How the Neurotoxin ß-N-Methylamino-l-Alanine Accumulates in Bivalves: Distribution of the Different Accumulation Fractions among Organs.

The environmental neurotoxin ß-methylamino-l-alanine (BMAA) may represent a risk for human health. BMAA accumulates in freshwater and marine organisms consumed by humans. However, few data are available about the kinetics of BMAA accumulation and detoxification in exposed organisms, as well as the organ distribution and the fractions in which BMAA is present in tissues (free, soluble bound or precipitated bound cellular fractions). Here, we exposed the bivalve mussel Dreissena polymorpha to 7.5 µg of dissolved BMAA/mussel/3 days for 21 days, followed by 21 days of depuration in clear water. At 1, 3, 8, 14 and 21 days of exposure and depuration, the hemolymph and organs (digestive gland, the gills, the mantle, the gonad and muscles/foot) were sampled. Total BMAA as well as free BMAA, soluble bound and precipitated bound BMAA were quantified by tandem mass spectrometry. Free and soluble bound BMAA spread throughout all tissues from the first day of exposure to the last day of depuration, without a specific target organ. However, precipitated bound BMAA was detected only in muscles and foot from the last day of exposure to day 8 of depuration, at a lower concentration compared to free and soluble bound BMAA. In soft tissues (digestive gland, gonad, gills, mantle and muscles/foot), BMAA mostly accumulated as a free molecule and in the soluble bound fraction, with variations occurring between the two fractions among tissues and over time. The results suggest that the assessment of bivalve contamination by BMAA may require the quantification of total BMAA in whole individuals when possible.

Gelsedine-type alkaloids: Discovery of natural neurotoxins presented in toxic honey.

The fascinating collection and evaluation of natural products with enormous structural and chemical diversity can contribute to ensure human health and inspire potential drug discovery. We reported the identification of 14-(R)-hydroxy-gelsenicine (HGE), a new component from poisonous honey, which has recently caused multiple serious intoxications and deaths up on consumption. The prevalence, toxicity, toxicokinetics and metabolic profile of HGE were evaluated through in vitro and in vivo analyses. HGE is a very toxic substance and shows significant gender difference with LD50 of 0.125 mg kg-1 and 0.295 mg kg-1 for the female and male mice, respectively. Toxicokinetics test indicates that HGE has good bioavailability in rats, and is metabolized extensively, in which hydroxylation, reduction, N-demethyl ether and glucuronication are the major metabolic pathways. Additionally, HGE shows specific neurotoxicity by enhancing the binding of γ-aminobutyric acid (GABA) to its receptors. We found that flumazenil, a selective antagonist of GABA receptor, could effectively increase the survival of the tested animals, which provides a potential therapy for future clinical applications.

Three-dimensional (3D) brain microphysiological system for organophosphates and neurochemical agent toxicity screening.

We investigated a potential use of a 3D tetraculture brain microphysiological system (BMPS) for neurotoxic chemical agent screening. This platform consists of neuronal tissue with extracellular matrix (ECM)-embedded neuroblastoma cells, microglia, and astrocytes, and vascular tissue with dynamic flow and membrane-free culture of the endothelial layer. We tested the broader applicability of this model, focusing on organophosphates (OPs) Malathion (MT), Parathion (PT), and Chlorpyrifos (CPF), and chemicals that interact with GABA and/or opioid receptor systems, including Muscimol (MUS), Dextromethorphan (DXM), and Ethanol (EtOH). We validated the BMPS platform by measuring the neurotoxic effects on barrier integrity, acetylcholinesterase (AChE) inhibition, viability, and residual OP concentration. The results show that OPs penetrated the model blood brain barrier (BBB) and inhibited AChE activity. DXM, MUS, and EtOH also penetrated the BBB and induced moderate toxicity. The results correlate well with available in vivo data. In addition, simulation results from an in silico physiologically-based pharmacokinetic/pharmacodynamic (PBPK/PD) model that we generated show good agreement with in vivo and in vitro data. In conclusion, this paper demonstrates the potential utility of a membrane-free tetraculture BMPS that can recapitulate brain complexity as a cost-effective alternative to animal models.

