LAAN-J-MS-E139
GC-MS
Gas Chromatograph Mass Spectrometer
Analysis of Volatile Toxic Substances
Using Headspace GC/MS Part.1
139 - Paint Thinner and Alcohol -
Forensic toxicologists in the police laboratories and forensic medicine departments of university measure a variety
of volatile substances in the course of investigating accidents, crimes, and other incidents.
Blood alcohol (ethanol) is measured to provide evidence in cases of traffic accidents caused by drinking, incidents
involving alcohol such as physical assault and injury, and acute alcohol poisoning.
Paint thinner is a solvent with toluene, methanol, and ethyl acetate as its main constituents. Paint thinner also has
anesthetic and stimulant properties. Paint thinner abuse by inhalation has become prevalent and laws have been
enacted to prevent the harmful effects of its abuse.
Used in industrial applications, cyanide and azide are compounds that are relatively easy to procure. This has
resulted in incidents of contamination by these toxic substances. After these incidents, testing regimes for poisons
were enhanced in order to determine the cause of such incidents, i.e. crime or suicide.
Blood alcohol and paint thinner can be measured relatively simply using headspace sampler, with measurements
performed on a routine basis at police laboratories and in university forensic departments.
There have been reports of the headspace method being used to measure cyanide and azide. These compounds
are normally measured by performing PFB derivatization, solvent extraction, and then liquid injection for GC/MS
analysis. However, the derivatization and extraction steps of this method are labor-intensive.
This two-part application presents details on the investigation of using headspace GC/MS to measure cyanide and
azide, as well as information on optimizing column conditions to allow simultaneous measurement of alcohol and
paint thinner. Of these two investigations, results obtained from measuring blood alcohol and paint thinner are
presented below. Please see Application Data Sheet No. 140 for part two.
Sample Preparation
Blood Ethanol Analysis
An aqueous solution of 1-propanol (0.5 mg/mL), which was to be added to blood samples as the internal standard,
was prepared by dissolving 1-propanol in distilled water. Standards for calibration curve of blood ethanol were
prepared by making up samples of hemolyzed equine blood with ethanol concentrations of 0.03, 0.1, 0.3, 1.0, and
2.0 mg/mL, then adding 0.5 mL of each of these samples and 0.5 mL of 1-propanol (0.5 mg/mL) internal standard
to a 20 mL headspace vial. Each vial was quickly sealed with a headspace cap and then agitated. To confirm that
2-propanol, which is used for sterilization during blood sampling, is separated from ethanol and 1-propanol on a
chromatogram, an aqueous solution of ethanol, 2-propanol, and 1-propanol was also prepared.
Paint Thinner Analysis
Paint thinner (5 L) was added to a 20 mL headspace vial, which was quickly sealed with a headspace cap.
HS-20 Headspace Sampler + GCMS-QP2020
139
Analytical Conditions
Table 1 shows the headspace and GC/MS analytical conditions. Alcohol, cyanide, and azide were all measured
using the same headspace and GC/MS conditions, and only paint thinner was measured with different conditions
after changing the split ratio and detector voltage. With the HS-20 headspace sampler, even when GC/MS
analysis conditions are changed, measurements can be performed within the same batch file by switching
methods, as long as the same headspace conditions are used. Changing the split ratio is an effective way of
analyzing both cyanides and azides that must be measured at trace quantities, and undiluted thinner solution that
contains high-concentration constituents.
For this application, results were collected using the GCMS-TQ8040 GC-MS/MS device, though the same
results can be obtained using the GCMS-QP2020 single-GC/MS device.
Table 1: Analytical Conditions
HS: HS-20
GC-MS: GCMS-TQ8040
[HS] [GC]
Headspace mode: Loop Column: Rtx-BAC2 (length: 30 m, 0.32 mm I.D., df = 1.2 m, Restek
Oven temp.: 60 C Corporation)
Sample line temp.: 100 C Column oven temp.: 40 C (5 min) (40 C /min) 200 C (1 min)
Transfer line temp.: 150 C Carrier gas: Helium
Vial pressurization gas pressure: 70 kPaCarrier gas control: Linear velocity (62.5 cm/sec)
Vial warming time: 10 min Injection mode: Split
Vial pressurization time: 0.5 min Split ratio: 10:1 (alcohol, cyanide, azide)
Loading time: 0.5 min 30:1 (paint thinner)
Loading equalization time: 0 min [MS]
Injection time: 0.5 min Interface temp.: 230 C Ion source temp.: 200 C
Needle flush time: 5 min Solvent elution time: 0.7 min Data acquisition time: 1 - 10 min
GC cycle time: 18 min Measurement mode: Scan Mass range: m/z 10 - 300
Event time: 0.2 sec Emission current: 60 A (standard)
*Note: The detection voltage and other conditions must be optimized since they can differ depending on equipment status.
