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Phytochemical, Antioxidant, Antimicrobial and Micropropagation Study of Species

Nita Thapa1, Janardan Lamichhane 1 & Dhurva P. Gauchan 1 1Department of Biotechnology, Kathmandu University, Dhulikhel, Kavre, Nepal

Abstract: 1. Introduction being a versatile provides numerous benefits to living beings are perennial that belong to comprising the environment. It is used in grass family in the subfamily household works for construction, support; Bambusoideae. Woody bamboos consist of provides fodder for animals, edible shoots 9 to 10 sub tribes, containing between 59 for human; helps environment by and 111 genera and at least 1447 species sequestering carbon dioxide from the [1]. It includes some of the fastest growing atmosphere and converting the carbon into plants in the world due to its unique plant fiber and this benefit continues. rhizome dependent system [2]. It does not Besides, individual plant parts such as stem, require replanting after harvest as it will leaves possess medicinal properties which is regenerate from its rhizome root structure. analyzed and evaluated through various Besides, it improves soil quality and helps studies of phytochemical, antioxidant and to rebuild eroded soil. The word bamboo is antimicrobial activity. Many people are derived from the Kannada benefitted economically by selling this plant term ‘bambu’ which was found to be so demand for its production is escalating introduced through Indonesian and Malay to day by day. Micropropagation is thus English [3]. It is likely that more than 1250 considered as crucial as this ensures species under 75 genera of bamboo are production throughout the year irrespective unequally distributed in the humid tropical, of season and hindrance caused by subtropical and temperate regions of the flowering of this plant. Among many genus world [4]. It is popularly known as “poor of this plant, Bambusa is considered as one man’s timber” since it is commonly used by of the important one and has many species rural people of many countries [5]. It serves under this genus. This review is thus an as a great relief for rural people especially attempt to explore studies conducted by of Asian countries in economical as well as many scholars and researchers of this genus social terms. It is used as building material which is intended to provide light to some of for construction purposes and people make the exceptional works done which money by selling bamboo as a whole, eventually is beneficial to humankind. handicrafts and bamboo shoot for edible purposes and many more. Bamboo is also known as the “green gold of the forest” for Keywords of its global demand and versatile uses [6]. Bamboos are present with 75 genera Bambusa, Phytochemical, Antioxidant, growing in a wide range of different habitats Antimicrobial, Micropropagation worldwide so there is much variation in the characteristics and behavior they exhibit. includes 509 scientific plant

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names of species rank for the phytochemicals they inhibit. In this review genus Bambusa out of which 130 are paper, phytochemical assessment, accepted species names. The Plant antioxidant activity, micropropagation List includes a list of botanical names of abilities and antimicrobial activity of plant species which was launched in 2010 various species of Bambusa genus is being and created by the Royal Botanic Gardens, reviewed. Kew and the Missouri Botanical Garden. These accepted species are also included in 2. Phytochemical Screening World Checklist of Selected Plant Families Plants are the richest resource of drugs of (WSCP). This list further includes a further traditional systems of medicine, modern 97 scientific plant names of infraspecific medicines, nutraceuticals, food supplements, folk medicines, rank for the genus Bambusa which is not pharmaceutical intermediates and chemical included in WSCP. It is a genus of large entities for synthetic drugs [14]. Plants are clumping bamboos in the major group capable of synthesizing a wide variety of angiosperms (flowering plants). They are chemical compounds that perform important inherent to Southeast Asia, China, Taiwan, biological functions and these compounds Himalayas, New Guinea, Melanesia and are also used to defend against attack from Northern Territory of Australia. They are predators such as insects, fungi and herbivorous mammals. Photochemical can also found to be acclimatized in other be defined as the non-nutritive chemical regions such as Africa, Latin America and compounds which occur naturally in plants different oceanic islands [7, 8]. Some where ‘phyto’ means plant in Greek. members of the genus Bambusa species has Extraction and characterization of several been found to provide numerous benefits to active phytocompounds from plants have science and communities [9, 10, 11]. given rise to some high activity