Objective:
You will investigate the relationship between increasing seawater concentration of CO2 dissolved in seawater and the resulting decreasing seawater pH on the weight of the shells and skeletal elements of the animals that make up a coral reef. 

Background:
Since the beginning of the industrial revolution, about one third of the CO2 released by human activities has been absorbed by the world’s ocean which plays a key role in moderating climate change. Without the ocean absorbing CO2, the CO2 in the atmosphere would be much higher, potentially increasing the effects of global climate change.

The burning of fossil fuels for energy production, manufacturing, and transportation has driven the level of carbon dioxide in the atmosphere to levels 30% greater than prior to the industrial revolution. Higher atmospheric levels of carbon dioxide have resulted in increasing amounts of carbon dioxide dissolving in the ocean leading to the reduction in pH or increase in acidity of the ocean. It should be noted that the ocean is not acidic (pH < 7) however the trend in declining pH has been observed and is referred to as “ocean acidification”. When carbon dioxide (CO2) dissolves in seawater, it creates carbonic acid (H2CO3), which releases bicarbonate ions (HCO31-) and hydrogen (H+) ions into the water. The hydrogen ions make the seawater more acidic (lowering its pH) and promoting the dissolution of CaCO3.In addition, some of the hydrogen ions (H+)react with carbonate ions (CO32-) already in the seawater to create more bicarbonate (Figure 1). This process reduces the amount of carbonate (an important mineral for building shells) dissolved in the seawater.

Figure 1. The process of ocean acidification.

Thus, the impacts of ocean acidification and lower carbonate ion concentrations on marine ecosystems include reduced growth of organisms that form calcareous skeletons or shells, such as corals, pteropods (swimming snails) and bivalves (such as oysters, clams and mussels). This results in reducing the rate at which marine animals can build new skeletons and shells while simultaneously increasing the rate of dissolution of existing shells. 

In this lab, we will focus on coral reefs with their enormous biodiversity, productivity and susceptibility to human impact We can’s investigate the rate at which coral reefs are being built in the context of this lab.  We can however investigate the impact of elevated CO2 on the dissolution of existing reefs. The vast bulk of a coral reef is composed of CaCO3  in coral skeletons and the shells of a vast array of animals inhabiting the reef and we can simply expose a composite sample of a reefs skeletons and shells to increased CO2 (decreased pH).  We can obtain a representative sample of coral skeletons and animal shells in an unlikely place, parrotfish poop. Check out the link Parrotfish poop.

Check out the poop.
The beach sand we will use today was collected from the beaches of the Florida Keys by previous SMCM students.  Place a small sample (single layer of grains) in a weigh boat and observe it under a dissecting microscope.  List the organisms that you can identify.  This web site will help. Coral beach sand.

Experimental Procedure Week 1
Set up the microcosms:

1. We will compare 3 CO2 concentrations (pHs) with respect to the rate of coral reef dissolution using coral reef beach sand as a surrogate for the reef.
1. The high pH will be 8.2 representing the average pH of the oceans around 1900
2. The mid pH will be 8.0 representing the projected average pH in 2050 (assuming  complete compliance with the Paris accord)
3. The low pH will be 7.75 the projected global oceanic pH in 2100 with out CO2 reduction.
4. Values from Bopp et al., 2013
2. Each team will need three microcosms (250ml BOD bottles) use lab tape to label the bottles pH 7.75, pH. 8.00 and pH 8.2 along with your team’s computer number.
3. Into each bottle weigh 5.00 g of beach sand and record the exact weight on the label, in your notebook and in google sheets.  Ocean acidification google sheet.  Be certain that you are on the tab for your section.

Prepare acidified seawater

1. Go to one of the pH station to obtain your seawater samples for your experimental bottles.
2. Pour about 250 ml of seawater (35ppt) into a plastic beaker and with the help of the TA or your instructor, decrease the pH of your sample to 8.2.  To accomplish this task, you will pipette acidified seawater into your stable seawater sample while gently stirring the sample with the pH probe.
3. Once you have acidified your seawater, go back to your lab bench and carefully pour the sample over the sand in your pH 8.2  bottle (over fill the bottle and tap out any bubbles).
4. Stopper the BOD bottle with a tapered glass stopper leaving the excess water in place.
5. Repeat for the pH 8.00 and the pH 7.75, being very careful to add the acidified water to the appropriate bottle.

When all of your samples have been prepared, double check your labels and store them on the shelves at the back of the lab until next time.

Questions

While you wait your turn to process your samples, answer the questions on todays work sheet and submit it in the assignments tab on BB
Provide your answers in blue font color below each question.
This must be submitted before you leave lab today

Experimental Procedure Week 2
Determine the weight of the remaining coral and shell material in each treatment pH
Caution:  take great care, at every step below, to prevent sand loss.

