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IJBPAS, July, 2015, 4(7): 4499-4511 ISSN: 2277–4998

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IJBPAS, July, 2015, 4(7): 4499-4511 ISSN: 2277–4998 IJBPAS, July, 2015, 4(7): 4499-4511 ISSN: 2277–4998 PHARMACOGNOSTICAL STANDARDIZATION AND CHROMATOGRAPHIC EVALUATION OF TECOMELLA UNDULATA STEM BARK PUNDIR S1*, TOMAR SK1, MISHRA SH2, ALBERT S3 AND JAIN M4 1: Research Scholar, School of Pharmaceutical Sciences, Shoolini University, Solan, India. 2: Herbal Drug Technology Laboratory, Pharmacy Department, Faculty of Technology and Engineering, M. S. University of Baroda, Kalabhavan, Vadodara-390001, Gujarat, India. 3: Seed Anatomy Laboratory, Department of Botany, M. S. University of Baroda, Gujarat, India. 4: Department of Pharmacognosy, Oriental University, Jakhya, Indore, Madhya Pradesh, India. *Corresponding Author: E Mail: [email protected]; Mob.: 09817837689 ABSTRACT Tecomella undulata (Family Bignoniaceae), commonly known as honey tree, ammora, rohida, desert teak or white cedar is widely used as an ethno medicine in India. Its stems have found extensive usage by some ethnic communities in reliving body pain and in treating snake bite. However, detailed scientific information is not available to identify the plant material and to ascertain its quality and purity. Therefore in this background meticulous pharmacognostical and chromatographic study of stem bark has been carried to establish the pharmacognostical standards and phytochemical evaluation. Physico-chemical parameters were studied for systemic identification and authentification of leaves. A qualitative fingerprinting of Tecomella undulata (TU), extracts have been performed by HPLC methods. Transverse Sections of stem bark shows presence of various characteristic features like single layer thick walled epidermis, some of its cell forming non- glandular and glandular hairs. In physico-chemical evaluation the water soluble ash 4.02±0.21% w/w is higher than acid insoluble ash 1.49±0.08% w/w. Alcohol & water soluble extractive value was found to be 8.97±0.93 and 9.31±0.54% w/w respectively. Elemental analysis shows presence of high amount of iron (618 ppm) as compare to other elements. Successive extractive value of methanolic extract 7.05±0.12 % w/w is higher than the other extractive values. TLC and HPLC chromatogram shows the presence of various chemical 4499 IJBPAS, July, 2015, 4(7) Pundir S et al Research Article constituents in different Rf value and retention time, which provide quality evaluation and standardization of crude drug. This study provides referential information for identification and standardization of Tecomella undulata stem bark and its extracts. Keywords: Tecomella undulata.; stem; Qualitative Chromatographic evaluation; Histological parameters; Physico-chemical evaluations; elemental analysis Abbreviations: TU- Tecomella undulata Linn; FAA - formalin: acetic acid: 70 % ethyl alcohol; HPLC-High Performance Liquid chromatography; PE-Pet ether extract ; TU- Toluene extract; CF-Chloroform extract; EA-Ethyl acetate extract; ME-Methanol extract. INTRODUCTION Traditional herbal medicines and their Authenticaton, pharmacognostic evaluation, preparations have been widely used for the phytochemical analysis and sensitive thousands of years in developing and chromatographic separation techniques developed countries owing to its natural (HPLC) provide a forefront for development origin and lesser side effects or dissatisfaction of rapid and cost effective methods for with the results of synthetic drug [1]. There standardization and characterization of herbal has been a growing interest in the analysis of drugs and to unknot novel phytoconstituents plant products which has stimulated intense [3, 4, 5]. research on their potential health benefits. Tecomella undulata (family Bignoniaceae), Analytical methods such as photometric commonly known as Ammora (in English) or analysis (UV, IR, MS, and NMR), thin layer Rohida and is locally known as honey tree, chromatography (TLC), high performance desert teak, marwar teak or white cedar, is liquid chromatography (HPLC), and gas widely distributed in Arabia, southern chromatography (GC) can be employed in Pakistan and North west India upto an order to establish the constant composition of elevation of 1200 meters. In Pakistan it is herbal preparations [2]. Majority of crude found in Baluchistan and Sind [6]. Due to the herbs come from nature like wild and it is presence of secondary metabolites with collected to evaluate quality parameters by therapeutic potential Tecomella undulata which presence of various phytochemicals gained importance in medicinal world. As a can be confirmed. Due to heterogenous result, significant number of articles has been composition in form of whole plant natural published traversing over the second half of product standardisation is not a