RICIN
TOXIN FROM CASTOR BEAN PLANT
Ricinus communis
Ricin is one of the most poisonous naturally occuring substances known.
Index
- Castor
Bean Plant, Poisoning, and Oil
- More about Ricin Toxin from Castor Beans
Introduction
The seeds from
the castor bean plant, Ricinus communis, are poisonous to people, animals and insects.
One of the main toxic proteins is "ricin",
named by Stillmark in 1888 when he tested the beans'
extract on red blood cells and saw them agglutinate. Now we know that the
agglutination was due to another toxin that was also present, called RCA (Ricinus communis agglutinin). Ricin is a potent cytotoxin
but a weak hemagglutinin, whereas RCA is a weak cytotoxin and a powerful hemagglutinin.
Poisoning by ingestion
of the castor bean is due to ricin, not RCA,
because RCA does not penetrate the intestinal wall, and does not affect red
blood cells unless given intravenously. If RCA is injected into the blood, it
will cause the red blood cells to agglutinate and burst by hemolysis.
Perhaps just one
milligram of ricin can kill an adult.
The symptoms of
human poisoning begin within a few hours of ingestion.
The symptoms are:
- abdominal pain
- vomiting
- diarrhea, sometimes bloody.
Within several
days there is:
- severe dehydration,
- a decrease in urine,
- and a decrease in blood pressure.
If death has not
occurred in 3-5 days, the victim usually recovers. It is advisable to keep
children away from the castor bean plant or necklaces made with its seeds. In
fact donít even have them in or around a house with
small children. If they ingest the leaves or swallow the seeds, they may get
poisoned. The highly toxic seeds beaded into necklaces, cause skin irritation
at the contact point.
If the seed is
swallowed without chewing, and there is no damage to the seed coat, it will
most likely pass harmlessly through the digestive tract. However, if it is
chewed or broken and then swallowed, the ricin toxin will
be absorbed by the intestines.
It is said
that just one seed can kill a child.
Children are more sensitive than adults to fluid loss due to vomiting and
diarrhea, and can quickly become severely dehydrated and die.
Castor bean plants
in a garden should not be allowed to flower and seed.
Castor
Bean Plant, Poisoning, and Oil [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
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Mechanism
of Toxic Action
Many cytotoxic proteins from a variety of plants have
been identified, and they are related to ricin both
in structure and function. They inhibit protein sythesis
by specifically and irreversibly inactivating eukaryotic ribosomes.
These "ribosome-inactivating
proteins" (RIPs) are typically N-glycosylated, 30 kDa monomers (Type
1 RIPs). However, in order to bind to the cell
surface galactosides and enter the cytosol to reach ribosomes, they
require a second monomer, a galactose-binding, 30 kDa lectin. The monomers are
joined by a disulfide bridge to form the toxic heterodimers
(Type 2 RIPs).
Some plants, such
as wheat andbarley, have only Type 1
RIPs, and are not poisonous, while others,
such as the castor bean plant seed, contain the Type 2 RIPs that are among the most potent cytotoxins in nature. 5% of the Ricinus
seed consists of ricin and RCA (Ricinus
communis agglutinin).
Ricin is a heterodimeric type 2
RIP. This ribosome-inactivating enzyme (32 kDa), also
known as the A chain, is linked by a disulfide bond to the galactose/N-acetylgalactosamine-binding
lectin (34 kDa), also
called the B chain.
Ricin Biosynthesis
Ricin and RCA are synthesized in the endosperm cells of
maturing seeds, and are stored in an organelle called the "protein
body", a vacuolar compartment. When the mature seed germinates, the toxins
are destroyed by hydrolysis within a few days.
Ricin begins sythesis as a prepropolypeptide that contains both A and B
chains. The signal sequence of the Nh3-terminus targets the nascent
chain to the endoplasmic reticulum (ER) and is then cleaved off. As the proricin polypeptide elongates it is N-glycosylated within the lumen of the ER. Protein disulfide isomerases catalyze disulfide bond formation as the proricin molecule folds itself. Proricin
undergoes further oligosaccharide modifications within the Golgi
complex and then is transported within vesicles to the protein bodies.
Ricin is not catalytically active until it is proteolytically cleaved by an endopeptidase
within the protein bodies. This splits the polypeptide into the A chain
and the B chain still linked by a single disulfide bond. Since ricin is inactive until then, the plant avoids poisoning
its own ribosomes in case some proricin
accidentally passes into the cytosol during synthesis
and transport.
