Modern civilization is facing hundreds of disorders associated with microorganisms. The natural phytochemicals from non-edible plants are gaining importance to fight against these disease The intention of this study is to evaluate the ethanol and dichloromethane extracts of flower of Couroupita guianensis (Lecithydaceae) for anti-tubercular activity. The anti-tubercular activity of all the extracts of Couroupita guianensishave been evaluated against Mycobacterium tuberculosis H73Rv strain using Microplate Alamar Blue Assay (MABA). The activity was documented within MIC range of 0.8 to 100μg/ml. The results of MABA showed that both ethanol and dichloromethane extract exhibited significant anti-tubercular activity. The present investigation suggests that Couroupita guianensis possess remarkable anti-tubercular activit

Aravind, D Sai

Babu, P Srinivas

Karthikeyan, R

Journal of Applied Pharmaceutical Research

English

Journal of Applied Pharmaceutical Research (JOAPR)

27     *For Correspondence: rkcognosy@gmail.com ©2017 The authors. This is an Open Access article distributed under the terms of the Creative Commons Attribution (CC BY NC), which permits unrestricted use, distribution, and reproduction in any medium, as long as the original authors and source are cited. No permission is required from the authors or the publishers.                               Journal of Applied Pharmaceutical Research 2017, 5 (1): 27 – 29      IN – VITRO ANTI TUBERCULAR ACTIVITY OF FLOWERS OF COUROUPITA GUANENSIS L. D. Sai Aravind, R. Karthikeyan*, P. Srinivas Babu  Department of Pharmacognosy, Vignan Pharmacy College, Vadlamudi, Guntur 522213, AP  Article Information  ABSTRACT Received: 17th Oct 2016  Modern civilization is facing hundreds of disorders associated with microorganisms. The natural phytochemicals from non-edible plants are gaining importance to fight against these disease The intention of this study is to evaluate the ethanol and dichloromethane extracts of flower of Couroupita guianensis (Lecithydaceae) for anti-tubercular activity. The anti-tubercular activity of all the extracts of Couroupita guianensishave been evaluated against Mycobacterium tuberculosis H73Rv strain using Microplate Alamar Blue Assay (MABA). The activity was documented within MIC range of 0.8 to 100μg/ml. The results of MABA showed that both ethanol and dichloromethane extract exhibited significant anti-tubercular activity. The present investigation suggests that Couroupita guianensis possess remarkable anti-tubercular activity.  Revised: 27th Nov 2016 Accepted: 5th Dec 2016    Keywords Couroupita guianensis flower, anti-tubercular activity, Microplate Alamar Blue Assay    INTRODUCTION There are many medicinal plants all over the world. The drug which is obtained from these plants has lesser side effects. Medicinal plants are age long agents for human beings as they were having many therapeutic values. Now a days plants are playing a crucial role in drug extraction. Plants are economically essential [1]. They contain May constituents that are useful in treatment of many human diseases. Plants are having rich secondary metabolites that are having most therapeutic use for many ailments. Some plants extracts are also used to prevent the antibiotic resistance. Couroupita guianensis is a large deciduous evergreen plant having a great medicinal value. It is having a height of 20 meters. Leaves are alternate oblongative up to 20 centimeters long. It shows racemose inflorescence arising from the trunk and large branches. Flowers are reddish with a yellow tinge on the outside, fragrant with stamens borne on an overarching androphore. Fruit is a large, reddish brown globose, 15-24 cm with woody capsule containing 200-300 seeds each. Each and every part of the plant is having the medicinal value. Tuberculosis (TB) is one of the leading infectious disease and health burden in the world. It has been estimated that, one third of world’s population including 40% from India estimated to be infected with tuberculosis. The leaves of Couroupita guianensisis used as an anti-depressant using in rats. The flowers extract of Couroupita guianensis has different pharmacological properties. Among them the most important activity is Anti tubercular activity as the disease Tuberculosis is the most threatening disease in the present scenario in the world [2].  Couroupita  guianensis has showed a broad spectrum of anti-bacterial and antifungal activities. The leaves of Couroupita guianensis yielded an aliphatic triterpene, it is used as an anti-depressant using in rats. The phenolic compounds of Couroupita guianensis are active in curing the kidney and stomach problems and helpful as anti-inflammatory in action [3]. The biological function of flavonoids, apart from their anti-oxidant properties, includes protection against allergies, inflammation, platelet aggregation, microbial ulcers, hepatotoxins virus and tumours. It is known that one of the active constituents of the medicinal plant Couroupita JOURNAL OF APPLIED PHARMACEUTICAL RESEARCH ISSN No. 2348 – 0335                                                                                             www.japtronline.com Karthikeyan et. al  In – vitro anti tubercular activity of flowers of Couroupita guanensis L.   