PUBLICATIONS - INFLIBNETshodhganga.inflibnet.ac.in/bitstream/10603/106806/20/20...as Sansevieria...

PUBLICATIONS

170

LIST OF PUBLICATIONS

I. Hanumanth kumar G and Pramoda kumari J (2015) Phytochemical

analysis of secondary metabolites and antimicrobial activity of Sansevieria

roxburghiana. Wr.J.Pha.Res, 4(2): 1072-1077. (SJIF Impact factor-5.99)

II. Hanumanth kumar G and Pramoda kumari J (2015) Lead effect on the

mineral composition in Sansevieria roxburghiana Schult. and Schult. f.

Int.J.Sci.Eng.Res, 6(1): 31-35. (Impact factor-3.2)

III. Hanumanth Kumar G and Pramoda kumari J (2014) Phytoextraction of

weed plants by survey and analysis in response to lead accumulation.

Int.J.Chem.Bio.Sci, 1: 11-1-8. (Impact factor-0.528)

IV. Hanumanth Kumar G and Saritha KV (2014) Lead stress-induced changes

of antioxidant enzymes and biochemical compounds in selected weeds and

their use for phytoremediation. Int.J.Bioinfo.Bio.Sci, 4 (1):1-8.

V. Hanumanth Kumar G and Saritha KV (2014) Lead uptake and its effects

on antioxidant defence system in Sansevieria roxburghiana schult & schult.

F. Int.J.Chem.Res.Pharm.Sci, 1(5): 33-39. (ISSN-2319-5169). (Impact

factor-0.5)

VI. Hanumanth kumar G and Pramoda kumari J (2015) A study on

phytoremediating efficiency of pb, cu and zn in Sansevieria roxburghiana

shult and shult.f under different metal contaminated soils. Int.J.

Pharm.Sci.Res, (Impact factor-2.5) (Accepted).

VII. Hanumanth Kumar G and Pramoda kumari J (2015) Evaluation of lead

toxicity in Sansevieria roxburghiana shult & shult.F under Hydroponic

Study. 3 Biotech. (Under review).

VIII. Hanumanth Kumar G and Pramoda kumari J (2015) Heavymetal Lead

Influative toxicity and its Assessment in Phytoremediating plants. Water soil

and Pollution research. (Under review).

www.wjpr.net Vol 4, Issue 2, 2015.

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Pramoda & Hanumanth. World Journal of Pharmaceutical Research

PHYTOCHEMICAL ANALYSIS OF SECONDARY METABOLITES

AND ANTIMICROBIAL ACTIVITY OF SANSEVIERIA

ROXBURGHIANA

G. Hanumanth kumar1 and J. Pramoda kumari

2*

1Department of Biotechnology, Sri Venkateswara University, Tirupati-517502, India.

*2Department of Microbiology, Sri Venkateswara University, Tirupati-517502, India.

ABSTRACT

In the present investigation, different solvent soluble fractions of

leaves of Acetone (F1), Ethanol (F2), methanol (F3), chloroform (F4)

and Ether (F5) of Sansevieria roxburghiana Schult. and Schult.f.

(Agavaceae) was evaluated for the preliminary analysis of phyto

chemicals and antimicrobial investigation against clinically significant

bacterial strains. Qualitative analysis of the selected parts confirmed by

the presence of various primary and secondary plant metabolites such

as alkaloids, terpenoids, flavonoids, saponins, steroids, phenols,

tannins, and quinone. Antimicrobial assays revealed significant

antimicrobial activity against bacteria such as Proteus vulgaris,

Salmonella typhi, Pseudomonas aeruginosa, Klebsiella pneumoniae,

and Escherichia coli. The current study suggests that Sansevieria roxburghiana contain

potential secondary metabolites with effective antimicrobial activities, but still research

findings needs to authenticate which compound is effective from the combination.

KEYWORDS: Sansevieria roxburghiana, phytochemicals, secondary metabolites,

Antimicrobial activity.

INTRODUCTION

Sansevieria roxburghiana belongs to Agavaceae family (L), is one of hemp species with

concave, short petioled leaves that are in part transversely banded with light and dark green,

also linearly striated with whitish to light green and dark green striations ( Jeeva et al., 2012).

This plant has long rhizomes with rapid rate of growth containing long fibrous roots and are

grown ornamentally (USDA, 2008). Nearly 70 numbers of species exists in Sansvieria such

World Journal of Pharmaceutical Research

SJIF Impact Factor 5.045

Volume 4, Issue 2, 1072-1077. Research Article ISSN 2277– 7105

Article Received on

29 Nov 2014,

Revised on 23 Dec 2014,

Accepted on 18 Jan 2015

*Correspondence for

Author

J. Pramoda kumari

Department of

Microbiology, Sri

Venkateswara University,

Tirupati-517502, India.


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as Sansevieria trifasciata, Sansevieria ehrenbergii, Sansevieria guineensis, Sansevieria

longiflora, Sansevieria zeylanica etc.

Medicinal plants continue to be an important therapeutic aid for alleviating the ailments of

humankind (Harini et al., 2011). In India, this plant has been traditionally used for several

medicinal purposes. The whole plant is traditionally used as a cardiotonic, expectorant,

febrifuge, purgative, tonic, in glandular enlargement and rheumatism (Aneesh, 2009). Recent

studies on S. roxburghiana rhizome demonstrated remarkable anti diabetic activity in STZ

(streptozotocin) induced diabetic rats (Pallab et al., 2010). The leaf sap is applied directly to

infected sores, cuts and grazes (Sardessai et al., 2010).

In current study, our effort is to identify potential bioactive secondary metabolites and

unveiling the possible antimicrobial activities of leaf extracts of Sansevieria roxburghiana.

MATERIAL AND METHODS

Plant material

Sansevieria roxburghiana was collected from Sri Venkateswara University, India. Botanical

identification of the plant was done by K.Madhavachetty, Assistant professor, Department of

Botany, Sri Venkateswara University, India.

Solvent extracts

10g of pulverized leaf material was mixed with 100ml of each solvent of acetone, ethanol,

methanol, chloroform and ether are kept in rotary shaker at 100 rpm overnight and filtered

with Whatman No.1 filter paper and concerted to dryness at 40°C, lyophilized and stored at

4°C until further use.

Antibacterial Activity

Dried fractions Acetone (F1), Ethanol (F2), Methanol (F3), Chloroform (F4) and ether (F5)

were dissolved in 5% DMSO (Dimethylsulphoxide) so as to get a concentration of 1 mg/ml.

500µl of this were applied on the disc and allowed to diffuse for half an hour at 4ºC and

incubated at 37ºC for 24hrs. Azithromycin (F6) served as control (C).The plates were

observed for the presence of inhibition of bacterial growth that was indicated by the clear

zone around the disc.


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Test for alkaloids

About 0.2 g of each of the fractions was warm with 2% H2SO4 for two minutes. The reactants

were filtered and added a few drops of Dragendroff’s reagent to each filtrate. Orange red

precipitate indicates the presence of alkaloids moiety.

Test for flavonoids

About 0.2 g of each extract was dissolved in diluted NaoH and few drops of HCl were added.

A yellow solution that turn colorless indicates the presence of flavonoids.

Test for tannins

A small quantity of each extract was mixed with water and heated on water bath and filtered.

A few drops of ferric chloride were added to each filtrate. A dark green solution indicates the

presence of tannins.

Test for saponins

0.2g of each extract was shaken with 5 ml of distilled water and heated to boiling. Frothing

(appearance of creamy miss of small bubbles) shows the presence of saponins.

Test for anthraquinone

About 0.5g of each extract was boiled with 10 % HCl for few minutes on water bath. The

reaction mixture was filter and allows to cool. Equal volume of CHCl3 was added to each

filtrate. Few drops of 10 % ammonia was added to each mixture and heated. Rose pink color

formation indicates the presence of anthraquinone.

Test for steroids

2ml of acetic anhydride was added to the mixture of 0.5g of each extract and H2SO4 (2ml).

The color change from violet to blue or green in some samples indicates the presence of

steroids.

Test for terpenoids

About 0.2g of each extract was mixed with 2ml of chloroform and concentrated H2SO4 (3ml)

was carefully added to form a layer. The formation of a reddish brown coloration at the

interface indicates positive results for the presence of terpenoids.


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Phenols

To 1ml of the extract, 2ml of distilled water followed by few drops of 10% ferric chloride

was added. Formation of blue or green color indicates presence of phenols.

RESULTS AND DISCUSSION

The phytochemical screening of the leaf parts of Sansevieria roxburghiana revealed the

presence of bioactive secondary metabolites such as alkaloids, saponins, steroids, terpenoids,

tannins, and phenols (Table 1).

Table 1. Phytochemical profile of Sansevieria roxburghiana

Phytochemical

components

Acetone Ethanol Methanol Chloroform Ether

Alkaloids ++ - +++ ++ +

Flavinoids - + - - +

Saponins - + + - -

Steroids + - ++ ++ ++

Terpenoids - - + -

Tannins ++ + ++ + -

Phenols - - +++ - -

Quinones - - + + +

+, ++, +++ Moderate, High, Very high

The phytochemical screening revealed that shown high presence of alkaloids in methanol

extract compared to acetone, chloroform and ether. Flavonoids are present in ethanol and

ether in moderate proportions; saponins are present in ethanol and methanol in moderate

proportions. Steroids shown higher proportions in methanol, chloroform and ether and

moderate in acetone; Terpenoids presence is shown in chloroform and absent in all rest of the

extracts. Tannins are high in Acetone and methanol and moderate in ethanol and chloroform.

Phenols are very high only present in methanol fractions, quinones are present in methanol,

chloroform and ether in moderate levels. Overall extracts of S. roxburghiana leaves showed

the strong presence of, alkaloids, steroids, terpenoids, tannins and phenols. All these have

potential health promoting effects, at least under different circumstances (Basu et al., 2007).

The antimicrobial activity by using disc diffusion method evaluated that methanolic extract

was found to have significant activity against all strains. All the bacteria showed better zone

of inhibition against 5 fractions (Table 2).


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Table 2. Antimicrobial activity of Sansevieria roxburghiana.