profile Among twenty priority species of bamboo drugs [15]. Many plant extracts and phytochemicals show antioxidant or free with economic importance which was listed radical scavenging properties [16]. These in ‘Priority Species of phytochemicals are divided into primary Bamboo and Rattan', compiled by the and secondary metabolites. Primary International Network for Bamboo and metabolites are essential to all plants such as Rattan (INBAR), there are seven species of sugars, fats and secondary metabolites; also Bambusa genus [12]. INBAR released a called phytochemicals are found in smaller range of plants and serve a specific purpose revised list that included 20 taxa (species since these have protective properties. and genera) of particular economic There has been conduction of many importance and a further 18 taxa of investigations regarding study of importance [13]. Species of Bambusa genus phytochemicals on various medicinal plants are gaining more popularity nowadays from long time ago. However, study of because of various properties it possesses these phytochemicals on bamboo leaves can whether be it on construction work or better be viewed as a novel practice and its interest is increasing more as each day passes. micropropagation capabilities. They also Bamboo leaves have therapeutic along with possess various antimicrobial capabilities nutritive properties so it is a good source of and antioxidant activity because of different fodder for livestock. Phytochemical

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investigation was performed on three acetone along with Fourier bamboo species B. vulgaris Schrad. ex J. C. Transform Infrared Spectroscopy (FTIR Wendl., B. ventricosa McClure and O. analysis). Aqueous solvent comparatively abyssinica (A. Rich.) by Coffie et.al., 2014 showed better presence of phytochemicals in order to check its safety for consumption while quinones were completely absent in in both the dry and wet ethanol extracted all the bamboo leaf extracts. Flavonoids and leaf samples [17]. All the leaves extract was coumarins were found to be present in seven found to contain saponin, general glycoside, extracts whereas glycosides in six extracts coumarin and cyanogenic glycoside and indicating their way for future use as extract of B. ventricosa and O. abyssinica antioxidant agents. The bamboo leaf had polyphenol and flavonoid as well. There contains 2% to 5% flavone and phenolic were no traces of alkaloid, carotenoid, compound that have the power to remove triterpenoid, steroid, anthraquinone and active free radicals, stop sub-nitrification anthracene glycoside in any of the species. and reduce blood fat [21]. The B. vulgaris was regarded as the safest phytochemical screening of leaf extracts of among these three species as it contained B. vulgaris revealed the presence of four classes of phytochemicals and phenolic compounds, flavonoids, contained relatively less saponin as these terpenoids, alkaloids, tannins, alkaloids can lyse red blood cells of larger herbivores when extracted on hexane, chloroform, and this is why plant extracts are not ethyl acetate and water solvents [22]. The directly injected into human beings [18]. plant extracts were good sources of different Also their study concluded that the classes of bioactive compounds. These phytochemicals present in leaves extract phytochemicals have therefore were not considerably affected by air drying demonstrated good potential for the of the leaves because of which these leaves expansion of modern chemotherapies could be fed to livestock either in the fresh against microbial infections as the crude state or in the dry state as hay. In the dry extracts considerably inhibited the growth season where there is low rainfall, lack of of fungi A. niger. fodder for livestock is still a great problem Not only leaves and stem, bamboo shoots of in developing countries. Bamboos can be a few species are considered equally better alternative for fodder as their leaves important since it has suitable health are carbohydrate rich when compared with benefits because of their rich contents in components of the leaves from several proteins, carbohydrates, vitamins, fibres, vegetables [19]. Besides, these are minerals and very low fat. Bamboo shoots nutritious, content of amino acid is high, has contain many minerals like potassium, less microbial susceptibility and possess calcium, manganese, zinc, chromium, long shelf-life of their meal. copper, iron and also the minor amounts of In the study performed by Joselin et.al, phosphorus and selenium [23]. which are 2014, phytochemical analysis was done on important for human body. Fresh bamboo the leaves extract of five different bamboo shoot also possesses several vitamins such species B. arundinacea (Retz) Willd, B. as Vitamin A, vitamin B1, vitamin B3, heterostachya (Munro) Holttum, B vitamin B6 [24]. Besides, bioactive ventricosa McClure, B. vulgaris Schrad. ex. compounds such as flavonoids, phenols and J. C. Wendl. and D strictus (Roxb.) Nees phenolic acids which are found to have [20]. Analysis was performed on six potential health benefits are also found in different solvents namely aqueous, bamboo shoot [25]. Its consumption as petroleum ether, chloroform, ethanol and healthy diet is increasing rapidly mainly in

