Active Properties

In the presentation of sake the koji is added to the steamed rice and water, and the solution, mixed with the insoluble residue of starch and cellulose, then acts upon the steamed rice. To study this action more readily it is more convenient to make use of a filtered aqueous extract of koji, for it has been ascertained that the active property of the koji, the "diastase", is dissolved out by contact with water. And first as to the nature of the solution. A sample of koji when powdered or rubbed down in a porcelain mortar and then digested with water for a short time gives, after filtration, a yellow liquid which contains dextrin, dextrose, albumenoid matter, and a small quantity of mineral matter. The proportions which the three first of these constituents bear to one another depend upon two things: 1.) The quantity of water used in proportion to the koji. 2) The duration of the digestion, whilst 3) the temperature at which the digestion is effected affects the amount of the total matter dissolved and the rapidity with which it enters into solution. Table 9 gives the results of experiments made at the ordinary temperature of the air, showing the truth of the first two of these statements.

In column II the volume of water used to dissolve the soluble matter of 100 grams of koji is given; in III, the time during which the water and the koji remained in contact; in IV, the number of grams of solid matter dissolved from 100 grams of koji by the amount of

Table 9: Amount of Solid Matter Dissolved by Water from 100 grams of Koji at 10-15° C.

I

II

III

IV

V

VI

VII

VIII

Number

Volume of water used

Time

Weight of solid matter in solution

Avg. weight of solids

Percent dextrose in solid matter

Specific rotatory power

Avg. specific rotatory power

1

500 cc

12 hours

17.7

60.0

65°

2

1000

18

25.7

61

3

"

24.2

55.7

57.6

4

"

23.0

56.0

5

12

33.3

49.0

65.3

6

"

33.3

50.9

65.4

7

"

29.4

27.0

45.0

62.9

8

"

28.6

46.5

67.7

64.6

9

"

26.8

53.0

61.4

10

"

22.5

53.0

64.5

11

"

22.2

54.0

65.0

12

4

28.0

61.4

13

2000

3

31.1

68.0

78.0

14

2500

"

32.2

58.0

68.1

15

"

"

32.5

31.4

70.0

65.2

69.3

16

"

"

30.7

65.0

73.8

17

"

"

30.1

68.0

70.2

18

5000

24

30.0

47.0

64.5

19

10000

"

40.0

66.0

60.5

water mentioned; column V gives the average percentage of solid matter in the experiments indicated; column VI gives the percentage of dextrose contained in the solid matter; column VII, the specific rotatory power of the solution, and VIII, the average specific rotatory power of the solutions indicated. In experiments 2 to 12 the amount of water used for 100 grams of koji was 1000 c.c. and these experiments include three differing periods of digestion, but there is no evidence that the time of digestion has much influence upon the quantity of matter dissolved, at least at the temperature 10-15° C. The average percentage of solid matter dissolved is 27.0. Experiments 14 to 17 show how much solid matter is dissolved when the amount of water used is 2500 c.c. to 100 grams of koji; the average percentage being 31.4. We see, therefore, that when a larger quantity of water is used the amount of solid matter obtained in solution is greater. It is not possible to draw any defi nite conclusions from single experiments, but the very large percentage dissolved when 100 grams of koji were digested with 10,000 c.c. of water, bears out the above observations.

We have next to consider the influence of time upon the nature of the soluble matter. We have seen that it does not after 3 or 4 hours at the ordinary temperature affect very much the total amount of solids dissolved. But column VIII, which gives the average specific rotatory power of three series of experiments lasting respectively 18, 12, and 3 hours, shows that at 18 hours the specific rotatory power is smaller than at 12 hours, and at 12 hours less than at 3 hours. What is the meaning of this variation? The specific rotatory power of the solution is made up of three factors. The specific rotatory power of dextrin is 216°, that of dextrose is 59°. If these were the only two substances present the specific rotatory power of the solution would lie between these two numbers having a value proportionate to the amount of each present. It will be seen however, that the average of the experiments at 18 hours is less than 59°, and this shows that something else is present which tends to lower the value of the specific rotatory power. The albumenoids which are held in solution have been ascertained by nitrogen determinations to have an average value of -40°, and it is owing to their presence that the specific rotatory power is so low as it is. The composition of the liquid in experiment 6, for example, will illustrate this more clearly. 100 c.c. of the solution contained 1.695 gram dextrose, 0.723 gram dextrin, and 0.914 of albumenoids (calculated by multiplying the nitrogen found by 6.3). This gives a composition in 100 parts shown in the table below.