Microorganism Zone of inhibition(mm)

Acetone

(500 µl)

mm

Ethanol

(500 µl)

mm

Methanol

(500 µl)

mm

Chloroform

(500 µl)

mm

Ether

(500µl)

mm

Azithromycin

(500µl) mm

Escherichia coli 5±0.2 7±0.34 10±0.13 8±0.65 6±0.5 5±0.4

Klebsiella

pneumoneae

8±0.3 11±0.12 13±0.17 7±0.22 8±0.1 6±0.1

Pseudomonas

aeuroginosa

7±0.7 8±0.56 12±0.24 3±0.19 3±0.6 3±0.4

Salmonella typhi 5±0.2 9±0.16 14±0.15 9±0.11 7±0.3 5±0.5

Proteus vulgaris 4±0.1 8±0.24 4±0.16 6±0.06 5±0.4 3±0.2

Mean ± SD, n=3

Escherichia coli showed 5±0.2, 7±0.34, 10±0.13, 8±0.65, 6±0.5; Klebsiella pneumoneae

showed 8±0.3, 11±0.12, 13±0.17, 7±0.22, 8±0.1; Pseudomonas aeuroginosa showed 7±0.7,

8±0.56, 12±0.24, 3±0.19, 3±0.6; Salmonella typhi showed 5±0.2, 9±0.16, 14±0.15, 9±0.11,

7±0.3; Proteus vulgaris showed 4±0.1, 8±0.24, 4±0.16, 6±0.06 and 5±0.4 in Acetone,

ethanol, methanol, chloroform and ether solvent extracts compared to Azithromycin acts as

control (5±0.4, 6±0.1, 3±0.4, 5±0.5 and 3±0.2). Among all extracts methanol leaf extract

exhibited high antimicrobial activity in all strains of about 14 ± 0.15 activity, comparable to

acetone (7±0.7) ethanol (11±0.12), chloroform (9±0.11) and ether (6±0.1).

Fig 1: Sansevieria roxburghiana leaf pure fractions (F1, F2, F3, F4 & F5 compared with

Azithromycin exhibiting zones of inhibition against five pathogenic microbes.

(A) Escherichia coli (B) Klebsiella pneumoneae (C) Pseudomonas aeuroginosa (D)

Salmonella typhi (E) Proteus vulgaris


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This antimicrobial activity of the 5 bacterial species implies that this plant can contribute

significantly therapeutic use. The information obtained above can be in the identification of

the plant. This study has revealed that the presence of important phytochemical analysis of

the plant leaves can be for therapeutic usage. These results support the medicinal use of the

plant, and in addition, unveil the possibility of its acting as a potential source of secondary

metabolites.

REFERENCES

1. Aneesh TP, Mohamed H, Sonal SM, Manjusree M, Deepa TV. International Market

scenario of Traditional Indian Herbal drugs- India declining, 2009; 184-190.

2. Yogita Sardessai, Gauri Pai Angle, Arun Joshi, Sonia Carvalho, Maya bhobe. Anti

microbial Activity of Methanolic Extract of the Rhizomes of Costus igneus. J Pharm

Chem Biol Sci, 2014; 2(3): 176-185.

3. Pallab K. Haldar, Biswakanth Kar, Sanjib Bhattacharya, Asis Bala R, Suresh Kumar B.

Antidiabetic activity and modulation of antioxidant status by sansevieria roxburghiana

rhizome instreptozotocin-induced diabetic rats. Diabetologia croatica, 2010; 39-4.

4. D. Jeya Sheela, S. Jeeva, I. M. Ramzan Shamila, N. C. J. Packia Lekshmi and J. Raja

Brindha. Antimicrobial activity and phytochemical analysis of Sanseiveria roxburghiana

leaf. Asian Journal of Plant Science and Research, 2012; 2(1): 41-44.

5. Dhanalakshmi D, Kumar S, Prasad MS, Koli V, Kumar BP., and Harani A. Eur. J. Exp.

Bio, 2011; 1(1): 103-105.

International Journal of Scientific & Engineering Research, Volume 6, Issue 2, February-2015 31 ISSN 2229-5518

IJSER © 2015 http://www.ijser.org

Lead Effect on the Mineral Composition in Sansevieria roxburghiana Schult. and Schult. f.

G.Hanumanth kumar and J.Pramoda kumari*

Abstract: The present investigation reported that mineral levels in plant Sansevieria roxburghiana Schult. and Schult. f. (Agavaceae) exposed to different lead (Pb) concentrations from 60, 120, 180, 240 and 300 mg/l under hydroponics for 40 day duration. Pb accumulation (13, 24, 55, 71 and 98 mg/kg DW) was reported in different concentrations. Results showed that the decrement in mineral levels sodium (598±0.3 to 454±0.2 mg/100g DW), potassium (18±0.2 to 6±0.4 mg/100g DW), calcium (245±0.3 to 122±0.2 mg/100g), zinc (0.14±0.5 to 0.7±0.4 mg/100g DW), copper (0.65±0.2 to 0.44±0.1 mg/100g DW), manganese (0.55±0.2 to 0.25 ±0.1mg/100g DW) and iron (2±0.3 to 0.73±0.4 mg/100g DW). Pb accumulated to a greater degree in plants resulted in oxidative stress, as evidenced by increased concentrations of malondialdehyde. This result clearly indicates potential lead toxicity affects the nutritive status of Sansevieria roxburghiana plant.

Keywords: Lead, Minerals, Nutritional potentiality, Oxidative stress, Sansevieria roxburghiana.

—————————— ——————————

1. INTRODUCTION

Industrialization has led to the increases several heavy metals like Cd, Pb, Zn, Cu, and Hg in the soil environment. High levels of heavy metals in the soil adversely affect plant growth. Among heavy metals; lead is an element that is easily accumulated in soil and sediments. The level of Pb in the environment is currently of great concern. Although lead is not an essential element for plants, it is absorbed and accumulated due to different man made polluted activities [1]. The absorption of metals from the soil by plants is

Department of Biotechnology,* Department of Microbiology, Sri Venkateswara University, Tirupati-517502, India. Corresponding author email:[email protected]

influenced by a variety of factors, including pH, temperature, soil ions, cation exchange capacity of soil, organic matter content of the soil, the type and concentration of metal and the species of plant [2].

The action of metals is seen at the whole plant level in reduced growth, and at the organ level in leaf symptoms. At a smaller scale, the effects of metals can be seen as cellular symptoms. Symptoms, both macro and micro cellular, and growth effects are side effects of the direct mode of action. Each metal has a different mode of action. However, in general, metal toxicity has been shown to reduce photosynthesis, affect enzyme and protein production and utilisation, alter nutrient transport and has negative effects on cellular functioning [3, 4].

IJSER



Sansevieria roxburghiana Schult. & Schult. f. (Agavaceae), called chaga in Telugu and Marul in Tamil, Indian bowstring hemp in English is an herbaceous perennial plant with short fleshy stem and stout rootstock, occurring in eastern coastal region of India, also in Sri Lanka, Indonesia and tropical Africa. In India, this plant has been traditionally used for several medicinal purposes. There are 16 essential nutrients in these plants. These are carbon (C), hydrogen (H), oxygen (O), nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg), iron (Fe), sulphur (S), zinc (Zn), manganese (Mn), copper (Cu), boron (B), molybdenum (Mo) and chlorine (Cl). These nutrient elements have to be available to the crops in quantities as required for a yield target. Any limiting or deficient nutrient (or nutrients) will limit crop growth [5]. These are helpful for the plant in photosynthesis and plant structure, stomatal function, enzyme activation, C4 metabolism etc.

The objective of this study is the levels of heavy metal lead influence on selected minerals such as sodium, potassium, calcium, manganese, iron, copper and zinc in Sansevieria roxburghiana, a medicinal plant.

2. MATERIAL AND METHODS

2.1 Plant material

Sansevieria roxburghiana was collected from Sri Venkateswara University, India. Botanical identification of the plant was done by K. Madhava chetty, Assistant professor, Department of Botany, Sri Venkateswara University, India.

2.2 Experimentation

Sansevieria roxburghiana plants (60 day-old plants) were hydroponically grown with 0, 60, 120, 180, 240 and 300 mg/l for 40 days period grown in Hoagland's medium (Hoagland and Arnon, 1950)[6]. The experiment was repeated for three times with six replicates. For the experimental procedure, plants were maintained with 16-h photoperiod (PAR 200 μmol m-2 s-1, temperature 25 ±1°C, relative air humidity 50 - 60%). The nutritional solution was changed every 7 days and aerated every day.

2.3 Determination of lead and mineral nutrients

Harvested plants were successively oven-dried at 1050C (for 30 min) and at 800C until they reached stable weight. These dry materials were analyzed for mineral nutrients (Pb) lead, calcium (Ca), sodium (Na) copper (Cu), iron (Fe), potassium (K), manganese (Mn) and zinc (Zn) quantities. The biomass of the oven-dried plant tissues was measured using an electronic scale. Then, the tissues were ground into a fine powder, digested in to a HNO3 and HClO4 solution (3:1; v/v), and heated at 1200C for over 3 h. All measurements were performed using an atomic absorption (Analyst 200, Perkin Elmer, UK) [7, 8].

2.4 Data analysis The significance of differences between the means of the treatments was evaluated by one way analysis of variance followed by Pearson correlation test at the significance level of P<0.05. The statistical SPSS version 12 was used for the analysis. 3. RESULTS AND DISCUSSION

IJSER



Pb accumulation from 60-300 mg was 13, 24, 55, 71 and 98 mg/kg DW. Pb accumulation to a greater degree inhibited the mineral nutrition. However, changes was found in root and shoot length also. Pb accumulation of the treatments with different doses indicates that the Pb uptake is dose dependent (Table-1).

Table-1. Accumulation levels of Pb in S.roxburghiana

Pb supplemented (mg/l) Pb accumulated (mg/kg DW)

Control 00

60 13

120 24

180 55

240 71

300 98

The mean concentration levels of mineral found in Sansevieria roxburghiana were summarized in Table -2. The mineral content shows potassium, zinc, manganese, copper and iron were main constituents of the watery mucilage part of this plant. Pb application significantly decreased in mineral content of treated plants when compared to controls. Mineral contents of all samples were reduced to Na (454±0.2), K (6±0.4), Ca (122±0.2), Zn (0.7±0.4), Cu (0.44±0.1), Mn (0.25 ±0.1) and Fe (0.73±0.4) from controls. Vast differences were established in Fe and K. These differences could probably be the result of lead toxicity. The ion balance in a cell is tightly linked with plant acclimation to heavy-metal phytotoxicity [9]. Furthermore, this is confirmed according to the Pearson correlation analysis, the significance was obtained as P< 0.05 (Table 2).

