Chondroprotective activity of a detoxicated traditional Chinese medicine (Fuzi) of Aconitum carmichaeli Debx against severe-stage osteoarthritis model induced by mono-iodoacetate

Ethnopharmacological relevance: Fuzi is an effective but toxic traditional Chinese medicine (TCM) derived from Aconitum carmichaeli. In our previous study, detoxicated Fuzi (d-Fuzi) has been originally developed with no toxicity but significant efficacy. However, whether d-Fuzi can be used for therapy of osteoarthritis (OA), remain unknown.Materials and methods: Severe OA model was established by intra-articular mono-iodoacetate (MIA) injection (1.25 mg) into rats and orally treated with 2 g/ml d-Fuzi at a dosage of 7 ml/kg body weight for 28 days. In vivo, the articular radiographic and histopathologic analyses were performed to qualitatively assess the chondroprotective effect of d-Fuzi, followed by quantitative measurements of bone density and Mankin scores. In vitro, such effect on chondrocyte viability after MIA attack was evaluated. Hybrid quadrupole time-of-flight mass spectrometry (QTOF-MS) was performed for chemical analysis of d-Fuzi.Results: d-Fuzi was demonstrated to possess chondroprotective activity on MIA-induced OA model by in vivo preventing the articular degeneration and the reducing of bone density and Mankin score, as well as by in vitro promoting the chondrocyte proliferation and inhibiting the MIA-induced chondrocyte damage. A total of 23 compounds were identified in d-Fuzi, most of which were deduced as the non-toxic derivatives of aconite alkaloids.Conclusions: This is the first report regarding chondroprotective effect and chemical profile of d-Fuzi, originally revealing its great anti-OA potential and thereby providing a promising TCM candidate for OA therapy.     

Safety evaluation of main alkaloids from Rhizoma Coptidis

Ethnopharmacological relevance

Rhizoma Coptidis (RC), a widely used traditional Chinese medicine, has been used for the treatment of heat-clearing and detoxifying, but there is very little information on its safety.

Aim of the study

To provide information on the safety of RC, we evaluated the toxicity of the crude RC and RC alkaloids (berberine, coptisine, palmatine and epiberberine) including cytotoxicity, acute toxicity in mice and sub-chronic toxicity in rats.

Materials and methods

The cytotoxicity of RC alkaloids was tested in HepG2 and 3T3-L1 cells by the MTT assay. The acute toxicity of RC alkaloids was tested in mice and the mortality was calculated at the end of experiment. For sub-chronic toxicity study, the rats were treated with the RC alkaloids at a dose of 156 mg/kg/day and RC at a dose of 521 mg/kg/day for 90 days. Mortality, clinical signs, body weight changes, organ weights, urinalysis and hematological parameters, gross necropsy and histopathology were monitored during the study period.

Results

The cell assay indicates that the IC₅₀ values of berberine, coptisine, palmatine and epiberberine in HepG2 cells were 48.17, 64.81, 112.80 and 120.58 μg/mL, which in 3T3-L1 cells were 41.76, 56.48, 84.32 and 104.18 μg/mL, respectively. In the acute toxicity assay, the LD₅₀ values of four alkaloids were 713.57, 852.12, 1533.68 and 1360 mg/kg, respectively. However, in the sub-chronic toxicity study, no mortality and morbidity were observed which could be related to RC alkaloids and RC treatment. Besides, there was no abnormality in clinical signs, body weights, organ weights, urinalysis, hematological parameters, gross necropsy and histopathology in any of the animals after the oral administration of RC alkaloids and RC.

Conclusions

Taking these results together, we came to the conclusion that the toxicity of berberine is the maximum and palmatine is the minimal in four RC alkaloids. The currently recommended doses of RC alkaloids and RC consumed are relatively safe.     

Acute and sub-chronic oral toxicity studies of an aqueous stem bark extract of Pterocarpus soyauxii Taub (Papilionaceae) in rodents

Pterocarpus soyauxii Taub (Papilionaceae) is used in Cameroonian traditional medicine and pharmacopoeia to treat hypertension, diabetes, gastrointestinal parasitizes and cutaneous diseases.

Aim of the study

The present investigation was carried out to evaluate the safety of an aqueous stem bark extract of Pterocarpus soyauxii by determining toxicity after acute and sub-chronic oral administration in male and female rodents.

Materials and methods

The acute toxicity test was conducted in mice. An aqueous extract of barks was administrated by gavage in single doses of 2.5-12.5 g/kg. General behaviour and mortality were examined for up to 7 days. The sub-chronic toxicity test was performed in rats. The plant extract was administered by daily gavage of 150-600 mg/kg for 42 days. Body weight, food and water intakes were followed weekly. Haematological, biochemical and organ parameters were determined at the end of the 42-day administration.

