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-   -   Do you know how to make a yeast starter? Then why not farm yeast and freeze it? (http://www.homebrewtalk.com/f163/do-you-know-how-make-yeast-starter-then-why-not-farm-yeast-freeze-269488/)

IsItBeerYet 01-29-2012 08:08 PM

1 Attachment(s)

Well,

After a failed attempt or two to get the yeast list into HTML format, guess I'll just have to attach it as a screenshot. These are my current working strains.

I do remember that I've tried 1028, 1728, and Brett with no or little success.

@BBL - the older strains aren't as large and contain more glycerin (I started at around a 50/50 mix by volume, and have since gone to a 1/3 glycerin by volume mix with water. As a matter of fact, I have some older tubes that are still liquid in my -20f freezer!) When I started, I got a lot of trub in there as well, but since, I have started washing and now, I put almost 100ml of dense yeast slurry in each 125ml tube. One thing that improved my viability (perceived, I have not tested with empirical tests) is a "cold rest". I let my last yeast step-up sit until fermentation is complete and all the yeast has settled, then put it in the refridgerator for a few days. At this point, I will decant as much wort as I can, mix the yeast slurry and remaing wort, and decant into sterilized tubes. I put the tubes into the fridge to settle, and once the slurry had settled and compacted, I decant the last of the wort and replace it with the glycerin/water mix.

A quick shake to mix the yeast and glycerin mix, then off to the freezer.

I have read some things in "the Yeast book" by Jamil and Chris White that talked about mixing up some fresh wort with the glycerin along with a touch of citric acid and using that as a storage medium, but haven't tried it.


Forkhead 11-07-2012 08:12 PM

2 Attachment(s)
Quote:
Originally Posted by Brewitt View Post
That all said, if one uses the cells to inoculate a starter, I think it is a moot point. An overnight starter should give you about a 10X increase in cell number which way exceeds the necessary cell number for a 5-10 gallon batch.
I agree that if you're doing a starter, it shouldn't matter much. However, I've been doing some playing around with the Wyeast calculator and have generated the following graph on the top showing the 'Expected Fold Expansion' vs. 'Starting Cell Concentration'. I think this is the most useful way to represent the data because it can be used for any culture as long as you know your volume and cell #. From it, you can estimate the expected final cell #. The second graph, titled 'Actual Fold Expansion' is data generated from my own cultures. It comes very close to the Wyeast data.

I'd like to point out a few interesting things. 1st, you get more expansion if you start at a lower conc. than at higher. For example, if you made a 1L starter with 100 billion cells, that'd be 100e6/ml, and you could expect the cells to expand 2 fold for a final # of 200 billion. If you made the same 1L starter with only 10 billion cells, i.e. 10e6/ml, you can expect a 7 fold expansion for a final # of 70 billion. This is definitely something to keep in mind when planning your step-ups.

My data pretty much confirms the Wyeast data, except I've been getting better expansion at low concentrations, and worse expansions at high concentrations. The Wyeast data also predicts cultures higher than 200e6/ml to roughly double, whereas I am unable to get hardly any expansion above that concentration, which is why I cut the graph off at that point. I'm pretty sure the Wyeast Pitch Rate Calculator is based on data from cultures with starting concentrations near the middle of the graph, and that the numbers it gives you for higher and lower concentrations are extrapolations, which are not accurate. I should also remind those who haven't been following along that I start all my cultures by oxygenating with pure oxygen and a diffusing stone, and then use a stirplate along with an air pump to provide continuous aeration.

Forkhead 11-07-2012 08:12 PM

2 Attachment(s)
Quote:
Originally Posted by Brewitt View Post
That all said, if one uses the cells to inoculate a starter, I think it is a moot point. An overnight starter should give you about a 10X increase in cell number which way exceeds the necessary cell number for a 5-10 gallon batch.
I agree that if you're doing a starter, it shouldn't matter much. However, I've been doing some playing around with the Wyeast calculator and have generated the following graph on the top showing the 'Expected Fold Expansion' vs. 'Starting Cell Concentration'. I think this is the most useful way to represent the data because it can be used for any culture as long as you know your volume and cell #. From it, you can estimate the expected final cell #. The second graph, titled 'Actual Fold Expansion' is data generated from my own cultures. It comes very close to the Wyeast data.

