Study of the Effect of Lead (Heavy Metal)

A major environmental concern due to dispersal of industrial and urban wastes generated by human activities is the contamination of soil. Controlled and uncontrolled disposal of waste, accidental and process spillage, mining and smelting of metalliferous ores, sewage sludge application to agricultural soils are responsible for the migration of contaminants into non-contaminated sites as dust or leachate and contribute towards contamination of our ecosystem. The present study was undertaken to study the effect of heavy metals on the growth of Brasicca juncea and thereby its utilization for phytoremediation. The minimum amount of lead (0 mg/gm) was found in plant treated in 0 % concentration of lead acetate and maximum amount of lead (0. 0011 mg/gm) was found in plant treated in (0. 6%) concentration of lead acetate. The percentage germination for control, 0. 2, 0. 4, 0. 6 percent lead acetate treated seedling was 89%, 88%, 86. 66% and 83% respectively. The length of shoot was found much less in plant treated in 0. 6% concentration of lead acetate (10 cm), as compared to the length of shoot in plant treated with 0% concentration of lead acetate (8 cm). Length of roots was found maximum in 0. 6% concentration of lead acetate (2 cm), as compared to the length of shoot in plant treated with 0% concentration of lead acetate (0. 5 cm). It was found that Brasica juncea tolerated highest concentration of heavy metals and can accumulate lead. Key Words: leachate, heavy metals, lead, phytoremediation, Brasicca juncea. Introduction Heavy metal contamination of soil, water and air has caused serious environmental hazard in the biosphere due to rapid industrialization and urbanization. Lead is probably one of the most frequently encountered heavy metals in polluted environment. The primary sources of this metal include mining and smelting of metalliferous ores, burning of leaded gasoline, disposal of municipal sewage and industrial wastes enriched in lead as well as using of lead-based paint (Kabata-Pendias and Pendias 1984; Seaward and Richard-son 1990). Relatively high levels of lead concentrations were recorded as high as 7,000 Â µg/ gm in roadside soil (Kabata-Pendias and Pendias 1984) and 13,380 Â µg/gm in mining district soil (Wick land 1990). In recent years it has been reported that some plant species known as hyper accumulator derived from heavy metal-contaminated areas have the ability to accumulate unusually high content of heavy metals without dramatically being impacted in their growth and development. (Reeves, Brooks and Malaises 1985; Baker and Brooks 1989). This raises the suggestion that these hyper accumulators may provide the basis for phytoremediation of heavy metal-contaminated sites (Baker et al. 1991). Phytoremediation potential of a few such species for heavy metal-contaminated soil and water has recently been detected (Brown et al. 994; Kumar et al. 1995; Dushenkov et al. 1995; Huang et al. 1997; Blaylock et al. 1997). Lead as a Heavy metal: Lead (Pb) exists naturally in many forms throughout the world & has a soil retention time of 150-5000 yrs. Lead is among those heavy metals which have no known biological function. Never the less, numerous investigations show that plants can accumulate lead via root and shoot, and that the lead concentrations in plant tissues are significantly related to the lead levels in environment (Kabata-Pendias and Pendias 1984; Nwosu et al. 995; Sawidis et al 1995; Xiong 1998). Excessive lead accumulated in plant tissue can be toxic to most plants, leading to decrease in seed germination, root elongation and biomass, inhibition of chlorophyll biosynthesis, as well as cell disturbance and chromosome lesion (Balsberg Pahlsson 1989; Kumar et al. 1991; Fargasova 1994; Xiong 1997c). In lead and other heavy metal-contaminated sites, the vegetation structure and biodiversity are usually reduced barren patches of soil occurring, and trees are sparse or absent (Wickland 1990). Material and Method Brasicca juncea seeds obtained from nursery were treated with lead acetate solution of concentrations ranging from 0. 0%, 0. 2%, 0. 4% & 0. 6% for control the seed were soaked in distilled water. After 24 hrs the seeds were transferred on wet filter paper (wet by the solutions of respective conc. ) in plastic trays. Seedlings were harvested after twenty days, the germination rate and the length of root and shoot was recorded. The amount of lead accumulated in the seedlings was also calculated. Stock solution of 0. 2%, o. 4% and 0. % was prepared in the following way: 20 mg of lead acetate in 1000 ml distilled water in volumetric flask gave 0. 2% of lead acetate solution. In the same way 40 mg & 60 mg lead acetate salt were dissolved to prepare 0. 4% & 0. 6%. stock solutions respectively. The DNA in control and the treated seedlings was estimated by Diphenylamine reaction. Optical density was read at 595 nm on a photoelectric colorimeter. Standard graph of O. D. on Y-axis and concentration of standard DNA on X-axis was plotted. DNA from sample was estimated with the help of standard graph. The Folin Lowry’s method of protein assay was used for protein estimation in control and treated seedlings. Optical density was read at 625 nm on photoelectric colorimeter. For the estimation of lead from the treated seedlings, 10 gm of the plant material was crushed in mortar and pestle in 40 ml distilled water and the solution was filtered. Another series was made by taking 10 ml sample solution from filtrate of all concentrations from 0. 