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Experiment to test contam resistant substrate
    #1520700 -

I have been thinking about this since I woke up with it in my head this morning. I have read that in Agar cultures, H202 kills mold spores, fungal spores, and bacteria. What I wonder now is whether it be possible to innoculate a peroxide-enriched substrate with live mycellium.

I know there is a recovery time for the mycelium to produce new enzymes when peroxide is introduced. Surely this will not be so severe as to starve the mycelium before it adapts.

My experiment is in the planning stages and is the reason for this post. I am not even a novice chemist, so I need a little advice. This may seem ridiculously simple, but I never had any formal chemistry trainint and am trying to learn as I go.

Stabilized H202 3% has a neutral pH? I assume so by stabilized but I do not
know for sure. If it is not neutral, what additive should be used to counter-act it
in the substrate?

The basic design of the experiment is to use PC Ecuador mycellium currently growing in a 4% by weight honey-water solution to inoculate 12 jars of varying peroxide contents.

Substrate Mixture will be 1/2 verm 1/4 brf (home-ground), and 1/4 water mixture.

Water mixtures for Jars 1-3 will be distilled water

Water mixtures 4-6: 2mL/L stabalized H2O2 3%

Water Mixtures 7-9: 4mL/L stabalized H2O2 3%

Water Mixtures 10-12: 6mL/L stabalized H202 3%

All jars will be sterilized and innoculated with 4CCs mycelial solution and incubated at 80-88 degrees. Mycelial growth in milimeters of width on jar wall will be recorded daily.

My H2O2 concentrations are completely arbitrary. Would different test concentrations be more reasonable?

Any thoughts, obvious problems, or even encouragement?

My initial research turned up a Dutch study that did something similar to this, but I could only find general information about this study and no research data of any kind was available.

The obvious goal of this experiment is to estimate the optimum concentration to ward off contamination without slowing down too much or stopping the mycelium growth.

-Seiko



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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1520789 -

Remember youb have to add the h2o2 after the substrate has been pc'd and has cooled substantially. Otherwise the heat will denature it.

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Re: Experiment to test contam resistant substrate [Re: CultyVader]
    #1520809 -

I didn't know that. Would pasturization denature the H2O2? I'm not even sure what denature means!

-Seiko

EDIT: It sounds sacreligious to say it, but would sterilization/pasturization be necessary for these jars. I hadn't considered that until now. I wonder how much
H2O2 it takes to keep ANYTHING from germinating w/o sterilization.

I think I'm going to attempt to sterilize substrate before, in a flow-hood, mixing in the H2O2, except on the 6mL/L test group. I am going to not sterilize in any way there just to see what happens.


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

Edited by Seiko (05/04/03 11:11 PM)

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521161 -

Peroxide decomposes at 140F so you cannot pasteurize it.
http://www.shroomery.org/index/par/23462
You have to add peroxide just before inoculation. When I'm making grain spawn I open each jar, inject about 5ml of 3% H2O2, close and shake to spread the peroxide and then open again and inoculate.

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Re: Experiment to test contam resistant substrate [Re: zeronio]
    #1521238 -

Has anyone added anti bacterial substances to the grow medium before?
antibiotics? im thinking of trying that on wood chips.Any recomendations?


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Re: Experiment to test contam resistant substrate [Re: Paid]
    #1521243 -

I didn't try that, but I once had a crazy idea of adding a natural antibiotics from other mushrooms, like Trametes versicolor. :tongue:

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521286 -

>Stabilized H202 3% has a neutral pH?

No. Peroxide solution is acidic(depending on the concentration pH from 2 - 6).

Oh, and:
Spelling, spelling, spelling..... Please read http://www.shroomery.org/forums/showflat.php?Cat=&Board=Forum2&Number=1045630 .


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Grow Mushrooms from Spores - PF-Tek for Simple Minds

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Re: Experiment to test contam resistant substrate [Re: Anno]
    #1521418 -

I, of course apologize for my spelling errors. A searchable database is useless if it is full of mispellings.

English is not my first or even second language. I have taken years to learn the ridiculously complex grammar, yet at times, the phonetical nature or my native language, Japanese, comes through.

