EFFECT OF LEAD ON THE FUNCTIONING OF LABORATORY MODEL WASTE STABILIZATION POND

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1 EFFECT OF LEAD ON THE FUNCTIONING OF LABORATORY MODEL WASTE STABILIZATION POND Mangala Veeresh, A.V. Veeresh* and B.B. Hosetti** Department of Zoology, *Department of Botany, S.P.Chowgule College of Arts & Science, Margao , Goa; ** Department of Applied Zoology, Bioscience Block, Kuvempu University, Shankarghatta , Shimoga Dist., Karnataka, India. ABSTRACT: Raw industrial waste water collected from Cosme Mathais Menezes industrial complex, Curti, Ponda, Goa served as the growth medium. Plastic tubs in duplicate were filled with wastewater and treated with different concentration of lead. To each of the experimental pond (laboratory model) seed material collected from the field stabilization pond was added and placed in open sunlight and were operated on batch system. The experiment was run for 15 days. Observations were made by collecting the known aliquot of sample from each pond on 5 th, 10 th and 15 th day of experiment and analyzed for ph, DO, BOD 5, catalase, phosphatase, total phosphate, total solids, total chlorophyll and total algal count. The study revealed that with increasing concentration of lead in the experimental ponds there was a decrease in the ph value, dissolved oxygen and chlorophyll content and also algal numbers on all the days of analysis. Also it was observed that with increasing concentration of lead, the activity of the enzymes catalase and phosphatase reduced and the percentage removal of inorganic phosphate and BOD 5 decreased. The total solids in the experimental ponds increased with increasing concentration of heavy metals. 1. INTRODUCTION: Water bodies are polluted due to the release of liquid pollutants like that of domestic sewage containing detergents and other wastes, agricultural wastes which contain various chemicals and industrial effluents rich in heavy metals. These pollutants have harmful effects on the aquatic flora and fauna. The functioning and life of the aquatic organisms is increasingly being threatened day by day due to the release of various extraneous pollutants containing lethal metals namely Lead, Copper, Zinc, Mercury, Cadmium and Chromium through industrial discharges. Due to various human activities, the level of pollutants and the types of pollutants released into the environment are increasing and ever changing continuously which are having deleterious effects on the organisms and especially on the self purification of the water bodies (Mason, 1989; Gautam, 1998; Hosetti and Kumar, 2001). 22 nd -24 th December 2010 Page 1

2 Verma et al., (1978), opined that with the increase in the levels of temperature, hardness and ph of the water medium, the toxicity of the heavy metals also increases. Physico-chemical and biological components of the water body are altered due to the disposal of heavy metals and other toxicants, and furthermore the transformed products of the toxicants have been proved to be more toxic to the aquatic biota (Madhyastha et al., 1996). Lead is one of the common industrial pollutants associated with Zinc, Cadmium, Nickel, Cobalt and Copper is known to affect the growth of aquatic organisms (Kaviraj and Konar, 1983; Hosetti 1988; Hosetti et al., 1993; Tharavathy and Hosetti, 1998). The wastes containing heavy metals when discharged into the stabilization ponds change their functioning and the organisms when exposed to these heavy metals behave differently and survival depends on their ability of tolerance. Some of the heavy metals are essential for the growth in traces as micronutrients, but the same metals are toxic in excess to the aquatic organisms. When DO level is low, it has been observed by Lloyd (1962) that toxicity of lead increased. The toxicity of a metal is affected by various physicochemical factors such as ph, hardness and nutrient levels of the medium (Hosetti et al., 1993). The proper functioning of the stabilization pond system depends upon the growth and biochemical activity of algae. The pollutants enter the pond water and inhibit the algal activity and thus impair the treatment process. In the present study, the raw waste was collected from the industrial complex of Cosme Mathias Menezes Ltd., Curti, Ponda, Goa. The liquid wastes produced by the different units were allowed to flow in a single channel and pumped to the aeration tank. From this tank, the wastes were allowed to flow through trickling filter beds to remove suspended matter before being released to the stabilization pond. The present investigation aims to understand the toxic effects of lead on the physicochemical and biological parameters in the laboratory model waste stabilization ponds. 2. MATERIAL AND METHODS: The raw waste collected from CMM s industrial complex, situated at Curti, Ponda, which is located at latitude 15 24'00" N and longitude 74 00'30"E at an altitude of 100 m served as the growth medium. Five sets of plastic ponds in duplicate were filled with 10 litres of raw waste. The first set served as the control without any metal, and to the remaining four sets lead was added in the form of lead acetate at the rate of 10, 20, 40 and 60 mg/l respectively. The above amount of lead acetate was added after range finding experiments were conducted. To each of the experimental ponds, seed material collected from the field stabilization pond was added at the rate of 50 ml/l. These experimental stabilization ponds were placed in the open sunlight outside the window and were operated on batch system under day and night conditions. 22 nd -24 th December 2010 Page 2

