~ "

IMPACT OF THERMAL EFFLUENT FROl\1 A COAL PO\VER

STATION ON TIlE VARIABILITY OF BENTHIC

DIATOMS COMPOSITION

Nurfarahaisha Binti Roslan

QH Bac.helor of Science with Honours90.8

(Aquati(.~ Resource Sciel1ce and .Management)846 2014

N974 2014

Impact of Thermal Effluent from a Coal Power Station on the Variability of Benthic

Diatoms Composition

Nurfarahaisha Binti Roslan (32301)

A report submitted in partial fulfilment of the requirements for the degree of Bachelor of

Science with Honours (Aquatic Resource Science and Management)

Supervisor: Prof Madya Dr Othman Bin Bojo

Aquatic Resource Science and Management

Department of Aquatic Science

Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

2013/2014

DECLARATION

I hereby declare that no portion of the work referred to in this dissertation has been

submitted of an application for another degree of qualification or any other university or

institution of higher learning.

Nurfarahaisha Binti Roslan

Aquatic Resource Science and Management

Department of Aquatic Science

Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

I

Acknowledgement

Alhamdulillah praise be to Allah, with His blessing all along I am able to get

through rough times in my life. I would like to thank the Faculty of Resource Science and

Technology (FRST) and UNIMAS for providing best facilities. Greatest gratitude to my

supervisor Assoc. Prof. Dr. Othman Bojo. To my supervisor, I thank you for all the great

advice, support, kindness, and the time you spent for guiding me. Thank you to Prof.

Shabdin Mohd. Long, Assoc. Prof. Dr. Norhadi Ismail, and all the lecturers from

Department of Aquatic Science with their non-stop support upon the completion of my

final year project and during my stay as an aquatic student.

To all precious lab assistants, Mr. Zaidi Ibrahim, Mr. Mohd Nor Azlan, Mr. Nasri

Latib, Mr. Zulkifli Ahmad, Mr. Harris Mustafa Kamal, Mr. Richard Toh, and Mrs. Lucy, I

really appreciate for all the help given to me. Not forgetting Mr. Alias, I thank you for

lending me your time and your boat during my fieldwork. To the humble UNIMAS staffs,

Mr Darus, Mr. Muhammad, Mr. Ismi, thank you for bringing me to my sampling site. To

Nur Najah Mustaffa, Nur Syazwani Abd Rahim, Nur Hanisah Zainal, my lab mates, course

mates, and many of them that too many to mention, thank you for always being supportive

and cheer for me through my up and down. To my beloved family that I missed so much,

thank you for the never-ending support. Last but not least, thank you Jabatan Perkhidmatan

Awam (JPA) for providing financial support throughout my studies here in UNIMAS. To

everyone, only Allah can repay the kindness that all of you had given to me.

II

Table of Contents

Acknowledgement......................................................................................................... I

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

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

List of Figures............................................................................................................... V

Lists of Tables............................................................................................................... VII

