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EFFECT OF LEAD (Pb)
EXPOSURE ON SKIN
ELASTICITY
CHAD-2010
ECOLE PRIVEE
INTERNATIONALE TCHADO-TURQUE
EDITOR and ADVISORY FAYSAL YILMAZ
NTIC Head of Biology Department [email protected]
ASSOCIATE EDITORS
Dayanat Mammadov
EPITT Computer Teacher
Yousouf Daoussa Deby Itno EPITT Student
(4emeClass) Younous Oumar Souni
EPITT Student (4emeClass)
Haggar Bechir Ali Abdramane Haggar EPITT Student
(5emeClass) Idriss Ramadan Erdebou
EPITT Student (4emeClass)
The Effect of Lead (Pb) Exposure on Skin Elasticity
ABSTRACT The goal of this project is to determine whether exposing chicken skin to lead metal for an extended period of time (1 week) in DI water will change the Young’s modulus of the skin. The aim of using lead in this experiment is to be able to apply any findings to real-world situations in which people become exposed to lead through their water pipes. If it is found that lead does have adverse effects on chicken skin, the findings can be used as a basis for further testing with human skin so that the negative health effects of lead can be further understood. This greater understanding could be used to generate more awareness of the possible harms of lead exposure and to develop treatments against lead poisoning. Based on literature findings on the composition of collagen and elastin fibers in the skin, it is hypothesized that lead exposure will make the skin more fragile and prone to breakage than normal, resulting in a lower Young’s modulus than unexposed skin. By comparing the Young’s moduli of the lead-exposed skin samples to the Young’s moduli of unexposed samples using a one-tailed, unpaired t-test in Excel, p<0.05 did reveal that the Young’s modulus of the exposed samples were significantly less than that of the unexposed samples.
INTRODUCTION
The goal of this project is to determine whether exposure to lead can change the elasticity of chicken skin, as roughly measured by Young’s modulus. It is widely known that the extracellular matrix of skin (and many other tissues) contains numerous collagen and elastin fibers. The fibers are constructed from individual collagen or elastin molecules which are covalently cross-linked. The cross-links form between the side chains of different lysine amino acids in the molecules and lend a higher tensile strength to the fibers and hence to the skin.
The lysine side chains have terminal –NH3+ groups which join by forming a bond
between the N atoms (1). In the process, H atoms are given up. However, if a sufficient number of these bonds are broken, the skin will become fragile and tear-prone (1). In order to break the bonds, H atoms must be reintroduced through a reduction reaction. One possible way of accomplishing this reduction might be to expose the tissue to lead, which can undergo the reaction Pb→Pb+2 + 2e-, thereby providing electrons to initiate the reduction reaction and break the bonds.
This project was developed to protect the public health by minimizing lead levels in drinking water. If water is too corrosive, it can cause lead to leach out of the plumbing materials and fixtures and enter the drinking water. Children are especially susceptible to high levels of lead, which can cause damage to the brain, red blood cells, and kidneys. Exposure to low levels of lead can cause low IQ, hearing impairment, reduced attention span and poor classroom performance. High exposure to copper can cause stomach and intestinal distress, liver and kidney damage, and complications from Wilson’s disease in genetically-predisposed people. High lead levels in adults have been linked to high blood pressure. Pregnant women and their fetuses are especially vulnerable to lead exposure, which can significantly harm the fetus, cause low birth weight and slow down normal mental and physical development.
Using lead could lend the experiment a practical objective, since exposing chicken
skin to lead in this experiment could simulate the exposure to lead that some people get from their tap water if the pipes or solders in their homes are old (pre-1987) and lead-containing (City of N`DJAMENA/Chad Water Department). Thus, if it is found that such exposure has the capability of changing the mechanical properties of the skin, further experiments could be done to determine whether human skin is also at risk from such exposure. In this project unexposed skin was used for comparison of the exposed skin.
EQUIPMENTS Major Equipment
Handmade Instron - The Instron was used to stretch the lead-exposed skin samples. The Excel software will record force/displacement data for each sample.
Lab Equipment
2 Large plastic containers- This was required to be large enough to soak chicken legs wrapped in lead sheeting prior to the actual experiment. Ten legs will need to be soaked in each container. Surgical scissors- These were used to separate the skin from the legs and cut out samples from the skin of set dimensions. Knife/scalpel- These were needed to help with cutting the skin off of the leg itself and to cut the skin into sample pieces. Like the marker, they could also be used to make small markings (in the form of cuts) where the skin should then be cut with the scissors. Forceps- These were useful for peeling the skin off the legs and for holding the skin in place when the samples are being cut.
