HPLC and GC Characterization of Dicliptera bupleuroides Aerial Parts and Evaluation of Its Anti-Inflammatory Potential in Vitro, in silico and in Vivo Using Carrageenan and Formalin Induced Inflammation in Rat Models.

The current study aimed to evaluate the anti-inflammatory activity of Dicliptera bupleuroides Nees aerial parts methanol extract and its different fractions namely hexane, chloroform, ethyl acetate and butanol in vitro using cyclooxygenase inhibitory assay (COX-2). In vivo anti-inflammatory evaluation was performed using carrageenan and formalin induced inflammation in rat models followed by molecular docking. High performance liquid chromatography (HPLC) and gas chromatography coupled with mass chromatography (GC/MS) analyses were used for chemical analyses of the tested samples. The tested samples showed significant inhibition in COX-2 inhibitory assay where methanol extract (DBM) showed the highest inhibitory potential at 100 µg/mL estimated by 67.86 %. At a dose of 400 mg/kg, all of the examined samples showed pronounced results in carrageenan induced acute inflammation in rat model at 4th h interval with DBM showed the highest efficiency displaying 65.32 % inhibition as compared to the untreated rats. Formalin model was employed for seven days and DBM exhibited 65.33 % and 69.39 % inhibition at 200 and 400 mg/kg, respectively approaching that of the standard on the 7th day. HPLC revealed the presence of caffeic acid, gallic acid and sinapic acid, quercetin and myricetin in DBM. GC/MS analysis of its hexane fraction revealed the presence of 16 compounds belonging mainly to fatty acids and sterols that account for 85.26 % of the total detected compounds. Molecular docking showed that hexadecanoic acid followed by decanedioic acid and isopropyl myristate showed the best fitting within cyclooxygenase-II (COX-II) while nonacosane followed by hexatriacontane and isopropyl myristate revealed the most pronounced fitting within the 5-lipoxygenase (5-LOX) active sites. Absorption, metabolism, distribution and excretion and toxicity prediction (ADMET/ TOPKAT) concluded that most of the detected compounds showed reasonable pharmacokinetic, pharmacodynamic and toxicity properties that could be further modified to be more suitable for incorporation in pharmaceutical dosage forms combating inflammation and its undesirable consequences.

Targeting Anti-Inflammatory Pathways to Treat Diabetes-Induced Neuropathy by 6-Hydroxyflavanone.

It is evident that inflammation and metabolic syndrome instigated by diabetes mellitus can precipitate diabetes-induced neuropathy (DIN) and pain. In order to find an effective therapeutic method for diabetes-related problems, a multi-target-directed ligand model was used. 6-Hydroxyflavanone (6-HF) carrying anti-inflammatory and anti-neuropathic pain potential due to its quadruplicate mechanisms, targeting cyclooxygenase-2 (COX-2), 5-lipoxygenase (5-LOX), and opioid and GABA-A receptors was investigated. The anti-inflammatory potential of the test drug was confirmed utilizing in silico, in vitro, and in vivo tests. A molecular simulation approach was utilized to observe the interaction of 6-HF with the inflammatory enzyme COX-2 as well as opioid and GABA-A receptors. The same was confirmed via in vitro COX-2 and 5-LOX inhibitory assays. In vivo tests were performed to analyze the thermal anti-nociception in the hot-plate analgesiometer and anti-inflammatory action in the carrageenan-induced paw edema model in rodents. The potential anti-nociceptive effect of 6-HF was evaluated in the DIN model in rats. The Naloxone and Pentylenetetrazole (PTZ) antagonists were used to confirm the underlying mechanism of 6-HF. The molecular modeling studies revealed a favorable interaction of 6-HF with the identified protein molecules. In vitro inhibitory studies revealed that 6-HF inhibited the COX-2 and 5-LOX enzymes significantly. The 6-HF at dosages of 15, 30, and 60 mg/kg substantially reduced heat nociception in a hot plate analgesiometer as well as carrageenan-induced paw edema in rodent models. The authors discovered that 6-HF had anti-nociception properties in a streptozotocin-induced diabetic neuropathy model. According to the findings of this study, 6-HF was demonstrated to diminish inflammation caused by diabetes as well as its anti-nociception effect in DIN.

