Caffeine Lab Report Final Draft Danella PDF

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Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

Extraction, Isolation and Identification of Caffeine using Gas Chromatography & Mass Spectrometry Coeli Danella Chem 110, Lab sec. 404 Spring 2020, University of Nebraska Lincoln

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

Abstract: There are four objectives of this caffeine lab: Use a solvent extraction method to separate a pure substance from a mixture, be able to determine the mass of the assigned caffeine sample through mass spectrometry and introduce gas chromatography as a way to separate compounds. Gas chromatography with mass spectrometry helps more accurately determine the caffeine sample.. Results from the sample of caffeine ended up being 71 mg/serving with a molar mass ranging from 194.17 – 55.09. Introduction: Caffeine is a member of the alkaloids family. Alkaloids are a natural nitrogen containing basic compound that are found in plants and typically have a bitter taste to them. Other compounds within the alkaloid family include nicotine, cocaine, and theobromine, however caffeine tends to be more stimulating than nicotine but less than that of cocaine (1). Caffeine tends to affect the blood sugar and the calcium-phosphate balance within the human body. A table below can show you the effects of caffeine in milligrams of the typical beverages that are consumed daily. Type of Caffeinated Drink in oz Amount in mg Coffee 100-150 Tea 60-75 Coca Cola 40-72 Coca 50 The focus of this lab experiment was to use a solvent extraction method to separate a pure substance from a mixture and then determine the mass of the sample through mass spectrometry. Extraction of the solvent happened through a separatory funnel using the sample and dichloromethane. After proper sit time for separation both the water and dichloromethane + sample, the bottom section (dichloromethane) in the funnel filtered into a pre-weighted beaker. From there the caffeine is isolated in the hood using a hot plate. After proper extraction, gas chromatography and mass spectrometry were used for analysis. The purpose of gas chromatography is to show how many different types of particles are in a solution as it moves its way through the column. From there a chromatogram is printed and allows the compounds (peaks on the chromatogram) to then be identified by a mass spectrometer. Mass spectrometry is a technique which involves measurements of mass-tocharge ratios of the ions formed when a sample is ionized (2). Therefore, the mass spectrometer allows analysis to include the parent molecule and fragmentation which explains the structure of said molecule. Taken together, the mass spectra and the chromatographic peaks allowed unambiguous identification of each component (3). Methods: Measured out 100 mL of caffeinated liquid sample into graduated cylinder and measured 2 grams of sodium carbonate on an electrical balance then added that to an Erlenmeyer flask . Poured the caffeine sample into a 500 mL Erlenmeyer flask and mixed solution until dissolved. Measured 25 mL of dichloromethane and once parafilm was prepared, added the dichloromethane to the sample and covered. Swirled the solution for about 7 minutes. Poured solution into funnel and left to sit and separate. Measured a 50 mL beaker and mass was recorded. The weighted beaker was placed underneath the funnel and liquid

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

extraction took place. Dichloromethane filtered out of the funnel. After all dichloromethane was in the beaker, isolation of caffeine continued under the hood. The beaker was placed on a hot plate measuring about 119F. Beaker remained on hot plate until only thin layer was left and then the mass was weighed. For gas chromatography and mass spectrometry analysis a vial sample from TA was received and 6 drops of DCM were added to sample beaker. A small sample from beaker was placed back into vial then returned back to TA for analysis in HaH 414. The actual procedure can be found in 110 Chemistry Lab Manual by J. Kautz & J. Periago. Results: Quantitative Analysis of Caffeine: Identity of Sample & Vial Number Volume of Sample Used mL Mass of Empty Beaker (g) Mass of Beaker & Caffeine (g) Mass of Caffeine (g) Mass of Caffeine per serving (mg/serving)

528 100.01 mL 27.166 g 27.237 g .071 g 71 mg/serving

Gas Chromatography Peak with Corresponding Retention Time & Identity of Molecule: Peaks Retention Time (min) Identity of Molecule 12 1 1 .705 minutes C8 H1014N416O2+ Mass Spectrum of Caffeine Results & Assigned Fragment Ions Peak m/Z Value A 194.17 B 165.15 C 137.16 D 109.14 E 82.11 F 67.08 G 55.09

