Aspergillus oryzae

Aspergillus flavus has the capacity to grow on many nutrient sources. It is mainly a saprophyte and grows on dead plant and animal tissue in the soil, which is important for the recycling of nutrients. Because it grows well on decaying vegetation, one common place you might find Aspergillus is in compost or on fallen leaves. However, this fungus is also a parasite and can use living organisms as hosts. Host species include corn, peanuts, nut trees, cotton, and even humans.

Typically this organism favors hot, dry conditions which makes it unique in comparison to other fungi. The best temperature for growth is 37°C (98.6°F). However, it can grow at any temperature between 12-48°C (54-118°F). Aspergillus flavus is omnipresent in nature but has a more limited growth indoors. Aspergillus molds thrive best in oxygen-rich environments and also on materials that are rich in carbon which they feed off of for nutrients.

This cycle is essential for producing various fermented foods, including sake, soy sauce, and miso. Here’s an overview of the cropping cycle:

1. Preparation of Substrate

• Selection of Substrate: Typically, polished rice, soybeans, or barley are used as substrates.
• Cleaning and Soaking: The selected substrate is thoroughly washed to remove impurities and soaked in water to hydrate.
• Steaming: The hydrated substrate is steamed to soften it, making it more suitable for mold growth.

2. Inoculation

• Cooling the Substrate: After steaming, the substrate is allowed to cool to a temperature favorable for mold growth (30-32°C or 86-90°F).
• Inoculation with Spores: Aspergillus oryzae spores are evenly sprinkled over the cooled substrate and mixed thoroughly to ensure even distribution.

3. Incubation and Growth

• Incubation Conditions: The inoculated substrate is placed in a warm, humid environment to promote mold growth. Ideal conditions are 30-32°C (86-90°F) with high humidity.
• Monitoring and Mixing: During the incubation period, the substrate is monitored and occasionally mixed to ensure uniform growth and prevent overheating.
• Duration: This stage typically lasts for 40 to 48 hours. During this time, the Aspergillus oryzae spores germinate and mycelium spreads throughout the substrate, converting it into koji.

4. Harvesting

• Completion of Growth: The koji is considered ready when the substrate is fully colonized by the mold, indicated by a white or yellowish mycelium covering the substrate.
• Harvesting: The fully grown koji is harvested and can be used immediately for fermentation processes or dried for later use.

5. Post-Harvest Processing

• Drying (Optional): If not used immediately, the koji can be dried to prolong its shelf life. Drying is done under controlled conditions to preserve the enzymatic activity.
• Storage: Dried koji should be stored in a cool, dry place to maintain its quality until it is ready to be used.

Ingredients and Equipment

• 1 kg (2.2 lbs) sushi rice
• 2 liters (2 quarts) water
• 1 teaspoon yeast (sake yeast or wine yeast)
• 5 grams (0.18 oz) Aspergillus oryzae (koji mold spores)
• Fermentation vessel (glass or food-grade plastic)
• Steamer or large pot for cooking rice
• Cloth or towel
• Thermometer
• Stirring utensil (sanitized)

Steps

1. Prepare the Rice

1. Rinse the Rice: Wash the sushi rice thoroughly in cold water until the water runs clear.
2. Soak the Rice: Soak the rice in water for about 4-8 hours, then drain.
3. Steam the Rice: Steam the rice for about 45 minutes or until it is fully cooked and sticky. Allow it to cool to room temperature.

2. Inoculate the Rice with Koji Mold

1. Spread the Rice: Spread the cooled steamed rice on a clean surface or a large tray.
2. Inoculate with Aspergillus oryzae: Sprinkle the koji mold spores evenly over the rice. Mix well to ensure even distribution.
3. Incubate the Rice: Place the rice in a shallow container and cover it with a cloth. Keep it in a warm place (30-32°C or 86-90°F) for 48 hours. Stir the rice every 12 hours to ensure even growth of the mold.

3. Create the Starter Mash (Moto)

1. Prepare a Portion of Koji Rice: Take about 200 grams (7 oz) of the koji rice.
2. Mix with Water and Yeast: Mix the koji rice with 1 liter (1 quart) of water and the yeast in the fermentation vessel.
3. Ferment the Moto: Allow this mixture to ferment for 5-7 days at room temperature, stirring daily.

4. Create the Main Mash (Moromi)

1. First Addition (Hatsuzoe):
   • Add another 300 grams (10.6 oz) of steamed rice and 1 liter (1 quart) of water to the moto. Mix well.
   • Allow this mixture to ferment for 1 day.
2. Second Addition (Nakazoe):
   • Add another 300 grams (10.6 oz) of steamed rice and 1 liter (1 quart) of water. Mix well.
   • Allow this mixture to ferment for 1 day.
3. Third Addition (Tomezoe):
   • Add the remaining 200 grams (7 oz) of steamed rice and 1 liter (1 quart) of water. Mix well.
   • Allow this mixture to ferment for about 20-30 days, stirring daily.