Development of Tofacitinib Loaded pH-Responsive Chitosan/Mucin Based Hydrogel Microparticles: In-Vitro Characterization and Toxicological Screening.

Tofacitinib is an antirheumatic drug characterized by a short half-life and poor permeability, which necessitates the development of sustained release formulation with enhanced permeability potential. To achieve this goal, the free radical polymerization technique was employed to develop mucin/chitosan copolymer methacrylic acid (MU-CHI-Co-Poly (MAA))-based hydrogel microparticles. The developed hydrogel microparticles were characterized for EDX, FTIR, DSC, TGA, X-ray diffraction, SEM, drug loading; equilibrium swelling (%), in vitro drug release, sol-gel (%) studies, size and zeta potential, permeation, anti-arthritic activities, and acute oral toxicity studies. FTIR studies revealed the incorporation of the ingredients into the polymeric network, while EDX studies depicted the successful loading of tofacitinib into the network. The thermal analysis confirmed the heat stability of the system. SEM analysis displayed the porous structure of the hydrogels. Gel fraction showed an increasing tendency (74-98%) upon increasing the concentrations of the formulation ingredients. Formulations coated with Eudragit (2% w/w) and sodium lauryl sulfate (1% w/v) showed increased permeability. The formulations equilibrium swelling (%) increased (78-93%) at pH 7.4. Maximum drug loading and release (%) of (55.62-80.52%) and (78.02-90.56%), respectively, were noticed at pH 7.4, where the developed microparticles followed zero-order kinetics with case II transport. Anti-inflammatory studies revealed a significant dose-dependent decrease in paw edema in the rats. Oral toxicity studies confirmed the biocompatibility and non-toxicity of the formulated network. Thus, the developed pH-responsive hydrogel microparticles seem to have the potential to enhance permeability and control the delivery of tofacitinib for the management of rheumatoid arthritis.

Development and Evaluation of Cellulose Derivative and Pectin Based Swellable pH Responsive Hydrogel Network for Controlled Delivery of Cytarabine.

In the present study, pH-sensitive, biodegradable, and biocompatible Na-CMC/pectin poly(methacrylic acid) hydrogels were synthesized using an aqueous free radical polymerization technique and encapsulated by cytarabine (anti-cancer drug). The aim of the project was to sustain the plasma profile of cytarabine through oral administration. Sodium carboxymethyl cellulose (Na-CMC) and pectin were cross-linked chemically with methacrylic acid (MAA) as a monomer, using methylene bisacrylamide (MBA) as cross-linker and ammonium per sulfate (APS) as an initiator. Prepared hydrogel formulations were characterized for their texture, morphology, cytarabine loading efficiency, compositional and structural properties, thermal nature, stability, swelling response, drug release profile (pH 1.2 and pH 7.4), and in-vivo pharmacokinetic evaluation. Cytarabine-loaded hydrogels were also evaluated for their safety profile by carrying out toxicity studies in rabbits. Results demonstrated efficient encapsulation of cytarabine into the prepared network with loading ranging from 48.5-82.3%. The highest swelling ratio of 39.38 and maximum drug release of 83.29-85.27% were observed at pH 7.4, highlighting the pH responsiveness of the grafted system. Furthermore, cytarabine maximum release was noticed over 24 h, ensuring a sustained release response for all formulations. Histopathological studies and hemolytic profiles confirmed that the prepared hydrogel system was safe, biocompatible, and non-irritant, showing no symptoms of any toxicities and degeneration in organs. Moreover, pharmacokinetic estimation of the cytarabine-loaded hydrogel showed a remarkable increase in the plasma half-life from 4.44 h to 9.24 h and AUC from 22.06 µg/mL.h to 56.94 µg/mL.h. This study revealed that the prepared hydrogel carrier system has excellent abilities in delivering the therapeutic moieties in a controlled manner.

pH Responsive Hydrogels for the Delivery of Capecitabine: Development, Optimization and Pharmacokinetic Studies.

