Gluten Free Brewing With Enzymes

[coeliacsurvivor]

Hi guys,

I have been following Thirstyboys threads on experimental brewing with gluten free grains and enzymes, and I am ready to give it a go.

I have gotten my hands on some enzymes, pullulanase, beta-glucanase, alpha-amylase, protease, and gluco-amylase(amyloglucosidase).

However, I don't have a lot of these as they are trial packs sent out from a company in Auckland, so I want to make sure I have the process right before I start.

I am aiming for 5L batches to start with, so my recipe and protocol is as follows, modified from Andrew (Millet man)

1/2 kg millet grain
1/2 kg sorghum syrup

Crush grain and mix with 1.0 lt water and add pullulanase, beta-glucanase, alpha-amylase, and protease.
30 min at 55C
30 min at 85C

Add cold water to reduce temperature and add gluco amylase
120 min at 65C

Then lauter and sparge, and continue as normal adding the 1/2kg of sorghum syrup to the kettle.

My problem is figuring out how much of each enzyme to add. I have calculated from the manufacturer and the internet that 1KNU(T) is the amount of alpha amylase which at pH7.1 37C dextrinises 5.26g starch dry substance per hour. So I will need 115 KNU(T)'s per kilo of grain. ~1g of my alpha amylase per kilo of grain. Will this work better or faster at a higher temp and lower pH?

But what about the others? Is there a way of calculating this or do I just play around and see what happens?

Is it ok to brew a 5L batch in a standard 30L fermenter or is there too much headspace?

Any ideas, tips, or tricks are much appreciated.

Colm

 

[Thirsty Boy]

Hi guys,

I have been following Thirstyboys threads on experimental brewing with gluten free grains and enzymes, and I am ready to give it a go.

I have gotten my hands on some enzymes, pullulanase, beta-glucanase, alpha-amylase, protease, and gluco-amylase(amyloglucosidase).

However, I don't have a lot of these as they are trial packs sent out from a company in Auckland, so I want to make sure I have the process right before I start.

I am aiming for 5L batches to start with, so my recipe and protocol is as follows, modified from Andrew (Millet man)

1/2 kg millet grain
1/2 kg sorghum syrup

Crush grain and mix with 1.0 lt water and add pullulanase, beta-glucanase, alpha-amylase, and protease.
30 min at 55C
30 min at 85C

Add cold water to reduce temperature and add gluco amylase
120 min at 65C

Then lauter and sparge, and continue as normal adding the 1/2kg of sorghum syrup to the kettle.

My problem is figuring out how much of each enzyme to add. I have calculated from the manufacturer and the internet that 1KNU(T) is the amount of alpha amylase which at pH7.1 37C dextrinises 5.26g starch dry substance per hour. So I will need 115 KNU(T)'s per kilo of grain. ~1g of my alpha amylase per kilo of grain. Will this work better or faster at a higher temp and lower pH?

But what about the others? Is there a way of calculating this or do I just play around and see what happens?

Is it ok to brew a 5L batch in a standard 30L fermenter or is there too much headspace?

Any ideas, tips, or tricks are much appreciated.

Colm

I think that you are likely to get a better, more definative answer from Millet Man.. but.

I suggest you will need less alpha amylase than that if you are using all those enzymes. The alpha knocks your amylase and amylopectin into smaller chunks, basically providing more chain ends for you sachrifying enzymes to work on. This would in a malt mash, normally be beta-amylase and be citing off maltose chunks.. but in your mash its going to predominantly be the amyloglucosidase biting off single glucose units. The pullulanase will also knock off some fermentables.

In your mash - the alpha amylase (is it heat stable bacterial AA or malt derived??) is going to be supplemented by the de-branching that the pullulanase (limit dextrinase) does. So I think you could get away with a lower dose.

BUT - your mash regime has nothing for your alpha amylase if it is malt derived with a 72ish optimum activity temp. You are too low at the 55 rest for it to do very much, and straight into kill it off territory at 85C.

Also - I don't know about thestuff you have, but the amylogucosidase and pullulanase I have been using is heat stable with an optimum temp of 80+ degrees, so cooling your mash down would not do it any good and would actually retard it.

