Friday 16 December 2011

THE ESTIMATION OF HEAVY METALS CONTENT ON SOME AQUATIC PLANTS AT LAKE


ABSTRACT
This study was conducted to determine the concentration of heavy metals (cadmium, lead, zinc, chromium and manganese.) in four species of plants that are mostly eaten by fish at Lake Gerio namely Pistia Specie, Ipomea Aquatica, Hydrila Verticilla Typha Latifolia. Atomic absorbance spectrophotometer was used in the study. Manganese was found as the highest in concentration in all the species of plants studied with a range of (12.5 – 28.50), Zinc was found as the second highest with a range of (20.00-25.00) this is followed by Lead with a range of (7.00 – 15.00) Chromium was not detected in all the four plants that were studied at Lake Gerio. Cadmium was found in all the four plants sampled at low concentration at range of (2.00-5.10). It can be said that the concentrations of the heavy metals in the studied plant were within acceptable limits as recommended by FAO.






CHAPTER ONE
1.0         Introduction
Heavy metals are toxic to man, animals and plants once their safe limits are exceeded. Their ability to accumulate in plants makes them particularly hazardous. Heavy metals are toxic to all aquatic biota and causes mortality to fish larva, fingerings and adult fish (Warner. et.al, 1991). Heavy metal has a relative high density and is toxic at relatively high concentration. It mainly includes transition metals, some metalloids, lanthanides and actinides. e.t.c.
In terms heavy metal has been called a misinterpretation in an IUPAC technical report due to contradictory definition and its lack of coherent scientific basis, heavy metal can include element lighter than carbon and can exclude some of the heaviest metals. Heavy metals are individual metals and metal compound that affect people health negatively when in large amount. Human beings have been exposed to heavy metal toxics for number of  years, due to the consumption of aquatic animals which has been affected by high content of heavy metals.
Plants are living organisms belonging to life kingdom plantae, they include familiar organisms such as herbs, grasses vites, ferns green algae, blue algae e.t.c
The high concentration of heavy metals in plants affects insectivorous fish                          and other aquatic animals which feed on those plants. As a result of high               content of such elements the aquatic habitat absorbs them through the skin of their feet and also through feeding which when consumed by human beings affect them negatively. (Vaishney C.K, 1991).
1.1         Plants Species Commonly Found In Lake Environment
Ecology is the study of interactions between organisms and their environment. various species living in the same place interacting among themselves and with their environment forms an ecosystem.
A healthy ecosystem has a variety of organism that play different role in various food chain. If the ecosystem losses one of its members it can be crippled, the major types of organism found  in aquatic environment is a fresh water life zone (Anon, 1984).
Aquatic plants are plants that have adopted living within aquatic enviromnent they are also referred to as hydrophytes or aquatic marcrophytes,these plants requires special adaptation for living in submerged water or in water surface.
Some of these plant species found in the lake environment includes:water lectuce,sea weeds,hydropytes,multicellular  marine algae,zooplankton.etc.
1.2         Plants Used As Food Material For Aquatic Animals
Hundred of species of planst are cultivated by humans under managed condition, however a remarkably small number of species contribute greatly to the global harvest of plants. crops are plants grown, tended and harvested by humans as a source of food.
Food is an essential part of living among all human activities. Eating undoubtedly has the greatest effect on health, after breathing eating is what we do most frequently during a life time.
Plant is one of the source of food for aquatic animals and research shows that plant based food are better than animal- based food for good health and longevity (Boyed c.e,1968).
Algae are single celled aquatic plants they are often the producers in aquatic food webs. algae are unique because although they form spheres, sheets, or filaments each cell acts like an  independent organism, They are the simplest  of all plants.
They are not considered to be plants by some scientist and are place in the kingdom protista.
The two common types found in our waters are green algae these are considered as basic biological elements of quality aquatic ecosystem and may consist of single cell or multiple cell colonies that give water a greenish colour.
Diatoms:- Are another group of beneficial silicon (glass- like) skeletal that does not break down over time.
Liverworts and Mosses: These plants are usually found growing in moist   soil along shorelines of water bodies. these simple plants grow in dense mats, usually on ground that has been disturbed. They are more complex than algae, the cells work together, but they do not have conducting cells to move water and food between cells. They readily take up water and hold soil together to help reduce soil erosion and this helps aquatic animals drastically.
Phytoplankton:- These are the base of the aquatic food chain. Small animals consume the phytoplankton and in turn are consumed by large animals.
1.3         Mode of Life of Some Plants in Lakes.
Life is thought to have originated in an aquatic environment’’ the lakes’’ living organisms have since adopted to numerous aquatic habitats.
Above 17 percent of known biological species live in oceans, marine species are described as pelagic organism or benthic organism.
Pelagic organisms or benthic organism. Pelagic organism lives on the lake bottom.
Zooplankton uses a variety of techniques to stay close to the water surface. These includes secretion of oily or waxy substance, possession of air filled sacs similar to the swim bladder of fish and special appendages that assist in floating ,some zooplankton even tread on water.
Cynobacteria as well as true algae are capable of causing various nuisance effects in lake such as excessive accumulators of   foams, scums and discoloration of the water, when a number of algae in a lake or a river increases explosively, an algal bloom is the result. lakes are susceptible to blooms (Duttai,et.al., 1999)
Adaptive Features Of Some Plants
Floating plants: In an outdoor body of water they receive more light than submerge plants do. They also rarely have to compete with another for sunlight. Submerge plants:- The leaves of submerge plants receives lower levels of sunlight because light energy diminishes while passing through a water column.
Water lilly
* Structural material to reach highest point and receive macro sunlight.