The objective of the current study was to achieve a sustained release profile of capecitabine (CAP), an anticancer agent frequently administered in conventional dosage form due to its short plasma half-life. A drug-loaded smart pH responsive chitosan/fenugreek-g-poly (MAA) hydrogel was synthesized by an aqueous free radical polymerization technique. The developed network was evaluated for capecitabine loading %, swelling response, morphology, structural and compositional characteristics, and drug release behavior. Significantly higher swelling and in vitro drug release rate were exhibited by formulations at pH 7.4 than at pH 1.2, demonstrating the pH responsive character of hydrogels. Swelling percentage and CAP loading ranged within 74.45-83.54% and 50.13-72.43%, respectively. Maximum release, up to 93%, was demonstrated over 30 h, evidencing the controlled release pattern of CAP from hydrogels. The optimized formulation was further screened for acute oral toxicity studies. No signs of oral, dermal, or ocular toxicities were noticed, confirming safety evidence of the network. Furthermore, pharmacokinetic analysis demonstrated the sustained release response of CAP from hydrogels as confirmed by a significant increase in plasma half-life (t1/2) (13 h) and AUC (42.88 µg h/mL) of CAP. Based on these findings, fabricated hydrogels are strongly recommended as a biocompatible carrier for colorectal delivery of active agents.

Evaluation of the Antihyperglycemic and Antihyperlipidemic Activity of Saussurea hypoleuca Root in Alloxan-Induced Diabetes in Rat Model and Correlation to Its Major Secondary Metabolites.

Saussurea hypoleuca belongs to the family Asteraceae, which has previously shown hepatoprotective, anticancer, and antioxidant activity. This study aimed to evaluate the antihyperglycemic and antihyperlipidemic activity of its root methanol extract and various fractions for the first time. This was performed using alloxan-induced diabetes in the rat model for both short, and long-term periods using different administration doses. Different biochemical parameters were studied and further consolidated by histopathological examination and in silico molecular modeling. The results showed that in the long-term study, at a dose of 400 mg/kg b.wt, the ethyl acetate fraction caused a pronounced reduction in fasting blood glucose level (FBG) and glycated hemoglobin (HbA1c) by 77.2% and 36.8%, respectively, compared to the diabetic group. This was confirmed by the histopathological examination of the animals' pancreatic sections. The ethyl acetate fraction also showed a reduction in total cholesterol (TC), total glycerides (TG), and low-density lipoprotein cholesterol (LDL-C) levels. It improved kidney and liver functions, causing a reduction in aspartate aminotransferase (AST), alkaline phosphatase (ALP), alanine transaminase (ALT), urea, and creatinine levels. This is mainly attributed to its richness in secondary metabolites. Molecular docking showed that all the tested compounds showed certain inhibitory potential towards human α-glucosidase (HAG) and ATP citrate lyase (ACL). Thus, Saussurea hypoleuca roots can help in the management of hyperglycemia, hyperlipidemia, and hepatic and kidney dysfunction.

Evaluation of The Antioxidant, Antimicrobial, and Anticancer Activities of Dicliptera bupleuroides Isolated Compounds Using In Vitro and In Silico Studies.

