Frozen yeast 5 years later, not good

[N3Bruce]

After a 3 year hiatus to take care of an aging parent, I am back to my brewing ways. One of the ongoing projects I left behind was a library of stored yeast kept in the freezer section of my beer fridge. While I was away I checked in with my neighbors in case there was a power failure, and to my knowledge there wasn't anything more than an hour or two. The yeast samples were stored in a glycerin solution in sealed test tubes and stored in an insulated box.

Yeah it could have been thicker, but I had pounds of homegrown hops vacuum sealed in there as well. The freezer section was also frost free, so that worked against me too. I still was able to bring back viable yeast after 2 years, but after 5, it seems my luck had run out.

One thing I noticed was a phase separation between the water and the glycerin, with the ice floating over the glycerin as seen in the right photo. The yeast settled into a pill sized deposit at the bottom of the vial, and was dark in color. I had 30 or 40 vials, and this was typical.

I have attempted to revive the yeast with 300 ml of extract at about 1.030 on a stir plate, but all I have after 2 days is basically dark colored starter with very little evidence of yeast growth. I guess I need to rebuild my library, and use or regrow the samples every couple of years. My brother is a biochemist tells me how they do it in their lab, but I can't afford to keep yeast samples on liquid nitrogen.

 

[day_trippr]

Before you toss the whole lot, try using a much weaker starter - like, 1.010 SG - and give it a week at a comfortable temperature...

Cheers!

 

[N3Bruce]

Before you toss the whole lot, try using a much weaker starter - like, 1.010 SG - and give it a week at a comfortable temperature...

Cheers!

I will try, I haven't tossed it yet.

Before you toss the whole lot, try using a much weaker starter - like, 1.010 SG - and give it a week at a comfortable temperature...

Cheers!

Just stepped down to the basement to check on the yeast, after 4 days I am seeing some signs of life from the bubbler. Following your suggestion. I transferred 300 ml of starter in a 500 ml flask into a liter flask adding 500 ml of distilled water from a sealed jug. Let's see how this plays out. Basement is a comfortable 69 degrees.

 

[Dog House Brew]

The yeast isn’t dead, just injured. Don’t be alarmed, shot in the arm ! A few days is nothing IMO when looking for signs. 7 days….I start to worry. I agree to start very week and little volume. I never like waking up to a trumpet! Once I see signs, I give it a little juice. Once it looks finished, I refrigerate, decant, and give it my normal starter run.

 

[deuc224]

Yeah just give it time, my 530 when being woken up takes about 5 days, they are some resilient critters.

 

[N3Bruce]

I have at least 8 or 9 strains, but I also want to check those that proof for off flavors.

 

[McMullan]

N3Bruce, I've had glycerol preps at -20℃ for over 5 years without observing the separation you've observed. It looks to me like there is actually a good chance they thawed at some point(s). Mine are in 1.5ml cryovials, though, and they were homogenised thoroughly before being stored. In your situation, I'd scale things down, say 10ml starter wort @ 1.020 with nutrients, zinc especially. I'd be surprised (assuming few freeze-thaw cycles) if there were no viable yeast cells. The dark colour is a bad omen, but I'd still expect some viable cells. By scaling things down hopefully you'll notice some biological (yeast) activity sooner. It might be that 300ml is a massive underpitch that's going to take a week or two to show any signs of life?

 

[McMullan]

Here's a vial I just pulled from the freezer in my yeast lab. Almost 5 years since it was prepped. (Note I normally culture, QC and restore my master bank after 3 years at the most.)

No signs of any separation either:

The key, apart from prepping small (<2.0ml) volumes, is to ensure the glycerol is well mixed with the yeast and the viable cells have had a little time (about 30 minutes or so, imo) to incorporate it as a substitute for water molecules, especially in the cell wall and membrane. This reduces the risk of hazardous ice crystal formation during freeze-thaw cycles, exactly why glycerol is used. The preps still freeze, however, even at -20℃.

 

[N3Bruce]

I was actually able to bring it back i think, as I said, my freezer is frost free so I have to worry about the freeze thaw cycles. I followed Day Tripper's advise and diluted the starter wort with distilled water, and it looks like I have some
healthy yeast I can build on.

 

[jtgoral]

how was the beer made using this yeast?