A pharmacokinetic model of lead absorption and calcium competitive dynamics.

Lead is a naturally-occurring element. It has been known to man for a long time, and it is one of the longest established poisons. The current consensus is that no level of lead exposure should be deemed "safe". New evidence regarding the blood levels at which morbidities occur has prompted the CDC to reduce the screening guideline of 10 µg/dl to 2 µg/dl. Measurable cognitive decline (reduced IQ, academic deficits) have been found to occur at levels below 10 µg/dl, especially in children. Knowledge of lead pharmacology allows us to better understand its absorption and metabolization, mechanisms that produce its medical consequences. Based upon an original and very simplified compartmental model of Rabinowitz (1973) with only three major compartments (blood, bone and soft tissue), extensive biophysical models sprouted over the following two decades. However, none of these models have been specifically designed to use new knowledge of lead molecular dynamics to understand its deleterious effects on the brain. We build and analyze a compartmental model of lead pharmacokinetics, focused specifically on addressing neurotoxicity. We use traditional phase space methods, parameter sensitivity analysis and bifurcation theory to study the transitions in the system's behavior in response to various physiological parameters. We conclude that modeling the complex interaction of lead and calcium along their dynamic trajectory may successfully explain counter-intuitive effects on systemic function and neural behavior which could not be addressed by existing linear models. Our results encourage further efforts towards using nonlinear phenomenology in conjunction with empirically driven system parameters, to obtain a biophysical model able to provide clinical assessments and predictions.

Tetrodotoxin in marine bivalves and edible gastropods: A mini-review.

Tetrodotoxin (TTX) is a potent neurotoxin responsible for countless human intoxications and deaths around the world. The distribution of TTX and its analogues is diverse and the toxin has been detected in organisms from both marine and terrestrial environments. Increasing detections seafood species, such as bivalves and gastropods, has drawn attention to the toxin, reinvigorating scientific interest and regulatory concerns. There have been reports of TTX in 21 species of bivalves and edible gastropods from ten countries since the 1980's. While TTX is structurally dissimilar to saxitoxin (STX), another neurotoxin detected in seafood, it has similar sodium channel blocking action and potency and both neurotoxins have been shown to have additive toxicities. The global regulatory level for the STX group toxins applied to shellfish is 800 µg/kg. The presence of TTX in shellfish is only regulated in one country; The Netherlands, with a regulatory level of 44 µg/kg. Due to the recent interest surrounding TTX in bivalves, the European Food Safety Authority established a panel to assess the risk and regulation of TTX in bivalves, and their final opinion was that a concentration below 44 µg of TTX per kg of shellfish would not result in adverse human effects. In this article, we review current knowledge on worldwide TTX levels in edible gastropods and bivalves over the last four decades, the different methods of detection used, and the current regulatory status. We suggest research needs that will assist with knowledge gaps and ultimately allow development of robust monitoring and management protocols.

Transfer of a cyanobacterial neurotoxin, ß-methylamino-l-alanine from soil to crop and its bioaccumulation in Chinese cabbage.

Most cyanobacteria can synthesize the notorious neurotoxin ß-methylamino-l-alanine (BMAA) that is transferred and bioaccumulated through natural food webs of aquatic ecosystems and ultimately arises the potential human health risks by the consumption of BMAA-contaminated aquatic products. Fertilization of cyanobacterial composts in farmlands may also lead to BMAA contamination in soil and its possible transfer and bioaccumulation within major crops, thereby threatening human health. In this study, we first detected a high level of BMAA (1.8-16.3 µg g-1) in cyanobacterial composts. In order to assess the health risks from cyanobacterial composts, we planted Chinese cabbage, a favourite vegetable in China, in BMAA-contaminated soil (4.0 µg BMAA/g soil) and detected the levels of free and protein-associated BMAA in soil and crop organs during the whole growth cycle by HPLC-MS/MS, respectively. Our results demonstrated that BMAA indeed transferred from soil to root, stem and leaf of Chinese cabbage during the growth cycle. The BMAA level finally accumulated in the edible portions was much higher than the initial level in soil, including 13.82 µg g-1 in leaf and 4.71 µg g-1 in stem. The discovery of the neurotoxin BMAA in this vegetable, an ending in human consumption, not only provides a BMAA transfer pathway in farmland ecosystems, but also is alarming and requires attention due to the potential risk of cyanobacterial composts to human health.

