A Comparison between Bulgarian Tanacetum parthenium Essential Oil from Two Different Locations.

Tanacetum parthenium L. (Asteraceae) is a perennial herbaceous plant with a long-standing historical use in traditional medicine. Recently Tanacetum parthenium L. essential oil has been associated with a promising potential for future applications in the pharmaceutical industry, in the cosmetics industry, and in agriculture. Investigations on the essential oil (EO) have indicated antimicrobial, antioxidant, and repellent activity. The present study aimed to evaluate the chemical composition of Bulgarian T. parthenium essential oil from two different regions, to compare the results to those reported previously in the literature, and to point out some of its future applications. The essential oils of the air-dried flowering aerial parts were obtained by hydrodistillation using a Clevenger-type apparatus. The chemical composition was evaluated using gas chromatography with mass spectrometry (GC-MS). It was established that the oxygenated monoterpenes were the predominant terpene class, followed by the monoterpene hydrocarbons. Significant qualitative and quantitative differences between both samples were revealed. Camphor (50.90%), camphene (16.12%), and bornyl acetate (6.05%) were the major constituents in the feverfew EO from the western Rhodope Mountains, while in the EO from the central Balkan mountains camphor (45.54%), trans-chrysanthenyl acetate (13.87%), and camphene (13.03%) were the most abundant components.

A Study of the Chemical Composition, Antioxidant Potential, and Acute Toxicity of Bulgarian Tanacetum vulgare L. Essential Oil.

Common tansy (Tanacetum vulgare L.) is a plant with medicinal properties that has traditionally been used in folk medicine for its anthelmintic, antispasmodic, and choleretic effects, for the treatment of diarrhea and digestive problems, and externally, as an insecticide in veterinary practices. In the current study, we investigated, for the first time, the chemical profile and antioxidant activity of essential oil from a wild population of T. vulgare L. growing in Bulgaria. Common tansy essential oil (EO), which is rich in bicyclic monoterpenes, was obtained using hydrodistillation and characterized by using gas chromatography-mass spectrometry (GC-MS). Thirty-seven compounds were identified in Bulgarian tansy EO. Among the major constituents were oxygenated monoterpenes, including compounds such as camphor (25.24%), trans-chrysantenyl acetate (18.35%), cis-verbenol (10.58%), thujone (6.06%), eucaliptol (5.99%), and α-campholenal (5.98%). The analysis results identified the essential oil from T. vulgare L. grown in the western Rhodope Mountains of Bulgaria as the camphor chemotype. Furthermore, its antioxidant activity was analyzed using the oxygen radical absorbance capacity (ORAC) method and was found to be 605.4 ± 49.3 µmol TE/mL. The essential oil was also tested for single-dose acute toxicity on Wistar rats and was found to be non-toxic by oral administration. The mean lethal dose by intraperitoneal administration was LD50 i.p. = 14.9 g/kg body weight. The results of the conducted study can serve as a basis for the evaluation and subsequent exploration of other pharmacotherapeutic effects of the essential oil obtained from the inflorescences of the Bulgarian species T. vulgare L.

A Study of the Chemical Composition, Acute and Subacute Toxicity of Bulgarian Tanacetum parthenium Essential Oil.

BACKGROUND: Tanacetum parthenium (L.) Sch.Bip. (T. parthenium) is an aromatic perennial plant belonging to the Asteraceae family, also known as feverfew. It is widely distributed in various regions of Europe and other parts of the world. The plant has a rich background in the traditional medicine of many nations and has been used as a remedy for fever, pain, inflammation, asthma, rheumatism, menstrual disorders, etc. Methods: GC-MS analysis was conducted to determine the chemical composition of the isolated essential oil (EO). Using the method proposed by Litchfield and Wilcoxon, the average lethal dose (LD50) of the EO on Wistar rats was determined for two routes of administration: oral (p.o.) and intraperitoneal (i.p.). The subacute toxicity of the EO was also tested by oral administration of a daily dose of 1.0 g/kg body weight (BW) for 28 days. The toxicity of the EO was evaluated by observing and evaluating changes in behavior, body weight, basic hematological and serum biochemical parameters, and histopathological changes of the internal organs. RESULTS: Thirty-seven volatile organic compounds representing 94.58% of the total oil composition were tentatively detected in the obtained T. parthenium EO. The dominant compounds were camphor (45.47%), trans-chrisantenyl acetate (21.65%), camphene (9.48%), and cis-isogeraniol (5.42%). The results showed that the EO was not toxic when administered in acute oral doses. The acute mean lethal dose for intraperitoneal administration was LD50 i.p. = 2.13 g/kg BW. In the subacute study involving administration of an oral dose of EO for 28 days, there were a number of changes in the hematological and serum biochemical parameters of the blood compared with the control group of animals. However, no symptoms of toxicity, changes in the body weight of the rats, death, or pathological changes in the histological indicators of the examined organs-brain, heart, stomach, liver, spleen and kidney-were found. Extrapolating the results obtained from the rat experiments, we can state that the EO is safe for use in doses below 1 g/kgBW for a period not exceeding one month.