Hydrilla
Elongates rapidly to reach water surface and branches out at water surface, more light can be obtained on the outer surface.
Their Relationship
-Ammensalism: This is a parasitic relationship between plants that has a destructive effects on one and no effect on the other.
-Biostatics: Is the study of relationship between structures and functions in plants.
-Diatropism: The capacity or tendency of some plants to adapt a position transverse to the line of force of an external stimulus.
1.4         Nutrient Content Of Some Aquatic Plants.
A study was conducted to evaluate nutritional potential of three local aquatic plants, namely:
-                Nymphaea alba —water lilly (Bum)
-                Nymphoids peltata (Khor)
-                Hydrilla species (khel)
Which grow abundantly in the dal and other lakes and water bodies of Kashmir. Proximate analysis revealed that all the three plants had dry matter below 10%. Crude protein content of Hydrilla, Nymphaea and Nymphoides was 17.10%, 20.28% and 21.87% respectively. Ether extract level of Hydrilla (2.79%), Nymphaea (1.5%) and Nymphoides (3.5%) was also estimated. Ash content varied from 8% in Nymhaea to 19.45% in Hydrilla. Calcium content was 1.29%, 1.10% and 1.56% in Hydrilla Nymphaea and Nymphoides respectively. Fiber content was lowest (13.36%) in Hydrilla and highest (21%) in Nymphoides. Animals fat with aquatic plants (Group A) produced about 3. litres macro milk per day per animal than straw fat  Group (C) animals, there was however no significant difference in daily milk yield of Group A and  (Group B) animals (Zolo vol. 5, No. 1 Article 53)
All the three plants had below 10%   which lies in agreement with reports of (Boyd, 1968). Crude protein content of Hydilla, Nymphaea and Nymphoides was 17.10%, 20.28% and 20.87% respectively which is comparable findings of  (Boyd, 1972) the lower values observed in Hydrilla which is a submerged plant, may be due to environmental differences as nutrient concentrations. Nymphaea alba has medicinal properties and perusal of literature reveals that all these plants contains traces of alkaloids and polyphenols but within a range. (Baherjec and Matai, 1990). Which will not produce any harmful effect to the animals.
1.5         Toxic Effects of Some Plants on Aquatic Animals.
Toxic substances are chemicals at some concentration can harm aquatic animals. Aquatic animals may be poisonous by pesticides residues that remain on food after spraying for example when aquatic animals enter sprayed seas shortly after spraying.
Widespread application of pesticides can eliminate food sources that certain types of animals can need causing the aquatic animals to change their diet or starve. Some pesticides can bioaccumulate or build up to toxic levels in the bodies of organisms that consume them over time. (Raloff, J Sept 5, 2007).
Fish and aquatic biota may be harmed by pesticide — contaminated water. Pesticides contaminated water can be highly lethal on aquatic life, sometimes killing all the fish in a particular lake.
Application of herbicides to bodies of water can cause fish kills when the dead plants rot and use up the water oxygen suffocating the fish. Some herbicides such as copper sulphate that are applied to water kill plants are toxic to fish and other aquatic animals at concentration similar to those used to kill plants.
More so, the application of herbicides to body of water can kill off plants on which  fish depends on  for their habitat (Bingham, S. 2007).
Pesticides can accumulate in bodies of water levels that kill off  zooplankton the main source of food foroung fish. It can also kill off insects on which fish feeds causing the fish to travel farther in search of food and disposing them to greater risk from predators. The faster a given pesticides breaks down in the environment the less threat it poses to aquatic life. (Lorenz. Eric S. 2009)
1.6         Aim and Objectives
The aim is to ascertain and find out whether the contents of  heavy metals on plants are above or below the required standard.
And the objective is to determine  the composition of heavy metals on plants.