Cutting board- This was used as the surface on which the skin is stripped from the legs and cut into samples
Calipers- These were used to measure the thickness of the skin samples and the distance between the loading clamps on the Instron once samples are loaded.
Weight set (500g, 1kg, 2kg) - These were used to verify that the load cell transducer in the Handmade-Instron is outputting correct measurements by making measurements for known applied loads.
Incubator- It was used to stimulate the human body by adjusting the temperature as 37oC
Supplies
20 freshly slaughtered chicken - The skin was exposed to lead and will act as the actual test subject in this experiment. 20 legs are needed for each group to use 10 in an experiment.
20 1 ft. x 1 ft. (1 ft = 30.48 centimeters) pieces of 1/32 inch (1 inch 2.54 cm) 99% pure lead sheeting- These are needed to act as the source of lead exposure for the skin.
Gloves- These were worn by anyone handling the lead or meat to avoid contamination of their hands by germs from the meat or by lead, which, if ingested, can be harmful.
DI water- This was necessary to soak the lead-covered legs in before the experiment.
Paper towels wet with DI water- These was needed to store the skin samples after they have been cut and before they are loaded into the Instron to prevent their drying out, which could cause changes in their mechanical properties.
Microscopes & Fixed Human Skin Preparets- To be able to understand better the anatomy of human skin, some specimens were observed unde microscopes.
METHODS
1. One week before the lab was to be run, separate the legs from corpse and wrap each of 10 chicken legs tightly in a 1 ft. x 1. ft. piece of lead sheet (maximize the amount of skin that is directly in contact with the lead, especially the region of skin just above the knee). Next, submerge all of the wrapped and non-wrapped legs in two large plastic containers independently and fill with DI water. Set the containers in an incubator (37oC) and let it sit for a week. BE SURE TO WEAR GLOVES DURING THIS STEP AND ALL STEPS IN WHICH THE CHICKEN SKIN OR LEAD IS HANDLED.
1a- Leg separation from corpse
1b- Lead sheet preparation
1c- Wrapping the legs
1d- Submerged two groups of legs
1e- Filling the containers with DI water
1f- Ready legs
1g- Putting the legs in incubator
1h- One week at 37oC
2. On the day of the experiment, set up the handmade-Instron and calibrate it. Generally:
• Verify the load transducer settings using the provided set of known weights. • Make sure the Instron is set to move up, or in other words, to stretch the samples.
2a- Main components of handmade-Instron
2b- Setting up the Instron
2c- Loading known weighs
2d- Stretching the sample
3. Remove the lead from the chicken legs and the skin from the legs. Cut the skin off so that the part above
the outside of the knee remains intact. While not working with the skins, place them in wet paper towels. Make sure you note in what orientation you place them.
3a- Dissection unit
3b- Getting ready for removing the skin
3c- Wetting the paper towel
3d- Cutting the skins off
3e- Placing them on wet paper towels
3f- Skin pieces are ready
4. Cut 2 samples of skin from each of the 20 specimens. The samples should be taken from just above the
outside of the knee and have dimensions of 2 cm x 4 cm (determined to be appropriate for Instron testing). Use the calipers to make measurements. Place the samples back in the paper towels until testing occurs.
4a- Cutting sample skins
4b- Using calipers
4c- Prepared GI (no lead) skins on towel
4d- Prepared GII (with lead) skins on towel
5. Taking one sample, clamp it in the Instron with the 2 cm sides in the clamps. Using the calipers,
measure the distance between the top of the bottom clamp and the bottom of the top clamp. This will act as the gage length in strain calculations and should be kept constant for all samples. Also, measure the width of the sample in the clamps and the thickness near the center of the sample. Multiply these to find the cross-sectional area of the sample. This was used to calculate stress. Pull the dynamometer for different weighs and measure the distance. Record all data.
5a- Clamping the skin
5b- Measuring the thickness & width
6. Pull the dynamometer for different weighs (10-20-30 N) and measure the strain. Record all data and
repeat it for all 40 samples.
