Mycelium Leather

The dutch company Ecovative grows Reishi mycelium into the air, separates it from the substrate and compresses it to make leather. I try to recreate this procedure.

Literature

Growth environment

To prevent fruiting, CO₂ concentration is kept at 5-7% and the temperature at +-30° C. A mister maintains the humidity above 99%. The hyphae grow up to escape CO₂ and get oxygen.

Airflow increases homogeneity by delivering humidity and/or minerals, by eliminating micro climates or by mechanical force.

Minerals in the mist increase growth speed by osmotic pressure.

A humid substrate means faster growth but lower density.

Growth substrates

From the patent "Increased homogeneity of mycological biopolymer grown into void space":

Corn stover	600 g
Poppy seeds	144 g
Maltodextrin	25 g
Calcium sulfate	8 g
Water	1600 g
Spawn	288 g

From the paper "MycelioTronics: Fungal mycelium skin for sustainable electronics"

dry beech (FR: hêtre) wood shavings	1 kg
organic full-grain spelt flour	50 g
fine plaster (CaSO4)	25 g
water	2 l
beech wood–based inoculum	500 g

Procedure

After compression, the mycelium is left for 3 days to strengthen, then dried at 43° C followed by heat pressing if desired.

My own experiments

First test in a monotub

First I made grain spawn out of bird seeds. Then I sterilized a box with rubbing alcohol, lined the bottom with a clean plastic film and spread sterilized substrate made of 0.75l hardwood pellets, 0.2l wheat bran and 0.75l water onto the film. I distributed some grain spawn on top of it, closed the box and waited.

After one day, the mycelium already comes out of the grain spawn.

The mycelium has almost colonized the whole surface, but probably got crushed by water droplets

The mycelium colonized the surface and grew about 1.5 cm thick as in the pictures. After four days I sprayed water on it to keep it humid, but the droplets crushed the fluffy mycelium and it stopped growing.

My first try to do mycelium leather

I let it dry and peeled it off the substrate. It was fused with the chunks of grain spawn, so I didn't get one big tissue but many small pieces, the biggest one shown in the picture. Depending on how much it was crushed it feels like paper (but more fragile) or like a very soft, light and flexible tissue. It is very thin and rips apart easily.

What to improve

The grain spawn should be broken into smaller pieces using disposable gloves, and evenly mixed with the substrate. Using unpopped popcorn as grains may be a good idea since it doesn't stick together as much as bird seeds.

Experimenting with substrates in petri dishes

On the 2.5.2023 I filled petri dishes with Agar, malt extract, maizena, yeast extract and gypsum according to the following table.

Nutrient tests on petri dishes

Ingredient	Substrate	Malt	Maizena	Yeast	Gypsum	8 days	12 days
1	Agar					very bad
2	Agar	0.5 g				good	completely covered dense
3	Agar			0.05 g		very bad	contam
4	Agar	0.5 g		0.05 g		bad	contam, bad
5	Agar	0.5 g		0.05 g	0.025g	bad	contam, dense
6	Agar		0.5 g			very bad
7	Agar		0.5 g	0.05 g		very bad	contam, jumped to the ceiling
8	Agar		0.5 g		0.025g	bad	contam
9
10	Pellets & Bran					good
11	Pellets & Bran		0.5 g			very good
12	Pellets & Bran			0.05 g		very good
13	Pellets & Bran				0.025g	very good

To accurately measure those values I dissolved the nutrients in water solutions and used a pipetter. However gypsum is not soluble so the quantities may be inaccurate, and Maizena clumped together, so I put clumps of it into the petri dishes.

Results

Results are in the above ingredients table. After 12 days the mycelium on wood (pellets) substrate clearly shows two zones: The first early zone near the inoculation point is very thin and abruptly a second zone starts where it is thick. I see the following hypotheses.

1. It first had to get used to wood again, since the grain spawn was popcorn, bird seeds and liquid culture based on malt exctract. This would explain why it doesn't happen on agar: malt extract is more similar to grains than to wood. Test by inoculating one wood dish with grain spawn and one with a piece of the thick zone. Compare the thickness of the growing mycelium.
2. It first grew fast due to the grain energy but then slowed down, and slower growth could allow more thickness. Test as before, also comparing the growth speed. Another test can be calculating the growth speed from pictures.
3. It oscillates between thick and thin, as Rachel observed on another mycelium in agar filled petri dishes. Test by waiting to see if another thin layer forms after the thick one.
4. Old mycelium has no purpose anymore and gives some of its mass to growing mycelium. Test by looking if the zone interface moves forward.
5. At some point there was enough mycelium to raise CO2 levels, and the growing mycelium grew not only forward but also up into the air to find oxygen. Test by putting an airtight dome over one dish the next time I open the box to take photos, and see if this mycelium gets even thicker.

Heated inoculation chamber

It would be nice to have a chamber that can heat sterilize itself and the substrate, and then keep the temperature constant at 25 or 30° C. I am buying the XH-W1209 thermostat: https://www.berrybase.ch/xh-w1209-digitales-thermostat/temperaturschalter-12v-frei-programmierbar

A very cheap CO₂ meter

Since an electronic CO₂ sensor costs 60 CHF I make an analog one. Henry's law states that the amount of a dissolved gas in water is proportional to the partial pressure in the air above the water. Furthermore, dissolved CO₂ lowers the ph of water. This means that by filling a small bowl with distilled water and adding a few drops of pH indicator liquid we use its color to guess the CO₂ concentration of the air. Since the math are complicated, we just assume a linear relation and calibrate with CO₂ made from baking soda and vinegar and an expensive sensor.

[CO₂] = a(T) + b(T)*pH

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