Thiol-yne click post-synthesis of phenylboronic acid-functionalized magnetic cyclodextrin microporous organic network for selective and efficient extraction of antiepileptic drugs.

Owing to their incomplete digestion in the human body and inadequate removal by sewage treatment plants, antiepileptic drugs (AEDs) accumulate in water bodies, potentially affecting the exposed humans and aquatic organisms. Therefore, sensitive and reliable detection methods must be urgently developed for monitoring trace AEDs in environmental water samples. Herein, a novel phenylboronic acid-functionalized magnetic cyclodextrin microporous organic network (Fe3O4@CD-MON-PBA) was designed and synthesized via the thiol-yne click post-modification strategy for selective and efficient magnetic solid-phase extraction (MSPE) of trace AEDs from complex sample matrices through the specific B-N coordination, π-π, hydrogen bonding, electrostatic, and host-guest interactions. Fe3O4@CD-MON-PBA exhibited a large surface area (118.5 m2 g-1), rapid magnetic responsiveness (38.6 emu g-1, 15 s), good stability and reusability (at least 8 times), and abundant binding sites for AEDs. Under optimal extraction conditions, the proposed Fe3O4@CD-MON-PBA-MSPE-HPLC-UV method exhibited a wide linear range (0.5-1000 µg L-1), low limits of detection (0.1-0.5 µg L-1) and quantitation (0.3-2 µg L-1), good anti-interference ability, and large enrichment factors (92.2-104.3 to 92.3-98.0) for four typical AEDs. This work confirmed the feasibility of the thiol-yne click post-synthesis strategy for constructing novel and efficient multifunctional magnetic CD-MONs for sample pretreatment and elucidated the significance of B-N coordination between PBA and N-containing AEDs.

Preparation of a ß-cyclodextrin grafted magnetic biochar for efficient extraction of four antiepileptic drugs in plasma samples.

Simultaneous monitoring of plasma concentration levels of multiple antiepileptic drugs (AEDs) is essential for dose adjustment in comprehensive epilepsy treatment, necessitating a sensitive technique for accurate extraction and determination of AEDs. Herein, a magnetic solid-phase extraction (MSPE) technique on the basis of modified biochar (BC) is investigated to extract four AEDs from plasma, in conjunction with high performance liquid chromatography. BC derived from Zizyphus jujuba seed shells was activated by phosphoric acid (PBC) and magnetized via coprecipitation to produce MPBC. The MPBCCD obtained after modification with ß-cyclodextrin (CD) was characterized and evaluated for adsorption. It exhibited fast adsorption kinetics based on second-order kinetics and satisfactory adsorption capacity for AEDs. Then it was employed as the MSPE adsorbent and the influencing parameters were optimized. The enrichment factor was 18.75. The validation analysis revealed a favorable linearity that ranged from 0.04 to 20 µg·mL-1 along with a low limit of detection of 6.85 to 10.19 ng·mL-1. The recovery of the AEDs ranged from 78.7 to 109.2 %, with relative standard deviations below 6.7 %. Using quantum chemistry theory calculations and experimental results analysis, the adsorption mechanism was investigated. It disclosed that the suggested strategy built upon MPBCCD was appropriate for the assessment of AEDs in plasma and expanded the usage of BC as the environmentally favorable matrix for the analysis of biological samples.

Chemical profiling, anticonvulsant and anxiolytic effects of the smoke constituents isolated from Leonotis leonurus (L.) R.Br.

ETHNOPHARMACOLOGICAL RELEVANCE: The use of medicinal plants for central nervous system (CNS)-related ailments, such as epilepsy and anxiety, is prevalent in South Africa. Plants from the Lamiaceae family are commonly used for their therapeutic benefits. Leonotis leonurus (L.) R.Br. has been reported in ethnobotanical literature to have anticonvulsant and anxiolytic effects through the inhalation of pyrolysis products obtained by combustion of the aerial parts. AIM AND OBJECTIVES: To explore the chemical profiles and CNS activity of the smoke extract and isolated constituents of L. leonurus in zebrafish larvae, through anticonvulsive and anxiolytic activity assays. MATERIALS AND METHODS: The smoke extract of L. leonurus was obtained through the combustion of the aerial parts of the plant using a custom-built smoke recovery apparatus. The chemical profile of the smoke constituents was determined using Ultra-Performance Liquid Chromatography coupled with Mass Spectrometry (UPLC-MS). Targeted compounds were subjected to preparative High-Performance Liquid Chromatography for separation before structure elucidation using Nuclear Magnetic Resonance (NMR). The maximum tolerated concentrations, as well as the anxiolytic activity of the smoke extract were determined in five days post fertilisation zebrafish larvae. Reverse-thigmotaxis and locomotor activity of larvae in the light/dark transition assay were used to determine anxiolytic activity. Zebrafish larvae at six days post fertilisation (dpf) were subjected to several concentrations of the smoke constituents of L. leonurus. The baseline locomotor activity of the larvae was tracked for 30 min, prior to addition of pentylenetetrazole (PTZ) to induce seizure-like behaviour in the larvae, after which the locomotor activity of the larvae was once again tracked for an additional 30 min. RESULTS: The UPLC-MS profiles of the smoke extract revealed the presence of two main compounds, leoleorin A and leoleorin B, which were targeted and isolated. Upon subjection to NMR spectroscopy for structure elucidation, the compounds were confirmed to be labdane diterpenoids. Both leoleorin A and leoleorin B, and the smoke extract displayed suppression of the PTZ induced seizure-like behaviour in zebrafish larvae. Under light and dark conditions, the smoke extract and compounds displayed potential anxiolytic activity at different concentrations. CONCLUSION: Our results suggest that the smoke constituents of L. leonurus may exert anticonvulsant and anxiolytic effects which align with the traditional indications and the mode of administration.

