An LC/MS/MS method for simultaneous quantitation of two homoisoflavones: Protosappanin B and brazilin with hypoglycemic activity in rat plasma and its application to a comparative pharmacokinetic study in normal and streptozotocin-treated rats

Ethnopharmacological relevance

The heartwood of Caesalpinia sappan L. (Leguminosae), a widely used Chinese medicine in folk, has been used for the treatment of traumatic injury, stasis pain, amenorrhea, dysmenorrheal, as well as stabbing pain in the chest, abdomen and so on. Protosappanin B and brazilin, as the major bioactive homoisoflavones of Sappan Lignum, are used as the marker components for the quality control of the herb in China Pharmacopoeia.

Aim of the study

To establish a sensitive LC/MS/MS method for investigating the pharmacokinetic properties of protosappanin B and brazilin in rats after oral administration of Sappan Lignum extract, and compare their pharmacokinetics difference between normal and streptozotocin-treated rats.

Material and methods

A rapid, selective and sensitive LC/MS/MS method was developed and validated for the simultaneous quantification of protosappanin B and brazilin in rat plasma. Normal and streptozotocin-treated rats were orally administered with the Sappan Lignum extract at the same dose of 2.83 g extract/kg body weight (equivalent to 35.56 mg/kg of protosappanin B and 52.25 mg/kg of brazilin), respectively.

Results

After oral administration of Sappan Lignum extract, a remarkable increase (p<0.05) in the value of AUC_0–24 h, AUC_0–∞, C_max and T_1/2 associated with protosappanin B and brazilin was observed in the streptozotocin-treated group. Compared with the normal rats, elimination of both compounds in the streptozotocin-treated rats was slower.

Conclusion

The established method was successfully applied to compare the pharmacokinetic behaviors of protosappanin B and brazilin in rat plasma after oral administration of Sappan Lignum extract between normal and streptozotocin-treated groups; the results might suggest the accumulation of both compounds in diabetic pathologic states and the adverse reaction should be considered when it was used.     

Neuroprotective effect of calycosin on cerebral ischemia and reperfusion injury in rats

Ethnopharmacological relevance

Radix Astragali has been commonly used as traditional herbal medicine in China for reinforcing vital energy, strengthening superficial resistance and promoting the discharge of pus and the growth of new tissue.

Aim of the study

The present study was to investigate the neuroprotective effect of calycosin isolated from the roots of Radix Astragali on cerebral ischemic/reperfusion injury.

Materials and methods

After 24 h of reperfusion following ischemia for 2 h induced by middle cerebral artery occlusion (MCAO), Sprague-Dawley rats were intragastrically administered different doses of calycosin (7.5, 15, 30 mg/kg, respectively). Neurological deficit, infarct volume, histopathology changes and some oxidative stress markers were evaluated after 24 h of reperfusion.

Results

Treatment with calycosin significantly ameliorated neurologic deficit and infarct volume after cerebral ischemia reperfusion. Calycosin also reduced the content of malondialdehyde (MDA), protein carbonyl and reactive oxygen species (ROS), and up-regulated the activities of superoxide dismutase (SOD), catalase and glutathione peroxidase (GSH-Px) in a dose-dependent manner. Moreover, calycosin can also inhibit the expression of 4-Hydroxy-2-nonenal (4-HNE).

Conclusion

These results suggest that calycosin has a neuroprotective effect against cerebral ischemia/reperfusion injury. The mechanism might be attributed to its antioxidant effects.     

Pharmacokinetics and tissue distribution of resveratrol,emodin and their metabolites after intake of Polygonum cuspidatum in rats

Ethnopharmacological relevance

The rhizome of Polygonum cuspidatum SIEB. et ZUCC. (Polygonaceae, PC), a widely used Chinese medicine, is commonly prescribed for the treatments of amenorrhea, arthralgia, jaundice, abscess, scald and bruises.

Aim of the study

PC contains various polyphenols including stilbenes, anthraquinones and flavonoids. This study investigated the pharmacokinetics and tissue distribution of emodin and resveratrol in PC.

