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

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

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

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

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?

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

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.

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

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

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.

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

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

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

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

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.

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

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

References http://www2.centralcatholichs.com/extracurr icular http://www.chlamy.org/info.html http://www.fcps.edu/islandcreekes/ecology/e uglena.htm http://geology.com/energy/hydraulic- fracturing-fluids/

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.

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.