Phytochemical investigation of chloroform fraction (DBC) and ethyl acetate fraction (DBE) of D. bupleuroides (Acanthaceae) resulted in the isolation of ß-sitosterol (1) from DBC and vanillic acid (2) from DBE, which were first to be isolated from D. bupleuroides. ß-Sitosterol (1) exhibited substantial antioxidant activity (IC50 = 198.87 µg/mL), whereas vanillic acid (2) showed significant antioxidant power (IC50 = 92.68 µg/mL) employing 1,1-diphenyl-2-picrylhydrazyl (DPPH*) radical scavenging capacity assay. Both compounds showed pronounced antimicrobial activity using the agar disc diffusion method, particularly against fungi showing MIC values of 0.182 and 0.02 concerning Candida albicans, respectively, and 0.001 mg/mL regarding Penicillium notatum. They revealed considerable antibacterial activity with MIC values ranging between 0.467 and 0.809 mg/mL. Vanillic acid (2) exhibited substantial anticancer potential displaying 48.67% cell viability at a concentration of 100 µg/mL using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-Diphenyl-2H-Tetrazolium Bromide) assay concerning HepG2 cell lines. These results were further consolidated by in silico studies on different enzymes, where vanillic acid displayed a high fitting score in the active pockets of DNA-gyrase, dihydrofolate reductase, aminoglycoside nucleotidyltransferase, and ß-lactamase. It also inhibited human cyclin-dependent kinase 2 (CDK-2) and DNA topoisomerase II, as revealed by the in silico studies. ADME/TOPKAT (absorption, distribution, metabolism, excretion, and toxicity) prediction showed that vanillic acid exhibited reasonable pharmacodynamic, pharmacokinetic, and toxicity properties and, thus, could perfectly together with D. bupleuroides crude extract be incorporated in pharmaceutical preparations to counteract cancer and microbial invasion, as well as oxidative stress. Thus, it is concluded that D. bupleroides could be a potential source of therapeutically active compounds, which would be helpful for the discovery of clinically effective and safe drugs.

Development and validation of single analytical HPLC method for determination of flavoxate HCl in bulk, tablets and biological fluids.

A simple, sensitive and precise high performance liquid chromatographic (HPLC) method was developed and validated for determination of flavoxate HCI in raw material, tablets and biological fluids. The method followed by using the Zorbax XDB-C18 column containing Di-isobutyl n-octadeceylsilane (4.6mm×150mm, 5µm). The mobile phase consisted of acetonitrile: methanol: 0.15M sodium perchlorate (17:35:48 v/v) having pH 3. UV detection was carried out at 229nm at 40°C. Results indicated that the method has successfully established and validated in accordance with ICH guidelines acceptance criteria for linearity (0.03-7.5µg), accuracy (101.18-101.28%), robustness of column age and column lot (peak area %CV<0.04, purity %CV< 0.006) and robustness of HPLC condition (%CV<0.02), precision (intra and inter day precision assay, %CV values for peak area and percent purity of flavoxate HCl<2%) and system suitability parameters. The average noise, theoretical LOD and LOQ were found to be 0.01 mAU, 0.03 mAU and 0.6ng, respectively. The Coefficient of determination (r2) ranging from 0.03µg to 7.5µg, 0.99 which was within acceptable criteria of r2 & gt 0.99. The spiked recoveries of samples were 101.28, 101.18 and 101.18% respectively. All data revealed that this method can be used for in-vitro & in-vivo determination of flavoxate HCI in various pharmaceutical preparations.

In vitro phytochemical and anticancer activity of Misopates orontium L. and Dicliptera bupleuroides Nees.

In the present study phytochemical analysis and anticancer activity of Misopates orontium L. and Dicliptera bupleuroides Nees was carried out. Methanolic extracts of M. orontium and D. bupleuroides were selected for phytochemical analysis. The present analysis showed the presence of phytochemical such as carbohydrates, proteins, tannins, glycosides, alkaloids, saponins, phenols and flavonoids in M. orontium and D. bupleuroides. Anticancer assays including MTT, Alamar Blue (AB), Neutral Red (NR) and lactate dehydrogenase (LDH) were employed on whole herb methanolic extract and all other fractions of both plants to calculate the % age of cell viability and cell cytotoxicity. The percentage of cell viability was highly significant in all anticancer assays for all fractions. Therefore, ethyl acetate and aqueous fractions showed the excellent profile in evaluation of cytotoxicity in each assay. All above findings indicated that the whole herb of both selected plants have strong anticancer activity.

Evaluation of antioxidant, antimicrobial and anticancer activities of compounds reported Saussurea hypoleuca Spreng. roots.