Material and methods

Male Sprague-Dawley rats were orally administered PC (2 and 4 g/kg) and blood samples were withdrawn at the designed time points via cardiopuncture. Moreover, after 7-dose administrations of PC (4 g/kg), brain, liver, lung, kidney and heart were collected. The concentrations of resveratrol and emodin in the plasma and tissues were assayed by HPLC before and after hydrolysis with β-glucuronidase and sulfatase.

Results

The glucuronides/sulfates of emodin and resveratrol were exclusively present in the plasma. In liver, kidney, lung and heart, the glucuronides/sulfates of resveratrol were the major forms. For emodin, its glucuronides/sulfates were the major forms in kidney and lung, whereas considerable concentration of emodin free form was found in liver. Neither free forms nor conjugated metabolites of resveratrol and emodin were detected in brain.

Conclusion

The sulfates/glucuronides of resveratrol and emodin were the major forms in circulation and most assayed organs after oral intake of PC. However, the free form of emodin was predominant in liver.     

Three Chinese herbal medicines promote neuroproliferation in vitro, and reverse the effects of chronic mild stress on behavior,the HPA axis,and proliferation of hippocampal precursor cell in vivo

Ethnopharmacological relevance

The present study investigated whether Chinese herbal medicines (CHMs) could reverse the effects of chronic mild stress (CMS) in a depression-like mouse model.

Materials and methods

The effects of three Chinese herbals, Rhizome Chuanxiong, Radix Scutellaria and Radix Phellodendri on promoting neuroproliferation were evaluated in vitro first and followed by in vivo study of mice which were received by an experimental setting of CMS for 14 days. The effects of the three CHMs on depression were evaluated using a behavioral test, named a forced swimming test (FST). The possible anti-depressive mechanisms of these three CHMs, including the modulation of HPA axis and promoting the hippocampal precursor cell proliferation, were evaluated by measuring plasma corticosterone levels and BrdU incorporation.

Results

The in vitro results of MTS assay showed that Rhizome Chuanxiong, Radix Scutellaria and Radix Phellodendri could promote the proliferation of neural stem cells (NSCs) in a concentration-dependent manner. The oral administration of these three CHMs for 14 days reversed not only the elevation of plasma corticosterone levels and body weight loss, but also the decreasing of hippocampal precursor cell proliferation and abnormal behavior in the CMS induced depression-like mouse model.

Conclusion

These results indicated that Rhizome Chuanxiong, Radix Scutellaria and Radix Phellodendri have the potential to ameliorate depression. The possible mechanisms were the inhibition of HPA axis hyperactivity and the increasing of hippocampal precursor cell proliferation. These findings supported the multicomponent and multitargeted approach of Chinese herbal medicine.     

Antihyperglycaemic activity of aqueous leaf extract of Combretum micranthum (Combretaceae) in normal and alloxan-induced diabetic rats

Aim of the study

To investigate antidiabetic effect of the leaves of Combretum micranthum G. Don, a medicinal plant used for treating diabetes in Northwestern Nigeria.

Materials and methods

Three doses (100 mg/kg, 200 mg/kg and 400 mg/kg) of the aqueous leaf extract of Combretum micranthum were administered to normal glucose loaded, subdiabetic and diabetic rats.

Results

Of the doses tested, 100 mg/kg of the extract was the most effective. It produces a significant hypoglycaemic and antidiabetic activity comparable to the effect of standard drug (0.6 mg/kg glibenclemide).

Conclusion

This study demonstrated the potential antidiabetic property of aqueous leaf extract of Combretum micranthum thus justifying its traditional usage.     

Pharmacokinetic study on costunolide and dehydrocostuslactone after oral administration of traditional medicine Aucklandia lappa Decne. by LC/MS/MS

Ethnopharmacological relevance

Radix Aucklandiae (RA), a well known traditional Chinese medicine, is widely used for treating various problems in digestive system. A selective and sensitive high-performance liquid chromatography coupled with mass spectrometry method was first developed and validated for simultaneous quantification of costunolide and dehydrocostuslactone in rat plasma with diazepam as internal standard after oral administration of RA extraction.

