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The doubly labeled water (DLW) method is the gold standard methodology for determination of free-living, total daily energy expenditure (TEE). However, there is no single accepted approach for either the sampling protocols (daily vs. two-point, in which samples are collected after dosing and at the end of the measurement period) or the calculations used in the determination of the rate of carbon dioxide production (rCO2) and TEE. Moreover, fluctuations in natural background abundances introduce error in the calculation of rCO2 and TEE. The advent of new technologies makes feasible the possibility of including additional isotope measures (17O) to account for background variation, which may improve accuracy. Sixteen subjects were studied for 7 consecutive days in a whole-room indirect calorimeter (IC) with concurrent measurement of TEE by DLW. Daily urine samples were obtained and isotope ratios were determined using off-axis integrated cavity output spectroscopy (OA-ICOS). We determined the best combination of approaches for estimating dilution spaces and elimination rates and calculated average daily volume of carbon dioxide production (VCO2) using six different published equations. Using this best combination, multi-point fitting of isotope elimination rates using the daily urine samples substantially improved the average precision (4.5% vs. 6.0%) and accuracy (−0.5% vs. −3.0%) compared with the two-point method. This improvement may partly reflect the less variable day-to-day chamber measurements of energy expenditure. Utilizing 17O measurements to correct for errors due to background isotope fluctuations provided additional but minor improvements in precision (4.2% vs. 4.5%) and accuracy (0.2% vs. 0.5%). This work shows that optimizing sampling and calculation protocols can improve the accuracy and precision of DLW measurements.

Summary The doubly labeled water (DLW) method measures total energy expenditure (TEE) in free-living subjects. Several equations are used to convert isotopic data into TEE. Using the International Atomic Energy Agency (IAEA) DLW database (5,756 measurements of adults and children), we show considerable variability is introduced by different equations. The estimated rCO2 is sensitive to the dilution space ratio (DSR) of the two isotopes. Based on performance in validation studies, we propose a new equation based on a new estimate of the mean DSR. The DSR is lower at low body masses (, Graphical Abstract, Highlights 5,756 doubly labeled water (DLW) measures highlight variation from calculation equation We derive here new equations for calculating CO2 production when using DLW These equations outperform previous equations in validation studies We recommend these equations should be adopted in future studies using DLW in humans, Speakman et al. use a large database of doubly labeled water measurements to show the choice of equation for the calculation of energy expenditure introduces significant variation into the final estimate. They then derive new equations that outperform previous equations in validation studies against chamber calorimetry.

When the doubly labeled water (DLW) method is used to measure total daily energy expenditure (TDEE), isotope measurements are typically performed using isotope ratio mass spectrometry (IRMS). New technologies, such as off-axis integrated cavity output spectroscopy (OA-ICOS) provide comparable isotopic measurements of standard waters and human urine samples, but the accuracy of carbon dioxide production (V̇co2) determined with OA-ICOS has not been demonstrated. We compared simultaneous measurement V̇co2 obtained using whole-room indirect calorimetry (IC) with DLW-based measurements from IRMS and OA-ICOS. Seventeen subjects (10 female; 22 to 63 yr) were studied for 7 consecutive days in the IC. Subjects consumed a dose of 0.25 g H218O (98% APE) and 0.14 g 2H2O (99.8% APE) per kilogram of total body water, and urine samples were obtained on days 1 and 8 to measure average daily V̇co2 using OA-ICOS and IRMS. V̇co2 was calculated using both the plateau and intercept methods. There were no differences in V̇co2 measured by OA-ICOS or IRMS compared with IC when the plateau method was used. When the intercept method was used, V̇co2 using OA-ICOS did not differ from IC, but V̇co2 measured using IRMS was significantly lower than IC. Accuracy (~1–5%), precision (~8%), intraclass correlation coefficients ( R = 0.87–90), and root mean squared error (30–40 liters/day) of V̇co2 measured by OA-ICOS and IRMS were similar. Both OA-ICOS and IRMS produced measurements of V̇co2 with comparable accuracy and precision compared with IC.