Abstract…………………………………………………………………………...… i

1.0 Introduction & Objectives................................................................................ 1

2.0 Literature Review.............................................................................................

2.1 Ecosystem at Thermal Effluent...................................................................

2.2 Phytoplankton Assemblages.......................................................................

2.3 Benthic Diatoms Community......................................................................

2.4 Physico - Chemical Parameters...................................................................

3

3

4

5

7

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

3.1 Description of the Study Area……………………………………………

3.2 Sampling Strategy and Laboratory Analysis……………………………..

3.3 Diatoms Permanent Slide Preparation…………………………………

3.4 Chlorophyll a Analysis………………………………………………...

3.5 Data Analysis…………………………………………………………..

8

8

10

11

12

13

4.0 Results…………………………...……………………………………………

4.1 Benthic Diatoms Community……………………………………...….

4.2 Physico – Chemical Parameters……………………………...……….

4.2.1 Temperature………………………………………………….

4.2.2 Dissolved Oxygen………………………………………...……

14

14

22

22

23

III

4.2.3 pH………………………………………………………………

4.2.4 Salinity…………………………………………………………

4.2.5 Chlorophyll a……………………………………………...…

4.3 Correlation Analysis…..………...……………………………………

25

26

27

28

5.0 Discussion……………………………………...…………………………... 30

6.0 Conclusion and Recommendations…...……………………………………… 35

7.0 References…………………………………………………...……………….. 36

8.0 Appendices………………………………...…………………………………. 41

IV

List of Abbreviations

APHA American Public Health Association

SESCO Sarawak Electricity Supply Corporation Berhad

WHO World Health Organization

V

List of Figures

Description Page

Figure 1 Map showing the location of study site of the Sejingkat Coal Power

Station at Kg. Goebilt, Kuching, Sarawak. (Adapted from Alfred,

2005).

9

Figure 2 Map showing the location of study site of the sampling points (adapted

from Google map).

9

Figure 3 Percentage occurrence (%) of five most common diatoms in sediments

(December 2013).

17

Figure 4 Percentage occurrence (%) of five most common diatoms on rocks

(December 2013).

17

Figure 5 Percentage occurrence (%) of five most common diatoms in sediments

(January 2014).

18

Figure 6 Percentage occurrence (%) of five most common diatoms on rocks

(January 2014).

18

Figure 7 Percentage occurrence (%) of five most common diatoms in sediments

(February 2014).

19

Figure 8 Percentage occurrence (%) of five most common diatoms on rocks

(February 2014).

19

VI

Description Page

Figure 9 Percentage occurrence (%) of five most common diatoms in sediments

(March 2014).

20

Figure 10 Percentage occurrence (%) of five most common diatoms on rocks

(March 2014).

21

Figure 11 Mean temperature (°C) for each sampling month. 23

Figure 12 Mean Dissolve Oxygen (mg/L) for each sampling month. 24

Figure 13 Mean pH for each sampling month 25

Figure 14 Mean salinity (ppt) for each sampling month 26

Figure 15 Mean Chlorophyll a (mg/m3) for each sampling month 28

VII

Lists of Tables

Description Page

Table 1 Coordinate for each station. 8

Table 2 List of taxa recorded during the observation from December 2013 to

March 2014 for sediment samples.

14

Table 3 List of taxa recorded during the observation from December 2013 to

March 2014 for rocks samples.

15

Table 4 Pearson's correlations between chlorophyll a and selected physico-

chemical parameters.

29

i

Impact of Thermal Effluent from a Coal Power Station on the Variability of Benthic

Diatoms Composition

Nurfarahaisha Binti Roslan

Aquatic Resource Science and Management Programme

Faculty of Resource Science and Technology

Universiti Malaysia Sarawak

ABSTRACT

Sejingkat Coal Power Station discharges a large volume of warm effluent into the river. This could give some

impact on the aquatic organisms. The objectives of the study are i) to identify diatoms taxa in the study area

for month of December 2013 until March 2014 ; ii) to estimate relative abundance of each benthic diatom

taxa found in the study area ; iii) to record selective water quality parameters (temperature, dissolved oxygen,

pH, salinity) and chlorophyll a concentration from December 2013 to March 2014 ; iv) to assess the

relationship between physico-chemical parameters with chlorophyll a in all the sampling stations. The

methods used were categorized into sampling strategy and laboratory analysis, cleaning of diatom frustules,

and preparation of diatoms permanent slide. The result of this study revealed that the trend that five most

common diatoms occurring in the sediment were Actinocyclus sp., Cyclotella sp., Manguinea sp., Synedra sp.,

and Coscinodiscus sp., while Navicula sp. and Synedra sp. were consistently inhibiting on rocks at the study

area. The water parameters (temperature, dissolved oxygen, pH, and salinity) were significantly different

along the sampling points. Even though there were significant changes happen to the physico-chemical

parameters of water, the benthic diatoms communities were still inhibiting the area.