CHAPTER TWO
2.0   LITERATURE REVIEW
Numerous studies have been carried out on the assessment of plants and scientists have it that plants in general is the major source of raw materials in the industry. Though the growth of plants is hindered by excessive concentration of both essential and non essential heavy metals which results in photo toxicity. Although the relative toxicity of different metals to plants varies with experimental condition. The metals which when present in excessive amount are most toxic to higher plants and micro organisms. This metals includes mecury,copper, nickel etc. (Kumar and Singh, 1980).
Heavy metals in the environment have a bad impact on the lives of both aquatic animals and aquatic plants. The presence of heavy metals in lake may bring about poisoning of aquatic life. (Eilli.K.V .1989).
The nutritive value of aquatic plants which often contains as much crude protein, crude fats and crude mineral matter could help to alleviate food shortages until lasting agriculture and social solutions are found. (Edie H. H and B.W.C, 1969).


2.1 Classification of Some Plants Specie Found In Lake
In order to study nature scientists have classified the lifespan or forms in plants by putting them into groups based on how they are related to each other. (Duttai,et.al.,1998).
Plants has been classified as follows:
1. CLASS              Angiosperm           These are plants that produces flowers.
2. SUBCLASS       Dicotyledons            These are plants with two seeds.
3. ORDER              Hammalidae          A group of related families developed from a common ancestors. 
4.FAMILY                                            each order is divided into sub families. 
5.GENUS                                                 the plant  that is most familiar.
6.SPECIE.                                               the level that defines an individual plant.
7. VARIETY                                           it is a plant that is different from other species of plants..
8. FORM                                                  a form is a plant between a specie
2.2 Plants Commonly Used By Fish and Snails as a Major Source of
Nutrient.
Photosynthesis is the process by which plants converts carbon dioxide and water with the help of light energy into glucose energy and oxygen gas. This process can be expressed with the equation below.
6C02 + 6H2O+ Sunlight     C6H12O6 + 602
Thus in an aquarial environment during the day plants uses the carbon dioxide produced by fish and water to produce oxygen and energy, the oxygen is used by fish for respiration. The main source of nutrient is the decayed, plants they produces the major source of oxygen for fish, snails and other aquatic habitat ( Bannerjee, G. C., 1990).
Plants specie such as algae has a reserve carbohydrate in form of starch and thus when being consumed by fish produces nutrient in their system or body (Anon, et.al,1998).
These are specie of plants used by fish , snails as a major source of nutrient:
-Fungi, Water lettuce,  Water pepper, Water spinach ,Sea weeds ,Legumes,Wheat,
-        Water cress , Sugar cane,  Sugarbeets ,Wild rice , Chestnut.
2.3 Nutrient Content of Some Edible Plants in Lakes.
What are nutrients? Food consist of different components of variable nutritional value called nutrients, these are carbohydrates, proteins, fat and oil, mineral salt, vitamins and water. (Boussingault, J.B 1999).
Carbohydrates, proteins and fats and oils are referred to as macro nutrients because they are needed in large quantity while vitamins and minerals are referred to as micro nutrients needed in small quantity. Carbohydrates functions mainly as a supplier of energy to carry out bodies function. Protein serves as a body building food it also replace worn out tissues. Fats and oils helps in absorption of fat solubles vitamins A, D, E and K. minerals forms part of the tissues and skeletons. Various edible fungi contains the nutrient necessary for the plants initial growth, some of these plants includes:
- Algae - rich in carbohydrates and minerals.
- Water lilly - rich in protein and vitamins.
- Water lettuce - rich in vitamins.
- Water spinach - rich in vitamins.
Alligator weed (Alterhanthera Philozeroides) has so much mineral content used by aquatic animals. It has a more improved with salt and salad dressing, this content of nutrient is really a source of minerals to aquatic animals found in lakes (J.F Hentages, 2004).
Table 2.3.1 : Chemical composition of some aquatic plants (%DM)
CONSTITUENTS
HYDRILLA
NYMPHAEA
NYMPHOIDES
Orgarnc matter
80.44
88.40
88.40
Crude protein
17.10
21.87
21.87
Ether extracts
2.79
3.50
3.50
Total Ash
19.45
11.60
11.60
Crude fiber
13.34
21.00
21.00
Nitrogen free extract
43.92
49.03
49.03
Calcium
1.29
1.56
1.56
Dry matter
6.12
8.12
7.72

