This research was supported by the National Institute of Justice (Award No. 2015-DN-BX-K058 &#38; 2018-75-CX-0035). The opinions, findings, and conclusions expressed here are those of the author and do not necessarily reflect those of the funding organizations.

1. General TV-SPME sample preparation and GC-MS analysis
NOTE: If the sample is already dissolved in a matrix, skip to Step 1.2.
<ol>
	<li>Extract or dissolve the solid sample in enough solvent (water, methanol, acetone, etc.) to reach the desired concentration. Liquid samples can be used &#8220;as is&#8221;. 
	NOTE: The amount of solid sample used depends on the desired concentration of the sample. Concentrations below 1 ppm (w/v) are recommended to avoid overloading the GC column. Analyte should be soluble in chosen organic solvent.
	<ol>
		<li>Ensure that the sample has fully dissolved.</li>
	</ol>
	</li>
	<li>Calculate the volume needed to fully vaporize the sample using Equation 3 at the chosen temperature. For example, if the experiment is to be performed at 60 &#176;C, calculate the volume needed to completely vaporize the solvent at 60 &#176;C.
	<ol>
		<li>Transfer this sample volume into a headspace vial and secure the cap. Acceptable methods for transferring liquid samples at the microliter scale include manually via glass syringe, an electronic glass syringe, or an autosampler robot capable of liquid transfers for sample preparation.</li>
	</ol>
	</li>
	<li>If derivatizing the sample, prepare the proper derivatization agent by placing ~1 mL of the agent into a headspace vial.
	<ol>
		<li>Choose the derivatization agent based on the type of derivatization needed: alkylation, acylation, or silyation. In this case, the recommended derivatization agent for the carboxylic acid and alcohol functional groups found on GHB is O-bis(trimethylsilyl) trifluoroacetamide (BSTFA). The derivatization agent may be used &#8220;as is&#8221; and does not require dilution. One mL of derivatization agent is enough to ensure complete saturation of the SPME fiber. 
		CAUTION: Derivatization agents are toxic and should be handled in a fume hood.</li>
	</ol>
	</li>
	<li>Set the proper incubation/extraction temperature based upon the calculation in step 1.2. This temperature ensures total vaporization, sufficient sample extraction, and complete derivatization (if necessary).
	<ol>
		<li>Select GC-MS parameters (oven temperature program, flow rate, inlet temperature, etc.) based on the class of the compound(s) of interest. See Step 3 for an example parameter set.</li>
		<li>Ensure that the proper inlet liner (e.g., 2 mm inner diameter or less) is in the GC inlet.</li>
	</ol>
	</li>
	<li>Ensure the SPME fiber has been properly conditioned and is in good working condition before beginning the analysis.
	<ol>
		<li>Vary the conditioning parameters based on the type of SPME fiber being used. Please see SPME fiber instructions for proper conditioning temperature and time. In general, analyzing several SPME fiber blanks until they are reproducible is sufficient to characterize a SPME fiber as fully conditioned.</li>
	</ol>
	</li>
</ol>
2. Gamma-hydroxybutyrate (GHB) and Gamma-butyrolactone (GBL) sample preparation
<ol>
	<li>Prepare a sample of GHB and/or GBL in water with a concentration of less than 1 ppm.</li>
	<li>Transfer 1 &#181;L of this sample to a 20 mL headspace vial using one of the methods described in 1.2.1.
	<ol>
		<li>Note that that the analysis of aqueous samples requires the lowest sample volumes. For example, one &#181;L of water will completely vaporize into a 20 mL headspace vial at 60 &#176;C.</li>
		<li>Cap the vial immediately.</li>
	</ol>
	</li>
	<li>Place ~1 mL of BSTFA + 1% trimethylchlorosilane (TMCS) into a separate 20 mL headspace vial and cap. 
	NOTE: GBL does not derivatize. A derivatization step is still required, however, is to ensure the water solvent derivatizes and does not interfere with the sample. 
	CAUTION: BSTFA is toxic and should be handled in a fume hood.</li>
</ol>
3. GC-MS parameters and setup for GHB and GBL in water
<ol>
	<li>Create a method using the following GC-MS parameters: 
	Initial oven temperature: 60 &#176;C held for 1 minute. 
	Oven program: 15 &#176;C/minute. 
	Final oven temperature: 280 &#176;C, held for 1 minute. 
	Flow rate: 2.5 mL/minute (speed optimized flow for a 0.25 mm i.d. column). 
	Inlet temperature: 250 &#176;C. 
	Transfer line temperature: 280 &#176;C.</li>
	<li>Ensure a narrow (2 mm i.d. or less) SPME inlet liner has been placed inside the GC inlet.</li>
	<li>Ensure the PDMS/DVB SPME fiber has been properly conditioned and is in good working order prior to analysis. 
	NOTE: PDMS/DVB SPME fibers should be conditioned in the GC inlet at 250 &#176;C for 30 minutes. PDMS/DVB SPME fibers should be an off-white color.</li>
	<li>Run the GC-MS on the sample.</li>
</ol>

