Bioaccumulation of toxic metals (Cr, Cd, Pb and Cu) by seeds of Euryale ferox Salisb. (Makhana)

Bioaccumulation of toxic metals (Cr, Cd, Pb and Cu) by seeds of Euryale ferox Salisb. (Makhana)

Chemosphere 46 (2002) 267±272 www.elsevier.com/locate/chemosphere Bioaccumulation of toxic metals (Cr, Cd, Pb and Cu) by seeds of Euryale ferox Sali...

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Chemosphere 46 (2002) 267±272

www.elsevier.com/locate/chemosphere

Bioaccumulation of toxic metals (Cr, Cd, Pb and Cu) by seeds of Euryale ferox Salisb. (Makhana) q U.N. Rai a

a,*

, R.D. Tripathi a, P. Vajpayee a, Vidyanath Jha b, M.B. Ali

a

Ecotoxicology and Bioremediation, National Botanical Research Institute, Lucknow 226 001, India b Department of Biotechnology, L.N. Mithila University, Darbhanga 846 004, India Received 6 September 2000; accepted 17 March 2001

Abstract The level of toxic metals Cr, Cd, Pb and Cu was determined in seeds, water and sediments collected from nine closed waterbodies of Darbhanga, north Bihar, used for cultivation of the edible aquatic macrophyte Euryale ferox Salisb. during harvesting season of the crop for two successive years (1996 and 1997). Seeds bioconcentrated appreciable amount of these toxic metals in the order Pb > Cr > Cu > Cd. The increased load of metal pollution due to domestic and municipal discharges threatened the habitats of the plant. The toxic metal contents in seeds were found positively correlated with the ambient concentration of metals in water and sediments. The importance of these ®ndings has been discussed for national water resource economy of the country and human health perspectives. Ó 2002 Elsevier Science Ltd. All rights reserved. Keywords: Euryale ferox; Edible aquatic macrophyte; Seeds; Toxic metals

1. Introduction Rapid urbanization, industrialization, fertilizer and pesticide use has resulted in toxic metal pollution of land and water resources. The increasing load of toxic metals has caused imbalance in aquatic ecosystems and the biota growing under such habitats accumulate high amounts of toxic metals (Cu, Zn, Cd, Cr, Fe, etc.) which, in turn, are being assimilated and transferred within food chains by the process of biomagni®cation (Pergent and Pergent-Martini, 1999). Further, high concentration of these metals in food chains results in various health hazards to animal and human beings (Revai et al., 1990).

q

NBRI Research Publication No. (503) NS. Corresponding author. Tel.: +91-522-205831; fax: +91-522205839. E-mail address: rai_un@redi€mail.com (U.N. Rai). *

Bioaccumulation of essential and non-essential metals by aquatic macrophytes is well documented in the literature (Vesk and Allaway, 1997; Khan et al., 2000). This property of bioaccumulation was found useful in biomonitoring and ameliorating the water bodies (Wang and Williams, 1988; Dunbabin and Bowmer, 1992; Whitton and Kelley, 1995; Vajpayee et al., 1995). However, a number of aquatic macrophytes are edible and possess medicinal properties, which could not be utilized for metal removal studies. Euryale ferox Salisbury (Makhana), of family Euryalaceae is a rooted macrophyte with large spiny ¯oating leaves. The genus is now represented only by this single species. This species is cultivated by marginal farmers as a cash crop in non-calcareus zone of the Kosi±Kamala belt of north Bihar and lower Assam in India. Bihar contributes 75% of the total `Makhana' production from India (Thakur and Jha, 1999) followed by Bengal (10%), Assam (7%), Uttar Pradesh (5%) and Madhya Pradesh (3%). In Bihar, Madhubani district has maximum share of Makhana production (40%) followed by Darbhanga

0045-6535/02/$ - see front matter Ó 2002 Elsevier Science Ltd. All rights reserved. PII: S 0 0 4 5 - 6 5 3 5 ( 0 1 ) 0 0 0 8 7 - X