The anticonvulsant effect of chronic treatment with topiramate after pilocarpine-induced status epilepticus is accompanied by a suppression of comorbid behavioral impairments and robust neuroprotection in limbic regions in rats.

Epilepsy is a widespread neurological disorder frequently associated with a lot of comorbidities. The present study aimed to evaluate the effects of the antiseizure medication topiramate (TPM) on spontaneous motor seizures, the pathogenesis of comorbid mood and cognitive impairments, hippocampal neuronal loss, and oxidative stress and inflammation in a rat model of temporal lobe epilepsy (TLE). Vehicle/TPM treatment (80 mg/kg, p.o.) was administered 3 h after the pilocarpine (pilo)-induced status epilepticus (SE) and continued for up to 12 weeks in Wistar rats. The chronic TPM treatment caused side effects in naïve rats, including memory disturbance, anxiety, and depressive-like responses. However, the anticonvulsant effect of this drug, administered during epileptogenesis, was accompanied by beneficial activity against comorbid behavioral impairments. The drug treatment suppressed the SE-induced neuronal damage in limbic structures, including the dorsal (CA1 and CA2 subfield), the ventral (CA1, CA2 and CA3) hippocampus, the basolateral amygdala, and the piriform cortex, while was ineffective against the surge in the oxidative stress and inflammation. Our results suggest that neuroprotection is an essential mechanism of TPM against spontaneous generalized seizures and concomitant emotional and cognitive impairments.

An experimental study on phytochemical composition and memory enhancing effect of Ginkgo biloba seed extract.

INTRODUCTION: The Ginkgo biloba L. tree is considered as one of the oldest species on Earth. It is known as a "living fossil" dating back approximately 200 million years. Both the leaves and seeds of this tree have been used for millennia in traditional Chinese medicine. AIM: To study the phytochemical profile of Gingko biloba seed extract (GBSE) and its memory enhancing effects. MATERIALS AND METHODS: Liquid chromatography with mass detection (LC-MS) was performed for phytochemical analyses of the extracts. For the in vivo experiments, male Wistar rats were divided randomly into 5 groups (n=8): saline; piracetam;  GBSE 50; 100, and 200 mg/kg b.w. Y-maze, T-maze, step-down passive avoidance and novel object recognition test (NORT) were performed. The observed parameters were: percentage of spontaneous alternations (% SA), working memory index, latency of reaction and recognition index, respectively. Statistical analysis was done using SPSS 19. RESULTS: LC-MS analysis showed the presence of the flavonoids quercetin, kaempferol and isorhamnetin (as aglycones), the ginkgolides A, B, C, J, and bilobalide. In Y-maze task, the groups treated with 50 and 100 mg/kg of GBSE significantly increased the % SA during the memory test compared to saline (p<0.05). In T-maze test, the three experimental groups with GBSE significantly increased the working memory index in comparison with that of the control group (p<0.05). In step-down test, the animals receiving 100 mg/kg b.w. GBSE, notably increased the latency during both retention tests (p<0.05 and p<0.01, respectively). In NORT, only the animals with the middle dose of GBSE ameliorated the recognition index when compared to saline (p<0.05). CONCLUSIONS: GBSE enhances spatial working memory, recognition memory, and short- and long-term recall in naïve rats due to the synergic effects of detected flavonoids and terpene lactones on brain functions. The brain structures involved are probably the hippocampus and prefrontal cortex.

Effects of Lacosamide Treatment on Epileptogenesis, Neuronal Damage and Behavioral Comorbidities in a Rat Model of Temporal Lobe Epilepsy.