Glutathione antioxidant system and methylmercury-induced neurotoxicity: An intriguing interplay.

Methylmercury (MeHg) is a toxic chemical compound naturally produced mainly in the aquatic environment through the methylation of inorganic mercury catalyzed by aquatic microorganisms. MeHg is biomagnified in the aquatic food chain and, consequently, piscivorous fish at the top of the food chain possess huge amounts of MeHg (at the ppm level). Some populations that have fish as main protein's source can be exposed to exceedingly high levels of MeHg and develop signs of toxicity. MeHg is toxic to several organs, but the central nervous system (CNS) represents a preferential target, especially during development (prenatal and early postnatal periods). Though the biochemical events involved in MeHg-(neuro)toxicity are not yet entirely comprehended, a vast literature indicates that its pro-oxidative properties explain, at least partially, several of its neurotoxic effects. As result of its electrophilicity, MeHg interacts with (and oxidize) nucleophilic groups, such as thiols and selenols, present in proteins or low-molecular weight molecules. It is noteworthy that such interactions modify the redox state of these groups and, therefore, lead to oxidative stress and impaired function of several molecules, culminating in neurotoxicity. Among these molecules, glutathione (GSH; a major thiol antioxidant) and thiol- or selenol-containing enzymes belonging to the GSH antioxidant system represent key molecular targets involved in MeHg-neurotoxicity. In this review, we firstly present a general overview concerning the neurotoxicity of MeHg. Then, we present fundamental aspects of the GSH-antioxidant system, as well as the effects of MeHg on this system.

Short-chain consensus alpha-neurotoxin: a synthetic 60-mer peptide with generic traits and enhanced immunogenic properties.

The three-fingered toxin family and more precisely short-chain α-neurotoxins (also known as Type I α-neurotoxins) are crucial in defining the elapid envenomation process, but paradoxically, they are barely neutralized by current elapid snake antivenoms. This work has been focused on the primary structural identity among Type I neurotoxins in order to create a consensus short-chain α-neurotoxin with conserved characteristics. A multiple sequence alignment considering the twelve most toxic short-chain α-neurotoxins reported from the venoms of the elapid genera Acanthophis, Oxyuranus, Walterinnesia, Naja, Dendroaspis and Micrurus led us to propose a short-chain consensus α-neurotoxin, here named ScNtx. The synthetic ScNtx gene was de novo constructed and cloned into the expression vector pQE30 containing a 6His-Tag and an FXa proteolytic cleavage region. Escherichia coli Origami cells transfected with the pQE30/ScNtx vector expressed the recombinant consensus neurotoxin in a soluble form with a yield of 1.5 mg/L of culture medium. The 60-amino acid residue ScNtx contains canonical structural motifs similar to α-neurotoxins from African elapids and its LD50 of 3.8 µg/mice is similar to the most toxic short-chain α-neurotoxins reported from elapid venoms. Furthermore, ScNtx was also able to antagonize muscular, but not neuronal, nicotinic acetylcholine receptors (nAChR). Rabbits immunized with ScNtx were able to immune-recognize short-chain α-neurotoxins within whole elapid venoms. Type I neurotoxins are difficult to isolate and purify from natural sources; therefore, the heterologous expression of molecules such ScNtx, bearing crucial motifs and key amino acids, is a step forward to create common immunogens for developing cost-effective antivenoms with a wider spectrum of efficacy, quality and strong therapeutic value.

