Lab 4 Manual-Using GC to Identify Unknowns - Jan 2024 PDF

CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography Identifying Unknown Compounds using Gas Chromatography A. Experimental Objectives  Measure and analyze the retention time of each of six substances as they pass through a Vernier Mini GC....

CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography Identifying Unknown Compounds using Gas Chromatography A. Experimental Objectives  Measure and analyze the retention time of each of six substances as they pass through a Vernier Mini GC. -  Measure and analyze the retention time of an unknown mixture of the substances.  Identify the substances present in the unknown mixture based on retention times. B. Introduction There are many different types of chromatography: paper, thin layer (TLC), liquid, high-pressure liquid (HPLC), and gas (GC). Chromatography is applied in many fields. Biochemists use liquid chromatography to separate proteins; chemists use GC, TLC, and HPLC to identify organic compounds. Forensic scientists and other specialties use gas chromatography for drug tests, toxin screens, and environmental analysis. All types of chromatography use the same principles that include a stationary phase and a mobile phase. The stationary phase is immobile on the column or the plate and the mobile phase travels from a start point to an end point. Compounds travel from the start to the end at a specific rate depending on their competing affinity for the mobile gas/liquid phase versus the stationary solid phase. Compounds adhere to the stationary phase through dipole interactions, dispersion forces, or ionic interactions. Gas Chromatography is a technique widely used to separate complex mixtures of substances. Compounds present in a volatile liquid or gaseous solute are isolated after traveling through a coated column based on the substance’s size and intermolecular interactions. If a compound tends to adhere to the column through intermolecular interactions, it takes a longer time to emerge compared with a compound that does not tend to stick onto the column. The overall strength of the interaction experienced between the substances and the column is determined based on the number and strength of intermolecular interactions between the two species. Substances that pass quickly through the column exhibit fewer intermolecular interactions with the column. Figure 1: Sample gas chromatogram The Vernier Mini GC uses a metal column with a nonpolar coating, called the stationary phase. A sample, consisting of one or more compounds, is injected into the column and is carried through the stationary phase by atmospheric air, which acts as the mobile phase. The nonpolar coating of the stationary phase most strongly retains solutes of the same polarity. Organic compounds flowing out of the chromatography column are then detected by a chemical sensor that produces electrical responses proportional to the concentration of the compounds. The presence of such a chemical at Page 1 of 5 CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography the detector is seen as a peak on a chromatogram, as shown in Figure 1. The unique time it takes for a compound to exit the column after it is injected is called the retention time. With a gas chromatograph, a compound can be identified from a mixture by its retention time. Several factors can affect a compound’s retention time. More volatile compounds (i.e., compounds with a lower boiling point) will move through the column faster because they are flowing in the mobile phase and not strongly attracted to the stationary phase. The surface functional groups present on the compound are also a factor. For example, alcohols may interact with a polar stationary phase more than esters because alcohols are capable of donating hydrogen bonds. The molecular weight of a compound may also play a role to a slight extent, although it is not a direct relationship between molecular mass and the time it will travel through a GC column. In this experiment you will explore the process of identifying one or more unknown species using gas chromatography. First, you will practice using a gas chromatograph by testing several known substances. You will then use this information to identify the substances present in an unknown mixture. C. Chemicals and Supplies Chemicals: Ethanol, propyl acetate, butyl acetate, 2-butanone, 4-methyl-2-pentanone, methanol, unknown mixture and distilled water. Supplies: Vernier Mini GC, Laptop with Vernier Instrumental Analysis installed, 1 L glass syringe and Kimwipe. D. Safety note  Methanol, ethanol, propyl acetate, butyl acetate, 2-butanone and 4-methyl-2-pentanone are flammable. Take care to avoid flames. Avoid breathing vapors from these substances.  4-methyl-2-pentanone is hazardous in case of skin and eye contact (irritant), ingestion or inhalation.  DO NOT inhale or swallow any chemicals. Handle all the chemicals with care.  Be sure you are wearing proper safety equipment.  Be sure you wash your hands with soap and water before leaving the lab. E. Waste disposal Dispose the chemicals in “Non-halogenated Organic Waste Container”. Page 2 of 5 CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography F. Procedure Part 1: Testing Known Compounds 1. Obtain a glass syringe and a set of vials containing the six known substances and one unknown mixture containing two substances to be identified. You will not only test methanol but use it to clean the syringe needle. 