Internal Diameter Column Research Articles

Development of microflow ultra high performance liquid chromatography-mass spectrometry metabolomic assays for analysis of mammalian biofluids

Introduction and objectivesThe application of untargeted metabolomics assays using ultra high performance liquid chromatography-mass spectrometry (UHPLC-MS) to study metabolism in biological systems including humans is rapidly increasing. In some of these studies there is a requirement to collect and analyse low sample volumes of biofluids (e.g. tear fluid) or low cell and tissue mass samples (e.g. tissue needle biopsies). The application of microflow, capillary or nano liquid chromatography (≤ 1.0 mm column internal diameter (i.d.)) theoretically should accomplish a higher assay sensitivity compared to analytical liquid chromatography (2.1–5.0 mm column internal diameter). To date, there has been limited research into microflow UHPLC-MS assays that can be applied to study samples of low volume or mass.MethodsThis paper presents three complementary UHPLC-MS assays (aqueous C18 reversed-phase, lipidomics C18 reversed-phase and Hydrophilic Interaction Liquid Chromatography (HILIC)) applying 1.0 mm internal diameter columns for untargeted metabolomics. Human plasma and urine samples were applied for the method development, with porcine plasma, urine and tear fluid used for method assessment. Data were collected and compared for columns of the same length, stationary phase and stationary phase particle size but with two different column internal diameters (2.1 mm and 1.0 mm).Results and conclusionsAll three assays showed an increase in peak areas and peak widths when applying the 1.0 mm i.d. assays. HILIC assays provide an advantage at lower sample dilutions whereas for reversed phase (RP) assays there was no benefit added. This can be seen in the validation study where a much higher number of compounds were detected in the HILIC assay. RP assays were still appropriate for small volume samples with hundreds of compounds being detected. In summary, the 1.0 mm i.d. column assays are applicable for small volume samples where dilution is required during sample preparation.

Honey, I Shrunk the Columns: Recent Advances in the Design and Use of Narrow Internal Diameter UHPLC Columns

The quest for increased sensitivity for low-concentration samples continues at an accelerated pace, especially in the pharmaceutical and clinical environments. High-end mass spectrometry instruments can enable the analyst to detect analytes at attomolar concentrations, but can be prohibitively expensive and require well-trained scientists to operate and interpret the data. Another approach to improve the sensitivity of high performance liquid chromatography (HPLC) assays is by decreasing the internal diameter (i.d.) of the column. Reducing the column’s internal diameter down from 4.6 mm to 2.1 mm has been in practice for several decades. However, extra caution is required when considering going down below 2.1-mm i.d. into the “capillary-flow” format. This paper describes advances in column design that enable the analyst to achieve high sensitivity and efficiency when reducing the internal diameter of the column. After an introduction outlining the theory of increased sensitivity with narrow and capillary-scale internal diameter columns, a novel 1.5-mm i.d. column format will be presented detailing how sensitivities comparable to capillary-like separations can be achieved with this column geometry. Application examples will demonstrate the performance benefits of utilizing a 1.5-mm i.d. column geometry.

A High-Sensitivity Low-Nanoflow LC-MS Configuration for High-Throughput Sample-Limited Proteomics.

This work demonstrates the utility of high-throughput nanoLC-MS and label-free quantification (LFQ) for sample-limited bottom-up proteomics analysis, including single-cell proteomics (SCP). Conditions were optimized on a 50 μm internal diameter (I.D.) column operated at 100 nL/min in the direct injection workflow to balance method sensitivity and sample throughput from 24 to 72 samples/day. Multiple data acquisition strategies were also evaluated for proteome coverage, including data-dependent acquisition (DDA), wide-window acquisition (WWA), and wide-window data-independent acquisition (WW-DIA). Analyzing 250 pg HeLa digest with a 10-min LC gradient (72 samples/day) provided >900, >1,800, and >3,000 protein group identifications for DDA, WWA, and WW-DIA, respectively. Total method cycle time was further reduced from 20 to 14.4 min (100 samples/day) by employing a trap-and-elute workflow, enabling 70% mass spectrometer utilization. The method was applied to library-free DIA analysis of single-cell samples, yielding >1,700 protein groups identified. In conclusion, this study provides a high-sensitivity, high-throughput nanoLC-MS configuration for sample-limited proteomics.

