IBU Assay: where to get chemicals?

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I need smallish amounts of iso-octane (trimethylpentane) and HCl for measuring bitterness units.

Where to buy inexpensive chemicals online? Thanks.

( I can't help but wonder if I could use a different common solvent than the iso-octane, like something in my garage).
 
I need smallish amounts of iso-octane (trimethylpentane) and HCl for measuring bitterness units.

Where to buy inexpensive chemicals online? Thanks.
Which procedure are you using? Absorbance measurement? You're best off using spectral-grade isooctane, and that's not cheap. You can order from places like Sigma Aldrich or Alfa Aesar


( I can't help but wonder if I could use a different common solvent than the iso-octane, like something in my garage).

I know you're mostly kidding, but no. You'd have to recalibrate the scale to the new solvent, which would have a different solubility for the iso-alphas that you want to measure.
 
There's another procedure that I've seen around that is far more bootstrap, but certainly good enough for homebrewing use.

Take 1oz of unknown hops, and boil for a specified length of time in a quart of water. Cool, strain, and successively dilute the solution by half until you can't taste bitterness. That puts you at ~5 IBUs, and you can work backwards to figure out the AA% of the hops.

You'd primarily use it for homegrown hops, but I imagine you could try it for a beer too.
 
Thanks for all the info dwarven. I'm planning on measuring absorbance @ 275nm per this ASBC method.

Argh, $50 for the iso-octane. Better than what I found previously, but still more than I wanted to spend on a whim.

I read that the anti-knock additives for fuel are mostly iso-octane, which was my point about finding something in my garage. Since absorbance is a ratio, I thought any minor differences between said garage chemical and the "spectral-quality" stuff would cancel out. Thoughts?
 
A small impurity (as %) could really screw up your assay if it has a high extinction coefficient in the wavelength range of interest, in your case 275nm.

I couldn't follow the link to your method, but it might be feasible to zero/blank out the background from your garage solvent, but if this is a high proportion of your total signal that makes it tough. Is this solvent just for measuring or is it also needed for extraction?
 
A small impurity (as %) could really screw up your assay if it has a high extinction coefficient in the wavelength range of interest, in your case 275nm.

That's exactly the problem. (that, and upsetting the Abs --> IBU equation that I mentioned above). For example, alcohols absorb very strongly in the UV range, and a mix of alcohols can get pretty wild. Iso-octane is used because its absorbance is known, and a small amount of additives can play havoc with your measurement. I think another additive in those anti-knock formulas is methanol, which will certainly screw up UV absorbance.


Is this solvent just for measuring or is it also needed for extraction?
It's used for extraction, but I imagine that's just for simplicity's sake, so that the whole procedure is done in one solvent.
 
A small impurity (as %) could really screw up your assay if it has a high extinction coefficient in the wavelength range of interest, in your case 275nm.

I couldn't follow the link to your method, but it might be feasible to zero/blank out the background from your garage solvent, but if this is a high proportion of your total signal that makes it tough. Is this solvent just for measuring or is it also needed for extraction?

My understanding (from the docs I linked to above) is that the isomerized hop oils are made "more hydrophobic" by lowering the pH of the beer (with HCl). When the non-polar solvent is added, I guess they are then they are extracted from the beer and dissolve into the solvent. The organic phase of this mixture is then measured for UV absorbance.

I'm not a chemist, but if I can get the chemicals I think I can take a stab at it. I'd then like to test some other solvents and acids to see if there is a simpler way to do this with household chemicals (i.e, mineral spirits and vinegar). Just for fun.
 
( I can't help but wonder if I could use a different common solvent than the iso-octane, like something in my garage).

There is indeed something you can use in your garage - gasoline. If you read the MOA carefully you will note that it says that triple distilled gasoline will do. I will happily pay the freight for spectrographic grade 2,2,4 trimethyl pentane before I will distill any gasoline in my lab and would advise you to do the same.

You are correct in your assumption that the HCl is there to lower the pH thus enhancing extraction into the gasoline phase.

