101-676: Anatoxin-a , also known as Very Fast Death Factor ( VFDF ), is a secondary, bicyclic amine alkaloid and cyanotoxin with acute neurotoxicity . It was first discovered in the early 1960s in Canada, and was isolated in 1972. The toxin is produced by multiple genera of cyanobacteria and has been reported in North America, South America, Central America, Europe, Africa, Asia, and Oceania. Symptoms of anatoxin-a toxicity include loss of coordination , muscular fasciculations , convulsions and death by respiratory paralysis . Its mode of action
202-615: A basic nitrogen atom with a lone pair . Formally, amines are derivatives of ammonia ( NH 3 ), wherein one or more hydrogen atoms have been replaced by a substituent such as an alkyl or aryl group (these may respectively be called alkylamines and arylamines; amines in which both types of substituent are attached to one nitrogen atom may be called alkylarylamines). Important amines include amino acids , biogenic amines , trimethylamine , and aniline . Inorganic derivatives of ammonia are also called amines, such as monochloramine ( NClH 2 ). The substituent −NH 2
303-530: A 100 fold lesser selectivity for these types of receptors than it has for nAchRs. Anatoxin-a also shows much less potency in the CNS than in neuromuscular junctions. In hippocampal and brain stem neurons, a 5 to 10 times greater concentration of anatoxin-a was necessary to activate nAchRs than what was required in the PNS. In normal circumstances, acetylcholine binds to nAchRs in the post-synaptic neuronal membrane, causing
404-523: A cell-free environment which allows easier manipulation. The first vertebrate ever to be cloned was an African clawed frog in 1962, an experiment for which Sir John Gurdon was awarded the Nobel Prize in Physiology or Medicine in 2012 "for the discovery that mature cells can be reprogrammed to become pluripotent". Additionally, four female African clawed frogs and stored sperm were present on
505-473: A conformational change in the extracellular domain of the receptor which in turn opens the channel pore. This allows Na and Ca ions to move into the neuron, causing cell depolarization and inducing the generation of action potentials , which allows for muscle contraction. The acetylcholine neurotransmitter then dissociates from the nAchR, where it is rapidly cleaved into acetate and choline by acetylcholinesterase . Anatoxin-a binding to these nAchRs cause
606-457: A final product. The first and most extensively explored approach used to synthesize anatoxin-a in vitro, cyclooctene cyclization involves 1,5-cyclooctadiene as its initial source. This starting substance is reacted to form methyl amine and combined with hypobromous acid to form anatoxin-a. Another method developed in the same laboratory uses aminoalcohol in conjunction with mercuric (II) acetate and sodium borohydride. The product of this reaction
707-655: A greater frequency and prevalence of cyanobacterial blooms. The World Health Organization in 1999 and EPA in 2006 both came to the conclusion that there was not enough toxicity data for anatoxin-a to establish a formal tolerable daily intake (TDI) level, though some places have implemented levels of their own. Anatoxin-a is not regulated under the Safe Drinking Water Act , but states are allowed to create their own standards for contaminants that are unregulated. Currently there are four states that have set drinking water advisory levels for anatoxin-a as seen in
808-459: A harmful invasive species. They can travel short distances to other bodies of water, and some have even been documented to survive mild freezes. They have been shown to devastate native populations of frogs and other creatures by eating their young. In 2003, Xenopus laevis frogs were discovered in a pond at San Francisco 's Golden Gate Park . Much debate now exists in the area on how to exterminate these creatures and keep them from spreading. It
909-420: A ketone intermediate. Addition of an organocuprate to the ketone produces an enol triflate derivative, which is then lysed hydrogenously and treated with a deprotecting agent in order to produce anatoxin-a. Similar strategies have also been developed and utilized by other laboratories. Iminium ion cyclization utilizes several different pathways to create anatoxin-a, but each of these produces and progresses with
1010-541: A more comprehensive discussion of the use of these frogs in biomedical research, see Xenopus . Xenopus laevis is also notable for its use in the first widely used method of pregnancy testing . In the 1930s, two South African researchers, Hillel Shapiro and Harry Zwarenstein, students of Lancelot Hogben at the University of Cape Town , discovered that the urine from pregnant women would induce oocyte production in X. laevis within 8–12 hours of injection. This
1111-458: A more pronounced cloaca and have hip-like bulges above their rear legs where their eggs are internally located. Both males and females have a cloaca , which is a chamber through which digestive and urinary wastes pass and through which the reproductive systems also empty. The cloaca empties by way of the vent which in reptiles and amphibians is a single opening for all three systems. African clawed frogs are fully aquatic and will rarely leave
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#17327875828631212-516: A mottled greenish-grey-brown in color, sometimes with yellowish botches, and with a pale white-cream belly. African clawed frogs have been frequently sold as pets, and are sometimes misidentified as African dwarf frogs . Albino clawed frogs are common and sold as animals for laboratories . Amphibians reproduce by fertilizing eggs outside of the female's body (see frog reproduction ). Of the seven amplexus modes (positions in which frogs mate), these frogs are found breeding in inguinal amplexus, where
1313-631: A permit in the following US states: Arizona, California, Kentucky, Louisiana, New Jersey, North Carolina, Oregon, Vermont, Virginia, Hawaii, Nevada, and Washington state. However, it is legal to own Xenopus laevis in New Brunswick (Canada) and Ohio. Feral colonies of Xenopus laevis exist in South Wales , United Kingdom . In Yunnan , China there is a population of albino clawed frogs in Lake Kunming , along with another invasive:
