Faculty of Resource Science and Technology Heavy metals in ... Metals in Fishes From Sarawak... ·...

Heavy metals in fishes from Sarawak River

Nur Hanisah binti Mohd Yusof

Bachelor of Science with Honours

(Aquatic Resource Science and Management)

2014

Faculty of Resource Science and Technology

HEAVY METALS IN FISHES FROM SARAWAK RIVER

NUR HANISAH BINTI MOHD YUSOF

This project is submitted in partial fulfilment of the requirements for the degree of Bachelor

of Science with Honours

(Aquatic Resource Science and Management)

Department of Aquatic Science


UNIVERSITI MALAYSIA SARAWAK

2014

I

ACKNOWLEDGEMENT

All praises to Allah for helping me in every moment in completing my final year project.

I would like to express my deep appreciation to my supervisor Prof. Madya Dr.

Othman Bojo for his guidance, help, encouragement and patience throughout my project.

Thanks to my co-supervisor, Dr Farah Akmal Idrus for her great help, advice and

information while dealing with any problems during work.

I also extend thanks to Unimas and Faculty of Resource Science and Technology for

the facilities and financial support throughout the project.

I would like to express my full appreciation and gratitude to laboratory assistants,

Mr. Zaidi bin Ibrahim, Mr. Azlan and Mr. Tomy for helping me to set up the apparatus and

instruments during the laboratory works.

Millions of thanks to my beloved parents, Mohd Yusof bin Osman and Che Om binti

Hamid for their endless love, support, blessing and advice to boost my strength and passion

throughout the project.

Not to forget to my partners Nur Syuhadah Kamarozaman, Nur Syazwani Abdul

Rahim and Masania Mohd Basri, my housemates, and my classmates for their help, thoughts

and recommendation.

Special thanks to all of individuals that involved in this project directly or indirectly

in completing this project. Thank you so much. May Allah bless you.

