This review (2003) examines the chemical, biological, and toxicological properties of the alkaloids contained in Amanita Muscaria (Fly Agaric) alongside the sociocultural context of its etymology. The principal substrates ibotenic acid and muscimol exert their psychotropic effects through stimulation of inhibitory glutamatergic and GABAergic neurotransmission.
The fly agaric is a remarkable mushroom in many respects; these are its bearing, history, chemical components and the poisoning that it provokes when consumed. The ‘pantherina’ poisoning syndrome is characterized by central nervous system dysfunction. The main species responsible are Amanita muscaria and A. pantherina (Amanitaceae) ; however, some other species of the genus have been suspected for similar actions. Ibotenic acid and muscimol are the active components, and probably, some other substances detected in the latter species participate in the psychotropic effects. The use of the mushroom started in ancient times and is connected with mysticism. Current knowledge on the chemistry, toxicology, and biology relating to this mushroom is reviewed, together with distinctive features concerning this unique species.
The fly agaric (Amanita muscaria) and the closely related A. pantherina are the principal species implicated in the so-called 'pantherina-muscaria' poisoning syndrome, a central nervous system disorder that is usually non‑fatal but can produce marked neuropsychiatric effects. Earlier work established the presence of isoxazole derivatives (notably ibotenic acid and muscimol) as the primary bioactive constituents, but uncertainties remained about the full chemical complement of the species, the biological mechanisms of action, variation in toxin content, and the cultural uses of the mushroom. Michelot sets out to synthesise current knowledge across multiple domains — medical, chemical, pharmacological, biological and ethnomycological — with particular emphasis on the chemistry and biological properties of the substances found in A. muscaria and related taxa. The review assembles clinical descriptions of poisoning, experimental toxicology, methods for detecting active compounds, pigment chemistry, and the mushroom's historical and cultural roles to provide a comprehensive account of its distinctive features.
Papers in Blossom that reference this study
Winkelman, M. J., Szabo, A., Frecska, E. · European Neuropsychopharmacology (2023)
Lerner, A. G. · The Israel Journal of Psychiatry and Related Sciences; Jerusalem (2015)
Geml, J., Laursen, G. A., Nusbaum, H. C. et al. · Molecular Ecology (2005)
This paper is a narrative review that compiles published mycological, chemical, pharmacological, toxicological and ethnographic findings relating to Amanita muscaria and A. pantherina. The extracted text indicates the authors drew on a broad range of primary studies, animal experiments, analytical chemistry reports and historical/ethnographic sources to construct their synthesis. The extracted material does not provide an explicit description of search strategy, inclusion/exclusion criteria, databases searched, date ranges, or formal methods for study selection or quality assessment. Instead, the review appears to be an expert synthesis organised by topic (species identification, clinical syndrome, chemical constituents, analytical detection, mode of action, pigments, miscellaneous biochemistry and ethnomycology) rather than a systematic review with reproducible methods.
Species and poisoning characteristics: The review identifies Amanita muscaria and A. pantherina as the main species responsible for the pantherina‑muscaria syndrome; a few other Amanita species have been suspected in some reports. A. muscaria is described as conspicuous (bright red, orange or yellow cap with white flecks) and mycorrhizal with trees such as birch and pine; A. pantherina has a grey to grey‑brown cap with white flakes and has been mistaken for edible species in some accidental poisonings. Clinical onset after ingestion is typically 30 minutes to 2 hours, with a period of confusion, dizziness, hypersensitivity of the senses, space and time distortion, pupillary dilatation and dry mouth; an initial period of excitation or altered cognition is often followed by marked drowsiness or deep sleep, and most poisoning episodes resolve within about 24 hours. Fatalities are described as very rare. Treatment and legal status: Management is primarily symptomatic. Gastrointestinal decontamination (induced emesis, gastric lavage, activated charcoal) is recommended early. The authors emphasise that atropine is contraindicated; physostigmine (eserine) has been recommended as an antidotal cholinesterase inhibitor in some cases. Sedatives such as diazepam or clonazepam have been used for convulsions, but diazepam has been suspected of potentiating muscimol's actions in some reports. Cooking does not reliably abolish toxicity because the active compounds are water‑soluble and may survive common preparation techniques. Legal control varies geographically; A. muscaria is not widely controlled in Europe, although some countries regulate hallucinogenic mushrooms and isolated psychoactive compounds. Primary chemical constituents: The review concludes that ibotenic acid and its decarboxylation product muscimol are the principal biologically active principles in A. muscaria and A. pantherina. Both are isoxazole derivatives: ibotenic acid is structurally related to glutamate and muscimol to GABA. Other related compounds described include muscazone (a photorearrangement product of muscimol with minor activity), tricholomic acid, stizolobic and stizolobinic acids (reported mainly in A. pantherina), and several additional amino‑acid derivatives. Muscarine is present only in minute amounts in these Amanita species and is not considered responsible for the characteristic syndrome. The authors also report a range of minor or disputed constituents (choline, acetylcholine, betaine, tropane alkaloids, bufotenine, and traces of amatoxins and phallotoxins reported by some assays) but note that many of these findings are contested or occur at trace levels. Pigments and miscellaneous biochemistry: Pigment chemistry is elaborated: muscarufin, muscaflavin and a series of betalain‑type muscaaurins are implicated in the red–orange coloration of caps. The mushroom is also notable for concentrating certain metals and inorganic complexes: high levels of vanadium (the vanadium complex amavadin), selenium and various