1414-493: A pyrrolidine iminium ion. The major differences in each pathway relate to the precursors used to produce the imium ion and the total yield of anatoxin-a at the end of the process. These separate pathways include production of alkyl iminium salts, acyl iminium salts and tosyl iminium salts. Enyne metathesis of anatoxin-a involves the use of a ring closing mechanism and is one of the more recent advances in anatoxin-a synthesis. In all methods involving this pathway, pyroglutamic acid
1515-865: A species of Pseudomonas , capable of biodegrading anatoxin-a at a rate of 2–10 μg ml d, has been identified. Biological (granular) activated carbon (BAC) has also been tested as a method of biodegradation, but it is inconclusive whether biodegradation occurred or if anatoxin-a was simply adsorbing the activated carbon. Others have called for additional studies to determine more about how to use activated carbon effectively. Chemical treatment methods are more common in drinking water treatment compared to biological treatment, and numerous processes have been suggested for anatoxin-a. Oxidants such as potassium permanganate , ozone , and advanced oxidation processes ( AOPs ) have worked in lowering levels of anatoxin-a, but others, including photocatalysis, UV photolysis , and chlorination , have not shown great efficacy. Directly removing
1616-428: A weak toxic effect at 10 μg/mL only after an extended period of contact (48 hours). No data on inhalation toxicity of anatoxin-a is currently available, though severe respiratory distress occurred in a water skier after they inhaled water spray containing a fellow cyanobacterial neurotoxin, saxitoxin . It is possible that inhalation of water spray containing anatoxin-a could pose similar consequences. Anatoxin-a
1717-649: Is a very powerful nicotinic acetylcholine receptor agonist and as such has been extensively studied for medicinal purposes. It is mainly used as a pharmacological probe in order to investigate diseases characterized by low acetylcholine levels, such as muscular dystrophy , myasthenia gravis , Alzheimer disease , and Parkinson disease . Further research on anatoxin-a and other less potent analogues are being tested as possible replacements for acetylcholine. Amine In chemistry , amines ( / ə ˈ m iː n , ˈ æ m iː n / , UK also / ˈ eɪ m iː n / ) are compounds and functional groups that contain
1818-561: Is able to fit into their mouths. Being aquatic, clawed frogs use their sense of smell and their lateral line to detect prey rather than eyesight like other frogs. However, clawed frogs can still see using their eyes and will stalk prey or watch predators by sticking their heads out of the water. Clawed frogs will dig through substrate to unearth worms and other food. Unlike other frogs, they have no tongue to extend to catch food, so clawed frogs use their hands to grab food and shovel it into their mouths. These frogs are particularly cannibalistic;
1919-422: Is an agonist of both neuronal α 4 β 2 and α 4 nicotinic acetylcholine receptors present in the CNS as well as the (α 1 ) 2 βγδ muscle-type nAchRs that are present at the neuromuscular junction . (Anatoxin-a has an affinity for these muscle-type receptors that is about 20 times greater than that of acetylcholine .) However, the cyanotoxin has little effect on muscarinic acetylcholine receptors ; it has
2020-477: Is anatoxin-a or another neurotoxin that causes the resulting deaths. Large amounts of sample material are also needed for such testing. In addition to the biological methods, scientists have used chromatography to detect anatoxin-a. This is complicated by the rapid degradation of the toxin and the lack of commercially available standards for anatoxin-a. Despite the relatively low frequency of anatoxin-a relative to other cyanotoxins, its high toxicity (the lethal dose
2121-475: Is because (+)-anatoxin-a, the s- cis enone conformation, has a distance a 6.0 Å between its nitrogen and carbonyl group, which corresponds well to the 5.9 Å distance that separate the nitrogen and oxygen in acetylcholine. Respiratory arrest , which results in a lack of an oxygen supply to the brain, is the most evident and lethal effect of anatoxin-a. Injections of mice, rats, birds, dogs, and calves with lethal doses of anatoxin-a have demonstrated that death
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#17327875828632222-449: Is called an amino group. The chemical notation for amines contains the letter "R", where "R" is not an element, but an "R-group", which in amines could be a single hydrogen or carbon atom, or could be a hydrocarbon chain. Compounds with a nitrogen atom attached to a carbonyl group , thus having the structure R−C(=O)−NR′R″ , are called amides and have different chemical properties from amines. Amines can be classified according to
2323-632: Is considered a more viable model for genetics, although gene editing protocols have now been perfected for. Roger Wolcott Sperry used X. laevis for his famous experiments describing the development of the visual system. These experiments led to the formulation of the chemoaffinity hypothesis . X. laevis have been used as a model organism in vertebrae cardiogenesis, human congenital heart defects, and in GWAS studies of congenital heart defects. Xenopus oocytes provide an important expression system for molecular biology . By injecting DNA or mRNA into
2424-408: Is determined by the connectivity of the substituents attached to the nitrogen: It is also possible to have four organic substituents on the nitrogen. These species are not amines but are quaternary ammonium cations and have a charged nitrogen center. Quaternary ammonium salts exist with many kinds of anions . Amines are named in several ways. Typically, the compound is given the prefix "amino-" or
2525-453: Is introduced into the birds via water contaminated with cyanobacterial mat communities that arise from the hot springs in the lake bed. The first biologically occurring initial substance for tropane expansion into anatoxin-a was cocaine , which has similar stereochemistry to anatoxin-a. Cocaine is first converted into the endo isomer of a cyclopropane, which is then photolytically cleaved to obtain an alpha, beta unsaturated ketone. Through
2626-523: Is no official guideline level for anatoxin-a, although scientists estimate that a level of 1 μg l would be sufficiently low. Likewise, there are no official guidelines regarding testing for anatoxin-a. Among methods of reducing the risk for cyanotoxins, including anatoxin-a, scientists look favorably on biological treatment methods because they do not require complicated technology, are low maintenance, and have low running costs. Few biological treatment options have been tested for anatoxin-a specifically, although
