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Shrimp (Heterocarpus ensifer) Exoskeleton as a Source of Chitin Component for

the Biosorption of Lead and Copper

An Investigatory Project Presented to


Department of Education (DepEd)

As an Entry to the
2010 – 2011 INTEL PHILIPPINE SCIENCE FAIR
(Regional Level) Cluster 2
Applied Science – Team

Proponents

Clarisse Anne Caganda


Alyanna Katrina Velasco
Anne Sherina Lazarte
IV-Curie

Miss Judah D. Tolosa


Research Adviser

RAMON MAGSAYSAY (CUBAO) HIGH SCHOOL


EDSA Corner Ermin Garcia St. Cubao, Quezon City

AUGUST 2010

1
IPSF OFFICIAL ABSTRACT FORM
Category
Shrimp (Heterocarpus ensifer) Exoskeleton as a Source of Chitin Pick one only-
Component for the Biosorption of Lead and Copper mark an “X” in
Clarisse Anne Caganda, Alyanna Katrina Velasco, Anne Sherina Lazarte
Ramon Magsaysay (Cubao) High School (EDSA corner Ermin Garcia Street Biochemistry 
Quezon City, Philippines)
Botany 
This study is conducted to determine the biosorption capability of Shrimp Chemistry 
(Heterocarpus ensifer) Exoskeleton in accumulating lead and copper. Computer Science
Based on the results, the addition of 300 mg and 500 mg biosorbent
material to the copper solutions decreased the concentration of copper, from 12, 
570 mg/L to 8,238.7 mg/L and 7,961.7 mg/L, respectively. The same thing
happened to the lead in the lead solutions by the addition of 300 mg and 500 mg Earth and Space 
biosorbent material decreased the concentration from 2, 590 mg/L to 221.7 mg/L
Sciences
and 162.7 mg/L. Moreover, addition of 500 mg of the biosorbent yielded the
lowest final concentration of copper and lead. Hence, as the amount of
biosorbent increases the concentration of copper in the solution decreases. Also, Engineering 
the sorption percentage increases as the amount of biosorbent added increases. Environmental 
From the results and findings, the researcher found out that addition of
Science
300 mg of the biosorbent material will result to the sorption percentage of 34.46
% and Qeq of 14.44 mg Cu/mg biosorbent while the addition of 500 mg
Mathematics 
biosorbent material will yield a sorption percentage of 36.66 % and an Qeq of
9.217 mg Cu/mg of biosorbent. The same thing is concluded in the biosorption of Medicine and 
lead. The addition of 300 mg of the biosorbent material will result to the sorption Health
percentage of 91.44 % and Qeq of 7.894 mg Pb/ mg of biosorbent, and the
addition of 500 mg of the biosorbent material will result to the sorption Microbiology 
percentage of 93.72 % and Qeq of 4.855 mg Pb/ mg of biosorbent.
Moreover, there is a significant difference in the initial and final Physics 
Zoology
concentration of copper and lead with and without the addition of biosorbent in
the copper and lead solutions. The final concentration of copper in the copper 
solution added with 300 mg and 500 mg biosorbent decreased to 8,238.7 mg/L
and 7,961.7 mg/L, respectively starting with the initial concentration of 12,570 Gerontology 
mg/L, and the final concentration of lead from the lead solutions also decreased
to 221.7 mg/L and 162.7 mg/L respectively, starting with the initial concentration
of 2,590 mg/L.
Also, there is a significant difference in the final concentration of copper
and lead by adding 300 mg and 500 mg of biosorbent. As the amount of

1. As a part of this research project, the student directly handled, manipulated, or interacted with (check ALL
that apply):
 human subjects  pathogenic agents  recombinant DNA

 nonhuman vertebrate animal controlled substances  human/animal tissues

2. Student independently performed all procedures as outlined in this abstract.  Yes  No

3. This project was conducted at a Registered Research Institution.  Yes  No


4. Is this project a continuation?  Yes  No
I/We hereby certify that the above statements are correct and the information provided in the Abstract is
the result of one year’s research. I/We also attest that the above properly reflects my/our own work.
2
______________________________________ ______________________
2010-2011 Intel Philippines Science Fair
RESEARCH PLAN
Required For All Projects
To be Submitted to IRB/SRC Before the Experimentation Begins

Clarisse Anne Caganda/ Alyanna Katrina 2 Category: Applied Science (Team)


Name: Velasco/ Anne Sherina Lazarte
Cluster :

Ramon Magsaysay (Cubao) High School


School:

School Address: Ermin Garcia St. Corner Edsa Cubao, Q.C. Tel. No.: 4144098

Title of Project:Shrimp (Heterocarpus ensifer) Exoskeleton as a Source of Chitin


Component for the Biosorption of Lead and Copper

Project Adviser: Miss Judah D. Tolosa

Statement of the Problem

This study aims to find out the biosorbent property of the exoskeleton of

shrimp in accumulating lead and copper from their solutions. Specifically the

investigatory project sought to answer the following questions.

1. Is there any significant difference in the concentration of copper and lead

without and with the application of biosorbent in the following solutions:

a. Pb(NO3)2 solution?

b. CuSO4 solution?

2. Is there a significant difference in the concentration of copper and lead

without and with the application of 0.3 g biosorbent?

3
3. Is there a significant difference in the concentration of copper and lead

without and with the application of 0.5 g biosorbent?

4. Is there a significant difference in the concentration of copper and lead

with the application of the following amounts of biosorbent:

a. 0.3 g

b. 0.5 g

Hypothesis

1. There is no significant difference in the concentration of copper and lead

without and with the application of biosorbent in the following solutions:

a. Pb(NO3)2 solution

b. CuSO4 solution

2. There is no significant difference in the concentration of copper and lead

without and with the application of 0.3 g biosorbent.

3. There is no significant difference in the concentration of copper and lead

without and with the application of 0.5 g biosorbent.

4. There is no significant difference in the concentration of copper and lead

with the application of the following amounts of biosorbent:

a. 0.3 g

b. 0.5 g

4
General Procedure

Collection of Shrimp (Heterocarpus ensifer) Exoskeleton


PHASE I:

Preparation of Soaking in distilled H2O for 24 hours and rinsing three times
the Biosorbent

Air drying for two days and storing at room temperature

Preparation of copper and lead solutions with initial concentrations of 12,570mg/L and
2, 590 mg/L.

PHASE II:

Preparation of Distribution of stock solution 100 mL stock solution in 9 different


the Copper Erlenmeyer flasks at room temperature.
and Lead
Solutions
Addition of 2 mL acetate buffer to each of the Erlenmeyer flasks.
Using a data logger to determine the pH of the solution

Determination of initial copper concentration using


spectrophotometer

Addition of different amounts of biosorbent (0.3 g, and 0.5 g) to 9


separate flasks containing Cu stock solution with constant initial
PHASE III: [Cu] and to 9 separate flasks containing Pb stock solution with
constant initial [Pb].
Varying
Shaking the samples for two hours and storing for 24 hours.
biosorbent to
Filtering the biosorbent using filter paper.
copper/lead
ratios
Determination of final copper and lead concentration using
spectrophotometer

Using a statistical tool to determine the significant difference in the


PHASE IV: concentration of copper and lead with and without the addition of
biosorbents in copper(II)sulfate and lead (II) nitrate solution with
Statistical test varying amounts.

5
Discussion

Based on Tables 3 and 4, the final concentration of copper decreased

upon the addition of varying amounts of biosorbent in the copper (II) sulfate and

lead (II) nitrate solutions. The result indicates that higher amount of biosorbent

lowers the concentration of copper and lead in their solutions.

Moreover, Tables 5 and 6 showed that as the amount of biosorbent added

increased, the sorption percentage (%S) also increased in both the copper and

lead stock solutions. This indicates that higher amount of biosorbent results to

higher sorption percentage (%S).

On the other hand, the specific uptake (Qeq) of copper with the addition of

300 mg and 500 mg of biosorbent yielded, 14.44 and 9.217 mg Cu/mg

biosorbent, respectively. The specific uptake (Qeq) of lead with the addition of

300 mg and 500 mg of biosorbent yielded, 7.894 and 4.855 mg Pb/mg

biosorbent, respectively. Hence, it shows that the biosorbent, exoskeleton of

shrimp (Heterocarpus ensifer) can accumulate heavy metals, such as copper and

lead.

The results obtained were supported by the findings of Guibal (1999),

Mcafee (2001) and Navarro (2000), that chitin/chitosan, which is produced

industrially from materials such as crab shells, can be effective at binding heavy

metals. Moreover, according to Niu and Volesky (2006), chitin/chitosan which is a

component of crustacean exoskeleton, function similarly to special anionic

exchange resins, in sequestering anions. This component contains chemical

active groups within their structures and it is often the NH2 group that is active.

6
Thus, it is possible that this amino group is responsible in accumulating the

heavy metals such as copper and lead.

On the other hand, based on the statistical test employed, there is a

significant difference between the concentration of copper in the copper (II)

solution without the biosorbent and the copper solution with 0.3 g biosorbent.

Likewise, there is a significant difference between the concentration of copper in

the copper (II) solution without the biosorbent and the copper solution with 0.5 g

biosorbent. Also, , there is a significant difference between the concentration of

copper in the copper (II) solution with 0.3 g biosorbent and the copper solution

with 0.5 g biosorbent Hence, the concentration of copper in the copper (II)

solutions with the addition of 0.30 g And 0.50 g biosorbent decreased the

concentration of copper.

The same results were obtained for lead; there is a significant difference

between the concentration of lead in the lead (II) nitrate solution without the

biosorbent and the lead solution with 0.3 g biosorbent. Likewise, there is a

significant difference between the concentration of lead in the lead solution

without the biosorbent and the lead solution with 0.5 g biosorbent. Also, there is a

significant difference between the concentration of lead in the lead solution with

0.3 g biosorbent and the lead solution with 0.5 g biosorbent Hence, the

concentration of lead in the lead solutions with the addition of 0.30 g And 0.50 g

biosorbent decreased the concentration of lead.

7
Bibliography

Books

• Aldor, I., Fourest, E., Volesky, B., & Can, J., (1995). Chem. Eng. Montreal.

• Davis, T.A., Mucci, A., & Volesky, B. (2003). A review of the biochemistry

of heavy metal biosorption by brown algae. Montreal.

• Fourest, E., & Volesky, B. (1996). Environ. Sci. Technol. Montreal.

• Guibal, E., Roulph, C., & Le Cloirec, P. (1992 ). Water Res.

• Hu, M.Z.-C., Norman, J.M., Faison, N.B., & Reeves, M. (1996) Biotechnol.

Bioeng.

• Kratochvil, D., & Volesky, B. (1998). Removal of trivalent and hexavalent

chromium by seaweed biosorbent. Montreal.

