Neuroprotective properties of Melissa officinalis after hypoxic-ischemic injury both in vitro and in vivo

Background

Brain ischemia initiates several metabolic events leading to neuronal death. These events mediate large amount of damage that arises after some neurodegenerative disorders as well as transient brain ischemia. Melissa officinalis is considered as a helpful herbal plant in the prevention of various neurological diseases like Alzheimer that is related with oxidative stress.

Methods

We examined the effect of Melissa officinalis on hypoxia induced neuronal death in a cortical neuronal culture system as in vitro model and transient hippocampal ischemia as in vivo model. Transient hippocampal ischemia was induced in male rats by tow vessel-occlusion for 20 min. After reperfusion, the histopathological changes and the levels inflammation, oxidative stress status, and caspase-3 activity in hippocampus were measured.

Results

Cytotoxicity assays showed a significant protection of a 10 μg/ml dose of Melissa against hypoxia in cultured neurons which was confirmed by a conventional staining (P<0.05). Melissa treatment decrease caspase3 activity (P<0.05) and TUNEL-positive cells significantly (P<0.01). Melissa oil has also inhibited malon dialdehyde level and attenuated decrease of Antioxidant Capacity in the hippocampus. Pro-inflammatory cytokines TNF-α, IL-1β and HIF-1α mRNA levels were highly increased after ischemia and treatment with Melissa significantly suppressed HIF-1α gene expression (P<0.05).

Discussion

Results showed that Melissa officinalis could be considered as a protective agent in various neurological diseases associated with ischemic brain injury.     

Anxiolytic activity of methanol leaf extract of Achyranthes aspera Linn in mice using experimental models of anxiety

Objective:

To study the anxiolytic activity of methanol extract of Achyranthes aspera Linn (Amaranthaceae).

Materials and Methods:

Male Swiss albino mice were used. Methanolic extract of Achyranthes aspera (MEAA) was administered in the doses of 100, 300 and 600 mg/kg p.o. Hole board (HB), open field (OF), elevated plus maze (EPM) and light/dark exploration (LDE) tests were used for determination of anxiolytic activity.

Results:

The methanolic extract of Achyranthes aspera significantly increased the number and duration of head poking in HB test. The extract also significantly increased the time spent and the number of entries in open arm in EPM. In LDE test, the extract produced significant increase in time spent and number of crossings and decreased the duration of immobility in light box. In OFT, the extract showed significant increase in number of rearing, assisted rearing and the squares crossed.

Conclusion:

In the present study, MEAA exhibited anxiolytic activity which might be attributed to its phyto-constituents viz. alkaloid, steroid and triterpenes. Since Achyranthes aspera is ubiquitous and abundantly grown, it could be a fairly economical therapeutic agent for management of anxiety disorders.KEY WORDS: Achyranthesaspera, anxiolytic activity, elevated plus maze, hole board, light/dark exploration, MEAA, open field     

In vivo investigation of the neuroprotective property of Convolvulus pluricaulis in scopolamine-induced cognitive impairments in Wistar rats

Aim:

To investigate the neuroprotective effect of Convolvulus pluricaulis aqueous extract (AE) against scopolamine (1 mg/kg body weight (bwt))-induced neurotoxicity in the cerebral cortex of male Wistar rats.

Materials and Methods:

The study was carried out on male Wistar rats (age matched, weight 250 ± 20 g). The present study investigated cognitive-enhancing property of AE using Elevated plus maze (EPM) (transfer latency [TL]) and Morris water maze (MWM). Besides evaluating the effect of extract on neurochemical enzymes, in vivo antioxidant and free radical scavenging activities were also screened. All the measured parameters were compared with rivastigmine tartrate (1 mg/kg bwt) which was taken as standard.

Results:

Pretreatment of rats with AE (150 mg/kg bwt) significantly reduced scopolamine-induced increase in the TL in EPM, whereas in MWM, administration of extract improved the impairment of spatial memory induced by scopolamine. The activity of acetylcholinesterase (AChE) was significantly inhibited by extract within the cortex and hippocampus. Reduced activities or contents of glutathione reductase, superoxide dismutase, and reduced glutathione within the cortex and hippocampus induced by scopolamine were elevated by the extract. Taken together, it could be postulated that extract may exert its potent-enhancing activity through both anti-AChE and antioxidant action.

