In the ninth grade I came across bioplastics, plastics derived from renewable biomass, for the first time. Since then I have spent hundreds of hours researching the space and coming up with an idea for a bioplastic made from duckweed. The thought process was that bioplastics today generally come from biomass like corn, potatoes, sugar cane, etc. but these take up acres of land to grow and need to be consumed by humans (especially with the rise in climate change causing a shortage in food supply). Thus, duckweed, although possessing a bit of an odd name, has been a large focus of mine for two years now.
Shortly after I discovered bioplastics, the CEO of Pond Biomaterials became a mentor of mine. Pond is a startup based out of Denmark and I have been working with them (virtually) for these two years. This summer, though, I finally got to come to Denmark in person and Pond set me up to work in three labs across Copenhagen for five weeks, including Novozymes, the Technical University of Denmark, and the University of Copenhagen.
The reason for three labs instead of just one is that to build this project requires work in many different areas of science, including microbiology, biochemistry, and chemistry.
Below, I will go over what I worked on in each lab as well as my biggest learnings / takeaways.
Novozymes was the first lab I worked in. This is a private company, and the world leader in industrial enzyme production. I had never worked on my project in a lab that was not in a university, so this was a very different experience. I worked closely with Ulla Rosenberg and Kim Borch, two individuals I am incredibly lucky to have met. I always had help in the lab and the support was unreal. My days were usually 9–11 hours long but there was always someone by my side sticking it out with me and it was honestly so much fun.
For the first week I worked on an optimized enzymatic procedure for the duckweed. We experimented with different parameters and got the highest glucose concentrations I have seen out of my duckweed. I learned a lot about enzymes themselves, dilution series, and how to run an assay.
The next few days, I worked on fermenting the sugar solutions where I was in the microbiology lab with Robin Dorau trying to produce lactic acid. Some of the things I learned about were buffering capacities, measuring the optical density of bacteria via a spectrophotometer, and how to ensure the bacteria had enough nutrients. We ended up getting very high lactic acid concentrations, although I also learned about some other compounds present in my mixtures that have led to more discoveries about the duckweed and experiments themselves.
We were working large scale, with hundreds of grams of duckweed, so we had to be innovative with our setups. It doesn’t matter what lab you are working in: you always have to do some improvisation. Here are some examples of us simply having to figure it out:
1.) We had to have our samples stirring at a certain temperature for the enzymes to optimally operate, however our system kept dropping in temperature. In order for the setup to be self sufficient, we improvised and used aluminum and towels for insulation, and poured boiling water into the tub our samples were in. It worked like a charm, but definitely looked a little interesting.
2.) Another scenario where we had to just figure it out was when our samples would not filter prior to fermentation. I have been working on a smaller scale in Ottawa so although filtering was an issue I wasn’t on as much of a time crunch and had less material to work with. What we ended up doing was going to the brewery lab and their pressure filters (used for beer) worked perfectly. This taught me another valuable lesson, which was the importance of interconnectedness and asking others — even if in a completely different space of work — for help. Working at Novozymes was so unique because there are so many individuals working on different things and yet everyone can help each other, even a beer brewery lab and a biotech lab.
One more valuable lesson for me was to not always trust the results you get. When we did colorimetric analysis (basically determining the concentration of a chemical compound using a colour reagent) we got very low lactic acid concentrations which was honestly very disappointing because I never would have expected my results to be in this ball park. Later, though, we ran an HPLC (high performance liquid chromatography, a method that is much more precise) and our results were over 20x higher than we had initially thought. This really changed the next steps that had to be taken, so it’s important to ensure that when doing a test like an assay or something generally less precise (usually because they are easier and cheaper) they can have large margins of error in the results they yield.
Technical University of Denmark
The second lab I worked in was at the Technical University of Denmark, DTU. Here I was working on something I had never done before: the separation of lactic acid from the fermentation mixture. This process was quite difficult and was certainly not a success, but I learned a lot about purification technologies and namely, their complexity.
My biggest takeaway was to not underestimate how much time something new can take. I would have figured I would have learned this by now but it’s easy to have too high of expectations in science. Since I had never worked on the purification side of things before, I did not fully comprehend the difficulty of it until I tried it out. Researching and ideating is one thing, and actually doing it is something else entirely! For example, we attempted something called an azeotropic distillation on my first day (see the photo below) which theoretically should have worked quite easily but in fact did not at all. Even removing the water without evaporating other compounds was hugely complex and when I researched what to do, I was greeted with hundreds of research papers.
Thus, I learned something else that was very important which was to keep my solutions as clean as possible. What I mean by this is that in prior steps to the purification I am adding enzymes, buffers, extracts, etc. to my mixtures, but I have to in reality remove everything to get pure lactic acid at the end. When working on prior steps it is very important to keep in mind the end goal and future steps that must also be optimized for.
Another takeaway was to utilize online videos for the use of analytical equipment because there is so much online that can help. I had to be much more independent in this lab, and through a video I learned how to do infrared spectroscopy which was very helpful.
I had a hard time coping with failure at this lab overall but it offered many insights into what comes next in this project, as this step will be the key to producing the bioplastic and will be a challenge on its own.
The University of Copenhagen
Finally, I worked at the University of Copenhagen with Peter Ulvskov, a professor in plant biology. This tied more into what I had been working on at Novozymes, but instead of optimizing the enzymes being added to the biomass we were working on what could happen even prior to this: cell wall degradation, mechanical methods of breakdown, etc. Honestly, I found this so much fun and full of learning. I truly got to understand the duckweed to a greater extent, like what exists in it that may inhibit bacterial growth, what polysaccharides it contains, and how the starch granules behave. For the first time I got to look at the duckweed under a microscope — microscopy was actually key in my work at KU. For example, before degradation of the cell walls, we could see the starch granules stuck in the duckweed cells (left image, where the granules are dark blue) and then after we could see the granules released from the cells (right image).
Something else we worked on was the removal of pigmentation from the duckweed to explore how this may hinder future steps in my procedures (see the below picture, where the rightmost beaker contains a much lower content of chlorophyll). I found I was looking at duckweed through a different lens because of how much unique experimentation I got to do at the university.
Likely my biggest takeaway was the significance of writing absolutely everything down and taking many pictures. This is of course something I have always had to do in the lab but Peter put a lot of emphasis on organization and I came up with a much better system for taking notes, keeping track of which photos are which, etc. It can be easy to recognize what a microscopy image is showing a few minutes after you’ve taken it, but a month later, the importance of that image to you may be gone.
My second takeaway was to keep in mind everything that is required for each part of my project and look at how this may translate to a real world protocol. For example, when we were grinding the duckweed we made sure to use equipment that could be scaled up easily as opposed to only something that works small scale.
Another one of my learnings was the importance of going back to the very beginning — the foundations of a project — because this is where the most improvement can transpire. Breaking down the cell walls to a greater extent was not helping me in the purification of the lactic acid, but eventually it could help me get more lactic acid to work with by creating improvements in glucose yields. Every step holds weight.
All in all, getting to spend time in Copenhagen was such a neat experience and I am so grateful to everyone who helped me. I also felt that I became a more independent person as I lived alone for around five days which I have never done before! Getting to meet my supervisor Martin from Pond was a huge highlight, as well as exploring the wonderful city (and the crêpe stands👩🏽🍳).
Thank you so much for reading this! I’m a 17-year-old learning about where sustainability meets technology, and am the author of the “Chronicles of Illusions” duology. If you want to see more of my work, connect with me on LinkedIn, Twitter, or subscribe to my newsletter!