Results

In the acute study in mice, oral administration of the aqueous extract of Pterocarpus soyauxii caused dose-dependent general behaviour adverse effects and mortality. The no-observed adverse effect level (NOAEL) of the extract was 5.0 g/kg. The lowest-observed adverse effect level (LOAEL) was 7.5 mg/kg. Mortality increased with the dose, LD₅₀ was > 10.75 g/kg for the mouse. In the sub-chronic study in rats, daily oral administration of the aqueous extract of Pterocarpus soyauxii did not result in death or significant changes in haematological or biochemical parameters, excepted increased hepatic catalase activity (P < 0.05) at the dose of 600 mg/kg. No alteration was observed in body weight, food and water intake. Liver, kidney, lung and pancreas histopathology did not reveal morphological alteration.

Conclusions

The results showed that the aqueous stem bark extract of Pterocarpus soyauxii Taub had very low toxicity in oral acute high dose administration and no toxicity in oral sub-chronic low dose administration and indicate that the plant could be considered safe for oral medication.     

The effects of Rhizoma Zingiberis on pharmacokinetics of six Aconitum alkaloids in herb couple of Radix Aconiti Lateralis−Rhizoma Zingiberis

Ethnopharmacological relevance

Radix Aconiti Lateralis (Fuzi in Chinese, derived from the lateral roots of Aconitum Carmichaeli Debx.) is widely used for the treatment of heart failure, internal cold, arthralgia, diarrhea and edema for thousands of years. It was usually prescribed in combination with Rhizoma Zingiberis (Ganjiang in Chinese, derived from the dry rhizome of Zingiber officinale Rosc.) to decrease toxicity and increase efficacy.

Aim of the study

In order to investigate the influence of Rhizoma Zingiberis on pharmacokinetics of six Aconitum alkaloids, i.e. aconitine (AC), hypaconitine (HA), mesaconitine (MA), benzoylaconine (BAC), benzoylhypaconine (BHA) and benzoylmesaconine (BMA), in Fuzi−Ganjiang herb couple, the comparative pharmacokinetics of six Aconitum alkaloids after oral administration of Fuzi and Fuzi−Ganjiang aqueous extract was carried out.

Materials and methods

A sensitive, specific and rapid LC−MS/MS method was developed to determine the six analytes in plasma. Then the rats were randomly divided into two groups and orally administered with Fuzi and Fuzi−Ganjiang aqueous extract. At designated time points after oral administration, the concentrations of the six Aconitum alkaloids in rat plasma were determined, and main pharmacokinetic parameters were investigated using 3P97 (Practical Pharmacokinetics Program Version 1.0).

Results

Comparing with Fuzi group, both T_1/2 and AUC_0−t of AC and HA decreased (P<0.05), while T_1/2, AUC_0−t and C_max of BAC, BHA increased (P<0.05) in Fuzi−Ganjiang group, which indicated that Ganjiang could promote the elimination of AC and HA and enhance the absorption of BAC, BHA and BMA.

Conclusion

The differences of pharmacokinetics of Aconitum alkaloids in rat plasma could support those of pharmacologics and toxicity in previous reports between Fuzi and Fuzi−Ganjiang herb couple. The results might be helpful in explaining the mechanism of combination of Fuzi−Ganjiang to decrease toxicity and increase efficacy.     

Evaluation of safety of the herbal formula Ojeok-san: Acute and sub-chronic toxicity studies in rats

Aim of the study

Ojeok-san (OJS; wuji powder in China and goshaku-san in Japan), a widely used herbal formula in traditional Korean medicine and Japanese herbal medicine (Kampo medicine), has been used to treat common cold and illnesses including fatigue and gastrointestinal disorders, but there is very little information on its safety. To provide information on the safety of OJS, we evaluated its acute and sub-chronic toxicity in rats.

Materials and methods

The single and sub-chronic toxicity of OJS was examined using male and female Sprague-Dawley rats. The rats were treated with the OJS extract orally at the highest dose level of 2000 mg/kg/day body weight. After single administration, signs of toxicity were observed every hour for the first 6 h and every day for two weeks. In the sub-chronic toxicity study, OJS was administered for 13 weeks. Mortality, clinical signs, body weight changes, food and water consumption, ophthalmologic findings, urinalysis, hematological and biochemical parameters, gross findings, organ weights and histological markers were monitored during the study period.