2.4 Mode of Life of Some Lake Plants
Fresh water aquatic plants are referred to as hydrophytes or aquatic macrophytes. These plants require special adaptations for living in submerged water or at the water surface. Aquatic plants can only grow in water.
These are certain adaptations undergone by lake plants which enables them to adapt to their more of life. Some of these adaptations includes:
* all floating plants have air spaces trapped in their roots or else air space in their bodies (Aerenchyma) to help them float in water thus receiving adequate sunshine.
* they have hair on their leaves that traps air.
* they have structural adaptations.
A simple organism known as blue-green algae appeared and spread across the seas, the blue-green algae uses sunlight and water to make food and in the process creates oxygen. As the blue-green algae in the earth’s seas they began to fill the atmosphere with oxygen. The oxygen blue-green algae produced made it possible for other types of organisms to develop. This oxygen helps them to carry out their mode of life processes of growth, feeding, responding and reproducing unlike the blue-green algae those organisms could not produce their own food. They needed oxygen to perform their lives processes of growth, feeding, responding and reproducing; in return they produce carbon dioxide which the algae needs to perfonn its life processes. A précised balance was established between the plants. Some of the lake plants produce their young ones asexually which yields reproduction. (Willard. I.1997).
CHAPTER THREE
3.0 Material And Method
3.1 Materials Required
3.1.1 Equipment and Apparatus
Beaker pyprex lOOml , Mortar and Pestle, Spatula, Conical Flasks, Filter Paper, Funnels, Hot Plate, Fumecupboard, Round bottom Flask, dessicators, Weighing balance, Atomic Absorption Spectrometer,
3.1.2  Chemicals / Reagents.
The Chemicals used for this analysis were of BDH grade and are listed below.
          Copper nitrate (Cu (NO3)2.4H2O)
          Iron sulphate (FeSO47H2O)
          Manganese Chloride (MnCl2)
          Cadmium Chloride (CdCl2)
          Calcium Chloride (CaCl2)
          Magnesium Chloride (MnCl26H2O)
          Chromium (iii) Chloride (CrCl3)
          Lead (iii) Nitrate Pb (NO3)2
          Zinc Nitrate Zn (NO3)26H2O
          Hydrochloride Acid (HCl) 36% W/WS.G 1.18
          Nitric Acid (HNO3) 43% W/WS.G 1.42
          Distilled water.
3.2 Sampling and Sample Preparation
3.2.1 Sampling
Sampling was done similar to that described by ledger (1974) sampling is a method of obtaining a small part or portion of large quantity of material which represents the whole amount of the material.
Four different species of plants were collected and the samples are named as follows.
- Pistia specie  (water lecttuce)
- Ipomea aquatica. (water spinach)
- Hydrilla verticillata (water weed)
- Typha latifolia (broad leaf cattail)
The samples are collected at random in the lake Gerio of Adamawa state. during the sampling the plants were uprooted from the root. Following the method described by (Shirmal and Vegas (1975).
3.2.2 Sample Preparation.
The samples collected were  air dried, grounded using mortal and pestle and then stored for analysis
3.3 Analytical studies of plants.
3.3.1Ashing of samples.
The crucible was placed in a hot plate it was then cooled in desiccators and weighed, Twenty grams of the sample was weighed into the crucible.
The crucible and the content was transferred to a furnace and heated at 600°C  for three hours to burn of f the organic matter  living the ash.  
3.3.2 Sample digestion
