The cold hard truth about rinsing yeast with boiled water

[EarlyAmateurZymurgist]

There is a problem with your microbiology here. The beer spoilage organisms you listed (Lacto, Pedio, Aceto) are NOT spore formers. The common sanitation practices are sufficient to avoid these organisms, as long as they are thorough. In fact, the ONLY beer spoilage organism that is a spore-former is from the genus Bacillus and it is sensitive to hops, and thus not a serious threat. Also, I am not sure if you read the source you posted, but they do not describe a single spore-forming bacteria involved in beer spoilage.

Your whole premise of avoiding bacterial spores is irrelevant. Spores are just not a concern, and mostly for the reasons you have already mentioned (particularly pH).

No, part of my premises is avoiding spores. Rinsing yeast with boiled water raises the pH of the culture, which is a concern. People who rinse yeast with boiled water tend to store it under boiled water. Are you telling me that that pH change is not going to allow any spores in the culture to turn into vegetative cells and that there are no flavor affecting bacteria that are spore formers?

 

[cervid]

"For brewing industry, beer spoilage bacteria have been problematic for centuries. They include some lactic acid bacteria such as Lactobacillus brevis, Lactobacillus lindneri and Pediococcus damnosus, and some Gram-negative bacteria such as Pectinatus cerevisiiphilus, Pectinatus frisingensis and Megasphaera cerevisiae."

Source: "Beer spoilage bacteria and hop resistance", Kanta Sakamoto and Wil N. Konings.

http://www.sciencedirect.com/science/article/pii/S0168160503001533

I know.. and we used to have Polio and TB ins this country too. I'll take my chances with the gram negative bacteria you provided..

But hey, thanks for stepping up to the challenge I proposed to EAZ..

 

[mattd2]

"For brewing industry, beer spoilage bacteria have been problematic for centuries. They include some lactic acid bacteria such as Lactobacillus brevis, Lactobacillus lindneri and Pediococcus damnosus, and some Gram-negative bacteria such as Pectinatus cerevisiiphilus, Pectinatus frisingensis and Megasphaera cerevisiae."

Source: "Beer spoilage bacteria and hop resistance", Kanta Sakamoto and Wil N. Konings.

http://www.sciencedirect.com/science/article/pii/S0168160503001533

But what is the significance of the species Cerevisiae? as it is a species of yeast, acetobacteria and Megasphaera.

 

[Bottoms_Up]

But hey, thanks for stepping up to the challenge I proposed to EAZ..

The suspense was killing me.... and I didn't think an answer was forthcoming

 

[EarlyAmateurZymurgist]

But what is the significance of the species Cerevisiae? as it is a species of yeast, acetobacteria and Megasphaera.

Saccharomyces cerevisiae is a yeast species. Megasphaera cerevisiae is a family of gram-negative anaerobic spoilage bacteria.

 

[paperairplane]

So 16 pages later and we determine that slants are better than a mason jar in the fridge - but as always, better is the worst enemy of good enough.

 

[cervid]

So 16 pages later and we determine that slants are better than a mason jar in the fridge - but as always, better is the worst enemy of good enough.

Well.. Not really.. According to EAZ those labs you slant from could be infected to as they are not completely sterile.

You might be slanting brett. I have a vial of it in my fridge now. Minus the fact there is a ton of liquid in it, the yeast look the same color to me..

So.. Is it not good that I make sauerkraut right next to my beer?

 

[aiptasia]

Gentleman and ladies, may I suggest ultraviolet sterilization? If we're really, really concerned over our contaminants in water, then why not just pass your water through a UV sterilizer unit with a small pump?

It seems to work well enough from microbiologists to waste treatment facilities and small units designed for aquaria are cheaper than an autoclave. Intense UV radiation even kills those nasty spores we're all so concerned about.

If you're overly worried about spores and surviving pathogens in your boiled 212 degree water, cool it off and run it through one of these.

 

[ColoHox]

there are no flavor affecting bacteria that are spore formers?

Exactly! There is one Bacillus species that forms spores and can grow in wort, but it is sensitive to ethanol and hops, so it is not a concern.

