Polyhydroxy-fullerenes as safe emergency treatment for organophosphate poisoning

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Organophosphates (OP) are widely used as insecticides but also as potent chemical warfare agents. OP compounds are rapidly absorbed, interfering with the regulation of neural transmission, and high doses can lead to breathing paralysis and death within minutes of exposure. The current treatments for OP poisoning can also present a risk of toxicity, prompting the search for safer alternatives. Dr Marion Ehrich and her collaborators from the Virginia-Maryland College of Veterinary Medicine have demonstrated that polyhydroxy-fullerenes are safe compounds that could protect against early acute OP toxicity.

Organophosphate (OP) compounds are neurotoxic substances often used in formulations for insecticides and chemical warfare agents. Under conditions of sufficient exposure, the nervous system is targeted by irreversible inhibition of the activity of acetylcholinesterase (AChE). This is an essential enzyme that is involved in the transmission of signals across nerve endings, specifically by breaking down or hydrolysing the neurotransmitter acetylcholine (ACh). Acute poisoning with organophosphates can occur within minutes of exposure, as these compounds are absorbed rapidly via inhalation, ingestion or through contact with the skin. The most common outcome of acute OP exposure is a ‘cholinergic crisis’ – overstimulation at a neuromuscular junction caused by excess ACh – which can lead to neuromuscular blockade and central nervous symptoms, such as breathing paralysis and death. Some cases of OP poisoning can lead to chronic neurological damage, including epilepsy and cognitive impairment.

The advent of 9/11 and other terrorist attacks led many to wonder if countries around the world have adequate defence plans in place in the eventuality of an attack involving the use of chemical weapons. Events in which large numbers of people are exposed, such as those that occurred when an OP compound was released in the subway system of Tokyo in 1995, have prompted the search for safe and stable products that could be applied to the skin by non-medical personnel.

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Naled insecticide molecule (top) and temephos larvicide (bottom) Sanpath Chindathong/Shutterstock.com

Standard treatment of cholinergic poisoning includes the administration of high doses of the anticholinergic drug atropine. However, atropine itself can be toxic and high doses of this agent should not be used unless serious or life-threatening symptoms of poisoning are present. Dr Marion Ehrich and her collaborators from the Virginia-Maryland College of Veterinary Medicine have conducted studies to investigate the beneficial effects of polyhydroxy-fullerene compounds for the treatment of OP poisoning. These are water-soluble, spherical structures containing 60-80 carbons which are capable of sequestering OP compounds, making them unavailable to bind to AChE and cause toxicity. Ehrich’s team conducted several experimental studies, the most recent published in 2020, indicating that topical and intraperitoneal administration of polyhydroxy-fullerenes in rodents did not result in toxicity at the concentrations required for action against OP compounds and that polyhydroxy-fullerenes are capable of decreasing the inhibition of AChE by OP compounds in vitro.

“Acute poisoning with organophosphates can occur within minutes of exposure, as these compounds are absorbed rapidly via inhalation, ingestion or through the skin.”

Fullerene hydroxyl derivatives as free radical scavengers

Since the current treatments and preventative approaches for OP poisoning can result in toxicity, there currently are no products that can be used safely for prophylaxis during threats of mass exposures, including substances that could be administered after exposure but before the onset of toxicity. The Ehrich laboratory has years of experience using fullerenes both as sequestering agents and as antioxidants. OP-induced toxicity can have a significant oxidative stress component, and fullerenes can be engineered into derivatives that have high antioxidant activity, acting as ‘free radical sponges’. ‘Native’ fullerenes have spherical structures that can be likened to empty cages made of 60-80 carbon atoms. This native structure makes fullerenes insoluble in water and prone to form aggregates that would be toxic for cells. In order to be made into water-soluble compounds, fullerenes are hydroxylated, which means that hydrophilic structures containing the –OH group are attached to the carbon cage.

OP exposure can cause neuromuscular blockage. This electron micrograph shows a cross-section through the neuromuscular junction (T: axon terminal, M: muscle fiber, arrow: junctional folds with basal lamina).

In 2011, Ehrich and her team published a study reporting on the free radical scavenging activity of polyhydroxy-fullerenes. Ehrich and collaborators monitored the production of free radicals in the presence and in the absence of hydroxyl-fullerenes in vitro. The experiments confirmed that the presence of polyhydroxy-fullerenes resulted in a marked reduction of free radicals in the reaction medium. The researchers confirmed that the scavenging effect was dependent on the concentration of fullerenes used and that the mode of action was consistent with the capture of free radicals rather than enzymatic inhibition of their production. The same study confirmed that polyhydroxy-fullerenes were able to scavenge free radicals, limiting both OP-induced AChE inhibition and the production of substances associated with oxidative stress.

Direct sequestration of organophosphates by polyhydroxy-fullerenes

As discussed above, OP-induced toxic effects can occur within a short time after exposure. Following the results of the 2011 study, the research team conducted a series of experiments to determine the binding affinity between OP compounds and polyhydroxy-fullerenes. The results, published in 2019, showed that the binding of OP compounds to polyhydroxy-fullerenes is transient and non-covalent, in contrast to the covalent, irreversible binding of OP compounds to the AChE active site. This seemed to suggest that fullerenes might have a minor role in reducing symptoms of severe systemic OP toxicity, including toxicity associated with OP nerve agents.