ICH Q14-inspired novel approach to establish an SFC-based purity method for carbamazepine.

The proposed ICH Q14 guideline "Analytical procedure development" describes science and risk-based approaches for development and maintenance of analytical procedures suitable for the assessment of the quality of drug substances and drug products. As a case study, the systematic development and validation of a supercritical fluid chromatography (SFC)-based purity method for carbamazepine is presented. Systematic analytical quality by design (AQbD) principles were applied using the software package Fusion QbD to the method development approach. The relationship between chromatographic parameters and the responses of interest were examined to improve the reliability of the method by understanding, reducing, and controlling sources of variability. Method performance qualification in terms of method robustness was finally carried out with the parameters that were classified as critical after method development and a validation study met previously set acceptance criteria. The developed SFC purity method for carbamazepine demonstrated readiness as a viable alternative to the official HPLC method published in the Ph.Eur. with improved peak resolution, improved peak symmetry, and faster analysis times (3 min vs. 80 min for the official method). Its inherent reliability illustrates the superiority of AQbD in method development and application for drug quality assurance.

Anticonvulsant effects of Cymbopogon giganteus extracts with possible effects on fully kindled seizures and anxiety in experimental rodent model of mesio-temporal epilepsy induced by pilocarpine.

ETHNOPHARMACOLOGICAL RELEVANCE: Epilepsy is a neurological disorder of the brain characterized by periodic and unpredictable occurrence of a transient behavior alteration due to the rhythmic, synchronous and disordered firing of brain neuron. Worldwide, approximately 50 million people currently live with epilepsy and close to 80% of people with epilepsy live in poor countries. However, it was noticed in many countries worldwide that people with epilepsy and their families suffer from stigma and discrimination and that situation exposes them to high psychological conditions such as depression and anxiety as well as more physical problems including bruising and fractures from injuries related to seizures. However, several plants-based products used for epilepsy and anxiety treatments in different system of folk medicine have exhibited a significant anti-epileptic and antianxiety activities using animal models with fewer side effects. AIM OF THE STUDY: The study aimed at evaluating the antiepileptic, status post-epilepticus and anxiolytic effects of Cymbopogon giganteus decoction in rat model induced by pilocarpine. MATERIALS AND METHODS: A total of 90 rats were partitioned into 7 groups and treated as follow: animals of groups I (normal control) and II (considered the negative control) received distilled water (10 mL/kg); while groups III, IV, V, and VI were treated with the C. giganteus extract at 34, 85, 170 and 340 mg/kg p.o, respectively; and the group VII (considered positive control) received sodium valproate at 300 mg/kg, i.p. After 40 min post-treatment, a single dose of n-methyl-scopolamine (1 mg/kg, i.p) was administered to animals of groups (II, III, IV, V, VI, VII) followed by pilocarpine (360 mg/kg, i.p). Animal of group I (normal group) received distilled water. Rats were further observed for 6 h to evaluate the severity and the duration of the acute seizures of epilepsy according to Racine scale. Anxious behavior status post-epilepticus was also assessed in the same rats used above in the Elevated Plus Maze and number of entries into the open or closed arms and the time spent on either open or closed arms of the platform were recorded. Animals were also evaluated on Open Field Test and the number of rearing, crossing, grooming, defecation and center time were registered. RESULTS: C. giganteus decoction significantly (P < 0.05) reduced the animal mortality, the number and duration of convulsions and effectively increased the latency of convulsions. The plant extract significantly (P < 0.05) improved GSH level and SOD activity, reduced MDA and CAT activity, increased GABA level and decreased GABA-t activity in hippocampus. The anxiety induced by pilocarpine was also significantly (P < 0.05) inhibited by the extract of the plant. CONCLUSIONS: Thus, C. giganteus has demonstrated its antiepileptic and anxiolytic activities in rat model and may be used as preventive measure for patients suffering from epilepsy seizures and anxiety.