Background The doubly-labeled water (DLW) method is the gold standard methodology for determination of free-living, total daily energy expenditure (TEE). However, there is no single accepted approach for either the sampling protocols (daily vs. two-point, in which samples are collected after dosing and at the end of the measurement period) or the calculations used in the determination of the rate of carbon dioxide production (rCO2) and TEE. Moreover, fluctuations in natural background abundances introduce error in the calculation of rCO2 and TEE. The advent of new technologies makes feasible the possibility of including additional isotope measures (17O) to account for background variation, which may improve accuracy. Methods Sixteen subjects were studied for 7 consecutive days in a whole-room indirect calorimeter (IC) with concurrent measurement of TEE by DLW. Daily urine samples were obtained and isotope ratios determined using off-axis integrated cavity output spectroscopy (OA-ICOS). Results We determined the best combination of approaches for estimating dilution spaces, elimination rates, and calculated average daily carbon dioxide production (VCO2) using six different published equations. Using the best combination, multi-point fitting of isotope elimination rates using the daily urine samples substantially improved the average precision (4.5% vs. 6.0%) and accuracy (−0.5% vs. −3.0%) compared with the two-point method. This improvement may partly reflect the less variable day-to-day chamber measurements of energy expenditure. Utilizing 17O measurements to correct for errors due to background isotope fluctuations provided additional but minor improvements in precision (4.2% vs. 4.5%) and accuracy (0.2% vs. 0.5%). Conclusions This work shows that optimizing sampling and calculation protocols can improve the accuracy and precision of DLW measurements.

[1] Measurements of δ2H and δ18O in isotope-based field studies have fundamentally improved our understanding of water flow and transport time scales in soils and headwater catchments. Until recently, however, technical constraints have limited the temporal resolution at which water samples could be collected and analyzed. We introduce a new sample acquisition system—consisting of a four-channel peristaltic pump, custom flow manifold, and CTC LCPAL auto-sampler—that is paired with a field-deployable laser spectrometer (LGR LWIA). Our system enables high-frequency (subhourly) measurement of δ2H and δ18O in as many as four water sources. We deployed the system at a field site in Corvallis, OR, USA, where we measured the δ2H and δ18O composition of precipitation and the drainage from two lysimeters. The system produced δ2H and δ18O time series for precipitation and drainage from each lysimeter at a temporal frequency of one sample every 34 min, which, on average, corresponded to 0.84, 0.63, and 0.48 mm of precipitation or lysimeter drainage per sample. The high-frequency data showed substantially greater short-term variability than observed when sampling at successively longer time intervals. The system and sampling configuration are versatile and can be adapted to sample multiple water flows at variable frequencies depending on the characteristic transit times of each source.

A compact, lightweight atmospheric gas analyzer has been integrated into and flown on the National Aeronautics and Space Administration (NASA) Sensor Integrated Environmental Remote Research Aircraft (SIERRA) unmanned aerial vehicle (UAV) and deployed to make highly accurate, 1 Hz measurements of methane, carbon dioxide, and water vapor. The analyzer was used to measure gas concentrations in flight and to demonstrate the system for providing measurements at altitudes as low as 10 m and in remote locations. The first flights were conducted at Crows Landing, CA, an agricultural site, with H 2 O concentrations showing distinct structure and sharp features that were well outside of the measurement noise. The instrument was then deployed in Svalbard, Norway prior to the NASA Characterization of Arctic Sea Ice Experiment (CASIE). During the Svalbard flight, there was minimal variation in the CO 2 and CH 4 concentrations, but the water concentration changed dramatically, oscillating as the aircraft moved repeatedly through its racetrack shaped flight pattern. The regions of high water concentration corresponded to low-lying areas which collect runoff from the nearby Vestre Broggerbreen glacier. This novel, integrated instrument-aircraft system allows more numerous and efficient measurements of carbon dioxide, methane, and water vapor concentrations at low-altitudes and in remote or dangerous locations.

