Hepatoprotective Formulation Research Articles

Combinatorial Herbal Extracts Alleviate Alcohol-Induced Hepatic Disorders.

Plant-derived compounds can be useful for the management of liver disease. Traditionally, hepatic disorders have been treated with herbal extracts. Although many herbal extracts in Eastern medicine have been shown to possess hepatoprotective activities, single-origin herbal extracts primarily demonstrate either antioxidant or anti-inflammatory activities. The current study investigated the effects of combinatorial herbal extracts on alcohol-induced hepatic disorders in an ethanol-fed mouse model. Sixteen herbal combinations were evaluated as hepatoprotective formulations; the active constituents in these herbal extracts were daidzin, peonidin-3-glucoside, hesperidin, glycyrrhizin, and phosphatidylcholine. RNA sequencing analysis showed that exposure to ethanol altered hepatic gene expression profiles (compared to those of the non-alcohol-fed group), resulting in 79 differentially expressed genes. A majority of the differentially expressed genes in alcohol-induced hepatic disorders were associated with dysfunction of the normal cellular homeostasis in the liver; however, these genes were repressed by treatment with herbal extracts. Moreover, following treatment with herbal extracts, there were neither acute inflammatory responses in the liver tissue nor abnormalities in the cholesterol profile. These results suggest that combinatorial herbal extracts may alleviate alcohol-induced hepatic disorders by modulating the inflammatory response and lipid metabolism in the liver.

RP-UHPLC and HPTLC Method Development and Validation for Analysis of Andrographolide from Herbal Hepatoprotective Formulation

This work aimed to create a speedy, precise, and selective HPTLC and RP-UHPLC method for analysing andrographolide in fi nished products and raw materials. Asian medicines are using Andrographis paniculata since long. It’s used to treat skin eruptions, boils, scabies, and chronic, unexplained fevers, all of which are caused by blood “abnormalities.” Liquid chromatographic methods were developed to separate andrographolide and its herbal dosage form. HPTLC chromatography employed a 10 by 10 cm aluminum plate coated with 0.2 mm of silica gel 60 F254 (E. Merck, Germany). Camag Linomat 5 applicator with 100 μL syringe was used to apply samples in 6 mm bands (Hamilton, Switzerland). 14 mm separated the two bands, and 150 nL sec-1 was applied. Mobile phase was dichloromethane, toluene, ethyl acetate, and formic acid (6:4:1:0.5). This chromatogram runs 80 mm. Camag TLC scanner measured density at 254 nm. Mean RF=0.69. The linear calibration curve covers 500–3000 ng/spot and has a 0.996 correlation coeffi cient. Limit of Detection: 31.5 ng; Limit of Quantitation: 95.48 ng. A validated RP-UHPLC method for quantifying andrographolide in extract and formulation has been established. UHPLC analysis was performed in isocratic mode, at room temperature, using acetonitrile: Water (0.2% acetic acid) (85:15, v/v) as mobile phase on a 250 mm 4.6 mm i.d., 5 μm Cosmosil C18 column. Detection was at 230 nm. Andrographolide has a 4.1 minute half-life. Between 10 and 60 g/mL, andrographolide was linear. The approach met or exceeded ICH’s linearity, precision, accuracy, and robustness

A Critical Appraisal of the Most Recent Investigations on the Hepatoprotective Action of Brazilian Plants.

Conventional treatments for liver diseases are often burdened by side effects caused by chemicals. For minimizing this problem, the search for medicines based on natural products has increased. The objective of this review was to collect data on the potential hepatoprotective activity of plants of the Brazilian native flora. Special attention was given to the modes of extraction, activity indicators, and identification of the active compounds. The databases were Science direct, Pubmed, and Google Academic. Inclusion criteria were: (a) plants native to Brazil; (b) studies carried out during the last 15 years; (c) high-quality research. A fair number of communications met these criteria. Various parts of plants can be used, e.g., fruit peels, seeds, stem barks, and leaves. An outstanding characteristic of the active extracts is that they were mostly obtained from plant parts with low commercial potential, i.e., by-products or bio-residues. The hepatoprotective activities are exerted by constituents such as flavonoids, phenolic acids, vitamin C, phytosterols, and fructose poly- and oligosaccharides. Several Brazilian plants present excellent perspectives for the obtainment of hepatoprotective formulations. Very important is the economical perspective for the rural producers which may eventually increase their revenue by selling increasingly valued raw materials which otherwise would be wasted.

