Dissolved atmospheric gas in xylem sap measured with membrane inlet mass spectrometry

A new method is described for measuring dissolved gas concentrations in small volumes of xylem sap using membrane inlet mass spectrometry. The technique can be used to determine concentrations of atmospheric gases, such as argon, as reported here, or for any dissolved gases and their isotopes for a variety of applications, such as rapid detection of trace gases from groundwater only hours after they were taken up by trees and rooting depth estimation. Atmospheric gas content in xylem sap directly affects the conditions and mechanisms that allow for gas removal from xylem embolisms, because gas can dissolve into saturated or supersaturated sap only under gas pressure that is above atmospheric pressure. The method was tested for red trumpet vine, Distictis buccinatoria (Bignoniaceae), by measuring atmospheric gas concentrations in sap collected at times of minimum and maximum daily temperature and during temperature increase and decline. Mean argon concentration in xylem sap did not differ significantly from saturation levels for the temperature and pressure conditions at any time of collection, but more than 40% of all samples were supersaturated, especially during the warm parts of day. There was no significant diurnal pattern, due to high variability between samples.     

Mass spectrometry as en ecological tool for in situ measurement of dissolved gases in sediment systems

Abstract The use of membrane-inlet quadrupole mass spectrometry, as a method for quantitative monitoring of dissolved gases in natural or semi-natural environments, is described. Its advantages over other methods lie in the fact that it provides an accurate, sensitive means for non-invasive, continuous analysis of several dissolved gases simultaneously. The potential of mass spectrometry as an ecological tool is illustrated by representative results from measurements made on undisturbed and experimentally amended estuarine and fresh-water sediments.
Dissolved gas profiles from the surface to a depth of 10 cm in the estuarine sediment showed that the dissolved oxygen decreased gradually until at 10 cm it was undetectable (< 0.25 μM); dinitrogen reached a maximum at 6 cm, where oxygen was 20 μM. In a fresh-water sediment, methane reached 1.5 mM at 10 cm depth. NO_x was also detected; quantitation of carbon dioxide necessitates a correction for the contribution of NO_x. Manipulation of conditions (gas phase, nitrogen and carbon sources) permitted ecological modelling.     

Sequential V(A)/Q distributions in the normal rabbit by micropore membrane inlet mass spectrometry.

We developed micropore membrane inlet mass spectrometer (MMIMS) probes to rapidly measure inert-gas partial pressures in small blood samples. The mass spectrometer output was linearly related to inert-gas partial pressure (r(2) of 0.996-1.000) and was nearly independent of large variations in inert-gas solubility in liquid samples. We infused six inert gases into five pentobarbital-anesthetized New Zealand rabbits and used the MMIMS system to measure inert-gas partial pressures in systemic and pulmonary arterial blood and in mixed expired gas samples. The retention and excretion data were transformed into distributions of ventilation-to-perfusion ratios (V(A)/Q) with the use of linear regression techniques. Distributions of V(A)/Q were unimodal and broad, consistent with prior reports in the normal rabbit. Total blood sample volume for each VA/Q distribution was 4 ml, and analysis time was 8 min. MMIMS provides a convenient method to perform the multiple inert-gas elimination technique rapidly and with small blood sample volumes.     

Observation of a chaotic multioscillatory metabolic attractor by real-time monitoring of a yeast continuous culture

We monitored a continuous culture of the yeast Saccharomyces cerevisiae by membrane-inlet mass spectrometry. This technique allows very rapid simultaneous measurements (one point every 12 s) of several dissolved gases. During our experiment, the culture exhibited a multioscillatory mode in which the dissolved oxygen and carbon dioxide records displayed periodicities of 13 h, 36 min and 4 min. The 36- and 4-min modes were not visible at all times, but returned at regular intervals during the 13-h cycle. The 4-min mode, which has not previously been described in continuous culture, can also be seen when the culture displays simpler oscillatory behavior. The data can be used to visualize a metabolic attractor of this system, i.e. the set of dissolved gas concentrations which are consistent with the multioscillatory state. Computation of the leading Lyapunov exponent reveals the dynamics on this attractor to be chaotic.     

Use of a portable quadrupole mass spectrometer for the measurement of dissolved gas concentrations in ovine rumen liquor in situ

The use of membrane-inlet mass spectrometry in the study of dissolved gas concentrations in the rumen was evaluated in order to assess the value of the technique as a tool for the study of microbial activity in ecosystems in situ. Four dissolved gases (CH₄, CO₂, H₂, and O₂) were measured simultaneously and continuously for short periods (up to 30 min) during the feeding period. These preliminary results have demonstrated the usefulness of the technique for monitoring microbial activity via gas production in a complex natural ecosystem.     