The sample each weighing 10g was digested in 15rml aqua regia (HNO3: HCL in the ratio of( 3:1)  it was poured inside a kjeldahl  flaskand swirl to wet the sample.stand over night.The next  day the flask was placed in the heating mantle and was heated for two hours  after which it was cooled  this was filtered and the volume of filtrated noted.


3.3.3 Preparation of standard stock solutions and serial dilutions used for Atomic Absorption Spectrophotometer.
(a)           Preparation of chromium stock solution. 3.050g of chromium (iii) chloride (CrCl3) was weighed using an electronic balance; it was transferred into a 1000cm3 volumetric flask and filled to the mark with distilled water. A serial dilution of this solution was done to obtain solution containing 10, 15, 20 and 25  ppm.for construction of the calibration curve.
(b)          Preparation of zinc stock solution. 4.550g of zinc nitrate Zn (NO3)26H2O was weighed serial dilution was 0.5,1.01.5, and 2  ppm.
(c)           preparation of  manganese stock solution.2.290g  of Manganese Chloride (MnCl2 ) was weighed using an electronic balance, the working  was prepared  as described above, the serial dilution was  1, 1.5, 2.5  and 3 ppm.
(d)          Preparation of Cadmium stock solution. 1.630g of dried cadmium was weighed using electronic balance; the working standard was prepared as described above. the serial dilution was 4, 6, 8,   and 10  ppm.  
(e)           Preparations of Lead stock solution. 1.600sg of lead (iii) Nitrate Pb (NO3)2 was weighed using an electronic balance. The working standard was prepared as described above, the serial dilution was 10, 20, 30,  and 40 ppm.
3.4.1 Determination of metals by Atomic Absorption Spectrophotometer
3.4.2 Principle of Atomic Absorption Spectrophotometer
The determination of heavy metals in plants, soil, water and in acid sediment digest of biological tissues are done in most analytical laboratories using atomic absorption spectrometer.
A number of authors including Bartram and balance (1996) have described the principles of AAS for metals analyzed in water and plants.
The principle is based on the fact that absorption of electromagnetic radiation by atoms is proportional to the number of atoms present in the material, here atomization of the element is achieved by introducing energy equal to the energy required for the transition of the analyte element  from ground state (EO) to an excited electrons state (E1). Associated radiation is passed through a monochromatic for isolation of spectral line and then into the photo detector. The absorbance being measured is a function of the analyte concentration and is given by the equation.
A=ebc     or
 = A= Log Io/It=ebc
Where
A= Absorbance
e= Absorbtivity constant
b= Path Length of light
C= Concentration
I0 and It = Intensities of incidence and transmitted radiation respcectively 
3.4.3 Method of Determination of Metals by AAS.
Absorbance readings for the working standards were obtained on the AAS, for the various metals using the Absorbance for the samples were obtained under similar instrumental condition.
Calibration curves was constructed for t the various metals from result absorbance reading reading of the working standards.
The concentration of the sample was obtained by extrapolating the absorbance reading on calibration curves.
Results obtained on the curves were in parts per million (ppm) which was then converted to mg/kg of the plant sample by using the formula.
mg/kg =                    
ppm= concentration in parts per million
v=volume of the digested plant
wt= weight of sample