II

TABLE OF CONTENT

Acknowledgement……………………………………………………………… I

Table of Contents………………………………………………………………. II

List of Abbreviations…………………………………………………………. IV

List of Tables…………………………………………………………………… V

List of Figures………………………………………………………………….. VI

Abstract……………………………………………………………………….... VII

1.0 Introduction and Objectives……......……………………………………. 1

2.0 Literature Review………………………………………………………... 3

2.1 Heavy metals……………………………………………………….. 3

2.1.1 Zinc……………………………………………………..... 3

2.1.2 Copper……………………………………………………. 4

2.1.3 Lead………………………………………………………. 5

2.1.4 Cadmium…………………………………………………. 5

2.1.5 Mercury…………………………………………………... 6

2.2 Heavy metals uptake mechanism…………………………………... 6

2.3 Bioaccumulation and biomagnification in different part of fish…… 7

2.4 Previous studies related to determination of heavy metals in fish…. 8

2.5 Concerning safety food for consumption…………………………... 11

3.0 Materials and Methods…………………………………………………... 12

3.1 Sampling site description…………………………………………... 12

3.2 Pre-cleaning and treatment for bottles……………………………... 13

III

TABLE OF CONTENT

3.3 Samples collection…………………………………………………. 13

3.4 Samples preparation………………………………………………... 13

3.5 Samples digestion…………………………………………………... 14

3.6 Analysis of samples………………………………………………… 14

3.7 Statistical analysis of data………………………………………….. 15

4.0 Result…………………………………………………………………….. 16

4.1 Concentration of heavy metals between parts……………………… 16

4.2 Concentration of heavy metals between species…………………… 25

5.0 Discussion………………………………………………………………….. 31

5.1 Concentration of heavy metals……………………………………... 31

5.2 Heavy metals contents in part of fish.……………………………… 38

5.3 Other factors influence rate of heavy metals accumulation in fish… 39

5.4 Comparison of heavy metals with FAO/WHO (1985)…………….. 40

6.0 Conclusion……………………………………………………………….. 42

7.0 References……………………………………………………………….. 43

8.0 Appendix...………………………………………………………………. 47

IV

LIST OF ABBREVIATIONS

Abbreviation Description

AAS Atomic Absorption Spectrophotometer

ANOVA Analysis of Variance

BDL Below Detection Limit

Cd Cadmium

Cu Copper

FAO/WHO Food and Agricultural Organization/World Health Organization

FIMS Flame Injection Mercury Spectrophotometer

HCl Hydrochloric acid

Hg Mercury

HNO3 Nitric acid

mg/L Milligram per litre

Pb Lead

Zn Zinc

V

LIST OF TABLES

Table 1: Recommended maximum consumption of heavy metals according to

FAO/WHO (1985) 11

Table 2: Details of fish caught with their length measurement and weight 16

Table 3: Mean concentration of heavy metals analysed in flesh, liver and gills

of the six species in mg/kg 17

Table 4: Range of heavy metals concentration (mg/kg)in all of studied parts

and species 20

Table 5: Total of heavy metals in each parts of fish studied 20

Table 6: Order pattern of accumulated heavy metals in parts of fish 21

Table 7: Total concentration of heavy metals in all body parts of fish species 25

Table 8: Comparison of heavy metals in various species and parts 35

VI

LIST OF FIGURES

Figure 1: Sampling site in main channel of Sarawak River near to Kuching

City 12

Figure 2: Mean concentration of Zinc (Zn), Copper (Cu), Cadmium (Cd),

Lead (Pb), and Mercury (Hg) in the flesh, gill and liver of

Hemibagrus sabanus

22



Hemibagrus planiceps

22



Osphronemus goramy

23


Lead (Pb), and Mercury (Hg) in the flesh, gill and liver of Tilapia

sp.

23


Lead (Pb), and Mercury (Hg) in the flesh, gill and liver of Channa

micropletes

24



Scatophagus argus

24

Figure 8: Zinc (Zn) content in different parts of Hemibagrus sabanus,

Hemibagrus planiceps, Osphronemus goramy, Tilapia sp., Channa

micropeltes, and Scatophagus argus

26

Figure 9: Copper (Cu) content in different parts of Hemibagrus sabanus,



27

Figure 10 Cadmium (Cd) content in different parts of Hemibagrus sabanus,



28

Figure 11: Lead (Pb) content in different parts of Hemibagrus sabanus,



29

Figure 12: Mercury (Hg) content in different parts of Hemibagrus sabanus,



30

VII

Heavy metals in fishes from Sarawak River

Nur Hanisah binti Mohd Yusof

Aquatic Resource Science and Management


University Malaysia Sarawak

ABSTRACT

The main sources of heavy metals are derived from human activities such as urbanization which includes

industrial and agricultural wastewater. There are also increasing of industrial development along the Sarawak

River which probably increase the heavy metals accumulation in the fishes. This study was done to determine

the level of heavy metals (Zn, Cu, Cd, Pb and Hg) in Hemibagrus sabanus, Hemibagrus planiceps,

Osphronemus goramy, Tilapia sp., Channa micropeltes, and Scatophagus argus from Sarawak River. Fish

samples (flesh, liver and gills) were digested with concentrated HNO3 (63%) and HCl (37%) on the hot plate

and heavy metals concentration in fish samples were analysed by using Flame Atomic Absorption

Spectrophotometer (FAAS) for Zn, Cu, Cd and Pb while Hg was analysed by using Flow Injection Mercury

System (FIMS). The accumulation was observed in three parts of fish which were flesh, gills and liver in order

liver > gill > flesh. The result revealed that Zn concentration was the highest in Hemibagrus sabanus (88.8 -

368.25 mg/kg), followed by Cu in Channa micropeltes (BDL - 173.08 mg/kg), Cd concentration in Tilapia sp.

(6.8 - 13.38 mg/kg), Pb in Hemibagrus sabanus (0.37 - 4.09 mg/kg) and Hg concentration in Channa

micropeltes (0.32 - 1.97 mg/kg). This study also found that heavy metals accumulation in Hemibagrus sabanus,

Hemibagrus planiceps, Tilapia sp. and Channa micropeltes were exceeded the permissible limit recommended

by FAO/WHO (1985) and not advisable for regular consumption.

Keywords: heavy metals, fish, Sarawak River.

