Kōji (food)

Fermentation starter

Kōji (ニホンコウジカビ, 日本麹黴, ‘nihon kōji kabi’) refers to various molds of the genus Aspergillus sp., which are traditionally used in East Asian cuisine for the fermentation of food. In Japanese, kōji refers to both the Aspergillus starter culture and mixtures of Aspergillus with wheat and soybean meal. It can be fried and eaten directly or processed to a sauce.[1]

## Characteristics

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Four Aspergillus species in a Petri dish. The bottom two are strains of A. oryzae

Various types of kōji are used, including yellow, black, and white.[2][1] The kōji is stored for two to three days at 30 °C under high humidity to allow A. oryzae to grow.[3] In this process, the starch from cereals such as wheat, buckwheat or barley as well as from sweet potato is split into glucose, creating a sweet taste. Due to the amino acids glutamic acid and to a lesser extent also aspartic acid split off from the proteins during fermentation, a strong umami taste is created on the human tongue when consumed.[4][5] Depending on the Aspergillus used, culture substrate and culture conditions (temperature, pH value, salt content, humidity), different products are created in terms of composition, flavour and odour.[6] Kōji can be freeze-dried and crushed to produce spores.[7] Dried kōji-spores can be stored and transported light-protected at room temperature.

### Yellow kōji

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Aspergillus sojae on soybeans and wheat

Yellow kōji is used, among other things, for the production of soy sauce,[3][4] miso,[8][9] sake,[10] tsukemono, jiang, makgeolli, meju, tapai, kōji-amazake, rice vinegar,[11] mirin, shio koji[12] and natto. Typically, for the production of soy sauce (shoyu), soybeans and sometimes also wheat are swollen in water, steamed, and possibly mixed with wheat bran roasted at 160–180 °C and ground. The enrichment with kōji creates a moist mash.[3]

There are three Aspergillus species that are used as yellow kōji:

* Aspergillus flavus var. oryzae[13] (キコウジキン / 黄麹菌 ‘ki kōji-kin’). The growth range of this species includes pH values from below 2 to above 8, a temperature optimum of 32 – 36 °C, a temperature minimum of 7 – 9 °C and a temperature maximum of 45 – 47 °C.[14] The colony color is initially yellow-green, later more or less brown.[14]
* Aspergillus sojae[15][16] (醤油麹菌 ‘shōyu-kōji-kin’)
* Aspergillus tamarii[3][4]

A. oryzae has three α-amylase genes, which allows it to break down starch relatively quickly into glucose.[3] In contrast, A. sojae has only one α-amylase gene under a weak promoter and the CAAT box has a gene expression attenuating mutation (CCAAA instead of CCAAT), but has a higher enzyme activity of endopolygalacturonase and glutaminase.[3] A too rapid release of glucose from starch at the beginning of fermentation inhibits the growth of the microorganisms in the maturation phase.[3] For the breakdown of proteins to amino acids, A. oryzae strain RIB40 has 65 endopeptidase genes and 69 exopeptidase genes, and A. sojae strain SMF134 has 83 endopeptidase genes and 67 exopeptidase genes.[3] Similarly, starch-degrading enzymes (glucosidases) are more strongly expressed and protein-degrading enzymes (proteases) less strongly expressed in A. oryzae, and the odour profiles differ significantly.[17] A. sojae has 10 glutaminase genes.[18] Various mutants of A. oryzae with altered properties were generated by irradiation[3] or by the CRISPR/CAS method.[19][20][21] Similarly, mutants of A. sojae with altered properties were generated by a variant of the CRISPR/Cas method[21] or chemical mutagenesis.[22]

### Black & white kōji

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Aspergillus niger on MEAOX-Agar Aspergillus tubingensis on Czapek-Agar

Black kōji produces citric acid during fermentation, which inhibits the growth of unwanted microorganisms.[2] It is typically used for the production of Awamori.[2][12]

There are three Aspergillus species that are used as black kōji:[2]

* Aspergillus luchuensis (synonym Aspergillus awamori, Aspergillus inuii, Aspergillus nakazawai and Aspergillus coreanus, クロコウジキン / 黒麹菌 ‘kuro kōji-kin’)
* Aspergillus niger (synonym Aspergillus batatae, Aspergillus aureus or Aspergillus foetidus, Aspergillus miyakoensis and Aspergillus usamii including A. usamii mut. shirousamii)
* Aspergillus tubingensis (synonym Aspergillus saitoi and A. saitoi var. kagoshimaensis)

White kōji (Aspergillus kawachii) is an albino variant of Aspergillus luchuensis.[6] It is typically used in the production of Shochu.

