We would like to thank the Ministry of Science and Technology of Taiwan (grant number: MOST 104-2628-M-009-003-MY4) for the financial support of this work.

This protocol assumes that all the steps are performed according to the relevant laboratory safety regulations. Some of the steps use commercial instruments - in those cases, manufacturer guidelines need to be followed. When handling toxic chemicals, MSDS guidelines need to be followed. The custom-made equipment4 must be operated cautiously; especially, when handling pressurized gases and live electric wiring.
1. Preparation of Standard Solution
<ol>
	<li>Prepare 6.2 x 10-2 M stock solution of limonene in ethanol by mixing 10 &#181;L limonene with 990 &#181;L ethanol.</li>
	<li>Prepare 10 mL of 6.2 x 10-5 M limonene solution by mixing 10 &#181;L 6.2 x 10-2 limonene, 490 &#181;L ethanol, and adding pure water to the final volume of 10 mL. Shake the volumetric flask thoroughly.</li>
	<li>Transfer the prepared standard solution to a 20-mL screw top headspace glass vial with septum cap. The diluted standard solution can be used for testing the system.</li>
</ol>
2. Preparation of Real Sample
<ol>
	<li>Obtain lime juice by squeezing fresh lime fruit (cut in half) on a kitchen squeezer.</li>
	<li>Prepare 10 mL of diluted lime juice by mixing 2 mL lime juice, 500 &#181;L ethanol, and adding pure water to the final volume of 10 mL. Shake the volumetric flask thoroughly.</li>
	<li>Transfer the prepared sample to a 20-mL screw top headspace glass vial with septum cap.</li>
</ol>
3. Spiking the Real Sample with Standard Solution
<ol>
	<li>First standard addition: Prepare 10 mL of spiked sample by mixing 2 mL lime juice, 10 &#181;L 6.2 x 10-2 M limonene solution, 490 &#181;L ethanol, and adding pure water to the final volume of 10 mL. Shake the volumetric flask thoroughly.</li>
	<li>Second standard addition: Prepare 10 mL of spiked sample by mixing 2 mL lime juice, 20 &#181;L 6.2 x 10-2 M limonene solution, 480 &#181;L ethanol, and adding pure water to the final volume of 10 mL. Shake the volumetric flask thoroughly.</li>
</ol>
4. Setting Up the Fizzy Extraction System
<ol>
	<li>Put the fizzy extraction system (Figure 1)4 next to the APCI source of the triple quadrupole mass spectrometer.</li>
	<li>Connect the carbon dioxide gas cylinder to the gas supply inlet of the fizzy extraction system. Open the valve in the gas regulator. Set the output pressure to 1.5 bar (150 kPa).</li>
	<li>Connect the extraction chamber outlet to the ion source inlet.</li>
	<li>Connect the fizzy extraction system to the 12-V power supply.</li>
	<li>Set up the data acquisition software of the triple quadrupole mass spectrometer (Figure 2). Operate the instrument with the APCI source, in the positive-ion multiple reaction monitoring (MRM) mode, with argon as collision gas.
	<ol>
		<li>Run the data acquisition software.</li>
		<li>Select the option &#34;LCMS8030 only&#34;.</li>
		<li>Select the option &#34;MS On/Off&#34;.</li>
		<li>Set the desolvation line temperature to 250 &#176;C, and the flow rate of drying gas to 15 L min-1. Wait until the value of every instrument parameter becomes the same as the preset value.</li>
		<li>Select the MS data acquisition method file.</li>
		<li>Make sure the collision voltage is -20 V, the precursor ion m/z is 137, and the fragment ion m/z are 81 and 95</li>
		<li>Click on the &#34;Start Single Run&#34; button.</li>
		<li>Type the file name.</li>
		<li>Select the file path.</li>
		<li>Move to section 5 (&#34;Performing fizzy extraction&#34;).</li>
		<li>Select the option &#34;MS On/Off&#34;.</li>
		<li>Close the software window.</li>