Clinically, temporal lobe epilepsy (TLE) is the most prevalent type of partial epilepsy and often accompanied by various comorbidities. The present study aimed to evaluate the effects of chronic treatment with the antiepileptic drug (AED) lacosamide (LCM) on spontaneous motor seizures (SMS), behavioral comorbidities, oxidative stress, neuroinflammation, and neuronal damage in a model of TLE. Vehicle/LCM treatment (30 mg/kg, p.o.) was administered 3 h after the pilocarpine-induced status epilepticus (SE) and continued for up to 12 weeks in Wistar rats. Our study showed that LCM attenuated the number of SMS and corrected comorbid to epilepsy impaired motor activity, anxiety, memory, and alleviated depressive-like responses measured in the elevated plus maze, object recognition test, radial arm maze test, and sucrose preference test, respectively. This AED suppressed oxidative stress through increased superoxide dismutase activity and glutathione levels, and alleviated catalase activity and lipid peroxidation in the hippocampus. Lacosamide treatment after SE mitigated the increased levels of IL-1ß and TNF-α in the hippocampus and exerted strong neuroprotection both in the dorsal and ventral hippocampus, basolateral amygdala, and partially in the piriform cortex. Our results suggest that the antioxidant, anti-inflammatory, and neuroprotective activity of LCM is an important prerequisite for its anticonvulsant and beneficial effects on SE-induced behavioral comorbidities.

Anticonvulsant Effects of Topiramate and Lacosamide on Pilocarpine-Induced Status Epilepticus in Rats: A Role of Reactive Oxygen Species and Inflammation.

BACKGROUND: Status epilepticus (SE) is a neurological disorder characterized by a prolonged epileptic activity followed by subsequent epileptogenic processes. The aim of the present study was to evaluate the early effects of topiramate (TPM) and lacosamide (LCM) treatment on oxidative stress and inflammatory damage in a model of pilocarpine-induced SE. METHODS: Male Wistar rats were randomly divided into six groups and the two antiepileptic drugs (AEDs), TPM (40 and 80 mg/kg, i.p.) and LCM (10 and 30 mg/kg, i.p.), were injected three times repeatedly after pilocarpine administration. Rats were sacrificed 24 h post-SE and several parameters of oxidative stress and inflammatory response have been explored in the hippocampus. RESULTS: The two drugs TPM and LCM, in both doses used, succeeded in attenuating the number of motor seizures compared to the SE-veh group 30 min after administration. Pilocarpine-induced SE decreased the superoxide dismutase (SOD) activity and reduced glutathione (GSH) levels while increasing the catalase (CAT) activity, malondialdehyde (MDA), and IL-1ß levels compared to the control group. Groups with SE did not affect the TNF-α levels. The treatment with a higher dose of 30 mg/kg LCM restored to control level the SOD activity in the SE group. The two AEDs, in both doses applied, also normalized the CAT activity and MDA levels to control values. In conclusion, we suggest that the antioxidant effect of TPM and LCM might contribute to their anticonvulsant effect against pilocarpine-induced SE, whereas their weak anti-inflammatory effect in the hippocampus is a consequence of reduced SE severity.

Immunomodulatory and Anti-Inflammatory Effects of Fucoidan: A Review.

Inflammation is the initial response of the immune system to potentially harmful stimuli (e.g., injury, stress, and infections). The process involves activation of macrophages and neutrophils, which produce mediators, such as nitric oxide (NO), prostaglandin E2 (PGE2), pro-inflammatory and anti-inflammatory cytokines. The pro-inflammatory cytokines interleukin-1ß (IL-1ß), interleukin 6 (IL-6), and tumor necrosis factor-α (TNF-α) are considered as biomarkers of inflammation. Even though it occurs as a physiological defense mechanism, its involvement in the pathogenesis of various diseases is reported. Rheumatoid arthritis, inflammatory bowel disease, Alzheimer's disease, and cardiovascular diseases are only a part of the diseases, in which pathogenesis the chronic inflammation is involved. Fucoidans are complex polysaccharides from brown seaweeds and some marine invertebrates, composed mainly of L-fucose and sulfate ester groups and minor amounts of neutral monosaccharides and uronic acids. Algae-derived fucoidans are studied intensively during the last years regarding their multiple biological activities and possible therapeutic potential. However, the source, species, molecular weight, composition, and structure of the polysaccharides, as well as the route of administration of fucoidans, could be crucial for their effects. Fucoidan is reported to act on different stages of the inflammatory process: (i) blocking of lymphocyte adhesion and invasion, (ii) inhibition of multiple enzymes, and (iii) induction of apoptosis. In this review, we focused on the immunemodulating and anti-inflammatory effects of fucoidans derived from macroalgae and the models used for their evaluation. Additional insights on the molecular structure of the compound are included.

Etifoxine does not impair muscle tone and motor function in rats as assessed by in vivo and in vitro methods.