El modelo de 6-hidroxidopamina y la fisiopatología parkinsoniana: nuevos hallazgos en un viejo modelo / The 6-hydroxydopamine model and parkinsonian pathophysiology: Novel findings in an older model

El neurotóxico 6-hidroxidopamina (6-OHDA) ha sido utilizado para generar modelos de la enfermedad de Parkinson (EP). A la fecha se ha establecido que si bien el modelo neurodegenerativo inducido por la 6-OHDA no reproduce la totalidad de síntomas de la enfermedad, sí replica procesos celulares tales como el estrés oxidativo, la neurodegeneración, la neuroinflamación y la muerte neuronal por apoptosis. En esta revisión se contempla el análisis de los factores que influyen en la vulnerabilidad de las neuronas dopaminérgicas, así como la estrecha relación entre el proceso neurodegenerativo, el neuroinflamatorio y la apoptosis ocasionada por la 6-OHDA. El conocimiento de los mecanismos involucrados en la neurodegeneración y la muerte celular en este modelo es relevante para definir posibles blancos terapéuticos para EP (AU)

The neurotoxin 6-hydroxydopamine (6-OHDA) is widely used to induce models of Parkinson's disease (PD). We now know that the model induced by 6-OHDA does not include all PD symptoms, although it does reproduce the main cellular processes involved in PD, such as oxidative stress, neurodegeneration, neuroinflammation, and neuronal death by apoptosis. In this review we analyse the factors affecting the vulnerability of dopaminergic neurons as well as the close relationships between neuroinflammation, neurodegeneration, and apoptosis in the 6-OHDA model. Knowledge of the mechanisms involved in neurodegeneration and cell death in this model is the key to identifying potential therapeutic targets for PD (AU)

Design of modified botulinum neurotoxin A1 variants with a shorter persistence of paralysis and duration of action.

Botulinum neurotoxins (BoNTs) are classified by their antigenic properties into seven serotypes (A-G) and in addition by their corresponding subtypes. They are further characterized by divergent onset and duration of effect. Injections of low doses of botulinum neurotoxins cause localized muscle paralysis that is beneficial for the treatment of several medical disorders and aesthetic indications. Optimizing the therapeutic properties could offer new treatment opportunities. This report describes a rational design approach to modify the pharmacological properties by mutations in the C-terminus of BoNT/A1 light chain (LC). Toxins with C-terminal modified LC's displayed an altered onset and duration of the paralytic effect in vivo. The level of effect was dependent on the kind of the mutation in the sequence of the C-terminus. A mutant with three mutations (T420E F423M Y426F) revealed a faster onset and a shorter duration than BoNT/A1 wild type (WT). It could be shown that the C-terminus of BoNT/A1-Lc controls both onset and duration of effect. Thus, it is possible to create a mutated BoNT/A1 with different pharmacological properties which might be useful in the therapy of new indications. This strategy opens the way to design BoNT variants with novel and useful properties.

Determination of selected neurotoxic insecticides in small amounts of animal tissue utilizing a newly constructed mini-extractor.

We developed a simple analytical method for the simultaneous determination of representatives of various groups of neurotoxic insecticides (carbaryl, chlorpyrifos, cypermethrin, and α-endosulfan and ß-endosulfan and their metabolite endosulfan sulfate) in limited amounts of animal tissues containing different amounts of lipids. Selected tissues (rodent fat, liver, and brain) were extracted in a special in-house-designed mini-extractor constructed on the basis of the Soxhlet and Twisselmann extractors. A dried tissue sample placed in a small cartridge was extracted, while the nascent extract was simultaneously filtered through a layer of sodium sulfate. The extraction was followed by combined clean-up, including gel permeation chromatography (in case of high lipid content), ultrasonication, and solid-phase extraction chromatography using C18 on silica and aluminum oxide. Gas chromatography coupled with high-resolution mass spectrometry was used for analyte separation, detection, and quantification. Average recoveries for individual insecticides ranged from 82 to 111%. Expanded measurement uncertainties were generally lower than 35%. The developed method was successfully applied to rat tissue samples obtained from an animal model dealing with insecticide exposure during brain development. This method may also be applied to the analytical treatment of small amounts of various types of animal and human tissue samples. A significant advantage achieved using this method is high sample throughput due to the simultaneous treatment of many samples. Graphical abstract Optimized workflow for the determination of selected insecticides in small amounts of animal tissue including newly developed mini-extractor.

Tissue uptake, distribution and excretion of brevetoxin-3 after oral and intratracheal exposure in the freshwater turtle Trachemys scripta and the diamondback terrapin Malaclemys terrapin.