ABSTRAK

Sumber utama logam berat berasal dari aktiviti manusia seperti ubanisasi yang termasuk air sisa industri dan

pertanian. Terdapat juga peningkatan pembangunan industri sepanjang Sungai Sarawak yang mungkin

meningkatkan akumulasi logam berat dalam ikan. Kajian ini dilakukan untuk mengetahui tahap logam berat

(Zn, Cu, Cd, Pb and Hg) dalam Hemibagrus sabanus, Hemibagrus planiceps, Osphronemus goramy, Tilapia

sp., Channa micropeltes, dan Scatophagus argus dari Sungai Sarawak. Sampel ikan (isi, insang dan hati)

dicerna dengan menggunakan HNO3 (63%) dan HCl (37%) yang pekat di atas hot-plate dan konsentrasi logam

berat dalam sample ikan dianalisa dengan Spektrofotometer Penyerapan Atom Nyalaan untuk Zn, Cu, Cd dan

Pb manakala Hg dianalisis menggunakan Injection Merkuri Spectrofotometer Nyalaan. Akumulasi itu telah

dilihat dalam tiga bahagian ikan iaitu isi, insang dan hati mengikut urutan hati > insang > isi. Hasil kajian

ini mendedahkan konsentrasi Zn adalah yang tertinggi dalam Hemibagrus sabanus (88.8 - 368.25 mg/kg),

diikuti dengan Cu dalam Channa micropeltes (BDL - 173.08 mg/kg), konsentrasi Cd dalam Tilapia sp. (6.8 -

13.38 mg/kg), Pb dalam Hemibagrus sabanus (0.37 - 4.09 mg/kg) dan konsentrasi Hg dalam Channa

micropeltes (0.32 - 1.97 mg/kg). Hasil kajian ini juga mendapati bahawa akumulasi logam berat dalam

Hemibagrus sabanus, Hemibagrus planiceps, Tilapia sp. dan Channa micropeltes telah melebihi had yang

dibenarkan oleh FAO/WHO (1985) dan tidak digalakkan untuk dimakan kerap.

Kata kunci: logam berat, ikan, Sungai Sarawak

1

1.0 INTRODUCTION

Increase of human population also increase the demand of food supply including fish

as they contain high-quality protein. Fishes are one of the main protein source besides meat

which comprise a lot of omega-3, fatty acids, amino acids, and vitamins which are important

in maintaining human health and well-being (James, 2013). The beneficial minerals and

vitamins include omega-3 fatty acids that are good for our body which is eicosapentenoic

acid (EPA) and docohexenoic acid (DHA), that are able to lower cholesterol level, lowering

blood pressure and maintain the heart in good condition (James, 2013). Fortunately in

Malaysia, there are a lot of fishes and seafood that easily found and can be consumed as our

daily protein supplement. Despite having a lot of nutritional value, fishes may also contain

certain amount of heavy metals accumulated in their body tissue and organs through

bioaccumulation and biomagnification processes as they are positioned in higher trophic

level in food chain (Neff, 2002).

Rapid urbanization and development in Malaysia, as well as increasing of human

population, the number of factories and housing projects are also rising. The discharge of

untreated wastewater from industries, housing and intensive agriculture activities may result

in water contamination which may also contribute to the detrimental heavy metals into the

river (Murtala et al., 2012). These heavy metal wastes from anthropogenic activities have

devastating effect to the environment and directly influence the river and aquatic ecosystem

including fishes.

Some of the heavy metals are essential for growth such as iron, copper and zinc.

There are also non-essentials heavy metals such as mercury, cadmium, lead and arsenic. If

the concentration of heavy metals are over limit, it may become toxic to human. The

2

concentration of heavy metals should not exceed the permitted value established. The heavy

metals accumulated from food chain may give detrimental effect not only to fish but also to

human health if we consume it regularly. For example, in Japan, the disease called ‘Itai-Itai’

caused by mass consumption of cadmium from Toyama Jintsu River was documented

(Sarejam, 2009). There were also ‘Minamata’ disease in Japan caused by mercury poisoning.

It is known as the Four Big Pollution Disease of Japan (Sarejam, 2009).

However, Sarawak River is expected to face the same problem of contamination that

may be contaminated with heavy metals discharge from untreated waste from industries,

housing nearby, boat and ferry services and also agricultural site. Heavy metals may affect

human health if the level of concentration consumed are high. Thus, this study is conducted

to ensure the safety of consuming fishes from the river. Besides, there are inadequate studies

about heavy metals in Sarawak River.

Therefore, the objectives of this study were:

(1) to determine the concentration of heavy metals (Hg, Cd, Pb, Cu and Zn) in liver, gills

and flesh of selected fishes from Sarawak River;

(2) to compare concentration of heavy metals between species;

(3) and to determine whether the fishes are safe for human consumption.