Materials and methods

Plasma samples were extracted via solid-phase extraction and detected by multiple-reaction monitoring mode under positive electrospray. Chromatographic separation was accomplished on an Agilent C₁₈ column (2.1 mm×150 mm, 5 µm), with 0.1% formic acid and acetonitrile (1:1) as the mobile phase at a flow rate of 0.5 mL/min.

Results

The quantification was performed using the transitions of m/z 233/187 for costunolide, m/z 231/185 for dehydrocostuslactone and m/z 285/193 for diazepam, respectively. Calibration curves were linear over the concentration range of 0.7–769.7 ng/mL for costunolide and 0.9–956.0 ng/mL for dehydrocostuslactone. The intra-day and inter-day precisions (RSD%) for two compounds was less than 8.76% and 9.70% and the accuracy (RE%) range from 6.14% to 5.35%. The time to reach the maximum plasma concentration (T_max) was 10.46 h for costunolide, 12.39 h dehydrocostuslactone. The elimination half-time (t_1/2) of costunolide and dehydrocostuslactone was 5.54±0.81 and 4.32±0.71 (h). The AUC of costunolide and dehydrocostuslactone was 308.83 and 7884.51 respectively (ng h/mL).

Conclusions

It was the first report for the study of pharmacokinetic profile of costunolide and dehydrocostuslactone in rat plasma after oral administration of RA extract. These results provided a meaningful basis for better understanding the absorption of traditional medicine, RA, and provide useful scientific data for clinical application.     

In vivo disposition and metabolism of madecassoside,a major bioactive constituent in Centella asiatica (L.) Urb.

Ethnopharmacological relevance

Centella asiatica (L.) Urb. herb is frequently used in traditional Chinese medicine for many indications, such as traumatic injuries, keloid and scar. Madecassoside is the main active ingredient of this herb drug with higher content than other triterpenoid constituents. Understandings of pharmacokinetic profiles of madecassoside should be beneficial for its development as a therapeutic agent.

Materials and methods

Sprague-Dawley rats were intravenously and orally administered madecassoside (100 mg/kg), respectively. Plasma, heart, liver, spleen, lung, kidney, brain, bile, urine and feces were collected at the designed time points. Madecassoside concentrations in biological samples were determined by a sensitive and well-validated liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) method. A liquid chromatography coupled with time-of-flight mass spectrometry (LC-TOF-MS) method was established to identify its major metabolites in feces. To further pursue the disposition characteristics of madecassoside in rats, the involvement of the hepatobiliary efflux transporters in biliary elimination were studied by combination with digoxin (P-glycoprotein inhibitor) or probenecid (multidrug resistance-associated protein inhibitor). A linked-rat model was also used to assess the role of enterohepatic circulation in the pharmacokinetics of madecassoside.

Results

After a single oral dosing, madecassoside was widely distributed in heart, liver, spleen, lung and kidney of rats, and the levels of madecassoside in liver and kidney were relatively higher than other organs. The excretions of madecassoside in bile, urine and feces were 7.16% (0–12 h), 0.25% (0–72 h) and 24.68% (0–72 h), respectively. The findings suggested that madecassoside might excrete mainly by metabolites. The combination with either digoxin or probenecid significantly attenuated the excretion of madecassoside as parent from bile, indicating that P-glycoprotein and multidrug resistance-associated protein might contribute to the hepatobiliary elimination of madecassoside. The presence of enterohepatic circulation, as implied by double-humped profiles in plasma and tissue concentration-time curves, was confirmed by a linked-rat model. Furthermore, three metabolities of madecassoside were indentified in rat feces and the possible metabolic pathways were proposed.

Conclusions

These findings provide valuable information regarding in vivo process of madecassoside, and help us to recognize the efficacy and safety of madecassoside itself, the relevant herbs or herbal preparations.     

Antidepressant effects of Radix et Caulis Acanthopanacis Santicosi extracts on rat models with depression in terms of immobile behavior

Objective

To investigate the antidepressant effects of the Radix et Caulis Acanthopanacis Santicosi extracts in response to the tail suspension test (TST) and forced swim test (FST) in rats.