• Kuyucak, N., & Volesky, B. (1990). In Biosorption of Heavy Metals.

Montreal.

• Mullen, M.D., Wolf, D.C., Beveridge, T.J., & Bailey, G.W. (1990) Sorption

of heavy metals by soil fungi Aspergillus niger and Mucor Rouxii

• Naja, G.M., & Volesky, B. (2006). Multi-metal biosorption in a fixed- bed

flow-through column. Montreal.

• Niu, C.H., & Volesky, B. (2006). Biosorption of chromate and vanadate

with waste crab shells. New York.

• Volesky. B. (2003). Sorption and Biosorption.

• Volesky, B. (2007). Biosorption and me. Montreal.

• Volesky, B., & Holan, Z.R. (1995). Biotechnol. Prog.

8
• Volesky, B., Yang, J., & Niu, H. (2001). Biosorption of metal cations and

anions. In: Ion Exchange and Solvent Extraction (Vol.14, pp. 119-

168).New York

• Volesky, B., Weber, J., & Park, J. M. (2003). Continuous-flow metal

biosorption in a regenerable Sargassum column

Journal

• Ahalya, N., Ramachandra, T.V., & Kanamadi, R.D. (2003). Biosorption of

Heavy Metals. Research Journal Of Chemistry And Environment, 7, 4.

Internet

• Copper - Cu. (1998-2009) Lenntech BV. Retrieved August 28 ,


2010, from
http://www.lenntech.com/periodic/elements/cu.htm#ixzz0xuL1RCXp

• Lead – Pb. (1998-2009) Lenntech BV. Retrieved August 28 ,2010,


from http://www.lenntech.com/periodic/elements/pb.htm

• Parvathi, K., Nagendran, R., & Naresh, R. (2007). Lead biosorption onto

waste beer yeast by-product, a means to decontaminate effluent

generated from battery manufacturing industry [Electronic version].

Journal of Biotechnology. Retrieved January 15, 2007, from

http://www.ejbiotechnology.info/content/vol10/issue1/full/13/index.html

• Volesky, B. (2010) bv-Publications. Retrieved January 2010, from

http://biosorption.mcgill.ca/

• Volesky, B. (2009, March 31). Biosorption. SciTopics. Retrieved July 19,

2010, from http://www.scitopics.com/Biosorption.html

9
• Wilson, L. (2010). COPPER TOXICITY SYNDROME. Retrieved August
28, 2010 from
http://www.drlwilson.com/articles/copper_toxicity_syndrome.htm

10
APPROVAL SHEET

This Science Investigatory Project entitled Shrimp (Heterocarpus ensifer)

Exoskeleton as a Source of Chitin Component for the Biosorption of Lead and Copper,

prepared and submitted by Clarisse Anne Caganda, Alyanna Katrina Velasco and Anne

Sherina Lazarte as part of the requirements in Research 2 and an entry to the 2010 –

2011 INTEL PHILIPPINE SCIENCE FAIR (Division Level) Cluster 2 Applied Science –

Team, has been examined and recommended for acceptance an approval for the

Science Congress.

_______________________

Miss Judah D. Tolosa


Research Adviser

Date of Congress:

Noted:

____________________`

Mrs. Araceli B. Señido


HT III, Science Department

11
TABLE OF CONTENTS

Page
TABLE OF CONTENTS.....................................................................................................i
APPROVAL SHEET………………………………………………………………………..........ii
ABSTRACT.........................................................................................................………..iii
TABLE OF CONTENTS...................................................................................................iv

Chapter
1 INTRODUCTION....................................................................................1
Background of the study.............................................................3
Conceptual Framework …………………………………………….4
Statement of the Problem...........................................................5
Hypotheses................................................................................5
Significance of the Study............................................................6
Definition of Terms ………………………………………………….8

2 REVIEW OF RELATED LITERATURE...................................................9

3 RESEARCH METHODOLOGY................................................................
Research Design…………………………………………………...24
Materials and Equipment………………………………………….24
Data Gathering Procedure………………………………………..27
Statistical Test………………………………………………………26

4 PRESENTATION, ANALYSIS, AND INTERPRETATION OF DATA


Results ……………………………………………………………..28
Statistical Results …………………………………………………30
Discussion ……………………………………………………… 31

5 SUMMARY OF FINDINGS, CONCLUSIONS, RECOMMENDATIONS


Summary of Findings …………………………………………… 34
Conclusions …………………………………………………….. 35
Recommendations:…………………………………………….. 36

BIBLIOGRAPHY………………………………………………………………… 37

APPENDICES

APPENDIX A ……………………………………………………………………. 39

APPENDIX B ……………………………………………………………………. 40

APPENDIX C ……………………………………………………………………. 41

APPENDIX D…………………………………………………………………….. 43

APPENDIX E ……………………………………………………………………. 45

APPENDIX F …………………………………………………………………… 47

12
APPENDIX G…………………………………………………………………………74

APPENDIX H………………………………………………………………….……….81

RESEARCHERS’ BACKGROUND…………………………………………………53

13
Abstract

This study is conducted to determine the biosorption capability of Shrimp

(Heterocarpus ensifer) Exoskeleton in accumulating lead and copper. This

investigatory is deemed to help in finding new ways to eliminate pollution

particularly in the contamination of water in the industries. Moreover it is

conducted to contribute new knowledge with regards to biosorption and to

explore a biosorbent material that has the capability to attract and sequester the

metals.

In the study, the exoskeleton were obtained from the market, rinsed with

distilled water and soaked at room temperature. The 12.57 g of copper (II) sulfate

anhydrous and 4.14 g of lead (II) nitrate was used to dissolve each to 1 L of

distilled water, to make the copper and lead solutions. The acetate buffer was

prepared using glacial acetic acid and the pH of the solution was adjusted using

sodium hydroxide solution. The Experimental groups were prepared by using 300

mg and 500 mg of biosorbent added to the copper and lead solutions. Three

trials were made for each amount of biosorbent and each solutions. The control

group was prepared by using the same concentration and amount of copper and

lead in the solutions but without the addition of biosorbent material. The initial

and final concentration of copper (mg/L) were determined using a

spectrophotometer.

14
Based on the results, the addition of 300 mg and 500 mg biosorbent

material to the copper solutions decreased the concentration of copper, 8,238.7

mg/L and 7,961.7 mg/L, respectively. The same thing happened to the lead in the

lead solutions by the addition of 300 mg and 500 mg biosorbent material

decreased the concentration to 221.7 mg/L and 162.7 mg/L. Moreover, addition

of 500 mg of the biosorbent yielded the lowest final concentration of copper and

lead. Hence, as the amount of biosorbent increases the concentration of copper

in the solution decreases. Also, the sorption percentage increases as the amount

of biosorbent added increases.

From the results and findings, the researcher found out that addition of

300 mg of the biosorbent material will result to the sorption percentage of 34.46

% and Qeq of 14.44 mg Cu/mg biosorbent while the addition of 500 mg

biosorbent material will yield a sorption percentage of 36.66 % and an Qeq of

9.217 mg Cu/mg of biosorbent. The same thing is concluded in the biosorption of

lead. The addition of 300 mg of the biosorbent material will result to the sorption

percentage of 91.44 % and Qeq of 7.894 mg Pb/ mg of biosorbent, and the

addition of 500 mg of the biosorbent material will result to the sorption

percentage of 93.72 % and Qeq of 4.855 mg Pb/ mg of biosorbent.

Moreover, there is a significant difference in the initial and final

concentration of copper and lead with and without the addition of biosorbent in

the copper and lead solutions. The final concentration of copper in the copper

solution added with 300 mg and 500 mg biosorbent decreased to 8,238.7 mg/L

and 7,961.7 mg/L, respectively starting with the initial concentration of 12,570

15
mg/L, and the final concentration of lead from the lead solutions also decreased

to 221.7 mg/L and 162.7 mg/L respectively, starting with the initial concentration

of 2,590 mg/L.

Also, there is a significant difference in the final concentration of copper

and lead by adding 300 mg and 500 mg of biosorbent. As the amount of

biosorbent increased, the final concentration of copper and lead also decreased.

Based on the statistical test employed, there is a significant difference between

the concentration of copper and lead in the following: solution without the

biosorbent and with 0.3 g biosorbent, solution without the biosorbent and with 0.5

g biosorbent, solutions with 0.3 g and 0.5 g biosorbent respectively.

Indeed, the exoskeleton of shrimp (Heterocarpus ensifer) can be used as

a biosorbent material for the removal of copper and lead.

16
Chapter I

THE PROBLEM AND ITS BACKGROUND

Introduction

Nowadays, different pollutants that are being released into the

environment has increased due to industrialization and therefore alarmed the

community. Heavy and toxic metals are most important because of their non-

biodegradability, with lead ions being the toxic and hazardous (Volesky 1990)

and has been classified in the USEPA’s Group B2 (probable human carcinogen)

Evangelou (1998).Thousands of lead are discharged from electric battery

manufacturing, lead smelting, internal combustion engines fuelled with leaded

petroleum, and mining activities (Cho and Kim 2003). Lead affects the human

central nervous system, the blood pressure and the reproduction (Gower 1993).

While copper being a widely used material causes serious toxicological concerns

(Nohoglu et al. 2002).

Biosorption is an economically feasible means for the removal and/or

recovery of heavy metals from industrial wastewaters (Volesky 1990). Biosorption

is the process that makes use of biological materials as adsorbents, and this

method has been studied by several researchers as an alternative technique to

conventional methods of heavy metal removal from wastewater (Jeon 2001, Sa

and Kutsal 2001, Volesky 2001, Yu 1999). Living or dead biomass can be used to

remove metals, nevertheless maintaining a living biomass during metal

17
biosorption is difficult because it requires a continuous supply of nutrients and

toxicity of metal for microorganism might take place. On the other hand, the use

of dead biomass can avoid these problems and the used cells can be easily

regenerated (Sudha and Abraham 2001, Yan and Viraraghavan 2001). The low

cost of biosorbents is a tangible advantage over other technologies, such as an

ion exchange and reverse osmosis (Bailey 1999). Compared this technology to

other standard methods, such as chemical precipitation and reverse osmosis, are

associated with high reagent consumption, high energy requirement and the

production of sludge containing metals, from which it is difficult to remove the

water and which also requires careful disposal (Aderhold 1996, Wilde and

Benemann 1993, Winter 1994). A variety of biomaterials such as bacteria, yeast,

algae and fungi have been successfully used as biosorbents for the removal of

heavy metals (Kapoor and Viraraghavan 1995, Volesky 1994) defined a low cost

sorbent as one that is abundant in nature, or is a by-product or waste material

from industries such as breweries and dairy products. The diffusion of the metal

from the bulk solution to active sites of biosorbents occurs by passive transport

mechanisms (Veglio and Beolchini 1997) and various functional groups such as

carboxyl, hydroxyl, amino and phosphate existing on the cell wall of biosorbents

can bind the heavy metals (Avery and Tobin 1993).

