Pharmacological Evaluation of Safoof-e-Pathar Phori- A Polyherbal Unani Formulation for Urolithiasis.

Safoof-e-Pathar phori (SPP) is an Unani poly-herbomineral formulation, which has for a long time been used as a medicine due to its antiurolithiatic activity, as per the Unani Pharmacopoeia. This powder formulation is prepared using six different plant/mineral constituents. In this study, we explored the antiurolithiatic and antioxidant potentials of SPP (at 700 and 1,000 mg/kg) in albino Wistar rats with urolithiasis induced by 0.75% ethylene glycol (EG) and 1% ammonium chloride (AC). Long-term oral toxicity studies were performed according to the Organization for Economic Co-operation and Development (OECD) guidelines for 90 days at an oral dose of 700 mg/kg of SPP. The EG urolithiatic toxicant group had significantly higher levels of urinary calcium, serum creatinine, blood urea, and tissue lipid peroxidation and significantly (p < 0.001 vs control) lower levels of urinary sodium and potassium than the normal control group. Histopathological examination revealed the presence of refractile crystals in the tubular epithelial cell and damage to proximal tubular epithelium in the toxicant group but not in the SPP treatment groups. Treatment of SPP at 700 and 1,000 mg/kg significantly (p < 0.001 vs toxicant) lowered urinary calcium, serum creatinine, blood urea, and lipid peroxidation in urolithiatic rats, 21 days after induction of urolithiasis compared to the toxicant group. A long-term oral toxicity study revealed the normal growth of animals without any significant change in hematological, hepatic, and renal parameters; there was no evidence of abnormal histology of the heart, kidney, liver, spleen, or stomach tissues. These results suggest the usefulness of SPP as an antiurolithiatic and an antioxidant agent, and long-term daily oral consumption of SPP was found to be safe in albino Wistar rats for up to 3 months. Thus, SPP may be safe for clinical use as an antiurolithiatic formulation.

Development and validation of a high-performance thin-layer chromatographic method for the quantitative analysis of vitexin in Passiflora foetida herbal formulations.

The aim of this study is the development of validated HPTLC method for the quantification of vitexin from Passiflora foetida commercial herbal formulations. The developed method was validated, in accordance with ICH guidelines for precision, accuracy, specificity and robustness. The plate was developed using ethyl acetate:methanol:water:formic acid 30:4:2:1(%, v/v/v/v) on 20 × 10 cm glass coated silica gel 60 F254 plates and the developed plate was scanned and quantified densitometrically at λ = 340 nm. Linear regression analysis revealed a good linear relationship between peak area and amount of vitexin in the range of 100-700 ng/spot. The amount of vitexin in nine commercial herbal formulations was successfully quantified by the developed HPTLC method. The developed and validated high performance thin layer chromatographic method offers a new sensitive and reliable tool for quantification of vitexinin in various herbal formulations containing Passiflora foetida.

Optimization of the extraction condition for benzyl isothiocyanate contents in Salvadora persica roots "Siwak".

Benzyl isothocyanate is the major active antibacterial metabolite in Salvadora persica roots "Siwak" beside two minor isothiocyanate derivatives namely; 3-methoxy benzyl isothiocyanate and 3-hydroxy benzyl isothiocyanate. The extraction condition effect on the amount of benzyl isothiocyanate was explored in detailed study. Both cold and hot extraction with different solvents was applied. The amount of benzyl isothiocyanate was estimated using HPLC and HPTLC. The results indicated that cold extraction of the fresh samples with chloroform offers the maximum amount of benzyl isothiocyanate. Drying process leads to great loss of the active component of Siwak.

HPTLC method for simultaneous determination of ascorbic acid and gallic acid biomarker from freeze dry pomegranate juice and herbal formulation.

A new rapid, simple, sensitive and high performance thin layer chromatography (HPTLC) has been established for the simultaneous determination of ascorbic acid and gallic acid in the freeze-dried pomegranate fruit juice and herbal formulation. HPTLC method was carried out using ethyl acetate: acetone: water: formic acid, 10:6:2:2 (%, v/v/v/v)) on 20 × 10 cm glass coated silica gel 60 F254 plates and scanned at 254 nm for ascorbic acid and gallic acid. Ascorbic acid and gallic acid in the freeze-dried pomegranate fruit juice were identified by comparing their single spot at Rf = 0.54 ±â€¯0.02 and Rf = 0.83 ±â€¯0.01 respectively. The value of regression equation (r2 ≥ 0.9992) revealed a good linear relationship between peak area and amount of ascorbic acid and gallic acid in the range of 100-800 ng/band. The method was validated for precision, accuracy, robustness LOD and LOQ. The method proposed can be useful for routine determination of ascorbic acid and gallic acid in various crude as well as herbal formulations as a quality control tool.

