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International Journal of Ayurveda and Pharma Research
Article · January 2018
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ISSN: 2322- 0902 (P) ISSN: 2322- 0910 (O) International Journal of Ayurveda and Pharma Research
Research Article
ANTIMICROBIAL AND PHYTOCHEMICAL EVALUATION OF CISSUS QUADRANGULARIS L.
Biresh Kumar Sarkar1*, Ravi Kumar2, Vikas Kumar3, Reeta3, Shashi Pal4, C. Murali Krishna5, S. C. Verma6, Ravindra Singh7, Ramaiah Maddi8 *1Assistant Director, 2Lab Technician, 3JRF, Central Ayurveda Research Institute for Respiratory Disorder, (CCRAS, Ministry of AYUSH, Govt. of India) Motibag Road, Patiala, Punjab, India. 4Assistant Professor, Manav Bharati University, Solan, HP, India. 5Regional Ayurveda Research Institute for Skin Disorders, (CCRAS, Ministry of AYUSH, Govt. of India),
Vijayawada, India. 6Pharmacopoeial Laboratory for Indian Medicine (PLIM) Ghaziabad, UP, India. 7Central Council for Research in Ayurvedic Sciences (CCRAS Ministry of AYUSH, Govt. of India), New Delhi. 8Dept. of Pharmacognosy, Hindu College of Pharmacy, Amaravathi Road, Guntur, A.P., India.
ABSTRACT The present investigation involves antimicrobial and phytochemical evaluation of Cissus quadrangularis L. The antibacterial activity of Cissus quadrangularis was performed using disk diffusion method. The Results of study proved prompt efficacy of herbal extract against S. aureus and E. coli. The concentration dependent antibacterial activity of extract was observed against both organisms. Study also involves phytochemical investigation of herbal extract using HPTLC, IR and UV-Visible spectrophotometer. The result of study indicated that the methanolic extract possessed most potent antibacterial activity as compared to other extract. The antibacterial activity increases with the concentration and results indicated that the diameters of zone inhibition of the extract were comparable with the standard drug. The antimicrobial potential of plant extract may be attributed to the presence of specific phytoconstituents. KEYWORDS: Cissus quadrangularis, Herbs, Antimicrobial, HPTLC. INTRODUCTION Cissus quadrangularis is plant belongs from standardization were lacking accuracy and precision Vitaceae family which is an edible plant occurs in therefore modern analytical techniques such as Malaya, Java, India, West Africa and Thailand.[1] It is HPLC, HPTLC and UV-Visible Spectroscopy came in used for strengthening of bone, reduces pain and help practice for the standardization of herbs and herbal to repair bone fractures therefore also called products. The high performance thin layer “Hadjod”.[2] Plant also used for the management of chromatography (HPTLC) play vital role towards the gout, tumors, piles, leucorrhoea and ulcers.[3] The analysis of compounds obtained from natural origin. plant also evaluated for its anti-inflammatory and This technique used for fingerprinting and antioxidant potential by various researchers. The quantification of marker compounds in herbs.[4-6] constituent of plant considered responsible for antimicrobial potential therefore present investigation involve antimicrobial evaluation of Cissus quadrangularis along with its phytochemical investigation using chromatographic and spectro- photometric methods.[1-3] Herbs and herbal products used extensively now a day’s as medicine but the quality concern is major issue regarding the natural products. The quality and composition of herbs and herbal formulation may be ensured using various techniques of standardization which also confirm presence and amount of active principles/marker of formulation. The traditional approaches of Figure 1: Cissus quadrangularis Plant
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Biresh Kumar Sarkar et al. Antimicrobial and Phytochemical Evaluation of Cissus Quadrangularis L. Materials and Methods spotted in the bands of width 8mm with a Camag Plant materials were collected and shade microlitre syringe on pre-coated silica gel glass plate. dried then homogenized to fine powder and stored in The sample loaded plate was kept in TLC twin trough airtight bottles for further use. developing chamber with mobile phase methanol: Preparation of Plant Extract ethyl acetate (6.5: 3.5) and the plate was developed up to the optimum level. Linear ascending Air dried powder was mixed with petroleum development was carried out and the developed plate ether (100ml) in a cotton wool plugged container was dried to evaporate solvents from the plate. The followed by frequent shaking. The supernatant was plate was scanned by UV Densitometric scanning on discarded and solvent allowed evaporating, the Camag scanner.