The purpose of our study is to evaluate the effects of the translocator protein (TSPO) ligand etifoxine on muscle tone and locomotor activity. In addition, the mechanism of action of etifoxine on the presynaptic membrane and neuromuscular junction is investigated. These effects of etifoxine were examined employing the following methods: 1) in vivo experiments using bar holding test and activity cage test, and 2) comparative in vitro studies with nifedipine on indirectly-elicited twitches of striated abdominal muscle preparations. Etifoxine in doses 50 mg/kg and 100 mg/kg i.p. does not produce any significant changes in locomotor activity and muscle tone of intact rats. Nifedipine (10-5 М) induces a significant decrease in the muscle force of striated muscle preparations. Etifoxine (10-8-10-4 М) has no significant effect on indirectly-elicited twitch tension. Results show that the TSPO ligand etifoxine has no myorelaxant effect. The activation of TSPO is not associated with a reduction in muscle tone and motor impairment. Etifoxine does not affect the presynaptic membrane and its influence on L-type Ca2+-channels is insignificant. Etifoxine does not act as a competitive antagonist of acetylcholine and does not impair the impulse transmission in the neuromuscular junction.

The Effect of Chronic Treatment with Lacosamide and Topiramate on Cognitive Functions and Impaired Emotional Responses in a Pilocarpine-induced Post-status Epilepticus Rat Model.

INTRODUCTION: Epilepsy and antiepileptic drugs can affect negatively the cognitive abilities of patients. AIM: The present study aimed to evaluate the effect of topiramate (TPM) and lacosamide (LCM) on the emotional and cognitive re-sponses in naive animals and in animals with pilocarpine-induced status epilepticus. MATERIALS AND METHODS: Male Wistar rats were randomly divided into 6 groups and status epilepticus was evoked in half of them by a single i.p. administration of pilocarpine (Pilo) (320 mg/kg): Pilo-veh, Pilo-TPM (80 mg/kg) and Pilo-LCM (30 mg/kg). Matched naive rats were treated with the same doses as follows: C-veh, C-TPM, and C-LCM. In a step-down passive avoidance test, the learning session was held for one day, the early retention test was conducted on day 2, and the long-term memory test - on day 7. Motor activity and anxiety were evaluated in an open field test. RESULTS: The Pilo-TPM and Pilo-LCM groups increased the time spent on the platform compared to Pilo-veh animals while the C-LCM animals decreased the time compared to C-veh animals during short- and long-term memory retention tests. TPM and LCM exerted an anxiolytic effect in naive rats. The two antiepileptic drugs were unable to alleviate the hyperactivity, but they alleviated the impulsivity associated with decreased anxiety level in epileptic rats. CONCLUSIONS: Our findings suggest that LCM and TPM have a beneficial effect on cognition both in naive and epileptic rats. While the two antiepileptic drugs can produce an anxiolytic effect in naive rats, they alleviate the impulsivity after pilocarpine treatment.

Enhancing effect of aerobic training on learning and memory performance in rats after long-term treatment with Lacosamide via BDNF-TrkB signaling pathway.

Aerobic training has a neuroprotective effect, reduces the risk of developing neurodegenerative diseases and facilitates functional recovery. The present study assesses the effect of aerobic training on cognitive functions, hippocampal BDNF/TrkB ligand receptor system expression and serum levels of BDNF and corticosterone in intact rats after chronic treatment with Lacosamide (LCM). Male Wistar rats were randomly divided into two groups. One group was exercised on a treadmill (Ex) and the other one was sedentary (Sed). Half of the rats from each group received saline (veh) while the other half - LCM. The rats underwent a month-long training and LCM treatment before being subjected to one active and two passive avoidance tests. Both trained groups increased significantly the number of avoidances compared with the sedentary animals during the learning session and on memory retention tests, while the number of avoidances of the LCM-treated rats was significantly lower in comparison with the saline-treated animals. Both passive avoidance tests revealed that trained animals spent more time in the lighted compartment or caused longer stay on the platform than did the sedentary rats during acquisition and short- and long-term memory retention tests. Aerobic training increased BDNF and TrkB hippocampal immunoreactivity. We found no significant difference between BDNF serum levels but corticosterone levels of the Sed-LCM rats were lower than those of the Sed-veh animals. Our results show that aerobic training increases the hippocampal BDNF/TrkB expression suggesting a role in preventing the negative effect of Lacosamide on cognitive functions in rats.

Chronic treatment with the new anticonvulsant drug lacosamide impairs learning and memory processes in rats: A possible role of BDNF/TrkB ligand receptor system.