Harmful algal blooms (HABs) occur nearly annually off the west coast of Florida and can impact both humans and wildlife, resulting in morbidity and increased mortality of marine animals including sea turtles. The key organism in Florida red tides is the dinoflagellate Karenia brevis that produces a suite of potent neurotoxins referred to as the brevetoxins (PbTx). Despite recent mortality events and rehabilitation efforts, still little is known about how the toxin directly impacts sea turtles, as they are not amenable to experimentation and what is known about toxin levels and distribution comes primarily from post-mortem data. In this study, we utilized the freshwater turtle Trachemys scripta and the diamondback terrapin, Malaclemys terrapin as model organisms to determine the distribution, clearance, and routes of excretion of the most common form of the toxin, brevetoxin-3, in turtles. Turtles were administered toxin via esophageal tube to mimic ingestion (33.48µg/kg PbTx-3, 3×/week for two weeks for a total of 7 doses) or by intratracheal instillation (10.53µg/kg, 3×/week for four weeks for a total of 12 doses) to mimic inhalation. Both oral and intratracheal administration of the toxin produced a suite of behavioral responses symptomatic of brevetoxicosis. The toxin distributed to all organ systems within 1h of administration but was rapidly cleared out over 24-48h, corresponding to a decline in clinical symptoms. Excretion appears to be primarily through conjugation to bile salts. Histopathological study revealed that the frequency of lesions varied within experimental groups with some turtles having no significant lesions at all, while similar lesions were found in a low number of control turtles suggesting another common factor(s) could be responsible. The overall goal of this research is better understand the impacts of brevetoxin on turtles in order to develop better treatment protocols for sea turtles exposed to HABs.

Brain Mitochondrial Subproteome of Rpn10-Binding Proteins and Its Changes Induced by the Neurotoxin MPTP and the Neuroprotector Isatin.

Mitochondria play an important role in molecular mechanisms of neuroplasticity, adaptive changes of the brain that occur in the structure and function of its cells in response to altered physiological conditions or development of pathological disorders. Mitochondria are a crucial target for actions of neurotoxins, causing symptoms of Parkinson's disease in various experimental animal models, and also neuroprotectors. Good evidence exists in the literature that mitochondrial dysfunction induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) influences functioning of the ubiquitin-proteasomal system (UPS) responsible for selective proteolytic degradation of proteins from various intracellular compartments (including mitochondria), and neuroprotective effects of certain antiparkinsonian agents (monoamine oxidase inhibitors) may be associated with their effects on UPS. The 19S proteasomal Rpn10 subunit is considered as a ubiquitin receptor responsible for delivery of ubiquitinated proteins to the proteasome proteolytic machinery. In this study, we investigated proteomic profiles of mouse brain mitochondrial Rpn10-binding proteins, brain monoamine oxidase B (MAO B) activity, and their changes induced by a single-dose administration of the neurotoxin MPTP and the neuroprotector isatin. Administration of isatin to mice prevented MPTP-induced inactivation of MAO B and influenced the profile of brain mitochondrial Rpn10-binding proteins, in which two pools of proteins were clearly recognized. The constitutive pool was insensitive to neurotoxic/neuroprotective treatments, while the variable pool was specifically influenced by MPTP and the neuroprotector isatin. Taking into consideration that the neuroprotective dose of isatin used in this study can result in brain isatin concentrations that are proapoptotic for cells in vitro, the altered repertoire of mitochondrial Rpn10-binding proteins may thus represent a part of a switch mechanism from targeted elimination of individual (damaged) proteins to more efficient ("global") elimination of damaged organelles and whole damaged cells.

Physiological and behavioural impacts of Pacific ciguatoxin-1 (P-CTX-1) on marine medaka (Oryzias melastigma).

