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Indium seals have been used extensively in ultra-high vacuum and cryogenic applications. Typically, these seals use indium alongside or in place of other metal gaskets in stainless-steel vacuum flanges, with some custom applications for flanges sealing directly with glass (optics or tubes). Here, we present the design and performance of three pressed indium seals (99.99% In) between aluminum and 0.5 in. diameter sapphire optics and aluminum and gold coated Kovar semiconductor packages (TO-66 and TO-39). Test fixtures were designed to mimic those of future tunable diode laser spectrometers for Earth, planetary, and manned spaceflight environmental monitoring applications. Successful high-hermeticity seals [<10-10 atm cc/s (He)] were achieved for all seals formed with sufficient pressure applied to allow indium to flow between mating surfaces. The hermeticity of the seals was maintained after temperature cycling (-10 to +80 °C, 20 cycles), with the optical seals surviving extended duration tests (-55 to +85 °C, per MIL-STD-883). Semiconductor packages (TO-39) subjected to these extended tests saw a moderate increase in leak rate [∼5 × 10-9 atm cc/s (He)]; however, further testing showed that either the glass-metal package seals or the indium were affected (the sample size was too small to draw firm conclusions for future applications). Overall, these results suggest long-term survivability of indium seals for Kovar-aluminum and sapphire-aluminum interfaces [>10 years at 10-10 atm cc/s (He)], where the coefficient of thermal expansion differs by approximately four times., (© 2024 Author(s). Published under an exclusive license by AIP Publishing.)

Capillary temperature control during capillary electrophoresis (CE) separations is key for achieving accurate and reproducible results with a broad array of potential methods. However, the difficulty of enabling typical fluid temperature control loops on portable instruments has meant that active capillary temperature control of in situ CE systems has frequently been overlooked. This work describes construction and test of a solid-state device for capillary temperature control that is suitable for inclusion with in situ instruments, including those designed for space missions. Two test articles were built, a thermal mass model (TMM) and a functional model (FM). The TMM demonstrated that temperature gradients could be limited using the proposed control scheme, and that our thermal modeling of the system can be relied on for future adaptations of physical geometries of the system. The FM demonstrated CE analytical performance while under active temperature control and that the device was compatible with the harsh thermal-vacuum environments that might be encountered during space flight., (© 2024 Wiley‐VCH GmbH.)

We report here the first fully automated capillary electrophoresis (CE) system that can be operated underwater. The system performs sample acquisition and analysis by coupling CE to contactless conductivity detection. Using 5 M acetic acid as the background electrolyte (BGE), inorganic cations and amino acids at concentrations as low as 5.2 μM can be separated and identified. This technology could be augmented to include a variety of other detection modes. This system serves as an early prototype for potential future underwater explorers on ocean worlds of the outer solar system such as Europa or Enceladus. This work documents the first step in the development of this general-purpose technology platform.

Capillary electrophoresis (CE) holds great promise as an in situ analytical technique for a variety of applications. However, typical instrumentation operates with open reservoirs (e.g., vials) to accommodate reagents and samples, which is problematic for automated instruments designed for space or underwater applications that may be operated in various orientations. Microgravity conditions add an additional challenge due to the unpredictable position of the headspace (air layer above the liquid) in any two-phase reservoir. One potential solution for these applications is to use a headspace-free, flow-through reservoir design that is sealed and connected to the necessary reagents and samples. Here, we demonstrate a flow-through high-voltage (HV) reservoir for CE that is compatible with automated in situ exploration needs, and which can be electrically isolated from its source fluidics (in order to prevent unwanted leakage current). We also demonstrate how the overall system can be rationally designed based on the operational parameters for CE to prevent electrolysis products generated at the electrode from entering the capillary and interfering with the CE separation. A reservoir was demonstrated with a 19 mm long, 1.8 mm inner diameter channel connecting the separation capillary and the HV electrode. Tests of these reservoirs integrated into a CE system show reproducible CE system operation with a variety of background electrolytes at voltages up to 25 kV. Rotation of the reservoirs, and the system, showed that their performance was independent of the direction of the gravity vector., (© 2023 Wiley-VCH GmbH. California Institute of Technology. Government sponsorship acknowledged. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.)