Results

We found no mortality and no abnormality in clinical signs, body weight, and necropsy findings for any of the animals in the acute and sub-chronic toxicity study following oral administration of OJS.

Conclusion

OJS may not have any single dose toxicity. The lethal dose with a 50% mortality rate (LD₅₀) was over 2000 mg/kg. The no-observed adverse effects level (NOAEL) was considered to be 2000 and 1000 mg/kg/day for male and female rats, respectively.     

Evaluation of toxicity of Calophyllum brasiliense stem bark extract by in vivo and in vitro assays

Ethnopharmacological relevance

Calophyllum brasiliense Camb., Clusiaceae, is commonly known as “guanandi” and its stem bark is used in Brazilian traditional medicine to treat rheumatism, vein problems, hemorrhoids and gastric ulcers. The aim of this study was to evaluate the toxicity of hexane extract of Calophyllum brasiliense stem bark (HECb) using in vitro and in vivo experimental models.

Materials and methods

In vitro toxicity was evaluated by Alamar Blue cytotoxicity assay and micronucleus test, using Chinese hamster ovary (CHO-k1) epithelial cells. in vivo toxicity was evaluated by oral acute and subchronic toxicity assays. In the oral acute toxicity screening, a single dose of HECb was administered to mice at doses ranging from 250 to 1000 mg/kg. In the subchronic study, HECb was administered orally for 30 days to Wistar rats at doses of 100 mg/kg and 500 mg/kg. Phytochemical analyses were performed by HPLC/UV–vis, secondary metabolites were quantified by spectrophotometric methods.

Results

HECb presented IC₅₀=119.94±4.31 µg/mL after a 24 h cytotoxicity test using CHO-k1 cells, showing low cytotoxicity. However, when the cells were exposed to HECb for 72 h, the IC₅₀ value was 8.39±2.00 µg/mL, showing in this case, a pronounced cytotoxic effect. In the oral acute toxicity studies, doses up to 500 mg/kg of HECb did not cause any changes in both male and female mice. At 1000 mg/kg, male mice showed signs typical of depression and stimulation that were reversed at 72 h. Besides, female mice were more sensitive to the toxic effect of HECb at 1000 mg/kg, which initially presented typical agitation signals, followed by depression signals, leading to death of all the animals at 24 h. In subchronic assay with rats, HECb administered orally at doses of 100 and 500 mg/kg did not cause significant changes in all clinical parameters evaluated. Histopathological analyses showed no deleterious effect in the vital organs of rats. Preliminary phytochemical analysis revealed the presence of phenolic compounds, steroids, and volatile coumarins. Analysis by HPLC showed two major peaks characteristic of chromanones.

Conclusions

In vitro toxicological tests showed that HECb exhibited cytotoxicity especially after 72 h of exposition, and mutagenicity on the highest tested dose. The in vivo studies demonstrated that HECb produced some toxicity signs at the highest dose tested, particularly, in the acute toxicity test but showed no significant signs of toxicity in the subchronic assay. Based on these and previous pharmacological studies, it is possible to say that HECb did not exhibit significant toxicity at its effective dose. This suggests that HECb is relatively safe in humans at its effective dose.     

Acute and sub-chronic toxicity of a lyophilised aqueous extract of Centaurium erythraea in rodents

Ethnopharmacological relevance

An aqueous concoction made from centaury (Centaurium erythraea (L.) Rafn., (Gentianaceae) whole plant is used in the Moroccan traditional medicine for the treatment of diabetes, as well as a number of other diseases. No systematic study of the potential toxicity of the plant has been described.

Aim of the study

The present investigation was carried out to evaluate the safety of an aqueous extract of Centaurium erythraea whole plant (CE-extract) by determining its potential toxicity after acute and sub-chronic administration in rats and mice.

Materials and methods

For the acute study, the lyophilised CE-extract was administered to adult IOPS OFA mice in single oral doses of 1-15 g/kg given by gavage, and single intraperitoneal (i.p.) doses of 1-14 g/kg. General behavioral adverse effects, mortality, and latency of mortality were determined for up to 14 days. In the sub-chronic dose study, the CE-extract was administered orally at doses of 100, 600 and 1200 mg/kg daily for 90 days to Wistar rats. Body weight and selected biochemical and hematological parameters were determined every 30 days and at the end of 90 days of daily administration; sections of liver and kidney were examined histologically for any signs of organ damage at the end of the treatment.

