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INDEX – GJRMI, Vol.2, Iss. 2, February 2013 Medicinal plants Research Natural Resource CHEMICAL COMPOSITION AND ANTIBACTERIAL ACTIVITY OF ESSENTIAL OIL OF ZIZIPHORA HISPANICA L. Bounar Rabah, Takia Lograda, Messaoud Ramdani, Pierre Chalard and Gilles Feguiredo
73–80
Agriculture COMPARISON OF COLCHICINE CONTENT BETWEEN HYSTERANTHOUS AND SYNANTHOUS COLCHICUM SPECIES IN DIFFERENT SEASONS Alirezaie Noghondar Morteza, Arouee Hossein, Shoor Mahmoud, and Rezazadeh Shamsali
81–88
Biological Science ECOLOGICAL AND MEDICINAL INTEREST OF TAZA NATIONAL PARK FLORA (JIJEL ALGERIA) BOUNAR Rabah, REBBAS Khellaf, GHARZOULI Rachid, DJELLOULI Yamna and ABBAD abdelaziz 89–101 Bio-Technology CONSERVATIVE PRODUCTION OF BIODIESEL FROM WASTE VEGETABLE OIL Chethana G S, Reddy K Dayakar, Vijayalakshmi
102–109
Indigenous medicine Ayurveda PHYTOCHEMICAL STUDIES ON SMILAX MACROPHYLLA LINN.; A SOURCE PLANT OF CHOPACHEENI Jyothi T, Acharya Rabinanaryan, Shukla C P, Harisha CR 110–117 SELECTION OF MEDICINAL PLANTS FOR THE MANAGEMENT OF DIABETIC FOOT ULCER; AN AYURVEDIC APPROACH Pampattiwar S P, Adwani N V, Sitaram Bulusu, Paramkusa Rao M
118–125
COVER PAGE PHOTOGRAPHY: DR. HARI VENKATESH K R, PLANT ID – FRUITS OF MALLOTUS PHILIPPENSIS (LAM.) MULL. ARG, OF THE FAMILY EUPHORBIACEAE PLACE – AGUMBE, SHIMOGA DISTRICT, KARNATAKA, INDIA
Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80 ISSN 2277-4289 | www.gjrmi.com | International, Peer reviewed, Open access, Monthly Online Journal
Research article CHEMICAL COMPOSITION AND ANTIBACTERIAL ACTIVITY OF ESSENTIAL OIL OF ZIZIPHORA HISPANICA L. Bounar Rabah1, 6, Takia Lograda2*, Messaoud Ramdani3, Pierre Chalard4 and Gilles Feguiredo5 1, 2, 3
Laboratory of Natural Resource Valorization, Sciences Faculty, Ferhat Abbas University, 19000 Setif, Algeria 4 Clermont Université, Université Blaise Pascal, BP 10448, F-63000 Clermont Ferrand 5 LEXVA Analytique, 460 rue du Montant, 63110 Beaumont, France 6 Department of Natural Sciences and Life, Faculty of Science, M'sila University, 28000 M’sila (Algeria) *Corresponding Author : tlograda63@yahoo.fr; +21336835894, +213 776243824; Fax : +21336937943
Received: 24/12/2012; Revised: 25/01/2013; Accepted: 31/01/2013
ABSTRACT The aerial parts of Ziziphora hispanica L. species were collected on April 2011 from Boussaâda localities in Algeria. The chemical compounds of the plant were isolated by hydrodistillation. A total of 28 constituents, representing more than 93.8% of the total oil, were identified by gas chromatograph/mass spectrometry (GC/MS). The most presented compounds of the essential oil of Z. hispanica were Pulegone (78.6%), limonene, menthofuran, trans-iso-pulegone and piperitenone are represented by low concentrations. The essential oil of aerial parts of Z. hispanica has a broad spectrum of antimicrobial activity. The sensitivity of bacteria and fungi tested with essential oil compounds was found to be very high. Key
words:
Ziziphora
hispanica
L.,
essential
oil,
Antibacterial
activity,
Algeria
Cite this article: Bounar R, Takia L, Messaoud R, Pierre C and Gilles F (2013), CHEMICAL COMPOSITION AND ANTIBACTERIAL ACTIVITY OF ESSENTIAL OIL OF ZIZIPHORA HISPANICA L., Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 73–80
Global Journal of Research on Medicinal Plants & Indigenous Medicine || GJRMI ||
Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80
INTRODUCTION Belonging to the family Lamiaceae, Ziziphora hispanica L. is an annual plant with very branchy erect stem. The leaves are all similar, ovate-lanceolate, and ciliate on margins. Spike-like inflorescences composed of verticillastres pauciflores; corolla long tubular structure (Quézel et Santa, 1962–1963). This plant is found in areas of the Saharan Atlas and the highlands. Some of Ziziphora species are used for their aperitif, carminative and antiseptic effects in treatment of various diseases (Ozturk and Ercisli, 2007), especially Z. taurica infusions (Tzakou et al., 2001; Gözde et al., 2006). Z. persica is an edible medicinal plant, it is frequently used as wild vegetable or additive in foods to offer aroma and flavour (Nezhadali et al., 2008, 2009, 2010). Z. clinopodioides, riche in monoterpene glucosides (Megumi et al., 2012), is used mostly in food and medicine (Maya, 2012). Z. hispanica is used as a substitute for Morocco pennyroyal (Mentha pulegium) (Bellakhdar, 1997). According to the population of Boussaâda Z. hispanica is used as an infusion to soothe the stomach pains, for the heart fatigue and added to the coffee for a better taste. A literature survey showed that the oil of Ziziphora species has been found to be rich in pulegone. The major components in Z. taurica, Z. vychodceviana and Z. persica are pulegone and isomenthone (Dembistikii et al., 1995; Sezik and Tumen, 1990; Nezhadali and Zarrabi, 2010). The major constituent found in the oil of Z. tenuior L. has been reported to be pulegone (Sezik et al., 1991). The chemical composition of Z. clinopodioides Lam. was analyzed, the major constituents were pulegone and piperitenone (Salehi et al., 2005; Sonbola et al., 2006; Verdian-Rizi, 2008; Xing et al., 2010; Soltani, 2012). The essential oil of Turkish Z. taurica subsp. clenioides was found to contain pulegone (Meral et al., 2002). The major constituents of essential of Z. pamiroalaica were pulegone and menthone (Xing et al., 2010). Z. capitata contained no oil; Z. persica, Z. taurica, Z. Tenuior and Z.
clinopodioide have a Pulegone as a major compound while Z persica has a major component the thymol (Hüsnü, 2002). The oil of Z. hispanica is characterised by pulegone (Velasco and Mata, 1986; Bellakhdar, 1997; Bekhechi et al., 2007). The essential oil of Ziziphora species has a broad spectrum of antimicrobial activity. The oil of Z. clinopodioides was found to exhibit interesting antibacterial activity against Staphylococcus epidermidis, S. aureus, Escherichia coli and Bacillus subtilis (Sonbola et al., 2006), the oil of Z. clinopodioides was tested against some human pathogenic bacteria, which showed good activity against all tested bacteria, except for Pseudomonas aeruginosa (Soltani, 2012). Investigation of the antimicrobial activity of the essential oil of the Turkish endemic Ziziphora taurica on eight bacterial strains and Candida albicans, indicate that the essential oil remarkably inhibited the growth of tested microorganisms except Candida albicans (Gozde et al., 2006). Z. tenuior oils had bactericidal and inhibitory effects of K. pneumoniae, It can be used as candidates for treatment of infectious diseases that is caused by this bacteria (Mahboubi et al., 2012). The oil from Z. pamiroalaica was better than that from Z. clinopodioides in antioxidant abilities (Xing et al., 2010). The insecticidal and ovicidal effects of essential oil of Z. clinopodioides were tested on adults and eggs of Callosobruchus maculatus (Lolestani and Shayesteh, 2009). The objective of this research is to determine the chemical composition of essential oil of Z. hipanica from the Boussaada region and evaluate its potential to be antimicrobial. MATERIALS AND METHODS Plant material Aerial parts of Ziziphora hispanica were collected during the flowering stage in October 2011 from Boussaâda localities in Algeria. Identified by Dr. Lograda Takia, the voucher specimen is deposited in the herbarium of the Department of Biology, Ferhat Abbas University, Algeria. Z. hispanica was submitted
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80
to hydrodistillation for 3h using a Clevenger apparatus (Lograda et al., 2013). The distilled essential oils were stored at +4 °C for further use. Essential oil Analysis: The essential oils were analysed on a Hewlett-Packard gas chromatograph Model 5890, coupled to a Hewlett-Packard model 5971, equipped with a DB5 MS column (30 m X 0.25 mm; 0.25 μm), programming from 50°C (5 min) to 300°C at 5°C/min, with a 5 min hold. Helium was used as the carrier gas (1.0 mL/min); injection in split mode (1:30); injector and detector temperatures, 250 and 280°C, respectively. The mass spectrometer worked in EI mode at 70 eV; electron multiplier, 2500 V; ion source temperature, 180°C; MS data were acquired in the scan mode in the m/z range 33–450. The identification of the components was based on comparison of their mass spectra with those of NIST mass spectral library (Masada, 1976; NIST, 2002) and those described by (Adams, 2001) as well as on comparison of their retention indices either with those of authentic compounds or with literature values (Adams, 2001).
agar supplemented with 5% sheep blood for fastidious bacteria were poured in Petri dishes, solidified and surface dried before inoculation. Sterile discs (6 mm Φ) were placed on inoculated agars, by test bacteria, filled with 10 μl of mother solution and diluted essential oil (1:1, 1:2, 1:5, and 1:10 v:v of Dimethylsulfoxide (DMSO). DMSO was used as negative control. Chloramphenicol for bacteria and amphotericin B for fungi were used as positive control. Bacterial growth inhibition was determined as the diameter of the inhibition zones around the discs. All tests were performed in triplicate. Then, Petri dishes were incubated at 37°C during 18–24 h aerobically (Bacteria) and at 25°C for 7 days (fungi). After incubation, inhibition zone diameters were measured and documented. RESULTS The essential oil, of Ziziphora hispanica L., isolated by hydrodistillation from the aerial parts, was obtained in yield of 1.01% (v/w). The chemical composition of essential oil, analyzed by gas chromatography/mass spectrometry (GC-MS), gave 28 constituents representing 93.82% of the total oil. The names of the corresponding compounds and their percentages are listed in table 1.
Antibacterial activity: Two Gram positive bacteria (Staphylococcus aureus ATCC25923 and Bacillus subtilis ATCC 6633) and seven Gram negative bacteria (Pseudomonas aeruginosa ATCC27853, Pseudomonas syringae pv. Tomato ATCC 1086; Escherichia coli ATCC 25922, klebsiella pneumoniae CIP 53-153, Salmonella enterica CIP 60-62T, Enterobacter sp. and Citrobacter sp.) and three fungi (Aspergelus flavus LBVM20, Aspergilus niger LBBM62 and Candida albicans ATCC 24433) were used in this study. The bacterial inoculums was prepared from overnight broth culture in physiological saline (0.8 % of NaCl) in order to obtain an optical density ranging from 0.08–01 at 625 nm. Muller-Hinton agar (MH agar) and MH
The oil is characterized by a high content of pulegone (78.6%). Other compounds a low rate were piperitenone (2.9%), 8-hydroxy-pmenthan-3-one (2.24%), menthofurane (1.26%), trans-isopulegone (1.09%) and limonene (1.4%). The analysis of Z. hispanica essential oil revealed the presence of a high percentage of ketone monoterpene with the pulegone (78.6%) its dominant compound. The ketone (3.33%) represents the second class of chemical oil, followed by the terpene oxide (1.31%). The monoterpene with 7 compounds, represent 2.37% of total oil with limonene as major compound, unlike sesquiterpene (0.65%), alkene (0.3%), Ether (0.98%) and monoterpene alcohol are poorly represented (Table 2).
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80
Table 1: Chemical composition of Ziziphora hispanica essential oil Compounds α-pinene Cyclohexanone-3methyl Sabinene β-pinene β-myrcene Isolimonene-trans Limonene Iso menthone 1,8-Cineole (-)-L-Isopulegol Camphor Trans-isopulegone Menthofuran DB5-785 neo-Menthol
KI 939 952
% Compounds 0.52 (3Z,5E)-1,3,5-undecatriene 0.24 1-Dodecene
976 980 987 983 1031 1130 1033 1145 1146 1157 1164 1166
0.11 0.5 0.3 0.1 1.4 0.11 0.1 0.1 0.06 1.09 1.26 0.05
α-terpineol Puleone Piperitenone 8-hydroxy-p-menthan-3-one 1,3-Dimethyl pyrogallate α-copaene β-bourbonene β-caryophyllene γ-cadinene Mint furanone-2 Caryophyllene oxide 2-Pentenoic acid, methyl ester, (E)
KI 1184 1192
% 0.41 0.36
1196 1237 1245 1256 1357 1376 1417 1425 1514 1520 1582 1592
0.71 78.6 2.9 2.24 0.98 0.2 0.1 0.4 0.1 0.59 0.11 0.18
Table 2: Chemical classes and dominant compound oil from Ziziphora hispanica Chemical class monoterpene terpene oxide monoterpene ketone monoterpene alcohol ketone ether alkan alkene sesquiterpene others
Nb 7 2 4 3 3 1 2 1 5 1
% 2.59 1.31 81.45 0.79 3.07 0.98 0.77 0.30 0.91 0.18
Dominant compound Limonene Menthofurane Pulegone α-terpeneol 8-Hydroxy-.delta.-4(5)-p-menthen-3-one Syringol (3Z, 5E)-1, 3, 5-undecatriene 3-nanone β-caryophyllene 2-Pentenoic acid, methyl ester, (E)-
The present research showed that sensitivity of bacteria Gram-positive, to the essential oil of Z. hispanica, is higher than that of Gramnegative bacteria. Antimicrobial activity results are shown in table 3. The essential oil of aerial parts of Z. hispanica has a broad spectrum of antimicrobial activity. Although this essential oil has remarkably inhibited the growth of all tested bacteria including medically important pathogen
% 1.06 1.26 78.6 0.71 2.24 0.98 0.41 0.30 0.40 0.18
Staphylococcus aureus ATCC 6538/P (inhibition zone is 40 mm). Essential oil has weakly inhibited the growth of Aspergelus flavus LBVM20 and A. niger LBBM62, while its action on Candida albicans ATCC 24433 is highly active. Anti-bacterial activities of Z. hispanica essential oil show the presence of Pulegone found as 77.53% in volatile oil and also Limonene and Piperitenone can be responsible in the activity.