Construction and performance of plug-in membrane inlet mass spectrometer for fermentor monitoring

An in situ sterilizable plug-in membrane inlet mass spectrometer for monitoring dissolved gases and volatiles in fermentors was constructed and tested. The design ensured a minimal distance to be traveled by analyte molecules from the bulk of the fermentation broth to the ionization chamber of the mass spectrometer. Apart from the specific cross talk due to overlapping mass peaks from different compounds, we found that carbon dioxide interfered unspecifically with all the mass peaks of other substances, changing them by the same factor. The interference changed slowly with time and could be positive or negative depending on the history of the mass spectrometer. Also, the general sensitivity of the instrument changed slowly with time. These effects can be neglected or corrected for empirically in short-term measurements. When the fermentor was aerated with a three-component gas mixture including carbon dioxide, a rapid change in the partial pressure of carbon dioxide in the gas mixture gave rise to a transient in the signal of a gas whose partial pressure was kept constant. This effect revealed a transient change in the composition of the gas mixture in the bubbles caused by net import or export of carbon dioxide during equilibration with the new gas mixture. An experimental method to determine the effective partial pressures of gases in the bubbles during steady-state transport of carbon dioxide was designed. The plug-in membrane inlet mass spectrometer was tried as a probe for oxygen and ethanol in an oxystatic culture of the yeast Pichia stipitis. We found that it was possible to keep a steady-state concentration of as little as 0.5 muM throughout the lifetime of the culture. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 54: 535-542, 1997.     

Simultaneous quantitation of chlormethiazole and two of its metabolites in blood and plasma by gas—liquid chromatography

A simple and rapid gas chromatographic method for the quantitation of chlormethiazole and two of its pharmacologically active metabolites, 5-acetyl-4-methylthiazole and 5-(1-hydroxyethyl)-4-methylthiazole, in plasma and blood is described. The total analysis time is less than 20 min for a single sample. The method requires 50–500 μl of plasma or 1 ml of blood. The compounds are detected with a nitrogen—phosphorus detector. An internal standard technique is used for the quantitation. Calibration data are linear over the range 32–2376 ng of chlormethiazole and a similar range of the metabolites in plasma. The method may be used for pharmacokinetic studies.     

Liquid-to-Gas Mass Transfer in Anaerobic Processes: Inevitable Transfer Limitations of Methane and Hydrogen in the Biomethanation Process

Liquid-to-gas mass transfer in anaerobic processes was investigated theoretically and experimentally. By using the classical definition of k(L)a, the global volumetric mass transfer coefficient, theoretical development of mass balances in such processes demonstrates that the mass transfer of highly soluble gases is not limited in the usual conditions occurring in anaerobic fermentors (low-intensity mixing). Conversely, the limitation is important for poorly soluble gases, such as methane and hydrogen. The latter could be overconcentrated to as much as 80 times the value at thermodynamic equilibrium. Such overconcentrations bring into question the biological interpretations that have been deduced solely from gaseous measurements. Experimental results obtained in three different methanogenic reactors for a wide range of conditions of mixing and gas production confirmed the general existence of low mass transfer coefficients and consequently of large overconcentrations of dissolved methane and hydrogen (up to 12 and 70 times the equilibrium values, respectively). Hydrogen mass transfer coefficients were obtained from the direct measurements of dissolved and gaseous concentrations, while carbon dioxide coefficients were calculated from gas phase composition and calculation of related dissolved concentration. Methane transfer coefficients were based on calculations from the carbon dioxide coefficients. From mass balances performed on a gas bubble during its simulated growth and ascent to the surface of the liquid, the methane and carbon dioxide contents in the gas bubble appeared to be controlled by the bubble growth process, while the bubble ascent was largely responsible for a slight enrichment in hydrogen.     