		<li>Tick the items &#34;Nebulizing Gas Off&#34;, &#34;DL Heater Off&#34;, &#34;Heat Block Off&#34;, and &#34;Dry Gas Off&#34;. Click &#34;OK&#34;.</li>
	</ol>
	</li>
</ol>
5. Performing Fizzy Extraction
<ol>
	<li>Place a sample vial in the fizzy extraction system by using the screw mount. The extraction system is operated at room temperature (~ 25 &#176;C).</li>
	<li>Press the &#34;Start&#34; button on the LCD shield of the fizzy extraction system.</li>
	<li>Wait while the automated fizzy extraction process is executed (Figure 3). Observe development of ion signals on the screen of the triple quadrupole mass spectrometer. 
	NOTE: The following steps are executed automatically: Sample headspace is flushed with carbon dioxide during 60 s. Sample is pressurized with carbon dioxide during 60 s. Stirrer motor is on. Sample is depressurized. Multiple bubbles are formed. In the later phase, stirrer motor is on to enhance bubbling.</li>
	<li>Take out (unscrew) the sample vial.</li>
	<li>Wipe the sample stirring spindle with cellulose tissue.</li>
	<li>Wash the stirring spindle with ethanol, and wipe it with cellulose tissue again.</li>
	<li>The system is ready for analysis of another sample (repeat steps 5.1-5.6).</li>
	<li>Switch off the power supply.</li>
	<li>Disconnect the fizzy extraction outlet tube from the ion source.</li>
	<li>Close the valve of the gas cylinder, and disconnect the gas tubing.</li>
</ol>
6. Data Analysis
<ol>
	<li>Export extracted ion currents for the m/z 81 from the triple quadrupole mass spectrometer&#39;s data acquisition software to ASCII files (Figure 4). 
	NOTE: The ion current at the m/z 95 is not used in this demonstration.
	<ol>
		<li>Run the data acquisition software. Select the option &#34;Postrun&#34;.</li>
		<li>Select the option &#34;Select Project (Folder)&#34;, and choose the data file.</li>
		<li>Click on the &#34;File&#34; menu, and select &#34;Export Data&#34; / &#34;Export Data as ASCII&#34;.</li>
		<li>Select &#34;Output File&#34;, and select the file path. Select &#34;MS Chromatogram (MC)&#34;.</li>
	</ol>
	</li>
	<li>Import the raw data sets into peak integration software, and measure peak areas (Figure 5). Settings: linear baseline; HVL function.
	<ol>
		<li>Run the peak integration software.</li>
		<li>Select the option &#34;Import&#34; from the &#34;File&#34; menu. Click &#34;Yes&#34; button.</li>
		<li>Select the data in X and Y column. Click &#34;OK&#34; button. Select the option &#34;AutoFit Peaks I Residuals&#34;.</li>
		<li>Fit the extraction peak semi-automatically. Make sure the fitted curve follows the experimental data points. Select the option &#34;List Peak Estimates&#34;. Select the option &#34;ASCII Editor&#34;.</li>
		<li>Copy the fitting results to &#34;Clipboard&#34;.</li>
	</ol>
	</li>
	<li>Input the measured peak areas into a spreadsheet in data analysis software (Figure 6).
	<ol>
		<li>Run the data analysis software.</li>
		<li>Input the concentration values in X column and peak area values in Y column. Select the option &#34;Symbol&#34; / &#34;Scatter&#34; from the &#34;Plot&#34; menu. Select the option &#34;Fitting&#34; / &#34;Fit Linear&#34; from the &#34;Analysis&#34; menu.</li>
	</ol>
	</li>
	<li>Calculate the concentration of limonene in the diluted real sample based on the formula: 
	<img alt="Equation 1" src="/files/ftp_upload/56008/56008eq1.jpg" /> 
	where I is the intercept of the linear function, while S is the slope.</li>
	<li>Compute the concentration of limonene in the original real sample (before dilution) based on the formula: 
	<img alt="Equation 2" src="/files/ftp_upload/56008/56008eq2.jpg" /> 
	where DF is the dilution factor (here, 5).</li>
</ol>

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The authors have nothing to disclose.