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis][Ricin Enzymatic Action] [Ricin Structure][Ricin Uptake] [Therapeutic
Applications of Immunotoxins][Toxigenic Ablation] [References]
·
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·
Ricin Enzymatic Action
·
The ricin A portion of
the heterodimer is the enzyme that binds and depurinates a specific adenine of the 28S rRNA. The adenine ring of the ribosome becomes
sandwiched between two tyrosine rings in the catalytic cleft of the
enzyme and is hydrolyzed by the enzymeís
N-glycosidase action. The target adenine is a specific RNA sequence that
contains the unusual tetranucleotide loop, GAGA.
Ricin is more active against animal than plant ribosomes, and intact bacterial ribosomes
are generally not susceptible.
·
Ricin Structure
·
·
This figure from
Lord et al, depicts a 3-dimensional ribbon drawing of ricin, modeled from X-ray crystallography data. The
upper right half, the dotted ribbon, is the A chain, and the lower left half,
the solid ribbon, is the B chain.
·
The A chain
(or RTA)is
a 267-amino acid globular protein. It has 8 alpha helices and 8 beta sheets.
The substrate binding site is the cleft marked by the substrate adenine ring.
·
The B chain
(or RTB) is a 262-amino acid protein that is shaped like a barbell. It has
a binding site for galactose at each end, (depicted
by lactose rings). These two sites allow hydrogen bonding to specific membrane
sugars (galactose and N-acetyl galactosamine).
A disulfide bridge (-S-S-) joins RTA with RTB (far-right, center). The spheres
are trapped water molecules.
·
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
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Ricin Uptake
The RTB portion of ricin
binds to both glycoproteins and glycolipids
at cell surfaces that terminate with galactose. It
has two binding sites for galactose, and 106 to 108 ricin molecules may bind per cell. However, just a
single ricin molecule that enters the cytosol can inactivate over 1,500 ribosomes
per minute and kill the cell.
As shown in the diagram, the pathway for
internalization of ricin involves:
- endocytosis by coated pits and vesicles or,
- endocytosos by smooth pits and vesicles. The vesicles fuse
with an endosome.
- Many ricin molecules are returned to the
cell surface by exocytosis, or
- the
vesicles may fuse to lysosomes where the ricin would be destroyed.
- If the ricin-containing vesicles fuse to
the Trans Golgi Network, (TGN), thereís still a chance they may
- return to the cell surface.
- Toxic action will occur when RTA, aided by RTB, penetrates the TGN
membrane and is liberated into the cytosol.
Once inside the cytosol,
the RTA catalyzes the depurination of the ribosomes, halting protein synthesis.
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
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Therapeutic Applications of
Immunotoxins
Ricin can be targeted to specific cells, such as cancer
cells, by conjugating the RTA subunit to antibodies or growth factors that
preferentially bind the unwanted cells. These immunotoxins
have worked very well for in vitro applications, e.g. bone marrow
transplants. Although they have not worked very well in many in vivo
situations, progress in this area of research shows promise for the future.
·
IN VITRO APPLICATIONS
In bone marrow transplant procedures,
RTA-immunotoxins have been used successfully
to destroy T lymphocytes in bone marrow taken from histocompatible
donors. This reduces rejection of the donor bone marrow, a problem
called "graft-vs-host disease" (GVHD). In
steroid-resistant, acute GVDH situations, RTA-immunotoxins
helped alleviate the condition. Also, in autologous
bone marrow transplantation, a sample of the patients own bone marrow
is treated with anti-T cell immunotoxins to destroy
malignant T-cells in T cell leukemias and lymphomas.
·
IN VIVO APPLICATIONS
"For the in vivo treatment of solid
tumors, considerable problems can arise due to poor access of the immunotoxin (IT) to the tumor mass; lack of IT specificity,
tumor cell heterogeneity, antigen shedding, breakdown or rapid clearance of the
IT, and dose-limiting side effects". (Lord et al.).
One common problem encountered in patients treated with ricin-immunotoxins
is the "vascular leak syndrome", in which fluids leak from blood
vessels leading to hypoalbuminemia, weight gain and
pulmonary edema. "Research efforts to expand and develop immunotoxins and therapies for clinical use in cancer
and AIDS are continuing with strategies utilizing recombinant DNA
technology (Lord et al.).
