Ricin

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Ricin (from Ricinus communis )
Ricin (from Ricinus communis)
Ribbon model of the rizins according to PDB  2aaI . The A chain is shown in blue, the B chain in orange.

Existing structural data : 2AAI

Mass / length primary structure 529 = 267 + 262 amino acids (A + B chain)
Secondary to quaternary structure Heterodimer A + B
Precursor (576 aa)
Identifier
External IDs
Enzyme classification
EC, category 3.2.2.22 ,  N-glycosylase
Response type Hydrolysis of an N-glycosyl bond
Substrate rRNA (28S) + H 2 O
Products defective rRNA (28S)

Ricin or ricin is an extremely poisonous protein from the seeds of the miracle tree (Ricinus communis) from the milkweed family . Chemically, ricin is a lectin that consists of a cell-binding and a toxin-promoting component. Its toxicity is attributed to an inhibition of the eukaryotic protein synthesis.

If the poison gets into the human organism, it causes the contaminated cells to die. For a fatal poisoning of a person, 0.3–20 milligrams of isolated ricin per kilogram of body weight are sufficient (in the case of oral ingestion) , corresponding to about eight seeds, the size and content of which, however, vary widely. In children, depending on their age and constitution, even half a seed can be fatal. However, it is also reported that there is a chance of survival even after ingesting 40 to 60 seeds. It depends on what time the vomiting starts. In the case of intravenous , inhalative or subcutaneous intake, significantly lower amounts are lethal , for example 43 μg / kg body weight with subcutaneous administration.

Ricin is listed in the war weapons list of the German War Weapons Control Act.

Occurrence

Ricin is found in the endosperm of the bean-like seeds of the miracle tree . The rice content in the seeds is around 1 to 5 percent of the protein content. In the castor oil , which is also obtained from the seeds, ricin is removed by extraction and heat inactivation. The seeds also contain the slightly poisonous ricinin , which is mostly in the seed coat.

Extraction

Castor seeds

Castor can be obtained from the by-products of castor oil production. In the aqueous phase of the castor oil extraction, the castor content is around five to ten percent. However, ricin will be inactivated if extraction is carried out with the application of heat, as usual.

Usually, the extraction of ricin takes place from the residues of the oil production after cold pressing and separation of the oil. As shown in a patent specification of the US Army described, ricin can from the residue in the weakly acidic pH range is extracted at pH 7 with a saturated sodium chloride - or sodium sulfate solution like are. The end product can be separated from the precipitant from the precipitate with the aid of carbon tetrachloride . However, it can be assumed that with the help of this manufacturing process, at best, a protein mixture and not pure ricin is obtained. Alternatively, ricin can be isolated and purified using chromatographic methods. After the fat fraction has been separated off, ricin is extracted with the aid of dilute acetic acid and precipitated with saturated ammonium sulfate solution . A gel filtration permits on the one hand, the separation of other proteins and the other the separation of Rizinuslectine in ricin and Rizinusagglutinin .

Alternatively, fully biologically active, recombinant ricin can be obtained with the help of genetically modified tobacco cells.

biochemistry

Superimposition of the structures of ricin and abrin (above: B-chain, below: A-chain)

Biochemical properties

Ricin is a fat-repellent and heat-sensitive protein. Structurally it is a heterodimeric, globular glycoprotein with a molar mass of 60 to 65 kDa . It consists of two different polypeptide chains , the A and the B chain, with molar masses of about 32 and 34 kDa, respectively, which are linked by a disulfide bridge . The A chain has characteristic α-helix and β-sheet structures and functions as an N- glycosidase . The B-chain is a lectin unit without pronounced secondary structures . In its structure, ricin is similar to the abrin of the paternoster pea .

biosynthesis

Ricin is formed in the endosperm of the seeds of the miracle tree. The primary product of the protein biosynthesis is of 576 amino acids existing precursor protein consisting of a signal peptide sequence (amino acids 1-35), the A chain (amino acids 36-302), a linker (amino acids 303-314) and the B-chain (amino acids 315-576) exists. The signal peptide sequence ensures transport into the endoplasmic reticulum . After the signal peptide sequence has been cleaved, the proprotein is glycosylated in the endoplasmic reticulum . A disulfide bridge is inserted between cysteines 294 and 318 with the aid of a disulfide isomerase. Another glycosylation takes place in the Golgi apparatus . After being transported further into the vacuoles , the proprotein is cleaved by an endopeptidase to form the finished ricin.

toxicology

Mode of action (toxicodynamics)

Ricin belongs to the group of ribosome inactivating proteins (RIP) of type 2 (RIP-II). Its toxic effect is due to a multi-stage process, which includes cell binding, transport through the cell, activation in the endoplasmic reticulum and ultimately a fatal inhibition of protein synthesis.