Several smart ways to deliver samples to a mass spectrometer were developed in the studies conducted during the past three decades (e.g., references8,9,10,11,12,13,14). One of the goals of those studies was to simplify preparation of samples for analysis. To achieve that goal, vario...

Various phenomena observed in nature and daily life are linked to gas-liquid phase equilibriums. Carbon dioxide is dissolved in soft and alcoholic drinks under elevated pressure. When a bottle of such a fizzy drink is opened, the pressure drops down, and gas bubbles rush to the liquid surface. In this case, effervescence improves organoleptic properties of beverages. The release of gas bubbles is also the main cause of decompression sickness ("the bends")1. Due to sudden decompression, bubbles form in divers' bodies. The persons suffering from the decompression sickness are treated in hyperbaric chambers.
Gas bubbles have various applications in analytical chemistry. Notably, sparging methods rely on passing gas bubbles through liquid samples to extract volatile compounds2. For example, a method called "purge-closed loop" is combined with gas chromatography to enable rapid analysis of dissolved volatiles3. While sparging can continuously extract volatiles over time, it does not confine them in space or time. The released gas-phase species need to be trapped, and-in some cases-concentrated by applying a temperature program or using sorbents. Thus, there is a need to introduce new on-line sample treatment strategies, which could reduce the number of steps, and-at the same time-concentrate volatile analytes in space or time.
To address the challenge of extracting volatile compounds from liquid samples, and performing analysis on-line, we recently introduced "fizzy extraction"4. This new technique takes advantage of the effervescence phenomenon. Briefly, a carrier gas (here, carbon dioxide) is first dissolved in the sample by applying overpressure and stirring the sample. Then, the sample chamber is decompressed abruptly. The sudden decompression leads to formation of numerous carrier gas bubbles in the sample liquid. These bubbles assist the release of dissolved analyte species from the liquid to the gas phase. The released analytes are immediately transferred to the mass spectrometer, producing signals in the time domain. Because the release of the analyte species is confined to a short period of time (a few seconds), the temporal signals have high amplitudes and high signal-to-noise ratios.
The pressures involved in the fizzy extraction process are very low (~150 kPa)4; much lower than in supercritical fluid extraction5 (e.g., &#8805;10 MPa). The technique does not require the use of any special consumable items (columns, cartridges). Only small volumes of solvents are used for dilution and cleaning. The extraction device can be assembled by chemists with medium technical skills using widely available parts4; for example, open-source electronic modules6,7. Fizzy extraction can be coupled on-line with modern mass spectrometers equipped with atmospheric pressure chemical ionization (APCI) interface. Because gas-phase extracts are transferred to the ion source, operation of fizzy extraction does not substantially contaminate vulnerable parts of the mass spectrometer.
The purpose of this visualized experiment article is to guide the viewers on how to implement fizzy extraction in a simple analytical task. While the core of the fizzy extraction system is as described in our previous report4, several improvements have been introduced to make the operation more straightforward. A microcontroller equipped with an LCD screen shield has been incorporated into the system to display the key extraction parameters in real time. All the functions are programmed in the microcontroller scripts, and there is no longer a need to use an external computer to control the extraction system.