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80
Table 3: Antibacterial activity of Ziziphora hispanica essential oil Strains Inhibition zone Cont (mm) rol [C] v/v 1 ½ ¼ Bacteria Bacillus subtilis ATCC 6633 22 22 20 24 Citrobacter sp. 20 20 18 22 Escherichia coli ATCC 25922 22 24 25 28 Enterobacter sp. 22 24 25 20 Klebsiella pneumoniae CIP 53-153 45 49 32 22 Pseudomonas aeruginosa ATCC 27853 38 42 44 26 Pseudomonas syringae pv. Tomato ATCC 1086 35 37 33 20 Staphylococcus aureus ATCC 25923 40 40 41 25 Salmonella enterica CIP 60-62T 45 43 40 30 Fungi Aspergelus flavus LBVM20 Aspergilus niger LBBM62 Candida albicans ATCC 24433
8 9 33
8 8 34
12 8 35
25 20 22
Inhibition zone (diameter of the disk, 6 mm, included), values represent average of 3determinations; Control: Chloramphenicol for Bacteria and Amphotericin B for fungi (10 μg/disk); CIP: Collection of Pasteur Institute, Algeria; ATCC: American Type Culture Collection; LBM: Laboratory of Biotechnology and Metagenomic, M’sila, Algeria
DISCUSSION The result of this research is in accordance with other earlier studies on Ziziphora species that are all found to be rich in pulegone and the review of the published literatures reveal that the composition of Ziziphora species oil shows large similarity in the major components, but relative concentrations have some difference (Gözde et al., 2006; Sonboli et al., 2006; Ozturk and Ercisli, 2006, 2007; Aghajani et al., 2008; Amiri, 2009; Maya, 2011; Ozturk et al., 2007 and Soltani, 2012).
The essential oil of Z. clinopodioides showed good activity against all test bacteria (Soltani, 2012). The antibacterial activity of the oil may be associated with the relatively high pulegone, piperitenone and 1- 8-cineole content. It has been reported that these components have significant antimicrobial activities (Sezik et al., 1991; Meral et al., 2002; Bakkali and Averbeck, 2008; Sonboli et al., 2006). CONCLUSION
The previous studies showed that Pulegone and Limonene are anti-bacterial (Maya, 2011). The results in this study are consistent with the other antibacterial study results of Ziziphora species and other pulegone rich plants. However, it has been reported that the essential oils of pulegone rich plants such as Micromeria silicica and Mentha suaveolens inhibited Candida albicans (Gözde et al., 2006).
In conclusion, the essential oil of the aerial parts of Z. Hispanica, remarkably inhibited the growth of all tested gram positive and gram negative bacteria and the fungus tested. The essential oil with a composition of pulegone (77.35%), piperitenone (2.90%) and limonene (1.06%) and its observed antibacterial properties show that the essential oil could be evaluated in the pharmaceutical industry as a possible new pulegone resource.
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 73–80
ACKNOWLEDGEMENTS
Blaise Pascal University (France) and MESRS of Algeria.
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activity of the essential oil of Salvia ringens. Planta Med. 67(1): 81–83. Velasco Negueruela A, Mata Rico M (1986). The volatile oil of Ziziphora hispanica L. Flavour and Fragrance Journal. 1(3): 111–113. Verdian-Rizi Mohammadreza (2008). Essential oil composition and biological activity of Ziziphora clinopodioides Lam. From Iran. Research Journal of Pharmacology, 2(2): 17–19. Xing
Si-lei，Zhang Pi-hong，Ji Qiaoling，Jia Hong-li，Wang Xue-hua (2010). Essential Oil Compositions and Antioxidant Activities of Two Ziziphora Species in Xinjiang. Food Sciences. 31(7): 154–159.
Tzakou O, Pitarokili D, Chinou I B, Harvala C (2001). Composition and antimicrobial
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Conflict of Interest: None Declared
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 81–88 ISSN 2277-4289 | www.gjrmi.com | International, Peer reviewed, Open access, Monthly Online Journal
Research article COMPARISON OF COLCHICINE CONTENT BETWEEN HYSTERANTHOUS AND SYNANTHOUS COLCHICUM SPECIES IN DIFFERENT SEASONS Alirezaie Noghondar Morteza1*, Arouee Hossein 2, Shoor Mahmoud 2, and Rezazadeh Shamsali 3 1*
PhD student, Ferdowsi University of Mashhad, Agriculture Faculty, Horticultural Sciences Department, Mashhad, Iran 2 Assistant Professor, Ferdowsi University of Mashhad, Agriculture Faculty, Horticultural Sciences Department, Mashhad, Iran 3 Assistant Professor, Institute of Medicinal Plants, Department of Pharmacognosy and Pharmaceutics, ACECR, Tehran, Iran *Corresponding author: Email: mortezaalirezaie@yahoo.com
Received: 13/12/2012; Revised: 24/01/2013; Accepted: 30/01/2013
ABSTRACT In order to compare of different phonological stages and seasonal changes of colchicine content between hysteranthous and synanthous colchicum species, amount of colchicine was determined in Colchicum speciosum Steven, C. kotschyi Bioss and C. robustum Stefanov, in different seasons, 2009–2010. The observations under wild conditions showed, that the leaves of appeared with flowers in the same stage of life cycle (synanthous) in C. robustum, while in case of C. kotschyi and C. speciosum flowers occurred first and leaves later, in another developmental stage (hysteranthous). Seed’s colchicine content in C. robustum, C. kotschyi and C. speciosum was obtained as 1.28, 0.46 and 0.92 mg g-1 dry weight, respectively. Corm’s colchicine content was higher in C. speciosum than the other species in all seasons. The highest colchicine content of corm in C. speciosum was obtained in winter and autumn (2.17 and 2.13 mg g-1 dry weight, respectively), while in C. robustum and C. kotschyi it was found in autumn, 0.49 and 0.77 mg g-1 dry weight, respectively. The lowest colchicine content of corms was obtained in summer, when the corms were dormant before flowering stage, in C. speciosum and C. kotschyi, 0.131 and 0.0058 mg g-1 dry weight, respectively, whilst in C. robustum obtained in winter, 0.08 mg g-1 dry weight, synchronous to flowering and vegetative growth. KEYWORDS: Colchicine content, Colchicum kotschyi, C. speciosum, C. robustum, Flowering stage, Hysteranthous, Root activity, Synanthous, Seasonal changes.
Cite this article: Alirezaie Noghondar Morteza, Arouee Hossein, Shoor Mahmoud, and Rezazadeh Shamsali (2013), COMPARISON OF COLCHICINE CONTENT BETWEEN HYSTERANTHOUS AND SYNANTHOUS COLCHICUM SPECIES IN DIFFERENT SEASONS., Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 81–88
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The genus Colchicum belongs to the family Colchicaceae, which comprises of 19 genera, and 225 species (Nordenstam, 1998). Plants of the genus Colchicum have been known for more than 2000 years for their marked beneficial and poisonous effects (Brickell, 1984). The modern medicine uses Colchicum as a source of therapeutically active alkaloids called colchicinoids. One of the most abundant alkaloid - colchicine, is known to have cancerostatic, antirheumatic, antimitotic, antiinflammatory, cathartic and emetic effects. It is also applied in plant breeding to induce polyploids (Komjatayova et al., 2000; Frankova et al., 2005). In addition to the genus Colchicum, colchicine was reported from species belongs to Merendera and Gloriosa genera, which belonging to the Colchicaceae family (Nordenstam, 1998).
and seeds. C. autumnale seeds contain 0.6-1.2%, while corms contain up to about 0.6%. Seeds are mainly used by the pharmaceutical industry for the extraction of colchicinoids (Trease & Evans, 1983). The content of colchicine alkaloid in corms, stems, leaves, and flowers of C. cilicicum were 0.05%, 0.01%, 0.01% and 0.20% (g% dry weight), respectively (Sütlüpinar et al., 1988). In another study by Alali et al. (2004), C. stevenii corms, flowers and leaves were reported to contain 0.17, 0.12 and 0.20 (wt/wt) g%, respectively, while C. hierosolymitanum corms and flowers were found to contain 0.13 and 0.09 (wt/wt) g%, respectively. Ondra et al. (1995), assayed corms of seven Turkish Colchicum species; namely: C. macrophyllum, C. turcicum, C. cilicicum, C. kotschyi, C. bornmuelleri, C. speciosum and C. triphyllum for their colchicinoid alkaloids. Colchicine content was found to be 222.3, 323, 300, 1058, 3063, 4245 and 958 µg g-1 dried drug, respectively.
Many factors are interfering in biosynthesis of secondary metabolites such as essential oils and alkaloids. The study conducted by Takia et al. (2013), has shown that essential oil composition and content in four populations of Pituranthos scoparius were different. Very little is known about the factors interfering with the biosynthesis of colchicine-like alkaloids. Results obtained by Sütlüpinar et al. (1988), indicated that the composition of tropolone alkaloids differs in different parts of the plants and varies during the different growth stages (Sütlüpinar et al., 1988). Presence and concentration of colchicine is determined by a variety of environmental factors including season (Vicar et al., 1993; Poutaraud and Girardin 2002; Alali et al., 2006) and resource availability (Hayashi et al., 1988; Pouraraud and & Girardin, 2005; Mróz, 2008) as well as genetic variations between populations and individuals (Poutaraud and Champay, 1995). Also colchicine content varies among different organs of the plant body (Sütlüpinar et al., 1988; Alali et al., 2004; Alali et al., 2006).
Colchicine variation in different organs of plant and during different growth stages has been studied by researchers. Colchicine and demecolcine were determined in raw and dried leaves, stems, mother and daughter corms of C. autumnale in four stages of its ontogenesis by Vicar et al. (1993). They found that colchicine content in raw material varies during plant growth. Colchicine content in C. brachyphyllum and C. tunicatum, was determined during different growth stages by Alali et al. (2006). Underground parts in both species and during different growth stages, always showed higher colchicine content than the above ground parts. In C. brachyphyllum, total colchicine content of underground parts during flowering stage was found to be about 0.15% (wt/wt), while that of aerial parts was only about 0.04% (wt/wt). In C. tunicatum, total colchicine content of underground parts was found to be 0.12% (wt/wt) and 0.13% (wt/wt) during flowering and vegetating stages, respectively, while that of aerial parts was only about 0.04% (wt/wt) and 0.02% (wt/wt), respectively (Alali et al., 2006).
Among all species of Colchicum, C. autumnale is the best source for colchicine. The richest plant parts in colchicine are the corms
Generally, geophytes are plants that survive by subterranean storage organ with renewal buds (Raunkiaer, 1934). They divide into two
INTRODUCTION
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 81–88
groups – synanthous and hysteranthous one. The leaves of synanthous geophytes coexist with flowers in the same stage of life cycle. In case of hysteranthous plants flowers occur in the first and leaves later, in another developmental stage (Dafni et al., 1981). A special case is the hysteranthous plant Colchicum tunicatum which perceive the photoperiodic signal when the dry bulb lies well below the soil surface (Halevy, 1990). C. speciosum, C. kotschyi Boiss, and C. robustum stefanov, are three wild growing Iranian Colchicum species (Presson, 1992). C. speciosum Steven and C. kotschyi Bioss are hysteranthous but C. robustum is a synanthous species (Presson, 1992). So far no study has been performed on colchicine content variation between synanthous and hysteranthous Colchicum species in different seasons, thus the aim of this study was to evaluate phenological changes and their relationship with corm and seed colchicine content variation among three Iranian native Colchicum species, under their habitat conditions. MATERIAL AND METHODS Plant Material The corms of three wild Colchicum species were collected in different seasons (spring, summer, autumn and winter during 2009–2010, and seeds were collected in spring 2010. Corms and seeds of C. speciosum, C. robustum and C. kotschyi were collected from Khalkhal-Asalem road, Ardabil province, at an altitude of 940m in Iran, Babaaman Mountain, North Khorassan Province, at an altitude of 1091m in Iran and Noghondar valley near Mashhad, Razavi Khorasan province, at an altitude of 1400m in Iran, respectively. The collected materials of three species were identified by Mohammad Reza Joharchi, Ferdowsi University of Mashhad Herbarium (FUMH). Voucher specimens of C. kotschyi (Herbarium Number: 39516), C. robustum (Herbarium Number: 39519) and C. Speciosum (Herbarium Number: 39531) were registered. These are kept in the herbarium of FUMH.