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2.0 LITERATURE REVIEW

2.1 Heavy metals

Heavy metals are the general term for metallic elements that have average density

greater than 5g/cm3 and high atomic weight (Duffus, 2002). According to Mudgal et al.

(2010), in order to maintain good condition of physiological and biochemical mechanism in

living organisms, around 30 of these elements are needed. These important elements are

called essential metals as they are very crucial for body system to be functioned well, for

example zinc and copper (Mudgal et al., 2010). But there are also non-essential heavy metals

that contaminated in fish such as cadmium, lead and mercury which are highly toxic to

human even in low concentration.

2.1.1 Zinc

Zinc has its importance in industry. It is used in batteries, roof-covering and as alloys.

It often used in manufacturing automotive engine parts and electrical equipment. In addition,

it is also used in steel coating to prevent corrosion. To sustain human being, small amount

of zinc is needed to forms part of specialized enzymes and other proteins. For example,

enzymes that contain zinc are used to digest food and make DNA and RNA, which are

molecules that store genetic information (Palo Alto Regional Water Quality Control Plant,

1999a).

4

The immediate exposed to zinc may cause abdominal pain, nausea, diarrhea and

vomiting. Intake of large amounts of zinc for longer periods may cause anaemia, nervous

system disorders and damage to the pancreas (Plum et al., 2010). However, zinc are not

causative agent of cancer (Plum et al., 2010).

2.1.2 Copper

Copper exists in reddish-brown, shiny, ductile and malleable metals that can

combining with other element to form compounds. Copper can be found in block form,

sheets, wire, tubing and powder form. It is used in manufacture of electrical conductors,

plumbing (Nayem et al., 2011), bronze, brass and other alloys manufacture, ammunition and

copper salts (Palo Alto Regional Water Quality Control Plant, 1999b)

A small amount of copper is enough for well-functioned body. It act as catalyst when

it combines with enzymes (Wilson, 2011). In addition, copper is also important in nervous

system to stimulates production of neurotransmitters epinephrine, norepinephrine and

dopamine. Besides that, it involve in transformation of melanin for pigmentation of the skin

and important for heart and arteries as it able to maintain and repair connective tissues

(Wilson, 2011). Thus, deficiency of copper may lead to suffer coronary heart disease. The

imbalance of copper in human body may also cause nervous system dysfunction,

miscarriages and infertility and liver damage (Wilson, 2011).

5

2.1.3 Lead

Lead is widely used in insecticides, cable coverings, ammunition, plumbing, battery

storage in vehicles and as industry waste. Lead can be found in numerous food especially

fish as it is highly subjected to industrial pollution. Besides that, there are also contamination

of lead in storage and manufacture such as canned food (Mudgal et al., 2010). Paint also

used lead to add colour, improve its ability to cover the surface and make it durable, and this

include painted toys and furniture.

The excess consumption of lead may cause damage to nervous system and disrupt

kidney permeability (Hanan et al., 2009). It can also cause high blood pressure and anaemia.

Exposure at very high level may cause damage to reproductive system, coma and eventually

death. Lead is carcinogenic to human and may cause cancer (Martin & Grisworld, 2009).

2.1.4 Cadmium

Cadmium is highly toxic even in low concentration. It is used in industrial processing

such as metals coatings, electroplating, batteries, control rods and shields in nuclear reactor

and so on (Martin and Grisworld, 2009; Mudgal et al., 2010).

The ingestion of cadmium may affect kidney, interrupts metabolism of bone which

cause bone fragility and risk of fractures. It also cause alter endocrine system as well as

reproductive system (Mudgal et al., 2010). Intake of high levels of cadmium can harshly

hurt stomach, vomiting and diarrhea. For long term period in low exposure of cadmium can

cause kidney damage by formation of kidney stone (Martin & Grisworld, 2009).

6

2.1.5. Mercury

The most toxic elements among all heavy metals studied is mercury (Alina et al.,

2012). It able to combine with other element to form organic and inorganic mercury

compound such as methyl mercury, a bioaccumulating substance (Martin & Grisworld,

2009). Mercury is widely used in dental fillings, batteries, light bulbs, thermometer and also

switches. In US, it is reported that coal-burning power plants are the most source of mercury

emission.