So - lacking the specific optimum and denature temps for the enzymes you have... if I were using my enzymes I would.

Mash in at 43C (beta glucanase) with a bit of all of your enzymes - immediately start raising the temp, rising through the protease activity range, till you get to 70 or 72C - rest a bit to give your Alpha amylase a chance to play (the AMG and pullulanase will be working by now too) slowly raise it to 85 and rest there to finish off conversion.

OR - just use the beta glucanase, the pullulanase and the AMG - mash in at 82-85 and rest there. Fair chance that might work.

I have also heard that sorghum brewers in Africa are having luck producing beers using only pullulanase - it seems to give a decent balance of fermentable vs non when used in isolation - but you would want to do a bit of googling before you took my word on that.Or try it out if you are feeling adventurous. Thats what I will be doing with my next enzyme brew. Pullulanase only. I let you know how it goes.

Wait for Millet Man though... I am well and truly pulling stuff out of my bum on this topic.

TB

 

[coeliacsurvivor]

Thanks TB,

I haven't actually got the enzymes yet, and I am hoping there will be some info with the packaging.

From what I can find online they are all produced with either bacteria or fungi and are 'extremely' heat stable. I think your idea of mashing in at 43 and then slowly raising to a 72 rest and then up to 85 would work.

What do you think about doing 5L brews in a 30L fermenter? I have 3 of these 30L barrels so can have 3 experiments at once, or I could go out and buy some smaller barrels.

Cheers

Colm

 

[raven19]

What do you think about doing 5L brews in a 30L fermenter? I have 3 of these 30L barrels so can have 3 experiments at once, or I could go out and buy some smaller barrels.

If you are pitching with an active yeast starter, extra headspace wont hurt.

With proper santiation, I dont see an issue regardless actually.

No point in buying more fermentors imo, buy more enzymes!

Good luck with it. :icon_cheers:

 

[Thirsty Boy]

You'll most likely be using dried yeast (GF) and a starter will be a bit pointless (not to mention hard to do GF) - but I still doubt you will have an issue. O2 isn't an issue until after fermentation starts, and after that its all being driven off by the C02 produced. Just consider them to be "open" fermenters that happen to have a lid on them and your mind will rest easy.

Let us know the details of those enzymes when they arrive

Cheers

TB

 

[coeliacsurvivor]

The enzymes have arrived, but there is no additional info with them regarding the optimal temps etc.

So I guess it's time for a little experimentation, which is where the fun really lies anyway.

Here is what I plan on doing (thanks to Thirsty Boy)

1/2 kg millet grain (not sure on roasting plan yet but will be ~ 30min at 150C and 30min at 200C)
1/2 kg sorghum syrup

Crush grain and mix with 1.0 lt water, mash in at 43C (beta glucanase activity) and add 1mL each of pullulanase, beta-glucanase, alpha-amylase, gluco-amylase (AMG), and protease.
Immediately start raising the temp, rising through the protease activity range, till I get to 70 or 72C.
30min rest to give Alpha amylase a chance to play (the AMG and pullulanase will be working by now too) slowly raise it to 85C and 30min rest to finish off conversion.

I will probably do an iodine test to check for conversion, as this will also give me an idea of the amount of enyme I need to add.

Then lauter and sparge to 5L, and continue as normal adding the 1/2kg of sorghum syrup to the kettle.

I am hoping that by using whole grain I won't have a problem with stuck sparge like TB did with the flour, but just in case I will probably do this through a rough gauze first then something more like t-shirt material.

I still have some fuggles left over from a brown ale I did so I will probably use that with an ale yeast.

This is in the pipeline for tomorrow night all going well.

 

[Thirsty Boy]

Only a couple of things really - I'd mash with 1.5-2.0L of water - 1L will give you only 2L/kg and thats ate the very low end, in my experience the millet absorbs a lot of water and I'd even say that 3:1 (or 1.5L in your mash) might give you too little liquid. BUT - I have only done it a few times... so I'm far from sure.

And remember to stir during your ramps of you will scorch your mash.

Have fun, hope it works well for you - I know it will work, so from here on in its just a matter of experimenting till you make it go.