Ciguatoxins (CTXs) are natural biotoxins produced by benthic dinoflagellates of the genus Gambierdiscus, which are bioaccumulated and biotransformed along food chains in coral ecosystems. They are neurotoxins that activate voltage-gated sodium channels and disrupt ion conductance in the excitable tissues. Pacific ciguatoxin-1 (P-CTX-1) is the most prevalent and potent CTX congener present in fishes from the Pacific Ocean. In this study, P-CTX-1 was administrated to larval marine medaka (2h post-hatch) via microinjection. Exposure to P-CTX-1 at sub-ppb levels led to adverse behavioural changes, altered physiological performances and reduced survivability of the larval marine medaka as early as 24h after exposure. P-CTX-1 decreased the rate of heartbeat and locomotion of the exposed larvae, probably owing to a series of physiological processes and morphological changes such as pericardial oedema, failure of swim bladder inflation and spinal curvature. The exposed larval marine medaka also demonstrated reduced, delayed and paralyzed responses to external stimulations. This may render them more susceptible to predation. P-CTX-1 could be effectively distributed from the yolk sac to all parts of the fish body, including head and trunk, 24h after exposure. Repeated low-dose P-CTX-1 exposure resulted in larval mortality comparable to that of a single high-dose exposure.

Biodistribution and Lymphatic Tracking of the Main Neurotoxin of Micrurus fulvius Venom by Molecular Imaging.

The venom of the Eastern coral snake Micrurus fulvius can cause respiratory paralysis in the bitten patient, which is attributable to ß-neurotoxins (ß-NTx). The aim of this work was to study the biodistribution and lymphatic tracking by molecular imaging of the main ß-NTx of M. fulvius venom. ß-NTx was bioconjugated with the chelator diethylenetriaminepenta-acetic acid (DTPA) and radiolabeled with the radionuclide Gallium-67. Radiolabeling efficiency was 60%-78%; radiochemical purity ≥92%; and stability at 48 h ≥ 85%. The median lethal dose (LD50) and PLA2 activity of bioconjugated ß-NTx decreased 3 and 2.5 times, respectively, in comparison with native ß-NTx. The immune recognition by polyclonal antibodies decreased 10 times. Biodistribution of ß-NTx-DTPA-(67)Ga in rats showed increased uptake in popliteal, lumbar nodes and kidneys that was not observed with (67)Ga-free. Accumulation in organs at 24 h was less than 1%, except for kidneys, where the average was 3.7%. The inoculation site works as a depot, since 10% of the initial dose of ß-NTx-DTPA-(67)Ga remains there for up to 48 h. This work clearly demonstrates the lymphatic system participation in the biodistribution of ß-NTx-DTPA-(67)Ga. Our approach could be applied to analyze the role of the lymphatic system in snakebite for a better understanding of envenoming.

Multifactorial theory applied to the neurotoxicity of paraquat and paraquat-induced mechanisms of developing Parkinson's disease.

Laboratory studies involving repeated exposure to paraquat (PQ) in different animal models can induce many of the pathological features of Parkinson's disease (PD), such as the loss of dopaminergic neurons in the nigrostriatal dopamine system. Epidemiological studies identify an increased risk of developing PD in human populations living in areas where PQ exposure is likely to occur and among workers lacking appropriate protective equipment. The mechanisms involved in developing PD may not be due to any single cause, but rather a multifactorial situation may exist where PQ exposure may cause PD in some circumstances. Multifactorial theory is adopted into this review that includes a number of sub-cellular mechanisms to explain the pathogenesis of PD. The theory is placed into an environmental context of chronic low-dose exposure to PQ that consequently acts as an oxidative stress inducer. Oxidative stress and the metabolic processes of PQ-inducing excitotoxicity, α-synuclein aggregate formation, autophagy, alteration of dopamine catabolism, and inactivation of tyrosine hydroxylase are positioned as causes for the loss of dopaminergic cells. The environmental context and biochemistry of PQ in soils, water, and organisms is also reviewed to identify potential routes that can lead to chronic rates of low-dose exposure that would replicate the type of response that is observed in animal models, epidemiological studies, and other types of laboratory investigations involving PQ exposure. The purpose of this review is to synthesize key relations and summarize hypotheses linking PD to PQ exposure by using the multifactorial approach. Recommendations are given to integrate laboratory methods to the environmental context as a means to improve on experimental design. The multifactorial approach is necessary for conducting valid tests of causal relations, for understanding of potential relations between PD and PQ exposure, and may prevent further delay in solving what has proven to be an evasive etiological problem.

Method development for quantification of the environmental neurotoxin annonacin in Rat plasma by UPLC-MS/MS and application to a pharmacokinetic study.