Radiation-induced degradation of polymeric materials occurs through numerous, simultaneous, competing chemical reactions. Although degradation is typically found to be linear in adsorbed dose, some silicone materials exhibit nonlinear dose dependence due to dose-dependent dominant degradation pathways. We have characterized the effects of radiative and thermal degradation on a model filled-PDMS system, Sylgard 184 (commonly used in electronic encapsulation and in biomedical applications), using traditional mechanical testing, NMR spectroscopy, and sample headspace analysis using solid-phase microextraction (SPME) followed by gas chromatography/mass spectrometry (GC/MS). The mechanical data and (1)H spin-echo NMR spectra indicated that radiation exposure leads to predominantly cross-linking over the cumulative dose range studied (0-250 kGy) with a rate roughly linear with dose. (1)H multiple-quantum NMR spectroscopy detected a bimodal distribution in the network structure, as expected from the proposed structure of Sylgard 184. The MQ NMR spectra further indicated that the radiation-induced structural changes were not linear in adsorbed dose and that competing chain scission mechanisms made a greater contribution to the overall degradation process in the range of 50-100 kGy (although cross-linking still dominated). The SPME-GC/MS data were analyzed using principal component analysis (PCA), which identified subtle changes in the distributions of degradation products (the cyclic siloxanes and other components of the material) as a function of age that provide insight into the dominant degradation pathways at low and high adsorbed dose.

Characterizing and classifying molecular variations within biological samples are critical for determining the fundamental mechanisms of biological processes. Toward these ends, time-of-flight secondary ion mass spectrometry (ToF-SIMS) was used to examine increasingly complex samples of biological relevance. The large, multivariate datasets were analyzed using five commonstatisticalandchemometrictechniques:principalcomponentanalysis(PCA),lineardiscriminantanalysis(LDA),partial least-squaresdiscriminantanalysis(PLSDA),softindependentmodelingofclassanalogy(SIMCA),anddecision-treeanalysisby recursivepartitioning.PCAwasfoundtoprovideinsightintoboththerelativegroupingsofsamplesandthemolecularbasisfor thosegroupings.Formonosaccharide,pureprotein,andcomplexproteinmixturesamples,LDA,PLSDA,andSIMCAallproduced excellent classification. For mouse embryo tissues, however, SIMCA did not classify samples as accurately. The decision-tree analysis was the least successful for all tested samples, providing neither as accurate a classification nor chemical insight. Based on these results we conclude that as the complexity of samples increases, so must the sophistication of the multivariate technique used to classify the samples. PCA is a preferred first step for understanding ToF-SIMS data that can be followed by either LDA or PLSDA for effective classification. This study demonstrates the strength of the combination of ToF-SIMS and multivariate analysis to classify increasingly complex biological samples. Applying these techniques to information-rich mass spectraldataopensthepossibilitiesfornewapplicationsincludingclassificationofsubtlydifferentbiologicalsamplesthatmay provide insights into cellular processes, disease progress, and disease diagnosis. Copyright c � 2008 John Wiley & Sons, Ltd.

Time-of-flight secondary ion mass spectrometry (ToF-SIMS) equipped with a gold ion gun was used to image mouse embryo sections and differentiate tissue types (brain, spinal cord, skull, rib, heart and liver). Embryos were paraffin-embedded and then deparaffinized. The robustness and repeatability of the method was determined by analyzing ten tissue slices from three different embryos over a period of several weeks. Using principal component analysis (PCA) to reduce the spectral data generated by ToF-SIMS, histopathologically identified tissue types of the mouse embryos can be differentiated based on the characteristic differences in their mass spectra. These results demonstrate the ability of ToF-SIMS to determine subtle chemical differences even in fixed histological specimens.

© 2014 Elsevier Ltd. Field-scale biostimulation and desorption tracer experiments conducted in a uranium (U) contaminated, shallow alluvial aquifer have provided insight into the coupling of microbiology, biogeochemistry, and hydrogeology that control U mobility in the subsurface. Initial experiments successfully tested the concept that Fe-reducing bacteria such as Geobacter sp. could enzymatically reduce soluble U(VI) to insoluble U(IV) during in situ electron donor amendment (Anderson et al., 2003; Williams et al., 2011). In parallel, in situ desorption tracer tests using bicarbonate amendment demonstrated rate-limited U(VI) desorption (Fox et al., 2012). These results and prior laboratory studies underscored the importance of enzymatic U(VI)-reduction and suggested the ability to combine desorption and bioreduction of U(VI). Here we report the results of a new field experiment in which bicarbonate-promoted uranium desorption and acetate amendment were combined and compared to an acetate amendment-only experiment in the same experimental plot. Results confirm that bicarbonate amendment to alluvial aquifer sediments desorbs U(VI) and increases the abundance of Ca-uranyl-carbonato complexes. At the same time, the rate of acetate-promoted enzymatic U(VI) reduction was greater in the presence of added bicarbonate in spite of the increased dominance of Ca-uranyl-carbonato aqueous complexes. A model-simulated peak rate of U(VI) reduction was ~3.8 times higher during acetate-bicarbonate treatment than under acetate-only conditions. Lack of consistent differences in microbial community structure between acetate-bicarbonate and acetate-only treatments suggest that a significantly higher rate of U(VI) reduction in the bicarbonate-impacted sediment may be due to a higher intrinsic rate of microbial reduction induced by elevated concentrations of the bicarbonate oxyanion. The findings indicate that bicarbonate amendment may be useful in improving the engineered bioremediation of uranium in aquifers.