Recording developmental stages To study the plant phenology in wild conditions observations were carried out for three species from three different locations during 2009–2010. Observations were including of developmental stages such as beginning of flowering, peak flowering time, root formation time, beginning of vegetative growth, fruiting and capsule formation and daughter corm formation in wild conditions. Extraction and Isolation The methods described by Rosso and Zuccaro (1998) and Alali et al. (2006), were adopted with some modifications. Acetonitrile, methanol and other reagents were of chromatographic grade and prepared from Panreac (Spain). Reference standard of colchicine was prepared from USP. The corms were sliced into small pieces and air-dried at room temperature together with the seeds. After drying, exact weight of 2 g of corms (collected in different seasons) and 2 g of seeds of three Colchicum species were grounded to powder in a laboratory mill and then used for extraction. Powdered material placed into 250 mL Erlenmeyer flasks and extracted with 100 mL of methanol in 35oC for 1h with ultrasonic apparatus. Afterwards, plant residues were filtered through Wattman filter paper and the filtrates were saved. Then plant residue was transferred into Erlenmeyer flasks again and extracted with 50mL of methanol in 35oC for 30min with ultrasonic apparatus and then filtered. Plant residues were washed with 10 mL of methanol and then filtered. The collected filtrates and washes were combined and transferred into a 250 mL separatory funnel and extracted with petroleum ether (30 mL × 3) with frequent shaking for 30 min in order to remove non-alkaloid substances. 10 mL of distillate water was added each time for better separation and creation of two separate phases. The resulting methanolic phase was transferred to an empty separatory funnel and extracted with chloroform (30 mL × 3) for 10 min. The chloroform phases obtained from three stages were collected and Sodium sulphate Anhydrous
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was added to the chloroformic solution for dewatering of it and then filtered through filter paper. The chloroformic extract was dried in vacuum and then dissolved in 5 ml of HPLC grade methanol and injected to the HPLC instrument. Injection volume at 50 μl, room temperature, detection at 243 nm. All analyses were done in duplicate. HPLC instrument was Knauer ® (Germany) equipped with auto sampler and column was Bondapak C18 (Technochrom) micrometer particles and 4.9 mm id and 250 mm in length. An UV detector K-2501 and a dynamic mixing chamber were employed. Mobile phase system
consisted from phosphate buffer pH=6 and acetonitrile (77: 23). For preparation of phosphate buffer 800 mg of NaH2Po4 and 200mg of Na2HPo4 were dissolved in 1000mL of HPLC grade water and the pH was adjusted on 6. The flow rate was adjusted to 2mL/min and detection was performed at a wavelength of 243nm. The stock solution of colchicine standard was prepared by accurately weighing of colchicine reference standard and then diluted using HPLC grade methanol to construct calibration curve of six –points (30, 50, 75, 90, 100 and 120 ppm). Figure 1 shows colchicine HPLC analysis standard curve.
Figure. 1. Colchicine HPLC analysis standard curve
RESULTS AND DISCUSSION Developmental stages Table 1 shows, beginning time of developmental stages in three colchicum species under their habitat conditions. The results showed that flowering started sooner in C. Speciosum (end of August) and C. kotschyi (middle of September) than to C. robustum (end of January). Fruiting and capsule formation started later in C. robustum (middle of April) than to C. speciosum (beginning of April) and C. kotschyi (end of March). In all species root activity got initiated in middle of autumn (Table 1).
Observations showed that C. kotschyi and C. speciosum were hysteranthous geophyte (flowers develop first and leaves later) and autumn-flowering species but C. robustum was a synanthous geophyte (leaves coexist with flowers in the same stage of the life cycle) and winter-flowering species. This type of obviously hereditary phenological behaviour is rather the rule in the genus Colchicum, in contrast to the onset of leaf growth which seems to be largely environmentally triggered (Burtt, 1970; Gutterman and Boeken, 1988; Persson, 1999).
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Table 1. Beginning of developmental stages of three colchicum species under natural conditions Different Species
Beginning of flowering
Pick time of flowering
Root formation
Vegetative growth E-Mar
Fruiting and capsule formation B-Apr
Daughter corm formation E-May
C. speciosum
Ea-Aug
E-Sep
E-Oct
C. kotschyi
M-Sep
B-Oct
B-Nov
B-Feb
E-Mar
M-May
C. robustum
E-Jan
M-Feb
B-Dec
B-Feb
M-Apr
B-May
Notes: a B, M and E indicate the beginning, middle and end of each month, respectively
Colchicine content The results of HPLC analysis of plant extracts are summarized in table 2. The level of colchicine varies in different seasons as well as species and plant parts. Seed’s colchicine content in C. robustum was higher than the other species. Seed’s colchicine content in C. speciosum, C. kotschyi and C. robustum was 0.92, 0.46 and 1.28 mg g-1 dry weight (DW), respectively (table 2). The amounts of corm colchicine in C. speciosum were higher than the other species in all seasons. Among different seasons the highest colchicine content of corm in C. speciosum was obtained in winter (2.17 mg g-1 DW), while in C. robustum and C. kotschyi it was found in autumn, 0.49 and 0.77 mg g-1 DW, respectively. The lowest colchicine content of corm was obtained in summer in C. speciosum and C. kotschyi was found to be about 1.31 and 0.058, respectively, while in C. robustum it was in winter, 0.08 mg g-1 DW. Corm’s colchicine content in C. speciosum and C. kotschyi (as hysteranthous species) in autumn and winter were higher than to spring and summer, while in C. robustum (as a synanthous species) the highest corm colchicine content was obtained in autumn. The lowest colchicine content in C. kotschyi and C. speciosum was obtained in summer, whilst in C. robustum, it was observed in winter.
Colchicine content in different species varies considerably during different seasons. Matching of the table related to developmental stages with seasonal variation of colchicine content indicates that corm colchicine content in the three colchicum species studied was high in autumn (at the time of root activity). The lowest colchicine content of corm in C. speciosum and C. kotschyi (as hysteranthous species) was observed when the corms were dormant, while in C. robustum (as a synanthous species) it was obtained during flowering and vegetative stages. During flowering stage and in the absence of leaves, the only source of colchicine in flowers could be due to the translocation of colchicine from corms and this may explain the slightly low corm colchicine content at flowering stage (Al-Fayyad et al., 2002). Seed colchicine content in C. robustum was higher than those of the other species. Previously reported that the amount of seed alkaloid and colchicine content is more in unripe seed and declines as the seeds mature (Poutaraud and Girardin, 2003; Alali et al., 2006). Since In the present study, the capsules and seeds of C. speciosum and C. kotschyi were formed sooner than those of C. robustum so it seems that, less colchicine content in C. speciosum and C. kotschyi seeds had been due to more mature their seeds than those of C. robustum.
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Table 2. Mean Colchicine content of different organs of the three Colchicum species (mg g-1 dry weight) during different seasons.
Season
organ
Species C. speciosum C. robustum
Winter
corm
2.17 ± 0.04a
0.08 ± 0.002
0.60 ± 0.03
Spring
corm
1.47 ± 0.03
0.21 ± 0.003
0.50 ± 0.03
Summer
seeds corm
0.92 ± 0.02 1.31 ± 0.02
1.28 ± 0.02 0.13 ± 0.002
0.46 ± 0.04 0.06 ± 0.003
autumn
corm
2.14 ± 0.04
0.49 ± 0.006
0.77 ± 0.03
C. kotschyi
Notes: a Colchicine content is expressed as mass of colchicine in 1 gram dry weight ± standard deviation, derived from the average of two extraction replicates, each run in duplicate
Alkaloids are responsible for the plant adaptation to its environment. It is known that alkaloids are efficiently used as defensive agents and they may be moved around the plant to those parts needing greater protection during growth and development (Harborne, 1997). As part of their defences against herbivores, many geophytes are toxic and unpalatable, or have developed different physical defences against herbivores (Lovegrove & Jarvis, 1986; Go´mez-Garcı´a et al., 2004). This is the case of different plant species of colchicum, which contain colchicine.
three species was found in autumn (the period of root activity). Thus the corms of these three species are better to be collected in autumn from their local habitats, to ensure that maximum of colchicine is achieved. The lowest corm colchicine content in C. robustum (as a synanthous species) was observed in winter (at flowering stage) whereas in C. speciosum and C. kotschyi (as hysteranthous species) in summer, when the corms are dormant. However, more species of colchicum need to be examined to determine a stronger relationship between developmental habit (specially flowering habit) and colchicine content.
CONCLUSION In conclusion, what the results suggest is that the highest corm colchicine content in the
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Source of Support: Nil
Conflict of Interest: None Declared
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Research article ECOLOGICAL AND MEDICINAL INTEREST OF TAZA NATIONAL PARK FLORA (JIJEL - ALGERIA) BOUNAR Rabah1,2*, REBBAS Khellaf2, GHARZOULI Rachid1, DJELLOULI Yamna3and ABBAD Abdelaziz4 1
Department of Biological Sciences, University of Ferhat Abbas Setif 19000, Algeria. Department of Nature and Life Sciences, University of M'Sila 28000, Algeria. 3 Department of Geography, University of Maine, 72085 Le Mans, France. 4 Faculty of Sciences, University Cadi Ayyad, Semlalia, BP 2390, Marrakech, Morocco. 2
*Corresponding Author: E-mail: br_s_dz@yahoo.fr
Received: 08/01/2013; Revised: 26/01/2013; Accepted: 29/01/2013
ABSTRACT The forest of Taza National Park (NP), located in North-Eastern Algeria, is characterized by a high floristic diversity. Analysis of the park flora showed 420 species belonging to 258 genera and 71 botanical families. Asteraceae (54 species), Fabaceae (37), Poaceae (34), Lamiaceae (26) and Brassicaceae (24) are the most dominant families. The endemism rate is around 12.38% (52 species); approximately 21% of endemic species of Algeria. Rare and very rare species were estimated to be 120 taxa representing 28.57% compared to the park flora. Analysis of global phytochoric spectrum shows dominance of native Mediterranean species (193 species). This floristic wealth contains a number of 205 species of medicinal interest. KEYWORDS: Floristic diversity, medicinal plants, Taza National Park, Algeria.
Cite this article: BOUNAR R, REBBAS K, GHARZOULI R, DJELLOULI Y and ABBAD A (2013), ECOLOGICAL AND MEDICINAL INTEREST OF TAZA NATIONAL PARK FLORA (JIJEL ALGERIA), Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 89–101
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INTRODUCTION Algeria, like all Mediterranean countries, has long been involved in the politics of preservation and conservation of biodiversity through the creation of several National Park’s. Currently, it counts eight NP’s including all original landscapes the main hot spots of plant biodiversity in the country (Benhouhou & Vela, 2007). Several research works mainly focused on the identification and mapping of the phytobiodiversity have been made in these hot spots: NP of Chrea (Zeraia, 1981), NP of El Kala (Stevenson et al., 1988; Belouahem et al., 2009), NP of Tlemcen (Yahi et al., 2007; Letreuch-Belarouci et al., 2009) and NP of Gouraya (Rebbas, 2002; Rebbas et al., 2011). These research works underlined the rich flora of these areas and highlighted panoply of endemic and/or rare species which must be placed in conservation priorities. This work also evoked the advanced state of degradation of these natural ecosystems and emphasized the importance of such an inventory list in the rational management of these natural ecosystems. Indeed, several authors evoked that the conservation and the development of a natural ecosystem pass by a good knowledge of its biodiversity (Daget & Poissonnet, 1971; Médail & Quezel, 1997; Véla & Benhouhou, 2007). In order to know the vascular flora of these natural environments, we are interested by the study the floristic diversity of one of the most original ecosystems, at a biogeographic and ecological level, of the Algerian North-eastern sector. It is about Taza NP which belongs to the small Kabylia sector of the Babors (Figure 1) and is regarded as the most wooden area in Algeria with a very high rate (Bensettiti & Abdelkrim, 1990). This work fills the gaps on the state of current knowledge on the vascular flora of the Taza National Park. Indeed, the only known floristic inventory work known in the area and concerned neighborhoods of the Park primarily (Gharzouli 1989; Gharzouli & Djellouli, 2005 Gharzouli, 2007, Bounar, 2003). Only work of
floristic synthesis which refers to all North Eastern Algeria remains very old and not updated (Khelifi 1987; Aouedi 1989; Aktouche et al., 1991). Other research made on some forest formations of the park remains very sketchy. As examples, we can mention phytosociological work of Zeraia (1981), Dahmani (1984) and Bensettiti & Abdelkrim (1990). Knowledge of the diversity of species of medicinal interest of this area allows us to offer solutions for conservation and recovery of these resources within the framework of sustainable development. I- Presentation of the study area Taza NP was created in 1984 on a total area of 3807 ha. It is located in the North-East of Algeria between geographical coordinates 36° 35'–36° 48' North latitude and 5° 29'–5° 40' West longitude. Taking part of the small Kabylia of Babors, it opens onto the Mediterranean Sea in the Gulf of Bejaia (Figure 1). According to the rainfall map established by the National Agency for Water Resources (NAWR, 1996), the study area is situated in annual sections ranging from 850 mm– 1750 mm. Average minimum temperature of the coldest month (January) varies between 6.1° C and 8.1° C. Maximum temperatures of the hottest month (July) is between 30.2° C and 34.8° C. Dry period varies from 3–5 months. High relative humidity of the air (80%) promotes the installation and maintenance of quite important plant diversity. Emberger pluviothermic quotient Q2 (Emberger, 1955) varies between 110 and 124 placing the Park in humid bioclimatic stages to sub-humid with variations to mild and warm winter (Daget & David, 1982). The Park presents a very rugged terrain including several mountain ranges oriented from east to west with altitude varying from 480 m to the highest point in the area (1121 m). These orographic elements give a general configuration in folds in North-eastern and South-western orientations. Geologically, the area is dominated by sedimentary grounds of sandstone and volcanic soils in North zones (Obert, 1970).
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 89–101
Figure 1 Localization map of the study area
These climatic and lithological characteristics determine a rich and diversified flora whose principal forest species are the zeen oak (Quercus canariensis Willd) , which covers more than 40%, the cork oak (Quercus suber L) with 39% and afares oak (Quercus afares Pomel) with only 5% (Bensettiti & Abdelkrim, 1990). According to Maire (1926), Quezel & Santa (1962-1963), Zeraia (1983), Barry et al. (1974), Quezel (1978) and Barbero et al. (2001), Taza NP is on the phyto-geographical region Mediterranean, North African Mediterranean area and belonging to the Numidian. II - METHODOLOGY Park flora was established by floristic surveys carried out, according to the phytosociological method, in different types of vegetation. Surfaces floristically homogeneous were defined on the basis of most common ecological parameters such as altitude, exposure and slope. Covering of the vegetation, by layer, was also taken into account. 63 floristic surveys were carried out. Survey surface varies according to vegetation types. It oscillates between 300–400 m² for forest
vegetation and between 5 and 10 m² for rupicolous vegetation. Surveys were conducted during years 2005 and 2008. The floristic surveys were carried out according to a subjective sampling in all vegetation types of the Park. Samples of plant species collected were determined in laboratory using different flora: Maire (1952-1987), Quezel & Santa (1962-1963), Fennane et al. (1999; 2007) and Valdes et al. (2002). Species nomenclature adopted was according to "MedCheklist, critical inventory of vascular plants of circum Mediterranean countries"(Greuter et al., 1984). Control samples of collected species were deposited in the laboratory of Setif University. Chorologic types of various identified taxa were assigned as indicated in consulted floras; special attention was given to endemic and/or rare species. Analysis of the floral study area and various ethnobotanical fieldwork in the Park surrounding regions, allowed us to have an extensive list of medicinal plants used by the neighboring population.
