.5 | <0.5 | <0.5 | <0.5 | | 410 / D | 3.6 | <0.5 | <0.5 | <0.5 | <0.5 | | 410 / E | <0.5 | * 0.5 | <0.5 | <0.5 | <0.5 |

I I _! _! _! _ J_L

20 urns The CIRF color of the syrup flowing from these columns b e s teinmes.

Table IV (e) i i i "| Product J_CIKF color in syrup_ [

II I I I

| For 0 hours | 24 hours after | 72 hours after |

I I (bleeding) I start I start I

II I I I

| 410 / C I 0.266 I 0.030 | 0.019 | | 410 / D | 0.247 | 0.036 | 0.020 | | 410 / E | 0.232 | 0.036 | 0.022 |

I 'I _! _! _ L

In comparison, the CIRF color of three samples of the food syrup used during this period was determined at 0.019, 0.012 and 0.014.

The color stability of the syrup flowing from these columns is determined.

Table IV (f) 21 1A7236 i i Γ I Product J_Colour stability of syrup__J_

II II I

| | 0 hours | 24 hours after | 72 hours after |

I I (bleeding) l start l start I

II II I

I 410 / C I 0.21 I 0.040 | 0.014 | I 410 / D | 0.21 in 0.050 | 0.017 | I 410 / E | 0.22 | 0.044 | 0.017 |

In comparison, the color stability of three samples of the food syrup used during this period was determined. The results were 0.004, 0.002 and 0.004.

The iron content of the enzyme products is determined before and after use.

Table IV (g)

I I I

| Product J_mg iron in with 20 g of enzyme packed column_ [

II I I

J [_1_By ^ jtart_L_ After 900 hours I

II I I

| 410 / C I 16 I 24 | | 410 / D | 36 | 42 | | 410 / E | 8 | 14 |

I 1 _! _ I

147236 22

It should be noted that the iron content after 900 hours is greater than at the beginning of the experiment. The enzyme therefore adsorbs iron from the food syrup. Since no iron was added to the food syrup used in these experiments, the iron adsorbed by the enzyme is derived from traces of iron naturally present in the solutions of crystalline glucose. Analysis of the iron content of the 45% by weight of redissolved glucose syrup yields less than 0.5 ppm of iron, approx. 0.1 ppm iron. During the 900 hours the trial of these columns was conducted, approx. 75 kg of syrup through each column containing 20 g of enzyme.

If the average iron concentration in the food syrup is 0.1 ppm, the total iron content in the food syrup becomes approx. 7.5 mg.

This corresponds well to the amount absorbed by the enzyme products during the test.

conclusion

The addition of magnesium oxide has a significant influence on the outlet pH in the period between 0 and 100 hours after the start of the experiment. With magnesium oxide (experiments 410 / B and 410 / D), the outlet pH is 0.5 - 1.0 units higher than without magnesium oxide (experiments 410 / A and 410 / E),

By the addition of iron salt (experiment 410 / D), the activity is increased without deterioration in stability, and a total increase in productivity of between 20 and 30% is obtained.

Adding smaller amounts of magnesium dioxide and iron salt (Experiment 410 / C) results in a smaller increase in the outlet pH and a smaller increase in productivity, but these increases are significant.

Example 5 23 147236

Comparison of ferrous and ferric salts.

A mixture of iron salt, glucose and magnesium oxide is added to samples of coarse granulated filter cakes prepared in an analogous manner as described in Example 1.

The mixture is then further treated by extrusion through a 0.8 mm hole screen and then by drying as a fluidized mass to a water content of approx. 10%. The product and the amount of the premix made up of iron salt, glucose and magnesium oxide are such that end products with the following compositions are obtained:

Table V (a) i i i i i i * | Product | Iron salt | Glucose | Magnesium oxide |

1. 1 1 I I I

III I I

| IG 403 II C I1.2% ferric sulphate | 8% | 1% | | IG 403 II D | 1.2% Ferrosulfate | 8% | 1% |

| IG 403 II E | lntet | 8% | 1% I

J_I_ | _I_L

Analysis of the products gives the following results for the actual iron content: IG 403 II C: 0.22% IG 403 II D: 0.27% IG 403 II E: 0.05%.

The isomerizations are carried out under the following conditions:

Syrup: 45% by weight of redissolved glucose

Inlet pH: 8.4-0.1

MG ++: 0.0016 M

Temperature: 65 ° C

Column size: height: 20 cm diameter: 2.5 cm volume: 100 ml Enzyme weight: 20 g.

The enzyme is soaked for 1 hour at room temperature in the above syrup and then packed in the column.

The following results are obtained:

Table V (b) i i i i i i Γ | Product | Max. measured | Total experimental [Half-life [Productivity | | | activity | time, hours [T ^, hours [after 2xT ^ hours |

111 1 1 I

in I I I I I I

[IG 403 II C i 290 | 755 | 482 1 2093 [| IG 403 II D | 274 | 755 | 467 | 1914 | | IG 403 II E | 254 | 755 | 431 | 1635 |

J _ [_! _ [_! _ L

The iron content of the enzyme products is determined before and after use.

