Advice on cheap tentative toxicity tests for diacetyl substitutes in butter flavorings

[Martin Baldan]

Hello,

Guys, first of all, please let me tell you what you are doing is really awesome. I haven't got my hands dirty with DIY biology yet, but the little I've learned is fascinating. Also, people tend to fear the unknown, so, by making all this knowledge available to everyone, you are paving the way for the wide (and responsible) adoption of these powerful technologies.

Now, my question. I'm starting a small business in the flavors industry and one thing I've noticed is that, apparently, a few assumptions about the safety of some food flavorings turned out to be wrong. The most dramatic example I've heard of is "popcorn lung", that is, bronchiolitis obliterans caused by hot butter flavoring vapors in butter-flavored popcorn factories. The industry then switched to diacetyl substitutes, but many of those, according to some tests, also turned out to be harmful. 

Now it seems clear that alpha-diketones are toxic for the airway epitelium cells, but I can't find information on what butter flavorings have been confirmed as nontoxic (for breathing). What I've read about acetoin (an alpha-hydroxy ketone) seems contradictory and not very conclusive. There seems to be a growing consensus that diacetyl toxicity is caused by reaction of the diketone group with the guanidine residue of the amino acid arginine. There's also talk about electronic affinity (EA) and other electrochemical properties of diacetyl and its derivatives, which may be involved in electron transfer (ET) phenomena, ultimately leading to pulmonar damage.

The issue seems very complex, but I decided to have a tentative look at the prospect of performing some kind of toxicity tests on candidate diacetyl substitutes, in order to pick the least problematic ones. First I found out about EpiAirway, a commercial human airway cell culture line by the company Mattek, which has been used to confirm diacetyl as toxic for the lungs. I contacted them asking for more information. 

In short, they said: " if we were to test 2 substitutes alongside of diacetyl at only 1 concentration each with endpoints of LDH assay and histology the price would be around $6,000-$7,000."

That's currently beyond my means as a very small startup. But even if I managed to raise the funds, I would have to pick my candidate substitutes very carefully!

So, I was thinking of ways to pre-select candidate substances, and this is one area where I'm hoping to get some useful pointers from DIY Biology experts. I would need to evaluate the reactivity of different "buttery" compounds (usually linear esters, lactones and fatty acids) with arginine, guanidine and/or some other molecule that reacts with diacetyl in the same way. Well, I guess that's chemistry rather than biology, but I'm sure you know some tricks to make it simpler and cheaper. Then I'd like to test the best candidates with, say, brine shrimps , C. elegans or other small invertebrates. Eventually, if that's not too hard or expensive, I'd like to test them on DIY cell cultures. At the end of this process, I would have one or two candidates for the really expensive tests.

Notice that I'm not talking about testing my own secret formulas, but standard, currently available aroma chemicals with a formula and a Flavis (and possibly FEMA) number. The results would be available to everyone, and that's why I hope others might be interested in helping me with this effort. 

Also, I'm not at all in a hurry. I'm just starting and I'm not in a position to spend much money right now, but I thought that it would be best to start learning about this challenge as eearly as possible.

Thanks in advance and I hope to hear from you guys soon!

Best,

Martin Baldan

[Mega [Andreas Stuermer]]

Hi Martin!

Sounds great, but as for me I haven't worked on anything related other than cytotox testing with hunan cell lines.

Drosophila and brine shrimps seem like good candidates, though AFAIK drosophila has no lung but trachaes. Does that make a difference?