METHODOLOGY

Location of Study Area

This research will be conducted in the municipality of Sibutad, Zamboanga del Norte.

This is an inland municipality of Zamboanga del Norte with sixteen barangays around forty-five

kilometers from Dipolog. Six of these are along the coastal areas, namely: Calube, Kanim,

Libay, Panganuran, Sawang and the island of Sinipay. Sibutad lies in the northern boundary of

the province of Zamboanga del Norte facing the islands of Bohol, Cebu, and Siquijor. It is

bounded in the north by the Murceillagos Bay, Dapitan City by the west, Barangay Potungan in

the south, and the municipality of Rizal in the east.

Description of Sampling Sites

There will be two sampling sites in this research which are at different distances from the

major drainage of the mining site in Lalab. The first sampling will be done in Purok 2 of

Barangay Libay (Figure 2) and is about 1 km from the mining site’s drainage. The second and

the last sampling will be done in Lalab (Figure 3) of Barangay Libay, where the effluent from the

mining site flow and is about 10 m away from it.

Oysters that will be collected from Purok 2 and Lalab of Barangay Libay were not

cultured. Oysters live in salty or brackish waters clustering on older shells, rock, piers, or any

hard, submerged surface. They fuse as they grow, forming rock-like reefs that provide habitat for

other marine animals and plants. Oysters from these areas are gathered as part of the people’s

daily meals they usually eat raw oysters and are sold in the markets as a source of livelihood.
Sampling and Sample Preparation

The samples of oysters from different sampling stations will be manually collected. With

the help of the local fisherman, oysters will be randomly collected from areas where appreciable

quantities were found. The samples will be washed thoroughly with seawater to remove dirt and

other debris will be rinsed several times using deionized water. Other plants attached will be

physically handpicked before it undergoes air drying. After which, all samples will be

homogenized using a blender then placed in labeled acid-washed plastic containers and stored in

a refrigerator and transported to the laboratory for preparations and analysis.

Acid Digestion of Sample

About 5 grams of the homogenized sample will be weighed and will be transferred to 125

mL of Erlenmeyer flask and will be added with 20 mL concentrated HNO 3. The samples will be

gently heated over a hot plate maintaining the solution temperature at 60ºⅭ in a well-ventilated

hood and will be allowed to froth. When the frothing ceased, it will be allowed to cool to room

temperature, and 10 mL of concentrated H2SO4 will be added to each sample. The sample will

then gently be heated over a hot plate also maintaining the solution temperature at 60ºⅭ for about

3 hours to ensure complete digestion. The color of the solution will be changed from brownish to

pale yellow. The sample will be subjected to strong heating until no browner NO 2 fumes will be

observed. The solution will be then allowed to cool to room temperature and will be transferred

into a 200 mL volumetric flask and will be diluted to the mark with deionized water. Two

replicates from each sampling site will be digested.

Blank Preparation
 Twenty millimeters of concentrated HNO3 will be poured into a 125 mL Erlenmeyer

flask. It was gently heated over a hot plate in a well-ventilated hood. Then it will be allowed to

cool to room temperature, after which 10 mL concentrated H2SO4 will be added. The mixture

will be then gently heated. After several minutes, it will be subjected to strong heating until no

browner NO2 will be observed. The solution will then be allowed to cool to room temperature

and will be transferred into a 100 mL volumetric flask and diluted to the mark with the deionized

water.

Mercury Analysis

Two hundred milliliters of digested samples will be poured into the BOD bottles. It will

be added with 2 drops of KMNO 4 solution and will be swirled and will be added with more

drops of the solution until the purple color persists. It will be then added with 5 mL of each of

56N HNO3 and 18N H2SO4 and will be swirled continuously. Five mL of 1.5% hydroxylamine

hydrochloride solution will be also added with swirling until the purple solutions become

colorless. Subsequently, 5 mL of 10% SnCl2 solution will be introduced, and the bubbler of Hg

analyzer will be immediately inserted into the BOD bottles containing the sample. The system

will be properly closed. At this stage, the instrument has been previously set at 0% transmittance

(T) and 100% T by pressing the 0% and 100%T switch, respectively. The peak hold and the

absorbance will also be pressed before aeration. The highest absorbance reading indicated by the

blinking of the peak hold signal light will be recorded. The bubbler will be removed and will be

washed with deionized water after recording the highest reading. The pump switch will be left on

to clear the system of mercury released from the previous sample.

The concentration of mercury in the sample was calculated using the equation:
 µg Hg |x|SCF mL digest 1000
= x x
Kg Sample g Sample mL aliquot 1 Kg

where:

ppb : µg Hg/Kg sample

Abs : Absorbance reading in mercury analyzer

SCF : standard calibration factor

mL digest : volume of digested sample (as submitted), in mL

mL aliquot : volume of digested sample used in analysis, in mL

Figure 2 Lalab
Figure 3 Purok 2, Libay