2727-493: Is not known for humans, but is estimated to be less than 5 mg for an adult male) means that it is still considered a serious threat to terrestrial and aquatic organisms, most significantly to livestock and to humans. Anatoxin-a is suspected to have been involved in the death of at least one person. The threat posed by anatoxin-a and other cyanotoxins is increasing as both fertilizer runoff, leading to eutrophication in lakes and rivers, and higher global temperatures contribute to
2828-516: Is not to be confused with guanitoxin (formerly anatoxin-a(S)), another potent cyanotoxin that has a similar mechanism of action to that of anatoxin-a and is produced by many of the same cyanobacteria genera, but is structurally unrelated. Anatoxin-a was first discovered by P.R. Gorham in the early 1960s, after several herds of cattle died as a result of drinking water from Saskatchewan Lake in Ontario, Canada, which contained toxic algal blooms . It
2929-467: Is preceded by a sequence of muscle fasciculations , decreased movement, collapse, exaggerated abdominal breathing, cyanosis and convulsions . In mice, anatoxin-a also seriously impacted blood pressure and heart rate, and caused severe acidosis . Many cases of wildlife and livestock deaths due to anatoxin-a have been reported since its discovery. Domestic dog deaths due to the cyanotoxin, as determined by analysis of stomach contents, have been observed at
3030-601: Is represented in this article by two dots above or next to the N. The water solubility of simple amines is enhanced by hydrogen bonding involving these lone electron pairs. Typically salts of ammonium compounds exhibit the following order of solubility in water: primary ammonium ( RNH 3 ) > secondary ammonium ( R 2 NH 2 ) > tertiary ammonium (R 3 NH ). Small aliphatic amines display significant solubility in many solvents , whereas those with large substituents are lipophilic. Aromatic amines, such as aniline , have their lone pair electrons conjugated into
3131-668: Is suspected to have also been the cause of death of one human. One study found that anatoxin-a is capable of binding to acetylcholine receptors and inducing toxic effects with concentrations in the nano-molar (nM) range if ingested. Dermal exposure is the most likely form of contact with cyanotoxins in the environment. Recreational exposure to river, stream, and lake waters contaminated with algal blooms has been known to cause skin irritation and rashes. The first study that looked at in vitro cytotoxic effects of anatoxin-a on human skin cell proliferation and migration found that anatoxin-a exerted no effect at 0.1 μg/mL or 1 μg/mL, and
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3232-529: Is through the nicotinic acetylcholine receptor (nAchR) where it mimics the binding of the receptor's natural ligand , acetylcholine . As such, anatoxin-a has been used for medicinal purposes to investigate diseases characterized by low acetylcholine levels. Due to its high toxicity and potential presence in drinking water, anatoxin-a poses a threat to animals, including humans. While methods for detection and water treatment exist, scientists have called for more research to improve reliability and efficacy. Anatoxin-a
3333-486: Is unknown if these frogs entered the San Francisco ecosystem through intentional release or escape into the wild. San Francisco officials drained Lily Pond and fenced off the area to prevent the frogs from escaping to other ponds in the hopes they starve to death. Due to incidents in which these frogs were released and allowed to escape into the wild, African clawed frogs are illegal to own, transport or sell without
3434-539: Is unstable in water and other natural conditions, and in the presence of UV light undergoes photodegradation , being converted to the less toxic products dihydroanatoxin-a and epoxyanatoxin-a. The photodegradation of anatoxin-a is dependent on pH and sunlight intensity but independent of oxygen, indicating that the degradation by light is not achieved through the process of photo-oxidation. Studies have shown that some microorganisms are capable of degrading anatoxin-a. A study done by Kiviranta and colleagues in 1991 showed that
3535-485: Is used as a starting material in conjunction with a Grubb's catalyst. Similar to iminium cyclization, the first attempted synthesis of anatoxin-a using this pathway used a 2,5-cis-pyrrolidine as an intermediate. Anatoxin-a is synthesized in vivo in the species Anabaena flos-aquae , as well as several other genera of cyanobacteria. Anatoxin-a and related chemical structures are produced using acetate and glutamate. Further enzymatic reduction of these precursors results in
3636-430: The platanna ) is a species of African aquatic frog of the family Pipidae . Its name is derived from the short black claws on its feet. The word Xenopus means 'strange foot' and laevis means 'smooth'. The species is found throughout much of Sub-Saharan Africa ( Nigeria and Sudan to South Africa ), and in isolated, introduced populations in North America, South America, Europe, and Asia. All species of
3737-486: The American bullfrog . Because this population is albino, it suggests that the clawed frogs originated from the pet trade or a laboratory. The African clawed frog may be an important vector and the initial source of Batrachochytrium dendrobatidis , a chytrid fungus that has been implicated in the drastic decline in amphibian populations in many parts of the world. Unlike in many other amphibian species (including
3838-593: The Delépine reaction , although this is rarely employed on an industrial scale. Selectivity is also assured in the Gabriel synthesis , which involves organohalide reacting with potassium phthalimide . Aryl halides are much less reactive toward amines and for that reason are more controllable. A popular way to prepare aryl amines is the Buchwald-Hartwig reaction . Disubstituted alkenes react with HCN in