QUALITY ASSESSMENT OF SOME MARKETED HEPATOPROTECTIVE POLYHERBAL FORMULATIONS

The present study was proposed for assessing the variation in quality of some hepatoprotective polyherbal products that are widely available in the market. Different brands of marketed hepatoprotective formulations were evaluated in the present investigation with respect to qualitative analysis by HPTLC, phytochemical evaluation and heavy-metal detection. Among all formulations, it was observed that formulation of Brand C showed maximum amount of phenolic content, tannin content, and flavonoid content. In HPTLC chromatograms of formulations of various brands, spots with RF values (0.46 ±0.3 and 0.86±0.3, respectively) were found, which confirmed the presence of andrographolide and phyllanthin in them. The increased risk of harmful side effects to the patients was indicated by the presence of heavy metals in some formulations above the permissible limits. The present study gives an insight into the fact that there is a requirement to prepare stricter quality control procedures and parameters for formulations for consumer safety.

Development and validation of RP HPLC method for simultaneous determination of picroside I, picroside II, phyllanthin and boeravinone B in a polyherbal hepatoprotective tablet formulation

Purpose: To develop and validate RP-HPLC method for simultaneous determination of four active ingredients: picroside I (PSI), picroside II (PSII), phyllanthin (PHY) and boeravinone-B (BVB) in a polyherbal hepatoprotective tablet formulation.Methods: The study was carried out using Waters X-Bridge, C18 (250 mm x 4.6 mm, 5 μm) column with mobile phase consisting of 5 mM ammonium acetate in 10 % methanol and acetonitrile, with gradient programme at dual wavelengths of 220 nm and 274 nm and flow speed of 1 mL.min-1. The procedure was validated with respect to specificity, linearity, precision, accuracy, system suitability, limit of detection (LOD) and of quantification (LOQ), and robustness in line with ICH specifications.Results: The method was linear within the concentration range of 25 to 200 %, and the values of correlation coefficients (R2) were > 0.999. Intra-day and inter-day RSDs of PAs and RTs were < 5.0 %, with recovery in the range of 100.0 - 106.0 %.Conclusion: The four active ingredients have with good resolution with regard to the method used. The method is rapid, simple, highly selective, sensitive and cost effective, which make it an efficient method for quality assurance.

Validated HPTLC Method to Estimate Silymarin in Marketed Herbal Formulation

Objectives: The objective of present research work is to develop simple, rapid and accurate high performance thin layer chromatographic method for quantitative estimation of silymarin in marketed hepatoprotective formulation. Methods: The separation was performed on silica gel 60 F254 precoated TLC aluminum plates and toluene: ethyl acetate: formic acid: methanol as mobile phase. Determination and quantitation were performed by densitometric scanning at 288 nm. Validation of method is carried out as per International Conference of Harmonization (ICH) guidelines. Results: Accuracy, precision and recovery were all within the required limits. Conclusion: In conclusion, the developed HPTLC method is precise and accurate for implementation in routine quality control. The selected marketed hepatoprotective formulation contains the sufficient amount of the active ingredient which is required for its therapeutic efficiency.

Ameliorative effect of enhanced Fischer ratio flaxseed protein hydrolysate in combination with antioxidant micronutrients on ethanol-induced hepatic damage in a rat model