An aqueous extract of Canarium schweinfurthii attenuates seizures and potentiates sleep in mice: Evidence for involvement of GABA Pathway.

About 30% of epileptic patients continue to have seizures. The present study investigates the anticonvulsant and sedative effects of an aqueous extract of C. schweinfurthii in mice. Anticonvulsant effects of C. schweinfurthii aqueous extract (0.01-300 mg/kg, p.o.) were tested against 4-aminopyridine (4-AP, 15 mg/kg, i.p.) -, pilocarpine (PILO, 380 mg/kg, i.p.) - and pentylenetetrazole (PTZ, 75 mg/kg, i.p.) -induced seizures, while sedative effects were tested on diazepam (35 mg/kg, i.p.)-induced sleep. Afterward, the most effective dose of the extract (11.9 mg/kg) was antagonized with N-methyl-ß-carboline-3-carboxamide or flumazenil. In another set of experiments, mice were sacrificed for the estimation of GABA content and GABA-T activity in the cerebral cortex. The dose of the extract that protected 50% of mice (ED50) against 4-AP, PILO, and PTZ was respectively 4.43 mg/kg (versus 12.01 for phenobarbital), 9.59 mg/kg (vs 8.67 for diazepam), and 2.12 mg/kg (vs 0.20 for clonazepam). Further, the ED50 of the extract that increased the duration of sleep was 0.24 mg/kg (vs 0.84 for phenobarbital). N-methyl-ß-carboline-3-carboxamide or flumazenil antagonized (p < 0.001) the anticonvulsant effect of C. schweinfurthii in PTZ-induced seizures and diazepam-induced sleep when compared to the negative control group. The extract at all doses increased (p < 0.001) the GABA content and decreased (p < 0.001) GABA-T activity. These findings suggest that C. schweinfurthii possesses anticonvulsant and sedative effects. These effects seem to be mediated via the modulation of the GABA neurotransmission. These data explain the use of this plant to treat epilepsy in Cameroon traditional medicine.

The Anticonvulsant Effect of Hydroethanolic Leaf Extract of Calotropis procera (Ait) R. Br. (Apocynaceae).

A number of currently used drugs have been obtained from medicinal plants which are a major source of drugs. These drugs are either used in their pure form or modified to a semisynthetic drug. Drug discovery through natural product research has been fruitful over the years. Traditionally, Calotropis procera is used extensively in the management of epilepsy. This study is conducted to explore the anticonvulsant effect of a hydroethanolic leaf extract of Calotropis procera (CPE) in murine models. This effect was evaluated using picrotoxin-induced convulsions, strychnine-induced convulsions, and isoniazid- and pilocarpine-induced status epilepticus in mice of both sexes. The results showed that CPE (100-300 mg/kg) exhibited an anticonvulsant effect against strychnine-induced clonic seizures by significantly reducing the duration (p = 0.0068) and frequency (p = 0.0016) of convulsions. The extract (100-300 mg/kg) caused a profound dose-dependent delay in the onset of clonic convulsions induced by picrotoxin (p < 0.0001) and tonic convulsions (p < 0.0001) in mice. The duration of convulsions was reduced significantly also for both clonic and tonic (p < 0.0001) seizures as well. CPE (100-300 mg/kg), showed a profound anticonvulsant effect and reduced mortality in the pilocarpine-induced convulsions. ED50 (~0.1007) determined demonstrated that the extract was less potent than diazepam in reducing the duration and onset of convulsions but had comparable efficacies. Flumazenil-a GABAA receptor antagonist-did not reverse the onset or duration of convulsions produced by the extract in the picrotoxin-induced seizure model. In isoniazid-induced seizure, CPE (300 mg kg1, p.o.) significantly (p < 0.001) delayed the onset of seizure in mice and prolonged latency to death in animals. Overall, the hydroethanolic leaf extract of Calotropis procera possesses anticonvulsant properties.

Anti-seizure activity of African medicinal plants: The identification of bioactive alkaloids from the stem bark of Rauvolfia caffra using an in vivo zebrafish model.