Chitin is a long-chain polymer of a N-acetyglucosamine a derivative of

glucose and is the main component of cell walls of the exoskeletons of

arthropods such as crustaceans like shrimp (Wagner 1994). It is also proven

useful for several medical and industrial purposes .Chitin is found out to be one

18
of the chemical groups that would attract and sequester the metals in biomass.

This research seeks the potential of chitin obtained from the exoskeleton of

shrimp as biosorbent for lead.

Background of the Study

In the present day, people are facing environmental crisis and one of these

is pollution: pollution in land, air and water. Overdoing of disposal of chemicals

and heavy metals from factories in industrial wastewaters can cause water

contamination. This concerns the people living near those sewage wastewaters.

Since heavy metals like lead and copper are harmful to human health and can be

associated with diseases in liver and kidney, gastrointestinal damage and mental

retardation in children. It is now time to research methods that would sequester

these metals before it will affect the health of several people.

Many different processes and technologies are being used to lessen the

contamination of these harmful substances in the wastewaters. Biosorption a

new technology that is being used today would likely be the most suitable

process that can sequester the metals since it uses the ability of biological

materials. The proponents prefer this method because of its effectiveness in

reducing the concentration of heavy metal ions to very low levels, the use of

inexpensive biosorbent materials, minimization of chemical and or biological

sludge, no additional nutrient requirement, and regeneration of biosorption and

possibility of metal recovery. It is also an effective alternative to physic-chemical

methods of separation in treating wastewater contaminated with heavy metal

19
ions. Extensive efforts have been made to explore new types of biosorbent

materials. Chitin is found out to be one of the chemical groups that would attract

and sequester the metals in biomass. It is mainly found in the exoskeleton of

arthropods such as crustaceans like shrimp. People usually just threw the shells

of shrimp after eating so it is good to make use of these to be a source of chitin

than just disposing it. In view of this the proponent believes that chitin obtained

from the exoskeleton of shrimp is effective as latent biosorbent for lead.

Conceptual Framework

Shrimp (Heterocarpus ensifer)

Exoskeleton Concentration of heavy


metals

Copper

Mass of Biosobent Lead

0.3 g

0.5 g

Fig 1. 1

In Fig 1.1 the research paradigm shows that the independent variable, which is

the varying amount of powdered shrimp exoskeleton, will affect the concentration

of Lead and Copper, in the lead nitrate and the copper sulfate solutions

respectively, which are the dependent variable under the study.

20
Statement of the Problem

This study aims to find out the biosorbent property of the exoskeleton of

shrimp in accumulating lead and copper from their solutions. Specifically the

investigatory project sought to answer the following questions.

1. Is there any significant difference in the concentration of copper and lead

without and with the application of biosorbent in the following solutions:

a. Pb(NO3)2 solution

b. CuSO4 solution

2. Is there a significant difference in the concentration of copper and lead

without and with the application of 0.3 g biosorbent.

3. Is there a significant difference in the concentration of copper and lead

without and with the application of 0.5 g biosorbent.

4. Is there a significant difference in the concentration of copper and lead

with the application of the following amounts of biosorbent:

a. 0.3 g

b. 0.5 g

Hypothesis

1. There is no significant difference in the concentration of copper and lead

without and with the application of biosorbent in the following solutions:

a. Pb(NO3)2 solution

b. CuSO4 solution

21
2. There is no significant difference in the concentration of copper and lead

without and with the application of 0.3 g biosorbent.

3. There is no significant difference in the concentration of copper and lead

without and with the application of 0.5 g biosorbent.

4. There is no significant difference in the concentration of copper and lead

with the application of the following amounts of biosorbent:

a. 0.3 g

b. 0.5 g

Significance of the Study

This investigatory project helps in finding new ways to eliminate pollution

particularly in the contamination of water in the industries. This can also help in

adding new knowledge with regards of biosorption and to explore a biosorbent

material that has the capability to attract and sequester the metals. This study

hopes to make contributions to the following:

Natural Environment

This study may help in our environment specifically in the industrial

wastewater by lessening the lead that contributes to contamination. The process

biosorption method used in the study is also eco-friendly since the study would

make use of biological materials that are not harmful to the environment. There is

also a possibility of metal recovery. This will also help in cleaning and avoiding

water pollution. Recovery of the deposited metals from saturated biosorbent can

be accomplished because they can often be easily released from the biosorbent

22
in a concentrated wash solution which also regenerates the biosorbent for

subsequent multiple reuse. Also it makes the process highly economical and

competitive particularly for environmental applications

Community

This finding of the study may aid people near those industrial sewage

areas in terms of their health since there is a lesser chance that people will get

different kinds of diseases in liver and kidney and mental retardation in children

from the lead contamination. The government can provide a colossal of

biosorbent material like this in each industrial wastewater since it is inexpensive.

Scope and Limitations of the Study

This study will be conducted for at least one week, including the testing of

the solutions in the Ultraviolet-visible Spectroscopy in Adamson University for the

comparing and analysis of the results.

The variables of the study specifically the lead and copper solutions and shrimp

will be obtained from Bambang and Balintawak wet market. The materials that

the researcher will use are six separate 50mL lead and copper solutions with

initial concentrations of roughly 8g Pb/L and 5g Cu/L will be prepared and

inoculated with shrimp shells. Glacial acetic acid will be used for the buffer and

0.1M NaOH will be used to adjust the buffer to the desired pH=4 ± 0.5. The

instruments will be used in the study were for filtration of the solutions and to

determine the amount of absorbed lead and copper. The sorption experiments

will be conducted in six 125mL Erlenmeyer flasks containing 100mL of the lead

23
nitrate solution and 100mL of the copper sulfate solution. And for the filtration

process a Büchner funnel, filter flask and a clamp. The determination of the

amount of copper and lead amount in the solutions will be the use of the

Ultraviolet-visible Spectroscopy.

Other properties of chitin and the metals are not being covered by the study.

Variation of contact time and other factors affecting the sorption of the metals are

not enclosed in the study. The proponent doesn’t have the control for the

possibility of metal recovery from the chitin.

Definition of Terms

The following terms are defined conceptually for the better understanding

in the study:

Biosorption – a new technology used in the study that uses chitin as biosorbent

material to remove heavy metals such as lead and copper from the industrial

wastewater.

Chitin –Obtained from the exoskeleton of shrimp that has a potential to attract

and sequester the metals in biomass.

Copper – the dependent variable in the study.

Heterocarpus ensifer – the independent variable in the study. Commonly known

as shrimp and where chitin was obtained from.

Lead – the dependent variable in the study.

24
Chapter 2

REVIEW OF RELATED LITERATURE AND STUDIES

Generally speaking, pollution in the environment has concerned itself of

how to stop and diminish the pollutants. Due to industrialization different

pollutants particularly heavy and toxic metals that are being released into the

environment has increased particularly in the industrial wastewaters. Using tools

and methods such as controlled laboratory experiments and the new technology

involving the removal of toxic metals from the wastewaters which is known as

biosorption, pollution can be reduced. Biosorption can be one solution to clean

up heavy metal contamination. Researchers have considered the advantages of

using this technology in sequestering the heavy metals.

Among the possible advantages of biosorption that is being low cost over

other technologies such as an ion exchange and reverse osmosis ,its high

efficiency to accumulate and sequester the metal ions, minimization of chemical

and or biological sludge, no additional nutrient requirement since it make use of

biological materials as adsorbents, regeneration of biosorbent and possibility of

metal recovery of the deposited metals from the saturated biosorbent because

they can often be easily released from the biosorbent in a concentrated wash

solution which also regenerates the biosorbent in a concentrated wash solution

which also regenerates the biosorbent for subsequent multiple reuse.(Kratochvil

and Volesky 1998). This makes the process highly economical and competitive

25
particularly for environmental applications in detoxifying effluents of metal-plating

and metal-finishing operations, mining and ore processing operations, battery

and accumulator manufacturing operations, thermal and nuclear power

regeneration. Therefore, new types of biomass have been searched and

modified physically, chemically and even biologically to improve the metal

sequestering performance.

A. Foreign Literature

I. Biosorption

Biosorption is the ability of biological materials to accumulate and sequester

heavy metals from industrial wastewater through metabolically mediated or

physico-chemical pathways of uptake (Fourest and Roux, 1992).

Research on biosorption is revealing that it is sometimes a complex occurrence

where the metallic species could be deposited in the solid biosorbent through

different sorption processes. In the case of precipitation, the metal uptake may

take place both in the solution and on the cell surface, It may be dependent on

the cell's' metabolism if, in the presence of toxic metals, the microorganism

produces compounds that favors the precipitation process. If it occurs after a

chemical interaction between the metal and cell surface, precipitation may not be

dependent on the cells' metabolism. (Ercole 1994)

26
In transport across cell membrane this kind of means is not related with

metabolic activity. Basically biosorption by living organisms consist of two steps.

First, a metabolism independent binding where the metals are bound to the cell

walls and second, metabolism dependent intracellular uptake, whereby metal

ions are transported across the cell membrane. Transport of the metal across the

cell membrane yields intracellular accumulation, which is dependent on the cell's

metabolism. The metal transport systems may become confused by the

presence of heavy metal ions of the same charge and ionic radius linked with

essential ions. (Costa, et.al., 1990, Ghourdon et.al., 1990, Huang et.al., 1990.,

Nourbaksh et.al., 1994).

While in physical adsorption takes place with the help of van der Waals'

forces that takes place through electrostatic interactions between the metal ions

in solutions and cell walls of microbial cells. (Kuyucak and Volesky 1990)

In ion exchange the cell walls of microorganisms enclose polysaccharides

and metal ions exchange with the counter ions of the polysaccharides. (Kuyucak

and Volesky 1988).

Whereas in complexation, after the interface between the metal and the

active groups, the metal removal from solution may also take place by complex

formation on the cell surface. (Aksu et al. 1992 )

27
Metals may be biosorbed or complexed by carboxyl groups originated in

microbial polysaccharides and other polymers. There are many factors that affect

the biosorption process. One of these is the the temperature that seems not to

influence the biosorption performances in the range of 20-35°C (Aksu et al.