Mercury is toxic and carcinogens to human even in small amount. It can caused

permanent damage to the brain, kidneys and developing foetus if exposed to high level.

Besides that, vapours of metallic mercury may cause nausea, vomiting, diarrhea, lung

damaged, increasing in blood pressure, eye irritation and skin rashes if exposed to high

amount in short term (Martin & Grisworld, 2009).

2.2 Heavy metals uptake mechanism

A huge amount of organic and inorganic chemicals from natural and anthropogenic

sources enter river environment. Increase input to the river of some of these chemicals

especially from anthropogenic activities can result in increased the concentration of the

chemicals and enhanced bioaccumulation in the tissue of the fish. Bioavailable chemicals

will be bioaccumulated and thus biomagnified in the fish food webs (Neff, 2002).

Bioavailability means the amount of heavy metals or chemicals that can enter living

organisms by absorbed or adsorbed actively or passively (Neff, 2002). These chemicals can

7

pass through the lipid membrane by dissolving it or pass through channels in the membrane

(Neff, 2002).

The uptake and retention of bioavailable chemicals (heavy metals) from potential

external sources is called bioaccumulation. The rate of uptake of chemicals is high, cause

decreasing gradient and eventually lead to slow the rate of elimination from the tissue

(passively or actively). Bioaccumulation factor is used to measure the ratio of concentration

in tissue and the surrounding environment of the organism (Neff, 2002).

Biomagnification is the process where chemicals from food pass through a food

chain or food web and the concentration is getting high yet greater in higher trophic level

animal. In other words, the concentration ratio of chemicals in consumer’s tissue is higher

than its food as chemicals are transferred by trophic transfer, known as biomagnification

factor (Neff, 2002). Briefly, prey is small in size compared to predator. So, the predator may

consumed a lot of small prey so that the food is enough. The problem is, the prey already

contain amount of heavy metals in body and the predator need to eat abundance of them

which indirectly take in a huge amount of heavy metals from the prey body. This is how the

heavy metals bioaccumulate and biomagnify in living organisms.

2.3 Bioaccumulation and biomagnification in different part of fish

Fish can act as bioindicator in polluted water as it tend to bioaccumulate the heavy

metals and can be used to reflect the condition of surrounding environment (Jezierska &

Witeska, 2006). Different metals have different affinity towards fish organ. Most of metals

are accumulate mainly in high concentration in liver and gills and lower concentration in

8

flesh (Jezierska & Witeska, 2006). Accumulation some of the metals are organ-specific

which mostly are essential metals such as zinc and copper. For example, zinc and copper

have high affinity towards gonads and liver respectively as in these organs they play their

main metabolic roles even at low concentration (Jezierska & Witeska, 2006).

Liver of fishes tends to accumulate higher concentration of heavy metals followed

by gills and flesh (Murtala et al., 2012; Taweel et al., 2011). But according to Eneji et al.

(2011), the accumulation of heavy metals are higher in kidney of Clarias gariepinus. This

basis is also supported by Victor et al. (2012) where the Cd and Pb are also higher in kidney

of the fishes studied. Besides, there were also reported that accumulation of heavy metals

are high in the liver of fishes (Senarathne & Pathiratne, 2007).

For fish flesh, studies found that accumulation of heavy metals is the lowest

compared to liver and gills (Akan et al., 2012; Eneji, et al., 2011; Victor et al., 2012). Taweel

et al. (2013) stated that concentration of heavy metals in gills reflect the concentration of

heavy metals in the water while the concentration of heavy metals in liver indicated the

accumulation or storage of them or how long the metals are accumulated (Taweel et al,

2013).

2.4 Previous study related to determination of heavy metals in fish.

Studies on heavy metals in commercial fishes are intensively done around the world

as to ensure safety of food supply and to reduce the potential hazards effect to human health.

In Malaysia, including Sarawak, even though there were some research done but the

available data on this study are still inadequate. Variety of fish species has been used in this

research in various area including freshwater, marine, lotic and lentic environment.

9

A research on common Tilapia zilli and Clarias gariepinus from Benue River in

north central of Nigeria was investigated by Eneji et al. (2011) in gills and muscles to

determine level of Cd, Cu, Pb, and Zn. They found that concentration of Zn was higher than

Cd, Pb and Zn in both fishes. A year after, another study was done in the same area but on

different fish species which were Districhodus rostarus, Heterotis niloticus, Lates niloticus

and Citharinus citharium (Akan et al., 2012). They revealed that gills had the highest

concentration of Zn and Cd while liver had the highest concentration of Cu.