3939-816: The Hinsberg reaction , is a chemical test for the presence of amines. Because amines are basic, they neutralize acids to form the corresponding ammonium salts R 3 NH . When formed from carboxylic acids and primary and secondary amines, these salts thermally dehydrate to form the corresponding amides . Amines undergo sulfamation upon treatment with sulfur trioxide or sources thereof: Amines reacts with nitrous acid to give diazonium salts. The alkyl diazonium salts are of little importance because they are too unstable. The most important members are derivatives of aromatic amines such as aniline ("phenylamine") (A = aryl or naphthyl): Anilines and naphthylamines form more stable diazonium salts, which can be isolated in
4040-476: The Space Shuttle Endeavour when it was launched into space on mission STS-47 on September 12, 1992, so that scientists could test whether reproduction and development could occur normally in zero gravity. Xenopus laevis also serves as an ideal model system for the study of the mechanisms of apoptosis. In fact, iodine and thyroxine stimulate the spectacular apoptosis of the cells of
4141-717: The mediobasal hypothalamus , then it stimulates seasonal testicular growth; if peripherally, then testicular regression and cold-season thermogenesis. These observations are regarded as widely applicable across vertebrate thyroid systems. The lipidomics of Xenopus oocytes have been studied by Tian et al 2014 and Phan et al 2015. In X. laevis , epigenetic methylation changes in neural-developmental genes associated with aging are analogous to aging related epigenetic changes in mammalian species. This finding suggests that, during their evolutionary divergence, patterns of epigenetic changes in neural-development genes during aging have been conserved between frogs and mammals C. In
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4242-473: The nitrogen inversion of the stereocenter is about 7 kcal/mol for a trialkylamine. The interconversion has been compared to the inversion of an open umbrella into a strong wind. Amines of the type NHRR' and NRR′R″ are chiral : the nitrogen center bears four substituents counting the lone pair. Because of the low barrier to inversion, amines of the type NHRR' cannot be obtained in optical purity. For chiral tertiary amines, NRR′R″ can only be resolved when
4343-481: The oocyte . Translation of proteins can be blocked or splicing of pre-mRNA can be modified by injection of Morpholino antisense oligos into the oocyte (for distribution throughout the embryo) or early embryo (for distribution only into daughter cells of the injected cell). Extracts from the eggs of X. laevis frogs are also commonly used for biochemical studies of DNA replication and repair, as these extracts fully support DNA replication and other related processes in
4444-568: The National Institutes of Health. The work rapidly expanded to include de novo reconstruction of X. laevis transcripts, in collaboration with groups around the world donating Illumina Hi-Seq RNA sequencing datasets. Genome sequencing by the Rokhsar and Harland groups (UC Berkeley) and by Taira and collaborators (University of Tokyo, Japan) gave a major boost to the project, which, with additional contributions from investigators in
4545-496: The Netherlands, Korea, Canada and Australia, led to publication of the genome sequence and its characterization in 2016. X. laevis oocytes are often used as an easy model for the artificially induced expression of transgenes . For example, they are commonly used when studying chloroquine resistance produced by specialized transporter mutants. Even so the foreign expression tissue may itself confer some alterations to
4646-645: The R, R', and R″ groups are constrained in cyclic structures such as N -substituted aziridines ( quaternary ammonium salts are resolvable). In aromatic amines ("anilines"), nitrogen is often nearly planar owing to conjugation of the lone pair with the aryl substituent. The C-N distance is correspondingly shorter. In aniline, the C-N distance is the same as the C-C distances. Like ammonia, amines are bases . Compared to alkali metal hydroxides, amines are weaker. The basicity of amines depends on: Owing to inductive effects,
4747-541: The United States, Canada, and Finland between 1980 and the present. A particularly interesting case of anatoxin-a poisoning is that of lesser flamingos at Lake Bogoria in Kenya . The cyanotoxin, which was identified in the stomachs and fecal pellets of the birds, killed roughly 30,000 flamingos in the second half of 1999, and continues to cause mass fatalities annually, devastating the flamingo population. The toxin
4848-930: The action potential. Later, Thomas et al., (1993) through his work with chicken α 4 β 2 nAChR subunits expressed on mouse M 10 cells and chicken α 7 nAChR expressed in oocytes from Xenopus laevis , showed that anatoxin-a is also a potent agonist of neuronal nAChR. Laboratory studies using mice showed that characteristic effects of acute anatoxin-a poisoning via intraperitoneal injection include muscle fasciculations , tremors, staggering, gasping, respiratory paralysis, and death within minutes. Zebrafish exposed to anatoxin-a contaminated water had altered heart rates. There have been cases of non-lethal poisoning in humans who have ingested water from streams and lakes that contain various genera of cyanobacteria that are capable of producing anatoxin-a. The effects of non-lethal poisoning were primarily gastrointestinal: nausea, vomiting, diarrhea, and abdominal pain. A case of lethal poisoning
4949-445: The amino acid lysine . The anionic polymer DNA is typically bound to various amine-rich proteins. Additionally, the terminal charged primary ammonium on lysine forms salt bridges with carboxylate groups of other amino acids in polypeptides , which is one of the primary influences on the three-dimensional structures of proteins. Hormones derived from the modification of amino acids are referred to as amine hormones. Typically,
5050-875: The amount of cyanotoxins in the surrounding water, threatening the health of both aquatic and terrestrial organisms. Some species of cyanobacteria that produce anatoxin-a don't produce surface water blooms but instead form benthic mats. Many cases of anatoxin-a related animal deaths have occurred due to ingestion of detached benthic cyanobacterial mats that have washed ashore. Anatoxin-a producing cyanobacteria have also been found in soils and aquatic plants. Anatoxin-a sorbs well to negatively charged sites in clay-like, organic-rich soils and weakly to sandy soils. One study found both bound and free anatoxin-a in 38% of aquatic plants sampled across 12 Nebraskan reservoirs, with much higher incidence of bound anatoxin-a than free. In 1977, Carmichael, Gorham, and Biggs experimented with anatoxin-a. They introduced toxic cultures of A. flos-aquae into