ETHNOPHARMACOLOGICAL RELEVANCE: Epilepsy is one of the major chronic diseases that does not have a cure to date. Adverse drug reactions have been reported from the use of available anti-epileptic drugs (AEDs) which are also effective in only two-thirds of the patients. Accordingly, the identification of scaffolds with promising anti-seizure activity remains an important first step towards the development of new anti-epileptic therapies, with improved efficacy and reduced adverse effects. Herbal medicines are widely used in developing countries, including in the treatment of epilepsy but with little scientific evidence to validate this use. In the search for new epilepsy treatment options, the zebrafish has emerged as a chemoconvulsant-based model for epilepsy, mainly because of the many advantages that zebrafish larvae offer making them highly suitable for high-throughput drug screening. AIM OF THE STUDY: In this study, 20 medicinal plants traditionally used in South Africa to treat epilepsy were screened for anti-epileptic activity using a zebrafish larvae model. MATERIALS AND METHODS: Toxicity triaging was conducted on 120 crude extracts, 44 fractions and three isolated compounds to determine the maximum tolerated concentration (MTC) of each extract, fraction or compound. MTC values were used to guide the concentration range selection in bioactivity studies. The effectiveness of crude extracts, fractions and isolated compounds from Rauvolfia caffra Sond. in suppression of pentylenetetrazole (PTZ) induced seizure-like behaviour in a 6-dpf zebrafish larvae model was measured using the PTZ assay. RESULTS: Following a preliminary toxicity triage and bioactivity screen of crude extracts from 20 African plants used traditionally for the treatment and management of epilepsy, the methanolic extract of Rauvolfia caffra Sond. was identified as the most promising at suppressing PTZ induced seizure-like behaviour in a zebrafish larvae model. Subsequent bioactivity-guided fractionation and spectroscopic structural elucidation resulted in the isolation and identification of two tryptoline derivatives; a previously unreported alkaloid to which we assigned the trivial name rauverine H (1) and the known alkaloid pleiocarpamine (2). Pleiocarpamine was found to reduce PTZ-induced seizures in a dose-dependent manner. CONCLUSIONS: Accordingly, pleiocarpamine represents a promising scaffold for the development of new anti-seizure therapeutic compounds. Furthermore, the results of this study provide preliminary evidence to support the traditional use of Rauvolfia caffra Sond. in the treatment and management of epilepsy. These findings warrant further studies on the anti-epileptic potential of Rauvolfia caffra Sond. using other models.

Pergularia daemia hydro-ethanolic extract protects against pentylenetetrazole kindling-induced seizures, oxidative stress, and neuroinflammation in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Current antiepileptic drugs fail to control approximately 30% of epilepsies. Therefore, there is a need to develop more effective antiepileptic drugs, and medicinal plants provide an attractive source for new compounds. Pergularia daemia (Asclepiadaceae) is used in Cameroon traditional medicine to treat stroke, anemia, inflammation, and epilepsy. Recently, traditional healers claim that an hydro-ethanolic extract of the roots of P. daemia is more effective than an aqueous extract on refractory seizures. AIM OF THE STUDY: The antiepileptic effect of P. daemia hydro-ethanolic extract was investigated on the pentylenetetrazole kindling model of temporal lobe epilepsy in mice and possible mechanisms of action. MATERIALS AND METHODS: Mice were divided into 8 groups treated as follows: normal group received distilled water (10 ml/kg, p.o.), control group received distilled water (10 ml/kg, p.o.), ethanol group received ethanol (5%, p.o.), positive control received sodium valproate (300 mg/kg, p.o.), and test groups received P. daemia hydro-ethanolic (HE) extract (1.6, 4, 8 and 16 mg/kg, p.o.). All groups were kindled by 11 injections of pentylenetetrazole (PTZ) (35 mg/kg, i.p.), once every alternate day (48 ± 2 h), until the development of kindling, i.e., the occurrence of stage 5 seizures for two consecutive trials. One week later, i.e., 29th day, mice were challenged with a single and lower dose of PTZ (25 mg/kg, i.p.) that does not induce seizures in normal mice but causes seizures in mice prone to seizures and behavioral alterations. After completion of the kindling procedure, Morris water maze, passive avoidance, and open field tests were performed. Afterward, animals were euthanized, and hippocampi were removed for the estimation of the levels of GABA-transaminase (GABA-T), L-glutamate decarboxylase (L-GAD), and γ-aminobutyric acid (GABA). Oxidative stress and neuroinflammation markers also were quantified. Finally, histological analysis of the hippocampus was carried out. RESULTS: PTZ-kindling induced myoclonic jerks and generalized tonic-clonic seizures in control mice. However, the HE extract of P. daemia (4-16 mg/kg), compared to sodium valproate, significantly protected mice against myoclonic jerks and generalized tonic-clonic seizures. Also, the HE extract (1.6-16 mg/kg) significantly increased the seizure score. Furthermore, the HE extract of P. daemia significantly reduced seizure-induced cognitive impairments. PTZ-kindling induced significant alterations in GABA, GABA-T, and L-GAD contents as well as oxidative stress, and neuroinflammation, and the HE extract significantly reversed these effects, suggesting possible mechanisms. All these activities of the HE extract were confirmed by its protective effect against neuronal loss in the hippocampus. CONCLUSIONS: The HE extract of P. daemia protected mice against kindled seizures and cognitive impairments, and these effects were greater than those of sodium valproate, a widely used antiepileptic drug. These effects may be mediated by neuromodulatory, anti-oxidant, and anti-inflammatory activities, thus suggesting a neuroprotective effect. These findings help to explain the beneficial use of these HE extracts of P.daemia in traditional medicine to treat epilepsy in Cameroon.