Current study was intended to isolate bioactive compounds from ethyl acetate fraction of Saussurea hypoleuca root extract and evaluation of their antioxidant, antimicrobial and anti-cancerous activities which might be helpful for their chemo preventive potential against selected bacterial strains. Column chromatography was done for isolation of compounds which were characterized on the basis of extensive spectroscopic analysis; Infra-red (IR), Electron Ionization (EI-Positive), Proton Nuclear Magnetic Resonance (1H-NMR) and Carbon Nuclear Magnetic Resonance (13C-NMR). Two compounds were identified, as sesquiterpenes (40mg) and linoleic acid (33mg) from 10 grams of ethyl acetate fraction. Both compounds have shown in vitro antioxidant activity which in regard; 2, 2- diphenyl-1-picrylhydrazyl (DPPH) scavenging potential was high in sesquiterpenes (261.81) as compared to linoleic acid (90.89). The minimum inhibitory concentrations (MIC) of both compounds were evaluated in various bacterial and fungal strains against respective controls. However, in human hepatocellular carcinoma (Hep G2 cell lines) sesquiterpenes exhibited strong anticancer potential than linolenic acid which might be its potential free radical inactivator in MTT assay. This paper directs the ethano medicinal worth of plant root as it possesses bioactive compounds which in our best knowledge these compounds isolated and reported first time from this plant root specie.

Pharmacological evaluation of the gabapentin salicylaldehyde derivative, gabapentsal, against tonic and phasic pain models, inflammation, and pyrexia.

Gabapentinoids are effective drugs in most animal models of pain and inflammation with variable effects in humans. The current study evaluated the pharmacological activity of gabapentin (GBP) and its salicylaldehyde derivative (gabapentsal; [2-(1-(((2-hydroxybenzylidene) amino) methyl) cyclohexyl) acetic acid]; GPS) in well-established mouse models of nociceptive pain, inflammatory edema, and pyrexia at doses of 25-100 mg/kg. GPS allayed tonic visceral pain as reflected by acetic acid-induced nociception and it also diminished thermally induced nociception as a mimic of phasic thermal pain. Antagonism of GPS-induced antinociceptive activities by naloxone (NLX, 1.0 mg/kg, subcutaneously, s.c), beta-funaltrexamine (ß-FNT, 5.0 mg/kg, s.c), naltrindole (NT, 1.0 mg/kg, s.c), and nor-binaltorphimine (NOR-BNI, 5.0 mg/kg, s.c), and pentylenetetrazole (PTZ-15 mg/kg, intraperitoneally, i.p) implicated an involvement of both opioidergic and GABAergic mechanisms. Tail immersion test was conducted in order to delineate the mechanistic insights of antinociceptive response. Inflammatory edema induced by carrageenan, histamine, or serotonin was also effectively reversed by GPS in a fashion analogous to aspirin (150 mg/kg, i.p), chlorpheniramine (1.0 mg/kg, i.p), and mianserin (1.0 mg/kg, i.p), respectively. Additionally, yeast-induced pyrexia was decreased by GPS in a comparable manner to acetaminophen (50 mg/kg, i.p). These observations suggest that GPS possesses ameliorative properties in tonic, phasic, and tail immersion tests of nociception via opioidergic and GABAergic mechanisms, curbs inflammatory edema, and is antipyretic in nature.

A novel gabapentin analogue assuages neuropathic pain response in chronic sciatic nerve constriction model in rats.