Cheers

TB

 

[Millet Man]

Colm,

Looks good and the only things I would change would be as follows:

• Use 2 lt of mash water as TB suggested, with my mashes I would mash in at least 2.3:1 but with infusions would end up with 4:1 for the conversion rest.
• Millet should gelatinise in the 75-80C range, you can see the colour and consistentcy change when it does, so 85C would be your max temp.
• Don't put the AMG in with the other enzymes, after the 85C rest cool the mash to 60C and then put the AMG in for a 1-2 hr rest.
• The AMG will work better if the mash has already been converted to dextrins.
• Will such a small mash it will be difficult to get a clear runoff from my experience so you may just have to accept that the wort will be a bit cloudy.

Have fun!

Cheers, Andrew.

 

[coeliacsurvivor]

Looking forward to it already.

Thanks for your advice TB and Andrew.

One question, if I don't have time to do the boil straight after the mash, can I store the lautered mix overnight? should this be in the fridge?

 

[Thirsty Boy]

Colm,

Looks good and the only things I would change would be as follows:

• Use 2 lt of mash water as TB suggested, with my mashes I would mash in at least 2.3:1 but with infusions would end up with 4:1 for the conversion rest.
• Millet should gelatinise in the 75-80C range, you can see the colour and consistentcy change when it does, so 85C would be your max temp.
• Don't put the AMG in with the other enzymes, after the 85C rest cool the mash to 60C and then put the AMG in for a 1-2 hr rest.
• The AMG will work better if the mash has already been converted to dextrins.
• Will such a small mash it will be difficult to get a clear runoff from my experience so you may just have to accept that the wort will be a bit cloudy.

Have fun!

Cheers, Andrew.

How come the AMG at the end Andrew? the AMG should be heat stable to 85 and at its maximum activity at 80-85 ?? and if you have it in the whole mash wouldn't that mean that you needed a shorter rest at the end?

Or maybe it depends on which AMG you get - the stuff I have is AMG 300L from novozymes - or do I have it arse about? Maybe the stuff isn't heat stable at all? I'll try to find out why I think it is.

 

[coeliacsurvivor]

I haven't gotten around to doing this yet, the roasted grain is in paper bags at the moment.

TB I have been looking around for info on the optimal temp for the AMG 300L, and found a couple of sites,

http://www.biotimes.com/en/Articles/2005/J...ovesstarch.aspx

This one compares this enzyme with another 'better' one and states that it has a higher optimal temp than AMG 300L at 65-70C.

However this site
http://books.google.co.nz/books?id=KA1sLFi...ved=0CB4Q6AEwBA#

states that AMG is very heat stable and can survive pasteurisation.

I think I will follow Andrews advice and reduce the temp to 60C before I add the AMG, just to be on the safe side.

 

[Millet Man]

How come the AMG at the end Andrew? the AMG should be heat stable to 85 and at its maximum activity at 80-85 ?? and if you have it in the whole mash wouldn't that mean that you needed a shorter rest at the end?

Or maybe it depends on which AMG you get - the stuff I have is AMG 300L from novozymes - or do I have it arse about? Maybe the stuff isn't heat stable at all? I'll try to find out why I think it is.

TB,

The AMG I have had in the past (don't use it anymore) was optimum around 60C IIRC, the alpha amylase I used was optimum at 85C. This is why I would do protein rest, then raise to 85C to convert, and then cool down for the AMG. The boil then kills off all the enzymes.

If you leave the wort for a while before boiling the AMG will keep working and make the beer drier.

Cheers, Andrew.

 

[Thirsty Boy]

Yep - thats why I said you should wait for Andrew.

Me speaky through bottomy.

A bit of Googling and a chat with the development brewer at work - and it's (AMG 300L) optimum temp is indeed 60C - The Promozyme Pullulanase is the heat stable one as are the alpha amylases like Thermamyl.

That second article is about adding the stuff to the fermenter.. where the maximum temp it experiences will be much lower than in a GF mash. It isn't ever boiled and is stable enough to maybe retain a little activity through a pasturiser.

Listen to Andrew - listen to me less, I don't know what I'm talking about... no really, I don't!!

On the other hand.... Novozymes has a product that might be interesting to you... its called Ceremix Sorghum and is specifically designed for brewing beer from unmalted sorghum and other grains. Might be worth having a chat to them about that.

