I posted this thread a while back. I am back in the growing scene and would like to continue active research on the topic. I am going to begin the first wave of experiments soon, and tackle the rest later when I have more time.
Purpose
To study the effectiveness of Potassium Metabisulfite (K2S2O5) as a sterilizing medium and bacterial contamination inhibitor.
Synopsis
Potassium Metasulphite (Kmeta) has long been used in the winemaking industry to sanitize equipment before brewing. Furthermore, it is used to prevent bacterial contaminations while brewing.
K2S2O5 + H20 = 2 KHS03
The Kmeta breaks down into two new molecules that have a Sulfite ion. It is also a mild acid. Some of the Kmeta also oxydizes and forms SO2 (sulphur dioxide). A portion of SO2 binds with H2O and forms H2SO4 (sulphuric acid)
Sulfites have long been used in the food industry as a preservative. Sulfites are extremely effective at inhibiting bacterial growth. Sulfites are not toxic to bacteria or fungus, but rather make the environment unfavorable to growth. In small doses, Kmeta is used to prevent bacterial contamination in winemaking. In larger doses (5% solutions), Kmeta releases a large amount of SO2. Sulphur dioxide disrupts electron transfer in both all forms of life (plant, fungus, bacterial, etc). This in turn causes the SO2 to be converted to H2S (Hydrogen Sulfide), which is toxic. Sulphur also forms a protective layer that prevents spore production in fungus. Lastly, Sulphuric acid corrodes cell walls of all life forms, although the dilute Sulphuric Acid does relatively little to combat contaminations. Sulphur has been used as a natural fungicide for many years.
At this point you may be wondering how such a toxic chemical could be of any use. The inspiration comes from the wine industry. Yeast is a fungus that is used to make alcohol. The winemaking industry has bred and isolated strains of yeast that are resistant to sulfites and sulphur, giving them an edge over the natural 'wild' yeasts.
The purpose of this experiment is two-fold. First, determine the nescessary (if possible) concentration of Kmeta needed to sterilize growth medium. Kmeta will not kill endospores. However, the availability of water in the Kmeta solution will allow the endospores to germinate. Once germinated, the bacteria will find itself in a highly unfavorable environment (sulfite and sulphur). It may not be possible to sterilize growth medium without using a concentration of Kmeta that is entirely toxic to fungus.
Secondly, the goal is to isolate and breed a strain of Cubensis that is tolerant to sulfites and sulphur, much like the wine industry has done with yeast. A sulfite resistant strain will allow growth medium to be lightly sulfited to further prevent bacterial contamination. This is similar to using hydrogen peroxide in growth medium, however it is longer lasting (hydrogen peroxide has a short half-life, whereas the sulfites and residual sulphur stay around much longer). This will provide active bacterial contamination prevention. Substrate jars, once sterilized, have no protection against contamination. They are easily colonated by contaminations. A light sulfiting will allow the substrate to actively 'fight' off some contaminations, increasing the chance that the grow is sucessful.
Procedure
**First of all, this is subject to change. I'm still figuring out how I want to try things.
The first area of study is to determine the nescessary (if possible) concentration of Kmeta needed to sterilize growth medium. As mentioned before, this may not be possible without using rediculously high concentrations of Kmeta. Even if full sterilization is not possible, it will give a general idea how effective Kmeta is at slowing bacterial growth in substrates.
1) Prepare 16 substrate jars. I will be using spelt flour and vermiculite in the ratio 1 part flour to 3 parts vermiculite.
2) Prepare one part water to 4 parts substrate mix. Add enough Kmeta to bring solution to 5%. Add this to one substrate jar. Repeat this for three more jars.
3) Repeat step 2 for solution percentages 10%, 15% and 20%
4) Label jars and leave open to air overnight. This is to ensure that some airborne contaminants make it into the jar)
5) Seal jars and place in incubator at 86F.
*Note on Kmeta solution: We are working with incredibly small amounts of liquid, which means reliable and accurate measure. I do not have a high precision gram scale. Therefore, I'm making the liquid in bulk. One teaspoon of Kmeta (6.2 Grams) to one gallon of water is a 5% solution (930 ppm). Two tsp to one gallon is 10% (1860 ppm). Three tsp to one gallon is 15% (2790 ppm). Four tsp to one gallon is 20% (3720 ppm)
The second area of study is to breed a strain of cubensis that is tolerant to sulfites. This will allow the substrate to be lightly (or moderately, depending on the results of the first experiment) sulfited and still allow fungus growth.
1) Prepare 5 agar media. Increase the sulfite solution in each dish (1%-5%)
2) inoculate, incubate. Begin strain isolation process of the highest (successful) sulfite percentage.
3) Once single strain has been isolated, inoculate 5 different identical (non sulfited) substrates.
4) Grow until fruitation, collect spores.
5) Repeat above procedures with new spores. Sulfite solutions should start at the original strain's percentage and increase from there.
If the experiment should ever stall (no growth at sulfite solution), continue fruiting the next highest (successful) strain for several generations. After strong growth is observed, continue increasing sulfite percentage.
Comments welcome.
|
Would the fruiting bodies contain or carry sulfites? Sulfite allergies are not uncommon and as one who suffers from the aforementioned allergy I wouldn't touch a sulfite laced mushroom with a ten foot pole.
--------------------
The smart way to keep people passive and obedient is to strictly limit the spectrum of acceptable opinion, but allow very lively debate within that spectrum - even encourage the more critical and dissident views. That gives people the sense that there's free thinking going on, while all the time the presuppositions of the system are being reinforced by the limits put on the range of the debate
-Noam Chomsky
|
Threaded
Previous
Index
Next
Bulk Cannabis Seeds 
Bulk Substrate 
Maeng Da Thai Kratom Leaf Powder 
Mono Tub Substrate 
Isolated Cubensis Liquid Culture For Sale 
Buy Kratom Extract 
All-In-One Bags That Don't Suck
Some of these posts are very old and might contain outdated information. You may wish to search for newer posts instead.
polyfractal 
Edit 
Reply 
Seiko
1 2 3 4 ... 12 13 all )
Hippie3
polyfractal
GnuBobo
Shaw