Keywords: Thermal effluent, benthic diatoms community, physico-chemical parameters

ABSTRAK

Stesen Arang Batu Sejingkat mengalirkan air yang mengandungi haba panas ke dalam sungai. Ia akan

memberi kesan kepada hidupan akuatik. Objektif kajian adalah untuk i) mengenal pasti taxa diatom di

kawasan tumpuan bermula bulan Disember 2013 hingga Mac 2014 ; ii) untuk mengenalpasti populasi

diatom di tempat kajian ; iii) untuk mencatat parameter air (suhu, larutan oksigen, pH, saliniti) dan klorofil

a bermula Disember 2013 hingga Mac 2014 ; iv) untuk menganalisa perkaitan antara klorofil a dengan

kualiti air (suhu, larutan oksigen, pH, dan saliniti). Kajian terbahagi kepada strategi pengambilan sampel,

melakukan analisa di makmal, pembersihan frustul diatom, dan penyediaan slaid mikroskop kekal. Kajian

mendapati lima genera yang sentiasa didapati di dalam sedimen semasa kajian dijalankan adalah

Actinocyclus sp., Cyclotella sp., Manguinea sp., Synedra sp., dan Coscinodiscus sp., manakala Navicula sp.

dan Synedra sp. didapati kekal di atas substrata batu. Kajian juga mendapati bahawa parameter air

mempunyai perkaitan yang signifikan di setiap stesen. Walaupun terdapat perubahan signifikan berlaku

kepada kualiti air, hidupan bentik diatom dapat hidup di kawasan kajian.

Kata kunci: Efluen termal, Komuniti diatom bentik, parameter fiziko-kimia

1

1.0 Introduction

Sejingkat Power Corporation Sdn. Bhd is situated 27 km north of Kuching City,

Sarawak and was commissioned in the year 1997. The location’s latitude is 1° 38' 15.36"

and longitude, 110° 27' 53.28". The power plant consists of two units which in total

produces 100 Megawatts at full operational capacity. Sarawak Electricity Supply

Corporation Berhad (SESCO), a body responsible for electricity supplies all over the

Sarawak, operates the power station.

At the power station, electricity is produced through the generator triggered by the

rotation of the steam turbine (Chhatwal et al., 1989). After the rotation, steam is converted

back into liquid by the condenser. Heat rejection is required for the steam to convert into

liquid in the condenser, hence there are available continuous liquid supplies to the boiler

(Chhatwal et al., 1989). Therefore, large volume of waters from the environment is

pumped into the condenser, helping the condenser to absorb heat (Chhatwal et al., 1989).

The hot water is then channeled back to the natural environment, known as the thermal

effluent (Chhatwal et al., 1989).

Temperature changes in the thermal effluent were higher as compared to natural

ambient waters. Temperature of thermal effluent is about 5°C-10°C (Choi et al., 2002) or

even warmer 8°C-12°C (Langford, 2001) than natural waters. In Sejingkat Coal Power

Station, preliminary study conducted by Noor (2004) found that the thermal effluent had

the highest temperature at 35.3°C.

Studies on phytoplankton related to thermal effluent impact have been conducted in

many places (Alfred, 2005; Lee, 2003; Chuang et al., 2009; Ma et al., 2011; Muhammad

Adlan et al., 2012; Noor, 2004; Parke and Gammon, 1986; Shiah et al., 2005; Takesue and

Tsuruta, 1978; Zargar and Ghosh, 2006). The phytoplankton research at the thermal

2

effluent normally focused on their diversity and abundance (Lee, 2003; Muhammad Adlan

et al., 2012) and on phytoplankton productivity (Takesue and Tsuruta, 1978). Benthic

diatoms also have been the subject by researchers to evaluate the effect of thermal effluent

(Chuang et al., 2009; Hickman, 1974; Hickman and Klarer, 1975).

Noor (2004) did a preliminary study on the effect of thermal effluent on the

abundance and diversity of diatoms at Sejingkat Coal Power Station. His findings

highlighted that diatoms such as Chaetoceros, Biddulphia, Pleurosigma, and Guinardia

can be used as biological indicators. He stated the four above mentioned taxa did not occur

at the thermal effluent only, but presence at other sampling points (Noor, 2004). Thus,

conducting a similar study in the same area will provide a comparison data on benthic

diatom diversity after a decade.

