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Ricin

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Not to be confused with raisin.
Castor oil plant, fruits
Castor beans

Ricin (/[invalid input: 'icon']ˈraɪsɪn/), from the castor oil plant Ricinus communis, is a highly toxic, naturally occurring protein. A dose as small as a few grains of table salt can kill an adult human. The LD50 of ricin is around 22 micrograms per kilogram (1.78 mg for an average adult, around 1⁄228 of a standard aspirin tablet/0.4 g gross) in humans if exposure is from injection or inhalation.[1] Oral exposure to ricin is far less toxic and a lethal dose can be up to 20–30 milligrams per kilogram.

Toxicity

Ricin is poisonous if inhaled, injected, or ingested, acting as a toxin by the inhibition of protein synthesis. It is resistant, but not impervious, to digestion by peptidases. By ingestion, the pathology of ricin is largely restricted to the gastrointestinal tract where it may cause mucosal injuries; with appropriate treatment, most patients will make a full recovery.[2] Because the symptoms are caused by failure to make protein, they emerge only after a variable delay from a few hours to a full day after exposure. An antidote has been developed by the UK military, although it has not yet been tested on humans.[3][4] An antidote has been developed by the U.S. military which has so far shown to be safe and effective when lab mice were injected with antibody-rich blood mixed with ricin, and has had some human testing.[5] Symptomatic and supportive treatment are available. Long term organ damage is likely in survivors. Ricin causes severe diarrhea and victims can die of shock. Death typically occurs within 3–5 days of the initial exposure.[6] Abrin is a similar toxin, found in the highly ornamental rosary pea.

Deaths from ingesting castor plant seeds are rare, partly because of their indigestible capsule, and because the body can, with difficulty, digest ricin.[7] The pulp from eight beans is considered dangerous to an adult.[8] A solution of saline and glucose has been used to treat ricin overdose.[9] Rauber and Heard have written that close examination of early 20th century case reports indicates that public and professional perceptions of ricin toxicity "do not accurately reflect the capabilities of modern medical management".[10]

==

Biochemistry

Ricin is classified as a type 2 ribosome inactivating protein (RIP). Whereas Type 1 RIPs consist of a single enzymatic protein chain, Type 2 RIPs, also known as holotoxins, are heterodimeric glycoproteins. Type 2 RIPs consist of an A chain that is functionally equivalent to a Type 1 RIP, covalently connected by a single disulfide bond to a B chain that is catalytically inactive, but serves to mediate entry of the A-B protein complex into the cytosol. Both Type 1 and Type 2 RIPs are functionally active against ribosomes in vitro, however only Type 2 RIPs display cytoxicity due to the lectin properties of the B chain. In order to display its ribosome inactivating function, the ricin disulfide bond must be reductively cleaved.[11]

Structure

Ribosome inactivating protein (Ricin A chain)
Ricin structure. The A chain is shown in blue and the B chain in orange.
Identifiers
SymbolRIP
PfamPF00161
InterProIPR001574
PROSITEPDOC00248
SCOP21paf / SCOPe / SUPFAM
Available protein structures:
Pfam  structures / ECOD  
PDBRCSB PDB; PDBe; PDBj
PDBsumstructure summary
Ricin-type beta-trefoil lectin domain (Ricin B chain)
Identifiers
Symbol?
PfamPF00652
Pfam clanCL0066
PROSITEIPR000772
SCOP21abr / SCOPe / SUPFAM
CAZyCBM13
Available protein structures:
Pfam  structures / ECOD  
PDBRCSB PDB; PDBe; PDBj
PDBsumstructure summary

The tertiary structure of ricin was shown to be a globular, glycosylated heterodimer of approximately 60–65 kDA.[7] Ricin toxin A chain and ricin toxin B chain are of similar molecular weight, approximately 32 kDA and 34 kDA respectively.

