Ricin Extracton

[Xanax]

I´ve got a lot of castor beans, which i have removed the hull and picket out the seed. Then smashed the seed and extract it with acetone, three times. It still remaining about 1,5 grams of powder, and I don´t think all of this is pure ricin. It´s also smells acetone and seems to be kind of oily.

So, what to do? Pure it with acetone a few times more?


Another question is; I also have revomeved the hulls from a lot of jequirity beans, which contain abrin, a ricin-like substanse. But is there any method to extraxt the abrin from this beans?

[Sarevok]

Read US3060165 if you want to extract ricin from castor beans.

You need water to extract it; acetone doesn't work because ricin is not soluble in acetone. Acetone only works to remove some of the oil. You also need sodium sulfate to precipitate the ricin. You need to keep the pH within a certain range during the extraction.

Just read the patent.

[mrcfitzgerald]

I am not sure patent 3060165 is as valid as many people assume, it appears that the extraction denatures much of the desired Ricin. (At least the writers of National Security Notes believe so...) More info can be found at: http://www.globalsecurity.org/org/nsn/nsn-040723.htm
However, I am not knowledgeable enough to know whether or not National Security Notes' assertion it true. Can anyone shed light on this mater?

[Sarevok]

What I know is that ricin is soluble in water and not soluble in acetone. Therefore, you need water to extract it from the castor beans, not acetone. Acetone can be used to de-oil the pulp, of course.

I will read this page now, mrcfitzgerald, but I do believe that both the patent and the NSN are probably saying half truths and half lies. :(

EDIT: The text is attacking the "cookbook recipes" for ricin (we already knew they were full of shit, anyways), not exactly the patent. They don't really say "the patent is wrong, they lie". They merely say it would perhaps not yield a pure product.

The patent may not be perfect, but it is still the best source on ricin we have.

EDIT2: Ok, let's suppose the procedure denatures the protein. Those who are concerned with ricin (I'm not) must then read books on proteins and biochemistry and then experiment and develop their own procedure.

[mrcfitzgerald]

The specific area of my concern is where the author of the text states: "Without going into even greater scientific detail, it is clear from a straight reading that the patent's intent was to produce a product for use in a biological weapon. It is equally obvious to the expert that as published it contains fundamental errors in the application of biochemical methods of protein purification. In fact, some of the methodology would actually denature an active protein rather than preserve it, a piece of information that is not at all to be seen in layman literature commenting upon it." -Now, certainly the assertion isnt that the patent "lies" but in that it does not produce a pure product. Going into further detail, the text tells us that the patented method yeilds a variety of assorted proteins -certainly more concentrated in ricin than a mashed pulp, but still somewhat denatured and not pure. The real question is, for those more acquainted with bio-chemistry than I, by what means would one achieve purity of the product?

[edit]- I didnt see Sarevok's last edit as I was writting this one, sorry :(

[Sarevok]

Certain conditions such as temperature and pH can denature a protein. Denatured proteins are proteins that have lost many of their most stable interactions, rendering them inactive or dysfunctional. Since the body acts to maintain a temperature of 37 degrees Celsius and a pH of 7 throughout its tissues, enzymes will function more efficiently in these conditions. If these conditions are disrupted, proteins will begin to denature Source (http://www.google.com.br/search?q=cache:oxGMYxm9BMMJ:www.sparknotes.com/nutrition/aminoacids/section1.html+denature+protein+pH&hl=pt-BR)

The patent says a pH of 3 or something should be used. Perhaps this denatures the protein. A pH of 7 would be more suitable, I think, if proteins are similar (i.e. if ricin requires the same pH range the proteins in our body require to survive). There are also other variables (chemicals used, temperature, etc) that may denature a protein. Like you, I don't know much of biochemistry.

Read this (http://www.google.com.br/search?q=cache:CBSlsSj1jQQJ:www.courses.psu.edu/fd_sc/fd_sc400_jnc3/proteins/denat.htm+denature+protein+pH&hl=pt-BR) too.

[Bugger]

Castor seeds contain, as well as ricin (the main protein/polypeptide, which when pure is a white powder), fatty acid triglycerides (lipids), the chief of which (85%) is that of ricinoleic acid, which is an unsaturated alpha-hydroxy acid, C17H32(OH)COOH, somewhat similar to oleic acid (8% of castor oil). Organic solvents such as acetone would dissolve the latter, but the ricin would be practically insoluble in all solvents because of its molecular size and internal bonding.

Because ricin is a liver poison, through containing a polypeptide sequence which cannot be metabolized by humans (toadstool and puffer-fish poisons work in the same way), it is most unlikely that "denaturation", e.g. by heating (as in the usual cooking of protein foods), which involves the deactivation of enzyme and hormone receptor sites in the protein molecule, would significantly reduce its toxicity.

I think that castor seeds also contain small amounts of other substances, e.g. water soluble starches, and, on the seed casing, waxes (long-chain alcohols).

Bugger.

[nbk2000]

Extremes of pH, temperature, and mechanical stresses would denature the protein.

Thus, you'd want to use moderation in all these things to minimize the losses. The acid/alkali precipitation process would be the main area, I'd think, for caution.

Temperatures should be as low as possible to prevent heat-denaturing, and mechanical stresses eliminated in the reduction to inhalable particles by using standing-wave acostic grinding.