1992). Another one is the pH that seems to be the most important factor in the

biosorptive process: it affects the solution chemistry of the metals, the activity of

the functional groups in the biomass and the competition of metallic ions (Friis

and Myers-Keith, 1986, Galun et al. 1987).Furthermore, Biomass concentration

in solution seems to manipulate the specific uptake: for lower values of biomass

concentrations there is an increase in the specific uptake (Fourest and Roux,

1992). Lastly biosorption is principally used to treat industrial wastewater where

more than one type of metal ions would be present; the removal of one metal ion

may be influenced by the presence of other metal ions. (Sakaguchi and

Nakajima, 1991). Some limitations in this process can be pointed out, such as

high cost, low efficiency, labour intensive operation, and lack of selectivity of the

precipitation process. In the comparison of ion exchange and biosorption

processes: The same equipment (i.e. pipes, columns, etc.) can be used with both

(a given treatment installation can be interchangeably used with both types of

sorbents),According to all estimates, biosorbents can be at least an order of

magnitude cheaper (1/10),Only a shorter life cycle can be assumed for

biosorbents. (Lee et al., 1998)

28
The mechanism of biosorption is complex, mainly ion exchange, chelation,

adsorption by physical forces, entrapment in inter and intrafibrilliar capillaries and

spaces of the structural polysaccharide network as a result of the concentration

gradient and diffusion through cell walls and membranes.

II. Biosorbent

There are several chemical groups that would attract and sequester the

metals in biomass: acetamido groups of chitin, structural polysaccharides of

fungi, amino and phosphate groups in nucleic acids, amido, amino, sulphydryl

and carboxyl groups in proteins, hydroxyls in polysaccharide and mainly

carboxyls and sulphates in polysaccharides of marine algae. (Ahalya,

Ramachandra and Kanamadi 2003). Bux and Kasan (1994) suggested that the

higher the biomass electronegativity, the greater the attraction and adsorption of

heavy metal cations.

Biosorbents must be hard enough to withstand the application pressures,

porous and/or “transparent” to metal ion sorbate species, and have high and fast

sorption uptake even after repeated regeneration cycles. Some of the biomass

types come as a waste by-product of large-scale industrial fermentations.

Biosorbents can accumulate the metal ions due to the metal binding capability of

various biological materials.

29
III. Chitin

Chitin (2-acetamido-2-deoxy-b-D-glucose–(N-acetylglucan), which is the

main structural component of mollusks, insects, crustaceans, fungi, algae and

marine invertebrates like crabs and shrimps. Since chitin contains positively

charged amine groups, it can be hypothesized that shrimp shells will be

particularly suitable for removing anionic metal complexes. (Chen and Chang,

1994;Ilyina et al.,1995

IV. Lead

According to Duffus, the term “heavy metal” is often used in the literature

as a group name for metals and semi-metals (metalloids) that have been

associated with contamination and potential toxicity. Lead because of its severe

toxicity, it is a major environmental concern (Volesky 1990) and has been

classified in the USEPA’s Group B2 (probable human carcinogen) (Evangelou

1998).The presence of lead in drinking water is known to cause serious health

problems including to death in cases of extreme exposure One of the health

problems are diseases in liver and kidney, gastrointestinal damage and mental

retardation in children. (Niu,1993).

Lead is a bluish-white lustrous metal. It is very soft, highly malleable,

ductile, and a relatively poor conductor of electricity. It is very resistant to

corrosion but tarnishes upon exposure to air.

30
Applications

Lead pipes bearing the insignia of Roman emperors, used as drains from the

baths, are still in service. Alloys include pewter and solder. Tetraethyl lead (PbEt4)

is still used in some grades of petrol (gasoline) but is being phased out on

environmental grounds.

Lead is a major constituent of the lead-acid battery used extensively in car

batteries. It is used as a coloring element in ceramic glazes, as projectiles, in

some candles to threat the wick. It is the traditional base metal for organ pipes,

and it is used as electrodes in the process of electrolysis. One if its major uses

are in the glass of computer and television screens, where it shields the viewer

from radiation. Other uses are in sheeting, cables, solders, lead crystal

glassware, ammunitions, and bearings and as weight in sport equipment.

Lead in the environment

Lead is usually found in ore with zinc, silver and copper. Lead occurs naturally in

the environment. However, most lead concentrations that are found in the

environment are a result of human activities. Due to the application of lead in

gasoline an unnatural lead-cycle has consisted. In car engines lead is burned, so

that lead salts (chlorines, bromines, and oxides) will originate.

These lead salts enter the environment through the exhausts of cars. The larger

particles will drop to the ground immediately and pollute soils or surface waters,

the smaller particles will travel long distances through air and remain in the

31
atmosphere. Part of this lead will fall back on earth when it is raining. This lead-

cycle caused by human production is much more extended than the natural lead-

cycle. It has caused lead pollution to be a worldwide issue.

Health effects of lead

32
Lead is a soft metal that has known many applications over the years. It

has been used widely since 5000 BC for application in metal products, cables

and pipelines, but also in paints and pesticides. Lead is one out of four metals

that have the most damaging effects on human health. It can enter the human

body through uptake of food (65%), water (20%) and air (15%). (Lenntech,

2009)

Foods such as fruit, vegetables, meats, grains, seafood, soft drinks and

wine may contain significant amounts of lead. Cigarette smoke also contains

small amounts of lead. (Lenntech, 2009)

Lead can enter (drinking) water through corrosion of pipes. This is more

likely to happen when the water is slightly acidic. That is why public water

treatment systems are now required to carry out pH-adjustments in water that will

serve drinking purposes. (Lenntech, 2009)

Lead can cause several unwanted effects, such as disruption of the

biosynthesis of hemoglobin and anemia, high blood pressure, kidney damage,

miscarriages and subtle abortions, disruption of nervous systems, brain damage,

declined fertility of men through sperm damage, diminished learning abilities of

children, behavioral disruptions of children, such as aggression, impulsive

behavior and hyperactivity. (Lenntech, 2009)

Not only leaded gasoline causes lead concentrations in the environment to

rise. Other human activities, such as fuel combustion, industrial processes and

solid waste combustion, also contribute. (Lenntech, 2009)

33
Moreover, lead is a particularly dangerous chemical, as it can accumulate

in individual organisms, but also in entire food chains. (Lenntech, 2009)

V. Copper

Copper is a reddish metal with a face-centered cubic crystalline structure.

It is malleable, ductile, and an extremely good conductor of both heat and

electricity. It is found in group IB of the periodic table. Copper has low chemical

reactivity. In moist air it slowly forms a greenish surface film called patina; this

coating protects the metal from further attack.

Copper may be found as a contaminant in food, especially shellfish, liver,

mushroom, nuts, and chocolate. Briefly, any processing or container using

copper material may contaminate the product, such as food, water or drink.

Copper is essential to human life and health but, like all heavy metals, is

potentially toxic as well (Nuhoglu et al. 2002).

Copper is a very common substance that occurs naturally in the

environment and spreads through the environment through natural phenomena.

Humans widely use copper. Copper is applied in the industries and in agriculture.

The production of copper has lifted over the last decades and due to this copper

quantities in the environment have expanded. .(Lenntech, 2009)

Copper can be found in many kinds of food, in drinking water and in air.

Because of that we absorb eminent quantities of copper each day by eating,

drinking and breathing. The absorption of copper is necessary, because copper is

a trace element that is essential for human health. Although humans can handle

34
proportionally large concentrations of copper, too much copper can still cause

eminent health problems. .(Lenntech, 2009)

People that live in houses that still have copper plumbing are exposed to

higher levels of copper than most people, because copper is released into their

drinking water through corrosion of pipes. .(Lenntech, 2009)

Health effects of copper

Long-term exposure to copper can cause irritation of the nose, mouth and

eyes and it causes headaches, stomachaches, dizziness, vomiting and diarrhea.

Intentionally high uptakes of copper may cause liver and kidney damage and

even death. Whether copper is carcinogenic has not been determined yet. .

(Lenntech, 2009)

Industrial exposure to copper fumes, dusts, or mists may result in metal

fume fever with atrophic changes in nasal mucous membranes. Chronic copper

poisoning results in Wilson’s Disease, characterized by a hepatic cirrhosis, brain

damage, demyelization, renal disease, and copper deposition in the cornea. .

(Lenntech, 2009)

When copper ends up in soil it strongly attaches to organic matter and minerals.

As a result it does not travel very far after release and it hardly ever enters

groundwater. In surface water copper can travel great distances, either

suspended on sludge particles or as free ions. .(Lenntech, 2009)

35
Copper does not break down in the environment and because of that it can

accumulate in plants and animals when it is found in soils. Copper can interrupt

the activity in soils, as it negatively influences the activity of microorganisms and

earthworms. The decomposition of organic matter may seriously slow down

because of this.(Lenntech, 2009)

Copper’s role in the body

Copper has a number of important functions in the human body. For

example, in bones and connective tissues, copper is required to fix calcium in the

bones and to build and repair all connective tissue. This includes the tendons,

ligaments, skin, hair, nails, arteries, veins and a few other tissues.Imbalances can

contribute to osteoporosis and bone spurs. (Wilson, 2010)

Copper is also needed in the final steps of the Krebs energy cycle called the

electron transport system. This is where most of our cellular energy is produced.

Any problem here causes fatigue, depression and other imbalances related to low

energy. (Wilson, 2010)

Copper must remain in balance with zinc. When imbalances occur, one is

more prone to all infections, in particular fungal and yeast infections that are so

common today. For example, most people have some intestinal yeast if they eat

sugars and most people have chronic sinus infections if they have common

symptoms such as post-nasal drip and others. (Wilson, 2010)

The thyroid gland is extremely sensitive to copper. In part this is due to its

nature and how easily it is influenced by the sympathetic nervous system.

36
Common conditions seen with copper imbalance include hypothyroidism and even

hyperthyroidism of a particular type that is very common that I all secondary

hyperthyroidism. Grave’s disease usually due to stress, copper imbalance and

often mercury as well. Anyone with a diagnosis of Grave’s disease or

hyperthyroidism should have a hair analysis performed at a lab that does not wash

the hair and properly interpreted. (Wilson, 2010)

Most often, the problem goes away with a properly designed nutritional

balancing program. Reducing all stress and balancing the body chemistry are both

required to resolve the condition naturally in my experience. Drugs may be needed

temporarily to control the symptoms. Surgery or radioactive iodine treatment and

too drastic and not needed, in my experience so far. (Wilson, 2010)

Copper is closely related to estrogen metabolism, and is required for

women's fertility and to maintain pregnancy. Imbalance can cause every

conceivable female organ-related difficulty such as premenstrual syndrome, ovarian

cysts, infertility, miscarriages, sexual dysfunctions and more. It affects men less

than women in this area, but it may affect men’s potency and sexual drive as well

as that of women. (Wilson, 2010)

Copper stimulates production of the neurotransmitters epinephrine,

norepinephrine and dopamine. It is also required for monoamine oxidase, an

enzyme related to serotonin production. (Wilson, 2010)

B. Foreign Studies

I. Biosorption

37
Biosorption is a property of certain types of inactive, dead, microbial

biomass to bind and concentrate heavy metals from even very dilute aqueous

solutions it can represent an attractive and cost-effective alternative for certain

types of industrial waste water treatment (Volesky, 1990). Furthermore,

biosorption processes do not generate the chemical sludge that is a major

problem in chemical precipitation methods for the removal of heavy metals

(Bailey 1999)In general, biomass considered could be an industrial waste or

naturally grown and collected and, therefore, cheaper than manmade ion

exchange resins. On the other hand, bioaccumulation is a metabolically

controlled process for the removal of metals by living organisms, which invariably

involves more complex cultivation and toxicity issues. (Zouboulis et al.,1997).