Table 2.Variations in mineral composition under 60-300 mg lead concentrations.

Critical Pb2+ activity caused decrease in K concentration in shoot of cowpea [10]. Ca concentrations in shoots of A. gangeticus with Pb were agreed with that of other workers. For

Values are mean ± SD of six replicates

example, Ca concentrations decreased in grain, straw and roots of rice, shoot and root of radish and leaf, stem and root of Indian spinach due to Pb application [11]. The calcium content of Sanseviera roxburghiana is comparable to those of Boerhavia diffusa, Commelina nudiflora and soybeans [12]. Zinc content of shoots and roots of A. gangeticus and roots of A. oleracea decreased significantly with increasing rate of Pb application showing a negative relation between Pb and Zn [2].

Copper deficiency or excess can have a negative impact on plants. Similar decrease was observed in rye plants according to Szatanik-kloc et al 2014.The Cd contamination decreased the nutrients in the order of Zn > Mn > Na for Phaseolus vulgaris [13].

Mineral Variation in mineral composition under different

lead concentrations

60 mg

Pb

120 mg

Pb

180 mg 240 mg 300 mg

Sodium 592±0.3 557±0.5 520±0.4 476±0.3 454±0.2

Potassium 15±0.1 12±0.2 10±0.1 8±0.2 6±0.4

Calcium 240±0.3 213±0.2 180±0.3 166±0.3 122±0.2

Zinc 0.13±0.5 0.11±0.5 0.9±0.5 0.6±0.5 0.7±0.4

Copper 0.61±0.2 0.58±0.1 0.52±0.2 0.48±0.5 0.44±0.1

Manganese 0.52±0.2 0.49±0.2 0.41±0.2 0.37±0.2 0.25±0.1

Iron 1.98±0.3 1.62±0.3 1.16±0.5 0.94±0.3 0.73±0.4 IJSER



Table-3.Mineral composition effected by lead for 40 day period Sansevieria roxburghiana

Minerals mg dw/100 g

(Controls)

Reduced

mg/kg

dw/100 g

% ge

Varied

Sodium 598±0.3 454±0.2 75.9

Potassium 18±0.2 6±0.4 33.3

Calcium 245±0.3 122±0.2 49.7

Zinc 0.14±0.5 0.7±0.4 50.0

Copper 0.65±0.2 0.44±0.1 67.6

Manganese 0.55±0.2 0.25 ±0.1 45.4

Iron 2±0.3 0.73±0.4 36.5

Values are means ± SD of six replicates

Mineral composition effected by lead for 40 day period Sansevieria roxburghiana was reported in table-3.The optimally balanced mode of mineral ions plays a central role in plant metabolism [14]. However, no consensus exists with regard to the effects of Pb on mineral absorption because there are inconsistent results due to discrepancies and interactions between heavy-metal ions and plant tissues.

4. CONCLUSION

Lead application in our study significantly decreased mineral conntent of S.roxburghiana. The reductions of minerals were of S.roxburghiana were Na (75%) K (33%) Ca (49%) Zn (50%) Cu (67%) Mn (45%) and Fe (36%), when compared with control (Table 3). A decrease in all minerals up to 120 mg lead was not affected severely but from 180–300 mg lead concentrations showing a drastic decrease in minerals of shoots of Sansevieria roxburghiana under Pb application.

REFERENCES

[1] Kabata–Pendias, A. Trace elements in soils and plants. 3rd Ed. CRC Press, Boca Raton, USA. Pp 413. 2011.

[2] Elegbede, J.A. “Legumes”. In: Osagie, A.U. and Eka, O.U. (eds). Nutritional Quality of Plant Foods. 120-133. 1998.

[3] Dim LA, Funtua II, Oyewale AO, Grass F, Umar IM, Gwozdz R, et al. Determination of some elements in Ageratum conyziodes, a tropical medicinal plant, using instrumental neutron activation analysis. J Radioanal Nucl Chem, 261: 225-228. 2004.

[4] Arzani A, Zinali H, Razmjo K. Iron and magnesium concentrations of mint accessions (Mentha spp.). Plant Physiol Biochem, 45: 323-329. 2007.

[5] Kovacik J, Babula P, Hedbavnya J, Svec P. Manganese induced oxidative stress in two ontogenetic stages of chamomile and amelioration by nitric oxide. Plant Sci, 1(10):215–216.2014.

[6] Hoagland, D.R., Arnon, D.I. The water-culture method for growing plants without soil. In: California agricultural experiment station, circ 347, 2nd edn.1950.

[7] Motsara M.R., Roy R.N. Guide to laboratory establishment for plant nutrient analysis. ISBN 978-92-5-105981-4.FAO publishers. 2008.

[8] Zhang X, Gao B, Xia H. Effect of cadmium on growth, photosynthesis, mineral nutrition and metal accumulation of banana grass and vetiver grass. Ecotoxicol Environ Saf 106:102–108. 2014.

[9] Xu L, Dong Y, Kong J, Liu S.Effects of root and foliar applications of exogenous NO on alleviating cadmium toxicity in lettuce seedlings. Plant Growth Regul, 72:39–50. 2014.

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[10] Kopittke, P.M., C.J. Asher, R.A. Kopittke and N.W. Menzies. Toxic effects of Pb2+ on growth of cowpea (Vigna unguiculata). Environmental Pollution, 150: 280-287. 2007.

[11] Kibria, M.G. Dynamics of cadmium and lead in some soils of Chittagong and their influx in some edible crops. Ph. D. Thesis, University of Chittagong, Bangladesh. 2008.

[12] Ujowundu, C.O., C.U. Igwe, Enemor V.H.A., Nwaogu, L.A. and Okafor, O.E.. Nutritional and Anti-Nutritive Properties of Boerhavia diffusa and Commelina nudiflora Leaves. Pak. J. Nutr. 7(1):90-92.2008.

[13] Vinod Kumar, A. and Chopra, K. Ferti-irrigational impact of sugar mill effluent on agronomical characteristics of Phaseolus vulgaris (L.) in two seasons. Environ Monit Assess 186:7877–7892. 2014.

[14] Xu L, Dong Y, Kong J, Liu S. Effects of root and foliar applications of exogenous NO on alleviating cadmium toxicity in lettuce seedlings. Plant Growth Regul, 72:39–50. 2014.

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IJCBS RESEARCH PAPER VOL. 1 [ISSUE 8] NOVEMBER, 2014 ISSN:- 2349–2724

© Virtu and Foi.

A Non-Paid Journal

16

www.ijcbs.org

PHYTOEXTRACTION OF WEED PLANTS BY SURVEY

AND ANALYSIS IN RESPONSE TO LEAD

ACCUMULATION

1Hanumanth Kumar.Gurijala Department of Biotechnology Sri Venkateswara University

Tirupati-517502, India Email: [email protected]

2Pramoda kumari.Jasti Department of Microbiology

Sri Venkateswara University Tirupati-517502, India

Corresponding author Email: [email protected]

ABSTRACT: Field experiments were conducted on lead contaminated spoil dump. Plants growing on spoiled dump were collected for phytoremediation (phytoextraction) studies in order to determine whether the plants would thrive in contaminated soil and undergo phytoremediation. Lead accumulation level in soil was 164 mg/kg. Pb in plants was determined by atomic absorption spectrophotometer. Lead (Pb) accumulation levels in dry weight (DW) of plant samples were Bidens pilosa (90.53±8.86 mg/kg), Parthenium histocarpus (81.53±3.37 mg/kg ), Spermococa pusilla (74.9±7.21 mg/kg), Tinospora cordifolia (74.53±3.31 mg/kg), Hemidesmus indicus (74.73±6.80 mg/kg), Atropa belladonna (73.26±5.88 mg/kg), Cyperus flavidus (51.33±4.71 mg/kg), Jatropha gossypifolia (55.2±5.55 mg/kg),Typha angustata (57.1±2.96 mg/kg), Holoptela integrefolia (70.4±6.78 mg/kg), Crotolaria verrucosa (43.53±2.43 mg/kg), Senna occidentalis (27.4±2.07 mg/kg), Alternanthera ficoidea (36.66±5.41 mg/kg), Amaranthus spinosus (66.13±1.10 mg/kg ), Sansevieria roxburghiana (84.1±1.02 mg/kg), Achnatherum hymenoides (70±3.01 mg/kg ), Eichorniafasculata (10± 0.97 mg/kg), Anisomilus molabarica (29± 3.69 mg/kg), Ipomea purpurea (72±4.34 mg/kg), Bromustectorum (77±4.48 mg/kg) and Calotropisprocera (90±2.55 mg/kg). It was found that all the plants were able to accumulate in Pb contaminated site. Among all the plants taken, Bidens pilosa, Calotropisprocera and Sansevieria roxburghiana showed highest lead accumulation and Eichornia fasculata showed lowest accumulation. Based on the atomic absorption spectrometric determination of the plant samples, it was confirmed that these suitable plant species are having potential for lead phytoremediation.

Keywords: Atomic absorption, thrive, lead phytoremediation INTRODUCTION

Phytoextraction is a green in situ technology

which aims to diminish the concentration of the

chemical element(s) (often synonymous with

heavy metals) of contaminated soils to such a

level that the soil can be used without danger for

agriculture, horticulture, forestry or amenity.

(Ernst et al., 2005). It has been demonstrated

that the success of phytoextraction depends on

the degree of soil contamination and on the

number of metals in surplus at the site, the

metal-resistance of the plant species.

In mine tailings and other metal-

contaminated soils the distribution of the heavy

metals is not homogenous. Heavy metal

contamination of soil is widespread and there is

a risk of transfer of toxic and available metals to

humans, animals, and agricultural crops. If they

are phytoavailable, some toxic metals are

potentially accumulated in some plants and may

pose a threat to humans and grazing animals.

Heavy metal contamination of soil varies

horizontally with soil depth (Whiting et al.,

2000; Haines, 2002; Podar et al., 2004;

Chehregani et al., 2009). Heterogeneity in plants

modifies growth pattern of roots and root

morphology, resulting in ecotype-specific root

production in soils with heterogeneously

distributed heavy metals (Hodge, 2009).

For present study, the major source of

heavy metal contamination is lead which is

possibly emitted from industrial area. These

emissions transported by air, sewage water and

industrial sewage effluents have been

considered to be responsible for the increased

heavy metal concentrations found in the soils of

the central studied area. Lead concentration up

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to 164 mg /kg was found in most pollutant area.