Conclusion:

AE possesses neuroprotective potential, thus validating its use in alleviating toxic effects of scopolamine.     

Anticataleptic and antiepileptic activity of ethanolic extract of leaves of Mucuna pruriens: A study on role of dopaminergic system in epilepsy in albino rats

Objective:

To assess the anticataleptic and antiepileptic activity of leaves of Mucuna pruriens in albino rats.

Materials and Methods:

Haloperidol-induced catalepsy (HIC), maximum electro-shock (MES) method, pilocarpine-induced Status epilepticus (PISE) and single-dose effect of M. pruriens were employed.

Results:

M. pruriens (100 mg/kg) had significant anticataleptic and antiepileptic activity in HIC, MES, and PISE.

Conclusions:

M. pruriens extract has the potential to be an anticataleptic and antiepileptic drug. Dopamine and 5-HT may have a role in such activity.     

Effects of Ocimum sanctum and Camellia sinensis on stress-induced anxiety and depression in male albino Rattus norvegicus

Objective:

The aim of this study was to study the ameliorative effects of Ocimum sanctum and Camellia sinensis on stress-induced anxiety and depression.

Materials and Methods:

The study was carried out using male albino rats (200 ± 50 g). The effect of O. sanctum and C. sinensis was evaluated for anxiety and depression using elevated plus maze (EPM) test, open field test (OFT), forced swim test (FST), and tail suspension test (TST).

Result:

Restraint stress (3 h/day for six consecutive days) induced a significant reduction in both the percentage number of entries and time spent in open arms in EPM, and these changes were reversed with post-treatment of aqueous extract of O. sanctum and C. sinensis (100 mg/kg for 6 days). Restraint stress-induced (a) increased latency and (b) decreased ambulation and rearing were also reversed by O. sanctum and C. sinensis in OFT. A significant increase in immobility period was observed in FST and TST after restraint stress. O. sanctum and C. sinensis significantly reduced the immobility times of rats in FST and TST.

Conclusion:

O. sanctum and C. sinensis possess anxiolytic and antidepressant activities.     

Kaurenic acid: An in vivo experimental study of its anti-inflammatory and antipyretic effects

Objective:

This study was designed to investigate the anti-inflammatory and antipyretic effects of kaurenic acid (KA), a tetracyclic diterpenoid carboxylic acid, using in vivo experimental animal models.

Material and Methods:

The anti-inflammatory activity of KA was evaluated in rats, using egg albumin-induced paw edema (acute test) and Freund’s complete adjuvant-induced paw edema (subacute test), whereas the antipyretic effect was studied in rabbits by peptone-induced pyresis. Acute and subacute toxicity of KA were analyzed in NMRI mice.

Results:

KA showed anti-inflammatory and antipyretic properties, and the effect caused was significantly dose-related (P < 0.001) in both cases. The mean lethal doses of KA were 439.2 and 344.6 mg/kg for acute and subacute toxicity, respectively.

Conclusion:

On the basis of these findings, it may be inferred that KA has an anti-inflammatory and antipyretic potential.     

Mentha piperita in nephrotoxicity – a possible intervention to ameliorate renal derangements associated with gentamicin

Objective:

Free radical generation has a strong role in the pathogenesis of renal damage associated with the use of gentamicin. Therefore, the present study was carried out to evaluate the renoprotective effect of Mentha piperita against gentamicin induced nephrotoxicity.

Materials and Methods:

A total of 24 male rabbits were divided into 4 groups receiving normal saline, gentamicin, M. piperita extract and co-therapy of extract and gentamicin respectively. Gentamicin was provided as 80 mg/kg/day intramuscularly and extract was given 200 mg/kg/day orally for a period of 21 days. Serum and urinary biochemical parameters and histological changes were studied for each group. The impact of the extract on the antibacterial action of gentamicin was also evaluated.

Results:

Animals treated with gentamicin showed derangements in serum and urinary biochemical parameters. These alterations were reversed by treatment with M. piperita extract. The histological changes showed in gentamicin group were also reverted by treatment with the extract. Further the plant did not influence the efficacy of gentamicin with respect to its antimicrobial properties.