Anticonvulsant activity of Valeriana edulis roots and valepotriates on the pentylenetetrazole-induced seizures in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: For many centuries, Mexican Valerian (Valeriana edulis ssp. procera) has been an important plant in folk medicine. It has been considered useful to control epilepsy; however, electroencephalographic evidence of its anticonvulsant activity is missing in literature. AIM OF THE STUDY: In the present study, in situ electroencephalographic (EEG) analysis was performed along with administration of a crude ethanol extract of V. edulis and its valepotriate fraction on the pentylenetetrazole (PTZ)-induced convulsive behavior in rats. MATERIALS AND METHODS: Experiments were performed using male Wistar rats with nail-shaped electrodes implanted in the frontal and parietal cortices for EEG recording. All animals received a single dose of PTZ (35 mg/kg, i.p.) to test the anticonvulsant activity of V. edulis crude extract and valepotriate fraction (100 mg/kg, i.p.) 15 and/or 30 min after administration. EEG recordings were obtained from the cortices and were evaluated to assess ictal behavior over 60-75 min. Chromatographic analysis of the valepotriate fraction and in silico predictions of pharmacodynamic properties were also explored. The latency, frequency and duration of seizures evaluated using EEG recordings from the frontal and parietal cortices of rats showed significant changes demonstrating the inhibition of paroxystic activity. RESULTS: The spectral analysis confirmed the reduction of excitatory activity induced by V. edulis extract, which was improved in the presence of the valepotriate fraction as compared to that induced by ethosuximide (a reference anticonvulsant drug). The presence of valepotriates such as: isodihydrovaltrate (18.99%), homovaltrate (13.51%), 10-acetoxy-valtrathydrin (4%) and valtrate (1.34%) was identified by chromatographic analysis. Whereas, not only GABAA receptor participation but also the cannabinoid CB2 receptor was found to be likely involved in the anticonvulsant mechanism of action after in silico prediction. CONCLUSIONS: Our data support the anticonvulsant properties attributed to this plant in folk medicine, due to the presence of valepotriates.

Dose dependent anticonvulsant activity of Morus nigra in strychnine induced seizures model.

Herbal medicines have gained tremendous surge of interest in recent years. M. nigra leaves are a rich source of phenolics which are well-known for their antioxidant property. Morus nigra popularly known as black mulberry is considered to be the most significant species of genus Morus. This study was designed to evaluate its activity on seizure model in different doses. Five groups were made comprising of n=10 animals in each group respectively. Group I was on distilled water, Group II was administered with reference drug diazepam and Group III, IV and V were on 125mg/kg, 250mg/kg and 500mg/kg dose of Morus nigra for 15 days prior to experiment. On day 16th all animals were administered with strychnine after 30 minutes of respective treatments and three parameters were recorded i.e. duration, frequency and onset of seizures. M. nigra treatment showed significant seizure protection as noted by delayed latency of seizures (P<0.05), decrease in frequency and jerk's duration (P<0.05) in comparison to control and reference standard. Most significant (P<0.05) anticonvulsant effects were observed with 500mg/kg dose. Anticonvulsant activity of M. nigra could be due to potentiation of both Gabaergic and glycinergic activities. Antiepileptic potential of extract could also be amplified due to its antioxidant activity. This could serve as a non-pharmacological treatment for seizure management.

Potential anti-epileptic phytoconstituents: An updated review.

ETHNOPHARMACOLOGICAL RELEVANCE: Epilepsy is one of the most commonly occurring non-communicable neurological disorder that affects people of all age groups. Around 50 million people globally are epileptic, with 80% cases in developing countries due to lack of access to treatments determined by high cost and poor availability or it can be defined by the fraction of active epileptic patients who are not appropriately being treated. The availability of antiepileptic drugs and their adjuvant therapy in such countries is less than 50% and these are highly susceptible to drug interactions and severe adverse effects. As a result, the use of herbal medicine is increasingly becoming popular. AIM OF THE STUDY: To provide pharmacological information on the active constituents evaluated in the preclinical study to treat epilepsy with potential to be used as an alternative therapeutic option in future. It also provides affirmation for the development of novel antiepileptic drugs derived from medicinal plants. MATERIALS AND METHODS: Relevant information on the antiepileptic potential of phytoconstituents in the preclinical study (in-vitro, in-vivo) is provided based on their effect on screening parameters. Besides, relevant information on pharmacology of phytoconstituents, the traditional use of their medicinal plants related to epilepsy and status of phytoconstituents in the clinical study were derived from online databases, including PubMed, Clinicaltrial. gov, The Plant List (TPL, www.theplantlist.org), Science Direct. Articles identified using preset searching syntax and inclusion criteria are presented. RESULTS: More than 70% of the phytoconstituents reviewed in this paper justified the traditional use of their medicinal plant related to epilepsy by primarily acting on the GABAergic system. Amongst the phytoconstituents, only cannabidiol and tetrahydrocannabinol have been explored for clinical application in epilepsy. CONCLUSION: The preclinical and clinical data of the phytoconstituents to treat epilepsy and its associated comorbidities provides evidence for the discovery and development of novel antiepileptic drugs from medicinal plants. In terms of efficacy and safety, further randomized and controlled clinical studies are required to understand the complete pharmacodynamic and pharmacokinetic picture of phytoconstituents. Also, specific botanical source evaluation is needed.