5151-417: The aromatic ring, and their positions relative to the amino group, also affect basicity as seen in the table. Solvation significantly affects the basicity of amines. N-H groups strongly interact with water, especially in ammonium ions. Consequently, the basicity of ammonia is enhanced by 10 by solvation. The intrinsic basicity of amines, i.e. the situation where solvation is unimportant, has been evaluated in
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#17327875828635252-416: The bacterial genus Pseudomonas was capable of degrading anatoxin-a at a rate of 2–10 μg/ml per day. Later experiments done by Rapala and colleagues (1994) supported these results. They compared the effects of sterilized and non-sterilized sediments on anatoxin-a degradation over the course of 22 days, and found that after that time vials with the sterilized sediments showed similar levels of anatoxin-a as at
5353-504: The bands appearing below 1600 cm , which are weaker and overlap with C-C and C-H modes. For the case of propyl amine , the H-N-H scissor mode appears near 1600 cm , the C-N stretch near 1000 cm , and the R 2 N-H bend near 810 cm . Alkyl amines characteristically feature tetrahedral nitrogen centers. C-N-C and C-N-H angles approach the idealized angle of 109°. C-N distances are slightly shorter than C-C distances. The energy barrier for
5454-423: The basicity of an amine might be expected to increase with the number of alkyl groups on the amine. Correlations are complicated owing to the effects of solvation which are opposite the trends for inductive effects. Solvation effects also dominate the basicity of aromatic amines (anilines). For anilines, the lone pair of electrons on nitrogen delocalizes into the ring, resulting in decreased basicity. Substituents on
5555-643: The benzene ring, thus their tendency to engage in hydrogen bonding is diminished. Their boiling points are high and their solubility in water is low. Typically the presence of an amine functional group is deduced by a combination of techniques, including mass spectrometry as well as NMR and IR spectroscopies. H NMR signals for amines disappear upon treatment of the sample with D 2 O. In their infrared spectrum primary amines exhibit two N-H bands, whereas secondary amines exhibit only one. In their IR spectra, primary and secondary amines exhibit distinctive N-H stretching bands near 3300 cm . Somewhat less distinctive are
5656-1699: The case of formaldehyde (R' = H), these products typically exist as cyclic trimers : RNH 2 + R 2 ′ C = O ⟶ R 2 ′ C = NR + H 2 O {\displaystyle {\ce {RNH2 + R'_2C=O -> R'_2C=NR + H2O}}} Reduction of these imines gives secondary amines: R 2 ′ C = NR + H 2 ⟶ R 2 ′ CH − NHR {\displaystyle {\ce {R'_2C=NR + H2 -> R'_2CH-NHR}}} Similarly, secondary amines react with ketones and aldehydes to form enamines : R 2 NH + R ′ ( R ″ CH 2 ) C = O ⟶ R ″ CH = C ( NR 2 ) R ′ + H 2 O {\displaystyle {\ce {R2NH + R'(R''CH2)C=O -> R''CH=C(NR2)R' + H2O}}} Mercuric ions reversibly oxidize tertiary amines with an α hydrogen to iminium ions: Hg 2 + + R 2 NCH 2 R ′ ↽ − − ⇀ Hg + [ R 2 N = CHR ′ ] + + H + {\displaystyle {\ce {Hg^2+ + R2NCH2R' <=> Hg + [R2N=CHR']+ + H+}}} An overview of
5757-514: The closely related western clawed frog ) where this chytrid fungus causes the disease Chytridiomycosis , it does not appear to affect the African clawed frog, making it an effective carrier. Invasive: The African clawed frog is considered invasive by the Centre of Invasive biology from Stellenbosh University with this species even going as far as predating on other species. There has even been
5858-509: The commencement of the experiment, while vials with non-sterilized sediment showed a 25-48% decrease. There are two categories of anatoxin-a detection methods. Biological methods have involved administration of samples to mice and other organisms more commonly used in ecotoxicological testing, such as brine shrimp ( Artemia salina ), larvae of the freshwater crustacean Thamnocephalus platyurus , and various insect larvae. Problems with this methodology include an inability to determine whether it
5959-493: The crystalline form. Diazonium salts undergo a variety of useful transformations involving replacement of the N 2 group with anions. For example, cuprous cyanide gives the corresponding nitriles: Aryldiazoniums couple with electron-rich aromatic compounds such as a phenol to form azo compounds . Such reactions are widely applied to the production of dyes. Imine formation is an important reaction. Primary amines react with ketones and aldehydes to form imines . In
6060-498: The cyanobacteria in the water treatment process through physical treatment (e.g., membrane filtration ) is another option because most of the anatoxin-a is contained within the cells when the bloom is growing. However, anatoxin-a is released from cyanobacteria into water when they senesce and lyse, so physical treatment may not remove all of the anatoxin-a present. Additional research needs to be done to find more reliable and efficient methods of both detection and treatment. Anatoxin-a
6161-510: The expression, and so findings may or may not be entirely identical to native expression: For example, iron has been found by Bakouh et al 2017 to be an important substrate for one such transporter in X. l. oocytes, but as of 2020 iron is merely presumptively involved in native expression of the same gene. Xenbase is the Model Organism Database (MOD) for both Xenopus laevis and Xenopus tropicalis . Xenbase hosts
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#17327875828636262-427: The family Pipidae are tongueless, toothless and completely aquatic. They use their hands to shove food in their mouths and down their throats and a hyobranchial pump to draw or suck things in their mouth. Pipidae have powerful legs for swimming and lunging after food. They also use the claws on their feet to tear pieces of large food. They have no external eardrums, but instead subcutaneous cartilaginous disks that serve
6363-400: The formation of anatoxin-a. Homoanatoxin, a similar chemical, is produced by Oscillatoria formosa and utilizes the same precursor. However, homoanatoxin undergoes a methyl addition by S-adenosyl-L-methionine instead of an addition of electrons, resulting in a similar analogue. The biosynthetic gene cluster (BGC) for anatoxin-a was described from Oscillatoria PCC 6506 in 2009. Anatoxin-a