simple task. the 20th century. Phytochemical studies of 4500 IJBPAS, July, 2015, 4(7) Pundir S et al Research Article plant parts has revealed presence of The M. S. University of Baroda, Vadodara, pharmacologically relevant compounds such India. Samples were authenticated in the as Iridoid glycosides [7], naphthaquinone [8, Botany Department and were deposited in the 9] phytosterols, flavanoid glycosides, flavonol Pharmacy Department of The M. S. [10], fatty alcohol [11], fatty acids [12] and University of Baroda. triterpenoids [13]. Tecomella undulata is well Chemicals and instruments known for its medicinal value in Ayurveda All the solvents viz. petroleum ether, benzene, [14, 15] and traditionally, it is extensively chloroform, acetone, ethanol (95%), n-butanol used for treatment of several diseases like and reagents viz. formalin, acetic acid, ethyl liver, spleen, internal tumours, diseases of alcohol, tertiary-butyl alcohol, paraffin wax, abdomen, wound healing, conjunctivitis, safranin, fast-green, potassium iodide were of hepatosplenomegaly, blood purifier, syphilis, analytical grade and were procured from E. gonorrhoea and rewarding in hepatitis [6]. Merck, India. HPLC (Shimadzu, Kyoto, Various reported Pharmacological activity of Japan) and HPTLC (Camag, Switzerland) are Tecomella undulata is anti-HIV [16], the major instruments used for the Antibacterial activity [17], Antimicrobial fingerprinting studies. Microscopic [18], analgesic and anti-inflammatory [19], photographs were taken using a Magnus antioxidants [20] and hepatoprotective [6]. In MIPS-4613 microscope attached with spite of numerous medicinal uses of TU, Magnus live USB 2.0 camera. standardization parameters for its stem bark Macroscopic and Microscopic analysis have not been reported. Hence, the present Morphological features of the stem bark were investigation is an attempt in this direction to examined after the collection. Fresh root bark evaluate morphological, microscopic and were collected from the plant and fixed in physico-chemical characters along with FAA (formalin: acetic acid: 70% ethyl phytochemical screening fluorescence alcohol) for the anatomical studies. Twenty analysis of powdered crude drug and HPLC four hours later, the samples were dehydrated analysis of its extracts. with a graded series of tertiary-butyl alcohol MATERIAL AND METHODS (TBA). Infiltration of the samples was carried Plant Material out by gradual addition of paraffin wax T. undulata stem bark was collected in the (melting point 58º – 60º C) until TBA month of September 2013 from the campus of solution attained super saturation. The sample 4501 IJBPAS, July, 2015, 4(7) Pundir S et al Research Article was then successively embedded into paraffin dissolved in 10 ml of 1 N HCl and then the blocks. solution was filtered and diluted to 50 ml with Sectioning distilled water and used for quantitative With the aid of an MC 930 advanced determination of heavy metals by the atomic precision rotary microtome the paraffin- absorption spectrophotometer (AAS) embedded samples were sectioned. Serial (SYSTRONIC 128), coupled with hydride sections were cut at thickness of 10 - 12 μm. generator and hollow cathode lamps for After that dewaxing of the sections was different elements including heavy metals. performed by treating the specimen slides Preliminary phytochemical screening sequentially with xylol, xylol + alcohol, water Preliminary phytochemical tests for the and finally, with staining fluid. Wherever presence of alkaloids, sugars, phenols, necessary, sections were also stained with flavanoids, saponins, steroids, tannins, fast-green, safranin and iodine in potassium coumarins, terpenoids and glycosides were iodide in order to evaluate cambium, phloem, performed on the petroleum ether, crystals and medullary rays. The Section was chloroform, methanol and aqueous extracts of examined and by using Leica research TU leaves by using standard procedures microscope using ×40 objective the described by Harborne, 1998 [26]. representative areas were photographed. The HPLC studies micro powder analysis was done according to The HPLC analysis of methanolic extract was the method [21, 22] carried with the chromatographic system Physico-chemical analysis (Shimadzu, Kyoto, Japan) consisted of a Physicochemical analysis i.e., percentage ash Shimadzu LC-20 AT Prominence solvent values and extractive values were determined delivery module, a manual Rheodyne injector according to the official methods described (Perkin Elmer, Mumbai, India) with a 20 mL [23] and the WHO guidelines for the quality fixed loop, and an SPD-20A Prominence control methods for medicinal plant materials (Shimadzu) UV-Vis detector. The separation [24]. Fluorescence analysis was carried out was performed on a Hypersil C18 column according to the method [25]. For estimation (particle
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