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AS245: Bachelor in Science Applied Chemistry

Title: Analysis of Animal Tissue


Course: Spectrochemical Methods of Analysis
(CHM580)
Name: Mohamad Nor Amirul Azhar b. Kamis
Matrix no:

2014647344 (AS245 3S)

Date of experiment: 1st April 2015


Date of submission: 29th May 2015
Group members: 1. Nurnailah bt. Noorazlan
2. Nor Amirah bt. Ahmad Azuan
3. Noramira bt. Saad
It is my responsibility as a student of UiTM to adhere
to truthfulness and avoid dishonesty, fraud or deceit
of any type in connection with write up and conduct
of this experiment.
Signature:__________
Date:

TITLE:

Determination of mercury (Hg) in animal tissue (anchovies) by using


mercury analyser.

ABSTRACT:
The sample used is the anchovies that had been treated with wet digestion by using
strong acid to extract the analyte (mercury) from the sample. The experiment was
done by using the Mercury analyser to know that amount of mercury in the sample.
The sample has been done triplicate to ensure the accuracy and getting more
accurate data. In order to make sure the amount of mercury in samples is in the
range of standard solutions, the samples had been diluted with dilution factor of 20.
The average amount of mercury in the diluted triplicate samples is 4.432ppb with
relative standard deviation of 1.213. The actual average concentration of samples is
88.64ppb. Smaller relative standard deviation value indicates the more precise data
between the triplicates.

INTRODUCTION:
Fishes are important for a healthy diet. Fish such as anchovies is lean, high-quality
protein, essential nutrients, low saturated fat, contain omega-3 fatty acids and lowcalorie source of protein for a good diet. However, some of the fishes may contain
high amount of harmful chemical such as mercury that is not safe to be consumed in
high amount for human. Mercury enters atmosphere from natural sources estimated
1600 to 4000 tonnes each year. Mercury is generated by industries, burning fossil
fuels and as by-product of some bacteria. High amount of mercury consumed may
harm an unborn baby or young childs developing nervous system. According to
Food and Drug Administration (FDA) and the Environmental Protection Agency
(EPA), fishes that contain high level of mercury that is not safe to be consumed are
shark, swordfish, king mackerel and tilefish. For this experiment, the sample used
was anchovies that is known been contaminated with mercury. On average, by
calculation made by the FDA, the average amount of mercury in anchovies is

0.04mg/kg. 0.5ppm to 1.0ppm is considered safe amount of mercury to be


consumed by human according to EPA.
OBJECTIVES:
To perform wet digestion method to extract mercury out from sample anchovies and
determine the amount of mercury in anchovies.

EXPERIMENTAL:
A. Preparation of reagent:
1. Preparation of 3% HCl solution:
i.
30mL of concentrated HCl solution was pipetted into 1000mL
volumetric flask and distilled water was used to mark the solution
until the calibration mark.
2. Preparation of 0.2% NaBH4 in 0.05% NaOH
i.
0.125g NaOH powder and 0.5g NaBH4 was poured in a beaker and
was dissolved with distilled water. The mixture then was transferred
into a 250mL volumetric flask and distilled water was used to mark
up the mixture until the calibration mark.
B. Preparation of Hg standards:
1. 5mL of 100ppm Hg stock solution was pipetted into a 50mL volumetric
flask and 3% acetic acid solution was used to mark up the solution until
the calibration mark to make a 10ppm Hg solution.
C1V1= C2V2
100V1 = 10(50)
V1 = 5mL
2. 5mL of 10ppm Hg solution was pipetted into a 50mL volumetric flask and
3% acetic acid solution was used to mark up the solution until the
calibration mark to make a 1ppm (1000ppb) Hg solution.
3. 5mL of 1000ppb Hg solution was pipetted into a 50mL volumetric flask and
3% acetic acid solution was used to mark up the solution until the
calibration mark to make a 100ppb Hg solution.
4. 100ppb Hg solution was pipetted into 6 different 50mL volumetric flasks to
make the following concentration:

TABLE 1:Preparation of standard solutions


Standard

Volume of Hg

concentration, ppb
100ppb, (mL)
0
0
2
1
4
2
6
3
8
4
10
5
Sample calculation: standard 1

Blank
Standard 1
Standard 2
Standard 3
Standard 4
Standard 5

C1V1= C2V2
100V1 = 2(50)
V1 = 1.0mL
5. Each of the volumetric flasks was mark up with 3% acetic acid solution
until the 50mL calibration mark.

C. Preparation of sample:
1. Day 1:
a. The anchovies were cut to smaller size and were dried in oven at
110C overnight to remove water moisture in the tissue.
2. Day 2:
a. 1.017g (A), 1.0885g (B) and 1.1291g (C) of the dried anchovies was
weighed and was mixed with 7mL nitric acid, then was let stand
overnight with cover.
3. Day 3:
a. The mixture was heated until the red fume was observed.
b. The mixture was let to cool at room temperature.
c. 1mL of 70% H2O2 was added into the mixture and was reheating again
until the mixture become concentrated.
d. 5mL of the concentrated sample mixture then was diluted into 100mL
volumetric flask and was mark up with 3% acetic acid solution until the
calibration mark.
Dilution factor calculation:
100
DF
=
5
= 20

D. Operational instrument:
1. Select method, new method, element (Hg), NaBH 4, fill title.
2. Select setting, time (12s), replicate (3).
3. Select sampler, speed pump 1 (100), equation (linear through zero).
4. Select standard concentration, fill id (std 1-5 and blank), fill concentration
5.
6.
7.
8.

for standards and blank, unit (ppb or g/L).


