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Report TemplateEXPERIMENT 2: ANALYSIS OF TOLUENE IN PETROL BY GAS CHROMATOGRAPHY-MASS SPECTROMETRY (GC-MS)IntroductionThe aim of this experiment is to accurately identify and determine toluene concentration in petrol using gas chromatography-mass spectrometry (GC-MS). Gas chromatography is a techniq...

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Report Template EXPERIMENT 2: ANALYSIS OF TOLUENE IN PETROL BY GAS CHROMATOGRAPHY-MASS SPECTROMETRY (GC-MS) Introduction The aim of this experiment is to accurately identify and determine toluene concentration in petrol using gas chromatography-mass spectrometry (GC-MS). Gas chromatography is a technique which is used to separate compounds in a sample by the vaporising the sample using heat, it then carries this gas to a GC column which is coated with a stationary phase that separates these compounds depending on the polarity of each part. They then leave the GC column and move to the mass spectrometer (MS). An electron beam at 70ev bombards the toluene molecule, results in a charged molecular ion. These fragments are then analysed by a quadrupole MS. Selected ion monitoring is also performed in this experiment to increase the selectivity for toluene. This experiment consists of preparing a blank, toluene standards and diluting unknown samples to measure in the GC-MS. A calibration curve is constructed with the results of the standards for peak area vs toluene concentration and peak height concentration. These calibration curves will then be used to calculate the concentrations of toluene in the unknown samples and these concentrations will have to be converted from ppm to % v/v. Experimental GC column full description: DB5 – MS, 4mm ultra-inert Liner (quartz wool), 30m x 0.25mm (0.25𝜇m film). 5% phenyl + 95% PMS Temperature program full description: Range of temperature -60℃ à 280℃. For final analysis: 40℃ for 4 minutes and increases by 30℃ per minute for 6 minutes until 180℃ is reached then held for 1 minute. Split ratio: 100:1 Injection volume: 1𝜇L Injector temperature: 250℃ Helium flow rate: 1mL/min

Results Part 1: Standard blank Abundance

TIC: Blank.D\data.ms .634

75000

70000

65000

60000

55000

50000

45000

40000

35000

30000

25000

20000

15000

10000

5000

Time-->

1.60 1.80 2.00 2.20 2.40 2. 60 2.80 3.00 3.20 3.40 3.60 3. 80 4.00 4.20 4.40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6.00 6.20

Figure 1: Standard blank GC mass chromatogram.

Part 2: Determination of SIM parameters and library search Abundance

TIC: 80 ppm.D\data.ms .635

Abundance

4.346

Ion 91.00 (90.70 to 91.30): 80 ppm.D\data.ms 4.346

35000 75000

30000 70000

25000 65000

20000 60000

15000 55000

10000 50000

5000 45000

40000

0 Time--> 1.60 1.80 2.00 2.20 2.40 2. 60 2.80 3.00 3.20 3.40 3.60 3. 80 4.00 4.20 4.40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6. 00 6.20 Abundance Ion 92.00 (91.70 to 92.30): 80 ppm.D\data.ms

35000 35000

30000 30000

25000 25000

20000 20000

15000 15000

10000 10000

5000 5000

Time-->

1.60 1.80 2.00 2.20 2.40 2. 60 2.80 3.00 3.20 3.40 3.60 3. 80 4.00 4.20 4. 40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6.00 6. 20

Figure 2: Total ion chromatogram for 80ppm standard

0 Time--> 1.60 1.80 2.00 2.20 2.40 2. 60 2.80 3.00 3.20 3.40 3.60 3. 80 4.00 4.20 4.40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6. 00 6.20

Figure 3: Extracted ion chromatogram for 80ppm standard (91 and 92m/z)