The objectives of the study were i) to identify diatoms taxa in the study area for

month of December 2013 until March 2014 ; ii) to estimate relative abundance of each

benthic diatom taxa found in the study area ; iii) to record selective water quality

parameters (temperature, dissolved oxygen, pH, salinity) which were done in-situ and

chlorophyll a concentration from December 2013 to March 2014 ; iv) to assess the

relationship between physico-chemical parameters with chlorophyll a in all the sampling

stations.

3

2.0 Literature Review

2.1 Ecosystem at Thermal Effluent

In the thermal effluent, temperature is one of the important key factors regarding

stress effect to the organisms (Alden, 1979). Many studies on thermal effluent have been

done to investigate the level of deterioration on aquatic organisms. Organisms are found to

be affected living at the thermal effluent (Muhammad Adlan et al., 2012). According to

Parker (1979), the effect of higher temperature than the tolerable range for a particular

organism can cause rapid protein denature. As a result, the mortality and the declined in

organisms living at the thermal effluent such as copepods (Alden, 1979; Choi et al., 2012;

Donald, 1976; Jiang et al., 2009), fish (Bernotas, 2002), and benthic populations (Kailasam

and Sivakami, 2004) are obvious. Other than organisms mentioned above, bacteria

production also was found low at the thermal effluent and hence the heterotrophic

nanoflagellates grazing rates on the bacteria eventually was affected too (Choi et al., 2002).

Nevertheless, chlorination activity also contributed to the negative impact on the

habitat resides in the thermal effluent. According to Poornima et al. (2005), even though

the temperature of the effluent were high, decline in phytoplankton population was greater

when the water is treated with chlorine to prevent biofouling. Choi et al. (2002) stated that

the water received a lot of heat and chemicals formed from antifouling activities and were

discharged back into the environment.

However, a few other organisms showed a reversible attitude at the thermal effluent

with an example of a mutualistic relationship and positive growth between blue green

algae Fischerella (cyanobacterium) and Legionella pneumophila (Legionnaires disease

bacterium) occur at the thermal effluent (David et al., 1980). In Namwan Bay, Taiwan, the

thermal effluent does not affect the organisms as its temperature was found lower

4

compared to the least affected area due to an upwelling phenomenon (Chen et al., 2004).

Due to the nature of the surroundings, previously non-abundant fish such as silver beam,

rudd, and tench has found increased at the thermal effluent as compared to before a power

plant was established at the study area (Bernotas, 2002).

2.2 Phytoplankton Assemblages at Thermal Effluent

The thermal effluent had also resulted in the degradation of phytoplankton (U.S

Department of Energy Laboratory, 1999), causes mechanical damage to the entrained

planktons like ruptured prosomes and broken antenna (Choi et al., 2012), and dropped of

biomass about 35% (Schneider and Kay, 1994).

All the green plants have chlorophyll a pigments including phytoplankton. Hence,

assays on the biomass of the phytoplankton can be estimated through chlorophyll a

analysis. Chlorophyll a concentration was found low indicates the density of

phytoplankton also low at the thermal effluent (Chuang et al., 2009; Muhammad Adlan et

al., 2012; Pitchaikani et al., 2010; Poornima et al., 2005; Takesue and Tsuruta, 1978). The

density of phytoplankton community at thermal effluent of Sultan Azlan Shah Power

Station (SASPS) in Manjung, Perak was lower than the inlet area, which is 48313.25

Cells/m3 compared to the inlet area 82398.53 Cells/m

3 (Muhammad Adlan et al., 2012).

The findings from studies of thermal effluent on the phytoplankton show that

diatom (Bacillariophyta) is the most abundant group (Lee, 2003; Muhammad Adlan et al.,

2012; Noor, 2004; Poornima et al., 2005; Zargar and Gosh, 2006). Muhammad Adlan et al.