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Southeast Asian and East Asian countries antioxidant capacity [34]. When plant [26, 27]. The most widely distributed derived phytochemicals, are consumed in a phytosterols, sitosterols are abundantly regular basis, Halliwell B, 1996 pointed found in fermented bamboo shoots [28]. that it may drift the balance toward an Fermented bamboo shoots have been eaten ample antioxidant status [35]. The study of by various tribes since ancient times [29, phytochemicals and potential antioxidant 30]. Likewise, dish prepared with fermented activity of plant extract is thus important bamboo shoots is taken as local delicacy in in medical world as well. Flavonoids are many ethnic communities. Phytochemical diverse group of phytonutrients found in screening of fermented B. balcooa shoot nearly all fruits and vegetables, grains, bark, revealed the presence of tannins, steroids, roots, stems, flowers, tea and wine with phenols, glycosides, flavanoids, more than 6,000 types. These are carbohydrates and proteins when done in responsible for providing all those vibrant methanol and ethyl acetate solvents [31]. colors in fruits and vegetables along with More phytochemical constituents were carotenoids. These have the capacity to shown by methanol solvent in comparision reduce free radical formation and scavenge to ethyl acetate. In study conducted by them so interest is growing on study of Srivastava, 1990, the concentration of total relationship of flavonoids and antioxidant phytosterols in dried samples of fresh shoots activity particularly for the reason of health of B. tulda and D. giganteus was 0.21- benefits. Besides, the antioxidant capacities 0.39% and 1.6-2.8% in samples of of many flavonoids are much found stronger fermented shoots which was determined than those of vitamins C and E [36]. Many colorimetrically [32]. Anaerobic digestion groups of flavonoids have also been used in was the cause for increase in level of treatment of human diseases [37]. Both the phytosterols in fermented shoots as it antioxidant and prooxidant effects of degrades the organic matter. Therefore, flavonoids are discussed by Procházková et. fermented bamboo shoots could be used as a al. 2011 in their review paper [38]. starting material for the production of Flavonoids cannot only be purely accounted steroidal drugs which can reduce for antioxidants as meanwhile under certain inflammation and pain. reaction conditions they can also display prooxidant activity promoting the oxidation of other compounds. Nonetheless, 3. Antioxidant Activity prooxidant or antioxidant properties of a An antioxidant is broadly defined as any particular flavonoid depend most of all on substance that delays or inhibits oxidative its concentration. Flavonoids have also been damage to a target molecule by trapping stated to as ‘nature’s biological response free radicals [33]. There is increment in the modifiers as they possess inherent ability to interest and study of antioxidant activity of alter the body’s reaction to allergies, viruses various bamboo leaves among scholars and and carcinogens [39]. researchers worldwide although Study of antioxidant activity of fifteen antioxidative compounds in bamboo leaves different bamboo species was performed are not fully known. Many age related by Wang et.al, 2007 among which eight human diseases are also the result of cellular species were of Bambusa [40]. Their damage by free radicals and antioxidants results revealed that the methanolic extract could play a crucial role in preventing such of B. textilis McClure possessed the diseases. The medicinal effects of bamboo highest antioxidant activity among the leaves are mostly attributed by their selected fifteen bamboo species. It’s

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extract was then further analysed by phenolic compounds and total flavonoids HPLC-UV and HPLC-micro-fractionation indicated that these compounds contributed to identify antioxidant compounds, where to the antioxidative activity. Phytochemical three antioxidant fractions namely especially flavonoid is responsible for isoorientin 4''-O-β-D-xylopyranoside, antioxidant activity and many species of isoorientin 2''-O-α-L-rhamnoside and Bambusa exhibit this property. Further isoorientin were isolated. Evaluation of research is still necessary for more total polyphenol and antioxidant activity of qualitative and quantitative analysis of these leaves extract of B. nutans and B. vulgaris compounds to derive potential use of was carried out by Tripathi et.al, 2015 bamboo