Identified Fragment Ion C81H1014N416O2+ 12 1 C7 H914N416O+ 12 1 C6 H714N316O+ 12 1 C5 H714N316O 12 1 C4 H614N216O 12 1 C3 H314N216O 12 1 C3 H514N116O 12

Mass Spectrum of Dichloromethane Results and Assigned Fragment Ions Peak m/Z Value Identified Fragment Ion 12 1 17 A 82.95 C H Cl2 12 1 B 84.94 C H217Cl2 12 1 C 86.93 C H417Cl2 Discussion: The questions proposed at the beginning of the lab included, “Was the solid actually caffeine?” and “Was it only caffeine?” The results showed that final mass collected after evaporation was 71 mg of caffeine was in the sample. If referencing the introduction caffeine table, it’s plausible to identify that type of caffeine could be found in either tea or soda. The liquid-liquid extraction created two separate layers, one with the caffeine and an aqueous layer

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

without caffeine. Results showed that caffeine was found in the DCM layer because it’s more soluble in DCM. The DCM and caffeine have a stronger attraction to each other than they do to water. This is why extraction through the funnel was simple. The Gas Chromatography peak confirmed that the solute tested was just pure caffeine, due to the one peak in the graph, located at .705 minutes. This is also confirmed using gas chromatography because the DCM was more mobile than the caffeine leading to caffeine to remain stationary through the column. The mass spectrometry showed fragments for both caffeine and DCM as well as their charge ratio. For caffeine results showed seven peaks, while the DCM results had three, which was initially expected. Human error should be accounted for in calculations of isotopes in the results tables, but otherwise it seemed like the experiment accurately proved the mass was caffeine and only caffeine. Conclusion: At the conclusion of this experiment, the measured caffeine sample ended up being .071 grams. Through the gas spectrometry it was proven that with only one peak, there substance was pure caffeine. Overall the procedure ended up being quite simple throughout the caffeine experiment. Separation of the caffeinated liquid sample in the funnel was interesting to see live as well as understanding the difference in molecular weight leading to a separation of elements. Overall, the experiment ran smoothly and the measurements and techniques were carried out in the correct manner with precision. Sources Used:

1. Kautz, J. and Peragio, J. Chemistry 110 Lab Manual 2018-2019. Hayden-McNeil, LLC: Plymouth, 2018. 1. Kautz, J. and Peragio, J. Chemistry 110 Lab Manual

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

2018-2019. Hayden-McNeil, LLC: Plymouth, 2018. 1. Kautz, J. and Peragio, J. Chemistry 110 Lab Manual 2018-2019. Hayden-McNeil, LLC: Plymouth, 2018. 1. Kautz, J. and Peragio, J. Chemistry 110 Lab Manual 2019-2020. Hayden-McNeil, LLC: Plymouth 2020. 2. Hoffman, Edmond de. Mass Spectrometry. John Wiley & Sons, Inc. 2005. 3. Jones, Mark. Gas Chromatography-Mass Spectrometry: A National Historic Chemical Landmark 2019. ACS.org. Supporting Information:

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

Image 1: Mass Spectrum of Caffeine Results & Assigned Fragment Ions

Image 2: Gas Chromatography of Caffeine Sample with Corresponding Retention Time

Extraction, Isolation & Identification of Caffeine Using Gas Chromatography & Mass Spectrometry

C :\X calibu r\d ata \C H E M 110 \S um m er2 0 15 \1 0

7/2 9 /2 0 15 5 :1 6:47 P M

1 0 # 2 3 0 R T : 2 .2 6 A V : 1 N L : 4 .5 4 E 6 T : + c F u ll m s [ 5 0 .0 0 -6 5 0 .0 0 ] 5 0 .9 6 100 95 90 85 80 75 70 65 8 2 .9 5

R e la tive A b u n d a n ce

60 55 50 45

8 4 .9 4

40 35 30 25 20 15 10

8 6 .9 3 5 5 2 .0 0

5 7 .6 2 5 9 .9 5

6 6 .9 7

6 2 .9 6

7 0 .1 6 7 3 .2 7 7 6 .2 0

8 1 .0 3

8 8 .0 4

0 50

55

60

65

70 m /z

75

80

85

90

Image 3: Mass Spectrum of Dichloromethane Results and Assigned Fragment Ions...