(2012) stated more than 80% of relative abundance was from the green algae

(Chlorophyta), blue green algae (Cyanophyta), and dinoflagellates (Dinophyta). Study

from Poornima et al. (2005) does not include any blue green algae. Study at the thermal

5

effluent also found that blue green algae, Oscillatoria, Phormidium and Mastigocladus can

live at 45°C-50°C (Savannah River Laboratory, 1979).

Studies on diatoms by Lee (2003) and Noor (2004) have similarities. Both studies

found that Coscinodiscus, Navicula, Pleurosigma and Rhizosolenia are common taxa in the

study area. Chaetoceros was not found in the thermal effluent of Sejingkat Power Station

(Noor, 2004). Muhammad Adlan et al. (2012) agree that the availability of Chaetoceros

was affected by the water temperature. Muhammad Adlan et al. (2012) found abundant of

Pseudonitzschia heimii at the control site, which is not affected by the thermal effluent.

However, Noor (2004) did not have any record of the genus.

2.3 Benthic Diatoms Community at Thermal Effluent

The diatoms can grow in many habitats such as freshwater, brackish water or marine

water, in acidic or alkaline, and could be in lentic and lotic waters (Chia et al., 2011). Chia

et al. (2011) stated diatoms can be either planktonic (living floating on the water column),

benthic (growing by attach itself on a substrate), or both planktonic and benthic. For

planktonic diatoms, the structures are made of fine frustules with or without long

appendices to help them to float in the water (Chia et al., 2011).

The benthic diatom (Bacillariophyta) communities are reliable as ecological

indicators (Birkett and Gardiner, 2005) because they received repeated fluctuations in

environmental conditions including light, temperature, and salinity (McKew, 2011). A

number of research done has stated the diatoms are useful as an environmental monitoring

tool because the diatoms are strongly influenced by their surrounding chemical and

structural environment (Gaiser et al., 2005). Research on diatoms mostly have used the

frustule morphology compared to their genetic affinity to study the diversity of diatoms

(Petrov et al., 2010).

6

The benthic diatoms studies can be sampled from different substrates such as

sediments, rock surface, and macrophytes (Bere, 2010). Rovira et al. (2009) in the study

of periphytic diatom community in salt wedge estuary stated in the findings that most

abundant genera found in the estuary were Cocconeis, Navicula, Nitzschia, and Tabularia.

Since the fluctuation of environmental conditions does contribute effects to the

benthic diatom community (Rovira et al., 2009), therefore, the benthic diatoms have been

used as well in the monitoring study for antrophogenic interference. In a thermal impact

study, the epiphytic algae show changes in their species composition and its primary

productivity (Hickman, and Klarer, 1975). Fonge et al. (2013) studied the diversity and

abundance of benthic algae in an agricultural wetland in response to the physico - chemical

parameters in the paddy fields. In an acidic condition of soils of the paddy fields (pH =

4.65 - 5.27), the most abundant division was the Bacillariophyta (62.29%) followed by the

Chlorophyta (16.98%), and the rest were the Euglenophyta, Pyrrophyta, and Rhodophyta

(1.87%) (Fonge et al., 2013) .

Habitat and structure preferences of the diatoms are in response to anthrophogenic

effect. Study of urban pollution in concerns with diatoms community revealed a

widespread of the diatoms distribution (Bere, 2010). Bere (2010) in the study found at

highly impacted area with urban pollution were characterized by species such as

Gomphonema sp., Nitzschia sp., Nupela sp., Rhoicosphenia sp., Sellaphora sp., Fallacia

sp., and Luticola sp. While the least impacted area were composed of diatoms species such

as Aulacoseira sp., Cymbopleura sp., Eunotia sp., Fragilaria sp., and Pinnularia sp. (Bere,

2010).

