The goal of this protocol is to form simulated hydrothermal chimneys via chemical garden injection experiments and introduce a thermal gradient across the inorganic precipitate membrane, using a 3D printable condenser that can be reproduced for educational purposes.
Deep sea hydrothermal vents are self-organizing precipitates generated from geochemical disequilibria and have been proposed as a possible setting for the emergence of life. The growth of hydrothermal chimneys in a thermal gradient environment within an early Earth vent system was successfully simulated by using different hydrothermal simulants, such as sodium sulfide, which were injected into an early Earth ocean simulant containing dissolved ferrous iron. Moreover, an apparatus was developed to sufficiently cool the ocean simulant to near 0 °C in a condenser vessel immersed in a cold water bath while injecting a sulfide solution at hot to room temperatures, effectively creating an artificial chimney structure in a temperature gradient environment over a period of a few hours. Such experiments with different chemistries and variable temperature gradients resulted in a variety of morphologies in the chimney structure. The use of ocean and hydrothermal fluid simulants at room temperature resulted in vertical chimneys, whereas the combination of a hot hydrothermal fluid and cold ocean simulant inhibited the formation of robust chimney structures. The customizable 3D printed condenser created for this study acts as a jacketed reaction vessel that can be easily modified and used by different researchers. It will allow the careful control of injection rate and chemical composition of vent and ocean simulants, which should help accurately simulate prebiotic reactions in chimney systems with thermal gradients similar to those of natural systems.
Hydrothermal chimneys are self-organizing chemical garden precipitates generated from geochemical disequilibria within deep-sea vent environments as heated, hydrothermally altered fluid seeps into a colder ocean. In an early Earth scenario, it has been proposed that the chimneys formed at ancient alkaline vents, and that transecting ambient pH/redox/chemical gradients could have driven reactions toward the emergence of metabolism1,2,3,4,5,6. Hydrothermal vents have also been postulated to exist on other planets including the ocean worlds, Europa and Enceladus7,8,9,10. Various experiments have been conducted to simulate aspects of proposed prebiotic hydrothermal chimney chemistry including precipitation of catalytic iron sulfide minerals that could reduce CO211,12, gradient-driven organic synthesis13,14,15, and incorporation of organics into chimney structures16. In creating experimental setups to mimic hydrothermal vents, whether on Earth or on other worlds, it is essential to consider the geochemical gradients and the open, far-from-equilibrium nature of the system to produce realistic simulations.
In addition to pH, redox, and chemical gradients, hydrothermal vents also impose a thermal gradient across the chimney membrane/wall due to the feed of heated vent fluid into a cold seafloor environment. Cold seafloor ocean temperatures can vary as a function of depth, solar penetration, and salinity; average seafloor ocean depths at vent sites (mostly at mid-ocean ridges) are in the range of 0-4 °C17. Depending on the type of vent, the thermal gradient between ocean and vent fluid can vary dramatically-from the milder gradients of alkaline vents, such as Lost City18,19 or the Strytan Hydrothermal Field where the vent fluid is 40-90 °C20,21, to the deep seafloor black smokers where the vent fluid can reach several hundred degrees Celsius22,23,24,25. From an origin-of-life perspective, simulation of thermal gradients in hydrothermal systems is significant as they could affect the mineralogy and chemical reactivity of chimney precipitates3,13 and/or could affect habitability as hydrothermal chimneys host microbes that take up electrons directly from mineral surfaces26. In a gradient across the chimney wall, a range of temperature conditions would be present over a short distance, and the chimney wall would represent a combination of minerals and reactions characteristic of all these thermal regimes.