  • Ricin A chain (RTA) is an N-glycoside hydrolase composed of 267 amino acids.[12] It has three structural domains with approximately 50% of the polypeptide arranged into alpha-helices and beta-sheets.[13] The three domains form a pronounced cleft that is the active site of RTA.
  • Ricin B chain (RTB) is a lectin composed of 262 amino acids that is able to bind terminal galactose residues on cell surfaces.[14] RTB form a bilobal, barbell-like structure lacking alpha-helices or beta-sheets where individual lobes contain three subdomains. At least one of these three subdomains in each homologous lobe possesses a sugar-binding pocket that gives RTB its functional character.

Many plants such as barley have the A chain but not the B chain. People do not get sick from eating large amounts of such products, as ricin A is of extremely low toxicity as long as the B chain is not present.

Entry into the cytosol

The ability of ricin to enter the cytosol depends on hydrogen bonding interactions between RTB amino acid residues and complex carbohydrates on the surface of eukaryotic cells containing either terminal N-acetylgalactosamine or beta-1,4-linked galactose residues. Additionally, the mannose-type glycans of ricin are able to bind cells that express mannose receptors.[15] Experimentally, RTB has been shown to bind to the cell surface on the order of 106-108 ricin molecules per cell surface.[16]

The profuse binding of ricin to surface membranes allows internalization with all types of membrane invaginations. Experimental evidence points to ricin uptake in both clathrin-coated pits, as well as clathrin-independent pathways including caveolae and macropinocytosis.[17][18] Vesicles shuttle ricin to endosomes that are delivered to the Golgi apparatus. The active acidification of endosomes are thought to have little effect on the functional properties of ricin. Because ricin is stable over a wide pH range, degradation in endosomes or lysosomes offer little or no protection against ricin.[19] Ricin molecules are thought to follow retrograde transport via early endosomes, the trans-Golgi network, and the Golgi to enter the lumen of the endoplasmic reticulum (ER).[20]

For ricin to function cytotoxically, RTA must be reductively cleaved from RTB in order to release a steric block of the RTA active site. This process is catalysed by the protein PDI (protein disulphide isomerase) that resides in the lumen of the ER.[21] Free RTA in the ER lumen then partially unfolds and partially buries into the ER membrane, where it is thought to mimic a misfolded membrane-associated protein.[22] Roles for the ER chaperones GRP94 [23] and EDEM [24] have been proposed prior to the 'dislocation' of RTA from the ER lumen to the cytosol in a manner that utilizes components of the endoplasmic reticulum-associated protein degradation (ERAD) pathway. ERAD normally removes misfolded ER proteins to the cytosol for their destruction by cytosolic proteasomes. Dislocation of RTA requires ER membrane-integral E3 ubiquitin ligase complexes,[25] but RTA avoids the ubiquitination that usually occurs with ERAD substrates because of its low content of lysine residues, which are the usual attachment sites for ubiquitin.[26] Thus RTA avoids the usual fate of dislocated proteins (destruction that is mediated by targeting ubiquitinylated proteins to the cytosolic proteasomes). In the mammalian cell cytosol, RTA then undergoes triage by cytosolic molecular chaperones that results in its folding to a catalytic conformation [23] that de-purinates ribosomes, thus halting protein synthesis.

Ribosome inactivation

Study of the N-glycosidase activity of ricin was pioneered by Endo and Tsurugi[27] who showed that RTA cleaves a glycosidic bond within the large rRNA of the 60S subunit of eukaryotic ribosomes. They subsequently showed RTA specifically and irreversibly hydrolyses the N-glycosidic bond of the adenine residue at position 4324 (A4324) within the 28S rRNA, but leaves the phosphodiester backbone of the RNA intact.[28] The ricin targets A4324 that is contained in a highly conserved sequence of 12 nucleotides universally found in eukaryotic ribosomes. The sequence, 5’-AGUACGAGAGGA-3’, termed the sarcin-ricin loop, is important in binding elongation factors during protein synthesis.[29] The depurination event rapidly and completely inactivates the ribosome, resulting in toxicity from inhibited protein synthesis. A single RTA molecule in the cytosol is capable of depurinating approximately 1500 ribosomes per minute.