heavy metals have been reported. Biologically active polysaccharides (a branched beta‑D‑glucan with reported antitumour activity in mice), lectins and haemolysins have been isolated from Amanita species. Analytical detection and toxin content: Multiple analytical methods have been described for detection and quantification, including paper chromatography, high‑performance thin‑layer chromatography (HPTLC), HPLC (including ion‑interaction and reverse‑phase methods), automated amino‑acid analysis and GC‑MS of derivatised muscimol. Reported concentrations vary by method, part of the fruit body and preparation. HPTLC values reported in the text include muscimol at about 0.19% dry weight in A. muscaria and 0.3% in A. pantherina, while muscarine is very low (about 0.009% in A. muscaria and <0.0005% in A. pantherina). Ion‑interaction HPLC values given are ibotenic acid approximately 990 mg kg‑1 in caps and 230 mg kg‑1 in stems, muscimol approximately 380 mg kg‑1 in caps and 80 mg kg‑1 in stems; a fresh average fruit body (60–70 g) may contain up to 70 mg ibotenic acid according to the review. The authors emphasise marked variability due to season, geographic origin and preparation (peeling, boiling, drying) because ibotenic acid can be decarboxylated to muscimol during processing or gastric digestion. Pharmacology, in vivo effects and mechanism of action: Ibotenic acid acts as an agonist at excitatory glutamate receptors (ionotropic glutamate receptors) and muscimol is a potent GABA‑ergic agonist; both can cross the blood–brain barrier and mimic endogenous neurotransmitters. Animal data summarised include oral muscimol active doses of ~7.5–10 mg, intraperitoneal LD50 values for muscimol of about 2.5 mg kg‑1 in mice and 3.5 mg kg‑1 in rats, and other dose‑dependent effects such as sedation, hypnosis, muscle twitching, pupil dilation and anorexia. Experimental work shows that repetitive ibotenic acid exposure can alter GABA receptor sensitivity and may produce selective neuronal lesions in basal forebrain cholinergic cells with lasting learning and memory deficits in animals. Ibotenic acid and muscimol can also alter brain monoamine levels (serotonin and dopamine), which may contribute to some clinical features. Ethnomycology and iconography: The review compiles ethnographic and historical material indicating sporadic ritual use of A. muscaria in northern Eurasia (for example among some Siberian tribes) and wide cultural prominence of the species in art, iconography and folklore. The authors report archaeological and iconographic suggestions of ancient mushroom use in several regions but stress that, unlike psilocybin mushrooms in Mesoamerica, A. muscaria has not been a dominant object of religious veneration in most traditions.
Michelot and colleagues interpret the accumulated evidence to position A. muscaria as a chemically distinctive and culturally prominent mushroom whose primary psychotropic effects are due to the interplay of ibotenic acid and muscimol. They emphasise that these compounds form a functional entity: ibotenic acid is the predominant fungal constituent while muscimol, produced by decarboxylation, is the principal neuroactive agent encountered after preparation or digestion. Relative to earlier research, the review consolidates pharmacological and analytical advances that clarify why classical muscarinic poisoning is not the principal mechanism for these Amanita species; muscarine plays only a minor role. The authors highlight important uncertainties and contested findings: reports of amatoxins, phallotoxins, bufotenine and tropane alkaloids are inconsistent across studies, and toxin content shows pronounced intraspecific and geographic variation. Likewise, many ethnographic and iconographic claims concerning antiquity and ubiquity of ritual use remain interpretive and not uniformly corroborated. The paper acknowledges limitations inherent in the evidence base: much of the toxicological work is experimental or veterinary and human data are largely observational case reports; the review itself does not report a systematic literature search, so comprehensiveness cannot be confirmed from the extracted text. Clinically, the authors caution that while typical poisonings do not produce organ failure, repeated exposure in animals can cause neuronal damage, suggesting potential long‑term risks to humans with chronic consumption. In practical terms, the authors recommend symptomatic management and intensive‑care support when needed, warn against the use of atropine, and note that common culinary processing does not reliably eliminate activity. They also observe that, despite the species' notoriety, A. muscaria is not a major target of drug control or trafficking in most jurisdictions and public awareness of its pharmacology and risks is limited.
Amanita muscaria occupies a singular place among fungi: its striking appearance and cultural symbolism are paralleled by a distinctive chemical profile dominated by ibotenic acid and muscimol. Consumption of A. muscaria and A. pantherina typically produces a transient central nervous system syndrome that is rarely fatal and usually resolves without organ damage, but experimental data indicate the possibility of neuronal injury after repeated exposure. The mushroom has driven advances in chemistry and pharmacology, yet it remains comparatively less regulated and less widely used recreationally than classic psilocybin‑containing species. Given the variability in toxin content and the potential for neurotoxicity with recurrent use, the authors recommend caution and appropriate clinical management for poisoning cases.
The fly agaric, Amanita muscaria, and the panther, A. pantherina, are the species mainly involved in the 'pantherina-muscaria' poisoning syndrome. Poisoning cases are sometimes accidental and mainly those caused by A. pantherina, since this mushroom might be mistaken for some other species; but, in some cases, intentional consumption of A. muscaria occurs for recreational purposes. Prognosis of the poisoning is generally minor ; although, very seldom lethal cases are mentioned. Central nervous system dysfunctions primarily characterize this poisoning. This review describes the unusual features associated with these species, medical, chemical, pharmacological, historical and phytogeographical and aims to synthesize current knowledge on these matter. The emphasis is, however, the chemical and biological properties of the substances found in the mushroom.