6a- Loading sample
6b- Loading weighs
6c- Measuring the strain
6d- Recording data
ANALYSIS 1. Move the raw force (N) and displacement (mm) data for each sample into an Excel worksheet. Divide
all the force data of a sample by the cross-sectional area of that sample and then divide by 1000 to calculate stress data in kPa. Divide the displacement data of the sample by the gage length to obtain stress data. Repeat for all samples. (Stress=Force/Area)
2. Find the Young’s modulus (E) for each of the skin samples. This was the slope of the fit line for the sample. (E= Stress/Strain)
3. Compare the Young’s moduli of the lead-exposed skin samples to the Young’s moduli of unexposed samples using a one-tailed, unpaired t-test in Excel. Since the variance of the Young’s modulus of the unexposed samples is much more than 5% of their mean, unequal variance is assumed. A p<0.05 will reveal that the Young’s modulus of the exposed samples is significantly less than that of the unexposed samples.
YOUNG MODULUS = STRESS/STRAIN
E= (F/A) / (∆L/L)
RESULTS Experimental Results
STRESS (F/A)
STRAIN (∆L/L)
YOUNG MODULUS (E=STRESS/STRAIN)
SAMPLE NUMBER
THICKNESS (mm)
WIDTH (mm)
CROSS SECTIONAL
AREA (A)(mm2)
1ST 10/A
2ND 20N/A
3RD
30N/A
GAGE LENGTH L0(mm)
1ST
EXTENT LENGTH ∆L(mm)
2nd
EXTENT LENGTH ∆L(mm)
3rd
EXTENT LENGTH ∆L(mm)
(E1) (E2) (E3) MEAN (E)
GROUP‐I
NO LEAD
1 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.80 1.70 3.20 21.88 20.59 16.41 19.62
2 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.60 1.50 3.00 19.44 15.56 11.67 15.56
3 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.75 1.56 3.10 20.74 19.94 15.05 18.58
4 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.80 1.70 3.20 21.88 20.59 16.41 19.62
5 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.75 1.56 3.10 20.74 19.94 15.05 18.58
6 1.10 20.00 22.00 0.45 0.91 1.36 28.00 0.65 1.60 3.40 19.58 15.91 11.23 15.57
7 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.60 1.50 3.00 19.44 15.56 11.67 15.56
8 1.13 20.00 22.57 0.44 0.89 1.33 28.00 0.68 1.58 3.24 18.24 15.70 11.49 15.14
9 1.16 20.00 23.29 0.43 0.86 1.29 28.00 0.61 1.50 2.90 19.71 16.03 12.44 16.06
10 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.60 1.50 3.00 19.44 15.56 11.67 15.56
11 1.24 20.00 24.71 0.40 0.81 1.21 28.00 0.54 1.23 2.84 20.98 18.42 11.97 17.12
12 1.27 20.00 25.43 0.39 0.79 1.18 28.00 0.50 1.00 2.20 22.02 22.02 15.02 19.69
13 1.31 20.00 26.14 0.38 0.77 1.15 28.00 0.20 0.80 1.95 53.55 26.78 16.48 32.27
14 1.34 20.00 26.86 0.37 0.74 1.12 28.00 0.19 0.75 1.60 54.87 27.80 19.55 34.07
15 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.80 1.70 3.20 21.88 20.59 16.41 19.62
16 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.75 1.56 3.10 20.74 19.94 15.05 18.58
17 1.00 20.00 20.00 0.50 1.00 1.50 28.00 0.70 1.20 2.90 20.00 23.33 14.48 19.27
18 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.75 1.56 3.10 20.74 19.94 15.05 18.58
19 1.10 20.00 22.00 0.45 0.91 1.36 28.00 0.65 1.60 3.40 19.58 15.91 11.23 15.57
20 1.30 20.00 26.00 0.38 0.77 1.15 28.00 0.21 0.90 1.96 51.28 23.93 16.48 30.57
STRESS (F/A)
STRAIN (∆L/L)
YOUNG MODULUS (E=STRESS/STRAIN)
GROUP‐II WITH LEAD
SAMPLE NUMBER
THICKNESS (mm)
WIDTH (mm)
CROSS SECTIONAL
AREA (A)(mm2)
1ST 10/A
2ND 20N/A
3RD
30N/A
GAGE LENGTH L0(mm)
1ST
EXTENT LENGTH ∆L(mm)
2nd
EXTENT LENGTH ∆L(mm)
3rd
EXTENT LENGTH ∆L(mm)
(E1) (E2) (E3) MEAN (E)
22 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.42 0.82 1.60 41.67 42.68 32.81 39.05
