CHAPTER I

THE PROBLEM: RATIONALE AND BACKGROUND

Bioaccumulation refers to the intake of substances or

chemicals by an organism in the body by ingestion,

respiration, or contact of the chemicals, which is more

concentrated than its normal amount. It occurs when there

is more ingestion of chemicals in the tissues of the

organism than its excretion. Heavy metals are examples of

these chemicals which are commonly present in bodies of

water. These are known to be pollutants that enter bodies

of water such as streams, lakes, rivers, and groundwater

through industrial and consumer waste, or even from acidic

rain that seeps down the soil (Lenntech B.V. Online, 1998-

2011). Heavy metals are of particular concern around the

world because they are environmentally persistent, undergo

biogeochemical recycling, and pose ecological risks (Gu et

al., 2015; Liu et al., 2015; Olmedo et al., 2013). Heavy

metals can be categorized as being potentially toxic (e.g.,

As, Cd, Cr, Hg, Pb), probably essential (Ni), or essential

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(e.g., Cu, and Zn) (Hedberg et al., 2014; Tuzen, 2009).

Toxic metals can be harmful even if low concentrations, are

ingested for a long time. Toxic effects can also be caused

by excessive intake of essential heavymetals (Gu et al.,

2015; Tuzen, 2009). Marine organisms such as crustaceans,

fish, and shellfish are considered to be healthy foods,

because they have high protein and low saturated fat

contents and because they contain omega fatty acids, which

support health (Ahmad et al., 2015; Olmedo et al., 2013;

Tuzen, 2009). Seafood consumption has been shown to

decrease the risk of coronary heart disease occurring,

decrease mild hypertension, and prevent certain cardiac

arrhythmias (Olmedo et al., 2013; Tuzen, 2009).

Nonetheless, marine organisms may be contaminated with

heavy metals from various sources, such as domestic and

industrial wastewater, and polluted natural runoff and

rivers (Ip et al., 2005; Pan and Wang, 2012). Seafood

consumption is regarded as being an important route through

which humans are exposed to a variety of chemical

contaminants. Pollution is known to be present everywhere;

it is present in the land, water, and air. As an effect of

such, heavy metals are known to be a pollutant which is

common to be present in waters. Heavy Metals can enter in

the waters by means of industrial and consumer waste, or

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even from acidic rain breaking down soils and releasing

heavy metals into streams, lakes, rivers, and groundwater

(Lenntech B.V. Online, 1998-2011). If that is so, the Heavy

metals can enter our body by drinking the water, eating the

foods that are found in that specific location where heavy

metals are plenty or by inhaling, skin and eye contact.

Bivalves which are filter feeders and can be present in

streams, oceans, and lakes or any water supply can absorb

heavy metals either directly from the water or indirectly

from their food. The ability of bivalves to concentrate

trace metals, such as zinc, cadmium, lead and copper from

seawater is well known (Martincic et.al, 1984). In today’s

time, many people fish bivalves for their food because

bivalves are easy to find, and is an important part of

Philippine shell industry, not knowing that they can

accumulate heavy metals which are harmful to our body.

Statement of the Problem

The main objectives of this study are to determine

the presence of cadmium in the tissues and organs of the

Asian green mussel, using the Atomic Absorption

Spectrophotometry; and determine the safety of the bivalves

for human intake in comparison to International / National

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Standards for Heavy Metals in Food by Dr YY Choi standards.

Specifically, the objectives will aim to answer the

following questions:

1. Is the heavy metal cadmium and lead present in the

Asian Green Mussel bivalve? If present then:

A. How much Lead content is present in the bivalve?

B. How much Cadmium content is present in the bivalve?

2. Are the Bivalves found in the shores of Tagbilaran

City specifically in the shore of Poblacion II safe to be

eaten by the people?

Theoretical and Conceptual Framework

INDEPENDENT VARIABLE DEPENDENT VARIABLE

A. Bivalves(Perna Concentration of Heavy

Viridis) metals in the bivalves

B. Location and organs:

season where the

A. Cadmium
bivalves will be
B. Lead
collected
C.
Figure 1.1. Paradigm of the Independent and Dependent

variables on the bioaccumulation of heavy metal (Cadmium

and Lead) by bivalve (Perna Viridis) from Poblacion II,

Tagbilaran City.