Cognitive impairment is considered a frequent side effect in the drug treatment of epilepsy. The objective of the present study was to investigate the effects of lacosamide (LCM) on learning and memory processes in rats, on the serum level of brain-derived neurotrophic factor (BDNF) and BDNF/TrkB ligand receptor system expression in the hippocampal formation. Male Wistar rats underwent long-term treatment with three different doses of lacosamide - 3 mg/kg (LCM 3), 10 mg/kg (LCM 10) and 30 mg/kg (LCM 30). All rats were subjected to one active and one passive avoidance tests. The BDNF/TrkB immunohistochemical expression in the hippocampus was measured and serum BDNF was determined. The LCM-treated rats made fewer avoidance responses than controls during acquisition training and in the memory retention test. The number of escapes in the LCM 10 and LCM 30 groups decreased throughout the test, while the rats in the LCM 3 group showed fewer escapes only in the memory test in the active avoidance task. In the step-down test, the latency time of the LCM-30 treated rats was reduced as compared with the controls during the learning session and the short- and long-term memory retention tests. Lacosamide induced a dose-dependent reduction of the hippocampal expression of BDNF and its receptor TrkB. We found no significant difference between BDNF serum levels in the test animals and controls. The results of the study suggest that LCM suppresses the learning and memory processes in rats, with the inhibition of hippocampal BDNF/TrkB ligand receptor system being one of the possible mechanisms causing this effect.

Retigabine diminishes the effects of acetylcholine, adrenaline and adrenergic agonists on the spontaneous activity of guinea pig smooth muscle strips in vitro.

PURPOSE: The aim of this study is to evaluate the effect of retigabine on the smooth muscle response to acetylcholine, adrenaline, α-and ß-adrenoceptor agonists. METHODS: We studied the change in the spontaneous smooth muscle contraction of guinea pig gastric corpus strips before and after 20-min treatment with 2µM retigabine. We also evaluated the effect of retigabine on the smooth muscle response to 10µM acetylcholine, 1 and 10µM adrenaline, 1µM methoxamine, 0.1µM p-iodoclonidine and 10µM isoproterenol. RESULTS: We observed a significant reduction in the effects of all studied mediators and agonists when they were added to organ baths in the presence of retigabine. Retigabine diminished the effect of acetylcholine on the spontaneous smooth muscle activity. The effect was fully antagonized by XE-991 (Kv7 channel blocker), which supports our hypothesis about the role of KCNQ channels in the registered changes. The increase in the contraction force after adding of 1µM adrenaline, methoxamine, and 0.1µM p-iodoclonidine was also significantly smaller in presence of retigabine. However, comparing the effect of 10µM adrenaline on the contractility before and after treatment with retigabine, we observed increased contractility when retigabine was present in the organ baths. CONCLUSION: A possible explanation for the observed diminished effects of mediators and receptor agonists is that the effect of retigabine on smooth muscle contractility is complex. The membrane hyperpolarization, the interaction between Kv7 channels and adrenoceptors, and the influence on signaling pathways may contribute to the summary smooth muscle response.

Effects of propolis and CAPE on proliferation and apoptosis of McCoy-Plovdiv cell line.

UNLABELLED: The mechanisms of action of propolis can be studied in detail by comparing the effects of propolis and the effects of its constituent components. AIM: To clarify and compare the effects of Bulgarian propolis and caffeic acid phenethyl ester (CAPE, a chemically synthesized component of propolis)--by using a set of cellular, molecular-biological and immunological techniques. MATERIAL AND METHODS: The McCoy-Plovdiv cell line was treated with propolis and CAPE in increasing concentrations (0.01, 0.1, 1.0, 10 mg/L, and 2.5, 4, 8, 16 mg/L, respectively). The expression of the proliferating cell nuclear antigen (PCNA) and the tumour-suppressor protein p53 was studied immunocytochemically. Apoptosis was measured using a highly sensitive microgel electrophoresis technique (comet assay). RESULTS: The results of the study showed corresponding changes in the expression of the examined proliferative antigens. PCNA was detected in all examined concentrations of the tested substances the expression being dose-dependent. Molecule localization changed from the nucleus to the cytoplasm. Treatment with CAPE brought about gradual attenuation of PCNA expression. High propolis concentrations induced increased synthesis of p53. No p53 expression was found when cells were treated with CAPE. The studied substances in their highest concentrations (10 mg/L propolis and 16 mg/L CAPE) had a cytotoxic effect. The comet assay showed DNA degradation kinetics characteristic for apoptosis. CONCLUSIONS: The present study demonstrates that high concentrations of propolis and CAPE cause apoptosis-induced cell death in McCoy-Plovdiv cells.