-Seiko


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521449 -

Quote:
I think I'm going to attempt to sterilize substrate before, in a flow-hood, mixing in the H2O2, except on the 6mL/L test group. I am going to not sterilize in any way there just to see what happens.



Don't forget to tell us the results - it will be interesting to see how much is mycellium slowed down by peroxide.

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521494 -

- Post History Deleted Upon User's Request -

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Re: Experiment to test contam resistant substrate [Re: ]
    #1521580 -

Obviously, you missed the humor of 'mispelling'.

And I DO have to worry about contaminated jars. And this experiment is not born from necessity but from a scientific curiosity

-Seiko


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521617 -

a few technical notes. first, a purely technical point, since everyone wants to point out errors in spelling, i must say something about the incorrect usage of denature. denaturization refers to the process by which one can find out what amino acids (and in what order) make up a protein molecule. like i said, purely technical. second, h2o2 is highly unstable and will decompose (oxidize) when exposed to excess oxygen (in the air) this is true even of the diluted kinds you can find in the first aid section of stores. the upshot of this being that even if you do add h2o2 to your casing or the grain during inoculation it will break down into molecular oxygen, o2, and water. in effect, your casing will only be resistant to contam (and not necessarily all contams like yeasts and certain members of fungi imperfecti) until the h2o2 breaks down, probably in the first day or two, so it is most likely, IMO, that using h2o2 would do very little, if any, good. IMO, you'd be better off improving the sterilitly of your environment that messing around with the peroxide

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Re: Experiment to test contam resistant substrate [Re: effwun]
    #1521726 -

aaaah !!! A japanese on the shroomery !!! Sugoi !!! Irashaimase !!!

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Re: Experiment to test contam resistant substrate [Re: effwun]
    #1521729 -

I see.  Well, it did seem rather obvious not to be common practice.  Thank you.  In light of this information, I don't think I will bother performing the experiment. 

Also, everyone keeps assuming that  my environment must be atrocious just because I am experimenting with contamination resistance.  I do not want this lazy dirty image of myself all over.  :smile:

-Seiko


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521730 -

please excuse me if it seems that i am peeing on your campfire, but this experiment has allready been done in spades. there is a huge amount of info on H2O2 and it's use on this forum, including from the guy who originated the proceedure. before you waste needless effort and money, you should do a thourogh search on the topic, here.

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Re: Experiment to test contam resistant substrate [Re: tchyted]
    #1521740 -

I assume you mean RR Wayne's method. I am aware of it. He uses agar plates and does bulk production, if I'm not mistaken. I was curious about whether this could easily be replicated in brf/verm PF-style jars.

Either way, I've scrapped it in light of effwun's comments.

Thank you all

-Seiko


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: KOPELANDIAA]
    #1521759 -

doumo sumimasen

-Seiko

PS I wonder if my spelling will be questioned now?  :smile: 


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Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1521889 -

- Post History Deleted Upon User's Request -

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Re: Experiment to test contam resistant substrate [Re: ]
    #1522158 -

Well, thank you for the generous compliments. When I was ten, my father forced the four business languages (Japanese, Spanish German, and English) into my studies. I would have cursed him then, but it has been immeasurably beneficial with regard to international correspondence.

MrMushrooms, I didn't mean to imply you were a jerk. Everyone's input has been helpful. If I was short with you it was because so many people were 'explaining' uses of peroxide irrelevant to my experiment.

I think I will still dabble with this. It seems unlikely to yield any useful new information, but to satisfy my curiosity, I think I will still run a slightly less scientific form of the study.

Results will be posted as they are recorded.

-Seiko


--------------------
Seiko Samurai -=- The Way, The One, The Force: Tao

Anything herein is a work of creative fiction not to be treated as even having a basis in fact

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Re: Experiment to test contam resistant substrate [Re: Seiko]
    #1523901 -

Most contam resistant substrates are low in nitrogen, low in available carbohydrates, but contain a large amount of carbon in a form that is not easily digested. Well aged and composted cow or horse manure is an example of a natural contam resistant substrate.

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