Cognition and memory impairment attenuation via reduction of oxidative stress in acute and chronic mice models of epilepsy using antiepileptogenic Nux vomica.

Ethnopharmacological relevance Processed Nux vomica seed extracts and homeopathic medicinal preparations (HMPs) are widely used in traditional Indian and Chinese medicine for respiratory, digestive, neurological and behavioral disorders. Antioxidant property of Nux vomica is well known and recent investigation has highlighted the anticonvulsant potential of its homeopathic formulation. AIM OF THE STUDY: To explore the anticonvulsant and antiepileptogenic potential of Nux vomica HMPs (6CH, 12CH and 30CH potency) in pentylenetetrazole (PTZ) induced acute and chronic experimental seizure models in mice and investigate their effects on cognition, memory, motor activity and oxidative stress markers in kindled animals. MATERIALS AND METHODS: Acute seizures were induced in the animals through 70 mg/kg (i.p.) administration of PTZ followed by the evaluation of latency and duration of Generalized tonic-clonic seizures (GTCS). Subconvulsive PTZ doses (35 mg/kg, i.p.) induced kindling in 29 days, which was followed by assessment of cognition, memory and motor impairment through validated behavioral techniques. The status of oxidative stress was estimated through measurement of MDA, GSH and SOD. RESULTS: HMPs delayed the latency and reduced the duration of GTCS in acute model signifying possible regulation of GABAergic neurotransmission. Kindling was significantly hindered by the HMPs that justified the ameliorated cognition, memory and motor activity impairment. The HMPs attenuated lipid peroxidation by reducing MDA level and strengthened the antioxidant mechanism by enhancing the GSH and SOD levels in the kindled animals. CONCLUSIONS: Nux vomica HMPs showed anticonvulsant and antiepileptogenic potency in acute and chronic models of epilepsy. The test drugs attenuated behavioral impairment and reduced the oxidative stress against PTZ induced kindling owing to which they can be further explored for their cellular and molecular mechanism(s).

Supramolecular Solvent-Based Microextraction of Selected Anticonvulsant and Nonsteroidal Anti-Inflammatory Drugs from Sediment Samples.

The increase in the production and consumption of pharmaceuticals increases their presence in the global environment, which may result in direct threats to living organisms. For this reason, there is a need for new methods to analyze drugs in environmental samples. Here, a new procedure for separating and determining selected drugs (diclofenac, ibuprofen, and carbamazepine) from bottom sediment and water samples was developed. Drugs were determined by ultra-high performance liquid chromatography coupled with an ultraviolet detector (UHPLC-UV). In this work, a universal and single-step sample treatment, based on supramolecular solvents (SUPRAS), was proposed to isolate selected anticonvulsants and nonsteroidal anti-inflammatory drugs (NSAIDs) from sediment samples. The following parameters were experimentally selected: composition of the supramolecular solvent (composition THF:H2O (v/v), amount of decanoic acid), volume of extractant, sample mass, extraction time, centrifugation time, and centrifugation speed. Finally, the developed procedure was validated. A Speedisk procedure was also developed to extract selected drugs from water samples. The recovery of analytes using the SUPRAS procedure was in the range of 88.8-115%, while the recoveries of the Speedisk solid-phase extraction procedure ranged from 81.0-106%. The effectiveness of the sorption of the tested drugs by sediment was also examined.

Targeted Strategy to Analyze Antiepileptic Drugs in Human Serum by LC-MS/MS and LC-Ion Mobility-MS.

Routine small-molecule analysis is challenging owing to the need for high selectivity and/or low limits of quantification. This work reports a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify 14 antiepileptic drugs (AEDs) in human serum. For the optimized LC-MS/MS method described herein, we applied the guidelines outlined in the Clinical and Laboratory Standards Institute (CLSI) LC-MS C62-A document and the U.S. Food and Drug Administration (FDA) Bioanalytical Method Validation Guidance for Industry to evaluate the quality of the assay. In these studies, AED linearity, analyte recovery, matrix effects, precision, and accuracy were assessed. Using liquid chromatography-drift tube ion mobility-mass spectrometry (LC-DTIM-MS), a qualitative method was also used to increase confidence in AED identification using accurate mass and collision cross section (CCS) measurements. The LC-DTIM-MS method was also used to assess the ability of drift tube CCS measurements to aid in the separation and identification of AED structural isomers and other AEDs. These data show that another dimension of information, namely CCS measurements, provides an orthogonal dimension of structural information needed for AED analysis. Multiplexed AED measurements using LC-MS/MS and LC-DTIM-MS have the potential to enable better optimization of dosing owing to the high precision capabilities available in these types of analytical studies. Taken together, these data also show the ability to increase confidence in small-molecule identification and quantification using these analytical technologies.