Detailed mycological data have already been published. The main responsible species are Amanita muscaria (Fig.) and A. pantherina. The cap of A. muscaria can be 50 cm diam and bright red, orange, or even orange or yellow, apart from the white fleck. Many species of the A. muscaria complex bear so-called crassospores. The species cannot be mistaken for any other perhaps except the edible A. caesarea. A. pantherina is also known as: 'panther cap ', 'panther Amanita ', 'panther agaric ' (North America), 'pantherschwamm ', 'kroˆtenschwamm ' (Germany), 'tignosa' (Italy), 'pantera ', ' pixaca ', ' hongo loco ', ' hongo malo ' (Spanish America) and 'amanite panthe`re' (French). The cap of A. pantherina is 5-10 cm diam and grey or grey-brown, grey yellow, paling when old, with small pure white flakes. This species, in some accidental poisoning cases, has been mistaken for the edible A. rubescens and A. spissa. These two species occur in many continentsand grow in deciduous woods, especially beech and birch, and also coniferous ones. Some related species are suspected of poisoning or produce the active components detected in these fungi) : A. regalisand A. strobiliformis. A few cases of poisoning by A. gemmata (Cornue´1961; syn. A. junquillea), A. crenulata), and A. cothurnata have also been reported, but their possible toxic potencies are disputed. Toxin content has been suggested to be related to the place of gathering, indicating chemical intraspecific variation). Surprisingly,mentioned large amounts of chlorocrotylglycine and amino-2-hexadien-4,5-oic acid, both derivatives of glycine, in A. smithiana. Noting the functional identity (' toxicophore ') of these latter substances, an enzyme inhibitor activity has to be suspected ; nephrotoxic poisonings by this species have been reported.
The poisoning syndrome due to Amanita muscaria and A. pantherina has been called 'mycoatropinic ', as the symptoms are similar to those induced by atropinic plants such as Datura stramonium, Atropa belladonna, and Hyosciamus niger, but tropanic alkaloids are not present. Chronosymptomatology is variable amongst subjects, depending on the rituals, and experimental and accidental poisonings. In rituals, the poisoned person (e.g. the shaman) is in search of a peculiar state of mind where autosuggestion is important. A 'delirium ' is not completely created but the chemicals enhance the intellect. Socio-cultural context and environment associated with the psychological and physiological state of the participant are pertinent. The 'pantherina-muscaria ' syndrome is atropine-like and in the number and severity of poisoning cases fatality is rare. In most cases recovery is virtually complete after 24 h without noticeable after-effects. In contrast to Cortinarius, or Gyromitra poisoning syndromes, the fly agaric effects have a short latency like the Coprinus syndrome. In most cases as little as one cap, a cup of saute´ed mushrooms, is a sufficient for psychotropic effects. Symptoms start 30 min to 2 h after ingestion. A state of confusion, dizziness, and tiredness, visual and auditory aesthesia (hypersensitivity), space distortion, and unawareness of time ; presents reinforced by the consumption of some psycho-sedative agents. Aggressive attitudes have not been reported. Dryness of the mouth and mydriasis (dilation of the pupils) have been mentioned. Hallucinations, vivid colour perception and a sense of time standing still, are disputed. A drowsiness period after 2 h follows, with vivid dreams. A deep sleep ends the poisoning, which lasts generally 8 h. Subsequent gastrointestinal disorders with vomiting are inconstantly reported and are not characteristic of the syndrome. No damage to organs has been reported although the active components may induce in vivo brain lesions. Regular consumption of the mushroom would probably be harmful, even though most human poisoning cases do not report any after-effects. Brain lesions in rodents treated with ibotenic acid and muscimol can occur. According to, the biological effects are related to the period of collection, those collected in September induce more marked nausea and less narcotic and visionary experience than those collected in August. Mushroom intoxication in dogs has been reported; the chronosymptomatology is consistent with that in humans (Naude´&. The symptomatology and chronology of poisoning events are summarized in Fig..
The appropriate treatment is mainly symptomatic. First, eliminate the toxic substances from the gastrointestinal tract out by vomiting (e.g. administration of ipecac syrup ; Benjamin 1992), stomach washout, or the administration of activated charcoal and purging, preferably in an intensive care emergency unit. The actions of the active principles, ibotenic acid and muscimol, have to be compared with that of atropine; consequently the use of the latter drug is strongly contraindicated. Physostigmine (eserine), a cholinesterase inhibitor, has been recommended as it counteracts the effects of poisoning by atropine and related antimuscarinic drugs. The intravenous dosage for adults and adolescents is 1-2 mg repeated when needed, the drug has also been used successfully in veterinary cases. The administration of sedatives such as diazepam or clonazepam, by mouth or intravenously, in case of convulsions, as well as phenobarbitone has been suggested. However, diazepam has been suspected as strengthening the action of muscimol). Contrary to some statements, cooking does not notably lower toxicity, demonstrating that the active components are not heat sensitive.
About 12 species of mushrooms are consumed for recreational purpose, but seldom Amanita muscaria (Samorini 1992). The Italian courts proscribe the use of psilocybin, psilocin and A. muscaria as well (' Stampa alternativa '). However, in Europe, interest in this type of drug is lower than in North America, where laws prohibit its use. The use of A. muscaria, even in Europe, is nevertheless growing. In some countries, possession of hallucinogenic mushrooms is not illegal, contrary to their purified substances and derivatives. In Europe, the current legislation mainly concerns Psilocybe semilanceata, and to a lesser extent A. muscaria.
The fly agaric represented a challenge to chemists, biochemists, and biologists. The goals were: (1) the detection and mode of action of the substances involved in the psychotropic effect that might ultimately lead to the design of new drugs; (2) the isolation and chemical identification of the pigments responsible for the characteristic hue of the cap ; and (3) the discovery of miscellaneous unusual components. Amazingly, most substances produced are chemically and biogenetically interrelated, a common molecular architecture linking most of the structures.