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(25%), Saharsa (20%), Katihar (7%), Purnea (5%) and Champaran (3%). The fried pops of seeds are held sacred and have been reported to have medicinal and nutritional value (Jha et al., 1991). Habitats of E. ferox are threatened due to increasing load of metal pollution (from municipal and domestic wastes, etc.). Therefore, the present study was conducted to evaluate the metal pollution of nine ponds situated in Darbhanga (north Bihar) used for exclusive cultivation of E. ferox and bioaccumulation of toxic metals by the seeds of species. The objective was to address the issue of transfer of metals into the human food chain from dietary components growing in contaminated habitat.

2. Materials and methods Nine large ponds under exclusive E. ferox (Makhana) cultivation (Kangawa Pond ± I; Chuna Bhatti Pond ± II; Sara Mohanpur Pond ± III; Diwana Takia Pond ± IV; Sakir Rahman Pond ± V; Namaki Pond ±VI; Adarsh Nagar Pond ±VII; Alalpatti Pond ± VIII; Bela Pond ± IX) situated in district Darbhanga (north Bihar) were randomly selected for the present study (Fig. 1). The

Fig. 1. Various sampling ponds situated in district Darbhanga, north Bihar used for exclusively E. ferox (Makhana) cultivation.

sites were located between 19°100 N latitude and longitude 117°240 E. Water, sediment and seed samples from the nine aforesaid ponds were collected during October 1996 and 1997, when the crop was being harvested by farmers. The collection of water (1 l) from each pond was done in acid-washed plastic containers. Samples were ®ltered, acidi®ed and brought to the laboratory for metal analysis in accordance with standard methods (APHA, 1989). Sediments were collected in polythene bags, while seeds were blotted dry and placed in polythene bags for transportation. To measure the toxic metal content in water, sediments and seeds, 50 ml water, 1 g each of dried samples of sediments and seeds from each pond in triplicate were digested in HNO3 :HClO4 (3:1, v/v) mixture at 80°C. Metal concentrations (Cr, Cd, Pb and Cu) in these samples were recorded on atomic absorption spectrophotometer (2380 Perkin Elmer, USA). The standard reference materials (E-Merck, Germany) of Cr, Cu, Pb, and Cd were used to provide calibration and quality assurance for each analytical batch. The eciency of digestion of plant, soil, and

Fig. 2. The concentrations of chromium (a), cadmium (b), lead (c) and copper (d) in water samples of ponds used exclusively for E. ferox cultivation. Maximum permissible limits provided by WHO for Cr ˆ 0.05 lg cm 3 , Cd ˆ 0.005 lg cm 3 , Pb ˆ 0.05 lg cm 3 , Cu ˆ 0.05 lg cm 3 ; values are mean …n ˆ 3†  S:E.

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sediment samples was determined by adding standard reference material of each metal. After addition of standard reference material of each metal, samples were digested and metals were estimated as above. Mean recoveries of Cr, Cu, Pb and Cd were 96  6%, 97  5%, 96  4% and 97  5%, respectively. The detection limits of chromium, copper, lead and cadmium were 0.002, 0.001, 0.005 and 0.0002 lg ml 1 , respectively. Replicate …n ˆ 3† analyses were conducted to assess the precision of the analytical techniques. Triplicate analyses for each metal varied by no more than 5%. A two-way ANOVA involving nine sites and two years of analysis in randomized complete block design was done to test the variability and validity of the data. A simple regression and correlation analysis was done to correlate the toxic metal contents in pond and their accumulation in seeds of E. ferox (Gomez and Gomez, 1984). 3. Results and discussion All the water samples from the ponds surveyed for heavy metal pollution were found contaminated with toxic metals (Fig. 2). However, their quantity varied