Investigation of antioxidant compounds in commercial pomegranate molasses products using matrix-solid phase dispersion extraction coupled with HPLC.

Pomegranate is a well known fruit for its unique flavor, taste and health benefits. The medicinal properties of this fruits directly associated with the phenolic content present, with great anti-oxidant potential. The research is intended to develop matrix solid phase dispersion method (MSPD) and HPLC quantification of four major anti-oxidant marker constituents (vitamin C, gallic acid, rutin & ellagic acid) in pomegranate molasses samples. The effects of several important experimental parameters like type of dispersant, sample-dispersant ratio, solvents and its volume, time of extraction were investigated. A C18 column with the specification (5 µm, 250 × 4.0 mm) was used for the separation. A gradient flow of mobile phase was selected after many trials containing 0.1%, v/v solution of orthophosphoric acid and acetonitrile. The flow rate was 1.0 mL/min; and the chromatograms were recorded at 254 nm. The validation parameters, like linearity (r2 = 0.9985, 0.9965, 0.9925 & 0.9986), accuracy (100.3, 99.5, 100.9 & 101.9%), intra-day precision (%RSD = 1.09, 1.02, 1.26 & 0.97), inter-day precision (%RSD = 1.32, 0.83, 1.07, & 1.15) LOD (0.07, 4.50, 0.45 & 0.40 µg/mL), LOQ (0.095, 9.50, 0.85 & 9.5 µg/mL) and robustness (% RSD = 0.92, 0.76, 0.81 & 0.83) respectively for vitamin C, gallic acid, rutin & ellagic acid, were found satisfactory as per ICH guidelines.

Quantitative Analysis of Benzyl Isothiocyanate in Salvadora persica Extract and Dental Care Herbal Formulations Using Reversed Phase C18 High-Performance Liquid Chromatography Method.

CONTEXT: Benzyl isothiocyanate is the active antimicrobial agent in Salvadora persica (siwak) widely used in Islamic countries for oral hygiene. AIMS: Quantification of benzyl isothiocyanate in the ethanol extract of S. persica and some dental care herbal formulations labeled to contain siwak. SETTINGS AND DESIGN: Simple and sensitive high-performance liquid chromatography method was designed. SUBJECTS AND METHODS: Separation was achieved on reverse phase C18 (250 mm × 4.6 mm, 5 µ) column with a mobile phase comprising acetonitrile and water (1:1). The detection was carried out at 190 nm using ultra violet-visible detector. The flow rate was kept at 1 mL/min. RESULTS: A sharp and well-defined peak was obtained at the retention time of 9.322 ± 0.3 min. Linear regression analysis data for the calibration plot showed a good linear relationship between response and concentration in the range of 0.5-500 µg/mL with a regression coefficient (r2) of 0.9977. The method was validated for accuracy, precision, robustness, and sensitivity. All the parameters examined met the current recommendations for the International Conference on Harmonization guidelines for method validation. CONCLUSIONS: The method was applied for the quantification of benzyl isothiocyanate in siwak extract, dental care powder, mouth wash, and toothpaste claimed to contain siwak. The developed method was found specific, simple, selective, and reliable for routine use in quality control analysis of different commercially available herbal care products. SUMMARY: A simple, accurate and precise method was developed for the analysis of the antimicrobial agent benzyl isothiocyanate in Salvadora persica (Siwak) extract and selected dental care herbal formulations using RP18 HPLCAmount of benzyl isothiocyanate will indicate the efficacy of Siwak productsThe method subject to ICH validation guidelines. Abbreviations used: RP18: Reversed phase C18, HPLC: High performance liquid chromatography, UV: Ultra violet, r2: regression coefficient, ICH: international conference on harmonization, TLC: Thin layer chromatography, CHCl3: Chloroform, v/v: volume/volume, RSD: Relative standard deviation, LOD: Limit of detection, LOQ: Limit of quantification.