6464-550: The full details and release information regarding the current v10 Xenopus laevis genome released in 2022. The clawed frog have been kept as pets and research subjects since as early as the 1950s. They are extremely hardy and long lived, having been known to live up to 20 or even 30 years in captivity. African clawed frogs are frequently mislabeled as African dwarf frogs in pet stores. Identifiable differences are: African clawed frogs are voracious predators and easily adapt to many habitats. For this reason, they can easily become
6565-456: The gas phase, but ten thousand times less so in aqueous solution. In aprotic polar solvents such as DMSO , DMF , and acetonitrile the energy of solvation is not as high as in protic polar solvents like water and methanol. For this reason, the basicity of amines in these aprotic solvents is almost solely governed by the electronic effects. Industrially significant alkyl amines are prepared from ammonia by alkylation with alcohols: Unlike
6666-399: The gas phase. In the gas phase, amines exhibit the basicities predicted from the electron-releasing effects of the organic substituents. Thus tertiary amines are more basic than secondary amines, which are more basic than primary amines, and finally ammonia is least basic. The order of pK b 's (basicities in water) does not follow this order. Similarly aniline is more basic than ammonia in
6767-615: The larval gills, tail and fins in amphibians metamorphosis , and stimulate the evolution of their nervous system transforming the aquatic, vegetarian tadpole into the terrestrial, carnivorous frog. Stem cells of this frog were used to create xenobots . Early work on sequencing of the X. laevis genome was started when the Wallingford and Marcotte labs obtained funding from the Texas Institute for Drug and Diagnostic Development (TI3D), in conjunction with projects funded by
6868-746: The lower North Island in New Zealand in 2005, in eastern France in 2003, in California of the United States in 2002 and 2006, in Scotland in 1992, in Ireland in 1997 and 2005, in Germany in 2017 and 2020. In each case, the dogs began showing muscle convulsions within minutes, and were dead within a matter of hours. Numerous cattle fatalities arising from the consumption of water contaminated with cyanobacteria that produce anatoxin-a have been reported in
6969-425: The male clasps the female in front of the female's back legs until eggs are laid, and the male fertilizes the egg mass with the release of sperm. African clawed frogs are highly adaptable and will lay their eggs whenever conditions allow it. During wet rainy seasons they will travel to other ponds or puddles of water to search for food and new ponds. During times of drought, the clawed frogs can burrow themselves into
7070-444: The male. This frog has smooth slippery skin which is multicolored on its back with blotches of olive gray or brown. The underside is creamy white with a yellow tinge. Male and female frogs can be easily distinguished through the following differences. Male frogs are small and slim, while females are larger and more rotund. Males have black patches on their hands and arms which aid in grabbing onto females during amplexus. Females have
7171-474: The mud, becoming dormant for up to a year. Xenopus laevis have been known to survive 15 or more years in the wild and 25–30 years in captivity. They shed their skin every season, and eat their own shed skin. Although lacking a vocal sac , the males make a mating call of alternating long and short trills, by contracting the intrinsic laryngeal muscles . Females also answer vocally, signaling either acceptance (a rapping sound) or rejection (slow ticking) of
7272-498: The name Very Fast Death Factor. They determined the LD50 for mice to be 250 μg/kg body weight. Electrophysiological experiments done by Spivak et al. (1980) on frogs showed that anatoxin-a is a potent agonist of the muscle-type (α 1 ) 2 βγδ nAChR. Anatoxin-a induced depolarizing neuromuscular blockade, contracture of the frog's rectus abdominis muscle, depolarization of the frog sartorius muscle, desensitization, and alteration of
7373-454: The nature and number of substituents on nitrogen . Aliphatic amines contain only H and alkyl substituents. Aromatic amines have the nitrogen atom connected to an aromatic ring. Amines, alkyl and aryl alike, are organized into three subcategories (see table) based on the number of carbon atoms adjacent to the nitrogen (how many hydrogen atoms of the ammonia molecule are replaced by hydrocarbon groups): A fourth subcategory
7474-678: The nitroaromatics. In industry, hydrogen is the preferred reductant, whereas, in the laboratory, tin and iron are often employed. Many methods exist for the preparation of amines, many of these methods being rather specialized. Aside from their basicity, the dominant reactivity of amines is their nucleophilicity . Most primary amines are good ligands for metal ions to give coordination complexes . Amines are alkylated by alkyl halides. Acyl chlorides and acid anhydrides react with primary and secondary amines to form amides (the " Schotten–Baumann reaction "). Similarly, with sulfonyl chlorides, one obtains sulfonamides . This transformation, known as
7575-444: The oocyte or developing embryo, scientists can study the protein products in a controlled system. This allows rapid functional expression of manipulated DNAs (or mRNA ). This is particularly useful in electrophysiology , where the ease of recording from the oocyte makes expression of membrane channels attractive. One challenge of oocyte work is eliminating native proteins that might confound results, such as membrane channels native to
7676-475: The oral minimum lethal dose (MLD) (of the algae, not the anatoxin molecule), for calves is roughly 420 mg/kg body weight. In the same year, Devlin and colleagues discovered the bicyclic secondary amine structure of anatoxin-a. They also performed experiments similar to those of Carmichael et al. on mice. They found that anatoxin-a kills mice 2–5 minutes after intraperitoneal injection preceded by twitching, muscle spasms, paralysis and respiratory arrest, hence