7

2.4 Physico-chemical Parameters of Thermal Effluent

At the study area, Environmental Impact Assessment (EIA) study has been done in

year 2002 and there were records of preliminary study conducted to assess the thermal

impact on aquatic organisms such as harpaticoid copepods (Alfred, 2005) and diatoms

populations (Noor, 2004) during the operation of the Sejingkat Coal Power Station.

From the study, the thermal effluent has water temperature of 35.3 °C (Noor, 2004)

and 35.8°C (Alfred, 2005). Langford (1990) stated that apart from thermal influence on the

waters, the incoming river water and tides have significant effects. Previously recorded

salinity level at the study area does fluctuate due to incoming low tides and high tides in

between 4.5 PSU to 28 PSU (Alfred, 2005; Noor, 2004).

For dissolved oxygen, study conducted by Muhammad Adlan et al. (2012) at

SASPS in Manjung, Perak shows a dissolved oxygen concentration of between 4.973mg/L.

The level of dissolved oxygen at the thermal effluent was slightly higher which are 5.58

mg/L (Alfred, 2005) and 6.98mg/L (Noor, 2004).

For pH, the estuary of the Muara Tebas near Sejingkat Coal Power Station has

slightly acidic (6.29 - 6.7) and neutral (7.08) in Noor (2004) record, while all neutral pH

(7.95) of the waters during samplings conducted by Alfred (2005). Langford (1990) in

reviewing the data on the pH level of the thermal effluent stated there could be an effect to

the marine organisms when the pH exposed to a reduction of 0.3 units. The pH was

affected from the desulphurization of wastes being discharged into the open water that will

endanger aquatic life due to increasing acidity (Langford, 1990). Desulphurisation of

wastes was required to reduce emission of harmful sulphur dioxide that eventually

contribute to the acid rain (Langford, 1990).

8

3. 0 Materials and Methods

3.1 Description of the Study Area

Sejingkat Coal Power Station is located at the latitude 1° 38' 15.36" and longitude,

110° 27' 53.28" of Sarawak River (Figure 1). For the study, 8 sampling stations was

established with a distance of 100 meters away from each other, parallel to thermal effluent

area. One sampling point served as control site (station 8) which was located at the jetty of

Kampung Goebilt. Global Positioning System (GPS) was used to coordinate the locations

(Table 1).

Table 1: Coordinate for each station

Station Latitude (N) Longitude (E)

1 1° 38’ 07.7” 110°27’47.1”

2 1° 38’ 08.3” 110°27’50.1”

3 1° 38’ 12.3” 110°27’55.0”

4 1° 38’ 16.0” 110°27’57.9”

5 1° 38’ 04.1” 110°27’45.8”

6 1° 38’ 03.0” 110°27’44.6”

7 1° 38’ 58.6” 110°27’41.1”

8 1° 38’ 54.9” 110°27’27.8”

9

Figure 1: Map showing the location of study site of the Sejingkat Coal Power Station at Kg. Goebilt, Kuching,

Sarawak. (Adapted from Alfred, 2005).

Figure 2: Map showing the location of study site of the sampling points (adapted from Google map).

10

3.2 Sampling Strategy and Laboratory Analysis

Field sampling were conducted monthly from December 2013 to March 2014.

Sediment samples were collected at all sampling stations to estimate the percentage

occurrence of benthic diatoms around the Sejingkat Coal Power Station . Sampling was

conducted during low tide condition. Physico-chemical parameters such as water

temperature, dissolved oxygen, and pH were measured using Hanna Instruments HI-9146,

while salinity was measured using Hand Refractometer Atago.