Methods

Male Wistar rats (9-week-old) were orally administered the Radix et Caulis Acanthopanacis Santicosi extracts (30, 100 and 300 mg/kg) daily for 14 consecutive days. Serum corticosterone level was measured by using commercially available kit. The c-fos immunoreactivity was analyzed using immunohistochemistry methods.

Results

The effect of Radix et Caulis Acanthopanacis Santicosi extracts on the immobility time was significantly decreased at high-dose (300 mg/kg) treated group in the TST and FST (47.7% and 69.6%, P<0.05), and decreased the hypothalamic-pituitary-adrenal (HPA) axis response to the stress, as indicated by an attenuated corticosterone response and decreased c-fos immunoreactivity in the hippocampal and hypothalamic paraventricular nucleus region.

Conclusion

Radix et Caulis Acanthopanacis Santicosi extracts have beneficial effects on depression behaviors and restore both altered c-fos expression and HPA activity. Thus, Radix et Caulis Acanthopanacis Santicosi extracts may be a novel agent for the treatment of stress-related disorders.     

UPLC/MS/MS method for quantification and cytotoxic activity of sesquiterpene lactones isolated from Saussurea lappa

Ethnopharmacological relevance

Saussurea lappa (Asteraceae) roots have been reputed for the usage in traditional medicinal systems of India, China and Japan for the treatment of various kinds of disorders such as anti-ulcer, anti-convulsant, anti-cancer, hepatoprotective, anti-arthritic and anti-viral activities.

Materials and methods

Compounds were isolated using a column chromatographic technique. The root extract, fractions and isolated compounds were tested for cytotoxicity against A549 (human lung carcinoma) and C-6 (rat glioma) cells using the Sulphorhodamine B assay. Chromatographic separations of active sesquiterpene lactones were accomplished on BEH-HSS-T3 column at 25 °C.

Results

Phytochemical investigation of Saussurea lappa root extract resulted in the isolation of isoalantolactone (1), β-cyclocostunolide (2) α-cyclocostunolide (3), 4-hydroxy-3,5-dimethoxycinnamyl-9-O-β-d-glucopyranoside (4), sucrose (5), and alantolactone (6). Their structures were determined by spectroscopic means. Ethanolic extract, chloroform fraction, compounds 1, 2, 3 and 6 possessed significant activity against both tested cells. The quantification was performed using the transitions of m/z 233/105 for isoalantolactone and m/z 233/105 for alantolactone respectively. Costunolide and dehydrocostus lactone were also characterised by comparison of MS/MS fragmentation pattern.

Conclusions

This is the first study on simultaneous quantification of isoalantolactone and alantolactone by the UPLC/MS/MS method in Saussurea lappa. Our study against A549 and C-6 cells showed higher cytotoxicity. It is suggested that roots of Saussurea lappa might be a potential source of anticancer compounds.     

Analysis of the restorative effect of Bu-zhong-yi-qi-tang in the spleen-qi deficiency rat model using H-NMR-based metabonomics

Ethnopharmacological relevance

Bu-zhong-yi-qi-tang (BT) is a classical formula for the treatment of spleen-qi descending, visceroptosis with hyposplenic qi, uterine prolapse, and rectal prolapse due to chronic diarrhea in traditional Chinese medicine (TCM) and has been identified as an effective drug for the treatment of TCM spleen-qi deficiency in clinical practice. The present study aimed to investigate the restorative effect and the potential mechanisms of Bu-zhong-yi-qi-tang in a rat spleen-qi deficiency model using ¹H-NMR-based metabonomics.

Materials and methods

The rat spleen-qi deficiency model was established as follows: oral administration of Radix Rhei extract (equivalent to 10 g/kg body weight of the crude drug), loaded swimming, and starvation for 24 h. Each of these treatments was administered consecutively every three days. Sixty male SD rats were randomly divided into five groups, and three of the groups received a different oral dose of the aqueous extract of Bu-zhong-yi-qi-tang during the last seven days of the three-week experimental period. The body weight and motor behavior of the rats were measured and recorded once a week. The endogenous metabolites in the plasma were analyzed using NMR in conjunction with multivariate and statistical techniques. In addition, the liver and spleen were removed and weighed.