2%, 0. 4%, 0. 6% and control. 25 ml of freshly prepared Ammonium solution and 0. 5 ml of Sodium sulfide solution was added to all the above test tubes. Similarly for standard graph, varying concentration (0. 25 – 1. 5ml) of working lead solution was prepared. To this 25 ml of freshly prepared Ammonium solution to make it alkaline was added; also 0. 5 ml of Sodium Sulfide was added. The volume of all solutions was equalized with distilled water. A blank was also prepared in the same manner without adding lead solution. Optical density was read at 430 nm on photoelectric colorimeter. The concentration of lead was calculated in terms of mg/litre from standard graph. Result & Discussion The minimum amount of DNA (4. 50 mg/gm) was found in seedlings treated in (0%) concentration of lead acetate after 20 days of treatment, and maximum minimum amount of DNA (4. 675 mg/gm) found in plant treated in (0. 6%) concentration of lead acetate after 20 days of treatment. The graph shows that the amount of DNA increases as the concentration of lead acetate goes on increasing. The minimum amount of Protein (86 mg/gm) was found in plant treated in (0%) concentration of lead acetate after 20 days of treatment, and maximum amount Protein (93. 5 mg/gm) found in plant treated in (0. 6%) concentration of lead acetate after 20 days of treatment 89. mg and 91. 5mg of protein was estimated in the seedlings treated with 0. 4% and 0. 6% lead acetate. From the graph it can be concluded that the amount of protein increased with the increase in the concentration of lead. The graphical representation was done for calculating the concentration of lead in the treated seedlings. The optical density was read at 430 nm for control, 0. 2%, 0. 4% and 0. 6% lead acetate treated seedlings was 0, 0. 013, 0. 016 and 0. 019 respectively. The graph plotted resulted into a straight line, indicating the increased accumulation of lead in the seedlings. The concentration of lead was calculated using graph. In 1gm of control, 0. 2, 0. 4 and 0. 6 percent lead acetate treated seedling sample had 0 mg, 0. 00078 mg, 0. 00094 mg and 0. 0011 mg of lead respectively. The minimum amount of lead (0 mg/gm) was found in plant treated in (0%) concentration of lead acetate and maximum amount of lead (0. 0011 mg/gm) was found in plant treated in (0. 6%) concentration of lead acetate. This suggests that Brasicca juncea could accumulate comparatively high doses lead. The percentage germination for 1 gm each of control, 0. 2, 0. 4, 0. percent lead acetate treated seedling was 89%, 88%, 86. 66% and 83% respectively. The germination with increase in concentration of lead acetate did not show any inhibitory effect on the germination of seeds but a steady decrease in the percentage of germination was observed. Maximum percentage of germination was found in control seeds. Out of 300 seeds soaked in 0. 2%, 0. 4% and 0. 6% solution of lead acetate showed 264, 260 and 249 seeds germinating respectively. When compared to the control the germination percentage of the treated seeds did not differ much. This indicated the capability of Brasicca juncea in tolerating the presence of heavy metals like lead. The shoot length for 1 gm each of control, 0. 2, 0. 4 and 0. 6 percent lead acetate treated seedling was 8 cm, 8. 5 cm, 8. 3cm, and 10 cm respectively; also the root length for 1 gm each of control, 0. 2, 0. 4 and 0. 6 percent lead acetate treated seedling was 0. 5 cm, 0. 8 cm, 1. 0 cm and 2. 0 cm respectively. The length of shoot was found maximum in plant treated in 0. 6% concentration of lead acetate (10 cm), as compared to the length of shoot in plant treated with 0% concentration of lead acetate (8 cm). Length of roots was found maximum in 0. 6% concentration of lead acetate (2 cm), as compared to the length of shoot in plant treated with 0% concentration of lead acetate (0. 5 cm). This indicates that Brasicca juncea was able to utilize the increased amount of lead to its advantage, suggesting the use of Brasicca juncea as a natural alternative for phytoremediation. Conclusion The present study indicates that plant species like Brasicca juncea can tolerate wide range of heavy metals concentration, it utilizes the heavy concentration for its all round growth . It can be used in removal of lead; and hence its use in phytoremediation in future. Phytoremediation is a fast developing field, since last ten years lot of field application were initiated all over the world, it includes Phytoremediation of Organic, Inorganic and Radio nuclides. This sustainable and inexpensive process is fast emerging as a viable alternative to conventional remediation methods and will be most suitable for a developing country like India. Most of the studies have been done in developed countries and knowledge of suitable plants is particularly limited in India. In India commercial application of Phytoremediation of Soil heavy metal or Organic compounds is in its earliest phase. Fast growing plants with high biomass and good metal uptake ability are needed. In most of the contaminated sites hardy, tolerant, weed species exist and phytoremediation through these and other non-edible species can restrict the contaminant from being introduced into the food web. Further studies need to be performed in order to establish the maximum amount of lead that the plants may tolerate, and the ability of Brasicca juncea to germinate and grow in media containing mixtures of heavy metals.

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