Fingerprinting and identification of isolated plastic polymers with pyrolysis comprehensive two-dimensional gas chromatography and flame ionization detection.

An approach using pyrolysis with comprehensive two-dimensional gas chromatography with flame ionization detection is introduced for identifying common isolated plastic polymers. A quadrupole mass spectrometer is employed as a parallel detector to aid method development and improve polymer identification in complex matrices. Common plastic polymers including polyethylene, polypropylene, polystyrene, polyvinyl chloride, polyamide, poly(methyl methacrylate), styrene-butadiene rubber, and polyethylene terephthalate are accurately identified within a total analysis time of 45min. A strategy to enhance compatibility of high-resolution capillary gas chromatography using a 150-µm internal diameter column technology and a larger internal volume microfurnace-based pyrolyzer is discussed. This strategy resulted in minimizing the band broadening effect caused by the pyrolyzer's internal volume and overcoming the slow pressure buildup when the sample is inserted into the furnace. Prolonged pressure buildup to reach a final pressure setting can cause a safety shutdown to the pneumatic control system. The developed approach is complementary to spectroscopic techniques by offering mass based, chemical composition analysis of plastics.

Implementing 1.5 mm internal diameter columns into analytical workflows

The use of smaller column diameters in liquid chromatography (LC) is often associated with capillary LC. Although there are many analytical benefits gained by adapting this format, routine use continues to be challenging due to column fragility and extra column dispersion. Bridging the gap between routinely used 2.1 mm columns and capillary bore columns allows for a sequential but far from insignificant increase in performance without the need for specialized equipment associated with very low dispersion LC systems. Moreover, an incremental decrease in column internal diameter (i.d.) allows for similar mass load (avoiding column overload that may be observed in much larger decreases in i.d. without trapping) and thus an increase in measured signal. As such, 1.5 mm i.d. columns provide an alternative intermediate dimension between the more regularly used 2.1 mm i.d. columns and 1 mm i.d. columns. These columns balance an increase in sensitivity compared to 2.1 mm i.d. columns (theoretically doubling the time-domain peak area in mass sensitive detectors for the same mass load), while mitigating the efficiency losses due to extra-column dispersion effects that are commonly observed with 1.0 mm i.d. columns. Here, the use of 1.5 mm i.d. columns was applied to LC/UV analysis of small molecules and LC/MS methods for the analysis of monoclonal antibodies. With equivalent mass load on column, the 1.5 mm i.d. columns provide two-to-threefold improvement in analyte peak area signal for small molecules as well as intact, subunit, and peptide levels of antibody analysis. Peak height was also increased using the 1.5 mm i.d. column, although the scale of increase varies between isocratic and gradient modes, likely due to differences in system dispersion effects and variation in electrospray ionization efficiency at different flow rates.

Optimal heating rate in constant pressure and constant flow gas chromatography

Optimal heating rate is the one resulting in the shortest analysis time for achieving a required separation performance of a column. The previously recommended default heating rate (RT,Def ) was optimal for temperature-programmed gas chromatography analyses in constant pressure mode. It has been shown herein that the same recommendation can be extended to constant flow mode with fixed heating rate (RT ). The numerical value of RT,Def has been herein rescaled from previous (10°C per void time) where tM was measured at 50°C, to with tM measured at 150°C-a round number in the middle of the gas chromatography temperature range, chosen as a reference temperature for numerical values of all temperature-dependent gas chromatography parameters. It has been experimentally found based on theory developed herein that is optimal for columns with ( is dimensionless film thickness, d and df are the column internal diameter and film thickness, respectively) in constant pressure mode and constant flow mode with fixed RT . Theory shows that, for arbitrary φ, . The theory also shows that the fixed RT is optimal for constant pressure mode. In constant flow mode, however, the optimal RT should gradually increase with increasing temperature (T). The optimal theoretical curves RT (T), different for different flow rates, were found. However, only the optimization of the fixed RT was experimentally evaluated due to limited capability of existing gas chromatography instrumentation and resources. It has been shown that the separation-time tradeoff in constant pressure mode is slightly better than that in constant flow mode. The experimental data are compiled in the Supporting information.