## History

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The process of making rice wine and fermented bean paste using moulds was first documented in the 4th century B.C.[23] In 725 AD the Japanese book Harima no Kuni Fudoki ('Geography and Culture of the Harima Province') first mentioned kōji outside of China and described that the Japanese produced kōji with fungal spores from the air.[24][25] Around the 10th century, the kōji production method underwent a change and moved from the natural sowing system in rice to the so-called tomodane. This involved cultivating kōji until spores were released and using the spores to start a new batch of production.[26] In the Meiji era, the integration of new microbiological techniques made it possible to isolate and propagate kōji in pure cultures for the first time. These advances facilitated the improvement of mushroom culture quality and the selection of desirable characteristics.[27]

It later became known that Kōji comprises different species of Aspergillus. Aspergillus oryzae was first described in 1878 as Eurotium oryzae Ahlb.[28] and in 1883 as Aspergillus oryzae (Ahlb.) Cohn.[29][30] Aspergillus luchuensis was first described in 1901 by Tamaki Inui at the University of Tokyo.[31][32][33] Genichiro Kawachi isolated a colourless mutant of A. luchuensis (black Kōji)[34][35] in 1918 and named it Aspergillus kawachii (white Kōji). Aspergillus sojae was first described as a distinct species in Kōji in 1944.[36][37] Initially, Aspergillus sojae was considered a variety of Aspergillus parasiticus because, unlike the other fungi of Kōji, it had never been isolated from the soil.[38]

## Literature

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* H. Kitagaki: Medical Application of Substances Derived from Non-Pathogenic Fungi and -Containing. In: Journal of fungi. Band 7, Nummer 4, März 2021, S. , doi:10.3390/jof7040243, PMID 33804991, PMC 8063943.
* J. E. Smith (6 December 2012). Aspergillus. Springer US. pp. 46ff. ISBN 978-1-4615-2411-3.

## References

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2. ^ a b c d S. B. Hong, O. Yamada, R. A. Samson: Taxonomic re-evaluation of black koji molds. In: Applied Microbiology and Biotechnology. Volume 98, Number 2, January 2014, p. 555–561, doi:10.1007/s00253-013-5332-9, PMID 24281756 (Review).
3. ^ a b c d e f g h i K. Ito, A. Matsuyama: Koji Molds for Japanese Soy Sauce Brewing: Characteristics and Key Enzymes. In: Journal of fungi. Volume 7, Number 8, August 2021, doi:10.3390/jof7080658, PMID 34436196, PMC 8399179.
4. ^ a b c C. Diez-Simon, C. Eichelsheim, R. Mumm, R. D. Hall: Chemical and Sensory Characteristics of Soy Sauce: A Review. In: Journal of agricultural and food chemistry. Volume 68, Number 42, October 2020, p. 11612–11630, doi:10.1021/acs.jafc.0c04274, PMID 32880168, PMC 7581291. (English)
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16. ^ William Shurtleff, Akiko Aoyagi: History of Koji – Grains And/or Soybeans Enrobed with a Mold Culture (300 BCE To 2012). Soyinfo Center, 2012, ISBN 978-1-928914-45-7.
17. ^ J. Li, B. Liu, X. Feng, M. Zhang, T. Ding, Y. Zhao, C. Wang: Comparative proteome and volatile metabolome analysis of Aspergillus oryzae 3.042 and Aspergillus sojae 3.495 during koji fermentation. In: Food research international. Band 165, März 2023, S. 112527, doi:10.1016/j.foodres.2023.112527, PMID 36869527.
18. ^ K. Ito, Y. Hanya, Y. Koyama: Purification and characterization of a glutaminase enzyme accounting for the majority of glutaminase activity in Aspergillus sojae under solid-state culture. In: Applied Microbiology and Biotechnology. Band 97, Nummer 19, Oktober 2013, S. 8581–8590, doi:10.1007/s00253-013-4693-4, PMID 23339014.
19. ^ J. I. Maruyama: Genome Editing Technology and Its Application Potentials in the Industrial Filamentous Fungus. In: Journal of fungi. Volume 7, Issue 8, August 2021, S. , doi:10.3390/jof7080638, PMID 34436177, PMC 8399504.
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31. ^ Inui Tamaki (1901). "Ryukyu awamori hakko kin chyosa houkokusyo". J. Chem. Soc. Japan. 4: 1421–1430.
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33. ^ Takeo Koizumi (2018-04-05). "黒麹菌の役割 発酵中の雑菌繁殖防ぐ". Okinawa Times. Archived from the original on 2023-04-04. Retrieved 2023-04-10.
34. ^ K. Kitahara, M. Kurushima: Studies on the diastaic enzyme systems of molds (Third report). About the Awamori white koji mold. Part 2. Is Asp. kawachii really a mutant of black Aspergillus? In: J. Ferment. Technol. (1949), Volume 27, p. 182–183. (Japanisch)
35. ^ K. Kitahara, M. Yoshida: Studies on the diastaic enzyme systems of molds (Third report). About the awamori white koji mold. Part 1. Morphological and several physiological characteristics. In: J. Ferment. Technol. (1949), Volume 27, p. 162–166.
36. ^ Katsumi Yuasa, Kazuya Hayashi, Takeji Mizunuma: A new criterion by which to distinguish Aspergillus sojae, a Kōji-mold, from related taxa producing echinulate conidia. In: Agricultural and biological chemistry. 1982, Volume 46, Nummer 6, p. 1683–1686 doi:10.1271/bbb1961.46.1683.
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