The Rizin's B chain, which carries out the lectin function, enables it to bind to the cell surface . The binding occurs unspecifically to terminal N -acetylgalactosamine residues or 1,4-linked galactose units of glycoproteins and glycolipids . In addition, ricin can bind to cells that carry the mannose receptor via its glycan chains . Under experimental conditions, up to 100 million ricin molecules reach the surface of a cell. The ricin bound to the cell surface is absorbed into the cell by endocytosis . Via endosomes , in which it is not broken down thanks to its pH stability and its enzymic resistance, the unit of A and B chains reaches the endoplasmic reticulum via the Golgi apparatus. The function of the A chain, which is initially blocked by the B chain, is released in the endoplasmic reticulum by cleaving the disulfide bridge between the two chains with the aid of a disulfide isomerase. With the help of chaperones , the A chain finally reaches the cytosol .

The A chain that has entered the cytoplasm, or ricin A , is a glycosidase that inactivates ribosomes . The exact effect is the cleavage of the adenine 4324 of the 28S-RNA of the ribosomes. On the one hand, this depurination has the effect that the formation of the initiation complex is impaired during the initiation of translation - this process of translation is greatly slowed down to a sixth of the usual speed. Adenine 4324 is also necessary for binding the elongation factors. Therefore, on the other hand, the translocation step is prevented during the elongation.

Because of their cytostatic properties, i.e. a growth-inhibiting effect on cells, toxins of the ricin type are now increasingly being investigated for their suitability as therapeutic agents for malignant tumors .

Toxicokinetics

Only a few toxicokinetic data , mostly based on animal experiments, are available for ricin . The transferability of the data to humans is only partially given. The uptake of ricin after injection , inhalation and oral intake is of toxicological relevance. After an injection, if it does not result in death, most of the rizine is excreted in the urine within 24 hours . As a protein, ricin has very little oral bioavailability . Therefore, in mice, for example, the lethal oral dose is 1000 times higher than the lethal dose after injection or inhalation. The toxicity of ricin after inhalation depends crucially on the particle size. The toxic effects after inhalation are also limited to the airways; a Systemic absorption following inhalation does not matter.

Consequences of poisoning

Since ricin is usually accidentally ingested through the consumption of castor seeds, cells of the digestive tract in particular are affected ( stomach , intestines , liver , kidneys ). Ultimately, ricin poisoning also leads to the destruction of red blood cells . Death occurs after 36 to 72 hours after the ingestion of a fatal dose.

After a latency period of several hours to days, the following symptoms may occur: nausea, vomiting, diarrhea , weakness, tachycardia , abdominal pain and acute fluid loss. In severe cases, mydriasis , cramps in the hands and legs, fever, and symptoms of liver necrosis and acute kidney failure can also occur. Death occurs from paralysis of the medullary centers, especially the respiratory center.

The poison can also be inhaled (inhaled as an aerosol ) or injected. The symptoms change accordingly: pulmonary edema and respiratory failure or severe paralysis are the result.

Symptoms

About four to eight hours after consuming the seeds:

Without treatment, death usually occurs from circulatory failure about 48 hours after poisoning. An agglutinating protein causes the red blood cells to clump together. There is no known effective antidote . According to Challoner and McCarron, the literature contains 424 cases of ricin poisoning, 8% of which were fatal if left untreated and only 0.4% when treated.

First aid

There is currently no antidote to poisoning with ricin . Substances under development, one of which protected up to 49 percent of the mice exposed to the deadly poison. The substance (Retro-2) must, however, be administered before the poison is absorbed and can therefore not be used as an antidote after poisoning.

Analytics

The reliable analysis of ricin in different matrices requires adequate sample preparation and can be done both by specific immunassays and by using the HPLC-mass spectrometry coupling . Another complementary method in analysis is the reliable detection of ricinine as a biomarker by gas chromatography with mass spectrometry coupling .

Use as a bio-weapon

Research and usage considerations

During the First World War , ricin was examined by the United States of America for its military potential. On the one hand, the coating of ammunition or explosive devices and, on the other hand, the spread of the poison as dust were considered. The spread of the poison via a cloud of dust could not be sufficiently developed and the use of ricin in weapons of war would have violated the Hague Land Warfare Regulations . So World War I passed before the United States was able to make weapons with ricin.