<table><tbody><tr><td>Water</td><td>Fisher</td><td>W6212</td><td>Diluent</td></tr><tr><td>Ethanol</td><td>Sigma-Aldrich</td><td>32221-2.5L</td><td>Diluent</td></tr><tr><td>(R)-(+)-Limonene</td><td>Sigma-Aldrich</td><td>183164-100ML</td><td>Standard</td></tr><tr><td>Carbon dioxide</td><td>ChiaLung</td><td>n/a</td><td>Carrier gas</td></tr><tr><td>Cellulose tissue, Kimwipes Kimtech</td><td>Kimberly-Clark</td><td>34120</td><td>Used for cleaning</td></tr><tr><td>Triple quadrupole mass spectrometer</td><td>Shimadzu</td><td>LCMS-8030</td><td>Detection system</td></tr><tr><td>Atmospheric pressure chemical ionization interface</td><td>Shimadzu</td><td>Duis</td><td>Ion source</td></tr><tr><td>20-mL screw top headspace glass vial with septum cap</td><td>Thermo Fisher Scientific</td><td>D-52379</td><td>Sample vial</td></tr><tr><td>LabSolutions software</td><td>Shimadzu</td><td>n/a</td><td>version 5.82</td></tr><tr><td>PeakFit software</td><td>Systat Software</td><td>n/a</td><td>version 4.12</td></tr><tr><td>OriginPro software</td><td>OriginLab</td><td>n/a</td><td>version 8</td></tr></tbody></table>

fizzy extraction of volatile organic compounds combined with atmospheric pressure chemical ionization quadrupole mass spectrometry

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to the gas phase, and efficiently transferred to a detection system. Fizzy extraction takes advantage of the effervescence phenomenon. First, a carrier gas (here, carbon dioxide) is dissolved in the sample by applying overpressure and stirring the sample. Second, the sample chamber is decompressed abruptly. Decompression leads to the formation of numerous carrier gas bubbles in the sample liquid. These bubbles assist the release of the dissolved analyte species from the liquid to the gas phase. The released analytes are immediately transferred to the atmospheric pressure chemical ionization interface of a triple quadrupole mass spectrometer. The ionizable analyte species give rise to mass spectrometric signals in the time domain. Because the release of the analyte species occurs over short periods of time (a few seconds), the temporal signals have high amplitudes and high signal-to-noise ratios. The amplitudes and areas of the temporal peaks can then be correlated with concentrations of the analytes in the liquid samples subjected to fizzy extraction, which enables quantitative analysis. The advantages of fizzy extraction include: simplicity, speed, and limited use of chemicals (solvents).

At the beginning, the fizzy extraction system is tested with a standard solution. Subsequently, the real sample and real sample spiked with standard are analyzed. The areas of the temporal peaks of extraction events are correlated with concentrations of the analytes in the liquid samples subjected to fizzy extraction, which enables quantitative analysis. Here, we performed double standard addition to demonstrate quantitative capabilities of the technique (Figure 7</st...

Watch this Scientific Journal Video about Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry at JoVE.com

Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry

Fizzy extraction is a new laboratory technique for analysis of volatile and semivolatile compounds. A carrier gas is dissolved in the liquid sample by applying overpressure and stirring the sample. The sample chamber is then decompressed. The analyte species are liberated to the gas phase due to effervescence.

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to ...

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7.6K Views.  National Chiao Tung University. Fizzy extraction is a new laboratory technique for analysis of volatile and semivolatile compounds. A carrier gas is dissolved in the liquid sample by applying overpressure and stirring the sample. The sample chamber is then decompressed. The analyte species are liberated to the gas phase due to effervescence.

Video: Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry

Characterizing Bacterial Volatiles using Secondary Electrospray Ionization Mass Spectrometry (SESI-MS)

Secondary electrospray ionization mass spectrometry (SESI-MS) is a method developed for the rapid detection of volatile compounds, without the need for sample pretreatment. The method was first described by Fenn and colleagues1 and has been applied to the detection of illicit drugs2 and explosives3-4, the characterization of skin volatiles5, and the analysis of breath6-7.
SESI ionization occurs by proton transfer reactions between the electrospray solution and the volatile analyte, and is therefore suitable for the analysis of hetero-organic molecules, just as in traditional electrospray ionization (ESI). However, unlike standard ESI, the proton transfer process of SESI occurs in the vapor phase rather than in solution (Fig. 1), and therefore SESI is best suited for detecting organic volatiles and aerosols.
We are expanding the use of SESI-MS to the detection of bacterial volatiles as a method for bacterial identification and characterization8. We have demonstrated that SESI-MS volatile fingerprinting, combined with a statistical analysis method, can be used to differentiate bacterial genera, species, and mixed cultures in a variety of growth media.8 Here we provide the steps for obtaining bacterial volatile fingerprints using SESI-MS, including the instrumental parameters that should be optimized to ensure robust bacterial identification and characterization.