Abundance

Scan 1071 (4.346 min): 80 ppm.D\data.ms 91

Abundance

TIC: Sample A.D\data.ms

36000

75000 34000

70000 32000

65000 30000

60000

28000

26000

55000

24000

50000 22000

45000 20000

40000 18000

35000 16000

14000

30000

12000

25000

10000

20000 8000

15000 6000

10000 65

4000

4.348

5.946 6.222

5000

2000

51

5.850 56

0 m/z-->

45

50

55

61 60

68 65

70

73

77 75

82 80

86 85

90

95

100 105 110

114 120 124 128 132135 139

95

100 105 110 115 120 125 130 135 140 145 150 155

6.829 6.899 7.168 7.057

145 149

Time-->

Figure 4: Representative mass spectrum of toluene standard (80ppm) Abundance

Ion 91.00 (90.70 to 91.30): Sample B.D\data.ms

12000 11000 10000 9000 8000 7000 6000 5000 4000 3000 2000 1000

2.00

2.50

3.00

3.50

4.00

2.00

2.50

3.00

3.50

4.00

4.50 5.00 5.50 6.00 6.50 7. 00 Ion 92.00 (91.70 to 92.30): Sample B.D\data.ms

7.50

8.00

8.50

9.00

9.50

7.50

8.00

8.50

9.00

9.50

13000 12000 11000 10000 9000 8000 7000 6000 5000 4000 3000 2000 1000 0 Time-->

4.50

5.00

5.50

6.00

6.50

7. 00

2.50

3.00

3.50

4.00

4.50

5.00

5.50

6.00

6.50

7.00

7.50

Figure 5: Representative TIC of Sample A

13000

0 Time--> Abundance

2.00

Figure 6: Representative EIC of Sample B (91 and 92 m/z)

8.00

8.50

9.00

9.50

Which ions have you chosen for SIM (selected ion monitoring) mode? Identify the molecular ion and fragment ion. 92 m/z à as shown in figure 7, this is the 91m/z à as shown in figure 8, this is the molecular ion where an electron is fragment ion where the electron and a removed, and it shows at 92m/z. hydrogen are removed.

Figure 7: Molecular ion of toluene. Drawn with MarvinSketch V20.1

Figure 8: Fragment ion of toluene. Drawn with MarvinSketch V20.1

Part 3. Construction of calibration curve and analysis of unknown Abundance

Ion 91.00 (90.70 to 91.30): 80 ppm.D\data.ms 4.346

35000

Abundance

Ion 91.00 (90.70 to 91.30): Sample B.D\data.ms

13000 12000

30000

11000 10000

25000

9000 8000

20000 7000 6000

15000 5000

10000

4000 3000

5000

2000 1000

0 Time--> 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20 4.40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6.00 6.20 Abundance Ion 92.00 (91.70 to 92.30): 80 ppm.D\data.ms

35000

0 Time--> Abundance

2.00

2.50

3.00

3.50

4.00

2.00

2.50

3.00

3.50

4.00

4.50 5.00 5.50 6.00 6.50 7.00 Ion 92.00 (91.70 to 92.30): Sample B.D\data.ms

7.50

8.00

8.50

9.00

9.50

7.50

8.00

8.50

9.00

9.50

13000 12000

30000

11000 10000

25000

9000 8000

20000

7000 6000

15000 5000 4000

10000 3000

5000

2000 1000

Time-->

0 1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20 4.40 4.60 4.80 5.00 5.20 5.40 5.60 5.80 6.00 6.20

Figure 9: Representative chromatogram SIM for 80ppm.

0 Time-->

4.50

5.00

5.50

6.00

6.50

7.00

Figure 10: Representative chromatogram of SIM for Sample B

Abundance

TIC: Sample A.D\data.ms

75000

70000

65000

60000

55000

50000

45000

40000

35000

30000

25000

20000

15000

10000 4.348

5.946 6.222

5000 5.850

6.829 6.899 7.168 7.057 Time-->

2.00

2.50

3.00

3.50

4.00

4.50

5.00

5.50

6.00

6.50

7.00

7.50

8. 00

8.50

9.00

9.50

Figure 11: TIC of unknown sample A.