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III-RESULTS AND DISCUSSION Specific richness Enumerated taxa were 420 species and subspecies belonging to 258 genera and 71 botanical families of vascular plants (phanerogams and vascular cryptogams); approximately 10% of the Algerian total flora estimated at 3139 species (Quezel & Santa, 1962; 1963). Phanerophytes (41 species) occupy 9% of the Park flora. On the total flora recorded at the Park, Asteraceae, Fabaceae, Poaceae, Lamiaceae, Brassicaceae, Caryophyllaceae and Rosaceae were best represented with more than 20 species each. These families represent nearly 40% of the total richness of the Park. Our results are consistent with those of Gharzouli & Djellouli (2005). This wealth places the Park among the most diversified ecosystems in the country, as is the case for all Small Kabylia (Gharzouli, 2007; Vela & Benhouhou, 2007). This floristic wealth of the Park is probably due to (i) its geographical position opening directly on the Mediterranean Sea and therefore exposed to the maritime influences of the North-West (ii) diversity of habitats resulting from climatic and edaphic heterogeneity and (iii) a relatively weaker
exploitation of the medium compared to other ecosystems. Chorological Type Floristic analysis shows the presence of several phytochoric units (Figure 2). Mediterranean one is the most representative with 193 species. This situation is common to most natural ecosystems of Algeria (Quezel, 1964; 2002) and the Mediterranean basin (Dahmani, 1984; Quezel & Barbero, 1990; Quezel & Medail, 2003). This whole Mediterranean is divided into several subsets: s.l. Mediterranean (114 species), western Mediterranean (42 species), Iberomediterranean (20 species), oro-mediterranean (8 species), central mediterranean (2 species) and eastern mediterranean (7 species). Northern chorologic species (Nordic) are relatively well represented in the Park, such as those of european element (20 species), eurasian (41 species), paleo-tempered (22 species), circumboreal (6 species), oro-european (01 species) and atlantic (14 species). Other species correspond to transition elements between chorological mediterranean and those neighbors such as the euro-Mediterranean (30 species), mediterranean-irano-turanian (6 species), macaronesian, mediterranean and asian mediterranean with 4 species each.
Figure 2 Chorological spectrum of Taza National Park
20% 1%
46%
Mediterranean Endemic
21%
Nordic 12%
Paleotropical Wide distribution
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Analysis of endemism 52 taxa were recorded, about 12.38% of total species of the Park and 9.47% compared to the total endemic flora of the country estimated at 549 species (Quezel, 1964) and nearly 12.7% of northern Algeria (Vela & Benhouhou, 2007). Endemism rate is relatively high compared to that recorded in several Parks in central and eastern Algeria such that of Belezma -Batna- (32 species), Gouraya Bejaia- (26 species) (Rebbas, 2002; Rebbas et al., 2011), Djurdjura (35 species) (Meribai, 2006) and Kala -Taref- (75 species) (Stevenson, 1988). Endemic flora of Taza Park consists mainly of endemic Algerian species (18 species), North Africa (22 species), Algerian-Moroccan (5 species), Algerian-Tunisian (7 species). 13.47% of the Park endemic taxa belonged to
Asteraceae and Lamiaceae families with 7 species each. Analysis of the rarity Relying on Quezel & Santa data (1962; 1963) nearly 120 species were reported as rare or very rare. On the basis of these data, the Taza NP records a 28% rarity rate of all its inventoried taxa and around 7% compared to rare species of northern Algeria and about 6.6% over the entire national territory. Compared to the phyto-geographical of Kabylia totaling approximately 487 rare species (Vela & Benhouhou, 2007), Taza NP occupies nearly 24.6% (Figure 3). Among the 129 Algerian taxa Red listed by the International Union for Nature Conservation (1980), 12 species belong to the Taza NP spread over the studied three types of formations (Tables 1 and 2).
Figure 3: Rare Plants in Taza National Park (Photos: K. Rebbas, 2011)
1. Phlomis bovei de NoĂŠ 2. Berberis hispanica Boiss. et Reut.,
3. Atropa belladonna L. 4. Crataegus laciniata Ucria.
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Table 1 : Number of rare and endemic species per botanical family Botanical families
Number of endemic species
Percentage (%)
Number of rare species
Percentage (%)
Asteraceae Lamiaceae Poaceae Caryophyllaceae Brassicaceae Fabaceae Scrofulariaceae Apiaceae Ranunculaceae Crassulaceae Campanulaceae Pinaceae Fagaceae Berberidaceae Geraniaceae Thymelaeaecea Violaceae Cistaceae Primulaceae Convolvulaceae Plantaginaceae Rubiaceae Caprifoliaceae Valerianaceae Linaceae Rosaceae Saxifragaceae
07 07 03 03 03 03 03 03 02 02 02 01 01 01 01 01 01 01 01 01 01 01 01 01 01 -
13.46 13.46 5.76 5.76 5.76 5.76 5.76 5.76 3.84 3.84 3.84 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 1.92 -
16 07 11 09 12 13 04 08 05 03 01 01 02 02 02 01 01 02 04 04 02 07 03
13.33 5.83 9.16 7.5 10 10.83 3.33 6.66 4.16 2.5 0.83 0.83 1.66 1.66 1.66 0.83 0.83 1.66 3.33 3.33 1.66 5.83 2.59
Total
52
120
100
Medicinal plants 205 species of medicinal interest were enumerated. Development of research in field of pharmacology and identification of species active principles will create economic activity in use of plants organized in a friendly safeguard flora. As in the majority of Algerian areas, some of these species are employed by inhabitants bordering the Park as traditional medicine and are marketed by herbalists (Alnus glutinosa L.,
Arbutus unedo L., Asphodelus microcarpus Salzm. & Viv., Asparagus officinalis L., Clematis flammula L., Ceterach officinarum Lamk, Crataegus laevigata (Poiret) DC, Crataegus laciniata Ucria, Mentha pulegium L., Mentha spicata L., Inula viscosa L., Mentha rotundifolia L., Myrtus communis L., Opuntia ficus indica (L.) Mill., Ficus carica L., Pistacia lentiscus L., Prunus avium L., Punica granatum L., Quercus suber L., Juniperus oxycedrus L., Nerium oleander L., Teucrium polium L., Thapsia garganica L., Ulmus campestris L).
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Table 2: Rate rarity by chorological origin Chorological origin
Mediterraneans Mediterranean Western Mediterranean Ibero-Mauritanian Euro-Mediterranean Central Mediterranean East Mediterranean endemics Algerian endemic North African Algerian-Moroccan Algerian-Tunisian Nordics Eurasiatic European Paleo-Temperate Circum-Boreal Oro-European paleotropicals Wide distribution Euro-Mediterranean Atlantic-Mediterranean MacaronesianMediterranean EurasiaticMediterranean Asiatic-Mediterranean Irano-TuranianMediterranean Eurasian-Macaronesian Mediterraneo-SaharanArabian diverse Total
Total number of species
Percentage rate (%)
193 114 42 20 08 02 07 52 18 22 05 07 90 41 20 22 06 01 02 83 30 14 04
Degree of rarity Total species rare and very rare
Percentage rate (%)
45.95
73
37.82
12.38
11
21.15
21.42
19
21.11
0.47 19.78
1 16
50 19.27
100
120
02 04 06 03 02 18 420
Many plants were subject (of) to phytochemical analysis and ethnobotanical studies in North Africa in general and in Algeria in particular. Majority of these plants appear in the floristic list of the study area like: Berberis hispanica Boiss. & Reut., Bupleurum montanum Coss, Cynodon dactylon L., Inula
crithmoides L., Inula viscosa L., Origanum glandulosum Desf., Olea europaea L., Pistacia lentiscus L., Phlomis bovei de NoĂŠ, Salvia verbenaca L., Teucrium polium L. Ricinus communis L (Chemli, 1997; Hmamouchi, 1997; Baba Aissa, 1999; Ruberto et al., 2002;
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Belarchaoui & Boukhadra, 2006; Boulaacheb, 2006; Sari et al., 2006; Hseini & Kahouadji, 2007; Liolios et al., 2007; Benguerba, 2008; Laouer et al., 2009; Hachicha et al., 2009 ; Derridj et al., 2009; Ouled Dhaou et al., 2010; Cahuzac-Picaud, 2010; Makhlouf et al., 2010; El Youbi, 2011; Rebbas et al., 2012; Lemoui et al., 2012; Sari et al., 2012; Hendel et al., 2012).
natural ecosystems, is subject to a worrying degradation. Indeed, human activities (anarchic collection of wood, cork exploitation, uprooting plants of interest) and uncontrolled pasture are seriously detrimental to the richness. To face these problems and to keep the ecological integrity of the Park, an integrated strategy for conservation of biodiversity must be installed.
The anarchy in exploitation of the species known for their therapeutic virtues constitutes a risk for their survival. Certain species are in danger of extinction because of their overexploitation (abusive pulling up). It is the case of Lamiaceae species which are uprooted (torn off with their roots), to be sold in towns and villages of the area, as: Teucrium polium L., Mentha rotundiflolia L., Origanum glandulosum Desf
This strategy must be focused primarily on tree forestation of the Park, especially with zeen oak (Quercus canariensis Willd), cork oak (Quecus suber L) and afares oak (Quercus afares Pomel) which constitute the essential structure of this natural ecosystem. These principal forest formations harbor several endemic and/or rare genera like Cyclamen, Corydalis. Many rare or endangered species of the Park deserve to be integrated in the Red List of the International Union for Conservation of Nature (IUCN). It is about Galium odoratum (L) Scop, Satureja juliana L., Viburnum lantana L., Hieracium ernest Maire, Convolvulus dryadum Maire, Stellaria holostea L, Chrysanthemum fontanesii L., Bupleurum montanum Coss, Quercus afares Pomel and Sedum pubescens Vahl. (Table 3).
CONCLUSION Analysis of the floristic diversity of Taza NP shows well its great richness and its ecological and phytogenetic originality. These data justify its classification with all small Kabylia as a hot spot in northern Algeria (Vela & Benhouhou, 2007). Despite legislative protection, this Park, like most Mediterranean
Table 3 : Rare and endangered species in Taza National Park. Species not listed in the IUCN Red List
Species listed in the IUCN Red List
Galium odoratum (L) Scop Satureja juliana L.
Arabis doumetii Coss. Saxifraga numidica Maire
Hieracium ernest Maire
Teucrium kabylicum Batt.
Viburnum lantana L.
Fedia sulcata Pomel.
Convolvulus dryadum Maire
Carum montanum (Coss & Dur.)Benth.
Stellaria holostea L.
Lonicera kabylica Rehder.
Chrysanthemum fontanesii L.
Teucrium atratum Pomel.
Bupleurum montanum Coss.
Epimedium perralderianum Coss.
Quercus afaresPomel
Phlomis bovei de NoĂŠ.
Sedum pubescens Vahl.
Sedum multiceps Coss & Durieu. Pimpinella battandieri Chabert Moehringia stellaroides Coss. IUCN :International Union for Conservation of Nature
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Increasing ethno botanical studies will allow a better potential understanding of this field, evaluate consequent risks to the use of certain toxic plants and adopt a new management approach for protection and preservation of natural resources (Lahsissene et al., 2010). A large number of spontaneous species of the study area are used in medicine and food like fodder. Culture of these species for economic interest, instead of anarchic gathering, can improve the income of local people while ensuring the conservation of plant diversity (Bounar et al., 2012). For the extraction of active ingredients, the creation of plots of medicinal plants selected, from lists established by floristic inventories, can replace the one gathered. In Algeria, the market for plants with medicinal properties is uncontrolled (Boulaacheb et al., 2006). Considering the various uses of these plants, a regulation seems necessary. So every country must define its own specifications (Veuillot, 2001). Rare and endemic species of the study area form a draft list of local red rare and
endangered flora. The protection and conservation of these formations are needed more than ever before and should receive strict protection. Tourism activities and grazing may be detrimental to the biodiversity of the Park. Urgent solutions must be found to ensure their survival. South of the Mediterranean, where the situation is much more serious, accomplishments are sporadic and generally ineffective. Only authoritative decisions taken by national leaders would likely aim at preserving some ecosystems or certain groups particularly at risk. It is this desire that has been taken in Rabat in 1987, at the meeting for the conservation of plant resources in the countries of North Africa (Quezel et Barbero, 1990). ACKNOWLEDGEMENTS We are very much grateful to all the personnel of Taza National Park; BOUAZID Tayeb (university of Setif); SARI Madani and HENDEL Noui from university of M’sila for their help.
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(Algeria). National Agronomy, 130p.