Depurination reaction

Within the active site of RTA, there exist several invariant amino acid residues involved in the depurination of ribosomal RNA.[19] Although the exact mechanism of the event is unknown, key amino acid residues identified include tyrosine at positions 80 and 123, glutamic acid at position 177, and arginine at position 180. In particular, Arg180 and Glu177 have been shown to be involved in the catalytic mechanism, and not substrate binding, with enzyme kinetic studies involving RTA mutants. The model proposed by Mozingo and Robertus,[30] based x-ray structures, is as follows:

  1. Sarcin-ricin loop substrate binds RTA active site with target adenine stacking against tyr80 and tyr123.
  2. Arg180 is positioned such that it can protonate N-3 of adenine and break the bond between N-9 of the adenine ring and C-1’ of the ribose.
  3. Bond cleavage results in an oxycarbonium ion on the ribose, stabilized by Glu177.
  4. N-3 protonation of adenine by Arg180 allows deprotonation of a nearby water molecule.
  5. Resulting hydroxyl attacks ribose carbonium ion.
  6. Depurination of adenine results in a neutral ribose on an intact phosphodiester RNA backbone.

Manufacture

Ricin is easily purified from castor oil manufacturing waste. The aqueous phase left over from the oil extraction process is called waste mash. It would contain about 5–10% ricin by weight, but heating during the oil extraction process denatures the protein, making the resultant seed cake safe for use as animal feed. From fresh seed, separation requires chromatographic techniques similar to other plant proteins.[citation needed]

Patented extraction process

A process for extracting ricin has been described in a patent.[31] The described extraction method is very similar to that used for the preparation of soy protein isolates.

The patent was removed from the United States Patent and Trademark Office (USPTO) database sometime in 2004.[32][33] Modern theories of protein chemistry cast doubt on the effectiveness of the methods disclosed in the patent.[34]

Potential medicinal use

Some researchers have speculated about using ricins in the treatment of cancer, as a so-called "magic bullet" to destroy targeted cells.[35] Because ricin is a protein, it can be genetically linked to a monoclonal antibody to target malignant cells recognized by the antibody. The major problem with ricin is that its native internalization sequences are distributed throughout the protein. If any of these native internalization sequences are present in a therapeutic, then the drug will be internalized by, and kill, untargeted epithelial cells as well as targeted cancer cells.

Some researchers hope that modifying ricin will sufficiently lessen the likelihood that the ricin component of these immunotoxins will cause the wrong cells to internalize it, while still retaining its cell-killing activity when it is internalized by the targeted cells. Generally, however, ricin has been superseded for medical purposes by more practical fragments of bacterial toxins, such as diphtheria toxin, which is used in denileukin diftitox, an FDA-approved treatment for leukemia and lymphoma. No approved therapeutics contain ricin.

A promising approach is also to use the non-toxic B subunit as a vehicle for delivering antigens into cells thus greatly increasing their immunogenicity. Use of ricin as an adjuvant has potential implications for developing mucosal vaccines.

Ricinine has some insecticidal effects on three insect pests as well as a hepatoprotective activity. Ricinine, when administered to mice at low doses has memory-improving effects. The signs of intoxication caused by ricinine can be used as chemical model of epilepsy in the screening of anticonvulsant drugs.[36]

Incidents involving ricin

Main article: Incidents involving ricin

Ricin has been involved in a number of incidents, including the high-profile assassination of Georgi Markov using a weapon disguised as an umbrella.

The ingestion of Ricinus communis cake is responsible for fatal ricin poisoning in animals.[37]

It was reported on April 17, 2013, that a letter addressed to President Barack Obama containing a suspicious substance was intercepted at an offsite mail facility. [38] Following anthrax scares in the wake of the Sept. 11, 2001 terror attacks, the White House moved all mail processing off-site.