leaves in medicinal world. where total phenolic content of the extracts was calculated spectrophotometrically 4. Antimicrobial Activity using a modified Folin-Ciocalteu method In today’s world, the discovery and and DPPH (2,2-diphenyl-1-picrylhydrazyl) development of antibiotics is considered as radical scavenging assay was used to one of the most successful and powerful assess antioxidant activity [41]. Their achievements of modern science and study revealed that B. nutans can be technology which is intended to control beneficial and has higher polyphenolic infectious diseases. There is increase in content and antioxidant activity than B. resistance of pathogens to traditional vulgaris. Therefore, it can be speculated antimicrobial agents. This resistance has that B. nutans can also be potentially indicated in need for isolation of microbial useful as a natural source of antioxidant or agents which is less susceptible to regular in medicine. The higher antioxidant and antibiotics; also allowing the recovery of superoxide scavenging activity in the resistant isolates during antibacterial chlorogenic acid (CGA) methyl ether therapy. Although there is much compounds than in other phenolic acids advancement in medical world, there is still and antioxidants is mediated through o- a critical need to control antimicrobial dihydroxyl and methoxyl properties of the resistance by upgraded antibiotic usage in molecules as well as inhibition of xanthine order to reduce hospital cross-infection [44]. oxidase [42]. In their study, three CGA The medicinal plants containing compounds derivatives from the leaves of bamboo P. with antimicrobial activity are found all edulis was isolated and identified as 3- O- around the world and many efforts have (3′-methylcaffeoyl)quinic acid, 5- O-caf- been made to discover novel effective feoyl-4-methoxylquinic acid and 3- O- antimicrobial compounds. Antimicrobial caffeoyl-1- methoxylquinic acid. CGA is screening is done to specify the presence of an ester of caffeic acid with quinic acid antimicrobial molecule in the plant against found naturally in various plants such as pathogen and microbes. coffee beans, apples and blueberries. Micro dilution assay showed that ethanolic DPPH radical scavenging activity, total extract of D. asper leaves extract was phenols and antioxidant activities was effective to inhibit all tested E. coli strains assessed and analyzed by Goyal et.al, 2010 [45]. The chemical identification using in Indian wild B. vulgaris "Vittata" pyrolysis GC/MS revealed that fatty acids, methanolic leaf extract [43]. There was together with esters, long chain alcohols and found presence of carbohydrates, reducing aldehydes were the major compounds in sugars, flavonoids, steroids, saponins, ethanolic extract. Also, antidiarrheal agent alkaloids, tannins, anthraquinones and could be extracted from D. asper leaves glycosides. The high contents of total extract. In similar experiment, antibacterial Available online: https://pen2print.org/index.php/ijr/ P a g e | 81

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activity of B. arundinaceae was tested Besides, additional work should also be against the strains of S. aureus, carried out in order to fractionate the P.aeruginosa , E. coli and Bacillus [46]. extracts to draw a better understanding of Ethanolic extract was moderately effective mechanism of those antioxidants against all four organisms, while aqueous responsible for antimicrobial activity. extract showed moderate effect against E. coli and S. aureus. Bamboo is considered as 5. Micropropagation a good source of phytosterols that are the It is a rapid multiplication of a selected precursors of various pharmaceutically plant by use of vitro culture techniques. active steroids found in plants and these also This technique utilizes modern plant tissue act as nutraceuticals. In the study, culture methods. Explants upon subsequent conducted by Wasnik and Tumane, 2014, subcultures result in a number of multiple they found that the ethanolic and methanolic shoots and these multiple shoots on leaves extract of B. bambose L. showed elongation allow rooting in vitro. After excellent inhibitory action against all tested rooting, they are in vitro hardened and ten multiple drug resistant bacteria and they transferred to field [49]. Traditionally, all were clinical isolates from wound bamboos were propagated with seeds, infection [47]. Here also ethanol solvent clump division, culm and rhizome cuttings comparatively showed better results like but due to gregarious flowering, low seed previous studies. Effectiveness of plant viability, high costs, problems faced during extract against microbial strains was long-distance transportation of vegetative determined by measuring zone of inhibition propagules and poor efficiency of plant (ZOI) in all the studies. Ethanolic extracts production alternative propagation methods of B. arundinacea and M. indica