Although the chemical constituents of this mushroom have been thoroughly investigated, a few details remain unclear ; they concern the relationships between the physicochemical properties of the active components and the mode of consumption. For instance, Mexican people eat the carpophore of Amanita muscaria without the cuticle that has been peeled off, and also discard the cooking water. In Italy, after boiling and rejecting excess water, the mushroom is preserved in brine prior to consumption. In North America, the red cuticle is peeled off, and the remainder dried, and then smoked. These latter procedures would eliminate or destroy the greater part of the active water-soluble substances. Muscimol and ibotenic acid are most likely to be the biologically active principles of the species of concern, but other active components are suspected. Different research groups performed almost simultaneous isolation and structure determination of the active components. The substances are isoxazole derivatives : ibotenic acid 1 (from 'iboten-gu-take ', the Japanese name of A. strobiliformis, previously called 'premuscimol ' ;, its decarboxilation gives muscimol 2 ; these compounds are only reported in A. muscaria and A. pantherina subspecies. Ibotenic acid (pantherin, agarin), i.e. a-amino-3hydroxy-5-isoxazoloacetic acid of the cooked mushrooms, or even after gastric digestion, would only contain the latter substance, and be the agent responsible for the main symptoms. The red skin of the cap and the yellow tissue beneath contain the highest amounts of these substances, so explaining practices in removing it. Muscimol, i.e. 5-(aminomethyl)-3-hydroxyisoxazole 2, isolated from A. muscaria, is colourless (crystals, C 4 H 6 N 2 O 2 , mol wt 114.10, mp 175 x decomposition) readily soluble in cold water. The structure of pantherine, isolated from A. pantherina, was found to be similar to that of 'agarin '. Muscazone, i.e. a-amino-2,3-dihydro-2-oxo-5oxazoleoacetic acid 3, is colourless (crystals, C 5 H 6 N 2 O 4 , mol wt 158.11, mp 190 xC decomposition) and was also detected and the structure confirmed by synthesis. This lactame isomer of muscimol would result from photo-rearrangement of the latter structures, and is produced during isolation. Muscazone exhibits minor pharmacological activities in comparison with the previous substances. Interestingly, cycloserine (myxomycin, seromycin, D-4-amino- 3-isoxazolidone) is an antimicrobial tuberculostatic agent that exhibits a similar carbon backbone as muscimol. It is known to induce effects on the central nervous system, but with a longer latency period : somnolence, confusion, and nervousness distinguish untoward effects) (Fig.). Several syntheses of ibotenic acid and muscimol have been developed via different routes. Thio-derivatives and even more potent inhibitors of GABA uptake have been prepared. Gram-scale preparation has been performed by regiospecific 1,3-dipolar cycloaddition/elimination. Thiomuscimol derivatives have been prepared ; they exhibit highly potent binding properties even able to displace GABA from its receptor. Other active substances (x)-R-4-hydroxy-pyrrolidone-(2) 4, whose structure is closely related to ibotenic acid and muscimol, was found in Amanita muscaria. This chemical frame is common in some micromycetes; they generally exhibit a potent biological activity against bacteria and other fungi (e.g. aureothrycin, equisetin). This additional compound, gives information on the biogenesis of ibotenic acid, muscimol, and muscazone. All probably originate from the same precursor, b-hydroxyglutamic acid ; ring closures and decarboxilation then determine the structures of these products (Fig.). Tricholomic acid 5, a closely related dihydro derivative of ibotenic acid, has been identified in Tricholoma muscarium, and its structure was confirmed by synthesis) (Fig.). Stizolobic 6 and stilozobinic acids 7, known in Stizolobium hasjoo, have been found in A. pantherina, but not A. muscaria, low levels have been detected in A. gemmata. Stizolobic acid, and to a lesser extent stizolobinic acid, exhibit an excitatory action in an isolated rat spinal cord. Biosynthetic studies show that both these amino acids originate from 3,4-dihydroxyphenylalanine (DOPA), and the enzymes implicated have been isolated and purified) (Fig.). Muscarine has long been considered as the active principle in A. muscaria, due to a presumed action on the central nervous system. This was after discredited. Nevertheless, some publications still wrongly assert muscarine's involvement in the action, although several analyses have demonstrated that this agent, which occurs in minute amount in the Amanita's of concern, is not responsible for the poisoning syndrome. The occurrence of only small amounts of muscarine 8, i.e. tetrahydro-4-hydroxy-N,N,N-5-tetramethyl-2-furanmethanaminium, (0.0002-0.0003%, f.w.; chloride: stout prisms, mp 180-181 x, readily soluble in water) have been established in A. muscaria. They have to be considered minor components in comparison to some Inocybe spp. (to 0.43 % in I. subdestricta) and Clitocybe spp. (to 0.15 % in C. dealbata). The two Amanita species would not be able to induce any muscarinic syndrome without excessive consumption. Equal quantities of the muscarine, showing the (+)-(2S,3R,5S) conformation, and (+)-(2S,3S,5S) epi-muscarine 9 have been detected in A. pantherina), as well as (x)-(2S,3R,5R) allo-muscarine 10 in A. muscaria. Although its occurrence was not demonstrated unambiguously, (+)-(2S,3S,5R) epiallo-muscarine 11 was not present in detectable amounts. Muscarine 8, epi-muscarine 9 and allo-muscarine 10 have been detected in the mycelium of A. muscaria culture. Chemical syntheses of these compounds have been performed : epiallo-muscarine 11, DL-muscarine and DL-allo-muscarine) (Fig.). Two additional active components reported in A. pantherina are (2R),(1R)-2-amino-3-(1,2-dicarboxyethylthio) propanoic acid 12 and (2R),(1S)-2-amino-3-(1,2-dicarboxyethylthio) propanoic acid 13. These are antagonists of N-methyl-Daspartic acid (NMDA) receptors KI-II-A and KI-II-B respectively. Actions of these substances could be an alternative explanation of some additional effects observed during Amanita poisoning (Fig.). Other active constituents detected in A. muscaria include choline, acetylcholine, betain, muscaridin, a quaternary trimethylammonium salt of 6-amino-2,3dihydroxy-hexan, hercynin, (x)-R-hydroxy-4 pyrrolidone, uracile, hypoxanthin, xanthin, adenosin, a carbolinic derivative, and b-D-n-butylglycopyranoside. Minor amounts of tropan alkaloids, atropine, hyoscyamine, scopolamine, and bufotenine have also been reported, although their occurrence is rejected by others. The occurrence of muscarine, muscaridine and choline was confirmed, when neither 1-hyoscyamine nor atropine were detected in Dutch specimens). The possible occurrence in A. muscaria of amatoxins and phallotoxins, the typical toxins of A. phalloides, were at first discounted). However, highly sensitive detection methods, radio immunoassays, demonstrated traces of amatoxins, and phallotoxins are also suspected. Another possibly active compound, bufotenine (5-hydroxy-N,N-dimethyltryptamine), has been proposed) but the occurrence of indolic hallucinogenic substances such as psilocybine frequently speculated, has not been confirmed.