<ol>
	<li>Pawliszyn, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Method+and+Device+for+Solid+Phase+Microextraction+and+Desorption.">Method and Device for Solid Phase Microextraction and Desorption.</a> United States patent. , (2005).</li><li>Pawliszyn, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Solid phase microextraction: theory and practice. , (1997).</li><li>Rainey, C. L., Bors, D. E., Goodpaster, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+optimization+of+a+total+vaporization+technique+coupled+to+solid-phase+microextraction.">Design and optimization of a total vaporization technique coupled to solid-phase microextraction.</a> Analytical Chemistry. 86 (22), 11319-11325 (2014).</li><li>Sinnott, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Chemical Engineering Design: Chemical Engineering. 6, (2005).</li><li>Davis, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Detection of Illicit Drugs in Various Matrices Via Total Vaporization Solid-Phase Microextraction (TV-SPME). , (2019).</li><li>Ash, J., Hickey, L., Goodpaster, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formation+and+identification+of+novel+derivatives+of+primary+amine+and+zwitterionic+drugs.">Formation and identification of novel derivatives of primary amine and zwitterionic drugs.</a> Forensic Chemistry. 10, 37-47 (2018).</li><li>Sauzier, G., Bors, D., Ash, J., Goodpaster, J. V., Lewis, S. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimisation+of+recovery+protocols+for+double-base+smokeless+powder+residues+analysed+by+total+vaporisation+(TV)+SPME/GC-MS.">Optimisation of recovery protocols for double-base smokeless powder residues analysed by total vaporisation (TV) SPME/GC-MS.</a> Talanta. 158, 368-374 (2016).</li><li>Bors, D., Goodpaster, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mapping+smokeless+powder+residue+on+PVC+pipe+bomb+fragments+using+total+vaporization+solid+phase+microextraction.">Mapping smokeless powder residue on PVC pipe bomb fragments using total vaporization solid phase microextraction.</a> Forensic science international. 276, 71-76 (2017).</li><li>Bors, D., Goodpaster, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+analysis+of+racing+fuels+using+total+vaporization+and+gas+chromatography+mass+spectrometry+(GC/MS).">Chemical analysis of racing fuels using total vaporization and gas chromatography mass spectrometry (GC/MS).</a> Analytical Methods. 8 (19), 3899-3902 (2016).</li><li>Beiranvand, M., Ghiasvand, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+optimization+of+the+VA-TV-SPME+method+for+ultrasensitive+determination+of+the+PAHs+in+polluted+water.">Design and optimization of the VA-TV-SPME method for ultrasensitive determination of the PAHs in polluted water.</a> Talanta. 212, 120809 (2020).</li></ol>

The authors have nothing to disclose.

TV-SPME has some benefits over liquid injection GC in that large sample sizes (e.g., 100 &#181;L) may be used without instrument modifications. TV-SPME also has some of the same benefits as headspace SPME. Headspace SPME does not require any extraction or filtration because any nonvolatile compounds will remain in the headspace vial and will not be adsorbed onto the fiber, yielding a clean sample. This method also helps to eliminate matrix effects due to this being a two-phase system (headspace and fiber) as opposed to a three-phase system (sample, headspace, and fiber) like standard headspace SPME. TV-SPME is like immersion SPME in that immersion SPME is also a two-phase system. With immersion SPME, a fiber is immersed into a liquid (typically aqueous) sample containing the analyte as opposed to extracting the analyte from its vapor. TV-SPME differs from immersion SPME because immersion SPME requires a polar/aqueous matrix in order to generate sufficient driving force for analytes to leave the solution phase and sorb to the fiber coating. In addition, immersion SPME requires much larger sample volumes (e.g., mL).
Many solvents may be used with TV-SPME including methanol, acetone, water, and acetonitrile. SPME fibers should not be exposed to or immersed in chloroform as these solvents can damage the fiber coating. Twenty mL screw cap glass headspace vials have been found to have the best performance with TV-SPME methods. It is recommended to use an autosampler with TV-SPME. While many parameters of the TV-SPME method may be adjusted as desired, the proper volume and extraction temperature must be used for each solvent. Sample volume and extraction temperature are proportional with one another and must be adjusted accordingly. For example, the extraction temperature of a method may be reduced, but the sample volume must also be reduced. This volume may be found by adjusting Equation 3.
Modifications to the derivatization procedure may be made. Derivatization may be performed pre- or post-extraction, at room temperature or heated in the agitator, by exposing the fiber to the vapor of the derivatization agent or by direct immersion of the fiber in the derivatization agent.
There are limitations to the TV-SPME method including the need for compounds to be soluble, thermally stable, and volatile. TV-SPME requires expensive SPME fibers which can have their coating stripped or broken during analysis. These limitations are outweighed by benefits such as large sample volumes relative to typical GC injection volumes, high sensitivity, and no need for filtration. TV-SPME is preferred to headspace SPME because more of the sample is extracted onto the fiber and matrix effects are reduced. TV-SPME is also preferred to immersion SPME because immersion SPME consumes much more sample than TV-SPME. TV-SPME allows for derivatization during the extraction process which reduces analysis time compared to methods such as liquid injection that require that the analyte to be derivatized prior to injection. TV-SPME also requires little to no sample preparation. TV-SPME is simple, efficient, and sensitive for the analysis of a wide variety of samples, including drugs, explosive materials, and racing fuels.