[Child-of-Bodom]

sorry for bringing this old thread up, I have some recomendations for working with proteins, which was part of my job.
If you want to extract proteins, you have to do this in a buffersolution, (like tris/HCl, mopso, or even a simple phosphate buffer) to keep the PH a little constant, that will keep the protein stable.
Once you have extracted anything, keep it cold all the time, proteins are not stable outside their natural envirement.
You can freeze them at -20°C rather good, but please think of it that activity gets lost when you warm it up...
If you intent to use it, and it is possible, add some DTT (dithiothreitol, C4H10O2S2) or B-mercaptoethanol to keep it as active as possible... It would be hard though to let someone eat something wich smells of H2S... :rolleyes:

[Child-of-Bodom]

sorry for bringing this old thread up, I have some recomendations for working with proteins, which was part of my job.
If you want to extract proteins, you have to do this in a buffersolution, (like tris/HCl, mopso, or even a simple phosphate buffer) to keep the PH a little constant, that will keep the protein stable.
Once you have extracted anything, keep it cold all the time, proteins are not stable outside their natural envirement.
You can freeze them at -20°C rather good, but please think of it that activity gets lost when you warm it up...
If you intent to use it, and it is possible, add some DTT (dithiothreitol, C4H10O2S2) or B-mercaptoethanol to keep it as active as possible... It would be hard though to let someone eat something wich smells of H2S... :rolleyes:

[Child-of-Bodom]

sorry for bringing this old thread up, I have some recomendations for working with proteins, which was part of my job.
If you want to extract proteins, you have to do this in a buffersolution, (like tris/HCl, mopso, or even a simple phosphate buffer) to keep the PH a little constant, that will keep the protein stable.
Once you have extracted anything, keep it cold all the time, proteins are not stable outside their natural envirement.
You can freeze them at -20°C rather good, but please think of it that activity gets lost when you warm it up...
If you intent to use it, and it is possible, add some DTT (dithiothreitol, C4H10O2S2) or B-mercaptoethanol to keep it as active as possible... It would be hard though to let someone eat something wich smells of H2S... :rolleyes:

[arnold]

can I prepare ricin from expired castor beans?

[BlackFalcoN]

I don't think the lye/acetone extraction is a very useful procedure at all.

Ricin is a protein, and most proteins are denatured when put into contact with acetone. All one would get from extracting or purifying with acetone is a denatured substance of protein.

The same goes for the treatment of the beans with lye.
Some extraction procedures ("recipes" by Kurt Saxon, Uncle Fester, crapbooks and others) recommend putting the castor beans into a solution of lye, to ease the removal of their hard outer shells.
While effective in aiding the removal of the shells, sodium hydroxide is a strong base which will not only "mush up" the hard outer shells, but will also come into contact with the fleshy inner part of the castor bean. This will inevitably denature a certain amount of the ricin present.

Breaking up the unsoaked hulls with pliers is maybe not as easy as first letting the beans soak in lye, but it will prevent the beans to come in contact with the lye which will cause denaturing. (Note that putting them in a grinder is also not recommended, since there will be heat involved in this process which can also denature the protein you are after )

Ricin is an albumen, which means it's a water soluble protein. Albuminoids can easily be extracted from a protein sludge, by using a sulfate water solution (sodium/ammonium/magnesium/... sulfates) which is basically what the patent US3060165 and other sources recommend.

The key is to get a relative oil-free pulp to start from, and acetone is not really an option according to my research.
Castor oil is a commercial product which has multiple uses and has been produced for ages. All one needs is to obtain a manual 'hobby' oil press to rid the beans of their oil.

I have attached a file that describes the process that was used in 1905 by Osborne, Mendel and Harris to extract ricin. It was this research that was eventually perfected into patent US3060165.
It also has observations on the lethality (LD50) of this preparate on various animals

Attached: 'A Study of an Extremely Pure Preparation of Ricin By Cyrus W. Field M.D.'

[Man Down Under]

Attachments take a long time to get approved, so posting them to rapidshare would seem to be a good thing, to get them out pending approval, which is what I'm doing now.

A direct link to the file:
http://www.jem.org/cgi/reprint/12/4/551.pdf


The minimal lethal dose of the preparation under discussion was as follows: for rabbits, o.oooi milligram per kilo; for guinea pigs, 0.0008 milligram per kilo; for dogs, 0.0006 milligram per kilo, for cats, o.ooo2 milligram per kilo; for goats, 0.003 milligram per kilo (death in three days).


Sounds like good stuff!

[BlackFalcoN]

Attachments take a long time to get approved

My mistake for posting so soon after New Year I guess ;)
[hangovers tend to have that effect on mods :p]

Anyway, after browsing heaps of patents, I found another method of extracting lectins. (proteins such as ricin, abrin, ... )

If one wants to obtain small quantities for individual targeting rather than a WMD approach, a very useful and chemical free way of extraction would be to utilise a laboratory microcentrifuge ( 5000x - 10.000x g range).
There would be no heat or strong acids/bases involved, so it's far less possible to denature the protein strings in the process.

After centrifuging the pulp in a solvent (5% acetic acid seems to be the consensus, to get a slightly acidic medium to optimize protein extraction), one would simply have to remove the supernate from the pellet formed in the Eppendorf tube with a pipette to obtain a crude lectin extract in water/solvent.

[HypocriticalBuddhist]

Because ricin is a liver poison, through containing a polypeptide sequence which cannot be metabolized by humans (toadstool and puffer-fish poisons work in the same way), it is most unlikely that "denaturation", e.g. by heating (as in the usual cooking of protein foods), which involves the deactivation of enzyme and hormone receptor sites in the protein molecule, would significantly reduce its toxicity.

It's not that simple at all, it enter's a cell, attaches to and rRNA molecule and tears it apart. Denaturing would have a huge affect.