Providing Enhanced Migration Time Reproducibility with a High-Voltage-Compatible Flow Sensor for Capillary Electrophoresis.

In capillary electrophoresis (CE), analyte identification is primarily based on migration time, which is a function of the analyte's electrophoretic mobility and the electro-osmotic flow (EOF). The migration time can be impacted by the presence of parasitic flow from changes in temperature or pressure during the run. Presented here is a high-voltage-compatible flow sensor capable of monitoring the volumetric flow inside the capillary during a separation with nL/min resolution. The direct measurement of both flow and time allows for compensation of flow instabilities. By expressing the electropherogram in terms of signal versus electromigration velocity instead of time, it is possible to improve the run-to-run reproducibility up to 25×.

Long-term thermal stability of fluorescent dye used for chiral amino acid analysis on future spaceflight missions.

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.

Stability of reagents used for chiral amino acid analysis during spaceflight missions in high-radiation environments.

The search for biosignatures on spaceflight missions requires in situ instrumentation capable of highly selective and sensitive organic analyses. To this end, CE-LIF is a uniquely promising technique, capable of determining the type, abundance, and chirality of amino acids present in environmental samples at nanomolar concentrations. However, this type of assay requires several reagents that have not yet been used on spaceflight missions. A key concern, particularly for future missions to Europa, is the survivability of these critical components for CE separation and LIF detection under high levels of radiation. Here we present an investigation of the chemical stability of the reagents and associated fused silica capillary after a total ionizing dose of 300 krad, exceeding the predicted total ionizing dose for the potential Europa Lander Mission payload by two-fold. Neither the fused silica capillary nor the fluorescent dye (5-carboxyfluorescein succinimidyl ester) showed significant change in performance following irradiation. Following the irradiation of the pre-mixed background electrolyte, both migration time and resolution were affected. However, when the reagents (sodium tetraborate, sodium taurocholate, and γ-cyclodextrin) and the acetonitrile solution were irradiated separately and mixed afterwards, there was no change in the separation performance.

Extraction of amino acids from aerogel for analysis by capillary electrophoresis. Implications for a mission concept to Enceladus' Plume.

Ocean worlds like Europa and Enceladus in the outer solar system are prime targets in the search for life beyond Earth. Enceladus is particularly interesting due to the presence of a water plume ejecting from the south polar region. The recent discovery of H2 in the plume, in addition to the presence of previously observed organic compounds, highlights the possibility of life in this moon. The plume provides materials from the underlying ocean that could be collected simply by flying through it. The presence of the plume means that material from the ocean is available for collection during a flyby, without the need for landing or complex sample handling operations such as scooping or drilling. An attractive approach to preserve the organics in particles collected during flyby encounters would be to utilize silica aerogel, the material used to collect particles at hypervelocity during the Stardust mission. Here we demonstrate amino acids can be extracted from aerogel simply by adding water. This simple liquid extraction method could be implemented during a mission prior to analysis with a liquid-based technique like capillary electrophoresis.

Enhanced Resolution of Chiral Amino Acids with Capillary Electrophoresis for Biosignature Detection in Extraterrestrial Samples.

Amino acids are fundamental building blocks of terrestrial life as well as ubiquitous byproducts of abiotic reactions. In order to distinguish between amino acids formed by abiotic versus biotic processes it is possible to use chemical distributions to identify patterns unique to life. This article describes two capillary electrophoresis methods capable of resolving 17 amino acids found in high abundance in both biotic and abiotic samples (seven enantiomer pairs d/l-Ala, -Asp, -Glu, -His, -Leu, -Ser, -Val and the three achiral amino acids Gly, ß-Ala, and GABA). To resolve the 13 neutral amino acids one method utilizes a background electrolyte containing γ-cyclodextrin and sodium taurocholate micelles. The acidic amino acid enantiomers were resolved with γ-cyclodextrin alone. These methods allow detection limits down to 5 nM for the neutral amino acids and 500 nM for acidic amino acids and were used to analyze samples collected from Mono Lake with minimal sample preparation.

Implementation of microchip electrophoresis instrumentation for future spaceflight missions.

We present a comprehensive discussion of the role that microchip electrophoresis (ME) instrumentation could play in future NASA missions of exploration, as well as the current barriers that must be overcome to make this type of chemical investigation possible. We describe how ME would be able to fill fundamental gaps in our knowledge of the potential for past, present, or future life beyond Earth. Despite the great promise of ME for ultrasensitive portable chemical analysis, to date, it has never been used on a robotic mission of exploration to another world. We provide a current snapshot of the technology readiness level (TRL) of ME instrumentation, where the TRL is the NASA systems engineering metric used to evaluate the maturity of technology, and its fitness for implementation on missions. We explain how the NASA flight implementation process would apply specifically to ME instrumentation, and outline the scientific and technology development issues that must be addressed for ME analyses to be performed successfully on another world. We also outline research demonstrations that could be accomplished by independent researchers to help advance the TRL of ME instrumentation for future exploration missions. The overall approach described here for system development could be readily applied to a wide range of other instrumentation development efforts having broad societal and commercial impact.

Capillary electrophoresis separation of the desamino degradation products of oxytocin.

Oxytocin (OT) is an endogenous and therapeutic hormone necessary for maternal health. It is also the subject of fast growing research in the field of behavioral science. This article describes a rapid CE method using UV detection at 214 nm for the determination of the deamidation products of OT. Deamidation is the most common degradation pathway of peptides and proteins and can lead to reduced therapeutic efficiency of biopharmaceuticals. To achieve a separation of the seven structurally similar desamino peptides from OT, 11 mM sulfobutyl ether ß-CD and 10% v/v MeOH were added to a BGE of 50 mM phosphate buffer at pH 6.0. The assay is linear within ≤5-100 µM for all species with a total analysis time of 12 min. The method was then applied to monitor the heat-stress degradation of OT at 70°C, where all seven desamino species were observed over a 96 h period.

Recent advances in the analysis of therapeutic proteins by capillary and microchip electrophoresis.

The development of therapeutic proteins and peptides is an expensive and time-intensive process. Biologics, which have become a multi-billion dollar industry, are chemically complex products that require constant observation during each stage of development and production. Post-translational modifications along with chemical and physical degradation from oxidation, deamidation, and aggregation, lead to high levels of heterogeneity that affect drug quality and efficacy. The various separation modes of capillary electrophoresis (CE) are commonly utilized to perform quality control and assess protein heterogeneity. This review attempts to highlight the most recent developments and applications of CE separation techniques for the characterization of protein and peptide therapeutics by focusing on papers accepted for publication in the in the two-year period between January 2012 and December 2013. The separation principles and technological advances of CE, capillary gel electrophoresis, capillary isoelectric focusing, capillary electrochromatography and CE-mass spectrometry are discussed, along with exciting new applications of these techniques to relevant pharmaceutical issues. Also included is a small selection of papers on microchip electrophoresis to show the direction this field is moving with regards to the development of inexpensive and portable analysis systems for on-site, high-throughput analysis.