Concerning the health risk of the population

bioavailability and mobility of heavy metals

seems to be of major importance, based on the

soil properties found in the study area. In order

to recuperate the affected lead contaminated

site by employing plants capable of

accumulating high levels of contaminants.

Periodical field surveys have been made to

identify the metal-tolerant species that are

spontaneously growing in the polluted soils, and

are able to uptake lead.

The objective of this study is to carry out a

preliminary assessment of lead accumulation in

weed plants potentially growing in Pb

contaminated site.

MATERIALS AND METHODS

Study Site

The experiment was conducted in lead

contaminated area near Tirupati. All the

experimental weed plants were grown under

natural conditions; neither agricultural practices

nor irrigation were carried out. The plants used

for these experiments were similar in size.

Soil sampling

A total of 6 soil samples were collected from of

the lead contaminated area. Another six samples

of references were also collected from an area of

20 kilometers away from the industrial complex.

Soil samples were collected from the surface of

the soil (0-20 cm deep) and preserved. From

each sampling points, five Soil samples were

gathered and mixed properly to obtain a

composite sample mixture. The soil sampling

spots were cleared of debris before sampling.

Each composite soil samples were placed in

cellophane bags labeled and were taken to the

laboratory for pre-treatment and analysis. In the

laboratory, bulk soil samples were spread on

trays and were air dried at ambient conditions

for two weeks. The samples were then grounded

by mortar and pestle, sieved through a 2 mm

mesh, and oven-dried at 50°C for about 48

hours. The samples were then stored in

polyethylene bags and re-homogenized before

being used (USEPA, 2010).

Plant sampling

Weed plants were collected from contaminated

site near Tirupati, India. The collected plant

samples were thoroughly cleaned under running

water, then with distilled water, dried and

stored. Six replicates of each species were

collected, although sometimes an exhaustive

sampling was impossible for some species. In

addition to that plant samples from an

uncontaminated area (Sri Venkateswara

University) were also collected as a target set.

The collected plant specimens on the site were

botanically identified by Dr.K.Madhavachetty,

Taxonomist at Sri Venkateswara University,

Tirupati, Andhra Pradesh, India.

Plant and soil sample digestion

Plant and soil samples (dried weight) were

digested in a HNO3-HClO4 (3:1,v:v) mixture

and Pb concentrations were determined by

atomic absorption spectrophotometer

(AAnalyst 200, Perkin-Elmer, UK)(Liu et al.,

2010).

Statistical analysis

Statistics were analyzed with SPSS version 11.0,

and ANOVA was performed, with significance at

p<0.05. All values were mean of six independent

replicates.

RESULTS AND DISCUSSION

Soil characteristics

Table- 1 illustrates the metal concentration in

the studied area. The soil used in this

investigation had a pH of 5.2 ± 0.08. The analysis

of this soil showed the following composition:

Texture: sandy loam; organic carbon

(0.13±0.05); available N (68±1.20); available P

(0.33±0.05); available K (72±0.33).The analysis

revealed the contaminated site contain Pb

164.5±2.29 mg /kg in comparison with controls

(1.27±0.27).


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Table1: Soil characteristics and lead present in contaminated area

The following species formed the predominant

vegetation in the contaminated area during the

collecting time Bidens pilosa, Calotropis procera,

Sansevieria roxburghiana, Parthenium

histocarpus, Bromus tectorum, Tinospora

cordifolia, Hemidesmus indicus, Spermococa

pusilla, Atropa belladonna, Ipomea purpurea,

Holoptela integrefolia, Achnatherum hymenoides,

Amaranthus spinosus, Typha angustata, Jatropha

gossypifolia , Cyperus flavidus, Crotolaria

verrucosa, Alternanthera ficoidea, Anisomilus

molabarica, Senna occidentalis, and Eichornia

fasculata. Plant species belonging to the

Asteraceae, Rubiaceae, Menispermaceae,

Agavaceae, Amaranthaceae, Poaceae, Fabaceae,

Lamiaceae, Convolvulaceae and Asclepiadaceae

families are the most frequently found in the

contaminated soil.

Pb uptake potential of plants

Among the twenty one collected plant species

selected for their ability to grow on the lead

contaminated soil, and for their efficiency in

accumulating lead, the range of accumulation

varied from 10-90 mg/ kg DW (Table 2).

Table 2 Total lead concentrations in the weed plants of contaminated area

Mean (range) concentrations of heavy metal lead (mg /kg DW) for the species sampled (n=6)


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Accumulation and distribution of heavy metals

in plant tissues are important aspects to

evaluate the role of plant in remediation of

metalliferous soils (Pichtel and Bradway, 2008).

Pb increased from the values obtained in the

order from maximum to minimum was Bidens

pilosa, Calotropis procera, Sansevieria

roxburghiana, Parthenium histocarpus, Bromus

tectorum, Tinospora cordifolia, Hemidesmus

indicus, Spermococa pusilla, Atropa belladonna,

Ipomea purpurea, Holoptela integrefolia,

Achnatherum hymenoides, Amaranthus spinosus,

Typha angustata, Jatropha gossypifolia, Cyperus

flavidus, Crotolaria verrucosa, Alternanthera

ficoidea, Anisomilus molabarica, Senna

occidentalis, Eichornia fasculata. Here with

phytoextraction technique the risk of pollutant

dispersal into the environment is minimized.

Moreover, it is noticeable that Bidens pilosa,

Calotropis procera, Spermococa pusilla,

Tinospora cordifolia and Hemidesmus indicus

removal efficiencies are similar, which is

indicative of admitting that the absorption of

lead into the bodies is constant.

Table: 3 Illustrates the lead accumulation levels of roots, shoots and leaves in all experimental

plants

Mean ±SD for concentrations of heavy metal lead (mg /kg DW) for the species sampled (n=6).

The heavy metal, lead concentrations in the

shoots of the plants collected on polluted soils

were significantly higher than those of the roots

and leaves. The highest accumulation levels in

roots were observed in Calotropis procera

(90±2.2mg/kg DW), shoots Achnatherum

hymenoides (85.8±2.02mg/kg DW) and leaves

were observed in Bidens pilosa (123.6±2.0

mg/kg DW).In the overall plant parts taken the

highest lead levels were observed in leaves

followed by roots and shoots. Establishing the

agronomic management practices to the above-

mentioned species allow us to utilize their


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whole possibilities for cleaning up of

contaminated lead.

CONCLUSIONS

The present work verifies the usefulness of

collecting and studying plant species on

contaminated soils in order to identify and

select those plants with high potential to be used

in the remediation of the affected area. This

investigation showed that the susceptibility of

plants to lead was correlated with their growth.

Weed plants tolerated Pb at 164 mg/kg at the

growth stage of 60 days after germination.

Among the 21 different plant species belonging

to 17 different families highlight as the most

promising lead accumulators to be used in the

remediation of the affected area. Based on the

above observations, it was inferred that this

study allowed us to conclude that the above

stated weed plants could be considered

potentially feasible to clean up Pb soils. Among

them, we found the most effective weed plants

for phytoremediation of lead were Bidens pilosa,

Calotropis procera and Sansevieria roxburghiana.

ACKNOWLEDGMENTS

Authors acknowledged to Dr.K.Madhavachetty,

Taxonomist at Sri Venkateswara University,

Tirupati, Andhra Pradesh, India for his helping

hand in botanical identification of different plant

species.

REFERENCES

1. Chehregani, A., Noori, M., Lari Yazdi, H.b

2009. Phytoremediation of heavy-

metal-polluted soils: Screening for new

accumulator plants in Angouran mine

(Iran) and evaluation of removal ability.

Ecotoxicology and Environmental

Safety., 72: 1349-1353.

2. Ernst, W. 2005. Phytoextraction of mine

wastes – Options and impossibilities.

Geochemistry., 65: 29-42.

3. Haines, B.J. 2002. Zincophilic root for

aging in Thlaspi caerulescens. New

Phytol., 155: 363–372.

4. Hodge, A. 2009. Root decisions. Plant

Cell and Environment., 32: 628-640.

5. Liu, W.T., Zhou, Q.X., An, J., Sun, Y.B., Liu,

R. 2010. Variations in cadmium

accumulation among Chinese cabbage

cultivars and screening for Cd-safe

cultivars. J Hazard Mater., 17:737-743.

6. Podar, D., Ramsey, M.H., Hutchings, M.J.

2004. Effect of cadmium, zinc and

substrate heterogeneity on yield, shoot

metal concentration and metal uptake

by Brassica juncea: implications for

human health risk assessment and

phytoremediation. New Phytol., 163:

313-324.

7. Pichtel, J., Bradway, D.J. 2008.

Conventional crops and organic

amendments for Pb, Cd and Zn

treatment at a severely contaminated

site. Bioresource Technology., 99: 1242-

1251.

8. USEPA. 2007. Basic Information, Lead in

Paint, Dust, and Soil.

http://www.epa.gov/lead/pubs.lead

info.htm#facts. Accessed 22 May 2009.

9. Whiting, N.S., Leake, R.J., McGrat,h P.S.,

baker, M.J.A. 2000. Positive response to

Zn and CD by roots of the Zn and Cd

hyperaccumulator Thlaspi caerulescens.

New Phytol., 145:199-210.

http://www.epa.gov/lead/pubs.lead

3

Lead stress-induced changes of antioxidant enzymes and Biochemical Compounds in Selected Weeds and their

contribution for phytoremediation

G. Hanumanth Kumar and K.V. Saritha*

Department of Biotechnology, Sri Venkateswara University, Tirupati, A.P -517502, India

*Corresponding author: [email protected]

Abstract

Investigation was carried out to identify tolerant plant species growing in lead contaminated soil. Lead effected Chlorophyll a, chlorophyll b and ascorbicacidoxidase. Induced malondialdehyde and starch contents with up-regulated activities of antioxidative enzymes like catalase and phenylalanineammonialyase were observed. Enzymatic activities have been enhanced in all plants taken indicating that free radical generation was accelerated due to Pb exposure. Concentrations of lead accumulated were Calotropis procera (90 mg/kg) and Bromus tectorum (77 mg/kg) are higher followed by Ipomea purpurea (72 mg/kg), Parthenium histocarpus (33 mg/kg), Anisomilius molabarica (29 mg/kg) and Eichornia fasculata (10 mg/kg). Inspite of accumulation all plants survived successfully. With our findings we can suggest these plants can be used for effective phytoremediation of lead.