Diatoms were sampled depending on the availability of the substrates at the

sampling site by using recommendations from the American Public Health Association

(APHA) (method 10300B) (1999). The substrates sampled were rocks and sediments. For

rocks, diatoms were sampled by brushing the surface of rocks with a toothbrush. The

samples were placed in small bottles. For sediment samples, a modified syringe (diameter=

2.9cm) was used to soak in about 1 cm of surface sediment. All the samples were placed in

a sealed plastic wrapper and was labeled properly. Each station was sampled in three

replicates for both rock and sediment samples. For each month, 24 sediment samples were

obtained from 8 stations, and only 6 samples were obtained from rock samples due to the

rocks substrates only available at station 1 (thermal effluent) and station 5. In the

laboratory, diatoms samples were preserved in Lugol’s solution. Samples were kept at 4°C

in refrigerator (Brand Acson International) prior to analysis.

https://www.google.com.my/url?sa=t&rct=j&q=&esrc=s&source=web&cd=1&cad=rja&uact=8&ved=0CCcQFjAA&url=http%3A%2F%2Fajph.aphapublications.org%2Ftoc%2Fajph%2F89%2F3&ei=yopqU8_xEMrprQebvYCQAQ&usg=AFQjCNFaagjHdkjqjxd75iIe_WRNcla05A&bvm=bv.66111022,bs.1,d.c2E

11

3.3 Diatoms Permanent Slide Preparation

Diatom samples were processed according to APHA (method 10300C) (1999).

Diatoms samples were treated with about 2 drops of hydrogen peroxide (H2O2) solution.

The process was aimed to clean the diatom frustules from organic matter (Birkett and

Gardiner, 2005).The samples were left overnight to allow the particles to settle at the

bottom. After 24 hours, the samples were rinsed with distilled water by taking out the

water slowly from the top in order not to accidentally take out the diatoms. The samples

were rinsed three to four times until the pH of the fluid on 7.

The cleaned diatom samples were ready for permanent slide preparation. Duran

Group 76 x 26 mm slide and 22 x 22 mm cover slip must be cleaned. One drop of cleaned

diatom samples were placed onto the cover slip. The cover slip with cleaned diatom

samples were dried through gentle heating on the hot plate. The glass slide was heated at

90°C. A drop of Naphrax as the mounting medium was used to mount the glass slide and

cover slip. The cover slip was turned over and placed on top of the slide. The glass slide

with mounted cover slip went heated for about 30 seconds to 1 minute. After that, the slide

was left to be cooled for a few minutes. A total of 96 slides from sediment samples and 24

slides from rock samples were made.

Diatoms were counted using Motic BA210 compound microscope. The diatom

valves were seen under 1000x magnification with oil immersions. 100 cells were counted

for each station. Identification of diatoms were based on Tomas (1997) and Cupp (1943)

up to genus level.

12

3.4 Chlorophyll a Analysis

Diatoms extraction for photosynthetic pigment followed the APHA

(spectrophotometric method 10200H) (1999) with some modification. Work was

conducted under subdued light to avoid degradation of the chlorophyll a. The sediments

were ground with 90% of acetone. After grinding was completed, the sample was

transferred into a screw- cap centrifuge tube. The grinder was rinsed with a little of 90%

acetone. After the work was done, the samples were kept at least 4 hours in 4°C. Samples

were centrifuged at 3000rpm for 10 minutes. Samples were analyzed using Lorenzen (1967)

spectrophotometric equation. Volume of the sediment samples were obtained through dry

weight of the samples divided with the density of sediment.

Chlorophyll a (mg/m3) = 26.7 (E0 - Ea) x V

Vs x L

Where,

E0 = absorbance before acidification at 665nm

Ea = absorbance after acidification at 665nm

V = Volume of water content of the samples with acetone

added

Vs = Volume of sediment samples

L = 1cm length of light path of the spectrophotometric cell

13

3.5 Data Analysis

Qualitative measurement is to be conducted by listing presence (+) or absence (-) of

diatom taxa in each sampling station and months as presented in Table 1. The counted

diatom cells were represented in percentage occurrence. One-way analysis of variance

(ANOVA) using statistical software Statistical Package for Social Science (SPSS) version

22 was used to test the significance differences among the eight stations for each physico-

chemical parameters like temperature, salinity, dissolved oxygen, and pH. Differences

between groups were considered significant at the p < 0.05 level. Correlation analysis with

Pearson’s Correlation was used to know the relationship of chlorophyll a with each of

selected physico-chemical parameters such as temperature, dissolved oxygen, pH and

salinity.