Thirsty

 

[coeliacsurvivor]

The brew is on!

I started trying to crush the grains with a rolling pin, but soon got tired of that, so I stuck them in a blender and pulsed them for about 30 sec.

I warmed the water up to 43C and added the grain and imL of all the enzymes except the AMG 300L.
the temp took about 30mis to rise to 70C and I let it rest there for about 30min. It was pretty hard keeping it at exactly 70-72 but I think I was close enough.
Then inc temp to 85C and rested for another 30min, again hard to keep constant but always 80-85C.

I added about 500mL of cold water to cool it to 60C and added 1mL of the AMG 300L, then let it rest at ~60 for about 2 hours.

After all that I strained it through a layer of muslin and got about 2.2L of muddy wort. see photo.



I then placed four layers of muslin on a kitchen sieve and passed all the wort through the grain again and more water up to 5L total.

photo of my lauter/sparge apparatus.



will post more as the brew progresses


Lautering is finished - took about an hour.

I took a gravity reading of the 5L and it was 1024. How do I calculate what I expected to get? Should I have run the wort through the grains again to increase the OG?

I have decided not to use my sorghum syrup as it might affect the taste, I would rather find out what the millet tastes like on its own. I have some liquid glucose that I will add instead and a small amount of candi suger SRM-80.

 

[Thirsty Boy]

what you should have got depends on the potential of your grain.

Normal malt has a potential of about 1.036-1.037 ppg (points per pound per gallon) and I have been working on millet having a potential of 1.036.

In your case that would mean that 5L of pre-boil wort @ 1.024 from 0.5kg of millet = a mash/lauter efficiency of 80%... which is really quite high. Well done.

Andrew will be able to tell you more accurately what potential you should actually expect from millet grains.. but I'll give you highs and lows.

If the potential is

1.034ppg - your efficiency was 83%
1.037ppg - your efficiency was 77%

either way... high! You must have gotten nice conversion and had an effective lautering process. Good work.

Adding 20% sugar will give you about a 1.040 post boil gravity with 4.25 of post boil volume & that wil give you about a 4% abv beer.

Cheers

TB

 

[SpillsMostOfIt]

Yep - thats why I said you should wait for Andrew.

Me speaky through bottomy.

<SNIP>

On the other hand, I tried a little of Thirsty Boy's GF brewing output last weekend and he is making very good beer. So, bottom-talking may be the answer...

 

[coeliacsurvivor]

On the other hand, I tried a little of Thirsty Boy's GF brewing output last weekend and he is making very good beer. So, bottom-talking may be the answer...

TB's bottomy talk is still far more intelligent than most peoples mouthy talk!

here is the final recipe for the brew

brought the 5L to a rolling boil;

7g Fuggles, pellet 4.5% (60 min)
65g Dark Candi Sugar (80 SRM) (25min)
265g glucose syrup (25min)
5g East Kent Goldings, pellet 4.8% (15 min)
1/4 tsp Irish Moss (10 min)
1/4 pkg SafAle English Ale (S-04) ~3g

cooled to 22C before pitching yeast.

I had about 3.5L postboil which I topped up to 5L with cold water in the fermenter. Should I do this? or is it better to have sparged with more water and let it boil off?

My OG was 1.047 in the fermenter which is higher than I had expected, I have read elsewhere that brewing with enzymes can cause the beer to ferment out more than normal, down to 1 in some cases, so this could end up ~6%

Smelled like wort and tasted like wort so here's hoping!

If TB's calculations are correct my efficiency was 77-83% I might try this again with less enzymes, try to find the minimum amount I have to use. I will also have a go with some other grains, buckwheat, amaranth, quinoa, and corn.

Thanks for all the advice guys.

 

[Thirsty Boy]

TB's bottomy talk is still far more intelligent than most peoples mouthy talk!

here is the final recipe for the brew

brought the 5L to a rolling boil;

7g Fuggles, pellet 4.5% (60 min)
65g Dark Candi Sugar (80 SRM) (25min)
265g glucose syrup (25min)
5g East Kent Goldings, pellet 4.8% (15 min)
1/4 tsp Irish Moss (10 min)
1/4 pkg SafAle English Ale (S-04) ~3g

cooled to 22C before pitching yeast.