Whereas in Turkey, Ozturk et al. (2009) investigated heavy metals in common carp

(Cyprinus carpio) from Avsar Dam Lake. Their study involved analysis of heavy metals in

the liver, gills and flesh. They found that Cu concentration was high in the liver while Cd

concentration was low in the muscle. In India, a research was done by Javed and Usmani

(2011) on Channa punctatus (murrel), Clarias gariepinus (cat fish) and Labeo rohita (carp)

which randomly purchased from market. They evaluated the concentration of Cu and Zn in

muscles, liver, gills. Zn and Cu concentration was high in liver of all species studied while

low accumulation in muscles. However, it was reported that metals accumulation in those

fishes were exceed permissible limit.

There was also a study of Pb, Cd, Cu and Zn in Mytus gulio (Anguluwa) from

Bolgoda Lake, Sri Lanka which serves as commercially important fisheries but at the same

time it received urban and industrial waste from various sources. Thus, raising up the

concerning on heavy metals contamination in the lake (Senarathne and Pathiratne, 2007).

They found that levels of Cu and Zn were slightly higher in gills compared to flesh. Similar

to a study done in North East Coast India by Kumar et al (2012), they revealed that Cu in

muscle tissue was higher in Pampus argentius compared to Zn which more accumulated in

Aritus sp. While the concentration of Hg was the lowest and observed in Trichiurus

10

trichiurus. In 2011, they also study the heavy metal of Cu, Zn, Cd and Hg in aquaculture

pond in India. In contrast with the following study, Zn was the highest concentration in fishes

studied followed by Cu, Hg and Cd. Another research done in coastal water near Gresik,

Indonesia revealed the levels of all heavy metals analysed in ponyfish (Leiognathus equlus),

anchovy (Coilia dusumieri), milk fish (Chanos chanos), mullet (Mugil vaigiensis) and sea

catfish (Arius leptonotacanthus) are low compared to other regions around the world

(Soegianto and Hamami, 2007).

In Malaysia, several studies on determining heavy metals concentration were

conducted in Langkawi, Selangor and Straits of Malacca. Irwandi and Farida (2009) studied

Cu, Zn, Cd, Pd and Hg in five species of fishes preferred by nearby community around the

Langkawi Island. They found that all species were high in Zn and lowest in toxic element of

Hg and Pb. In Bangi area, Taweel et al. (2011) conducted a research to compare the

concentration of heavy metals (Pb, Cd, Cu, and Zn) between natural and industrial cultured

pond of Tilapia fish (Oreochromis niloticus).They discovered that all heavy metals

concentration were highest in liver of the fishes from river. Another research they had done

in 2013 determined Cu, Cd, Zn and Pb in Tilapia fish (O. niloticus) from Langat River and

Engeneering Lake located near to Universiti Kebangsaan Malaysia (Taweel et al., 2013).

They revealed that concentration of Cu was highest in liver while Zn was highest in gills and

muscles. Whereas, Ismaniza and Idaliza (2012) investigated concentration of Cd, Cu, Pb and

Zn in Tilapia sp. from several stations in Mutiara Lake, Puchong as this lake is subjected to

illegal waste dumping from industrialization, previous mining operation and sewage

treatment plant may contain toxicant elements that accumulate in fish. They found that Cu

was highest only in Station 1 that near to sewage treatment plant and paint factory.

11

Alina et al. (2012) investigated level of four toxic heavy metals which are Cd, Pb,

As and Hg in twelve seafood include shellfish from the Straits of Malacca as there are major

fishery industries along this area. Yet, the Straits of Malacca contained high level of heavy

metals as it is one of the busiest shipping routes in the world and various industrial activities

along the West Region of Peninsular Malaysia. However, it was reported that all species

studied were safe to eat.

2.5 Concerning safety food for consumption

Due to a lot of pollution from various anthropogenic activities into aquatic

environment that may also give impact to human health, FAO/WHO suggests the

permissible limits set by FAO/WHO for Cu, Zn, Cd, Pb and Hg which is shown in Table 1.