A high performance thin layer chromatographic method for the estimation of colchicine in different formulations.

BACKGROUND: Colchicine is a main alkaloid present in bitter and sweet variety of colchicum (Colchicum luteum Baker), which have been reported to possess anti-rheumatic, anti-gout, and anticancer potential. Colchicum is an important ingredient of several Unani and Herbal formulations. Quantification of colchicine will play a great role in quality control of these formulations. Hence, a high-performance thin layer chromatographic (TLC) method has been developed for the analysis of colchicine in Unani formulations of various dosage forms such as hubb (tablet) and capsules. MATERIALS AND METHODS: The samples were applied on aluminum TLC plates precoated with silica gel 60-F254 and developed using mobile phase toluene-dichloromethane-methanol in equal proportions. Quantification was done by densitometric scanning at 350 nm, which showed a linear response in the range of 50-500 ng/spot. The developed method was validated as per the International Conference on Harmonization guidelines for linearity, precision, accuracy, specificity, robustness, limit of detection, and limit of quantification. RESULTS AND CONCLUSION: The developed method was applied for quantitative estimation of colchicine in different Unani and Herbal formulations. The method was found simple, selective, accurate with a wide range of linearity, hence suitable for the quality control of different formulations and varieties of colchicum with respect to colchicine content.

Quality control and in vitro antioxidant potential of Coriandrum sativum Linn.

BACKGROUND: Coriandrum sativum Linn., commonly known as coriander, is a well-known spice and drug in India. It has various health-related benefits and used in various Unani formulations. In this present study, quality assessment of coriander fruits was carried out by studying anatomical characters, physicochemical tests, and chemoprofiling using high performance thin layer chromatography (HPTLC) and gas chromatography-mass spectroscopy (GC-MS) along with in vitro antioxidant potential. MATERIALS AND METHODS: Standardization was carried out as per the pharmacopeial guidelines. Estimation of heavy metals, pesticides, and aflatoxins was carried out to ascertain the presence of any contaminant in the sample. Chemoprofiling was achieved by thin layer chromatography (TLC) by optimizing the mobile phase for different extracts. The most of the pharmacological activities of coriander are based on volatile oil constituents. Hence, GC-MS profiling was also carried out using hexane-soluble fraction of hydro-alcoholic extract. The total phenolic contents and in vitro antioxidant efficacy were determined using previously established methods. RESULTS: The quality control and anatomical studies were very valuable for the identification whereas good antioxidant potential was observed when compared to ascorbic acid. The drug was found free of contaminant when analyzed for pesticides and aflatoxins whereas heavy metals were found under reported limits. CONCLUSION: The work embodied in this present research can be utilized for the identification and the quality control of the coriander fruit.

Matrix Solid-Phase Dispersion Extraction and Quantification of Alpinetin in Amomum Seed using Validated HPLC and HPTLC Methods.

Alpinetin is a flavonoidal constituent of seeds of Amomum subulatum Roxb., recently reported to possess vasorelaxant and antiHIV activities. Simple, accurate and precise HPLC and HPTLC methods were developed for the analysis of alpinetin in A. subulatum seed extracts and extraction technique was optimized to get maximum yield using conventional, ultrasonic and matrix solid phase dispersion extraction. HPLC was performed on a C18 column with methanol and water (70:30, v/v) as mobile phase at a flow rate of 1.0 ml/min whereas HPTLC on silica aluminum sheet (60F254) using toluene, dichloromethane and ethyl acetate as solvent system. A sharp peak was obtained for alpinetin at a retention time (Rt) of 5.7 min by HPLC and retardation factor (Rf) of 0.48 by HPTLC. Both methods were validated as per the ICH guidelines and the content of alpinetin was estimated in different extracts. Matrix solid phase dispersion technique was found most suitable for extracting alpinetin as compared to other techniques. Validation data are indicative of good precision and accuracy and proved the reliability of the methods.

Metabolic diversity in Coleus forskohlii Briq. of Indian subcontinent.

