PennScience Spring 2018 Issue: The Environment

Volume 16 • Issue 2 • Spring 2018

The Environment Protecting our Earth

Coloring Deserts Green

Bushmeat

Tech and Species Biomagnification Sustainable Cities Conservation

Can we transform The environmental How plastics have invaded the untapped deserts into consequences of energy sources? bushmeat consumption world’s waters

How close are Revolutionizing we to a greener the tracking of urban landscape? endangered species

PennScience Spring 2018

Volume 16 Issue 2

PennScience is a peer-reviewed journal of undergraduate research published by the Science and Technology Wing at the University of Pennsylvania and advised by a board of faculty members. PennScience presents relevant science features, interviews, and research articles from many disciplines, including the biological sciences, chemistry, physics, mathematics, geological sciences, and computer sciences. PennScience is funded by the Student Activities Council. For additional information about the journal including submission guidelines, visit www.pennscience.org or email pennscience@gmail.com.

EDITORIAL STAFF EDITORS-IN-CHIEF Mia Fatuzzo Grace Ragi WRITING MANAGERS Kenny Hoang Darsh Shah EDITING MANAGERS Aaron Zhang Rachel Levinson DESIGN MANAGERS Chigoziri Konkwo Abhi Motgi BUSINESS MANAGERS Donna Yoo Jenny Wang TECHNOLOGY MANAGERS Rounak Gokhale FACULTY ADVISORS Dr. M. Krimo Bokreta Dr. Jorge Santiago-Aviles

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WRITING

EDITING

DESIGN

BUSINESS

Hiab Teshome Rosie Nagele Asha Dahiya Eric Teichner Neelima Paleti Roshni Kailar Tamsyn Brann Piyush Pillarisetti Amanda Paredes-Barbeito

Brian Zhong Elly Choi Karbi Choudhury Miriam Minsk Sapna Nath Sarah Fendrich Sumant Shringari Vera Lee William Hasley Kathy Wang Kelly Liang Lily Zekavat Mimi Lu

Abigail Szabo Grace Wu Olivia Myer Lauren Kleidermacher

Catherine Ruan Justin Lish Felipe Carvao Rounak Gokhale

PENNSCIENCE JOURNAL | SPRING 2018

TABLE OF CONTENTS FEATURES

Biomagnification: A Closer Look into Marine Food Chains & Pollution • 6

by Neelima Paleti, design by Grace Wu

Bushmeat: An Environmental Crisis and a Threat to Public Health • 8

by Asha Dahiya, design by Abigail Szabo

Merging Technology and Species Conservation • 10

by Hiab Teshome, design by Abhi Motgi

Biofuels: A New Fuel in a Near Future? • 12

by Eric Teichner, design by Lauren Kleidermacher

Amine Scrubbing for Carbon Sequestration • 14

by Piyush Pillarisetti, design by Abigail Szabo

Coloring Deserts Green: Using Deserts for Energy and Vegetation • 16

by Roshni Kailar, design by Olivia Myer

Sustainability in Urban Design • 18

by Rosie Nagele, design by Chigoziri Konkwo

Biodiversity: The Key to Human Survival • 21

by Tamsyn Brann, design by Chigoziri Konkwo

INTERVEW Dr. Frances K. Barg • 23

by Amanda Paredes-Barbeito

RESEARCH Activated Glia May Induce Developmentally Regulated Inflammation and Synaptic Remodeling in a TSC Mouse Model of Epilepsy • 25

by Karbi L. Choudhury

Theoretical Analysis of Bond Dissociation Enthalpy: A DFT Study of Various Antioxidants with Hydroperoxyl and Hydroxyl Radicals • 33

by Vraj Rasesh Shroff (व्रज रसेश श्रॉफ)

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LETTER FROM THE EDITORS Dear Reader, We are thrilled to present our second issue of Volume 16 of the PennScience Journal of Undergraduate Research. We are incredibly grateful to all our staff who collectively created this issue, undergraduate students who submitted their research findings to our Journal, and the Penn undergraduate community for engaging in scientific discourse on campus. In this issue, Neelima Pateli explores marine food chains and pollution.

Asha Dahiya discusses the potential health concerns of bushmeat in Africa. Hiab Teshome examines new technology in conservation efforts. Eric Teichner takes a closer look at biofuels. Piyush Pillarisetti investigates new technology in carbon sequestration. Roshni Kailar proposes deserts as a new source of vegetation.

Rosie Nagele provides a detailed look at sustainability in urban design. Finally, Tamsyn Brann delves into the issue of invasive species. We are also proud to present the original research of two undergraduates – Karbi Choudhury and Vraj Shroff.

We have greatly enjoyed our semester leading PennScience, and we would like to extend a sincere thank you to the many groups and individuals who have made PennScience possible. First, we would like to thank our incredible journal staff--our writers, editors, and design and business members--for their hard work, dedication, and enthusiasm. Our publication is entirely student run and relies on the efforts of our scientifically curious undergraduate members. We want to recognize two leading Penn scientists, Dr. Marsha Lester and Dr. Eric Schelter, who helped us promote scientific discourse on campus by speaking at our coffee chats. We would also like to thank Dr. Frances Barg, who was interviewed for our jounal by writing committee member Amanda Paredes-Barbeito.We owe our funding to the Science and Technology Wing of the King’s Court College House and Student Activities Fund, in allowing us to publish a high-quality journal every semester. We would also like to thank our faculty mentors, Krimo Bokreta and Jorge Santiago-Aviles, for their guidance and support. Finally, we want to thank you for reading PennScience--enjoy our latest issue!

Sincerely, Mia Fatuzzo (C’ 19) and Grace Ragi (C’ 19), Co-editors in Chief

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PENNSCIENCE JOURNAL | SPRING 2018

CALL FOR SUBMISSIONS Looking for a chance to publish your research?

PennScience is accepting submissions for our upcoming Fall 2018 issue!

Submit your independent study projects, senior design projects, reviews, and other original research articles to share your work with fellow undergraduates at Penn and beyond.

Email submissions and any questions to pennscience@gmail.com

Research in any scientific field will be considered, including but not limited to:Â

Biochemistry Biological Sciences Biotechnology Chemistry Computer Science Engineering Geology Mathematics Medicine Physics Psychology

PREVIOUS ISSUES Visit the PennScience website

www.pennscience.org

to see previous issues and for more information.

Fall 2017

Spring 2017

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FEATURES

BIOMAGNIFICATION:

A CLOSER LOOK INTO MARINE FOOD CHAINS & POLLUTION NEELIMA PALETI

S

tretches of deep blue waves of water. Shining grains of cream-colored sand. A plethora of marine life. It’s quite easy to appreciate the divine beauty of the beaches lining the world’s coastlines, yet the fisheries of these coasts see beyond this facade. After scraping through the shells of oysters and organs of fish washing up on the shore, fishermen and researchers have found a wealth of information hiding amongst these sea dwellers. From clams to larger breeds of fish, each organism holds an abundance of one common ingredient tucked away in its tissues: plastic.

With approximately 1.4 billion pounds of waste entering the ocean each year, it comes as no surprise that eight million metric tons of plastic waste comprise these waters.1 Ranging from cosmetics and face washes to plastic bags and lighters, there are many ways that these plastics are deposited into the vast abyss of the world’s oceans. To make situations worse, UV rays from the sun constantly break down each piece of plastic waste into small particles of microplastics, which can then filter through cell membranes and accumulate inside organs of aquatic species.2 While this threatens the health and sustainability of marine organisms, it also points to health risks for the larger array of human consumers. Scientists have traced increased levels of mercury and other microplastic-associated pollutants down to even a single strand of human hair, largely due to biomagnification.3

Biomagnification refers to the accumulation of pollutants across organisms of a food chain. As plastics break down into microscopic particles and chemical pollutants, they are often absorbed by plankton and concentrated heavily amidst their cells.4 Since plankton serve as the food source for many larger fish, this accumulation of pollutants in the fish is readily transferred into the tissues and organs of higher trophic levels

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PENNSCIENCE JOURNAL | SPRING 2018

organisms. Over time, this process extends its effects over larger populations with higher concentrations of pollution and higher numbers of exposed plankton. Not limited to microplastics, biomagnification applies to pollutants of any nature that can infiltrate different food chains, including insoluble pollutants.2 The rule of ten further illustrates this flow of energy and biomass. As biomass is consumed from one trophic level to the next, only 10% of that energy is passed on.

“DUE TO THE INSOLUBILITY OF THESE POLLUTANTS IN FAT TISSUE, THE CHEMICALS REMAIN INDEFINITELY WITHIN THE ORGANISM’S TISSUE” Due to this decreasing energy obtained, higher-level organisms need to consume greater amounts of organisms of lower trophic levels in order to compensate for their energy needs. Eventually, the harmful pollutants that were absorbed and concentrated by plankton accumulate at toxic levels in these fish as they consume hundreds of plankton every day. Due to the insolubility of these pollutants in fat tissue, the chemicals remain indefinitely within the organism’s tissues.5 The low chemical concentration within planktons may have minimal effects, yet for organisms ranked higher up in the food chain, these chemical concentrations are magnified to the point of tangible health harm.4 In due time, humans, who preside at the highest level of the food chain, are the consumers of this accumulated pollution through

FEATURES the bodies of these sea organisms. This gradual process of face-wash products, as these predominantly comprise the biomagnification is now affecting many types of organisms microplastics found in oceans.7 With the United States now on board as well, this ban is looking to gain global traction over and food chains and it is only expected to increase with time. the upcoming

months.8 As supermarkets and Legislation common consumers are conflicts that now observing these effects may have in their commercially sold once caused seafood, the push to develop a standstill technology to eliminate between parties contamination in the oceans in the United has been more rampant than States must ever. The Ocean Cleanup temporarily is a nonprofit startup that be halted in demonstrates an effort to order to reach reverse the effects of pollution bipartisan deals for future generations. on this urgent Through this initiative, issue. Ultimately, Dutch innovator Boyan while new Slat intends to clean up the technical garbage patches infiltrating solutions may the oceans.6 By using a flexible pipe, anchors, and arise over the The Ocean Cleanup. “The Largest Cleanup in History.” The Ocean Cleanup, AkzoNobel, 2015 detectors, this technology will next few years, float across the top layer of the ocean, picking up any debris the time and efficiency spent on approving and instituting of plastic or other solid pollutants. Instead of using nets to these new measures becomes just as imperative to solving this capture the waste, the device will employ a solid sheet of finely- problem. Although we may not return to the untouched deep netted screening material that works with the currents to blue waves, we can attempt to restore these oceans to plasticattract pollutant particles without disrupting other organisms.6 free ecosystems with continuous collaborative efforts.

This serves to consolidate the collected trash, preparing it for disposal onto a support vessel nearby. Using this technology, Slot hopes to reduce 50% of the garbage patch in five years References starting in early 2018. Compared to other similar technologies, 1. Ocean pollution Ocean pollution | National Oceanic and this project would ideally work with higher efficiency with Atmospheric Administration. minimal disruption of the natural environment.6 2. Li, W., Tse, H. and Fok, L. (2016). Plastic waste in the marine

environment: A review of sources, occurrence and effects. Science

“IT IS THE MANNER WITH WHICH GOVERNMENTS ENCOURAGE SUCH INNOVATIONS THAT GREATLY DETERMINES FUTURE PROGRESSIONS”

While this range of new technology is being developed, different countries around the world are re-evaluating their environmental policies to minimize water pollution. The manner in which governments encourage technological innovations, both politically and financially, will greatly determine future progression in the field. The United Kingdom has taken the primary initiative to ban microbeads found in

of The Total Environment 566-567, 333–349.

3. Bełdowska, M. and Falkowska, L. (2016). Mercury in marine fish, mammals, seabirds, and human hair in the coastal zone of the southern Baltic. Water, Air, & Soil Pollution 227.

4. IUPUI Department of Biology. “Ecosystems.” Ecosystems, IUPUI Department of Biology, 19 Apr. 1999.

5. Remoundou, K. and Koundouri, P. (2009). Environmental Effects on Public Health: An Economic Perspective. International Journal of Environmental Research and Public Health6, 2160– 2178.

6. AkzoNobel partners with The Ocean Cleanup for largest clean-up in history (2017). AkzoNobel partners with The Ocean Cleanup for largest clean-up in history | AkzoNobel.

7. Knapton, S. (2017). 193 nations sign pledge to tackle ‘global crisis’ of plastic in the oceans . The Telegraph.

8. Laws & Regulations - The Microbead-Free Waters Act: FAQs U S Food and Drug Administration Home Page.

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FEATURES

Bushmeat: An Environmental Crisis and a Threat to Public Health

By Asha Dahiya

T

he Ebola crisis of 2014-2016 ravaged West Africa, killing over 10,000 people and crippling the economies of two of the poorest African countries, Liberia and Sierra Leone.1 Shockingly, the epidemic can be traced back to a single patient: a two-year old Guinean child from the village of Gueckedou. Patient Zero, as the child is called, contracted the virus after consuming bat meat.2 The practice of eating bushmeat – bats, but also primates, antelopes, rats, and other non-agricultural wildlife – is a longstanding tradition in West Africa and the Congo Basin and has been a major part of the diet there for centuries, with ties to medicinal beliefs and even circumcision rituals.3 But bushmeat consumption has a hidden, deadly risk for humans: primates and fruit bats are vectors for fatal viruses such as Ebola.