Results

In the acute study in mice, there were no deaths or any signs of toxicity observed after oral administration of single doses of the CE-extract at any dose level up to the highest dose tested (15 g/kg), which was the no-observed-adverse-effect level (NOAEL). However, the mortality rate as well as the acute toxicity of the i.p. administered CE-extract increased progressively with increasing dose. The NOAEL for the i.p. dose was 6 g/kg while the lowest-observed-adverse-effect level (LOAEL) was 8 g/kg; the calculated acute toxicity (LD₅₀) of i.p. administered CE-extract in mice was 12.13 g/kg.In sub-chronic studies in rats, the CE-extract (administered orally at daily doses of 100, 600 and 1200 mg/kg for 90 days), did not cause any changes in hematological and biochemical parameters, except a small reduction of mean corpuscular volume, and a decrease in serum glucose and triglyceride levels at the higher doses. Histopathological examination of the liver and kidneys at the end of the study showed normal architecture suggesting no morphological disturbances.

Conclusions

Because of the lack of toxicity of the CE-extract given by the oral route, and relatively high NOAEL values for the i.p. dose in the acute study in mice, as well as lack of mortality or clinically significant adverse changes in the biological and hematological parameters, and the morphology of liver and kidneys in rats after 90 days of daily dosing, it may be concluded that the CE-extract is relatively non-toxic. Also, in view of the doses consumed empirically in traditional medicine in Morocco, there is a wide margin of safety for the therapeutic use of Centaurium erythraea.     

In vitro and in vivo safety evaluation of Acer tegmentosum

Ethnopharmacological relevance

Acer tegmentosum, which contains salidroside and tyrosol, has been used for the treatment of hepatic disorders in eastern Asia. However, little is known about its safety.

Aim of the study

To determine the safety of Acer tegmentosum, we evaluated its acute oral toxicity and genotoxicity profiles.

Materials and methods

Salidroside and tyrosol present in Acer tegmentosum were quantified using high-performance liquid chromatography. Acute oral toxicity testing of Acer tegmentosum was performed in rats. Genotoxicity of Acer tegmentosum was assessed by bacterial reverse mutation, chromosomal aberration, and bone marrow micronucleus tests. All the tests were conducted in accordance with the good laboratory practices.

Results

The amounts of salidroside and tyrosol in Acer tegmentosum were found to be 85.01±1.21 mg/g and 3.12±0.04 mg/g, respectively. In the bacterial reverse mutation test, Acer tegmentosum increased the number of revertant Salmonella typhimurium TA98 colonies, regardless of metabolic activation by S9 mixture. In contrast, Acer tegmentosum application did not significantly increase the number of chromosomal aberrations in Chinese hamster ovary (CHO)-K1 cells and micronucleated polychromatic erythrocytes in mice. In the acute oral toxicity test, the median lethal dose (LD₅₀) of Acer tegmentosum was found to be >2000 mg/kg in rats.

Conclusion

Take together, Acer tegmentosum exhibits mutagenicity, which was evident from the bacterial reverse mutation test. Further studies are needed to identify the components responsible for such an effect and the underlying mechanisms.     

Acute toxicity studies of the leaf extract of Ficus exasperata on haematological parameters,body weight and body temperature

Introduction

The plant, Ficus exasperata is popularly used in Nigeria and in several parts of Africa for a variety of ailments.

Aim of study

This study was thus mapped out to investigate the toxicity profile of the aqueous leaf extract (AET) on haematological parameters, body weight and body temperature in mice.

Methodology

In the present study, AET was evaluated for acute toxicity over 24 h and 14-day periods. The LD₅₀ was assessed via oral and intraperitoneal administration.

Results

The LD₅₀ was indeterminable via the oral route but was determined to be 0.54 g/kg i.p. In the 24 h and 14 days single dose study, oral administration of 2.5, 5, 10 and 20 g/kg of AET produced neither mortality nor changes in behavior or any other physiological activity in mice. Body weights and body temperatures were not significantly altered. Haematological analysis showed no marked differences in any of the parameters examined (WBC count, platelet and haemoglobin estimation) in either the control or treated groups. However, the 14 days daily dose study showed significant increase in body temperature (p < 0.05) and a significant decrease in the red blood cell count, haemoglobin count and haematocrit values (p < 0.05), while other parameters remained unchanged.

Conclusion

In summary, AET was found to be relatively safe on short-term oral administration. However, chronic toxicity studies are required for the support of the safe use of this plant.     

Anticonvulsant,anxiolytic and sedative activities of the aqueous root extract of Securidaca longepedunculata Fresen.

Aim of the study

The objective of this study is to investigate the anticonvulsant, anxiolytic and sedative activities of the aqueous root extract of Securidaca longepedunculata.