Solid-Phase Microextraction (SPME) is a sampling technique in which a liquid or solid sample is placed into a headspace vial and a SPME fiber, coated with a polymeric material, is then introduced into the sample headspace (or immersed in a liquid sample). The analyte is sorbed onto the fiber and then the fiber is placed inside the GC inlet for desorption1,2. Total Vaporization Solid-Phase Microextraction (TV-SPME) is a similar technique as immersion SPME but completely vaporizes a liquid sample before analytes are adsorbed onto the fiber. This allows for partitioning of the analyte between only the solvent vapor and the coating of the fiber, allowing for more of the analyte to be adsorbed onto the fiber and resulting in good sensitivity3. There are various SPME fibers available and the fiber should be chosen based on the analyte of interest, solvent/matrix, and derivatization agent. See Table 1 for established TV-SPME analytes.
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Sample</td>
			<td>Analyte(s)</td>
			<td>Recommended SPME Fiber</td>
			<td>Reference(s)</td>
		</tr>
		<tr>
			<td>Human Hair</td>
			<td>Nicotine, cotinine</td>
			<td>Polydimethylsiloxane/divinylbenzene (PDMS/DVB), polyacrylate (PA)</td>
			<td>3</td>
		</tr>
		<tr>
			<td>Smokeless Powder</td>
			<td>Nitroglycerin, diphenylamine</td>
			<td>Polydimethylsiloxane (PDMS), polyethylene glycol (PEG)</td>
			<td>7, 8</td>
		</tr>
		<tr>
			<td>Racing fuel</td>
			<td>Methanol, nitromethane</td>
			<td>PEG</td>
			<td>9</td>
		</tr>
		<tr>
			<td>Water</td>
			<td>Polycyclic aromatic hydrocarbons</td>
			<td>PDMS</td>
			<td>10</td>
		</tr>
		<tr>
			<td>Beverages</td>
			<td>&#611;-Hydroxybutyric acid, &#611;-butyrolactone</td>
			<td>PDMS</td>
			<td>This Work</td>
		</tr>
		<tr>
			<td>Solid Powder</td>
			<td>Methamphetamine, amphetamine</td>
			<td>PDMS/DVB</td>
			<td>Unpublished</td>
		</tr>
	</tbody>
</table>
Table 1. Recommended SPME fibers with established TV-SPME analytes.
To perform TV-SPME, analytes are dissolved in a solvent and an aliquot of this mixture is placed into a headspace vial. Samples do not need to be filtered because only the solvent and volatile analytes will vaporize. Specific volumes of liquid samples must be used to ensure total vaporization of the sample. These volumes are determined by using the Ideal Gas Law to calculate the number of moles of a solvent multiplied by the molar volume of the liquid (Equation 1). 
<img alt="Equation 1" src="/files/ftp_upload/61880/61880equ01.jpg" /> Equation 1
where Vo is the volume of the sample (mL), P is the vapor pressure of the solvent (bar), Vv is the volume of the vial (L), R is the ideal gas constant (0.083145 <img alt="Equation 1" src="/files/ftp_upload/61880/61880equ04.jpg" />), M is the molar mass of the solvent (g/mol), T is temperature (K), and <img alt="Equation 5" src="/files/ftp_upload/61880/61880equ05.jpg" /> is the density of the solvent (g/mL).3
In order to use the correct vapor pressure, the Antoine equation (Equation 2) is used to account for the influence of temperature:4 
<img alt="Equation 2" src="/files/ftp_upload/61880/61880equ02.jpg" /> Equation 2
where T is temperature and A, B, and C are the Antoine constants for the solvent. Equation 2 may be substituted into Equation 1, yielding: 
<img alt="Equation 3" src="/files/ftp_upload/61880/61880equ03v2.jpg" /> Equation 3
Equation 3 gives the volume of the sample (Vo) that can be completely vaporized as a function of the temperature and solvent used.
To perform derivatization with TV-SPME, the SPME fiber is first exposed to a vial containing the derivatization agent for a predetermined amount of time depending on the analyte. The SPME fiber is then exposed to a new vial containing the analyte of interest. This vial is heated inside of a heated agitator. The analyte is then adsorbed onto the fiber with the derivatization agent. The derivatization of the analyte and/or the matrix takes place on the fiber before being inserted into the GC inlet for desorption. Figure 1 shows a depiction of the TV-SPME process with derivatization.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/61880/61880fig01.jpg" /> 
Figure 1: Depiction of the TV-SPME process with derivatization.&#160;The SPME fiber first enters the derivatization vial where the derivatization agent (yellow circles) sorb onto the fiber. The fiber is then introduced to the sample (blue circles) and heated. Formation of the derivative (green circles) takes place on the fiber during the extraction time. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
TV-SPME is beneficial because it allows for the analyte to be derivatized during the extraction process which reduces analysis time. Other methods, such as liquid injection, require that the analyte react with the derivatizing agent in solution prior to being injected into the GC. TV-SPME also requires little to no sample preparation. A matrix containing an analyte may be placed directly into the headspace vial and analyzed. Many compounds of interest are compatible with TV-SPME. Compounds must be soluble in a solvent and sufficiently volatile to allow for vaporization. Additionally, compounds must be thermally stable to be analyzed by GC-MS. TV-SPME has been used to analyze drugs and drug metabolites, racing fuels, polycyclic aromatic hydrocarbons, and explosive materials3,5,6,7,8,9,10.