Laboratory-grown hydrothermal chimneys in thermal gradients were simulated to explore the effects of the cold ocean and hot hydrothermal fluid on this potential prebiotic environment. Generally, because growing simulated hydrothermal chimneys via an injection method with a heated interior and cold exterior presents practical challenges, the most accessible chimney experiments are those done at ambient pressure (therefore not requiring costly and complicated reactors). Previous attempts at lab-grown chimneys in a thermal gradient have not able to produce both a hot/warm hydrothermal fluid and a cold ocean. In an effort to keep the entire chimney at high temperature for long durations to form reactive minerals that can drive organic reactions, some studies heated the whole experiment (ocean and hydrothermal fluid) to ~70 °C using either a heating jacket or a hot bath13,14. Another type of chimney precipitate formation experiment, in a "fuel cell" apparatus, formed the chimney wall simulant on a flat membrane template; these experiments have also been heated in bulk by submerging the fuel cell gradient apparatus in a hot water bath27,28. Previous studies have formed simulated hydrothermal chimneys from hot hydrothermal fluids (heated to ~70 °C using various methods) injected into a room-temperature ocean3,12; however, a cold ocean has not been attempted.
This work advances methods for prebiotic chimney growth laboratory simulations4 to create a realistic thermal gradient from a cold (0-5 °C) ocean to a heated hydrothermal fluid in which to synthesize chimney materials and test properties of interest. To date, there have been no prebiotic chimney experiments successfully conducted with a realistic temperature gradient for alkaline vents: with the interior vent solution held at ~70 °C and the exterior ocean solution chilled to ~5 °C. Furthermore, in the few heated chimney experiments that have been conducted, the experimental setup is complex and can be costly. Chemical garden experiments have great potential to yield insights about the processes that may have taken place in hydrothermal vents on the early Earth. Hence, the ability to quickly set up multiple variations of a chimney experiment is advantageous, as is the ability to have a simple apparatus that is inexpensive, non-fragile, easily modified, and ideal for students to work with. Presented here is a novel apparatus (Figure 1) designed to facilitate growth of a simulated hydrothermal chimney while maintaining and monitoring a realistic thermal gradient between the cold ocean and heated hydrothermal fluid simulant. This experimental apparatus is similar in design to a jacket reactor, but is a three-dimensional (3D) printed condenser that can be easily produced by any research group interested in conducting similar experiments (see Supplementary printable file). Using this 3D printed condenser, thermal gradient chimney experiments were conducted to test the utility of this apparatus for maintaining robust temperature gradients and to test the effects of temperature gradients on chimney structure and morphology.
1. Safety considerations
2. Setup for injection experiments
3. Preparation of solutions for chemical garden growth
4. Setting up the thermistor
5. Setting up the ice bath
6. Prepping for injection
7. Monitoring the temperature and the experiment
NOTE: Once the water is circulating through the condenser, the thermistor temperature probe will begin to display the fall in temperature within the ocean. The goal is for the temperature to reach near 0 °C. See Table 2 for the precise temperature (thermal) gradient settings.
8. Ending the experiment
As in previous studies1,2,13,29; once the hydrothermal fluid simulant reached the ocean vial, a mineral precipitate structure began to form that grew thicker and taller for the duration of the injection. The iron sulfide chimneys were delicate structures that were not very robust and were easily disaggregated if the ocean vial or injection was physically disturbed. This is consistent with results from previous studies3. The chemical concentration of the sulfide solution also played a vital role in the morphology of the sulfide chimneys. More concentrated solutions of sulfide allowed for taller and sturdy mineral precipitates, as shown in Figure 5, whereas lower concentrations of sulfide solutions produced weak chimney structures. In some cases, no structure was formed, only a liquid sulfide-mineral "soup" was created, that would eventually settle out as a sediment (Figure 3D). This occurred in both thermal and non-thermal gradient conditions.
In thermal gradient chimney experiments with iron sulfide, solid chimney structures generally did not coalesce as well as they did at room temperature. Figure 3E–H shows the morphology of an iron sulfide chimney grown between a cold ocean and room-temperature hydrothermal fluid. The chimneys in the temperature gradient were string-like and tenuous in nature, whereas non-thermal gradient results (Figure 3A-D) show more semi-permanent structures. The same was true when the hydrothermal fluid was heated (Figure 4). The exception was at higher sulfide and iron concentrations (Figure 5) where a solid iron sulfide chimney was formed between a room-temperature hydrothermal solution and cold ocean simulant.