We use time-of-flight secondary ion mass spectrometry (TOF-SIMS) to image and classify individual cells on the basis of their characteristic mass spectra. Using statistical data reduction on the large data sets generated during TOF-SIMS analysis, similar biological materials can be differentiated on the basis of a combination of small changes in protein expression, metabolic activity and cell structure. We apply this powerful technique to image and differentiate three carcinoma-derived human breast cancer cell lines (MCF-7, T47D, and MDA-MB-231). In homogenized cells, we show the ability to differentiate the cell types as well as cellular compartments (cytosol, nuclear, and membrane). These studies illustrate the capacity of TOF-SIMS to characterize individual cells by chemical composition, which could ultimately be applied to detect and identify single aberrant cells within a normal cell population. Ultimately, we anticipate characterizing rare chemical changes that may provide clues to single cell progression within carcinogenic and metastatic pathways.

The method of choice for measuring total energy expenditure in free-living individuals is the doubly labeled water (DLW) method. This experiment examined the behavior of natural background isotope abundance fluctuations within and between individuals over time to assess possible methods of accounting for variations in the background isotope abundances to potentially improve the precision of the DLW measurement.In this work, we measured natural background variations in (2)H, (18)O and (17)O in water from urine samples collected from 40 human subjects who resided in the same geographical area. Each subject provided a urine sample for 30 consecutive days. Isotopic abundances in the samples were measured using Off-Axis Integrated Cavity Output Spectroscopy.Autocorrelation analyses demonstrated that the background isotopes in a given individual were not temporally correlated over the time scales of typical DLW studies. Using samples obtained from different individuals on the same calendar day, cross-correlation analyses demonstrated that the background variations of different individuals were not correlated in time. However, the measured ratios of the three isotopes (2)H, (18)O and (17)O were highly correlated (R(2)=0.89-0.96).Although neither specific timing of DLW water studies nor intraindividual comparisons were found to be avenues for reducing the impact of background isotope abundance fluctuations on DLW studies, strong inter-isotope correlations within an individual confirm that use of a dosing ratio of 8‰:1‰ (0.6 p.p.m.: p.p.m.) optimizes DLW precision. Theoretical implications for the possible use of (17)O measurements within a DLW study require further study.

Stable isotopes of water have long been used to improve understanding of the hydrological cycle, catchment hydrology, and polar climate. Recently, there has been increasing interest in measurement and use of the less-abundant (17)O isotope in addition to (2)H and (18)O. Off-axis integrated cavity output spectroscopy (OA-ICOS) is demonstrated for accurate and precise measurements δ(18)O, δ(17)O, and (17)O-excess in liquid water. OA-ICOS involves no sample conversion and has a small footprint, allowing measurements to be made by researchers collecting the samples. Repeated (514) high-throughput measurements of the international isotopic reference water standard Greenland Ice Sheet Precipitation (GISP) demonstrate the precision and accuracy of OA-ICOS: δ(18)OVSMOW-SLAP = -24.74 ± 0.07‰ (1σ) and δ(17)OVSMOW-SLAP = -13.12 ± 0.05‰ (1σ). For comparison, the International Atomic Energy Agency (IAEA) value for δ(18)OVSMOW-SLAP is -24.76 ± 0.09‰ (1σ) and an average of previously reported values for δ(17)OVSMOW-SLAP is -13.12 ± 0.06‰ (1σ). Multiple (26) high-precision measurements of GISP provide a (17)O-excessVSMOW-SLAP of 23 ± 10 per meg (1σ); an average of previously reported values for (17)O-excessVSMOW-SLAP is 22 ± 11 per meg (1σ). For all these OA-ICOS measurements, precision can be further enhanced by additional averaging. OA-ICOS measurements were compared with two independent isotope ratio mass spectrometry (IRMS) laboratories and shown to have comparable accuracy and precision as the current fluorination-IRMS techniques in δ(18)O, δ(17)O, and (17)O-excess. The ability to measure accurately δ(18)O, δ(17)O, and (17)O-excess in liquid water inexpensively and without sample conversion is expected to increase vastly the application of δ(17)O and (17)O-excess measurements for scientific understanding of the water cycle, atmospheric convection, and climate modeling among others.