<table>
	<tbody>
		<tr>
			<td>10 &#181;L Syringe</td>
			<td>Gerstel</td>
			<td>100111-014-00</td>
			<td></td>
		</tr>
		<tr>
			<td>BSTFA + 1% TMCS (10 x 1 GM)</td>
			<td>Regis Technologies Inc.</td>
			<td>50442882</td>
			<td></td>
		</tr>
		<tr>
			<td>eVol XR Sample Dispensing System Kit</td>
			<td>ThermoFisher Scientific</td>
			<td>66002-024</td>
			<td></td>
		</tr>
		<tr>
			<td><img alt="Equation 6" src="/files/ftp_upload/61880/61880equ06.jpg" />-Butyrolactone (GBL)</td>
			<td>Sigma-Aldrich</td>
			<td>B103608-26G</td>
			<td></td>
		</tr>
		<tr>
			<td><img alt="Equation 7" src="/files/ftp_upload/61880/61880equ06.jpg" />-Hydroxy Butyric Acid (GHB)</td>
			<td>Cayman Chemicals</td>
			<td>9002506</td>
			<td></td>
		</tr>
		<tr>
			<td>Headspace Screw-Thread Vials, 18 mm</td>
			<td>Restek</td>
			<td>23083</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnetic Screw-Thread Caps, 18 mm</td>
			<td>Restek</td>
			<td>23091</td>
			<td></td>
		</tr>
		<tr>
			<td>Optima water for HPLC</td>
			<td>Fisher Chemical</td>
			<td>W71</td>
			<td></td>
		</tr>
		<tr>
			<td>SPME Fiber Assembly Polydimethylsiloxane (PDMS)</td>
			<td>Supelco</td>
			<td>57341-U</td>
			<td></td>
		</tr>
		<tr>
			<td>SPME Fiber Assembly Polydimethylsiloxane/Divinylbenzene (PDMS/DVB)</td>
			<td>Supelco</td>
			<td>57293-U</td>
			<td></td>
		</tr>
		<tr>
			<td>Topaz 2.0 mm ID Straight Inlet Liner</td>
			<td>Restek</td>
			<td>23313</td>
			<td></td>
		</tr>
	</tbody>
</table>

gas chromatography-mass spectrometry paired with total vaporization solid-phase microextraction as a forensic tool

Gas Chromatography &#8211; Mass Spectrometry (GC-MS) is a frequently used technique for the analysis of numerous analytes of forensic interest, including controlled substances, ignitable liquids, and explosives. GC-MS can be coupled with Solid-Phase Microextraction (SPME), in which a fiber with a sorptive coating is placed into the headspace above a sample or immersed in a liquid sample. Analytes are sorbed onto the fiber which is then placed inside the heated GC inlet for desorption. Total Vaporization Solid-Phase Microextraction (TV-SPME) utilizes the same technique as immersion SPME but immerses the fiber into a completely vaporized sample extract. This complete vaporization results in a partition between only the vapor phase and the SPME fiber without interference from a liquid phase or any insoluble materials. Depending upon the boiling point of the solvent used, TV-SPME allows for large sample volumes (e.g., up to hundreds of microliters). On-fiber derivatization may also be performed using TV-SPME. TV-SPME has been used to analyze drugs and their metabolites in hair, urine, and saliva. This simple technique has also been applied to street drugs, lipids, fuel samples, post-blast explosive residues, and pollutants in water. This paper highlights the use of TV-SPME to identify illegal adulterants in very small samples (microliter quantities) of alcoholic beverages. Both gamma-hydroxybutyrate (GHB) and gamma-butyrolactone (GBL) were identified at levels that would be found in spiked drinks. Derivatization by a trimethylsilyl agent allowed for conversion of the aqueous matrix and GHB into their TMS derivatives. Overall, TV-SPME is quick, easy, and requires no sample preparation aside from placing the sample into a headspace vial.