The effect of a thermal gradient on the growth of iron hydroxide chimneys was also tested. The results showed patterns that were similar to those of the iron sulfide chimney: while the room-temperature iron hydroxide experiment resulted in a more robust chimney precipitate, the thermal gradient experiment between the warm hydrothermal fluid and the cold ocean resulted in a smaller mound of chimney material that did not coalesce vertically (Figure 6). In contrast to the tall upright structures of iron hydroxide chimneys observed in previous work (in room-temperature experiments)29, our thermal gradient experiment showed a different morphology.
Figure 1: Thermal gradient chimney apparatus. Please click here to view a larger version of this figure.
Figure 2: 3D printed condenser. (A) Schematic of a 3D printed condenser showing condenser dimensions. (B) Placement of a glass ocean vessel inside the condenser to cool the ocean simulant. Please click here to view a larger version of this figure.
Figure 3: A variety of thermal and non-thermal gradient chimneys. (A–D) Non-thermal gradient control experiment from room-temperature hydrothermal fluid (HTF) to room-temperature ocean simulant. (A) 10 mM Na2S•9H2O HTF and 20mM FeCl2·4H2O ocean simulant. (B) 20 mM Na2S•9H2O HTF and 10 mM FeCl2·4H2O ocean simulant. (C) 20 mM Na2S•9H2O HTF and 20mM FeCl2·4H2O ocean simulant. (D) 20 mM Na2S•9H2O HTF and 20mM FeCl2·4H2O ocean simulant. (E–H) Thermal gradient chimney experiment from room-temperature HTF simulant to a cold ocean reservoir (~5-10 °C). (E) 20 mM Na2S•9H2O HTF and 10 mM FeCl2·4H2O ocean simulant. (F) 10 mM Na2S•9H2O HTF and 20 mM FeCl2·4H2O ocean simulant. (G) 20 mM Na2S•9H2O HTF and 10 mM FeCl2·4H2O ocean simulant. (H) 10 mM Na2S•9H2O HTF and 20 mM FeCl2·4H2O ocean simulant. Please click here to view a larger version of this figure.
Figure 4: Thermal gradient experiment. Experiment performed with warm (~35-40 °C) 20 mM Na2S•9H2O solution injected into a cold (~5-10 °C) 20 mM FeCl2·4H2O ocean simulant, producing small chimney strands. Please click here to view a larger version of this figure.
Figure 5: Effect of concentration of ocean simulant on chimneys. Higher concentrations (~50 mM Na2S•9H2O, 10 mM FeCl2·4H2O, and 200 mM NaCl) of anoxic ocean simulants produced more structurally robust, taller chimneys. Room-temperature sulfide solution was injected into 2-10 °C ocean simulant. Please click here to view a larger version of this figure.
Figure 6: Simultaneous growth of thermal and non-thermal gradient chimneys. (A) 100 mM FeCl2·4H2O + 100 mM FeCl3·6H2O ocean solution with a 200 mM NaOH hydrothermal fluid (HTF) fluid simulant at room temperature. (B) Thermal gradient experiment with the same concentrations with warm HTF at ~35-50 °C into cold ocean simulant at ~5-10 °C. Please click here to view a larger version of this figure.
Hydrothermal Fluid Chemistry (Injection) | Ocean Chemistry (Reservoir) |
50 mM Na2S | 10 mM FeCl2·4H2O + 200 mM NaCl or NaHCO3 |
20 mM Na2S | 10 mM FeCl2·4H2O + 200 mM NaCl or NaHCO3 |
10 mM Na2S | 20 mM FeCl2·4H2O + 200 mM NaCl or NaHCO3 |
200 mM NaOH | 100 mM FeCl2·4H2O + 100 mM FeCl3·6H2O |
Table 1: Concentration matrix for both simulated ocean and hydrothermal fluid injection solutions.