The bright colour of Amanita muscaria has prompted investigations in several research groups; nowadays most of the structural features of the pigments have been elucidated. Muscarufin 14 is the principal component ; it is probably a putative derivative of a terphenylquinone; but efforts by different groups to reproduce these evidently premature results failed) (Fig.). Musso also proposed a quite novel structure, muscaflavin 15, for a yellow pigment, a result corroborated by a biosynthetic pathway that includes prior divergent oxidative cleavages of L-DOPA, further cyclization steps generating muscaflavin together with betalamic acid, stizolobic acid, and stizolobinic acid. The structure has been confirmed by synthesis). Muscaflavin has also been detected in red-fruited Hygrocybe species) (Fig.). Different compounds, the muscaaurins I-VII 16, have been proved to be the pigments responsible for the characteristic red-orange colour of the caps of several of other Amanita species such as A. caesarea). An elegant chromatographic procedure enables the separation of these pigments). Interestingly, ibotenic and stizolobic acids are essentially elements added to the whole structure of muscaaurins. The muscaaurins are betalaines and also originate from the transformation of DOPA. Muscapurpurin (purple) and muscarubin (red-brown), whose structures are closely related to the muscaaurins, have been isolated from Amanita muscaria) (Fig.).
Dioleine 1,3 17, a-diester of glycerol and oleic acid ; has been considered the putative fly attractant action of Amanita muscaria) (Fig.). Mushrooms are known for differential metal bioaccumulation. A. muscaria may concentrate vanadium to 200 ppm (30 times those reported in living organisms). Isolation, structure determination and synthesis of the pale blue vanadium complex, amavadin 18, was confirmed by crystallography. Similarly, unusually high levels of selenium (to 17.8 ppm; Watkinson 1964) and heavy metals have been reported : cadmium 13.9, cobalt 2.6, chromium 1.7, lead 33.3, mercury 61.3, and nickel 7.5 ppm d.w.) (Fig.). A lectin (APL) has been isolated from A. pantherina through Fast Protein Liquid Chromatography (FPLC), the molecular mass was estimated at 43 kDa and consists of two identical subunits. A b-(1p6) branched (1p3)-beta-D-glucan (AM-ASN) was isolated from A. muscaria. AM-ASN exhibited antitumour activity against Sarcoma 180 in mice. Phallolysin, a typical haemolysin initially found in A. phalloides and able to agglutinate red blood cells has also been detected in A. muscaria.
Several techniques provided information on the active components in fungi ; they are useful for both forensic science and chemiotaxonomy. Due to efforts to obtain pure muscarine, the estimation of the total muscarinic activity and potency of mushrooms was formerly performed by bioassays on rats (salivation and lacrymation ;. Detection and quantitative analysis of muscarine in Inocybe species has been undertaken by paper chromatography (eluent: n-butanol/methanol/water 10/3/2 v/v, staining Thies & Reuther's 1954 reagent ;. The isomer ratios were detected in the same genus by gas chromatography of the norbases. Muscarine has been identified in the red caps of Mexican fly agarics by cellulose column chromatography and identification as muscarinetetrachloroaurate. Isoxazole derivatives have been explored in various Amanita species by two-dimensional paper chromatography (eluents : 1, n-butanol/acetic acid/water 1/1/1; 2, butanol/pyridine/water 1/1/1 v/v, staining : ninhydrin ;. Ibotenic acid and muscimol were detected in A. muscaria and A. pantherina, and in intergrades of A. pantherina and A. gemmata, but not in A. strobiliformis by this method. Some probable isoxazole derivatives were detected in A. solitaria. Subsequently,suggested that the latter substances are chemiotaxonomical markers for the differentiation of species and even subspecies. High performance thin layer chromatography (HPTLC) on silica gel allows the detection of muscarine (eluent: n-butanol/ethanol 95 %/acetic acid/ water 80/20/10/30, chromogenic reagent: modified Dragendorff ) and muscimol (eluent 2-butanol/ethanol 95 % acetic acid/water 75/25/5/25, a chromogenic reagent often used for amino acids). HPTLC permits an evaluation of muscimol equivalents since the experimental procedure, extraction with formic acid, facilitates the decarboxilation of ibotenic acid. The levels of muscimol detected in the samples were of 0.19 % d.w. (A. muscaria), and 0.3 % d.w. (A. pantherina).made clear the amounts of muscarine were very low (0.009 % for A. muscaria and lower than 0.0005 % for A. pantherina). On the other hand, some Inocybe and Clitocybe species, responsible for the typical muscarine syndrome contain this substance in the range of 0.1-0.3 % d.w. A very sensitive analysis of muscimol and ibotenic acid can detect down to 30 ng of the compound. This technique makes use of an automated amino acid analyser equipped with a microbore column. The highest content of toxins is reported to be in the yellow flesh beneath the cuticle, ibotenic acid : 548 nmol g x1 f.w. and muscimol : 366 nmol g x1 f.w.. HPLC has been developed for A. muscaria content analysis. Ibotenic acid was assayed on two columns connected in series: (1) Ibotenic acid, LiChrosorb-NH 2 10 m column (Merck, Darmstadt, Germany) and a Nucleosil 5CN (Macherey Nagel, Du¨ren Germant) with 0.05 M sodium acetate buffer pH 4 as eluent ; (2) muscimol on LiChrosorb-NH 2 10 m column, 0.05 M with sodium acetate buffer pH 4/methanol 1/9 v/v as eluent. The amount of ibotenic