ARTICLE
Scanner image analysis to improve the EZ arsenic test kit from Hach
Mattias Frid
Received 11 December 2006
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The EZ arsenic test kit from Hach Company is used to determine arsenic levels in water and soil
samples around the world. Two of the advantages is that the test is cheap compared to advanced
lab equipment and that it is designed as a field test kit and therfore is fast and easy to use. The big
disadvantage is that the detection limit is poor because of individual visual determination of the
color on the test strip. This experiment set out to find a way to more accurate determine the color
on the test strip by scanning it and letting a software decide the RGB-values. Two calibration
curves were made. One for the concentration range 0-20 µg L-1 and one for 20-100 µg L-1. These
two were chosen because 10 µg L -1 and 50 µg L-1 are the concentration levels used to determine if
the water from a well is suitable or not for drinking in the US respectively Bangladesh. It is also
the ranges that is hardest to determine with the test kit because of the weak color. The experiment
showed that the low concentrations can be determined more accurate with the method of scann ing
the test strip compared to individual visual determination. The experiment also showed that there
was a significant improvement with using the 24 hour test compared to the 30 min test for the
lowest concentration range 0-20 µg L-1. The importance of chosing a good scanner became
obvious when comparing the result from the Epson scanner with the HP scanner.
Introduction
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There are several methods of detecting arsenic in ground
water and soil. Some of the more sophisticated ones are the
ones that are made in a lab with techniques such as ICP-MS,
ICP-AES, HPLC-ICP-MS, TOF SIMS and X-Ray
Fluorescence (1). But this is not an option for countries in the
third world where money is an issue. It is also important to
have a quick and reliable test out on the field so that a quick
decision can be made for water wells. One visual method for
detection of arsenic is the test kit from Hach. It is an
inexpensive (compared to the techniques mentioned above)
and quick method to determine if arsenic is present in for
example ground water or pressure treated wood.
In the test kit, sulfamic acid and powdered zinc reacts to
create strong reducing conditions in which inorganic arsenic is
reduced to arsenic gas (AsH 3). The arsine gas then reacts with
mercuric bromide impregnated test paper to form mixed
arsenic/mercury halogenides. These compounds discolor the
test strip depending upon the concentration of the arsenic in
the sample. The color change is from white to yellow to tan
to brown. Most tests for arsenic, including the Hach method,
rely on the conversion of arsenic to arsine gas.
As already described, the Hach Arsenic Test Kit uses a
colored test strip to determine how much arsenic there is
present in the sample. This is a highly individual way of
determine the arsenic level, because it is up to the observer to
decide “how yellow” or “how orange” the test strip is. The
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The EZ arsenic test kit from Hach has a calibrated color
map with the semi-quantitative steps of 0, 10, 30, 50, 70, 300
and 500 µg L-1. This is a wide range, especially if considering
that the World Health Organization recommends a maximum
contaminant level of 10 µg L -1 and the steps in the Hach
method are 0, 10 and 30 around the recommended 10 µg L -1level. The accepted arsenic concentration level used to
determine if a water well is consumable or not in Bangladesh
is 50 µg L-1. This project aims to evaluate a method to make
the color map more sophisticated by developing a calibration
curve so the arsenic concentration can be determined more
accurate in the two concentration levels of interest, 10 ppb
and 50 µg L-1 (2, 3).
Experimental
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Department of Chemistry, University of Massachusetts,USA
E-mail: frid@chem.umass.edu
aim of this project is to find a better way to determine the
arsenic level in a sample using the EZ arsenic test kit. The
method that will be evaluated is to use a scanner to scan the
test strip and then use software to determine exactly for
example how yellow or how white the test strip actually is by
making calibration curves of the RGB-values. This way the
software will decide it and the human factor decreases.
All experiments performed with the Hach Arsenic test kit
were carried out according to the procedure outlined in the
test kit except for the 24 hour test that was longer than the
instructions which are 30 min.
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Instrumentation
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The EZ arsenic test kit uses a reaction vessel with a special
cap that focuses the arsine gas on the test strip and therefore
forces the arsine gas to react with the mercuric bromide on the
test strip. This minimizes the toxic arsine gas that leaves the
vessel and leaves almost none exposure to the operator.
