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OBJECTIVE: To carry out HPTLC and HPLC fingerprint analysis of 18 batches of Ganoderma samples using two kinds of reference substance of Ganoderma extract, G. lucidum Extract Reference Substance(CZERS) and G. sinense Extract Reference Substance(ZZERS). METHODS: HPTLC Fingerprint was used to analyze triterpene acids and sterols in Ganoderma with chloroform-acetonitrile-methanol-formic acid (13∶2∶0.5∶0.5, develop 3 times) and cyclohexane-ethyl acetate-methanol-formic acid (15∶5∶0.5∶0.5, develop 2 times) respectively. HPLC Fingerprint analysis was conducted using Kromasil 100-5 C18 column (4.6 mm×250 mm, 5 μm) kept at 25 ℃. Mobile phase A was acetonitrile and B was 0.02% phosphoric acid; gradient elution procedure was as follows: 0-40 min, 29%→33% A; 40-70 min, 33%→65%A; 70-105 min, 65%→100%A; 105-120 min, 100% A; flow rate was 1.0 mL•min-1. DAD detector was adopted with detection wavelength set at 244 nm. The injection volume was 10 μL. RESULTS: By using ERS and fingerprint analysis, G. lucidum, G. sessile and G. lucidum could be distinguished. The components of G. lucidum in different species and growth patterns were different. CONCLUSION: There are many varieties of G. lucidum, which can be divided into wild and artificial cultures, and the culture media of artificial culture are different, which leads to the difference of individual components of different G. lucidum. Fingerprint analysis based on ERS of specific varieties are more suitable for the overall quality control of G. lucidum.
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Objective: To investigate the photochemical constituents present in methanol extract of martynia annua seeds using Gas Chromatography-Mass Spectroscopy(GC-MS), High-Performance Thin–Layer Chromatography(HPTLC) analysis and study antioxidant activity. Methods: Methanol extract of Martynia annua seeds were subjected to GC-MS and HPTLC analysis. HPTLC analysis was carried out using GAMAG system with a linomate5 applicator, system mobile phase (Toluene: Chloroform: Ethanol (4:4:1 V/V/V)), two different volume of extract was applied 2 µl and 5 µl. GC-MS analysis was carried out on JEOL GC MATE ΙΙ, column HP 5 MS and Quadruple double focusing mass analyzer. Antioxidant activity was determined by DPPH assay. Results: GC-MS analysis provided 17 peaks indicating the presence of seventeen different phytochemicals in methanol extract of martynia annua seeds. HPTLC fingerprint showed 6 peaks at both size 2 µl and 5 µl at 254 nm whereas 4 peaks, 9 peaks were detected at 366 nm for 2 µl and 5 µl respectively. After derivatization with 10 % methanolic sulphuric acid, 8 peaks, 11 peaks were detected for 2 µl and 5 µl respectively when the derivatized plate was scanned at 540 nm. DPPH free radical scavenging result showed EC50 value of 44.1±1.1 µg/ml. Conclusion: The GC-MS analysis showed the presence of fatty acids, ester, aldehydes and ketones whereas in HPTLC different peaks at different UV-lights before and after derivatization were observed. Maximum percentage inhibition using DPPH assay was found 74 at concentration of 50 µg/ml.
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Macrothelypteris torresiana is a fern species belonging to the family Thelypteridaceae. The present study was conducted to evaluate hepatoprotective potential of ethanol extract from M. torresiana aerial parts (EEMTAP) and detect the polyphenolic compounds present in the extract using high performance thin layerchromatography (HPTLC). Hepatoprotective potential of EEMTAP were tested at doses of 300 and 600 mg/kg, per os (p.o.), on Wistar albino rats. The extract and silymarin treated animal groups showed significant decrease in activities ofdifferent biochemical parameters like serum glutamic oxaloacetic transaminase(SGOT), serum glutamate-pyruvate transaminase (SGPT), and alkaline phosphatase (ALP), which were elevated by carbon tetrachloride (CCl4) intoxication. The levels of total bilirubin and total protein along with the liver weight were also restoredto normalcy by EEMTAP and silymarin treatment. After CCl4 administration, the levels of hepatic antioxidant enzymes such as glutathione (GSH) and catalase (CAT) were decreased whereas the level of hepatic lipid peroxidation (LPO) was elevated. The levels of these hepatic antioxidant enzymes were also brought to normalcy by EEMTAP and silymarin treatment. Histological studies supported the biochemical findings, and treatment with EEMTAP at doses of 300 and 600 mg/kg, p.o. was found to be effective in restoring CCl4-induced hepatotoxicity in rats. A simple HPTLC analysis was conducted for the detection of polyphenolic compounds in EEMTAP, and the result revealed the presence of caffeic acid as phenolic acid and quercetin as flavonoid. The proposed HPTLC method is simple and concise and provides a good resolution of caffeic acid and quercetin from other constituents present in EEMTAP.