Spores are not little magic nuggets of bacteria that can just start growing whenever, like in cold water for instance. You can pitch your yeast stock (however you wash, store, harvest, crop, whatever), and it will have spores (not beer spoilage spores, mind you) in it no matter what you do, but the conditions of your beer prevent the growth.

I agree with your assertions that washing with boiled water is not necessary, although certainly not harmful. Are spores what you should be worried about when preparing your yeast? No.

 

[EarlyAmateurZymurgist]

I know.. and we used to have Polio and TB ins this country too. I'll take my chances with the gram negative bacteria you provided..

But hey, thanks for stepping up to the challenge I proposed to EAZ..

You seem to think that you are the only person I have to reply to on this forum. I am fending off attacks from multiple directions. Since you are a doubting Thomas, I challenge you to a store a culture rinsed with boiled water under under boiled water and an unrinsed sample from the same culture under beer. In two months, I want to you send the culture to a reputable lab for analysis. If there is no difference in the bacteria or mold counts between the cultures, I will acquiesce and never discuss this topic again.

 

[AnOldUR]

Rinsing yeast with boiled water is an unnecessary step that does little more than introduce one or more infection vectors.

Infection vector? That must be like an Arctic Vortex. I swear, people must sit around making this stuff up.

But it is entertaining.

 

[boydster]

I know.. and we used to have Polio and TB ins this country too. I'll take my chances with the gram negative bacteria you provided..

So cervid, you took your chance and didn't get a polio vaccine?

Clearly, one can take the attitude that doing the extra work of boiling and cooling water to store the yeast under is good enough to take the chance, or one could take the approach that there is less work involved and fewer chances to be taken by keeping the yeast under green beer. I guess I don't understand why someone would "take [one's] chance" and add in additional steps, given that the alternative seems less involved. I guess I probably don't care to understand, either, because doing more work to take a greater risk seems foolish to me. What am I missing here? Seems like an easy "necessary risk vs. unnecessary risk" question to me.

 

[mattd2]

Saccharomyces cerevisiae is a yeast species. Megasphaera cerevisiae is a family of gram-negative anaerobic spoilage bacteria.

Not what I was meaning - Where all these discovered by some guy called Cerevis or something?

 

[Bottoms_Up]

There seems to be a lot of hypotheticals. I would like to see the results of further tests, such as the one EAZ suggested.

 

[EarlyAmateurZymurgist]

Exactly! There is one Bacillus species that forms spores and can grow in wort, but it is sensitive to ethanol and hops, so it is not a concern.

Spores are not little magic nuggets of bacteria that can just start growing whenever, like in cold water for instance. You can pitch your yeast stock (however you wash, store, harvest, crop, whatever), and it will have spores in in no matter what you do, but the conditions of your beer prevent the growth.

I agree with your assertions that washing with boiled water is not necessary, although certainly not harmful. Are spores what you should be worried about when preparing your yeast? No.

What about vegetative cell growth while stored under boiled water? I have never had spores germinate in crop that I stored under beer; however, I have had spores germinate while stored under boiled water. Either that, or vegetative cell pickup occurred during the transfer. I am fairly anal about liquid culture transfers. I try to get positive pressure going in the culture and I flame or 95% ethanol sterilize all pouring surfaces.

In the ten year period spanning 1993 through 2003, I tried every way to store yeast that was practical in a home brewery environment. While I have experienced okay results with freezing at -20C, my best long-term results have been with slants. I found that it's just easier to maintain a pure culture using slants and basic aseptic lab technique. Every other technique that I used resulted in lower than desired long-term viability. My wife suggested purchasing a desktop lyophilizer. I think she was kidding, but they are starting to reach the affordable point (affordable is a relative term).

 

[EarlyAmateurZymurgist]

Infection vector? That must be like an Arctic Vortex. I swear, people must sit around making this stuff up.

But it is entertaining.

Actually, it's a real term.

 

[EarlyAmateurZymurgist]

Well.. Not really.. According to EAZ those labs you slant from could be infected to as they are not completely sterile.

I did not claim that all liquid cultures are infected. I claimed that yeast manufacturers have a statistically-controlled acceptable level of contamination, which is a fact. Have you ever plated a commercial yeast culture on selective or differential media?