Table V (c) and 147236

ZD

I I I

| Product J_mg iron in with 20 g of enzyme packed column_I

II I I

j_I_Wedjstart __J_Eft ^ _755_year ____ l_

II I I

| IG 403 II C | 44 | 52 | | IG 403 II D | 54 | 68 | | IG 403 II E | 10 | 16 |

J _! _! _ L

The iron content increases slightly during the experiment, indicating that the products adsorb iron from traces of iron present in the redissolved glucose.

conclusion

By adding either ferrous or ferric sulfate, the activity and productivity of the enzyme preparation is increased.

Example 6 Detection of Iron Saturation.

A coarse granulated filter cake prepared in an analogous manner as described in Example 1 is used for the following products: 26 147236 415 / A: No addition.

415 / B: Approx. 10 parts by weight of mixture 2 is added to approx. 90 parts by weight of the filter cake, based on dry matter.

The filter cake contained approx. 77% water. Mixture 2 consists of 100 parts glucose, 10 parts magnesium oxide and 12 parts ferric sulfate.

Mixtures 415 / A and 415 / B are then extruded through a 0.8 mm hole filter and then dried in a fluidized state to a water content of approx. 10%.

The iron content of the two final products is determined: 415 / A: 0.03% 415 / B: 0.26%.

Isomerizations are performed with products 415 / A and 415 / B using the following conditions:

Syrup: 45% by weight of redissolved glucose

Inlet pH: 8.3-0.1

Mg ++: 0.0016 M

Fe content: 0.00007 M (4 ppm)

Temperature: 65 ° C

Column size: height: 20 cm diameter: 2.5 cm volume: 100 ml Z z weight: 20 0 g.

The enzyme is soaked in the syrup for 1 hour at room temperature and then packed in the column. The following results are obtained:

Table VI (a) 27 147236 in Γ "iiiii | Product | Maximum measured | Time to up | | Total for | | Half | | Productivity | | | activity | attainment of | search time, | call time | after | | | | max active- | hours \ Τψ hours 12x1 ^, hours | | | | tet, hours | | | |

J I I - I I I I

III I II I

| 415 / A I 272 I 160 | 906 | 611 | 2560 | | 415 / B | 275 | 20 | 906 | 547 | 2260 |

J_I_I_ | _I _! _ L

The concentration of iron in the syrup flowing from these columns is determined.

Table VI (b) _ j | | Product | Fe (ppm) in expiring syrup at |

I J_L

III I II I

| ! 0 hours | 20 hours | 70 hours 1350 hours | 900 hours |

I 1 (bleeding) I I II I

| 415 / A | <0.5 | <0.5 | <0.5 | <0.5 | 0.5 | | 415 / B | 7 | <0.5 | <0.5 | <0.5 | 0.6 |

J_I_I _! _ I I_L

The iron content of the enzyme preparations is determined before and after use.

Table VI (c) 28 147236

I l I

| Product J_mg iron in with 20 g of enzyme packed column_ |

II I I

I at start I after 900 hours I

II I I

I 415 / A I 6 | 320 | I 415 / B | 52 | 380 |

Than L1 _! _ L

conclusion

Test 415 / A gives 13% higher productivity than experiment 415 / B. However, it should be noted that Experiment 415 / B contains approx. 10% by weight of non-enzyme material. Thus, calculated on the basis of filter cake containing the original enzyme, both products give approximately the same productivity.

The activity of Experiment 415 / A is increased during the first 160 hours of the experiment. This is in contrast to Experiment 415 / B, which provides maximum activity after 20 hours. This indicates that during experiment 415 / A, iron from the food syrup is slowly adsorbed with a consequent slow activation. This slow activation is also the cause of the longer exponential half-life found for Experiment 415 / A, i.e. activation and exponential decomposition occur simultaneously.

During the 900 hours the trial lasts, approx. 90 kg of syrup through each column containing 20 g of enzyme product. The iron content of this syrup is 4 ppm.

Therefore, 90 kg of syrup contains 360 mg of iron. The iron content of the two columns is increased by 314 and 328 mg. Thus, most of the iron in the food syrup is removed by the enzyme products.

The results show that after 900 hours the iron level in the expiring syrup begins to increase.

This could indicate that the enzyme products are approaching the limit of their ability to absorb iron.

Claims (4)

147236 Dried, particulate cell mass product of glucose isomerase, characterized in that, based on the dry matter content of the cell mass product, at least 0.05% by weight of iron in the form of a non-toxic water-soluble iron salt is incorporated herein. Process for the preparation of the cell mass product according to claim 1, characterized in that at least 0.05% by weight of iron in the form of a glucose isomerase-containing cell mass product is incorporated in the form of a non-toxic, water-soluble iron salt and that the cell mass product is converted into a cell mass product. dried. Process according to claim 2, characterized in that the iron salt is incorporated in a solid state in the cell mass product of glucose isomerase. Process according to claims 2-3, characterized in that a cellular mass product of glucose isomerase is produced which is made up of a concentrate of whole or partially disrupted cell material made from microorganisms of Bacillus coagulans, which concentrate has a dry matter content of 3 to 30% by weight per by volume, from 0.01 to 1.0 parts by weight of glutaraldehyde per unit volume. part by weight of dry matter content of the concentrate to form a coherent solid product, after which water is removed and the glucose isomerase product is removed.