General Information. There are six experiments:

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1 General Information Welcome to Chemistry 339. In this course you will learn advanced organic synthesis techniques including the handling of air sensitive reagents and reactions, vacuum distillation, and silica column chromatography. You will perform several classic named reactions, such as a Diels-Alder reaction, a Parikh-Doering oxidation, a Wittig reaction, and a Friedlander synthesis. In addition to the laboratory techniques, you will learn to read and interpret experimental procedures from the chemical literature, search the chemical literature to devise multi-step synthetic procedures for a particular structural target, assess the potential safety concerns of a procedure, and plan and carry out multistep synthetic sequences in the lab safely and efficiently. The lecture component of the course will be used as a discussion section to address challenges encountered in the laboratory in the previous week and to introduce new techniques and relevant safety information for upcoming experiments. There are six experiments: 1. Experiment 1. The Schlenck line 2. Experiment 2. Synthesis of quinolinyl benzamides 3. Experiment 3. Synthesis of nitrophenylethynes 4. Experiment 4. Synthesis and coupling of benzoxazoles 5. Experiment 5. Synthesis of a dihydronaphthalene derivative 6. Experiment 6. Synthesis of vinyl quinolines Laboratory time is limited and it is important that you use your time efficiently. There will be times when you will have to wait to use a piece of equipment. Try to find something else that needs to be done (use a different instrument, or perform a different part of an experiment) while you re waiting. We will also be sharing the vacuum & nitrogen manifolds and it will be important to plan and coordinate your experiments. Tasks like running a column, collecting NMR data, or recrystallizing a product can be done anytime, but setting a reaction up on the vacuum line will need to be done on a day when your group is scheduled to use the manifolds. Learning to plan your experiments and use your time efficiently is a critical laboratory skill, and one that you will be graded on in this course. There are more experiments and syntheses assigned in this course than you can (and are expected to) complete! It is important that you focus your efforts on the quality of your work, not the quantity. Chem 339, Spring 2016, Hoover 1

2 Laboratory Safety Appropriate laboratory attire will be enforced. Safety goggles and a lab coat or apron must be worn at all times in the lab. Closed-toe shoes and long pants or long skirts are required. Bare skin should be kept to a minimum. Eating and drinking in the lab is forbidden. Cell phones, headphones, and radios are not allowed in the lab. Long hair should be tied back. Wash your hands often in lab, and always wash thoroughly before leaving. Do not put your hands, pens, or pencils in your mouth while working in the lab. You should be familiar with the emergency procedures in this laboratory. Know the location of all exits. In the case of a fire alarm, turn off any open flames, grab your valuables and leave the building as soon as possible. Learn the location and operation of the safety showers, emergency eyewashes and fire extinguishers in the laboratory. Report any accidents to your TA or instructor. There are a number of hazards in this (and any) laboratory. Safety is an important aspect of this class, and you should think about safety as you read through your laboratory manual, as you plan your experiments, and as you work in the lab. The most important safety rule is to think! Think about what you are doing at any given time while in the laboratory. If there is anything that doesn t seem right or doesn t make sense to you, stop what you are doing and ask the instructor or your TA. Working with Chemicals Read the label (contents and hazards) before using any reagent. Take only as much as you need. When taking reagents, transfer the amount you need to a clean vial or other appropriate container for taking the material back to your bench or hood. Always replace the cap. Never return unused reagent to their storage containers: if you take more than you need, dispose of the excess in the appropriate manner. Clean up spills immediately: the next person to come along has no way of knowing if the clear liquid or white powder on the lab bench in harmless or hazardous. Neutralize acid spills with sodium bicarbonate before cleaning them up. An important part of safety and the safe handling of chemicals is their disposal. Chemical wastes must be managed and discarded in the most responsible and environmentally sound method available. Dispose of all waste in the labeled waste containers provided. Chemicals may not go in the trash cans or down the drain. If you are unsure of how to dispose of a particular material, ask your TA or instructor. Chem 339, Spring 2016, Hoover 2