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Assumptions

The following assumptions will serve as the bases

of the study:

I. The bivalve (Perna Viridis) will be fresh.

II. The chosen panelist will be physically and visually

healthy.

Hypothesis

Based on the research, the researcher formulated

a null hypothesis:

 The heavy metal (Cadmium and Lead) haven’t been found

in the bivalve (Perna Viridis) so the place is contaminated

with heavy metal cadmium and lead.

Significance of the Study

This study recognizes the usefulness of the

bivalve Asian green mussel that can be attained in the area

of Tagbilaran City, specifically in the shore of Poblacion

II. Since Bohol province has a sufficient supply of Asian

green mussel that can be eaten by the people, the

researcher came up with an idea of making sure if the

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bivalves which are abundant in the nearby shores of

Poblacion II in Tagbilaran City are safe to eat. The

researcher looked forward that the bivalve is safe to be

eaten by the people near the sea or the people buying it

from the market, and this can also help the people in their

future researches which involve concerning the

bioaccumulation of bivalves. Furthermore, the results of

this study can also serve as a reference for future studies

concerning bioaccumulation, and it can help to determine if

the intake of the bivalves from Poblacion II in Tagbilaran

City, meets the standard requirement of heavy metals in the

body.

Scope and Limitations of the Study

This study deals only with the bivalve Asian Green

Mussel (Perna Viridis). The bivalves will be taken from the

coastal area in Poblacion 2, Tagbilaran City. The

researcher would only test for the heavy metals Cadmium and

Lead.

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Definition of Terms

Bivalves - Bivalvia, in previous centuries referred to as

the Lamellibranchiata and Pelecypoda, is a class of marine

and freshwater mollusks that have laterally compressed

bodies enclosed by a shell consisting of two hinged parts.

Bivalves as a group have no head and they lack some usual

molluscan organs like the radula and the odontophore. They

include the clams, oysters, cockles, mussels, scallops, and

numerous other families that live in saltwater, as well as

a number of families that live in freshwater. The majority

are filter feeders.

Bioaccumulation - The general term describing a process by

which chemicals are taken up by an organism either directly

from exposure to a contaminated medium or by consumption of

food containing the chemical. (U.S. Environmental

Protection Agency, 2010)

Heavy Metals - It refers to any metallic chemical element

that has a relatively high density and is toxic or

poisonous at low concentrations. (Lenntech B.V. Online,

1998-2011)

Environmental Protection Agency (EPA) - The Environmental

Protection Agency identifies contaminants to regulate in

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drinking water. The Agency sets regulatory limits for the

amounts of certain contaminants in water provided by public

water systems. These contaminant standards are required by

the Safe Drinking Water Act (SDWA).

Atomic Absorption Spectrophotometry (AAS) – The Atomic

Absorption Spectrophotometry is an analytical technique

that measures the concentration of elements. Atomic

absorption is so sensitive that it can measure down to

parts per billion of a gram (µg dm–3) in a sample. The

technique makes use of the wavelengths of light

specifically absorbed by an element.

Lead - 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. Lead isotopes are the

end products of each of the three series of naturally

occurring radioactive elements.

Cadmium - Cadmium (Cd) is a soft, malleable, bluish white

metal found in zinc ores, and to a much lesser extent, in

the cadmium mineral greenockite. In 2011, U.S. production

of cadmium was estimated at 600 metric tons, down

approximately 40% from the production levels 20 years ago

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(1992). Most of the cadmium produced today is obtained from

zinc byproducts and recovered from spent nickel-cadmium

batteries. Common industrial uses for cadmium today are in

batteries, alloys, coatings (electroplating), solar cells,

plastic stabilizers, and pigments.

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 CHAPTER 2

REVIEW OF RELATED LITERATURE

Bioaccumulation. An important process through which

chemicals can affect living organisms is bioaccumulation.

