Breeding for Ethanol tolerance

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LG11
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Breeding for Ethanol tolerance

Post by LG11 »

Hi sorry for the delay in my return!
I have been involved in a local primary school that wanted to bring enhanced science lessons to primary age children (9-11), so I have kinda been flat out making class material and lesson plans!
Anyway I had a PM asking if I would shed some light on how yeast can be bred for Ethanol tolerance............ Unfortunately this is normally a contentious issue so I will explain what I know and shed light on a few facts.
If you studied the material in the other thread I posted about brewing, you will no doubt be fully aware that yeast is a member of a GROUP of organism responsible for the production of alcohols in brewing. For the sake of clarity will just look at the role of the yeast we put in on purpose and disregard the other organisms.
First lets be really clear what we are talking about here, Ethanol is a type of Alcohol but it is by no means the only produced by fermentation. It has a set tolerance to Ethanol and while you can get yeast that claims to produce >14% Alcohol they are not claiming > 14% ETHANOL, however this dosnt mean you cant breed yeast that can produce a cleaner brew. The process is pretty straight forward and requires little apart from some selective petri dishes, microscope and some alginate (or agar at a push). The trick is to be utterly sterile when starting the process, make sure EVERYTHING is sterile and sealed when in use. We take daily samples from all breeding and testing vessels and plate them up, we look for bacterial contamination mainly and often find it in commercial yeast mix's. The expectation when buying a yeast strain is you are getting a pure strain, however this is pretty rare and the norm is a mix of different yeast strain's. As long as the bacteria are kept out this isnt normally a problem.
Personally most of the best results I have seen in 35+ years is by starting with a low concentration of yeast, dont aim to brew a batch just use a sugar like GLUCOSE, try and avoid white sugars at all cost as these tend to mess with ATP production and lead to side chain alcohols like amyl alcohol. Use RO water or distilled water, and thats it..............Dont add anything else as your brewing yeast and not making a beverage!
Use a temperature of between 72f-79f and the more stable the better, the idea is to keep as many variables under control as you can. Keep telling yourself to resist the temptation to mess!! And remind yourself your not making [insert drink here], if you want to test for contamination then get some of the venflon type valve attachments (I will come back with a link) and take samples with sterile syringe via a small Venous tube and valve. DONT think taking the lid off for a couple of mins once a day wont matter, over a 6-8 week period it will matter and will likely lead to contamination.
Once all gas has st0pped wait 4 days the start looking at the cells on the bottom of the container, dont look deeper than the top 1-2mm there is little point. The live cells that were the last to stop will be found at the top of the pile not the bottom, the ones further down may be viable and strong but they gave out first. What your looking for is single or double cells in the process of splitting, if you dont find any (you should) then look for the very very top cells that look like 4 cells inside a single cell.
You dont need many (a single one will do) and plate it up on selective agar, this is the start for the next generation. If you keep strict records then after 3-4 times doing this (no one said it was quick) you should see your Ethanol becoming cleaner (less other alcohols present), we use a gas chromatograph to test this but if you do it correctly this shouldnt be needed. I would say smell will tell you but then I will be asked what it should smell like, this is a problem as its a bit subjective but you get to know the right smell.
Anyway thats a really quick outline, I intend to do a full post on this and maybe a video showing the steps. But it should do in answer to the question I was asked, its worth having a go and if you stick at it and pay close attention to Aseptic technique etc then after 6 generations you should start to have yeast cultures way way better than you can buy.
One thing to keep in mind and something of a well kept secret is the fact that many TOP distilleries (scotch whiskey), use yeast from batches started many years ago and spend ALOT of money keeping the strains pure and vigorous. Some of the work we do is investigating strain die outs from when things go wrong, however these days most distilleries pay companies like mine to keep yeast banks for them.
I will also do a video and post on how to ferment without using yeast :D
Regards
Dr Jason Bell
InglisHill
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Re: Breeding for Ethanol tolerance

Post by InglisHill »

Dr Bell,

Do you have access to a GC.

If you ever found the time would you be able to run some distillations using some different seals that are in use to see if they leech.

PTFE, Silicone, flour paste, viton and there are some others as well.

It has been difficult for the armature distillers to get any clear information on these things and would be of great benefit to the distilling community at large if we could access some real data about it.

Cheers.
LG11
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Joined: Sat Nov 29, 2014 2:20 pm

Re: Breeding for Ethanol tolerance

Post by LG11 »

Yes I have access both at home and work to full analytical equipment including a GC with Mass spectrometer, sure I dont mind running some test for you.
I have results we have run many times for industry but to make it relevant to yourselves I am more than willing to run any tests you want. One small note at work we use Hydrogen as the carrier gas and at home I use Helium instead as the carrier gas, while the difference is minimal I should point that out.
Give me a full list of what you want and how long you want the exposure to ethanol etc to be.
InglisHill
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Re: Breeding for Ethanol tolerance

Post by InglisHill »

Awesome, absolutely awesome! I do not think I would be the only one interested in this. I shall get some groundwork done for you :-)
yakattack
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Re: Breeding for Ethanol tolerance

Post by yakattack »

You sir are a god send. We can now get some hard facts to what we have come to understand, and from a safety stand point this is very important.

Im not sure where you are located but being from Canada if there is anything I can do to aide you from here don't hesitate to ask. Same will probably be said for all the regulars on the forum.

Yak
HDNB wrote: The trick here is to learn what leads to a stalled mash....and quit doing that.
InglisHill
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Re: Breeding for Ethanol tolerance

Post by InglisHill »

It is not so much just exposure to ethanol, but all the various fractions that would come across in a typical run made by the hobby distiller. There is a real lack of information about this available.

If you already have results of the aforementioned substances and EPDM as well, that have been subjected to a full still run condition, well that would be a great starting point.

As to methods I think we can knock something together to test. Would you use glass to run the samples? Or would you use a still that is already setup? Another one that would be well worth exploring is having copper in the downward path, as there has been some conjecture about this, and, all these other things with no emperical evidence to back it up.

As I say I will piut something together over the next few days and we can vet it to see if the membership agrees that it would be sound and go from there.

Awesome!
LG11
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Re: Breeding for Ethanol tolerance

Post by LG11 »

Ahh, you want to see what else is in your brew!! I do that kind of thing 100 times a day! I can make very very small washes up if you like (I cant share commercial data) and test those. I can also post papers that give you a rough idea whats in there, I dont mind doing this at all but keep in mind that each batch is different and be prepared for a shock! Even the cleanest tasting best distilled will have azeotropes in including methanol. The simple fact is commercial stills are that size (height ) for a reason, it takes alot of energy (chemical bond ) to break apart azeotropes and that requires large surface area stills and very good design. The shape of commercial scotch whiskey stills is so that 92% of vapor gets refluxed and returned to the pot, so dont be surprised at the results and dont think its a fail just because a recipe gives more side alcohols than you thought.
I dont drink but I do get to taste, and the cleanest batches arnt always the best. The trick to it is having a flavor profile in your head first, then try and create the flavor by manipulating the wash etc. Depending how its done isoamyl alcohol can taste like banana, but you wouldnt want it in large amounts ;).
Its funny that most people shudder at the mention of methanol, yet fail to realize that a bad hangover is methanol poisoning! Ethanol is reduced in the liver to methanol, and high levels make you feel dreadful the headache comes from dehydration.
The only reason I dont drink is because I am intolerant to methanol abd really suffer even with small amounts of booze
butterpants
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Re: Breeding for Ethanol tolerance

Post by butterpants »

Was any data ever published from this inquiry? Just wondering. Interesting as hell
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