269

with pond (ANOVA, P < 0:05; Table 1). It is pertinent to mention that out of the nine ponds (I±IX) surveyed for water quality, pond-IV was found to be highly contaminated with toxic metals. In the year 1996, maximum contamination of chromium was found in water samples of pond II (0.135 lg cm 3 ) while maximum level of Cd (0. 052 lg cm 3 ), Pb (0.912 lg cm 3 ) and Cu (1.590 lg cm 3 ) were recorded from water samples of ponds IV, IX and III, respectively. It seems that the pollution load was escalating, as in 1997 the respective contamination in these ponds subsequently increased (ANOVA, P < 0:05; Table 1). A comparison of data from permissible limits provided by WHO (1995) showed that the concentration of chromium in water of ponds II, III, IV and V was above permissible limits, however, it was under permissible limit (0.05 lg cm 3 † in the remaining ponds. The concentrations of Cd, Pb and Cu in all the ponds surveyed were alarming as the concentrations of these metals were always above the recommended permissible limits. It has been observed that sediments of ponds act as a sink for toxic metals as concentrations of metals were higher in sediment samples than water (Fig. 3). A comparative study revealed that concentration of lead was high in almost all the ponds than other metals

Table 1 Two-way analysis of variance of data on metal contamination of water, sediment and seed samples of E. ferox Source of variance

df

F value Cr

Water Replications Treatment Year (A) Pond (B) AB Error Total

12 17 1 8 8 34 53

8.44 9135.33 507.89 1057.72 26.71

Sediment Replications Treatment Year (A) Pond (B) AB Error Total

2 17 1 8 8 34 53

1.28 500.33 323.17 1016.36 6.44

Seed Replications Treatment Year (A) Pond (B) AB Error Total

2 17 1 8 8 34 53

50.90 1871.39 1191.76 3804.67 23.06

a

Insigni®cant at 5% level.

Cd 8.82 329.04 528.86 621.54 115.60

3.59 40.00 15.34 82.44 0.76a

28.04 1334.55 4767.64 1966.79 284.43

Pb

Cu

6.94 197.95 163.28 392.20 7.71

64.93 1226.88 109.24 2593.08 0.39a

87.28 1099.39 1740.64 2092.38 26.24

6.18 182.29 27.11 7.50 0.50a

113.18 1176.73 536.82 2419.68 13.77

57.59 20 671.97 8095.36 1463.34 139.39

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Fig. 3. The concentrations of chromium (a), cadmium (b), lead (c) and copper (d) in sediment samples of ponds exclusively used for E. ferox cultivation; values are mean …n ˆ 3†  S:E.

Fig. 4. Chromium (a), cadmium (b), lead (c) and copper (d) accumulation in seeds of E. ferox collected from di€erent ponds of district Darbhanga, north Bihar; values are mean …n ˆ 3†  S:E.

(Cu, Cd, Cr) estimated during 1996 and 1997. However, the concentrations of all the metals in sediments were higher in the year 1997 than 1996 (ANOVA, P < 0:05; Table 1). Seeds of E. ferox were found enriched with chromium, cadmium, lead and copper (Fig. 4). Increased bioaccumulation of these toxic metals in seeds of E. ferox was observed in the year 1997. Chromium concentration in seeds collected during 1996 from pond II, was maximum (550.17 lg g 1 DW) which increased to 650.20 lg g 1 DW in the year 1997 (Fig. 4). Likewise, seeds have also shown a hike in the concentration of other metals during the year 1997 (ANOVA, P < 0:05; Table 1). The concentrations of toxic metals in seeds of E. ferox were found positively correlated (correlation coecient …r† signi®cant at 5% level; df ˆ 7) with their concentration in water and sediment samples of pond during the years 1996 and 1997 (Table 2). Lead is considered to be potentially toxic to the plants and other life forms (Singh et al., 1997). The higher concentration of Pb in water, sediments and seed samples of pond was probably due to excessive use of phosphate fertilizers and pesticides, which are sources of lead contamination. High concentration of lead (1.4 lg cm 3 ) in pond system was also reported by Chandra