Dextrose 50.9%

Dextrin 21.7%

Albumenoids 27.4% 100%

The observed specific rotatory power was 65.4°. The calculated specific power was obtained in the following way:

(0.509 x 59) + (0.217 x 216) + (0.274 x -40) = 30.031 + 46.872 - 10.96 = 65.94

The calculated number thus agrees very well with the observed number and we may, therefore, assume the specific rotatory power of the albumenoids to be expressed by the number -40°.

Taking the series of experiments which lasted for 3 hours we find that the average specific rotatory power is 69.3°, about 10° higher than that of pure dextrose; the average specific rotatory power of those at 12 hours is 64.6°, about 5° higher than that of dextrose, and that of the experiments at 18 hours 57.6°, about 1.4° lower than that of pure dextrose. This diminution occurs because the amount of albumenoids in solution is greater when the spe cific rotatory power is less, their left handed rotation partially neutralizing the right handed rotation of the dextrin and dextrose. But why is it that the amount of albumenoids is greater when the treatment with water is longer continued? The most probable explanation is that as the albumenoids exist in the koji, they are not entirely soluble; a portion is already soluble in water, but the rest is only brought into solution by the action of the water itself, and perhaps also, through the agency of the albumenoids at first dissolved. It is in fact a chemical reaction which takes time for its completion, and probably, if sufficient time were allowed, the whole of the nitrogenous matter of the rice would be degraded and brought into solution. This is a point of importance to brewers of sake, for we shall see that the power which the koji possesses of transforming rice into dextrose, capable of undergoing alcoholic fermentation, is due to the presence of these albumenoids in solution.

The effect of heating a mixture of koji and water is to bring the matter into solution much more rapidly than at a low temperature.

Table 10: Action of Water at Higher Temperatures Upon 100 grams of Koji

Exp.

Time and Temperature

c.c. of water / 100 gr. koji

Solid matter dissolved

Dextrose % or solids

Specific rot. power

1

2 hours at 50° + 18 hours at 15° C.

1700

51.80

68.0

68°

2

1/2 hour at 45° C.

2000

31.80

84.9

76.1°

3

2 hours at 45° C.

2000

61.6

68.5

58.5°

4

1/2 hour at 50° C.

5000

37.2

66.0

63.2°

5

24 hours at 15° + 2 hours at 100°

10000

49.2

58.0

73.8°

With the exception of Experiment 2, the percentage of matter dissolved by the water is greater than in the experiments conducted at a lower temperature, and as a rule the percentage of dextrose in the solid matter is also greater. We shall, however, learn something by comparing experiments 2 and 3 with an experiment made at the ordinary temperature with the same sample of koji. In every respect the conditions of the three experiments were the same except as regards time and temperature.

Table 11: Action of Water on Koji

Exp.

Time and Temperature

c.c. of water / 100 gr. koji

Solid matter dissolved

Dextrose % or solids

Specific rot. power

1

18 hours at 10-12° C.

2000

29.2

69.3

66.3°

2

1/2 hour at 45°

2000

31.8

84.9

76.1°

3

2 hours at 45°

2000

61.6

68.5

53.5°

The above comparison shows that the amount of solid matter dissolved when the contact between koji and water is for 18 hours at a low temperature and for 1/2 hour at a high temperature is very nearly the same, but the percentage of dextrose and the specific rotatory power of the solution indicate that the proportions in which the three ingredients are present are very different. If we assume the specific rotatory power of the albumenoids to be -40° we may ascertain the composition of the solid matter, and referring it to a fixed amount of dextrose we get per 100 parts of dextrose:

Table 12: Composition of Solid Matter per 100 Parts Dextrose

Exp.