Results

All of the rats in the spleen-qi deficiency group presented pasty loose stools, inactiveness, grouping, a decrease in swimming endurance, and lackluster, loose, and disorderly behavior in addition to a significant decrease in body weight, spleen weight, and liver weight. In contrast, the abovementioned demonstrations were reversed to a certain extent in the rats treated with Bu-zhong-yi-qi-tang compared with the model group (p<0.05, p<0.01). A significant separation was determined between the control and model groups in the PCA score plot, which indicates that the spleen-qi deficiency model was successfully duplicated. The changes in the levels of endogenous metabolites in the plasma included lower levels of valine, leucine, and O-acetyl-glycoprotein and a higher concentration of lactate in the spleen-qi deficiency group compared with the control group. Treatment with Bu-zhong-yi-qi-tang at least partially returned the levels of these metabolites to the normal levels.

Conclusions

The restorative effects of Bu-zhong-yi-qi-tang in rats with spleen-qi deficiency were confirmed, and four endogenous metabolites were identified as potential biomarkers of the symptoms of spleen-qi deficiency and most likely play roles in the changes observed in certain metabolic pathways, such as the energy, protein, and glycolytic metabolisms.     

Identification and pharmacokinetics of multiple constituents in rat plasma after oral administration of Yinchenzhufu decoction

Ethnopharmacological relevance

Yinchenzhufu decoction (YCZFD) is a classical Chinese herbal formula and has been used to treat severe jaundice in chronic liver injuries since the Qing Dynasty (18th century CE). To identify the components absorbed into the blood in YCZFD and explore their pharmacokinetic profile for understanding the effective ingredients of YCZFD.

Materials and methods

After rats were given YCZFD by intragastric administration, the plasma was processed by precipitation of protein. The compounds in YCZFD extract and the plasma were identified by using high-resolution mass spectrometry with a database-directed strategy. The pharmacokinetics of multiple compounds from YCZFD in rat plasma was studied by using the established UPLC-MS/MS method.

Results

Forty compounds in YCZFD extract and 21 prototype compounds with 11 metabolites in rat plasma were detected after oral administration. The pharmacokinetic parameters of glycyrrhizic acid, glycyrrhetic acid, cinnamic acid, ononin, atractylenolide III, and liquiritin from YCZFD were obtained in rats.

Conclusions

The identified constituents and the pharmacokinetic features of YCZFD are helpful for understanding the material bases of its therapeutic effects.     

Effects of drying process of Yuanzhi (Radix Palygalae) on its bioactive ingredients

Objective

To study the effects of the drying processing in terms operational parameters on the bioactive constituents of six Yuanzhi (Radix Palygalae) samples across China.

Methods

Six Yuanzhi (Radix Palygalae) samples were investigated using thermogravimetry analysis. The heating courses were set in two ways: the temperature-programmed process from room temperature to 150°C, and the constant-temperature course at 50°C, 70°C and 90°C.

Results

The peak temperature of six Yuanzhi (Radix Palygalae) samples ranged from 78°C to 88°C. The mass loss rate of Yuanzhi (Radix Palygalae) alcohol-soluble extract was significantly increased when heated at 90°C. Four types of bioactive ingredients were detected in volatile oils of Yuanzhi (Radix Palygalae) sample from Shanxi province by Gas Chromatography-mass spectrometry analysis. Fourier Transform Infrared Spectroscopy results showed that the drying temperature exerted a great influence on types and amount of ingredients of Yuanzhi (Radix Palygalae). The kinetic study showed that the constant-temperature drying process of Yuanzhi Radix Palygalae) samples could be well described by the Page Model, especially for the drying process at 50°C, in which R² and SD values were more than 0.98 and less than 0.04, respectively. The drying constant k of three Yuanzhi (Radix Palygalae) samples from Shanxi, Gansu and Shaanxi provinces in China was corresponding to the Arrhenius equation, and their activation energies were 28.07, 25.38 and 21.48 kJ/mol, respectively.

Conclusion

The drying process of Yuanzhi (Radix Palygalae) was very important for bioactive ingredients improvement in Yuanzhi (Radix Palygalae). Temperature was a thermodynamic property significantly affecting the process.     

