One Fixation Method

Nostoc Pink algal bloom

Sampling days in the field have been long yet extremely productive. We have collected nitrogen fixation data on primary producers found in four streams with varying temperatures. We can calculate the rate that primary producers are fixing nitrogen using a technique called Acetylene Reduction Assay or ARA. This is one method of measuring fixation which allows us to compare how rates of fixation change across a temperature gradient. You may be thinking “Whoa! Back up. What is this girl talking about?” Let me explain. Some primary producers, for example, one of my favorites, Nostoc Pink, can take nitrogen gas (N₂), which is abundant in the air, and utilize the nitrogen for growth and other critical cellular processes. But for the organism to use the nitrogen from N₂  gas, it must be converted into a usable form of nitrogen. One particular enzyme known as nitrogenase allows for some organisms to be able to do just that! This enzyme breaks the bonds in N₂  gas, or as we call it, "fixes N₂  gas",  and allows for nitrogen to be transformed into ammonia. Ammonia is a nitrogen-containing compound that can be incorporated into the cell.

Saturating chambers with acetylene gas

ARA, the technique that we have been performing on the streams, mimics the process of breaking the bonds in N₂ gas. With this technique, we use acetylene gas in place of N₂  gas. Because N₂  gas is found in such high concentrations in the environment, it is extremely difficult to detect a change in concentration. Luckily, acetylene is a molecule which has similar properties to that of nitrogen gas. I like to think of them as cousins. Because of their similar properties, the nitrogenase enzyme can break the bond in the acetylene molecule and allows us to detect rates of fixation. When performing Acetylene Reduction Assay, we saturate chambers containing primary producer samples with acetylene gas and take an initial gas sample. We let the algae “do its thing” for 2 hours and then take a final gas sample.

Charlie and Me

Back at the lab, we use an instrument called a gas chromatograph to detect gases and concentrations of gases that are present in the samples we collect. We can determine differences between the initial gas sample and final gas samples to calculate a rate of acetylene fixation. We then can use a known conversion factor which allows us to determine the rate at which  N₂ gas is fixed by each primary producer based on how fast they fixed acetylene gas. This data allows us to determine differences in fixation rates across streams and temperatures. It is very exciting to watch the data compile and observe “fixation” right before my eyes! I have spent many hours working with the gas chromatograph; one might even say we have become very close friends. Today I am spending my time in the lab running gas samples with our gas chromatograph which has been named Charlie by some of the other team members. I intend to find out the story behind Charlie's name, and I will be sure to let the story be known!