New economic, easy, specific, accurate, robust, validated high performance thin layer chromatography (HPTLC) and high performance liquid chromatography methods with good range of linearity and sensitivity were developed for quantification of forskolin in ten samples collected from different regions of Indian subcontinent, which showed a large variation among samples (0.074-0.282%, w/w). Metabolic diversity of all the samples using HPTLC fingerprint method showed a total of 16 well separated spots. There is no significant metabolic diversity among the samples collected from different locations of Indian subcontinent, which was obtained from HPTLC fingerprinting. The results of locational variation showed highest content of forskolin in Bengaluru sample by both analytical methods. The validated quantification methods and fingerprint profile together can act as a good authentication tool for coleus as well as for other medicinal plants.

Validated HPLC method for the simultaneous determination of taxol and ellagic acid in a Punica granatum fruit extract containing combination formulation.

A simple, rapid, precise and accurate isocratic reversed-phase high performance liquid chromatography (HPLC) method was developed and validated for the simultaneous determination of paclitaxel and ellagic acid in a combination nanoformulation. Separation was achieved using a 25 x 4.6 mm column, particle size 5 microm C18 reverse phase column (Luna), with a mobile phase consisting of methanol and 0.05% H3PO4, in gradient elution mode with a mobile phase flow rate of 1 mL/min, using UV visible detection at 230 nm. Sharp and well defined peaks were obtained at retention times of 13.75 min. and 11.6 min. for paclitaxel and ellagic acid, respectively. Regression analysis showed a good linear relationship (r2 = 0.996 +/- 0.0011) and (r2 = 0.993 +/- 0.0011) over wide ranges of 5-500 microg/ml and 1-500 microg/ml for paclitaxel and ellagic acid, respectively. LOD and LOQ of paclitaxel were 30 ng/ml and 100 ng/ml, respectively, while for ellagic acid LOD and LOQ were 300 ng/ml and 1 microg/ml, respectively. The accuracy of the method was determined by recovery studies using the standard addition method and was found to be in the range of 99.61-101.21% and 98.70-102.22% for paclitaxel and ellagic acid, respectively. The relative standard deviation (% RSD) for precision, repeatability and robustness was less than 2%. The ellagic acid content in fruits of Punica granatum and combination formulation with paclitaxel was analyzed and found to be 0.04% w/w and 0.0012%w/w, respectively. The proposed, developed and validated HPLC method for the simultaneous quantification of ellagic acid and paclitaxel can be used for the quality control and standardization of several crude drugs and different combination formulations, in which ellagic acid is present.

Rapid RP-HPLC method for the quantification of glabridin in crude drug and in polyherbal formulation.

A simple, economic, selective, precise and robust method has been developed and validated for the analysis of glabridin in crude drugs and polyherbal formulations. Reversed-phase chromatography is performed on a C18 column with water and acetonitrile as mobile phase in gradient elution method at a flow rate of 1 mL/min. Detection is performed at 230 nm and a sharp peak is obtained for glabridin at a retention time of 14.9 ± 0.02 min. Linear regression analysis data for the calibration plot showed a good linear relationship between response and concentration in the range of 1-500 µg/mL; the regression coefficient is 0.9992 and the linear regression equation is y = 26.683x - 142.17. The method is validated for accuracy, precision, reproducibility, robustness and detection and quantification limits, in accordance with International Conference on Harmonization guidelines. Statistical analysis proved that the method is precise, reproducible, selective and accurate for the analysis of glabridin. The proposed, developed and validated high-performance liquid chromatography method for the quantification of glabridin can be used for the quality control and standardization of licorice (Glycyrrhiza glabra Linn.) and different herbal formulations in which licorice is present as a constituent.

Simultaneous HPTLC determination of strychnine and brucine in strychnos nux-vomica seed.