Antihyperglycemic and hypolipidemic activity of aqueous extract of Cassia auriculata L. leaves in experimental diabetes

Ethnopharmacological relevance

Cassia auriculata L. (Caesalpiniaceae) is widely used from ancient period to treat diabetes mellitus. The leaves of Cassia auriculata are having potential in the development of drug for diabetes due to its antihyperglycemic and lipid-lowering activity.

Aim of the study

The present study was to evaluate antihyperglycemic and hypolipidemic activity of aqueous extract of Cassia auriculata leaves (CLEt) in streptozotocin (STZ)-induced mild diabetic (MD) and severe diabetic (SD) rats.

Materials and methods

Male Albino rats were rendered diabetic by STZ (45 mg/kg, intraperitoneally). CLEt was orally administered to MD and SD rats at 100, 200 and 400 mg/kg doses for 1 day to determine antihyperglycemic activity. The 400 mg/kg dose was administered daily for 3 weeks to assess glycemic control and hypolipidemic effect.

Results

CLEt showed dose dependant fall in fasting blood glucose (FBG). After 5 h of extract administration at 400 mg/kg dose, FBG was reduced by 13.9% and 17.4% in MD and SD rats respectively. After 3 weeks treatment, CLEt produced significant reduction in FBG and glycosylated haemoglobin (GHb) in both MD and SD rats. Serum lipid levels were reversed towards normal in extract fed MD and SD rats.

Conclusions

The results demonstrate that CLEt possesses potent antihyperglycemic and hypolipidemic activity in both MD and SD rats.     

Toxicity evaluation of standardized extract of Gynostemma pentaphyllum Makino

Ethnopharmacological relevance

To evaluate the safety of standardized extract of Gynostemma pentaphyllum in rats.

Materials and methods

The water extract of Gynostemma pentaphyllum was prepared and standardized, the dry powder yielded 6% gypenosides. In the acute oral toxicity test, the single oral dose of 5000 mg/kg of Gynostemma pentaphyllum extract was given to female Sprague–Dawley rats. In subchronic toxicity test, the oral dose of 1000 mg/kg/day of the extract was given to rats in treatment and satellite groups for 90 days. Satellite groups of both sexes were kept for additional 28 days after 90-day treatment. Control rats received distilled water.

Results

Standardized extract of Gynostemma pentaphyllum did not cause death or any toxic signs in rats. The daily administration of the extract for 90 days did not produce lethal or harmful effects. Although certain hematological and blood chemistry values (i.e., neutrophil, monocyte, glucose, and serum alkaline phosphatase levels) were found to be statistically different from the control group, however; these values were within the ranges of normal rats.

Conclusion

Standardized extract of Gynostemma pentaphyllum did not produce mortality or any abnormality in rats.     

LC-MS/MS determination and pharmacokinetic study of four lignan components in rat plasma after oral administration of Acanthopanax sessiliflorus extract

Ethnopharmacological relevance

Acanthopanax sessiliflorus (Rupr. et Maxim.) Seem. is a shrub mainly present in China, Japan and Korea, the root bark of which is considered as one of the sources of Wujiapi and widely used for its various pharmacological effects.

Aim of the study

A selective and sensitive UPLC–MS/MS method was developed and validated for the determination and pharmacokinetic study of asarinin, sesamin, helioxanthin and savinin in rat plasma.

Materials and methods

Sample preparation involved a liquid–liquid extraction of the analytes with methyl tert-butyl ether (MTBE). LC separation was achieved on a UPLC C₁₈ column at 30 °C with a mobile phase consisting of methanol–2 mM ammonium acetate (68:32, v/v). The detection was accomplished by multiple-reaction monitoring (MRM) scanning with electrospray ionization (ESI) source operating in the positive ionization mode. The optimized mass transition ion-pairs (m/z) monitored for asarinin, sesamin, helioxanthin, savinin and IS were 372.2/233.0, 372.2/233.0, 349.1/319.0, 352.9/334.9 and 180.0/109.7, respectively.