Part 4: Data processing and reporting Table 1: Table containing raw data for GC-MS of toluene Sample Identity

toluene retention time (min)

toluene area (A min)

toluene height (A)

10 ppm toluene

4.344

86153

33727

20 ppm toluene

4.344

169527

68134

40 ppm toluene

4.344

361429

147654

80 ppm toluene

4.341

671540

284421

120 ppm toluene

4.341

1037485

440622

Unknown Sample A

4.348

12439

6465

Unknown Sample B

4.341

48817

28363

A = abundance

Calibration curve

Toluene peak area vs concentration 1200000 y = 8572x + 2336.5 R² = 0.999

Peak area/A min

1000000 800000 600000 400000 200000 0 0

20

40

60

80

100

120

140

Concentration/ppm

Figure 12: Calibration curve for toluene peak area vs concentration

Toluene peak height vs concentration 500000

y = 3678.9x - 3748 R² = 0.9994

450000 400000

Peak height/A

350000 300000 250000 200000 150000 100000 50000 0 0

20

40

60

80

100

120

Concentration/ppm

Figure 13: Calibration curve for toluene peak height vs concentration

140

Determination of unknown Sample A diluted concentration calculations:

1. Peak area vs. concentration 𝑦 = 8572𝑥 + 2336.5 𝐸𝑞𝑢𝑎𝑡𝑖𝑜𝑛6𝑜𝑏𝑡𝑎𝑖𝑛𝑒𝑑6𝑓𝑟𝑜𝑚6𝑐𝑎𝑙𝑖𝑏𝑟𝑎𝑡𝑖𝑜𝑛6𝑐𝑢𝑟𝑣𝑒6(𝑓𝑖𝑔𝑢𝑟𝑒612) 𝑥=

𝑦 − 2336.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚

(𝑆𝑎𝑚𝑝𝑙𝑒6𝐴) = 124396𝐴6𝑚𝑖𝑛 𝑦 → 𝑝𝑒𝑎𝑘6𝑎𝑟𝑒𝑎6𝑓𝑜𝑟6𝑡𝑜𝑙𝑢𝑒𝑛𝑒6𝑓𝑜𝑢𝑛𝑑6𝑖𝑛 6𝑡𝑎𝑏𝑙𝑒616 𝑥=

123496𝐴6𝑚𝑖𝑛 − 2336.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚 𝑥=

10012.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚

𝑥 = 1.1686𝑝𝑝𝑚

2. Peak height vs. concentration 𝑦 = 3678.9𝑥 − 3748 𝐸𝑞𝑢𝑎𝑡𝑖𝑜𝑛6𝑜𝑏𝑡𝑎𝑖𝑛𝑒𝑑6𝑓𝑟𝑜𝑚6𝑐𝑎𝑙𝑖𝑏𝑟𝑎𝑡𝑖𝑜𝑛6𝑐𝑢𝑟𝑣𝑒6(𝑓𝑖𝑔𝑢𝑟𝑒613) 𝑥=

𝑦 + 37486𝐴 𝐴 3678.96 𝑝𝑝𝑚

(𝑆𝑎𝑚𝑝𝑙𝑒6𝐴 ) = 64656𝐴 𝑦 → 𝑝𝑒𝑎𝑘6ℎ𝑒𝑖𝑔ℎ𝑡6𝑓𝑜𝑟6𝑡𝑜𝑙𝑢𝑒𝑛𝑒6𝑓𝑜𝑢𝑛𝑑6𝑖𝑛 6𝑡𝑎𝑏𝑙𝑒 616 𝑥=

64656𝐴 + 37486𝐴 𝐴 3678.96 𝑝𝑝𝑚

𝑥=

102136𝐴 𝐴 3678.96𝑝𝑝𝑚

𝑥 = 2.7766𝑝𝑝𝑚

Sample B diluted concentration calculations: 1. Peak area vs. concentration 𝑦 = 8572𝑥 + 2336.5 𝐸𝑞𝑢𝑎𝑡𝑖𝑜𝑛6𝑜𝑏𝑡𝑎𝑖𝑛𝑒𝑑6𝑓𝑟𝑜𝑚6𝑐𝑎𝑙𝑖𝑏𝑟𝑎𝑡𝑖𝑜𝑛6𝑐𝑢𝑟𝑣𝑒6(𝑓𝑖𝑔𝑢𝑟𝑒612) 𝑥=