5. Pressing and Filtering

1. Press the Mash: After the fermentation period, strain the mash through a fine mesh or cheesecloth to separate the liquid (sake) from the solid rice particles.
2. Filter the Sake: You can further filter the sake through activated charcoal if desired to improve clarity.

6. Pasteurization (Optional)

1. Heat the Sake: To pasteurize, heat the sake to about 60°C (140°F) for a few minutes. This step is optional but helps to stabilize the sake and prevent spoilage.

7. Bottling and Aging

1. Bottle the Sake: Transfer the sake into clean, sanitized bottles.
2. Age the Sake: Allow the sake to age for a few weeks to a few months in a cool, dark place to develop its flavors.

1. Dissolve the malt extract and agar in distilled water.
2.  Sterilize the solution by autoclaving at 121°C for 15-20 minutes.
3. Pour the sterilized media into petri dishes or other containers under sterile conditions.
4. The Yeast is optional, but you can add it to any recipe for nutritions, same for Peptone.

Aspergillus oryzae thrives on substrates such as polished rice, barley, and soybeans, with polished rice being the most commonly used. To make a starter mix, combine 1 kg of polished rice with Aspergillus oryzae spores (typically 0.1-0.2 grams or according to the supplier's instructions). The rice should be washed, soaked for 6-8 hours, steamed for 45-60 minutes, and then cooled to 30-32°C before inoculating with the spores. Mix thoroughly to ensure even distribution, then incubate in a warm, humid environment for 40-48 hours until the rice is fully colonized by the mold.

1. Nutrient-Rich Additives

• Yeast Extract: Provides additional vitamins and amino acids to support mold growth.
• Peptone: Supplies nitrogen and promotes enzymatic activity.
• Soy Flour: Enhances protein content and supports robust mold development.

2. Mineral Supplements

• Magnesium Sulfate (Epsom Salt): Essential for enzyme activation and overall growth.
• Potassium Phosphate: Supports cellular energy and growth processes.

3. Sugars

• Glucose: Simple sugar that provides an immediate energy source for the mold.
• Malt Extract: Rich in carbohydrates and nutrients, supporting vigorous growth.

4. pH Adjusters

• Calcium Carbonate: Helps maintain a stable pH, creating a favorable environment for mold growth.
• Citric Acid: Can be used in small amounts to adjust pH levels as needed.

5. Moisture Enhancers

• Humectants (e.g., Glycerol): Help retain moisture within the substrate, providing a consistent environment for growth.

Example Additive Mix

For a 1 kg batch of substrate (polished rice), you might add:

• 5 g yeast extract
• 5 g peptone
• 2 g magnesium sulfate
• 2 g potassium phosphate
• 10 g glucose

The biological efficiency of Aspergillus oryzae is largely determined by its ability to convert substrates into valuable products through fermentation. This mold is highly efficient in breaking down complex carbohydrates, proteins, and lipids into simpler compounds, thanks to its robust enzymatic profile, which includes amylases, proteases, and lipases. These enzymes facilitate the rapid and thorough degradation of substrates like rice, barley, and soybeans, resulting in the production of koji, a critical ingredient in traditional Japanese fermentations such as sake, miso, and soy sauce. The efficiency of Aspergillus oryzae in this process is not only crucial for high yield and quality of the final products but also for ensuring the economic viability of fermentation operations. Its effectiveness in converting substrates to valuable end products showcases its significant role in both traditional and modern biotechnological applications.

Aspergillus flavus var. oryzae is a pathogenic fungus whose spores are ubiquitous in nature. It can cause aspergillosis in animals and humans.

Aspergillus oryzaecan often be found growing inside dirty air conditioners inside of buildings or in your own home. This has even been known to be a problem in some hospitals. Aspergillus may also grow on or inside walls in homes, especially if the house is damp or has been damaged by flooding.

Conidiophores:

• Conidiophores are the specialized structures that bear the spores. They are typically long, smooth, and terminate in a vesicle where the spores are formed.

Colony Morphology:

• Colonies of Aspergillus oryzae on agar media typically appear velvety or powdery, starting white, then becoming yellowish, and finally greenish-yellow as the conidia mature.

Microscopic Features:

• Under a microscope, Aspergillus oryzae shows septate hyphae, conidiophores with a swollen vesicle at the tip, and chains of conidia.

Aspergillus oryzae strength is that it produces a rich, fruity, and refreshing flavor, so it is still used by some producers despite the difficulty and great skill it requires.

Aspergillus oryzae, commonly known as koji mold, is a filamentous fungus used extensively in fermentation processes, particularly in Asian cuisine. Its composition includes a variety of nutritional and biochemical components that make it effective in breaking down complex organic materials into simpler forms.