Comparison of existing strategies for keeping symmetrical peaks in on-line Hydrophilic Interaction Liquid Chromatography x Reversed-Phase Liquid Chromatography despite solvent strength mismatch

In two-dimensional liquid chromatography, the combination of hydrophilic interaction liquid chromatography (HILIC) and reversed-phase liquid chromatography (RPLC) is very attractive due to the complementarity of their separation mechanisms. On-line comprehensive HILIC x RPLC is well-known to give rise to a large retention space coverage when dealing with ionisable compounds. However, method development in on-line HILIC x RPLC is challenging due to the reversed solvent strength between both dimensions, which can greatly affect the peak shapes in the second RPLC dimension, and thus the separation quality and the method sensitivity. In the present contribution, we compared four strategies designed to avoid this problem: (1) flow splitting, which consists in reducing the injection volume in the second dimension (2D), (2) on-line dilution with a make-up flow and (3) on-line dilution with Active Solvent Modulation (ASM), which both consist in reducing the solvent strength of the injected fractions, and (4) Total Breakthrough Strategy, which we recently proposed. Unlike the three preceding strategies, this latter one consists in injecting large volumes of strong solvent in 2D. The performance of each strategy was evaluated for sub-hour separations of a tryptic digest in on-line HILIC x RPLC. In this work, we considered the critical case for which the same column internal diameters (i.e. 2.1 mm here) are used in both dimensions. Peak capacity, peak shapes and peak intensities were considered for this evaluation. The highest peak capacity could be achieved with Total Breakthrough Strategy while the lowest one with on-line dilution using ASM. Peak intensities were usually higher with on-line dilution approaches (make-up flow and ASM). However, despite the presence of breakthrough, peak intensities were approximately 7-fold higher with Total Breakthrough Strategy than with flow splitting.

Heavy metals bioremediation by shells dust and chitosan derived from Belamya javanica Snail, an Eco-friendly biosorbent

The aim of this study is to utilize the shells dust of Belamya javanica snail (BSD) and produce chitosan from the same shells, then evaluate the ability of the shells dust and chitosan to remove heavy metals Cu2+, Cd2+, and Pb2+ from aquatic systems. Chitosan produced were characterized based on the FTIR spectra, the degree of deacetylation (DD), pH, water, and ash content. The adsorption capacity of chitosan and shells dust was evaluated by using it as a package in 150mm high, 17.5mm internal diameter columns. The heavy metals used were prepared at approximately 10mg/L using primary standards, then passed through the columns for 30 minutes. The results indicated that chitosan and BSD had shown the potential to remove copper, cadmium, and lead with adsorption percentage up to 95%, in which was also at the second elution The results obtained from the experiment were that chitosan and the BSD produced could be used as an efficient and environmental friendly biosorbent for copper, cadmium, and lead from aqueous solution.

Hyphenation of packed column supercritical fluid chromatography with mass spectrometry: where are we and what are the remaining challenges?

Over the last decades, packed column supercritical fluid chromatography (pSFC) using carbon dioxide (CO2) as supercritical fluid has gained interest as a complementary separation technique to liquid chromatography (LC). Various commercial solutions for the hyphenation to atmospheric pressure ionization (API) including electrospray (ESI) and atmospheric pressure chemical ionization (APCI) have been described using split-flow or full-flow introduction approaches. This review discusses various aspects of the hyphenation using these two approaches. It also illustrates the difference between LC-MS and SFC-MS. The benefits and challenges of the decoupling of the separation mobile phase from the ionization conditions are also pointed out. The effect of CO2 on ESI performance and the adduct reduction are also discussed. Finally, limitation of current hardware and the limited use of smaller column internal diameters (i.d.) are discussed. Graphical abstract.

