Bioactive Metabolites of Rhizosphere Fungi Associated With Cymbopogon Citratus (DC.) Stapf
Bioactive Metabolites of Rhizosphere Fungi Associated With Cymbopogon Citratus (DC.) Stapf
Bioactive Metabolites of Rhizosphere Fungi Associated With Cymbopogon Citratus (DC.) Stapf
E-ISSN: 2278-4136
P-ISSN: 2349-8234 Bioactive metabolites of rhizosphere fungi associated
JPP 2017; 6(6): 2289-2293
Received: 14-09-2017 with Cymbopogon citratus (DC.) Stapf
Accepted: 16-10-2017
Mosma N Shaikh
Department of Botany,
Mosma N Shaikh and Digambar N Mokat
Savitribai Phule Pune
University, Pune, Maharashtra, Abstract
India Rhizosphere soil samples of commercially explored aromatic and medicinal plant,
Cymbopogon citratus (DC.) Stapf (Var. OD-19) were collected from Anand Agriculture University,
Digambar N Mokat Gujarat. A total of 62 fungal isolates representing 9 genera and 15 different species were isolated and
Department of Botany, identified. The most dominant fungal species were Aspergillus niger (20.96%), Aspergillus terreus
Savitribai Phule Pune
(12.90%), Aspergillus flavus (9.67%), Rhizopus oryzae (8.06%), Rhizopus nigricans (8.06%), Mycelia
University, Pune, Maharashtra,
India
sterila (8.06%), Aspergillus fumigatus and Trichoderma viride (6.64%) followed by Rhizopus stolonifer
(3.22%), Penicillium sps.(3.22%), Curvularia lunata (3.22%), Fusarium oxyporum (3.22%), Mucor
racemosus (3.22%), Curvularia inaequalis (1.61%) and Alternaria tenuis (0.62%). These fungal species
were cultured on Potato Dextrose broth and bioactive compounds were extracted using ethyl acetate and
identified by using GC-MS. Out of 62 fungal isolates Trichoderma viride was found to synthesize Citral,
in Potato Dextrose Broth supplemented with 2 g/L geranyl pyrophosphate (GPP) as an universal
precursor of monoterpenes. Besides Citral, the two other important compounds were isolated include 1,
2-Benzenediol and 2,6-Octadien-1-ol, 3,7-dimethyl-, (Z)-respectively. The results indicated that
rhizosphere fungi isolated from Cymbopogon citratus could be a potential source for isolation and
purification of bioactive compounds which may have potential pharmaceutical applications. This is the
first report of the rhizosphere fungi isolated from Cymbopogon citratus synthesizing Citral and other
bioactive compounds.
Introduction
Medicinal plants are a rich source of bioactive metabolites (Toussaint et al. 2007) [34], and are
considered to be safe environmentally compared to the synthetic chemicals for the treatment
number of human ailments (Nema et al. 2013) [18]. The cost of production, side effects and the
development of resistance by disease causing agents against synthetic chemicals, necessitated
the use of chemicals derived from medicinal plants for human and animal applications. In view
of these, there is a need for sustainable cultivation and continuous production of naturally
available resources for variety of applications. Cymbpogon citratus (Lemon grass) is an
important medicinal and aromatic plant due to its high medicinal value. The oil extracted from
C. citratus is used for skin care and cosmetic products, such as soaps, deodorants, shampoos,
lotions, and tonics (Lawless, 1995) [15]. Citral is the main active compound extracted from
lemon grass used in perfumery industries and has strong antimicrobial and pheromonal effects
in insects (Robacker & Hendry, 1977) [27].
The role of microbes in plant growth, nutrient availability, disease resistance, yield and quality
of medicinally important compounds is demonstrated in medicinal plants. Nowadays there are
increasing research in the interaction between medicinal plant and their rhizosphere microbes
for the improvement of medicinal plants. Hiltner (1904) [9] introduced the term rhizosphere for
the soil zone just adjacent to plant roots. Fungal abundance is 10–20 times higher in the
rhizosphere than in the bulk soil (Morgan et al. 2005) [16]. The rhizosphere microbes play an
important role in improving medicinal values of medicinal plants. The inoculation of
rhizosphere fungi is a sustainable technology to enhance the quantity and quality of the
medicinal plant compounds. Overall the study will help to explore the possibilities of
enhancing the active contents in the selected medicinal plant. This interest is also linked to
environmental concerns for reduced use of chemicals as well as an appreciation for utilization
Correspondence of biological and organics in agriculture.
Digambar N. Mokat Meager work has been done in area of screening of rhizosphere fungi and bioactive
Department of Botany, compounds isolated from these rhizosphere fungi associated with selected important medicinal
Savitribai Phule Pune plant.
University, Pune, Maharashtra,
India
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Indigofera aspalathoides, Eclipta prostrata and Indigofera major peak was confirmed as 2, 6-Octadien-1-ol, 3,7-
tinctoria. Five genera of AM fungi were identified in the dimethyl-, (Z)- which is a is a monoterpene used in
rhizosphere of 3 medicinal plant species viz., Mitragyna perfumery. Third major and important peak was found as
parvifolia, Withania coagulans and Leptadenia reticulate by Citral which is either a pair, or a mixture of terpenoids with
Panwar and Tarafdar, 2006 [20]. the molecular formula C10H16O. Citral is an aroma compound
There was an enormous variation in the AM fungi spore which is used in perfumery and flavoring. It has
population and root colonization in the rhizosphere of ten strong antimicrobial qualities, and pheromonal effects in
medicinal plant species viz., Aloe barbadensis, Emblica insects.
