Compound Interest: An Antidote to Sarin

A rotating molecular model of Sarin. Animation: NaturphilosophieAbout Sarin

Sarin is a deadly compound.  Colourless, odourless, and fatal even at low concentrations. A new drug designed to fight against the deadly effects of organophosphorous nerve agents, like sarin, is in sight. 


Mass Destruction

Sarin was originally developed as a pesticide in Germany in 1938.  The compound was named in honour of its discoverers: Schrader, Ambros, Gerhard Ritter, and von der Linde.

Sarin was further developed as a chemical weapon of mass destruction during World War II.  In 1939, the chemical formula was passed onto the chemical warfare section of the German Army Weapons Office for inclusion into artillery shells.

Estimates for total sarin production by Nazi Germany range from 500 kilograms to 10 tonnes.  However, the agent was never deployed against the Allies.


Some Chemistry…

A 2-dimensional molecular conformation of Sarin isopropyl methylphosphonofluoridate.
The 2D molecular conformation of sarin (isopropyl methylphosphonofluoridate)

Sarin is a man-made nerve agent, also known as GB, phosphonofluoridic acid, isoproposmethylphosphonyl fluoride, or isopropyl methylphosphonofluoridate.  Sarin is heavier than air and lingers at ground level.

It is more lethal at higher temperatures, but it degrades faster with a rise in humidity.

Organophosphorous poisons (OP) – including nerve agents, as well as lots of pesticides with widespread civilian use – are built around a central phosphorus atom containing four substituents:

  • one double-bounded oxygen atom,
  • two organic residues (e.g. alkyl- or alkoxy-groups) and
  • one leaving group whose composition might be highly variable (e.g. fluorine, cyano, para-nitrophenyl).


Chemical Structure

The chemical structure determines stability and toxicity of the compound, mainly influenced by the nature of the leaving group.

In terms of biological activity, its mechanism mimics that of some commonly used carbamate insecticides, and some medicines used to treat such diverse conditions as muscle weakness, glaucoma and delayed gastric emptying.

Sarin is a colourless, odourless liquid or gas, and it is often fatal.

Even at very low concentrations, sarin attacks the nervous system by interfering with the electrochemical signals between the central nervous system and muscle fibres.

Death can result from sarin exposure within 1 to 10 minutes.


To understand why that is, we first need to get to grips with what goes on during skeletal muscle contraction…


Understanding Skeletal Muscle Contraction and the Synaptic Cleft

A time-lapse multiple exposure photograph showing a European tree frog jumping from a blade of grass into the air. Photograph: MNN
Muscle contractions underlie movement: Contractions of skeletal muscles allow vertebrate animals such as this European tree frog (Hyla arborea) to move. Source:

In vertebrates, skeletal muscle contractions are neurogenic as they require a synaptic input from motor neurons to produce muscle contractions.

The contraction produced can be described as a twitch, summation, or tetanus, depending on the frequency of the action potentials.

A neuro-muscular junction is a chemical synapse formed at the intersection of a motor neuron and a muscle fibre.  It is the site in which a motor neuron transmits a signal to a muscle fibre to initiate muscle contraction.

In a healthy nervous system, nerve cells carry information by sending electro-chemical impulses to the muscles and organs via neurotransmitters.

Neurotransmitters bind with a receptor and send an impulse to the muscle before being broken down by an enzyme, thereby stopping the process.


Muscular Contraction: A Chain of Chemical Events


A diagram describing the electro-chemical process of skeletal muscle contraction at the synaptic cleft.
Skeletal muscle contraction begins at the synaptic cleft. Source:

The sequence of events that results in the depolarization of the muscle fibre at the neuromuscular junction begins when an action potential is initiated in the cell body of a motor neuron.  The action potential is then propagated by saltatory conduction along its axon towards the neuromuscular junction.

Once it reaches the terminal bouton, the action potential causes a Ca2+ ion influx into the terminal by way of the voltage-gated calcium channels.  The Ca2+ influx causes synaptic vesicles containing the neurotransmitter acetylcholine to fuse with the plasma membrane, releasing acetylcholine into the synaptic cleft between the motor neuron terminal and the neuromuscular junction of the skeletal muscle fibre.

Acetylcholine diffuses across the synapse and binds to and activates nicotinic acetylcholine receptors on the neuromuscular junction.  Activation of the nicotinic receptor opens its intrinsic sodium/potassium channel, causing sodium to rush in and potassium to trickle out.