Bushmeat hunting takes a toll on not only vulnerable species like great apes but also on the entire forest ecosystem.

Primates and fruit bats are both known carriers of the Ebola virus. Although the meat poses little risk once thoroughly cooked, anyone who is exposed to the raw meat’s bodily fluids are at significant risk of transmission. Although scientists do not fully understand the mechanism of transmission, we do know that humans often contract the disease from a primate intermediate.4 Transmission can 8

PENNSCIENCE JOURNAL | SPRING 2018

also occur directly from ras meat to people, as evidenced

by Patient Zero’s case, or via contact with animal feces. The virus spreads from infected blood to tissue cells, hijacking the human cells to replicate itself and killing the cells in the process while provoking a massive inflammatory response that causes the characteristic heavy bleeding of Ebola. There is no cure for the virus – it can only be treated supportively by supplying fluids and monitoring fever.2 One out of every two people who contract the disease will die.1 This fatality rate was exacerbated by the scarcity of healthcare resources in West African countries such as Guinea and Sierra Leone, where it is not uncommon for hospitals to run out of basic tools like gloves and IV needles during a crisis.5 In addition to threatening human lives, bushmeat consumption can have a long-term impact on the targeted animal populations. Over 6 million tons of bushmeat are trafficked in the Congo Basin each year.3 Coupled with a swelling human population and expanding deforestation, this extensive hunting of wild animals has become a serious threat to forest ecosystems. Large carnivorous animals – crocodiles, large cats, raptors – are considered “keystone species” because of the important role they play as predators in the balance of population sizes of the ecosystem. Unfortunately, these larger animals are easy and appealing targets for hunters due to their size. Furthermore, bushmeat hunting affects threatened primate species such as the great apes, which are particularly vulnerable to population decrease due to their slow reproductive rate.

Primate population of Gabon has decreased by 50% in the past 20 years, primarily due to hunting and trapping.6 Even when hunters are not targeting primates, these animals are often caught in wire traps set for smaller animals, resulting in an agonizingly slow death. Bushmeat hunting takes a toll not only on vulnerable species like the great apes but

FEATURES also on the entire forest ecosystem. The overall population humans and the environmental threat to animals may density of hunted areas within the Congo Basin is lower involve helping hunters choose safer prey by promoting than that of non-hunted areas.6 awareness of which species are least vulnerable, which animal populations, such as great apes, should be spared Though the hazards of bushmeat are clear, it remains a at all costs, and which animals pose the greatest risk of pervasive element of West and Central African diet and transmitting deadly diseases. While in the long-term, culture. Several anti-bushmeat task forces exist, primarily sustainable agricultural practices could eliminate the need the Bushmeat Crisis Task Force, an independent coalition for bushmeat consumption altogether, the more near-term of scientists and conservation groups, and the Bushmeat solutions must involve compromise. As Guinea native Sâa Working Group, founded by international agreement and Fela Léno remarked to the Guardian, “Banning bushmeat consists of non-profit groups and government delegates.7,8 means a new way of life, which is unrealistic.”9 However, millions of people depend on bushmeat as a primary source for protein – is it truly feasible to eliminate it? The near-term solutions to both the health threat for

References: 1. Ebola Virus Disease: Fact Sheet(2018). World Health Organization: Media Centre. (http://www.who.int/ mediacentre/factsheets/fs103/en/) 2. Hogenboom, M. (2019). Ebola: Is bushmeat behind the outbreak? British Broadcasting Corporation.

3. Velde, B. V. (2014). 10 things you didn’t know about bushmeat in Africa. Forest News.

4. Phillips, A.(2014). Why West Africans keep hunting and eating bush meat despite Ebola concerns. Washington Post .

5. Belluz, J. (2015). 11 things you need to know about Ebola. Vox. 6. Nasi, R., Taber, A. and Vliet, N. V. (2011). Empty forests, empty stomachs? Bushmeat and livelihoods in the Congo and Amazon Basins. International Forestry Review13, 355–368.

7. Bushmeat Crisis Task Force (BCTF) Poverty and Conservation: The information portal of the Poverty and Conservation Learning Group. (https://www.povertyandconservation.info/en/org/o0163) 8. https://www.cites.org/eng/disc/what.php 9. Ebola risk unheeded as Guinea’s villagers keep on eating fruit bats (2014). The Guardian.

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FEATURES

MERGING TECHNOLOGY AND SPECIES CONSERVATION

by Hiab Teshome

H

uman activity and overpopulation are reducing Earth’s biodiversity at an alarming rate. According to the Intergovernmental Panel on Climate Change, 100 to 1,000 species per million are lost each year. This decline in biodiversity is largely due to the rapid increase of anthropogenic habitat destruction over time.1 The main difficulty conservationists face in their efforts to curb environmental destruction is tracking, recording, and monitoring species as their populations decline. New advancements in genetics and artificial intelligence within the past year have significantly improved the ability of the scientific community to collect and share valuable data about millions of species. These emerging technologies have revolutionized how scientists interact with the environment by simplifying and expanding the range of tools used to monitor and collect information on endangered species.

Advancements in species conservation technology this past year have also included new developments in remote monitoring. These methods combine photography and sound detection systems with computer-assisted species identification programs to facilitate long-term systematic data collection

An important aspect of mitigating this rapid decline in biodiversity is genetic conservation. Decreased genetic diversity has been associated with reduced biological fitness, diminished population growth, and ultimately, higher risk for extinction.2 Collecting genetic material and sequencing DNA from endangered species has been a very challenging task for researchers since DNA is sensitive to environmental conditions and is slow to replicate in the lab. These difficulties are especially prominent in remote locations where biodiversity loss is characteristically greatest. Recently, researchers have focused on creating portable field labs that allow scientists and conservationists to quickly decode the genetic makeup of plants and animals and determine genetic variability in these locations.

Nanopore Sequencing

A team of researchers in the Expeditionlab Project have developed a portable lab called GENE to immediately decode DNA in challenging environmental conditions where extreme humidity, aridity, and temperatures would typically destroy sensitive DNA samples. In the past year, the Expeditionlab team brought the GENE lab to the tropical forests of the Congo to collect DNA from one of the most diverse, but poorly studied woodlands in Africa.3 The DNA sequencer uses nanopore sequencing technology that allows researchers to selectively choose the DNA fragments to be sequenced, increasing the efficiency of this process from days to around 10 minutes.4 In nanopore sequencing technology, scientists pass an ionic current through a nano-scale hole which measures the changes in current as biological molecules pass through the nanopore.

This process allows researchers to directly record the base pairs of a DNA fragment. Because this process does not require DNA amplification, it reduces the possible errors associated with sequencing and increases the speed of the process.5

without encroaching on the habitats of these organisms.6 The use of this technology was first applied in endangered species of lemurs in Madagascar. In 2012, the International Union for Conservation of Nature listed Madagascar lemurs as the most endangered mammal on Earth.7 A team of lemur biologists and computer scientists at George Washington University recently developed a semi-automated facial recognition system called LemurFaceID that could identify lemurs of the same species in Madagascar in order to alleviate the difficulty conservationists face in tagging and collecting data on these species.

LemurFaceID can correctly identify individual lemurs based on their facial features with 98.7% accuracy without imposing any harm or stress on the organisms.8 The ability to harmlessly study individuals and populations over long periods of time will provide valuable information on both the mortality rates and population fluctuations of these species. This technology

10 PENNSCIENCE JOURNAL | SPRING 2018

http://www2.technologyreview.com/news/427677/nanopore-sequencing/

FEATURES is currently being developed to identify other endangered organisms such as primates and non-primate species with variable facial hair and skin patterns.

and management plans, researchers must understand the distribution and biological fitness of endangered species in a particular area. In the last year, incredible advancements have been made to increase the speed and accuracy of a range of different data collection tools. The data collected through this new technology can be used to create effective and long lasting conservation plans that reflect the specific needs of a particular endangered species. Through improving data collection researchers can save a wide variety of organisms based on their specific distribution patterns, bringing us one step closer to healing our environment.

Ring-tailed lemurs (Lemur catta)

https://www.newscientist.com/article/2154674-madagascars-lemurs-close-to-extinction-after-population-crash/

Researchers are not only developing facial recognition technologies, but are also introducing new bioacoustic models that are able to collect information on the distribution and abundance of various endangered species. Researchers have historically used automatic sound recorders to gather long-term information on species with relatively low effort. One pitfall of this method is that processing all the information requires large amounts of time and resources. Recently, a team led by Dr. Sebastián‐González from the University of Hawaii created an automatic detection algorithm to improve conservationists’ ability to acquire data about a species population from sounds emitted by animal species.9 In their first case study, they applied their algorithm to detect the abundance of an endangered bird species in Hawaii called the ‘Amakihi. The researchers were able to program their algorithm to detect ‘Amakihi songs and distinguish their sounds from that of other organisms by adding a support vector machine (SVM) to their algorithm.

A SVM is a classification tool that uses a multidimensional space to separate elements into distinct groups.10 The algorithm recognizes various sounds in the environment and divides the sounds into “non-‘Amakihi” and “‘Amakihi” sounds. They were then able to calculate the relative abundance of the ‘Amakihi by counting the number of detections from the SVM per minute. This information may be used to assess which areas should be prioritized in conservation plans or to evaluate the temporal trends in the population of a species.11 The SVM had an accuracy of 86.5% in identifying ‘Amakihi, which is very high considering the great variance in the ‘Amakihi songs and the interference from other organisms’ sounds.12 The ability to obtain information on species distribution and relative abundance has made Dr. Sebastián‐González’s algorithm a stepping stone for using other animal sounds to understand more about endangered species.

Data collection is an important part of the conservation effort.

‘Amakihi In order to effectively prepare http://www.mauiforestbirds.org/articles/4/ and implement conservation

Song Classification Using SVM

(A) Bad selection, type 1: song poorly selected/very overlapped with other sounds; (B) medium selection, type 2: ‘Amakihi song not too precisely selected/some overlap; (C) good selection, type 3: song well selected/no overlap, class 3; (D) picture of the Hawai’i ‘Amakihi; (E) picture of a songmeter tied to a tree. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4670053/ References: 1. IPCC - Intergovernmental Panel on Climate ChangeIPCC - Intergovernmental Panel on Climate Change. (http://www.ipcc-data.org/observ/index.html) 2. Frankham, R.(2005). Genetics and extinction. Frankham, Richard (2005-11-01).

“Genetics and extinction”.126, 131–140.

3. Rugged innovation: Meeting the challenges of bringing high tech DNA analysis to the field (2017). Conservation news. (https://news.mongabay.com/wildtech/2017/05/ rugged-innovation-meeting-the-challenges-of-bringing-high-tech-dna-analysis-tothe-field/) 4. DNA: Nanopore SequencingOxford Nanopore Technologies. (https://nanoporetech. com/applications/dna-nanopore-sequencing) 5. A nanopore is a very small hole. How it works. (https://nanoporetech.com/how-itworks) 6. Crouse, D., Jacobs, R. L., Richardson, Z., Klum, S., Jain, A., Baden, A. L. and Tecot, S. R.(2017). LemurFaceID: a face recognition system to facilitate individual identification of lemurs. BMC Zoology2.

7. Furness, D.(2017). Facial-recognition software may help save Earth’s most endangered mammals. Digital Trends.

8. LemurFaceID: a face recognition system to facilitate individual identification of lemurs (2017). (https://bmczool.biomedcentral.com/articles/10.1186/s40850-0160011-9) 9. Sebastián-González, E., Pang-Ching, J., Barbosa, J. M. and Hart, P.(2015). Bioacoustics for species management: two case studies with a Hawaiian forest bird. Ecology and Evolution5, 4696–4705.

10. Sebastián-González, E., Pang-Ching, J., Barbosa, J. M. and Hart, P.(2015).

Bioacoustics for species management: two case studies with a Hawaiian forest bird.

Ecology and Evolution5, 4696–4705.

11. Pearce J., and Ferrier S.. 2001. The practical value of modelling relative abundance of species for regional conservation planning: a case study. Biol. Conserv. 98:33–43.

12. Sebastián-González, E., Pang-Ching, J., Barbosa, J. M. and Hart, P.(2015).

Bioacoustics for species management: two case studies with a Hawaiian forest bird.

Ecology and Evolution5, 4696–4705.

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FEATURES

Biofuels: A New Fuel in a Near

By Eric Teichner Future?

T

he thought of going to the gas station and filling your car up with fuel made from algae may seem absurd. However, this possibility is closer than you think.

Emerging concerns over security and energy issues have led to a global increase in demand for sustainable fuel sources.

Contrary to other renewable energy sources, such as wind and solar, biomass can be converted directly into liquid fuels, known as biofuels. Today, the two most common types of biofuel are ethanol and biodiesel. Currently, these early forms of biofuel are made from starches and sugars. However, scientists are developing new technology in order to produce ethanol from other sources.