Gabapentin (GBP) is an established drug that has been used in the management of symptoms of neuropathy but it is associated with unwanted side effects such as sedation and motor incoordination. The goal of the study was to find out a drug with greater efficacy and safety for the treatment of neuropathic pain. Our previously synthesized GABA analogue (Gabapentsal, GPS) was tested (25-100 mg/kg, i.p) in chronic constriction injury (CCI) induced nociceptive model of static allodynia, dynamic allodynia, thermal hyperalgesia, mechanical hyperalgesia and cold allodynia in rats (Sprague Dawley). Open field and rotarod tests were performed to assess the impact of GPS on the motor performance of the animals. GBP (100 mg/kg, i.p) was used as a standard for comparison. GPS dose dependently reduced static (P <0.001) and dynamic allodynia (P <0.001), thermal hyperalgesia (P <0.001), mechanical hyperalgesia (P < 0.001) and cold allodynia (P < 0.001). In comparison to GBP, GPS failed to alter any significantly the motor performance of rats in both the open field and rotarod assays. These results suggest that GPS is effective in alleviating nociception in CCI neuropathic pain model but free from the side effect of motor discoordination seen in the treatment with GBP. In conclusion, GPS may prove to be a prospectively more effective and safer option in the management of neuropathic syndromes.

Muriolide, a novel antioxidant lactone from Ranunculus muricatus.

Muriolide (1), a new aromatic lactone, has been isolated from the ethyl acetate fraction of Ranunculus muricatus. The compound was structurally characterized with the help of UV, IR, mass, 1D- and 2D-NMR data. It was tested in vitro for antioxidant and lipoxygenase inhibitory potential. Compound 1 showed good DPPH radical scavenging activity (IC50=56.9 µM), however it was moderately active against lipoxygenase enzyme (IC50=68.3 µM).

The antioxidant N-(2-mercaptopropionyl)-glycine (tiopronin) attenuates expression of neuropathic allodynia and hyperalgesia.

The current pharmacotherapy of neuropathic pain is inadequate as neuropathic pain involves varied clinical manifestations with multifactorial etiology, modulated by a cascade of physical and molecular events leading to different clinical presentations of pain. There is an accumulating evidence of the involvement of oxidative stress in neuropathy, and antioxidants have shown promise in mitigating neuropathic pain syndromes. To explore the evidence supporting this beneficial proclivity of antioxidants, this study investigated the antinociceptive effectiveness of N-(2-mercaptopropionyl)glycine or tiopronin, a well-recognized aminothiol antioxidant, in a refined chronic constriction injury (CCI) rat model of neuropathic pain. Tiopronin (10, 30, and 90 mg/kg, i.p.) and pregabalin (30 mg/kg, i.p.) were administered daily after CCI surgery. The neuropathic paradigms of mechanical/cold allodynia and mechanical/heat hyperalgesia were assessed on days 3, 7, 14, and 21 post-nerve ligation. At the end of study, malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) levels were estimated in the sciatic nerve, dorsal root ganglion, and spinal cord for assessing the extent of oxidative stress. The expression of neuropathic nociception was attenuated by tiopronin which was observed as a significant attenuation of CCI-induced allodynia and hyperalgesia. Tiopronin reversed the neuronal oxidative stress by significantly reducing MDA, and increasing SOD, CAT, and GSH levels. Pregabalin also showed similar beneficial propensity on CCI-induced neuropathic aberrations. These findings suggest prospective neuropathic pain attenuating efficacy of tiopronin and further corroborated the notion that antioxidants are effective in mitigating the development and expression of neuropathic pain and underlying neuronal oxidative stress.

6-Methoxyflavanone abates cisplatin-induced neuropathic pain apropos anti-inflammatory mechanisms: A behavioral and molecular simulation study.