A GBL volume study was performed to demonstrate the sensitivity of TV-SPME compared to headspace and immersion SPME. A 100ppmv sample of GBL in water was prepared and placed into 20 mL headspace vials with volumes of 1, 3, 10, 30, 100, 300, 1000, 3000, and 10,000 &#181;L. The phase ratio of the samples allowed for TV-SPME (1-3 &#181;L), Headspace SPME (10 &#8211; 3,000 &#181;L) and Immersion SPME (10,000 &#181;L). All samples were analyzed in triplicate and the average peak area was plotted against the sample volume. Overall, sample volumes that allowed for TV-SPME demonstrated more sensitivity than headspace or immersion SPME for GBL in water as shown in Figure 2. A comparison of the chromatograms for each method is shown in Figure 3.
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/61880/61880fig02.jpg" /> 
Figure 2: Graph of average peak area versus sample volume for GBL in water.&#160;A GBL volume study was performed to demonstrate the efficacy of TV-SPME compared to headspace and immersion SPME. A 100ppmv sample of GBL in water was prepared and placed into 20 mL headspace vials with volumes of 1, 3, 10, 30, 100, 300, 1000, 3000, &#38; 10000 &#181;L. All samples were analyzed in triplicate and error bars correspond to standard deviation of the mean. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/61880/61880fig03.jpg" /> 
Figure 3: Total Ion Chromatograms for GBL in water.&#160;(100 ppm) for 3 &#181;L (blue), 300 &#181;L (red), and 10,000 &#181;L (green). <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Realistic samples of wine spiked with an effective dose of GHB and GBL are shown in Figure 4 and Figure 5, respectively. These samples also show the interconversion of GBL and GHB. When TV-SPME is performed properly, a sharp, abundant peak will result as shown in Figure 6. TV-SPME has good sensitivity and therefore proper concentrations should be used as to not overload the column. When high concentrations are present, peak asymmetry will result as shown in Figure 5 and Figure 7. In these cases, diluting the sample or using a split injection can improve peak shape.
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/61880/61880fig04.jpg" /> 
Figure 4: Realistic sample of GHB in wine with an 8 mg/mL concentration.&#160;Peaks: 1) GBL, 2) hexanoic acid-TMS, 3) GHB-TMS2, 4) benzoic acid-TMS, 5) octanoic acid-TMS, 6) glycerol-TMS3, * denotes a cyclic siloxane (fiber/column bleed). <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/61880/61880fig05.jpg" /> 
Figure 5: Realistic sample of GBL in wine with a 10 mg/mL concentration.&#160;Peaks: 1) GBL, 2) hexanoic acid-TMS, 3) Siloxane, 4) trimethyl(2-phenylethoxy) silane, 5) GHB-TMS2. TIC shows GBL converting to GHB. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig05large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/61880/61880fig06.jpg" /> 
Figure 6: Total Ion Chromatogram for GBL in water with a 0.1-ppm concentration.&#160;Results following the TV-SPME method previously described for GBL in water. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig06large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/61880/61880fig07.jpg" /> 
Figure 7: Total Ion Chromatogram for GBL in water with a 10-ppm concentration.&#160;Results following the TV-SPME method previously described for GBL in water. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/61880/61880fig07large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
When derivatizing, the analyst should ensure that the method allows for the analyte(s) to be fully derivatized prior to being desorbed into the GC. Partial derivatization can result in a peak representing the derivatized analyte and a peak representing the underivatized analyte. Partial derivatization will also result in a lower sensitivity for the analyte as less of it may adsorb onto the fiber.

Watch this Scientific Journal Video about Gas Chromatography-Mass Spectrometry Paired with Total Vaporization Solid-Phase Microextraction as a Forensic Tool at JoVE.com

Gas Chromatography-Mass Spectrometry Paired with Total Vaporization Solid-Phase Microextraction as a Forensic Tool

Total Vaporization Solid Phase Microextraction (TV-SPME) completely vaporizes a liquid sample whilst analytes are sorbed onto a SPME fiber. This allows for partitioning of the analyte between only the solvent vapor and the SPME fiber coating.

Gas Chromatography &#8211; Mass Spectrometry (GC-MS) is a frequently used technique for the analysis of numerous analytes of forensic interest, including ...

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6.9K Views.  Indiana University - Purdue University Indianapolis. Total Vaporization Solid Phase Microextraction (TV-SPME) completely vaporizes a liquid sample whilst analytes are sorbed onto a SPME fiber. This allows for partitioning of the analyte between only the solvent vapor and the SPME fiber coating.

Video: Gas Chromatography-Mass Spectrometry Paired with Total Vaporization Solid-Phase Microextraction as a Forensic Tool

Large Scale Non-targeted Metabolomic Profiling of Serum by Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS)

Non-targeted metabolite profiling by ultra performance liquid chromatography coupled with mass spectrometry (UPLC-MS) is a powerful technique to investigate metabolism. The approach offers an unbiased and in-depth analysis that can enable the development of diagnostic tests, novel therapies, and further our understanding of disease processes. The inherent chemical diversity of the metabolome creates significant analytical challenges and there is no single experimental approach that can detect all metabolites. Additionally, the biological variation in individual metabolism and the dependence of metabolism on environmental factors necessitates large sample numbers to achieve the appropriate statistical power required for meaningful biological interpretation. To address these challenges, this tutorial outlines an analytical workflow for large scale non-targeted metabolite profiling of serum by UPLC-MS. The procedure includes guidelines for sample organization and preparation, data acquisition, quality control, and metabolite identification and will enable reliable acquisition of data for large experiments and provide a starting point for laboratories new to non-targeted metabolite profiling by UPLC-MS.

Large Scale Non-targeted Metabolomic Profiling of Serum by Ultra Performance Liquid Chromatography-Mass Spectrometry UPLC-MS

Non-targeted metabolite profiling by ultra performance liquid chromatography coupled with mass spectrometry (UPLC-MS) is a powerful technique to investigate metabolism. This article outlines a typical workflow utilized for non-targeted metabolite profiling of serum including sample organization and preparation, data acquisition, data analysis, quality control, and metabolite identification.

Non-targeted metabolite profiling by ultra performance liquid chromatography coupled with mass spectrometry (UPLC-MS) is a powerful technique to ...

