CHAPTER 3: METHODOLOGY

3.1 Introduction

The method used to treat the POME by using Microbial Fuel Cell is discussed in detail

throughout this chapter. List of apparatus, material and chemicals are also listed in order to

justify all the procedure needed in this wastewater treatment process. Moreover, this chapter

discussed about the materials used including the preparation, methods to prepare and how the

research is being conducted where the job scopes were divided into two parts, the preparation of

microbial fuel cells and analysis. Finally, the treated POME were analysed in for the removal

suspended solid, BOD, COD and electricity generation. The performance of the analysis will be

compared in term of percentage removal of all parameters before and after the treatment.

Effect of total no of electrode with electricity generation

Effect of electrode size with electricity generation

Double Chamber Reactor

Experiment Parameter

(BOD, COD, PH and TSS)

Analysis

POME (waste water)

3.2 Experimental Procedure

pH Electricity Generation

CODBOD

TSS

3.3 Apparatus, Equipment and Material

3.3.1 Apparatus

The apparatus used in the experiment were:

a) Beaker (500mL).

b) Plastic container with sealable lids

c) Plastic pipe (polyethylene or PVC)

d) Carbon cloth

e) Sealing material

f) Wires with alligator clips

g) BOD bottles

h) Incubator

3.3.2 Equipment

The equipment used in this experiment were:

a) pH meter (B-811 Buchi Extraction System, Switzerland).

b) COD reactor (A 32, Germany).

c) Spectrophotometer DR 2800 (Hach, USA).

d) Multimeter XY-360TR

3.3.3 Chemicals

The chemical used in this experiment were:

a) Salt

b) Industrial wastewater (POME)

c) COD reagent (Low range, 3 mg/L to 150 mg/L, Hach)

d) Nitrification inhibitor

3.4 Sampling of Palm Oil Mill Effluent (POME)

The wastewater (POME) used for this experiment was collected at Serting Palm Oil Mill,

Bandar Seri Jempol, Negeri Sembilan. The POME was collected at the drain point before the

wastewater was sent to the treatment pond. 10 L of raw POME was taken and stored in the dark

plastic bottle to preserve the wastewater from exposed to the sunlight.

Figure 3.1 Sampling point of the palm oil mill effluent

For the preservation of the wastewater sample, the POME collected was kept in the fridge

at 4ºC to ensure that no biological and chemical changes will occur to the POME since it will be

continuously used for the experiment. After the POME collection, the initial characteristic of the

wastewater were analyzed. A portion of the POME was taken to determine it pH, suspended

solid, BOD and COD values.

3.5 Preparation of Microbial Fuel Cells

In anode chamber, 100 ml of salt solution (pure NaCl), would be added to each of

the 500 ml of wastewater samples to make the mixture electrically conductive (Pranab and

Deka, 2010). This mixture would be placed in a sealed chamber to stop entering of oxygen, thus

forcing the microorganism to use anaerobic respiration. An electrode which is carbon cloth

coated with platinum would then be placed in the solution that would act as the anode.

In the second chamber of the MFC there would be placed 500 ml of distilled water and

another electrode. According to Pranab and Deka (2010), this electrode, called the cathode would

be positively charged and would be the equivalent of the oxygen sink at the end of the electron

transport chain, only now it would be external to the biological cell. An aquarium pump was

used to supplied continuous oxygen towards the cathode compartment.

Connecting the two electrodes would be a wire and completing the circuit and connecting

the two chambers there would be a salt bridge. In place of commercially available electrodes,

different size of carbon cloth is been used (0.5cm x 0.5cm, 1cm x 1cm and 2cm x 2cm), the tips

of which had been soldered to copper wires travelling from one chamber to the other.

For preparation of salt bridges, a solution containing salt was allowed to boil. The rope

was been using as a medium to transfer the electron. The setup was thereafter allowed to cool for

nearly 2 hours. The salt bridges were thus ready for use. Lastly, external circuit is connected

through a voltmeter and voltage output is measured. The electricity was been recorded for 8

hours. In every 2 hours, the reading was recorded to analyze the maximum voltage been generate

by the microorganism in the microbial fuel cell.