HTF °C | Ocean Simulant Temperatures °C | |
~23 | ~23 | 5-10 |
~35-50 | ~23 | 5-10 |
Table 2: Thermal gradient experimental matrix. The hydrothermal fluid (HTF) temperature refers to the temperature of the fluid in the syringe; the actual temperature at the inlet to the ocean vial was between 20 and 35 degrees lower than the temperature within the syringe (~70 °C) (see Supplementary Appendix 1, Figure 3, and Figure 4).
Supplementary printable file. Please click here to download this file.
Supplementary Appendix 1. Please click here to download this file.
Supplementary Appendix 2. Please click here to download this file.
Supplementary Appendix 3. Please click here to download this file.
Effect of thermal gradients on simulated chimney growth: This experimental apparatus yielded several variations in chimney morphologies that were due to several experimental parameters. Chimneys of iron sulfide and iron hydroxide formed tall upright structures at room temperature, but formed more tenuous, stringy precipitates or flat mounds in the thermal gradient experiments. This was consistent with the findings of Herschy et al. where wispy, non-erect chimney precipitates were formed from a hydrothermal fluid heated to 70-80 °C and injected into room-temperature ocean simulant33. There are various possible explanations for this: convective heat transfer can cause more natural buoyant forces (along with the forced pumping of the injection) to make the precipitate flow rapidly towards the top of the ocean vessel as it is forming. Alternatively, heating the syringe fluid makes the hydrothermal simulant less dense and thus more prone to rise vertically than to stabilize on top of the injection point. It is possible that this effect could be mitigated by changing the syringe injection rate to slower rates to allow the growth of a more stable structure. White et al. examined iron sulfide chimney growth with the hydrothermal simulant injected at extremely slow rates (0.08 mL/h), and although the chimney took days to coalesce, it was structurally stable13. As Herschy et al. used peristaltic pumps at injection rates of 10-120 mL/h, which is several orders of magnitude faster than the rates used in our thermal gradient experiments, it is not surprising that they also produced string-like chimney structures33.
Higher concentrations of precipitating reactants in the ocean and vent solutions can also yield more robust chimneys in thermal gradients. Higher chemical concentrations of precipitating ions (sulfide or hydroxide) in the hydrothermal fluid or ocean simulant can lead to higher overall precipitate mass, thus creating a stronger structure. As Herschy et al. and White et al. used lower concentrations of sulfide in the hydrothermal fluid (10 mM), their structures were smaller than the ones produced in this work using higher (20-50 mM) sulfide concentrations. Additionally, some studies of iron sulfide chimney growth have also included silica in the hydrothermal fluid along with the sodium sulfide, which can help produce more robust chimneys3,13,33. Silica chemical garden structures have also been used to simulate aspects of hydrothermal chimney growth34, and these tend to produce very robust structures that can be removed from the tube/vial for physical analysis. However, the effects of temperature gradients on silica injection structures are not known and will be an area of further study.
Considerations for future chimney simulation experiments: The 3D printed condenser created in this study to cool the ocean vessel acted like a jacketed reaction vessel, but with some practical improvements: 1) the open top allowed sampling of the chimney and maintaining the anoxic ocean headspace; 2) the 3D printed part conferred easy reproducibility; 3) as the designs can be digitally edited, the apparatus can be quickly modified and re-printed if desired; and 4) the use of inexpensive materials made each condenser more cost-effective than the actual glass-jacketed reaction vessels. These 3D printed condensers are a flexible and easily shared experimental apparatus that could be a useful way to standardize platforms for simulated hydrothermal chimney experiments across different research groups, allowing better comparison of samples and data. Files of the condenser can be sent to colleagues to print on their own for their educational or scientific purposes (see Supplemental 3D printing file of the condenser used in this work). This inexpensive setup could also be used as an undergraduate laboratory experiment for chemical gardens or chemobrionics29,35.