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Obert, D., (1970). The North Eastern extremity of the Babors chain and Djebel Taounart. Bull. School. Hist. Nat. of Northern Africa. 16 (1–2), 39–47. Ouled Dhaou, S, Jeddi, K., Chaieb, M., (2010). Poaceaein Tunisia: systematic and therapeutic utility. Phytotherapy 8,145– 152. Quezel, P., (1964). Endemism in the flora of Algeria. C. R. Soc. Biogéogr., 361, 137– 149 Quezel, P., (1978). Analysis of the flora of Mediterranean and Saharan Africa. Ann. Missouri Bot. Gard., 65, 479–534. Quezel, P., (2002). Reflections on the evolution of the flora and vegetation in the Maghreb Mediterranean. Paris: Ibis Press, 117 p. Quezel, P., Barbero, M., (1990). Mediterranean forests: problems posed by their historical meaning, ecological and conservation. Acta Bot. Malacit., 15, 145–178. Quezel, P., Medail, F., (2003). Ecology and biogeography of Mediterranean Basin forests. Paris: Elsevier, 573 p. Quezel, P., Santa, S. (1962). Newflora of Algeria and southerly desert regions. Paris: CNRS. 1,1–565 Quezel, P., Santa, S. (1963). New flora of Algeria and southerly desert regions. Paris: CNRS. 2,571–1091 Rebbas, K., (2002). Contribution to the study of vegetation of Gouraya National Park (Algeria): phytosociological study. Magister thesis, Ecology: Ecosystems Management. Setif (Algeria): University of Setif, 100 p. Rebbas, K., Vela, E,. Gharzouli, R., Djellouli,Y., Alatou, Dj., Gachet, S., (2011a). Characterization
phytosociological vegetation of the Gouraya National Park(Bejaia, Algeria). Journal of Ecology (Earth Life), 66,267– 289 Rebbas, K., Boutabia, L., Touazi,Y., Gharzouli, R., Djellouli, Y., Alatou, D., (2011b). Inventory of lichens of the Gouraya National Park(Bejaia, Algeria). Phytotherapy, (9) 4, 225–233. Rebbas, K., Bounar, R., Gharzouli, R., Ramdani, M, Djellouli, Y., Alatou, D., 2012. Medicinal and ecological plants interest in the region of Ouanougha (M'Sila, Algeria). Phytotherapy, 10, 1–12 Ruberto, G., Baratta, M.T., Sari, M., Kaabeche, M., (2002). Chemical composition and antioxidant activity of essential oils from Algerian Origanum glandulosum Desf. Flavour and Fragrance Journal, 17, 251–254 Sari, M., Biondi, D., Kaâbeche M., Mandalari, G., D'Arrigo, M., Bisignano, G., Saija, A., Daquino, C., Ruberto, G. (2006). Chemical composition, antimicrobial and antioxidant activities of the essential oil of several populations of Algerian Origanum glandulosum Desf. Flavour and Fragrance Journal, 21, 890–898 Sari, M., Hendel N., Boudjelal, A., Sarri, Dj., (2012a). Inventory of medicinal plants used for traditional treatment of eczema in the region of Hodna (M’Sila – Algeria). Global J Res. Med. Plants & Indigen. Med., (1) 4, 97–100 Sari, M., Sarri, D., Hendel, N., Boudjelal, A., (2012b). Ethnobotanical study of therapeutic plants used to treat arterial hypertension in Hodna region of Algeria. Global J Res. Med. Plants & Indigen. Med., 1 (9), 411–417 Stevenson, A.C., Skinner, J., Hollis, G.E., Smart, M., (1988). The El Kala National Park and Environs, Algeria: An Ecological Evaluation. Environmental Conservation, 15, 335–348.
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International Union for Conservation of Nature (IUCN).Lists of rare and endangered plants of the Mediterranean basin states. IUCN: Athens, 1980 Valdés, B., Rejdali, M., El Kadmiri, A.A., Jury, S.L., Montserrat, J.M., (2002). Catalogue of Vascular Plants of northern Morocco including identification keys. Biblioteca De Ciencias, Consejo Superior De Investigaciones Cientificas (Csic). Madrid (Edit.), 2 Volumes. 1007 p. Vela, E., Benhouhou, S. (2007).Evaluation of a new hotspot of plant biodiversity in the Mediterranean Basin (North Africa).C.R.Biologics,330,589–605 Veuillot, M. (2001).Plants: uses and legal statutes. TheCourierEnvironment[online],
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44, 23 p. (page consulted on02/12/2012) http://www.inra.fr/dpenv/so.htm#ce44 Yahi, N., Djellouli, Y., De Foucault, B. (2008). Floristic diversity and biogeography of cedar forests of Algeria. Acta Bot. Gallica, 155 (3), 403–414 Zeraia, L., (1981). Trial comparative interpretation of ecological data, phenological and production Suberowoody in the forests of cork oak crystalline Provence (Southern France) and Algeria. PhD thesis. Ecology. Marseille: University of Aix-Marseille, 367 p. Zeraia, L., 1986. Phytosociological study of plant communities foresters of Chréa Park. Cheraga, Algiers: INRF. 1, 23–52. – (Ann. Rech. Forest)
Conflict of Interest: None Declared
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Research article CONSERVATIVE PRODUCTION OF BIODIESEL FROM WASTE VEGETABLE OIL Chethana G.S1*, Reddy K Dayakar2, Vijayalakshmi3 1
Research Associate, R & D, Sri Sri Ayurveda Trust, 21st Km from Bangalore, Kanakapura Road, Bangalore-82, Karnataka, India 2, 3 Department of Biotechnology, Oxford College of Science, HSR Layout, Bangalore, Karnataka, India *Corresponding Author; Email Id: gschethana@gmail.com
Received: 08/01/2013; Revised: 11/02/2013; Accepted: 15/02/2013
ABSTRACT Biodiesel can be made only from oils and fats which are triglycerides and not from any other kinds of oil (such as engine oil). Chemically, triglyceride consists of three long chain fatty acid molecules joined by a glycerin molecule. Waste oil is more appealing than using new oil because refined fats and oils have a free fatty acid (FFA) content of less than 0.1%, in contrary with used and waste oil, where FFA contents are high. FFAs are formed by cooking, the oil longer and hotter the oil has been cooked, the more FFAs it will contain. The study reports on biodiesel production from waste vegetable oil procured from markets where a catalyst (lye) was used to break off the glycerin molecule and combine each of the three fatty acid chains with a molecule of methanol or ethanol, creating mono-alkyl esters, or Fatty Acid Methyl Esters (FAME)—biodiesel. In this process of Transesterification, the glycerin sunk to the bottom and was removed. FFAs interfere with the Transesterification process inhibiting biodiesel formation. With waste oil more lye had to be used to neutralize the FFAs. KEY WORDS: Biodiesel, Waste vegetable oil, Triglyceride, FFAs, Transesterification
Cite this article: Chethana G.S, Reddy K Dayakar, Vijayalakshmi (2013), BIODIESEL PRODUCTION FROM WASTE VEGETABLE OIL, Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 102–109
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INTRODUCTION Petroleum was formed by geologic processes dating from the Cretaceous and Jurassic periods, 90 to 150 million years ago, when vast amounts of zooplankton, algae, and other organic material were deposited on ocean floors. However, the majority of petroleum now extracted in the range of 85% is used to produce fuels. Most of these are transportation fuels such as gasoline, diesel fuel, and jet fuel, while some, such as fuel oil, liquefied petroleum gas, and propane, are used for heating and power generation. Petroleum accounts for more than 90% of transportation fuel, but only for 2% of electricity generation. Increasing worldwide demand for petroleum will affect the transition in important ways. Global petroleum demand is currently at 84 million barrels per day, and it is predicted to increase by 1% to 2% per year, reaching 116 million barrels per day by 2030. Much of this increasing demand will occur in developing nations (Howard Frumkin et al., 2009). Air quality data generated by the Central Pollution Control Board (CPCB) for 2007 under the National Air Quality Monitoring Programme (NAMP) presented deadly facts about air pollution levels in Indian cities. Centre for Science and Environment has analysed the official data to assess the state of air quality and trend in Indian cities. The most widely monitored pollutants in India are particulate matter (PM), nitrogen dioxide (NO2), sulphur dioxide (SO2), and on a limited scale carbon monoxide. Some of the worst forms of air pollutions are found in Indian cities. The Central Pollution Control Board (CPCB) considers air to be ‘clean’ if the levels are below 50 per cent of the prescribed standards for pollutants (Centre for science and environment, 2012). Biodiesel is an alternative fuel source made from renewable resources such as vegetable oil or animal fat, which is simple to use, gives clean burning, biodegradable, non toxic, and essentially free of sulfur and aromatics. Biodiesel is meant to be used in standard diesel
engines and is thus distinct from the vegetable and waste oils used to fuel converted diesel engines. Biodiesel contains no petroleum, but it can be blended with petroleum diesel to create a biodiesel blend. It can be used in diesel engines with no major modifications. Biodiesel is registered as a fuel and fuel additive with the U.S.Environmental Protection Agency (EPA) and meets clean diesel standards established by California Air Resources Board (ARB). Neat (100 percent) biodiesel has been designated as an alternative fuel by the U.S. Department of Energy (DOE) and the U.S. Department of Transportation (DOT) (California energy commission, 2012).Since the passage of the Energy Policy Act of 2005, biodiesel has been increasing in the U.S. In Europe, the renewable Transport Fuel Obligation obliges suppliers to include 5% renewable fuel in all transport fuel in the EU by 2010. This study was undertaken to awaken the utility of waste Vegetable oil which is a trashed product from hotels, canteens etc. which after little chemical treatment can be used as an efficient bio-diesel. Chemically, triglycerides contained in vegetable oil or animal fat consists of three long chain fatty acid molecules joined by a glycerin molecule. Waste oil is more appealing than using new oil because refined fats and oils have a free fatty acid (FFA) content of less than 0.1%, in contrary with used and waste oil, where FFA contents are high. FFAs are formed by cooking, the oil longer and hotter the oil has been cooked, the more FFAs it will contain. Hence this study was conducted to use these waste oils where lye was used to break the Glycerin chain. MATERIALS (Keith Addison, 2012)
1 liter of fresh vegetable (sunflower) oil and waste vegetable oil from a local canteen was procured. 4.5 g of potassium hydroxide (also known as lye) 200 ml of ethanol (ethyl alcohol) 10 ml isopropyl alcohol Glass or plastic container that is marked for 1 liter
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The production of biodiesel (Keith Addison, 2012)
METHODOLOGY Basic titration
For processing used oil it is essential to titrate the oil to determine the free fatty acid (FFA) content and calculate how much extra lye will be required to neutralize it. Phenolphthalein indicator was used. 1g of pure Potassium Hydroxide lye (KOH) was dissolved in 1 liter of distilled water (0.1% W/V KOH solution) In a small beaker, 1 ml of dewatered waste vegetable oil was dissolved in 10 ml isopropyl alcohol. The beaker was gently warmed on a hot water bath; stirred until all oil dissolved in the alcohol and the mixture turns clear. 2 drops of phenolphthalein indicator was added. Using graduated syringe, 0.1% KOH solution was added drop by drop to the Oil-alcoholphenolphthalein indicator, stirring all the time, kept stirring. The lye solution was added until the solution stays pink for 15 seconds. The number milliliters of 0.1% lye solution used was noted and added to the 3.5 grams of lye (the basic amount of lye needed for fresh oil). So the total quantity of lye used to process the Waste vegetable oil per liter is 4.5 gms (Venkata Ramesh Mamilla et al., 2011; C.V. Sudhir et al., 2007).
Fresh Sunflower oil & Waste Vegetable oil were taken to which the amount of catalyst to be added was calculated as 4.5 for both. 200 ml ethanol was poured into glass blender pitcher. Blender was turned on to its lowest setting and slowly 4.5 g of potassium hydroxide (lye) was added. This reaction produced potassium methoxide. Ethanol and potassium hydroxide was mixed until the potassium hydroxide has completely dissolved (about 2 mins), 1 liter of waste vegetable oil was added to this mixture. Similar procedure was followed for new vegetable oil. This mixture (on low speed) was blended continuously for 20 mins to 30 mins. After completing the procedure the oils were kept for observation. The bottle of oil was kept for 2 days, uncovered inside a rack.
Purification step Purification of the resultant bio-diesel was done in accordance with the method explained by Y. Zhang et al., 2003 & Arjun B. Chhetri et al., 2008.
Figure 1: The apparatus used for Biodiesel production
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The confirmatory test:
Methanol test
Wash test
25 ml of biodiesel was dissolved in 225 ml of methanol in a measuring glass. The biodiesel got dissolved completely in methanol. ―The biodiesel should be fully soluble in the methanol, forming a clear bright phase (figure 6) (Jan Warnqvist, 2005).
150 ml of unwashed biodiesel (settled for 12 h or more, with the glycerin layer removed) was taken in a half liter glass jar or PET bottle. 150 ml of water (at room temperature), was added. Screwed the lid on tight and shaken it up and down violently for 10 seconds and was let to settle (figure 2) (Keith Addison, 2012).
Figure 2: Wash test for biodiesel
Figure 3: Picturing shows Methanol test carried out for biodiesel, along with biodiesel
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Figure 4: The total amount of Biodiesel obtained
RESULTS
Wash Test
Production results
The biodiesel should separate from the water in half an hour or less, with amber (and cloudy) biodiesel on top and milky water below. After a violent 10-second shaking; biodiesel and water separated cleanly within minutes. This is quality fuel, a complete product with minimal contaminants. It was observed that the clear water was at the bottom and biodiesel was on the top. This indicates the positive result for biodiesel for wash test. This tells that the biodiesel got purified, that is the oil which underwent incomplete reaction was removed by wash test. The biodiesel which is purified stands on the top leaving clear water at the bottom (figure 5).
Biodiesel was obtained after processing the waste vegetable oil and new sunflower oil. After 2 days of observation, it was observed that the biodiesel was on top of the glycerin, which settled at the bottom. The amount of biodiesel obtained from waste vegetable oil was 540 ml from 1 liter and 850 ml of biodiesel from 1 liter sunflower oil (figure 4). The purification step
This step was done by washing biodiesel with water. This was done to remove the impurities and the incomplete reaction products like soap. 10 ml of normal tap water was added to 100 ml of biodiesel, shaken vigorously, allowed for some time and the water was removed. This was done until we got clear water indicating that most of the impurities were removed.
Results of Methanol test: Biodiesel dissolves easily in methanol, where as vegetable or animal oils and fats (triglycerides) does not dissolve in methanol. Any uncovered oil left in the biodiesel will settle out at the bottom of the tank. 25 ml of biodiesel was added in 225 ml of methanol. A clear solution indicates a positive result for biodiesel (figure 6).