Annonacin is an environmental neurotoxin identified in the pulp of several fruits of the Annonaceae family (for example in Annona muricata, Asimina triloba), whose consumption was linked with the occurrence of sporadic atypical Parkinsonism with dementia. Pharmacokinetic parameters of this molecule are unknown. A method for its quantification in Rat plasma was developed, using its analogue annonacinone as an internal standard. Extraction from plasma was performed using ethylacetate with a good recovery. Quantification was performed by UPLC-MS/MS in SRM mode, based on the loss of the γ-methyl-γ-lactone (-112amu) from the sodium-cationized species [M+Na](+) of both annonacin and internal standard. The limit of quantification was 0.25ng/mL. Despite strong matrix effects, a good linearity was obtained over two distinct ranges 0.25-10ng/mL and 10-100ng/mL. The intra- and inter-day precisions (RSD) were lower than 10%, while accuracy was within ±10%. This method was applied to a pharmacokinetic study in the Rat. After oral administration of 10mg/kg annonacin, a Cmax of 7.9±1.5ng/mL was reached at Tmax 0.25h; T1/2 was 4.8±0.7h and apparent distribution volume was 387.9±64.6L. The bioavailability of annonacin was estimated to be 3.2±0.3% of the ingested dose.

A switch-on MRI contrast agent for noninvasive visualization of methylmercury.

This communication presents the first Gd(III)-based T1 MRI contrast agent, o-MeHgGad, for noninvasive visualization of CH3Hg(+). o-MeHgGad showed a relaxivity enhancement of 62% in the presence of 1 equiv. of CH3Hg(+). Moreover, a noticeable contrast enhancement was recorded in the liver, kidney, and intestine of mice exposed to CH3Hg(+). Thus, the newly designed contrast agent has the potential to be used for in vivo bio-imaging of CH3Hg(+) and could be useful for biomedical applications.

Sweet Poisons: Honeys Contaminated with Glycosides of the Neurotoxin Tutin.

Poisonings due to consumption of honeys containing plant toxins have been reported widely. One cause is the neurotoxin tutin, an oxygenated sesquiterpene picrotoxane, traced back to honeybees (Apis mellifera) collecting honeydew produced by passionvine hoppers (Scolypopa australis) feeding on sap of the poisonous shrub tutu (Coriaria spp.). However, a pharmacokinetic study suggested that unidentified conjugates of tutin were also present in such honeys. We now report the discovery, using ion trap LC-MS, of two tutin glycosides and their purification and structure determination as 2-(ß-d-glucopyranosyl)tutin (4) and 2-[6'-(α-d-glucopyranosyl)-ß-d-glucopyranosyl]tutin (5). These compounds were used to develop a quantitative triple quadrupole LC-MS method for honey analysis, which showed the presence of tutin (3.6 ± 0.1 µg/g honey), hyenanchin (19.3 ± 0.5), tutin glycoside (4) (4.9 ± 0.4), and tutin diglycoside (5) (4.9 ± 0.1) in one toxic honey. The ratios of 4 and 5 to tutin varied widely in other tutin-containing honeys. The glycosidation of tutin may represent detoxification by one or both of the insects involved in the food chain from plant to honey.

From naturally-occurring neurotoxic agents to CNS shuttles for drug delivery.

Central nervous system (CNS) diseases are hard to diagnose and therapeutically target due to the blood brain barrier (BBB), which prevents most drugs from reaching their sites of action within the CNS. Brain drug delivery systems were conceived to bypass the BBB and were derived from anatomical and functional analysis of the BBB; this analysis led researchers to take advantage of brain endothelial membrane physiology to allow drug access across the BBB. Both receptors and carriers can be used to transport endogenous and exogenous substances into the CNS. Combining a drug with substances that take advantage of these internalization mechanisms is a widely exploited strategy for drug delivery because it is an indirect method that overcomes the BBB in a non-invasive way and is therefore less dangerous and costly than invasive methods. Neurotoxins, among other naturally-occurring substances, may be used as drug carriers to specifically target the CNS. This review covers the current delivery systems that take advantage of the non-toxic components of neurotoxins to overcome the BBB and reach the CNS. We hope to give insights to researchers toward developing new delivery systems that exploit the positive features of substances usually regarded as natural hazards.