3 The experiment was run for 15 days. Observations were made by collecting the required amount of sample from each pond on 5 th,10 th and 15 th day and analyzed for ph, dissolved oxygen (DO), biological oxygen demand (BOD 5 ), phosphate (PO 4 ) content, chlorophyll content, and algal numbers as per the standard procedures given in Chemical and Biological Methods for Water Pollution Studies (Trivedi and Goel 1990) and Standard Methods for the Examination of Water and Wastewater published by APHA, AWWA, and WEF (1998). Catalase activity was measured according to the procedure of Euler and Josephson (1927) and as adopted by Hosetti and Frost (1994). Phosphatase activity was determined according to the method given by Verstraete et al. (1976) and later modified by Hosetti (1987). Algal members were identified by referring to the keys (Ramaswamy and Somashekar 1982; Hosmani 2002; Kumar et al. 2002). Relationship between various physico-chemical parameters and biological properties was measured by Pearson s correlation coefficient (r). 3. RESULTS: The physico-chemical and biological characteristics of the raw wastes are printed in Table 1. From the data it is obvious that on zero day a minimum DO (1.15 mg/l) was recorded with a ph of 6.5. The Biological Oxygen Demand of the raw waste was 95.4 mg/l and the phosphate content 6.85 mg/l. The activity of catalase and phosphatase was and 1.21 units respectively. However the chlorophyll content was not analyzed as there were no algae in the raw waste. The characteristics of effluents from laboratory model waste stabilization ponds are shown in Table 2. The ph was maximum in the control ponds on the 5 th day of analysis and it decreased with increasing number of days and also with increasing concentration of lead. (Table 2 ). The DO levels in the control pond sample were high on all the days of analysis. It gradually increased and it was maximum on day 15, whereas in case of ponds treated with Lead the DO levels decreased with the increase in metal toxicity. The comparative study on DO levels revealed that it was comparatively low in metal treated ponds (Table 2). The organic strength measured in terms of BOD 5 was reduced to a maximum of 89.3 % on day 15 th in control ponds. The efficiency of BOD 5 removal was comparatively low in the ponds treated with the metal. The BOD 5 removal was 69.1 %, 52.7 %, 45.1 % and 36.2 % in the ponds treated with metal levels 10, 20, 40 and 60 mg/l of Lead respectively (Table 3). The enzymes catalase and phosphatase recorded a minimum of 15.8 & 1.16; 13.3 & 1.05; 11.7 & 0.89; 9.2 & 0.8; and 5.0 & 0.52 units in the pond samples control, 10, 20, 40 and 60 mg/l of Lead 22 nd -24 th December 2010 Page 3

4 levels respectively on day 5. It was interesting to note that the activities of both the enzymes increased gradually in the consecutive days, however there was not much of a hike in the activity of ponds treated with 20, 40 and 60 mg/l of Lead. In case of control and lowest metal treated pond, the activity increased gradually and recorded maximum on day 10 and declined thereafter. The catalase activity was recorded maximum 32.5 & 28.2 units in the control and 10 mg/l Lead treated ponds respectively. Similarly highest activity of phosphatase was 1.25, 1.1, 0.92, 0.83 and 0.58 units in the respective ponds on day 10 (Table 2). The inorganic phosphate recorded to a maximum of 6.85 mg/l in the raw sewage on zero day, was reduced in the pond samples. It was reduced to maximum extent in the control ponds than in other ponds. It was recorded 5.6, 3.5 and 2.2 mg/l on day 5, 10 and 15 in control ponds respectively. Similarly the trend was also followed in the Lead treated ponds. However the phosphate uptake was affected in the presence of Lead and it was severe in the higher concentrations (Table 2). The total solids increased from 285 mg/l to 650 mg/l in the case of experimental pond treated with 60 mg/l of Lead on the 15 th day of analysis which was the maximum. The determination of algal counts and study on algal diversity revealed that both recorded higher values in the control pond and the pond treated with 10 mg/l of Lead. The algal count and the algal diversity both were affected in the ponds treated with higher levels of Lead (Table 4). Chlorophyll content is given in Table 2. The control pond had maximum chlorophyll on all the days of analysis when compared to the experimental pond. In the case of control pond it increased from 97.5 mg/l on the 5 th day of analysis to mg/l on the 15 th day, similarly the experimental pond treated with 10 mg/l of Lead showed the same trend (94.17 mg/l to mg/l) whereas in the other treatments it was seen that the chlorophyll content decreased with increasing number of days. Algal counts also showed the same trend as that of chlorophyll (Table 2). The density and diversity of algae was more in the control and 10 mg/l of Lead experimental pond (Table 4) when compared to the higher treatments of Lead. Also it was observed that the density wise Cyanophyceae members were more but diversity wise Euglenophyceae was rich in the laboratory experimental ponds. With increasing number of days and higher concentration of Lead, it could be seen that only a few algae like Merismopedia tenuissima, Spirulina species and Chlorella vulgaris were able to survive even though their number reduced. 4. DISCUSSION: The ph value of raw sewage usually varied around neutral (7.0). In the hitherto studies the raw waste is a combination of 3 types of industrial wastes and it includes effluents from pharmaceutical, chemical and cosmetic industries. It is due to this reason that the raw waste was having a slightly acidic 22 nd -24 th December 2010 Page 4