Manish Gupta,1 J Brian Leen,1 Elena Simone Franklin Berman,1 Aldo Ciambotti2 1Los Gatos Research, Mountain View, CA, USA; 2Consiglio per la Ricerca e la Sperimentazione in Agricoltura, Centro di Ricerca per l'Enologia, Asti, Italy Abstract: Wine counterfeiting is an international, multi-billion dollar issue, with some estimates suggesting that up to 5% of wines sold at auctions or secondary markets are fraudulent. Isotope ratio mass spectrometer (IRMS) measurements of the 18O/16O stable isotope ratio (δ18O) of water-in-wine have been used for wine authentication; however, these analyses are time-consuming and costly. In this preliminary study, off-axis integrated cavity output spectroscopy (OA-ICOS) is used to quantify δ18O in wines. This laser-based method has been extensively used to study water isotopes for hydrological and medical applications. Recently, the development of a spectral contaminant identifier (SCI) has extended the application of these OA-ICOS analyzers to contaminated water samples (eg, plant, soil, and leaf waters). Here, we utilize OA-ICOS with the SCI to characterize wine samples (9%–15% ethanol), and show that the laser-based instrument provides a δ18O measurement precision of ±0.07‰ (1σ) and agrees with IRMS to within ±0.63‰ (1σ). Moreover, by training the SCI on isotopically-characterized wines, the agreement with IRMS improves to within ±0.30‰ (1σ). The utility of the instrument is demonstrated by measuring watered and mixed wines. The method presented here can be readily extended to address other food authentication applications. Keywords: wine isotopes, wine fraud, counterfeit wines, OA-ICOS

Developments in cavity-enhanced absorption spectrometry have made it possible to measure water isotopes using faster, more cost-effective field-deployable instrumentation. Several groups have attempted to extend this technology to measure water extracted from plants and found that other extracted organics absorb light at frequencies similar to that absorbed by the water isotopomers, leading to δ(2)H and δ(18)O measurement errors (Δδ(2)H and Δδ(18)O). In this note, the off-axis integrated cavity output spectroscopy (ICOS) spectra of stable isotopes in liquid water is analyzed to determine the presence of interfering absorbers that lead to erroneous isotope measurements. The baseline offset of the spectra is used to calculate a broadband spectral metric, m(BB), and the mean subtracted fit residuals in two regions of interest are used to determine a narrowband metric, m(NB). These metrics are used to correct for Δδ(2)H and Δδ(18)O. The method was tested on 14 instruments and Δδ(18)O was found to scale linearly with contaminant concentration for both narrowband (e.g., methanol) and broadband (e.g., ethanol) absorbers, while Δδ(2)H scaled linearly with narrowband and as a polynomial with broadband absorbers. Additionally, the isotope errors scaled logarithmically with m(NB). Using the isotope error versus m(NB) and m(BB) curves, Δδ(2)H and Δδ(18)O resulting from methanol contamination were corrected to a maximum mean absolute error of 0.93 [per thousand] and 0.25 [per thousand] respectively, while Δδ(2)H and Δδ(18)O from ethanol contamination were corrected to a maximum mean absolute error of 1.22 [per thousand] and 0.22 [per thousand]. Large variation between instruments indicates that the sensitivities must be calibrated for each individual isotope analyzer. These results suggest that the properly calibrated interference metrics can be used to correct for polluted samples and extend off-axis ICOS measurements of liquid water to include plant waters, soil extracts, wastewater, and alcoholic beverages. The general technique may also be extended to other laser-based analyzers including methane and carbon dioxide isotope sensors.