I'd like to point out a few interesting things. 1st, you get more expansion if you start at a lower conc. than at higher. For example, if you made a 1L starter with 100 billion cells, that'd be 100e6/ml, and you could expect the cells to expand 2 fold for a final # of 200 billion. If you made the same 1L starter with only 10 billion cells, i.e. 10e6/ml, you can expect a 7 fold expansion for a final # of 70 billion. This is definitely something to keep in mind when planning your step-ups.

My data pretty much confirms the Wyeast data, except I've been getting better expansion at low concentrations, and worse expansions at high concentrations. The Wyeast data also predicts cultures higher than 200e6/ml to roughly double, whereas I am unable to get hardly any expansion above that concentration, which is why I cut the graph off at that point. I'm pretty sure the Wyeast Pitch Rate Calculator is based on data from cultures with starting concentrations near the middle of the graph, and that the numbers it gives you for higher and lower concentrations are extrapolations, which are not accurate. I should also remind those who haven't been following along that I start all my cultures by oxygenating with pure oxygen and a diffusing stone, and then use a stirplate along with an air pump to provide continuous aeration.

Forkhead 11-07-2012 09:53 PM

1 Attachment(s)
Quote:
Originally Posted by Brewitt View Post
I just assume about 1-2x10e8/ml as max density when brewing.
I think that's a safe assumption as long as you're starting your culture at a concentration higher than 20e6/ml, as illustrated when you express the data as below.

Golddiggie 12-08-2012 04:30 PM

1 Attachment(s)
Quote:
Originally Posted by BBL_Brewer View Post
My yeast always settle out in the fridge before freezing. Not a problem.
Good to know. Now I'm not worried about them being good when I go to use them. Thinking about getting a tiny cooler (saw them at Lowes) to keep them in the freezer. Might put an ice pack in there too. If nothing else, it will help prevent any warmer air exposure (however brief) when getting into the freezer...

BTW, these things were SOLID within a few hours of being put into the freezer. Basically, in the time between my posts (~7 hours) they went from liquid to SOLID. I'll take some pics of the vials either tonight or tomorrow and post them up.

Picture of the vials... Graded A, B and C from left to right.

ldave 01-14-2014 03:08 AM

Proposed: increase 4 deg C storage time with glycerol contact to 3 days
 
3 Attachment(s)

So, I've recently discovered this thread. I, too, have had an interest in freezing yeast and am assembling the gear to do it. But the knowledge... that's been a little trickier. This thread has been a gold mine of data for me and I'd like to throw in something I came across as it seems relevant.

When I suspect that a good deal is known about something pretty technical but nobody technical is talking, I turn to patent searches. They can refuse to give *you* any info, but if it's a coveted process it gets spelled out in a patent. So, I came across this patent:

1) Granted in 1997. It deals with baker's yeast and the cryoprotectant properties of adding glycerol in various concentrations and freezing at -21 deg C. Concentrations were .2 M, .3 M, and .4 M. The .4 M concentration had the greatest improvement of yeast viability .4 M translates to just 3.7% glycerol in water concentration by weight (2.8% by volume), an interesting level of concentration given Forkhead's experiment indicating that, at this freezing temp, he found 7.5% by volume concentration ideal. I think this brackets the range we're looking at: 3% to 8%. Pretty small doses.

2) This excerpt shows glycerol uptake by yeast over a period of days. Interesting, huh? Remember, though, glycerol is yeast poison, so these extended contact times, while increasing glycerol uptake, don't translate necessarily to better viability. There must a sweet spot.