acid was estimated at 100 ppm f.w. and that of muscimol <3 ppm. Mushrooms extracts treated with acetic acid had muscimol at 40 ppm, demonstrating that ibotenic acid is the major component in the fungus, while muscimol is the real psychotropic substance resulting from decarboxilation occurring during the preparation of the meal and the gastric digestion. Therefore, the two substances must be considered as a functional entity, so-called 'pilzatropine '. An ion-interaction HPLC method permitted the simultaneous detection of muscimol and ibotenic acid. The stationary phase was a reverse C 18 matrix, and the mobile phase aqueous octylammonium o-phosphate 5 mm. The amounts of both compounds were higher in caps than in stems; the mean values for ibotenic acid and muscimol respectively were (f.w.) : 990 mg kg x1 (caps), 230 mg kg x1 (stems), and 380 mg kg x1 (caps), and 80 mg kg x1 (stems). These substances were not detected in A. mappa. A fresh average size fruit body of A. muscaria (60-70 g) contains up to 70 mg of ibotenic acid. Reverse phase HPLC (column : Shim-Pack P-NH 2 /S 2054 allows the detection of ibotenic acid (limit: 18 ppm) and muscimol (limit : 10 ppm). Muscimol as its trimethylsilylor trifluoroacetyl derivativeshas been detected by gas chromatography coupled with mass spectrometry (GC-MS).
The 'insecticidal ' effect A large number of publications traditionally refer to the insecticidal properties of Amanita muscaria. Early investigations simultaneously described such an unusual property for ibotenic acid, 'agarin ', with qualitative experiments (Amanita Factor B ;. Moreover a tripeptide, 'pantherine ', with such a biological ability has been found in the fungus. However, quantitative experiments of, who named the substance 'pra¨muscimol ', showed that diptera were weakly sensitive to any toxic action by contact or ingestion. These results were confirmed by. A putative use of A. muscaria components for house fly (Musca domestica) control has been discussed with regard to the increasing importance of resistance. Such a reputed characteristic must be rationalized because of the names of the mushroom in different countries and arising comes from oral tradition, tales, paintings, and folklore.
Besides experimental data on animals, there are a few veterinary reports of accidental cat poisonings, with early symptoms 15-30 min after ingestion. A state of excitement, which lasts up to 4 h, quickly follows a brief period of drowsiness. The animal then passes into a deep sleep, and recovery is usually within 24 h. Considering the diagnosis, biochemical changes develop 30 min after peritoneal injection of aqueous extracts of Amanita muscaria, A. pantherina and A. rubrovolvata into male rats. These are decreases of acetylcholine esterase activity, liver glycogen, blood urea nitrogen, together with increase of blood glucose level ; serum transaminase activities were not affected. The values returned to normal within 6 h. The latter data demonstrate that the poisoning is not serious and that vital organs like liver and kidneys are not affected. Vegetative functions are hardly influenced by the toxins. A bioassay was designed to monitor the efficiency of extraction and purification procedures, the prolongation of sleeping time in mice after administration of a short-acting narcotic. Further studies show that a muscimol active dose given orally is 7.5-10 mg, and the LD 50 (i.p.) is 2.5 mg kg x1 for mice and 3.5 mg kg x1 for rats). After intraperitoneal injection and oral administration of muscimol (4-8 mg kg x1 ) and ibotenic acid (16 mg kg x1 ), the diameter of mouse pupils is broadened. Both drugs induce a marked anorexogenic action on mice (2-3 mg kg x1 oral) with sedation, hypnosis, muscle twitching and catalepsy. The drug produces electroencephalogram (EEG) alterations that are different from those provoked by hallucinogenic substances such as LSD or mescaline. The EEG patterns resemble those induced by anticholinergic drugs such as atropine and Ditran (Scotti de. The effects are slightly affected by physostigmine (eserine). Bilateral injections of ibotenic acid into the nucleus basalis of rats cause fewer problems than kainic acid. Most probably, after ingestion, the low pH gastric fluid hydrolyses ibotenic acid into muscimol ; afterwards, the active component proceeds to the brain or is eliminated via the systemic circulation. The remnant of the active component in the urine accounts for the tradition of drinking the urine of the shaman or of deers who consumes the mushrooms in some Siberian tribes in order to get a 'second-hand ' stimulus). Both ibotenic acid and muscimol are found in human urine 1 h after consumptions ; this observation was confirmed experimentally on mice. Thus, a priori, an acidic decarboxylation step in the gastrointestinal tract would produce muscimol that afterwards enters the central nervous system via the systemic circulation. Ibotenic acid has been shown to be the key component of A. muscaria and A. pantherina. No direct toxic action on typical target systems implicated in mushroom poisonings (i.e. liver, kidneys, essential metabolism) has been reported). Nevertheless, repetitive injections of ibotenic acid induce a transient change in GABA receptor sensitivityand might produce lesions in the basal forebrain as it destroys a small percentage of cholinergic cells; the consequence in vivo would be a significant and prolonged learning and memory deficit. In the case of young mammals, muscimol also disturbs the development of the hypothalamic noradrenaline system.