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The aim of the study was to establish the finger printing profile and evaluation of in vitro antidiabetic potential of C. obtusa. In vitro antidiabetic activity was carried out according to the method adopted by Miller, 1959. HPTLC studies were carried out using CAMAG HPTLC system equipped with Linomat 5 applicator, TLC scanner 3 and Win cats-4 software for the active fractionation of aqueous-methanolic leaf extracts of C. obtusa. Among the various plant parts analyzed, leaf exhibited efficient inhibitory activity against α-amylase and α-glucosidase enzymes. Therefore, the leaf extract was further fractionated using various solvent systems petroleum ether, chloroform, ethylacetate, butanol and water and were subjected to in vitro antidiabetic activity. Among the fractions analyzed chloroform fraction exhibited remarkable antidiabetic activity by inhibiting α-amylase and α- glucosidase enzymes. Furthermore, the active leaf fractions were analyzed with HPTLC to develop fingerprint profiles and these fractions revealed the presence of 13 and 22 major spots of alkaloids and flavonoids respectively with different Rf values. The results of the present study thus claim the folkloric usage of the plant in diabetic related maladies.
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Objective: To carry out the physicochemical and phytochemical standardization with high performance thin layer chromatography fingerprinting of Piper nigrum L. (P. nigrum) fruits in order to ascertain the standard pharmacognostical parameters of this king of spices. Methods: Many standardization parameters like extractive values, total ash value, water soluble ash value and acid insoluble ash, moisture content, loss on drying and pH values of P. nigrum L. fruits were analyzed. The method of Harborne was adopted for the preliminary phytochemicals screening. Analysis of total phenolic and flavonoid contents, pesticides residues, aflatoxin and heavy metals were also performed. CAMAG-high performance thin layer chromatography system was used for fingerprinting of methanolic extract of P. nigrum L. fruits. Results: The results of phytochemicals testing indicated the presence of carbohydrates, phenolic compounds, flavonoids, alkaloids, proteins, saponins, lipids, sterols and tannins in various solvent extracts. Total phenolic and flavonoid contents in methanolic extract were found to be 1.728 1 mg/g and 1.087 μg/g, respectively. Heavy metals concentrations were found to be within standard limits. Aflatoxins and pesticides residues were absent. Conclusions: The outcome of this study might prove beneficial in herbal industries for identification, purification and standardization of P. nigrum L. fruits.
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There is increasing global interest in herbal and other forms of traditional medicines. Herbs have long been an important source of numerous effective drugs. As per World Health Organization recommendations, there is a need for investigation of traditional medicinal plants for their potential therapeutic efficacy. The bark of Alstonia scholaris (L.) R. Br. (Family: Apocynaceae) locally known as ‘Sapthaparni’ or ‘Satwid’, is reported to have anticancer, antihelminthic, antidiarrhoeal, antiasthamatic, antimalarial etc. The present work embodies the study carried out for quality control of herbal drugs which comprises of macroscopy, microscopy, physicochemical properties, phytochemical analysis, fluorescence analysis and HPTLC fingerprint. The anatomical markers present were found to be stone cells, sclereids, cork cells, fibers and prismatic crystals of calcium oxalate. Methanol soluble extractive value was found to be higher than Water, Ethanol and Petroleum ether soluble extractive values. Preliminary phytochemical analysis revealed the presence of tannins, alkaloids, steroids, amino acids, fats, fixed oil, glycosides, proteins, starch and flavonoids. A unique HPTLC fingerprint for A. scholaris (L.) R. Br. bark was developed. Results of the present study on pharmacognostical and phytochemical investigation of A. scholaris (L.) R. Br. bark will be helpful in developing standards for quality, purity and sample identification of this plant.
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Laghu Gokshur (Tribulus terrestris) and Brihat Gokshur (Pedalium murex) are well known drugs used in Ayurveda as diuretics. These are classified under mishrak varga as ‘Dashmoola’ in Ayurveda and in chemotaxonomy as Saponin glycosides. The objective of the work is to find out the diagnostic tool to identify the two varieties of Gokshur. The powder was studied for macroscopic, microscopic and physicochemical parameters. For HPTLC Stationary phase was Pre-coated silica gel GF 254 and mobile phase was Toluene: Ethyl acetate: Formic acid (7:2:1 v/v/v). The plate was scanned and quantified at 254 nm for Diosgenin.Results shows that microscopic characters like trichomes, stomata and crystals show some difference in the two varieties while physicochemical parameters show difference in extractive values. Phytochemical screening also shows similar findings. HPTLC analysis carried out using Diosgenin as reference standard revealed the presence of steroidal Saponin “Diosgenin”. Quantitative estimation for Saponins found marked variation in the two varieties, where Laghu gokshur had 16% of Saponins while Brihat gokshur had 13% Saponins. The study can be used as a diagnostic tool for identification of these two varieties of Gokshur.