3 Working with Equipment and Glassware Do not leave a Bunsen burner or other heating apparatus unattended. Do not pick up hot objects with your bare hands. Do not use chipped or cracked glassware. Do not adjust glass tubing connected to rubber stoppers. Severe cuts or puncture wounds may result. Do not force pipette bulbs onto pipettes. Apply just enough pressure to maintain a seal between the pipette and bulb. Forcing the bulbs may cause the pipette to slip and break, leading to sever cuts or puncture wounds. Lab Notebook You should keep a good laboratory notebook in this course. Use a bound 8 x 10 laboratory notebook that pages cannot be removed from. Update your notebook before each laboratory session (not each experiment) with a description of the experiment you will perform. You can revise this procedure as you work, to reflect your actual procedure. Your notebook is a record of what you did and it should be written and revised as you are working. Your notebook is the place for recording your plan of attack, all of your experimental results, measurements, observations, and conclusions. It should include numerical data (weights, volumes, times, etc.), procedures (A was added to B dropwise over 5 minutes using a pipette), and observations (a white precipitate formed after half of A was added to B). The most important features of a good lab notebook are clarity and completeness. You should never remove a page or plan to go back and fill something in later. Your notebook is your record of what you did and it should be written as you are working. Do not make notes on scratch paper and transcribe them into your notebook later. If necessary you can cross something our or recopy it for clarity, just indicate why and make sure the original is still there. Your notebook should enable you (or another researcher) to reconstruct what you did, including both good and bad aspects of the procedure. Your TA or instructor may check your notebook at anytime during the semester. Readings It is crucial that you carefully read (and understand!) the experimental description before entering the lab. Even more than reading, you should think through what you will be doing and what data you will need to collect. Use the literature procedure(s) to draft your own, more detailed, experimental procedure in your notebook. If the experiment references a procedure that you re not familiar with, or haven t used in a while (such as an aqueous extraction, or TLC analysis), then be sure to also read the appropriate sections in the Appendix or laboratory text before coming to class. Think through any nitrogen Chem 339, Spring 2016, Hoover 3

4 and vacuum manipulations that your synthesis will require. Make a note of any safety concerns associated with your synthetic procedure. These preparations are critical for safely working in the lab and to manage your time efficiently. Laboratory Reports Each of the laboratory reports will be due at the end of the semester. For each of Experiments 2-6, you will hand in a brief but formal laboratory report that consists of: (1) a ChemDraw scheme of the synthesis you completed, (2) a detailed experimental procedure for each step completed that is modeled after a scientific journal (including a ChemDraw structure of the intermediate or product made, your isolated yield, and your spectroscopic characterization data), (3) 2 copies of the 1 H NMR data of any intermediates you made, (4) 2 copies of the 1 H NMR, 13 C NMR, and IR spectral data of any final product(s) you made, and (5) your isolated compound(s) in a vial labeled with your name, the date, the name of the product, and the structure of the product. For Experiment 6, you will also be required to plan your own synthetic procedure. Your proposed synthesis will be due in lecture on Thursday March 19 th. Your laboratory reports and your final compounds will be graded on quality (purity, spectral data, laboratory technique used, writing, etc.) not quantity. It will be better for you to complete three steps of a synthesis to isolate a clean crystalline intermediate than to complete five steps and isolate a brown goop! Plagiarism. According to the Office of the Provost Academic dishonesty includes the following: plagiarism; cheating and dishonest practices in connection with examinations, papers, and projects; and forgery, misrepresentations, and fraud. ( Academic dishonesty on any assignment will result in a grade of zero for that assignment. Laboratory reports that are plagiarized will be given a grade of zero. This includes copying parts or all of your report from published materials or other students. Any material taken directly from another source should be referenced appropriately. Chem 339, Spring 2016, Hoover 4

