CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)

An Online International Journal Available at http://www.cibtech.org/cjm.htm

2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article
PRELIMINARY PHYTOCHEMICAL ANALYSIS AND IN VITRO
INVESTIGATION OF ANTIMICROBIAL ACTIVITY OF EICHHORNIA
CRASSIPES (MART.) SOLMS. AGAINST POULTRY PATHOGENS
*Kayathri B., Kanimozhi K. and Panneerselvam A.
Department of Botany and Microbiology, A.V.V.M Sri Pushpam College (Auto.), Poondi. Thanjavur (Dt.),
Tamil Nadu, India
*Author for Correspondence

ABSTRACT
Eichhornia crassipes (Mart.) Solms (Waterhyacinth) is an aquatic macrophyte, monocotyledon of the
family pontederiaceae. It is listed as one of the most productive plant on earth and is considered the
world’s worst aquatic weed. The preliminary phytochemical analysis of leaf extracts of Eichhornia
crassipes (Mart.) Solms. reveals the presence of alkaloids, amino acids, carbohydrate, fats, flavonoids,
glycosides, phenols, protein, saponins, sterols, tannins and terpenoids. The antimicrobial efficacy of
acetone, distilled water, ethanol, ethyl acetate, methanol and n-Butyl alcohol extracts of E. crassipes were
analyzed by well agar method. The leaf extracts of E. crassipes (Mart.) Solms., standard antibiotics
streptomycin and fluconazole exhibited variable degrees of antimicrobial activity. Among the six extracts
tested, n-Butyl alcohol extract showed significant antimicrobial activity against the poultry pathogens.

Keywords: Eichhornia Crassipes, Leaf Extracts, Antimicrobial Activity, Poultry Pathogens

INTRODUCTION
The world is rich with natural and unique medicinal plants. Medicinal plants are now getting more
attention than ever because they have potential of myriad benefits to society or indeed to all mankind,
especially in the line of medicinal and pharmacological aspect. Traditional use of the medicinal plants
have fewer side effect over allopathic medicine such promising fact leads the development of herbal
derive medicines whole over the world (Pal and Shukla, 2003). Plants contains flavonoids, alkaloids,
Tannins, phenols etc., which have biological significance in terms of medicine development and extracts
of aqueous, methanol and ethanol are good source of antiviral, antitumor and antibacterial agents (Cowan,
1999).
Eichhornia crassipes (Mart.) Solms. commonly known as Water hyacinth is a warm water aquatic plant
belonging to the family Pontideriaceae. It is native to Brazil. Plants are thought to have been first
introduced into the United States at the 1884 Cotton States Exposition in New Orleans, LA.
Waterhyacinth is a floating, flowering, perennial weed, form dense rafts in the water and mud (Mane et
al., 2011). It can quickly grow to very high densities (over 60kg m-2); thereby completely clogging water
bodies, which in turn may have negative effects on the environment, human health and economic
development (Jayanthi et al., 2011).
It is listed as one of the most productive plant on earth and is considered the world worst aquatic weed
(Grodowitz, 1998).
The ‘beautiful blue devil’, water hyacinth is recognised by its lavender flowers and shining bright leaves
which spread at an alarming rate. Its habitat ranges from tropical desert to subtropical or warm temperate
rain forest zones, and tolerates a temperature range of 21.1 to 27.2ºC (Lata and Venapani, 2010). It can be
used as compost, paper, fuel, and animal feed and water purification (Kristie, 2012). It is also an excellent
source of biomass; and use to make furniture, hand bags and ropes in East Africa. Its flowers are used as a
medication on skin of horses and a tonic (Wikipedia, 2012).
Waterhyacinth possesses phytochemicals (Lata and Dubey, 2010; Jayanthi et al., 2011; Jayanthi and
Lalitha, 2011) which are of medicinal importance (Sayeed Ahmad, 2010). The methanol extract of leaves
of this plant aids in wound healing process (Ali et al., 2010) and has tumour inhibition potential (Ali et
al., 2009). In addition, the extracts of this plant showed antimicrobial activity (Shanab et al., 2010).
© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 19
 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article
The uses of plant-derived products as disease control agents have been studied, since they tend to have
low mammalian toxicity, less environmental effects and wide public acceptance. Hence the present
investigation reports the phytochemical and antimicrobial activity of Eichhornia crassipes against poultry
pathogens.