23 1.10 20.00 22.00 0.45 0.91 1.36 28.00 0.25 0.74 1.40 50.91 34.40 27.27 37.53
24 1.10 20.00 22.00 0.45 0.91 1.36 28.00 0.25 0.74 1.40 50.91 34.40 27.27 37.53
25 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.35 0.78 1.55 44.44 39.89 30.11 38.15
26 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.42 0.82 1.60 41.67 42.68 32.81 39.05
27 1.30 20.00 26.00 0.38 0.77 1.15 28.00 0.12 0.46 0.98 89.74 46.82 32.97 56.51
28 1.40 20.00 28.00 0.36 0.71 1.07 28.00 0.08 0.42 0.85 125.00 47.62 35.29 69.30
29 1.10 20.00 22.00 0.45 0.91 1.36 28.00 0.25 0.74 1.40 50.91 34.40 27.27 37.53
30 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.26 0.90 1.90 44.87 25.93 18.42 29.74
31 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.42 0.82 1.60 41.67 42.68 32.81 39.05
32 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.35 0.78 1.55 44.44 39.89 30.11 38.15
33 1.00 20.00 20.00 0.50 1.00 1.50 28.00 0.30 0.75 1.20 46.67 37.33 35.00 39.67
34 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.26 0.90 1.90 44.87 25.93 18.42 29.74
35 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.35 0.78 1.55 44.44 39.89 30.11 38.15
36 0.90 20.00 18.00 0.56 1.11 1.67 28.00 0.35 0.78 1.55 44.44 39.89 30.11 38.15
37 1.30 20.00 26.00 0.38 0.77 1.15 28.00 0.12 0.46 0.98 89.74 46.82 32.97 56.51
38 1.20 20.00 24.00 0.42 0.83 1.25 28.00 0.26 0.90 1.90 44.87 25.93 18.42 29.74
39 1.00 20.00 20.00 0.50 1.00 1.50 28.00 0.30 0.75 1.20 46.67 37.33 35.00 39.67
40 0.80 20.00 16.00 0.63 1.25 1.88 28.00 0.42 0.82 1.60 41.67 42.68 32.81 39.05
Table: Results of experiment and data analyzing
Statistical t-Test Results
The results of unpaired t-test
t= 7.91 Standard deviation= 8.08 Degrees of freedom = 38
Group A: Number of items= 20 29.7 29.7 29.7 29.7 37.5 37.5 37.5 38.1 38.1 38.1 38.1 38.1 39.0 39.0 39.0 39.0 39.7 56.5 56.5 69.3 Mean = 40.0 95% confidence interval for Mean: 36.33 thru 43.65 Standard Deviation = 9.92 Hi = 69.3 Low = 29.7 Median = 38.1 Average Absolute Deviation from Median = 5.43
Group B: Number of items= 20 15.1 15.6 15.6 15.6 15.6 15.6 16.1 17.1 18.6 18.6 18.6 18.6 19.3 19.6 19.6 19.6 19.7 30.6 32.3 34.1 Mean = 19.8 95% confidence interval for Mean: 16.12 thru 23.43 Standard Deviation = 5.69 Hi = 34.1 Low = 15.1 Median = 18.6 Average Absolute Deviation from Median = 3.42
Degrees of
Freedom
Probability, p
0.1 0.05 0.01 0.001
28 1.70 2.05 2.76 3.67
29 1.70 2.05 2.76 3.66
30 1.70 2.04 2.75 3.65
40 1.68 2.02 2.70 3.55 60 1.67 2.00 2.66 3.46
120 1.66 1.98 2.62 3.37
infinity 1.65 1.96 2.58 3.29
CONCLUTION Our t value (7.91) was bigger than the table value (2.03). That proves lead exposed chicken skins had
lost their flexibility by showing low Young module values. It was known the effects of lead on internal organs, but any scientific research about skin was not done before. We hope this project would help to understand the effects of lead exposure better and to aware people who still use lead pipe in their installation.
References
(1) Alberts, Bruce, et al. (2002). Molecular Biology of the Cell. (4th ed., pp. 1099,1102). New York: Garland Science.
(2) Zumdahl, Steven S. (2005). Chemical Principles. (5th ed., pp. 468). (3) Boston: Houghton Mifflin Company. (4) Bains, A., Baumann, B., Connie, C., & Wang, E. (2007). Tensile Testing: Elastic Properties. (5) Meeting the Lead Standards. City of Philadelphia Water Department. Retrieved April 23,
2007 from http://www.phila.gov/water/water_quality2.html#bk_leadstandard.