OBJECTIVE: A simple, sensitive, and specific thin layer chromatography (TLC) densitometry method has been developed for the simultaneous quantification of strychnine and brucine in the seeds of Strychnos nux-vomica. MATERIALS AND METHODS: The method involved simultaneous estimation of strychnine and brucine after resolving it by high performance TLC (HPTLC) on silica gel plate with chloroform-methanol-formic acid (8.5:1.5:0.4 v/v/v) as the mobile phase. RESULTS: The method was validated as per the ICH guidelines for precision (interday, intraday, intersystem), robustness, accuracy, limit of detection, and limit of quantitation. The relationship between the concentration of standard solutions and the peak response was linear within the concentration range of 50-1000 ng/spot for strychnine and 100-1000 ng/spot for brucine. The method precision was found to be 0.58-2.47 (% relative standard deviation [RSD]) and 0.36-2.22 (% RSD) for strychnine and brucine, respectively. Accuracy of the method was checked by recovery studies conducted at three different concentration levels and the average percentage recovery was found to be 100.75% for strychnine and 100.52% for brucine, respectively. CONCLUSIONS: The HPTLC method for the simultaneous quantification of strychnine and brucine was found to be simple, precise, specific, sensitive, and accurate and can be used for routine analysis and quality control of raw material of S. nux-vomica and several unani and ayurvedic formulations containing this as an ingredient.

Development and validation of HPLC method for simultaneous estimation of piperine and guggulsterones in compound Unani formulation (tablets) and a nanoreservoir system.

An attempt has been made to develop and validate a simultaneous HPLC method for novel approach of drug release via oil-in-water (o/w) nanoemulsion formulation and Habb-e-Khardal Unani tablet containing piperine and guggul sterones E and Z as main ingredients. Nanoemulsion was prepared by titration method using sefsol-218 as an oily phase, cremophor-EL as a surfactant, transcutol as a co-surfactant and distilled water as an aqueous phase. The formulation was optimized on the basis of thermodynamic stability and dispersibilty test. The nanoformulation was evaluated for particle size, surface morphology, electrical conductivity and viscosity determination. The in vitro dissolution was carried out by dialysis bag method. Drugs were quantified using an HPLC method developed in-house with a C(18) column as stationary phase and acetonitrile and water as mobile phase at λ(max) of 240 nm. The optimized formulation showed higher drug release, lower droplet size and less viscosity as compared with the conventional Habb-e-Khardal Unani tablet. The present study illustrated the potential of nanoemulsion dosage form in improving biopharmaceutic performance of piperine and guggul sterone. The HPLC method was also found to be quite sufficient for the routine quality control of formulations containing piperine and guggul sterone E and Z as ingredients and also for in vitro drug release studies.

Khamiras, a natural cardiac tonic: An overview.

The Unani system of Medicine (Unanipathy), which originated in Greece, is based on the principles proposed by Galen, a Greek practitioner. Since then, many Arab and Persian scholars have contributed to the system. Among them Ibn-e-Sina, an Arab philosopher and Physicist who wrote 'Kitab-al-shifa' are worth mentioning. This system has an extensive and inspiring record in India. It was introduced in India around the tenth century A.D with the spread of Islamic civilization. At present, Unanipathy has become an important part of the Indian system of Medicine. Unani medicines have been used since ancient times, as medicines for the treatment of various ailments. In spite of the great advances observed in modern medicine in recent decades, Unani drugs still make an important contribution to healthcare. The Unani system of medicine is matchless in treating chronic diseases like arthritis, asthma, mental, cardiac, and digestive disorders, urinary infections, and sexual diseases. The medicines administered go well with the temperament of the patient, thus speeding up the process of recovery and also reducing the risk of drug reaction. The Unani system of medicine recognizes the influence of the surroundings and ecological conditions on the state of health of human beings. The system aims at restoring the equilibrium of various elements and faculties of the human body. It has laid down six essential prerequisites for the prevention of diseases and places great emphasis, on the one hand, on the maintenance of proper ecological balance, and on the other, on keeping water, food, and air free from pollution. These essentials, known as 'Asbab-e-Sitta Zarooriya', are air, food, and drink, bodily movement and repose, psychic movement and repose, sleep and wakefulness, and excretion and retention. The Unani system is a secular system in temperament and is popular among the masses. In Unani medicine, although the general preference is for single drugs, compound formulations are also used in the treatment of various complex and chronic disorders. In the light of the present knowledge, this review is a small effort to discuss the efficacious nature of 'Khamira', a semi-solid preparation, which is traditionally used for cardiac ailments, such as, palpitations, weakness of the heart, and so on. On the basis of their constituents these are named as, Khamira Aabresham, Khamira Gaozaban, Khamira Marwareed, and so on. Khameeras are also used as general tonics for other vital organs like the liver and brain. In view of the increasing number of cardiac diseases, a thorough evaluation of this ancient work on Khamira is of special significance.