CHAPTER FOUR
4.0   RESULT AND DISCUSSTION
4.1 RESULT FOR  HEAVY METAL CONTENT OF AQUATIC PLANTS.
The Table 4.1 below shows the result for the concentration of heavy metals of different species of plants, mostly eaten by fish at lake gerio. The results are the mean values of four determinations by AAS.
Table 4.1 Results of analysis of some plants species             
Plant type          Concentration of heavy metals (mg/kg)
                                   Mn
Cd
Cr
Pb
Zn
Pistia specie
28.25
5.10
ND
15.00
21.25

Ipomea aquatica
28.50
5.00
ND
12.50
25.00

Hydrilla Verticilata
12.50
3.00
ND
10.00
20.00

Typha specie
28.00
4.00
ND
7.00
23.75












ND=Not detected



4.1   DISCUSSIONS   
The table above shows the concentration of the heavy metals of different specie of plants.  Five heavy metals were determined in the four different plants mostly eaten by fish and all the  essential heavy metals determined were present with the exception of cromium which was not detected in all the plants studied.
The metals of high concentration are Mn, Zn and Pb in an increasing order. Cadmium is of low concentration. The result, however are within acceptable values of world standard.
Manganese is the most abundant in the species of plants with a range of (12.5 – 28.50) Zinc as indicated is the second most abundant followed by Lead.
Cadmium as indicated is the least abundant in all the species of plants with a range of (2.00 – 5.10), chromium was not detected in all the species of plants studied.
The resuls are however within acceptable values of plants as reported by Hardy, et al (1991).

CHAPTER FIVE
5.0   Conclusion and Recommendation
5.1   Conclusion
Four elements were detected from the five elements on study by atomic absorption spectrophotometer. From the result obtained, we can conclude that manganese has the highest value of heavy metal which is followed by zinc then lead. Cadmium has the lowest value of heavy metals. Chromium was not detected. Therefore the plants species do not have high levels of heavy metals that are of fearful pollution load in lake Gerio especially there effect on animal life.
5.2   Recommendation
This study considers five heavy metals contents of plants, further work could be carried out on some heavy metals like Arsenic, Mercury and Bismuth.
The use of more sophisticated instruments is recommended to ascertain the levels of heavy metals on plants like X-ray fluorescence and ICP-OES.