The use of biosorption technology to remove toxic heavy metal ions using

naturally abundant commodities such as seaweeds, especially brown algae, has

been greatly envisaged, showing high metal uptakes and selectivities (Davis et

al., 2003). Biosorption studies involving the removal of heavy metal ions have

most often made use of nitrate salts to prepare experimental solutions. These are

less likely to form complexes with metals that can thus be present as free metal

ions, simplifying the system studies. However, real industrial solutions may

contain anions other than nitrate, for example, chloride and sulphate. (Diniz et al.,

2001; Diniz and Volesky, 2005)

38
Some studies reported in the literature have examined the effect of anions

on the removal of metal ions from aqueous solutions. However, very little

attention has been paid to the effect of metal speciation on their biosorption

uptake in order to explain sometimes unexpected anomalies. (Ahuja et al.,

1999a, b; Kuyucak and Volesky, 1989; Palmieri et al., 2003; Pulsawat et al.,

2003).

Biosorption of metals is not based on only one mechanism. It consists of

several ones that quantitatively and qualitatively differ according to the type of

biomass, its origin and its processing. The biosorption process involves a solid

phase (sorbent or biosorbent; biological material) and a liquid phase (solvent,

normally water) containing a dissolved species to be sorbed (sorbate, metal ions)

(Ahalya, Ramachandra and Kanamadi 2003).Non-metabolism dependent

biosorption, metal uptake is by physico-chemical interaction between the metal

and the functional groups present on the microbial cell surface. This is based on

physical adsorption, ion exchange and chemical sorption, which is not dependent

on the cells' metabolism (Ahalya et.al.,2003).

Recovery of the deposited metals from saturated biosorbent can be

accomplished because they can often be easily released from the biosorbent in a

concentrated wash solution which also regenerates the biosorbent for

subsequent multiple reuse (Volesky 2007). It should be pointed out that there is a

potential added benefit of metal-recovery as an additional source of revenue

39
generated by a water treatment that must be carried out anyway (from a

regulatory and environmental point of view). This cost reduction applies to

“cheap” metal as well as “expensive” metal no matter what the economic indices

may be. The majority of biosorption systems are bench scale, treating very low

flows of acidic drainage. (Volesky and Naja 2005)

Sorption a chemical occurence is based on the Langmuir isotherm (114. It

is assumed that the forces exerted by chemically unsaturated surface atoms

(total number of binding sites) do not extend further than the diameter of one

sorbed molecule and that therefore sorption is restricted to a monolayer. In the

simplest case the following assumptions are made: all sorption sites are uniform,

there is only one sorbate, one sorbate molecule reacts with one active site, there

is no interaction between sorbed species. The rate of ad sorption is proportional

to the rate constant of the forward reaction, the number of free sites, and the

number of sorbate molecules hitting the surface per unit time.

When it comes to an ion exchange process, at least one ion from within

the molecular structure of the sorbent is exchanged for another one coming from

outside. This leads to ever-changing conditions in the sorption system due to the

stream of ‘exchanged’ ions also leaving the sorbent into the liquid environment.

That is until the sorption equilibrium is established. (Naja, Volesky and Murphy

2009).

40
There are three basic types of sorption solid/liquid contact systems: the

packed-bed column (and modifications), the fluidized-bed system and the

completely mixed system, often combined with a solid concentration operation.

II.Biosorbent

Biomass can come from: fast-growing organisms that are specifically

cultivated or propagated for biosorption purposes (crab shells, seaweeds).

(Ahalya, Ramachandra and Kanamadi 2003).Biomass exhibits a property, acting

just as a chemical substance, as an ion exchanger of biological origin (Yun, Park

and Volesky 2001). The quality of the sorbent material is judged according to

how much sorbate it can attract and retain in an ‘immobilized’ form. For this

purpose it is customary to determine the metal uptake by the biosorbent as the

amount of sorbate bound by the unit of solid phase (by weight, volume, etc.).

Correspondingly, the amount of metal bound by the sorbent which ‘disappeared’

from the solution can be calculated based on the mass balance for the sorbent in

the system

Biosorbents can accumulate in excess of 25% of their dry weight in

deposited heavy metals: Pb, Cd, U, Cu, Zn, even Cr and others. Due to higher

affinity of the sorbent for the sorbate species, the latter is attracted and bound

there by different mechanisms. The process continues till equilibrium is

established between the amount of solid-bound sorbate species and its portion

remaining in the solution. (Volesky 2000).

41
While there is a preponderance of solute (sorbate) molecules in the

solution, there are none in the sorbent particle to start with. This imbalance

between the two environments creates a driving force for the solute species. The

heavy metals adsorb on the surface of biomass thus, the biosorbent becomes

enriched with metal ions in the sorbate. The degree of sorbent affinity for the

sorbate determines its distribution between the solid and liquid phases.

The importance of any given group for biosorption of a certain metal by a

certain biomass depends on such factors as the number of sites in the biosorbent

material, the accessibility of the sites, the chemical state of the sites (i.e.,

availability), and the affinity between the site and the metal (i.e., binding

strength). Both chemical pretreatments, such as contacting cells with acids,

alkali, and organic compounds and physical pretreatments, such as heat

treatment, autoclaving, freeze drying, and boiling and showed enhancement in

metal biosorption by microorganisms.

III. Chitin

Chitin is the world’s second most abundant naturally occurring

polysaccharide. Much of it is disposed of as waste from seafood crustaceans,

mainly crabs, shrimps, prawns, and lobsters (Niu and Volesky 2007).

Chitin/chitosan, which is produced industrially from materials such as crab shells,

can be effective at binding heavy metals (Guibal 1999, Mcafee 2001, Navarro

42
2000). Similarly to special anionic exchange resins, biosorbents that are capable

of sequestering anions contain chemical active groups within their structures. It is

often the NH2 group that is active and ubiquitous in fungal cell walls as well as in

the chitin/chitosan components of crustacean exoskeletons (Niu and Volesky,

2006).

IV. Lead

Out of 106 identified elements, about 80 of them are called metals. The

chemical speciation of a metal depends upon its oxidation state and its

interactions with other components in the system as well as on other parameters

including pH, redox potential, ionic strength, salinity and alkalinity. These metallic

elements can be divided into two groups: those that are essential for survival,

such as iron and calcium, and those that are nonessential or toxic, such as

cadmium and lead.

Depending on the angle of interest and the environmental impacts, metals

can be divided into four major categories: (1) Toxic heavy metals, (2) strategic

metals, (3) precious metals, and (4) radionuclides (51).The classification of

metals for biosorption applications. In terms of environmental threat it is mainly

categories 1 and 4 that are of interest for removal from the environment.

Heavy metal pollution in the aquatic system has become a serious threat

today and of great environmental concern as they are non-biodegradable and

43
thus persistent. Heavy metals form positive ions in solution and they have a

density five times greater than that of water. Further work with biosorbents

identified for their high metal uptake could best be directed to derivation of

engineering process scale-up parameters for application in the clean-up of two

most ubiquitous types of metal-contaminated industrial effluents: Acid Mine

Drainage (AMD) and electroplating effluents (Macaskie, 1997; Tabak et al.,

2003).

Main sources of heavy metal contamination include urban industrial

aerosols, solid wastes from animals, mining activities, industrial and agricultural

chemicals. Heavy metals also enter the water supply from industrial and

consumer water or even from acid rain which breaks down soils and rocks,

releasing heavy metals into streams, lakes and ground water. The metals

increase in concentration at every level of food chain and are passed onto the

next higher level–a phenomenon called bio-magnification (Paknikar et al., 2003).

It has been established that dissolved metals (particularly heavy metals)

escaping into the environment pose a serious health hazard. They accumulate in

living tissues throughout the food chain, which has humans at its top, multiplying

the danger. Lead is a cumulative general poison, with fetuses, infants, children

up to six years of age, and pregnant women (because of their fetuses) being

most susceptible to adverse health effects. Overt signs of acute intoxication

include dullness, restlessness, irritability, poor attention span, headaches, muscle

44
tremor, hallucinations, and loss of memory, with encephalopathy occurring at

blood lead levels of 100–120 μg dL-1 in adults and 80–100 μg dL-1 in children.

The toxicity of metal ion is owing to their ability to bind with protein molecules and

prevent replication of DNA and thus subsequent cell division (Kar et al.,

1992).Lead affects the human central nervous system, the blood pressure and

the reproduction. (Gower 1993).

Imbalances or excessive amounts of a metal species along this lead to

toxicity symptoms, disorders in the cellular functions, long-term debilitating

disabilities in humans, and eventually death. One of the most insidious effects of

inorganic lead is its ability to replace calcium in bones and remain there to form a

semi permanent reservoir for long-term release well after the initial absorption.

Such on-site treatment of waste streams contributing a load of heavy

metals would be more efficient than treating the large volumes of mixed, diluted

metal-contaminated waste water in a general (municipal) sewage treatment plant

V. Copper

Copper, one of the most widely used heavy metals, is mainly employed in

electrical and electroplating industries and in larger amounts and is extremely

toxic to living organisms. The presence of copper (II) ions, cause serious

toxicological concerns, it is usually known to deposit in brain, skin, liver, pancreas

and myocardium (Davis et al., 2000).

Copper may be found as a contaminant in food, especially shellfish, liver,

mushroom, nuts, and chocolate. Briefly, any processing or container using

45
copper material may contaminate the product, such as food, water or drink.

Copper is essential to human life and health but, like all heavy metals, is

potentially toxic as well (Nuhoglu et al. 2002).

B. Local Literature

Successive treatment of loaded biomass can enable recovery of valuable

element or further containment of highly toxic species. Heavy metals from liquid

wastes and results in safe environmental discharge (Mamaril, Foser and

Alphante 1997). Biosorbents are one material that binds with metals (Mojica and

Palafox 2004).

D .Local Studies

Biosorption is at industrial interest because of the removal of toxic heavy

metals from liquid wastes. Chitin is a polymer of N-acetyl B.D. glucosamine.

Presence of the amine and hydroxyl groups in chitin serve as binding sites for

lead ions. (Mojica and Palafox 2004).

46
Chapter 3

RESEARCH METHODOLGY

This chapter presents the research procedures that will be employed. The

research design, product, formation, treatment, sampling, instrumentation,

procedure in gathering data and statistical tool.

Research Design

For the study, the researchers will use the control group design because

this study uses only two groups consisting of the experimental and control

groups.