Bioaccumulation means an increase in the concentration of a

chemical in a biological organism over time, compared to

the chemical's concentration in the environment. Compounds

accumulate in living things any time they are taken up and

stored faster than they are broken down (metabolized) or

excreted. Understanding the dynamic process of

bioaccumulation is very important in protecting human

beings and other organisms from the adverse effects of

chemical exposure, and it has become a critical

consideration in the regulation of chemicals (Extension

Toxicology Network Online, 1993). A 15-month experiment

combining a geochemical survey of Cad, Cu, Zn and Hg with a

bioaccumulation study for three filter-feeding bivalve

species (oysters, Crassostreagigas; cockles,

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Cerastodermaedule; and clams, Ruditapesphilippinarum) was

conducted in a breeding basin of the Nord Medoc salt

marshes connected to the Gironde estuary, which is affected

by historic polymetallic pollution. (Baudimont M. et. al.,

2004) This study utilizes bivalves for the bioaccumulation

of heavy metals. Bioaccumulation of contaminants from the

surrounding media or food can cause concealed toxic

responses in the affected organisms or toxicity to

organisms higher in the food chain. Such propagation

through the food chain leads to human health concerns.

(Ejazul Islam et.al, 2007)

Asian green mussel (Perna Viridis). Also known as

Philippine Green Mussel also known as Amahong or Tahong in

Cebuano and bisaya, this bivalve belongs to the family

Mytilidae. It generally ranges from 80 to 100 millimeters

in length and may occasionally reach 165 mm. Its shell ends

in a downward-pointing beak. Their smooth outer shell

(periostracum) is dark green, becoming increasingly

brownish towards its point of attachment (umbo), where it

is lighter. Younger mussels are bright green in color and

become darker as they ages. The shell’s interior has a pale-

blue sheen. The mussel has a large mobile foot which it

uses to climb vertically should it be covered by sediments.

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It also produces byssus to help it attach to its substrate.

Green mussels fertilize externally and have separate sexes

there are only few functional mussels which is

hermaphrodite. It is harvested for food but is also known

to harbor toxins and cause damage to submerged structures

such as drainage pipes. Asian Green Mussel is a bivalve

that is only found in marine environment.

Taxonomy of Perna Viridis: (Linnaeus, 1758)

Domain: Eukaryota

Kingdom: Metazoa

Phylum: Mollusca

Class: Bivalvia

Subclass: Pteriomorphia

Order: Mytiloida

Superfamily: Mytiloidea

Family: Mytilidae

Genus: Perna

Species: Perna viridis

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 Plate 2.1: Asian Green Mussel (Perna Viridis)

Heavy Metals. Heavy Metals, they are the natural components

of the Earth’s crust and can’t be degraded. To a small

extent they can enter our bodies via food, water, air. Heavy

metals are dangerous because they tend to bioaccumulate in

bodies of organisms.

Cadmium. Cadmium (Cd) is a soft,

malleable, bluish-white metal found

in zinc ores, and in the cadmium

mineral greenockite. Most of the Plate 2.1: Cadmium 1

cadmium produced today is obtained

from zinc byproducts and recovered from spent nickel-

cadmium batteries. First discovered in Germany in 1817,

cadmium found early use as a pigment because of its ability

to produce brilliant yellow, orange, and red colors. It is

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used in making batteries, alloys, coating (electroplating),

solar cells, plastic stabilizers, and pigments. Cadmium is

also used in nuclear reactors where it acts as a neutron

absorber. China, South Korea, and Japan are the leading

producers of cadmium in the world, followed by North

America.

Cadmium and its compounds are highly toxic and exposure to

this metal is known to cause cancer and targets the body’s

cardiovascular, renal, gastrointestinal, neurological,

reproductive, and respiratory systems (Occupational Safety

& Health Administration Online, U.S. Department of Labor,

n.d.). Human uptake of cadmium takes place mainly through

food.

Foodstuffs that are rich in cadmium can greatly increase

the cadmium concentration in human bodies. Examples are

liver, mushrooms, shellfish, mussels, cocoa powder and

dried seaweed. An exposure to significantly higher cadmium

levels occurs when people smoke. Tobacco smoke transports

cadmium into the lungs. Blood will transport it through the

rest of the body where it can increase effects by

potentiating cadmium that is already present from cadmium-

rich food.