5 EO Standard Synthetic Techniques A. Thin Layer Chromatography (TLC) Adapted from J. Chem. Educ. 2012, 89, You will need TLC plates, a pencil, a ruler, forceps, multiple capillary tubes, and a TLC developing chamber. Prepare your TLC Plates: Notice that the TLC plates are two sided; one side is glass (or sometimes plastic) while the other is the silica adsorbant (the stationary phase). On each TLC plate, with a pencil, gently draw a line approximately 1-2 cm from the bottom. Mark three points, evenly spaced, on the line, and label the marks. A center mark can be used as a co-spot. The purpose of the co-spot is to assess two different samples (for example a test sample and a reference sample) under the same conditions. Lane: STD CO Apply the Samples: Using a small capillary tube, apply a small drop of your sample solution to the appropriate mark. When applying the drop, be sure to gently touch the tube tip to the plate and quickly remove it. The spot should be not more than a few millimeters in diameter. Using a second small capillary tube, apply a small drop of the standard solution, or different sample to the other mark. You can practice applying small spots to a paper towel first. Develop your Plate: Obtain a wide mouth jar. Prepare 10 ml of the TLC solvent (mobile phase). The mobile phase is most commonly a mixture of hexanes and ethyl acetate, such as 10% ethyl acetate in hexanes, or equal volumes of hexanes and ethyl acetate. Pour approximately 10 ml of TLC solvent into the wide mouth jar (TLC chamber). Cap the jar. The volume of solvent inside the jar should be enough to cover the entire bottom of the jar, but not too much! The solvent level should be below the spotted material. Remove solvent if the level is at or above the line you drew on the TLC plate. Handling only the top of the plate or with forceps, place the spotted TLC plate into the wide-mouth jar. (Silica side up, spotted material near solvent surface.) Cap the jar. Do not move the developing chamber when the plate is in it! Allow the solvent to elute up the plate until it travels ~2 cm from the top. Stop the elution by removing the plate from the jar using your forceps. Quickly, with a pencil, mark the solvent front on the plate. Visualize the Plate: Gently circle with a pencil any spots that are visible (colored) in the three sample lanes. Samples that contain UV absorbing compounds can be visualized using a hand held UV lamp. UV absorption indicates that the molecule is conjugated (has delocalized π electrons). CAUTION: Do not look into the UV light! It will damage your Chem 339, Spring 2016, Hoover 5

6 eyes! Circle the spots that are colored (glowing) when exposed to UV light. In those circles place a dash, -, to represent those spots as being visualized by UV-Vis. Iodine (I 2 ) reacts reversibly with π bonds of a molecule to form a yellow- brown product. Samples that contain an iodine- reactive compound can be visualized using an iodine chamber. (Closed chamber contains iodine crystals.) Place the developed TLC plate into the iodine chamber in the hood. Close the lid and gently shake the iodine crystals. You will notice a coloration of those spots that represent compounds reacting with the iodine. Remove the TLC plate using forceps after a few minutes. (Do not add or remove the TLC plates from the iodine chamber using your hands!) Circle with a pencil those spots that were stained by the iodine. (Do not wait to mark the spots as the coloration will disappear over time.) Place an x in those circled spots that were visualized by iodine. Some spots will be visualized by both the iodine and UV-Vis techniques. The retention factor, R f, value is the distance traveled (in mm) by the spot divided by the distance traveled by the solvent (in mm). Measure the distances travelled at the center of the spot. Calculate the R f values for each circled spot in each sample lane. More polar substances will travel less, while the more nonpolar substances will travel further up the plate. Compounds in a sample with very similar polarity can travel the same distance up the plate. Compare the three sample lanes, both in the number and location of the spots. Sketch the TLC plate in your notebook. An example of a TLC sketch is shown below. This sketch should include the origin mark, the drawing of all spots seen in each lane, and the distance traveled by each spot and the solvent. Representative TLC plates before and after solvent development. B. Column Chromatography Flash Chromatography is described as an air pressure driven form of chromatography that is optimized for rapid separations. Flash chromatography differs from normal phase column chromatography in the use of a finer silica gel of mesh and also by the Chem 339, Spring 2016, Hoover 6

7 application of positive air pressure to the head of the column to flush the solvent through. The overall result is a simple, rapid analytical technique, allowing for high resolution. Find your Conditions: Select a column of the appropriate size. As a rule of thumb, the amount of adsorbent should be 25 to 30 times, by weight, the amount of material to be separated. It is also recommended that the column have a height-to-diameter ratio of about 8:1. These are just a rule of thumb and you will find that the difficulty of the separation (how close your spots are on your TLC plate) will be a key factor in choosing the right column. Choose a solvent system. Your solvent system should separate the mixture and produce an Rf on TLC of approximately 0.2 for the compound of interest. The most common solvents are mixtures of hexanes / ethyl acetate, although dichloromethane, and methanol / dichloromethane mixtures are also common solvents. For trickier separations, a gradient mobile phase system may be used. In this type of separation, you begin your separation with a relatively non-polar solvent, such as hexane : ethyl acetate (9:1). As you continue your separation, you alter the ratio of the two solvents so that the concentration of hexane decreases, while that of ethyl acetate increases. Your final solvent system is then much more polar, such as 4:1 ethyl acetate: hexanes. Pack the Column: Insert a plug of cotton into the end of your column, using a long stick or a small pressure of air to secure it in place. This protects the base of the column and prevents the silica gel from getting out. Add sand to the column to a depth of ~5 mm to obtain a flat surface onto which the stationary phase is added. Next, you will pack the column. There are many different techniques for packing and loading a column, a few of them are described below. The properties of your compounds will help you determine which method will be best. Dry Packing a Column In the fume hood, pour silica gel through a long-stemmed funnel into the column to a height of approximately 55 mm, whilst gently tapping the column vertically on the bench to ensure sufficient packing of the stationary phase. A layer of 5 mm of sand is then added to the top of the silica layer to ensure the top of the column remains flat throughout the isolation procedure. Using a Pasteur pipette, condition the stationary phase by filling the column with hexane (or the non-polar solvent of your solvent system). Apply light pressure to the top of the column (typically with an air or N 2 inlet, but for smaller columns a pipette bulb can be used. Increasing the air pressure in the column forces the hexane through the silica, removing any trapped air pockets from the column. Continue to flush the hexane through the column until the silica layer appears homogenous. This method is ideal for small columns but is not ideal for larger columns. Chem 339, Spring 2016, Hoover 7