Journal of Applied Pharmaceutical Research  January – March 2017  Volume 5 Issue 1  28 guianensis namely isatin, is known to exert cytotoxic activity against certain cancer cell lines, being a potential source of new chemo therapeutic agents. In a study performed using extracts of Couroupita guianensis against human promylocytic leukemia (HL60) cells, isatin showed antioxidant activity and was cytotoxic to the HL60 cells due to induction of apoptosis, a natural cell death. These results suggest that isatin can be further evaluated to be used as a prophylactic agent to prevent the free radical induced cancer and as a chemotherapeutic agent to kill the cancer cells [4].   The intention of this study is to evaluatethe ethanol and dichloromethane extracts of flower of Couroupita guianensis (Lecithydaceae) for antitubercular activity.   EXPERIMENTAL METHODS Collection and processing of plant material  The flowers are collected from the plant Couroupita guianensis and they are dried under the shade for 2 weeks and they are powdered finely into powder. The powder was passed through sieve number 40 and the obtained powder was collected.   Extraction of plant materials The powder so obtained weighed 200g and divided into two equal parts and taken in two iodine flask they were added with ethanol and dichloromethane respectively. The two flasks were kept on orbital shaker for 48 hours to obtain the extract. The solvents were evaporated and the two solvents extracts were obtained with the yield of 1.4% and 1.8% respectively.   Microplate Alamar Blue assay (MABA) The anti-mycobacterial activity of compounds were assessed against M. tuberculosis using microplate Alamar Blue assay (MABA).This methodology is non-toxic, uses a thermally stable reagent and shows good correlation with proportional and BACTEC radiometric method. Briefly, 200µl of sterile deionized water was added to all outer perimeter wells of sterile 96 wells plate to minimized evaporation of medium in the test wells during incubation. The 96 wells plate received 100 µl of the Middlebrook 7H9 broth and serial dilution of compounds were made directly on plate. The final drug concentrations tested were 100 to 0.2 µg/ml. Plates were covered and sealed with parafilm and incubated at 37ºC for five days. After this time, 25µl of freshly prepared 1:1 mixture of Almar Blue reagent and 10% tween 80 was added to the plate and incubated for 24 hrs.  A blue color in the well was interpreted as no bacterial growth, and pink color was scored as growth.  The MIC was defined as lowest drug concentration which prevented the color change from blue to pink. [5-7]  RESULTS AND DISCUSSION The results obtained from the microplated almar blue assay for the assesment of anti tubercular activity was shown in figure no. 1. The minimum inhibitory concentration (MIC)of both ethanol and dichloromethane extracts exhibiting minimum inhibitory concentration 1.6μg/ml of respective extracts. The activity may be due to the presence of alkaloids and terpenoids present in the flowers of Couroupita guanensis [8-10].         Table 1:  Inhibitory response of both the extracts of Couroupita guanensis Concentration Ethanol extract Responce Dichloromethane extract Responce 0.8 N Y 1.6 N Y 3.2 Y Y 6.4 Y Y 12.5 Y Y 25 Y Y 50 Y Y     Figure 1: Microplate shows minimum inhibitory concentration of ethanol and dichloro methane extracts of Couroupita guanensis  CONCLUSION The activity was documented within MIC range of 0.8 to 100μg/ml. The results of MABA showed that both ethanol and dichloromethane extract exhibited significant antitubercular activity at 0.8 μg/ml of ethanol extract showed the inhibition and 3.126μg/ml of dichloromethane extract showed its inhibition. The present investigation suggests that Couroupita guanensis possess remarkable anti tubercular activity with increase in concentration of extract. This drug could be Karthikeyan et. al  In – vitro anti tubercular activity of flowers of Couroupita guanensis L.   Journal of Applied Pharmaceutical Research  January – March 2017  Volume 5 Issue 1  29 considered for isolation of responsible bioactive molecule for anti tubercular activity.  REFERENCES  1. Nöthlings UTE, Lynne R, Brian E, Laurence N.  Flavonols and Pancreatic Cancer Risk:  The Multiethnic Cohort Study, American Journal of Epidemiology,  2005: 166( 8) ,   924-931. 2. Maria C S, Lourenco M V, Desouza N, Alessandra, Monica  A. Evaluation of anti-Tubercular activity of nicotinic and isoniazid analogues, ARKIVOC, 2007; (xv), 181-191. 3. Kavitha R, Kamalakannan P, Deepa T, Elamathi R, Sridhar S,Suresh Kumar J. In vitro Antimicrobial Activity and Phytochemical Analysis of Indian Medicinal Plant Couroupita guianensis Aubl, Pharmacologyonline 2011; 3(6), 115-121.  4. Shah GN., Shete SA., Patil VS., Patil KD, Killedar SG. 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In – vitro anti tubercular activity of flowers of Couroupita guanensis L

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