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Figure 5: Positive result for biodiesel by wash Test
Figure 6: Confirmation of biodiesel by Methanol test
DISCUSSION Biodiesel is an alternative fuel similar to conventional or fossil diesel. Biodiesel can be produced from straight vegetable oil, animal oil/fats, tallow and waste cooking oil. The process used to convert these oils to biodiesel is called Transesterification (Ulf Schuchardt et al. 1997). Biodiesel has many environmentally beneficial properties. The main benefit of biodiesel is that it can be described as ‘carbon neutral’. This means that the fuel produces no net output of carbon in the form of carbon dioxide. The (figure 7) below shows the chemical process for methyl ester biodiesel. The reaction between the fat or oil and the alcohol is a reversible
reaction and so the alcohol must be added in excess to drive the reaction towards the right and ensure complete conversion.
Mixing of alcohol and catalyst The catalyst used was typically potassium hydroxide. It was dissolved in alcohol which acts and enhances the reaction with the oil to form esters (figure 7) which is nothing but the crude biodiesel which is in compliance with the study done by Venkata Ramesh Mamilla et al., 2011. Excess of catalyst was used to convert the oil completely into esters. The reaction happens with vigorous agitation, done using a mixer. The recommended reaction time was 20 minute to 1 hour. The so formed biodiesel is kept ideal for 24 to 48 hours under observation.
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Figure no 7: Transesterification of Fatty acids
Our results on separation step are similar to the study conducted by Y. Zhang et al., 2003) which shows the clear separation of the biodiesel on the top, from the glycerine at the bottom which is much denser than the biodiesel. Once its separated from glycerine, biodiesel is is sometimes purified by washing with warm water to remove residual catalyst or soaps, dried and sent to storage, which marks the end of the process. The process results in the clear amber-yellow liquid.
pollution caused by petroleum products because Biodiesel is a biodegradable, non toxic and virtually free from sulfur and aromatics. A number of studies have found that biodiesel biodegrades much more rapidly than conventional diesel. In this respect, its action is similar to petroleum diesel fuel. However, biodiesel does not have the toxicity and the solvent action that diesel fuel has, so its effects on animals are expected to be less severe. A lot of research and development is needed in this aspect to make the biodiesel easily available to everyone.
CONCLUSION
ACKNOWLEDGEMENTS
In the present situation where the natural resources in the form of fossil fuel are getting exhausted, it has become very important to think the alternate source of energy. So biodiesel is one of the alternate solutions which are ecofriendly. It is also advantageous over the
Authors are thankful to the Head of the Department, Biotechnology, Dr.Vedamurthy Anakalbasappa and other lecturers for their support in completion of this work successfully.
Separation
REFERENCES Arjun B. Chhetri, K. Chris Watts and M. Rafiqul Islam (2008), Waste Cooking Oil as an Alternate Feedstock for Biodiesel Production., Energies; 1, 3– 18; DOI: 10.3390/en1010003
C.V. Sudhir, N.Y. Sharma and P.Mohanan., (2007) Potential of waste cooking oils as biodiesel feed stock., Emirates Journal for Engineering Research, 12 (3): 69–75
California energy commission, (2012), California energy commission retrieved from http://www.consumerenergycenter.org/t ransportation/afvs/biodiesel.html
Gerhard Knothe, Robert O. Dunn and Marvin O. Bagby, (1997) The Use of Vegetable Oils and Their Derivatives as Alternative Diesel Fuels. Oil Chemical Research, National Center for Agricultural Utilization Research, Agricultural Research Service, U.S. Department of Agriculture, Peoria, Volume: 666; Chapter, 10; pp-172–208
Centre for science and environment, (2012), Air pollution, State of Air Pollution in Indian cities, retrieved from http://www.cseindia.org/node/207
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Howard Frumkin, MD, Dr P HJeremy Hess, MD, MPH Stephen Vindigni., (2009), Energy and Public Health: The Challenge of Peak Petroleum., Public Health Rep; 124(1): 5–19. Jan
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Warnqvist, (2005). AGERATEC AB Biofuel mailing list Re: Quality Test, retrieved on 12.12.2012 from http://www.mwilarchive.com/biofuel@sustainablists.org/ msg53363.html Addison (2012), JOURNEY TO FOREVER, HONG KONG TO CAPE TOWN TO OVERLAND, retrieved from http://www.Journeytoforever.org
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Ulf Schuchardt , Ricardo Serchelia, and Rogério Matheus, (1998). Esterification of Oils :a review., J Braz, Chem. Soc. Vol 9, No. 1, 199–200. Venkata Ramesh Mamilla, M.V. Mallikarjun,Dr.G Lakshmi Narayana Rao., (2011), Preparation of Biodiesel from Karanja Oil., Internal journal of Mechanical & Production Engineering Research & Development.Vol 1. (1): 51–69., Y. Zhang, M.A. Dubee, D.D. McLean, M. Kates. (2003) Biodiesel production from waste cooking oil: 1. Process design and technological assessment., Bioresource Technology 89:1–16.
Conflict of Interest: None Declared
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 110–117 ISSN 2277-4289 | www.gjrmi.com | International, Peer reviewed, Open access, Monthly Online Journal
Research article PHYTOCHEMICAL STUDIES ON SMILAX MACROPHYLLA LINN.; A SOURCE PLANT OF CHOPACHEENI Jyothi T1*, Acharya Rabinanaryan2, Shukla C P3, Harisha CR4 Research assistant, ALN Rao Memorial Ayurvedic medical college, Koppa – 577126, Chikkamagalur District, Karnataka, India 2 Associate Professor, Department of Dravyaguna. IPGT& RA, Gujarat Ayurved University, Jamnagar361008, Gujarat, INDIA 3 Head, Department of Pharmaceutical Chemistry, IPGT& RA, Gujarat Ayurved University, Jamnagar361008, Gujarat, INDIA, 4 Head, Pharmacognosy Laboratory, IPGT& RA, Gujarat Ayurved University, Jamnagar- 361008, Gujarat, INDIA *Corresponding Author: ayurjyothi.n@gmail.com 1
Received: 10/01/2013; Revised: 02/02/2013; Accepted: 05/02/2013
ABSTRACT Chopacheeni is an important herbal drug widely used in Ayurveda. Chopacheeni is one among in the red list and in top 20 highly traded medicinal plants in India. Commonly known as Sarsaparilla, various species are available in the market in the name of Chopacheeni and are rarely Smilax china, the official source. The plant is considered as a remedy for Syphilis, Rheumatism, Skin diseases and Gout. Botanically authenticated drug is Smilax china but due to unavailability many adulteration is coming in the market to avoid this it is attempt to look for substitute of the drug of same genus so Smilax macrophylla Linn. was used for the phytochemical analysis. TLC of alcoholic extract of drug on silica gel "G" plate using Toluene (6.5 ml): Ethyl acetate (3.5 ml): Glacial acetic acid (0.2 ml) showed one spot under 366 nm UV, in 254 nm UV no spots, After spraying with Liebermann Burchard reagent followed by heating and then was visualized in day light which showed 2 prominent spots are seen.TLC using Chloroform (9.5 ml): Methanol (0.5 ml) showed two spots under 366 nm UV ,in 254 nm UV no spots were seen. After spraying, it showed one prominent spot. In HPTLC chromatogram showed 2 prominent spots in short wave UV 254 nm, one prominent spot in long wave UV 356 nm and 3 prominent spots at 400 nm. KEYWORDS: Smilax macrophylla, Chopacheeni, Madhusnuhi, Phytochemical Analysis
Cite this article: Jyothi T, Acharya Rabinanaryan, Shukla C P, Harisha CR (2013), PHYTOCHEMICAL STUDIES ON SMILAX MACROPHYLLA LINN.; A SOURCE PLANT OF CHOPACHEENI, Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 110–117
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INTRODUCTION Smilax china Linn. of the Family Smilacaceae) is an important herbal drug widely used in Ayurveda. It is used for the treatment of Phiranga roga, Upadamsha, Vatavyadhi, Vrana (Vaidya Bapalal, 2005). It is said that the drug is similar to Ashwagandha (Sharma PV, 2005) in its properties and action. It is one among the red listed plants and in top 20 highly traded medicinal plants in India (http://www.megforest.gov.in). Now a day, due to commercialization and other economic interests, the pharmaceutical industry uses various source plants for the drug Chopacheeni. The availability of genuine samples is a burning issue since it is enlisted as endangered species (www.iucnredlist.org) and demand surpasses production causing confusion in the end users about the quality and safety also therapeutic ambiguity. Various species of Smilax are used as source plant for Chopacheeni; S. China Linn. , S. macrophylla Linn., S. zeylanica Linn. , S. regelli to name a few. S. china is formerly considered as the authentic identification as per The Wealth of India (Anonymous, 1950). Due to non availability and possible adulteration of cheaper substitutes, the therapeutic efficacy is obscure. Hence there is an urgent need to explore the raw drugs for their quality through phytochemical investigations to establish the authenticity and logical reasoning behind multiple source plants of Chopacheeni. In the current research, roots and rhizomes of Smilax macrophylla Linn. which is a potential source for Chopacheeni was evaluated phytochemically for available active ingredients and their strength. Pharmacognostical investigation with macerate and powder study details and HPTLC finger printing of the rhizome and roots of Smilax macrophylla Linn. which helps in identification of crude drug is not available. Hence the present study has been carried out with following Aims & Objectives; Pharmacognostical and Phytochemical analysis of Smilax macrophylla Linn. The vernacular names (Prajapati ND et al., 2003) of Chopacheeni (Smilax china) are:
Sanskrit- Chopacheeni, Dvipantharavacha, Madhusnuhi (Chopra RN et al., 1956); EnglishSarsaparilla, China root; Hindi- Jangli, Austibab, Chopachini; Marathi- Guti; TamilMalaith, Tamarai, Parangichekkarda; TeluguKondata mora, Malkaltamora; KannadaKaduhambu; Gujarati- Chopachini; AssameseAslussini; Bengali- Topachini Smilax macrophylla Linn. is a large more or less prickly climber (Haines HH, 2000) growing in Himalaya eastwards from Kumaon at Assam, Bengal, Burma & South to Central Konkan extending to Java. The Stem is Smooth, striate (lines or several angled), armed with a few small distant prickles or almost unarmed; roots are rope-like originate from a short rhizome (Haines HH, 2000). MATERIALS & METHODS Plant Material: Fresh roots and rhizomes were collected from the forests of Odisha during the month of November 2009. Botanical identification was done by expert taxonomists using local floras (Haines HH, 2000) and found to be Smilax macrophylla Linn. Voucher Ref no. IPGT&RA - 301). The collected samples were washed with potable water and chopped in to small pieces which were dried in shade, powdered and used for scientific evaluation.
Pharmacognostical Study (Kokate CK, et al., 2008, Anonymous 2000, Trease and Evans, 2009): Morphological, Organoleptic, Microscopic and Histochemical study of the powdered drug was done as per the guidelines of Ayurvedic pharmacopoeia of India.
Phytochemical study (Kasture AV et al., 2009, Anonymous 2000, Stahl E. 2005): Smilax macrophylla Linn. were analyzed by Physicochemical, Qualitative, Quantitative parameters. Chromatographic fingerprinting and Ultra-Violet Spectroscopy were carried out. TLC is mentioned as a primary tool for identification as part of monographs on all medicinal plants. Methanolic extract was used for the spotting of the TLC plate
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(Silica gel G Pre-coated plates). The solvent systems used in the study were Chloroform: Methanol and Toluene: Ethyl acetate: acetic acid. The sample extract was made to run on silica plate in various ratios. The ratio of 9.5:0.5 and 6.5:3.5:0.2 respectively has given good separation on trial method. Hence these systems are adopted for Chromatographic evaluation of the sample.
Acetic acid was added for the second system for better separation. The resulting TLC pattern was viewed under long wave UV light at 366 nm and Short wave at 254 nm. Then after spraying with the Liebermann Burchard reagent and drying in a hot air oven and the number of spots viewed under daylight (Table no.4).
Picture No. 1 Pharmacognostical Study of Smilax macrophylla Linn
Picture No. 2 Pharmacognostical Study of Smilax macrophylla Linn.
A.Rhizome, B. Root, C. Cork, D. Acicular Crystals, E. Scalariform Vessels, F. Scleroids, G. Tannins & Starch Grains, H. Fiber, I. Parenchyma, J. Tracheids, K. Pitted Vessels, L. Reticulate Vessel
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chloroform, acetone soluble extractive values and for pH value (Table No. 1). The percentage of moisture content was 9.40%w/w, total ash 2.45%w/w, acid insoluble ash 0.15%w/w; Water soluble extractives 31.25%w/w, Alcohol soluble extractives 19.30%w/w, Chloroform soluble extractives 0.1%w/w and Acetone soluble extractives 8.58%w/w. pH was 5.28. Low total ash and Acid insoluble ash signifies low levels of inorganic matter and silica content. The high solubility of the sample in water denotes that drug is best suited for extraction with water or water based preparations. The negligible presence of Volatile oils is also in favor of thermal extractions with water.
RESULTS & DISCUSSION: Pharmacognostical Study: (Picture No.2)
Morphological Study: Rhizomes were 5–6 × 3–4 cm in size, Root 20–23 × 1.5–2 cm in length Cylindrical & tapering towards apex, externally brownish and internally pinkish in color with rough and woody surface, and fracture coarsely fibrous.
Organoleptic characters: The powder was reddish brown in color, with characteristic odor, slight bitter in taste and fibrous in texture.
Powder Microscopy: The dried powder was mounted with distilled water to detect the Starch Grains, Cork, Simple Fiber, Acicular crystals, Scalariform vessels, Scleroids, Reticulate Vessels, Pitted Vessels, Parenchyma, Tracheids and Tannin. When stained with Iodine solution, Dil. FeCl, Conc. HCl and Phloroglucinol with Conc. Hcl, Showed Starch grains (Blue), Tannins (Bluish black), Crystals (Effervescence) and Lignified cells (Pink) respectively.
