Chemical Gardens as Flow-through Reactors Simulating Natural Hydrothermal Systems

The overall goal of the following experiment is to simulate natural chimneys precipitated at submarine hydrothermal vents of early earth using chemical garden experiments to generate the self assembling structures on a small scale. This is achieved by first preparing two solutions. One solution simulates the composition of early earth seawater, and one solution simulates an alkaline hydrothermal fluid produced by water rock reactions.

Next, an apparatus is set up so that the hydrothermal like fluid can be injected into a reservoir of the simulated ocean under an anoxic atmosphere. As a third step, electrodes are set up to measure the voltage between hydrothermal and ocean solutions during the injection. Finally, the hydrothermal like solution is slowly injected into the base of a vial containing the simulated ocean solution, mimicking the natural seepage of vent fluid out of the ocean crust and into the surrounding sea water.

The results show that a mineral precipitate forms at the interface of the ocean and hydrothermal solutions, and this develops into a self assembling chemical garden structure resembling a natural hydrothermal chimney Submarine. Hydrothermal chimneys are just like natural chemical gardens. They were generated, we think on earth about 4.4 billion years ago, and they're made of iron sulfides and iron hydroxides, to the most part with a little bit of silica.

And they behave like a skin they're generated where alkaline solutions come up into the ocean, which is somewhat acidic. And at the interface between the two, they precipitate these minerals, as I say, looking rather like a skin. In fact, one could compare them to life as we know it today.

And that is life tends to have an alkaline interior as we do, for example. And it tends to rather like acidic exteriors. I mean, I can say flippantly perhaps that we like a pH shampoo of pH 5.5.

We like acidic soda pops for example. But also we like to have alkaline drinks to keep us healthy because after all, our interiors are healthy, just like natural chemical gardens. And what we've seen in the lab today is the fact that the interior of these chemical gardens is alkaline.

The exterior is acid. It's somewhat oxidized, and we have, what happens is these chemical gardens impose a chemical gradient across the two, that chemical gradients of pH and Redox, and they summed to about half a volt, which is about the amount of electrochemical energy required to drive life. And we feel that this was just right to drive life to be into being in the first place in one of these chemical gardens on the ocean floor 4 billion years ago.

This experiment allows us to simulate different planetary conditions like oceans, hydrothermal fluids, atmospheres, but also allows us to simulate the effects of changing specific experimental parameters and what effect that will have on the system. Originally, we were studying chemical gardens that grew directly from crystals inside test tubes, but we weren't able to measure the membrane potential because we couldn't get a wire directly inside the membrane itself. So in order to do this, we developed a new method.

It's an injection method where we injected hydrothermal simulant into ocean fluid in a reservoir. This allowed us to use an automated data logger to measure the membrane potential over the growth of the membrane. Those who are New to this method may struggle for growing nicely structured chemical gardens and positioning virus into the chi needs for voltage measurement.

However, by troubleshooting, they can solve all these problems. Begin by using a glass cutter to cut the bottom off of a 100 milliliter clear glass crimp top serum bottle for uses the injection vial. Also collect a 20 millimeter septum, 20 millimeter aluminum crimp seal, and a 0.5 to 10 microliter plastic pipette tip.

Using a 16 gauge syringe needle, carefully puncture a hole through the center of the septum, then remove and discard the needle in the appropriate sharps waste container. Next, insert the pipette tip into the needle hole and push the pipette tip through the septum so that it pokes out the other side. Crimp seal the septum with the pipette tip onto the injection vessel.

To make a watertight seal wind sealed, push the pipette tip further through the septum so that it protrudes to the outside. Then affix a one 16th inch inner diameter, clear, flexible, and chemical resistant tube to the pipette tip. Slide the tubing up onto the pipette tip In order to create a watertight seal, ensure that the tubing is long enough to reach from the injection vial to the syringe pump.

Next, clamp the injection vials on a stand inside a fume hood so that the injection will feed in from the bottom of the vial. Cut two lengths of insulated wires so they are long enough to reach from inside the reaction vessels to the data logger. Leave a little bit of slack in the wires for positioning.

Next, strip about three millimeters of insulation from the end of each wire that will be located inside the reaction vial at the other ends. That will be connected to the multimeter leads. Strip about one centimeter of the wire insert one wire the wire that will be inside the chemical garden close to the opening of the pipette tip to ensure contact with the injection solution, but not so far that it will clog the injection flow.

And then secure both ends of the wires in place using tape. Then place the outside wire so that it will contact the solution reservoir, but not the chemical garden precipitate. Next, attach the free ends of the wires to the data logger to keep the headspace above the solution reservoir.