Materials and methods

The anticonvulsant effect of the aqueous root extract (100, 200 and 400 mg/kg) was evaluated in mice using the strychnine- and picrotoxin-induced seizure models. Its anxiolytic activity was evaluated using the elevated plus maze (EPM) and the Y maze (YM) methods (14 and 32) while the hexobarbitone induced sleep and the hole board models were used to evaluate the sedative and exploratory activities in mice respectively. The acute toxicity studies and phytochemical analysis of the extract were also carried out.

Results

The extract (100–400 mg/kg) produced a significant (P < 0.01) dose dependent increase in onset of convulsion compared to the control for strychnine- and picrotoxin-induced seizures. It also produced a significant (P < 0.01) dose dependent prolongation of the cumulative time spent in the open arms of the elevated plus maze and Y maze compared with the control. The extract (100–400 mg/kg) produced significant (P < 0.01) reduction in the time of onset of sleep induced by hexobarbitone. The prolongation of hexobarbitone sleeping time by the extract (200 mg/kg) was comparable to that produced by diazepam (3 mg/kg). At doses of 100–400 mg/kg, the extract produced a dose dependent decrease in exploratory activity of the mice. The reduction in exploratory activity produced by the extract (400 mg/kg) was greater than that of chlorpromazine (1 mg/kg). The results obtained from the experiments indicate that the extract has central nervous system depressant and anxiolytic activities. The LD₅₀ obtained for the acute toxicity studies using both oral and intraperitoneal routes of administration were 1.74 g/kg and 19.95 mg/kg respectively.

Conclusion

These findings justify the use of Securidaca longepedunculata in traditional medicine for the management of convulsion and psychosis.     

Toxicology evaluation of standardized methanol extract of Gynura procumbens

Aims of the study

Gynura procumbens (Merr.), which is known as “Sambung nyawa”, is widely used in South East Asian countries in the traditional treatment of many ailments. However, there is little toxicological information available regarding safety following repeated exposure.The present investigation describes the toxicity of a methanol extract of Gynura procumbens leaves in rats.

Materials and methods

For acute toxicity studies, a methanol extract of Gynura procumbens was orally administered to Sprague–Dawley rats (female and male) at a dose range of 1000–5000 mg/kg. For sub-chronic toxicity studies, the rats were orally administered the methanol extract of Gynura procumbens at the doses of 125, 250, and 500 mg/(kg day) for a period of 13 weeks. The animals were sacrificed, followed by examination of their organs and blood serum.

Results and conclusions

Administration of the methanol extract from Gynura procumbens leaves at 1000–5000 mg/kg did not produce mortality or significant changes in the general behaviour, body weight, or organ gross appearance of rats. There were no significant differences in the general condition, growth, organ weights, haematological parameters, clinical chemistry values, or gross and microscopic appearance of the organs from the treatment groups as compared to the control group. Therefore, the NOAEL for the Gynura procumbens methanol extract is 500 mg/(kg day) administered orally for 13 weeks.     

Anticonvulsant activity of Benkara malabarica (Linn.) root extract: In vitro and in vivo investigation

Aim of the study

To systematically investigate the anticonvulsant activity of methanol extract of Benkara malabarica roots and to provide a biochemical basis elucidating its mode of action.

Methods

The median lethal dose (LD₅₀) of Benkara malabarica extract was determined. The anticonvulsant activity of the extract was assessed in strychnine-induced and isoniazide-induced convulsion models; phenytoin (20 mg/kg) and diazepam (1 mg/kg) were used as standards, respectively. Percentage protection provided by the drug was accounted as decrease in the number of convulsions within 8 h of observation. Mechanism of action was studied by performing GABA transaminase (GABA-T) assay, isolated from rat brain. Active constituent was isolated and characterized from the plant extract.

Results

The median lethal dose (LD50) of Benkara malabarica was found to be more than 500 mg/kg. It demonstrated 30% and 35% protection against strychnine-induced convulsions and 60% and 80% protection against isoniazide-induced convulsions, at doses of 25 mg/kg and 50 mg/kg, respectively. Enzyme assay results revealed that Benkara malabarica extract possesses GABA-T inhibitory activity (IC50 = 0.721 mg/ml). Scopoletin which was identified as the major constituent of the extract was found to be an inhibitor of GABA-T (IC50 = 10.57 μM).

Conclusions

The anticonvulsant activity of the plant extract is predominantly GABA mediated and may be due to the action of scopoletin alone or is a result of synergy of different compounds in the extract in which scopoletin is the major constituent.     