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Adobe Photoshop by using bicubical interpolation to 1x1
pixels which gave the average of all the pixels in the image.
The RGB-value was determined with the RGB-value function
in Adobe Photshop.
Results and discussion
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30 min test
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The average results from the three 30 min experiments
scanned with the Epson scanner at 600 dpi can be seen in Figure
1 and it is clear that the arsenic concentration do not have
sufficient impact on the red and the green color in lower
concentrations around 10 µg L -1 and 50 µg L-1. The same
experiment with the HP ScanJet 5300c at 300 dpi can be seen
in Figure 2.
Fig. 1. The EZ arsenic test kit from Hach ().
0-250 ppb As(III) Epson 600 dpi - 30 min
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The test kit comes with two reaction vessels, a container with
100 test strips and a color map, two bags of reagents for 100
tests.
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In the EZ arsenic test kit, arsine gas is generated by
reduction with zinc metal and a strong acid. Strong acids are
dangerous and difficult to work with, especially if the work
cannot be done in the lab. The Hach test kit uses powdered
reagents for safety. For example, the sulfamic acid (H3NSO3)
is in powdered form. This minimizes the reagent hazard. The
zinc powder is packaged in a unit dose form for convenience
and to minimize handling. Lead acetat solution for filtration
of interfering hydrgogen sulfide gas is also available in the
test kit.
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Red
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Green
Blue
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Fig. 1. RGB-values for 0-250 µg L-1 with the Epson scanner after 30 min
reaction.
0-250 ppb As(III) HP 300 dpi - 30 min
Procedure
Standard solutions of arsenic (III) were prepared from
NaAsO 2 (s) and deionized water in the following
concentrations: 5, 10, 15, 20, 25, 50, 100 and 250 µg L -1.
Nine vessels were filled with 50 ml of each concentration.
Sulfamic acid powder (from unit dose) was added to the
vessels that were swirled for 1 min to get the acid solved in
the solution. Then the zinc powder was added (also from unit
dose) and the reaction that converts arsenic to arsine gas
started. The reaction was held for 30 min and the vessels
were gently swirled after 10 and 20 min during the reaction
time. After that the test strips were removed and scanned in
two different scanners (HP ScanJet 5300c and Epson). The
experiment was repeated 3 times in two days and after the
second round the vessels were left over night to react for 24 h.
The images from the scanners were processed in Adobe
Photoshop. The colored circle on the test strip allowed a
picture of 3x3 mm to be cut out without interference from the
fade edges of the ring. For the HP scanner a 300 dpi (dots per
inches) resolution was used and for the Epson scanner a 600
dpi resolution was used. Since 1 inch is 25.4 mm, the image
size from the HP scanner was 1250 pixels and from the Epson
scanner 5000 pixels. The image was then diminished in
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Conc. As(III) (ppb)
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RGB-values
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RGB-values
Reagents
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Red
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Green
Blue
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Conc. As(III) (ppb)
Fig. 2. RGB-values for 0-250 µg L-1 with the Epson scanner after 30 min
reaction
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After the decision to work with the blue color, a magnified
graph around the two concentration ranges of interest was
made. Figure 3 shows the graph for the blue color around 10
ppb when the test strip was scanned at 600 dpi with the Epson
scanner and Figure 4 shows the result from the HP ScanJet
5300c at 300 dpi.
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Blue average 0-20 ppb - Epson 30 min 600 dpi
Blue average 20-100 ppb - Epson 30 min 600 dpi
y = -1,1x + 253,4
R2 = 0,9302
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Blue value
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Blue value
y = -1,2669x + 248,51
R2 = 0,9716
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Conc. As(III) (ppb)
Conc. As(III) (ppb)
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Fig. 5. Blue value with max and min values for each concentraion at the
range 20-100 µg L-1 with the Epson scanner after 30 min reaction.
Fig. 3. Blue value with max and min values for each concentraion at the
range 0-20 µg L-1 with the Epson scanner after 30 min reaction.