Sample injection is a critical step in a capillary electrophoresis (CE) analysis. Electrokinetic injection is the simplest approach and is often selected for implementation in portable CE instruments. However, in order to minimize the effect of sample matrix upon the results of a CE analysis, hydrodynamic injection is preferred. Although portable CE instruments with hydrodynamic injection have been reported, injection has always been performed at the grounded end of the capillary. This simplifies fluidic handling but limits coupling with electrochemical detectors and electrospray ionization-mass spectrometry (ESI-MS). We demonstrated previously that injection at the high-voltage (HV) end of the capillary could be performed using an HV-compatible rotary injection valve (fixed-volume injection). However, the mismatch between the bore sizes of the channels on the rotor-stator valve and the separation capillary caused peak tailing and undesired mixing, impairing analytical performance. In this work, we present an HV-compatible hydrodynamic injection approach that overcomes the issues associated with the fixed-volume injection approach reported previously. The performance of the CE instrument was demonstrated by analyzing a mixture of 13 amino acids by CE coupled to laser-induced fluorescence, which showed relative standard deviations for peak area and migration time below 5% and 1%, respectively, for triplicate analysis. Additionally, replicate measurements of a mixture of amino acids, peptides, nucleobases, and nucleosides by CE coupled to electrospray ionization-mass spectrometry (CE-ESI-MS) were performed to evaluate peak tailing, and results were similar to those obtained with a commercial CE-ESI-MS setup., (© 2023 Jet Propulsion Laboratory, California Institute of Technology. Government sponsorship acknowledged.)

Capillary electrophoresis (CE) systems have undergone extensive development for spaceflight applications. A flight-compatible high voltage power supply and the necessary voltage isolation for other energized components can be large contributors to both the volume and mass of a CE system, especially if typical high voltage levels of 25-30 kV are used. Here, we took advantage of our custom CE hardware to perform a trade study for simultaneous optimization of capillary length, high voltage level, and separation time, without sacrificing method performance. A capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C 4 D) method recently developed by our group to target inorganic cations and amino acids relevant to astrobiology was used as a test case. The results indicate that a 50 cm long capillary with 15 kV applied voltage (half of that used in the original method) can be used to achieve measurement goals while minimizing instrument size., (© 2022 Wiley-VCH GmbH. California Institute of Technology. Government sponsorship acknowledged.)

Fatty acids are a well-established class of compounds targeted as biosignatures for future missions to look for evidence of life on ocean worlds such as Europa and Enceladus. In order to establish their abiotic or biotic origin, we need to separate and quantify fatty acids to determine their relative abundances within a sample. In this study, we demonstrate the high potential of capillary electrophoresis coupled to mass spectrometry (CE-MS) for the efficient separation and sensitive detection of a wide variety of fatty acids. Three derivatization strategies were evaluated to allow the detection of fatty acids by positive ionization mode MS. Furthermore, CE-MS conditions were optimized to provide maximum separation efficiencies and detection sensitivities for the analysis of saturated and unsaturated fatty acids with even- and odd-numbered carbon chain lengths. Optimum separation and detection were obtained using a background electrolyte of 2 M acetic acid in 45% acetonitrile, after derivatization of the fatty acids with 2-picolylamine or N , N -diethylethylenediamine. The limits of detection for the derivatized fatty acids using the optimized method ranged from 25 to 250 nM. The optimized method was also used for the analysis of fatty acids in cell cultures and natural samples. Two distinctive biosignatures were obtained for the microorganisms Halobacillus halophilus and Pseudoalteromonas haloplanktis . In addition, multiple fatty acids were detected in a natural sample from Mono Lake, California.