Characterizing Bacterial Volatiles using Secondary Electrospray Ionization Mass Spectrometry SESI-MS

Secondary electrospray ionization mass spectrometry (SESI-MS) enables the detection of volatile organic compounds (VOCs) without the need for any sample pretreatment. This protocol provides instructions for the rapid (within minutes) characterization of bacterial VOCs using SESI-MS.

Secondary electrospray ionization mass spectrometry (SESI-MS) is a method developed for the rapid detection of volatile compounds, without the need for ...

Bioengineering

An Inverse Analysis Approach to the Characterization of Chemical Transport in Paints

The ability to directly characterize chemical transport and interactions that occur within a material (i.e., subsurface dynamics) is a vital component in understanding contaminant mass transport and the ability to decontaminate materials. If a material is contaminated, over time, the transport of highly toxic chemicals (such as chemical warfare agent species) out of the material can result in vapor exposure or transfer to the skin, which can result in percutaneous exposure to personnel who interact with the material. Due to the high toxicity of chemical warfare agents, the release of trace chemical quantities is of significant concern. Mapping subsurface concentration distribution and transport characteristics of absorbed agents enables exposure hazards to be assessed in untested conditions. Furthermore, these tools can be used to characterize subsurface reaction dynamics to ultimately design improved decontaminants or decontamination procedures. To achieve this goal, an inverse analysis mass transport modeling approach was developed that utilizes time-resolved mass spectroscopy measurements of vapor emission from contaminated paint coatings as the input parameter for calculation of subsurface concentration profiles. Details are provided on sample preparation, including contaminant and material handling, the application of mass spectrometry for the measurement of emitted contaminant vapor, and the implementation of inverse analysis using a physics-based diffusion model to determine transport properties of live chemical warfare agents including distilled mustard (HD) and the nerve agent VX.

In this paper, a procedure for quantifying the mass transport parameters of chemicals in various materials is presented. This process involves employing an inverse-analysis based diffusion model to vapor emission profiles recorded by real-time, mass spectrometry in high vacuum.

The ability to directly characterize chemical transport and interactions that occur within a material (i.e., subsurface dynamics) is a vital component in ...

Analysis of Volatile and Oxidation Sensitive Compounds Using a Cold Inlet System and Electron Impact Mass Spectrometry

This video presents a protocol for the mass spectrometrical analysis of volatile and oxidation sensitive compounds using electron impact ionization. The analysis of volatile and oxidation sensitive compounds by mass spectrometry is not easily achieved, as all state-of-the-art mass spectrometric methods require at least one sample preparation step, e.g., dissolution and dilution of the analyte (electrospray ionization), co-crystallization of the analyte with a matrix compound (matrix-assisted laser desorption/ionization), or transfer of the prepared samples into the ionization source of the mass spectrometer, to be conducted under atmospheric conditions. Here, the use of a sample inlet system is described which enables the analysis of volatile metal organyls, silanes, and phosphanes using a sector field mass spectrometer equipped with an electron impact ionization source. All sample preparation steps and the sample introduction into the ion source of the mass spectrometer take place either under air-free conditions or under vacuum, enabling the analysis of compounds highly susceptible to oxidation. The presented technique is especially of interest for inorganic chemists, working with metal organyls, silanes, or phosphanes, which have to be handled using inert conditions, such as the Schlenk technique. The principle of operation is presented in this video.

This video presents a protocol for the mass spectrometrical analysis of volatile and oxidation sensitive compounds using electron impact ionization. The presented technique is especially of interest for inorganic chemists, working with metal organyls, silanes, or phosphanes which have to be handled using inert conditions, such as the Schlenk technique.