Note that you will also need some octanol (or hexanol). This is only used to keep the beer from foaming when it is picked up in the pipet (the tip is dipped into octanol and then the bit remaining in it shaken off before dunking it into the beer). Accordingly, the blank also has a wee bit of octanol in it.

I suppose you could take plain old gasoline, add a bit of octanol and measure the absorbance against distilled water. If it's not too great you could use that as a reagent blank and get readings but I wouldn't be too comfortable doing that.

Also note that you sometimes need a centrifuge as the protein (I assume) in the beer can form a sort of slush which you need to get out of the way to pipet out the solvent phase.

WRT other solvents - when you ask for spectrographic grade you pay a high price for that. Spectrographic grade methanol (used as a dilutant when determining hops alpha and beta acids) is expensive as is the toluene which is used for the extraction.
 
I assume you are using a spectrometer to measure this stuff...where did you get one that you can just do this stuff at home?

I traded a function generator for it (I have a lot of idle / redundant equipment at home). I also have unlimited access to another spec (that I use for UV already) and a UV power meter and thermopile. I need to trade something for some chemicals and a little knowhow :eek:
 
The know-how is free. You can get it here. It would be best,however, if you got a copy of the MOA. ASBC now publishes an abbreviated set of MOA's for craft brewers at a price which is much more reasonable than the full set (which is only available on line at this point anyway). If you don't want to buy those or can't find them I can go through the procedure in detail here. It's not really complicated at all for someone familiar with how to use a spectrophotometer.

There is one clunker in there though. After putting beer, acid and iso-octane into a 50 mL centrifuge tube it has to be shaken vigorously for 15 minutes. This can be done by hand (I even met a guy who does this for a living shaking by hand) but a mechanical shaker (it's obvious from the MOA that ASBC had Burell in mind) makes life much easier if you intend to do many anyalyses.
 
The know-how is free. You can get it here. It would be best,however, if you got a copy of the MOA. ASBC now publishes an abbreviated set of MOA's for craft brewers at a price which is much more reasonable than the full set (which is only available on line at this point anyway). If you don't want to buy those or can't find them I can go through the procedure in detail here. It's not really complicated at all for someone familiar with how to use a spectrophotometer.

There is one clunker in there though. After putting beer, acid and iso-octane into a 50 mL centrifuge tube it has to be shaken vigorously for 15 minutes. This can be done by hand (I even met a guy who does this for a living shaking by hand) but a mechanical shaker (it's obvious from the MOA that ASBC had Burell in mind) makes life much easier if you intend to do many anyalyses.

Lab Methods for Craft Brewers is $140 on Amazon. If I intended on taking this much further than a weekend experiment, I'd buy that. If I decide to go further and do a lot more analysis, it's a no brainer.

But right now I'm just looking for some fun between brew days. So, yes AJ, if you wouldn't mind detailing this one test, I'd appreciate it. The links I posted above have some broad strokes; I suspect you have further detail for a non-chemist like me. Thanks. :mug:
 
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Didn't think the book was that expensive!

What you need:

1. Spectrophotometer with UV capability
2. Quartz cuvets (expensive)*
3. 10 mL disposable pipets
4. 50 mL centrifuge tubes
5. Spectrographic grade 2,2,4 trimethyl pentane
6. 3 N hydrochloric acid
7. Octanol
8. Wrist action shaker (Burell) or wrists with good action
9. Centrifuge.
10. Pipetter (bulb) for 10 mL pipets
11. Smaller pipetter and tips

What you do:

1. Place a sample of cold, carbonated beer in a small beaker or glass
2. Dip the tip of a 10 mL pipet into octanol. Withdraw it and shake the alcohol off the end. A wee bit will remain and that's what you want
3. Lower the pipet into the beer and draw up 10 mL. The octanol will keep it from foaming so you can read the level accurately
4. Pipet into a 50 mL centrifuge tube
5. Pipet in 1 mL 3 N hydrochloric acid
6. Pipet in 20 mL 2,2,4 trimethyl pentane (use a new 10 mL pipet twice or if you have a 20 mL pipet use that).
7. Cap tube and shake vigorously for 15 minutes preferrably on a shaker
8. While extraction is taking place (do this and the next step before everything else if you are shaking manually) dip a toothpick or micropipet tip into octanol, shake off and touch to the inside of your blank cuvet.
9. Pipet 2 or so mL 2,2,4 trimethyl pentane into blank cuvet. Invert to mix.
10. At the conclusion of extraction remove tube from shaker and examine. There will be 1, 2 or 3 phases. One is the beer, the second, which may or may not be present, is a sort of slush. The third is clear solvent which should be on top and separate from the second. If it is skip to Step 12
11. Centrifuge at top speed** for three minutes to separate second and third phases.
12. Pipet 2 or so mL of the clear 3rd phase into the sample cuvet.
13. 0 photometer at 275 nm using the blank cuvet.
14. Read absorption at 275 nm for the sample cuvet.
15. Multiply absorption by 50 e.g if A275 = 0.500, the IBU = 25

*There are plastic disposable cuvets that claim decent transmission into the UV. Much less expensive, of course, but I've never tried them.
As the iso octane is quite volatile and as many of the less expensive instruments have lots of plastic parts (and even caveats against the use of volatile solvents in them) I use cuvets that have a screw on cap.

**It takes lots of g's to separate these two if they form together. A pretty hefty centrifuge is required to do the job properly but I've found a trick that lets me do it in even and old clinical centrifuge. After three minutes cetrifugation the 2nd phase will pack down somewhat. It's now like a wet-snow slush. If you step in that the water snow part collapses leaving the water. The same works here. Take a tip from the smaller pipetter and poke at the slush gently or smash it against the side of the tube wall. This will leave you with clear liquid which you can pipet out. In extreme cases you may have to go back into the centrifuge for another couple of minutes.
 
*There are plastic disposable cuvets that claim decent transmission into the UV. Much less expensive, of course, but I've never tried them.

I've tried them, and was not impressed. OTOH, I have access to those $200 quartz cuvettes so it doesn't matter how not impressed I am. Great writeup, btw.
 
Find the graduate student TAs that run the CHEM 101 labs at the local university and offer them some homebrew. I think that should get you all the chemicals you need.
 
Find the graduate student TAs that run the CHEM 101 labs at the local university and offer them some homebrew. I think that should get you all the chemicals you need.

Right. I've got a 20-yr old boy taking chem in college starting next week. He has his instructions. The plan is coming together. :D

I'll buy the cuvetts and other small stuff from cynmar.com.
 
I've tried them, and was not impressed. OTOH, I have access to those $200 quartz cuvettes so it doesn't matter how not impressed I am. Great writeup, btw.

I guess I should have said I haven't tried them at UV or in particular for this MOA. I use them all the time for color determination and they are fine for that. The concerns with this MOA would be differences in absorption between the blank and sample cuvets and the fact that these are open to the air but I suppose one could put parafilm over the tops. For color determination one eliminates the differential problem by using the same cuvet for blank (DI water) and sample.
 
Right. I've got a 20-yr old boy taking chem in college starting next week. He has his instructions. The plan is coming together. :D

I'll be interested to hear about how that works out. Regulations (anti terror, environmental, drug precursor...) have beer proposed and in some cases implemented that impose accounting requirements on users of chemicals that would practically prevent a university from teaching lab courses (I would think). I know this is just gasoline (very expensive gasoline but still gasoline) but the world is now just a crazy place.
 
Due to the anti-terror environmental and blah blah blah regulations most of my chem labs are built round household consumer products for cleaning and cooking. They just happen to use all the fancy glass and electronic whizbang gizmos.

I was actually looking at doing this testing for a local brewery as a school project. It was tough to get anyone to agree to purchasing the necessary reagents for the reaction. The brewery isn't tiny but they're not big and $300 in alcohols was not entirely interesting to them. The school wanted to allow me to have the experience but didn't want to buy the $300 in alcohols either. I'd still like to get some experience doing the testing I guess I'll just have to wait.

I want to come hang out at your house AJ. I'd love to geek it up beer science wise.
 
I guess I should have said I haven't tried them at UV or in particular for this MOA. I use them all the time for color determination and they are fine for that.