Canada and the United States studied the use of ricin in cluster munitions during World War II . Although plans for mass production were already in place and several field experiments were carried out with various bomblets , it was concluded that the use of phosgene was more economical. This conclusion was based on a comparison of the weapons developed and not on the toxicity of the substances used. The interest in ricin still held for a short time on the Second World War also, but dried up with the onset of research at Sarin by the Chemical Corps of the US Army. Ricin falls under both the Biological Weapons Convention of 1972 and the Chemical Weapons Convention of 1997 and is listed there as a substance that can itself be used as a biological or chemical weapon or is used in the manufacture of such weapons.

Since it also works via the respiratory tract, it was tested by the British Army for its usability as a warfare agent , but its use was discarded and the corresponding supplies destroyed, especially since it is difficult to distribute as an aerosol and is more suitable for attacks on individuals. Despite its unsuitability for an attack with the aim of mass killing, ricin is listed on List 1 of the Chemical Weapons Convention (CWC), which contains the most toxic toxins, and also in the last version of the Biological and Toxin Weapons Convention (BTWC).

Actual Uses

The first known use of ricin as a weapon was in the umbrella attack in 1978 , when Bulgarian journalist and dissident Georgi Markov was in London by Bulgarian intelligence agents on the street with an umbrella, the tip of which was prepared with a 1.52 millimeter ball containing 40 micrograms of the toxin was attacked and stabbed in the lower leg. Markov died a few days later in the hospital of circulatory failure as a result of the poisoning.

In 1991, several members of the right-wing extremist group Patriot's Council were arrested in Minnesota for making an amount of ricin sufficient to kill over 100 people for an attack on federal police. Four of them were found guilty under the Biological Weapons Anti-Terrorism Act of 1989 and were the first ever convicted under this law.

In 1995, a man who was also part of the right-wing extremist camp was arrested on the Alaska border while trying to smuggle 130 grams of powdered ricin into Canada.

The London Times reported on November 16, 2001 that manufacturing instructions for ricin had been found in abandoned Al Qaeda houses in Kabul , but not ricin itself.

In August 2002, US authorities announced that the Islamist terrorist organization Ansar al-Islam had carried out experiments with ricin and other chemical and biological warfare agents in northern Iraq.

On January 9, 2003, the dpa reported that small quantities of ricin and equipment for its production had been found in London. Six Algerians were arrested in connection with this. In April 2005, all but one of the participants were acquitted. One defendant was sentenced to life imprisonment for the murder of a police officer whom he stabbed to death during a house search. Contrary to earlier reports, the investigating authorities stated in the proceedings that they had not found any ricin, but only amateurish instructions for its production.

In February 2008, ricin was found in a hotel room in Las Vegas . The local police said that a seriously poisoned person had been hospitalized and that his life was in danger. Despite the discovery of weapons and "anarchist literature" one does not believe in a terrorist background. A 57-year-old was sentenced to three years in prison in November 2008 for possession of 4 grams of the poison, which he had already made in 1998, and a banned silencer .

On August 12, 2011, the New York Times reported intelligence about an attempt by Al-Qaeda’s regional branch in Yemen to make ricin . Accordingly, the US government was concerned that ricin could be produced there for attacks against the US.

On April 16, 2013, a letter with ricin was intercepted to the Republican US Senator Roger Wicker and to the US President Barack Obama . On May 30, 2013, it became known that on May 24 and 26, 2013, two letters with ricin to the incumbent New York City Mayor Michael Bloomberg had been intercepted. Actress Shannon Guess Richardson was identified as the sender of the letters to Bloomberg and Obama ; she confessed this. She was sentenced to 18 years in prison and $ 367,222 in compensation.

At the beginning of September 2014, poorly secured ricin residues from weapons experiments during World War I were found in a US laboratory at the National Institutes of Health near Washington, together with plague and botulism pathogens.

In December 2017, a senior citizen was arrested in a Vermont home for testing ricin on her roommates.

In May 2018, police arrested two Egyptian brothers in France. One of them remained in custody for allegedly planning an attack with ricin.

In June 2018, the Cologne police found substances for the production of ricin in an apartment of a Tunisian and assessed this as preparation for the construction of a biological weapon . The Federal Public Prosecutor started the investigation on suspicion of a serious criminal offense. (See: Castor found in Cologne ). In March 2020, the State Security Senate in Düsseldorf sentenced the 31-year-old Tunisian to 10 years in prison, and in June 2020 his co-defendant was sentenced to 8 years in prison.

literature

  • Lutz Roth, Max Daunderer, Kurt Kormann: Poison Plants - Plant Poisons. Ecomed, Landsberg / Lech 1994, ISBN 3-933203-31-7 .
  • Hermann Stillmark : About ricin, a poisonous ferment from the seeds of Ricinus comm. L. and some other Euphorbiaceen. Dorpat, 1888.