5 ph (6.5). It gradually increased and shifted to alkaline side in both control and metal treated ponds. The functioning of the pond was also affected by the hydrogen ion concentration that is the ph. The ph of the raw waste on zero day was 6.5, which increased to 9.0 on the 5 th day in the control pond and later decreased to 8.8 and 8.2 on the 10 th day and 15 th day respectively. Whereas in the metal treated ponds the ph went on decreasing with increasing concentration on all days of analysis. The higher ph in the control ponds was due to the photosynthetic activity of algae which utilizes CO 2 and releases O 2. At higher concentrations due to the lower density of algae the photosynthetic activity was reduced thus reducing the ph which is significantly correlated (Table 5 to 7). The dissolved oxygen analysis showed a declining trend with the increasing levels of the metal treatment throughout the period of the experiment. The maximum concentration of DO was recorded on the 15 th day of analysis in the control pond (Table 2), whereas minimum dissolved oxygen was recorded from the pond treated with highest concentration of Lead (60 mg/l). This was evidenced from the light brownish colour of the water in the ponds treated with 40 and 60 mg/l of Lead. Photosynthetic activity of the pond was reduced at higher concentrations of Lead treatment which affected the growth of algae as they could not tolerate the toxic effects of the metal, thus low amount of DO was produced at high metal concentrations (Patil et al., 1986). The gradual increase in the DO values as the days progress in all the experimental ponds showed that algal activity was increasing day by day and the breakdown of organic matter was taking place till the 15 th day of analysis. The raw waste on zero day had a BOD 5 value of 95.4 mg/l, which drastically reduced in the control and 10 mg/l Lead treated ponds. In the case of control it reduced to 10.2 mg/l and in the 10 mg/l Lead treated ponds, it reduced to 29.5 mg/l with a retention period of 15 days. In the control ponds BOD 5 was reduced by 39 % on the 5 th day and to 89.3 % on 15 th day of analysis. In the case of 10 mg/l Lead treated ponds it reduced by 33.4 % and 69.1 % on the 5 th and 15 th day respectively (Table 3). At the highest concentration of Lead treatment the maximum reduction (36.2 %) in BOD 5 that took place was on the 15 th day. This showed that the bacterial activity which was responsible for the breakdown of organic matter was affected by the toxicity of Lead and also the availability of the DO for their metabolic activity (Whitton, 1970; Whitton and Shehata, 1982). Furthermore the DO production and BOD 5 removal are correlated to one another and are significantly related to each other which can be evident from the correlation matrix (Tables 5 to 7). Studies on the activity of enzymes have been suggested for the evaluation of the quality of effluents (Hosetti and Patil, 1987; Gaddad and Hosetti, 1989) and river water (Parthasarthy et al., 1982; Hosetti and Birasal, 1989). Toxic secondary metabolites like alcohol, hydrogen peroxide, thiols etc. are produced by aerobic cells during oxidative respiration and are also known to release certain enzymes is the waste water which would breakdown and detoxify the toxic secondary byproducts produced during 22 nd -24 th December 2010 Page 5