1. Carefully decant most of the seawater from each BOD bottle.
2. Label a filter papers with your team number and the experimental pH.
3. Carefully weigh the filter paper and record its weight in your notebook and on the
   the Ocean acidification google sheet
4. Place the filter paper in a vaccume filtration apparatus (label side down) and reassemble it, as demonstrated.
5. Shake out as much sand as you can into the filter funnel
6. Now using a squeeze bottle of dionized water, rinse the remaining sand into the filter.
7. Using the vaccume pumps provided suck the water through the filter
8. Now rinse the filter funnel, sand and filter paper with dionized water – suck dry and repeat three times.
9. Fabricate a foil pas as demonstrated and label each with your group # and the pH
10. Disassemble the filter and very carefully transfer the filter with the sand into the appropriate aluminum foil pan.  Double check to insure the label reflects the pH of the sand treatment in the pan.
11. Repeat for each pH.
12. Carefully transfer the foil pans into a desicator.

Experimental Procedure Week 3
Final weighing an visualization of your  data
 (To Be Completed Today--Your ticket out the door!)
Procedures:

1. Weigh each of your filter papers with sand and transfer this data to the google sheet. Ocean acidification google sheet and input your data for the dry weights into the appropriate cells.  Check that all of the other data has been entred and that you have selecting the appropriate cells. Note: Colums for calculated data are included on the Google sheet. Do not enter thes calculated values on this spread sheet.  Calculate them on your own excel sheet.

1. Next,  download a copy of the data (Ocean acidification google sheet) onto the desktop of your computer as an .XLS spreadsheet.  Rename the spreadsheet with a proper file name.

 

1. Calculate the final weight of the sand by subtracting the filter paper weight from the total filter paper + sand for all of the samples on the data sheet.  This is your sample weigth at time t1

1. Calculate the percent change of all of the sand samples at all three pHs from Time T0 to T1  Use excel on the lab computers NOT Google sheets

Percent Change (%) =  

1.  Now calculate the descriptive statistics (average, standard deveation, and standard error of th mean) for the % disolution at each ph.  Remember Lab 2?  Same calculations. Ask for help if you need it.

1. Create a bar plot in Excel that shows the percent change from Time 0 to Time 2. This is the % disolution.  Make the plot aesthetically pleasing, ensuring that the final product will look good when imported into a WORD.
1. Checklist:
1. Bars on the plot should compare T0 vs. T1 (as a % weight loss)  for each experimental pH
2. Is the text large enough to read?
3. Are both axes labeled?
4. Are the tick marks a reasonable distance apart on the y-axis?
5. Are the y-axis tick mark labels appropriate and legible?

1. Open a new MSWord document with a proper file name that includes “AcidFinalRept” along with your name.  Remember for group reports only the team leaders name need be in the file name.   Remember that other team members names need to be in the report.
1. Copy and paste or import your figure from Excel (as a PNG or JPG) into this MSWord document
2. Write a Figure 1 caption.  Remember, captions should include all the information so the reader will have a solid understanding of what the figure represents if it was a stand alone product.

Part B. Addressing your hypothesis (Final Report)
Consider the hypothesis you created during the first week of the Ocean Acidification Lab. You now have access to the entire class data set.  Using this complete data set, visualize the data needed to address your hypothesis.  You will need to consider how best to visualize the data and be able to discuss why your hypothesis was upheld or not. You may need to do some more research and seek help from your instructor to TA on how to accomplish this task.  You can also complete some simple statistical analyses as well, that your instructor or TA could help you with.   Remember “Hypothesis testing from the stats lab? . If not take the time to take the time to work with your TA or instructor.

Your rubric and expectations are below. Let me know if you have questions.

Expectations of final report
You are to turn in a typed document, which includes all of the components of a scientific paper. You may want to reread the lab manual for lab 1 This document is to be formatted like a formal lab report, with Introduction, Methods, Results and Discussion.

Introduction	1. Includes the background information that you acquired in the first week.  This information should lead to your hypothesis. 2. States hypothesis that is based on knowledge of what you know about ocean acidification	15
Methods	Logical description of methods, that includes all steps taken to collect ALL the data. Also, description of replicates (each lab team did the exact same experiment and collected the same data. why did you do that?) Be sure to include everything we did over the three week long research project, that includes, weighing, seawater, pH...drying...etc...!	15
Results	Describes what happened in words and uses tables and/or graph type which highlights relationships between the data points or other relevant aspects of the data. To ilustrate the text. The graphs have has informative, concise and complete captions and axis labels. The results of hypothesis testing statistics The statistical analysis is appropriate, correct and clearly explained It includes a description of what constitutes a significant value and why that value was chosen as the threshold (e.g., p < 0.05).	15
Discussion	-Lead section with the most exciting or interesting result -Provide support from published literature as to why you observed the results you observed.--(Hint: use google scholar to look for ocean acidification papers relevant to your shell or shells considered in your hypothesis) -Conclusions address and logically refute or explain conflicting data	15
Background Info	Completed on first day of lab (already turned in)	15