Use as a chemical or biological warfare agent

The United States investigated ricin for its military potential during World War I.[39] At that time it was being considered for use either as a toxic dust or as a coating for bullets and shrapnel. The dust cloud concept could not be adequately developed, and the coated bullet/shrapnel concept would violate the Hague Convention of 1899 (adopted in U.S. law at 32 Stat. 1903), specifically Annex §2, Ch.1, Article 23, stating "... it is especially prohibited ... [t]o employ poison or poisoned arms".[40] World War I ended before the United States weaponized ricin.

During World War II the United States and Canada undertook studying ricin in cluster bombs.[41] Though there were plans for mass production and several field trials with different bomblet concepts, the end conclusion was that it was no more economical than using phosgene. This conclusion was based on comparison of the final weapons, rather than ricin's toxicity (LCt50 ~40 mg·min/m3). Ricin was given the military symbol W or later WA. Interest in it continued for a short period after World War II, but soon subsided when the U.S. Army Chemical Corps began a program to weaponize sarin.

The Soviet Union also possessed weaponized ricin. There were speculations that the KGB used it outside of the Soviet bloc; however, this was never proven. In 1978, the Bulgarian dissident Georgi Markov was assassinated by Bulgarian secret police who surreptitiously "shot" him on a London street with a modified umbrella using compressed gas to fire a tiny pellet contaminated with ricin into his leg.[2][42] He died in a hospital a few days later; his body was passed to a special poison branch of the British Ministry of Defence (MOD) that discovered the pellet during an autopsy. The prime suspects were the Bulgarian secret police: Georgi Markov had defected from Bulgaria some years previously and had subsequently written books and made radio broadcasts which were highly critical of the Bulgarian communist regime. However, it was believed at the time that Bulgaria would not have been able to produce the pellet, and it was also believed that the KGB had supplied it. The KGB denied any involvement although high-profile KGB defectors Oleg Kalugin and Oleg Gordievsky have since confirmed the KGB's involvement. Earlier, Soviet dissident Aleksandr Solzhenitsyn also suffered (but survived) ricin-like symptoms after a 1971 encounter with KGB agents.[43]

Despite ricin's extreme toxicity and utility as an agent of chemical/biological warfare, it is extremely difficult to limit the production of the toxin. The castor bean plant from which ricin is derived is a common ornamental and can be grown at home without any special care, and the major reason ricin is a public health threat is that it is easy to obtain.[citation needed]

Under both the 1972 Biological Weapons Convention and the 1997 Chemical Weapons Convention, ricin is listed as a schedule 1 controlled substance. Despite this, more than 1 million tonnes of castor beans are processed each year, and approximately 5% of the total is rendered into a waste containing negligible concentrations of undenatured ricin toxin.[44]

Ricin is several orders of magnitude less toxic than botulinum or tetanus toxin, but the latter are harder to come by. Compared to botulinum or anthrax as biological weapons or chemical weapons, the quantity of ricin required to achieve LD50 over a large geographic area is significantly more than an agent such as anthrax (tons of ricin vs. only kilogram quantities of anthrax).[45] Ricin is easy to produce, but is not as practical nor likely to cause as many casualties as other agents.[2] Ricin is inactivated (the protein changes structure and becomes less dangerous) much more readily than anthrax spores, which may remain lethal for decades. Jan van Aken, a German expert on biological weapons, explained in a report for The Sunshine Project that Al Qaeda's experiments with ricin suggest their inability to produce botulinum or anthrax.[46]