7777-1262: The original structure of the amino acid is modified such that a –COOH, or carboxyl, group is removed, whereas the –NH 3 , or amine, group remains. Amine hormones are synthesized from the amino acids tryptophan or tyrosine . Primary aromatic amines are used as a starting material for the manufacture of azo dyes . It reacts with nitrous acid to form diazonium salt, which can undergo coupling reaction to form an azo compound. As azo-compounds are highly coloured, they are widely used in dyeing industries, such as: Most drugs and drug candidates contain amine functional groups: Aqueous monoethanolamine (MEA), diglycolamine (DGA), diethanolamine (DEA), diisopropanolamine (DIPA) and methyldiethanolamine (MDEA) are widely used industrially for removing carbon dioxide (CO 2 ) and hydrogen sulfide (H 2 S) from natural gas and refinery process streams. They may also be used to remove CO 2 from combustion gases and flue gases and may have potential for abatement of greenhouse gases . Related processes are known as sweetening . Xenopus laevis X. boiei Wagler 1827 The African clawed frog ( Xenopus laevis ), also known as simply xenopus , African clawed toad , African claw-toed frog or
7878-599: The presence of a nickel catalyst. Suitable groups include nitriles , azides , imines including oximes , amides, and nitro . In the case of nitriles, reactions are sensitive to acidic or alkaline conditions, which can cause hydrolysis of the −CN group. LiAlH 4 is more commonly employed for the reduction of these same groups on the laboratory scale. Many amines are produced from aldehydes and ketones via reductive amination , which can either proceed catalytically or stoichiometrically. Aniline ( C 6 H 5 NH 2 ) and its derivatives are prepared by reduction of
7979-479: The presence of strong acids to give formamides, which can be decarbonylated. This method, the Ritter reaction , is used industrially to produce tertiary amines such as tert -octylamine . Hydroamination of alkenes is also widely practiced. The reaction is catalyzed by zeolite-based solid acids . Via the process of hydrogenation , unsaturated N-containing functional groups are reduced to amines using hydrogen in
8080-427: The properties of primary and secondary amines. For example, methyl and ethyl amines are gases under standard conditions, whereas the corresponding methyl and ethyl alcohols are liquids. Amines possess a characteristic ammonia smell, liquid amines have a distinctive "fishy" and foul smell. The nitrogen atom features a lone electron pair that can bind H to form an ammonium ion R 3 NH . The lone electron pair
8181-403: The reaction of amines with alcohols the reaction of amines and ammonia with alkyl halides is used for synthesis in the laboratory: In such reactions, which are more useful for alkyl iodides and bromides, the degree of alkylation is difficult to control such that one obtains mixtures of primary, secondary, and tertiary amines, as well as quaternary ammonium salts. Selectivity can be improved via
8282-440: The reactions of amines is given below: Amines are ubiquitous in biology. The breakdown of amino acids releases amines, famously in the case of decaying fish which smell of trimethylamine . Many neurotransmitters are amines, including epinephrine , norepinephrine , dopamine , serotonin , and histamine . Protonated amino groups ( –NH 3 ) are the most common positively charged moieties in proteins , specifically in
8383-447: The same effects in neurons. However, anatoxin-a binding is irreversible , and the anatoxin-a nAchR complex cannot be broken down by acetylcholinesterase . Thus, the nAchR is temporarily locked open, which leads to overstimulation due to the constant generation of action potentials. Two enantiomers of anatoxin-a, the positive enantiomer , (+)-anatoxin-a, is 150 fold more potent than the synthetic negative enantiomer, (−)-anatoxin-a. This
8484-416: The same function. They use their sensitive fingers and sense of smell to find food. Pipidae are scavengers and will eat almost anything living, dying, or dead and any type of organic waste. It is considered an invasive species in several countries , including across Europe. These frogs are plentiful in ponds and rivers within the south-eastern portion of Sub-Saharan Africa. They are aquatic and are often
8585-409: The state of Washington implemented a recreational advisory level for anatoxin-a of 1 μg/L in order to better manage algal blooms in lakes and protect users from exposure to the blooms. The Canadian province of Québec has a drinking water Maximum Accepted Value for anatoxin-a of 3.7 μg/L. New Zealand has a drinking water Maximum Accepted Value for anatoxin-a of 6 μg/L. As of now, there
8686-692: The stomach contents of feral clawed frogs in California have revealed large amounts of the frog's larvae. Clawed frog larvae are filter feeders and collect nutrients from plankton, allowing adult frogs that consume the tadpoles to have access to these nutrients. This allows clawed frogs to survive in areas that have little to no other food sources. Clawed frogs are nocturnal and most reproductive activity and feeding occurs after dark. Male clawed frogs are very promiscuous and will grab onto other males and even other species of frogs. Male frogs that are grasped will make release calls and attempt to break free. If not feeding, clawed frogs will just sit motionless on top of
8787-445: The stomachs of two young male calves, and observed that muscular fasciculations and loss of coordination occurred in a matter of minutes, while death due to respiratory failure occurred anywhere between several minutes and a few hours. They also established that extensive periods of artificial respiration did not allow for detoxification to occur and natural neuromuscular functioning to resume. From these experiments, they calculated that
8888-590: The substrate or floating, legs splayed below, at the waters surface with their nostrils and eyes sticking out. The clawed frog liver responds to low temperatures by increasing production of type II iodothyronine deiodinase through increased food intake . This in turn spurs the thyroid to increase T 3 to increase body temperature . (This T 3 increase also induces germ cell apoptosis , mediated through genes left over from tadpole metamorphosis.) The effects of provocation of T hormone release are broadly differentiated by where it starts: If centrally, within
8989-452: The suffix "-amine". The prefix " N -" shows substitution on the nitrogen atom. An organic compound with multiple amino groups is called a diamine , triamine , tetraamine and so forth. Lower amines are named with the suffix -amine . Higher amines have the prefix amino as a functional group. IUPAC however does not recommend this convention, but prefers the alkanamine form, e.g. butan-2-amine. Hydrogen bonding significantly influences