Appraisal of antinociceptive and anti-inflammatory potential of extract and fractions from the leaves of Torreya grandis Fort Ex. Lindl

Torreya grandis (Taxaceae) was studied for antinociceptive and anti-inflammatory effects. Leaves were extracted with 80% ethanol at 80 °C for 3 h and fractionated with petroleum ether, chloroform, ethylacetate and n-butanol. The aqueous ethanolic extract (aq.EE), ethylacetate fraction (EaF) and butanol fraction (BtF) at the doses of (100 and 200 mg/kg, i.g.) body weight were used for study. Evaluation of antinociceptive activity was carried out by acetic acid-induced writhing response and formalin-induced paw licking time in the first and second phases of mice. The paw edema induced by formalin- and xylene-induced ear edema were used to assess anti-inflammatory activity. It was found that Torreya grandis extract and fractions at the doses of (100 and 200 mg/kg, i.g.) were significantly attenuated the writhing responses induced by acetic acid and second phase of pain response induced by subplantar injection of formalin in mice. In addition, these extract and fractions inhibiting the formaldehyde-induced arthritis as well as xylene-induces edema prolifically. From acute oral toxicity studies no mortality was pragmatic even at highest dose (2500 mg/kg, i.g.). Furthermore, our phytochemical studies indicated that the aq. ethanolic extract of leaves contains alkaloids, flavonoids, tannins, terpenoids and saponins. The results provide justification for the folkloric uses of Torreya grandis in the treatment of analgesic and inflammatory-based diseases across the China.     

In vivo and in vitro toxicological evaluation of the hydroalcoholic leaf extract of Ageratum conyzoides L. (Asteraceae)

Ethnopharmacological relevance

In African traditional medicine, Ageratum conyzoides has been used as purgative, febrifuge, anti-ulcer and wound dressing. To date there is no safety information about long term use of Ageratum conyzoides which contains pyrrolizidine alkaloids, a class of hepatotoxic and carcinogenic phytochemicals. This study aims to evaluate the 90 days subchronic toxicity and in vitro toxicity of Ageratum conyzoides.

Materials and methods

Three groups of 8 rats (4 males and 4 females) received distilled water (control), 500 and 1000 mg/kg of the extract daily for 90 consecutive days by oral gavage. The animals were observed daily for abnormal clinical signs and death. Body weight, relative organ weight, haematological and biochemical parameters of blood as well as heart, kidney, liver and spleen tissues histology were evaluated.

Results

After 90 days administration, Ageratum conyzoides increased significantly (p<0.05) the relative weight of the liver, the spleen and kidney as compared to control group. Ageratum conyzoides increased also significantly (p<0.05) ALP, ALT, AST and blood glucose. Furthermore, an increase in the number of platelets associated with a normocytic and normochromic anaemia was observed. The cytotoxicity, determined by the MTT test and neutral red assay, has shown that the cytotoxicity of hydroalcoholic extract of Ageratum conyzoides and its total alkaloids was very close.

Conclusions

Our results have shown that Ageratum conyzoides at 500 and 1000 mg/kg can induce liver, kidney and haematological disorders. These toxics effects can be attributed to its total alkaloids especially to pyrrolizidine alkaloids which are present in this plant.     

Evidence of the anti-Helicobacter pylori, gastroprotective and anti-inflammatory activities of Cuphea aequipetala infusion

Ethnopharmacological relevance

Cuphea aequipetala (Lythraceae) is a medicinal plant highly appreciated in Mexico to treat stomach ailments such as pain and burning sensation, stomach infections, ulcers, diarrhea, dysentery, and different types of tumors and bruises. In this work, the infusion of aerial parts of this plant (CAI) was investigated for its polypharmacological potential.

Materials and methods

In vitro anti-Helicobacter pylori activity was assessed by broth dilution method. Pharmacological studies included acute toxicity in mice using Lorke´s model, anti-inflammatory activity by xylene and TPA induced ear edema assay, as well as gastroprotection with ethanol-induced gastric ulcer model. DPPH and ABTS assays were used to determine antioxidant capacity. Polyphenols and flavonoid contents were determined by Folin–Ciocalteu method and AlCl₃ reaction, respectively.