Effects of acetone in heparin on the multiple inert gas elimination technique

We have detected acetone in several brands of heparin. If uncorrected, this leads to errors in measuring acetone in blood collected in heparinized syringes, as in the multiple inert gas elimination technique for measuring ventilation-perfusion ratio (VA/Q) distributions. Error for acetone retention [R = arterial partial pressure-to-mixed venous partial pressure (P-V) ratio] is usually small, because R is normally near 1.0, and the error is similar in arterial and mixed venous samples. However, acetone excretion [E = mixed expired partial pressure (P-E)-to-P-V ratio] will appear erroneously low, because P-E is accurately measured in dry syringes, but P-V is overestimated. A physical model of a homogeneous alveolar lung at room temperature and without dead space shows: the magnitude of acetone E error depends upon the ratio of blood sample to heparinized saline volumes and acetone partial pressures, without correction, acetone E can be less than that of less soluble gases like ether, a situation incompatible with conventional gas exchange theory, and acetone R and E can be correctly calculated using the principle of mass balance if the acetone partial pressure in heparinized saline is known. Published data from multiple inert gas elimination experiments with acetone-free heparin, in our labs and others, are within the limits of experimental error. Thus the hypothesis that acetone E is anomalously low because of physiological mechanisms involving dead space tissue capacitance for acetone remains to be tested.     

A numerical study of the nonsteady transport of gases in the pulmonary capillaries

A mathematical model is formulated for simulating the unsteady transport of gases in the blood flowing through the pulmonary capillaries. The formulation takes into account the transport mechanisms of molecular diffusion, convection and facilitated diffusion of the species due to haemoglobin. A time dependent situation is created by allowing to vary suddenly the partial pressures of the gases either in the venous blood or in the alveolar air. A numerical technique is described to solve the resulting time-dependent system of nonlinear coupled partial differential equations with the physiologically relevant boundary, entrance and initial conditions. The time required by the gases to achieve equilibrium is computed. It is shown that the dissolved oxygen takes longest in reaching equilibration whereas the carbon dioxide is the fastest. The various physiologically relevant unsteady situations have been examined.     

Determination of Natural In Vivo Noble-Gas Concentrations in Human Blood

Although the naturally occurring atmospheric noble gases He, Ne, Ar, Kr, and Xe possess great potential as tracers for studying gas exchange in living beings, no direct analytical technique exists for simultaneously determining the absolute concentrations of these noble gases in body fluids in vivo. In this study, using human blood as an example, the absolute concentrations of all stable atmospheric noble gases were measured simultaneously by combining and adapting two analytical methods recently developed for geochemical research purposes. The partition coefficients determined between blood and air, and between blood plasma and red blood cells, agree with values from the literature. While the noble-gas concentrations in the plasma agree rather well with the expected solubility equilibrium concentrations for air-saturated water, the red blood cells are characterized by a distinct supersaturation pattern, in which the gas excess increases in proportion to the atomic mass of the noble-gas species, indicating adsorption on to the red blood cells. This study shows that the absolute concentrations of noble gases in body fluids can be easily measured using geochemical techniques that rely only on standard materials and equipment, and for which the underlying concepts are already well established in the field of noble-gas geochemistry.     

Direct measurement of dissolved gases in microbiological systems using membrane inlet mass spectrometry

Membrane inlet mass spectrometry is a novel technique that has been used to measure concentrations of dissolved gases and volatile compounds of microbiological interest. This technique is compared with other methods of measuring dissolved gases. Applications to some microbiological processes (respiration, photosynthesis, fermentation, nitrogen fixation and methanogenesis) are discussed in greater detail. The advantages of the technique and possible future developments are presented; its major attraction is that a number of different gases can be simultaneously and continuously monitored directly and non-invasively in cell suspensions.     

Stopped flow mass spectrometry: applications to the carbonic anhydrase reaction

Membrane inlet mass spectrometry is combined with a stopped flow technique in order to allow the recording of moderately fast transients in concentrations of dissolved gases. The method was used for direct measurements of CO2 transients in the spontaneous and enzyme catalyzed hydrations of carbon dioxide. Rate constants and activation energies were determined.     

P blood group regulation of glycosphingolipid levels in human erythrocytes.

The neutral glycosphingolipid content of normal human erythrocytes was analyzed by a new method which utilizes high performance liquid chromatography. This rapid and accurate technique permits the quantitation of each of the major neutral glycolipids from individual blood samples. A correlation between the P blood group and the relative quantities of neutral glycosphingolipids is demonstrated. Erythrocytes from P1 individuals are shown to contain more globotriaosylceramide and less lactosylceramide than do erythrocytes from P2 individuals. The results of these experiments suggest the existence of a new phenotype in the P blood group system, and have further implications regarding the biosynthesis of the P blood group glycosphingolipids.     

The methane mono-oxygenase reaction system studied in vivo by membrane-inlet mass spectrometry.

