What (the heck) is Tryptophan-like Fluorescence?

The summer I transitioned from undergrad into grad school, I started working on a coastal field project with my future co-advisor and project-PI, Dr. Natalie Nelson. As we prepared for our field campaigns, we began assembling our new sondes—new vocabulary swirled right over my head. Starting grad school on a niche research focus - can feel really inaccessible at first. I remember thinking: Will I ever understand all these words? After four years of writing, presenting, and working with sondes and tryptophan-like fluorescence, I can confidently say—yeah, I get it now. (BTW - It feels ridiculous saying “tryptophan-like-fluorescence” over and over again during a seven-minute presentation.)

Let’s start with sondes. “Sonde” is basically a fancy (French) word for a water quality tool that holds a bunch of sensors so you can take measurements for a bunch of things at once. Some of these sensors measure things that might sound familiar: Dissolved Oxygen (DO) – how much oxygen is in the water (important for fish and aquatic life - because if DO gets too low, they won’t be able to breathe and could die); pH – a measure of acidity or basicity (from chemistry classes and people test pool pH); Temperature – self-explanatory; Salinity/Conductivity – measures how salty the water is; Turbidity – how clear or cloudy the water is. 

There were also sensors I wasn’t familiar with, like the optical/fluorescence sensors. These included: Total Algae – detects two types of algae (though they showed similar readings for our study); Fluorescent Dissolved Organic Matter (fDOM); Tryptophan-like Fluorescence (TLF). Did I lose you? Undergrad Julia would definitely be lost hearing all of that. She would think, “Oh no, I should know what this means!…but I don’t.” So let PhD Candidate Julia try to break it down.

Natural waters—rivers, lakes, estuaries—contain a lot of organic matter, both dissolved and suspended in the water. This matter comes from decaying plants, algae, and other living and formerly things. It can give the water color and includes compounds that support life higher up the food chain. (Yummy!)  Some of this organic matter is fluorescent, meaning it absorbs light and then re-emits it as a different wavelength. When we shine a specific light on this material and it emits light back, we can measure it—that’s fluorescence! A sensor that does this gives us fDOM readings.

So, fDOM is a subset of DOM (dissolved organic matter), and within fDOM, we can break things down even further. For example: Humics – think potting soil or decomposed plant material; Algae – some types fluoresce too; Amino acids – like tryptophan, which is where we get TLF.

Let’s talk about tryptophan. If you’ve heard of it, it was probably in the context of turkey and post-Thanksgiving naps—tryptophan is an amino acid that’s been linked to sleepiness in people.

Because tryptophan is a building block of proteins, its presence in water suggests there’s protein-rich organic material around. Think of plant material like reeds and marsh grass—mostly carbohydrates, low in protein. Bacteria, on the other hand, are protein-rich. So when TLF is high, it could indicate elevated bacterial concentrations. That’s why TLF is interesting: it can indicate when there is a lot of bacteria in water.

In our research, we were interested in using TLF in coastal waters—places where people swim or harvest shellfish—to detect possible fecal contamination that could make people sick. The idea is: TLF is relatively easy to measure and has been shown to correlate with bacteria, so it could be useful to tell us when waters would be unsafe.

But in practice?! In our experience - our TLF sensor was tricky. It was finicky during calibration, gave inconsistent readings, and varied more than expected even in supposedly stable conditions. AND since TLF just measures the presence of tryptophan, which is present in all kinds of protein-rich material—not just bacteria, and also not just fecal bacteria which is what we’re most interested in from a public health protection point-of-view. Estuarine environments like those in North Carolina are incredibly rich and productive. Tons of microscopic organisms live there—eggs, larvae, zooplankton—all of which may contribute to TLF (there’s not a ton of research on it – and if there is and you understand it – PLEASE EXPLAIN IT TO ME!).