As a result, the sarcolemma reverses polarity and its voltage quickly jumps from the resting membrane potential of -90mV to as high as +75mV as sodium enters.  The membrane potential then becomes hyperpolarized when potassium exits and is then adjusted back to the resting membrane potential.


Exposure to Sarin and Organophosphorous Poisons

Two photographs showing the dramatic effect of Sarin on the pupil of a rabbit's eye.
Pupil constriction is one of the symptoms of the nervous system overstimulation due to sarin exposure.  Source: Journal of Medical, Chemical, Biological, and Radiological Defense

The class of organophosphorous poisons (OP) comprises nerve agents as well as many pesticides licensed for widespread civilian use.

As a nerve agent, sarin is a highly toxic and deadly compound.  High purity OP are nearly odourless and colourless.  Therefore, a compound like sarin is hardly ever recognised sensorily, thus allowing unnoticed incorporation, which contributes to its special risk potential.

Lipophilicity – the ability of a chemical to dissolve in lipids – enables rapid resorption by penetration of the skin (percutaneous incorporation).

Intake of aerosolized and gaseous sarin via the respiratory tract by inhalation, as well as incorporation through the eyes represent realistic routes of poisoning in military and terrorist attack scenarios.

Gastro-intestinal invasion is more typical of pesticide poisoning in suicidal attempts by swallowing liquid solutions of the OP compounds.  (Indeed, over 200,000 fatal cases per year of pesticide poisoning are observed worldwide – most of them occurring in rural areas of developing countries.)

Intravenous intake is typical of controlled toxicological studies on animals.  (See next section.)


Pathophysiology at the Neuromuscular Junction

A diagram explaining the molecular toxicology of organophosphorus poisons. Source: Medical Corps International Forum
Molecular Toxicology of Organophosphorous Poisons: Patophysiology and Bioanalysis Source: Medical Corps International Forum

Sarin attacks the nervous system by interfering with the degradation of the neurotransmitter acetylcholine (ACh) at neuromuscular junctions.  In vertebrates, acetylcholine is the neurotransmitter used at the neuromuscular junction, where signals are transmitted between neurons from the central nervous systems to the muscle fibres – the synaptic cleft.

Death usually occurs as a result of asphyxia due to the sudden inability of the metabolism to control the muscles involved in the breathing function.

Specifically, sarin is a potent inhibitor of acetylcholinesterase (AChE) – an enzyme that degrades the neurotransmitter acetylcholine after it is released into the synaptic cleft.

Acetylcholine is normally released in the body from the neurons to stimulate the muscles, after which it becomes degraded by acetylcholinesterase, thereby allowing the muscle to relax.

However, a build-up of acetylcholine at the synaptic cleft, due to the inhibition of cholinesterase, means the neurotransmitter continues to act on the muscle fibre, so that any nerve impulses are effectively continually transmitted.

This causes a deadly over-stimulation of the nervous system.

Sarin poisoning causes visual disturbance, frothing from the mouth, vomiting, and serious breathing difficulties.  Eventually, untreated sarin exposure leads to death.


Sarin Toxicity

A black and white photograph showing sarin testing being carried out on a caged rabbit.
Sarin toxicity was determined using animal testing. Source: Wikimedia

Sarin is highly toxic, whether by respiratory or dermal exposure.  The toxicity of organophosphorous poisons is based on the chemical derivatization (or phosphylation) of AChE.

OP undergo a nucleophilic substitution reaction with AChE.  While binding to the hydroxyl-group of the serine amino acid residue of the catalytic triade of the esteratic centre, the leaving group of the OP is released.

Phosphylation causes enzyme inhibition thus preventing esteratic cleavage (deactivation) of the neurotransmitter at the synaptic clefts.  As a result, agonistic ACh binding to muscarinic and nicotinic receptors causes permanent over-stimulation of effector cells, and thus cholinergic crisis.


Clinical Symptoms

The clinical signs and symptoms include miosis and muscle fasciculation, as well as excessive secretion of diverse body fluids like saliva, sweat, urine and tears, which is considered as the most important clinical indicator for AChE inhibition.  The fatal outcome results mainly from central and peripheral respiratory paralysis.

The toxicity in humans is determined on calculations from studies involving animal subjects.

The lethal concentration of sarin in air is approximately 35 mg per cubic metre per minute for a two-minute exposure time by a healthy adult breathing normally (i.e. exchanging 15 litres of air per minute).