6 oxidative respiration. If these byproducts had not been broken down and degraded, they would have accumulated in the water and become lethal to the micro-organisms themselves. Catalase produced by the micro-organisms present in the wastewater are known to break down hydrogen peroxide into water and oxygen and thus detoxify which otherwise would have accumulated and become lethal to the cell itself. The enzymatic activity of catalase is taken into consideration for the evaluation of the water and wastewater quality by the previous workers (Gaddad et al., 1982; Hosetti and Patil, 1992; Hosetti and Frost, 1994; Tharavathy and Hosetti, 1998). The raw waste on the zero day showed units of catalase activity, which decreased to units on the 5 th day of analysis and suddenly increased on the 10 th day to the maximum and again decreased thereafter. Similar was the case of the enzyme activity in the ponds treated with increasing concentration of the Lead. The catalase activity in the treated pond samples was much lower than the raw waste, on the 15 th day of analysis especially in ponds treated with 20 mg/l, 40 mg/l and 60 mg/l of Lead. This shows that the enzymatic activity was inhibited by higher concentration of the metal which might be due to the formation of the metal complexes in the pond and also due to the change in the ph levels (Gaddad, 1983; Tharavathy and Hosetti, 1998). Higher catalase activity in the control ponds and treated with low amount of Lead was due to the increased photosynthetic activity which would further enhance the bacterial and fungal activity due to the availability of dissolved oxygen. The catalase activity showed a significant correlation with BOD 5 values on all the days of analysis. The phosphatase activity in the raw waste was 1.21 units on the zero day and in the control ponds it was 1.16 units, 1.25 units 0.91 units on 5 th,10 th and 15 th days of analysis respectively. With increasing concentration of the metal the phosphatase activity decreased which did not recover at all till the end of the experiment. This showed that the enzyme phosphatase is more sensitive to the metal toxicity even at low concentrations. The percent reduction of PO 4 was maximum in the control ponds on the 15 th day (Table 3) and the least in the treated pond which showed that the removal of phosphate was affected at higher concentration due to lower density of algae. Phosphate was absorbed by the proliferating algae which reduced its content in the water (Kosaric et al., 1974). The total solids increased with increasing concentration of the metal on all the days of analysis due to the precipitation and formation of metal complexes at higher concentration and not because of the biomass. At the minimum concentration of 10 mg/l of treatment, the growth of the organisms was normal and comparable to the control ponds, but with increasing concentration of Lead, the algal growth was inhibited. Successful functioning of the waste stabilization ponds depends on the photosynthetic activity of algae which would be affected by the toxic substances like heavy metals present in the wastewaters (Moshe et al., 1972; Mara and Pearson, 1986). 22 nd -24 th December 2010 Page 6

7 The laboratory study showed that there was an exponential growth in the control ponds than in the treated ones. In the control pond Merismopedia tenuissma and Chlorella vulgaris were abundant which also decreased in density with increasing concentration of Lead. Along with the algal density, chlorophyll content also decreased with increasing concentration of Lead and it was at its minimum on the 15 th day of analysis at the highest concentration of Lead treatment. Maximum diversity and density of algae was observed in the control and the ponds treated with 10 mg/l of Lead on all days of treatment. With increasing concentration of treatment and time, it was observed that the number of species reduced and also the density of those tolerant algal species was less when compared to the control ponds. The study revealed that algae are sensitive to the metal toxicity in terms of diversity and to certain extent in density also. 6. ACKNOWLEDGEMENTS: The authors would like to thank the authorities of Cosme Mathias Menezes Ltd., Curti, Ponda, Goa for permitting to carry out the study on their stabilization pond; the Principal and Heads of the Departments of Botany and Zoology of S.P. Chowgule College of Arts and Science, Margao, Goa for the facilities extended. We also thank Dr. B.D. Huddar and late Dr. P.R. Naik for the kind help in identifying the algal species. 7. REFERENCES: APHA: Standard Methods for Examination of Water and Wastewater, 20 th Edition, APHA, AWWA and WEF, New York, USA Euler, H.V. and Josephson, K.: Catalase II, Leibigs Ann., 1,455, Gaddad and Hosetti: Enzymes as indicators of domestic wastewater quality, In Trends in Environmental Pollution and Pesticide Toxicology, Eds. Shashikanth S., Vohra S and Sahai N., Jagamandir Book House, Delhi, , Gaddad, S.M., Jayaraj, Y.M. and Rodgi, S.S.: Catalase and protease activities in relation to BOD removal and bacterial growth in sewage, Ind. J. Environ. Health, 24, , Gaddad, S.M.: Studies on the enzymes in relation to microbial growth and activity in sewage and stabilization ponds, Ph.D. Thesis, Gulburga University, Gulburga, India, nd -24 th December 2010 Page 7

