Stressbaby's Sanitizer Experiments

Discussion in 'Yeast, Additives & Wine Making Science' started by Stressbaby, Nov 3, 2018.

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  1. Nov 3, 2018 #1

    Stressbaby

    Stressbaby

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    There has been debate in other threads about sanitizers, contact time, and rinsing. I went to Amazon and bought 20 tryptic soy agar plates to test some of these ideas myself. This is the thread in which I'll post my experiments and results. Feel free to suggest experiments. I'll run the tests any way I can until such time as I run out of plates.

    An important assumption in all of these experiments is that the pathogens grown on the tryptic soy agar do not differ substantially in their response to the sanitizer interventions than the wine pathogens with which we are concerned. If someone has thoughts on how to verify this assumption, please post below.
     
    Last edited: Nov 3, 2018
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  2. Nov 3, 2018 #2

    Stressbaby

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    Quickie experiment #1 - StarSan on the hands

    One plate, four quadrants. I made two fingerprints in each quadrant of the agar plate. Each quadrant got fingerprints from different fingers of each hand. So quadrant 1 was index left and pinkie right; quadrant 2 was middle left and ring finger right, etc.
    Quadrant 1 - no sanitizer
    Quadrant 2 - StarSan, immediate (minimal contact time)
    Quadrant 3 - StarSan, 1 min contact time
    Quadrant 4 - StarSan, 1 min contact time, rinsed

    Will post back in a few days.
     
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  3. Nov 3, 2018 #3

    Stressbaby

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    Experiment #2

    In this experiment I'll try to more formally mimic what happens when sanitizing winemaking equipment. The design I have in mind looks like this: I have some leftover country wine, pH 3.3 and SO2 <10 by titrette method. I'll take soda straws and uniformly expose them to pathogens by rolling them in my hands and on the work surface. Straw 1 goes into test tube #1 with 10ml of wine, that's the control. Then each straw will be treated according to a sanitization protocol and go into it's own test tube. All the straws will be swirled thoroughly and 0.1ml of wine drawn from the test tube and placed on the agar plate.

    1. Control
    2. StarSan, minimal contact time
    3. StarSan, 1 min contact time
    4. StarSan, 3 min contact time
    5. StarSan, left to dry
    6. StarSan, 1 min contact time, brief tap water rinse
    7. StarSan, 1 min contact time, extended tap water rinse

    I would then repeat this with KMS, altering the contact times.
    I would also repeat this using a wine with adequate SO2.

    I've set up a proof of concept plate to 1) assure my wine is not contaminated; 2) assure that this method yields a reasonable number of control colonies; and 3) to see how much variability there is in the number of colonies produced. I don't want to go through the entire experiment only to find that the control doesn't yield enough colonies to tell whether the interventions had any effect or that there is so much variability that the results aren't reliable. So I put 0.1ml of wine without any other contamination or intervention into quadrant 1 of a fresh agar plate. Then I contaminated a straw and placed it in 10ml of wine and swirled. From that I took 0.3ml and placed 0.1ml in each of the other 4 quadrants. What I'm hoping for here is minimal to no growth in quadrant #1 and some decent number of colonies in each of the other 4 quadrants. Failing to get that, we'll have to revise the methods.

    Everyone is welcome to post suggestions.
     
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  4. Nov 11, 2018 at 2:42 PM #4

    Stressbaby

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    IMG_1633.jpg Update on experiment #1: All four quadrants grew lots of colonies. In this experiment, there doesn't appear to be any lower counts using the StarSan on the hands. The quadrant with the rinse looks worse, but the colony count isn't really that much different; some other bug showed up here and these larger colonies just overgrew the smaller ones.
     
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  5. Nov 12, 2018 at 12:52 PM #5

    vacuumpumpman

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    Thanks for doing this !

    There are so many ideas about the proper way to santize, I do believe this is a great wato test those theories
     
  6. Nov 12, 2018 at 12:55 PM #6

    Stressbaby

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    Thanks. The proof of concept test plate for experiment #2 didn't work out. Grew one colony on each section, so that is not sufficient numbers. Will need to redesign that. Considering actually inoculating the straws. Stay tuned.
     
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