Synthetic Methodology for Asymmetric Ferrocene Derived Bio-conjugate Systems via Solid Phase Resin-based Methodology

Early detection is a key to successful treatment of most diseases, and is particularly imperative for the diagnosis and treatment of many types of cancer. The most common techniques utilized are imaging modalities such as Magnetic Resonance Imaging (MRI), Positron Emission Topography (PET), and Computed Topography (CT) and are optimal for understanding the physical structure of the disease but can only be performed once every four to six weeks due to the use of imaging agents and overall cost. With this in mind, the development of &#8220;point of care&#8221; techniques, such as biosensors, which evaluate the stage of disease and/or efficacy of treatment in the clinician&#8217;s office and do so in a timely manner, would revolutionize treatment protocols.1 As a means to exploring ferrocene based biosensors for the detection of biologically relevant molecules2, methods were developed to produce ferrocene-biotin bio-conjugates described herein. This report will focus on a biotin-ferrocene-cysteine system that can be immobilized on a gold surface.

The synthesis of asymmetric species of ferrocene is challenging using solution techniques. This report focuses on the methods carried out to produce a ferrocene-biotin bioconjugate using facile and clean reactions accomplished via solid-phase synthesis. Incorporation of a thiolate moiety is shown to impart the ability for immobilization on gold surfaces.

Early detection is a key to successful treatment of most diseases, and is particularly imperative for the diagnosis and treatment of many types of cancer. ...

Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

This manuscript outlines how to visualize mass spectrometry ambient ionization sources using schlieren photography. In order to properly optimize the mass spectrometer, it is necessary to characterize and understand the physical principles of the source. Most commercial ambient ionization sources utilize jets of nitrogen, helium, or atmospheric air to facilitate the ionization of the analyte. As a consequence, schlieren photography can be used to visualize the gas streams by exploiting the differences in refractive index between the streams and ambient air for visualization in real time. The basic setup requires a camera, mirror, flashlight, and razor blade. When properly configured, a real time image of the source is observed by watching its reflection. This allows for insight into the mechanism of action in the source, and pathways to its optimization can be elucidated. Light is shed on an otherwise invisible situation.

This paper presents a protocol for the visualization of gaseous streams of an ambient ionization source using schlieren photography and mass spectrometry.

This manuscript outlines how to visualize mass spectrometry ambient ionization sources using schlieren photography. In order to properly optimize the mass ...

Coulomb Explosion Imaging as a Tool to Distinguish Between Stereoisomers

This article shows how the COLTRIMS (Cold Target Recoil Ion Momentum Spectroscopy) or the "reaction microscope" technique can be used to distinguish between enantiomers (stereoisomers) of simple chiral species on the level of individual molecules. In this approach, a gaseous molecular jet of the sample expands into a vacuum chamber and intersects with femtosecond (fs) laser pulses. The high intensity of the pulses leads to fast multiple ionization, igniting a so-called Coulomb Explosion that produces several cationic (positively charged) fragments. An electrostatic field guides these cations onto time- and position-sensitive detectors. Similar to a time-of-flight mass spectrometer, the arrival time of each ion yields information on its mass. As a surplus, the electrostatic field is adjusted in a way that the emission direction and the kinetic energy after fragmentation lead to variations in the time-of-flight and in the impact position on the detector.
Each ion impact creates an electronic signal in the detector; this signal is treated by high-frequency electronics and recorded event by event by a computer. The registered data correspond to the impact times and positions. With these data, the energy and the emission direction of each fragment can be calculated. These values are related to structural properties of the molecule under investigation, i.e. to the bond lengths and relative positions of the atoms, allowing to determine molecule by molecule the handedness of simple chiral species and other isomeric features.

For small chiral species, Coulomb Explosion Imaging provides a new approach to determine the handedness of individual molecules.

This article shows how the COLTRIMS (Cold Target Recoil Ion Momentum Spectroscopy) or the "reaction microscope" technique can be used to distinguish ...

Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

Solid-phase synthesis has been used to obtain canonical and modified polymers of nucleic acids, specifically of DNA or RNA, which has made it a popular methodology for applications in various fields and for different research purposes. The procedure described herein focuses on the synthesis, purification, and characterization of dodecamers of RNA 5&#39;-[CUA CGG AAU CAU]-3&#39; containing zero, one, or two modifications located at the C2&#39;-O-position. The probes are based on 2-thiophenylmethyl groups, incorporated into RNA nucleotides via standard organic synthesis and introduced into the corresponding oligonucleotides via their respective phosphoramidites. This report makes use of phosphoramidite chemistry via the four canonical nucleobases (Uridine (U), Cytosine (C), Guanosine (G), Adenosine (A)), as well as 2-thiophenylmethyl functionalized nucleotides modified at the 2&#39;-O-position; however, the methodology is amenable for a large variety of modifications that have been developed over the years. The oligonucleotides were synthesized on a controlled-pore glass (CPG) support followed by cleavage from the resin and deprotection under standard conditions, i.e., a mixture of ammonia and methylamine (AMA) followed by hydrogen fluoride/triethylamine/N-methylpyrrolidinone. The corresponding oligonucleotides were purified via polyacrylamide electrophoresis (20% denaturing) followed by elution, desalting, and isolation via reversed-phase chromatography (Sep-pak, C18-column). Quantification and structural parameters were assessed via ultraviolet-visible (UV-vis) and circular dichroism (CD) photometric analysis, respectively. This report aims to serve as a resource and guide for beginner and expert researchers interested in embarking in this field. It is expected to serve as a work-in-progress as new technologies and methodologies are developed. The description of the methodologies and techniques within this document correspond to a DNA/RNA synthesizer (refurbished and purchased in 2013) that uses phosphoramidite chemistry.

Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

This article provides a detailed procedure on the solid-phase synthesis, purification, and characterization of dodecamers of RNA modified at the C2&#39;-O-position. UV-vis and circular dichroism photometric analyses are used to quantify and characterize structural aspects, i.e., single-strands or double-strands.

Solid-phase synthesis has been used to obtain canonical and modified polymers of nucleic acids, specifically of DNA or RNA, which has made it a popular ...

Electrochemical Impedance Spectroscopy as a Tool for Electrochemical Rate Constant Estimation

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). In the case of fast reversible electrochemical processes, current is predominantly affected by the rate of diffusion, which is the slowest and limiting stage. EIS is a powerful technique that allows separate analysis of stages of charge transfer that have different AC frequency response. The capability of the method was used to extract the value of charge transfer resistance, which characterizes the rate of charge exchange on the electrode-solution interface. The application of this technique is broad, from biochemistry up to organic electronics. In this work, we are presenting the method of analysis of organic compounds for optoelectronic applications.

Electrochemical impedance spectroscopy (EIS) of species that undergo reversible oxidation or reduction in solution was used for determination of rate constants of oxidation or reduction.

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). ...

Color Spot Test As a Presumptive Tool for the Rapid Detection of Synthetic Cathinones

Synthetic cathinones are a large class of new psychoactive substances (NPS) that are increasingly prevalent in drug seizures made by law enforcement and other border protection agencies globally. Color testing is a presumptive identification technique indicating the presence or absence of a particular drug class using rapid and uncomplicated chemical methods. Owing to their relatively recent emergence, a color test for the specific identification of synthetic cathinones is not currently available. In this study, we introduce a protocol for the presumptive identification of synthetic cathinones, employing three aqueous reagent solutions: copper(II) nitrate, 2,9-dimethyl-1,10-phenanthroline (neocuproine) and sodium acetate. Small pin-head sized amounts (approximately 0.1-0.2 mg) of the suspected drugs are added to the wells of a porcelain spot plate, and each reagent is then added dropwise sequentially before heating on a hotplate. A color change from very light blue to yellow-orange after 10 min indicates the likely presence of synthetic cathinones. The highly stable and specific test reagent has the potential for use in the presumptive screening of unknown samples for synthetic cathinones in a forensic laboratory. However, the nuisance of an added heating step for the color change result limits the test to laboratory application and decreases the likelihood of an easy translation to field testing.

Here we present a simple, inexpensive, and selective chemical spot test protocol for the detection of synthetic cathinones, a class of new psychoactive substances. The protocol is suitable for use in various areas of law enforcement that encounter illicit material.

Synthetic cathinones are a large class of new psychoactive substances (NPS) that are increasingly prevalent in drug seizures made by law enforcement and ...

Solid-phase Synthesis of [4.4] Spirocyclic Oximes

A convenient synthetic route for spirocyclic heterocycles is well sought after due to the molecule&#39;s potential use in biological systems. By means of solid-phase synthesis, regenerating Michael (REM) linker strategies, and 1,3-dipolar cycloaddition, a library of structurally similar heterocycles, both with and without a spirocyclic center, can be constructed. The main advantages of the solid-support synthesis are as follows: first, each reaction step can be driven to completion using a large excess of reagents resulting in high yields; next, the use of commercially available starting materials and reagents keep the costs low; finally, the reaction steps are easy to purify via simple filtration. The REM linker strategy is attractive because of its recyclability and traceless nature. Once a reaction scheme is completed, the linker can be reused multiple times. In a typical solid-phase synthesis, the product contains either a part of or the whole linker, which can prove undesirable. The REM linker is &#34;traceless&#34; and the point of attachment between the product and the polymer is indistinguishable. The high diastereoselectivity of the intramolecular 1,3-dipolar cycloaddition is well documented. Limited by the insolubility of the solid support, the reaction progression can only be monitored by a change in the functional groups (if any) via infrared (IR) spectroscopy. Thus, the structural identification of intermediates cannot be characterized by conventional nuclear magnetic resonance (NMR) spectroscopy. Other limitations to this method stem from the compatibilities of the polymer/linker to the desired chemical reaction scheme. Herein we report a protocol that allows for the convenient production of spirocyclic heterocycles that, with simple modifications, can be automated with high-throughput techniques.

Here we present a protocol to demonstrate an efficient method for the synthesis of spirocyclic heterocycles. The five-step process utilizes solid-phase synthesis and regenerating Michael linker strategies. Generally difficult to synthesize, we present a customizable method for the synthesis of spirocyclic molecules otherwise inaccessible to other modern approaches.

A convenient synthetic route for spirocyclic heterocycles is well sought after due to the molecule&#39;s potential use in biological systems. By means of ...