This video presents a protocol for the mass spectrometrical analysis of volatile and oxidation sensitive compounds using electron impact ionization. The ...

Quantitative Detection of Trace Explosive Vapors by Programmed Temperature Desorption Gas Chromatography-Electron Capture Detector

The direct liquid deposition of solution standards onto sorbent-filled thermal desorption tubes is used for the quantitative analysis of trace explosive vapor samples. The direct liquid deposition method yields a higher fidelity between the analysis of vapor samples and the analysis of solution standards than using separate injection methods for vapors and solutions, i.e., samples collected on vapor collection tubes and standards prepared in solution vials. Additionally, the method can account for instrumentation losses, which makes it ideal for minimizing variability and quantitative trace chemical detection. Gas chromatography with an electron capture detector is an instrumentation configuration sensitive to nitro-energetics, such as TNT and RDX, due to their relatively high electron affinity. However, vapor quantitation of these compounds is difficult without viable vapor standards. Thus, we eliminate the requirement for vapor standards by combining the sensitivity of the instrumentation with a direct liquid deposition protocol to analyze trace explosive vapor samples.

Trace explosive vapors of TNT and RDX collected on sorbent-filled thermal desorption tubes were analyzed using a programmed temperature desorption system coupled to GC with an electron capture detector. The instrumental analysis is combined with direct liquid deposition method to reduce sample variability and account for instrumentation drift and losses.

The direct liquid deposition of solution standards onto sorbent-filled thermal desorption tubes is used for the quantitative analysis of trace explosive ...

PTR-ToF-MS Coupled with an Automated Sampling System and Tailored Data Analysis for Food Studies: Bioprocess Monitoring, Screening and Nose-space Analysis

Proton Transfer Reaction (PTR), combined with a Time-of-Flight (ToF) Mass Spectrometer (MS) is an analytical approach based on chemical ionization that belongs to the Direct-Injection Mass Spectrometric (DIMS) technologies. These techniques allow the rapid determination of volatile organic compounds (VOCs), assuring high sensitivity and accuracy. In general, PTR-MS requires neither sample preparation nor sample destruction, allowing real time and non-invasive analysis of samples. PTR-MS are exploited in many fields, from environmental and atmospheric chemistry to medical and biological sciences. More recently, we developed a methodology based on coupling PTR-ToF-MS with an automated sampler and tailored data analysis tools, to increase the degree of automation and, consequently, to enhance the potential of the technique. This approach allowed us to monitor bioprocesses (e.g. enzymatic oxidation, alcoholic fermentation), to screen large sample sets (e.g. different origins, entire germoplasms) and to analyze several experimental modes (e.g. different concentrations of a given ingredient, different intensities of a specific technological parameter) in terms of VOC content. Here, we report the experimental protocols exemplifying different possible applications of our methodology: i.e. the detection of VOCs released during lactic acid fermentation of yogurt (on-line bioprocess monitoring), the monitoring of VOCs associated with different apple cultivars (large-scale screening), and the in vivo study of retronasal VOC release during coffee drinking (nosespace analysis).

Proton Transfer Reaction Time of Flight Mass Spectrometry allows high-sensitivity, rapid and non-invasive analysis of volatile organic compounds. To demonstrate its potential, we give three examples: lactic acid fermentation of yogurt (on-line bioprocess monitoring), different apple genotypes (large-scale screening), and retronasal space after drinking coffee (nosespace analysis).

Proton Transfer Reaction (PTR), combined with a Time-of-Flight (ToF) Mass Spectrometer (MS) is an analytical approach based on chemical ionization that ...