2. Prepare the Vernier Mini GC for data collection. a. Turn on the Mini GC. b. Connect the Mini GC to the USB port on your Laptop. c. Open Vernier Instrumental Analysis software in your laptop, and Click on “Gas Chromatography” d. Set the Temperature-Pressure values to: Start temperature 35C Hold time 1 min Ramp rate 10C/min Final temperature 65C Hold time 6 min Total length 10.0 min Pressure 4.0 kPa e. Click “Apply” to initiate the Mini GC warm up. When the Mini GC is ready for injection in Step 6, the message will read, “Vernier Mini GC is ready, Inject and select Collect simultaneously”, and the LED will turn to green in the GC f. Continue with Step 3 during GC warm up. 3. Follow the steps below to clean and flush the syringe with methanol. Important: The glass syringe is fragile. Be careful not to bend the needle or bend the plunger. Never pull the plunger back more than 50% of its total volume. Be careful not to bend the plunger as you press it down. a. Depress the plunger fully. b. Submerge the tip of the syringe needle into the vial of methanol. c. Pull back the plunger to fill the barrel about 1/3 full of methanol. Examine the barrel of the syringe and estimate the amount of methanol in the barrel. d. Expel the liquid onto a Kimwipe e. Repeat steps a–d at least two times, until you are comfortable pulling up a liquid into the syringe and measuring the volume in the syringe barrel. Use a Kimwipe to carefully pat around the tip of the syringe needle. 4. Follow the process in Step 3 to clean and flush the syringe with propyl acetate. Page 3 of 5 CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography 5. Collect a volume of propyl acetate for injection. a. Submerge the tip of the syringe needle into the vial of propyl acetate one last time. b. Draw up 0.10 L of liquid. It is important that the volume be very close to 0.10 L and that you inject the same volume for each test. c. After collecting your sample, gently wipe the needle from barrel to tip, with a Kimwipe®. 6. Prepare for injection and the start of data collection. a. The Mini GC should now have reached the correct start temperature and pressure and the LED turned to green. b. To insert the needle of the syringe into the injection port of the Mini GC, hold the syringe with one hand and steady the needle with your other hand, as shown in Figure 2. Insert the needle into the injection port until the needle stop is fully seated. If the needle sticks, rotate it slightly while inserting. Do not move the plunger yet. c. Simultaneously, depress the syringe plunger and select Collect to begin data collection. Pull the needle out of the injection port immediately. 7. While the data collection proceeds, repeat Step 3 to thoroughly clean the syringe and needle. It may take more than three flushes to feel the syringe plunger move smoothly again, which is your indicator that the syringe and needle are both suitably clean. 8. The data collection will end after ten minutes. You may stop the data collection early if you are certain that the entire injected sample has passed through the detector. Figure 2 9. Determine the retention time for your chromatogram: a. Select and integrate the peak. To do this, drag from a little before the peak to a point far enough to the right that includes the entire peak. b. Click on Peak Integration tool at the bottom of the chart. c. Peak 1 with retention time and peak area will be displayed in the data table. Replace peak 1 with the name of the compound and click ok. 10. Save your run of data with the compound name. 11. Follow the steps below to test the next sample in your set of substances. a. Select New Experiment from the Untitled menu b. Click on “Initiate Data Collection” on top of the screen to bring up the Temperature-Pressure profile again. This profile will be the same as for your previous run. Select “Apply” to initiate the Mini GC profile. c. While the Mini GC adjusts to its Temperature-Pressure profile, repeat Steps 4 and 5 with the next sample. d. After the Mini GC is ready, repeat Steps 6–10 to test your new sample. 12. Repeat Steps 4–10 for the remaining four known substances. Page 4 of 5 CHEM 211 Lab – Organic Chemistry Lab # 4 – Identifying unknown compounds by gas chromatography Part 2: Testing Unknown Mixture 13. Select New from the File menu:  In Vernier Instrumental Analysis program, proceed directly to Step 14. 14. Repeat Step 4–10 to test your unknown mixture, but this time inject 0.2 L of the unknown mixture. When the run is completed, integrate the peaks starting from the left-most peak adding one by one to the data table. 15. After you have completed the test of your unknown mixture, save your file for use at a later time. Save the file on the program, as directed by your instructor. 16. Turn off the Mini GC. You will need to refer to the various chromatograms to answer the Data Analysis questions. Print a copy of your graph and/or save your data, as directed by your instructor. References: 1) Engel R. G. et al. 2011. Introduction to Organic Laboratory Techniques: A Microscale Approach, fourth Edition. Brooks/Cole Cengage Learning. Page 797-817. 2) Pavia D. L. et al. 2007. Introduction to Organic Laboratory Techniques: A small-Scale Approach, Third Edition. Brooks/Cole Cengage Learning. Page 277-296. 3) Some of the information in this experiment are obtained, edited and modified from the websites: http://www.vernier.com/experiments/ http://en.wikipedia.org/wiki/Gas_chromatography Page 5 of 5