In conclusion, this work describes a novel experimental apparatus using 3D printing to facilitate the growth of simulated hydrothermal chimneys in temperature gradient environments. The 3D printed condenser is able to cool the ocean simulant to near-freezing temperatures, similar to the seawater near seafloor hydrothermal systems. Meanwhile, a heated syringe was used to simulate the high-temperature hydrothermal fluid injecting into this cold ocean. The morphologies and structures of iron sulfide and iron hydroxide chimneys were affected by the thermal gradient: when both the ocean and the hydrothermal fluid simulants were at room temperature, the chimneys formed vertically oriented structures, but when the hydrothermal fluid was heated and the ocean was cooled, the formation of robust chimney structures was inhibited. For accurately simulating prebiotic reactions in such chimney systems with thermal gradients analogous to those of natural systems, it will be necessary to carefully control parameters such as injection rate and chemical composition of both vent and ocean simulants. The custom and inexpensive 3D printed condenser created for this study is similar in function to a jacketed reaction vessel and can be easily modified and distributed electronically to various research and educational groups for use in many types of chemobrionic experiments.
The authors have nothing to disclose.
This research was carried out at the Jet Propulsion Laboratory, California Institute of Technology, under a contract with NASA, supported by the NASA Astrobiology Institute Icy Worlds. Dr. Gabriel LeBlanc was supported in part by a Research Initiation Grant (2017-34) through the Oklahoma NASA EPSCoR Cooperative Agreement (NNX15AK42A). We would like to thank Heather Whitehead for assistance with the initial 3D printed condenser design, Kalind Carpenter for assistance with 3D printing, John-Paul Jones for helpful discussion on condenser vessels, Laura Rodriguez for help with temperature data analysis, and Erika Flores with laboratory assistance. Copyright 2020 California Institute of Technology.
3/8-Inch Clear Vinyl Tubing | Watts | SVIG10 | Cut to desired length for experiment |
40-pin Male to Female Wire Jumper Multicolored Ribbon Cables | EDGELEC | ED-DP_L30_Mix_120pcs | These wires will require stripping of plastic ends and carefully removing one of the 2 plastic casings |
Aluminum seals | Fisher | 0337523C | Thermo Scientific National Headspace 20 mm Crimp Seals |
Ferric chloride hexahydrate | Fisher | I88-100 | Ferric Chloride Hexahydrate (Lumps/Certified ACS) |
Ferrous chloride tetrahydrate | Fisher | I90500 | Ferrous Chloride Tetrahydrate (Crystalline/Certified) |
Gear Hose Clamps | Glarks | 40Pcs | |
Gray butyl stoppers | Fisher | 0337522AA | Thermo Scientific National 20 mm Septa for Headspace Vials |
Pipette tips | VWR | 53511-682 | pipette tips 0.5-10 microliters |
Serum bottles | Sigma-Aldrich | 33110-U | Vials, crimp top, serum bottle, size 100 mL, clear glass, outer diameter x height 51.7 mm x 94.5 mm. For these experiments, the bottom of the serum bottle should be cut off. |
Sodium hydroxide | Sigma-Aldrich | S5881 | reagent grade, ≥98%, pellets (anhydrous) |
Sodium sulfide nonahydrate | Fisher | S425212 | Sodium Sulfide Nonahydrate (Crystalline/Certified ACS). Store at -20 °C. Only open in a glove box or fume hood. Releases toxic H2S gas; all sulfide-containing solutions must be kept in a glove box or fume hood. |
syringe heater | Syringepump.com | HEATER-KIT-5SP | Clamp gear hose clamps around heating blanket |
Syringe needles (16 gauge) | Fisher | 14-826-18B | BD General Use and PrecisionGlide Hypodermic Needles, 16 G x 1.5 in. (38 mm) |
Syringe Pump | Syringepump.com | NE-4000 | Dual or multiple channel, depending on desired number of simultaneous experiments |
Syringes (10 mL) | Fisher | 14-823-16E | BD Syringe with Luer-Lok Tips (Without Needle) |
Tubing | Cole Parmer | EW-06407-71 | Tygon Lab Tubing, Non-DEHP, 1/16" internal diameter X 1/8" outer diameter |
Water Circulation Pump | Bayite | BYT-7A015 | May need two people to help prime pump |