The permissible limit for essential metals are 10 mg/kg and 150 mg/kg for Cu and Zn

respectively while for non-essential metals are 0.2 mg/kg, 1.5 mg/kg and 0.44 mg/kg for Cd,

Pb and Hg respectively (FAO/WHO, 1985).

Table 1: Recommended maximum consumption of heavy metals according to FAO/WHO (1985).

Heavy metals Concentration (mg/kg)

Cu 10

Zn 150

Cd 0.2

Pb 1.5

Hg 0.44

12

3.0 MATERIALS AND METHODS

3.1 Sampling site description

Sg. Sarawak has two branches namely Sg. Sarawak Kiri and Sg. Sarawak Kanan.

The two branches meet at Batu Kitang, approximately 34 km upstream to Kuching. From

Batu Kitang, the river mainstream meanders across a wide coastal flood plain and through

the city of Kuching. The industrial area is in the eastern part of Kuching. It covers the area

from Sg. Kuap to further downstream at Loba Batu Belat before discharge at Muara Tebas

at South China Sea (Sarawak Government/DANCED, 2001) (Figure 1). A barrage was

constructed to control the river water flow and to prevent intrusion of seawater.

Kuching City Sampling site

Figure 1: Sampling site in main channel of Sarawak River near to Kuching City. (Adapted from

Google Map)

Legend

Sarawak River

13

3.2 Pre-cleaning and treatment for bottles

All apparatus and plastic bottles were decontaminated overnight with 10%

Hydrochloric acid and soaked overnight. They were rinsed with distilled water and air-dried

in laminar flow to prevent contamination (see Appendix A).

3.3 Samples collection

A total of 15 fish from 6 species were caught directly from Sg. Sarawak near to

Kuching waterfront at coordinate N 01033’35.6”, E 110020’10.6” with the help of a

fisherman. The fishes namely Hemibagrus sabanus, Hemibagrus planiceps and

Osphronemus goramy, were caught using gill net, Tilapia sp. and Scatophagus argus were

caught using cast net and Channa micropeltes was caught by fishing. All fishes were put in

polyethylene resealable zipper plastic bag and were kept in a cooler box to maintain its

freshness. Upon arrival at laboratory, the samples were kept in refrigerator at -20o until

further analysis.

3.4 Samples preparation

The samples were allowed to thaw and rinsed with distilled water to removed

absorbed metals on skin. They were first identified by species (see Appendix B – E), then

the length measurement (cm) and wet weight (g) were recorded individually before

dissection as shown in Table 2. Their parts (gills, liver and flesh) were separated immediately

using ceramic knife before keeping in the oven at 55oC for 48 hours and until constant weight

was obtained.

14

3.5 Samples digestion

Hot plate digestion technique were used to digest the samples. The dried samples

were ground with mortar and pestle after allowed to cool in room temperature. 0.5 g of

ground samples were digested in triplicate with 63% nitric acid (HNO3) and 30%

hydrochloric acid (HCl) in ratio 3:1 (Beetseh & Abrahams, 2013) for approximately 2 hours

by using hot plate until the solution colour almost clear. The samples were left to cool at

room temperature. The digestion was done in fume cupboard to prevent contamination and

inhalation of hazardous chemicals. The samples were diluted with 0.1M HNO3 to 50 ml. The

residues were filtered by using 0.47 µm pore size of Whatman filter paper.

3.6 Analysis of samples

The filtered solution were analysed by Flame Atomic Absorption Spectrophotometry

(Thermo Scientific, AAS iCE3500) to measure the concentration of Cu, Zn, Pb and Cd in

each species and each part of fish. For Hg concentration, the Flow Injection Mercury System

(Perkin Elmer, FIMS 400) was used. After calibration of the instrument by using standard

solution, the system was optimized and set based on recommendation by the instrument. The

true metal content result from metals analyses were calculated as follows:

Metal

content

(mg/kg)

Concentration in solution from AAS result (mg/L) x volume of dilution (L)

Weight of dry samples (kg)

=

15

3.7 Statistical analysis of data

Descriptive statistics such as average, range, and standard error values were

calculated. Statistical analyses was performed using one-way analysis of variance (ANOVA)

technique using SPSS (PASW Statistic Version 18) between species and part of fish. P<0.05

was set to check significant difference among species and concentration of metals.

Faculty of Resource Science and Technology Heavy metals in ... Metals in Fishes From Sarawak... ·...

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