had higher were introduced [50]. Micropropagation of inhibition on both gram positive as well as bamboo is the only viable method [51] and gram negative bacteria indicating both are it has has allowed in developing new type of potential source of natural antioxidants, ornamental or commercial bamboo. The phytochemicals and antibacterials. [48]. M. plantlet following this method can be indica showed more therapeutic capacity. In produced in any time of the year without this study also, ethanolic and methanolic depletion in its quality. This method is extracts of both the tested plants were more regarded as the best available technique and effective than aqueous extracts. will serve as standard for mass scale Antimicrobial screening of the n-hexane, propagation of bamboo in the future [52]. chloroform and ethyl acetate extracts of B. For tissue culture of bamboo, the use of vulgaris showed that the crude extracts starting material i.e. seeds or adult plants inhibited the growth of fungi A. niger. This including the choice of the propagation supports the ethno medicinal use of aqueous method are crucial [53]. In comparision to extracts of this species to treat sexually other medicinal plants, tissue culture in transmitted diseases and for wounds [22]. bamboo was practiced lately. The aseptic However, although bamboo leaves possess germination of bamboo seeds in D. strictus antibacterial activity there is not much was stated by Alexander and Rao, 1968 [54] studies and research conducted in order to and this is reported to have indicated the find its efficacy and usefulness till date. start of tissue culture of bamboo in White, Further investigations are still needed to 1964 major and minor elements [55]. identify their vital components needed for Following this, this medium has been then their effective role as therapeutic plant for used by Nadguada et al, 1990 in B. those already studied extracts as well. arundinacea and D. strictus; Mascarenhas

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et. al, 1988 in D. strictus; Ravikumar et. al., Shirin and Rana for B. glaucescens, Islam 1998 in D. strictus for zygotic embryo and Rahman and Yasodha et. al for B. germination [56-58]. After use of this nutans, Mishra et. al for B. tulda, Ndiaye medium, Murashige and Skoog ,1962 (MS) et.al for B. vulgaris [68-76]. Likewise, basal medium which is more popular these axillary shoots was used as explant by for B. days was regularly used in embryo atra and axillary bud for B. vulgaris ‘striata’ germination of B. arundinacea where half [77-78]. strength of MS basal medium was used, in Nodal explant is considered as the best B. nutans and D. membranaceus and in B. starting material for micropropagation tulda. MS medium is fortified with where multiple shoots are preferred or cytokinins, auxins, glucose source, gelling required as this is capable of providing agent and carbon source according to the such. An efficient micropropagation type of explant cultured as well as plant protocol for multiplication of B. species involved [59-62]. Contamination balcooa from nodal explants was and browning of explant is one of the main established where 88% success rate of reasons of failure of tissue culture acclimatization was achieved when done in experiment. The severity of browning has liquid MS media [79]. In the same species, varied according to species, tissue or organ, Patel et.al achieved survival rate of 74.66% and nutrient medium and other tissue when nodal segments were cultured on variables [63]. However, many techniques both solid and liquid MS media, Sharma have already been evolved and studied to and Sarma, achieved 100% success in field overcome these problems. Besides above transfer, 90% success rate achieved by mentioned, there are many other researchers Mudoi and Borthakur whereas Ansari et. al who have accomplished successfully in achieved 90% success rate when culture propagating various bamboo species from was initiated in solid MS media [80-83] . zygotic embryo. Some of these are in B. In similar experiment, Raju and Roy arundinacea in Nitsch and Nitsch medium, established an efficient and reproducible 1967; in MS medium; in B. vulgaris in MS micropropagation protocol for mass medium [64-67]. Various plant parts such as production and generation of uniform clones nodal segments, axillary buds were then for B. bambos (L.) Voss where explant used used as explants as this ensured large scale was nodal segments containing unsprouted propagation of bamboo. Micropropagation axillary buds in liquid MS media [84]. They through axillary bud proliferation by nodal recorded 100% survivability from well- explants did not interfere with the established rooted plantlets during production of true to type plant. This acclimatization process. method also does not interrupt the callus B. tulda is another species which phase and thus maintains the clonal fidelity researchers show great interest