Post-Lab 5: Gas Chromatography: Blood Alcohol Content (BAC) Analysis: Determination of Ethanol Concentration in a Blood Sample PDF

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Post lab 5 for CHM 3120C...

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Post-Lab 5: Gas Chromatography: Blood Alcohol Content (BAC) Analysis: Determination of Ethanol Concentration in a Blood Sample Christian Claros 3/18/21 CHM 3120C – 002 Group A Quantitative Observation Area Ratio =

𝐴𝑟𝑒𝑎 𝑈𝑛𝑑𝑒𝑟 𝐸𝑡ℎ𝑎𝑛𝑜𝑙 𝑃𝑒𝑎𝑘

463321

𝐴𝑟𝑒𝑎 𝑈𝑛𝑑𝑒𝑟 𝐼𝑆 𝑃𝑒𝑎𝑘

125011

y = 762344x + 27121

= 3.706

463321 = 762344x + 27121 490442 = 762344x x = 0.643

% error =

|𝑣𝑎𝑙𝑢𝑒 𝑜𝑏𝑠𝑒𝑟𝑣𝑒𝑑 − 𝑒𝑥𝑒𝑐𝑡𝑒𝑑 𝑣𝑎𝑙𝑢𝑒| 𝑒𝑥𝑝𝑒𝑐𝑡𝑒𝑑 𝑣𝑎𝑙𝑢𝑒

× 100 %

𝑔 𝑑𝐿

|0.643 − 0.63| 0.63

× 100 % = 2.06 %

Table 1. Flas k

Estimated Concentratio n of Ethanol 𝑔 ( ) 𝑑𝐿

1

0.63

Ethanol Retentio n Time (Min)

1.112

46332 1 33636 8 24460 4 13411 5

1.01

35011

1.074 2

0.32 1.106

3

0.24 1.105

4 5

Area Under Ethano l Peak (1)

0.12

IS Retentio n time (Min)

1.812 1.844 1.843 1.848

0.06 1.841

Area under IS Peak (2)

Area Rati o (1) ÷ (2)

Actual Ethanol Concentratio 𝑔 n * 𝑑𝐿

% error (%)

12501 1 15678 4 14660 6 13310 7 13411 5

3.70 6 2.14 5 1.66 8 1.00 8 0.26 1

0.643

2.06

0.406

26.86

0.285

18.75

0.140

16.67

0.010

83.33

6

0.03

U

1.01 1.109

N/A

B

N/A

1.838

16589 33454 4 26928 5

12872 6 12132 8 -

1.839 1.848 -

0.12 9 2.75 7 -

The best fit line for ethanol (y = mx + b): y = 762344x + 27121 Concentration of the ethanol in the unknown blood sample: 0.318

𝑔 𝑑𝐿

Graph 1. External Standardization-based Calibration Curve

Area Under Ethanol Peak (1)

600000 y = 762344x + 27121 R² = 0.9343

500000 400000 300000 200000 100000 0 0

0.1

0.2

0.3

0.4

0.5

Estimated Concentration of Ethanol (g/dL)

Table 2. Data points for graph 1 Estimated Area Concentration Under of Ethanol Ethanol (g/dL) Peak (1) 0.63 463321 0.32 336368 0.24 244604 0.12 134115 0.06 35011 0.03 16589

0.6

0.7

-0.014 0.403

146.6 7 N/A

0.318

N/A

Precision R² = 0.9343

Accuracy % error = Flask

|𝑣𝑎𝑙𝑢𝑒 𝑜𝑏𝑠𝑒𝑟𝑣𝑒𝑑 − 𝑒𝑥𝑒𝑐𝑡𝑒𝑑 𝑣𝑎𝑙𝑢𝑒| 𝑒𝑥𝑝𝑒𝑐𝑡𝑒𝑑 𝑣𝑎𝑙𝑢𝑒

Estimated Actual Concentration Ethanol of Ethanol Concentration 𝑔 𝑔 ( ) * 𝑑𝐿 𝑑𝐿

× 100 % % error (%)

1

0.63

0.643

2.06

2

0.32

0.406

26.86

3

0.24

0.285

18.75

4

0.12

0.140

16.67

5

0.06

0.010

83.33

6

0.03

-0.014

146.67

U

N/A

0.403

N/A

B

N/A

0.318

N/A

|0.643 − 0.63| 0.63

× 100 % = 2.06 %

Research Question 1. (¶) Provide a component diagram sketch of a typical gas chromatography instrument setup and discuss the importance of each component.

The gas cylinder, regulator, and traps are used to hold the gas that will transport the sample through the system. The autosampler or syringe is how you get the sample into the system. The inlet allows the sample to be injected into the system without losing the mobile phase. The column is where the sample goes to elute in the oven during the stationary phase. The detector gives the signal from the elution of the sample from the chemical components to the computer where the chromatogram is produced from the data. (1)

2. What is an internal standard and how is it utilized?

Internal standard is a component in chromatography used to determine concentrations of analytes by calculating a response factor. To utilize an internal standard, you need to add a known amount to each sample you are testing, then the calibration will use the ration of the responses between the analyte and the internal standard. (1)

3. Describe how an FID detector and one other commonly used detector operate and what type of analytes they are useful for detecting.

Flame ionization detector measure analytes in gas streams. The ions formed are detected during the combustion of the organic compounds in a flame of hydrogen. (3) A katharometer is a chemical specific detector that senses changes in thermal conductivity of the eluent column which is compared to the carrier gas. They can be used in lung function testing, detecting helium loss in a helium vessel, and quantify the Caloric amount of methane in biogas samples. (4)

4. In a calibration curve, you obtain the best fit line and R2 value. What does the R2 value tell you about your data? The R2 value is the coefficient determination, it is the proportion of the dependent variable that can be predicted by the best fit line. If an R2 value is 0.9000 it means that 90% of the values for x would wall onto the line for y.

5. (¶) In the context of this experiment, GC is used for forensic analysis but is applicable to many other fields. Perform a literature search for new advancements in GC techniques. Referencing one paper, summarize the work and make sure to provide proper ACS citations.

The identification of full complement of metabolites in a biological sample is the goal of metabolomics. Innovations in liquid chromatography and gas chromatography column technologies improved the numbers of metabolites that can be detected using mass spectrometry. Each of the methods extends the coverage of the metabolome by the number of metabolites maintained and divided in the system. The new advancements allow the capabilities for functional analysis of metabolome by combining of the methodologies. Metabolomics will reach its potential as omics technique, giving an analytical tool for the illumination of new insights into biology systems through the instrumental and technological advancements. (5)

References: (1)

Technology Networks. Gas chromatography – how a gas chromatography machine works, how to read a chromatograph and GCxGC https://www.technologynetworks.com/analysis/articles/gas-chromatography-how-a-gaschromatography-machine-works-how-to-read-a-chromatograph-and-gcxgc-335168 (accessed Mar 18, 2021).

(2)

When Should an Internal Standard be Used? https://www.chromatographyonline.com/view/when-should-internal-standard-be-used-0 (accessed Mar 18, 2021).

(3)

The flame ionization detector https://www.chromatographyonline.com/view/flameionization-detector (accessed Mar 18, 2021).

(4)

Daynes, H. A.; Shakespear, G. A. The Theory of the Katharometer. Proc. R. Soc. Lond. A Math. Phys. Sci. 1920, 97 (685), 273–286.

(5)

Haggarty, J.; Burgess, K. E. V. Recent Advances in Liquid and Gas Chromatography Methodology for Extending Coverage of the Metabolome. Curr. Opin. Biotechnol. 2017, 43, 77–85....