Preparation of Biosorbent and the Copper(II) sulfate and Lead (II) nitrate
Solutions

The exoskeleton of shrimp (Heterocarpus ensifer), used as biosorbent in

this study, were collected from the market. The biosorbent were soaked in

distilled water for 24 hours and rinsed three times with distilled water. The

biosorbent was transferred to a container, air dried for two days and stored at

room temperature. The initial concentration of copper and lead are of 5g/L, and

5g/L respectively. The entire one liter of Cu and Pb stock solutions were

distributed into 18 different Erlenmeyer flasks, 100mL of Cu stock solution in nine

different flasks, and 100 mL Pb stock solution in nine different flasks and stored

at room temperature. Glacial acetic acid was diluted to make a 0.5M acetate

buffer with the pH=3.9, and 0.1M NaOH was used to adjust the buffer to the

47
desired pH=4 ± 0.5. The acetate buffer was used to maintain the acidity of the Cu

and Pb solutions at the pH=4 ± 0.5. Two milliliters of the acetate buffer was

added to each sample before the biosorbent was added. The pH of the solutions

were determined using a data logger.

Varying Biosorbent to Copper and Lead Ratios

All samples were treated at room temperature, and with pH=4 ± 0.5. The

initial and final copper concentrations [Cu] and lead concentrations [Pb], in units

of milligrams of Cu and Pb per liter of solution, were determined by

Spectrophotometer.

The initial [Cu] and [Pb] of each sample was determined. Different

amounts of biosorbent (0.3g, and 0.5g) were added to 12 separate flasks

containing Cu and Pb stock solution with constant initial [Cu] and [Pb]. Different

amounts of biosorbent represent different biosorbent to Cu ratio (BS:Cu) (Table

1) and Pb ratio (BS:Pb). BS:Cu and BS:Pb were calculated using the equation,

BS: Cu = mass of biosorbent = mass of biosorbent


mass of copper initial [Cu] x volume of sample
BS: Pb = mass of biosorbent = mass of biosorbent
mass of lead initial [Pb] x volume of sample

The samples were shaken for two hours and were stored at room

temperature for 24 hours. After the treatment, the biosorbent was filtered out

using filter paper. Although no biosorbent was added to the untreated sample, it

was filtered before the determination of the final [Cu] and [Pb] to control for any

copper and lead sorption caused by the filtering processes. The filtrate was

48
collected, then the final [Cu] and [Pb] were determined. The calculated initial and

final [Cu] and [Pb] were used to determine the specific uptake (Qeq), in units of

milligram of Cu removed per milligram of biosorbent used, and the sorption

percentage (%S). Qeq and %S were calculated using the equations,

Qeq = (initial [Cu/Pb] – final [Cu/Pb]) x volume of sample


mass of biosorbent

% S = (initial [Cu/Pb] – final [Cu/Pb]) x 100


Initial [Cu/Pb]

Statistical Tool

The study will use the t-test as its statistical tool. The t-test will be used by

the researcher to compare the difference of the mean of the populations of each

trial the researcher made. The t-test will determine the differences in the

concentration of copper and lead in the control and experimental groups.

49
Figure 2: Schematic diagram

Collection of Shrimp Exoskeleton


PHASE I:

Preparation of Soaking in distilled H2O for 24 hours and rinsing three times
the Biosorbent

Air drying for two days and storing at room temperature

Adding 31.57 g of copper (II) sulfate anhydrate in 1L distilled H2O


with initial [Cu], 5g/L
PHASE II:

Preparation of Distribution of stock solution 100 mL stock solution in 9 different


the Copper Erlenmeyer flasks at room temperature.
and Lead
Solutions
Addition of 2 mL acetate buffer to each of the Erlenmeyer flasks.
Using a data logger to determine the pHn of the solution

Determination of initial copper concentration using


spectrophotometer

Addition of different amounts of biosorbent (0.3 g, and 0.5 g) to 9


separate flasks containing Cu stock solution with constant initial
PHASE III: [Cu] and to 9 separate flasks containing Pb stock solution with
constant initial [Pb].
Varying Shaking the samples for two hours and storing for 24 hours.
biosorbent to Filtering the biosorbent using filter paper.
copper ratios

Determination of final copper and lead concentration using


spectrophotometer

PHASE IV: Using a statistical tool to determine the significant difference in the
concentration of copper with and without the addition of
Statistical test biosorbents in copper(II)sulfate solution and with varying amounts.

50
Chapter 4

PRESENTATION, ANALYSIS, AND INTERPRETATION OF DATA

Result

Table 1. Varying Amount of biosorbent (mg), and the BS:Cu

Amount of Biosorbent (mg) BS: Cu

0.00 0.00

300 0.02387

500 0.03978

Table 1 shows the amount of biosorbent used (300 and 500) mg and the BS: Cu.

The values for the BS: Cu were computed using the formula in chapter 3.

0.05

0.04

0.03 BS:Cu vs
Amount of
0.02 Biosorbent

0.01

0
0g 0.3 g 0.5 g

Figure ___. Relationship of BS: Cu and the amount of biosorbent

The Line Graph shows the relationship between the BS:Cu and the amount of

the biosorbent. If the amount of Biosorbent increases, the BS:Cu also increases.

51
Table 2. Varying Amount of biosorbent (mg), and the BS:Pb

Amount of Biosorbent (mg) BS: Pb

0.00 0.00

300 0.1158

500 0.1930

Table 2 shows the amount of biosorbent used (300 and 500) mg and the BS: Pb.

The values for the BS: Pb were computed using the formula in Chapter 3.

0.2

0.15

0.1 BS:Pb vs
Amount of
Biosorbent
0.05

0
0g 0.3 g 0.5 g

Figure ___. Relationship of BS: Pb and the amount of biosorbent

The Line Graph shows the relationship between the BS:Pb and the amount of the

biosorbent. If the amount of Biosorbent increases, the BS:Pb also increases.

Table 3. Initial and Final Concentration of Copper(mg/L) With and Without the
Addition of Biosorbent

Amount of Biosorbent (mg) Initial [Cu] Final [Cu]

52
0.00 12570 12570
300 12570 8238.7
500 12570 7961.7

14000
12000
10000
8000 Initial Cu
6000 Final Cu

4000
2000
0
0 300 500

Figure ___. The figure shows the difference of the initial and the final

concentration of Cu in the solutions.

Table 4. Initial and Final Concentration of Lead (mg/L) With and Without the
Addition of Biosorbent

Amount of Biosorbent (mg) Initial [Pb] Final [Pb]

0.00 2590 2590

300 2590 221.7

500 2590 162.7

53
3000

2500

2000
Initial Pb
1500
Final Pb
1000

500

0
0 300 500

Figure ___. The figure shows the difference of the initial and the final

concentration of Pb in the solutions.

Tables 3 and 4 show the initial and final concentration of copper and lead (mg/L)

without and with the application of varying amount of biosorbent in their solutions.

Table 5 Varying BS:Cu. Amount of biosorbent (g), BS:Cu, Sorption percentage


(%S), Qeq specific uptake (mgCu removed / mg biosorbent).

Amount of Biosorbent (mg) BS:Cu %S Qeq

0.00 0.000 0.00 0.00

300 0.02387 34.46 14.44

500 0.03978 36.66 9.217

Table 5 shows the sorption percentage (%S) and the specific uptake (Qeq)of

without and with the addition of varying amounts of biosorbent in the copper(II)

sulfate solutions.

54
16 40

14 35

12 30

10 25

8 20

6 15

4 10

2 5

0 0
0.00 g 0.3 g 0.5 g
amount of biosorbent

BS:Cu Qeq %S

Figure ___ shows the relationship between the amount of Biosorbent, BS:Cu,

Percentage sorption(%S) and the Specific uptake (Qeq) of the Cu metal.

Table 6 Varying BS:Pb. Amount of biosorbent (g), BS:Pb, Sorption percentage


(%S), Qeq specific uptake (mg Pb removed / mg biosorbent).

Amount of Biosorbent (mg) BS:Pb %S Qeq

0.00 0.00 0.00 0.000

0.30 0.1158 91.44 7.894

0.50 0.1930 93.72 4.855

55
Table 6 shows the sorption percentage (%S) and the specific uptake (Qeq)of

without and with the addition of varying amounts of biosorbent in the lead (II)

nitrate solutions.

9 100

8 90

80
7
70
6
60
5
50
4
40
3
30
2
20

1 10

0 0
0.00 g 0.3 g 0.5 g
amount of biosorbent

BS:Pb Qeq %S

Figure ___ shows the relationship between the amount of Biosorbent, BS:Pb,

Percentage sorption(%S) and the Specific uptake (Qeq) of the Pb metal.

Statistical Results

Table 7: Comparison of [Cu] using 0.00 g and 0.3 g biosorbent using T-test

Group Amount of BS (g) [Cu]mg/L Tcomp vs Tcrit Decision

Experimental 0.00 12,570 3.873 > 2.015 Reject Null Hypothesis

Experimental 0.30 8, 238.7

56
Table 8: Comparison of [Cu] using 0.00 g and 0.5 g biosorbent using T-test

Group Amount of BS (g) [Cu]mg/L Tcomp vs Tcrit Decision

Experimental 0.00 12,570 3.873 > 2.015 Reject Null Hypothesis

Experimental 0.50 7, 961.7

Table 9: Comparison of [Cu] using 0.30 g and 0.50 g biosorbent using T-test

Group Amount of BS (g) [Cu]mg/L Tcomp vs Tcrit Decision

Experimental 0.30 8, 238.7 124.99 > 2.015 Reject Null Hypothesis

Experimental 0.50 7,961.7

Table 10: Comparison of [Pb] using 0.00 g and 0.30 g biosorbent using T-test

Group Amount of BS (g) [Pb]mg/L Tcomp vs Tcrit Decision

Experimental 0.00 2, 590 3.873 > 2.015 Reject Null Hypothesis

Experimental 0.30 221.7

Table 11: Comparison of [Pb] using 0.00 g and 0.50 g biosorbent using T-test

Group Amount of BS (g) [Pb]mg/L Tcomp vs Tcrit Decision

Experimental 0.00 2,590 3.873 > 2.015 Reject Null Hypothesis

Experimental 0.50 162.7

Table 12: Comparison of [Pb] using 0.30 g and 0.50 g biosorbent using T-test

Group Amount of BS (g) [Pb]mg/L Tcomp vs Tcrit Decision

Experimental 0.30 221.7 314.67 > 2.015 Reject Null Hypothesis

Experimental 0.50 162.7

57
Discussion

Based on Tables 3 and 4, the final concentration of copper decreased

upon the addition of varying amounts of biosorbent in the copper (II) sulfate and

lead (II) nitrate solutions. The result indicates that higher amount of biosorbent

lowers the concentration of copper and lead in there solutions.