Figure 3.2: A Microbial Fuel Cell

3.6 Analysis of pH

The pH of the Palm Oil Mill Effluent (POME) was measured by using pH meter to

determine the initial pH of the wastewater. The pH meter is standardized by means of the

standard solutions provided. pH is one of parameter that measure the acidity or alkalinity of

liquid solution (Kelly et.al., 2004). This pH can be measure by using pH paper or pH meter. In

this study, pH meter has been used to measure pH of the sample. 50 ml of sample is put into the

glass beaker and the sample is left to stand for a minimum of one hour to allow the temperature

to stabilize, stirring it occasionally while waiting. Noted that the sample has been in freezer

overnight, thus the pH of the sample was allowed to take after the temperature was same with

surrounding temperature.

The electrode of the pH meter is immersed into the waste sample. The electrode is

required immersion of 30 seconds or longer in the sample before reading to allow the meter to

stabilize. This to avoid any error while reading the meter. The electrode is rinsed well with

distilled water and then dabbed lightly with tissues to remove any film formed on the electrode.

The pH value was recorded before and after the experiment had been done.

Figure 3.3: pH Testing Using pH Meter

3.7 Suspended Solid Testing

Spectrophotometer DR 2800 was used to test the suspended solid for POME treatment.

Suspended solid was the actual measure of mineral and organic particles transported in the water.

It also refers to small solid particles which remain in suspension in water. DR 2800 was used by

using photometric method to analyze. The sample was put into the sample cell to the required

level that indicates to the sample cell. For the blank preparation, distilled water was used as the

blank and the blank was insert into the cell holder then the button zero was press and it will

display will 0 mg/L TSS. Then the suspended solid of POME treatment was measured by insert

the sample cell into DR 2800 and the read button was pressed.

Figure 3.4 Suspended solid test using DR 2800

3.8 Chemical Oxygen Demand (COD)

Chemical Oxygen demand is an important parameter for determining the amount of

organic pollution in water. The COD test uses a strong chemical oxidant in an acid solution and

heat to oxidize organic carbon to carbon dioxide (CO2) and hydrogen (H2O). It is expressed in

milligrams per litre (mg/L), which indicates the mass of oxygen consumed per litre of solution.

Mercuric sulphate (HgSO4), potassium dichromate (K2Cr2O7), concentrated sulphuric acid

(H2SO4) and silver sulphate (Ag2SO4) is used as content of COD reagent. The COD vial that was

used in this experiment was the high range which range from 0 mg/L – 15000 mg/L.

First step, the COD reactor was turned on and preheats for 150ºC. After that, 2mL of

wastewater sample was carefully pipette into the vial. The vial was closed and cleaned at the

outside of surface vial. Then, the vial was invert for several times to mix. The vial was heated

into the COD reactor for two hours. The blank sample was analyzed to ensure that the initial

reading is ZERO. After two hours the COD vial was insert into cell holder in DR 2800 and the

reading was measured by pressing READ button.

Table 3.7: Sample Concentration Range (Source: Principles of Water Quality Management,

1990).

Sample Concentration Range (mg/L) COD Digestion Reagent Vial Type

0-150 Low range

0-1500 Medium range

0-15000 High range

(a) (b)

Figure 3.5 (a) COD vial was heated for 2 hours (b) Vial was test using DR

2800

3.9 Analysis of Biological Oxygen Demand (BOD)

Put at each of 300 mL of BOD bottles with 0.16 g of nitrification inhibitor. One 300mL

of BOD bottles was put with 100mL of wastewater sample and was added with 200mL of

distilled water. One 300 mL of BOD bottles was put with 200mL of wastewater sample and was

added with 100mL of distilled water. One 300 mL of BOD bottles was put with 300 mL of

wastewater sample. One 300 mL of BOD bottles was put with 10 mL of wastewater sample and

was added with 290 mL of distilled water. One 300 mL of BOD bottles was put with 50mL of

wastewater sample and was added with 250mL of distilled water. Check all the initial DO in the

BOD bottle by using DO meter as seen in Figure 3.6. Make sure no bubble trapped and close the

bottle cap. The entire BOD bottle was put in the thermostat that has been set to 20 oC for 5 days.

The final DO was measured after 5 days and BOD5 value was calculated. BOD formula is used

to determine the BOD in mg/L by calculation (Barnes, et. al. 1981).

Figure 3.6: Analyzing DO Value