REFERENCES
Alloyway B.J Ed (1990): Heavy metals in plants Halsted press New York pp.4- -100’
Anon. (1984): making aquatic weeds useful: some perspective for developing countries. National Academy of sciences Washington D.C, 175.
Banerjec, A and metal .S.. (1990): Composition of Indian aquatic plants in relation to utilization as animal forage: J. Aquatic plant manage 28: 69-73
Bingham, S. (2007): Pesticides in rivers and lakes. Environment Agency
retrieved on September 15, 2007.
Boussingualt J.B (1847): Academic of science. A history of nutrition
Boston hough teon niffin.
Boyd, C. E. (1968): Fresh water plants: a potential source of protein economic Botany 22: pp 359-368.
Duitta, A. C. Botany and biology, cotton college Guahati.
Ellis, K. V white G. Warn (1998): Surface water population and its
control pp.23-26
Karl .M. (2002): Agency for toxic substance and disease registry (AYSDR)
Lorenz, Eric S.” Potential Health effects of pesticides “Ag Communications and marketing (2009) pp. 1-8
Osiei Y. Ababio ((1990): New school chemistry new edition African. Feb .Publisher limited.
Raloff .J. (September 5.1998): Common pesticide clobbers amphibians, science News, and volume. 154, Number 10, PP, 150 — 158. Retrieved 2007-10-15
Vaishney, ck (1991); heavy metal in aquatic environment by k. c. pillar in lake. Wiley eastern L.t.d new Delhi India. Pp, 70-74.
Willard. L. (1974): the plant root and its environment 2”’ edition university press of Virginia
William Lramsey(1982),Holt earth science Pp.215-217.s.



Appendix 1;Absorbance readings of standard solutions for calibration curves.

Plant samples
Absorbance
reading (nm)
Concentration
Mn(ppm)
Absorbance (nm)
  Pistia species               
    Ipomea aquatica
Hydrilla verticillata
Typha latifolia
0.998
1.o99

o.397

1.166
1
1.5
2.5
3
0.076
0.144
0.275
0.389


Cd (ppm)
Absorbance (nm)
 Pistia species
Ipomea aquatic
Hydrilla verticillata
Typha latifolia
0.030
0.021
0.017

0.025 
4
6
8
10
0.921
1.035
1.062
1.226


Pb (ppm)
Absorbance (nm)
Pistia specie
Ipomea aquatic
Hydrilla verticillata
Typha latifolia
0.05
o.04
0.03

0.02
10
20
30
40
0.051
0.244
0.540
0.872


Zn (ppm)
Absorbance (nm)
Pistia specie
Ipomea aquatic Hydrilla verticillata
Typha latifolia
1.154
1.250
1.056

1.181
2
3
4
5
0.069
0.139
0.172
0.242


Cr (ppm)
Absorbance (nm)
Pistia specie
Ipomea aquatica
Hydrilla verticillata
Typha specie
ND
ND
ND

ND
10
15
20
25
0.010
0.016
0.025
0.030



Appendix 2: Calibration curve for the various heavy metals
Absorbance (nm)
      1.25

      1.20

      1.15

      1.10

      1.05


      1.00

      0.50

      0.00
                                           2.0                               4.0                          6.0                          8.0                          10.0
                                                                                Concentration (ppm)
Calibration curve for Cd
                                                                   

                                                        


 Calibration curve for Lead (Pb)
Absorbance (nm)


            1.0


            0.90

            0.80

            0.70

             0.60


             0.50

              0.40
    
             0.30


             0.20                        

             0.10

              0.00
                                                   10                         20                           30                           40                           50                                                                                                              concentration (ppm)
Calibration curve for Zinc (Zn)
Absorbance (nm)


          0.30

          0.25

         0.20

         0.15     
                                     
         0.10

        0.05

         0.00
                                                1.0                          2.0                          3.0                          4.0                          5.0
                                                                                        Concentration (ppm)


Calibration curve for Chromium (Cr)

Absorbance (nm)
                                                                               
    0.034

    0.028

     0.022

     0.016

     0.010

     0.000                                                                                                                                                                                               
                                              5.0                            10.0                        15.0                        20.0                        25.0
                                                                                Concentration (ppm)

                                                         






Calibration curve for Manganese (Mn)

Absorbance (nm)


       0.45


       0.35


      0.25

      0.15


      0.05

      0.00
                                               1.0                           1.5                          2.0                          2.5                          3.0
                                                                                Concentration (ppm)