Yeah, same. I use the plastic ones all the time for photocatalysis studies, but whenever I go into UV I use the quartz.
 
Something funny there. Most beers have pH's in the 4's. But that's a separate question. There is no 'target pH' for the test. It just needs to be low enough that none of the bittering acid molecules are dissociated i.e. they are not charged and therefore more soluble in the gasoline phase than in the water (beer) phase. One cc of 3 N acid would deliver (3/1000) moles to 11/1000 liter of non gasoline phase i.e. 3/11 moles/L which is enough to set the pH to something like 0.6. That's obviously plenty low enough.
 
Something funny there. Most beers have pH's in the 4's. But that's a separate question. There is no 'target pH' for the test. It just needs to be low enough that none of the bittering acid molecules are dissociated i.e. they are not charged and therefore more soluble in the gasoline phase than in the water (beer) phase. One cc of 3 N acid would deliver (3/1000) moles to 11/1000 liter of non gasoline phase i.e. 3/11 moles/L which is enough to set the pH to something like 0.6. That's obviously plenty low enough.

ajdelange,
Thanks for your reply. I have muriatic acid (31.45 HCl) which should be about 10.2 N. I can go from there with your answer.

I measured the pH of the beers right out of the tap without degassing them. I'll pull another sample of one and let it go flat and then measure the pH again. Don't know if that is the reason that my pH doesn't match your experience (i.e., ~4 pH).

Paul
 
Carbonic acid lowers the pH so a degassed sample will read slightly higher than one that still has CO2 dissolved.

Suggest you review https://www.homebrewtalk.com/f128/ph-meter-calibration-302256/ to make sure there isn't something you are overlooking there. Your mash should read 5.4 or so and the pH should be a bit lower than that in each successive step of the brewing process finishing well under 5 (how much depending on the beer).
 
For reference, I've used the disposable plastic (both PS and PMMA) in my QC chem lab for all sorts of routine analyses without issue, so long as the detection wavelength is in the Vis range - turbidity, color matching, etc. For true UV applications (anything below 300 nm) the quartz cuvettes are unfortunately a necessity. I don't have the comparative absorbances in front of me, but the PMMA will pretty much dampen any uv light to useless levels below 300 nm. Pity, because they're so cheap.
 
Carbonic acid lowers the pH so a degassed sample will read slightly higher than one that still has CO2 dissolved.

Suggest you review https://www.homebrewtalk.com/f128/ph-meter-calibration-302256/ to make sure there isn't something you are overlooking there. Your mash should read 5.4 or so and the pH should be a bit lower than that in each successive step of the brewing process finishing well under 5 (how much depending on the beer).

I let a small sample of beer sit overnight to go flat. The next morning it had dropped from 6.0 to 5.5 pH.

I checked the calibration of my pH meter when I first got it a few weeks ago. It read the 4.0 buffer as 4.0.; the 7.0 as 7.0; and the 10.0 buffer as 10.1. Close enough.
 
I let a small sample of beer sit overnight to go flat. The next morning it had dropped from 6.0 to 5.5 pH.

I checked the calibration of my pH meter when I first got it a few weeks ago. It read the 4.0 buffer as 4.0.; the 7.0 as 7.0; and the 10.0 buffer as 10.1. Close enough.

Maybe you breathed on it a lot.

The guy at my fish store says the pH of his tank drops when he has a lot of people in his house, or after a party. He says the CO2 from respiration of his guests affects the pH of the tank. Hence, he now has the fans over his tank pulling air from outdoors.

It's all very odd since we keep the alkalinity of our tanks relatively high to prevent pH swings.
 
There are other factors involved that can royally mess up IBU testing. I won't get into it right now, but I will in a few weeks.
Btw, getting the iso octane is extremely hard unless you are a business with an industrial address (homeland defense, and all...).
 
There are other factors involved that can royally mess up IBU testing. I won't get into it right now, but I will in a few weeks.
Btw, getting the iso octane is extremely hard unless you are a business with an industrial address (homeland defense, and all...).

Yes, I know, I've long since abandoned the testing idea.
 
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