References

  1. ^ "EFSA Scientific Opinion: Ricin (from Ricinus communis) as undesirable substances in animal feed [1] - Scientific Opinion of the Panel on Contaminants in the Food Chain". Efsa.europa.eu. Retrieved 2010-09-01.
  2. ^ a b c Schep, L. J.; Temple, W. A.; Butt, G. A.; Beasley, M. D. (2009). "Ricin as a weapon of mass terror—separating fact from fiction". Environ Int. 35 (8): 1267–71. doi:10.1016/j.envint.2009.08.004. PMID 19767104.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  3. ^ Rincon, Paul (2009-11-11). "http://news.bbc.co.uk/1/hi/sci/tech/8351666.stm". BBC News. Retrieved 2010-09-01. {{cite news}}: External link in |title= (help)
  4. ^ "Human trial proves ricin vaccine safe, induces neutralizing antibodies; further tests planned". University of Texas Southwestern Medical Center. 2006-01-30. Retrieved 2012-05-07.
  5. ^ Karen Fleming-Michael (2005-09-01). "http://www.dcmilitary.com/dcmilitary_archives/stories/090105/36813-1.shtml". Dcmilitary.com. Retrieved 2010-09-01. {{cite web}}: External link in |title= (help)
  6. ^ "The Emergency Response Safety and Health Database: Biotoxin: RICIN". Centers for Disease Control and Prevention (CDC). 2008-08-01. Retrieved 2011-02-17.
  7. ^ a b Aplin, P. J.; Eliseo, T. (1997). "Ingestion of castor oil plant seeds". Med. J. Aust. 167 (5): 260–1. PMID 9315014.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  8. ^ Wedin, G. P.; Neal, J. S.; Everson, G. W.; Krenzelok, E. P. (1986). "Castor bean poisoning". American Journal of Emergency Medicine. 4 (3): 259–61. doi:10.1016/0735-6757(86)90080-X. PMID 3964368.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  9. ^ Kopferschmitt, J.; Flesch, F.; Lugnier, A.; Sauder, P.; Jaeger, A; Mantz, J. M. (1983). "Acute voluntary intoxication by ricin". Human Toxicology. 2 (2): 239–42. doi:10.1177/096032718300200211. PMID 6862467.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  10. ^ Rauber, A.; Heard, J. (1985). "Castor bean toxicity re-examined: a new perspective". Veterinary and Human Toxicology. 27 (6): 498–502. PMID 4082461.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  11. ^ Wright, H. T.; Robertus, J. D. (1987). "The intersubunit disulfide bridge of ricin is essential for cytotoxicity". Arch Biochem Biophys. 256 (1): 280–4. doi:10.1016/0003-9861(87)90447-4. PMID 3606124.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  12. ^ Olnes, S.; Pihl, A. (1973). "Different biological properties of the two constituent peptide chains of ricin, a toxic protein inhibiting protein synthesis". Biochemistry. 12 (16): 3121–26. doi:10.1021/bi00740a028. PMID 4730499.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  13. ^ Weston, S. A.; Tucker, A. D.; Thatcher, D. R.; et al. (1994). "X-ray structure of recombinant ricin A-chain at 1.8 A resolution". J Mol Biol. 244 (4): 410–22. doi:10.1006/jmbi.1994.1739. PMID 7990130. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  14. ^ Wales, R.; Richardson, P. T.; Robers, L. M.; Woodland, H. R.; et al. (1991). "Mutational analysis of the galactose binding ability of recombinant ricin b chain". J Biol Chem. 266: 19172–79. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  15. ^ Magnusson, A. S.; Kjeken, R.; Berg, T. (1993). "Characterization of two distinct pathways of endocytosis of ricin by rat liver endothelial cells". Exp Cell Res. 205 (1): 118–25. doi:10.1006/excr.1993.1065. PMID 8453986.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  16. ^ Sphyris, N.; Lord, J. M.; Wales, R.; et al. (1995). "Mutational analys is of the ricinus lectin b-chains: Galactose-binding ability of the gamma subdomain of ricinus communis agglutin b-chain". J Biol Chem. 270 (35): 20292–97. doi:10.1074/jbc.270.35.20292. PMID 7657599. {{cite journal}}: Unknown parameter |author-separator= ignored (help)CS1 maint: unflagged free DOI (link)