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y = -0,64x + 250
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R = 0,8889
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Blue average 20-100 ppb - HP 30 min 300 dpi
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y = -0,7537x + 246,99
R2 = 0,9735
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Blue value
Blue value
Blue average 0-20 ppb - HP 30 min 300 dpi
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Conc. As(III) (ppb)
Conc. AS(III) (ppb)
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One obvious conclusion was made when looking at the two
graphs for the 30 min test scanned with the Epson and the HP
scanner. The Epson scanner gave a better result because, for
example, it could distinguish between the blue value
corresponding to 0 µg L -1 and the blue value corresponding to
5 µg L-1 which the HP scanner can not. The correlation
coefficient was 0.965 for the Epson graph and 0.943 for the
HP graph, so the line between the dots was straighter for the
graph made with the Epson scanner. The slope on the Epson
graph was steeper than the HP-graph, -1.10 µg L-1 per blue
value for the Epson and 0.64 µg L -1 per blue value for the HP.
When the same evaluation was made with the concentration
range around 50 µg L -1 (20-100 µg L-1), the pattern continued.
In Figure 5 and 6 it can be seen that the Epson scanner gives a
deeper slope (-1.3 µg L-1 per blue value) than the HP scanner
(-0.75 µg L-1 per blue value) but between these concentrations
the correlation coefficient was the same (0.98).
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Fig. 6. Blue value with max and min values for each concentraion at the
range 20-100 µg L-1 with the HP scanner after 30 min reaction.
A wider concentration range (0-100 µg L-1) can be seen for
the Epson scanner (Figure 7) and the HP scanner (Figure 8),
but because of the variations and the digression from a
straight calibration curve it is more accurate to construct a
calibration curve in smaller intervals.
The suggested
concentration ranges for the two calibration curves are 0-20
µg L-1 and 20-100 µg L-1.
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Blue average 0-100 ppb - Epson 30 min 600 dpi
y = -1,3328x + 253,61
R2 = 0,9803
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Blue value
Fig. 4. Blue value with max and min values for each concentraion at the
range 0-20 µg L-1 with the HP scanner after 30 min reaction.
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Conc. As(III) (ppb)
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Fig. 7. Blue value with max and min values for each concentraion at the
range 0-100 µg L-1 with the Epson scanner after 30 min reaction.
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Blue average 0-100 ppb - HP 30 min 300 dpi
0-250 ppb As(III) HP 300 dpi - 24 h
y = -0,7953x + 250,24
R2 = 0,9795
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RGB-values
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Blue value
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Red
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Green
Blue
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Fig. 8. Blue value with max and min values for each concentraion at the
range 0-100 ppb with the HP scanner after 30 min reaction.
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Conc. As(III) (ppb)
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Fig. 10. RGB-values for 0-250 µg L-1 with the HP scanner after 24 h
reaction.
24 hour test
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The average results from the three 24 hour experiments
scanned with the Epson scanner at 600 dpi can be seen in
Figure 9 and ones again it is clear that the arsenic
concentration do not have sufficient impact on the red and the
green color in lower concentrations around 10 ppb and 50
ppb. Same experiment with the HP ScanJet 5300c at 300 dpi
can be seen in Figure 10.
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After the decision to work with the blue color for the 24
hour test as well, a magnified graph around the two
concentration ranges of interest was made. Figure 11 shows
the graph for the blue color around 10 µg L -1 when the test
strip was scanned at 600 µg L -1 with the Epson scanner and
Figure 12 shows the result from the HP ScanJet 5300c at 300
µg L-1.
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Blue average 0-20 ppb - Epson 600 dpi 24 h
0-250 pp As(III) Epson 600 dpi - 24 h
y = -5,64x + 255,8
R2 = 0,9536
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Blue value
RGB-values
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Red
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Green
Blue
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Fig. 11. Blue value with max and min values for each concentraion at the
range 0-20 µg L-1 with the Epson scanner after 24 h reaction.
Conc. As(III) (ppb)
Fig. 9. RGB-values for 0-250 µg L-1 with the Epson scanner after 24 h
reaction.
Blue average 0-20 ppb - HP 300 dpi 24 h
y = -3,38x + 250,6
R2 = 0,9575
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Blue value
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Conc. As(III) (ppb)
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Conc. As(III) (ppb)
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Fig. 12. Blue value with max and min values for each concentraion at the
range 0-20 µg L-1 with the HP scanner after 24 h reaction.