International audience; The stable isotopes of hydrogen (δ(2)H) and oxygen (δ(18)O) in human urine are measured during studies of total energy expenditure by the doubly labeled water method, measurement of total body water, and measurement of insulin resistance by glucose disposal among other applications. An ultrasensitive laser absorption spectrometer based on off-axis integrated cavity output spectroscopy was demonstrated for simple and inexpensive measurement of stable isotopes in natural isotopic abundance and isotopically enriched human urine. Preparation of urine for analysis was simple and rapid (approximately 25 samples per hour), requiring no decolorizing or distillation steps. Analysis schemes were demonstrated to address sample-to-sample memory while still allowing analysis of 45 natural or 30 enriched urine samples per day. The instrument was linear over a wide range of water isotopes (δ(2)H = -454 to +1702 ‰ and δ(18)O = -58.3 to +265 ‰). Measurements of human urine were precise to better than 0.65 ‰ 1σ for δ(2)H and 0.09 ‰ 1σ for δ(18)O for natural urines, 1.1 ‰ 1σ for δ(2)H and 0.13 ‰ 1σ for δ(18)O for low enriched urines, and 1.0 ‰ 1σ for δ(2)H and 0.08 ‰ 1σ for δ(18)O for high enriched urines. Furthermore, the accuracy of the isotope measurements of human urines was verified to better than ±0.81 ‰ in δ(2)H and ±0.13 ‰ in δ(18)O (average deviation) against three independent isotope-ratio mass spectrometry laboratories. The ability to immediately and inexpensively measure the stable isotopes of water in human urine is expected to increase the number and variety of experiments which can be undertaken.

Characterizing the molecular contents of individual cells is critical for understanding fundamental mechanisms of biological processes. Imaging mass spectrometry (IMS) of biological systems has been steadily gaining popularity for its ability to create precise chemical images of biological samples, thereby revealing new biological insights and improving understanding of disease. In order to acquire mass spectral images from single cells that contain relevant molecular information, samples must be prepared such that cell-culture components, especially salts, are eliminated from the cell surface and that the cell contents are accessible to the mass spectrometer. We have demonstrated a cellular preparation technique for IMS that preserves the basic morphology of cultured cells, allows mass spectrometric chemical profiling of cytosol, and removes the majority of the interfering species derived from the cellular growth medium. Using this protocol, we achieve high-quality, reproducible IMS images from three diverse cell types: MCF7 human breast cancer cells, Madin-Darby canine kidney (MDCK) cells, and NIH/3T3 mouse fibroblasts. This preparation method allows rapid and routine IMS analysis of cultured cells, making possible a wide variety of experiments to further scientific understanding of molecular processes within individual cells.

Characterizing the molecular contents of individual cells is critical for understanding fundamental mechanisms of biological processes. Imaging mass spectrometry (IMS) of biological systems has been steadily gaining popularity for its ability to create precise chemical images of biological samples, thereby revealing new biological insights and improving understanding of disease. In order to acquire mass spectral images from single cells that contain relevant molecular information, samples must be prepared such that cell-culture components, especially salts, are eliminated from the cell surface and that the cell contents are accessible to the mass spectrometer. We have demonstrated a cellular preparation technique for IMS that preserves the basic morphology of cultured cells, allows mass spectrometric chemical profiling of cytosol, and removes the majority of the interfering species derived from the cellular growth medium. Using this protocol, we achieve high-quality, reproducible IMS images from three diverse cell types: MCF7 human breast cancer cells, Madin-Darby canine kidney (MDCK) cells, and NIH/3T3 mouse fibroblasts. This preparation method allows rapid and routine IMS analysis of cultured cells, making possible a wide variety of experiments to further scientific understanding of molecular processes within individual cells.

[1] A high-frequency, field-deployable liquid water isotope analyzer was developed that is capable of quantifying δ18O and δ2H to better than ±0.17 and ±0.32‰, respectively, on over 90 samples/d. The instrument was deployed for 4 contiguous weeks in the H. J. Andrews Experimental Forest Long-term Ecological Research site in western Oregon, where it was used for real-time measurement of the isotope ratios of precipitation and stream water during three large storm events. We were able to document fine-scale changes in rainfall composition and damping effects in the stream channel continuously through these periods. We also performed a rain-on-snow experiment where we sampled leachate from a melting snow core continuously at 2 min intervals for 5 h. These data show remarkable fine-scale patterns of internal rain-snow mixing, patterns that would not have been detected without such high-frequency sampling. These two preliminary applications show proof of concept of the new field analyzer, a device that will ultimately provide hydrologists with insight into water flow dynamics with unprecedented frequency over long time scales.