Solid Phase 11C-Methylation, Purification and Formulation for the Production of PET Tracers

Routine production of radiotracers used in positron emission tomography (PET) mostly relies on wet chemistry where the radioactive synthon reacts with a non-radioactive precursor in solution. This approach necessitates purification of the tracer by high performance liquid chromatography (HPLC) followed by reformulation in a biocompatible solvent for human administration. We recently developed a novel 11C-methylation approach for the highly efficient synthesis of carbon-11 labeled PET radiopharmaceuticals, taking advantage of solid phase cartridges as disposable &#34;3-in-1&#34; units for the synthesis, purification and reformulation of the tracers. This approach obviates the use of preparative HPLC and reduces the losses of the tracer in transfer lines and due to radioactive decay. Furthermore, the cartridge-based technique improves synthesis reliability, simplifies the automation process and facilitates compliance with the Good Manufacturing Practice (GMP) requirements. Here, we demonstrate this technique on the example of production of a PET tracer Pittsburgh compound B ([11C]PiB), a gold standard in vivo imaging agent for amyloid plaques in the human brains.

Solid Phase 11C-Methylation, Purification and Formulation for the Production of PET Tracers

We report an efficient carbon-11 radiolabeling technique to produce clinically relevant tracers for Positron Emission Tomography (PET) using solid phase extraction cartridges. 11C-methylating agent is passed through a cartridge preloaded with precursor and successive elution with aqueous ethanol provides chemically and radiochemically pure PET tracers in high radiochemical yields.

Routine production of radiotracers used in positron emission tomography (PET) mostly relies on wet chemistry where the radioactive synthon reacts with a ...

In situ FTIR Spectroscopy as a Tool for Investigation of Gas/Solid Interaction: Water-Enhanced CO2 Adsorption in UiO-66 Metal-Organic Framework

In situ infrared spectroscopy is an inexpensive, highly sensitive, and selective valuable tool to investigate the interaction of polycrystalline solids with adsorbates. Vibrational spectra provide information on the chemical nature of adsorbed species and their structure. Thus, they are very useful for obtaining molecular level understanding of surface species. The IR spectrum of the sample itself gives some direct information about the material. General conclusions can be drawn concerning hydroxyl groups, some stable surface species and impurities. However, the spectrum of the sample is &#34;blind&#34; with respect to the presence of coordinatively unsaturated ions and gives rather poor information about the acidity of surface hydroxyls, species decisive for the adsorption and catalytic properties of the materials. Furthermore, no discrimination between bulk and surface species can be made. These problems are solved by the use of probe molecules, substances that interact specifically with the surface; the alteration of some spectral features of these molecules as a result of adsorption provides valuable information about the nature, properties, location, concentration, etc., of the surface sites.
The experimental protocol for in-situ IR studies of gas/sample interaction includes preparation of a sample pellet, activation of the material, initial spectral characterization through the analysis of the background spectra, characterization by probe molecules, and study of the interaction with a particular set of gas mixtures. In this paper we investigate a zirconium terephthalate metal organic framework, Zr6O4(OH)4(BDC)6 (BDC = benzene-1,4-dicarboxylate), namely UiO-66 (UiO refers to University of Oslo). The acid sites of the UiO-66 sample are determined by using CO and CD3CN as molecular probes. Furthermore, we have demonstrated that CO2 is adsorbed on basic sites exposed on dehydroxylated UiO-66. Introduction of water to the system produces hydroxyl groups acting as additional CO2 adsorption sites. As a result, CO2 adsorption capacity of the sample is strongly enhanced.

In situ FTIR Spectroscopy as a Tool for Investigation of Gas/Solid Interaction: Water-Enhanced CO2 Adsorption in UiO-66 Metal-Organic Framework

The use of FTIR spectroscopy for investigation of the surface properties of polycrystalline solids is described. Preparation of sample pellets, activation procedures, characterization with probe molecules and model studies of CO2 adsorption are discussed.

In situ infrared spectroscopy is an inexpensive, highly sensitive, and selective valuable tool to investigate the interaction of polycrystalline solids ...

Gas Chromatography-Mass Spectrometry Paired with Total Vaporization Solid-Phase Microextraction as a Forensic Tool

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Chapters in this video

Large Scale Non-targeted Metabolomic Profiling of Serum by Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS)

Synthetic Methodology for Asymmetric Ferrocene Derived Bio-conjugate Systems via Solid Phase Resin-based Methodology

Visualization of Ambient Mass Spectrometry with the Use of Schlieren Photography

Coulomb Explosion Imaging as a Tool to Distinguish Between Stereoisomers

Protocol for the Solid-phase Synthesis of Oligomers of RNA Containing a 2'-O-thiophenylmethyl Modification and Characterization via Circular Dichroism

Electrochemical Impedance Spectroscopy as a Tool for Electrochemical Rate Constant Estimation

Color Spot Test As a Presumptive Tool for the Rapid Detection of Synthetic Cathinones

Solid-phase Synthesis of [4.4] Spirocyclic Oximes

Solid Phase ¹¹C-Methylation, Purification and Formulation for the Production of PET Tracers

In situ FTIR Spectroscopy as a Tool for Investigation of Gas/Solid Interaction: Water-Enhanced CO₂ Adsorption in UiO-66 Metal-Organic Framework