Select method Ed, save as, name of method.
Select samp info, fill sample id (1-3), file, sample info file, save as (name).
Select manual, FIAS, calibration, result (on ribbon), on pump.
Analyse blank, standards and finally samples.

RESULT:
TABLE 2:
ID

Result from instrumental analysis

Entered

Calculated

Mean signal

concentration (g/L)

concentration

(Abs)

Blank
Standard 1
Standard 2
Standard 3
Standard 4
Standard 5

0.0
2.0
4.0
6.0
8.0
10.0

(g/L)
0.000
2.257
4.443
6.346
7.974
9.523

0.0000
0.0198
0.0390
0.0557
0.0700
0.0836

Sample 1
Sample 2

4.404
4.494

0.0386
0.0394

Sample 3

4.398

0.0386

Graph of entered concentration vs calculated concentration


10
9 f(x) = 0.95x + 0.33
R = 0.99
8
7

calculated concentration (g/L)

6
5
4
3
2
1
0
0

10

12

entered concentration (g/L)

FIGURE 1:

Graph of entered concentration versus calculated concentration

Graph of mean signal vs calculated concentration


0.09
0.08

f(x) = 0.01x - 0
R = 1

0.07
0.06
0.05
mean signal (g/L) 0.04
0.03
0.02
0.01
0
0

10

calculated concentration (g/L)

FIGURE 2:

Graph of mean signal versus calculated concentration

Calculation for samples:

Mean =

Mean =

N
N
4.404+ 4.494+ 4.398
3

= 4.432
Actual concentration of sample

= mean calculated concentration dilution factor


= 4.432 20
= 88.64ppb = 0.08864ppm

Standard deviation =

1
( x imean)2
N 1 i

Standard deviation =

1
2
2
2
[( 4.4044.432 ) + ( 4.4944.432 ) + ( 4.3984.432 ) ]
31

= 0.054

Relative standard devation, %RSD

standard deviation
mean

0.054
4.432

100%

100%

= 1.213
Mean weight of sample (instrumental)

= 0.08864mg/L 0.1L
= 0.008864mg

Mean sample weight (actual weight)

1.017+ 1.0885+ 1.1291


3

= 1.0782g = 1078.2mg
w/w percentage

0.008864 mg
1078.2mg 100

= 8.221 10-4
DISCUSSION:
Figure 1 shows the graph of entered concentration into the instrument versus the
calculated concentration from the instrument. From this graph, it can show how the
standard solutions were prepared. The value of calculated concentration of wellprepared standard solutions will be only minor error with the entered concentration. It
still needs improvement in the preparation of the standard solutions. The laboratory
skills like pipetting and diluting need improvement. In figure 2, graph of mean
absorbance signal versus calculated concentration of the standard solution. The
graph shows a straight line passing all the points showing the accurate reading. The
R2 value of 1 is the additional information of a best fit for all points are on the straight
line. The triplicates samples were ensure the precision by calculating the relative
standard deviation. The smaller value of the relative standard deviation indicates the
more precise data between the triplicates. The mean concentration of the triplicates
samples is 88.64ppb. This value falls within the safe amount of mercury to be
consumed by human. By referring to EPA, safe levels of mercury to be consumed
are between 0.5ppm to 1.0ppm of mercury in the fish. The mercury analyser
instrument is used in this experiment to analyse the mercury because it has wider
dynamic range can be achieved as compared to AAS.
The mercury analyser has two configurations which are employing simple atomic
fluorescence and employing gold amalgamation to pre-concentrate mercury prior to
measurement by atomic fluorescence. It can be concluded that, mercury can analyse
mercury analyte better compared to AAS.

CONCLUSION:

The concentration of mercury in anchovies sample is 88.64ppb that is equal to


0.08864ppm and the %w/w is 8.221 10 -4. Since the value is in the range of safe
amount, the anchovies are safe to be consumed.

REFERENCES:
1. Fish

Consumption

Advice,

Retrieved

April

14,

from

http://www.epa.gov/mercury/advisories.htm
2. What You Need to Know about Mercury in Fish and Shellfish, United States
Environmental

Protection

Agency,

Retrieved

April

14,

from

http://water.epa.gov/scitech/swguidance/fishshellfish/outreach/advice_index.c
m
3. Should I be concerned about mercury in fish and what fish are safe to eat?,
Retrieved

April

14,

tname=george&dbid=103
4. Mercury:
how
much

from
is

http://www.whfoods.com/genpage.php?
safe?,

Retrieved

April

15,

from

https://www.greenleft.org.au/node/17431
5. K. Scoffin (2013), Mercury Analyzer in the Laboratory, Retrieved April 16, from
http://ww.labcompare.com/10-Featured-Articles/133134-Mercury-Analyzer-inth-Laboratory/
6. D. Pfeil (2012), Mercury Analysis, Which Techmique is right for you?,
Retrieved

April

17,

from

http://info.teledyneleemanlabs.com/blog/bid/123607/Mercury-Analysis-WhichTechnique-is-right-for-you