MATERIALS AND METHODS

Collection of Plant Material
Fresh leaves of E. crassipes (Mart.) Solms. free from disease were collected at local pond in
Nagapattinam District (Figure 1). The leaves were washed thoroughly 2-3 times with running water and
once with sterile distilled water, leaf material was then air-dried on sterile blotter under shade.
Solvent Extraction
Acetone, ethanol, ethyl acetate, methanol and n-Butyl alcohol extracts of E. crassipes (Mart.) Solms. were
prepared according to the methodology of Indian Pharmacopoeia (Anonymous, 1966). The aqueous
extraction achieved through the percolation method. These extracts were concentrated to dryness in flash
evaporator under reduced pressure and controlled temperature (40-50ºC). The extracts were put in air
tight containers stored in a refrigerator.
Preliminary Phytochemical Analysis
Preliminary phytochemical screening of the leaf extracts of E. crassipes (Mart.) Solms. was carried out as
per standard procedure (Kokate et al., 2005; Harborne, 2005).
Screening of Antimicrobial Activity
The antimicrobial activity of E. crassipes (Mart.) Solms. was evaluated by well agar method (Perez et al.,
1990).
Selection of Microorganisms
Totally ten chicken pathogenic microorganisms namely five bacterial strains such as, Escherichia coli,
Proteus vulgaris, Salmonella pullorum, Staphylococcus aureus and Streptococcus pyogenes and five
fungal strains such as, Aspergillus flavus, A. fumigatus, A. ochraceus, Candida albicans and Trichophyton
megnini were selected for the present investigation. The chicken pathogenic bacteria and fungi were
originally obtained from Microbial Germ Plasm Culture Collection Unit (MGPCCU), Sri Gowri Biotech
Research Academy, Thanjavur and used for the present investigation.
Antimicrobial Activity
The antimicrobial activity of ethanol, acetone, ethyl acetate, methanol, n-Butyl alcohol and distilled water
extracts of E. crassipes (Mart.) Solms. were tested against the selected bacterial and fungal strains. The
sterilized nutrient agar and potato dextrose agar medium was poured into each sterile petriplates and
allowed to solidify. By using a sterile cotton swabs, a fresh bacterial and fungal cultures were spread over
the plates by following spread plate technique. One well of 5mm size made into the agar plates with the
help of sterile cork borer, the wells were loaded with 200µl of ethanol, acetone, ethyl acetate, methanol,
n-Butyl alcohol and distilled water extracts of E. crassipes were loaded in to separated wells. The plates
were incubated for 24 hours at 37oC for bacteria and 28 oC for 48 - 72 hours for fungi. After the incubation
period, the results were observed and the diameter of the inhibition zone was measured around the wells.
The antibiotic sensitivity test was investigated using standard antibiotics Streptomycin (10 µg/ disc) for
bacteria and Fluconazole (10 µg/ disc) for fungi.

RESULTS AND DISCUSSION

Phytochemicals are responsible for medicinal activity of plants (Savithramma et al., 2011). In the present
study, the preliminary phytochemical screening of the leaf extracts of E. crassipes (Mart.) Solms.
revealed the presence of alkaloids, amino acids, carbohydrate, fats, flavonoids, glycosides, phenols,
protein, saponins, sterols, tannins and terpenoids (Table 1). These results of phytochemical screening of
study plant were in concurrence with other reports (Thamaraiselvi et al., 2012; Jayanthi and Lalitha,
2013; Aravind et al., 2013). There were a similarity between their results and the obtained results in this
study.