𝑦 − 2336.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚

(𝑆𝑎𝑚𝑝𝑙𝑒6𝐵) = 488176𝐴6𝑚𝑖𝑛 𝑦 → 𝑝𝑒𝑎𝑘6𝑎𝑟𝑒𝑎6𝑓𝑜𝑟6𝑡𝑜𝑙𝑢𝑒𝑛𝑒6𝑓𝑜𝑢𝑛𝑑6𝑖𝑛 6𝑡𝑎𝑏𝑙𝑒616 𝑥=

488176𝐴6𝑚𝑖𝑛 − 2336.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚 𝑥=

46480.56𝐴6𝑚𝑖𝑛 𝐴6𝑚𝑖𝑛 85726 𝑝𝑝𝑚

𝑥 = 5.4226𝑝𝑝𝑚

2. Peak height vs. concentration 𝑦 = 3678.9𝑥 − 3748 𝐸𝑞𝑢𝑎𝑡𝑖𝑜𝑛6𝑜𝑏𝑡𝑎𝑖𝑛𝑒𝑑6𝑓𝑟𝑜𝑚6𝑐𝑎𝑙𝑖𝑏𝑟𝑎𝑡𝑖𝑜𝑛6𝑐𝑢𝑟𝑣𝑒6(𝑓𝑖𝑔𝑢𝑟𝑒613) 𝑥=

𝑦 + 37486𝐴 𝐴 3678.96 𝑝𝑝𝑚

𝑦 → 𝑝𝑒𝑎𝑘6ℎ𝑒𝑖𝑔ℎ𝑡6𝑓𝑜𝑟6𝑡𝑜𝑙𝑢𝑒𝑛𝑒6𝑓𝑜𝑢𝑛𝑑6𝑖𝑛6𝑡𝑎𝑏𝑙𝑒 (𝑆𝑎𝑚𝑝𝑙𝑒6𝐵) 616 = 283636𝐴 𝑥=

283636𝐴 + 37486𝐴 𝐴 3678.96𝑝𝑝𝑚

𝑥=

321116𝐴 𝐴 3678.96 𝑝𝑝𝑚

𝑥 = 8.7286𝑝𝑝𝑚

Table 2: Table containing calculations of original concentrations of both Sample A and Sample B and conversion to %v/v Peak area vs concentration

Peak height vs concentration

1.1686𝑝𝑝𝑚 ∗ 2000

2.7766𝑝𝑝𝑚 ∗ 2000

23366𝑝𝑝𝑚 𝑚𝑔 23366 𝐿

55526𝑝𝑝𝑚 𝑚𝑔 5552 𝐿

Sample A (1:2000)

𝐶𝑜𝑛𝑣𝑒𝑟𝑠𝑖𝑜𝑛6𝑓𝑟𝑜𝑚6𝑝𝑝𝑚6𝑡𝑜6%𝑣/𝑣 𝐶𝑜𝑛𝑣𝑒𝑟𝑠𝑖𝑜𝑛6𝑓𝑟𝑜𝑚6𝑝𝑝𝑚6𝑡𝑜6%𝑣/𝑣

23366

𝑚𝑔 16𝑚𝐿 6× 863.66𝑚𝑔 𝐿

𝑚𝐿 6 ÷ 1000 𝐿 𝐿 0.00273976 × 100 𝐿 2.7397

55526

𝑚𝑔 16𝑚𝐿 6× 863.66𝑚𝑔 𝐿

𝑚𝐿 6 ÷ 1000 𝐿 𝐿 0.00642896 × 100 𝐿 6.4289

0.2740%6𝑣/𝑣

0.6429%6𝑣/𝑣

5.4226𝑝𝑝𝑚 ∗ 1000

8.7286𝑝𝑝𝑚 ∗ 1000

54226𝑝𝑝𝑚 𝑚𝑔 5422 𝐿

87286𝑝𝑝𝑚 𝑚𝑔 8728 𝐿

Sample B (1:1000)