The transition to an ethanol-based biofuel stems from the discovery that methyl tert-butyl ether (MTBE), commonly found in gasoline throughout the 20th century, was severely contaminating groundwater. Hence, it was found that ethanol could replace this ether by providing a much more sustainable method of fuel1. Today, when you go to nearly any gas station, there is about 10% of ethanol by volume. Most of this ethanol is derived from natural sources, mainly corn. In the United States, it is mandated that any gasoline-powered engine is able to run autonomously on 10% ethanol. However only specific engines are able to be powered by a mixture of fuel greater than this standard number. Cars labeled as “flexible-fuel vehicles” can run on E15, 15% ethanol, or E85, an 85% ethanol mixture typically sold only in the Midwest2.

Biodiesel is another realm of biofuel that is crucial to the 12 PENNSCIENCE JOURNAL | SPRING 2018

industry. Biodiesel is the stereotyped “biofuel,” often made from vegetable oils, fats, or greases. Notably, biodiesel can be used in nearly all standard diesel engines without any mechanical replacement. Biodiesel is highly sustainable, biodegradable, and non-toxic for the environment. Algal biofuel is particularly attractive, as well are other common biofuel sources such as corn and sugarcane. Algae fuel releases carbon dioxide when burned, similar to fossil fuels. However, because algae perform photosynthesis to absorb carbon from the atmosphere, the net carbon output is zero. Furthermore, algae can be cultivated in areas unsuitable for agriculture, hence any “wasted” space could easily become a breeding ground for a highly sustainable fuel3.

Chemically, biodiesel has some noteworthy properties of interest. Specifically, biodiesel are produced using a method known as “transesterification.” This organic process exchanges the the R’’ of an ester with the organic group of R’ of an alcohol.

Essentially, this allows the conversion conversion of one alcohol to another alcohol via an ester. In this process, researchers are looking at ways to optimize the the esterification of biodiesel feedstocks (natural plant oils and animals fats)4.

Microalgae are another important biodiesel feedstock systems -- they can provide a large amount of crude lipid which, through an industrial process utilizing hydrogenations and transesterification, can be converted into biodiesel. Algae is particularly attractive, as they have an unusually high photosynthetic rate, which if extracted efficiently would result in a large fuel obtainment4.

FEATURES It is important that we make a distinction between biofuel and biodiesel. Not only do the fuels differ chemically, but the corresponding engines differ as well. Diesel engines do not rely on spark plugs, but rather the “compression stroke” in the engine is designed to have a more intense compression of the gasses. As a result of this, the heating is much greater and the ignition is spontaneous. Chemically, diesel is easier to ignite due to its lower autoignition temperature (265 C vs 280 C). Diesel fuel contains mostly hydrocarbons, hence it is a bigger molecule, which means that it evaporates more slowly at ambient conditions, but requires more heating/compression before it spontaneously ignites5.

Biodiesel has a long way to go to become an easily accessible and inexpensive fuel. Various companies, such as Solazyme and Algenol, have made broken promises in 2009 to mass produce millions of gallons of ethanol from algae on ground conditions that are not favorable. Nothing close these millions of gallons has been achieved, and the pricing it not competitive for market use. Because algae contain up to 40 percent lipids, the future seems promising, but we must consider other capital needs such as water, energy balancing, growing, collecting, before determining if it a viable source of fuel6.

References 1. Goettemoeller, J. and Goettemoeller, A.(2007). Sustainable ethanol: biofuels, biorefineries, cellulosic biomass, flex-fuel vehicles, and sustainable farming for energy independence. Maryville, MO: Prairie Oak Publ.

2. Biofuels: Ethanol and Biodiesel Explained. (https://www.eia.gov/ energyexplained/index.cfm?page=biofuel_home) 3. Amin, S.(2009). Review on biofuel oil and gas production processes from microalgae. Energy Conversion and Management50, 1834– 1840.

4. Owolabi, R. U., Adejumo, A. L. and Aderibigbe, A. F.(2012).

Biodiesel: Fuel for the Future (A Brief Review). International Journal of Energy Engineering2, 223–231.

5. Turner, B. (2015). To diesel or not to diesel? Pros and cons of diesel vs. gas. Driving. (http://driving.ca/volkswagen/golf/auto-news/news/ to-diesel-or-not-to-diesel-that-is-the-question) 6. Wesoff, E.(2017). Hard Lessons From the Great Algae Biofuel Bubble. gtm.

Much more research and industrialization is needed before biofuels and biodiesel becomes a widespread source of energy.

Hopefully, this is something that will occur in our lifetime.

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FEATURES

Amine Scrubbing for Carbon Sequestration

by Piyush Pillarisetti

T

en miles north of the United States-Canada border, in a small town enriched by a long tradition of coal mining, power generation, and gas production, the sound of thousands of tons of carbon dioxide gas being stripped from power plant emissions pervades the air. That sound is coming from the SaskPower Boundary Dam Power Station in Estevan, Saskatchewan, one of the first deployers of the amine-scrubbing technology, an inthe-works carbon sequestration technique that hopes to strip the atmosphere of its accumulating greenhouse gases.

The New York Times “Technology to Make Clean Energy from Coal is Stumbling in Practice”

The global carbon cycle is one of many environmental equilibria that is regulated by chemical synergism, positive and negative feedback, disruption by human activity, and adaptive response.

Carbon exchanges within the biosphere were synchronized prior to the burning of fossil fuels and the resultant increase in atmospheric CO2 levels. Carbon used to flow between terrestrial and biological reservoirs such as the atmosphere, ocean, geological formations, soil, and living organisms such that the rate of CO2 uptake by the atmosphere (a process facilitated by “CO2 sinks” such as the forest and ocean) equaled the rate of CO2 released into the atmosphere.1 Human carbon-emitting activities have disrupted this balance as CO2 sinks are unable to 14 PENNSCIENCE JOURNAL | SPRING 2018

compensate for the 100 ppm increase in atmospheric CO2 levels over the last 250 years.2 According to the U.S. Climate Change Science Program, the net efflux of CO2 into the atmosphere by fossil fuel emissions was about 1.6 gigatons while the net intake of atmospheric CO2 by carbon sinks was only about 0.5 gigatons in 2003.2 To address this issue, ecologists and scientists are developing new carbon sequestration techniques that divert CO2 produced by fossil fuel power plants away from the atmosphere. A variety of carbon sequestration technologies are currently being developed but perhaps the most costeffective and practical is aqueous amine solvent-based amine scrubbing.3,4 Amine scrubbing is a post-combustion process that uses an aqueous amine solution consisting of a weak base alkanolamide to extract acidic substances such as hydrogen sulfide and CO2 from flue gas.5,6 Flue gas is exhaust gas, which contains greenhouse gases produced by power plants. In an amine scrubbing apparatus, flue gas is first passed through an absorber column containing the aqueous amine solution.7 Currently, the most commonly used amine is monoethanolamine but other amines with unique gaseous extraction efficiencies are used for different industrial processes. The aqueous solution absorbs and extracts acidic constituents of the flue gas via exothermic soluble salt reactions.7 During these reactions, CO2 reacts with the amine to form an ionic substance containing positively

CO2 sinks are unable to compensate for the 100 ppm increase in atmospheric CO2 levels over the last 250 years

and negatively charged ions. The processed CO2-free flue gas is then transported to the atmosphere and the amine solution containing the CO2 is moved to a stripper unit in which the initial salt-forming reaction is reversed by the addition of heat. As such, the CO2 is separated from the alkanolamide

FEATURES and is dehydrated and compressed.5 Amine scrubbing is also gas processing facilities because they (1) are modularly designed effective in reducing the harmful gaseous products generated such that new improvements in a particular compartment can by petrochemical facilities.6 be easily implemented into preexisting amine systems; (2) are subject to limited “manufacturing and commissioning times” because of standardized basic operating principles and design schemes; (3) have proven records for extraction efficiency Operational reports have found that the impleand performance; (4) easy retrofitting of amine systems with mented amine scrubbing technology can sedifferent amine gas sweetening solutions and (5) “built in liquid quester up to 1.3 million tons of carbon dioxide containment to reduce environmental hazards”.9

from flue gas

The Boundary Dam Carbon Capture Project, which began in 2014, was created to implement post-combustion amine scrubbing technology into all the flue gas expulsion sites at the Boundary Dam Power Station in Saskatchewan, Canada.

Operational reports have found that the implemented amine scrubbing technology can sequester up to 1.3 million tons of carbon dioxide from flue gas.8 Although there are other pre-combustion and oxyfuel based methods for reducing CO2 emission associated with natural gas and coal-fired power plants, post-combustion amine scrubbing “allows for easy and intuitive retrofitting of already existing CO2 sources”.7 Other post-combustion carbon capture technologies include the application of chemical and physical aqueous solvents, nanoparticle-based hybrid membranes, and solid sorbents.3 Amine-scrubbing, however, continues to be used in

Of course, amine scrubbing still faces challenges in terms of implementation and optimal efficiency. In particular, flue gases that enter absorber units are generally at a high temperature and can cause amine evaporation. Amine evaporation is of particular concern because alkanolamides such as methyldiethanolamine have been found to be volatile and environmentally corrosive.7 As anthropogenic CO2 levels continue to rise quickly, the need for flue gas post-combustion technologies such as amine-scrubbing that can be easily retrofitted, constantly improved, and readily implemented are necessary and present promising opportunities for a cleaner atmosphere.

References

“Amine-Based CO2 capture technology development from the beginning of 2013” by Dutcher, Fan, and Russell

1. Riebeek, H. (2011). The Carbon Cycle : Feature Articles.

NASA. 2. Sundquist, E., Burruss , R., Faulkner , S., Gleason , R., Harden , J., Kharaka , Y., Tieszen , L. and Waldrop, M. (2008). Carbon sequestration to mitigate climate change.

U.S. Geological Survey 1–4.

3. Park, Y., Lin, K.-Y. A., Park, A.-H. A. and Petit, C. (2015).

Recent Advances in Anhydrous Solvents for CO2 Capture: Ionic Liquids, Switchable Solvents, and Nanoparticle Organic Hybrid Materials. Frontiers in Energy Research 3, 42.

4. Rochelle, G. T. (2009). Amine scrubbing for CO2 capture.

Science 325, 1652–1654.

5. Amine Scrubbing Process Amine Scrubbing Process | Global CCS Institute.

6. Amine Scrubbing Technology Amine Scrubbing Technology | Global CCS Institute.

7. Dutcher, B., Fan, M. and Russell, A. G. (2015). AmineBased CO2 Capture Technology Development from the Beginning of 2013—A Review. ACS Applied Materials & Interfaces 7, 2137–2148.

8. Boundary Dam Carbon Capture Project SaskPower.

9. Amine Treating | Amine Gas Sweetening | CO2 & H2S

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FEATURES

Coloring Deserts Green: Using Deserts for Energy and Vegetation

By Roshni Kailar

A

Mehran Moalem from UC Berkeley, solar panels in just 1.2% of the Sahara Desert could power all of the world’s energy needs for one-tenth the cost of power that comes from electric plants.1

Deserts are typically considered arid regions that have no real utility. However, most deserts, like the Sahara, receive approximately 12 hours of unobstructed sunlight per day. Moreover, because deserts stretch on for miles, a cloudy area can be offset by other sunny areas within the desert itself. This makes deserts an ideal place for harnessing solar energy. The large area of deserts provides an immense amount of space to place solar panels to capture solar energy. According to Dr.

Although there is a general consensus that the use of solar panels in deserts is scientifically and economically viable, there are major challenges. First is that some politicians do not necessarily agree on climate issues and are reluctant to bear the costs, although minimal, to go about implementing such a large-scale plan.2 Another obstacle is the fact that deserts have a large amount of dust which also decreases the efficiency of solar panels by about 20% per month.

Therefore, before progress can be made, scientists have to find a way to limit dust deposition and degradation of the panels themselves in the desert.3 Other challenges are that solar panels may hurt the natural wildlife of the deserts and in fact perpetuate the lack of biodiversity.4

s climate change decreases habitable land, population growth outstrips agricultural production, and the Earth’s resources are depleted, scientists all over the world have proposed an instrumental solution: using deserts as sources of energy. The large surface area of deserts, combined with new technologies, makes formerly untapped deserts into a playground for researchers explore solution to the problems we are facing through desert energy and agriculture.

According to Dr. Mehran Moalem from UC Berkeley, solar panels in just 1.2% of the Sahara Desert could power all of the world’s energy needs for one-tenth the cost of power that comes from electric plants.

16 PENNSCIENCE JOURNAL | SPRING 2018

FEATURES in China, scientists have manufactured the same paste, carboxylmethyl cellulose (CMC), that plants use in their cell walls for protection. The scientists have applied this paste to over 70 varieties of crops. By utilizing this paste to convert sand into a soil-like material, plants can grow even in high temperatures and dry conditions. This is termed the “soilization” of sand, in which the top layer of sand is given properties of soil in order to allow plants to flourish in the arid conditions of the desert.