Time and Temperature

Dextrin

Albumenoids

1

8 hours at 10-12° C.

21.2

23.10

2

1/2 hour at 45°

14.7

3.06

3

2 hours at 45°

14.6

25.30

After 18 hours at a low temperature the amount of dextrin is 21.2 parts for every 100 parts of dextrose, but after both 1/2 hour and 2 hours at 45°, it remains practically the same and about two-thirds of the amount in the former case. The most interesting fact to be observed is the variation in the amount of the albumenoids; after 18 hours at 10-12° C it is very little different from the amount dissolved out in 2 hours at a temperature of 45° C, but after only 1/2 hour at 45° C the quantity in solution is only about one-eighth as much as in the two other experiments. This bears out the observations made at lower temperatures viz. that the amount of albumenoid matter dissolved is mainly affected by the duration of the experiment. It is not only dependent upon that, for we see that influence of a higher temperature in dissolving the albumenoids more rapidly, 2 hours at 45° C being more than equivalent to 18 hours at 10-12°C. Thus we are again led to the conclusion that the greater part of the nitrogenous matter in koji is insoluble in water, but that it is in such a state that the prolonged contact with water renders it soluble.

Although the effect of heat upon the mixture of koji and water is thus marked, when the clear solution has been separated by filtration from the undissolved grains it is not so rapidly changed either by exposure to heat or by longer standing at the ordinary temperature of the air. It is important for us to examine the change in composition of the solution on heating, as in the experiments upon starch-paste to be presently described it is the filtered solution of koji which is used. Table 13 gives the results of a number of experiments made by Watanabe Yuzuruy, graduate, on the effect of heating filtered solutions of koji for one hour at the specified temperatures, the same solution being examined for comparison after standing at the ordinary temperature for the same time.

Below 45° C the change in the composition of the liquid is so small that it may practically be neglected, but between 45°C and 60°C the effect is much more marked. An increase in the amount of solid matter and in the dextrose occurs, accompanied by a decrease in the specific rotatory power. These results are caused by an absorption of water by the dextrin which is converted into dextrose and thus the amount of solid matter in a given volume of the liquid is increased which, together with the smaller specific rotatory power of the dextrose, lowers the specific rotatory power of the solution.

Table 13: Action of Heat on Filtered Solutions of Koji

Temp.

Solids in 100 c.c. of solution

Dextrose in 100 c.c. of solution

Specific rotatory power

Unheated

Heated

Increase

Unheated

Heated

Increase

Unheated

Heated

Increase

30° C.

4.88

-

-

2.97

3.015

0.045

-

-

-

35°

4.88

-

-

2.97

3.062

0.092

-

-

-

40°

4.89

-

-

2.98

3.079

0.099

-

-

-

45°

4.92

4.98

0.06

2.92

3.412

0.494

74.0°

70°

50°

4.95

5.02

0.07

2.793

3.285

0.492

70°

67.1°

2.9°

55°

4.92

5.00

0.08

2.918

3.463

0.545

74°

68.9°

5.1°

60'

4.95

5.02

0.07

2.793

3.30

0.507

70°

67.8°

2.2°

65'

4.89

-

-

2.98

3.081

0.101

-

-

-

70°

4.89

-

-

2.98

3.075

0.095

-

-

-

The alteration is greatest at the temperature of 55° C, above which it rapidly diminishes. At 65° C and at 70° the effect produced is very much the same as at ordinary temperatures, so far as the composition of the liquid itself is concerned, but a very great change in the active properties of the liquid is brought about by heating it to these temperatures. The liquid becomes turbid, so much so that its specific rotatory power cannot be determined with any accuracy, an effect caused by the precipitation of a certain proportion of the albu-menoids which have been rendered insoluble by heating. We shall see that at some temperature between 60°C and 70°C the liquid loses its power of transforming starch into sugar, and reasons will appear connecting this loss of activity with the precipitation of the albumenoids.

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