In situ missions of exploration require analytical methods that are capable of detecting a wide range of molecular targets in complex matrices without a priori assumptions of sample composition. Furthermore, these methods should minimize the number of reagents needed and any sample preparation steps. We have developed a method for the detection of metabolically relevant inorganic and organic anions that is suitable for implementation on in situ spaceflight missions. Using 55 mM acetic acid, 50 mM triethylamine, and 5% glycerol, more than 21 relevant anions are separated in less than 20 min. The method is robust to sample ionic strength, tolerating high concentrations of background salts (up to 900 mM NaCl and 300 mM MgSO 4 ). This is an important feature for future missions to ocean worlds. The method was validated using a culture of Escherichia coli and with high salinity natural samples collected from Mono Lake, California., (© 2021 Wiley-VCH GmbH.)

The in situ search for chemical signatures of life on extraterrestrial worlds requires automated hardware capable of performing detailed compositional analysis during robotic missions of exploration. The use of electrophoretic separations in this search is particularly powerful, enabling analysis of a wide range of soluble organic compounds potentially indicative of life, as well as inorganic compounds that can serve as indicators of habitability. However, to detect this broad range of compounds with a single electrophoresis instrument, a combination of different detection modes is required. For detection of any ionizable species, including organic compounds that do not participate in terrestrial biology (i.e., "unknown unknowns"), mass spectrometry (MS) is essential. Inorganic ions, or any dissolved charged species present, can be analyzed using capacitively coupled contactless conductivity detection (C 4 D). Additionally, for the trace analysis of compounds of key interest to astrobiology (particularly, amino acids), laser-induced fluorescence (LIF) detection holds unique promise, due to the fact that it has the highest demonstrated sensitivity of any form of detection. Here, we demonstrate a fully automated, portable capillary electrophoresis analyzer that is capable of all these modes of detection. The prototype system developed here addresses the three most significant challenges for doing electrophoretic separations: precise sample injection, HV isolation, and automation of all operational steps. These key challenges were successfully addressed with the use of custom-designed rotor-stator valves with optimized operational sequences incorporating gas purging steps, rinses, and HV application.

Silver ions (Ag+) have been proposed as a biocide to treat the water in NASA's next generation of human space exploration vehicles/habitats. One advantage of Ag+ is that it is effective as a biocide in a range (200 to 500 ppb) safe for human consumption. So, monitoring Ag+ is essential to ensure the safety and health of the crew. Here we present two analytical methods based on capillary electrophoresis and capacitively coupled contactless conductivity detection (CE-C4D) to address the need to monitor Ag+ levels in water. Using 5 M acetic acid as a background electrolyte (BGE), 100 ppb of Ag+ could be detected in a simulant of the International Space Station (ISS) water. In addition to Ag+, other inorganic cations (K+, Ca2+, Na+, Mg2+, Ni2+, and Zn2+) frequently found in the ISS potable water can be detected simultaneously using this BGE in less than 4.5 min. By using a BGE composed of 0.5 M acetic acid, levels of Ag+ as low as 25 ppb could be detected in the ISS water simulant in less than 2.5 min. Although in this condition none of the other cations interfered with the detection of Ag+, some of them co-migrated, which could prevent obtaining additional information about the sample composition.

This work investigated microorganism survival under temperature and ultraviolet (UV) radiation conditions found at the surface of ice-covered ocean worlds. These studies were motivated by a desire to understand the ability of resilient forms of life to survive under such conditions as a proxy for potential endogenic life and to inform planetary protection protocols for future missions. To accomplish this, we irradiated Bacillus subtilis spores with solar-like UV photons at temperatures ranging from room temperature down to 11 K and reported survival fractions with respect to fluence. We observed an increase in survival at low temperatures and found that the inactivation rate follows an Arrhenius-type behavior above 60 K. For solar-photon fluxes and surface temperatures at Europa and Enceladus, we found that Bacillus subtilis spores would be inactivated in less than an hour when in direct sunlight.

