Category Archives: DDW Effects On Life

tardigrade1, Water Type Effects on Organism Growth

Experimental Ideas for Effects of D2O on Life

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Reading some papers by Lewis and others at the time got me thinking about cool experiments. There is one that I want to replicate where it was stated that plant cells become hypertonic (cell shrivels up) in the presence of D2O (Brooks et al, 1937 which I need to find). This leads me to want to test red blood cells and human cheek cells (because they are easy to get a hold of) for similar effects.

Reading the paper by Lewis revealed some funny results when dealing with mice (he reports an intoxication effect when one mouse drinks heavy water) that intrigue me. But also gave me an idea. I would like to test the effects of D2O on the Tardigrade which is  a microorganism in the animal kingdom. This thing (nicknamed the water bear) is known to survive the most extreme conditions. From Wikipedia:

Some can survive temperatures of close to absolute zero (−273 °C (−459 °F)), temperatures as high as 151 °C (304 °F), 1,000 times more radiation than other animals, and almost a decade without water. In September 2007, tardigrades were taken into low Earth orbit on the FOTON-M3 mission and for 10 days were exposed to the vacuum of space. After they were returned to Earth, it was discovered that many of them survived and laid eggs that hatched normally.

So you can see why I’d want to put this thing in D2O and see what’s up. I found one place that sells them, but I get the impression they sell only one per order and I would like to grow them (which requires two surprisingly, because of the egg thing). I have read that they grow on moss and lichens so maybe I can get some live samples over at the Rio Grande.

In an email correspondence, Koch pointed out that testing with paramecia might be a worthwhile venture as well. They may exhibit visible response in the presence of toxins or at least will definitely slow down and die quickly.

As an aside, in biology class in 9th grade we were given some paramecia to examine and I watched as mine spontaneously exploded in front of my eyes. I told the teacher about this and she couldn’t care less. Thanks a lot Mrs. Cuesta (here’s a Google link for you).

Bill Hooker on friendfeed also suggested an interesting experiment:

You can D-replace prokaryotes… what would happen if you did that for 100, or 100 million, generations, then switched ’em back to regular water? Can you H-replace ’em using D-depleted water? I’m trying to come up with ways to adapt some enzyme or other to D, wondering if you could get it sensitive enough that adding D to a system using that enzyme would act as a switch…

Which got me thinking that maybe I could get these creatures to live in D2O by slowly integrating more deuterium to regular water over time. I read a paper by Keith Hobson where they analyzed the source of hydrogen in quail tissues by feeding a group water with D2O mixed in. By combining their results (which I’ll need to reread) with what Bill says, maybe I can get a species (of anything really) to live in D2O and analyze the effects of regular water on them. Could be fun.

Wow I can get carried away. I started this post as an aside where I was just going to list some quick thoughts and then started rambling. It’s all good.

Also if anyone has read to the end of this post, can you tweet this post? I want to check the ability of Disqus to search tweets and include them in the comments (the Reactions section). Also try sharing via facebook, or whatever other social media that you may use these days. Thanks in advance.

DDW1, Water Type Effects on Organism Growth

Updated Preliminary Tobacco Seed Growth “Results”

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Those are images of the first batch of samples (Dark Virginia seeds, not pre-soaked), I’ll take pictures of everything and post it next week. But here are the preliminary reports of what different water types do to tobacco seeds:

  • Every seed submerged in deuterium depleted water (DDW) sprouted little hairs on the initial root (the radicle). The interesting thing is this happened almost immediately after emerging from the seed coat.
  • Typically the seeds submerged in deionized water (DI water) germinated the slowest. More will come on this when I replicate the Crumley experiment.
  • Little hairs sprouted inconsistently on the seeds in tap and DI water but are more prevalent on the DI water seedlings. They are not as abundant on these seedlings as they are on the ddw seedlings. If I had to give an analogy (and I do) then I would say the ddw seedlings can grow a nice ‘fro, while the other seeds exhibit male pattern baldness.
  • The little hairs remain localized on the end of the root and aren’t distrubted along the hypocotyl (early stem, and I almost said axon, lol!) so I’m inclined to believe that this is an early root system that is developing because of a lack of nutrients in the water (seen on ddw, most DI, and almost no tap water seeds). But I’m no botanist so I’m just guessing. The fact that the hairs are really prominent in the ddw seeds might suggest the plant recognizes the lack of deuterium, but I’m not willing to make that leap yet.

I setup a new photography system for these seeds. Dr. Koch lent me his Nikon D40 dSLR camera and I purchased some magnification lenses for it. I have the camera setup on an optical post and use a cylindrical lens holder to mount the seed samples (in cuvettes). The picture quality is much better now, with a much higher resolution. I’ll be looking into microscope images soon. Soon I’ll be developing a reliable way to measure the germination, but let’s not jump the shark now. All will be revealed in due time.

Also I was going to measure the pH of the samples at this stage of their development to gain some insight into whether the germination event drastically alters the pH, but the probe in the lab is too big and I don’t have enough sample volume. So I’m thinking that next week I combine the volumes of the water (of each type) to do one “average” measurement. There are four samples of each water type, two for each seed species, and each is filled about 2ml which would give me 8mL of combined volume for each water type. I’m just waiting for the pre-soaked samples to reach full germination (ie shed the seed coat). Now I’m not saying this will work, and it may not be reliable, but hopefully it is a decent approximation for expectations for now until I learn a little bit more.

DDW1, Water Type Effects on Organism Growth

Unsoaked tobacco seed growth

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In most of these samples there is one seedling that shed the seed coat, and in few that seedling is floating at the water surface. It also seems that the branching in the second DD water sample (the Virginia Gold species) has been shed on one of the seedlings (perhaps the one that shed the seed coat). There is a stringy thing floating in the background, which I’m not sure is visible in the image but definitely there in the sample.

