Chapter 4. Action of Koji Extract Upon Some Carbohydrates

Brown and Heron have shown that when an aqueous solution of malt is allowed to remain in contact with a solution of cane sugar, a change takes place by which the cane sugar is made to take up water and is thereby converted into invert sugar, a mixture of dextrose and levulose. The diastase of malt is said by them to exert its maximum effect upon sugar at 55°C; its action is considerably weakened at 60°C and almost destroyed at 66°C.

Experiment shows that the extract of koji also possesses the property of causing cane sugar to become inverted, but I am not able to define the limits of its action. The following experiments will suffice to prove this point.

Experiment 1: 1.974 gram of dry cane sugar was dissolved in 25 cc of koji extract, then diluted with water to 100 cc. The amount of rotation was found to be 15.8 divisions, and the calculated number 15.5 divisions.

1.974 grams case sugar dissolved in 100 c.c. gives rotation	= 12.1 div.
25 c.c. koji solution diluted to 100 c.c. gives rotation	= 3.4
	15.5 div.

After being allowed to stand for 18 hours at about 10 to 12°C the rotation was found to have diminished to 5 divisions, and the solution contained 1.67 grams of glucose. Deducting 0.36 gram contained in 25 cc of koji solution, the amount, formed from the cane sugar was 1.31 gram, equivalent to 1.2445 gram cane sugar and hence 0.7294 gram of unaltered cane sugar was present. We thus find the calculated number of divisions rotated by the inverted solution to be +5.43 against 5 divisions actually observed.

Unaltered cane sugar (0.7294 grams in 100 c.c.)	+4.4 div.
Koji extract (25 c.c.) in 100 c.c. of water)	+3.4 div
Invert sugar formed	-2.37 div.
	+5.43 div

Calculated in degrees of arc the specific rotatory power of the can sugar has been reduced from 74° to 10°.

Experiment 2: A solution of case sugar containing 5.41 grams in 100 cc, and giving a rotation in a 200 mm tube of 33.1 divisions, equal to [a] = 74°, was employed. 75 cc of this solution was mixed with 25 cc of a solution of koji which contained in 100 cc 1.46 gram of solid matter, 1.0125 gram of glucose, and which gave in a 200 mm tube an optical rotation of 8 divisions. It may be remarked that from this and other experiments made with the same solution of koji, it was found to be exceptionally weak in its converting power. The observations are as follow, after deducting the optical rotation due to the presence of the koji solution:

	Optical rotation	Specific rotatory power
At starting	24.8 div.	74°
After 1-1/2 hours at 15°C	23.7	70.6
After 20-1/4 hours at 10-12°	21.0	62.6
After 3/4 hour more at 40°C	20.2	60.2

50 cc of this mixture and 25 cc of koji were treated as below. The numbers given are corrected for the koji present:

	Optical rotation	Specific rotatory power
After 1-1/4 hours at 40°C	11.2 div.	50°
After 2 hours at 45-50°	4.0	17.8

The experiment was not carried further than this. At low temperature the converting action of this particular extract of koji is very slow, but at higher temperatures, and especially at from 45° to 50°C it is much more rapid. In this respect, therefore, koji extract resembles malt extract.

Action Upon Maltose

The maltose employed was obtained from ame, a kind of sweetmeat prepared by the action of malt in solution upon the starch contained in millet or in rice. Various specimens of ame contained from 68 to 94% maltose, which was separated according to the process described by O'Sullivan³. The specimens employed were in the crystalline state, and contained water sufficient to reduce the specific rotatory power from 150° to 144.5°.

Table 14: Action of Koji on Maltose

Time	Solids in 500 cc after deducting koji	Rotation after deducting koji	Specific rot. power of maltose prods.
10 minutes	2.826	5.8 div.	124.2°
30 minutes		5.3	111.1
1 hour	2.886	4.7	98.5
2 hours		3.7	77.6

The specific rotatory power therefore, fell from 144.5° to 77.6° in 2 hours, and would doubtless have fallen to 59° if the solution had not been used up after 2 hours. The action may be represented in the form of a curve using time and specific rotatory power as abscisse and ordinates respectively (See Figure 3).

The curve shows very clearly how regular the action is, and leaves no doubt about the power of extract of koji to effect the hydration of maltose. It is especially important to establish this, because this property marks in the sharpest manner the difference between malt extract and koji extract. Brown and Heron's experiments leave no doubt about the inability of malt extract to convert maltose into dextrose, and these experiments, I think, establish conclusively the ability of koji extract to do this.

Action Upon Dextrin

Experiment 6: 100 cc of a solution of commercial dextrin containing 5.56 grams of solid matter were diluted to 250 cc and then gave a specific rotatory power [a]j = 174°. It was, therefore, impure, and contained a considerable percentage of dextrose.

50 c.c. of this solution were mixed with 50 c.c. of a solution of koji and heated to 45°C for 1-1/2 hour. After being diluted to 250 c.c. the solution contained 4.145 grams of solid matter in 250 c.c., and deducting 1.285 gram contained in the 50 cc of koji solution added, we get 2.86 grams of solids formed from the 50 c.c. of dextrin solution, instead of 2.78 grams originally present. After making allowance for the rotation caused by the koji solution, the specific rotatory power of the dextrin products was 92°, instead of 174° that of the substance at starting. The koji solution had become exhausted, because when an additional amount of koji solution was added, and the mixture heated for a longer time, the specific rotatory power further diminished to 85°. This experiment leaves no doubt concerning the gradual absorption of water by dextrin under the solution of koji.

² Jour. Chem. Soc. 1879. Trans., p. 621.

³ Journ. Chem. Soc. 1876, ii, p. 127.