Unambiguous detection of chemical and physical signatures of microbial life on Mars or other solar system bodies requires differentiation between signals produced by biotic and abiotic processes; instruments aimed at generalized in situ extant life detection would therefore increase the science return of a life-detection mission. Here, we investigate Bioelectrochemical Systems (BES) as a technique to measure microbial metabolism (which produces electrical current and redox changes) and distinguish between potential abiotic and biotic responses in environmental samples. Samples from inhabited niches should contain everything necessary to produce current, that is, catalysts (microorganisms) and fuel (nutrients). BES can also probe for inactive organisms in less energetically rich areas by adding a fuel to drive metabolism. A commercial potting soil and a Mars simulant soil were inoculated in the anodic chamber of microbial fuel cells, and current was monitored over time. Addition of a fuel (electron donor) source was tested for metabolic stimulation of endogenous microbes. Redox reactions between Mars simulant soil and the introduced electron donor (lactate) produced false-positive results, emphasizing the importance of careful interpretation of signals obtained. The addition of lactate to both soils resulted in enhanced biologically produced current, allowing stimulation and detection of dormant microbes. Our results demonstrate that BES provide an approach to detect metabolism in samples without prior knowledge of the organisms present, and that thorough electrochemical analyses and experimental design are necessary to determine if signals are biotic.

Future spaceflight missions focused on life detection will carry with them new, state-of-the-art instrumentation capable of highly selective and sensitive organic analysis. CE-LIF is an ideal candidate for such a mission due to its high separation efficiency and low LODs. One perceived risk of utilizing this technique on a future mission is the stability of the chemical reagents in the spaceflight environment. Here, we present an investigation of the thermal stability of the fluorescent dye (5-carboxyfluorescein succinimidyl ester) used for amino acid analysis. The dye was stored at 4, 25, and 60°C for 1 month, 6 months, 1 year, and 2 years. When stored at 4°C for 2 years, 25°C for 6 months, or 60°C for 1 month there was no effect on CE-LIF assay performance due to dye degradation. Beyond these time points, while the dye degradation begins to interfere with the analysis, it is still possible to perform the analysis and achieve the majority of amino acid biosignature science goals described in the science definition team report for the potential Europa Lander mission. This work indicates that thermal control of the dye at ≤4°C will be needed during transit on future spaceflight missions to maintain dye stability., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

With growing interest in exploring ocean worlds, such as Europa and Enceladus, there is a fundamental need to develop liquid-based analytical techniques capable of handling high salinity samples while performing both bulk and trace species measurements. In this context, CE with capacitively coupled contactless conductivity detection (CE-C 4 D) has tremendous potential. One of its advantages is that this combination allows the detection of a wide number of charged species (both organic and inorganic) without the need of derivatization. Amino acids are an example of organic targets that are powerful biosignatures in the search for life beyond Earth. Simultaneous information on the inorganic cations in a sample helps with assessing the habitability of an extraterrestrial environment, as well as providing sample context for any measurements of trace amino acids. In this work, we present a series of flight-compatible methods capable of simultaneously measuring inorganic cations and amino acids in samples of varying salinity by CE-C 4 D. Regardless of the sample total salinity, 5.0 M acetic acid was selected as the optimum BGE. The methods were evaluated by analyzing natural samples of low and high salinity from Hot Creek Gorge, Mono Lake, and Santa Monica beach. Prospects for mission implementation are also discussed., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