8 The Slurry Method For this method, pour silica gel slowly into a beaker containing you solvent system, while swirling. Add enough silica gel to form a thick, but flowing, slurry. Add solvent to your column to fill it about half full. Open the stopcock to allow solvent to slowly drain out, then add the slurry slowly, portion-wise, to the column. A wide funnel will be helpful here. Be sure to swirl the slurry before each addition. While adding the slurry, you should gently tap the column continuously to remove air bubbles and encourage even packing of the silica gel. Never let the column run dry while packing. When enough silica has been added, flush some solvent through the column to remove air bubbles and form a flat surface on top. Load the sample: Again there are many methods for loading the sample onto the column. The most common, is to add the sample as a very concentrated solution, or if your sample is a liquid, to apply the liquid directly to the column. To do this, dissolve a solid sample in a minimum amount of polar solvent. I like to use chloroform. Transfer this solution (or your neat liquid) to the top of the column, slowly, using a Pasteur pipette. Rinse your flask or vial with a few drops of the polar solvent and load this solution onto the column. Apply positive air pressure to the column to force the extract onto the stationary phase. Top the column with ~5 mm of sand to prevent disturbance of your sample as you run your column. Run the Column: Add your solvent mixture to the top of the column, trying not to disturb your sample. Apply light pressure to the column to force the solvent through the stationary phase. Collect one column volume of solvent in a beaker or Erlenmeyer flask. This first column volume should contain only solvent, and none of the compounds that you are trying to separate. Then begin collecting fractions. The size of fractions that you will collect, will depend on the size of the column and the difficulty of the separation. It is common to monitor the progress of your column by TLC. Continue to collet fractions until all of your material has come off of the column. Identify the Pure Fractions: Apply a spot of each fraction collected onto a TLC plate. Develop the plate in a mobile phase of your choice. Visualize the plate (by UV, KMnO 4 stain, etc) to identify the fractions that contain your compound of interest. See the TLC section above for detailed TLC instructions. Accurately weigh a clean, dry round-bottomed flask. From the TLC plate, identify the fractions containing pure compound of interest and combine these fractions in the empty flask. Attach the flask to the rotary evaporator to remove the volatile solvent. Once all of the solvent has been removed, reweigh the flask and calculate the yield of your compound. It is a good idea to save your column fractions until you take an NMR spectrum of your isolated material, just in case you have not collected what you think you have! Other Tips Do not plug the column too tightly with cotton as solvent will fail to pass through. Ensure complete conditioning of the column prior to loading of the product sample. Should cracks in the column appear, flush more non-polar solvent through the column. Load your sample slowly onto the top of the column. Chem 339, Spring 2016, Hoover 8