Individual evidence

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  19. ^ KR Challoner, MM McCarron: Castor bean intoxication: review of reported cases. In: Ann. Emerg. Med. 19 (10), 1990, pp. 1177-1183, PMID 2221525 .
  20. ↑ Looking for a ricin antidote: New protection against the super poison. In: n-tv . April 20, 2010.
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  25. ↑ Vera Coopman, Marc de Leeuw, Jan Cordonnier, Werner Jacobs: Suicidal death after injection of a castor bean extract (Ricinus communis L.) . In: Forensic Science International . tape 189 , no. 1 , 2009, p. e13 – e20 , doi : 10.1016 / j.forsciint.2009.04.019 .
  26. ↑ Augerson, William S .; Spektor, Dalia M .; United States Dept. of Defense, Office of the Secretary of Defense, National Defense Research Institute (US) (2000). A Review of the Scientific Literature as it Pertains to Gulf War Illnesses. Rand Corporation, ISBN 978-0-8330-2680-4 .
  27. ^ The Avalon Project - Laws of War: Laws and Customs of War on Land (Hague II); July 29, 1899 . Avalon.law.yale.edu. Retrieved September 1, 2010.
  28. ^ Gupta R: Handbook of Toxicology of Chemical Warfare Agents . Academic Press, Boston 2009, ISBN 978-0-12-374484-5 .
  29. ↑ a b Chronology of Incidents Involving Ricin ( Memento of December 16, 2014 in the Internet Archive ) CNS WMD Terrorism Research Project, James Martin Center for Nonproliferation Studies (CNS), Monterey Institute of International Studies
  30. ↑ Jeffrey M. Bale, Anjali Bhattacharjee, Eric Croddy, Richard Pilch: Ricin Found in London: An al-Qa'ida Connection? ( Memento from September 11, 2015 in the Internet Archive ) Monterey Institute of International Studies, Center for Nonproliferation Studies, Chemical and Biological Weapons Nonproliferation Program, In: CNS-Reports . February 29, 2008.
  31. ↑ Bioweapons in London: Al-Qaeda also experimented with ricin. Spiegel Online , January 8, 2003, accessed November 16, 2017 .
  32. ↑ Police: Man in critical condition after exposure to ricin (English) . In: CNN , February 2, 2008. Retrieved May 5, 2010. 
  33. ↑ Man who had Ricin sentenced to prison , AP report on nbcnews.com dated November 17, 2008, accessed May 2, 2019.
  34. ^ Mary Manning: Man with ricin in Las Vegas hotel room sentenced , Las Vegas Sun, November 17, 2008, accessed May 2, 2019.
  35. ↑ Qaeda Trying to Harness Toxin for Bombs, US Officials Fear In: NYTimes . August 12, 2011.
  36. ↑ Deadly poison ricin found in letter to US Senator In: Süddeutsche Zeitung . 17th April 2013.
  37. ^ Poison attack on Michael Bloomberg ( Memento from June 7, 2013 in the Internet Archive ) In: Tagesschau . May 30, 2013.
  38. ^ Poison attack by Shannon Guess Richardson In: Focus . December 11th 2013.
  39. ↑ 18 years imprisonment for poison letter to Obama In: Süddeutsche Zeitung . 17th July 2014.
  40. ↑ Deadly plague and botulism microbes found in US lab. BBC , September 6, 2014.
  41. ↑ 70-year-old tests ricin on roommates. In: Spiegel Online . December 2, 2017, accessed May 2, 2019 .
  42. ↑ Caroline Piquet: Nouvel attentat déjoué en France: l'un des deux Égyptiens mis en examen, l'autre relâché , Le Figaro from May 18, 2018, accessed on May 2, 2019 (French).
  43. ↑ Two brothers arrested: French authorities apparently prevent a poison attack. In: Focus Online . May 18, 2018, accessed June 15, 2018 .
  44. ^ Frank Jansen: Apparently Islamist attack in Cologne foiled . In: Tagesspiegel Online . June 13, 2018 ( tagesspiegel.de [accessed June 13, 2018]).
  45. ↑ David Keller, Robert Baumanns: Terror-Alarm in Köln: Poison in small amount fatal - what was the suspect planning? In: Express.de . June 13, 2018 ( express.de [accessed June 13, 2018]).
  46. ↑ http://www.generalbundesanwalt.de/de/showpress.php?newsid=776
  47. ↑ https://www.radioberg.de/artikel/endung-im-rizin-prozess-546815-amp.html
  48. ↑ https://www.spiegel.de/panorama/justiz/koeln-rizin-bombenbauerin-zu-langer-haft-verendungt-a-960050f0-5c22-4b73-ab8e-3ae19f074749