Conclusion:

Co-therapy of M. piperita with gentamicin successfully attenuated biochemical kidney functioning derangements and morphological changes associated with gentamicin.KEY WORDS: Aminoglycosides, gentamicin, Mentha piperita, nephroprotective, peppermint     

Psychopharmacological effects and safety of styryl‐2‐pyrones and dihydrostyryl‐2‐pyrones‐rich fraction from Polygala sabulosa: absence of withdrawal syndrome and tolerance to anxiolytic‐like and anticonvulsant effects

Objectives

To investigate whether mice develop tolerance to the anxiolytic‐like and anticonvulsant effects of subchronic treatment with EA (the styryl‐2‐pyrones and dihydrostyryl‐2‐pyrones‐rich fraction of Polygala sabulosa ), as well as any withdrawal symptoms after abrupt discontinuation; to compare the effects of EA with those of diazepam (DZP ) on withdrawal‐induced anxiety; and to evaluate the toxicity of EA according to OECD guidelines.

Methods

Male or female mice were acutely or subchronically treated with EA or DZP , and their tolerance to anxiolytic (evaluated in the elevated plus maze, EPM ) and anticonvulsant effects (measured against pentylenetetrazole (PTZ )‐induced convulsions) were investigated. Other groups received EA or DZP for 28 days followed by withdrawal, being the anxiety‐like behaviour evaluated in the EPM .

Key findings

Both acute and subchronic treatments with EA induced an anxiolytic effect in the EPM . The anticonvulsant activity of DZP , but not EA , was reduced by protracted treatment. EA withdrawal retained the anxiolytic profile, while DZP withdrawal induced anxiogenesis. EA counteracted the anxiogenic‐like actions of DZP withdrawal. EA has low toxicity as it did not cause any changes in the biochemical, haematological and histopathological markers.

Conclusions

EA avoids the development of tolerance to its anxiolytic‐like and anticonvulsant actions, and does not promote withdrawal syndrome. EA does not cause relevant toxic effects in rodents.

     

Screening of anti-inflammatory and antipyretic activity of Vitex leucoxylon Linn

Objective:

This study was designed to evaluate the anti-inflammatory and antipyretic activity of ethyl acetate extract of Vitex leucoxylon Linn. in various animal experimental models.

Materials and Methods:

Ethyl acetate extract of V. leucoxylon Linn. evaluated for anti-inflammatory activity in carrageenan, mediator-induced rat paw edema, and cotton pellet-induced granuloma model. The antipyretic activity was evaluated by yeast-induced pyrexia model.

Results:

Single administration of the ethyl acetate extract of V. leucoxylon Linn. at dose of 500 mg/kg p.o. showed significant (P < 0.001) inhibition of rat paw edema. The ethyl acetate extract showed significant antipyretic activity in brewer yeast-induced pyrexia in rats throughout the observation period of 4 h.

Conclusion:

This study shows that ethyl acetate extract of V. leucoxylon Linn. has significant anti-inflammatory and antipyretic activity.     

Effect of Morus alba L. (mulberry) leaves on anxiety in mice

Objective:

The aim of the present work is to evaluate the anxiolytic effect of a methanolic extract of Morus alba L. leaves in mice.

Materials and Methods:

The hole-board test, elevated plus-maze paradigm, open field test, and light/dark paradigm were used to assess the anxiolytic activity of the methanolic extract of M. alba L. Morus alba extract (50, 100, and 200 mg/kg, i.p.) and diazepam (1 mg/kg, i.p.) were administered 30 min before the tests.

Results:

The results showed that the methanolic extract of M. alba significantly increased the number and duration of head poking in the hole-board test. In the elevated plus-maze, the extract significantly increased the exploration of the open arm in similar way to that of diazepam. At a dose of 200 mg/kg i.p. the extract significantly increased both the time spent in and the entries into the open arm by mice. Further, in the open field test, the extract significantly increased rearing, assisted rearing, and number of squares traversed, all of which are demonstrations of exploratory behavior. In the light/dark paradigm, the extract produced significant increase in time spent in the lighted box as compared to vehicle. The spontaneous locomotor activity count, measured using an actophotometer, was significantly decreased in animals pretreated with M. alba extract, indicating a remarkable sedative effect of the plant.

Conclusion:

The results of the present study suggest that a methanolic extract of M. alba leaves may possess an anxiolytic effect.     