Development and validation of high-performance liquid chromatographic method for quantification of Irinotecan and its active metabolite SN-38 in colon tumor bearing NOD/SCID mice plasma samples: application to pharmacokinetic study

Irinotecan (IRT) is an antineoplastic agent widely used in the treatment of various cancers primarily in colorectal cancer. A new, simple and sensitive high-performance liquid chromatography (HPLC) method coupled with fluorescence detector was developed and validated to quantify IRT and its active metabolite SN38 in the plasma of non-obese diabetic/severe combined immune-deficient mice (NOD/SCID) mice bearing colon tumor. The plasma samples were extracted by precipitation method using acetonitrile with 0.1% formic acid. The chromatographic separation was achieved using mobile phase consisted of water and acetonitrile (57:43 v/v) pH 3 at the flow rate of 0.8 mL/min in C18 column (internal diameter, 250 × 4.6 mm; pore size, 5 μm). The method was validated according to the bioanalytical guidelines defined by Food and Drug Administration (FDA) and European Medicine Agency (EMA). A regression (R2) value of 0.999 and 0.997 for IRT and SN38 suggested the good linearity in the range of 0.1–10 μg/mL and 5–500 ng/mL, respectively. The calculated lower limit of quantification (LLOQ) and limit of detection (LOD) for IRT were 0.1 and 0.065 μg/mL, respectively. However, for SN38, LLOQ and LOD were 5 and 2 ng/mL, respectively. The intra-day and inter-day variations (coefficient of variance; % CV) observed during the validation were found to be within the set limit of 15%. Both accuracy and percentage recovery analyzed and calculated from the quality control samples were in the between the defined range of 85–115%. Plasma samples were found to be stable when stored at room temperature for 2 h, after 2 freeze–thaw cycles and at −80 °C for 2 months. The developed method was successfully applied to study the plasma elimination profile of IRT in NOD/SCID mice with tumor. The results from plasma concentration time profile and pharmacokinetic parameter analyzed suggested the rapid elimination of IRT and SN38 from the plasma of NOD/SCID mice.

Multi-particle frits for packed capillary columns in electrochromatographic use

Further development of capillary electrochromatography (CEC) is to some extent restricted by column technology. Packed capillary columns are most commonly used in CEC because of higher sample capacity and higher sensitivity than open tubular columns, and better reproducibility and uniformity than monolithic columns. Fritting is the core component for the fabrication of packed capillary columns. Therefore various fritting technologies attract continuous attention from analytical community. A convenient fritting method was developed in this study. In contrast to the reported single-particle fritting, this method applies a multi-particle fritting technology to hold stationary phase in capillary columns. Several large porous particles smaller than the column internal diameter were tapped in and immobilized in inlet and outlet of the packing material bed due to direct and indirect keystone effect. Except the smallest permeability by the sintered frit and the lowest breaking pressure by the single-particle frit, EOF mobility and peak dispersion of the multi-particle frit were at the same levels with the formers. In addition, all the three frits avoided bubble formation. It demonstrated that the multi-particle frits met the general requirements of permeability, mechanical resistance and efficiency for packing capillary microcolumns as well as the sintered frits and the single-particle frits. Thus, peak efficiency around 4.3 μm plate height (approximately 232,560 plates per meter) was obtained by multi-particle fritted capillary columns with electrokinetic packing, compared to 6.5 μm plate height by single-particle fritted columns with slurry pressure packing. In particular, the multi-particle fritting was simpler, faster, and more reproducible than the sintered fritting, and the multi-particle fritting was suitable to pack capillary columns of various capillary bore sizes owing to rich commercial resources of fritting particles of various diameters. Multi-particle fritted columns offered satisfactory run-to-run and column-to-column reproducibilities for retention factor, plate height and peak area (precisions of 4.1–9.3% for 10 replicated columns, and precisions of 0.9–3.1% for 30 replicated trials). They also had consistent CEC performance (differences of 1.1–3.9% between the first and 31 st day) during a lifetime more than one month. Application examples in packing capillary columns of 75 and 100 μm i.d. and CEC analysis of sixteen polycyclic aromatic hydrocarbons prove the diversity and reliability of the multi-particle fritting in column technology.