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Other high exposures can occur with people who live near

hazardous waste sites or factories that release cadmium

into the air and people that work in the metal refinery

industry. When people breathe in cadmium it can severely

damage the lungs. This may even cause death. Another

important source of cadmium emission is the production of

artificial phosphate fertilizers. Part of the cadmium ends

up in the soil after the fertilizer is applied on farmland

and the rest of the cadmium ends up in surface waters when

waste from fertilizer productions is dumped by production

companies.

Cadmium can be transported over great distances when it is

absorbed by sludge. This cadmium-rich sludge can pollute

surface waters as well as soils.

Cadmium is first transported to the liver through the

blood. There, it is bond to proteins to form complexes that

are transported to the kidneys. Cadmium accumulates in

kidneys, where it damages filtering mechanisms. This causes

the excretion of essential proteins and sugars from the

body and further kidney damage. It takes a very long time

before cadmium that has accumulated in kidneys is excreted

from a human body. Animals and humans eating or drinking

cadmium sometimes get high blood-pressures, liver disease

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and nerve or brain damage. (Lenntech B.V. Online, 1998-

2011)

Lead. Lead (Pb) is a naturally-

occurring neurotoxin. Contamination

still occurs today mostly by drinking

lead-contaminated water, breathing

lead-polluted air, and living in or

Plate 2.3: Lead
near older painted buildings and certain toxic industrial

areas even though many lead containing products (such as

gasoline and house paints) were banned in the 1970s. It is

a highly toxic substance, exposure to which can produce a

wide range of adverse health effects. Both adults and

children can suffer from the effects of lead poisoning, but

childhood lead poisoning is much more frequent. Even today,

at minimum more than four hundred thousand children under

the age of six have too much lead in their blood. There are

many ways in which humans are exposed to lead: through

deteriorating paint, household dust, bare soil, air,

drinking water, food, ceramics, home remedies, hair dyes

and other cosmetics. Much of this lead is of microscopic

size, invisible to the naked eye. More often than not,

children with elevated blood lead levels are exposed to

lead in their own home. Young children under the age of six

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are especially vulnerable to lead's harmful health effects,

because their brains and central nervous system are still

being formed. For them, even very low levels of exposure

can result in reduced IQ, learning disabilities, attention

deficit disorders, behavioral problems, stunted growth,

impaired hearing, and kidney damage. At high levels of

exposure, a child may become mentally retarded, fall into a

coma, and even die from lead poisoning. In adults, lead can

increase blood pressure and cause fertility problems, nerve

disorders, muscle and joint pain, irritability, and memory

or concentration problems. When a pregnant woman has an

elevated blood lead level, that lead can easily be

transferred to the fetus, as lead crosses the placenta. In

fact, pregnancy itself can cause lead to be released from

the bone, where lead is stored—often for decades—after it

first enters the blood stream (Dr. K.M. Gopinathan et. al)

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 CHAPTER 3

METHODOLOGY

Materials

Asian Green Bivalves

Equipment

Mortar and Pestle

Warm Concentrated Nitric Acid

Hydrogen Peroxide

Volumetric Flask

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 Atomic Absorption Spectrophotometry

Procedure

I. Collecting and Gathering of Bivalve Samples

The Bivalve (Perna Viridis) will be collected from the

coastal waters in Tagbilaran City specifically Poblacion 2

with the help of some local fisherman. The collected live

bivalves will be submitted to University of San Carlos-

Talamban campus then the organs and tissues will be

carefully separated from the shell. These dissected parts

will be air-dried. The air-dried samples will be mixed

together and using mortar and pestle, it will be made into

fine powder by grounding.

II. Digestion of Samples

The powdered bodies will be first processed with warm

concentrated nitric acid (50 mg dry weight/ml). The

temperature will be raised to near boiling. After cooling,

an additional 0.25 ml of HNO3 will be added to the

solution. The solution will be heated again until it begins

to change to brown. After a third addition of 0.1 ml HNO3,

the volume will be reduced to approximately 0.5 ml.