Analyzing the Photo-oxidation of 2-propanol at Indoor Air Level Concentrations Using Field Asymmetric Ion Mobility Spectrometry

We demonstrate a versatile protocol to be used for determining the effectiveness of photocatalysts in degrading indoor air concentration (ppb) volatile organic carbons (VOCs), illustrating this with a titanium dioxide based catalyst, and the VOC 2-propanol. The protocol takes advantage of field asymmetric ion mobility spectroscopy (FAIMS), an analysis tool that is capable of continuously identifying and monitoring the concentration of VOCs such as 2-propanol and acetone at the ppb level. The continuous nature of FAIMS allows detailed kinetic analysis, and long-term reactions, offering a significant advantage over gas chromatography, a batch process traditionally used in air purification characterization. The use of FAIMS in photocatalytic air purification has only recently been used for the first time, and with the protocol illustrated here, the flexibility in allowing alternative VOCs and photocatalysts to be tested using comparable protocols offers a unique system to elucidate photocatalytic air purification reactions at low concentrations.

A protocol for determining the effectiveness of photocatalysts in degrading indoor air concentration (ppb) model volatile organic carbons such as 2-propanol is described.

We demonstrate a versatile protocol to be used for determining the effectiveness of photocatalysts in degrading indoor air concentration (ppb) volatile ...

Real-time Breath Analysis by Using Secondary Nanoelectrospray Ionization Coupled to High Resolution Mass Spectrometry

Exhaled volatile organic compounds (VOCs) have aroused considerable interest, since they can serve as biomarkers for disease diagnosis and environmental exposure in a non-invasive manner. In this work, we present a protocol to characterize the exhaled VOCs in real time by using secondary nanoelectrospray ionization coupled to high resolution mass spectrometry (Sec-nanoESI-HRMS). The homemade Sec-nanoESI source was readily set up based on a commercial nanoESI source. Hundreds of peaks were observed in the background-subtracted mass spectra of exhaled breath, and the mass accuracy values are -4.0-13.5 ppm and -20.3-1.3 ppm in the positive and negative ion detection modes, respectively. The peaks were assigned with accurate elemental composition according to the accurate mass and isotopic pattern. Less than 30 s is used for one exhalation measurement, and it takes approximately 7 min for six replicated measurements.

A protocol for characterizing chemical composition of exhaled breath in real time by using secondary nanoelectrospray ionization coupled to high resolution mass spectrometry is demonstrated.

Exhaled volatile organic compounds (VOCs) have aroused considerable interest, since they can serve as biomarkers for disease diagnosis and environmental ...

Profiling Volatile Compounds in Blackcurrant Fruit using Headspace Solid-Phase Microextraction Coupled to Gas Chromatography-Mass Spectrometry

There is an increasing interest in measuring volatile organic compounds (VOCs) emitted by ripe fruits for the purpose of breeding varieties or cultivars with enhanced organoleptic characteristics and thus, to increase consumer acceptance. High-throughput metabolomic platforms have been recently developed to quantify a wide range of metabolites in different plant tissues, including key compounds responsible for fruit taste and aroma quality (volatilomics). A method using headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) is described here for the identification and quantification of VOCs emitted by ripe blackcurrant fruits, a berry highly appreciated for its flavor and health benefits.
Ripe fruits of blackcurrant plants (Ribes nigrum) were harvested and directly frozen in liquid nitrogen. After tissue homogenization to produce a fine powder, samples were thawed and immediately mixed with sodium chloride solution. Following centrifugation, the supernatant was transferred into a headspace glass vial containing sodium chloride. VOCs were then extracted using a solid-phase microextraction (SPME) fiber and a gas chromatograph coupled to an ion trap mass spectrometer. Volatile quantification was performed on the resulting ion chromatograms by integrating peak area, using a specific m/z ion for each VOC. Correct VOC annotation was confirmed by comparing retention times and mass spectra of pure commercial standards run under the same conditions as the samples. More than 60 VOCs were identified in ripe blackcurrant fruits grown in contrasting European locations. Among the identified VOCs, key aroma compounds, such as terpenoids and C6 volatiles, can be used as biomarkers for blackcurrant fruit quality. In addition, advantages and disadvantages of the method are discussed, including prospective improvements. Furthermore, the use of controls for batch correction and minimization of drift intensity have been emphasized.

