Luciferin - Luciferase Experiment - Part I

I wanted to see the Luciferin - Luciferase reaction that produces bioluminescence and Mrs. Keel helped me to get this experiment going. She pointed me to a biological supplies store (Carolina Biological Supply) from where I got Sea Firefly lanterns (Cypridina hilgendorfii), gave me instructions on how to perform the experiment (how to separate out the luciferin and luciferase from the firefly lanterns), and allowed me to use her room to perform the experiment one morning before school. Thank you, Mrs. Keel!

Sea Firefly Lanterns

This short video clip shows the experiment that I performed: http://youtu.be/827Obd_Vm-s

I prepared two firefly lantern extracts, one of luciferin and the other of luciferase. I crushed some of the lanterns and placed them on an evaporation plate. I added distilled water to the crushed mixture and immediately a reaction started taking place. Luciferase acted upon luciferin and bioluminescence was produced. The luminescence slowly started dissipating as the luciferin in the extract got used up. When the luminescence completely disappeared, what I had left on the evaporation plate was a pure luciferase extract.

To create the pure luciferin extract, I added crushed firefly lanterns to boiling water. What this accomplished was to denature the luciferase. Luciferase does not automatically denature even when returned to room temperature.

I know have two separate containers, one with pure luciferase extract and the other with pure luciferin extract. Part II of my experiment will involve mixing these two together to see if they produce bioluminescence.

Video clip of why Fajardo bay is bioluminescent by our tour guide from Island Kayaking Adventures

http://youtu.be/FynpUTJ2kEM

This video clip was taken in the bioluminescent bay in Fajardo, Puerto Rico. It was given by our tour guide from Island Kayaking Adventures and contains a short description of why the bay is bioluminescent.

There is really nothing much to see in the video as it was shot completely in the dark, but you can see occasional flashes of green/blue as someone in our group put their paddle into the water. The audio is not the best quality either as there were other tour groups around us and you can hear their conversation in the background.

Why do bioluminescent bays glow?

Bioluminescence in the bioluminescent bays is caused by microscopic single-celled creatures called dinoflagellates. The conditions in these bays are such that allow an abundance of these organisms to thrive. There are about 750,000 tiny dinoflagellates per gallon of water that light up when they are touched.

The bilomuniscent dinoflagellates Pyrodinium bahamense are a photosynthesis-using plankton. They are one celled and measure about 1/500th of an inch. A tiny burst of light it gives off is a hundred times bigger than itself. Each dinoflagellate bursts into light when it feels pressure against its cell wall. The light is given off in an instantaneous process; when you add the light bursts of 750,000 dinoflagellates per cubic foot of water together, the effect is spectacular.

Dinoflagellates are organisms that are part of the Protista kingdom. Most dinoflagellates are algae, so they can produce their own food through photosynthesis. They possess tiny flagellates (Latin for whip), which are tail-like appendages that propel them through the water. Photosynthesis in dinoflagellates involves light being captures by their chlorophyll, which has bluish-green pigment (as opposed to plants which have green chlorophyll). What sets this cholorophyll apart is that it becomes luminescent when agitated.

The bioluminescent bay in Fajardo has three key attributes for attracting dinoflagellates. First it shallow waters and a narrow exit to the sea, allowing for the organisms to concentrate in its shallow refuge. Two, because the bay is small in size, water does not rush out of it quickly. Three, it has a high concentration of mangroves. Mangroves are important to dinoflagellates because they are a good source of vitamin B12, which is essential for dinoflagellates to survive. Mangroves release a large amount of vitamin B12 and so dinoflagellates conglomerate in the bay.

Kayaking in the Bioluminescent Bay in Fajardo, Puerto Rico

Last December, my family and I went to Puerto Rico on vacation. My uncle and his family joined us, too, from Boston. I had been looking forward to this vacation for quite some time as this was going to be my first scuba-diving trip. But I was even more excited when I found out that among the top 10 bioluminescent bays in the world, 3 of them were in Puerto Rico, and one of them was right in Fajardo, the place we were staying at in Puerto Rico! The bay is located in the Las Cabezas de San Juan Nature Reserve and is called Laguna Grande or the Big Lagoon.

We booked our trip to see the bay through Island Kayaking Adventures (http://www.ikapr.com/). Our tour started at 8:30 pm but we got there about an hour early so we could sign in, go through the informational presentation, and get instructions on how to kayak. To get to the bay, we had to go paddle through a narrow channel, lined with mangroves on either side. That was quite an experience! We were in 2-person kayaks and I was in one with my cousin, Sheela. Both of us were novices at this so we ended up zig-zagging across the channel and bumping into the roots of the trees multiple times. Added to that, it was pretty dark - the only light we had was a small green flashing light on the kayak in front of us.

Here's a map of the route we took through the mangrove channel to get to the bioluminescent bay.

Our trip guide asked us all to be really quite when we got to the opening of the bay. As we slowly paddled into Laguna Grande, I noticed a shimmering glow each time my paddle pushed through the water. I really didn't think much of it until we were in the middle of the bay and the tour guide made us all stop paddling. He then told us to scoop a handful of water into our hands. My hand was glowing green and it looked like I had shimmering stars in the palm of my hand! I kept running my fingers through the water and marveling at the incredible burst of green each time I did that. I threw a scoop of water at Sheela and there were glowing points of light on her chest.

We spent about 15 minutes in the bay before we had to paddle back. I could have spent all night there if I could.