Alcohol abuse causes severe metabolic abnormalities inducing hepatic damage and malnutrition. Since higher Fischer ratio proteins have therapeutic value in liver diseases, an investigation was undertaken to study the ameliorative effect of the enhanced Fischer ratio flaxseed protein hydrolysate (EFR-FPH) alone and in combination with antioxidant micronutrients on ethanol-induced hepatotoxicity in a rat model. The EFR-FPH was prepared by dual enzymatic hydrolysis and charcoal treatment of flaxseed protein. The ratio of the branched-chain:aromatic amino acids (Fischer ratio) was found to be 7·08. The EFR-FPH, characterised using LC-MS/MS, showed the abundance of free leucine and isoleucine compared with phenylalanine and tyrosine. The matrix-assisted laser desorption/ionisation-time of flight MS analysis revealed the larger peptides present in EFR-FPH with mass 2·3 kDa. The EFR-FPH improved the nutritional status, liver function and antioxidant defense in the ethanol hepatotoxicity-induced rat model. The hepatoprotective effect of EFR-FPH was significantly enhanced when combined with selenium or vitamin E. Ethanol-induced changes in the liver tissue were effectively suppressed in the groups receiving EFR-FPH. Flaxseed-based hepatoprotective dietary supplement was formulated incorporating an optimum level of EFR-FPH (10 %) based on sensory acceptability and was fortified with selenium and vitamin E. The hepatoprotective formulation significantly lowered aspartate transaminase, alanine transaminase, alkaline phosphatase and bilirubin by 47, 61, 55 and 78 %, respectively, and improved the antioxidant defense in the ethanol hepatotoxicity-induced rat model. The current investigation suggests that EFR-FPH in synergy with antioxidant micronutrients is potent in ameliorating ethanol-induced hepatotoxicity and has a potential to form a hepatoprotective dietary supplement.

Evaluation of acute oral toxicity of a broad-spectrum anti-mycotoxin and hepato-protective formulation

Mycotoxins are biologically active, toxic metabolites produced by toxigenic fungi mainly belonging to the genera Aspergillus, Fusarium and Penicillium.They are ubiquitous in poultry feeds. Mix-01-Vil is a broad-spectrum mould inhibitor and mycotoxin binder, recommended for complete protection against multiple mycotoxins and for better production in poultry, cattle and pigs. A study was undertaken to evaluate the acute oral toxicity potential of Mix-01-Vil (M/s Ayurvet Limited, India) according to OECD 423 guidelines. Six (3 male and 3 female) Swiss albino mice were used for the study, where each animal served as its own control. Following the oral administration of the test substance, the animals were observed for the manifestation of toxic effects and mortality. No toxic effects or mortalities were observed till 14 days and Mix-01-Vil was found to be safe for oral use.

Pharmacological screening of polyherbal hepatoprotective -‘Doctor’s LIVAFIT’

Considering the importance of ayurvedic herbs, science behind their potential in treating diseases and advantages over synthetic medicines in limiting side effects, there is an urgent need to employ such sacred science to produce medicines that effectively cure diseases and to distribute them to the needy. Many herbs mentioned in ayurveda are used to treat dreadful hepatic diseases especially, cirrhosis which is caused by the liver damage due to toxins, chronic alcoholism, synthetic drugs etc. In the present study, poly herbal hepatoprotective formulation, LIVAFIT, had been prepared and standardized for determining its quality and safety. Methanol herbal extracts were formulated according to a formula with Phyllanthus neruri, Picorrhiza kurroa, Andrographis paniculata as active ingredients and various other herbs helpful for possible hepatic regeneration and preventing hepatotoxicity. Physico chemically, microscopically and chemically standardized extracts were used in the study. Following the results of acute toxicity studies and after confirming the safety of extracts, hepatoprotective activity of formulation was evaluated by CCL4 induced hepatotoxicity method in albino wistar rats. It exhibited a dose dependent significant (P<0.001) reduction in hepatotoxicity when compared with that of standard, Silymarin and a pre-marketed ayurvedic product.

Bioactivity-guided isolation, characterization, and estimation of esculetin – A Potential Marker from Launaea pinnatifida cass

Background: Loss of expertise in Sanskrit language and medicinal plant identification as per ayurvedic texts has resulted in the creation of “Controversial Drugs,” i.e., plants with different botanical identities being used by the same name Launaea pinnatifida Cass. It is one of the species of one such traditional Indian ayurvedic herb known as “Gojihva.” As per the ayurvedic and modern research, Gojihva possesses hepatoprotective, diuretic, galactagogue, and blood-purifying properties. The present study was aimed at bioactivity-guided isolation, characterization, and estimation of a marker compound obtained from L. pinnatifida. Materials and Methods: Fractions were obtained through the successive solvent extraction technique and subjected to in vitro antioxidant and hepatoprotective bioassays which signified that the methanol extract of leaves had significantly better pharmacological activity. Hence, the methanol extract was subjected to further isolation of the active compound. Estimation of the isolated compound was carried out by high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography fingerprinting. Results: A marker compound was isolated from chloroform:ethyl acetate (90:10) fraction and elucidated to be esculetin based on its spectral analysis (Fourier transform infrared, liquid chromatography–mass spectroscopy, 13C nuclear magnetic resonance, and ultraviolet). The percentage purity and yield of esculetin in methanol extract of leaf fractions was found to be 99.75% and 0.927%, respectively, by HPLC. Conclusion: Esculetin has been reported for the first time in L. pinnatifida. Hence, it can be used as the bioactive marker for this plant. This marker can also be useful in the development of hepatoprotective formulations as well as standardization and quality control of L. pinnatifida formulations.

