Eisen Lab Blog

So I meant to post this yesterday, but I couldn’t figure out why it wouldn’t let me blog. Apparently my computer signs me off wordpress every so often so I have to sign back in.. So this information is from my time in the lab yesterday.

Dissolved oxygen, nitrate, and ammonia tests came with no reagents… really? It’s almost as if they expect us to have an abundance of reagents for nitrate test before we have the tools to run the test. Oops, our bad! So that’s a bit frustrating, but we ordered more reagents for everything which should come by tomorrow hopefully.

The Phosphorous and Nitrite kits came with a couple reagents (still annoying), but this allowed us to run a couple trial runs with them on salt water from one of the tanks. Alex did a few tests in the morning (check out her blog entry) and I did a few in the afternoon.
I’m glad I familiarized myself with the tests because I had to make a few modifications to the procedure, because it was just… not good. For example in these tests, you have to blank the meter (makes sense, right?) but then you have to add the reagents from a small packet into a small cuvette (harder than it sounds) and mix for at least two minutes according to the instructions. However, the meter turns off after two minutes. I solved this by having two cuvettes: one to blank the meter and one with the reagent in it.

Phosphorous gave a reading of 200 ppb, which leads me into our next problem. The meter maxes out at 200, so we can assume phosphorous is at a level higher than 200. We need to get another kit with a meter that measures a higher concentration of phosphorous. I measured Nitrite at 70 ppb, which was similar to what Alex measured.

I also measured pH at 7.46 and salinity at 48.3 mS

David and I hoped that the hardness and alkalinity tests would give similar results since they both test for CaCO3. If we had gotten the same result, we could eliminate one of the tests and save time. I measured alkalinity at 114ppm CaCO3. When I ran the hardness test, it didn’t work. I attempted twice and had the same result: failure. We’re thinking the test just doesn’t work with salt water? We’re going to look into it.

It was really good to familiarize myself with all the equipment because now when we begin our intensive sampling and testing of the succession of the coral ponds, I’ll be ready to go with those water chemistry tests!

And now we wait! We wait for an email telling us they’re going to load the water into the containers for the coral ponds. At this point we have a lot of sampling to do! I’ve decided to call it Occupy Bio Labs since we’ll be spending quite a bit of time in there 🙂

Most of our new gadgets and gizmos for water chemistry have finally come in, so we have started testing them out. Some of the kits we ordered include tests for nitrate, nitrite, phosphorus, chlorine, hardness, sulfate, and iron. At first, we were testing the kits out on tap water to get comfortable with all the tests, and then Matt collected sea water samples from one of the tanks so we started using those.

Today I tried the nitrate kit, which has never been used before. It was a surprisingly simple and quick test, and I found 79.5 ppm (mg/L) nitrate in the sea water. Unfortunately, I do not know the significance of this value, so David suggested that I try the test on tap water for comparison. The amount of nitrate in the tap water sample was 23.2 ppm (mg/L). According to Wikipedia, marine aquariums are only supposed to have trace amounts of nitrate in order to be healthy, so I do not know what our values mean.

A few of the kits we ordered do not have the range to collect values from sea water, so we are thinking of diluting our sea water samples with DI water. We will then use that mixture to conduct the tests that did not work (i.e. the phosphorus and hardness tests). In order for this plan to work, however, there must be small amounts of chemicals in the DI water or our data will be skewed. I checked the level of phosphorus in the DI water and the value came out to be 34 ppb. I am not sure if this means that our dilutions will work, but I am sure we will be able to figure out how to analyze our data after doing some more research.

The water chemistry kits seem to be pretty straightforward, with a few nuances here and there. I conducted the titration-based tests as well as the tests to detect phosphorous and hardness. For the tests I used DI water to compare values, and so far the values I have been getting are comparable to values other people are getting, as well as make sense, which is good. 

So a few tips. The phosphorous meter turns off after a few minutes, and it will forget the blank sample. So it is a good idea to make the reagent+sample tube before you begin. 

Also, it is good to choose the low range of testings, because the high range is too much for our samples. 

We finally got all our equipment for gathering metadata on the water, and decided to do some sample testing using tap water and saltwater from the tank. We ended up with the following:

Tap water –

NO2-N levels: 65 ppm

NO2 levels: 154.63 ppm

Phosphorus: 105 ppb

Saltwater –

NO2-N levels: [TRIAL ONE] 65 ppm, [TRIAL TWO] 57 ppm

NO2 levels: [TRIAL ONE] 213.85 ppm, [TRIAL TWO] 187.53 ppm

Phosphorus: [TRIAL ONE] 48 ppb, [TRIAL TWO] 200 ppb

Some problems we encountered: When I did the trial one on the saltwater phosphorus test, I used the wrong reagent (phosphate reagent is used for the nitrite scanner, while phosphorus reagent is used for the phosphorus scanner). In addition, the phosphorus scanner maxes out at 200 (Andrew later confirmed this by doing additional scans on the saltwater), which means we’ll need a broader range scanner.

First off I hope everyone had a great Thanksgiving break!
If you read Andrew’s previous posts regarding the project, you would know that we have decided to scrap all of the samples we have extracted DNA from and start from the beginning. This is so we will have water chemistry data collected at the same time as DNA is collected thus providing the most consistent and accurate data.

On Tuesday, we received a portion of our water chemistry kit, which tests Hardness, Sulfites, Alkalinity, Iron, pH, and Chloride. We decided to do a practice run on a couple of the tanks so we can familiarize ourselves with the reagents as well as fine tune our sampling procedure. The results are listed below:

Freshwater Tank A
Hardness Test: 93 ppm CaCO3
Sulfite Test: 2 ppm Na2SO3
Alkalinity Test: 90 ppm CaCO3
Iron Test: No detectable amount
pH: 7.3
Chloride Test: 20 ppm Cl-

Saltwater 1088
Iron Test: No detectable amount

We learned a couple of important points through this test run that will speed up our water chemistry sampling process in the future. For every single test we did, we started using the high concentration detection procedure, but found all of the concentrations in the tanks were extremely low, and had to redo it using the low concentration detection procedure. For our real samples that we will hopefully will be taking in the next week, we can save reagents and time and just jump right to the low detection procedures. We also noticed that the Hardness and Alkalinity tests detected the same molecule (CaCO3) and also had similar concentrations and have thus decided to use only one of the tests. (I will get back to you with the chemistry behind this reasoning, which I didn’t really understand). For both the Freshwater and Saltwater Test, we were not able to get a detectable amount of Iron and will likely scrap that water chemistry test. Lastly our pH meter results were a little different from Russell’s highly sensitive pH meter (pH=8.3) that takes continuous measurements and Tweets them. (Eisen is probably going to like the idea of that!) We will either scrap our pH meter and just use his or will have to verify if our pH meter is giving is accurate readings, by putting it in solutions of known low acidity. This is just an idea of mine, not sure if it’s a good way to check for its accuracy.

That’s where we are in the project as of Tuesday. I will get back to you about the differences between Hardness and Alkalinity and also update when we start taking samples again!