Vaccine developments to ricin toxin

Ricin is a potent lethal cytotoxin derived from castor seeds and combatting its toxicity is a tedious task.Currently no effective antidote has ever been formulated so far and makes therapy difficult.It also places the victims survival purely as a matter of chance.As chemotherapy andd other mehods fail to produce productive results,scientists have started to employ vaccines as a possible tool to tackle the toxin.

US researchers from Army Laboratory in Frederick have found a vaccine containing an inactivated toxin that selectively retains an activity to boost the immune system without inducing lethal toxicity to mount systemic defence response in the body by protein engineering princples and found to be effective on animal models.The US Army has reported that currently trials are underway in 30 human volunteers and plans to conduct clinical trials as well at Fort Detrick.

Vaccines are powerful tools to produce solutions in a timely consistent and biologically safe way.Historical credits to vaccines in global health are something extraordinary  with reference to pathogenic diseases and attempts such as this to focus on vaccine vehicles and expanding their horizon to toxins neutralization would most probably yield a fruitful result in future to address the problems of global threat from castor toxin.

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Catalytic transformation of castor cake by Penicillium fungus

Castor cake is a by product from castor seed but remains as a substance with restricted application potential due to its allergenic and toxic properties.People traditionally have treated the castor cake by wild fermentation for the catalytic transformation of anti-metabolites into their corresponding non-toxic forms for improving its utility.

This age-old practice was scientifically approached by the Brazilian researchers where they subjected castor cake to solid state fermentation.Penicillium simplicissimum fungus was chosen as the catalytic factor because of its potency to manipulate in castor cake substrate and generate lipid metabolizing lipase enzymes.Solid state fermentation(dessicated fermentation) was performed for the cultivation of fungus and the products, enzyme and fermented castor cake were partitioned by solid-liquid phase differential extraction.Enzyme lipase magnitude in crude isolated fraction was determined from its activity against synthetic substrate while the debris cake fermented was evaluated for its toxic protein contents and functional activities by immunoassays and immuno sensitive cell culture assays respectively.The results proved to be promising as the toxin titre and allergens induced cell degranulation were seen to be minimum and the microbial enzyme production was noticed to a significant extent as well from specific activity studies.

Results turn out to be promising but the experiment would have been better if an utility for the fermented castor cake in biology was defined.The solid state technology by fungus seems to be a viable one on commodities like castor cake to generate high value products when time factor is not signified to a greater extent.

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Medium formulation in ricin vaccine

Ricin is a noxious cytotoxin present in castor bean.People have long been pursuing research to understand its functional activities,physical structure and quantitative magnitude in beans etc.In spite of such extensive studies, no effective solution has been formulated to ricin intoxication. However researchers have found vaccine therapy as a promising route to combat the toxin.In ricin vaccinology, the wild toxin has been modified in such a way so that it tends to retain its antigenicity to boost the host defence system but not the toxic activity, called it as “ricin toxoid”.Genetic engineering  is a standardized technique to precisely produce variants in ricin and the candidate with expected property are subsequently chosen.

US scientists have identified an elite ricin toxoid from earlier studies and made an evaluation on vaccine formulation to achieve enhanced antigenicity and stability.Initially they treated the protein in phosphate buffer solution and found structural and functional tolerance in protein after cyclic exposures to extreme temperature ranges.To further improve its adjuvant adherence and related antigenicity, toxoid was treated with aluminium hydroxide adjuvant in succinate buffer and evaluated by differential centrifugation and immuno assays.The results indicated that aluminium hydroxide showed a good compatibility with toxoid by co-sedimentation of protein with gel matrix and also elicited a strong immune response, confirmed by antibody titre studies.Functional confirmation did not seem to be significantly affected by conjugation with adjuvant and Fourier transform infrared spectroscopy was applied to understand it.

Scientists in this study have made a complete profile in ricin vaccine formulation and its influence in conferring active immunity to primitive model systems but the confirmation is to be arrived from human clinical trials as extrapolation of data from mice to humans is difficult .

For reference:”ricin+breakth

Mapping of ricin by tools of molecular biology

Ricin toxin from castor seeds has been well established as a cytotoxin interferring with ribosomal functions.The activity of any protein generally is conferred by the domains present in it. This made scientists to probe the domains and  critical amino acids for the cytotoxic activity of ricin. Researchers in USA have made molecular analysis of castor bean toxin at the genetic level where tools of genetic engineering are exploited to address their domain mapping studies.The experimentation involved synthetic mutation process in genetic determinants of ricin protein for hypothetical amino acid, glutamate to alanine which basically is distant to the former constituent in terms of its physio-chemical properties and makes it an ideal substituent to understand the magnitude of glutamate in the ricin function.Their data in the end from mutant protein’s activity indicated that the target glutamate is not of primary significance in ricin’ toxicological function.