Ameliorative effect of flaxseed (Linum usitatissimum) and its protein on ethanol-induced hepatotoxicity in Wistar rats.

Hepatoprotective effect of flaxseed and its protein on ethanol-induced hepatotoxicity in adult Wistar rats was investigated. The rats were divided into eight groups of which two served as control (group I: Control for AIN-93M diet groups and group II: Control for cereal-pulse diet groups) and six groups received ethanol orally every day. After 10days along with ethanol, the rats received AIN-93M diet (group III); AIN-93M diet and commercial hepatoprotective formulation (CHF) (group IV); casein replaced by flaxseed protein in AIN-93M diet (group V); cereal-pulse diet (group VI); cereal-pulse diet and CHF (group VII); cereal-pulse diet containing flaxseed (group VIII) for four weeks. The flaxseed and its protein significantly prevented the elevation of plasma markers of hepatic damage, lowered lipid peroxidation, mitigated changes in antioxidant enzymes, and suppressed histopathological signs of hepatic damage. The hepatoprotective effect of flaxseed and its protein was comparable to CHF. These findings implicate the ameliorative effect of flaxseed and its protein on ethanol-induced hepatotoxicity. PRACTICAL APPLICATIONS: Owing to globalization and an increase in earning capacity, alcohol consumption is becoming a part of social life and gradually transforming to addiction. Binge drinking is highly prevalent among low socioeconomic status population, which poses severe risks to health. Alcohol abuse is a public health problem causing three million deaths annually worldwide. Alcohol consumption is known to be a major cause of liver damage worldwide and has contributed to 44% of deaths from liver disease. As abstaining from alcohol is a challenging task, there is an escalating need to formulate potential hepatoprotective agents to prevent alcohol-induced hepatic damage. This study investigates the efficacy of flaxseed and its protein in conferring protection to the liver against ethanol-induced hepatotoxicity. This study also explores the benefits of incorporating flaxseed in the staple cereal-pulse diet. Findings of this study suggest that incorporation of flaxseed or its protein in food formulations can prevent hepatotoxicity and improve the overall quality of life among alcoholics.

EVALUATION OF IN VITRO CYTOTOXIC AND ANTICANCER ACTIVITY OF HEPATOPROTECTIVE POLYHERBAL FORMULATION IN CELL LINE MODEL

Introduction: The use of natural products as anticancer agents has a long history that began with folklore medicine and through the years has been incorporated into traditional and allopathic medicine. Several drugs currently used are derived from medicinal plants.
 Objective: The main objective of this study is to investigate the cytotoxic potential of hepatoprotective polyherbal formulation in normal and cancer cell lines.
 Methods: A 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was utilized to screen the cytotoxic activity.
 Results: The results revealed that the formulation does not induce much mortality in normal liver and kidney cell lines, and LC50 value of liver cell lines was found 1716.355 μg/ml and kidney cell lines 2464.910 μg/ml. The in vitro anticancer activity was performed on liver, colon, and prostate cancer cell lines, and IC50 values are found 2.077, 3.850, and 11.989 μg/ml, respectively, which show excellent anticancer activity.
 Conclusion: Based on the results obtained, the hepatoprotective polyherbal formulation is safe for normal cells and cytotoxic for cancer cells. Further, identification and quantification of phytoconstituents responsible for the activity are in progress.

Herb-Drug Interaction between the Traditional Hepatoprotective Formulation and Sorafenib on Hepatotoxicity, Histopathology and Pharmacokinetics in Rats.