Results

The current LC–MS/MS assay was validated for linearity, intra-day and inter-day precisions, accuracy, extraction recovery and stability and was suitable for pharmacokinetic studies of the four lignans after oral administration of Acanthopanax sessiliflorus extract. The time to reach the maximum plasma concentration (T_max) was 2.50 ± 0.15 h for asarinin, 1.94 ± 0.28 for sesamin, 2.22 ± 0.48 h for helioxanthin and 2.83 ± 0.29 h for savinin. The elimination half-time (t_1/2) of asarinin, sesamin, helioxanthin and savinin was 6.08 ± 1.10, 11.69 ± 0.50, 7.16 ± 0.52 and 6.26 ± 0.57 h, respectively.

Conclusion

This paper described a simple, sensitive and validated UPLC–MS/MS method for simultaneous determination of four lignans in rat plasma after oral administration of Acanthopanax sessiliflorus extract, and investigated on their pharmacokinetic studies as well.     

Beneficial effect of the administration of Hemidesmus indicus against bromobenzene induced oxidative stress in rat liver mitochondria

Aim of the study

To study the beneficial effect of the prior administration of an aqueous extract of Hemidesmus indicus against bromobenzene induced oxidative damage in rat liver mitochondria.

Materials and methods

Oxidative stress was induced in rats with bromobenzene (10 mmol/kg body wt.). The rate of respiration, P/O ratios, lipid peroxides, protein carbonyls and sulphydryls were studied.

Results

When the rats were administered with bromobenzene, the rate of respiration was decreased significantly and the P/O ratio was completely abolished. There was a significant increase on the levels of lipid peroxide and protein carbonyl and a significant decrease on total sulphydryl groups when compared with control. Administration of rats with an aqueous extract (100 mg/kg) prior to bromobenzene administration showed significant beneficial effects like, stimulation in respiration, prevented the rise in lipid peroxides and protein carbonyls, increased the level of sulphydryl groups back to control level. Administration of vitamin E could not reverse as effectively as Hemidesmus indicus.

Conclusions

This study demonstrates a good protective effect of Hemidesmus indicus against the bromobenzene induced oxidative stress.     

Metabolites characterization of chamaechromone in vivo and in vitro by using ultra-performance liquid chromatography/Xevo G2 quadrupole time-of-flight tandem mass spectrometry

Ethnopharmacological relevance

Stellera chamaejasme L. (Thymelaeaceae) was a toxic perennial herb and widely used as pesticide and dermatological agents in China. Chamaechromone was a major component in the dried roots of Stellera chamaejasme with anti-HBV and insecticidal activity. Analysis of metabolic profile in vivo and in vitro plays a pivotal role to unravel how TCM works. And the metabolites of chamaechromone might influence the effects and toxicity of Stellera chamaejasme. Moreover, the metabolic routes of chamaechromone provide an important basis for toxicological safety evaluation. Until now, little is known about the metabolism of chamaechromone. The current study was designed to characterize the whole metabolic pathways of chamaechromone in vitro and in vivo.

Materials and methods

Twenty-four rats were randomly divided into four groups, including two oral administration groups (100 mg kg⁻¹), one intravenous injection group (5 mg kg⁻¹), and one control group. The metabolites in rat urine and feces and bile were identified by UPLC/Q-TOF MS analysis and β-glucuronidase hydrolysis. Moreover, the possible metabolic mechanism was further confirmed by Phase I and Phase II metabolism and catechol-O-methyltransferase methylation in rat liver S9 fraction and degradation in rat intestinal bacteria.

Results

A total of 24 metabolites from chamaechromone were detected and identified in vivo and in vitro, 20 of which were novel. And the major metabolic processes were hydroxylation, methylation, glucuronation, acetylation, dehydroxylation and degradation.

Conclusions

The present study revealed the whole metabolic pathways of chamaechromone in rat through both in vitro and in vivo experiments for the first time. And chamaechromone could undergo extensive phase I and phase II metabolism in rat. These findings would provide an important basis for the further study and clinical application of chamaechromone. In addition, the results of this work have showed the feasibility of the UPLC/Q-TOF-MS approach for rapid and reliable characterization of metabolites.