3) Finally, this excerpt shows proof times after freezing then thawing and proofing the yeast. Using the proof times of baker's yeast is a clever way of assessing yeast viability without high tech equipment, I think. Bottom line: 3 days at 4 deg C is optimum data point for yeast viability. Based on this, I would recommend that the 4 deg C storage time be increased from 48 hours to 72 hours.

A lovely creation this thread and it's associated tutorial is.

Thanks to all.


ldave 01-14-2014 03:08 AM

Proposed: increase 4 deg C storage time with glycerol contact to 3 days
 
3 Attachment(s)

So, I've recently discovered this thread. I, too, have had an interest in freezing yeast and am assembling the gear to do it. But the knowledge... that's been a little trickier. This thread has been a gold mine of data for me and I'd like to throw in something I came across as it seems relevant.

When I suspect that a good deal is known about something pretty technical but nobody technical is talking, I turn to patent searches. They can refuse to give *you* any info, but if it's a coveted process it gets spelled out in a patent. So, I came across this patent:

1) Granted in 1997. It deals with baker's yeast and the cryoprotectant properties of adding glycerol in various concentrations and freezing at -21 deg C. Concentrations were .2 M, .3 M, and .4 M. The .4 M concentration had the greatest improvement of yeast viability .4 M translates to just 3.7% glycerol in water concentration by weight (2.8% by volume), an interesting level of concentration given Forkhead's experiment indicating that, at this freezing temp, he found 7.5% by volume concentration ideal. I think this brackets the range we're looking at: 3% to 8%. Pretty small doses.

2) This excerpt shows glycerol uptake by yeast over a period of days. Interesting, huh? Remember, though, glycerol is yeast poison, so these extended contact times, while increasing glycerol uptake, don't translate necessarily to better viability. There must a sweet spot.

3) Finally, this excerpt shows proof times after freezing then thawing and proofing the yeast. Using the proof times of baker's yeast is a clever way of assessing yeast viability without high tech equipment, I think. Bottom line: 3 days at 4 deg C is optimum data point for yeast viability. Based on this, I would recommend that the 4 deg C storage time be increased from 48 hours to 72 hours.

A lovely creation this thread and it's associated tutorial is.

Thanks to all.


ldave 01-14-2014 03:08 AM

Proposed: increase 4 deg C storage time with glycerol contact to 3 days
 
3 Attachment(s)

So, I've recently discovered this thread. I, too, have had an interest in freezing yeast and am assembling the gear to do it. But the knowledge... that's been a little trickier. This thread has been a gold mine of data for me and I'd like to throw in something I came across as it seems relevant.

When I suspect that a good deal is known about something pretty technical but nobody technical is talking, I turn to patent searches. They can refuse to give *you* any info, but if it's a coveted process it gets spelled out in a patent. So, I came across this patent:

1) Granted in 1997. It deals with baker's yeast and the cryoprotectant properties of adding glycerol in various concentrations and freezing at -21 deg C. Concentrations were .2 M, .3 M, and .4 M. The .4 M concentration had the greatest improvement of yeast viability .4 M translates to just 3.7% glycerol in water concentration by weight (2.8% by volume), an interesting level of concentration given Forkhead's experiment indicating that, at this freezing temp, he found 7.5% by volume concentration ideal. I think this brackets the range we're looking at: 3% to 8%. Pretty small doses.

2) This excerpt shows glycerol uptake by yeast over a period of days. Interesting, huh? Remember, though, glycerol is yeast poison, so these extended contact times, while increasing glycerol uptake, don't translate necessarily to better viability. There must a sweet spot.

3) Finally, this excerpt shows proof times after freezing then thawing and proofing the yeast. Using the proof times of baker's yeast is a clever way of assessing yeast viability without high tech equipment, I think. Bottom line: 3 days at 4 deg C is optimum data point for yeast viability. Based on this, I would recommend that the 4 deg C storage time be increased from 48 hours to 72 hours.

A lovely creation this thread and it's associated tutorial is.

Thanks to all.



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