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The blue color for the 24 hour test shows an interesting
improvement in the concentration range 0-20 µg L-1 compared
to the 30 min test. The slope of the calibration curve for the
HP scanner improved (became steeper) from -0.64 µg L-1 per
blue value to -3.38 µg L-1 per blue value and the correlation
coefficient improved from 0.94 to 0.97. For the Epson
scanner the slope improved from -1.1µg L-1 per blue value to 5.64 µg L-1 per blue value and the correlation coefficient
improved from 0.96 to 0.98.
The same evaluation was made with the concentration
range around 50 µg L -1 (20-100 µg L-1). In this concentration
range, the 24 hour test did not show any improvement. The
slope for the HP scanner decreased from -0.75 µg L-1 per blue
value to -0.55 µg L-1 per blue value (Figure 13). The Epson
scanner showed a decrease in slope from -1.1 µg L-1 per blue
value to -1.05 µg L-1 per blue value (Figure 14). The
correlation coefficient for the HP scanner was almost the same
(0.98) and for the Epson scanner it was the same as well
(0.98).
Blue average 0-100 ppb - Epson 600 dpi 24 h
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Blue average 0-100 ppb - HP 300 dpi 24 h
y = -1,0044x + 223,25
R2 = 0,7553
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Blue average 20-100 ppb - HP 300 dpi 24 h
y = -0,5533x + 192,23
R2 = 0,9608
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Fig. 16. Blue value with max and min values for each concentraion at the
range 0-100 µg L-1 with the HP scanner after 24 h reaction.
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Conc. As(III) (ppb)
Fig. 13. Blue value with max and min values for each concentraion at the
range 20-100 µg L-1 with the Epson scanner after 24 h reaction.
Blue value
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Fig. 15. Blue value with max and min values for each concentraion at the
range 0-100 µg L-1 with the Epson scanner after 24 h reaction.
y = -1,0491x + 159,9
R2 = 0,9686
Conc. As(III) (ppb)
A test was done with the Epson scanner to see if there was
any improvement of using 600 dpi instead of 300 dpi. The
RGB-values were exactly the same for the two resolutions so
no improvement could be seen. Therefore it is no use
spending extra time to scan with a higher resolution.
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Conclusions
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Conc. As(III) (ppb)
Fig. 14. Blue value with max and min values for each concentraion at the
range 20-100 µg L-1 with the Epson scanner after 24 h reaction.
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Conc. As(III) (ppb)
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y = -1,7929x + 211,05
R2 = 0,7825
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Blue value
Blue value
Blue average 20-100 ppb - Epson 600 dpi 24 h
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to construct a calibration curve in smaller intervals. The
concentration range 20-100 µg L- 1 gave not any improvement
with the 24 hour test compared to the 30 min test but the
lower concentration range did.
Blue value
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A wider concentration range (0-100 µg L-1) can be seen for
the Epson scanner (Figure 15) and the HP scanner (Figure 16)
for the 24 hour test. But because of the variations and the
digression from a straight calibration curve it is more accurate
The suggested approach with scanning the test strip
compared to visual individual determination seems to be a
more accurate way of analysing low arsenic concentrations
with the test kit from Hach. First the individual error in
determine the color on the test strip is minimized and second
the concentration can be established with lower detection limit
since it is almost impossible to decide the color with
significant precision in the low concentrations by looking at
the test strip and compare it to the color map because of the
weak color.
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It seems to be of big importance to choose a good scanner.
The experiment clearly shows that the slope of the calibration
curve is steeper with the Epson scanner than the HP scanner.
The scanned images shows this in the way that the scanned
images from the Epson scanner is much clearer and brighter
than the ones from the HP scanner. A large improvement of
the 24 hour test contra the 30 min test could be seen in the
low concentration range 0-20 µg L-1, but not for the
concentration range from 20 to 100 µg L -1.
References
(1) Robert D Morrison; Brian L. Murphy. Environmental
Forensics. Academic Press. 2006. 89-106.
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(3) U.S. Environmental Protection Agency,
http://www.epa.gov/safewater/arsenic (accessed
10/11/06)
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Acknowledgements
The author thanks Professor Julian Tyson for his support and
for making this project possible. The author would also like
to thank Maura Mahar for her support in the lab.
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(2) Hach Company, www.hach.com (accessed 12/09/06)
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