et al. (1993). Similarly, toxic levels of chromium in pond water and soil used for Makhana cultivation might also be due to anthropogenic activities which may cause various disorders in human beings (Satyakala and Jamil, 1992). Copper is an essential metal but at high concentration it is toxic to plants and may a€ect the productivity of the crop (Dutta et al., 1986). Comparatively lower concentration of cadmium was recorded in water, sediments and seed samples tested during present investigations. But it has been reported that cadmium exposure at lower concentrations changes the reticulocytes and thrombocyte counts in human beings (Nath, 1986). Cadmium toxicity to humans caused by Cd-contaminated rice was reported from Japan (Kanneta et al., 1983). However, in Indians, daily intake of cadmium ranges between 7.8±16.5 mg/person without any chronic illness (Revai et al., 1990). Although the Cd content in the seeds was signi®cantly low, it could be considered alarming due to the highly toxic nature of the metal. The toxic metal concentrations in seeds of E. ferox were positively correlated with their concentration in water or sediments of pond. Plant metal levels were also found highly correlated with metal levels in the sediments during a study conducted by Jackson and

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Table 2 Simple linear regression analysis and simple correlation coecient (r) of toxic metal accumulation in seeds of E. ferox and their level in water or sediments of ponds used for cultivation of crop Year

Metal

1996

Cr Cd Pb Cu

1997

Cr Cd Pb Cu

a

Signi®cant at P < 5:00 at df …n

Linear regression equation

Correlation coecienta …r†

Water Sediments Water Sediments Water Sediments Water sediments

Y Y Y Y Y Y Y Y

ˆ 79:38 ‡ 3573:56X ˆ 83:32 ‡ 5:16X ˆ 1:46 ‡ 513:16X ˆ 1:75 ‡ 116:27X ˆ 0:41 ‡ 1138:39X ˆ 134:49 ‡ 4:37X ˆ 23:61 ‡ 83:53X ˆ 8:84 ‡ 1:96X

0.960 0.958 0.821 0.989 0.915 0.865 0.949 0.816

Water Sediments Water Sediments Water Sediments Water Sediments

Y Y Y Y Y Y Y Y

ˆ 100:61 ‡ 3013:08X ˆ 83:32 ‡ 5:16X ˆ 4:81 ‡ 1146:92X ˆ 1:19 ‡ 222:94X ˆ 316:64 ‡ 981:92X ˆ 43:96 ‡ 4:57X ˆ 60:33 ‡ 142:15X ˆ 12:71 ‡ 2:41X

0.960 0.958 0.915 0.994 0.913 0.830 0.899 0.884

2† ˆ 7.

Kal€ (1993). Metals in sediment are known to vary at variety of scales (Morrisey et al., 1994; Vesk and Allaway, 1997) and similar variation might be expected to apply to plant metal concentration. Our results support the view of earlier workers. The factors that explain variation of plant metal concentrations, and are likely to play role in a€ecting these concentrations, include metal speciation of the sediments, plant growth form, pH, redox potential, sediments, organic matter, etc. (Campbell et al., 1985; Jackson and Kal€, 1993; Jackson et al., 1993). Occurrence of toxic metals in pond, ditch and river water a€ect the lives of local people which depend upon these water sources for their daily requirements (Rai et al., 1996). Consumption of such aquatic food stu€ enriched with toxic metals may cause serious health hazards through food-chain biomagni®cation. During the present investigation, seeds of E. ferox were found highly contaminated with Pb followed by Cr, Cu and Cd. High accumulation values were also shown by some other aquatic vegetables such as Ipomoea aquatica, Marsilea minuta, Nelumbo nucifera and Trapa natans (Gommes and Muntau, 1981; Rai et al., 1996; Rai and Sinha, 2001). It may be concluded that cultivation of E. ferox in polluted water bodies may lead to serious health hazards by the bioaccumulation of toxic metals in its edible parts. Thus the results are suggestive of the limitation in the exploitation of aquatic resources to meet the ever increasing demand of food, however, a more detailed investigation is required to assess the level of metal contamination and also its impact on nutraceutical potential of the plant.