This sand retains water and helps plants grow as they would in normal conditions. This demonstrates that deserts can provide a large habitat on Earth for vegetation to combat deforestation and climate change. Research conducted on about 550 square meters of land has shown that a mixture containing just 2% of CMC can increase yields of crops like corn, potatoes, and radishes by about 50%.8 Vegetation in deserts provides an immense opportunity for reclaiming biodiversity as well as potentially reversing some climate change effects.9 Several solar panels concentrated on one solar tower

However, in the Sahara desert, there is an alternative method of collecting solar energy that might resolve some of the challenges present in solar panels. A line up of mirrors aim the collected sunlight into a tower of molten salt. Then, the molten salt stores the solar energy as thermal energy, which can be used to heat up water into steam to drive turbines, creating electrical energy.

TuNur, a solar power export project, has implemented this method in the Sahara desert.5 Moreover, the amount of energy stored can be discharged dynamically based on the demand for that energy. TuNur plans to transfer this energy from the Sahara to Europe through a connector cable at a loss of only 5% per 1000 kilometers.6 Although this is a long connection, according to Tony Patt, a professor of climate policy at the Swiss Federal Institute of Technology in Zurich, a single power cable can transmit energy of about 1000 miles, meaning this connection between Tunisia and the European grid is feasible.7 Scientists would like to implement the same types of ideas in other deserts to further supplement the supply of energy.

Another way deserts can be utilized is for planting new vegetation. Deserts can be turned into fertile land by various mechanisms. For example, at Chongqing Jiaotong University

Shrubbery growing naturally in the Sahara Desert in Algeria

Deserts have an unrealized potential regarding energy and agricultural resources. Thus far, researchers have investigated the possibility of solar energy and desert agriculture; however, future energy sources that deserts may adopt include geothermal and wind energy. TuNur provides an alternative energy plan, suggesting an alternative policy change for the future. Further research regarding CMC pastes and desert vegetation are

Vegetation in deserts provides an immense opportunity for reclaiming biodiversity as well as potentially reversing some climate change effects. also promising solutions to deforestation as well as a greener environment. Despite the general perception that the desert is wasted land, these recent findings suggest that deserts provide unforeseen potential to solve our environmental challenges.

References

1. Moalem, M.(2016). We Could Power The Entire World By Harnessing Solar Energy From 1% Of The Sahara. Forbes.

2. Darby, M.(2017). Giant Tunisian desert solar project aims to power EU. Climate Home News.

3. Saidan, M., Albaali, A. G., Alasis, E. and Kaldellis, J. K.(2016).

Experimental study on the effect of dust deposition on solar photovoltaic panels in desert environment. Renewable Energy92, 499–505.

4. Lovich, J. E. and Ennen, J. R.(2011). Wildlife Conservation and Solar Energy Development in the Desert Southwest, United States. BioScience61, 982–992.

5. Darby, M.(2017). Giant Tunisian desert solar project aims to power EU. Climate Home News.

6. Tu NurTu Nur: Overview.

7. Knies, G., Patt, T., Egbe, D. and Curry, H. A.(2015). Should we solar panel the Sahara desert? BBC News.

8. Yi, Z. and Zhao, C.(2016). Desert “Soilization”: An Eco-Mechanical Solution to Desertification. Engineering 2, 270–273.

9. Spaen, B.(2017). China Researchers Find Way To Transform Deserts Into Fertile Land. Green Matters.

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FEATURES

Sustainability in Urban Design

A

By Rosie Nagele

gust of wind sends leaves rustling along a shady street. The pale, porous pavement has soaked up the morning’s rainfall, and cyclists are out once again with the sun. There are few cars for them to compete with; many commuters rely on low emission subway lines or traverse tree-lined sidewalks on foot. Above, solar panels and windmills perch on the roofs of apartments and office buildings, powering the city beneath.

renewables. Temperature regulation, for instance, accounts for a significant portion of energy use. The outer barrier of a building, the building envelope, has the potential to insulate from cold and heat.10 Refining envelope design can considerably help reduce the use of high-energy HVAC systems. Solar shading devices and architectural designs that enhance passive heating from the sun also contribute to lower energy costs.11

With the human population rising and the consequences of irresponsible land use looming closer, scientists, government officials, and concerned city-dwellers alike are striving to make urban spaces more sustainable. Burlington, New Jersey’s 100% renewable energy supply, Melbourne’s sustainable building designs, and New York City’s 100 acres of urban gardens are just three examples.1, 2, 3 Cities around the world are taking steps to lower their environmental impact.

Installing energy efficient appliances and maximizing natural lighting can also reduce a building’s energy requirements. Adding innovations, however, does not always translate linearly into more energy savings.12 Variables like climate and building type mediate the interaction of different strategies.

In a building designed to exploit natural lighting, for example, installing light-dimming technology will do little to reduce energy use. Other technologies are synergistic, increasing energy savings by more than the sum of each innovation alone. In other words, the most effective sustainable measures vary by circumstance.

An urban area’s environmental impact depends on its built environment as well as the movement of people and materials within and across its borders. The built environment includes surfaces, namely streets and sidewalks, and the buildings atop them. Currently, impervious asphalt covers most cities.

Water runs over the surface, carrying pollutants to local waterways. Vegetation can reduce runoff pollution. As water moves through soil or evaporates off of plants, it leaves behind chemical contaminants. While there are many places for vegetation to expand, cities require a certain amount of hard, stable surfaces for pedestrians and cars. “Permeable pavements” offer a compromise. Simultaneously porous and solid, these materials form firm surfaces that maintain some functions of vegetative cover--namely the absorption, filtering, and storing of stormwater.4, 5, 6, 7 Permeable pavements are becoming more common in driveways, parking lots, and walkways but they are not yet suitable for the high-stress conditions of roads.

“Within the built environment, there is a constant flow of people and materials.”

Towering over streets, skyscrapers, mall complexes, and other buildings significantly contribute to urban energy expenditures.8 Worldwide, buildings account for around 40 percent of primary energy consumption and 50 percent of greenhouse gas emissions (GHG).9 In future construction and retrofitting projects, improved technology and design can lower this burden by reducing energy demands and utilizing 18 PENNSCIENCE JOURNAL | SPRING 2018

Within the built environment, there is a constant flow of people and materials. While individual ownership of electric vehicles still faces economic and logistical hurdles, some cities are investing in electric systems of public transportation.13 In order for the efficiency of public transportation to have an impact, however, it needs to provide reliable service and ample destination options. The Hong Kong Mass Transit Railway (MTR) provides a model for accessible, reliable, and affordable public transportation.

Even without widespread utilization of renewables, MTR’s consideration of

FEATURES land use and functional efficiency already goes a long way to lowering its environmental impact.14, 15 Urban landscapes are largely barren of the resources needed to meet their population’s daily demands, with food and water at the forefront. Few urban populations have access to potable water locally. Instead, extensive infrastructure diverts water from its natural course to be distributed throughout the city.

Moreover, droughts, floods, and pollution push clean sources of water further from dense population centers.16 Food, like water, is mostly imported from outside of cities, incurring large carbon footprints as fossil fuels burn in the engines involved in transporting it. Community farms not only eliminate emissions associated with shipping, but also those associated with the machinery and fertilizers common to industrial agriculture.17 Urban farming has grown in popularity and efficiency in recent years, though barriers remain.18 Staple grains, for example, have yet to penetrate small-scale gardening and local climate imposes limitations on crop type and growing season.

Still, fresh vegetables and fruits--which are more costly and complicated to transport than grains and meat--can be a valuable resource to urban Urban communities.19 farms also contribute to vegetative land cover and increase biodiversity.20 While currently not in a position to feed entire cities, local agriculture is an important element of urban sustainability.

As people consume food, they also produce waste--wrappers, cans, bottles, not to mention discarded food itself.

Traditional landfills occupy valuable space and don’t allow those materials to be recycled back into production.21, 22 Though concerns over pollution have long impeded the use of incineration plants, new technology has the potential to not only make this landfilling alternative safe, but also productive.23

Waste-to-energy plants sort through waste rather than burning it indiscriminately. While incinerating the wastes, the plants utilize the heat generated to produce electricity.24 Further refinements to this technology enable extraction of carbon dioxide and other gases as the material is burned.25 This additional step reduces the plants’ contribution to greenhouse gas emissions and produces compounds that can then be converted to natural gas and used as fuel.26 This technology is still in limited use, partly from lingering concerns about pollution and partly from the cost of construction.27, 28

“Ecosystem sustainability depends on the cyclical transformation of material and conservative processing of energy.”

Improving individual aspects of a city’s impact on the environment, however, doesn’t address the fundamental flaw in the way cities occupy space. Ecosystem sustainability depends on the cyclical transformation of material and conservative processing of energy. Plants use solar energy to build complex organic molecules out of inorganic materials from the surrounding environment. In consuming plants, animals convert plant material into yet new forms as they grow and derive energy from the organic molecules. After death, organic material decomposes back into inorganic material, diffusing throughout the environment to be reused by photosynthesizing plants. The energy is ultimately lost to the system as heat, but only after cycling through multiple chemical, mechanical, and kinetic forms and powering life forward. Every step in the process, each conversion of matter and energy, is connected to every other.

In cities, on the other hand, material flows linearly; building materials, food, water, and commodities are imported rather than derived from the local environment. Activity within a city depends for the most part on the combustion of fossil fuels, a process that uses up energy stores immediately. In order to become sustainable, different elements of a city--the buildings, roads, railways, urban gardens, public parks, pavements, energy grid, and waste facilities--must be considered as a single system. How does the design of an office building affect urban gardening? What impact does the type of pavement used have on the function of a waste processing plant? Considering potential connections may provide insight into how a city can function as a self-sustaining ecosystem.

References 1. Cities in action Cities in action. (https://www.cdp.net/en/research/ global-reports/cities-infographic-2017/cities-in-action#f6037614bf5 8e91d271ee6f6108ebc8a) 2. Sustainable Building City of Melbourne. (http://www.melbourne. vic.gov.au/building-and-development/sustainable-building/Pages/

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FEATURES sustainable-building.aspx) 3. Palmer, L.(2018). Urban agriculture growth in US cities. Nature Sustainability 1, 5–7.

4. Rowe, A. A Case Study: EPA’s Permeable Pavement Parking Lot. 5. Brown, R. A. and Borst, M. (2014). Quantifying evaporation in a permeable pavement system. Hydrological Processes 29, 2100–2111.

6. Scholz, M. and Grabowiecki, P. (2007). Review of permeable pavement systems. Building and Environment 42, 3830–3836.

7. Urban Watershed Management Branch, Water Supply and Water Resources Division and O’Connor, T. P. Update to permeable pavement research at the Edison Environmental Center.

8. Chwieduk, D.(2003). Towards sustainable-energy buildings.

Applied Energy 76, 211–217.

9. Ramesh, T., Prakash, R. and Shukla, K. (2010). Life cycle energy analysis of buildings: An overview. Energy and Buildings 42, 1592– 1600.

10. Li, D. H., Yang, L. and Lam, J. C. (2013). Zero energy buildings and sustainable development implications – A review. Energy 54, 1–10.

11. Bellia, L., Falco, F. D. and Minichiello, F.(2013). Effects of solar shading devices on energy requirements of standalone office buildings for Italian climates. Applied Thermal Engineering 54, 190–201.

12. Chidiac, S., Catania, E., Morofsky, E. and Foo, S.(2011).

Effectiveness of single and multiple energy retrofit measures on the energy consumption of office buildings. Energy 36, 5037–5052.

13. Sharma, R. and Newman, P.(2017). Urban Rail and Sustainable Development Key Lessons from Hong Kong, New York, London and India for Emerging Cities. Transportation Research Procedia 26, 92–105.

14. Sustainable mobility: Asian and European Cities lead the way Arcadis. (https://www.arcadis.com/en/global/news/latestnews/sustainable-mobilityasian-and-european-cities-lead-theway/2145479/) 15. The City Unlimited: Sustainability Report 2016(2016). Mass Transit Railway.

16. Prosser, I. (2011). Water: Science and Solutions for Australia.

17. Kulak, M., Graves, A. and Chatterton, J.(2013). Reducing greenhouse gas emissions with urban agriculture: A Life Cycle Assessment perspective. Landscape and Urban Planning 111, 68– 78.

18. Specht, K., Siebert, R., Hartmann, I., Freisinger, U. B., Sawicka, M., Werner, A., Thomaier, S., Henckel, D., Walk, H. and Dierich, A.(2013). Urban agriculture of the future: an overview of sustainability aspects of food production in and on buildings.

Agriculture and Human Values 31, 33–51.

19. Zezza, A. and Tasciotti, L.(2010). Urban agriculture, poverty, and food security: Empirical evidence from a sample of developing countries. Food Policy35, 265–273.

20. Lin, B. B., Philpott, S. M. and Jha, S.(2015). The future of urban agriculture and biodiversity-ecosystem services: Challenges and next steps. Basic and Applied Ecology 16, 189–201.

21. Landfills, Municipal Solid Waste EPA. 22. Laner, D., Crest, M., Scharff, H., Morris, J. W. and Barlaz, M. A.(2012). A review of approaches for the long-term management of municipal solid waste landfills. Waste Management 32, 498–512.