The efficacy of the ozonation process in the presence of activated carbon impregnated with magnesium oxide in the removal of benzene from the air stream

The main purpose of this study was to investigate the catalytic ozonation process with GAC–MgO in removal of benzene in air streams. The flow diagram of the experimental bench-scale setup included a glass column (internal diameter = 3 cm) packed for 10 cm with granular activated carbon coated with magnesium oxide. Different initial concentrations of benzene (80, 200 and 400 ppm) were used to evaluate the removal efficiency. The synthesized granular activated carbon coated by magnesium oxide was a microporous adsorbent with the BET specific surface area of 1028 m2/g. Hydroxyl, methylene, methyl, carboxylic groups and aromatic C=C bonds were some of the several functional groups identified on the its surface. In the catalytic ozonation process, the breakthrough time for the inlet concentration of 200 ppm increased from 36 h for granular activated carbon to 53 h for granular activated carbon ozonation. In other words, the concomitant use of activated carbon and ozone enhanced (from 36 h to 53 h) the removal efficiency of the system by 17 h in comparison with activated carbon alone. In the ozonation process for the bed, the benzene breakthrough time of the bed, for the inlet concentration of 200 ppm, increased from 44 h in the granular activated carbon coated by magnesium oxide system to 78 h in the granular activated carbon coated by magnesium oxide and ozonation. Eventually, the results showed the ozonation process in combination with the granular activated carbon coated by magnesium oxide catalyst.

Effect of a sweeping air stream and gas–phase aspect ratio of an isothermal Stefan diffusion column on the experimental estimation of binary gas diffusivities

The Stefan column consists of liquid A evaporating into an inert/stagnant gas B with a sweeping B stream at the top. It was designed to estimate binary gas diffusivities, DAB’s, but “end effects” such as gas mixing at the top and interfacial curvature have been either ignored or uncorrelated to the operational settings. This study’s hypothesis is that gas mixing at the top and the gas–phase aspect ratio affect DAB estimation in the acetone (A)-ambient air (B) system at 50 °C. The sweeping stream Reynolds number (Re) and the gas–phase aspect ratio (AR = initial gas phase height to column internal diameter) were the variables tested. Isothermal evaporation-diffusion experiments were conducted in which the temporal interfacial descent was tracked. The settings were 492 ≤ Re ≤ 5378 and AR between 5 and 15. A 1D transport model allowed determination of the experimental diffusivity, DAB,exp, by nonlinear regression. For Re < 600, the DAB,exp errors relative to DAB,CE (predicted by the Chapman–Enskog kinetic theory for low-density gases) were small and unrelated to AR, while for Re > 600 the errors increased considerably with Re and were inversely proportional to AR. This study is the first to relate the column’s operational settings to the DAB estimation errors. The column should be operated at low sweeping gas Re and large AR for accurate DAB,exp’s. The low Re region deserves further study, while the present transport model may have to be replaced by computational fluid dynamics simulations to account for the multidimensional gas flow patterns.

Greener Chromatographic Approaches for Dissolution Testing of Solid Pharmaceutical Formulations

Dissolution testing is a critical enabler for formulation process development as well as determination of product quality at release and on stability for solid oral dosage forms. In the pharmaceutical industry worldwide, this type of test is performed in the range of millions annually. We developed an improved chromatographic protocol combining the utilization of smaller internal diameter (i.d.) columns, superficially porous column technology, injection cycle time for gradient re-equilibration, system dwell volume understanding, and basic separation concepts for optimization as a greener, faster, yet robust way to conduct dissolution testing. Comparing this approach to standard analysis using a conventional approach, this methodology provides 70–80% reduction in solvent consumption and waste generation, as well as run times with equivalent accuracy, precision, and robustness. Feasibility of this approach was demonstrated by applying it to multiple drug products and those head-to-head comparisons showed that dissolution profiles and overall variability are comparable to those obtained by the conventional chromatographic approaches.