Afterwards 0.1 ml of 30 % hydrogen peroxide will be added.

After reduction of the volume by heating, additional

aliquots of H2O2 will be added until the solution attains

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 transparency. Again, the volume will be reduced to 0.5 ml

and 0.1 ml concentrated hydrochloric acid will be added. By

heating, the volume will be reduced to 0.25 ml. lastly the

solution will be added with 2 mL water in a volumetric

flask. (K. Boch, et. al, 2002)

III. Atomic Absorption Spectrophotometry

By using Atomic Absorption Spectrophotometry (AAS), the

calibration and measurement of the heavy metal, cadmium

will be determined.

IV. Comparison of the Results to the Known Standards

The results gathered would be compared to the International

/ National Standards for Heavy Metals in Food by Dr. YY

Choi to determine if the bivalves are safe for human

intake. (Choi, 2011)

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 CHAPTER 4

RESULTS AND DISCUSSION

Results

LABORATORY CODE 2019-03-0628

Bivalve Asian
CLIENT’S CODE METHOD
Green Mussel
PARAMETER/UNIT

Cadmium(mg Cd/Kg) less than 0.6500 AAS, Flame Method

Lead (mg Pb/Kg) less than 1.76 AAS, Flame Method

Table 4.1 Laboratory Results of the heavy metals found in

the bivalves using AAS method from USC-Talamban Campus

After getting the data the researcher compared it to

the International / National Standards for Heavy Metals in

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Food by Dr YY Choi to see if the data is within the

standards or not.

Maximum
permitted
Metal Description of food concentration
in parts per
million

All food in solid form 6

Lead (Pb)
All food in liquid form 1

Cereals and vegetables 0.1

Fish, crab-meat, oysters,

Cadmium (Cd) 2
prawns and shrimps

Meat of animal and poultry 0.2

Figure 4.2 International / National Standards for Heavy

Metals in Food by Dr YY Choi for Cadmium and Lead

The results from the laboratory of the accumulated heavy

metals in the bivalves’ tissues and organs are within the

International / National Standards for Heavy Metals in Food

by Dr YY Choi. This leads the rejection of the null

hypothesis because the heavy metals are present in the

bivalves but is within the standards of heavy metals in

food by Dr. YY Choi. The researcher thought that this is

what the results had shown because the gathering of the

bivalves was done near summer where the metal

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concentrations generally being the highest in the coldest

part of the year (late winter and early spring), and the

lowest during the summer and early fall. Therefore the

biological availability of trace metals is highly dependent

on seasonal meteorological factors which affect their

concentration and physical and chemical forms in the sea

water, as well as the metabolic rate of mussels, especially

in shallow coastal areas up to 10 m deep where the

influence of waves, wind and nutrients is the most

intensive. (Ivana Ujević et. al, 2015)

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 CHAPTER 5

SUMMARY, CONCLUSION AND RECOMMENDATIONS

Summary

The main objectives of this study are to determine

the presence of cadmium in the tissues and organs of the

Asian green mussel, using the Atomic Absorption

Spectrophotometry; and determine the safety of the bivalves

for human intake in comparison to International / National

Standards for Heavy Metals in Food by Dr YY Choi.

Specifically, the objectives will aim to answer the

following questions:

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1. Is the heavy metal cadmium present in the Asian Green

Mussel bivalve? If present then:

A. How much Lead content is present in the bivalve?

B. How much Cadmium content is present in the bivalve?

2. Are the Bivalves found in the shores of Tagbilaran

City specifically in the shore of Poblacion II safe to be

eaten by the people?

Conclusions

Results showed that the Asian Green Mussel is still safe to

eat and the area is not that polluted of heavy metals like

Cadmium and Lead.

The availability of trace metals is highly dependent on

seasonal meteorological factors which affect their

concentration and physical and chemical forms in the sea

water.

Recommendation

The researcher recommends the use of another bivalve to be

experimented on to determine the presence of a heavy metal.

The researcher also recommends the use of another heavy

metal to determine and the place to and the replication of

the study in the area. Lastly, the researcher recommends a

longer period of experiment and study, and patience.

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