 

[ColoHox]

What about vegetative cell growth while stored under boiled water? I have never had spores germinate in crop that I stored under beer; however, I have had spores germinate while stored under boiled water. Either that, or vegetative cell pickup occurred during the transfer. I am fairly anal about liquid culture transfers. I try to get positive pressure going in the culture and I flame or 95% ethanol sterilize all pouring surfaces.

In the ten year period spanning 1993 through 2003, I tried every way to store yeast that was practical in a home brewery environment. While I have experienced okay results with freezing at -20C, my best long-term results have been with slants. I found that it's just easier to maintain a pure culture using slants and basic aseptic lab technique. Every other technique that used resulted in lower than desired long-term viability. My wife suggested purchasing a desktop lyophilizer. I think she was kidding, but they are starting to reach the affordable point (affordable is a relative term).

It is very unlikely that storage under water was the reason for contamination, and also unlikely that bacterial spores were the source, for the reasons I have mentioned. However, even with the use of a hood, a biocontainment suit, and practiced sterile technique, contamination with vegetative cells from any number of locations is very likely, in all microbiological situations. Assigning the source of the contamination is notoriously difficult. It is just so easy to setup a lovely experiment testing the two storage conditions, bump the tip of your pipette on whatever, and throw all the results off.

I agree that plating or slanting is the best technique for the maintenance of pure cultures. However, bulk storage is best achieved via freezing. Bulk stocks are produced from your pure cultures on your plate and are, with proper technique, clean. For example, storing 50ml of frozen culture sourced from a plated culture is the proper method. Yes, your slant is pure, but upscaling is time consuming and introduces more chances for contamination. Slanting and freezing are different tools for different jobs.

Fortunately I have access to autoclaves, hoods and a lyophilizer but they would sure be more convenient in my basement.

Cheers to sticking to your guns, but scientists (even amateur ones) recognize the value of an open mind and incorporating new information. Send me some of your Ringwood so I can bulk culture it, would ya.

 

[EarlyAmateurZymurgist]

Not what I was meaning - Where all these discovered by some guy called Cerevis or something?

No, cerevisiae is derived from cerevisia, which is the Latin name for beer. Sacchormyces cerevisiae basically translates to sugar fungus of beer.

 

[mattd2]

No, cerevisiae is derived from cerevisia, which is the Latin name for beer. Sacchormyces cerevisiae basically translates to sugar fungus of beer.

Thanks, thought it would be something like that!

 

[EarlyAmateurZymurgist]

I agree that plating or slanting is the best technique for the maintenance of pure cultures. However, bulk storage is best achieved via freezing. Bulk stocks are produced from your pure cultures on your plate and are, with proper technique, clean. For example, storing 50ml of frozen culture sourced from a plated culture is the proper method. Yes, your slant is pure, but upscaling is time consuming and introduces more chances for contamination. Slanting and freezing are different tools for different jobs.

Are you using a true cryogenic freezer? I did not have great success with cryopreservation at -20C using a home freezer. No matter what I did, I always experienced cell rupture. I believe that problem lies in my inability to control the freeze/thaw rate accurately, especially the freeze rate.

Send me some of your Ringwood so I can bulk culture it, would ya.

Sadly, I no longer have that culture. I left the hobby for decade. I should have deposited it, but I did not feel the need at the time (I was fairly burned out from a decade of intense brewing and playing with yeast cultures). The brewery from which I obtained the culture was an early Pugsley-built brewery. The owners sold the brand name to another local brewery and sold the brew house to an out of state interest.

With that said, there is a early Pugsley-built brewpub in operation in my area that still uses their original yeast culture and brew house. I have attempted to talk the brewmaster into giving me crop, but have had no luck thus far. Getting my old crop was as simple as taking a 4oz sterile baby food with me when I visited the defunct Pugsley-built brewery. I asked the brewer if I could have one of his hydrometer samples, and he said, "Yes." It was dumb luck.

 

[EarlyAmateurZymurgist]

Thanks, thought it would be something like that!

What's weird is that only one Latin-based language has a name for beer that is even close is Spanish.