Keywords: Free radicals, Lead, Up-regulated, Malondialdehyde, Phytoremediation.

Heavy metal pollution of soils has been increasingly becoming a global problem which poses several risks to human health, decreased soil microbial activity, fertility and yield losses (McGrath, 1995). Inspite of all expensive extraction technologies like chelating and land filling, Phytoremediation is an emerging cheap cleanup technology, which uses green plants to remove, contain or render harmless environmental contaminants.

In heavymetals Lead (Pb) is one of the most abundant heavy metal pollutants in terrestrial and aquatic environments. Lead induces free radicals in various plant species by induction of lipid peroxidation and generation of reactive oxygen species which inhibits metabolic processes such as nitrogen assimilation, photosynthesis, respiration, water uptake and transcription (Boussama et al., 1999). Lead causes two types of unfavorable processes in biological systems: (I) it inactivates several

International Journal of Bioinformatics and Biological Sciences: v.4 n.1, p. 1-8. June 2014

14 International Journal of Biological Sciences: v.1 n.2 p. 9-17. June 2014

Kumar and Saritha

enzymes and (II) it can intensify the processes of reactive oxygen species (ROS) production leading to oxidative stress (Prasad et al., 1999). Reactive oxygen species produced can damage photosystem reaction center proteins by which photosynthesis could decrease and cause viability loss in the cells.

Plants initiate a signal transduction pathway that triggers enzymatic and non-enzymatic antioxidative systems that detoxify the cells (Dalton, 1995). In this mechanisms a wide series of enzymes exist in plants that serve to remove ROS, such as peroxidase, superoxidedismutases, ascorbate peroxidase, catalase, phenylalanine ammonialyase, ascorbicacidoxidase etc. (Verma and Dubey, 2003) thereby preventing the formation of OH radicals. These non-enzymatic and enzymatic systems in plant together keep the reactive oxygen species in low levels and not to be injured by accumulation of ROS. The purpose of this research is to exploit the ability of various plant species to thrive in high lead environments and its affect on antioxidative defense mechanism.

Materials And Methods

Study site

Lead contaminated industrial site (estimated production of about 12 tons per annum) and Pb free site (nearby forest) located at Tirupati, India is selected as a study site which is occupied by luxuriant natural vegetation. Selected weeds with abundant growth (a) Calotropis procera (b) Ipomea purpurea (c) Eichornia fasiculata (d) Parthenium histocarpus (e) Bromus tectorum (f) Anisomilus molabarica were collected from lead contaminated industrial area and nearby lead free site and samples of spoils were taken for laboratory analysis. Pb-contaminated soils were collected from an industrial site. The soil was screened to pass through a 1.0 cm sieve and thoroughly mixed before use. The following procedures were used to characterize the soil. Soil pH was measured using 1:1 soil/ water ratio; total soil Pb was determined by the atomic absorption spectrophotometer, organic matter content was measured by the Walkley Black method and particle size was measured by the hydrometer method. The selected physical and chemical properties of the Pb-contaminated soils are presented in Table 1.

Total chlorophyll content

Chlorophyll extraction was done according to method described by Wintermans and De Mots (1965) and expressed in mg g-1 FW.

TBARS assay

A TBARS (Thiobarbituric acid reactive substances) assay was performed following the method of Heath and Packer (1968).

International Journal of Biological Sciences: v.1 n.2 p. 9-17. June 2014 15

Lead stress-induced changes of antioxidant

Catalase activity

The Catalase activity was assayed by the method of Chance and Maehly (1955).

Phenylalanine ammonialyase activity

Phenylalanine ammonialyase activity was done as described by Dickerson et al., (1984).

Ascorbic acid oxidase activity

Determination of ascorbic acid oxidase was done according to method of Oberbacher and Vines (1963).

Total starch content

The total starch content was estimated by the anthrone–sulphuric acid method according to Hodge and Hofreiter (1962).

AAS analysis

The properties of soil sample were measured according to Liu (2000). Total metal concentration of Soil and plant metal concentration (digested by 5:1 with concentrated HNO3 and HClO4 were determined by atomic absorption spectrophotometry.


Statistical analysis was performed using spss version 11.5. All data were analyzed using one-way analysis of variance (ANOVA). Different letters in graphs or tables indicate significant differences at P<0.05. All values were presented as means ± standard error of the mean (SE), with a minimum of three replicates.

Results and Discussion

Lead uptake

In the present study Pb accumulation by different plants varied significantly (Anova, P < 0.05). Among the plants taken Calotropis procera showed highest amount of accumulation (90.5 mg/kg). This was about 5 times higher than in Eichornia fasculata (10.8 mg/kg) followed by Bromus tectorum (77.1 mg/kg), Ipomea purpurea (72.1 mg/kg), Parthenium histocarpus (33.7 mg/kg) and Anisomilius molabarica (29.7 mg/kg).

The Organic carbon content of the soils was determined as moderate which indicated that metals were less likely to be bound to organic matter to form metal-chelate complexes. This will make metals into available form for easy accessibility


Kumar and Saritha

to the weed plants. The P, K and N help the plants in up taking the metals in bounded forms. Soil particles are sandy and slightly acidic in both control and contaminated soils. Compared to control soils organic carbon (0.23±0.22) and phosphorous content (0.41±3.29) decreased in contaminate soils (0.13±0.35) (0.33±0.15) but nitrogen (55±2.26) and potassium contents (65±1.27) increased (68±1.20) (72±0.03) (Table 1).

Physical and Chemical properties of soil

Table 1: Physical and Chemical properties of soil

Parameters Control Soil Contaminated soil

Soil Texture Sandy loam Sandy loam

Soil PH 5.7±0.05 5.2±0.08

Organic Carbon (gm/kg) 0.23±0.22 0.13±0.35

Available N (gm/kg) 55±2.26 68±1.20

Available P (gm/kg) 0.41±3.29 0.33±0.15

Available K (gm/kg) 65±1.27 72±0.03Total Pb (mg/kg) 1.27 mg/kg 164.5 mg/kg

Effect of Pb on chlorophyll content

Chlorophyll content in plants is important factor in determining photosynthetic activity. In our present study, heavy metal Pb decreased chlorophyll content gradually by increased proportionality of metal concentration. Pb significantly effected phototosynthetic pigments (chlorophyll a and chlorophyll b) in weeds. Chlorophyll-a content was decreased from 62% to 34% in all plants when compared to controls with high decline in Anisomilius molabarica (62%), Ipomea purpurea (52%) and Eichornia fasculata (52%) followed by Parthenium histocarpus (47%), Calotropis procera (34%) and Bromus tectorum (40%) in comparision with controls (Fig. 1). Chlorophyll-b decreased from 81% (Bromus tectorum) to 36% (Calotropis procera) in all plants followed by Ipomea purpurea (76%), Anisomilius molabarica (75%), Parthenium histocarpus (63%) and Eichornia fasculata (56%) in comparision with controls (Fig. 2). The content of chlorophylls a, b chlorophyll was significantly decreased corroborates with findings of Salvinia minima wild by Jeffrey l. Gardner and Safaa h. al-hamdani (1997) and Zengin and Munzuroglu (2005) in phaseolus vulgaris L due to Pb stress. Decreased chlorophyll content associated with heavy metal stress may be the result of inhibition of the enzymes responsible for chlorophyll biosynthesis.



Effect of Pb on MDA content

During present study, a significant increase in malondialdehyde content from 25% to 137% was observed. Here it was observed excess of Pb promoted lipid peroxidation with excessive production of malondialdehyde content in contaminated over controls. The higher activity was observed in Calotropis procera (137%) and Eichornia fasculata (0.83%) followed by Anisomilius molabarica (42%), Bromus tectorum (33%), Parthenium histocarpus (26%) and Ipomea purpurea (25%) (Fig. 3). An enhanced level of lipid peroxidation observed may be due to the generation of toxic oxygen free radicals under metal stress. The ROS attack on free radicals and the polyunsaturated fatty acid components of membrane lipids initiated lipid peroxidation, an autocatalytic process that changes membrane structure and function. Our findings were similar to Qureshi et al., (2005) in Artemisia annua and Withania somnifera (Khatun et al., 2008) under copper toxicity.

Fig. 1 and 2: Effects of Pb concentration on Chlorophyll a ,Chlorophyll b content

Effect of Pb on Catalase activity

The Catalase increased from 63% to 19% compared to controls in all plants. Highest enhancement was observed in Calotropis procera (63%) and lowest in Bromus tectorum (19%) followed by Anisomilius molabarica (0.47%), Ipomea purpurea (0.37%), Eichornia fasculata (0.35%) and Parthenium histocarpus (0.25%) (Fig. 4). Elevated Catalase would lower H2O2 levels, which reduces the lipid peroxidation degree and lessen membrane damage by converting H2O2 into water and oxygen. Our results of increasing concentration of catalase activity with the increasing concentration of Pb corroborate with the findings of Verma and Dubey (2003) in rice plants and Pinus radiate plants (Jarvis and Leung 2002).


Kumar and Saritha

Fig. 3 and 4: Effects of Pb concentration on Catalase activity, and Phenylalanineammonialyase activity

Effect of Pb on Phenylalanineammonialyase activity

Increased Phenylalanineammonialyase activity could be a response to the cellular damage provoked by higher Pb concentrations. Unsurprisingly, phenylalanineammonialyase was more activated by the presence of lead. Significant increase was observed by 20% to 48% in all the plants in compared with controls. Parthenium histocarpus showed highest increase in phenylalanineammonialyase activity by 48% and lowest is seen in Ipomea purpurea (20%) followed by Anisomilius molabarica (46%), Calotropis procera (42%), Eichornia fasculata (37%) and Bromus tectorum (36%) compared with controls (Fig. 5). Overall phenylalanineammonialyase activity was elevated in all plants. Activation of PAL and lignin content increase is considered as common plant responses to various stress factors, including heavy metals (Yang et al., 2007; Kovacik and Klejdus 2008). So it seems that in weed plants the increase of PAL activity is efficient in catalyzing deamination reaction of the amino acid phenyl alanine at the gateway from the primary metabolism into the important secondary phenylpropanoid / phenolic metabolism in plants (Hahlbrock and Scheel 1989). Overall, Pb had the high inducement on Phenylalanineammonialyase. Similar observations were recorded in previous study under Nacl stress according to Simaei et al (2012) in Soybean Plants, rice under cadmium toxicity (Ting Hsu and Huei Kao 2004) and Luffa cylindrical under lead toxicity (Jiang et al., 2010).