I had about 3.5L postboil which I topped up to 5L with cold water in the fermenter. Should I do this? or is it better to have sparged with more water and let it boil off?

My OG was 1.047 in the fermenter which is higher than I had expected, I have read elsewhere that brewing with enzymes can cause the beer to ferment out more than normal, down to 1 in some cases, so this could end up ~6%

Smelled like wort and tasted like wort so here's hoping!

If TB's calculations are correct my efficiency was 77-83% I might try this again with less enzymes, try to find the minimum amount I have to use. I will also have a go with some other grains, buckwheat, amaranth, quinoa, and corn.

Thanks for all the advice guys.

Topping back up is fine - I usually do it during the boil - drip feeding it water out of the hot tap as it boils off. But thats just being a bit paranoid about sanitation.

You shouldn't get a too fermentable beer strictly from your enzymes - a lot of people who use them put them in the fermenter, where they keep on working basically forever, so they can dry a beer completely out. You are denaturing your enzymes in your boil, so you have control. If your beers are too dry, you just use a little less enzyme, or shorten your sachrification rest time. Visa versa for not dry enough.

Your beer in the other hand... probably will be pretty dry. You have use a lot of simple sugar as compared to grain. So, if you beer ferments like a normal beer - you could expect to get about 75% apparent attenuation out of the grain portion.. and basically 100% attenuation for the sugar portion.

So your beer had a gravity of 1.024 for 5L with only the grain. 75% AA with that would give you an FG of 1.06. Even with the sugar, your FG will still be around 1.06, maybe a little higher. Therefore your final abv will be around 5.2 - 5.3 ish percent.

If the grain based portion of your gravity was either very fermentable, or very unfermentable - that will throw the numbers a little in either direction.

I'd expect the beer to be quite light bodied and a bit thin for its strength. I'd be a little reluctant to use much more than 20% simple sugars for that reason.

But who knows?? this is a first go...so if its drinkable in any way you have done pretty damn well in my book.

TB

 

[Millet Man]

Millet is a bit lower in starch content at around 70% compared to barley which is closer to 80%. I have never had lab measurements done but doing small scale EBC standard test for extract (adjusted for decoction mash) I get a potential extract of 1.032. When I mash on a home brew scale using a fly sparging process I get close to 100% efficiency with that number (mash @ 4.0 lt/kg, sparge @4.0 lt/kg) , on a commercial scale with a mash filter press I get 90% efficiency (mash @ 5.7 lt/kg, sparge @ 0.5 lt/kg).

Glad to hear the brew has gone well so far.

Maybe we should start up a thread for recipies?

Cheers, Andrew.

what you should have got depends on the potential of your grain.

Normal malt has a potential of about 1.036-1.037 ppg (points per pound per gallon) and I have been working on millet having a potential of 1.036.

In your case that would mean that 5L of pre-boil wort @ 1.024 from 0.5kg of millet = a mash/lauter efficiency of 80%... which is really quite high. Well done.

Andrew will be able to tell you more accurately what potential you should actually expect from millet grains.. but I'll give you highs and lows.

If the potential is

1.034ppg - your efficiency was 83%
1.037ppg - your efficiency was 77%

either way... high! You must have gotten nice conversion and had an effective lautering process. Good work.

Adding 20% sugar will give you about a 1.040 post boil gravity with 4.25 of post boil volume & that wil give you about a 4% abv beer.

Cheers

TB

 

[coeliacsurvivor]

I have just repeated this experiment using 1/2 mL of each enzyme instead of 1mL and I got an OG of 1.021. While this is slightly less than the first batch I think there were some subtle differences in my process, I did not grind the grain as fine as I did the first time which resulted in a much faster sparge, and I was not quite so accurate with my water measurement so I think it was slightly more than 5L.

I am happy to conclude that 1/2mL does pretty much the same job as 1mL. I can't accurately measure less than 1/2mL so I think my next millet experiment will be using 1kg of grain and 1/2mL of enzymes.

I am going to get some buckwheat and try that next. I figure I should work out the efficiency of each grain on its own before I mix them, this should give me more accurate measurements to estimate my expected grravity in the future.