23. Waste incineration and public health(2000). Washington: National Academy Press.

24. Waste-to-Energy (Municipal Solid Waste)Waste-to-Energy (Municipal Solid Waste) - Energy Explained, Your Guide To Understanding Energy - Energy Information Administration.

25. Mangalapally, H. P. and Hasse, H.(2011). Pilot plant experiments

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for post combustion carbon dioxide capture by reactive absorption with novel solvents. Energy Procedia4, 1–8.

26. Lombardia, L., Corti, A., Carnevale, E., Baciocchi, R. and Zingaretti, D.(2011). Carbon dioxide removal and capture for landfill gas up-grading. Energy Procedia4, 465–472.

27. Waste-to-Energy (Municipal Solid Waste)Waste-to-Energy (Municipal Solid Waste) - Energy Explained, Your Guide To Understanding Energy - Energy Information Administration.

28. Municipal Waste Treatment Graph 2015 CEWEP Municipal Waste Treatment Graph 2015 Comments.

FEATURES

Gene Drive

By Tamsyn Brann

Solving the Problem of Invasive Species, One Gene at a Time

M

ost people think that the job of the U.S. Customs and Border Protection is to protect our country from those who want to harm us. And indeed, CBP officers dutifully search all luggage for explosives, weapons, and drugs, among other contraband. However, it is not only mankind that must be protected from whatever it is that people aim to bring across borders, it is for nature, too.

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FEATURES A single seed, should the deft hands of a customs official not remove it from the possession of the perpetrator, could wreck an entire ecosystem, and nowhere is there a more pertinent paradigm of the fragile nature of native species than the Galápagos Islands. The species in these islands are not found anywhere else in the world. If in the public imagination this region has become tantamount to a paradise of untouched ecological harmony, the reality is quite different: all of the acclaimed huge tortoises, famous finches, and beautiful sea turtles are currently being threatened by invasive species. On the island of Floreana, for example, the first documented invader was a marooned sailor who grew vegetables to survive. Today, there is a larger and more serious problem: rodents.

occurs.3 However, much of this research is still theoretical and is not yet applicable in eliminating the invasive species of Floreana. Despite laboratory successes, there remains a significant problem regarding the resistance of organisms to the efforts of gene drive. To successfully integrate their mutation of choice (the production of offspring of a single sex, for example) researchers will need to dull the effects of the gene drive until the mutation has been incorporated in the entire population. Then, once the mutation has been successfully integrated, researchers could theoretically “switch it on.”4 The weakness of this solution lies in the strength of mother nature itself: though humans may be able to tamper with a genome that has been evolving for millions upon millions of years, they are still yet no match for the powers of true nature.

“Floreana has one of the highest rates of extinction due to invaders, making it one of the most significant targets for efforts to improve biodiversity in the entire world.“ Floreana has one of the highest rates of extinction due to invaders, making it one of the most significant targets for efforts to improve biodiversity in the entire world. The invasive rodents that eat eggs straight from the nests of native reptiles and birds are a target for elimination—but a current plan to rid the island of these pests involves rat poison, and 400 tons of it. This would negatively affect the lives of pets and livestock and could even endanger children.

“...researchers are hoping to dull the effects of the gene drive for long enough so that a certain mutation can spread throughout the population.”

However, another plan is beginning to take shape involving genetic manipulation using a technique called gene drive. Gene drive involves manipulation of the DNA of mice to alter the chance that the offspring will be of a certain sex; this can be mediated by a few molecular mechanisms.2 Alleles can naturally evolve through these mechanisms to develop a chance of transmission of greater than 50%. In the realm of synthetic gene drive, modules with similar properties are being developed as part of a technique for editing the genomes of laboratory populations. This can be done using CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats), a system used to permanently modify the genes in live cells. For example, gene drive could be used to alter mice such that they produce offspring that are only male. This female-lacking population would not be able to reproduce and would rapidly die out.

There are potential ideas regarding the building of gene drives that could affect several places in the same gene or even multiple genes, in order to slow the rate with which resistance 22 PENNSCIENCE JOURNAL | SPRING 2018

References 1. Hall, S. S. (2017). Could Genetic Engineering Save the Galápagos? Scientific American 317, 48–57.

2. FAQs: Gene drives Wyss Institute - Harvard University 3. Regalado, A. (2017). Can CRISPR restore New Zealand’s ecosystem to the way it was? MIT Technology Review.

4. Callaway, E. (2017). Gene drives thwarted by emergence of resistant organisms. Nature 542, 15–15.

Additional Works Referenced Questions and Answers about CRISPR (2018). Broad Institute.

Gene-edited mosquitoes could end malaria in Africa (2016).

STAT. Mutant mosquitoes. Gene drive. And a bold vision to eradicate Zika (2018). STAT.

An Interview with

Dr. Frances K. Barg

CONDUCTED BY AMANDA PAREDES-BARBEITO What type of research do you normally do?

I do a lot of community-focused research and mixed methods research.

What is mixed methods research?

It is a method where we use both qualitative and qualitative approaches to research and try to have them inform each other.

What prompted you to start investigating the effects of asbestos in Ambler, PA?

Dr. Frances Barg is a medical anthropologist and professor of Family Medicine and Community Health at the University of Pennsylvania. She collaborates on mixed-method and qualitative research across six schools at Penn. The scope of her research includes topics such as emergency contraception, ADHD treatment, depression, the role of self-reliance in managing depressing among older African American adults, and communities affected by hazardous waste. The people of Ambler, Pennsylvania made up one such community, as they had been exposed to asbestos- a naturally-occurring mineral linked to lung cancers such as mesothelioma. Dr. Barg and her colleagues then designed a study targeted to help the residents of Ambler understand the current and longterm impacts of exposure to asbestos.

About 7 or 8 years ago, some members of the Ambler community came to Penn, and spoke with one of my colleagues, Dr. Edward Emmett, who is an occupational and environmental health medicine physician. They were very concerned about the asbestos situation. At the time, the problem had been identified and the EPA was starting to make plans to alleviate the site, but this particular group of community residents were not sure that their concerns were being heard, and they wanted help.

So Dr. Emmet came to me, since I’m a medical anthropologist, and together we designed as study where we went out into the community and tried to identify from the community members’ perspective, what they key issues were. We heard a lot of different things, and a lot of different concerns that the residents were thinking about, they were certainly just afraid and concerned about being exposed to asbestos. They had questions like, “What happens to asbestos when it gets in the air?”, “What happens to asbestos when it gets into the water?”, “If I’ve been exposed to asbestos what can I do about it?”, “Why does mesothelioma (which is a type of cancer you can get from asbestos) seem to cluster in some families and not others?”, “Is there anything preventative you can do about mesothelioma?”.

These were all questions that members in the community had, and what we did is we took each of those questions and we brought scientists together here at Penn, to write what is called a center-grant- a large grant that has a lot of different research projects associated with it. Each of the projects that we designed really was linked to one of the questions that the community members had. So we applied for funding, and got it, from the NIEHS which is an institute of the NIH. Did you encounter any unexpected challenges along the way?

There’s always challenges…depending upon the project. The project that I lead is an epidemiologic project, where we’re looking at the distribution of mesothelioma cases. We’re trying to get some idea of what’s the genuine risk for developing

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23

mesothelioma as well as other asbestos-related diseases, and also to trying to get a sense of how where you live or where you work affects the likelihood of developing mesothelioma. It’s been a really interesting and wonderful project but recruiting people to participate is hard… and with most research studies, that’s always a concern- one that we’re dealing with- but it’s certainly a concern. And in terms of unexpected problems, it actually is proceeding as we’d hoped. Research takes time, community members want answers tomorrow, so it does take time, and that’s always a concern, but it’s actually going very well.

level attention to the problem. It’s no longer legal to manufacture asbestos products in the United States, but there is asbestos mining going on in other parts of the world. Those places face problems. But I think more than just education, we really need to understand how to appropriately remediate places like the Ambler site, and also how to appropriately monitor the site at the time, so that the remediation efforts are sustained.

Why was it so important to you that the residents of Ambler were heard?

We’re still doing the epidemiologic study, and then all the other research projects that are involved are still going on as well.

That’s something that’s really fundamental to this kind of research. We feel that the entire success of our center is based on the fact that we’re studying a problem that matters to the people who are being affected, and are involved in the research. That is really key to any kind of research. At the same time, there’s lots of people who are affected and not all those people are the same. So, different groups of people have different concerns.

Why do you think educating the residents of Ambler about asbestos is so crucial?

I think educating them is very important, but I don’t know that that’s the answer. It’s certainly important for people to understand the risks they’re under, and it’s important to have people recognize what is and isn’t dangerous. At the same time, education isn’t enough. In a situation like we’ve faced with asbestos, we really need larger policy

So, you mentioned you’re still working on the case, what type of work are you doing now?

Have you seen any other communities facing similar problems? If so, how do these compare to Ambler?

There’s a lot of parallel communities around the world.

There’s one very large naturally-occurring asbestos site in Libby, Montana, where asbestos was actually mined. Here, in Ambler, it’s a different situation since it’s one where there was a manufacturing company that made asbestos products and then dumped asbestos waste out into the community. In Libby, it’s different since the asbestos is naturally-occurring. That’s a very different kind of situation, but we’re working very closely with the folks in Libby to collaborate on a number of different research projects there as well. There are also places in Australia and South Africa that have environmental asbestos that we’ve been in touch with.

A playground at the Ambler asbestos NPL site in 1984. Photo courtesy of the EPA 24 PENNSCIENCE JOURNAL | SPRING 2018

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Activated Glia May Induce Developmentally Regulated Inflammation and Synaptic Remodeling in a TSC Mouse Model of Epilepsy Karbi L. Choudhury, University of Pennsylvania, Philadelphia, PA 19104 Brain development requires an initial excess formation of synapses, which is fol¬lowed by synaptic elimination (pruning) before adult synaptic connections are established. Both processes are necessary for proper maturation and function of neuronal networks. There is growing evidence that glial cells play an important role in both synaptic formation and pruning, and several age-specific signaling pathways have been identified. Glial-dependent synaptic modeling occurs mainly during the first 3 postnatal weeks in mice, but may extend during adolescence (3-8 weeks). In addition activated microglia and reactive astrocytes represent a major source of pro-inflammatory cytokines in the brain, which can enhance excitatory synaptic transmission, while suppressing the inhibitory synaptic responses.

Here, I evaluated whether there are specific changes in glial cell numbers, morphology, and expression profiles during key steps of synaptogenesis in a Tuberous Sclerosis Complex (TSC) mouse model of epilepsy. The activation status of mTOR signaling pathway was assessed by Western blotting for phospho-S6, while glial cell densities were assessed by immunohistochemistry for glial cell markers GFAP, CD68, ALDH1L1, and Iba-1. I found that mTOR pathway was activated in the TSC mutants at all ages studied, in both neurons and astrocytes. This was accompanied by increased densities of GFAP-expressing reactive astrocytes, but no significant changes were observed in the densities of CD68expressing activated microglia. However, there was a marked increase in CD68 expressing-cells and a striking change in microglial morphology, suggestive of a phagocytic phenotype. The counts of neither ALDH1L1-nor Iba-1-expessing cells were significantly altered, demonstrating no change in total numbers of astrocytes and microglia. A better understanding of altered glial responses in TSC may help develop new therapies for this disorder.

Introduction Tuberous sclerosis complex (TSC) is a rare genetic disorder in which mutations of the TSC1 and TSC2 genes lead to formation of benign tumors throughout the body.

The upregulation of the mammalian target of rapamycin (mTOR) pathway in response to TSC1/TSC2 inactivation has a primary role in the pathology of TSC. The neurologic symptoms of the disease include seizures, intellectual disability and autism (DiMario et al., 2015). Epilepsy is the most devastating symptom for TSC patients, as 90% of them will develop seizures within their first year of life, which are often therapy-resistant. Effective seizure control in TSC is critical due to the strong association between early seizure onset, refractory epilepsy and poor developmental outcome.1 To date, agents inhibiting mTOR signaling pathway have emerged as the most promising therapeutic agents for TSC; however, their potential significant side effects may limit their clinical use.2 Thus, novel approaches to treat TSC related epilepsy are in high demand.

In addition to its direct role in regulating protein synthesis, mTOR is well known for regulating many functions related to immune defense and inflammation.3 Indeed, analysis of human and TSC mouse model brain tissue has shown glial cells activation and upregulation of proteins involved in complement and cytokine signaling.4, 5, 6, 7., 8 A highly simplified model of the mTOR pathway and its relation to inflammation is presented in Figure 1.

Neuroinflammation is involved in the pathophysiology of a variety of neurological diseases such as multiple sclerosis, Parkinson’s disease, Alzheimer’s disease, autism, and epilepsy.9, 10 Brain inflammation may be involved in seizure generation and development of epilepsy, and conversely seizures themselves can provoke inflammation in the central nervous system.