Ibotenic acid, and particularly muscimol, have to be regarded as the substances responsible for the psychotropic action of Amanita muscaria. The effects of both substances are similar but not identical to the effect of the fungus in toto. Ibotenic acid and muscimol are members of a distinctive class of alkaloids, the excitatory amino acids. This group includes kainic acid, domoic acid, tricholomic acid, quisqualic acid, yunaine, and petalonine ; all naturally occurring amino acids. Ibotenic acid and muscimol are respectively conformationally restricted derivatives of glutamic acid 19 and gamma-aminobutyric acid (GABA) 20. Consequently, they act like neurotransmitters involved in the control of neuronal activity in the mammalian central nervous system, operating on spinal neurones. Ibotenic acid is known to act on glutamic acid receptors, and muscimol acts on GABA receptors in the snail Helix aspersa. The same effect is observed in cats ; ibotenic acid and excitant action, muscimol and depressant action). Nevertheless, unlike glutamic acid and GABA, ibotenic acid and muscimol cross the blood-brain barrier, most probably by active transport, counterfeiting endogenous neurotransmitters and causing brain disorders (Fig.). Glutamic acid is a major excitatory neurotransmitter in mammalian central nervous systems and its receptors are implicated in neurological disorders such as epilepsy and Huntington's disease. Inhibitory glutamate receptors (IGluRs) constitute a class of ion channel proteins equivalent to glycine and GABA receptors. Ibotenic acid acts on IgluRs. Considering the minimum energy of the mono-anion, ibotenic acid displays a rigid structural analogue of L-glutamate. Hence it presents the suitable shape of glutamate agonists confirmed by molecular orbital theory. The central inhibitor receptor necessitates an effector displaying two highly charged sites of opposite signs separated by approximately 6 A ˚; this conformation was confirmed by X-ray analysis. AMPA, a synthetic analogue of ibotenic acid, displays the same typical moiety and was found to be more effective and specific glutamate agonist than the parent compound. Intraperitoneal injections of muscimol and ibotenic acid produce an increase of serotonin and dopamine in the brain of mice and rats. These variations might result from reduced turnover of neurotransmitters, therefore elevations could contribute to psychotonic actions and even accentuate some of them (e.g. anorexia, central pupil dilatation). There is evidence that damaged function of GABAmediated inhibitory synapses is implied in experimental and clinical seizure disorders. Decreased concentration, and then activity of GABA, may have a role in human epilepsy. Muscimol is a potent conformational analogue and biologically active bioisostere (Krogsgaard et al. 2000) (Fig.). This toxin tightly binds with the gamma-aminobutyrate receptor. Moreover, it is also an inhibitor of neuronal and glial GABA uptake and a substrate for the GABA-metabolizing enzyme : GABA transaminase. Subsequently, some research teams tried to improve the potency of drugs. Lipophilicity is of considerable interest for the prediction of the transport, adsorption and distribution properties of molecules, and so is an important factor in drug design. More lipophilic bioisosteres of muscimol and GABA have been synthesized, such as Tiagabine 21. This drug was marketed as a therapeutic agent for the treatment of epilepsy, Gabatril 1 (Krogsgaard-Larsen 1977,. The above biological and pharmacological activities account for the psychotropic effects of A. muscaria.
Among all mushrooms species, the fly agaric has long been a source of interest and attractiveness ; it embodies the concept of the 'mushroom ' and is probably the most illustrated one. Despite an evil reputation, this mushroom, unique in its striking appearance, is the most widely used pictogram for wild mushrooms. Its uses include illustrations of tales for children, and strip cartoons. Christmas tree decorations, pastries, and even tattoosas well as the artefactsare responsible for this renown. Because of its vivid colour, white spots, sturdy habit and its psychoactive properties, the fly agaric still exerts great fascination. Shamans, sorcerers, and witches made use of it, all classes of officiating ' priests ' in different ethnic groups which are geographically and culturally distant. The ' priests ' try to escape the human condition and use the mushroom in ' religious ' rituals. Beside the chroniclers of the 18th and 19th century in northern Europe and Asia,forcefully argued that this mushroom represents the ' soma ' of some ancient Aryan tribes. This concept designates a 'plant ' that produces a juice when squeezed, that is thereafter filtered and handled as a divine inebriating agent. He stated that several parts of the Rig Veda conveyed that ancient peoples extensively consumed the mushroom for its psychoactive properties. Besides various symbols that might correspond to A. muscaria and originate from northern and southern Asian traditions, some may also be discerned in Buddhist myths. They appear to be echoed in Germanic tradition, possibly in some characteristics related to the god Odin. A few publications argue that Christianity originates from a cult of A. muscaria, Jesus was supposedly being invested with the energy of the mushroom), but other religious and secular biblical scholars discredit this. Nowadays, reliable sources report that Ostyak and Vogul tribes in western Siberia, and Kamchadal, Koryak, Chukchi tribes in eastern Siberia, still use A. muscaria for shamanistic rites. However, in contrast to hallucinogenic mushrooms, mainly species of Psilocybe used in Mesoamerican culture, the fly agaric is not a true object of religious and ritual veneration. Two attitudes coexist in world cultures towards mushrooms consumption : mycophilic and mycophobic. They are present traditionally among the word cultures. Some groups (e.g. German, Celtic, British) have a dislike for mushrooms, while others (e.g. Slavic, Lithuanian, French, Italian, Spanish, and other Mediterranean ones) are, to a certain extent, fond of them. The mycophobic attitude possibly corresponds to remnants of a primitive worship/fear of mushrooms connected with 'folk wisdom'. This feeling is conveyed in language and etymology, possibly starting from Neolithic and Palaeolithic times. Unconsciously, some people make a clear-cut distinction between 'safe ' mushrooms and suspicious mushrooms for instance mushrooms vs toadstools in English, and setas, hongos vs hongos venenosos in Spanish. The different attitudes are documented bywho travelled widely in Latin America and various steps in the mycophobicity/ mycophily scale. Moreover, educated people are careful about mushrooms, when those of less developed cultures tend to consume them more readily.