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 20

 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article
In the present study, acetone, distilled water, ethanol, ethyl acetate, methanol and n-Butyl alcohol extracts
of E. crassipes (Mart.) Solms. and standard antibiotics streptomycin and fluconazole exhibited variable
degrees of antimicrobial activity. The results were given in Table 2, 3, 4 & 5 and Plate 1, 2, 3 & 4. The
inhibition activity of the plant extracts were compared with standard antibiotics streptomycin and
fluconazole. The maximum antibacterial activity was exhibited by n-Butyl alcohol extract against
Streptococcus pyogenes. The acetone extract showed maximum zone of inhibition against Aspergillus
ochraceus. These results of antimicrobial activity of different extracts of E. crassipes (Mart.) Solms. were
in coincidence with results of Mahavir and Sandeep (2013) who reported that, the leaves of E. crassipes
exhibited significant antibacterial activity. Similar works were done by Thamaraiselvi et al., (2012) and
Jayanthi and Lalitha (2013) who demonstrated that leaf extracts of E. crassipes exhibited considerable
antibacterial and antifungal activity.
Waterhyacinth which is often considered a weed has in recent days been taken up for many researches
concerning the phytochemical and pharmaceutical applications. From the present study it can be
concluded that the leaf extracts of Waterhyacinth showed significant antimicrobial activity against all the
microorganisms tested.

Table 1: Qualitative phytochemical analysis of leaf extracts of Eichhornia crassipes (Mart.) Solms
S. Phytocompounds Acetone Distilled Ethanol Ethyl Methanol n-Butyl
No water acetate alcohol
1. Alkaloids + + - - + +
2. Amino acids - - + + - -
3. Carbohydrates - + + + - -
4. Fat - - - - - +
5. Flavonoids - - - + + +
6. Glycosides - + - - + -
7. Phenols + - + - - +
8. Protein - + + - - +
9. Saponins - + - + + -
10. Sterols + - + + + -
11. Tannins + - - - + -
12. Terpenoids + + + - + +
+: Presence - : Absence

Table 2: Antibacterial activity of Eichhornia crassipes (Mart.) Solms

S.No Bacterial Zone of inhibition (diameter in mm)
pathogens Acetone Distilled Ethanol Ethyl Methanol n-Butyl alcohol
water acetate
1. Escherichia 10 8 9 - 9 10
coli
2. Proteus 11 9 10 - - -
vulgaris
3. Salmonella 8 - - - 8 10
pullorum
4. Staphylococcus - - 10 - 10 15
aureus
5. Streptococcus 9 - 9 10 8 20
pyogenes

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 21

 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article
Table 3: Antifungal activity of Eichhornia crassipes (Mart.) Solms
S.No Fungal Zone of inhibition (diameter in mm)
pathogens Acetone Distilled Ethanol Ethyl Methanol n-Butyl alcohol
water acetate
1. Aspergillus 10 9 10 - - -
flavus
2. A. fumigates 14 10 13 8 - 10
3. A. ochraceus 15 9 12 - - 10
4. Candida 10 9 8 - - 11
albicans
5. Trichophyton 10 9 8 9 - 10
megnini

Table 4: Effect of standard antibiotics on bacterial pathogens

S.No Bacterial pathogens Zone of inhibition (diameter in mm)
Streptomycin
1. Escherichia coli 12
2. Proteus vulgaris 10
3. Salmonella pullorum 13
4. Staphylococcus aureus 9
5. Streptococcus pyogenes 19

Table 5: Effect of standard antibiotics on fungal pathogens

S.No Bacterial pathogens Zone of inhibition (diameter in mm)
Fluconazole
1. Aspergillus flavus 10
2. A. fumigatus 19
3. A. ochraceus 18
4. Candida albicans 20
5. Trichophyton megnini 9

Figure 1: Leaves of Eichhornia crassipes (Mart.) Solms

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 22
 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article

Plate 1: Antibacterial activity of Eichhornia crassipes (Mart.) Solms

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 23
 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article

Plate 2: Antibacterial activity of Eichhornia crassipes (Mart.) Solms

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 24

 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article

Plate 3: Antifungal activity of Eichhornia crassipes (Mart.) Solms

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 25
 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article

Plate 4: Antifungal activity of Eichhornia crassipes (Mart.) Solms

REFERENCES
Ali H, Lata N, Ahi J and Ganesh N (2010). Evaluation of wound-healing activity of Eichornia
crassipes: A novel approach. Drug Invention Today 2 212-214.
Ali H, Patel M, Ganesh N and Ahi J (2009). The world’s worst aquatic plant as a safe cancer medicine
“Antitumor activity on melanoma induced mouse by Eichhornia crassipes: in vivo studies. Journal of
Pharmacy Research 2 1365-1366.

