VeritasC&E
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How to chemically digest meat into AAs?
How would one go about chemically digesting meat into individual amino acids in the lab? Can it be done without resorting to complex proteases?
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zed
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Dunno. I've got a procedure for Casein.
http://www.orgsyn.org/demo.aspx?prep=CV2P0612
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unionised
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For most of the amino acids, boiling the protein with 20% HCl under reflux will do the job.
Separating them is a amore complex problem.
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Fyndium
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So what we have here is a 20% beef jerky hydrochloride. Reflux for 4 hours.
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VeritasC&E
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Thanks a lot for your contributions!
The HCl route sounds more interesting than resorting to the likes of HF.
@Unionised: When you write "for most amino acids" would you perhaps know to list amino acids this certainly excludes? Or list amino avids this
certainly includes?
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Fulmen
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See if you can get pepsin, it's one of the primary digestive enzymes.
We're not banging rocks together here. We know how to put a man back together.
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karlos³
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Quote: Originally posted by VeritasC&E  |
The HCl route sounds more interesting than resorting to the likes of HF.
@Unionised: When you write "for most amino acids" would you perhaps know to list amino acids this certainly excludes? Or list amino avids this
certainly includes? |
Well, I searched quickly, and that makes sense to me, that tryptophan will not stand these conditions, thats why a few percent of thioglycolic acid
are added to protect that.
Threonine and serine also can partially decompose during acidic hydrolysis.
Have a look here: https://www.researchgate.net/post/Can_anyone_recommend_a_sim...
Someone else there also recommend pepsin, and it is very cost effective apparently(20mg per gram of protein only), and its pretty mild and quick.
Basic hydrolysis is also possible, even in the microwave, and then it seems to be quite fast as well.
The link gives more thorough informations, I just summarised what I found there.
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unionised
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| Quote: Originally posted by karlos³ | | Quote: Originally posted by VeritasC&E |
The HCl route sounds more interesting than resorting to the likes of HF.
@Unionised: When you write "for most amino acids" would you perhaps know to list amino acids this certainly excludes? Or list amino avids this
certainly includes? |
Well, I searched quickly, and that makes sense to me, that tryptophan will not stand these conditions, thats why a few percent of thioglycolic acid
are added to protect that.
Threonine and serine also can partially decompose during acidic hydrolysis.
Have a look here: https://www.researchgate.net/post/Can_anyone_recommend_a_sim...
Someone else there also recommend pepsin, and it is very cost effective apparently(20mg per gram of protein only), and its pretty mild and quick.
Basic hydrolysis is also possible, even in the microwave, and then it seems to be quite fast as well.
The link gives more thorough informations, I just summarised what I found there. |
... what he said. :-)
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Dr.Bob
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I have cleaved peptides with 2N HCl at 60C for several hours, there are many variations of that routine, some used sealed tubes to keep in the HCl,
others just use a huge amount of acid. using enzymes first will speed things up, but will add some extra amino acdis, as they can also degrade to
amino acids. There are a few ways to then quantify the amino acids, but none are perfect. There are whole bookjs on this topic.
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UC235
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Enzymes are my day job. I remember seeing them in procedures like this and thinking they're some sort of voodoo, but from the other side, they're
pretty straightforward.
It's also pretty clear that at the time (1930), the enzymes involved were not well understood. Pancreatin has the benefit of containing both trypsin
and chymotrypsin. Trypsin is restricted to cleavage at arginine and lysine bonds. Chymotrypsin cleaves at hydrophobic amino acids tyrosine,
tryptophan, and phenylalanine and to a lesser extent, leucine and methionine. The chymotrypsin is really the important part here, but the trypsin and
probable presence of carboxypeptidases as well gives a well-rounded digestion of the protein in question. Pancreatin is going to have the most enzyme
activity around pH 8. Sodium carbonate is going to be closer to 10. Bicarbonate is probably the way to go (and is the buffer present in the small
intestine when pancreatic enzymes are doing their thing). Calcium also improves enzymatic stability for trypsin at least and will reduce the tendency
to digest themselves.
Pepsin requires acidic pH for activity (it is present and active in stomach acid) and has overlap with chymotrypsin in specificity though it is more
broad in action. pH is important. If you exceed pH 8, it is irreversibly denatured.
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Tsjerk
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I would start with something like caseïne instead of straight up meat, as that contains many many other things that will probably produce a big mess.
What about proteinase K? It is widely used so you might be able to find it relatively cheap and it cuts about anything. You can speed up the reaction with
temperature. You could do a basic or acidic hydrolysis on the peptides afterwards.
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Amos
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Every grocery store should contain some kind of powdered meat tenderizer, which will contain natural enzymes that break down protein. You could also
obtain papain from green papaya juice/latex. These will help in addition to providing the proper pH.
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karlos³
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Papain, and also bromelain from pineapples, can both easily purchased online too, usually in the form of capsules.
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