officinalis, Mimosa pudica, Rauvolfia tetraphylla, Centella In ethyl acetate extract of Monochaetia kansensis isolated
asiatica, Sapindus trifoliatus, Euphoria longan, Rauwolfia from Rhododendron sp. showed presence of 2- tetradecene
serpentina, Smilax sp. and Trachyspermum copticum, in spite (22.00%), cyclodecane (19.39%), phenol, 2, 4-bis (1, 1-
of their growth in similar climatic conditions (Hussain and dimethyl ethyl) (9.78%), E-15-Heptadecenal (8.70%) and 1-
Srinivas 2013) [10]. Khamna et al. (2009) [13] reported a total of octadene (12.29%) (Yogeswari, 2012) [37]. Rajalakshmi and
445 actinomycete isolates from 16 medicinal plant Mahesh (2014) [24] reported that ethyl acetate extract of
rhizosphere soils. The AM fungal community in the Aspergilus terreus isolated from the rhizosphere of medicinal
rhizosphere of Phellodendron amurense showed three general plant showed presence of Cyclooctasiloxane,
groups of Glomus, Hyponectria and Scutellospora Hexadecamethyl, Heptasiloxane, hexadecamethyl- 1,2-
respectively (Cai et al. 2009) [2]. Benzenedicarboxylic acid, butyl 2-methylpropyl ester, 9,12-
Octadecadienoic acid (Z,Z)-, 9,12-Octadecadienoic acid,
Linolsaeure,10,12-Hexadecadien-1-ol, Docosane (cas) n-
Docosane, Hexacosane (CAS) n-Hexacosane, Tetracontane
and n-Hexacosane. GCMS analysis of volatile metabolites in
the ethyl acetate extract of the Colletotrichum gloeosporioides
isolated from Phlogacanthus thyrsiflorus revealed the
presence of Phenol,2,4-bis (1,1-dimethylethyl), 1-
Hexadecene, 1-Hexadecanol, Hexadecanoic acid,
octadecanoic acid methyl ester and 1-Nonadecene (Devi and
Singh, 2013) [5]. The GC-MS results of ethyl acetate extract of
Fusarium proliferatum FP85 revealed different metabolites
such as 3-ethyl-2, 5-dimethyl-; Pyridine, 3-butyl-; Pyrazine,
Stearic acid; Palmitic acid; Adipic acid; Oleic acid;
Piperitenone oxide; butyl isobutyl ester and Phthalic acid,
(Rasekhi, 2014) [26]. The ethyl acetate extract of Aspergillus
Fig 1: Percentage of fungal isolates per species
terreus MP15 isolated from Swietenia macrophylla leaf
showed major compound, di-n-octyl phthalate with 80%
2. Gas Chromatography Mass Spectrometry (GC-MS)
matching factor (Yin et al., 2015) [36]. GC-MS analysis of
Analysis: GC-MS analysis was carried out to identify the
ethyl extract of Irpex lacteus isolated from Ocimum sanctum
possible compounds in the bioactive fraction of Aspergillus
showed presence of 8 compounds in the ethyl acetate extract
niger, Aspergillus terreus, Aspergillus flavus, Aspergillus
were (Azaphenanthrene, 2-methyl-3-(2-phenylethenyl)
fumigatus, Rhizopus oryzae, Fusarium oxyporum and
(47.67%), Cycloheptasiloxane, tetradecamethyl) (22.18%),
Trichoderma viride extracts.
(Linolelaidic acid, methyl ester) (22.35 %), Androst-5-en-17-
The GC-MS analysis showed the presence of 3-Methylbut-3-
carboxylicacid (20.14%), (68.64%) Androst-5-en-17-
en-2-ol, Toluene, Methane, sulfinylbis-, Benzeneethanol,
carboxylic acid. N-Trifluoroacetyl-histidine, methyl ester or
Benzene, 1,3-bis (1,1-dimethylethyl)-, 1-dodecanol, 1-
Methyl 3-(1H-imidazol5yl) 2-[(trifluoroacetyl) amino]
Dodecene in Aspergillus niger extract, 3-Methylbut-3-en-2-ol,
propanoate and Thiocarbamic acid, N,N-dimethyl, S-1,3-
Toluene, Benzeneethanol, 1-Dodecene, 1-dodecanol, (trans)-
diphenyl-2-butenyl ester, (Chaudhary and Tripathy, 2015) [3].
2-nonadecene in Rhizopus oryzae extract. In Aspergillus
terreus ethyl extract showed presence of 3-Methylbut-3-en-2-
ol, 1-Undecanol, 1-Dodecene, 1-dodecanol. Important
biologically active compounds namely Nerol and Citral with
3-Methylbut-3-en-2-ol, Toluene, Phenylethyl Alcohol, 1-
Dodecene, 1-dodecanol were found to be present in
Trichoderma viride ethyl extract. Mycelia sterilia showed
presence of 1, 2-Benzenediol, 2,6-Octadien-1-ol, 3,7-
dimethyl-, (Z)- while as Aspergillus flavus showed presence
of 3-Methylbut-3-en-2-ol, Toluene, 1-Dodecene, 1-dodecanol.
Fusarium oxyporum was found to be synthesized 3-
Methylbut-3-en-2-ol, Methane, sulfinylbis-, 1-dodecanol and Fig 2: GC-MS chromatogram of Tichoderma viride shows citral
1-Dodecene. Figure 2 shows the GC-MS chromatogram of peak after 21 days incubation in a PDB medium at 28 ºC.
ethyl acetate extracts of Trichoderma viride and Fig 3 shows
the MS spectrum of Citral isolated from Tichoderma viride
extract. In ethyl acetate extract of given fungus, a total of 3
major peaks were observed on the chromatogram. Among the
possible compounds identified, given major groups of
compounds detected were 1,2-Benzenediol, which is
an aromatic compound comes under class phenol. Second
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