A membrane-inlet mass spectrometer connected to an open-system cuvette was used for direct measurement of dissolved methane and O2 in bacterial samples of strain OU-4-1, a type II methanotrophic bacterium. A technique was applied for keeping the concentration of dissolved methane or O2 in the sample constant while the concentration of the other dissolved gas was varied. This allowed the reaction mechanism of methane mono-oxygenase to be studied in vivo. The enzyme was found to follow a random bi-reactant mechanism with respect to binding of methane and O2. Binding of one substrate decreased the affinity for the other. The true binding constants were 1 microM for methane and 0.14 microM for O2. Studies of HCN inhibition confirmed the random bi-reactant mechanism. HCN was found to be a non-exclusive inhibitor with a binding constant of 0.4 microM.     

Adjustments in oxygen transport during head-out immersion in water at different temperatures

Respiratory gas exchange was investigated in human subjects immersed up to the shoulders in water at different temperatures (Tw = 25, 34, and 40 degrees C). Cardiac output (Qc) and pulmonary tissue volume (Vti) were measured by a rebreathing technique with the inert gas Freon 22, and O2 consumption (VO2) was determined by the closed-circuit technique. Arterial blood gases (PaO2, PaCO2) were analyzed by a micromethod, and alveolar gas (PAO2) was analyzed during quiet breathing with a mass spectrometer. The findings were as follows. 1) Immersion in a cold bath had no significant effect on Qc compared with the value measured at Tw = 34 degrees C, whereas immersion in a hot bath led to a considerable increase in Qc. Vti was not affected by immersion at any of the temperatures tested. 2) A large rise in metabolic rate VO2 was only observed at Tw = 25 degrees C (P less than 0.001). 3) Arterial blood gases were not significantly affected by immersion, whatever the water temperature. 4) O2 transport during immersion is affected by two main factors: hydrostatic pressure and temperature. Above neutral temperature, O2 transport is improved because of the marked increase in Qc resulting from the combined actions of hydrostatic counter pressure and body heating. Below neutral temperature, O2 transport is altered; an increase in O2 extraction of the tissue is even calculated.     

Rapid analysis of pharmaceuticals and excreted xenobiotic and endogenous metabolites with atmospheric pressure infrared MALDI mass spectrometry

Atmospheric pressure (AP) infrared (IR) matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) was demonstrated for the rapid direct analysis of pharmaceuticals, and excreted human metabolites. More than 50 metabolites and excreted xenobiotics were directly identified in urine samples with high throughput. As the water content of the sample was serving as the matrix, AP IR-MALDI showed no background interference in the low mass range. The structure of targeted ions was elucidated from their fragmentation pattern using collision activated dissociation. The detection limit for pseudoephedrine was found to be in the sub-femtomole range and the semi-quantitative nature of the technique was tentatively demonstrated for a metabolite, fructose, by using a homologous internal standard, sucrose. A potential application of AP IR-MALDI for intestinal permeability studies was also explored using polyethylene glycol.     

Placental gas exchange in the guinea-pig: fetal blood gas tensions following the reduction of maternal oxygen capacity with carbon monoxide

The effect of CO hypoxia on the placental exchange of respiratory gases was studied in anaesthetized pregnant guinea-pigs near term. Fetal PO2 and PCO2 were measured by mass spectrometry from a blood gas catheter in the right atrium. Administration of 5 ml CO over 65 s reduced maternal oxygen capacity by 26%. There was a rapid fall in fetal arterial PO2 and a more gradual rise in fetal PCO2. It was shown in separate experiments that the carboxyhaemoglobin content of fetal blood did not alter greatly in the first few min. after CO administration, which is the interval within which fetal PO2 was seen to fall. The alteration in fetal gas tensions can therefore be ascribed to the increased oxygen affinity and reduced oxygen capacity occasioned by the presence of carboxyhaemoglobin in the maternal blood. The alteration in placental oxygen transfer was calculated from the experimental findings, using a mathematical model of placental gas exchange in the guinea-pig. The total reduction in the oxygen transfer was 32% of the initial value. It was calculated that the reduction in maternal oxygen capacity was responsible for about two-thirds of this decrease, the remainder being due to the increased oxygen affinity of maternal blood.     

Detection of volatile metabolites produced by bacterial growth in blood culture media by selected ion flow tube mass spectrometry (SIFT-MS)

To achieve faster bacteremia diagnosis, selected ion flow tube mass spectrometry (SIFT-MS) measured metabolic gases in the headspaces of BacT/ALERT blood culture bottles. Pseudomonas aeruginosa, Streptococcus pneumoniae, Escherichia coli, Staphylococcus aureus and Neisseria meningitidis growth and trace gas patterns were detected from 10 colony forming units after 6 h.