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 26

 CIBTech Journal of Microbiology ISSN: 2319-3867 (Online)
An Online International Journal Available at http://www.cibtech.org/cjm.htm
2015 Vol. 4 (1) January-March, pp.19-27/Kayathri et al.
Research Article
Anonymous (1966). Pharmacopoeia of India (Ministry of Health Govt. of India Publication) New Delhi.
Aravind R Kurup, Divya Rajan, Jency Blesson, Sruthy Chandran, Thampatty AR and Veena PV
(2013). Detailed analysis on phytochemicals, antioxidants, antimicrobial activity of Eichhornia crassipes.
International Journal of Scientific Research 2(2) 17-19.
Cowan MM (1999). Plant products as antimicrobial agents. Clinical Microbiology Reviews 12(4) 564–
582.
Grodowitz MJ (1998). An active approach to the use of insect biological control for the management of
non native aquatic plants. Journal of Aquatic Plant Management 36 5761-5763.
Harborne JB (2005). Phytochemical Methods a Guide to Modern Techniques of Plant Analysis, edition 2
(Chapman & Hall) London.
Jayanthi P and Lalitha P (2013). Antimicrobial activity of solvent extracts of Eichhornia crassipes
(Mart.) Solms. Der Pharma Chemica 5(3) 135-140.
Jayanthi P and Lalitha P (2011). Determination of the in vitro reducing power of the aqueous extract of
Eichhornia crassipes (Mart.) Solms. Journal of Pharmacy Research 4 4003-4005.
Jayanthi P, Lalitha P and Shubashini KS (2011). Phytochemical investigation of the solvents extracts
and fractionates of Eichhornia crassipes. Journal of Pharmacy Research 4 1405-1406.
Kokate CK, Purohit AP and Gokhale SB (2005). Pharmacognosy, 39th edition (Nirali Prakashan) Pune
607-611.
Kristie T (2012). Plants, Garden Plants by name Water hyacinth. Available: http://www.ehow.com/facts.
Lata N and Dubey V (2010). Preliminary phytochemical screening of Eichhornia crassipes: The world’s
worst aquatic weed. Journal of Pharmacy Research 6 1240-1242.
Lata N and Venapani D (2010). Preliminary phytochemical screening of Eichhornia crassipes: The
world’s worst aquatic weed. Journal of Pharmacy Research 3(6) 1240-1242.
Mahavir Joshi and Sandeep Kaur (2013). In vitro evaluation of antimicrobial activity and
phytochemical analysis of Calotropis procera, Eichhornia crassipes and Datura innoxia leaves. Asian
Journal of Pharmaceutical and Clinical Research 6(5) 25-28.
Mane CP, Arjun BB and Pandurang AK (2011). Archieves of Applied Science Research 3(1) 222-229.
Pal SK and Shukla Y (2003). Herbal Medicine: Current Status and the Future. Asian Pacific Journal of
Cancer Prevention 4(4) 281-8.
Perez C, Pauli M and Bazerque P (1990). An antibiotic assay by the well agar method. Acta Biol Med
Experim 15 113–115.
Sayeed Ahmad (2010). Introduction of plant constituents and their tests, personal communication,
Available: http://nsdl.niscair.res.in/bitstream/123456789/708/1.pdf.
Shanab SMM, Shalaby EA, Lightfoot DA and El-Shemy HA (2010). Allelopathic Effects of Water
Hyacinth [Eichhornia crassipes]. Plus One 5 1-8.
Thamaraiselvi P, Lalitha and Jayanthi P (2012). Preliminary studies on phytochemicals and
antimicrobial activity of solvent extracts of Eichhornia crassipes (Mart.) Solms. Asian Journal of Plant
Science and Research 2(2) 115-122.

© Copyright 2014 | Centre for Info Bio Technology (CIBTech) 27