1. Let it Snow: POP effects on
micro-plastic uptake in marine
snow
Honors Thesis Presentation
Meghan Danley

2. Marine Snow
Heteroaggregates formed by water
movement
Important food source for bivalves
Detritus accumulators for the environment
Nano/micro particle magnets
“Sticky” composition

3. Micro-plastics
Plastic particles <5mm
Sources: medicine,
cosmetics, industry,
degradation
Large SA:V
Binding and leaching of toxic
organic pollutants
Bioaccumulation
http://voices.nationalgeographic.com/2013/04/12/new-concerns-about-
plastic-pollution-in-great-lakes-garbage-patch/

4. Persistent Organic Pollutants (POPs)
Additives in manufacturing
PCBs, BFRs, BPA, phthalates, PAHs
Leaching, binding, transfer
To environment, plastics or organismal cells
Make plastics more “attractive”

5. PAHs
Polycyclic Aromatic Hydrocarbons
Do not break down easily
Hydrophobic, lipophilic
Often carcinogenic
EPA “bad actors”
http://www.sigmaaldrich.com/content/dam/sigma-
aldrich/structure6/100/mfcd00004136.eps/_jcr_content/renditions/mfcd
00004136-large.png
Pyrene

6. The Question
Will the presence of POPs in microplastics
affect marine snow formation?
Size
Shape
Number
Composition

7. Hypotheses
H0: The presence of PAHs in micro-plastics
does not affect marine snow formation
Ha: The presence of PAHs in micro-plastics
does affect marine snow formation

8. Experiment Design
Plastic selection, preparation and spiking
Creation of marine snow
Size and shape analysis using Image J software
Composition analysis using flow cytometry

9. Plastic and PAH selection
Cospheric Fluroescent Red,
Low-Density Polyethylene
(LDPE) micropheres:10-45μm
AccuStandard PAH mix: 18
compounds in acetonitrile at
various concentrations. None
water-soluble
http://www.cospheric.com/resize/images/UVPMS-
BR_375.jpg?lr=t&bw=550&w=550&bh=550&h=550

10. Microbead preparation
Natural surfactant through seawater aging
“PAH”: spiked stock to 100,000μg/L PAH mix
“VPE” (virgin polyethylene): spiked stock with
equivalent volume of acetonitrile
“Blank”: seawater stock spiked with
acetonitrile, no beads

11. Rolling Experiments

12. Rolling Experiments
Artificial marine snow creation
Water collection and analysis
Spiked bottles to 100 x104
bds/mL
Rolled for 3 days
Bottle pictures taken
Collected visible aggregates
and final water

13. Image J analysis
PAH
VPE

14. Image J Analysis

15. Image J Results
Figure 1. Area of aggregates plotted over
circularity (unitless) for two treatments of
microbeads. A circularity of 1 indicates a
perfect circle, and 0 an elongated elipse.
VPE demonstrates more even and
tighter distribution, where PAH
distribution varies greatly. Low circularity
can be correlated with larger size for
PAH aggregates, but not in VPE. p
values <0.05 for both size and circularity.
p-values
Area
0.00046006
Circularity
0.0000962

16. Image J Results
Figure 1. Area of aggregates plotted over
solidity (unitless) for two treatments of
microbeads. A solidity of 1 indicates a
rounder edges, and a 0 more “ruffled”
edges. VPE demonstrates more even
and tighter distribution, where PAH
distribution varies greatly. Low circularity
can be correlated with larger size for
PAH aggregates, but not in VPE. p
values <0.05 for both size and circularity.
p-values
Area
0.00046006
Solidity
0.0000002

17. Aggregate Numbers
PAH: 213
VPE: 186
Bins(mm^2): 0.0-5 5.0-10 >10
PAH: 40.37 27.69 27.23
VPE: 30.04 30.98 26.29

18. Flow Cytometry

19. Flow Cytometry: “Blank”

20. Flow Cytometry: “VPE”

21. Flow Cytometry: “PAH”

22. Flow Cytometry Results

23. Flow Cytometry Comparison

24. Discussion
Differences in appearance were found to
be statistically significant
Reject H0: The presence of PAHs does
affect marine snow formation
Difference between the two suggests PAH
binding was successful

25. Discussion
PAH: Linear, wider distribution of sizes,
greater number, obscured algae
fluorescence
VPE: Circular, clustered size range,
“fluffy”, algae obscured plastic
fluorescence
Closer to a natural aggregate

26. Future Directions
Determine PAH concentration in beads
GC/MS
Image analysis of natural snow
New/added replicates for cytometry
Surface of beads
PAH effects?
Quantify surfactant

27. Ecological Impacts
Odd size distribution can lead to greater
dispersion
Multiple small, toxic particles worse than one
bigger non-toxic one
Less algae accessibility
Even less “food-like” composition
Hydrophobic, lipophilic
Attractive to other micro plastics and promotes
leaching into cells

28. Acknowledgements
Dr. Steev Sutton, College of
Pharmacy
Dr. Steve Zeeman, Marine
Sciences Dept.
Dr. J. Evan Ward, UCONN
John Doyle, UCONN
Dr. Srinidi Mohan, College of
Pharmacy
UNE “mini-grant”

29. References
• Cole M., et al., 2013. Microplastic ingestion by zooplankton.
Environmental science and
• Technology 47: 6646−6655.
• Collignon A., et al. 2012. Neustonic microplastic and zooplankton in
the north western
• Mediterranean sea. Marine Pollution Bulletin 61: 861-864.
• Collins JF, Brown JP, Alexeeff GV, Salmon AG. 1998. Potency
Equivalency Factors for
• Some Polycyclic Aromatic Hydrocarbons and Polycyclic Aromatic
Hydrocarbon Derivatives. Regulatory Toxicology and
Pharmacology 28(1): 45-54.
• Engler R.E., 2012. The Complex Interaction between Marine Debris
and Toxic
• Chemicals in the Ocean. Environmental Science and
Technology. 46:
• 12302−12315.

30. Questions?

31. Appendix: Blank

32. Appendix: PAH

33. Appendix:VPE

34. Appendix: TSS & Alkalinity
Blank/VPE
0.580906684
0.628327623
VPE/PAH
0.27192912
0.268480841
PAH/Blank
0.643187718
0.4735534
0
1
2
3
4
5
6
7
8
9
0
0.02
0.04
0.06
0.08
0.1
0.12
0 0.5 1 1.5 2 2.5 3
pH
GranFunctionf(V)
Acid Volume (mL)
f(V)
pH
0
1
2
3
4
5
6
7
8
9
0
0.02
0.04
0.06
0.08
0.1
0.12
0 0.5 1 1.5 2 2.5 3
pH
GranFunctionf(V)
Acid Volume (mL)