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
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Toxigenic Ablation
·
TOXIGENES
"Toxigenes
are DNA fusions in which DNA encoding a potent toxin, e.g. RTA, is placed under
the transcriptional control of a tissue- or developmental stage-specific
promoter and/or enhancer. When expressed intracellularly,
the toxigene product causes cell death. The
introduction and expression of a toxigene in
transgenic animals or plants may lead to cell type-specific ablation,
which can be used to
- study developmental cell lineages or to
- generate animal models of degenerative diseases." (Lord et al.)
·
SUICIDE
TRANSPORT
Diagram shows injection of ricin into vagal nerve and subsequent
destruction of neurons (dashed neurons destroyed, solid neurons unaffected).
Neuroscientists can selectively destroy
neurons by injecting ricin into nerves. Retrograde
axonal transport mechanisms bring the toxin to the neuronal cell bodies where
the ribosomes are localized.
Ultrastructural analysis reveals that ricin
first causes the dispersion of polyribosomes, and
then the rough endoplasmic reticulum disorganizes into smooth vesicles. The
cell bodies (perikaryon) swell, the nuclei degenerate
and the entire neuron disintegrates.
Since ricin is a N-acetyl galactosamine-binding lectin, it can be used with different lectins
that have different specificities tomap
neuronal patterns of glycosylation. When suicide
transport is observed after injection of the toxin, it confirms the presence of
N-acetyl galactosamine residues on the
neuronal cell surface. Strategies in suicide transport work very well in
studies of adult peripheral sensory and motor neurons because they are
sensitive to ricin.
Neurons in the central nervous system of
adults are resistant to ablation by ricin, whereas
young developing brains are sensitive, suggesting that brain development
involves changes in glycosylation of CNS neurons. The
galactose terminal residues may be either clipped or
masked by addition of sialic acids residues.
In suicide transport experiments, often some
ricin leaks out of the nerve, causing systemic
poisoning of the animal. This problem can be avoided by simultaneously
administering a ricin antiserum.
The value of using suicide transport
strategies is summarized (from Wiley and Oeltmann):
- anatomical mapping of neurons
- modeling of motor neuron degenerative diseases
- studying consequences of peripheral nerve damage and repair
mechanisms
- mapping cellular neurotransmitter receptors
- disease-related applications including
- eradication of latent herpes simplex virus in
trigeminal sensory neurons
- production and analysis of glial
fibrillary bundles
- treatment of equine neuromas
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
Return to Cornell poison
plant home page
References
Alber, J.I., and D.M. Alber. (1993) Baby-Safe Houseplants and Cut Flowers: A
Guide to Keeping Children and Plants Safely Under the Same Roof. Story Communications Inc.,
Cooper, M.R., and A.W.
Johnson. (1994) Poisonous Plants
and Fungi: An Illustrated Guide. CAB International Bureau of Animal Health, Weybridge;
Czapla, T.H., and I.A. Johnston. (1990) Effect of plant lectins
on the larval development of European corn borer (Lepidoptera:pyralidae) and southern corn rootworm (Coleoptera:chrysomelidae). J.Econ.
Entomol,
Frankel, A.E., (1993) Immunotoxin
Therapy of Cancer. Oncology (
Knight, B. (1979) Ricin-a potent homicidal poison. Br. Med. J. 278:350-351.
Lord, J.M., Roberts, L.M., and J.D. Robertus. (1994). FASEB J. Feb; 8(2):201-8.
Matthews, R.W., and J.R.
Matthews (1978). Insect Behavior, pub. Wiley and Sons, Inc.
Okoye, JOA, Enunwaonye,
CA. Okorie, A.U. and F.O.I. Anugwa
(1987). Pathological effects of
feeding roasted castor bean meal Ricinus communis to chicks. Avian Pathol. 16(2):283-290.
Olaifa, J.I., Matsumura,F., Zeevaart, J.A.D.,
Mullin, C>A>, and P. Charalambous. (1991)
Lethal amounts of ricinine in green peach aphids myzus-persicae suzler fed on
castor bean plants. Plant Sci. (
Purushotham, N.P., Rao,
M.S., and G.V. Raghavan (1986). Utilization of castor-meal in the
concentrate mixture of sheep. Indian J. Anim. Sci. 56(10):1090-1093.
Robertus, J.D. (1988). Toxin Structure.
Cancer Treat. Res. 37:11-24.
Robertus, J. D. (1991) The structure
and action of ricin, a cytotoxic
N-glycosidase. Sem. in Cell Biol. 2:23-30.
Vitetta, E.S. and P.E. Thorpe, (1991) Immunotoxins
containing ricin or its A chain, Sem.
in Cell Biol. 2:47-58.