Cisplatin is used as a first line therapy in treating cancers. However, its use is often accompanied with the development of peripheral neuropathy. 6-Methoxyflavanone (6-MeOF) is a positive allosteric modulator at GABAA receptors and is known for attenuating diabetes-induced neuropathic pain. Neuropathy was induced in male Sprague-Dawley rats (150-250 g), via intraperitoneal injection of cisplatin (3 mg/kg) once a week for four consecutive weeks. 6-MeOF (25, 50 and 75 mg/kg, i.p) and gabapentin (75 mg/kg, i.p) were administered 30 min before each cisplatin injection. Static and dynamic allodynia were assessed using von Frey filaments and cotton buds. The anti-inflammatory activity was analyzed with plethysmometer. Body weights were also measured each week. The binding affinity of 6-MeOF with chloride channel, Cyclooxygenase-1 (COX-1) and Cyclooxygenase-2 (COX-2) was studied using docking approach. The in vitro COX-1 and COX-2 inhibitory effect of 6-MeOF was conducted with COX colorimetric assay. Administration of cisplatin for four consecutive weeks induced static (decreased paw withdrawal threshold; PWT) and dynamic allodynia (decreased paw withdrawal latency; PWL). Co-administration of 6-MeOF for four weeks significantly attenuated the cisplatin-induced expression of nocifensive behaviors observed as significant increase in PWT and PWL. Moreover, it also prevented the body weight loss induced by cisplatin administration. In silico studies depicted a good interaction of 6-MeOF with chloride ion channels and COX-1 and COX-2 enzymes. The in vitro study confirmed the inhibitory activity of 6-MeOF for COX-1 and COX-2. 6-MeOF may be effective in attenuating cisplatin-induced allodynia, probably through interaction with GABAergic receptors and reducing inflammation.

Hepatoprotective and antioxidant activity of Dicliptera bupleuroides Nees. extracts on paracetamol induced hepatotoxicity in rats.

Aim of this study to evaluate the safety profile, hepatoprotective and in-vivo antioxidant activities of Dicliptera bupleuroides Nees. Toxicity studies were conducted in human RBCs and DNA by using standard procedures. Acute hepatoprotective investigation was carried out in albino rats by treated with all six fractions of D. bupleuroides 350 mg/kg/day. ALT, AST, ALP and total bilirubin (TB) were performed. The n-hexane fraction (200 mg/kg/day) exhibited appropriate hepatoprotective activity hence subjected to chronic study (14 days). Paracetamol induced the hepatotoxicity (350mg/kg) and silymarin (50 mg/kg) was standard drug. Liver function tests, liver peroxidation tests and histopathological examination were performed at the end. Hexane fraction showed significant decrease in the level of ALT (88.1±7.8), AST (93.8±7.6), ALP (136.3±8.4) and TB (0.6±0.03) as compared to the standard drug (p>0.05). Rats treated with ethyl acetate fraction showed decrease in MDA (42.8±0.7) while GSH was found to be increased (107.7±1.8) against the toxic group (51.3±2.9), (73.6±4.0) respectively. All the drug extracts decreased the oxidative stress and protect the DNA from free hydroxyl radicals. DNA damage protection activity of these fractions is due to phytochemicals present in these fractions. These results indicate that the plant fractions possess significant hepatoprotective and antioxidant activities with no toxic effects.

Acute and sub-acute toxicity study of a Pakistani polyherbal formulation.

BACKGROUND: Herbology is the prevailing system among the nationally-accepted alternative or complementary systems of medicine. The system is due to its general and patient-oriented methodology, is widely used in the general population exposing them to the risk of the side effects of the herbal medicines. METHOD: The aim of study was to assess the acute and sub-acute toxicity of the polyherbal formulation Hab-e-Kabad Noshadri tablets. In the acute arm of the study, a single dose of 2000 mg/kg was administered to Swiss Albino mice which were observed for physical symptoms and behavioral changes for 72 h. In sub-acute toxicity study repeated doses of the polyherbal preparation was administered to Wistar rats of both genders, separately. The animals received three doses of polyherbal product (50 mg/kg/day, 100 mg/kg/day and 200 mg/kg/day) for a period of 28 days. On 28th day of experiment, blood sampling of animals was done for hematological and biochemical analysis i.e. liver and renal function parameters, lipid profile and then sacrificed for histopathological examination of liver and kidney. RESULT: There was no morbidity and mortality noticed with single dose administration in acute toxicity study in mice. In sub-acute toxicity study, morphological changes with some damage in liver and kidney tissues of male and female animals were recorded at dose of 100 mg/kg/day and 200 mg/kg/day. CONCLUSIONS: It was found that prolonged use at higher dose i.e. 200 mg/kg/day of this polyherbal formulation should be avoided and practitioners should cautiously prescribe this formulation in patients with hepatic and renal impairment.