Adaptogenic effect of Morus alba on chronic footshock-induced stress in rats

Objective:

The objective of the present study was to evaluate the adaptogenic property of the ethyl acetate-soluble fraction of methanol extract of Morus alba roots against a rat model of chronic stress (CS).

Materials and Methods:

Rats were exposed to stress procedure for 21 days. The stress procedure was mild, unpredictable footshock, administered for 1 h once daily for 21 days. Rats were administered with the ethyl acetate soluble fraction of methanol extract of M. alba roots (25, 50 and 100 mg/kg p.o) 1 h before footshock for 21 days and behavioral parameters were evaluated for cognitive dysfunction and depression using elevated plus maze and despair swim test, respectively. On day 21, rats were sacrificed immediately after stress and blood was collected for biochemical estimation. The adrenal gland and spleen were dissected for organ weight and the stomach was dissected for ulcer score.

Results:

CS significantly induced cognitive deficit, mental depression and hyperglycemia and increased blood corticosterone levels, gastric ulcerations and adrenal gland weight, but decreased the splenic weight. Pre-treatments with the ethyl acetate soluble fraction of methanol extract of M. alba roots (25, 50 and 100 mg/kg, p.o.) significantly attenuated the CS-induced perturbations. Diazepam (1 mg/kg, p.o.) was used as the standard antistress drug.

Conclusion:

The results indicate that M. alba possesses significant adaptogenic activity, indicating its possible clinical utility as an antistress agent.     

Antidiabetic activity of flower buds of Michelia champaca Linn

Objective:

To identify the antihyperglycemic activity of various extracts, petroleum ether (60-80°), chloroform, acetone, ethanol, aqueous and crude aqueous, of the flower buds of Michelia champaca, and to identify the antidiabetic activity of active antihyperglycemic extract.

Materials and Methods:

Plant extracts were tested for antihyperglycemic activity in glucose overloaded hyperglycemic rats. The effective antihyperglycemic extract was tested for its hypoglycemic activity at two-dose levels, 200 and 400 mg/kg respectively. To confirm its utility in the higher model, the effective extract of M. champaca was subjected to antidiabetic study in alloxan induced diabetic model at two dose levels, 200 and 400 mg/kg respectively. The biochemical parameters, glucose, urea, creatinine, serum cholesterol, serum triglyceride, high density lipoprotein, low density lipoprotein, hemoglobin and glycosylated hemoglobin were also assessed in the experimental animals.

Results:

The ethanolic extract of M. champaca exhibited significant antihyperglycemic activity but did not produce hypoglycemia in fasted normal rats. Apart from this extract, the crude aqueous and petroleum ether extracts were found active only at the end of the first hour. Treatment of diabetic rats with ethanolic extract of this plant restored the elevated biochemical parameters significantly (P<0.05) (P<0.01) and the activity was found dose dependent.

Conclusion:

This study supports the traditional claim and the ethanolic extract of this plant could be added in traditional preparations for the ailment of various diabetes-associated complications.     

Renal effects of Mammea africana Sabine (Guttiferae) stem bark methanol/methylene chloride extract on L-NAME hypertensive rats

Objective:

The present study aims at evaluating the effects of methanol/methylene chloride extract of the stem bark of Mammea africana on the renal function of L-NAME treated rats.

Material and Methods:

Normotensive male Wistar rats were divided into five groups respectively treated with distilled water, L-NAME (40 mg/kg/day), L-NAME + L-arginine (100 mg/kg/day), L-NAME + captopril (20 mg/kg/day) or L-NAME + M. africana extract (200 mg/kg/day) for 30 days. Systolic blood pressure was measured before and at the end of treatment. Body weight was measured at the end of each week. Urine was collected 6 and 24 h after the first administration and further on day 15 and 30 of treatment for creatinine, sodium and potassium quantification, while plasma was collected at the end of treatment for the creatinine assay. ANOVA two way followed by Bonferonni or one way followed by Tukey were used for statistical analysis.

Results:

M. africana successfully prevented the rise in blood pressure and the acute natriuresis and diuresis induced by L-NAME. When given chronically, the extract produced a sustained antinatriuretic effect, a non-significant increase in urine excretion and reduced the glomerular hyperfiltration induced by L-NAME.

Conclusions:

The above results suggest that the methanol/methylene chloride extract of the stem bark of M. africana may protect kidney against renal dysfunction and further demonstrate that its antihypertensive effect does not depend on a diuretic or natriuretic activity.     