Figure 1. mTOR/TSC Signaling Pathway Although it is known that there is a significant increase in inflammation in TSC brain tissue, the relationship between glial cell activation, synaptic development, seizure susceptibility and epilepsy has not been studied. We hypothesized that in TSC, activated microglia and reactive astrocytes may partially retain an immature molecular profile, which will allow them to express a variety of factors/proteins required for synaptic remodeling (formation and/or elimination of synapses). This may contribute to the maintenance of a form of immature synaptic function characterized by increased neuronal excitability and decreased synaptic plasticity.11 Therefore, in this study I investigated the time course of altered glial cell responses in TSC, and I also seek to explore possible correlations between the degree of inflammation and the weakening or strengthening of synapses in future experiments. We SPRING 2018 | PENNSCIENCE JOURNAL 25

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Table 1. Antibodies Used in Western Blotting and Immunohistochemistry Experiments anticipate that these studies will reveal new mechanisms and time windows of vulnerability for epilepsy development in TSC. These mechanisms could serve as targets for novel treatments, which, when administered within the optimal time window, could prevent or cure epilepsy in TSC. In addition, such anti-inflammatory treatments can be conducted even at an early age without the side effects of mTOR inhibitors.

Methods Animals Tsc1cc Nestin-rtTA+ tet-OP-cre+ mice were produced by 26 PENNSCIENCE JOURNAL | SPRING 2018

timed inactivation of Tsc1 in neuronal progenitor cells at embryonic day (E)13.6 This was achieved by treating pregnant dams with doxycycline to induce recombination at the Tsc1 conditional (c) allele in the pups. To assess recombination levels, DNA was isolated from a small portion of the cortex from each mouse that received doxycyline.

Levels of recombination were determined by quantitative copy number analysis of the conditional Tsc1c and null Tsc1 alleles through a multiplex ligation-dependent probe assay (MLPA). Only mice with over 12% recombination were used as TSC model.

Tissue Collection and Perfusions

RESEARCH Animals were anesthetized with Nembutol (3 uL per gram). Once the animal reached a surgical plane of anesthesia, it was perfused with ice cold PBS. Brain tissue collected for Western blot was dissected and frozen directly after perfusion with PBS. The animal was further perfused with 4% paraformaldehyde if the brain was being collected for fixation. Fixed brains were post fixed overnight in 4% paraformaldehyde and then cryoprotected in 30% sucrose for 2 days.

Western Blot Tissue was homogenized with a glass dounce homogenizer. Concentrations of protein in homogenized tissue samples were determined using a Bradford assay with standards from 0-2 mg/ml and the Microplate Manager Software. Samples were then equalized with 1X Lysis buffer and 4X Sample Buffer to .8µg/ 1uL in a total volume of 35 uL. 30uL of each sample was loaded into a precast 4-20% Tris-Glycine precast gel and run at 115 V for 90 minutes. Afterwards, the gel was transferred to a membrane overnight. The membrane was probed with markers of the mTOR signaling pathway (See Table 1). The primary antibodies were applied overnight at 4°C. Actin or S6 were used as loading controls (Table 1). Blots were incubated for 1 hour at room temperature in peroxidase- conjugated mouse or rabbit secondary antibody at a 1:2000 dilution.

Tissue Sectioning and Immunocytochemistry Fixed tissue was mounted in Tissue-Tek, sectioned at 60 μm using the cryostat, and processed as free floating sec-

tions. Antigen retrieval was performed on the free-floating sections using Citrate Buffer and washed in PBS. Sections were double labeled in primary (See Table 1) and corresponding mouse or rabbit secondary antibody (1:1000) using 5% goat serum in PBS and 0.1% Triton. Upon staining, the sections were mounted onto slides. Slides were then cover slipped using Fluoromount with DAPI, a blue nuclear stain, and viewed using an epifluorescence microscope. Images were captured at 10x and 20x magnification (n=5/brain). Single and double labeled cells were counted for every image using ImageJ software.

Statistical Analyses For Western blot analysis, TSC-specific measures were normalized to the appropriate age-matched control samples run on the same blot. To determine the statistical significance, group differences were tested using t-tests within individual age groups. For quantitative immunohistochemistry analysis, an average cell count per mm2 was calculated for each brain, and cell densities where compared using two-way ANOVA adjusted by multiple comparisons.

Results Differential expression of mTOR activity marker phosphoS6 in TSC mutants and controls during early postnatal brain development The first step in our research was to confirm a time-sensitive activation of the mTOR pathway in TSC mutant mice compared to wild-type controls. Utilizing 3 time points in

Figure 2. Developmental Profile of mTOR activation in TSC mutants

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RESEARCH the mouse developmental period (P7, P15, and P30), we assessed levels of phosphorylated ribosomal protein S6 (pS6), a protein downstream of TSC1/TSC2 and mTOR (Figure 2; mutants are represented by the red bar graphs and controls by the blue bar graphs). PS6 has been widely implicated in cell proliferation and growth, and the phosphorylated form over the total protein (S6) levels constitutes an established readout of mTOR activity. At the P7 time point, we see an increase in the levels of pS6 normalized to S6 in the mutants (403±163%, n=5) over the controls (100±41%, n=5), although this increase is not significant (p=0.109). At P15, however, the overall increase in pS6 to S6 ratio is significantly higher in the TSC mutants (324±15%, n=8; p<0.0001) over controls (100±9%, n=8). Finally, P30 mutant mice exhibit also a significantly elevated pS6 to S6 ratio (350±10%, n=6; p<0.0001 over controls (100±12%, n=6).

Next, we sought to establish where exactly pS6 is expressed in a cell-specific manner in the cortex of both TSC muFigure 3. Cell-Specific pS6 Expression in TSC Control & tants and wild-type controls (Figure 3). Again, pS6 elevaMutant Cortexmutants tion indicates a general increase in activation of the mTOR pathway. Using immunohistochemical staining methods and P30-P50). and epifluorecsence imaging, we qualitatively visualized expression of pS6 (green) in the TSC and control brain at In contrast, the total number of astrocytes (Panel E), both P7 (Panels A-F) and P15 (Panels G-I). In Panels A-F, was not significantly different in the mutants relative to sections were double labeled with NeuN (red), a neuronal age-matched controls (p>0.05), but there was an obvimarker. Panels G-I show double labeling with GFAP (red), ous decreasing trend from P7 to P30 in both genotypes an astrocyte-specific marker. In Panels A-C, we see nor- (P7 control vs mutant: 1080 cells/mm2, n=2 vs 897 cells/ mal expression of pS6 in control cortical neurons. In con- mm2, n=2; P15 control vs mutant: 459 cells/mm2, n=2 trast, the levels of pS6 are elevated in a dramatic fashion vs 623 cells/mm2, n=2; P30-50 control vs mutant: 461 in the TSC cortical neurons, as indicated by Panels D-F, cells/mm2, n=1 vs 485 cells/mm2, n=1). The correspondalthough the total number of neurons appears unchanged ing representative images show that TSC mutants do not (Panel A and D). Panels G-I show that pS6 induction is seem to express any significant changes in total number not limited to neurons, but is also quite robust in astro- of astrocytes from their respective age-matched controls cytes in the TSC mutants. Magnification values for each (Panels F, G, & H). image are indicated in the upper left corner.

Finally, regardless of age, we observed a significant change Age-specific astroglial phenotypes in TSC mutants and con- in astrocytes morphology. In control mice, most astrocytes displayed a stellated shape with relatively small cell bodies trols As activation of mTOR in TSC astrocytes may impact their and thin processes, denoting a resting state. In contrast, in normal development, in this set of experiments I aimed to the TSC mice, astrocytes showed enlarged cell bodies and assess potential changes in astrocytes numbers, along with shorter, thicker processes (Panels B-D & F-H). alterations in cell morphology and expression profile indicative of cellular activation. Therefore, I have performed Characterization of microglial phenotypes in TSC mutants qualitative and quantitative immunohistochemical analy- and controls during brain development sis at different ages, spanning from ages P7 to P50 (Figure Although Tsc1 levels are preserved in microglia in our 4). Reactive astrocytes (in response to mTOR upregula- mouse model, and we did not see mTOR activation in this tion) were identified by GFAP staining, while total astro- cell type, we also studied microglial reaction in the TSC cytes were identified by expression of ALDH1L1. Panel and control mice at the same ages, as this is a very sensitive A shows that at all time points analyzed, the densities of indicator of brain inflammation. Activated microglia can GFAP+ reactive astrocytes are significantly higher in the be visualized by probing for CD68 in the immunohistocortex of TSC mutants (p<0.001), compared to controls chemistry, while total microglia are measured by visual(P7 control vs mutant: 166 cells/mm2, n=3 vs 817 cells/ ization of Iba1. mm2, n=4; P15 control vs mutant: 287 cells/mm2, n=4 vs 857 cells/mm2, n=4; P30-50 control vs mutant: 268 cells/ In Figure 5, the quantification of activated microglia with mm2, n=4 vs 1183 cells/mm2, n=3). In the accompanying CD68 (Panel A) indicates no significant changes in cell images (Panels B, C, & D) we see that the controls (left) densities in TSC mutants over controls (p>0.05), also they express significantly less GFAP than the corresponding trended higher in mutants at P15 (P7 control vs mutant: age-matched mutants (right) at every time point (P7, P15, 324 cells/mm2, n=2 vs 361 cells/mm2, n=5; P15 control vs 28 PENNSCIENCE JOURNAL | SPRING 2018

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Figure 4. Reactive Astrocytes and Total Astrocytes in TSC Controls and Mutants. Note. Scale bar represents a length of 25 microns. mutant: 325 cells/mm2, n=2 vs 541 cells/mm2, n=2; P3050 control vs mutant: 397 cells/mm2, n=2 vs 381 cells/ mm2, n=2). In the corresponding images, however, this difference is much more stark and convincing. The mutants (right) at all time points express much more CD68, a lysosomal protein associated with phagocytosis, than their age-matched controls (Panels B, C, & D), suggesting a robust phagocytic microglial phenotype in the mutants. This is accompanied by striking morphological changes (hy-

pertrophied, amoeboid cell bodies), which may allow for the engulfment and elimination of targeted brain structures (Schafer et al., 2012). However, whether these microglial cells indeed have phagocytic properties requires further investigation.

For total microglial densities (Figure 5, Panel E), there seem to be no differences between mutants and controls, however, there does seem to be more sparse Iba-1+ cells at

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Figure 5. Activated Microglia and Total Microglia in TSC Controls and Mutants. Note: Scale bar represents a length of 25 microns.

P30-50 in the mutants compared to controls (P7 control vs ing a possible contributing role of these cells in seizure mutant: 594 cells/mm2, n=2 vs 535 cells/mm2, n=5; P15 generation. Much of the trends presented here, however, control vs mutant: 539 cells/mm2, n=2 vs 632 cells/mm2, will require an increase in the sample size (n), and such n=2; P30-50 control vs mutant: 449 cells/mm2, n=5 vs 308 adjustments will need to be made before conclusions can cells/mm2, n=7). The corresponding immunostaining be drawn. Due to the limited time available for this project throughout the semester, the sample sizes of each age also depicts this trend (Panels F, G, & H). cohort in both mutants and controls were quite low. In addition, garnering statistical significance for every asDiscussion pect of inflammation is an important goal in neurologiOverall, the preliminary data presented here seem to sup- cal research. Many of the graphs, including ALDH1LI port a trend of increased glial activation in TSC, preced- and CD68, failed to achieve statistical significance due to ing the age-related onset of seizures (P21-P20), suggest- biological variability in the small number of samples. An 30 PENNSCIENCE JOURNAL | SPRING 2018

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Figure 6. Astrocyte and Microglial Cell-Mediated Phagocytosis and Pruning of Weak Synapses interesting follow-up to this data would be a study of inflammatory trends in human TSC tissue. Also, rectifying the low sample sizes would bolster the validity of the patterns seen in this paper. Such favorable trends, in all, look promising and are an exciting next step for new findings in epilepsy and TSC. Astrocytes and microglia have been implicated in the pathophysiology of many neurodevelopmental diseases such as epilepsy, autism and schizophrenia, which are all characterized by defects in synaptic function. Thus, these cell types may be targets for the development of new therapeutic agents to treat these diseases. However, the mechanism by which glial processes mediate synaptic dysfunction and overall cognitive decline in these disorders are poorly understood. There have been many hypotheses suggested regarding astrocytes inducing formation of aberrant glutamatergic connections, or microglia and astrocyte cooperatively pruning the unwanted synapses.11 In Tuberous sclerosis and epilepsy, for instance, astrocytes and microglia may undergo similar mechanism of activation leading to dysfunctional synaptic remodeling. The impact of glial cell activation on neuronal function and plasticity in TSC is likely multifold. Astrocytes control the number, effectiveness and stability of synapses through a variety of secreted signals (Figure 6), including the extracellular matrix proteins thrombospondins (TSPs), secreted protein acidic and rich in cysteine (SPARC) family proteins and glypicans.12, 13, 14 Microglia, the resident CNS immune cells, represent key regulators of developmental

structural spine plasticity (Tremblay et al., 2010). Microglia are able to phagocyte synaptic elements, a restricted developmental process mediated by immune molecules, including complement proteins C1q and C3 (Figure 6).