Acknowledgements We thank the Director, NBRI, Lucknow, for providing the required research facilities and to the CSIR, New Delhi for the award of visiting associateship to V.N. Jha. References APHA, 1989. Standard Methods for Examination of Water and Wastewaters, fourteenth ed. Washington, DC, USA. Campbell, P.G.C., Tessier, A., Bisson, M., Bougie, R., 1985. Accumulation of copper and zinc in the yellow water lily Nuphar variegatum: relationships to metal partitioning in the adjacent lake sediment. Can. J. Fish. Aquat. Sci. 42, 23±32. Chandra, P., Tripathi, R.D., Rai, U.N., Sinha, S., Garg, P., 1993. Biomonitoring and amelioration of non point source pollution in some aquatic bodies. Water Sci. Tech. 28, 323± 326. Dunbabin, J.S., Bowmer, K.H., 1992. Potential use of constructed wetlands for treatment of industrial waste waters containing metals. Sci. Total Environ. 111, 151±168. Dutta, R.N., Jha, U.N., Jha, S.N., 1986. Relationship of biomass yield of Makhana (E. ferox) with soil properties and water quality. Plant Soil 95, 345±350. Gomez, K.A., Gomez, A.A., 1984. Statistical Procedures for Agricultural Research. Wiley, New York. Gommes, R., Muntau, H., 1981. Determination in situ des vitesses d'absorption de metaux lourds par les macrophytes du Lago Maggiore. Mem. Ist. Ital. Idrobiol. 38, 347±377. Jackson, L.J., Kal€, J., 1993. Patterns in metal content of submerged aquatic macrophytes: the role of plant growth form. Freshwater Biol. 29, 351±359. Jackson, L.J., Kal€, J., Rasmussen, J.B., 1993. Sediment, pH and redox potential a€ect the bioavailibility of Al, Cu,

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Singh, R.P., Tripathi, R.D., Sinha, S.K., Maheshwari, R.K., Srivastava, H.S., 1997. Response of higher plants to lead contaminated environment. Chemosphere 34, 2467±2493. Thakur, Y.P., Jha, V.N., 1999. Trials on ®sh±makhana yield enhancement in ponds of Darbhanga division. Final Technical Report submitted to the Department of Science and Technology, Govt. of India, New Delhi, p. 82. Vajpayee, P., Rai, U.N., Sinha, S., Tripathi, R.D., Chandra, P., 1995. Bioremediation of tannery e‚uent by aquatic macrophytes. Bull. Environ. Contam. Toxicol. 55, 546±553. Vesk, P.A., Allaway, W.G., 1997. Spatial variation of copper and lead concentrations of water hyacinth plants in a wetland receiving urban run-o€. Aquat. Bot. 59, 33±44. Wang, W., Williams, J., 1988. Screening and biomonitoring of industrial e‚uents using phytotoxicity tests. Environ. Toxicol. Chem. 7, 645±652. Whitton, B.A., Kelley, M.G., 1995. Use of algae and other plants for monitoring rivers. Aust. J. Ecol. 20, 45±56. WHO, 1995. Guidelines for Drinking Water Quality, vol. 3. Geneva. Dr. U.N. Rai is a senior scientist working in the Ecotoxicology and Bioremediation group of Environmental Science Division at the National Botanical Research Institute, Lucknow. He has worked on various aspects of applied phycology and environmental remediation using plants and has published about 65 papers, mostly in the journals having high science citation index. In 1995, he worked at the University of Liverpool, Liverpool (UK) on isolation and characterization of metal-binding peptides in plants. Currently, he is engaged in di€erent research activities on metal toxicity and tolerance, pollution monitoring and biological water puri®cation, environmental impact assessment, to use phytoremediation system for treatment of industrial wastes, and planning remedial measures for di€erent industrial units.