Moreover, Tables 5 and 6 showed that as the amount of biosorbent added

increased, the sorption percentage (%S) also increased in both the copper and

lead stock solutions. This indicates that higher amount of biosorbent results to

higher sorption percentage (%S).

On the other hand, the specific uptake (Qeq) of copper with the addition of

300 mg and 500 mg of biosorbent yielded, 14.44 and 9.217 mg Cu/mg

biosorbent, respectively. The specific uptake (Qeq) of lead with the addition of

300 mg and 500 mg of biosorbent yielded, 7.894 and 4.855 mg Pb/mg

biosorbent, respectively. Hence, it shows that the biosorbent, exoskeleton of

shrimp (Heterocarpus ensifer) can accumulate heavy metals, such as copper and

lead.

The results obtained were supported by the findings of Guibal (1999),

Mcafee (2001) and Navarro (2000), that chitin/chitosan, which is produced

industrially from materials such as crab shells, can be effective at binding heavy

metals. Moreover, according to Niu and Volesky (2006), chitin/chitosan which is a

component of crustacean exoskeleton, function similarly to special anionic

exchange resins, in sequestering anions. This component contains chemical

active groups within their structures and it is often the NH2 group that is active.

58
Thus, it is possible that this amino group is responsible in accumulating the

heavy metals such as copper and lead.

On the other hand, based on the statistical test employed, there is a

significant difference between the concentration of copper in the copper (II)

solution without the biosorbent and the copper solution with 0.3 g biosorbent.

Likewise, there is a significant difference between the concentration of copper in

the copper (II) solution without the biosorbent and the copper solution with 0.5 g

biosorbent. Also, , there is a significant difference between the concentration of

copper in the copper (II) solution with 0.3 gbiosorbent and the copper solution

with 0.5 g biosorbent Hence, the concentration of copper in the copper (II)

solutions with the addition of 0.30 g And 0.50 g biosorbent decreased the

concentration of copper.

The same results were obtained for lead, there is a significant difference

between the concentration of lead in the lead (II) nitrate solution without the

biosorbent and the lead solution with 0.3 g biosorbent. Likewise, there is a

significant difference between the concentration of lead in the lead solution

without the biosorbent and the lead solution with 0.5 g biosorbent. Also, , there is

a significant difference between the concentration of lead in the lead solution with

0.3 g biosorbent and the lead solution with 0.5 g biosorbent Hence, the

concentration of lead in the lead solutions with the addition of 0.30 g And 0.50 g

biosorbent decreased the concentration of lead.

59
Chapter 5

SUMMARY OF FINDINGS, CONCLUSIONS AND RECOMMENDATIONS

Summary

The study was focused mainly in determining the biosorbent capability of

shrimp (Heterocarpus ensifer) exoskeleton.

The exoskeleton were obtained from the market, rinsed with distilled water

and soaked at room temperature. The 12.57 g of copper (II) sulfate anhydrous

and 4.14 g of lead (II) nitrate was used to dissolve each to 1 L of distilled water,

to make the copper and lead solutions. The acetate buffer was prepared using

glacial acetic acid and the pH of the solution was adjusted using sodium

hydroxide solution. The Experimental groups were prepared by using 300 mg

and 500 mg of biosorbent added to the copper and lead solutions. Three trials

were made for each amount of biosorbent and each solutions. The control group

was prepared by using the same concentration and amount of copper and lead in

the solutions but without the addition of biosorbent material. The initial and final

concentration of copper (mg/L) were determined using a spectrophotometer.

Based on the results, the addition of 300 mg and 500 mg biosorbent

material to the copper solutions decreased the concentration of copper, 8,238.7

mg/L and 7,961.7 mg/L, respectively. The same thing happened to the lead in the

lead solutions by the addition of 300 mg and 500 mg biosorbent material

decreased the concentration to 221.7 mg/L and 162.7 mg/L. Moreover, addition

of 500 mg of the biosorbent yielded the lowest final concentration of copper and

60
lead. Hence, as the amount of biosorbent increases the concentration of copper

in the solution decreases. Also, the sorption percentage increases as the amount

of biosorbent added increases.

Based on the statistical test employed, there is a significant difference

between the concentration of copper and lead in the following: solution without

the biosorbent and with 0.3 g biosorbent, solution without the biosorbent and with

0.5 g biosorbent, solutions with 0.3 g and 0.5 g biosorbent respectively.

Indeed, the exoskeleton of shrimp (Heterocarpus ensifer) can be used as

a biosorbent material for the removal of copper and lead.

Conclusions

From the results and findings, the researcher found out that addition of

300 mg of the biosorbent material will result to the sorption percentage of 34.46

% and Qeq of 14.44 mg Cu/mg biosorbent while the addition of 500 mg

biosorbent material will yield a sorption percentage of 36.66 % and an Qeq of

9.217 mg Cu/mg of biosorbent. The same thing is concluded in the biosorption of

lead. The addition of 300 mg of the biosorbent material will result to the sorption

percentage of 91.44 % and Qeq of 7.894 mg Pb/ mg of biosorbent, and the

addition of 500 mg of the biosorbent material will result to the sorption

percentage of 93.72 % and Qeq of 4.855 mg Pb/ mg of biosorbent.

Moreover, there is a significant difference in the initial and final

concentration of copper and lead with and without the addition of biosorbent in

the copper and lead solutions. The final concentration of copper in the copper

61
solution added with 300 mg and 500 mg biosorbent decreased to 8,238.7 mg/L

and 7,961.7 mg/L, respectively starting with the initial concentration of 12,570

mg/L, and the final concentration of lead from the lead solutions also decreased

to 221.7 mg/L and 162.7 mg/L respectively, starting with the initial concentration

of 2,590 mg/L.

Also, there is a significant difference in the final concentration of copper

and lead by adding 300 mg and 500 mg of biosorbent. As the amount of

biosorbent increased, the final concentration of copper and lead also decreased.

Therefore, the researchers concluded that the exoskeleton of shrimp

(Heterocarpus ensifer) can be used as biosorbent material in removing copper

and lead in water.

Recommendations

The researchers would like to recommend some points for the further

improvement of the study:

a) Application of Shrimp exoskeleton as a biosorbent in Cu removal in other

conditions such as varying pH and temperature in the treatment.

b) Variation and comparison of more than one heavy metals.

c) A variation of contact time but a constant amount of biosorbent.

d) Testing of other factors that affect the biosorption process like metal

binding capacity, biosorbent affinity and biomass electronegativity.

62
Bibliography

Books

• Aldor, I., Fourest, E., Volesky, B., & Can, J., (1995). Chem. Eng. Montreal.

• Davis, T.A., Mucci, A., & Volesky, B. (2003). A review of the biochemistry

of heavy metal biosorption by brown algae. Montreal.

• Fourest, E., & Volesky, B. (1996). Environ. Sci. Technol. Montreal.

• Guibal, E., Roulph, C., & Le Cloirec, P. (1992 ). Water Res.

• Hu, M.Z.-C., Norman, J.M., Faison, N.B., & Reeves, M. (1996) Biotechnol.

Bioeng.

• Kratochvil, D., & Volesky, B. (1998). Removal of trivalent and hexavalent

chromium by seaweed biosorbent. Montreal.

• Kuyucak, N., & Volesky, B. (1990). In Biosorption of Heavy Metals.

Montreal.

• Mullen, M.D., Wolf, D.C., Beveridge, T.J., & Bailey, G.W. (1990) Sorption

of heavy metals by soil fungi Aspergillus niger and Mucor Rouxii

• Naja, G.M., & Volesky, B. (2006). Multi-metal biosorption in a fixed- bed

flow-through column. Montreal.

• Niu, C.H., & Volesky, B. (2006). Biosorption of chromate and vanadate

with waste crab shells. New York.

• Volesky. B. (2003). Sorption and Biosorption.

• Volesky, B. (2007). Biosorption and me. Montreal.

• Volesky, B., & Holan, Z.R. (1995). Biotechnol. Prog.

63
• Volesky, B., Yang, J., & Niu, H. (2001). Biosorption of metal cations and

anions. In: Ion Exchange and Solvent Extraction (Vol.14, pp. 119-

168).New York

• Volesky, B., Weber, J., & Park, J. M. (2003). Continuous-flow metal

biosorption in a regenerable Sargassum column

Journal

• Ahalya, N., Ramachandra, T.V., & Kanamadi, R.D. (2003). Biosorption of

Heavy Metals. Research Journal Of Chemistry And Environment, 7, 4.

Internet

• Copper - Cu. (1998-2009) Lenntech BV. Retrieved August 28 ,


2010, from
http://www.lenntech.com/periodic/elements/cu.htm#ixzz0xuL1RCXp

• Lead – Pb. (1998-2009) Lenntech BV. Retrieved August 28 ,2010,


from http://www.lenntech.com/periodic/elements/pb.htm

• Parvathi, K., Nagendran, R., & Naresh, R. (2007). Lead biosorption onto

waste beer yeast by-product, a means to decontaminate effluent

generated from battery manufacturing industry [Electronic version].

Journal of Biotechnology. Retrieved January 15, 2007, from

http://www.ejbiotechnology.info/content/vol10/issue1/full/13/index.html

• Volesky, B. (2010) bv-Publications. Retrieved January 2010, from

http://biosorption.mcgill.ca/

• Volesky, B. (2009, March 31). Biosorption. SciTopics. Retrieved July 19,

2010, from http://www.scitopics.com/Biosorption.html

64
• Wilson, L. (2010). COPPER TOXICITY SYNDROME. Retrieved August
28, 2010 from
http://www.drlwilson.com/articles/copper_toxicity_syndrome.htm

65
APPENDIX A

Gantt Chart

July July August August August August August


Events
20-25 26-30 1-5 6-10 11-15 16-20 21-25

Working on Chapter 1-3


Gathering Information
and Details
Solution preparation and
biosorbent preparation
Preparation of Buffer
Solutions
for initial of pH level
Inoculation of Biosorbent
in the diluted solutions
Filtration of the diluted
Solutions
Testing of samples using
the Spectrophotometer
Gathering the data
after the experiment

Working on Chapter 4-5

Statistical Analysis

Finalization of Research
Paper

66
APPENDIX B

Financial Report

Material Cost
Shrimp ₱305
Lead Nitrate ₱1800
Glacial Acetic Acid ₱120
Filter Paper ₱75
Sample Test (Spectrophotometer) ₱3200
Total cost ₱5500