Characterization of 6-methoxyflavanone as a novel anxiolytic agent: A behavioral and pharmacokinetic approach.

Benzodiazepines are regularly prescribed for treatment of anxiety though there are side effects. Flavonoids have selective affinity for GABAA receptors implicated in anxiolytic-like activity in rodents, but are devoid of the unwanted side effects of benzodiazepines. In this study, 6-methoxyflavanone (6-MeOF), a positive allosteric modulator of γ-amino butyric acid (GABA) responses at human recombinant GABAA receptors, was evaluated for its behavioral profile in the elevated plus-maze as well as the staircase- plus and open-field tests in mice. In addition, the distribution of 6-MeOF in selected brain areas involved in anxiety (amygdala and cerebral cortex) was also examined using a validated high performance liquid chromatography ultraviolet detection (HPLC/UV) method. 6-MeOF (10, 30 and 50mg/kg) exerted an anxiolytic-like effect, increasing entries and time spent in the open arm and the central platform, as well as head-dipping frequency in the mouse elevated plus-maze assay. It also decreased rearing incidence without suppressing the number of steps ascended in the staircase test. Whereas, in the open-field anxiety test, 6-MeOF had no effect on locomotor activity at lower doses, a decrease was observed at the highest dose (100mg/kg). 6-MeOF additionally produced an anxiolytic-like increase in the time spent at the center of the open-field apparatus. These effects were preferentially antagonized by pentylenetetrazole (15mg/kg). Furthermore, pharmacokinetic studies disclosed a rapid appearance of 6-MeOF in the plasma and discrete brain areas. Taken together, our findings suggest that 6-MeOF readily crosses the blood brain barrier (BBB) generating anxiolytic activity, mediated through the GABAergic system.

6-Methoxyflavanone attenuates mechanical allodynia and vulvodynia in the streptozotocin-induced diabetic neuropathic pain.

BACKGROUND: Diabetic neuropathy is the most prevalent, persistent and debilitating complication of diabetes mellitus often coupled with vulvodynia that may present as an isolated symptom or as a part of constellation of other neuropathic abnormalities. OBJECTIVE: Flavonoids have selective affinity for GABA receptors and 6-methoxyflavanone (6-MeOF) is a positive allosteric modulator of GABA responses at human recombinant GABAA receptors. GABAergic and opioidergic system inhibition have been shown to facilitate neuropathic pain. METHODS: 6-MeOF was evaluated for analgesic effect in the hot plate test and streptozotocin-induced diabetic neuropathic pain in female rats using von Frey hairs. The possible involvement of opioidergic and GABAergic mechanisms was investigated using naloxone and pentylenetetrazole (PTZ) antagonists, respectively. The biodistribution of 6-MeOF in plasma and CNS was examined using a validated HPLC/UV analytical method. The binding affinity of 6-MeOF with opioid and GABA receptors was studied using molecular docking simulation approach. RESULTS: 6-MeOF (10 and 30mg/kg) attenuated the acute phasic thermal nociception in the hot plate test while in the case of streptozotocin-induced diabetic neuropathy model, 6-MeOF (10 and 30mg/kg) produced static/dynamic anti-allodynic (increased paw withdrawal threshold and latency) as well as static/dynamic anti-vulvodynic effects (increased flinching response threshold and latency), when compared to the vehicle and standard gabapentin (75mg/kg). In silico studies depicted the preference of 6-MeOF for the delta- and kappa-opioid and GABAA receptors. Moreover, the pharmacokinetic profile revealed a quick appearance of 6-MeOF in the systemic circulation and brain areas with maximum concentration observed after 30min in the amygdala, brain stem and cerebral cortex. CONCLUSION: 6-MeOF readily crosses the blood brain barrier and may be effective in attenuating the diabetes-induced allodynia as well as vulvodynia, probably through interactions with the GABAergic and opioidergic systems.