Physicochemical Parameters: The sample was evaluated for physicochemical parameters like Loss on drying, Total Ash value, Acid insoluble ash, water, alcohol,
Qualitative chemical tests: Qualitative chemical tests for different functional units were estimated using water, methanol, chloroform and acetone soluble extractives of Smilax macrophylla Linn. Carbohydrates, Reducing sugars, proteins, Amino acids, saponins, alkaloids, Flavonoids and Tannin were qualitatively investigated. All functional units were present in water soluble extractive except amino acids and Flavonoids as these are the basic functional units necessary for metabolism in herbs. (Table No.2)
Table No: 1 Physico Chemical Parameters Sr. No. 1 2 3 4 5 6
Parameter Loss on Drying Total Ash Acid Insoluble Ash Water Soluble Extractives Alcohol Soluble Extractives pH
Smilax macrophylla Linn. 9.40%w/w 2.45%w/w 0.15%w/w 31.25w/w 19.30%w/w 5.28
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Table No. 2 Qualitative chemical tests of Smilax macrophylla Linn. for different functional units in various solvent systems Sr. No. 1 2 3 4 5 6 7 8
Test
Water
Methanol
Carbohydrates-Molish’s test Reducing Sugars-Fehling’s test Proteins-Biuret test Amino acid- Ninhydrin test SaponinsFoam test Flavonoids-Shinoda test Alkaloids- Wagner’s test Tannins- FeCl3
Positive
Positive
Positive
Positive
Strongly Positive
Strongly Positive Negative Negative Positive
Positive
Positive
Negative Negative Negative
Negative Negative Negative
Positive Positive Positive
Negative Negative Negative
Negative Negative Positive
Positive Negative Moderately Positive Negative Positive Strongly Positive
Chloroform Acetone
Table No.3 Quantitative estimation Sr. No. 1. 2 3 4
Parameter Total Volatile oils Total Alkaloids Total Tannins Total Saponins
Quantitative estimation: Traces of total Volatile oils, 0.08%w/w of total Alkaloids, 8.28%w/w of total Tannins and 22.85%w/w total Saponins were observed in the sample. Saponins are high molecular weight glycosides, consisting of a sugar moiety linked to a triterpene, steroid or steroid alkaloidal aglycone (Natural Remedies). Aglycone portion of the saponin is called as sapogenin. Triterpene saponins are the most common type in the plant kingdom. They show hemolytic activity and have bitter taste. Majority of the pharmacological and clinical action may be linked to these saponins in the case of Chopacheeni. (Table No.3).
Thin Layer Chromatography Study: Thin layer chromatography of Methanol Extract of Smilax macrophylla Linn. Powder of the sample weighing 5 g were
Smilax macrophylla Linn. Trace 0.08%w/w 8.28%w/w 22.85%w/w
taken with 100 ml of alcohol and kept for twenty-four hours. Filtrate was prepared and evaporated till it was dried in a flatbottomed shallow dish and concentrated on water bath to volume of requirement. TLC of alcoholic extract of drug on silica gel "G" plate using Toluene (6.5 ml): Ethyl acetate (3.5 ml): Glacial acetic acid (0.2 ml) showed one spot under 366 nm UV at Rf 0.23. Where as in 254 nm UV no spots were seen. After spraying with Liebermann Burchard reagent followed by heating and then was visualized in day light which showed 2 prominent spots at Rf 0.70 and 0.82. TLC using Chloroform (9.5 ml): Methanol (0.5 ml) showed two spots under 366 nm UV at Rf 0.04 and 0.83. Where as in 254 nm UV no spots were seen. After spraying, it showed one prominent spot at Rf 0.83.
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Table No. 4 Thin Layer Chromatography of Smilax macrophylla Linn. Mobile Phase
Detection condition
Chloroform: Methanol (9.5:0.5)
254 nm UV 366 nm UV After spray with Liebermann Burchard reagent 254 nm UV 366 nm UV After spray with Liebermann Burchard reagent
Toluene: Ethyl acetate: acetic acid (6.5:3.5:0.2)
No.of Spots 0 2 1
Rf Value 0.04, 0.83 0.83
0 1 2
0.23 0.70,0.82
Table No. 5 High Performance Thin Layer Chromatography of Smilax macrophylla Linn. Mobile Phase
Detection condition
Chloroform: Methanol (9.5:0.5)
254 nm UV 366 nm UV 400 nm UV
No.of Spots 2 1 3
Rf Value 0.08, 0.56 0.08 0.10, 0.28, 0.70
Table No.6 UV Spectrophotometry of Smilax macrophylla Linn. Sample
Peak
Absorbance
1
Wavelength in nm 236
Smilax macrophylla Linn.
1
275.6
3.464
Picture No. 3 HPTLC of Smilax macrophylla Linn.
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3.161
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Picture No. 4 UV Spectrophotometry of Smilax macrophylla Linn
High-Performance Thin Layer Chromatography Study: Methanol extract of Smilax macrophylla Linn. was spotted on pre-coated silica gel GF 60254 aluminium plate as 5 mm bands, 5 mm apart and 1 cm from the edge of the plates, by means of a Camag Linomate V sample applicator fitted with a 100 μL Hamilton syringe. Chloroform (9.5 ml) and Methanol (0.5 ml) (v/v) (20 ml). The Chamber was saturated for 45 min, Development Time taken was 20 min and the development distance was 4.8cm. After development, Densitometric scanning was performed with a Camag T.L.C. scanner III in reflectance absorbance mode at 254 nm, 366 nm and 400 nm under control of win CATS software (V 1.2.1 Camag) (Picture No.2). The slit dimensions were 6 mm x 0.45 mm and the scanning speed was 20 mm s-1. However, chromatogram showed 2 prominent spots at Rf 0.08 and 0.56 in short wave UV 254 nm, one prominent spot at Rf 0.08 in long wave UV 356 nm and 3 prominent spots at 0.10, 0.28, 0.70 at 400 nm. (Table No.5 and Picture No. 3)
UV Spectrophotometry: The spectrum was measured by placing the sample solution into the Shimadzu UV-160 Double beam spectrophotometer. Based on the UV Spectrophotometric analysis, the peaks, wavelengths and absorbance are shown in Table No.6. (Picture No.4)
CONCLUSION Presence of more acicular crystals, Scalariform vessels, Scleroids, Reticulate Vessels, Pitted Vessels, is the identified character of Smilax macrophylla. The preliminary phytochemical analysis of the rhizome and root of Smilax macrophylla revealed the presence of Carbohydrates, Reducing sugar, saponin, protein, alkaloids and Tannins. The sample has got highest solubility in water followed by methanol. Hence drug is best suited for extraction with water or water based preparations. The Chromatographic finger printing was developed which could be useful for researchers to carry out further researches. The study is expected to be useful for quality control of sample and also will be a useful guide in deciding the source for Chopacheeni looking in to lack of availability and endangered status of the official source plant Smilax china Linn.
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REFERENCES Anonymous. (2000), Protocol for testing of Ayurveda, Siddha & Unani medicines. Ghaziabad: Pharmacopoeial laboratory for Indian medicines, Department of AYUSH, ministry of health & family welfare, Government of India. Anonymous. (1950), The wealth of India, Vol. VIII. New Delhi: Council of Scientific and Industrial Research;. pp 365. Chopra RN, Nayar SL, Chopra IC, (1956), Glossary of Indian Medicinal Plants. New Delhi: Council of Scientific and Industrial Research; pp 228. Haines HH. (2000), Botany of Bihar and Orissa. Part V-VI. Dehradun: Bishan singh Mahendrapal Singh; pp 1085. http://www.megforest.gov.in/activity_medic_pl ants.htm accessed on 3/23/2011 IUCN (2009), IUCN Red List of Threatened Species. Version 2009.2. <www.iucnredlist.org>. Downloaded on 01/29/2010 Kasture AV, Mahadik Kr, More HN, Wadodkar SG. (2009), Pharmaceutical Analysis. 18th Ed. Vol.II, Pune: Nirali Prakashan
Source of Support: Nil
Kokate CK, Purohit AP, Gokhale SB. (2008), Chapter 6. Pharmacognosy. 42nd Ed. Pune: Nirali Prakashan; pp 6.1–6.45. Natural
Remedies, Master Document on Tribulus terrestris, Quality control department, Natural Remedies, Bangalore
Prajapati ND, Purohit SS, Sharma AK, Kumar T, (2003), A Handbook of Medicinal Plants – A Complete source book, 1st Ed. Jodhpur: Agrobios; pp 477. Sharma PV, (2005), Dravyaguna Vijnana, Vol. 2, Varanasi, Chaukhambha Bharatiya Academy; 804 Stahl E. (2005), Thin-Layer Chromatography. 2nd Ed. Berlin: Springer; Taylor, L., The Healing Power of Rainforest Herbs downloaded from http://www.rain-tree.com/book2.htm. On 10/29/2010. Trease and Evans. (2009), Pharmacognosy. 16th Ed. New York: Elsevier Saunders; pp 309–10. Vaidya Bapalal (2005), Nighantu Adarsha, Varanasi, Chaukhambha Orientalia,Vol. II: 643. Conflict of Interest: None Declared
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 118–125 ISSN 2277-4289 | www.gjrmi.com | International, Peer reviewed, Open access, Monthly Online Journal
Review article SELECTION OF MEDICINAL PLANTS FOR THE MANAGEMENT OF DIABETIC FOOT ULCER; AN AYURVEDIC APPROACH Pampattiwar S P1*, Adwani N V2, Bulusu Sitaram3, Paramkusa Rao M4. P.G. Scholar – final Year, P.G. Dept. of Dravyaguna, T.T.D‟S S.V. Ayurveda College, Tirupati, Andhra Pradesh, India 3, 4 Professor (UG), Dept. of Dravyaguna, T.T.D‟S S.V. Ayurveda College, Tirupati, Andhra Pradesh, India *Corresponding Author: E mail: vd.yash.728@gmail.com; Mobile: +91 9700307493 1, 2
Received: 05/01/2013; Revised: 08/02/2013; Accepted: 10/02/2013
ABSTRACT Diabetic foot ulcer is one of the major complications of Diabetes mellitus. It can lead to amputation of leg also. Diabetes mellitus is one such metabolic disorder that impedes normal wound healing because of altered protein and lipid metabolism and abnormal granular tissue. This literary review was done to provide an effective management in cases of non healing ulcer. It is proposed that for the treatment of such patients common herbs explained in “Prameha Chikitsa”, “Kushtha Chikitsa” and “Vrana Chikitsa” can be useful. This is a new approach by which one can select the herbs for the treatment of diabetic foot ulcer. Due to this approach new formulations can be formulated for diabetic foot ulcer which can be beneficial to them. A list of drugs mentioned in treatment of above diseases is prepared from Ashtanga Hridaya and their activity checked with ongoing clinical research. KEYWORDS: Diabetic foot ulcer, Prameha, Kushtha, Vrana ABBREVIATIONS: A.H. SU- Ashtanga Hridaya Sutrasthana; WHO – World Health Organization
Cite this article: Pampattiwar S P, Adwani N V, Bulusu Sitaram, Paramkusa Rao M (2013), SELECTION OF MEDICINAL PLANTS FOR THE MANAGEMENT OF DIABETIC FOOT ULCER; AN AYURVEDIC APPROACH, Global J Res. Med. Plants & Indigen. Med., Volume 2(2): 118–125
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INTRODUCTION Diabetic foot is the most common complication of diabetes greater than retinopathy, neuropathy, heart attack and stroke combined. (Marvin E Levin, 1994) According to WHO, the foot of Diabetic patient has potential risk of pathological consequences including infection, ulceration and or dysfunction of deep tissues associated with neurological abnormalities, various degrees of peripheral vascular disease and metabolic complications of diabetes in lower limb. (Robert G Frykberg, 2000) The Diabetic foot is essentially vulnerable to amputation because of frequent complications of peripheral neuropathy (PN), infection and peripheral arterial disease (PAD). (Marvin E Levin, 1994) A combination of this triad leads to gangrene and amputation. Most of these are result of PN (peripheral neuropathy) and insensate foot which leads to painless trauma and ulceration. The relation between Diabetic neuropathy, the insensitive foot and foot ulceration was recognized by Pryce, a century ago. He stated that, “It was abundantly evident that the actual cause of perforating ulcer was peripheral nerve degeneration and that diabetes itself played active part in causation of perforating ulcer”. (Pryce TD, 1887) For instance wound infection has been one of the major impediments in the process of wound healing and after invention of antibiotics; it was thought that this problem would be conquered. Since then several antibiotics in form of systemic and local use have been tried but problems of wound healing remains as such. Apart from this, antibiotics have their adverse side effects. There are list of complications occurring as a result of taking hypoglycemic drugs like chest pain, irregular heartbeat, difficulty in breathing and erectile dysfunction. (Seppo Lehto, 1996). To avoid above complications, it would be better to go with herbal drugs for the management of Diabetic foot. Keeping in view
of aforesaid problems, ancient literature was explored to throw light regarding the wound and its management with the help of medicinal plants. MATERIALS Though the direct description of Diabetic foot ulcer is not available in Samhitas but it is found in „Vrana Paratishedha Adhyaya‟ of AstangaHridaya (Tripathi, 2007). Here Vagbhata denoted that if the patient of Vrana (ulceration) is suffering from Kushtha (skin disease) or malnutrition or poisoning or Prameha (diabetes) then that Vrana (ulcer) is difficult to treat. This quotation clearly verdicts the reference of Diabetic foot ulcer has been manifested and said to be almost incurable. Pathogenesis sequence of diabetic foot ulcer Classically all the Hetus (causative factors) described for Prameha are responsible for vitiation of Kapha, Mutra, Meda. So, Prameha is vitiated Kapha predominant disease (Tripathi, 2007). Vitiated Kapha vitiates Kleda (moisture), Sweda (sweat), Medo-dhatu (fat), Rasa Dhatu (plasma) and Mamsa Dhatu (muscle) in body. As vitiated kapha vitiates Mamsa Dhatu (muscle) it loses its „SAARATA‟ (consistency). This thing creates problem in healing of Diabetic foot ulcer. „Twachi Swapa‟ (numbness) and „Vrananam Shighrotpatti Chirastithi‟ (immediate onset of ulcers and become chronic), these two symptoms are present in „Kushtha nidana‟ (etiological factors of Skin diseases) as Purvarupa (Premonitory symptoms) of Kushta (Tripathi, 2007). These two can be correlated with insensate foot and rapid spreading with fixed nature symbolising necrotising fascitis involving deeper related tissues. Hence, vitiation of Rakta Dhatu (blood) occurs in Diabetic foot. All these references denote various complications of Diabetic foot ulcer with bad prognosis nature. There are some common factors found in „Prameha‟ (Diabetes), „Kushta‟ (skin disease) and „Vrana‟ (ulcer) with respect to Hetus (causative factors), Doshas (humors) and Dushyas
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(Vitiated tissue). The common causative factors are intake of excess curd, new cereals, jaggary, meat and milk. The common vitiated dosha (humors) in Prameha and Kushta is mostly Kapha and common vitiated tissues are Rakta (blood), Mamsa (muscles) and Lasika (lymph). From above description, it is clear that there is much similarity present between Prameha, Kushtha, and Dushta vrana. Therefore for the treatment of Diabetic foot ulcer common herbs described in treatment of foresaid diseases should be selected. Vitiation of Kleda (moisture) is also an important factor in the Samprapti (pathology) of Diabetic foot. Selection of herbal drugs for the treatment of Diabetic foot ulcer depends on following treatment principles like Vrana shodhana (purifying the wound), Vrana ropana (wound
healing purpose) and blood purifier. (Tripathi, 2007) Common herbal drugs used in the treatment of Prameha, Kushtha and Vrana In the treatment of Prameha pidika, Eladi gana is used for Vranaropana (wound healing purpose), Aragwadhadi gana is used for Udwartrana (rubbing herbal powder against body), Asanadi gana is used for Parisheka (pouring) and Vatsakadi gana is used for internal administration. (Tripathi, 2007). Surasadi and Aaragwadhadi ganas are indicated for Kshalana purpose (Tripathi, 2007). These ganas (groups) can hold an important place in the treatment of diabetic foot ulcer. All the Ganas explained above are commonly indicated in Prameha, Kushtha and Dushta vrana.