Sorafenib has been used as a standard therapy for advanced hepatocellular carcinoma (HCC). In Asia, patients with HCC are potentially treated with the combination of sorafenib and Chinese herbal medicines to improve the efficiency and reduce the side effects of sorafenib. However, limited information about the herb-drug interactions is available. We hypothesize that the Chinese herbal medicine may exert hepatoprotective effects on the sorafenib-treated group. The aim of this study is to investigate the pharmacokinetic mechanism of drug-drug interactions of sorafenib including interacting with hepatoprotective formulation, Long-Dan-Xie-Gan-Tang formulation (LDXGT) and with two cytochrome P450 3A4 (CYP3A4) inhibitors, grapefruit juice and ketoconazole. Liver enzyme levels and histopathology of liver slices were used to evaluate sorafenib-induced hepatotoxicity and the potential hepatoprotective effects of the LDXGT formulation on subjects treated with the combination of sorafenib and the herbal medicine. In this study, a validated HPLC-photodiode array analytical system was developed for the pharmacokinetic study of sorafenib in rats. As the result of the pharmacokinetic data, pretreatment with the LDXGT formulation did not significantly interact with sorafenib compared with sorafenib oral administration alone. Furthermore, grapefruit juice and ketoconazole did not significantly affect sorafenib metabolism. Furthermore, pretreatment with variable, single or repeat doses of the LDXGT formulation did not suppress or exacerbate the sorafenib-induced hepatotoxicity and histopathological alterations. According to these results, the LDXGT formulation is safe, but has no beneficial effects on sorafenib-induced hepatotoxicity. A detailed clinical trial should be performed to further evaluate the efficacy or adverse effects of the LDXGT formulation in combination with sorafenib in humans.

Hepatoprotective potential of kumaryasava and its concentrate against CCl4-induced hepatic toxicity in Wistar rats.

Objective:Kumaryasava (KS) is a marketed Ayurvedic formulation containing Aloe vera as the main ingredient. It has been used widely for the treatment of liver disorders; however, there is a lack of modern scientific data on hepatoprotection. The recommended dose of KS is high and up to 60 mL/day. The present study describes the preparation of new KS concentrate and evaluation of comparative hepatoprotective activity of KS and prepared KS concentrate at one-third of KS dose against CCl4-induced hepatic toxicity.Materials and Methods:Animals were divided into different groups (n = 6). The first group received normal saline (control) 1.0 mL/Kg/day p.o. for 10 days. The second group (toxicant) was given normal saline 1.0 mL/Kg/day p.o. for 10 days with CCl4 in olive oil (1:1 v/v) at 1.0 mL/Kg/day p.o. Third, fourth, and fifth groups received KS, KS concentrate and a marketed formulation as standard) at doses of 5.0 mL/Kg/day p.o., 1.6 mL/Kg/day p.o., and 100 mL/Kg/day p.o. (tablet suspended in water using 0.1% carboxymethyl cellulose) respectively for 10 days along with CCl4 as given to the toxicant group. On the 11th day, blood was withdrawn from retro-orbital plexus and serum was separated for biochemical estimation of serum glutamic-oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), alkaline phosphatase (ALP), and albumin levels. Later, animals were sacrificed under high dose of anesthesia to remove liver tissue, which were removed and washed with ice cold saline for the estimation of lipid peroxidation. Liver tissue from each group was also fixed in 10% formalin for histopathological analysis.Results:Results demonstrated that both KS and KS concentrate showed the protection against CCl4-induced hepatic toxicity. This was evident from the reduction in serum SGOT, SGPT, ALP levels, and elevation in serum albumin levels observed post treatment of CCl4 treated rats with KS and KS concentrate, which were supported by histopathological data.Conclusion:KS concentrate can be a useful hepatoprotective formulation which may help in reducing the high dose of KS to approximately one-third of the recommended dose.