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) is utilized to examine the mass spectra and fragmentation patterns of seven isomeric monosaccharides. Multivariate statistical analysis techniques, including principal component analysis (PCA), allow discrimination of the extremely similar mass spectra of stereoisomers. Furthermore, PCA identifies those fragment peaks that vary significantly between spectra. Heavy isotope studies confirm that these peaks are indeed sugar fragments, allow identification of the fragments, and provide clues to the fragmentation pathways. Excellent reproducibility is shown by multiple experiments performed over time and on separate samples. This study demonstrates the combined selectivity and discrimination power of TOF-SIMS and PCA and suggests new applications of the technique including differentiation of subtle chemical changes in biological samples that may provide insights into cellular processes, disease progress, and disease diagnosis.

The complex Ru(dipa)(2)(2+) (dipa = di-2-pyridylmethanamine) has been prepared, yielding approximately a statistical ratio of the meso and rac isomers. The electronic spectra of both isomers show pyridyl pi --pi transitions in the UV region and MLCT bands in the visible region. The solvent dependence of the spectra provides evidence of hydrogen bond formation between the solvent and the NH(2) site on the ligand. The electrochemical properties of the two isomers are identical; each undergoes a reversible one-electron oxidation in acetonitrile (E(1/2) = 0.933 V vs Ag/AgCl) and in aqueous solution below pH 3 (E(1/2) = 0.786 V vs Ag/AgCl). In aqueous solution above pH 3, one-electron oxidation of the ruthenium center is followed by deprotonation of the ligand NH(2) site yielding a reactive amidoruthenium(III) species. The ruthenium-bound dipa ligand possesses structural constraints that prevent the usual oxidative dehydrogenation reaction, which would yield exclusively the corresponding imine. Instead the amidoruthenium(III) intermediate finds alternative reaction routes leading to multiple products.

Cavity ring-down spectroscopy (CRDS) is an extremely sensitive absorption technique that has been applied primarily to gas samples, which are characterized by having narrow absorption features. Recently, CRDS has also been applied to liquid samples, which have broad absorption features. The use of small inexpensive diode lasers as light sources for liquid samples is demonstrated. The low cost coupled with the ease and technical straightforwardness of application gives this technique wide appeal.

The kinetics of methylene blue reduction by ascorbic acid in acetonitrile was investigated by cavity ring-down spectroscopy. Because of our high sensitivity we were able to use very low concentrations (1−10 nM) of the dye. Under these conditions, we observed a second-order loss of dye as well as a competing back reaction with dissolved oxygen. The use of an inexpensive diode laser and a relatively simple setup should make ultratrace kinetic studies more accessible.

Cavity ring-down spectroscopy is applied to the liquid phase by placing the target solution directly into the optical cavity. We demonstrate that solutions in the cavity can be stirred and more importantly monitored in a flow. We report a minimum detectable absorption of 10(-6) cm(-1) for a range of organic solvents. This detection limit corresponds to picomolar concentrations for strong absorbers.

Characterizing chemical changes within individual cells is important for determining fundamental mechanisms of biological processes that will lead to new biological insights and improved disease understanding. Analyzing biological systems with imaging and profiling mass spectrometry (MS) has gained popularity in recent years as a method for creating chemical maps of biological samples. To obtain mass spectra that provide relevant molecular information about individual cells, samples must be prepared so that salts and other cell culture components are removed from the cell surface and that the cell contents are rendered accessible to the desorption beam. We have designed a cellular preparation protocol for imaging/profiling MS that removes the majority of the interfering species derived from the cellular growth medium, preserves the basic morphology of the cells, and allows chemical profiling of the diffusible elements of the cytosol. Using this method, we are able to reproducibly analyze cells from three diverse cell types: MCF7 human breast cancer cells, Madin-Darby canine kidney (MDCK) cells, and NIH/3T3 mouse fibroblasts. This preparation technique makes possible routine imaging/profiling MS analysis of individual cultured cells, allowing for understanding of molecular processes within individual cells.