C1q, the initiating protein of the classical complement cascade, localizes to synapses during synaptic pruning, and its expression is stimulated by transforming growth factor (TGF)- β released from astrocytes.16 The final step of synapse elimination is mediated by an interaction between the complement component C3 and its high-affinity receptor CR3 expressed on microglia.17, 18 In addition, activated microglia and reactive astrocytes represent the main source of pro-inflammatory cytokines in the brain, which can have a significant impact on synaptic function.

For example, the excitatory synaptic transmission is potentiated by in vitro application of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF- α), concomitant with suppression of inhibitory synaptic transmission, with most potent effects noted for the pro-inflammatory cytokine IL-1β.19 The role of these processes in epilepsy and Tuberous sclerosis, however, has not been extensively studied. This is a great avenue for research for precisely this reason. How synaptic remodeling can precisely lead to the disease phenotype of epilepsy in TSC is a future direction that my lab hopes to further comprehend.

Acknowledgements I would like to thank my mentor and principal investigator, Dr. Delia Talos, for assisting me in my project and

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RESEARCH helping it come to fruition.

References 1. DiMario, F.J., Jr., M. Sahin, and D. Ebrahimi-Fakhari, Tuberous sclerosis complex. Pediatr Clin North Am, 2015.

62(3): p. 633-48.

2. Maroso, M., Balosso, S., Ravizza, T., Iori, V., Wright, C.I., French, J., Vezzani, A. Interleukin-1beta Biosynthesis Inhibition Reduces Acute Seizures and Drug Resistant Chronic Epileptic Activity in Mice. Neurotherapeutics, 2011.

3. Saxton, R.A. and D.M. Sabatini, mTOR Signaling in Growth, Metabolism, and Disease. Cell, 2017. 168(6): p.

960-976.

4. Boer, K., Crino, P.B., Gorter J.A., Nellist, M., Jansen FE, Spliet WG, van Rijen PC, Wittink FR, Breit TM, Troost D, Wadman WJ, Aronica E. Gene expression analysis of tuberous sclerosis complex cortical tubers reveals increased expression of adhesion and inflammatory factors. Brain Pathol, 2010. 20(4): p. 704-19.

5. Boer, K., Jansen F., Nellist M., Redeker S., van den Ouweland A.M., Spliet W.G., van Nieuwenhuizen O., Troost D., Crino P.B., Aronica E. Inflammatory processes in cortical tubers and subependymal giant cell tumors of tuberous sclerosis complex. Epilepsy Res, 2008. 78(1): p. 7-21.

6. Goto, J., Talos D.M., Klein P., Qin W., Chekaluk Y.I., Anderl S., Malinowska I.A., Di Nardo A., Bronson R.T., Chan J.A., Regulable neural progenitor-specific Tsc1 loss yields giant cells with organellar dysfunction in a model of tuberous sclerosis complex. Proceedings of the National Academy of Sciences of the United States of America, 2011. 108(45): p. E1070-9.

7. Zhang, B., Zou J., Han L., Rensing N., Wong M., Microglial activation during epileptogenesis in a mouse model of tuberous sclerosis complex. Epilepsia, 2016. 57(8): p.

1317-25.

8. Zhang, B., Zou J., Rensing N.R., Yang M., Wong M.. Inflammatory mechanisms contribute to the neurological manifestations of tuberous sclerosis complex. Neurobiol Dis, 2015. 80: p. 70-9.

9. Vezzani, A., Aronica E., Mazarati A., Pittman Q.J.. Epilepsy and brain inflammation. Exp Neurol, 2013. 244: p.

11-21.

10. Vezzani, A., French J., Bartfai T., Baram T.Z. The role of inflammation in epilepsy. Nat Rev Neurol, 2011. 7(1): p. 31-40.

11. Clarke, L.E. and B.A. Barres, Emerging roles of astrocytes in neural circuit development. Nature Neuroscience, 2013. 14: p. 311-21.

12. Liauw J., Hoang, S., Choi, M., Eroglu, C., Choi, M., Sun, G., Percy, M., Wildman-Tobriner, B., Bliss, T., Guzman, R.G., Barres, B.A. Thrombospondins 1 and 2 are necessary for synaptic plasticity functional recovery after stroke. Journal of Cerebral Blood Flow and Metabolism.

2008 Oct 1; 28 (10): 1722-32.

13. Christopherson, K.S., Ullian, E.M., Stokes, C., Mullowney, C.E., Hell, J.W., Agah, A., Lawler, J., Mosher, D.F., Bornstein, P., Barres, B.A. Thrombospondins are astrocyte-secreted proteins that promote CNS synaptogenesis. Cell. 2005 February 11; 120(3): 421-433.

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14. Kucukdereli, H., Allen, N.J., Lee, A.T., Feng, A., Ozlu, M.I., Conatser, L.M., Chakraborty, C., Workman, G., Weaver, M., Sage, E.H., Barres, B.A., Eroglu, C. Control of excitatory CNS synaptogenesis by astrocyte-secreted proteins Hevin and SPARC. PNAS. 2011 Jun 21; 108(32): 440-449.

15. Allen N.J., Bennett M.L., Foo L.C., Wang G.X., Chakraborty C., Smith S.J., Barres B.A. Astrocyte glypicans 4 and 6 promote formation of excitatory synapses via GluA1 AMPA receptors. Nature. 2012 May 27; 486(7403): 410-4..

16. Bialas, A.R. & Stevens, B. TGF-β signaling regulates neuronal C1q expression and developmental synaptic refinement. Nature Neuroscience. 2013 October 27; 16: 1773-82.

17. Schafer, D.P., Lehrman, E.K., Kautzman, A.G., Koyama, R., Mardinly, A.R., Yamasaki, R., Ransohoff, R.M., Greenberg, M.E., Barres, B.A., Stevens, B. Microglia sculpt postnatal neural circuits in an activity and complementdependent manner. Nature. 2012 May 24; 74(4): 691-705.

18. Stevens B., Allen N.J., Vazquez L.E., Howell G.R., Christopherson K.S., Nouri N., Micheva K.D., Mehalow A.K., Huberman A.D., Stafford B. The classical complement cascade mediates CNS synapse elimination. Cell.

2007 Dec 14; 131(6): 1164-78.

19. Kawasaki, Y., Zhang, L., Cheng, J.K., Ji, R.R. Cytokine mechanisms of central sensitization: distinct and overlapping role of interleukin-1beta, interleukin-6, and tumor necrosis factor-alpha in regulating synaptic and neuronal activity in the superficial spinal cord. J Neurosci, 2008.

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Theoretical Analysis of Bond Dissociation Enthalpy: A DFT Study of Various Antioxidants with Hydroperoxyl and Hydroxyl Radicals Vraj Rasesh Shroff (व्रज रसेश श्रॉफ), University of Pennsylvania, Philadelphia, Pennsylvania, U.S.A. Antioxidants are known to treat many chronic diseases. The goal of this research is to find the most efficient antioxidant among alpha-Tocopherol, Zeaxanthin, Kaempferol, and Idebenol. Each of the aforementioned antioxidants is supposed to represent the natural, non-enzymatic subfamily of the antioxidants they come from. Their effectiveness is measured by calculating their bond dissociation enthalpy when they are oxidized by Hydroperoxyl radical and Hydroxyl radical.

Bond dissociation enthalpy is calculated after geometrically optimizing the molecule using General Atomic and Molecular Electronic Structure System (GAMESS) Density Functional Theory (DFT) Routine (B3LYP 6-31G(d)) basic set. DFT spin-polarized technique was used for the radicals. The results suggest that the antioxidants in question have efficiency in the following order: Zeaxanthin < Kaempferol < alpha-Tocopherol < Idebenol.

Literature Analysis Introduction to Antioxidants Antioxidants are molecules that can donate one (or more) of their electrons to neighbouring molecules to stabilize them. At the molecular level, antioxidants scavenge free radicals or charged molecules in their environment and stabilize them by giving away their electron. Such processes help to counteract the effect of oxidative stress in the human body. In the absence of antioxidants, the charged molecules grab electrons from any molecule in their vicinity; consequently, other molecules become charged.

Hence, the resulting chain reaction will destroy the atom and often the entire molecule.1 Biologically, such radicals will destroy the DNA, which in turn damages the nucleus and the entire cell. Abundance of oxidative stress can cause serious damage because it is known to mutate DNA and damage the self-induced apoptosis checkpoints, which can further the damage done to the cell. As a result, antioxidants play a very crucial role in regulating the equilibrium in the cell.2 Antioxidants are known to treat many chronic diseases.

For instance, antioxidants play a significant role in preventing medical conditions such as cancers, macular degeneration, Alzheimer’s disease, and arthritis-related diseases. Conversely, oxidative stress plays a significant role in the progression of diseases like atherosclerosis, inflammatory conditions, and the process of aging. As a result, it is beneficial to further understand their structures and exploit antioxidants as a treatment for many of these diseases.3 Function of Antioxidants The first group of antioxidants synthesizes more antioxidants and provides compensation for the used up electron donors. Antioxidants are constantly being prepared in anticipation of oxidative stress being produced in the body.

Since oxidative stress is generated in the body due to sun radiation, consumption of food, pollution, metabolism and many other processes, the body strives to maintain the balance between the amount of oxidative stress generated and antioxidants synthesized to counteract that stress.

The second group of antioxidants prevent generation of reactive oxygen species (ROS) -- charged molecules like

peroxides, superoxide, Hydroxyl radical, and singlet oxygen.

The third group of antioxidants neutralize ROS and break the chain reactions of free radicals by donating electrons to them. They respond to the damage as it is affecting the cell. This last group of antioxidants function as the repair team. They detoxify and repair the damage done to the cell by these free radicals. All these different types of antioxidants work together to maintain homeostasis in the body and protect it from any damage.⁴ Classification of Antioxidants While antioxidants can be grouped by their function, it is convenient to categorize them by their chemical structure, origin, and properties to make a broad observation about their functionality. At first, based on their source, they can be divided into natural antioxidants and synthetic compounds.

Natural antioxidants are either synthesized in the human body through metabolic process or are supplemented from other natural sources. They are further divided into two categories: enzymatic antioxidants and nonenzymatic antioxidants.

Enzymatic antioxidants are subdivided into primary and secondary antioxidants. Primary enzymatic antioxidants include superoxide dismutase, catalase, and glutathione peroxidase, and secondary enzymatic antioxidants include glutathione reductase and glucose-6-phosphate dehydrogenase.

Nonenzymatic antioxidants are classified into vitamins, carotenoids, polyphenols, and other antioxidants. These antioxidants are essential to metabolism but they are not naturally produced in the bodies so they have to be supplemented.⁵ This research will focus on four natural, non-enzymatic antioxidants: alpha-Tocopherol (vitamin), Zeaxanthin (carotenoid), Kaempferol (polyphenol), and Idebenol (others / non-protein). These antioxidants chosen from each category will be used to better understand the correlations between efficiency and the molecular structure.

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RESEARCH Role of Reactive Oxygen Species (ROS) This research will involve reduction of Hydroxyl and Hydroperoxyl radicals. There are several types of radicals, but those of most concern in biological systems are derived from oxygen and are collectively known as reactive oxygen species. Oxygen has two unpaired valence electrons, and such structure makes oxygen susceptible to radical formation. Oxygen-derived radicals are generated constantly during day-to-day activities. For instance, they are formed in mitochondria by the electron transport chain.

Oxygen radicals are overproduced in white blood cells to kill pathogens, in cells exposed to hypoxia or hyperoxia environments, and/or in cells exposed to radiation.

Although radicals are generated in many essential reactions, they are toxic to cells. Their possession of unpaired electrons make them highly reactive and consequently, they are potent to damage lipids, proteins, nucleic acids and other molecules.

One of the best known toxic effects of oxygen radicals is damage to cellular membranes by the process of lipid peroxidation. The following reaction depicts peroxidation of a fatty acid. Such reactions are usually chain reactions: as shown in Figure 2, a hydrogen atom is taken away from the fatty acid by a Hydroxyl radical, leaving a carbon-centered radical as part of the fatty acid. That fatty acid then reacts with oxygen to yield the peroxyl (hydroperoxyl) radical, which can react with other molecules, increasing the proportion of radicals in the cell.

Density Functional Theory (DFT) B3LYP with the basic set routine: 6-31G(d) was used. DFT spin-polarized technique was used for radicals.

Generic equations 1) Antioxidant + Hydroperoxyl Radical â&#x2020;&#x2019; Oxidized Antioxidant Molecule + Hydrogen Peroxide.

2) Antioxidant + Hydroxyl Radical â&#x2020;&#x2019; Oxidized Antioxidant Molecule + Water.

This research focuses on four antioxidants and two radicals: alpha-Tocopherol, Zeaxanthin, Kaempferol, and Idebenol with Hydroperoxyl and Hydroxyl radicals. Bond dissociation enthalpy was used to compare the efficiency among different antioxidants for reducing these radicals.

Results [Figures on next page] Discussion The calculations show that antioxidants can stabilize reactive oxygen species while they become less stable. The antioxidant should not become more unstable than its radical form becomes stable. In other words, the amount of energy gained by the antioxidant can be at most equal to the amount of energy released by the radical. If the amount of energy gained by the antioxidant is greater than the amount of energy released by the radical, then the redox reaction will likely not occur in the cell.