Determination of Quercetin a Biomarker in Hepatoprotective Polyherbal Formulation through High Performance Thin Layer Chromatography

Background: Quercetin was determined in bioactive fractions of Ocimum gratissimum, Butea monosperma, Bauhinia variegate and polyherbal hepatoprotective formulation by HPTLC method. Methods: Polyherbal hepatoprotective formulation was developed by using five bioactive fractionated extracts of three plants namely Butea monosperma, Bauhinia variegate and O. gratissimum. All three plants contain quercetin. Chromatographic separation was performed on aluminium foil plates coated with 200 μm silica gel 60F254 Linear ascending development with toluene:ethyl acetate:formic acid, 5:4:0.1 (v/v/v) was performed at room temperature (25 ± 2°C) in a twin-trough glass chamber saturated with mobile phase vapor. Compact bands (Rf=0.38) were obtained for quercetin. Spectro densitometric scanning was performed in fluorescence mode at 380 nm. The method was validated for precision, recovery, specificity, detection and quantification limits. Results: Linear regression analysis of the calibration plots showed a good linear relationship (R²=0.9843 ± 0.0001) between peak area and concentration in the range 0.5-2.5 μg/band, respectively. The limits of detection and quantification were 0.089 and 0.26 μ/band. The recovery of the method was 97.33-99.11%. Conclusion: The above method was a rapid and cost effective quality-control tool for routine analysis of quercetin in herbal extracts and in pharmaceutical dosage form.

Design and evaluation of herbal hepatoprotective formulation against paracetamol induced liver toxicity

AimTo isolate and identify the quercetin from polyherbal hepatoprotective formulation. Polyherbal formulations were developed by using five bioactive fractionated extracts of Butea monosperma, Bauhinia variegata and Ocimum gratissimum for treatment of liver disorders by exploiting the knowledge of traditional system of medicine and evaluated for hepatoprotective activity using acute liver toxicity model of paracetamol induced liver damage in rats. MethodsMajor active fractions were isolated by solvent fractionation and quantified by HPTLC method. Two polyherbal tablet formulations were developed by the wet granulation method using microcrystalline cellulose, aerosil and other excipients and subjected for physicochemical evaluation to assess physical stability followed by pharmacological screening. The prepared tablets were finally subjected to stability testing to assess its shelf-life. The rats were monitored for change in liver morphology, biochemical parameters like serum glutamate pyruvate transaminase (SGPT), serum glutamate oxaloacetate transaminase (SGOT), alkaline phosphatase (ALP) and total bilirubin for polyherbal tablet formulation at 50 mg/kg and polyherbal tablet formulation at 100 mg/kg. ResultsActive principle was isolated, quantified by HPTLC and characterized with IR. Both formulations showed significant hepatoprotective activity. The histological studies were also support the biochemical parameters. From the results of biochemical analysis and histopathological studies, it can be accomplished that polyherbal tablet formulation at 100 mg/kg can be effectively formulated into a suitable dosage form with added benefit of no side effects for control and cure of chronic ailments like liver disorders. A comparative histopathological study of liver exhibited almost normal architecture as compared to toxicant group. ConclusionBiochemical marker showed improved results for polyherbal tablet formulation at 100 mg/kg. Polyherbal tablet formulation contains a potent hepatoprotective agent suggested to be a flavone concentrated in polyherbal formulation which may find clinical application in amelioration of paracetamol induced liver damage.

Development and validation of RP-HPLC method for simultaneous analysis of andrographolide, phyllanthin, and hypophyllanthin from herbal hepatoprotective formulation

Summary A simple, sensitive, and accurate liquid chromatographic method with photodiode array detector was developed for the determination of andrographolide, phyllanthin, and hypophyllanthin. The separation was carried out on a reverse-phase 250 mm × 4.6 mm, 5μ symmetry C8 column (Waters). The gradient was prepared from 0.1% orthophosphoric acid (solvent A) and (1:1) acetonitrile:methanol (solvent B) as mobile phase delivered at a flow rate of 1 mL min−1. A linear behavior was observed between observed peak area response, and concentration of analytes was investigated, with good correlation coefficient. The method established was successfully applied to quantify andrographolide, phyllanthin, and hypophyllanthin from the herbal hepatoprotective formulation.