CFD simulation of a slurry bubble column: Effect of population balance kernels

In this work, CFD simulations of a slurry bubble columns were performed and validated with experimental data. The superficial gas velocities investigated in the experiments were 5 cm/s and 8 cm/s for a solid loading varying from 7% to 14% in a 0.14 m to 0.26 m internal diameter column. A population balance model (PBM) was implemented and solved numerically using the bubble class method. The closure kernels involved Luo's coalescence model as well as two different breakup model: Luo's and Lehr's models. The kinetic theory of granular flows (KTGF) was used to calculate the local solid viscosity. The Eulerian framework was selected for multi-phase calculations and the interphase momentum transfer for gas–liquid included drag, lift, and turbulent dispersion terms. The liquid–solid interphase involved both drag and turbulent dispersion terms. Results showed that: (a) A combination of the Luo coalescence and Luo breakup kernels (Luo–Luo) can predict similar trends for the radial profile of solid axial velocity with regards to experiments. However, at the highest superficial gas velocity in the small column, a discrepancy was observed in the results. (b) A combination of the Luo coalescence and Lehr breakup kernels (Luo–Lehr) predicted similar trends for the solid velocities when compared to empirical data, however, this combination also showed discrepancy at the highest superficial velocity tested in the small column. (c) The total gas holdup in the Luo–Luo and Luo–Lehr models are under and over predicted respectively when compared with the experimental results. (d) The mesh sensitivity results showed that a 3 mm mesh size is realistic to capture the flow fields. (e) The sensitivity analysis of the number of bubble classes showed that the radial profiles were independent to a number of bubble classes. However, the highest number of classes cover the formations of the largest bubbles.

Characterization of ion exchange functionalized cyclic olefin polymer open tubular columns

We describe both the cation-exchange (CEX) and anion-exchange (AEX) capacity measurements of functionalized cyclic olefin polymer (COP) open tubular (OT) columns. COP capillaries were sulfonated to provide CEX functionality and then coated with AEX latex to provide AEX functionality. We measure functionalization uniformity along the column by (a) separately measuring capacities of two halves of a column, (b) measuring retention factors for several ions in successive 10 cm segments of a column in multiple injections and (c) measuring retention variance of a single ion in different segments of a column from a single injection. Data were collected in both flow directions. Errors in CEX capacity determination arises primarily from uncertainties in titration end point location. AEX capacities are typically 10x higher, reducing the relative measurement error. In the AEX case, both titration end point location error and variations in column internal diameter contribute to the overall uncertainty. Although sulfonation of COP results in nonuniform CEX functionalization, AEX latex coating of this surface results in uniform AEX coverage. Frontal displacement chromatography (using both admittance and optical detection) and acid-base titrimetry were compared for capacity measurement. In a pooled standard deviation based t-test for the data on 8 columns, at the 95% confidence level, CEX capacity differed significantly between two halves of a column. But AEX capacity of AEX columns prepared by coating the same CEX columns with AEX latex did not.

Mass transfer efficiency of a tall and low plate free area liquid pulsed sieve-plate extraction column

Mass transfer performance is studied in a tall, thin, and low plate free area liquid pulsed sieve-plate extraction column. The 5.0 cm internal diameter column consists of eighty sieve plates with percent free area of only 13.5. The effects of pulsation intensity (product of amplitude and frequency) and dispersed phase velocity are studied on the extraction efficiency of the column for the acetic acid–kerosene–water system. A mass transfer correlation for the measurement of overall mass transfer coefficient is developed that best-fits the experimental data obtained in the present study. Mathematical analysis of the column is carried out that shows the insignificance of axial diffusivities in the column.

High sensitivity and selectivity in quantitative analysis of drugs in biological samples using 4-column multidimensional micro-UHPLC-MS enabling enhanced sample loading capacity

Sensitivity is often a critical parameter in quantitative bioanalyses in drug development. For liquid-chromatography-based methods, sensitivity can be improved by reducing the column diameter, but practical sensitivity gains are limited by the reduced sample loading capacity on small internal diameter (I.D.) columns. We developed a set-up that has overcome these limitations in sample loading capacity. The set-up uses 4 columns with gradually decreasing column diameters along the flow-path (2.1 → 1 → 0.5 → 0.15 mm). Samples are pre-concentrated on-line on a 2.1 mm I.D. trapping column and back flushed to a 1 mm I.D. UHPLC analytical column and separated. The peak(s) of interest are transferred using heartcutting to a second trapping column (0.5 mm I.D.), which is back-flushed to a 0.15 mm I.D. micro-UHPLC analytical column for orthogonal separation. The proof of concept of the set-up was demonstrated by the simultaneous analysis of midazolam and 1′-hydroxy midazolam in plasma by injection of 80 μL of protein precipitated plasma. The 4-column funnel set-up proved to be robust and resulted in a 10–50 times better sensitivity compared to a trap-elute approach and 250–500 fold better compared to direct micro-UHPLC analysis. A lower limit of quantification of 100 fg/mL in plasma was obtained for both probe compounds.