Anticonvulsant and muscle relaxant activity of the ethanolic extract of stems of Dendrophthoe falcata (Linn. F.) in mice

Objective:

To investigate the anticonvulsant and muscle relxant activity of ethanolic extract of stems of Dendrophthoe falcata in mice.

Materials and Methods:

The ethanolic extract of stems of D. falcata (100, 300 and 500 mg/kg, p.o.) was studied for its anticonvulsant effect on maximal electroshock-induced seizures and muscle relaxant activity at the same dose level using rota rod and traction test in mice.

Results:

Preliminary phytochemical analysis revealed presence of proteins, carbohydrates, glycosides, steroids, triterpenes, flavonoids, tannins and phenolic compounds. D. falcata ethanolic extract (DFEE) (100, 300 and 500 mg/kg, p.o.) significantly (P<0.001) inhibited seizures induced by MES, reduced the duration of Hind limb tonic extensor phase (HLTE) and a decline in motor coordination.

Conclusion:

The ethanolic extract possesses anticonvulsant activity and muscle relaxant activity.KEY WORDS: Anticonvulsant, Dendrophthoe falcate, mice, muscle relaxant     

Acute and sub acute toxicity and efficacy studies of Hippophae rhamnoides based herbal antioxidant supplement

Objectives:

Present study was carried out to evaluate acute and subacute toxicity and efficacy of Seabuckthorn (Hippophae rhamnoides) based herbal antioxidant supplement (HAOS).

Materials and Methods:

In vivo toxicity studies were performed in male balb ‘C’ mice by oral administration. Acute toxicity study was done at doses ranging from 2000 to 10 000 mg/ kg while in subacute studies, HAOS was given at doses of 2000, 4000, and 8000 mg/kg body weight. Animals were observed for any toxic sign and symptoms periodically. At completion of study animals were sacrificed; their hematological, biochemical parameters were analyzed and histopathology of vital organs was done. In vivo efficacy studies in human volunteers were done and the levels of vitamin A and Vitamin C in blood samples were analyzed in comparison to a similar commercially available formulation.

Results:

No mortality and any clinical signs of toxicity were found in HAOS administered group of animals. There were no significant alterations in hematological and biochemical parameters. Histopathological analysis of vital organs showed normal architecture in all the HAOS administered groups. Human studies showed an increase of 32% and 172% in Vitamin A and Vitamin C levels respectively in term of bioavailability.

Conclusion:

The data obtained indicate no toxicity of this antioxidant supplement up to the highest dose studied. Efficacy in terms of increased bioavailability of vitamin A and C in human volunteers indicates the clinical usefulness of the supplement.KEY WORDS: Acute and subacute toxicity, herbal antioxidant supplement, seabuckthorn, histopathology and bioavailability     

Antimicrobial clerodane diterpenoids from Microglossa angolensis Oliv. et Hiern

Objective:

To identify the antimicrobial components present in Microglossa angolensis following fractionation of the methylene chloride extract of the aerial part of this plant.

Materials and Methods:

The plant was dried and extracted by percolation with methylene chloride. The dry extract was fractionated and purified by silica gel column chromatography. The isolated compounds were identified by comparison of their Nuclear Magnetic Resonance (NMR) spectral data with those reported in the literature. Antimicrobial activity was assayed by broth macro dilution method.

Results:

The crude extract of M. angolensis displayed significant antifungal and antibacterial activities (MIC = 312.50-1250μg/ml). 6β-(2-methylbut-2(Z)-enoyl)-3α,4α,15,16-bis-epoxy-8β,10βH-ent-cleroda-13(16),14-dien-20,12-olide and spinasterol were the most active compounds (MIC = 1.56-100μg/ml) and the most sensitive microorganisms were Enterococcus faecalis and Candida tropicalis for bacteria and yeasts respectively.

Conclusion:

The isolation of these active antibacterial and antifungal principles supports the use of M. angolensis in traditional medicine for the treatment of gastro-intestinal disorders.     

Studies on anti-inflammatory effect of aqueous extract of leaves of Holoptelea integrifolia, Planch. in rats

Objectives:

The purpose of the present study was to investigate the anti-inflammatory properties of aqueous extract of the leaves of H. integrifolia, Planch.