For decades, the Martian regolith has stymied robotic mission efforts to catalog the organic molecules present. Perchlorate salts, found widely throughout Mars, are the main culprit as they breakdown and react with organics liberated from the regolith during pyrolysis, the primary extraction technique attempted to date on Mars. This work further develops subcritical water extraction (SCWE) as a technique for extraction of amino acids on future missions. The effect of SCWE temperature (185, 200, and 215°C) and duration of extraction (10-120 min) on the total amount and distribution of amino acids recovered was explored for three Mars analog soils (JSC Mars-1A simulant, an Atacama desert soil, and an Antarctic Dry Valleys soil) and bovine serum albumin (as a control solution of known amino acid content). Total amounts of amino acids extracted increased with both time and temperature; however, the distribution shifted notably due to the destruction of the amino acids with charged or polar side chains at the higher temperatures. The pure bovine serum albumin solution and JSC Mars 1A also showed lower yields than the Atacama and Antarctic extractions suggesting that SCWE may be less effective at hydrolyzing large or aggregated proteins. Changing solvent from water to a dilute (10 mM) HCl solution allowed total extraction efficiencies comparable to the higher temperature/time combinations while using the lowest temperature/time (185°C/20 min). The dilute HCl extractions also did not lead to the shift in amino acid distribution observed at the higher temperatures. Additionally, adding sodium perchlorate salt to the extraction did not interfere with recoveries. Native magnetite in the JSC Mars-1A may have been responsible for destruction of glycine, as evidenced by its uncharacteristic decrease as the temperature/time of extraction increased. This work shows that SCWE can extract high yields of native amino acids out of Mars analog soils with minimal disruption of the distribution of those amino acids, even in the presence of a perchlorate salt., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This article discusses a microfluidic subcritical water extraction (SCWE) chip for autonomous extraction of amino acids from astrobiologically interesting samples. The microfluidic instrument is composed of three major components. These include a mixing chamber where the soil sample is mixed and agitated with the solvent (water), a subcritical water extraction chamber where the sample is sealed with a freeze valve at the chip inlet after a vapor bubble is injected into the inlet channels to ensure the pressure in the chip is in equilibrium with the vapor pressure and the slurry is then heated to ≤200 °C in the SCWE chamber, and a filter or settling chamber where the slurry is pumped to after extraction. The extraction yield of the microfluidic SCWE chip process ranged from 50% compared to acid hydrolysis and 80%-100% compared to a benchtop microwave SCWE for low biomass samples.

One of the most habitable environments in the Solar System outside of Earth may exist underneath the ice on Europa. In the near future, our best chance to look for chemical signatures of a habitable environment (or life itself) will likely be at the inhospitable icy surface. Therefore, it is important to understand the ability of organic signatures of life and life itself to persist under simulated europan surface conditions. Toward that end, this work examined the UV photolysis of Bacillus subtilis spores and their chemical marker dipicolinic acid (DPA) at temperatures and pressures relevant to Europa. In addition, inactivation curves for the spores at 100 K, 100 K covered in one micron of ice, and 298 K were measured to determine the probability for spore survival at the surface. Fourier transform infrared spectra of irradiated DPA showed a loss of carboxyl groups to CO2 as expected but unexpectedly showed significant opening of the heterocyclic ring, even for wavelengths>200 nm. Both DPA and B. subtilis spores showed identical unknown spectral bands of photoproducts after irradiation, further highlighting the importance of DPA in the photochemistry of spores. Spore survival was enhanced at 100 K by ∼5× relative to 298 K, but 99.9% of spores were still inactivated after the equivalent of ∼25 h of exposure on the europan surface.

The successes of the Mars exploration program have led to our unprecedented knowledge of the geological, mineralogical, and elemental composition of the martian surface. To date, however, only one mission, the Phoenix lander, has specifically set out to determine the soluble chemistry of the martian surface. The surprising results, including the detection of perchlorate, demonstrated both the importance of performing soluble ion measurements and the need for improved instrumentation to unambiguously identify all the species present. Ion chromatography (IC) is the state-of-the-art technique for soluble ion analysis on Earth and would therefore be the ideal instrument to send to Mars. A flight IC system must necessarily be small, lightweight, low-power, and have low eluent consumption. We demonstrate here a breadboard system that addresses these issues by using capillary IC at low flow rates with an optimized eluent generator and suppressor. A mix of 12 ions known or plausible for the martian soil, including 4 (oxy)chlorine species, has been separated at flow rates ranging from 1 to 10 μL/min, requiring as little as 200 psi at 1.0 μL/min. This allowed the use of pneumatic displacement pumping from a pressurized aluminum eluent reservoir and the elimination of the high-pressure pump entirely (the single heaviest and most energy-intensive component). All ions could be separated and detected effectively from 0.5 to 100 μM, even when millimolar concentrations of perchlorate were present in the same mixtures.

