Kombucha Bacterial Cellulose

Kombucha tea is a traditional health-promoting fermented beverage that has been around for several thousand years. It is produced by fermenting sweetened tea with a symbiotic colony of bacteria and yeast. Kombucha is a traditional beverage consumed for its many health benefits. The cellulose matrix formed in the culture medium can be used as biocellulose fabric to create clothing or biopaper. But it can also be used in cosmetology for its anti-inflammatory, antioxidant and anti-cancer properties.
Bacteria: Acetobacter xylinum, A. xylinoides, A. aceti, A. pasteurianus, Bacterium gluconicum. Yeasts: Schizosaccharomyces pombe, Kloeckera apiculata, Saccharomycodes ludwigii, Saccharomyces cerevisiae, Zygosaccharomyces bailii, Brettanomyces bruxellensis, B. lambicus, B. custersii and Pichia.

Protocol: how to start a bacterial cellulose culture?
Instructions: Boil 1.5L of distilled water and add 10-15g of tea bags (approximately 2gr of tea per bag). Allow to infuse for 5 minutes and add 1/2 cup sugar, glucose, fructose (1/2 cup agave, etc). Allow to cool to room temperature and add 100mL of a previous batch (liquid medium and a piece of the cellulosic matrix, the "mother" or baby kombucha). Seal the bottle and let the culture ferment and grow at room temperature for 5-7 days. After this period you can restart and/or take out the fermented tea and start a second fermentation with citrus, hibiscus flower, ginger, etc. in another bottle for 2-3 days. Optional: add 100mL of cider vinegar. The final pH reading should be between 2.5 and 3.2 (you can buy PH paper stripe in the pharmacy), to avoid contamination of the symbiotic culture with undesirable micro-organisms.
The culture does not need too much oxygen. Yeasts hydrolyse sucrose into glucose and fructose, producing ethanol and carbon dioxide as metabolites. Acetic acid bacteria convert glucose to gluconic acid and fructose to acetic acid.

Content: What is a biofilm?
In fact, the kombucha sheet forming on top of the culture is a type of biofilm. Biofilms are the most common way in which microorganisms exist in nature. They are very resistant and have very interesting physical properties.
A biofilm is any group of microorganisms in which cells stick to each other and often these cells adhere to a surface. These adherent cells are frequently embedded within a self-produced matrix of extracellular polymeric substance (EPS). Biofilm extracellular polymeric substance, which is also referred to as slime (although not everything described as slime is a biofilm), is a polymeric conglomeration generally composed of extracellular DNA, proteins, and polysaccharides (like the bacterial cellulose we find in kombucha). Biofilms may form on living or non-living surfaces and can be prevalent in natural, industrial and hospital settings.

LATEST EXPERIMENTS

We got some great 'jun' kombucha from Patrik d'Haeseleer and here is the current protocol.

These make very clean and light-colored bacterial cellulose biofilms.

With 10% glycerol, one can even get the 'dried' scoby to not crumble away, and have a more leather like texture.

We also did some mITS1/4 pcr, and got sequence for at least one scoby member - a bavarian brewer's yeast. (also known as a wine spoiling yeast, here in swiss romande! =P)

Previous documentation here:

aim: to grow big scobies in food trays.

expt1) trays tested English Breakfast vs earl gray in Sept 2021, but Javier said need to have more oxygen and we flipped lids.

mold contam!

also had bottle that kept a good smell, but no scoby...

expt2) 27jan2022 RA

test Mg++ for aiding them grow?

Try 2 trays of tea, english breakfast. about 5L per tray + 500ml each kombucha tea and half of the fresh part of current scoby (from Javier's batch, 27jan2022)... check pH and maybe add vinegar!

one try will get 2mM extra MgSO4 (make 2M MgSO4 with 12g/50ml - add 5ml to one of the trays. let cool before adding scoby and have lids not sealed, so o2 can get in)

- have 633g of sugar, so about 3.5L per tray in the end (afdd 3.5ml of 2M MgSo4 to one of them. with a big +) - both covered with checkered cloth... (to protect from light)

about pH3...

have 1/4 scoby for each and 1/2 to big jar with more fresh sweet tea to incubator...

To Note: lab room gets colder than incubator! check temp when arriving tomorrow...