9 1.5 Spectroscopy A. Infrared (IR) Spectroscopy There are many methods for acquiring an IR spectrum, most of which are outlined below. In this course, you will analyze your sample using an ATR (attenuated total reflectance) probe. For Liquid Samples Thin films The fastest sample preparation technique is simply to place a drop of liquid sample between two salt plates (KBr) and squeeze gently. If this is done properly, the film has enough surface tension to hold the plates together. Caution should be used to prevent air from getting back into the sample after it has been compressed. If the spectrum is too concentrated (many peaks bottoming out at 0 %T) try adding a much smaller volume of sample to the salt plate. For Solid Samples ATR Please see your TA if you have not had instruction on this apparatus. Grind a small amount of your solid (~100 mg) in a mortar & pestle. Carefully place this solid on top of the small zinc selenide crystal in the center of the apparatus. Do not contact the crystal with metal (your spatula) or paper products (e.g. Kim Wipes). Use a cotton Q-tip to maneuver your solid to fully cover the crystal. Once the solid is in place gently lower the press onto your sample/crystal (rotate the 2 black circular knobs). Scan the spectra as you would for a liquid. For clean up, carefully sweep up your solid reside with the brush and dispose of it in the solid waste container. Finish cleaning the zinc selenide crystal using a Q-Tip dipped in 2-propanol (Not Kim Wipes) Potassium bromide pellets In this method, a 1 mg solid sample is mixed with 80 mg of potassium bromide (located in the oven) and pressed between two stainless steel bolts in a threaded barrel. The two materials are ground to a find powder using a mortar and pestle. Screw one of the bolts into the barrel of the KBr press leaving one to two turns left. Pour the mixture into the open end of the pellet press and tap lightly on the benchtop to evenly distribute on the face of the bolt. The second bolt is then carefully screwed in until it is finger tight. Place the head of the bolt into the hexagonal hole that is attached to the benchtop. Using the torque wrench, making sure direction indicator on the head of the wrench is pointed to the R, tighten the bolt system until the wrench makes a loud click for the first time which is at 120 in/lb. Keep the bolts tight under pressure for approximately 60 seconds so that the crystals "settle". Be sure not to tighten the bolts too much--be firm, don't give Chem 339, Spring 2016, Hoover 9

10 it too much muscle. If you heard the loud click, not the softer clicks of the ratchet mechanism, you are done! Reverse the wrench by switching to l and rotating in the opposite direction. Remove both bolts and place the cylindrical chamber containing your pellet into the sample holder within the IR equipment. Some pellets will appear white. You may have used too much sample. If the pellet is more than 1-2 mm thick, you probably should regrind and remake it with less material. Your pellet could also have the freckled look. Tiny but distinct spots are apparent throughout the pellet. This arises from insufficient grinding. Methylene chloride solution Dissolve ~100 mg of your solid in a small amount of methylene chloride (approx 1 ml). You may heat the solution to help it completely dissolve. Add 3-5 drops of this solution to a salt plate and let the methylene chloride evaporate. Once evaporated, it should leave a thin film of your solid on the plate that is ready to be analyzed. Mineral oil (Nujol) mulls A relatively simple sampling method for softer organic samples is a mull. The proper approach is to use an agate mortar and pestle. Place a few milligrams of the sample into the mortar and grind it until it looks like a thin film. At this point, add a drop of mineral oil and continue to grind into a thick homogeneous paste. Scoop up enough of this paste to fill the center of a salt plate and sandwich it between a second salt plate. The particle size must be reduced before the sample can be lubricated. Mineral oil has a considerable spectrum of its own, being a hydrocarbon of high molecular weight. See the sample mineral oil spectrum located by the IR. They really are made of salt, so clean with methylene chloride and a Kim Wipe. Never clean with water! B. Nuclear Magnetic Resonance (NMR) Spectroscopy Spectroscopy deals with the interaction of light with matter. The energy of an atom or molecule is quantized, that is only a limited number of energy levels are allowed. Spectroscopies examine the gaps in energy between these energy levels, which can be quite informative as to the nature and structure of the molecule. In Chem 339 you will use a number of different kinds of spectroscopies, most if not all of which, you have used in other classes. A key issue to always keep in mind in almost all spectroscopic measurements is that your sample is in a vessel and often in a solvent. The spectrometer measures so your spectrum shows the entire sample, including the vessel and the solvent. For instance, NMR solvents often have residual protons that will show up in a 1 H NMR spectrum. Chloroform-d 1 is not 100% CDCl 3 ; it is probably about 99% CDCl 3 and 1% CHCl 3 Chem 339, Spring 2016, Hoover 10

11 To prepare your NMR sample: Place ~ 10 mg, of your sample into an NMR tube and add enough deuterated solvent (which deuterated solvent should you use?) to reach about 1½ inches (three fingers) up from the bottom of your tube. Shake the tube until your sample dissolves. If necessary, try a new tube with a different solvent to see if you get better dissolution. [A 13 C NMR spectrum may require a more concentrated sample.] When your NMR tube is ready, label it with your name and the NMR solvent used and ask your TA or instructor to help you collect a spectrum. Chem 339, Spring 2016, Hoover 11