Effect of Pb on Ascorbic acid oxidase activity

In our experiment, the exposure of plants to lead doses resulted, in decreased ascorbicacidoxidase content from 68% to 6% in all the plants compared with



controls. Calotropis procera and Ipomea purpurea showed highest decrease in ascorbicacidoxidase activity by 68% and lowest is seen in Parthenium histocarpus (6%) followed by Anisomilius molabarica (67%), Eichornia fasculata (63%) and Bromus tectorum (59%).compared with controls (Fig. 6). Our results accord with previous researches under Pb stress in ascorbicacidoxidase from findings of

Fig. 5 and 6: Effects of Pb concentration on

Ascorbic acid oxidase activity and lipid peroxidation levels

Anuradha et al., (2007) in radish, Ibrahim et al., (2013) in alfalfa under cobalt and copper toxicity and cucumber plants due to arsenate toxicity according to Czech et al., (2008). Ascorbicacidoxidase seems more important due to its role in plants not only as an antioxidant, but also as a signal molecule in many developmental and defense responses.

Fig. 7: Effects of Pb concentration on Starch content


Kumar and Saritha

Effect of Pb on Starch content

The increase in the starch content might be caused by excessive α-amylosis of the components. In our experiment significant increase of starch accumulation by lead occurred by 14% to 54% in comparision with controls (Fig. 7). Highest increase was observed in Eichornia fasiculata (54%) and lowest in Calotropis procera (14%) compared to controls followed by Ipomea purpurea (18%), Bromus tectorum (6%), Parthenium histocarpus (4%) and Anisomilius molabarica (2%). Increased starch levels were directly proportional with high concentration of heavy metal Pb. Nutrient deficiency and heavy metal toxicities are known to produce starch accumulation within leaves (Vazquez et al., 1987). C4 photosynthesis when compared to C3 allows fast biomass accumulation with high nitrogen and water use efficiency (Leegood and Edwards 1996; Sage 2004) which is desired set of trait to increase the productivity of crop plants (Matsuoka et al., 1998) may be the cause for increase in starch and a required character for successful phytoremediation. Similar findings were observed according to verma and dubey (2001) due to cadmium toxicity in rice and in Tilia argentea and Quercus cerris by Tzvetkova and Kolaro (1996). This may be due to higher resistance of their photosynthetic apparatus and low starch export from the mesophyll.

From the results it has been clear that the concentration of Pb toxicity on the plants posed high oxidative stress on activity of antioxidant enzymes and biochemical compounds in weed plants and both played a vital role in combating oxidative stress in plants for the plant susceptibility and tolerance. Among the plants calotropis procera and Bromus tectorum are efficient in combating the oxidative stress and accumulating higher amount of Lead than other species. Finally, data which is generated through this study will be very helpful in finding high tolerant plant species to thrive in high lead environments.

Acknowledgement

Author wants to thank Dr K.V. Saritha, Biotechnology Department of the Sri Venkateswara University, India for her technical assistance that enabled successful execution of this study.

References

Anuradha, S., Rao, S.S.R. 2007. The effect of brassinosteroids on radish (Raphanus sativus L.) seedlings growing under cadmium stress. Plant Soil and Environment, 53(11): 465-472.

Boussama, N., Quariti, O., Ghorbal, M.H. 1999. Changes in growth and nitrogen assimilation in barley seedlings under cadmium stress. Journal of Plant Nutrition, 22: 731-752.

Czech Viktoria, Palma Czovek, Jozsef Fodor, Karoly Boka, Ferenc Fodor., Edit Cseh 2008. Investigation of arsenate phytotoxicity in cucumber plants. Acta Biologica Szegediensis, 52(1): 79-80.



Chance, B., Maehly, A.C. 1955. Assay of catalases and peroxidases. Methods Enzymology, 2: 764-775.

Dickerson, D.P., Pascholati, S.F., Hagerman, A.E., Butler, L.G ., Nicholson, R.L. 1984. Phenylalanine ammonia-lyase and hydroxyl cinnamate: CoA ligase in maize mesocotyls inoculated with Helminthosporium maydis or Helminthosporium carbonum. Physiology and Plant Pathology, 25: 111-123.

Dalton, D.A. 1995. Antioxidant defenses of plants and fungi In: Sami A (ed) Oxidative stress and antioxidant defenses in biology. pp 298–354. Chapman and Hall, New York.

Hodge, J.E., Hofreiter, B.T. 1962. In: Methods in Carbohydrate Chemistry (Eds. Whistler RL, Be Miller JN): Academic Press New York.

Heath, R.L., Packer, L. 1968. Photoperoxidation in isolated chloroplasts. I. Kinetics and stoichiometry of fatty acid peroxidation. Archives of Biochemistry and Biophysics, 25: 189-198.

Hahlbrock, K., Scheel, D. 1989. Physiology and molecular biology of phenylpropanoid metabolism. Annual Review of Plant Physiology and Plant Molecular Biology, 4: 347–369.

Ibrahim Zeid, M., Ghazi, S.M., Nabawy, D.M. 2013. Alleviation of Co and Cr toxic effects on alfalfa. International journal of Agronomy and Plant Production, 4 (5): 984-993.

Jeffrey gardner I., Safaa al-hamdani H. 1997. Interactive Effects of Aluminum and Humic Substances on Salvinia. Journal of Aquatic Plant Management, 35: 30-34.

Jiang nan, xia luo. jin zeng, Zhi-rong yang, Lin-yong zheng., Song-tao wang. 2010. Lead toxicity induced growth and antioxidant responses in luffa cylindrica seedlings. International journal of agriculture and Biology, 12: 205-210.

Jarvis, M.D., Leung, D.W.M. 2002. Chelated lead transport in Pinus radiata: an ultrastructural study. Environmental and Experimental Botany, 48: 21–32.

Kovacik , J., Klejdus, B. 2008. Dynamics of phenolic acids and lignin accumulation in metal-treated Matricardia chamomilla roots. Plant Cell Reports 27: 605–615.

Khatun Serida, Mohammad Babar Ali, Eun-Joo Hahn., Kee-Yoeup Paek. 2008. Copper toxicity in Withania somnifera: Growth and antioxidant enzymes responses of invitro grown plants. Environmental and Experimental Botany 64: 279–285.

Qureshi Irfan, M., Israrb, M., Abdinb, M.Z., Muhammad Iqbal. 2005. Responses of Artemisia annua L. to lead and salt induced oxidative stress. Environmental and Experimental Botany, 53: 185-193.

Liu, D., Jiang, W., Liu, C., Xin, C., Hou. 2000. Uptake and accumulation of lead by roots, hypocotyls and shoots of Indian mustard (Brassica juncea L.). Bioresource Technolology, 71: 273-277.

Leegood, R.C., Edwards, G.E. 1996. Photosynthesis and the environment, Vol. 5, Kluwer Academic Publishers, Dordrecht.The Netherlands.

McGrath, S.P, Chaudri, A.M., Giller, K.E. 1995. Long-term effects of metals in sewage sludge on soils, microorganisms and plants. Journal of Industrial Microbiology, 14: 94-104.

PHARMACEUTICAL SCIENCES Int.J.Curr.Res.Chem.Pharma.Sci.1(5):33-39

© 2014, IJCRCPS. All Rights Reserved 33

RESEARCH ARTICLE

LEAD UPTAKE AND ITS EFFECTS ON ANTIOXIDANT DEFENCE SYSTEM IN SANSEVIERIAROXBURGHIANA SCHULT & SCHULT. F.

G. HAUMANTH KUMAR1 AND K.V. SARITHA*

1*Department of Biotechnology, Sri Venkateswara University, Tirupati-517502, A.P, India

*Corresponding Author: [email protected]

Introduction

Environmental pollution has become a burningproblem throughout the world over the past severaldecades. Many industrial processes are majorcontributors to extensive environmental pollution(Cunningham et al., 1995). Clean-up of Pb-contaminated soils is difficult. Existing methodssuch as mechanical removal and chemicalengineering are expensive, and are oftenincompatible with maintaining soil structure andfertility. Phytoremediation, i.e. the use of plantsystems to remove toxic elements from the soils,has recently attracted a great deal of attention as analternative means of soil decontamination, since it isa cost-effective, environmentally-friendly approach,applicable to large areas. Among all pollutants leadplays a predominant role and it exists in many formsin natural sources throughout the world. Accordingto the USA Environmental Protection Agency, Pb isone of the most common heavy metal contaminantsin aquatic and terrestrial ecosystems and can have

adverse effects on growth and metabolism ofplants, due to its direct release into the atmosphere(Watanabe, 1997).

The effect of lead depends on the concentration,type of salts, soil properties and plant speciesinvolved (Patra et al., 2004). There have been manyreports of Pb toxicity in plants (Choudhury & Panda,2005), including disturbance of mitosis (Wierzbicka,1998; Jiang & Liu, 2000), inhibition of root andshoot growth (Liu et al., 2009), induction of leafchlorosis (Pandey et al., 2007), reduction ofphotosynthesis (Xiao et al., 2008) and inhibition oractivation of several enzyme activities (Verma &Dubey, 2003; Sharma & Dubey, 2005; Liu et al.,2009). Plants use a diverse array of enzymes likesuperoxide dismutase (SOD; EC 1.15.1.1), catalase(CAT; EC 1.11.1.6), guaiacol peroxidase (POD; EC1.11.1.7), ascorbate peroxidase (APX; EC1.11.1.1), as well as low molecular weight

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Abstract

The effects of lead (Pb) stress on plant growth, lipidperoxidation and on the activity of antioxidant enzymes were studiedin Sansevieria roxburghiana Schult. & Schult. F., grown under hydroponical conditions in the absence and in thepresence of various concentrations (0.5, 1.0, 1.5, 2.0 and 2.5 g/l) of lead nitrate. The activity of antioxidant enzymesAscorbate peroxidase (APX), Guiacol peroxidase (GPX), and Glutathione-s-hydrogenase (GSH) was increased in plantsby lead treatment in a dose-related manner. The relative increase in enzyme activities demonstrated that Sansevieriaroxburghiana is more tolerant to Pb. Lipid peroxidation was enhanced in stressed Sansevieria roxburghiana with theincreased Pb concentration. The maximum accumulation of Pb (142.6 mg/kg DW, 322.0 mg/kg DW , 323.3 mg/kg DW,385.3 mg/kg DW and 536.6 mg/kg DW) occurred in plants followed by increase in enzyme activities proved this plantcan be effectively useful in removal of Pb from Pb contaminated sites..Keywords: Antioxidant enzymes, Lead, Sansevieria roxburghiana and dose-related.



antioxidants like cysteine, nonprotein thiol, andascorbic acid to scavenge different types of reactiveoxygen species (ROS), thereby protecting cell injuryand tissue dysfunction (Halliwell, 1987).