  17. ^ Moya, M.; Dautry-Varsat, A.; Goud, B.; et al. (1985). "Inhibition of coated pit formation in Hep2 cells blocks the cytotoxicity of diphtheria toxin but not that of ricin toxin". J Cell Biol. 101 (2): 548–59. doi:10.1083/jcb.101.2.548. PMC 2113662. PMID 2862151. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  18. ^ Nichols, B. J.; Lippincott-Schwartz, J. (2001). "Endocytosis without clathrin coats". Trends Cell Biol. 11 (10): 406–12. doi:10.1016/S0962-8924(01)02107-9. PMID 11567873.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  19. ^ a b Lord, M. J.; Jolliffe, N. A.; Marsden, C. J.; et al. (2003). "Ricin Mechanisms of Cytotoxicity". Toxicol Rev. 22 (1): 53–64. doi:10.2165/00139709-200322010-00006. PMID 14579547. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  20. ^ Spooner, R. A. (2006). "Retrograde transport pathways utilised by viruses and protein toxins". Virology Journal. 3: 26–35. doi:10.1186/1743-422X-3-26. PMC 1524934. PMID 16603059. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)CS1 maint: unflagged free DOI (link)
  21. ^ Spooner, R. A. (2004). "Protein disulphide-isomerase reduces ricin to its A and B chains in the endoplasmic reticulum". Biochem. J. 383 (Pt 2): 285–293. doi:10.1042/BJ20040742. PMC 1134069. PMID 15225124. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)
  22. ^ Mayerhofer, P. U. (2009). "A chain insertion into endoplasmic reticulum membranes is triggered by a temperature increase to 37(degrees)C". Journal of Biological Chemistry. 284 (15): 10232–10242. doi:10.1074/jbc.M808387200. PMC 2665077. PMID 19211561. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)CS1 maint: unflagged free DOI (link)
  23. ^ a b Spooner, R. A. (2008). "Cytosolic chaperones influence the fate of a toxin dislocated from the endoplasmic reticulum". Proceedings of the National Academy of Sciences. 105 (45): 17408–17413. doi:10.1073/pnas.0809013105. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)
  24. ^ Slominska-Wojewodzka, Monika (2006). "EDEM Is Involved in Retrotranslocation of Ricin from the Endoplasmic Reticulum to the Cytosol". Mol Biol Cell. 17 (4): 1664–1675. doi:10.1091/mbc.E05-10-0961. PMC 1415288. PMID 16452630. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)
  25. ^ Li, S. (2010). "Folding-competent and folding-defective forms of ricin A chain have different fates after retrotranslocation from the endoplasmic reticulum". Molecular Biology of the Cell. 21 (15): 2543–2554. doi:10.1091/mbc.E09-08-0743. PMC 2912342. PMID 20519439. {{cite journal}}: Unknown parameter |coauthors= ignored (|author= suggested) (help)