Aqueous extract of Terminalia arjuna attenuates tert‐butyl hydroperoxide–induced oxidative stress in HepG2 cell model

Arjuna (Terminalia arjuna) is a medicinal plant used in many polyherbal hepatoprotective formulations. Although widely claimed to be antioxidant, data supporting such actions of Arjuna are limited. In the present study, we have investigated the efficacy of the aqueous extract of T. arjuna (AETA) using a standard pro-oxidant [tertiary butyl hydroperoxide (TBHP)] in HepG2 cells. Cells were incubated with AETA (5-100 µg/ml) for a range of time points (4-24 h) with or without TBHP (500 μM), and biochemical markers of oxidative stress (OS) were determined. Cells incubated with TBHP showed the significant induction of OS response in cytosol manifested as lipid hydroperoxide (76%-198%) and the generation of reactive oxygen species (60%-127%). Diminished levels of reduced glutathione (35%-60%) and total antioxidant capacity (20%-61%) suggested an altered redox state. Significant perturbations in the activities of antioxidant enzymes such as catalase (30%-56%), superoxide dismutase (25%-68%), glutathione S-transferase (29%-67%), glutathione peroxidase (24%-68%) and glutathione reductase (38%-49%) were discernible suggesting the ongoing OS in the cells. However, cells treated with AETA (100 µg/ml) along with TBHP offered significant protection by reducing levels of lipid hydroperoxide (33%-62%) and ROS (69%) and by increasing antioxidant capacity (54%-81%) and levels of reduced glutathione (49%-82%). Further, it also enhanced the activities of endogenous antioxidant enzymes (superoxide dismutase, 60%; catalase, 35%-82%; glutathione peroxidase, 42-65 %; glutathione reductase, 48%-62%; and glutathione S-transferase, 22%-100%). Taken together, these data suggest that Arjuna can protect against the oxidative damage induced by TBHP and may be effectively used as a hepatoprotective adjuvant to abrogate OS in vivo.

Scientific validation of polyherbal hepatoprotective formulation against paracetamol induced toxicity

ObjectiveThis polyherbal formulation has been traditionally used in the Indian system of medicine as a chief formulation for the treatment of hepatic diseases as hepatoprotective. The aim of the study was to study hepatoprotective activity which will be scientific validation of traditional knowledge claimed about this polyherbal formulation. MethodsHepatotoxicity was induced by administration of paracetamol (300mg/kg) to the animals. The levels of liver enzymes (SGOT, SGPT, Alkaline phosphatase, Serum Bilirubin), lipid profiles (triglyceride, cholesterol, HDL, LDL), creatinine, urea levels and histopathological parameters were measured in order to evaluate hepatoprotective activity of polyherbal formulation. ResultsThe polyherbal formulation produced a significant hepatoprotective activity of the decoction of polyherbal formulation. The polyherbal formulation (PHF = 1) shows good hepatoprotective activity by lowering the levels of SGOT, alkaline phosphatase, bilirubin parameters (P<0.05), lipid profiles – cholesterol, triglyceride, LDL and histopathological evaluations shows that PHF = 1 and PHF = 3 formulations have significantly hepatoprotective activity (P<0.05). ConclusionsThe study validates that polyherbal formulation has a good hepatoprotective activity. Further standardization processes may be performed in order to make it a beneficial hepatoprotective formulation.

Determination of Phyllanthin and Gallic Acid In Herbal Hepatoprotective Formulation By TLC-Densitometry Analysis

Introduction: Phyllanthus niruri, Andrographis paniculata and Embilica officinalis are most important and high value medicinal plant known for its hepatoprotective activity. Several analytical methods including HPTLC and HPLC are reported for respective phytoconstituents. However, there is no simultaneous estimation of phyllanthin and gallic acid reported. The Objective of present work is to develop and validate HPtlc method for simultaneous determination of phyllanthin and gallic acid in polyherbal hepatoprotective formulation. Method: The method is employed tlc alumina plates precoated with silica gel 60F-254 as a stationary phase and toluene: ethyl acetate: formic acid (5:3.5:0.5 v/v/v) as a mobile phase. The standard markers namely phyllanthin and galic acid were identified in polyherbal formulation containing P.niruri and E.officinalis by Rf value. Densitometric analysis of phyllanthin and gallic acid was carried out at 254nm. Results: The precision of method was confirmed by relative standard deviation (RSD) which was lower than the 2%. This method was found to give proper separation of phyllanthin and gallic acid. The linear regression data for calibration plot shown good relationship with r2=0.998 (phyllanthin) and 0.9991(gallic acid) in the concentration range 5-9 µg/band. The method was validated for precision, accuracy, sensitivity, and specificity. Conclusion: statistical analysis proves that method is accurate and reproducible. The method is economical and can be employed for routine analysis of marketed polyherbal hepatoprotective formulation containing P.niruri and E.officinalis.