𝐶𝑜𝑛𝑣𝑒𝑟𝑠𝑖𝑜𝑛6𝑓𝑟𝑜𝑚6𝑝𝑝𝑚6𝑡𝑜6%𝑣/𝑣 𝐶𝑜𝑛𝑣𝑒𝑟𝑠𝑖𝑜𝑛6𝑓𝑟𝑜𝑚6𝑝𝑝𝑚6𝑡𝑜6%𝑣/𝑣

54226

16𝑚𝐿 𝑚𝑔 6× 𝐿 863.66𝑚𝑔

16𝑚𝐿 𝑚𝑔 6× 𝐿 863.66𝑚𝑔

𝐿 0.00627846 × 100 𝐿

𝑚𝐿 6 ÷ 1000 𝐿 𝐿 0.01010656 × 100 𝐿

0.6278%6𝑣/𝑣

1.011%6𝑣/𝑣

6.2784

𝑚𝐿

87286

𝐿

6 ÷ 1000

Density of Toluene = 863.6 mg/mL (SciFinder)

10.1065

Discussion Questions 1) Comment on any peaks present in the standard blank. Only the solvent peak is showing on the standard blank chromatogram. The solvent peak shows early at approximately 1.70 minutes. 2) Why is it important to perform the standard blank? It is important to perform a standard blank to understand what is in the background which consists of interferences such as contaminants and it also calibrates the chromatographer. We can also see where the solvent peak is and other possible contaminants. 3) Comment on the quality of your calibration curve, e.g. is it linear, does it pass though the origin, were there any outliers, etc. If the calibration curve is inadequate, provide possible explanations. For the toluene peak area vs concentration calibration curve (figure 12) the linearity is high as shown by the correlation coefficient (R2) of 0.999. It does not pass through the origin, however, passes through at 2336.5 A min, this is relatively close. The graph overall is of high quality as the correlation coefficient shows how well the concentration of toluene is proportional to the peak area and there are no outliers. For the toluene peak height vs concentration calibration curve (figure 13) the correlation coefficient is high with a value of 0.9994 it shows that the graph has a high linear graph. The y-intercept is further away from the peak area curve with a value of -3748 A min. This graph is of high quality due to the high correlation coefficient showing the relationship between concentration and peak height is strong. The high quality of these graphs show they are reliable for use to determine the unknown concentrations of the samples. 4) Is there a difference in your unknown concentration using the two methods? Which method is superior, peak height or peak area? Looking at table 2. For sample A, peak area and peak height values are 0.2740% v/v and 0.6429% v/v, respectively. These values are different. For sample B, peak area and peak height values are 0.6278% v/v and 1.011% v/v, respectively. These values also differ from each other. The peak area is known as the more superior method as it is less affected by interferences from the machine or the environment like the peak height is. affected Conditions such as temperature causes the peak heights to distort and can give incorrect values (Guiochon & Guillemin 1988).

5) Why did you use SIM mode for quantification of the petrol? Selected ion monitoring (SIM) is used for quantification of petrol because there are many compounds in petrol that can be detected by the mass spectrophotometer. Using SIM mode increases the selectivity of the mass spectrophotometer and speeds up the process instead of having to wait for the mass spectrometer to measure for other components of other compounds. 6) Do you think that it is acceptable to quantify in scan mode in combination with EIC? Scan mode used with extracted ion chromatogram (EIC) can be used to quantify a certain component depending on the mass over charge value for the desired ion/s. Though it will not be as sensitive and high quality as SIM mode, it can still be sensitive for the chosen values, therefore can be acceptable to use the values produced.

References Guiochon, G. & Guillemin, CL. (eds) 1988, 'Quantitative Analysis By Gas Chromatography Measurement of Peak Area and Derivation of Sample Composition', Journal of Chromatography Library, vol. 42, pp.629-659...