Jual Culture Aspergillus oryzae, Aspergillus sojae
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Aspergillus oryzae and Aspergillus sojae strains used for shoyu production have focused on
comparing these two fungal species and improving their enzyme-producing
abilities. A study of the enzymatic differences between 11 strains of A. oryzae
and 20 strains of A. sojae showed that the activities of neutral, acid, and
alkaline proteases, xylanase, pectin lyase, phosphatase, and aminopeptidase
were not significantly different. However, acid carboxypeptidase activity and
α-amylase activity were higher from A. oryzae when compared with A. sojae
strains, while endopolygalacturonidase activity was much higher from A. sojae
than from A. oryzae (Terada et al., 1980). The ratios between α-amylase
activity and endopolygalacturonidase activity of 0.5–2 for A. sojae and 20–2000
for A. oryzae were suggested as a differentiation criterion for the species
(Terada et al., 1980). Hayashi et al. (1981) compared the performance of these
two fungi in shoyu production. They found that the activities of protease,
acetic carboxypeptidase, and α-amylase were lower and those of
endopolyglueuronidase and glutaminase were higher in koji made with A. sojae.
In the moromi stage, the proportions of NH3 nitrogen (N), glutamic acid N, and
total Ν were higher, and viscosity and heat residue were lower with A. sojae.
The resulting concentrations of citric and succinic acids in the shoyu were
significantly higher (p < 0.001) with A. sojae than with A. oryzae. Ishihara
et al. (1996) compared the volatile components in commercial koikuchi shoyus from
different factories using either A. oryzae or A. sojae and found that the
concentrations of 1- and 2-propanol, furfuryl and benzyl alcohols,
ethyl-benzoate, and lactate, acetate, pyrazines, carbonyl compounds such as
ethanal, maltol, and phenyl acetaldehyde, phenol, and others, were higher in
the shoyu from factories using the latter fungus, but concentrations of
2-methyl- and 3-methyl − 1-butanol, 2-phenyl ethanol, 2-methyl- and
3-methyl-butanoic acid, 3-methylthio − 1-propanol, HEMF, 4-ethyl guaiacol, 4-ethyl
phenol, and others were greater in shoyu from factories using the former
fungus. These results have prompted factory managements to use A. sojae for
koji production.
Using an unusual
system, Yasui et al. (1982) tested a range of koji fungal strains for
glutaminase production and found that, when a strain showing 16% higher
glutaminase activity than its parent strain was compared with its parent in the
production of shoyu, the final glutamic acid concentration was 10% higher.
In the early
1950s, A. sojae KS was irradiated with X-rays by Iguchi to produce strain X-816
of A. sojae (Sekine et al., 1970). Sekine et al. (1970) obtained seven strains
with superior alkaline phosphatase activity (130–190%) and highly active
protease, peptidase, cellulase, and amylase activities that were better at
decomposing soybean protein. Yokoyama and Kadowaki (1983) UV-irradiated A.
sojae strain Η and obtained mutant strains with total protease activities 2.5
times that in wheat bran and soy sauce kojis. The mutant strains were
diploidized and combined with natural mutants from Μ strains, and strains TH
and D-15 were produced that possessed higher total protease activities than the
Μ strains, and grew well. However, UV irradiation may stimulate the production
of toxic elements in otherwise safe fungi. Kalayanamitr et al. (1987)
UV-irradiated A. flavus var. columnaris Raper and Fennel (ATCC44310) to obtain
mutant strains with high protease and amylase activities, and light-colored
conidia. Some selected mutant strains were found to be acutely toxic to
weanling rats, even though they were negative for aflatoxin production. The
investigators suggested that the toxic compound could be one of four
substances: maltorhyzine, aspergillic acid, kojic acid, or cycoopiazonic acid.
Furuya et al.
(1983) fused, with an efficiency of 1%, protoplasts derived from two strains of
A. oryzae, one with a high growth rate and the other producing high levels of
protease. Two strains derived from successful fusions showed high stability,
fast growth, and abundant sporulation and produced 2.3 times more protease than
the parent fast-growth strain.
The growth and
development of microorganisms on defatted soybean and ground wheat koji
prepared with A. sojae were studied by electron microscopy by Kitahara et al.
(1980). Growth of the mold on the surface of the soybean was rapid up to 24 h,
at which point formation of sporing bodies began, and spores were released
within 40 h. However, very little fungal growth was seen on the wheat surface,
but yeasts were seen growing on the wheat. Growth of Micrococcus species became
noticeable after 16 h, as did multiplication of lactobacilli. These
observations on the growth of the koji mold are at odds with the observation
that 10–20% of the dry matter in koji is lost in the koji stage (Takeuchi et
al., 1968) and the observations below on the significant consumption of
carbohydrate during the koji stage. I suggest that significant penetration of
the wheat endosperm should have been seen.
During koji
production, carbohydrate is consumed by the fungus, thus leaving less
carbohydrate available to provide flavor compounds for the final shoyu produced
(Furuya et al., 1985). This carbohydrate consumption is positively correlated
with α-amylase activity in koji culture. To overcome the depletion of
carbohydrate before the moromi stage, Furuya et al. (1985) derived mutants that
utilized 10–50% less carbohydrate during preparation of koji than the parent
strain, with about 1/3, 1/20, and 1/150 of the α-amylase activity of the parent
strain of A. oryzae. Significantly increased amounts of carbohydrate-derived
compounds were found in the resulting shoyu made with these mutants.
Enhanced
glutaminase activity in koji is desirable to increase glutamic acid production
in soy sauce, and reduced conidial production in the koji reduces contamination
of the air with floating conidia (Ueki et al., 1994a). A mixed tane koji of two
koji fungi, A. oryzae strains K2 and HG, increased glutaminase activity of the
mixture to 11.3 units · g− 1 dry koji, which was higher than the 4.7 or 4.4
units · g− 1 dry koji produced by the K2 strain or HG strain, respectively, and
conidia production was reduced tenfold (Ueki et al., 1994a). The mixed tane
koji was used in the manufacture of soy sauce, and the resulting mixed koji
made with 3.6 tons of defatted soybean and of wheat grain showed high
glutaminase activity (5.5 units · g− 1 dry weight koji) when compared to strain
K2 alone (1.8 units · g− 1 dry weight koji). In addition, the number of conidia
in the mixed culture was 2.5 × 107 g− 1 dry koji, which was lower than 1.3 ×
108 g− 1dry koji produced by strain K2 alone. The glutamic acid content of the
raw soy sauce was 1.25 times higher than the glutamic acid level found in
normal soy sauce (Ueki et al., 1994b).
Kim and Cho
(1975) investigated soy sauce production in Korea using a soy–wheat koji
prepared with A. sojae, using natto, a soy bean product prepared with Bacillus
natto, and using a mixture of the two in varying proportions. The natto–brine
mixture had protease activity twice as high as the koji alone, and this was
reflected in the protease activities found in mixtures of the natto and koji.
On comparing the organoleptic qualities of soy sauces fermented for 3 months,
the koji:natto at a ratio of 6:4 had the best flavor, followed by koji alone.
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