There is urgency in identifying effective leadphytoremediating species suitable for the particularecological condition of lead dumping sites. In thepresent study, Sansevieria was selected as suitablepb accumulator because of its survival in nearby Pbcontaminated sites. A number of species such asSansevieria cylindrica, Sansevieria ehrenbergii,Sansevieria guineensis, Sansevieria longiflora,Sansevieria roxburghiana, Sansevieria trifasciataand Sansevieria zeylanica are grown as ornamentalplants (USDA.2008). Sansevieria roxburghianaSchult. & Schult. F. (Agavaceae), called murva inSanskrit and Hindi, Indian bowstring hemp inEnglish, is a herbaceous perennial plant with shortfleshy stem and stout rootstock, occurring in theEastern coastal region of India, also in Sri Lanka,Indonesia and tropical Africa (Eggli, 2002; Prakashet al., 2008).

The present experiments addressed the effect oflead on the antioxidant activity of ascorbateperoxidase (APX), guaiacol peroxidase (GPX), andglutathione-s-hydrogenase (GSH) includingaccumulative potential of lead in Sansevieriaroxburghiana grown in hydroponic culture.

Material and Methods

Plant material

Sansevieria roxburghiana plants were collectedfrom Sri Venkateswara University campus andgrown in green house. The specimen was identifiedby Dr. K.Madhavachetty, taxonomist at SriVenkateswara University, Tirupati, Andhra Pradesh,India. Plants were hydroponically grown for 1 wkwithout Pb and then exposed to nutrient solutionswith 0, 0.5, 0.1, 1.5, 2.0 and 2.5 g/l for 90 daysperiod. The experiment was repeated for threetimes with five replicates. For the experimentalprocedure, 90 d-old plants were grown inHoagland's medium (Hoagland and Arnon, 1950),with 16-h photoperiod (PAR 200 μmol m-2 s-1,temperature 25 ±1 °C, relative air humidity 50 - 60%). The nutritional solution was changed every 7days and aerated every day.

Preparation of enzyme extracts

Plant samples were homogenized for 30 sec in achilled mortar with 50 mM potassium phosphate

buffer (pH 7.0) with 2% Polyclar AT added.Homogenates were centrifuged at 15,000 g for 30min at 4°C and the supernatant was used forenzyme assays.

Determination of Ascorbate Peroxidase Activity(EC 1.11.1.11)

The activity of APX was measured according toNakano and Asada (1987). The reaction mixturecontained 50 mM potassium-phosphate buffer, 0.5mM L-ascorbate (AsA), 0.1 mM H2O2 and theenzyme extract. H2O2-dependent oxidation of AsAwas followed by a decrease in absorbance at 290nm. APX activity was expressed as the absorbancedecrease units mg-1 protein.

Determination of Guaiacol peroxidase Activity(EC 1.11.1.7)

GPX activity was estimated according toHammerschmidt et al. (1982). The reaction mixturecontained 25 mM potassium phosphate buffer(pH7.0), 0.2 mM guaiacol, 0.09 mM H2O2 and theenzyme extract. H2O2-dependent oxidation ofguaiacol was followed by an increase ofabsorbance at 470 nm. Enzyme activity wascalculated as Mm-1 cm-1.

Determination of glutathione-s-hydrogenase

Glutathione-S-Hydrogenase (GSH) content wasdetermined by the recycling method of Anderson(1985). Fresh plant material (0.5 g) washomogenized in 3.0 ml of 5% (w/v) sulfosalicylicacid under cold conditions and was centrifuged at10,000 rpm for 10 min. Half ml aliquot was taken ina microfuge tube, to which 0.5 ml reaction buffer[0.1 M phosphate buffer (pH 7.0), 3 mMethylenediaminetetraacetic acid (Na2EDTA)] and50µl of 50 dithio-bis-(2-nitrobenzoic acid) (0.15%DTNB) were added. After 5 min, absorbance fordetermination of GSH was read at 412 nm usingUV–Vis spectrophotometer (Shimazdu, Japan). Thelevel of GSH was expressed in nmol g-1freshweight.

Lipid Peroxidation

The lipid peroxidation products in samples areexpressed as 2-thiobarbituric acid-reactivemetabolites (TBA-rm) (mainly MDA). MDA wasdetermined in crude extracts with thiobarbituric acid(Peever and Higgins 1989). The absorbance of the



supernatant at 532 nm was measured andcorrected for unspecific turbidity by substracting theabsorbance at 600 nm. The blank was 0.5% TBA in20% TCA. The lipidperoxidation levels wereexpressed as mol g-1 FW.

All enzymes were assayed spectrophotometrically(Shimazdu UV-VIS, Japan) by tracing the change inabsorbance at 27 °C.

Plant digestion

The roots, stems and leaves were digested in aHNO3-HClO4 (3:1,v:v) mixture and Pbconcentrations were determined by atomicabsorption spectrophotometry (AAnalyst 200,Perkin-Elmer, UK).


Statistics were analyzed with SPSS version 11.0,and ANOVA was performed, with significance at p <0.05. All values were mean of five independentexperiments.

Results

Pb accumulation

Sansevieria roxburghiana plants grown in thepresence of 0.5, 1.0, 1.5, 2.0 and 2.5 g/l of leadnitrate showed accumulation of 142.6 mg/kg DW,322.0 mg/kg DW, 323.3 mg/kg DW, 385.3 mg/kgDW and 536.6 mg/kg DW. Pb accumulationincreased according to dose dependent manner.The enhancement of metal accumulation in similartrend observed in Chromolaena odorata andLespedeza davidii exposed to lead according toTanhan et al. (2011) and Zheng et al. (2011).

Effect of Pb on ascorbateperoxidase activity

Ascorbateperoxidase (APX) was involved in thedestruction of H2O2. Thus, a rapid increase in APXactivity was observed on 30 days (6.06±1.25 unitsmg-1 protein), 60 days (6.58±0.83 units mg-1 protein)and 90 days (6.738±1.14 units mg-1 protein)compared to controls (3.48±0.074 units mg-1protein)(Fig. 1). Similar increase in the activity of anti-oxidative enzyme APX in P. vulgaris was observed(Chaturvedi et al. 2009).

Effect of Pb on guiacolperoxidase activity

In the present experiment, the activity of guaiacolperoxidase (GPX) increased in S.roxburghiana

plants exposed to Pb. The GPX activities rangedhigher for a treatment period of 90 days(10.69±0.63 Mm-1 cm-1) compared to 30 days(10.49±0.55 Mm-1 cm-1) and 60 days (10.52±0.55Mm-1 cm-1). Activities of guaiacol peroxidase (GPX)were not sensitive to Pb treatment and significantcorrelation was found between GPX activities andlead accumulation (Fig. 2). It can be seen clearlythat very low GPX activity at initial concentrations(p<0.05). Increased Gpx activity also observed inLemna minor according to Monika et al. (2007).

Effect of Pb on glutathione-s-hydrogenase

GSH, another enzyme of AGC (ascorbate-glutathione cycle), showed a significant increase inits activity after treatment with Pb. Enhancement inglutathione -S-hydrogenase (GSH) in all treatmentswas observed with increasing concentration andtime. The increased GSH activity was highest at 2.0g/l (1.95±0.61 nmol g-1 fresh weight) and 2.5 g/l(2.32±0.75 nmol g-1 fresh weight) Pb concentrations(Fig. 3). Glutathione also increased showing theactive participation in ROS detoxification as

reported earlier in case of Pb, Cu and Cd toxicity intea plant (Rennerberg, 1982; Asada and Takahashi,1987).

Effect of Pb on lipidperoxidation

The effects of lipidperoxidation increased withincreasing lead concentration and treatment time forS.roxburghiana species. The concentrations of lipidperoxides on days 30 (37.56±0.46 mol g-1 FW), 60(51.012±0.52 mol g-1 FW) and 90 days (65.61±0.57mol g-1 FW) compared to controls (26.83±0.75 molg-1 FW) were observed. There was a positivecorrelation between Pb-induced stress and leadaccumulation (Fig. 4). Similar Pb exposure inducedgeneration of reactive oxygen species (ROS) andincreased the level of lipid peroxidation,accompanied by upregulation of antioxidativeenzymes in Zea mays (Gupta et al. 2009) observed.Pb supplied at a toxic level caused a burst ofreactive oxygen species (ROS) in root cells ofSedum alfredii (Huang et al. 2012).

Discussion

In the present experiment, increase in APX followedby GPX clearly indicated that antioxidant enzymessuch as APX and GPX can scavenge free radicalsin synchrony. GSH and GPX are known to beresponsive to biotic and abiotic stresses, but they



Figure 1. Effect of Pb on ascorbateperoxidase activity

Figure 2. Effect of Pb on guiacolperoxidase activity

Figure 3. Effect of Pb on glutathione-s-hydrogenase



Figure 4. Effect of Pb on lipidperoxidation

are well characterized with respect to theirantioxidative roles in plants. This provided evidenceof activation of enzymatic defense mechanisms,such as APX and GPX in the S.roxburghiana,limited the production of free radicals and H2O2. Pbtreatment caused a increase in both GSH activityand GPX activity in S.roxburghiana plants. Theincrease in the activities of these enzymes mightexplain the inhibition of the production of MDA.Since ascorbate is the primary antioxidant and H2O2

is the major stable oxidant, the ratio of these redoxcomponents is indicative of the redox balance withinthe tissue. As Kingston-Smith et al., (1997)described the relative amounts of ascorbate andH2O2 may be used in determining the effectivenessof AGC. GSH content enhanced upon exposure toPb stress. Elevated GSH concentration is correlatedwith the ability of plants to withstand a metal-induced oxidative stress (Freeman et al., 2004).Mohan et al., (1997) suggested that the increase ofperoxidase activity may be an effect of acceleratedsenescence, connected with enhanced formation ofhydrogen peroxide (H2O2) or secondary metabolitessuch as phenolic compounds.