This number represents the estimated lethal concentration for 50% of exposed victims, the LCt50 value.


Comparison with Other Toxins

There are many ways to make relative comparisons between toxic substances.  The list below compares some current and historic chemical warfare agents with sarin, with a direct comparison to the respiratory Lct50:

 – Sarin is 28 times more lethal

  • Phosgene, 1500 mg-min/cubic meter

 – Sarin is 43 times more lethal

  • Hydrogen cyanide, 2860 mg-min/cubic meter

 – Sarin is 81 times more lethal

  • Chlorine, 19000 mg-min/cubic meter

 – Sarin is 543 times more lethal !!


The Antidote?

A rotating model showing sarin residues targeted by mutagenesis and their relation to its proposed structure.
The integration of DFT calculations with X-ray crystallography is a powerful combination that provides unexplored possibilities for computational chemistry and structural biology. The combination of techniques allowed us to determine a structure that may provide a starting point for analysis of reaction pathways and structure-based design of improved nerve agent antidotes. Source: Allgardsson et al (2016) – Click to animate the picture –

Even at very low concentrations, sarin can be fatal.  After direct inhalation of a lethal dose, death may follow within 1 to 10 minutes.

However, drugs against nerve agent poisoning have been used for a long time.  The over-stimulation of the nervous system can be stopped if an antidote is administered quickly.

Typically, those antidotes are atropine and pralidoxime.  Effective treatment of poisoning with nerve agents requires immediate administration of such antidotes.

Drugs are applied in auto-injectors – medical devices that allow the user effective self-treatment by automated intramuscular administration, and are an essential part of a soldier’s military kit.

Still, it has until now been unclear how they actually work.


Optimising the Dose

As we have seen, organophosphorous nerve agents destroy the function of acetylcholinesterase in the nervous system.  As long as the nerve agent is bound to the protein, the breakdown of an important signalling substance is prevented.

Optimisation of drug doses is an objective of pharmacokinetic studies that require time-dependent quantification of therapeutics in blood (plasma or serum).  Detailed knowledge of Administration, Distribution, Metabolism, and Excretion of drugs (ADME) helps to improve the chemical structure of antidotes, advance therapeutic success and speed up physical reconstitution.

As soon as OP have entered the body, the compounds are systemically distributed and undergo diverse chemical reactions that either cause poisoning on the molecular level, as described above, or can induce detoxification by hydrolysis or binding to scavenger molecules.

A computer model showing the interaction between sarin and its antidote HI6.
A model of how sarin and HI-6 are positioned in the protein acetylcholinesterase (AChE) just before HI-6 removes sarin and restores the function of the protein. The model was developed by a combination of X-ray crystallography and quantum chemical calculations. Sarin is coloured in magenta, HI6 in green, oxygen in red, phosphorus in orange and nitrogen in blue. Source: FOI, Swedish Defence Research Agency

All therapeutics and poisons, as well as their bio-transformation products, pass through the liver and kidneys, and are nearly exclusively excreted in urine.

The elucidation and monitoring of toxicological ­processes demand ­qualitative and quantitative determination of the relevant compounds.

Recently, a ground-breaking study was published in PNAS (Proceedings of the National Academy of Sciences), which describes in detail how such a drug works. Researchers at the Swedish Defence Research Agency, Umeå University and in Germany are behind the study.

The antidote HI-6 removes the nerve agent, and restores the normal function of the nervous system.

After seven years work using a variety of techniques, Anders Allgardsson and his team have finally been able to bring this to a successful close and can show a uniform picture of how HI-6 approaches sarin.

It opens up new opportunities of finding antidotes to sarin and other nerve agents by structure-based molecular design.

A photograph showing the grief-stricken relatives of the victims of the Eastern Ghouta chemical attack.
The tragic aftermath of the chemical weapons attack on the Damascus suburb of Ghouta, Syria, on Wednesday, August 21, 2013. The bodies of men, women and children victims, killed by nerve gas, lie-in-wait before the mass burial. The World was outraged, but not enough to take military action…  Image:

As of April 1997, the production and stock-piling of sarin was outlawed by the Chemical Weapons Convention of 1993, and classified as a Schedule 1 substance.  Nevertheless, the use of sarin in an attack on civilian populations was still being recorded as recently as in 2013, during the ongoing Syrian civil conflict, causing a death toll between 322 to 1,729 according to different sources.

Nerve agents are without doubt dreadful weapons.

The hope is that these results will lead to the creation of new improved drugs against them.