to mass [51]. This purpose of micropropagation was propagate as its main purpose is on well satisfied by MS media so almost all construction works and has high demand. researchers relied on using this medium as An efficient and rapid method of in vitro this media supplemented all the essential propagation of this species from aseptic nutrients required for development of new seedlings was studied and achieved 90% plantlet invitro. Nodal explants were used success rate upon field transfer when shoots by Arya et. al for B. arundinacea and B. from 3-week-old seedlings in liquid medium vulgaris, Das and Pal and Islam and were used to initiate the cultures [85] Rahman, for B. balcooa, Arya and Sharma whereas Mishra et.al had 98% rooting in for B. bamboos, Lin et.al for B. edulis, liquid MS media [86]. In same species,

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Waikhom and Louis had 81.81% success [2] llustrated Oxford Dictionary. Great Britain: when culture was initiated on liquid MS Oxford University Press. 1998. media [87]. They take the privilege of p. 69. ISBN 140532029-X. [3] Farrelly, David (1984). The Book of Bamboo. successfully developing protocol which Sierra Club Books. ISBN 0-87156-825-X. could produce rhizome in half-strength MS [4] Rai, P. K. (2009). Comparative assessment of medium for the first time of this soil properties after bamboo flowering and death species. Survival rate was 100% when in a tropical forest of Indo-Burma hot semisolid MS medium was used for culture spot. AMBIO: A Journal of the Human of nodal explant [88]. B. vulgaris Schrad. ex Environment, 38(2), 118-120. [5] Keshari, V. P. (2005). Bamboo: From poor Wendl has been promoted in order to solve man’s timber to green gold. Hamro Kalpana the deforested environments and economic Brikshya, 164, 10-4. problems [89]. During their study they [6] Sharma, Y. M. L. (1980). Bamboos in the Asia attained 75% survival where their explant Pacific Region. In Bamboo research in Asia: was inter-nodal region of stem and culture proceedings of a workshop held in Singapore, medium was solid MS media. B. nutans is 28-30 May 1980. IDRC, Ottawa, ON, CA. [7] Chapman, G. P. (1997). The bamboos. edible ornamental bamboo and is also In Linnean Society Symposium Series (No. 19). propagated in vitro by many researchers for [8] Clayton, W. D., & Renvoize, S. A. (1986). both study and commercial purpose. 95% Genera graminum. Grasses of the world. Genera survival rate was obtained by Mudoi et. al graminum. Grasses of the World., 13. whereas 100% success rate was observed by [9] Tripathi, Y. C. (1998). Food and nutrition Sharma and Sarma on field transfer when potential of bamboo. MFP News, 8(1), 10-11. nodal explants were used in MS medium [10] Bao, J. (2006). The nutrition and bioactive function of bamboo shoots. Food Nut in [90,91]. Research on other species of this China, 4, 2-3. genus is also being carried out which can [11] Chongtham, N., Bisht, M. S., & Haorongbam, S. ultimately contribute in mass production of (2011). Nutritional properties of bamboo shoots: bamboo. potential and prospects for utilization as a health food. Comprehensive Reviews in Food Science 6. Conclusion and Food Safety, 10(3), 153-168. [12] Benton, A. (2015). Priority species of bamboo. There have been many developments in In Bamboo (pp. 31-41). Springer, Cham. research techniques and equipment which [13] Rao, A. N., Ramanatha Rao, V., & Williams, J. has allowed for the assessment and T. (1998). Priority species of bamboo and rattan. evaluation of many constituents of plant [14] Hammer, K. A., Carson, C. F., & Riley, T. V. (1999). Antimicrobial activity of essential oils molecule and same applies for bamboo and other plant extracts. Journal of applied plant also. It is undoubtedly a versatile plant microbiology, 86(6), 985-990. and can serve to wellbeing of living beings [15] Mandal, V., Mohan, Y., & Hemalatha, S. (2007). in various ways. There has been conduction Microwave assisted extraction—an innovative of lot of works to find the beneficial active and promising extraction tool for medicinal plant constituents of some species, but still there research. Pharmacognosy reviews, 1(1), 7-18. [16] Larson, R. A. (1988). The antioxidants of higher is a long way to go to assess the true plants. Phytochemistry, 27(4), 969-978. potential of all the species of this plant. [17] Parekh, J., & Chanda, S. (2007). In vitro antimicrobial activity and phytochemical 7. References analysis of some Indian medicinal plants. Turkish Journal of Biology, 31(1), 53-58. [1] Ohrnberger, D. (1999). The bamboos of the [18] Coffie, G. Y., Antwi-Boasiako, C., & Darkwa, world: annotated nomenclature and literature of N. A. (2014). Phytochemical constituents of the the species and the higher and lower taxa. leaves of three bamboo (Poaceae) species in Elsevier. Ghana. Journal of Pharmacognosy and Phytochemistry, 2(6).

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