[12] finally obtained dry powder was mixed with 100ml of methanol and rotated again to obtained methanolic Ultraviolet visible absorption (UV) extract. The extracts were centrifuged at 5000rpm The methanol extract of plant was analyzed in and supernatant was collected, solvent evaporated UV-Visible range between 200-780nm using UV- and dry extract was stored in airtight bottles. The Visible Spectrophotometer (UV-1800, Shimadzu). same procedure was repeated for chloroform and Infra-red spectroscopy (IR) water extract. Petroleum ether extract (PE), Under the influence of IR radiations the methanolic extract (ME), chloroform extract (CE) and molecules shows various mode of vibration, which water extract (WE) were use further for gives different absorption spectrum selective to the antimicrobial evaluation.[7, 8] functional group present in molecule. The spectrum Antibacterial Activity of compound is unique feature of molecular Microbes were grown in broth 37°C for 24h. framework. The IR spectra of methanol extract of 100μl of the inoculums was added to each plate Cissus quadrangularis was scanned on FT-IR containing agar. Four different concentrations of the spectrophotometer over the frequency range from sample were used for antimicrobial activities. The 4000-400 cm-1. sterile filter paper disks (6mm in diameter) were Results and Discussion saturated with 50μl of each sample concentration. The yield and appearance of different extract The plates were incubated at 37°C for 24h and the of Cissus quadrangularis were reported in table 1. The diameters of inhibitory zones were measured. The high yield of methanolic extract indicated presence of assay was carried out three times. Disks containing polar component in plant. The antibacterial activity different concentrations of antibiotics were used as of compound performed to evaluate its antimicrobial reference to compare the sensitivity of each tested potential using two different microbial strains i.e., bacterial species.[9-11] S.aureus and E.coli. Results of antibacterial study HPTLC analysis showed that the tested compound inhibited the HPTLC systems equipped with Linomat-5 growth of E. coli and S. aureus significantly. The result applicator with WinCATS-4 software were used. All of study indicated that the methanolic extract the solvents used for HPTLC analysis were of possessed most potent antibacterial activity as selective grade. The dried methanolic extract was compared to other extract (figure 1). The found to be most potent antimicrobial agent antibacterial activity increases with the therefore it was subjected to further analysis. Extract concentration as mentioned in figure 2 and results was dissolved in methanol and used for HPTLC indicated that the diameters of zone inhibition of the analysis. Mobile phase was selected by trial and error extract were comparable with the standard drug. The basis using various combinations of different solvent antimicrobial potential of plant extract may be according to their polarity using precoated Silica Gel attributed to the presence of specific phyto- G-60 plate as stationary phase. The samples (10μl) constituents. Table 1: % Yield and colour of different extract S. No. Extract % Yield Colour 1 Petroleum Ether Extract 45 Dark yellow 2 Methanolic Extract 69 Brown 3 Chloroform Extract 57 Gray 4 Water Extract 62 Reddish Brown
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Int. J. Ayur. Pharma Research, 2018;6(7):24-28
% Inhibition 80 70 60 50 40 % Inhibition 30 20 10 0 PE ME CE WE
Figure 2: Comparative antimicrobial potential (% zone of inhibition) of petroleum ether extract (PE), methanolic extract (ME), chloroform extract (CE) and water extract (WE)
100
50 % Inhibition 0 25 mg 50 mg 75 mg 100 mg
Figure 3: Antimicrobial potential (% zone of inhibition) of different concentration of methanolic extract (ME) UV-Visible spectra of extract are shown in figure 3. The UV spectrum of Cissus quadrangularis showed absorption maxima at 273nm and 255nm.
Figure 4: UV-Visible spectra of extract
IR spectra of plant extract are shown in figure 4. The infrared region approximately 4000–400 cm-1 was used to study the different modes of vibration spectrum. Study confirmed presence of aromatic, carbonyl, C-H stretching and bending vibration along with characteristic peaks of alcoholic and amino group.
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Biresh Kumar Sarkar et al. Antimicrobial and Phytochemical Evaluation of Cissus Quadrangularis L.