Bacillus subtilis spores were deposited in high-density single layers on metal, glass, and polymer substrates using vacuum filtration followed by a wetted filter transfer step. Quantitative analysis of spore transfer was performed using culture-based and germinability assays, and spore distributions were observed with electron microscopy., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The reaction between the hydroperoxy radical, HO(2), and acetone may play an important role in acetone removal and the budget of HO(x) radicals in the upper troposphere. We measured the equilibrium constants of this reaction over the temperature range of 215-272 K at an overall pressure of 100 Torr using a flow tube apparatus and laser flash photolysis to produce HO(2). The HO(2) concentration was monitored as a function of time by near-IR diode laser wavelength modulation spectroscopy. The resulting [HO(2)] decay curves in the presence of acetone are characterized by an immediate decrease in initial [HO(2)] followed by subsequent decay. These curves are interpreted as a rapid (<100 μs) equilibrium reaction between acetone and the HO(2) radical that occurs on time scales faster than the time resolution of the apparatus, followed by subsequent reactions. This separation of time scales between the initial equilibrium and ensuing reactions enabled the determination of the equilibrium constant with values ranging from 4.0 × 10(-16) to 7.7 × 10(-18) cm(3) molecule(-1) for T = 215-272 K. Thermodynamic parameters for the reaction determined from a second-law fit of our van't Hoff plot were Δ(r)H°(245) = -35.4 ± 2.0 kJ mol(-1) and Δ(r)S°(245) = -88.2 ± 8.5 J mol(-1) K(-1). Recent ab initio calculations predict that the reaction proceeds through a prereactive hydrogen-bonded molecular complex (HO(2)-acetone) with subsequent isomerization to a hydroxy-peroxy radical, 2-hydroxyisopropylperoxy (2-HIPP). The calculations differ greatly in the energetics of the complex and the peroxy radical, as well as the transition state for isomerization, leading to significant differences in their predictions of the extent of this reaction at tropospheric temperatures. The current results are consistent with equilibrium formation of the hydrogen-bonded molecular complex on a short time scale (100 μs). Formation of the hydrogen-bonded complex will have a negligible impact on the atmosphere. However, the complex could subsequently isomerize to form the 2-HIPP radical on longer time scales. Further experimental studies are needed to assess the ultimate impact of the reaction of HO(2) and acetone on the atmosphere.

The self-reactions and cross reactions of the peroxy radicals C2H5O2 and HO2 were monitored using simultaneous independent spectroscopic probes to observe each radical species. Wavelength modulation (WM) near-infrared (NIR) spectroscopy was used to detect HO2, and UV absorption monitored C2H5O2. The temperature dependences of these reactions were investigated over a range of interest to tropospheric chemistry, 221-296 K. The Arrhenius expression determined for the cross reaction, k2(T) = (6.01(-1.47)(+1.95)) x 10(-13) exp((638 +/- 73)/T) cm3 molecules(-1) s(-1) is in agreement with other work from the literature. The measurements of the HO2 self-reaction agreed with previous work from this lab and were not further refined. The C2H5O2 self-reaction is complicated by secondary production of HO2. This experiment performed the first direct measurement of the self-reaction rate constant, as well as the branching fraction to the radical channel, in part by measurement of the secondary HO2. The Arrhenius expression for the self-reaction rate constant is k3(T) = (1.29(-0.27)(+0.34)) x 10(-13)exp((-23 +/- 61)/T) cm3 molecules(-1) s(-1), and the branching fraction value is alpha = 0.28 +/- 0.06, independent of temperature. These values are in disagreement with previous measurements based on end product studies of the branching fraction. The results suggest that better characterization of the products from RO2 self-reactions are required.