Anyone notice anything else?

DDW1, Water Type Effects on Organism Growth

Pre-soaked tobacco seeds early stage growth

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Note: There is some wordpress gallery error that won’t display the second and third picture captions. They are: 2. Dark V in Tap Water. 3. Dark V in DI Water

These images are from the presoaked tobacco seeds. I soaked them in their respective buffers (deuterium depleted water (DD water), tap water, and deionized water (DI water)), and then stored them in the fridge (4C) for 5 days in case there is any drastic chemical exchange between the seed and the water (which would change the buffer). It appears that the deuterium depleted water has started sprouting first in both cases, which is speculative but noteworthy. Also the branching that I noted in the last post appears again here, but only in the DD water samples, which is interesting. I wonder what that could be.

 

DDW1, Water Type Effects on Organism Growth

Setup of Tobacco Seed Growth in Different Types of Water

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I am growing 2 different species of tobacco seed (Virginia Gold #1 and Dark Virginia purchased from The Tobacco Seed Company, as an aside I find it strange that we bought seeds from a company in England that gets their seeds from the United States) in different water buffers: regular tap water, 18MΩ deionized water (DI water), and deuterium depleted water (DD water).

I place 3 seeds of each species in a cuvette (I actually have no clue what company these are from because they are from a former student in the lab, but USA Scientific has a comparable type) and add one type of water. So there are a total of 6 cuvettes (3 for each species).

Seeds in water in cuvettes.

The experiment has way more to consider than I initially suspected which presents some interesting challenges. On top of that I don’t know all that much (right now) about how deuterium interacts with the environment and the seeds, and I don’t know the biochemistry of seed growth in general. Because of this I started a second set of experiments that are identical except that the seeds were presoaked in their respective buffers in the refrigerator in case the initial stages of growth dramatically changed the water solution.

Finally I’m in the process of figuring out how to accurately record growth rates and right now I’m using very primitive macro photography (my camera phone and a big magnifying lens), which will develop into more advanced macro photography (Dr. Koch’s personal DSLR with 10x magnification lens) and hopefully eventually evolve into the microscope and camera system. Here is my current photography setup:

Cuvette photography system.

I have a 2in lens with a focal length of about 3.5in setup on an adjustable post (which is mounted on a rail). I place cuvettes on a cylindrical lens holder (the clampy thing in the back) and adjust the lens height and distance to get the best picture. Most parts are opto-mechanics purchased from Thor Labs.

DDW1, DDW2, DDW3, Water Type Effects on Organism Growth

Effect of Deuterium Depeleted Water on Life

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Hydrogen has several isotopes and one of them, deuterium, exists quite naturally in water to form D_{2}O. In previous experiments and several papers by Gilbert Lewis, it has been found that life is hindered in the presence of D_{2}O. While this may be true, my PI Steve Koch wondered if life had found a use for it because naturally occurring water has about a 17mM (millimolar) concentration of deuterium.

To put that number into perspective, when I do a typical polymerase chain reaction of DNA I add 10mM of each base of DNA (which is less than the amount of naturally occurring deuterium) to create millions of copies of a DNA template from an amount that is 1000x less then what the reaction yields. In fact most chemicals in most of my buffers on the order of the amount of naturally occurring deuterium.

So you can see it isn’t a stretch to think that nature has found a use for D_{2}O since it is quite abundant and life has been constantly evolving for billions of years. I want to test this hypothesis in a variety of different organisms:

  1. Tobacco Seeds – to act as a foil to Lewis’ experiments in which he grew tobacco seeds in pure D_{2}O.
  2. Mustard Seeds – from what I’m told mustard seeds are the powerhouse of the botanical genetics world much like Drosophila and S. cerevisiae are in their respective genetic fields.
  3. Escherichia coli – another molecular biological powerhouse that is very easy to grow and may be easy to see results with. We just got the facilities to be able to grow E. coli and damn it I want to use them!
  4. Saccharomyces cerevisiae (Yeast) – I know a guy who grows yeast for his experiments and I’m sure it wouldn’t be a stretch to get him to do so in deuterium depeleted water.

So the idea would be to try to grow these in regular water and in deuterium depleted water (no D_{2}O), and in the case of E. coli and yeast, perhaps in pure D_{2}O because I don’t think those experiments have been carried out yet. Hopefully I will be able to conclusively state whether or not life has developed a need/use for D_{2}O which would be a very interesting discovery indeed!

DDW1, Water Type Effects on Organism Growth

Preliminary Results of How Water Effects Organism Development

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This is a little backwards because I’m announcing some preliminary results of a project that I haven’t even discussed yet. It’s also a little awkward because I still don’t know what to call this project (at least in a way that would fit in a blog post) and I have even less of a clue what to categorize this project as. With all that said, I say let’s forget all that and just get into what we all came here for:

The results!

I put some tobacco seeds in different water buffers (types of water?). My next post will explain all this, but the short is I have seeds in tap water, filtered 18MΩ (deionized water), and deuterium depleted water (water contains H_{2}O, but it also contains a hydrogen isotope called Deuterium that can form D_{2}O). The lab has been working with D_{2}O a lot and found (along with research done in the 1930’s) that life is greatly hindered under pure D_{2}O. So Koch has been thinking that life evolved to find a use for D_{2}O. The hypothesis is that the seeds won’t grow as fast in water that has no D_{2}O compared to regular water. Let’s see what happened in the first trial of this experiment.

Now remember these are preliminary results so I don’t want to make claims that aren’t accurate, but it appears that the roots have sprouted tiny offshoots in the Deuterium Depleted water (DD water). The other option is that it is mold, I’m not sure. Hopefully this will be more clear in a few days.