Domain mapping by mutations is an iterative process but an intense technique to make a complete scrutiny of the sub components in a complex molecule and their implications to the ultimate emergence of functions from bulk.

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Interpretation of castor bean and Ricin as reproductive toxins.

Ricin toxin is well to act as a sub- acute toxin in humans to produce lethality between 2-5 days on the basis of concentration and portals of administration of toxin.However its role in reproductive toxicity has not been documented well and Jordanian scientists have made an attempt to assess the role for ricin in reproductive toxicity in pregnant female rats.

Rats intraperitoneally treated with ricin and castor bean extracts displayed symptoms of body weight loss and death of foetuses at 5-9 days of pregnancy while attrition of implantation sites was noticed with first 6 consecutive days of pregnancy.The toxicological function was not significantly reversed with gonadotrophin hormone in experimental subjects exposed to toxin.Hence the researchers were able to conclude that castor bean constituents including ricin have anti implantation and ovulation properties through their overlapped reproductive toxicological studies.

Results of this study is difficult to interpret as multiple parameters including strain,species,sex,age,feed,time,concentration of the toxin,mode of administration,animal size of the experimental study have a profound influence in the outcome of the study.Adopting International standard guidelines in toxicology  will greatly improves the precision and accuracy of the experiment.

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Marker assisted probing of castor toxin.

Molecular fingerprinting of DNA proves to be the most reliable of all biological assays applied today in spite of an extensive application of other biomolecules as markers in different functional evaluation studies.Ricin, Castor toxin profiling is not an exception as researchers from USA have devised a protocol for the detection of ricin in food as an accidental or intentional contaminant on the basis of a DNA based molecular marker, unique to castor.The experimental result indicated a sensitivity of 5 microgram of toxin per unit gram of ground beef which is promising according to scientists. They claim further that their methodology’ sensitivity is well below the threshold oral toxicity levels of toxin.

Molecular marker are highly polymorphic and hence the fingerprint that they generate will display a minimum probability for  mismatch but requires a high technical skill.It also has a critical limitation that any insignificant contamination interfering with experimental template DNA  would readily produce a magnified artifact and lead to misinterpretations.

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Valuable metabolite probe from castor meal

Castor meal is a concentrated residue matter produced after extraction of oil from castor seeds.Its application is so restricted due to ricin and poor molecular biochemical knowledge about the material.A surprising breakthrough has been made by the discovery of galactinol, a carbohydrate derivative associated with raffinose metabolism in plants to an appreciable extent on mass basis.

Experimental designs including chromatography and mass spectrometry were applied to isolate and confirm the purity of galactinol in castor meal.The scientists further stated that it is exciting as research in past in the concerned field is hindered by a lack of availability of commercial source of galactinol.

Galactinol discovery in castor meal illuminates the potential of castor meal which is currently applied as a field manure in most of the castor growing regions.Identification of high value metabolites would tremendously add the commercial value to this product on an international scale.

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Analysis of sub-cellular components in castor seed

Castor seed is an evolved propagative material with a great molecular complexity in endosperm. Biochemical analysis of protein bodies in castor seed endosperm has been made.Protein bodies account for a major fraction in the total protein content in castor seed and is thought to involve in an important role in the mobilization of nutrients during seed development and germination.Scientists from an University in USA have isolated them from endosperm tissues and identified two principal units namely globoids and crystalloids embedded in an amorphous matrix.A protein fingerprint of buffer and detergent soluble matrix and crystalloid fractions from protein bodies was examined and revealed complexes and modified proteins including the toxin Ricin present in the soluble fraction.The principal techniques applied in this experimental study were gel electrophoresis and chromatography for resolution and analytical purposes.

Studies of such type are noteworthy to identify novel members of proteins along with their sub-cellular localization and biological functions.

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Toxicological analysis of ricin protein.

Ricin is a toxic protein in Castor.Italian researchers have conducted an ocular toxicological study of ricin in rabbits.They administered variable concentrations of the toxin onto the rabbit’s eyes and evaluated its toxic function.The experimental study revealed that the toxin induced inflammation response in eyes of experimental animals and they even removed the eyes and noticed microscopic pathological manifestations of the toxin.As a way of identifying the neutralizer to the toxin,they evaluated phosphate buffer and 10% lactose solutions with the former had no effect while latter exhibited a partial toxin neutralization activity.The dead subjects were also analyzed.Results indicated that an internal injury to the organs was also imposed by the toxin which further made scientists to conclude that the inability of the neutralizer solutions was due to the rapid absorption  and circulation of the toxin in the system.

This research information is of immense significance as history has witnessed the castor toxin as a biological weapon in Georgi Markov’ assassination.So research studies would be valuable tools to best combat the toxin under the conditions of emergency.

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