67
APPENDIX C

68
69
APPENDIX D

Biosorbent to Copper/Lead ratio

BS: Cu = mass of biosorbent = mass of biosorbent


mass of copper initial [Cu] x volume of sample

BS:Cu of the control set-up

BS:Cu= __ _ 0mg
12,570mg/L x 1L

=___0mg
12,570 mg

= 0.00

BS:Cu of 300 mg set-up

BS:Cu=___300mg___
12,570mg/L x 1L

=__300mg__
12,570 mg

= 0.02387

BS:Cu of 500 mg set-up

BS:Cu=___500mg____
12,570mg/L x 1L

=__500mg__
12,570 mg

=0.03978

70
BS: Pb = mass of biosorbent = mass of biosorbent
mass of lead initial [Pb] x volume of sample

BS:Pb of the control set-up

BS:Pb=____00mg___
2,590mg/L x 1L

=__00mg_
2,590mg

= 0.00

BS:Pb of 300 mg set-up

BS:Pb=____300mg___
2,590mg/L x 1L

=__300mg_
2,590mg

= 0.1158

BS:Pb of 500 mg set-up

BS:Pb=____500mg___
2,590mg/L x 1L

=__500mg_
2,590mg

= 0.1930

71
APPENDIX E

Sorption Percentage of Copper/Lead

% S = (initial [Cu] – final [Cu]) x 100


Initial [Cu]

% S of the control set-up (Cu)

% S = (12,570 mg/L- 12,570 mg/L) x 100 %


12,570 mg/L

% S = __0 mg/L__ x 100%


12,570 mg/L

% S = 0.00 x 100%

%S=0%

% S of the 300 mg set-up (Cu)

% S = (12,570 mg/L- 8,238.7 mg/L) x 100 %


12,570 mg/L

% S = 1,881.3 mg/L x 100%


12,570 mg/L

% S = 0.34457 x 100%

% S = 34.46 %

% S of the 500 mg set-up (Cu)

% S = (12,570 mg/L- 7,961.7 mg/L) x 100 %


12,570 mg/L

% S = 4,608.3 mg/L x 100%


12,570 mg/L

% S = 0.36661 x 100%

% S = 36.66 %

72
% S = (initial [Pb] – final [Pb]) x 100
Initial [Pb]

% S of the control set-up (Pb)

% S = (2,590 mg/L- 2,590 mg/L) x 100 %


2,590 mg/L

% S = __0 mg/L__ x 100%


2,590 mg/L

% S = 0.00 x 100%

%S=0%

% S of the 300 mg set-up (Pb)

% S = (2,590 mg/L- 221.7 mg/L) x 100 %


2,590 mg/L

% S = _2,368.3 mg/L__ x 100%


2,590 mg/L

% S = 0.91440 x 100%

% S = 91.44 %

% S of the 500 mg set-up (Pb)

% S = (2,590 mg/L- 162.7 mg/L) x 100 %


2,590 mg/L

% S = _2,427.3 mg/L__ x 100%


2,590 mg/L

% S = 0.93718 x 100%

% S = 93.72 %

73
APPENDIX F

Specific uptake (Qeq) of the Copper/Lead

Qeq = (initial [Cu] – final [Cu]) x volume of sample


mass of biosorbent

Qeq of the control set-up (Cu)

Qeq = (12,570 mg/L – 12,570 mg/L) x 1L


0mg

= 0 mg Cu
0 mg of biosorbent

= n/a

Qeq of the 300mg set-up (Cu)

Qeq = (12,570 mg/L – 8,238.7 mg/L) x 1L


300mg

= 4,331.3 mg Cu
300 mg of biosorbent

= 14.438 mg Cu/mg of biosorbent

Qeq of the 500 mg set-up (Cu)

Qeq = (12,570 mg/L – 7,961.7 mg/L) x 1L


500mg

= 4608.3 mg Cu
500 mg of biosorbent

= 9.217 mg Cu/mg of biosorbent

74
Qeq = (initial [Pb] – final [Pb]) x volume of sample
mass of biosorbent

Qeq of the control set-up (Pb)

Qeq = (2,590 mg/L – 2,590 mg/L) x 1L


0mg

= 0 mg Pb
0 mg of biosorbent

= n/a

Qeq of the 300 mg set-up (Pb)

Qeq = (2,590 mg/L – 221.7 mg/L) x 1L


300mg

= 2368.3 mg Pb
300 mg of biosorbent

= 7.894 mg Pb/mg of biosorbent

Qeq of the 500 mg set-up (Pb)

Qeq = (2,590 mg/L – 162.7 mg/L) x 1L


500mg

= 2,427.3 mg Pb
500 mg of biosorbent

= 4.855 mg Pb/mg of biosorbent

75
APPENDIX G

Computation for the Value of t for the Statistical Analysis (using the t-test for
dependent samples)

t= D
SD
√N

tcritical = 2.015

0g vs 0.3g

[Cu] mg/L 0g 0.3 g

Initial 12570 12570

Final 12570 8238.7

Difference 0 4,331.3

(D-D) -2,165.55 2,165.55

(D-D)2 4,689,606.80 4,689,606.80

∑(D-D)2 = 9,379,213.61
n=6
D = 4,331.3
2
= 2,165.55
SD = ∑(D-D)2
n-1
= 9,379,213.61
6-1
= 9,379,213.61
5
= 1,875,842.72
= 1,369.61

76
t= 2, 165.55
1,369.61
√6
= 2, 165.55
559.14
= 3.873
tcomputed(0g vs 0.3g) vs tcritical
3.873 > 2.015

0g vs 0.5g

[Cu] mg/L 0g 0.5 g

Initial 12570 12570

Final 12570 7,961.7

Difference 0 4,608.3

(D-D) -2,304.15 2,304.15

(D-D)2 5,309,107.22 5,309,107.22

∑(D-D)2 = 10,618,214.45
n=6
D = 4,608.3
2
= 2,304.15
SD = ∑(D-D)2
n-1
= 10,618,214.45
6-1
= 10,618,214.45
5
= 2,123,642.89

77
= 1,457.27
t= 2,304.15
1,457.27
√6
= 2,304.15
594.93
= 3.873
tcomputed(0g vs 0.5g) vs tcritical
3.873 > 2.015

0.3g vs 0.5g

[Cu] mg/L 0.3g 0.5 g

Initial 12570 12570

Final 8238.7 7,961.7

Difference 4,331.3 4,608.3

(D-D) -138.5 138.5

(D-D)2 19,182.25 19,182.25

∑(D-D)2 = 38,364.5
n=6
D = 4,331.3 + 4,608.3
2
= 8,939.6
2
= 4,469.8
SD = ∑(D-D)2
n-1
= 38,364.5
6-1

78
= 38,364.5
5
= 7,672.9
= 87.60
t= 4,469.8
87.60
√6
= 4,469.8
35.76
= 124.99
tcomputed(0.3g vs 0.5g) vs tcritical
124.99 > 2.015

[LEAD]
0g vs 0.3g

[Pb] mg/L 0g 0.3 g

Initial 2590 2590

Final 2590 221.7

Difference 0 2368.3

(D-D) -1,184.15 1,184.15

(D-D)2 1,402,211.22 1,402,211.22

∑(D-D)2 = 2,804,422.45
n=6
D = 2,368.3
2
= 1,184.15
SD = ∑(D-D)2
n-1

79
= 2,804,422.45
6-1
= 2,804,422.45
5
= 560,884.49
= 748.92
t= 1,184.15
748.92
√6
= 1,184.15
305.75
= 3.873
tcomputed(0g vs 0.3g) vs tcritical
3.873 > 2.015

0g vs 0.5g

[Pb] mg/L 0g 0.5 g

Initial 2590 2590

Final 2590 162.7

Difference 0 2427.3

(D-D) -1,213.65 1,213.65

(D-D)2 1,472,946.32 1,472,946.32

∑(D-D)2 = 2,945,892.65
n=6
D = 2,368.3
2
= 1,213.65

80
SD = ∑(D-D)2
n-1
= 2,945,892.65
6-1
= 2,945,892.65
5
= 589,178.53
= 767.58
t= 1,213.65
767.58
√6
= 1,213.65
313.36
= 3.873
tcomputed(0g vs 0.5g) vs tcritical
3.873 > 2.015

0.3g vs 0.5g

[Pb] mg/L 0.3g 0.5 g

Initial 2590 2590

Final 221.7 162.7

Difference 2,368.3 2427.3

(D-D) -29.5 29.5

(D-D)2 870.25 870.25

∑(D-D)2 = 1,740.5
n=6
D = 2,368.3 + 2,427.3
2

81
= 2,397.8
SD = ∑(D-D)2
n-1
= 1,740.5
6-1
= 1,740.5
5
= 348.1
= 18.66
t= 2,397.8
18.66
√6
= 2,397.8
7.62
= 314.67
tcomputed(0g vs 0.5g) vs tcritical
314.67 > 2.015

82
APPENDIX H

83
84
85
86
Name: Alyanna Katrina C. Velasco
Batch: 2011
Address: 83 K-9th Street, Kamias, Quezon City
E-mail Address: irma.lair_tala@yahoo.com
Cell phone Number: 09261984137

Secondary
Ramon Magsaysay (Cubao) High School S.Y. 2007-2011
Ermin Garcia Street, Brgy. Pinagkaisahan, Cubao, Quezon City
Primary
St. Isidore Catholic School S.Y. 2001-2005
Quirino Elementary School S.Y. 2005-2007

Personal Data

Gender: Female
Age: 15 yrs. old
Birthdate: July 22, 1995
Favorite Color: Blue, Black
Favorite Band: Paramore, All Time Low

Affiliations:

RMCHS Glee Club


The Apprentice
Undecided (Band)

87
Name: Clarisse Anne S. Caganda
Batch: 2011
Address: 1480 Pipit St., Unit V, Brgy. Comm.,
Quezon City
E-mail Address: caganda999@yahoo.com
Telephone No.: 9720416

Educational Background
Secondary
Ramon Magsaysay (Cubao) High School
S.Y. 2007-2011
Ermin Garcia Street, Brgy. Pinagkaisahan,
Cubao, Quezon City
Primary
Diliman Preparatory School
S.Y. 2001-2007

Personal Data
Gender: Female
Age: 16 yrs. Old
Birthdate: August 24, 1994
Favorite Color: Gray, Pink, Blue
Favorite Band: Chicosci, Paramore

Affiliations:
Science Club member (SY 2009-2010)
Ang Tanglaw
Swimming Club

88
Name: Anne Sherina D. Lazarte
Batch: 2011
Address: 23 Barrio Panopio Compound Cubao,
QC
E-mail Address: aslazarte02@yahoo.com
Cell phone Number: 09266337152

Secondary
Ramon Magsaysay (Cubao) High
School S.Y. 2007-2011
Ermin Garcia Street, Brgy.
Pinagkaisahan, Cubao, Quezon City
Primary

Personal Data
Gender: Female
Age: 15 yrs. Old
Birthdate: February 28, 1995
Favorite Color: None
Favorite Band: Mayday Parade, Paramore

Affiliations:

The Apprentice
Science Club
English Club
CMLI JC-REB NCR 2009

89