9090-561: The super short generation time , or genetic simplicity generally desired in genetic model organisms , it is an important model organism in developmental biology , cell biology , toxicology and neurobiology . X. laevis takes 1 to 2 years to reach sexual maturity and, like most of its genus, it is tetraploid . It does have a large and easily manipulated embryo , however. The ease of manipulation in amphibian embryos has given them an important place in historical and modern developmental biology. A related species, Xenopus tropicalis ,
9191-532: The table below. On October 8, 2009 the EPA published the third Drinking Water Contaminant Candidate List (CCL) which included anatoxin-a (among other cyanotoxins), indicating that anatoxin-a may be present in public water systems but is not regulated by the EPA. Anatoxin-a's presence on the CCL means that it may need to be regulated by the EPA in the future, pending further information on its health effects in humans. In 2008
9292-494: The use of diethyl azodicarboxylate, the ketone is demethylated and anatoxin-a is formed. A similar, more recent synthesis pathway involves producing 2-tropinone from cocaine and treating the product with ethyl chloroformate producing a bicyclic ketone. This product is combined with trimethylsilyldiazylmethane, an organoaluminum Lewis acid and trimethylsinyl enol ether to produce tropinone. This method undergoes several more steps, producing useful intermediates as well as anatoxin-a as
9393-676: The water except to migrate to new water bodies during droughts or other disturbances. Clawed frogs have powerful legs that help them move quickly both underwater and on land. Feral clawed frogs in South Wales have been found to travel up to 2 kilometres (1.2 mi) between locations. The feet of Xenopus species have three black claws on the last three digits. These claws are used to rip apart food and scratch predators. Clawed frogs are carnivores and will eat both living and dead prey including fish, tadpoles, crustaceans, annelids, arthropods, and more. Clawed frogs will try to consume anything that
9494-654: The wide breadth of Xenopus research stems from the additional fact that cell-free extracts made from Xenopus are a premier in vitro system for studies of fundamental aspects of cell and molecular biology. Thus, Xenopus is the only vertebrate model system that allows for high-throughput in vivo analyses of gene function and high-throughput biochemistry. Xenopus oocytes are a leading system in their own right for studies of various systems, including ion transport and channel physiology. Xanthos et al 2001 uses oocytes to uncover T-box expression earlier than previously found in vertebrates. Although X. laevis does not have
9595-555: The wild - Found at higher densities in artificial water bodies such as ponds, dams and irrigation canals, rather than in natural lagoons or streams or rivers. - There is no evidence of predation on native anurans, but rather on their own larvae. - They face predation from native birds. Cause of concerns from African clawed frogs - They are reaching both lower and higher altitudes than formerly estimated. - They are able to migrate overland to colonise other water bodies, causing ecological disruption and spreading diseases. X. laevis in
9696-472: The wild are commonly infected by various parasites , including monogeneans in the urinary bladder . Xenopus embryos and eggs are a popular model system for a wide variety of biological studies, in part because they have the potential to lay eggs throughout the year. This animal is widely used because of its powerful combination of experimental tractability and close evolutionary relationship with humans, at least compared to many model organisms. For
9797-614: The wild, X. laevis are native to wetlands , ponds, and lakes across arid/semiarid regions of Sub-Saharan Africa . X. laevis and X. muelleri occur along the western boundary of the Great African Rift . The people of the sub-Saharan are generally very familiar with this frog, and some cultures use it as a source of protein, an aphrodisiac , or as fertility medicine . Two historic outbreaks of priapism have been linked to consumption of frog legs from frogs that ate insects containing cantharidin . African clawed frogs in
9898-642: Was isolated in 1972 by J.P. Devlin from the cyanobacteria Anabaena flos-aquae . Anatoxin-a is a neurotoxin produced by multiple genera of freshwater cyanobacteria that are found in water bodies globally. Some freshwater cyanobacteria are known to be salt tolerant and thus it is possible for anatoxin-a to be found in estuarine or other saline environments. Blooms of cyanobacteria that produce anatoxin-a among other cyanotoxins are increasing in frequency due to increasing temperatures, stratification , and eutrophication due to nutrient runoff. These expansive cyanobacterial harmful algal blooms , known as cyanoHABs, increase
9999-499: Was reported in Wisconsin after a teen jumped into a pond contaminated with cyanobacteria. Ingestion of drinking water or recreational water that is contaminated with anatoxin-a can pose fatal consequences since anatoxin-a was found to be quickly absorbed through the gastrointestinal tract in animal studies. Dozens of cases of animal deaths due to ingestion of anatoxin-a contaminated water from lakes or rivers have been recorded, and it
10100-401: Was transformed into an alpha, beta ketone and oxidized by ethyl azodicarboxylate to form anatoxin-a. This method for anatoxin-a production was one of the first used that does not utilize a chimerically analogous starting substance for anatoxin formation. Instead, a racemic mixture of 3-tropinone is used with a chiral lithium amide base and additional ring expansion reactions in order to produce
10201-528: Was used as a simple and reliable test up through to the 1960s. In the late 1940s, Carlos Galli Mainini found in separate studies that male specimens of Xenopus and Bufo could be used to indicate pregnancy Today, commercially available hCG is injected into Xenopus males and females to induce mating behavior and to breed these frogs in captivity at any time of the year. Xenopus has long been an important tool for in vivo studies in molecular, cell, and developmental biology of vertebrate animals. However,
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