Analytical Results of Blood Ethanol
Fig. 1 shows the total ion current (TIC) chromatogram obtained when analyzing a standard of 0.3 mg/mL ethanol
to which 2-propanol had been added—a compound that is used for sterilization during blood collection. The
chromatogram shows complete separation of 2-propanol and that quantitative values were not affected by 2-
propanol.
(1,000,000)
TIC
5.0 3. 1-propanol
(ISTD)
4.0
3.0 2. 2-propanol
2.0
1. Ethanol
1.0
1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50
Fig. 1: Total Ion Current Chromatogram of 2-Propanol Added to 0.3 mg/mL Ethanol Standard
139
The chromatograms obtained when analyzing a blank and a 0.03 mg/mL standard are shown in Fig. 2. The
calibration curve (0.03, 0.1, 0.3, 1.0, 2.0 mg/mL) obtained after internal standard correction is shown in Fig. 3.
The calibration curve correlation coefficient (R) was 0.9999 or above, showing that linearity was good.
Table 2 shows the reproducibility (n = 8) of measuring 0.3 mg/mL, which is the reference concentration used as
the basis of the breath test in Japan's Road Traffic Act and Order for Enforcement of the Road Traffic Act. Table
3 shows the reproducibility (n = 8) of measuring 0.03 mg/mL, which is 1/10 the reference concentration. The
accuracy of quantitative measurements of the 0.3 mg/mL concentration sample was 100.2 - 100.9 % (average
100.5 %) with a reproducibility relative standard deviation (%RSD) of about 0.2 %. The accuracy of quantitative
measurements of the 0.03 mg/mL concentration sample was 89.4 - 97.1 % (average 95.2 %) with a
reproducibility relative standard deviation (%RSD) of about 2.8 %.
(10,000) (10,000)
45.00 45.00
31.00 31.00
7.5 7.5
5.0 5.0
2.5 2.5
0.0 0.0
1.75 2.00 2.25 2.50 1.75 2.00 2.25 2.50
Fig. 2: Mass Chromatograms of Ethanol when Analyzing a Blank Sample and a 0.03 mg/mL Standard Sample
Left: Blank, Right: 0.03 mg/mL Standard
Area Ratio
R2 = 0.99987
3.0 R = 0.99993
2.5
2.0
1.5
1.0
0.5
0.0
0.00 0.50 1.00 1.50 Concentration Ratio
Fig. 3: Ethanol Calibration Curve (Concentration: 0.03 - 2.0 mg/mL)
Table 2: Reproducibility for 0.3 mg/mL (n = 8) Table 3: Reproducibility for 0.03 mg/mL (n = 8)
Concentration Accuracy Concentration Accuracy
Area Ratio Area Ratio
(mg/mL) (%) (mg/mL) (%)
1st 0.502 0.302 100.5 1st 0.130 0.029 96.1
2nd 0.501 0.301 100.4 2nd 0.131 0.029 96.8
3rd 0.500 0.301 100.2 3rd 0.131 0.029 96.8
4th 0.504 0.302 100.8 4th 0.129 0.029 95.5
5th 0.504 0.303 100.9 5th 0.131 0.029 97.1
6th 0.503 0.302 100.7 6th 0.131 0.029 97.1
7th 0.502 0.301 100.4 7th 0.121 0.027 89.4
8th 0.502 0.301 100.5 8th 0.126 0.028 92.9
Average 0.502 0.302 100.5 Average 0.129 0.029 95.2
Standard Deviation Standard Deviation
0.001 0.001 0.227 0.004 0.001 2.716
(SD) (SD)
%RSD 0.226 0.226 0.226 %RSD 2.849 2.853 2.853
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Analytical Results of Paint Thinner
The total ion current chromatogram obtained after analyzing commercially available paint thinner is shown in
Fig. 4. The mass spectra of each constituent of paint thinner (methanol, ethyl acetate, and toluene) are shown in
Fig. 5. The analysis conditions used in this investigation allowed for separation of the three main constituents of
paint thinner (methanol, ethyl acetate, and toluene) in 10 minutes.
(1,000,000)
8.0 TIC
3. Toluene
7.0 2. Ethyl acetate
6.0
5.0 1. Methanol
4.0
3.0
2.0
1.0
1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0
Fig. 4: Total Ion Current Chromatogram Obtained after Analysis of Undiluted Paint Thinner
1. Methanol 2. Ethyl acetate
% %
31 43
100 100
50 29 50
45 61 70
29 88
0 0
25 50 75 100 25 50 75 100
m/z m/z
3. Toluene
%
91
100
50
65
39 45 51
0
25 50 75 100
m/z
Fig. 5: Mass Spectra of Main Constituents Obtained after Analysis of Paint Thinner
GCMS-TQ and GCMS-QP are registered trademarks of Shimadzu Corporation.
Rtx is a registered trademark of Restek Corporation.
First Edition: March, 2018
Shimadzu Corporation
www.shimadzu.com/an/
© Shimadzu Corporation, 2018