And IC set up nitrogen gas lines. Split the gas feed from a nitrogen source tank into several tubes so that there is one feed for each injection vial. Prepare 200 milliliters of the reservoir solution, also known as the ocean simulant for each reaction vial by first bubbling deionized water with nitrogen gas for about 15 minutes per 100 milliliters.

This ensures that the experiment will remain anoxic throughout the injection. Then weigh out and add 25 millimolar of iron, two chloride tetra and 75 millimolar of iron. Three chloride hexahydrate to the water.

Stir the mixture gently to dissolve the ions without introducing oxygen into the mixture. After the ions are dissolved immediately resume light bubbling of the solution with nitrogen gas. While the injections are prepared, choose any two of the primary injection solutions from the table shown here and prepare 10 milliliters of each solution.

Then fill a 10 milliliter syringe to the seven milliliter mark with each of the solutions, cap them and set them aside. Next, prepare 20 milliliters of 50 millimolar sodium sulfide in the fume hood as the secondary injection. Fill two 10 milliliter syringes to the seven milliliter mark With the solution, replace the needle caps and set them aside.

Secure the primary injection syringes onto a programmable syringe pump that is located inside the fume hood. Set the injection rate to one milliliter per minute and turn on the pump until both syringes begin to drip into the waste beaker. Then stop the injection.

Next, insert the syringes containing deionized water into the two plastic injection tubes and inject so that the water fills the clear tubing up to the aperture where it enters the main reservoir. Then place the syringes on the stand above the injection vials. Then carefully pour 100 milliliters of the iron containing reservoir solution into each vial.

Carefully create an airtight seal over the vials using perfil and insert a nitrogen feed into each vial. Adjust the flow of the nitrogen gas lines as desired to keep the experiment anoxic for the duration of the injections. Next, bring the water filled syringes one at a time down next to the primary injection syringes.

Carefully slide off the plastic injection tubing and immediately transfer it directly onto one of the primary injection syringe needles. Taking care not to puncture the wall of the tubing. Then set the syringe pump to run at two milliliters per hour for three hours and start the injection at the same time.

Start the recording of the membrane potential. Hit stop on the syringe pump after three hours. Once the chemical garden structures have formed, carefully remove the primary injection syringes from the syringe pump, but leave them connected to the tubing so the structures are not disturbed.

Set them on the stand above the level of the fluid in the vials so that the fluid does not flow back into the syringe. Next, secure the secondary injection syringes, which in this case consist of sodium sulfide to the syringe pump and prime the needle until both begin to drip dispose of the sulfide solution in a special hazardous waste container. Then remove the syringes one at a time from the syringe pump and transfer the tubing from the primary injection syringe to the sodium sulfide containing syringe.

Reprogram the syringe pump to inject the solution at two milliliters per hour and hit start to inject the sodium sulfide solution at this rate for three hours. Once the primary injection solution is fed into the reservoir solution, a chemical garden precipitate begins to form at the fluid interface, and this structure continues to grow over the course of the injection under environmental scanning electron microscopy, the primary precipitates formed in the presence of alanine appeared more rounded in amorphous, whereas pure iron oxy hydroxide precipitates as well as those containing pyrophosphate appeared more crystalline following the primary injection. The potential tended to peak immediately around 0.45 to 0.55 volts, and then decreased for about an hour before stabilizing around 0.1 to 0.2 volts for the rest of the primary injection.

When the primary syringes were switched to the secondary syringes containing sodium sulfide, the chemical garden continued to grow except that visible new growths were now black iron sulfide. Rather than contributing to the existing walls, the black sulfide portions of the chemical garden appeared to branch off and grow separately. As soon as the sulfide injection solution reached the chemical garden, the membrane potential immediately jumped to about 0.9 volts.

There are many possible combinations of solutions that you can use to grow your chemical gardens. It is also possible to change the injection rate from a faster rate to a slower rate and change the primary injection to a secondary one at any time during the experiment, as long as your chemical garden structure is completely formed. Sodium sulfide solutions release hydrogen sulfide gas, which is extremely hazardous.

Open sodium sulfide containers and sulfide containing solutions must be used inside of fume hood and handled according to material safety data sheet guidelines both used and unused solutions must be disposed of as hazardous waste. Hydrothermal chimneys we've just seen being grown in the lab are made of porous minerals, basically iron sulfides and iron hydroxides. And they thi, we think they'll act as catalysts on early life, and they're very similar in structure to the catalysts and enzymes we have even in our own skin to this day.

This experiment can be used to simulate different geological environments, but is also applicable from a material science because there's a lot of interest in learning how to control the growth of these self assembling structures by manipulating the parameters of the experiment, and also perhaps for creating structures and materials that could act as catalysts for different reactions.