We can add that this possibility does not apply toascorbate peroxidase, an enzyme of the Halliwell-Asada pathway, but only to GPX activity thus wecan say increased activity of APX and GPX showthat they are competed to remove H2O2. In ourmodel conditions the activity of guaiacol peroxidase(GPX) increased in S.roxburghiana plants exposedto Pb, but there was no effect on Glutathione-S-hydrogenase (GSH). There is some evidence thatmodification of membrane structure could activate

production of oxygen free radicals and induce lipidperoxidation (Gupta et al., 1999; Elstneret et al.,1988). Lipid peroxidation mediated by activatedoxygen species should be accompanied bychanges in activities of antioxidative enzymesinvolved in oxygen metabolism. Enhancement oflipid peroxidation of membranes through oxygenfree radicals by lead is also supported by elevatedenzyme activity. One of the most damaging effectsof these active oxygen free radicals and theirproducts in cells is the peroxidation of membranelipids. Here Lead may have shown lowphytoavailability and restricted transport within theplants according to Kabata-Pendias and Sharmaand Dubey (1999, 2005).

The measured ratios of ascorbate to H2O2 arealways higher in the treated plants in comparisonwith the control, which indicated that the cycle isrobust and it is not disordered by Pb-stress. Thehighest dose (2.5g/l) was slightly toxic toS.roxburghiana: the plants were chlorotic and freshweight decreased threefold (results not included);their accumulation capacity probably wasexhausted, but according to the Kabata-Pendias etal. (1999) reports on the positive effect of lead (II)nitrate (V) on plant growth, although data showing aphysiological justification for this phenomenon arelacking. In our study, lead at concentrations lower at0.5 g/l did not harm the growth of S.roxburghiana,and even stimulated it slightly. Here the enzymesGPX, GSH, and APX are involved in thedetoxification of O2•–, and H2O2 respectively bypreventing the formation of •OH radicals.



The results suggest that the Pb-induced increase inantioxidative enzymes in the S.roxburghiana plantsmay represent a secondary defensive mechanismagainst oxidative stress that is not as direct as theprimary defensive responses such asphytochelatins and vacuolar compartmentalization(Sanita de Toppi and Gabbrielli 1999). Theinduction of APX, GPX, and GSH provided effectivedefense against metal toxicity.

Conclusion

This study established a link between leadaccumulation and antioxidant potential in thespecies of Sansevieria roxburghiana in tolerance tometal stress. Lead concentration increased activityof antioxidant enzymes Ascorbate peroxidase(APX), Guiacol peroxidase (GPX), and glutathione -S-hydrogenase (GSH) including MDA content in adose-related manner. Inspite of high leadconcentration a little physiological damage wasobserved like death of root tips. The increase in theactivities of these enzymes explains the inhibition ofthe production of lipidperoxidation. The plantS.roxburghiana taken in the present experiment isan ornamental plant so this plant will be the betteroptions as phytoremediator will not affect foodchain. This evidences suggests that the Sansevieriaroxburghiana was tolerant towards Pb and be usefulfor phytoremediation of lead-contaminated soils.

References

Anderson, M.E., 1985. Determination of glutathioneand glutathione disulfides in biological samples.Methods Enzymol. 113:548–554.

Asada, K. and Takahashi, M. 1987. ‘Production andscavenging of active oxygen in photosynthetictissue’, in Causes of Photooxidative Stress andAmelioration C. H. Foyer, and P. M. Mullneux(eds), of Defense System in Plants,CRC Press,Boca Raton, FL.

Cunningham, S.D., and Berti, W.R., Huang, J.W.,1995. Phytoremediation of contaminated soils.Trends Biotech. 13:393–397.

Choudhury, S., and Panda, S. K., 2005. Toxiceffects, oxidative stress and ultrastructuralchanges in moss Taxithelium nepalense(Schwaegr.) Broth. under chromium and leadphytotoxicity. Water Air Soil Poll. 167: 73–90.

Chaturvedi Ashish, K., Priti Bhardwaj., and Prasad,P. 2009. Effect of Enhanced Lead and Cadmiumin soil on Physiological and Biochemical

attributes of Phaseolus vulgaris L. Nature andscience 7(8).

Elstner, E.F., Wagner, G.A., and Schutz, W., 1988.Activated oxygen in green plants in relation tostress situations. Curr Top Plant BiochemPhysiol. 7:159–187.

Eggli, U.S., 2002. Illustrated Hand Book ofSucculent Plants: Monocotyledons. Berlin,Heidelberg: Springer.

Freeman, J.I., Persans, M.W., Nieman, K., Albrecht,C., Peer, W., Pickering, I.J., and Salt, D.E.,2004. Increased glutathione biosynthesis playsa role in nickel tolerance in Thalpsi nickelhyperaccumulator. Plant Cell. 16:2176–2191.

Gupta, M., Cuypers, A., Vangrosveld, J., Clijsters,H., 1999. Copper affects the enzymes of theascrobate-glutathione cycle and its relatedmetabolites in the roots of Phaseolus vulgaris.Physiol. Plant. 106:262–267.

Gupta, D.K., Huang, H.G., Yang, X.E.,Razafindrabe, B.H.N., Inouhe, M. 2010. Thedetoxification of lead in Sedum alfredii H. is notrelated to phytochelatins but the glutathione. JHazard. Mater. 177:437–444.

Huang, H.G., Gupta, D.K., Tian, S.K., Yang, X.E.,Li, T.X., 2012. Lead tolerance and physiologicaladaptation mechanism in roots of accumulatingand non-accumulating ecotypes of Sedumalfredii. Environ. Sci. Poll. Res. 19:1640–1651.

Hoagland, D.R., and Arnon, D.I., 1950. Thewaterculture method for growing plants withoutsoil. Calif. Agric. Exp. Sta. Circ. 347: 1–32.

Hammerschmidtr, nuclesem, and kucj. 1982.Association of enhanced peroxidase activity withinduced systemic resistance of cucumber toColletotrichum lagenarium. Physiological PlantPathology 20: 73–82.

Halliwell, B., 1987. Oxidative stress. FEBS Lett.216(1): 170–171.

Jiang, W.S., and Liu, D.H., 2000. Effects of Pb2+ onroot growth, cell division, and nucleolus of Zeamays L. B. Environ. Contam. Tox. 65: 786–93.

Kingston-Smith, A.H., Thomas, H., and Foyer, C.H.,1997. Chlorophyll a fluorescence, enzyme andantioxidant analysis provide evidence for theoperation of alternative electron sink during leafsenescence in a stay-green mutant of Festucapratensis. - Plant Cell Environ. 20: 1323–1337.

Kabata-pendiasa, and Pendiash. 1999.Biogeochemistry of trace elements. PWN,Warsaw. (In Polish).

Liu, D., Zou, J., Meng, Q., Zou, J., Jiang, W., 2009.Uptake and accumulation and oxidative stress in



garlic (Allium sativumL.) under leadphytotoxicity. Ecotoxicology 18: 134–43.

Mohan, B.S., and Hosettib, B. 1997. Potentialphytotoxicity of lead and cadmium to Lemnaminor grown in sewage stabilization ponds.Environmental Pollution. 98: 233–238.

Nakano, Y., and Asada, K., 1987. Purification ofascorbate peroxidase in spinach chloroplast, itsinactivation in ascorbate – depleted medium andreactivation by monodehydroascorbate radical.Plant and Cell Physiology. 28: 131–140.

Peever, L., and Higgins, V., 1989. Electrolyteleakage, lipoxygenase, and lipid peroxidationinduced in tomato leaf tissue by specific andnon-specific elicitors from Cladosporium fulvum.Plant Physiol. 90:867–875.

Patra, M., Bhowmik, N., Bandopadhyay, B., andSharma, A., 2004. Comparison of mercury, leadand arsenic with respect to genotoxic effects onplant systems and the development of genetictolerance. Environ. Exp. Bot. 52: 199–223.

Pandey, S, Gupta, K, and Mukherjee, A.K., 2007.Impact of cadmium and lead on Catharanthusroseus: a phytoremediation study. J. Environ.Biol. 28: 655–62.

Prakash, J.W., Raja, R.D.A, Anderson, N.A.,Williams, C., Regini, G.S., Bensar, K., Jajeev,R., Kirula, S., Jeeva S, Das, S.C.M., 2008.Ethnomedicinal plants used by Kani tribes ofAgastiyarmalai biosphere reserve, SouthernWestern Ghats. Indian J Trad Knowledge 7:410–413.

Rennenberg, H. 1982, Glutathione metabolism andpossible role in higher plants. Phytochemistry.28: 2771.

Sanita di Toppi, L., and Gabbrielli,R. , 1999.Response to cadmium in higher plants. -Environ. exp. Bot. 41: 105–130.

Sharma, P., and Dubey, R.S., 2005. Lead toxicity inplants. Braz. J. Plant Physiol. 17: 35–52.

Tanhan, P., Pokethitiyook P., Kruatrachue, M.,Chaiyarat, R., Upatham, S. 2011. Effects of soilamendments and EDTA on lead uptake byChromolaena odorata: Greenhouse and fieldtrial experiments.Int. J. Phytorem. 13:897–911.

USDA, 2008. USDA, ARS, National GeneticResources Program. Germplasm ResourcesInformation Network-(GRIN) [Online Database].National Germplasm Resources Laboratory,Beltsville, Maryland.

Verma, S., and Dubey, R.S., 2003. Lead toxicityinduces lipid peroxidation and alters theactivities of antioxidant enzymes in growing riceplants. Plant Sci. 164: 645–655.

Watanabe, M.A., 1997. Phytoremediation on thebrink of commercialization. Environ. Sci.Technol. 31: 182–86.

Wierzbicka, M., 1998. Lead in the apoplast of Alliumcepa L. root tips – ultrastructural studies. PlantSci. 133: 105–9.

Xiao, W., Hao, H., Liu, X.Q., Liang, C., Chao, L.,Su, M.Y., and Hong, F.H., 2008. Oxidativestress induced by lead in chloroplast of spinach.Biol. Trace. Elem. Res. 126: 257–68.

Zheng, L.J., Liu, X.M., Lutz-Meindl, U., Peer, T.2011. Effects of lead and EDTA-assisted leadon biomass, lead uptake and mineral nutrientsin Lespedeza chinensis and Lespedeza davidii.Water. Air. Soil. Poll. 220:57–68.

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