1. The Effects of Fracking Fluid on Algal Growth
John Peilert
Grade 11
Central Catholic High School

2. Problem
Do chemicals used in Fracking have a negative effect on the algae?

3. Euglena gracilis Algal-like protist
Survives in both fresh and salt water
Performs photosynthesis and endocytosis
Becomes dominant in negative conditions
No cell wall
Common experimental model

4. Chlamydomonas reinhardtii
Flagellated unicellular green algae
Commonly found in fresh water
Generally light sensitive
No cell wall
Common experimental model

5. Fracking Background
Fracking is used to access natural gas from shale that is deep underground.
Hydraulic fracturing is effective, however it may be unhealthy for the environment.
The root of the problem is the chemical solution used in mining the gas.
Are living organisms at risk from fracking?

6. Fracking Fluid Ingredients
Due to strict regulations, couldn’t obtain actual fluids, a researcher at CMU provided a simulation formula mix which was created along with the help of a chemistry instructor

7. Hypothesis
The Fracking fluid will significantly affect algal growth, especially in higher concentrations.
Null: the Fracking fluid will not significantly affect algal growth in either Chlamydomonas reinhardtii or Euglena gracillis.

8. Materials Euglena gracillis Chlamydomonas reinhardtii
Test tubes (borosilicate culture tubes)
Test tube rack
Carolina Spectrophotometer
Shop light
Pipettes
Soil Water and Spring Water
Fracking chemical solution

9. Procedure
32 tubes were filled to the following concentrations. (4 replicates) 0%
.2%
20%
40%
Algae
2ml
Soil Water
1ml
Spring Water
1.9ml
0ml
Fracking Fluid
0.1ml
Total
5ml

10. Procedure (cont.)
The tubes were placed equidistant from a fluorescent shop light and kept at 22˚C
The tubes were then inverted and had their absorbency read with the Carolina spectrophotometer at 430mn.
This process was repeated every day for fourteen days.

11. Effects of Fracking Fluid on Euglena
Absorbency
P-value<1e-8
P-value<1e-8
Day

12. Effects of Fracking Fluid on Chlamydomonas
Absorbency
P-value<1e-8
P-value<1e-8
Day

13. Statistical Analysis ANOVA
-Compares variation within groups to variation between groups.
-If the p-value is <0.05, than the null hypothesis can be rejected.
Dunnett’s Test
-Compares each group to the control individually.
-That result is then compared to a t-crit to determine significance

14. Dunnett's Test All values are greater than the t-crit of 3.29
Concentration
Day 6 Euglena
Day 6 Chlamydomonas
Day 14 Euglena
Day 14 Chlamydomonas
0.2%
9.175
17.473
8.922
19.976
20%
23.354
27.761
13.951
25.528
40%
33.362
35.273
18.655
30.415
All values are greater than the t-crit of 3.29

15. Conclusion
The ANOVA suggests that the null should be rejected with a high level of confidence.
The Dunnett's test shows that every concentration significantly affected algal growth.

16. Limitations Number of replicates Environmental regulation/monitoring
Only one exposure time
Only two models tested
Only tested for growth
Only one type of exposure

17. Expansion Elodea model to test oxygen production
Synergistic effects of multiple variables
Longer exposure time

18. References http://www2.centralcatholichs.com/extracurr icular
uglena.htm
fracturing-fluids/

19. Day 6 ANOVAs Euglena Chlamydomonas
-Interpretation: Both ANOVAs have a p-value close to 0. This indicates that
there is variance in both data sets.

20. Day 14 ANOVAs Euglena Chlamydomonas
-Interpretation: Both ANOVAs have a p-value close to 0. This indicates that
there is variance in both data sets.