Acteoside Isolated from Colebrookea oppositifolia Smith Attenuates Epilepsy in Mice Via Modulation of Gamma-Aminobutyric Acid Pathways.

The present study was aimed to evaluate the anticonvulsant activity of acteoside and explore its mechanism of action. Initially, the acteoside was evaluated in maximal electroshock (MES) and pentylenetetrazole (PTZ)-induced convulsions, and later it was evaluated against N-methyl-D-aspartic acid (NMDA)-induced mortality in Swiss albino mice. Based on the response in these models, further evaluations were performed to explore the mechanism of action. In the results, the acteoside (10, 25, and 50 mg/kg) has shown significant anticonvulsant activity in the PTZ model (p < 0.01 for all doses); however, there was no protection observed in MES and NMDA models. Therefore, further mechanism-based studies were performed on the PTZ model, and the outcomes have revealed that there was a significant reduction in GABA (p < 0.01 for both regions) and elevation of glutamate (p < 0.01 for both regions) in the cortex and hippocampus regions of PTZ-treated animals. Further, the antioxidant levels (SOD, catalase, GPx, GR, GSH, LPO) were altered significantly (p < 0.01 for all parameters), with reduced GABAA mRNA levels (p < 0.01) in the PTZ control compared with the normal control. Interestingly, co-administration of acteoside (25 mg/kg) (p < 0.01 for all parameters) has restored all the PTZ-induced alterations compared to PTZ-control. Moreover, the anti-PTZ action of acteoside was completely blocked in the presence of flumazenil, and thus confirmed the GABAergic mechanism behind the anticonvulsant activity of acteoside. Besides, actophotometer and rotarod tests have confirmed that the acteoside is free from central side effects like motor incoordination and locomotor deficits.

Understanding the basics of cannabidiol from cannabis to apply to therapeutics in epilepsy.

The compounds present in cannabis have been in use for both recreational and medicinal purposes for many centuries. Changes in the legislation in South Africa have led to an increase in the number of people interested in using these compounds for self-medication. Many of them may approach their general practitioner as the first source of information about possible therapeutic effects. It is important that medical professionals are able to give patients the correct information. Cannabidiol (CBD) is one of the main compounds in cannabis plants, and there is evidence that it can successfully treat certain patients with epilepsy. This review looks at the most recent evidence on the use of CBD in the treatment of epilepsy and explores the mechanisms behind these beneficial effects.

Anti-convulsant effects of cultures bear bile powder in febrile seizure via regulation of neurotransmission and inhibition of neuroinflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: Natural bear bile powder (NBBP) has been used to treat seizures for thousands of years, but its application is greatly restricted due to ethical reasons. Cultured bear bile powder (CBBP), which is produced by biotransformation, may be an appropriate substitute for NBBP. However, the anti-convulsant effects of CBBP and its mechanisms remain unclear. AIM OF THE STUDY: This study aimed to investigate the anti-convulsant effects and possible mechanisms of CBBP in a febrile seizure (FS) rat model. MATERIALS AND METHODS: FS was induced by placing the rats in a warm water bath (45.5 °C). The incidence rate and latency of FS, and hematoxylin-eosin staining (HE) were conducted for neurological damage. The levels of 4 bile acids and 8 main neurotransmitters in vivo were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The expression of bile acid related transports, neurotransmitter receptors, inflammatory factors, neurotrophic factors and glial fibrillary acidic protein (GFAP) in hippocampal tissues were detected by real-time PCR, western blotting, and immunohistochemistry. RESULTS: Pre-treatments with CBBP and similarly, NBBP, significantly reduced the incidence rate and prolonged the latency of FS. Additionally, CBBP alleviated the histological injury induced by FS in the rat hippocampus tissue. LC-MS/MS analyses revealed that CBBP markedly increased the levels of tauroursodeoxycholic acid (TUDCA), taurochenodeoxycholic acid (TCDCA), ursodeoxycholic acid (UDCA), and chenodeoxycholic acid (CDCA) in FS rats. Furthermore, the content of gamma-aminobutyric acid (GABA) was up-regulated in rats pre-treated with CBBP whereas GFAP was down-regulated. CBBP also significantly suppressed the expression of interleukin -1ß (IL-1ß), tumor necrosis factor α (TNF-α), nuclear factor kappa B (NF-κB), and brain-derived neurotrophic factor (BDNF) and its TrkB receptors, and improved the expression of GABA type A receptors (GABAAR) and farnesoid X receptors (FXR). CONCLUSIONS: The present study demonstrated that CBBP had anti-convulsant effects in a FS rat model. CBBP may protect rats against FS, probably by up-regulating FXR, which was activated by increasing brain bile acids, up-regulating GABAergic transmission by inhibiting BDNF-TrkB signaling, and suppressing neuroinflammation by inhibiting the NF-κB pathway.