Structure of neurotransmitter, neuromuscular junction, synaptic vesicle, axon and cleft

This prompted the team to conduct more experiments to establish if hydroxyl-fullerenes could be used after dermal exposure to OP compounds when signs of toxicity had not appeared before treatment could commence. The rationale for further experimentation was that when absorbed through the skin, OP compounds have a slower absorption rate, and the use of fullerene derivatives could ameliorate, at least temporarily, the adverse effects of OP-induced toxicity, especially in light of their low toxicity profile when compared to atropine.

“Polyhydroxy-fullerenes are water-soluble spherical nanoparticles, which are effective in delaying and reducing the adverse effects of poisoning with organophosphate compounds.”

Ehrich and her collaborators investigated polyhydroxy-fullerenes for their potential to provide a safe intervention that could be applied topically at or after exposure to OP compounds. The OP test compound used in this 2020 study was paraoxon. Topical route of paraoxon administration was explored as it provides a slower rate of absorption than ingestion or injection. Topical administration also has the advantage of providing a real-world relevant route of OP exposure and a safe and useful route for application of a countermeasure. The researchers observed that topical administration of solubilised fullerenes delayed the onset of paraoxon-induced clinical signs in mice. The polyhydroxyl-fullerenes used were water-soluble and did not generate systemic toxicity after application on the skin. Other investigators reported that, even when given intravenously, the fullerenes did not cause toxicity at doses higher than the 4 mg/kg of topical product applied in the experiments.

The Ehrich laboratory is engineering fullerenes into derivates that have high antioxidant activity.

The researchers pointed out that, given their low toxicity profile, the fullerenes could be safely applied by non-medical personnel to victims that, unknown at the time of exposure, may not be poisoned enough to show cholinergic signs. If such application to potential victims resulted in even a 20-minute delay until the onset of poisoning symptoms, this could provide emergency medical personnel with critical time to provide effective support to seriously ill patients.

Concluding remarks

Polyhydroxy-fullerenes are water-soluble spherical nanoparticles with 60-80 carbons and 14-20 hydroxyl groups, which are effective in delaying and reducing the adverse effects of poisoning with organophosphate (OP) compounds, neurotoxic substances often used as insecticides or as chemical warfare agents. Since OP poisonings often involve dermal exposure, the availability of products with low toxicity profiles, which could be applied to the skin by non-medical personnel, has significant value in cases of mass poisonings, such as agricultural accidents or terrorist attacks. The Ehrich laboratory has published several studies that demonstrated the effectiveness of polyhydroxy-fullerenes against the biochemical manifestation of OP toxicities in cultured cells and tissue homogenates. These water-soluble substances also delayed clinical manifestations of OP toxicities in mice. The mechanism of action behind the protective effect involves the sequestration of the OP compounds within the hollow, cage-like structure of the polyhydroxy-fullerenes, making the OP molecules unavailable to initiate toxicity.

The Tokyo subway sarin attack in 1995.

What are the next steps in your research?

Next steps in research are to promote formulations of safe products that effectively protect against dermal exposure to cholinesterase-inhibiting compounds. The best of these substances will be effective at low concentrations, relatively inexpensive, stable when stored in hot and humid temperatures, and capable of being safely and easily used by non-medical personnel.



  • Ehrich, M., Hinckley, J., Werre, S. R., Zhou, Z. (2020). Effects of polyhydroxyfullerenes on organophosphate-induced toxicity in mice. Toxicology, 445, 152586. Available at: https://doi.org/10.1016/j.tox.2020.152586
  • Ehrich, M., Van Tassell, R., Li, Y., Zhou, Z., Kepley, C. L. (2011). Fullerene antioxidants decrease organophosphate-induced acetylcholinesterase inhibition in vitro. Toxicology in vitro, 25(1), 301–307. Available at: https://doi.org/10.1016/j.tiv.2010.09.010
  • Magnin, G., Bissel, P., Council-Troche, R. M., Zhou, Z., Ehrich, M. (2019). Studies Exploring the Interaction of the Organophosphorus Compound Paraoxon with Fullerenes. ACS omega, 4(20), 18663–18667. Available at: https://doi.org/10.1021/acsomega.9b02587

Research Objectives

Dr Ehrich researches organophosphate (OP)-induced toxicities.


The research was funded by the CounterACT Program, National Institutes of Health, Office of the Director


  • Zhiguo Zhou, Luna Nanoworks


Marion Ehrich , Virginia-Maryland College of Veterinary Medicine, teaches pharmacology/toxicology, conducts toxicology research, and serves as a relief pharmacist and veterinary toxicology consultant. Degrees are from South Dakota State University, University of Chicago, and University of Connecticut. She was president of the Society of Toxicology, received its Merit Award, and served on many national committees.

Marion Ehrich

Marion Ehrich
Virginia-Maryland College of Veterinary Medicine
205 Duck Pond Drive
Virginia Tech
Blacksburg VA 24061

E: marion@vt.edu
T: +1 (540) 231-4938
W: https://vetmed.vt.edu/research.html

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