8 Gautam, A.: Conservation and management of aquatic resources, Published by Daya Publishing House, Delhi, Hosetti, B.B.: Studies on the use of some important groups of microbes in the treatment of wastewaters, Ph.D. Thesis, Karnataka University, Dharwad, Hosetti, B.B.: Effects of Zinc toxicity of Chlorella vulgaris in sewage, Environment and Ecology, , Hosetti, B.B. and Birasal, N.R.: Catalase activity: An indicator of self purification in River Kali. J. Nature Conservation, 1, , Hosetti, B.B. and Frost, S.: Catalase activity in water and wastewaters, Water Res. 28, , Hosetti, B.B. and Kumar A: A Text Book of Applied Aquatic Biology, Daya Publishing House, New Delhi- 35, India Hosetti, B.B. and H.S. Patil: Effect of cobalt and zinc on the growth of algae, Chlorella vulgaris and Scenedesmus quadricauda in Sewage. In Environment and Pesticide Toxicity. Edited by : R.C. Dalela, Shashikant, and S.Vohra. Published by the Academy of Environmental Biology, India, , Hosetti, B.B. and Patil, H.S.: Enzymes activity : An indicator of effluent quality of sewage stabilization pond, Bioresource Technology, 39, , Hosetti, B.B., Shivaraj, K.M. and Patil, H.S.: Toxicity of cobalt chloride on Scenedesmus quadricauda (Turp) Breb during sewage purification. Indian Journal of Experimental Biology, 31 : (7) , Hosmani, S. P. (2002). Ecological diversity of algae in freshwater. In B. B. Hosetti (Eds.), Wetland Conservation and Management (pp ). Jaipur: Pointer Publications, Kaviraj, A. and Konar, S.K.: Impact of mixture of mercury, chromium and cadmium on aquatic ecosystem, Environment and Ecology, 1, 159, Kosaric, N., Nguyen, H.T. and Bergougnou, M.A.: Growth of Spirulina maxima in the effluents from secondary wastewater treatment plants, Biotechnol. Bioeng., 16, , nd -24 th December 2010 Page 8

9 Kumar, A., Prasad, U., & Mishra, P. K. : Mathematical modeling for pollution assessment in aquatic environment of coal fields of Jharkhand (India). Ecology of Polluted Waters, 2, , Lloyd, R.: Factors that affect the tolerance of fish to heavy metal poisoning. In Biological Problems in Water Pollution, US Publ. Health Ser., 999, WP. 25, USA, Mason, C.F.: Biology of Freshwater Pollution. Ireland: Longmann Scientific and Technical Publishers, Madhyastha, M.N., Rao, I.J. and Hosetti, B.B.: Studies on some heavy metals on Netravati River, Ind. J. Environ. Health, 38, , Mara, D.D. and Pearson, H.: Artificial freshwater environment waste stabilization ponds, Biotechnology, 8, 177, VCH Weinheim, Moshe, M., Betzer, N. and Ketty, Y.: Effect of industrial effluents on oxidation pond performance. Water Res., 6, 1165, Parthasarathy, U.R., Prasad, D.Y., Pankhapekasan, B., Shankar, G.V. and Raju, N.: Catalase activity measurement: self purification capability of a river receiving kraft mill effluents, Ind. J. Environ. Health, 24, , Patil, H.S., Hosetti, B.B. and Vijayakumar, M.: Effect of cobalt on the growth and biochemical activities of cobalt on the growth and biochemical activities of Chlorella vulgaris in sewage. Environment and Ecology, 4, , Ramaswamy, S. N., & Somashekar, R. K.: Ecological studies on algae of electroplating wastes. Phykos, 21, 83 90, Tharavathy, N.C. and B.B. Hosetti: Laboratory model stabilization ponds for assessing the lead toxicity to micro-organisms, Int. J. Envi Edu. Inf., 17: (4) , UK, Trivedi, P.K. and P.K. Goel: Chemical and biological methods of water pollution studies. Environmental publications, Karad, India Verma, S.R., Tyagi, A.K. and Bansal, A.K.: Environmental Biology, Technical Papers, Proc. Of the Symp., Muzaffarnagar, NDTA, AEB, India, nd -24 th December 2010 Page 9