  26. ^ Deeks, E. D.; Cook, J. P.; Day, P. J.; et al. (2002). "The low lysine content of ricin A chain reduces the risk of proteolytic degradation after translocation from the endoplasmic reticulum to the cytosol". Biochemistry. 41 (10): 3405–13. doi:10.1021/bi011580v. PMID 11876649. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  27. ^ Endo, Y.; Tsurugi, K. (1987). "RNA N-glycosidase activity of ricin A-chain: mechanism of action of the toxic lectin ricin on eukaryotic ribosomes". J Biol Chem. 262 (17): 8128–30. PMID 3036799.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  28. ^ Endo, Y.; Tsurugi, K. (1998). "The RNA N-glycosidase activity of ricin A chain". J Biol Chem. 263 (18): 8735–9. PMID 3288622.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  29. ^ Sperti, S.; Montanaro, L.; Mattioli, A.; et al. (1973). "Inhibition by ricin of protein synthesis in vitro: 60 S ribosomal subunit as the target of the toxin". Biochem J. 136 (3): 813–5. PMC 1166019. PMID 4360718. {{cite journal}}: Unknown parameter |author-separator= ignored (help)
  30. ^ Monzingo, A. F.; Robertus, J. D. (1992). "X-ray analysis of substrate analogs in the ricin A-chain active site". J Mol Biol. 244 (4): 410–22. doi:10.1006/jmbi.1994.1739. PMID 7990130.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  31. ^ "Preparation of Toxic Ricin", U.S. patent 3,060,165, assigned to the U.S. Secretary of the Army, inventors: Harry L. Craig, O. H. Alderks, Alsoph H. Corwin, Sally H. Dieke, and Charlotte Karel (granted October 23, 1962)
  32. ^ "Harry L. Craig, O. H. Alderks, Alsoph H. Corwin, Sally H. Dieke, and Charlotte Karel, US Patent 3,060,165, "Preparation of Toxic Ricin", granted October 23, 1962". V3.espacenet.com. Retrieved 2010-09-01.
  33. ^ "Ricin Patent". Cryptome.org. 2004-03-12. Retrieved 2010-09-01.
  34. ^ John Pike. "http://www.globalsecurity.org/org/nsn/nsn-040723.htm". Globalsecurity.org. Retrieved 2010-09-01. {{cite web}}: External link in |title= (help)
  35. ^ . doi:10.2165/00139709-200322010-00006. {{cite journal}}: Cite journal requires |journal= (help); Missing or empty |title= (help)
  36. ^ Liu, Xiao; Li, Duan (2006). "Biological activity of ricinine and outlook of its applied development". Chinese Journal of Pharmacology and Toxicology. 20 (1): 76–8.
  37. ^ Soto-Blanco, B.; Sinhorini, I. L.; Gorniak, S. L.; Schumaher-Henrique, B. (2002). "Ricinus communis cake poisoning in a dog". Vet Hum Toxicol. 44 (3): 155–6. PMID 12046967. {{cite journal}}: Unknown parameter |month= ignored (help)CS1 maint: multiple names: authors list (link)
  38. ^ http://usnews.nbcnews.com/_news/2013/04/17/17794663-letter-sent-to-obama-tests-positive-for-ricin-officials-say?lite
  39. ^ Augerson, William S.; Spektor, Dalia M.; United States Dept. of Defense, Office of the Secretary of Defense, National Defense Research Institute (U.S.) (2000). A Review of the Scientific Literature as it Pertains to Gulf War Illnesses. Rand Corporation, ISBN 978-0-8330-2680-4
  40. ^ "The Avalon Project — Laws of War : Laws and Customs of War on Land (Hague II); July 29, 1899". Avalon.law.yale.edu. Retrieved 2010-09-01.
  41. ^ Gupta, Ramesh C. (2009). Handbook of Toxicology of Chemical Warfare Agents. Academic Press. ISBN 978-0-12-374484-5.
  42. ^ "Ricin and the umbrella murder". CNN. January 7, 2003. Retrieved 2008-03-15.
  43. ^ D. M. Thomas. Alexander Solzhenitsyn: A Century in His Life. pp. 368–378.
  44. ^ "Cornell University Department of Animal Science". Ansci.cornell.edu. Retrieved 2012-05-07.
  45. ^ Kortepeter, M. G.; Parker, G. W. (1999). "Potential biological weapons threats". 5 (4). U.S. Army Medical Research Institute of Infectious Diseases: 523–527. {{cite journal}}: Cite journal requires |journal= (help)CS1 maint: multiple names: authors list (link)
  46. ^ van Aken, Jan (2001). "Biological Weapons: Research Projects of the German Army". Backgrounder Series #7. The Sunshine Project.
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