Wiley, R. G., and T. N. Oeltmann,
(1991) Ricin and Related Plant Toxins: Mechanisms of
Action and Neurobiological Applications; In, Handbook of Natural Toxins, Vol.6,
ed. R.F.Keeler and A.T.Tu,
Marcel Dekker, Inc.,
[Castor
Bean Plant, Poisoning, and Oil] [Ricin] [Mechanism
of Toxic Action] [Ricin Biosynthesis] [Ricin Enzymatic Action] [Ricin Structure] [Ricin Uptake] [Therapeutic
Applications of Immunotoxins] [Toxigenic Ablation] [References]
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Ricin as a Bioterrorist Agent
Agent: Ricin, a glycoprotein toxin derived
from castor plant beans, has great potential as a biological agent due to its
wide availability. The toxin is quite stable over long periods of time.
Disease: Ricin intoxification
Incubation Period: 4-8 hours
Signs/Symptoms: Symptoms will depend on the dose and route of exposure.
Initial symptoms following inhalation include weakness, fever, cough, dyspnea, nausea, chest tightness, and arthralgia.
These are usually followed by sweating, pulmonary edema, and cyanosis.
Necrotizing, suppurative airway lesions may be noted
in conjunction with rhinitis and laryngitis. If left untreated, respiratory
failure and cardiovascular collapse due to inhalation of the agent can lead to
death after 36-72 hours.
Ingestion will be followed by rapid onset of nausea, vomiting, abdominal
cramps, and severe diarrhea. Other symptoms include fever, thirst, headache,
sore throat, and dilation of the pupils. Death may occur on the third day or
later and is usually due to vascular collapse.
Diagnosis:
Differential Diagnosis: For inhalational
exposure, similar symptoms in large numbers of patients might suggest several
respiratory pathogens. Influenza, Q fever, tularemia, plague, and respiratory
illnesses due to exposure to staphylococcal enterotoxin
B (SEB) and chemical agents such as phosgene should be included in the
differential diagnosis. SEB intoxication would likely have a more rapid onset
and lower mortality. Acute lung injury induced by phosgene would progress much
faster that caused by ricin. Nerve agent intoxication
would be characterized by acute onset of cholinergic crisis with dyspnea and profuse secretions.
The differential diagnosis for patients who have ingested ricin
would include disease due to all the major enteric pathogens. These should be
ruled out with culture.
Diagnostic Tests: Early postexposure (0-24
hours) nasal or throat swabs and induced respiratory secretions may be
collected for toxin assay. Blood for serum may be collected in a tiger-top
(SST) or red top tube. Toxin assays and measurement of antibody response can be
performed on serum.
Supportive Tests: Patients with aerosol exposure to ricin
may have bilateral infiltrates on chest x-ray, arterial hypoxemia, and neutrophilic leukocytosis. A
bronchial aspirate rich in protein compared to plasma is characteristic of high
permeability pulmonary edema. Endoscopic evaluation
may reveal necrotizing suppurative lesions in
conjunction with tracheitis and bronchitis/bronchiolitis.
Treatment: Management of patients is supportive. Acetaminophen for
fever, and cough suppressants may make the patient more comfortable. Hydration
is important. For those with pulmonary intoxification,
respiratory support may be necessary. Pulmonary edema may need to be treated
with positive end expiratory pressure ventilation and diuretics. Standard
management techniques for oral poisoning should be used if the toxin is
ingested.
Infection Control/Decontamination: Standard precautions should be used by
healthcare workers. Decontaminate exposed skin by washing with soap and water
and/or 0.1% sodium hypochlorite (1 part household bleach added to 49 parts
water).
RICIN
http://www.tdh.state.tx.us/bioterrorism/facts/ricin.html
http://www.ansci.cornell.edu/plants/toxicagents/ricin/ricin.html#ricinintro
http://www.nbc-med.org/SiteContent/MedRef/OnlineRef/FieldManuals/medman/Ricin.htm
http://www.dhfs.state.wi.us/healthtips/BCD/Ricin.htm
http://waynesword.palomar.edu/plmar99.htm
Detection
http://www.plab.ku.dk/tcbh/interlec20Tang.htm
http://www.osborn-scientific.com/PDF/ricin_insert_0518.pdf
http://www.tricareregion10.org/Readiness/Smart%20Card%20Ricin.pdf
http://www.cbwinfo.com/Biological/Toxins/ricin.html
3/7/2003, J.Ruprich