A headspace solid-phase microextraction-gas-chromatography platform is described here for fast, reliable, and semi-automated volatile identification and quantification in ripe blackcurrant fruits. This technique can be used to increase knowledge about fruit aroma and to select cultivars with enhanced flavor for the purpose of breeding.

There is an increasing interest in measuring volatile organic compounds (VOCs) emitted by ripe fruits for the purpose of breeding varieties or cultivars ...

Biochemistry

Capturing Actively Produced Microbial Volatile Organic Compounds from Human-Associated Samples with Vacuum-Assisted Sorbent Extraction

Volatile organic compounds (VOCs) from biological samples have unknown origins. VOCs may originate from the host or different organisms from within the host&#39;s microbial community. To disentangle the origin of microbial VOCs, volatile headspace analysis of bacterial mono- and co-cultures of Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, and stable isotope probing in biological samples of feces, saliva, sewage, and sputum were performed. Mono- and co-cultures were used to identify volatile production from individual bacterial species or in combination with stable isotope probing to identify the active metabolism of microbes from the biological samples.
Vacuum-assisted sorbent extraction (VASE) was employed to extract the VOCs. VASE is an easy-to-use, commercialized, solvent-free headspace extraction method for semi-volatile and volatile compounds. The lack of solvents and the near-vacuum conditions used during extraction make developing a method relatively easy and fast when compared to other extraction options such as tert-butylation and solid phase microextraction. The workflow described here was used to identify specific volatile signatures from mono- and co-cultures. Furthermore, analysis of the stable isotope probing of human associated biological samples identified VOCs that were either commonly or uniquely produced. This paper presents the general workflow and experimental considerations of VASE in conjunction with stable isotope probing of live microbial cultures.

This protocol describes the extraction of volatile organic compounds from a biological sample with the vacuum-assisted sorbent extraction method, gas chromatography coupled with mass spectrometry using the Entech Sample Preparation Rail, and data analysis. It also describes culture of biological samples and stable isotope probing.

Volatile organic compounds (VOCs) from biological samples have unknown origins. VOCs may originate from the host or different organisms from within the ...

Atmospheric-pressure Molecular Imaging of Biological Tissues and Biofilms by LAESI Mass Spectrometry

Ambient ionization methods in mass spectrometry allow analytical investigations to be performed directly on a tissue or biofilm under native-like experimental conditions. Laser ablation electrospray ionization (LAESI) is one such development and is particularly well-suited for the investigation of water-containing specimens. LAESI utilizes a mid-infrared laser beam (2.94 &mu;m wavelength) to excite the water molecules of the sample. When the ablation fluence threshold is exceeded, the sample material is expelled in the form of particulate matter and these projectiles travel to tens of millimeters above the sample surface. In LAESI, this ablation plume is intercepted by highly charged droplets to capture a fraction of the ejected sample material and convert its chemical constituents into gas-phase ions. A mass spectrometer equipped with an atmospheric-pressure ion source interface is employed to analyze and record the composition of the released ions originating from the probed area (pixel) of the sample. A systematic interrogation over an array of pixels opens a way for molecular imaging in the microprobe analysis mode. A unique aspect of LAESI mass spectrometric imaging is depth profiling that, in combination with lateral imaging, enables three-dimensional (3D) molecular imaging. With current lateral and depth resolutions of ~100 &mu;m and ~40 &mu;m, respectively, LAESI mass spectrometric imaging helps to explore the molecular structure of biological tissues. Herein, we review the major elements of a LAESI system and provide guidelines for a successful imaging experiment.

Laser ablation electrospray ionization (LAESI) is an atmospheric-pressure ion source for mass spectrometry. In the imaging mode, a mid-infrared laser probes the distributions of molecules across a tissue section or a biofilm. This technique presents a new approach for diverse bioanalytical studies carried out under native experimental conditions.