Figure 5: IR spectra of plant
High Performance Thin Layer Chromatography (HPTLC) is one of the efficient techniques for separation and identification of plant material which gives better precision and accuracy. The HPTLC fingerprinting of study are shown in figure 5 & 6. Various characteristics peaks were observed in HPTLC fingerprint of extract.
Figure 6: Densitometric chromatogram of Methanolic Extract
Figure 7: HPTLC Chromatogram of Methanolic Extract
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Int. J. Ayur. Pharma Research, 2018;6(7):24-28 CONCLUSION The plant Cissus quadrangularis was AITBS publisher and distributors, Delhi 2002; 28- evaluated for quality parameters using instrumental 30, 38-40, 64-73. techniques of analysis. The results of IR and HPTLC 6. J. B. Harborne, Phytochemical Methods, firsted., may be used further for identification of plant Chapman and Hall Ltd., 1973; pp.10–11. material as fingerprinting tool. Study also involves 7. Yadav M. and Chatterji S., 2014. Preliminary antimicrobial evaluation of Cissus quadrangularis L. phytochemical screening of six medicinal plants using disk diffusion method. The study proved used in traditional medicine, International J. of prompt efficacy of herbal extract against S.aureus and Pharmacy and Pharmaceutical Sciences., Vol. 6, E.coli. The concentration dependent antibacterial Issue 5. activity of extract was observed against both 8. Divya Paikaraa, Bhawana Pandey and Sheetal Singh organisms. Phytochemical Analysis and Antimicrobial Activity REFERENCES of Catharanthus Roseus Indian J. Sci. Res. 12 (2): 1. Shirwaikar A, Khan S, Malini S. Antiosteoporotic 124-127, 2017. effect of ethanol extracts of C. quadrangularis Linn. 9. Kothari V, Seshadri S, Mehta, P, Fractionation of on ovariectomized rat. J Ethnopharmacol 2003; Antibacterial Extracts of Syzygium cumini 89:245-250. (Myrtaceae) Seeds. Biotechnol. Res., 2011; 2(6): 53- 2. Nagani KV, Kevalia J, Chanda SV. Pharmacognostical 63. and phytochemical evaluation of stem of C. 10. M Amareswarareddy, B. Kameswararao. S, quadrangularis L. Int J Pharmaceut Sci Res 2011; Antibacterial Activity of Syzygium cumini in Herbal 2:2856-2862. Tooth Paste Geethika Priscilla, Int. J. Inv. Pharm. 3. Sumitra Chanda, Yogesh Baravalia, Krunal Nagani, Sci., 2014; 2(3): 724-729. Spectral analysis of methanol extract of Cissus 11. Chandrasekaran M, Venkatesalu V. Antibacterial quadrangularis L. stem and its fractions Journal of and Antifungal Activity of Syzygium jambolanum Pharmacognosy and Phytochemistry, 2013; 2(4): Seeds. J. Ethnopharmacol. 2004; 91: 105–108. 149-157. 12. Biresh Kumar Sarkar, Ravi Kumar, Prince Kumar, 4. J.K.Lalla,C.I.Jolly, P.D.Hamrapurkar, Standardization Palash Mandal, Vidya Bhusan Pal, M. Devgan, by GMP Standards for Ayurvedic medications, 48th Formulation and Evaluation of Anti-Inflammatory India National Congress, Madras, 1996; p.14. Herbal Topical Formulation of Cissus 5. Quality controls methods for medicinal plant Quadrangularis L. World Journal of Pharmaceutical materials. World Health Organization, Geneva Research, 2016; 5: 681-689. *Address for correspondence Cite this article as: Dr Biresh Kumar Sarkar, Biresh Kumar Sarkar, Ravi Kumar, Vikas Kumar, Reeta, Shashi Pal, C. Murali Assistant Director, Krishna, S. C. Verma, Ravindra Singh, Ramaiah Maddi. Antimicrobial and Phytochemical Evaluation of Cissus Quadrangularis L. International Journal of Central Ayurveda Research Ayurveda and Pharma Research. 2018;6(7):24-28. Institute for Respiratory Disorder, Source of support: Nil, Conflict of interest: None Declared (CCRAS, Ministry of AYUSH, Govt. of India) Motibag Road, Patiala, Punjab, India. Cell: 8427239267 Email: bireshsarkar@gmail.com
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