 

[gwapogorilla]

Water boils at 212F/100C at standard atmospheric pressure (atm). If we raise the pressure under which water is boiled to 15 pounds per square (psi) inch above normal atmospheric pressure, the boiling point increases to 250F/121C. Basically, the water inside of a 15 psi pressure cooker operated at sea level experiences a little more two atms.

Is it the water that raises above 212*...or the steam? Everything I've learned through the years says water gets to 212*, and that's it. Steam on the other hand is hotter than water, "when pressurized". In a pressure cooker, it's the steam due to the increased pressure that raises in temp, not the water itself.

 

[harrke]

The steam and water are in a saturated environment (meaning that there is as much vapor becoming liquid as there is liquid becoming vapor). In this case, the "steam" and "water" are at the same temperature.

 

[mtnagel]

So cervid, you took your chance and didn't get a polio vaccine?

Clearly, one can take the attitude that doing the extra work of boiling and cooling water to store the yeast under is good enough to take the chance, or one could take the approach that there is less work involved and fewer chances to be taken by keeping the yeast under green beer. I guess I don't understand why someone would "take [one's] chance" and add in additional steps, given that the alternative seems less involved. I guess I probably don't care to understand, either, because doing more work to take a greater risk seems foolish to me. What am I missing here? Seems like an easy "necessary risk vs. unnecessary risk" question to me.

But what I haven't seen answered is how to store harvested slurry for several months without agar slants and freezing? Many people have used rinsed yeast successfully up to a year later. But everything I've read says to not use the harvested yeast slurry after 2-4 weeks. I don't brew enough to do that. How can I make my slurry last longer without agar slants and freezing (which I may look at in the future)?

 

[cervid]

If i didn't ask, i don't know, 5 times for am answer from you that would be one thing. If you didn't directly respond to me twice after i asked the question and still be unable to answer said question, well that's a different matter. Just say you don't know.

I got all my vaccines, you can't go to college if you don't have them. I don't think I got MMR though, if I did, I didn't pass my titer test 6 years ago to get operating room access privileges. I've risked my life many times, as I'm sure everyone else has. I'm not so uptight that I'm afraid to risk a little 5 gallon batch of beer. Our a pint of yeast.

But, if you're the expert on thus, why haven't you performed said experiment yet? I think the onus I'd on you to do that. Maybe you should've done that before debating in such a know it all manner, speaking as if you have facts and are dealing with certainties.

Oh and, I don't think anyone is attacking you, it's a debate. You brought it on yourself. If you're going to argue against the way people have been doing something for so long, the burden of proof is on you.

Note: on phone working, sorry if spellcheck got me.

 

[masonsjax]

But what I haven't seen answered is how to store harvested slurry for several months without agar slants and freezing? Many people have used rinsed yeast successfully up to a year later. But everything I've read says to not use the harvested yeast slurry after 2-4 weeks. I don't brew enough to do that. How can I make my slurry last longer without agar slants and freezing (which I may look at in the future)?

It means you shouldn't use only the slurry after four weeks but should instead make a starter from that slurry. You can safely store it much longer than a month but you should make a starter before using it.

 

[AnOldUR]

Infection vector? That must be like an Arctic Vortex. I swear, people must sit around making this stuff up.

Actually, it's a real term.

But obscure enough that when Googled I only get reference to computer viruses. You're definitely a bright guy, but have this flaw of latching on to stuff like "boiled water isn't sterile" and become obsessed with it. You revel in spitting out your interpretation of science in the faces of people knowing that they don't have the education to contradict you. Like I said, taking it in with an open mind this is entertaining, but can also be educational. But what you say has to be taken with a grain of salt. Is it useful, practical information to us as homebrewers or is it someone’s attempt to boost their ego? I think a little of both.

Please don't take this as a personal attack. It's my honest observation. I'll take my blue collar opinions into lurk mode and only observe from here on out.

 

[masonsjax]

Infection vector, as in another site for infection to take hold. Makes sense to me. *shrug*

 

[rpkincaid]

Does anyone know what goes into white labs tube or wyeast packs with the yeast? Is it fermented wort/media or rinse water? Anyone know the pH of the liquid in those packages?