Efficiency of alpha-Tocopherol Alpha-Tocopherol gains 0.615 Ha of energy while it gets oxidized by Hydroperoxyl radical. However, when the Hydroperoxyl radical gets reduced by alpha-Tocopherol, it releases around 0.630 Hartrees (Ha) of energy . Hence, this system overall attains a lower energy state and becomes more stable. The products are more stable by approximately 0.016 Ha, or 41.03 kJ/mol. Such negative bond dissociation enthalpy shows that the redox reaction will be favored and net stability will be achieved. Such property of alpha-Tocopherol makes it an efficient antioxidant in scavenging Hydroperoxyl radicals.

Figure 1 shows the reduction of a fatty acid and formation of Hydroxyl and Hydroperoxyl radicals which are known to damage cells.

Methods Bond dissociation enthalpy was calculated for each antioxidant, radical, and product. The molecule was geometrically optimized using Web Molecular Orbital (WebMO) calculations. General Atomic and Molecular Electronic Structure System (GAMESS) engine was used for these ab initio and semiempirical calculations. Furthermore, 34 PENNSCIENCE JOURNAL | SPRING 2018

Furthermore, alpha-Tocopherol is also efficient in neutralizing Hydroxyl radicals. Similar calculations show that the products are more stable by 0.067 Ha or 175.82 kJ/ mol. It should be noted that alpha-Tocopherol is better at scavenging Hydroxyl radicals. It can make its products more stable by almost 0.051 Ha or 134.79 kJ/mol for the Hydroxyl reaction.

Efficiency of Idebenol Idebenol is an efficient antioxidant as well. It makes the Hydroperoxyl radical and antioxidant system more stable by 0.018 Ha or 47.87 kJ. It also makes Hydroxyl and antioxidant system more stable by 0.070 Ha or 182.66 kJ/mol.

It should be noted that Idebenol is a better antioxidant at neutralizing either Hydroperoxyl or Hydroxyl radical in

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Table 1 shows the net bond dissociation enthalpy for each antioxidant to reduce the Hydroperoxyl radical.

Table 2 shows the net bond dissociation enthalpy for each antioxidant to reduce the Hydroxyl radical. comparison to alpha-Tocopherol.

Efficiency of Kaempferol Kaempferol is truly efficient at scavenging only the Hydroxyl radicals. Reducing Hydroxyl radicals makes this system more stable by 0.051 Ha or 133.86 kJ/mol. But after becoming oxidized by Hydroperoxyl radicals, the radicalantioxidant system does not get any more stable. In fact, if results were utopical, then the system ends up a little more unstable, by 0.0004 Ha or 0.94 kJ/mol. However, conclusions at such extremely small values should be viewed with skepticism. Such change in enthalpy is more likely a byproduct of theoretical calculations of self-contained systems than any infinitesimally small real change in the enthalpy of the system. Regardless of the reason, Kaempferol can not efficiently neutralize Hydroperoxyl radicals due to the net positive enthalpy of the system. It should be noted that such property of Kaempferol is in contrast

of alpha-Tocopherol and Idebenol and the reason is likely due to its molecular structure and orbital energies.

Efficiency of Zeaxanthin In contrast to the above antioxidants, Zeaxanthin is truly inefficient at scavenging Hydroperoxyl radicals. The products have a positive enthalpy of 0.031 Ha or 81.10 kJ/mol.

Such results indicate that Zeaxanthin will quite likely not participate in such reaction. In contrast, the products of redox reaction of Zeaxanthin and Hydroxyl have a significantly negative enthalpy of -0.020 Ha or -53.69 kJ/mol. So, Zeaxanthin will reduce Hydroxyl radicals to reach a more stable system.

Comparison of Efficiency of All the Antioxidants Results point out that alpha-Tocopherol and Idebenol have similar effects. While Kaempferol comes somewhere in between, being neutral at best (in terms of reduction of

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Figure 2 shows the net bond dissociation enthalpy for each antioxidant reaction to reduce the respective radical.

Hydroperoxyl radicals). However, Zeaxanthin has a very unlikely reaction potential in the same regards. In terms of reduction of either radical, the following is the comparison of the efficiencies of all the antioxidants studied in this research: Zeaxanthin < Kaempferol < alpha-Tocopherol < Idebenol.

Conversion of Harmful Products by Antioxidants Reduction of reactive oxygen species by antioxidants led to formation of some sort of hydrogen oxide. When Hydroperoxyl was reduced, H2O2 formed; Hydroxyl made H2O on reduction. While H2O is a safe byproduct of such redox reaction, H2O2 can be quite harmful for the molecule or the cell. Furthermore, UV light or metal catalysis can convert H2O2 to Hydroxyl radical, bringing the system back to the beginning. As a result, the cell requires it to be transformed to something else.

Consequently, cell uses another antioxidants, namely Catalase (CAT), to transform H2O2 into H2O and O2. Similarly, glutathione (GSH) can be used to reduce H2O2 to H2O and lipid hydroperoxide.â ˇ

36 PENNSCIENCE JOURNAL | SPRING 2018

Recycling of Antioxidant Radicals It is unlikely that such redox reactions between an antioxidant and the radical will happen in isolation. In fact, most reactions in the cell take place as part of a cyclic process of neutralizing the radical and regenerating the antioxidant.

Often, even if the direct conversion of radical to a stable molecule is not favorable, the reaction will still take place because, in the larger scheme of reactions, the reaction comprises just one of the parts of a step-by-step reaction which leads to a more stable system.

When alpha-Tocopherol reduces the Hydroperoxyl or Hydroxyl radical, it becomes oxidized and alpha-Tocopherol itself becomes a radical. However, alpha-Tocopherol is a lot more stable in comparison to Hydroperoxyl or Hydroxyl radical. Specifically, it is 1133.36 kJ/mol and 1208.51 kJ/ mol lower in energy in comparison to Hydroperoxyl and Hydroxyl radicals, respectively. Nevertheless, it is still a radical which needs to be reduced.

Figure 12 shows one of the ways cells regenerate alphaTocopherol from alpha-Tocopheroxyl, with the help of ascorbic acid, another antioxidant. First, the Hydroperoxyl radical gains an electron as discussed earlier. Then, the same process happens for alpha-Tocopheroxyl which helps it transform back to alpha-Tocopherol. In this process, ascorbic acid is reduced to dehydroascorbic acid which is still a stable product. Hence, through this process, the radical is completely eliminated through the system. This loop continues until levels of reactive oxygen species is back to normal.â ¸

RESEARCH

Figure 3 shows a part of the cyclic chain reaction of reducing radicals and regenerating antioxidants. Here, ROO is Hydroperoxyl radical, ROOH is hydrogen peroxide, αTOH is alpha-Tocopherol, αTO is alpha-Tocopheroxyl, A- is ascorbate radical, AH- is ascorbic acid, and A is dehydroascorbic acid.

In similar ways, other antioxidant radicals are reduced with the help of other antioxidants in the presence of various antioxidant-radical systems.

Conformity to Previous Researches There is an abundance of scientific studies that measure the efficiency of various antioxidants. Different methods are often used, frequently leading to contrasting comparisons. An ideal experimental setup is still lacking because of certain properties of antioxidants and free radicals. Antioxidants act as radicals themselves upon oxidation, and radicals are very reactive, having a very short lifespan.

Consequently, experiments inherently have uncontrolled variables, which lead to different results depending on the method used. In such situations, theoretical analysis is useful to compare antioxidants in an as much controlled environment as possible.

Research of Stocker et al. on Ubiquinol-10 and alphaTocopherol produced similar results as this current study.

They researched the rate of consumption of Ubiquinol-10 and alpha-Tocopherol antioxidants when they were kept in proximity and exposed to radicals. They found that Ubiquinol-10 was used up first and later concluded that Ubiquinol was a better antioxidant than alpha-Tocopherol. Ubiquinol-10 and Idebenol are very similar in structures, thus supporting our results.⁹ Husain and his lab researched several antioxidants of the flavonoids category (i.e. Kaempferol). They produced Hydroxyl radicals under stimulated conditions and then observed the percentage of diminution of radicals with chromatogram and other techniques. They concluded that Kaempferol was second worst at scavenging Hydroxyl radicals. In their experiment, the worst antioxidant was Flavone which was not a part of this current experiment.10 Such results are expected because of the almost 0 kJ/mol enthalpy released. Since the overall system does not become more stable, such redox reactions are unlikely to take place.

Narayan and his team found similar results when they directly tested the reduction of lipid peroxidation upon addition of certain antioxidants. For their experiment, Kaempferol was the second worst antioxidant as well.11 Their worst antioxidant was not part of our experiment either. Such experiments support the results of our study.

Hennekens and his team performed a study using a group of around 22,000 people to see the effect of beta-carotene, a compound of the same class as Zeaxanthin. The group was randomly divided into two subgroups, one with placebo and one with beta-carotene. They noticed that there were no significant differences in the number of cases of lung cancer (82 in the beta-carotene group vs. 88 in the placebo group); the number of deaths from cancer (386 vs. 380), deaths from any cause (979 vs. 968); deaths from cardiovascular disease (338 vs. 313); the number of men with myocardial infarction (468 vs. 489); or the number with stroke (367 vs. 382).12 Such results prove that Zeaxanthin is not an effective antioxidant. It does not have the properties of a typical antioxidant which has clinical value or which can serve as a treatment for many chronic diseases. Such results concur with this experiment since the results indicate that the redox reaction by Zeaxanthin is unlikely due to the positive enthalpy of the system.

It should be noted that the underlying reasons and pathways of how antioxidants function are still unknown. It is possible that Zeaxanthin can be an efficient antioxidant if it were to reduce some other radical. Even changes to environment may improve its efficiency. However, such parameters are outside the scope of this experiment.

Caveat of Bond Dissociation Enthalpy Calculating efficiency with bond dissociation enthalpy/ energy is commonplace in research studies. However, several differences exist in such computations. Some experiments dissolve antioxidants in a given solvent like water or alcohol and then perform calculations. Others use a molecule’s bond dissociation energy as a fixed point and then compare other molecules on a relative basis. Still others use a different temperature, such as the room temperature. Along with these, there are many other variations.13, 14 Hence, the exact enthalpy is of less importance than the relative comparison and the molecular structure of the most efficient antioxidant.

This is not to say that bond dissociation enthalpy is the sole indicator of efficiency. Several other techniques exist; strictly experimental research investigations have compared rates of reactions of different antioxidants. However, most experiments use some combination of O-H bond dissociation enthalpy (BDE), ionization potential (IP), proton dissociation enthalpy (PDE), proton affinity (PA), and/or electron transfer enthalpy (ETE).

Conclusion and Future Prospects This research concludes that all the antioxidants in question -- Zeaxanthin, Kaempferol, alpha-Tocopherol, Idebenol -- are efficient at scavenging Hydroxyl radicals while only alpha-Tocopherol and Idebenol are efficient at tackling Hydroperoxyl radicals. Both radicals were best reduced by Idebenol and least reduced by Zeaxanthin. Such conclusions can be drawn back to their subclasses as well.

Classification of antioxidants by structure and origin al

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RESEARCH lows us to cautiously match the efficiency of the representative antioxidant to its subclass. Thus, the key to finding an efficient antioxidant is to look for structures similar to Idebenol.15 Research on antioxidants has become a focus because of its use as a treatment for many chronic diseases. Because of that, experiments like this are valuable. More specific functional groups can be pinned down which are known to increase antioxidant properties. Currently, research on this topic is very scattered and contrasting. The role of antioxidants is often debated because of their fickle behavior.16 After finding an efficient group, other factors can be optimized to increase its antioxidant properties. For instance, it is known that the atmospheric pressure (or the pressure of the system) also plays a role in the determination of bond dissociation enthalpy. With that, even the bioavailability of the molecule will play a significant role in its effects. Factors like these will become the question of the future.[17, 18] The novel antioxidant should be able to reduce any or most radicals so it can be used as a multi-purpose molecule in a cell. Moreover, it should not be overlooked that antioxidants often work in a cyclic process and need the help of other antioxidants. This means that the novel antioxidant will, in fact, be a group of antioxidants which complement each other and cover a wider range of radicals. Also, they will need to self-regenerate like vitamin E with vitamin C.[19, 20] Such formation ensures that the antioxidant system will be able to last the overproduction of oxidative stress and help to lower the levels of reactive oxygen species. The goal is to control the level of free radicals in a cell, not to completely eliminate oxidative stress. Moderate levels of oxidative stress help to regulate cell activities.

Oxidative stress in itself is not a problem, but its increased levels are. One of the biggest challenges will be finding the tipping point where oxidative stress starts to damage the molecule.[21, 22] Although there are still a lot of barriers in the way, research is in great momentum. At such pace, with further research, it will not be long until we will be able to use antioxidants to treat chronic diseases and save millions of lives.

Acknowledgements The researcher greatly appreciates the guidance of Dr. Andrew Rappe. This project would not have been possible without his dedication and constant support.

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