TABLE NO. 1 The properties of these ganas are as follows: Name of Gana
Indications
Action
It purifies blood. It has good healing property. Prameha ch dushta It should be used vranashodhana… externally. But, can be used orally also. Asanadi Gana Shwitrakushthakapha…krimin… It is indicated orally. Prameha ch medo dosha But, in “prameha pidika (Tripathi,2007) nibarhana… chikitsa” it is used for (A.H.SU. 15/19-20) parisheka. Its action is on rasa Vatsakadi Gana kapha meda dhatu, and medo dhatu. (Tripathi,2007) Therefore it is indicated (A.H. SU. 15/33-34) orally. Anti-microbial. Surasadi Gana shleshma medakriminishudana… It can be used externally. (Tripathi,2007) vranashodhana (A.H. SU 15/30-31) Eladi Gana (Tripathi,2007) (A.H. SU 15/43-44) Aragwadhadi Gana (Tripathi,2007) (A.H. SU 15/17-18)
Vrana prasadana, Kandupidikakota nashana
Arkadi Gana krimikushta vranashodhana (Tripathi,2007) (A.H. SU 15/28-29)
visheshat Anti-microbial. It can be used externally.
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TABLE NO. 2 The properties of these ganas are as follows: Most of the herbs present in this Nyagrodhradi Gana vranya… gana are astringent in taste. (Tripathi,2007) Bhagna sadana… Therefore they are Rakta (A.H.SU 15/41-42) shodhaka (Blood purifier) Kledashoshaka (absorbs secretions) and „Vranaropaka‟ (wound healers) Herbs found in this gana are Padmakadi Gana brihmana… Vata-pittashamaka and (Tripathi,2007) Raktaprasadaka (Blood (A.H.SU 15/12) purifiers) The above (Table no. 1) Ganas (group of drugs) explained can be used in initial stages of Diabetic foot when secretions are present. For deeply penetrated Diabetic foot ulcers, Nyagrodhradi and Padmakadi Ganas (Table no. 2) are indicated. Healing of ulcer (Vrana Ropana), secretions have ceased and infections stage is over can be done with the use of these Ganas. These ganas will help in the regeneration of healthy tissue. Most of the herbs explained in
above Ganas are not available or controversial like Madhurasa, Kadar etc. Therefore, good result may hardly be achieved using one gana in treatment. Hence, to overcome the above drawback new method can be implemented. For that instead of using whole gana with the help of common herbs present in above explained ganas (Eladi, Aragwadhadi, Asanadi Gana, Vatsakadi Gana, Surasadi Gana, Arkadi, Nyugrodhradi Gana, Padmakadi Gana) may be used. (Table no. 3).
TABLE NO. 3 common herbs present in above explained ganas and their properties: Name / Latin name
Ayurvedic literature
1) Patha Cissampelos pareira Linn. (From Aragwadhadi, vatsakadi)
Bhagnasandhankrit.. (Vaidya Bapalal,2007) Kushthanu…(VaidyaBa palal, 2007) Kriminut(Chunekar,201 0) Sandhaniya (Sastri,2008) Vranaan..(Chunekar, 2010) Vranam hanta…(Vaidya Bapalal,2007) Krimim hanta…(Vaidya Bapalal,2007) Kushtam… (Vaidya Bapalal,2007) Kushta twak dosha vrana nashana….(Vaidya Bapalal,2007)
2) Karanja Pongamia glabra Vent. (From Aaragwadhadi, Arkadi)
3) Chirabilva Holoptelia integrifolia Planch. (From Aragwadhadi, Arakadi, Asandi)
Prayojyanga with active principle Moola (root) – pelosine or Berbeerine 0.5%
Current Researches with references Blood purifying (Khare) Anti-inflammatory, anti-microbial (N. Savithramma et al.,2011)
Moolatwak –(rootbark) Karajin/ Demethoxy karanjin (Sangwan et al., 2010) Puspa(flower) – Pongamin/ Quercitin
Anti-microbial, Anti-inflammatory, Anti-hyperglycemic, Anti lipid peroxidase, decreases in level of blood sugar. Increase in glucose 6phosphatase activity and enhancing anti-oxidant status Flowers are used in diabetes. (Khare) Seed powder dissolved in water showed hypoglycemic activity in alloxinised rabbits. (Khare,) Anti-microbial, anti-oxidant, antiinflammatory properties of leaves (Sharma et al., 2009)
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Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 118–125 4) Kramuka Areca catechu Linn. (From Surasadi, Asanadi) 5) Vidanga Embelia ribes Burm.f. (From Surasadi, Vatsakadi) 6) Murva Marsdenia tenacissima W.& A. (from Aragwadhadi, Vatsakadi) 7) Arjuna Terminalia arjuna (From Nyagrodhradi, asanadi)
kledamalapaha…(Sastri , 2005)
Catechin, Arecoline (0.17%), Arecaidine (Patil et al.,2009)
Krimighna,
Embeline (2-3%), Christembine Haq et al.,2002)
8) Palasa Butea frondosa Koen.ex Roxb. (Asanadi, Nyagrodhadi) 9) Indrajava Holarrhena antidysenterica Wall. (From Argwadhadi, Asanadi)
Kushthanut… (Shastri, 2005) Pramehanut. (Shastri, 2005)
10) Bharangi Clerodendron serratum Spreng. (From Arkadi, Surasadi, Vatsakadi) 11) Agaru Aquillaria agallocha Roxb. (From Asanadi, Eladi)
Vranakrimighni… (Vaidya Bapalal, 2007)
Moola twak (stem bark) – Phenolic glycoside, saponine (Kajaria et al., 2011)
Kushthanut… (Vaidya Bapalal, 2007)
Heartwood-Sesquiterpene alcohol (Bhuiyan et al., 2009)
12) Ela Elettaria cardamomum Maton. (From Vatsakadi, Eladi) 13) Rohisha trina Cyambopogon martini(Roxb) Wats. (From Eladi, Surasadi)
(K.
Mehanut (Chunekar,2010) Kushthapaha.(Chuneka r,2010) Medomehavranam hanti (Chunekar, 2010)
Twak (stem bark) – Arjuentine, Frideline Glycoside, β-cystocetrol (Chander Ramesh et al., 2004) Beeja (seed) – Palasonin Twak (stem bark) – Kinotannic acid (Borkar, 2010) seeds – Conessin, Kurchicine (Lather, 2008)
Stimulation of nervous system, anti-oxidant activity, anti-microbial activity, hypoglycemic activity. (Patil et al.,2009) Wound healing activity, antiDiabetic activity , seeds-Blood purifying (M. Chitra, 1980) API recommends bark in lipid disorders, Anti-hypoglycemic activity -glucosidase inhibitor (Bacchawat, 2011) -glucosidase inhibitor (Bacchawat 2011) Anti-inflammatory and immuno modulator. (Halder et al., 2009) Fruit: Hypoglycemic and Hypo lipidemic activity Anti-oxidant (Miriyala et al., 2008)
Seeds: Anti-Diabetic activity, reduces LDL, VLDL, Elevation of glucose-6-phosphatase activity, reduces blood sugar level help in stimulation of -cells of islets of langerhans. Antimicrobial, antioxidant, antiDiabetic (Shrivastava et al.,. 2007)
Oil – Cineol, terpineol, terpinene, Limonene, Sabinene (Atta et al., 2000)
Antioxidant
Oil - Geraniol (80-94%), (Ginger (Dubey et al.,2003)
Antiseptic, wound healer.
DISCUSSION Diabetic foot ulcer is a common complication in Diabetic patients which is prevailing and disturbing the individual‟s routine and certainly lowering quality of life.
oil)
To avoid complications like gangrene and amputation, there is a need to develop a new treatment protocol which is simple and cost effective.
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Diabetic foot ulcer is not directly explained in Samhitas (Classical lexicons). Its Samprapti lies between Prameha, Kushtha and Dushta Vrana. Common herbs explained in the treatment of the above mentioned diseases would be better to use for the treatment of Diabetic foot ulcer. These common herbs are Patha (Cissampelos pareira Linn.), Karanja (Pongamia glabra Vent.), Chirabilva (Holoptelia integrifolia Planch.), Kramuka (Araca catechu Linn.), Vidanga (Embelia ribes Burm.f.), Murva (Marsdenia tenacissima W.& A), Arjuna (Terminalia arjuna), Palasa (Butea frondosa Koen.ex Roxb.), Indrajava (Holarrhena antidysenterica Wall.), Bharangi (Clerodendron serratum Spreng), Agaru (Aquillaria agallocha Roxb.) and Rohisha Trina (Cyambopogon martini(Roxb) Wats). These herbs can be used as a single drug or in combination in the treatment of Diabetic foot ulcer. These can be used externally (Shrivastava et al., 2011) orally or both.
CONCLUSION Management of Diabetic foot should be multipronged attack like controlling blood sugar level, preventing infection and avoiding peripheral nerve tissue damage is crucial. Hence in this study, an attempt has been made with available herbal drugs have been proved since time memo rid thoroughly and classified symptomatically keeping complications of Diabetic foot ulcer in mind. Regarding the treatment of an ulcer, two steps in Ayurveda are very important which are the Shodhana and Ropana and they have similar concept with debridement, dressing and elevation of wound as mentioned in modern medicine. Common herbs explained in the treatment of Prameha, Kushtha and Dushta Vrana were reviewed because they have potent medicinal property, less or negligible adverse effect. This study or various aspects of diabetic foot ulcer with single drug regimen specifically on various intricacies of disease particularly could prove beneficial to mankind.
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V. S. (2010), Evaluation of antihelminthic activity of Butea monosperma, International Journal of Phytomedicine; 2 (1): 555–610.
Chander Ramesh, Singh Kavita, Khanna A K, Kaul S M, Puri Anju, Saxena Rashmi, Bhatia Gitika, Rizvi Farhan, A K. Rastogi (2004), Antidyslipidemic and antioxidant activities of different fractions of Terminalia arjuna stem bark,Indian Journal Of Clinical Biochemistry, 19(2): Page no. 141–148. Chunekar K C, (2010) Bhavaprakasha Nighantu, Chukhambha Bharati Academy Varanasi. Dubey Vinod Shanker, Ritu Bhalla and Rajesh Luthra (2003), Sucrose mobilization in relation to essential oil biogenesis during palmarosa (Cymbopogon martinii Roxb. Wats. var. motia) inflorescence development, J. Biosci; 28 (4): 479–487
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Global Journal of Research on Medicinal Plants & Indigenous Medicine || GJRMI ||
Global J Res. Med. Plants & Indigen. Med. | Volume 2, Issue 2 | February 2013 | 118â&#x20AC;&#x201C;125
Shastri Ambikadatta, (2005), Susruta Samhita, Chaukhambha Sanskrit Sansthana Page no. 202 Shrivastava Neeta, Tejas Patel (2007), Clerodendrum and Heathcare: An Overview,Medicinal and aromatic plant science and biotechnology Singh Anil Kumar, Shrivastava Prabhat Kumar, Shukla Vijay Kumar (2011). Evaluation
Source of Support: Nil
of Nimba taila and Manjishha churna in non healing ulcer. IRJP 2(5), 201â&#x20AC;&#x201C;210 Tripathi Bramhananda, (2007), Ashtang Hridayam, 1st Edn, Chaukhambha Sanskrit Pratishthan, Varanasi. Vaidya Bapalal, (2007) Nighantu Adarsha Purvardha, Chaukhamba Bharati Academy Varanasi.
Conflict of Interest: None Declared
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