10 Verstraete, W., Votes, J.P. and Von Lanker, P.: Evaluation of some enzymatic methods to measure the bioactivity of aquatic environments, Hydrobiologia, 49,257, Whitton, B.A.: Toxicity of zinc, copper and lead to Chlorophyta from flowing waters, Arch. Microbiol., 72, 353, Whitton, B.A. and Shehata, F.H.A.: Influence of cobalt, nickel, copper and cadmium in the blue green alga Anacystis nidulans, Environ. Pollut., 27, 275, nd -24 th December 2010 Page 10

11 Table 1: Physico - chemical and biological characteristics of the raw waste on day zero 1. hydrogen minute per 2. nitrophenol per minute per Parameters Values ph 6.5 DO (mg/l) 1.15 BOD (mg/l) 95.4 Catalase (units) Phosphatase (units) Inorganic Phosphate (mg/l) 6.85 Total solids (mg/l) 285 µ moles of peroxide decomposed per 100 ml of sample µ g of p- phosphate liberated 100 ml of sample 22 nd -24 th December 2010 Page 11

12 Sl. No. Days of analysis 5th day (concentration of metal in mg/l) 10th day (concentration of metal in mg/l) 15th day (concentration of metal in mg/l) Parameters C C C ph DO (mg/l) BOD (mg/l) Catalase (units) Phosphatase (units) Inorg. Phosphate (mg/l) Total solids (mg/l) Total chlorophyll content (mg/l) Total algal cells x 1000/ml Table 2 : Changes in the physico - chemical and biological properties of the wastewater treated with different concentrations of lead 1- µ moles of hydrogen peroxide decomposed per min. per 100 ml of sample 22 nd -24 th December 2010 Page 12

13 2- µ g of p-nitrophenol phosphate liberated per min. per 100ml of sample 22 nd -24 th December 2010 Page 13

14 Sl. No. Parameter Days of analysis Concentration of metal in mg/l C BOD Phosphate Table 3 : Percent reduction in BOD 5 & Phosphate levels in ponds treated with lead 22 nd -24 th December 2010 Page 14

15 Sl. Days of analysis 5th day (concentration of metal in 10th day (concentration of metal in 15th day (concentration of metal in No. mg/l) mg/l) mg/l) Name of the Algae C C C Class:Cyanophyceae 1. Merismopedia tenuissima Spirulina sp Class: Chlorophyceae 1. Chlorella vulgaris Class: Euglenophyceae 1 Euglena elongata E. gracilis E. oxyuris Phacus curvicauda P. caudatus P. longicauda Trachelomonas volvocina Class: Bacillariophyceae 1 Navicula sp nd -24 th December 2010 Page 15

16 Table 4 : Effect of lead on the density and diversity of algae in the experimental stabilization pond. (No. of algal cells x1000/ml) Inorganic 88 Total Parameters799ooo777 DO888 BOD888 Catalase77 Phosphatase5 Phosphate Total Solids ph88888 Chlorophyll Total algal cells DO *0.86 *0.84 BOD Catalase *0.82 *0.79 Phosphatase *0.85 *0.83 Inorganic Phosphate Total Solids * nd -24 th December 2010 Page 16

17 ph Total Chlorophyll Total Algal Cells 1 Table 5 : Correlation matrix for lead treatment on 5th day * : not significant Inorganic Total Total Total algal Parameters DO BOD Catalase Phosphatase Phosphate Solids ph Chlorophyll cells DO *0.84 * nd -24 th December 2010 Page 17

18 BOD *-0.81 *-0.8 Catalase Phosphatase Inorganic Phosphate *-0.84 *-0.83 Total Solids *-0.86 *-0.85 ph 1 *0.86 *0.86 Total Chlorophyll Total Algal Cells 1 Table 6 : Correlation matrix for lead treatment on 10th day * : not significant 22 nd -24 th December 2010 Page 18

19 Inorganic 8 Total Parameters799ooo777 DO8888 BOD888 Catalase77 Phosphatase5 Phosphate Total Solids 88 ph 88 Chlorophyll Total algal cells DO * * *0.82 BOD Catalase Phosphatase *0.69 *0.83 *0.85 Inorganic Phosphate *-0.84 Total Solids 1 *-0.69 *-0.83 *-0.85 ph 1 *0.86 *0.75 Total Chlorophyll Total Algal Cells 1 Table 7 : Correlation matrix for lead treatment on 15th day 22 nd -24 th December 2010 Page 19

20 * : not significant 22 nd -24 th December 2010 Page 20

21 22 nd -24 th December 2010 Page 21