Results

CAI showed good anti-Helicobacter pylori activity with a MIC of 125 μg/mL. The infusion was not toxic according to Lorke's model with a LD₅₀ greater than 5 g/kg. CAI exhibited low anti-edematogenic action in the models assayed. Oral administration of 300 mg/kg CAI significantly reduced gastric lesions by 87.9%. The effect was reversed only by indomethacin and N-ethylmaleimide demonstrating the role of endogenous prostaglandins and sulfhydryl compounds in gastroprotection. Total phenolic and flavonoid contents of CAI were 109.9 mg GAE/g DW and 28.1 mg QE/g DW, respectively, and the infusion exhibited a good antioxidant activity that is thought to play a role in its biological activity. The analysis of a preliminary fractionation of the infusion indicates that the complete extract conserves all its pharmacological activities in contrast to fractionated extracts.

Conclusions

Cuphea aequipetala is a promising native herb in an integral therapy for the treatment of bacterial or non-bacterial gastric ulcer because it possesses some anti-inflammatory properties, as well as exhibits good gastroprotective and antibacterial effects. It represents an important source for the isolation of anti-Helicobacter pylori compounds. This work provides ethnopharmacological evidence that supports the traditional use of this species.     

Acute and subchronic oral toxicity of Coriolus versicolor standardized water extract in Sprague-Dawley rats

Ethnopharmacological relevance

Coriolus versicolor, which is known as Yun Zhi, is one of the commonly used Chinese medicinal herbs. Recent studies have demonstrated its antitumor activities on cancer cells which led to its widespread use in cancer patient. However, little toxicological information is available regarding its safety. The present study evaluated the potential toxicity of Coriolus versicolor standardized water extract after acute and subchronic administration in rats.

Materials and methods

In acute toxicity study, Coriolus versicolor water extract was administered by oral gavage to Sprague-Dawley (SD) rats (6 males, 6 females) at single doses of varying concentrations 1250, 2500 and 5000 mg/kg. In subchronic toxicity study, the extract was administered orally at doses of 1250, 2500 and 5000 mg/kg/day for 28 days to male and female SD rats respectively. General behavior, adverse effects and mortality were determined throughout the experimental period. Haematological and biochemical parameters, relative organ weights and histopathological were evaluated at the end of the experiment.

Results

There were no mortality and signs of toxicity in acute and subchronic toxicity studies. In the single dose acute toxicity and repeated dose 28-day subchronic toxicity studies, there were no significant difference in body weight, relative organ weight, haematological parameters, clinical chemistry, gross pathology and histopathology between treatment and control groups.

Conclusions

Coriolus versicolor water extract did not cause remarkable adverse effect in SD rats. The oral lethal dose of Coriolus versicolor water extract is more than 5000 mg/kg and no-observed-adverse-effect level (NOAEL) of the extract for both male and female rats is 5000 mg/kg per day for 28 days.     

Safety assessment of aqueous extract from leaf Smallanthus sonchifolius and its main active lactone, enhydrin

Ethnopharmacological Relevance

Leaves of Smallanthus sonchifolius (Poepp. & Endl.) H. Robinson (yacon) have been used since pre-Columbian times in the Andean region to prepare medicinal herbal tea with beneficial health properties. However, there are still disagreements about the safe use. This work was carried out to evaluate the toxicity profile of both, 10% decoction of yacon leaves and their major active lactone, enhydrin.

Materials and methods

In vitro cytotoxicity assays were performed with Hep-G2, COS1, CHO-K1 and Vero cell lines using a test of metabolic competence based upon assessment of mitochondrial performance. In vivo toxicity study was performed in adult Wistar rats. In the acute oral toxicity each group of rats was orally given a single dose of 10% decoction or enhydrin. General condition, behavior and mortality were recorded for up to 14 days post treatment. In subchronic toxicity studies, both products were given orally for 90 days to rats. Body weight and food intakes were observed weekly. Hematological, clinical chemistry parameters and organ weight were determined in all animals at the end of the experimental period.

Results

Cell viability decreased in a concentration dependent fashion when cells were incubated with 2–200 μg of 10% decoction and 0.015–7.5 μg of enhydrin. In acute study in rats, there were no deaths or signs of toxicity observed after oral administration of single doses of 10% decoction or enhydrin at any dose level up to the highest dose tested (14.0 g/kg and 0.32 g/kg, respectively). In subchronic studies in rats, both products administered orally for 90 days at daily doses of 0.07, 0.14 and 0.28 g 10% decoction/kg and 0.4, 0.8 and 8.0 mg enhydrin/kg, did not caused haematological, biochemical and histological alterations.

Conclusions

The results presented in this paper lead us to the conclusion that the use of 10% decoction and enhydrin is safe in rat at doses in which it is demonstrated the hypoglycaemic effect.     