Glaucacetalin E and galphimidin B from Galphimia glauca and their anxiolytic activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Galphimia glauca is a Mexican medicinal plant used to treat anxiety, fear, phobia and stress as it possesses sedative properties which produce a calming effect. Although some chemical and pharmacological studies have already been carried out on G. glauca, there are still new chemical entities from this plant whose anxiolytic activity should be established. AIM OF THE STUDY: To validate the use of G. glauca growing in Cuernavaca, Morelos, as an anti-stress agent, through the purification and structural identification of its extracts' chemical constituents; the analysis of the biogenetic relationship of its chemical compounds, and its biological evaluation to demonstrate its traditional use as anxiolytic agents. MATERIALS AND METHODS: The structures of all isolated compounds were established based on their spectroscopic and spectrometric data. The structure of compound 2 was corroborated through X-Ray. The anxiolytic and sedative-like activities were assessed by the open-field, hole-board and exploration cylinder test. RESULTS: The nor-triterpenes glaucacetalin E (1) and galphimidin B (2) were isolated for the first time along with seven other known compounds, one of them galphimidin (3), from the CHCl3 fraction of the aerial parts of Galphimia glauca. The biogenesis of the natural nor-triterpenes isolated from Galphimia glauca is delineated for the first time starting from the taraxasteryl cation. Oral administration of CHCl3 fraction and 1-3 compounds produced significant attenuation in the anxiety-response in cylinder activity, decrease in the ambulatory activity and in head dipping when compared to the vehicle. However, only the extract enhanced the pentobarbital-induced hypnosis. Diazepam was used as a positive control. CONCLUSION: Our results suggest that G. glauca growing in Cuernavaca, Morelos, exerts anxiolytic-like activity due to the presence of the nor-triterpenes 1-3. These results reinforce the potential use of this species in the treatment of anxiety.

Antiseizure potential of the ancient Greek medicinal plant Helleborus odorus subsp. cyclophyllus and identification of its main active principles.

ETHNOPHARMACOLOGICAL RELEVANCE: Ethnopharmacological data and ancient texts support the use of black hellebore (Helleborus odorus subsp. cyclophyllus, Ranunculaceae) for the management and treatment of epilepsy in ancient Greece. AIM OF THE STUDY: A pharmacological investigation of the root methanolic extract (RME) was conducted using the zebrafish epilepsy model to isolate and identify the compounds responsible for a potential antiseizure activity and to provide evidence of its historical use. In addition, a comprehensive metabolite profiling of this studied species was proposed. MATERIALS AND METHODS: The roots were extracted by solvents of increasing polarity and root decoction (RDE) was also prepared. The extracts were evaluated for antiseizure activity using a larval zebrafish epilepsy model with pentylenetetrazole (PTZ)-induced seizures. The RME exhibited the highest antiseizure activity and was therefore selected for bioactivity-guided fractionation. Isolated compounds were fully characterized by NMR and high-resolution tandem mass spectrometry (HRMS/MS). The UHPLC-HRMS/MS analyses of the RME and RDE were used for dereplication and metabolite profiling. RESULTS: The RME showed 80% inhibition of PTZ-induced locomotor activity (300 µg/ml). This extract was fractionated and resulted in the isolation of a new glucopyranosyl-deoxyribonolactone (1) and a new furostanol saponin derivative (2), as well as of 20-hydroxyecdysone (3), hellebrin (4), a spirostanol glycoside derivative (5) and deglucohellebrin (6). The antiseizure activity of RME was found to be mainly due to the new furostanol saponin (2) and hellebrin (4), which reduced 45% and 60% of PTZ-induced seizures (135 µM, respectively). Besides, the aglycone of hellebrin, hellebrigenin (S34), was also active (45% at 7 µM). To further characterize the chemical composition of both RME and RDE, 30 compounds (A7-33, A35-37) were annotated based on UHPLC-HRMS/MS metabolite profiling. This revealed the presence of additional bufadienolides, furostanols, and evidenced alkaloids. CONCLUSIONS: This study is the first to identify the molecular basis of the ethnopharmacological use of black hellebore for the treatment of epilepsy. This was achieved using a microscale zebrafish epilepsy model to rapidly quantify in vivo antiseizure activity. The UHPLC-HRMS/MS profiling revealed the chemical diversity of the extracts and the presence of numerous bufadienolides, furostanols and ecdysteroids, also present in the decoction.