The designation of the species as 'fly agaric', ' bug agaric', is surprisingly equivalent all over the world: 'amanite tue-mouche ' (French), 'Fliegenpilz ' (German), 'muchomor ' (Russian), ' moscario' (Italian), 'hongo mosquero ', 'hongo matamoscas ' (Spanish). The name of Amanita strobiliformis in Japanese is 'haetorimodashi ', fly-killer). At the beginning of the last century,mentioned the practice concerning a preparation of the mushroom to kill flies. The presumed insecticidal effect against flies should, however, be discounted since experiments show that a percentage of flies exposed to the juice of the mushroom did not die, but were rather subjected to transient behaviour disorder (i.e. became intoxicated). However, a few experiments do show that some species of Amanita inhibit the growth of Drosophila melanogaster larvae fed with powdered fruit bodies. The 'fly ' name may be explained in another way, since it bears the same semantic sound in several languages. Going back to ancient times in many countries, ' fly' has been identified with madness or supernatural possession. In the Middle Ages, it was believed that flies or other insects could penetrate the head of a person and cause mental illness. This is evidenced in Hyeronymus Bosch's painting 1510 'Paradise and Hell ': on the left panel, one can see very well high above in the sky the rebellious angels, who flew from heaven as a crowd of hideous insects. The Prado Museum in Madrid exhibits another painting by the same artist ' The garden of earthly delight ' 1504. A. muscaria can be distinguished on the left-hand panel of the work). In Europe, some other terms naming the mushroom are reminiscent of madness, for example 'Narrenschwamm ' (German), 'oriol fol ' (Catalan), 'mjioulo folo ' (Toulouse dialect), 'coucourlo foulo ' (langue d'oc), and 'ovolo matto' (northern Italian). Conversely, the fly-possession link could be rather beneficial to shamans. The Bible is no exception ; 'Evil One ', Belzebuth, means 'Lord of the flies ' and is synonymous with a false God adored by drunkards. It is likely that A. muscaria was named 'mushroom of the flies ' because it is able to induce divine or evil states of possession similar to those that flies were supposed to produce in human brains. In Mexico, a Quiche idiom that describes hazardous, frightful potentialities is 'itzel ocox ', meaning the diabolical mushroom. The original meaning was evidently forgotten or misinterpreted over time, giving way to the so-called insecticidal activity. With respect to rituals and beliefs, the habitat of the fly agaric is pertinent. The tree has long been considered the ' Tree of Life ', identified with the 'Weed of Immortality ', and later the 'Tree of Knowledge ' and with the 'Forbidden Fruit ' in the Genesis. Thus, it appears to be at the origins of religious practices. Its roots were said to 'swallow the pool of water of life ' and nourish the ' sacred mushroom ', 'his mind is the mind of a woman who offers her milk to anybody that comes towards her '. A. muscaria is often mycorrhizally associated with Betula, and also Abies, Cystus, Picea, Pinus, and Quercus species. The appearance of the mushroom is unique, colourful, red and covered by whitish patches. It would be possible to consider this aposematic as it is seen in ladybirds, dendrobate frogs, etc. The conspicuous markings make it easily recognizable and warn wouldbe predators that it contains toxins.
Historical documents, such as wall paintings, woodcarvings, and sculptures suggest that the psychotropic properties of some mushrooms have been known from ancient times and that consumption was for religious, social and therapeutic purposes, and occurred in all continents. Representations of Amanita mushrooms, most probably A. muscaria, have been reported in polychromatic rock paintings in the Sahara; such works date from the Palaeolithic 9000-7000 BC. The American surgeon generalwas particularly interested in extrasensory perception after World War II and searched for psychotropic substances able to induce latent abilities among ' sensitive' subjects. He investigated the mycology, biology, physiology, ethnology, etymology, and comparative linguistics of A. muscaria and its psychotropic properties. Further, he attempted to prove there were cults in ancient Egypt equivalent to those of the Indo-European transmigrations, and present today in eastern Siberia. Puharich also noted the comparison with cults in Central America related to hallucinogenic mushrooms. His conclusions confirm those ofrelating to other parts of north Africa. Among the various pieces of evidence he produced is an unambiguous distinction between a mushroom, a sunshade, and a lotus flower on seals illustrating a ritual ceremony in the pre-Snefrou period (before 2200 BC).
Amanita muscaria occupies a unique position amongst all mushrooms. Its emblematic aspect merges with the psychotropic effects and the chemical load. As far as toxic effects are concerned, consumption of A. muscaria and A. pantherina does not induce any critical organ damage. Thus, in the case of such a poisoning, the victims are not considered endangered, but intensive care is recommended. Nevertheless, severe neuron and even brain lesions could be anticipated in cases of recurrent consumption. A. muscaria has contributed to the progress of chemistry and pharmacology. Noteworthy, is that in most European countries, the possession, sale and consumption of the species are not yet subject to legislation such as in place for the more widely recognized hallucinogenic species (i.e. Psilocybe, Panaeolus and other species containing psilocybin-related toxins). A. muscaria is not yet the object of any drug-traffic, an aspect reflected in the lack of awareness amongst possible consumers or applicants for recreational purposes. Considering the effects so far reported, A. muscaria has a low psychotropic action, but still a toxic one.
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