In vivo antimalarial activities of Enantia polycarpa stem bark against Plasmodium berghei berghei in mice

Ethnopharmacological relevance

Enantia polycarpa (PC) Engl. Et Diels (Annonaceae) is used in traditional medicine as an antimalarial remedy in Southern Nigeria.

Aim of the study

The antimalarial activities of ethanolic stem bark extracts of Enantia polycarpa was studied in vivo, in mice infected with Plasmodium berghei berghei.

Materials and methods

The ethanolic stem bark extract of Enantia polycarpa was administered at doses ranging from 200 to 600 mg/kg/day to Plasmodium berghei infected mice in both early and established models of antiplasmodial studies.

Results

The extract of Enantia polycarpa exhibited promising antimalarial activity against both early and established infections. At a dose of 600 mg/kg the extract achieved a 75.8% and 72% chemosuppression of parasitaemia in the study of acute and established infections, respectively. The extract also prolonged mean survival time of Plasmodium berghei infected mice during the study of established infection. The mean survival time of mice administered Enantia polycarpa extract at 600 mg/kg/day (27 days) was significantly longer than infected/untreated control (12 days). For the acute toxicity study the extract had an intraperitoneal LD₅₀ of 186 mg/kg but caused no mortality when administered orally at doses as high as 2,000 and 4,000 mg/kg.

Conclusions

Collectively, the results indicate that Enantia polycarpa is safe when administered orally and possesses promising antimalarial activity, thus supporting its use in traditional medicine for the treatment of malaria.     

Acute and subacute toxicity of Schinus terebinthifolius bark extract

Ethnopharmacological relevance

Schinus terebinthifolius Raddi (Anacardiaceae) has long been used in traditional Brazilian medicine, especially to treat inflammatory and haemostatic diseases.

Aim of the study

The objective of this study was to evaluate the acute and subacute toxicity (45 days) of Schinus terebinthifolius via the oral route in Wistar rats of both sexes.

Materials and methods

For the acute toxicity test, the dried extract of Schinus terebinthifolius bark was administered in doses from 0.625 to 5.0 g/kg (n = 5/group/sex) and in the subacute toxicity test the following doses were used: 0.25, 0.625 and 1.5625 g/kg/day (n = 13/group/sex), for 45 consecutive days.

Results

In the acute toxicity test, Schinus terebinthifolius did not produce any toxic signs or deaths. The subacute treatment with Schinus terebinthifolius did not alter either the body weight gain or the food and water consumption. The hematological and biochemical analysis did not show significant differences in any of the parameters examined in female or male groups, except in two male groups, in which the treatment with Schinus terebinthifolius (0.25 and 0.625 g/kg) induced an increase of mean corpuscular volume values (2.9 and 2.6%, respectively). These variations are within the physiological limits described for the specie and does not have clinical relevance.

Conclusion

The acute and subacute administration of the dried extract of Schinus terebinthifolius bark did not produced toxic effects in Wistar rats.     

Toxicity and potential anti-trypanosomal activity of ethanolic extract of Azadirachta indica (Maliacea) stem bark: An in vivo and in vitro approach using Trypanosoma brucei

Aim of the study

To determine the toxicity and anti-trypanosomal activity of the ethanolic extract of Azadirachta indica (Maliacea) stem bark, through in vivo and in vitro approach using Trypanosoma brucei brucei.

Materials and methods

Graded concentrations (100, 200, 400, 800, 1600 and 3200 mg/kg) of the crude stem bark ethanolic extract of Azadirachta indica, Hochst ex. A. Dc. (Maliacea) was tested for acute toxicity in 35 out bred Swiss (Wister) adult albino rats of both sexes. Secondly, the in vitro activity in test tubes and in vivo activity of the extract in 30 out bred Swiss (Wister) adult albino rats against Trypanosoma brucei brucei strain NITR/14 (Federe) was evaluated in a graded dose manner.

Results

The calculated intra-peritoneal LD₅₀ of the extract was 870 mg/kg and produced toxicity at high doses (>800 mg/kg). Graded concentrations of the ethanolic extract produced remarkable in vitro activity against Trypanosoma brucei brucei within seconds of inoculation. It also suppressed the establishment of parasitaemia at 100 mg/kg when administered simultaneously with infection in vivo. Similarly, at 200 and 400 mg/kg, the extract administered at the onset of parasitaemia for 4 consecutive days reduced parasitaemia, modulated declined packed volume (PCV) changes by day 48 post-infection in vivo.

Conclusion

The results confirm that the folkloric medicinal application of the extract of Azadirachta indica (Maliacea) has a pharmacological basis. Further investigation is however, needed to optimize the effectiveness of the extract.