Materials and Methods:

The hind paw edema was produced in rats by subplanter injection of carageenan. The aqueous extract of H. integrifolia, Planch. (AHI) at dose (250 and 500 mg/kg p.o) was given to observe % inhibition of paw edema which were comparable with indomethacin (10 mg/kg p.o) used as a reference drug.

Results:

The extract administered orally at doses of 250 and 500 mglkg p.o produced a significant (P < 0.05) dose dependent inhibition of edema formation

Conclusions:

A significant % inhibition of paw edema by the aqueous extract of leaves of H. integrifolia, Planch. and its almost nearby same % inhibition with indomethacin suggest its usefulness as an anti-inflammatory agent.     

Effect of Tinospora crispa on thioacetamide-induced liver cirrhosis in rats

Objectives:

This study was conducted to determine the effect of ethanolic extract of the dried stems of Tinospora crispa in a male rat model of hepatic fibrosis caused by the hepatotoxin, thioacetamide.

Materials and Methods:

The extract was gavaged daily to the rats, at doses of 100 and 200 mg/kg along with thioacetamide at a dose of 200 mg/kg twice weekly. To assess the effectivity of extract, against thioacetamide, the activity of aminotransferases (alanine aminotransferase, aspartate aminotransferase), alkaline phosphatase (AP); and bilirubin were measured, together with morphological and histopathological indices in the liver of healthy and thioacetamide-treated rats.

Results:

A significant increase in the activity of liver enzymes, bilirubin and G-glutamyl transferase and gross and histopathological changes were determined. Although previous in vitro study established that this extract had strong antioxidant activity, this in vivo study establishes that this extract contains hepatotoxins whose identity may be quite different from those compounds with antioxidant properties.

Conclusion:

The study confirms that complete reliance on data obtained using in vitro methodologies may lead to erroneous conclusions pertaining to the safety of phytopharmaceuticals.     

Preliminary isolation and in vitro antiyeast activity of active fraction from crude extract of Gracilaria changii

Objective:

To isolate the active fraction from crude extract of Gracilaria changii and to determine its in vitro antifungal activity.

Materials and Methods:

The active fraction was isolated from the crude extract of G. changii by various purification procedures such as column chromatography, thin layer chromatography, bioauthograph etc. The in vitro antifungal activity (Candida albicans) of the active fraction (1.00, 0.50, and 0.25 mg/ml) was studied by disc diffusion method and the effect of the active fraction on the morphology of yeast was done by scanning electron microscope (SEM) studies.

Results:

An active fraction with remarkable antifungal activity was separated from the crude extract. The active fraction was effective as a fungicide against C. albicans and showed a dose-dependent antifungal activity. A Scanning Electron Microscope (SEM) study confirmed the fungicidal effect of G. changii active fraction on C. albicans, by changing the normal morphology of C. albicans.

Conclusion:

From G. changii crude extract, an active fraction with remarkable in vitro antifungal activity has been isolated.     

Calcium antagonistic activity of Bacopa monniera in guinea-pig trachea

Objective:

To demonstrate the calcium antagonistic property of ethanol extract of Bacopa monniera in guinea-pig trachea.

Materials and Methods:

The dose response curves of CaCl₂ (1 × 10^-5 to 1 × 10^-1 M) were constructed in the absence and presence of ethanol extract of Bacopa monniera (100, 500 and 700 μg/ml) or nifedipine (1 × 10^-6 M) in guinea-pig trachea in calcium free high K⁺-MOPS-PSS (3-(N-morpholino)-propanesulphonic acid physiological salt solution). The data was analyzed by ANOVA followed by least significant difference test or by Student''s ‘t’ test for unequal variance when appropriate. A probability of at least P < 0.05 was considered statistically significant.

Results:

The plant extract (500 and 700 μg/ml) significantly (P < 0.05) depressed and shifted the calcium concentration-response curves (1 × 10^-3- 1 × 10^-1 M) to rightward similar to that of nifedipine.

Conclusions:

Bacopa monniera extract exhibited calcium channel blocking activity in guinea-pig tracheal smooth muscles that may rationalize its relaxant action on guinea-pig trachea and its traditional use in respiratory disorders.KEY WORDS: Bacopa monniera, calcium channel antagonist, smooth muscle relaxation