Removal of hydrogen sulfide in air using cellular concrete waste: Biotic and abiotic filtrations

The objective of this study was to investigate the removal of hydrogen sulfide (H2S) present in air using cellular concrete waste as the packing material. Air filtration was performed under biotic and abiotic conditions. Experiments were carried out in a laboratory-scale PVC column (internal diameter of 300mm) filled with a volume of 70L of cellular concrete (1m height). The polluted air flow was generated at 4m3h−1 corresponding to an Empty Bed Residence Time (EBRT) of 63s. In dry conditions without biomass (abiotic conditions), cellular concrete can be an effective medium for the treatment of H2S in air. For an H2S concentration of 100ppmv, the removal efficiency was around 70% (Elimination Capacity (EC) of 5.6gm−3h−1). This finding can be explained by the physicochemical reactions that can take place between H2S and the cellular concrete components (mainly CaO, CaCO3 and Fe2O3). However, interactions between cellular concrete and H2S are not yet fully understood. Used as a packing material for H2S biofiltration (biotic conditions), cellular concrete waste efficiently treated (Removal Efficiency=100%) high concentrations of H2S (up to 133ppmv corresponding to an EC of up to 10.5gm−3h−1). Physicochemical and biological mechanisms explaining H2S removal seem to occur simultaneously in the biofilter. At an EBRT of 63s, the maximal elimination capacity (ECmax) was 17.8gm−3h−1. A packed bed of cellular concrete also presents a satisfactory mechanical behavior with low pressure drops.

Determination of thyroid hormones in mouse tissues by isotope-dilution microflow liquid chromatography-mass spectrometry method.

Thyroid hormones (THs) play a critical role in the regulation of many biological processes such as growth, metabolism and development both in humans and wildlife. In general, TH levels are measured by immunoassay (IA) methods but the specificity of the antibodies used in these assays limits selectivity. In the last decade, several analytical methods using liquid chromatography-mass spectrometry (LC-MS) and tandem mass spectrometry (LC-MS/MS) have been developed to measure THs. These new techniques proved to be more accurate than the IA analysis and they were widely used for the determination of TH level in different human and animal tissues. A large part of LC-MS/MS methods described in literature employed between 200 and 500mg of sample, however this quantity can be considered too high especially when preclinical studies are conducted using mice as test subjects. Thus an analytical method that reduces the amount of tissue is essential. In this study, we developed a procedure for the analysis of six THs; L-thyroxine (T4), 3,3',5-triiodo-l-thyronine (T3), 3,3',5'-triiodo-l-thyronine (rT3), 3,5-diiodo-l-thyronine (rT2), 3,3'-diiodo-l-thyronine (T2), 3-iodo-l-thyronine (T1) using isotope ((13)C6-T4, (13)C6-T3, (13)C6-rT3, (13)C6-T2) dilution liquid chromatography-mass spectrometry. The major difference with previously described methods lies in the utilization of a nano-UPLC (Ultra Performance Liquid Chromatography) system in micro configuration. This approach leads to a reduction compared to the published methods, of column internal diameter, flow rate, and injected volume. The result of all these improvements is a decrease in the amount of sample necessary for the analysis. The method was tested on six different mouse tissues: liver, heart, kidney, muscle, lung and brown adipose tissue (BAT). The nano-UPLC system was interfaced with a quadrupole time-of-flight mass spectrometer (Q-TOF2-MS) using the positive ion mode electrospray ionization. In our analytical method the instrumental calibration curves were constructed from 0 to 100pgμL(-1) and all of them showed good linearity (r(2)>0.99). The limit of quantification was from 2.5 to 5pg injected into the column. The method recoveries calculated using spiked mouse liver and spiked mouse muscle were between 83% and 118% (except T1 and rT2 at high concentration) with a coefficient of variation (CV) of <10% for all derivatives. The new methodology allows us to measure T4 and T3 concentrations in a range from 21 to about 100mg and give a more extensive insight on thyroid hormone concentration in different mouse tissue.