Experiment 5

Liquids: Distillation, Boiling Points, and the Fermentation and Distillation of Ethanol


Distillations are performed to isolate volatile chemicals from non-volatile or less volatile chemicals. This experiment addresses how distillations (both simple and fractional) can be used to isolate chemical in pure form.  In addition, boiling points and other physical properties can be determined, including refractive index and density of purified materials. We also setup a fermentation to isolate ethanol from these mixtures.


Parts A and B will be performed during the first laboratory period.  Part C, which is the distillation of ethanol from a fermentation mixture, will be performed during the second laboratory period.

Techniques and procedures that you will perform during this experiment include:

This experiment is performed in three parts over two days.  Parts A & B are performed during the first day.  Part C is performed during the second day of lab.  The fermentation setup is done one week prior to the beginning of Part C.

Part A: Purify 2-propanol by simple distillation. This technique shows how to purify, by distillation, a volatile organic compound.  This procedure is good for purification of volatile compounds from compounds that are not volatile, but not very successful for mixtures of compounds, each of which is volatile.  Part B describes how to fractionate these types of mixtures using fractional distillation.

Part B: Separate ethanol from water by fractional distillation.  Compounds which are volatile will each distill at temperatures well below their actual boiling points (e.g., remember the vapor pressure of water?).  We need to have a procedure that allows for more efficient separation of these volatile liquids, and this is the process of fractional distillation.

Part C: We will prepare a fermentation of glucose to produce ethanol.  The ethanol from this fermentation will be isolated by fractional distillation (using the procedure described in Part B).  You must repeat your fractional distillation, in order to remove water, which, unfortunately predominates during the first fractional distillation.  Once you have repeated your fractional distillation, you can determine the amount of alcohol in your final distillate sample.

In many of the previous experiments that we have performed, (e.g., the experiments involving extraction, separation, Re-crystallization, and chromatography), you have learned that a chemist must be able to exploit the specific physical properties of the components of a mixture in order to efficiently separate and purify the desired chemical compound. The most common method for separating and purifying volatile liquids is distillation, which makes use of the specific boiling points of the liquid components in the mixture.

When there is only one volatile liquid, or when one of the liquids has a boiling point well below the others, a simple distillation can be used. However, if there are two or more liquid components, which have boiling points near each other, a fractional distillation must be used (the theory of distillations is discussed in Chapter 35 of Zubrick).

In Part A of this experiment you will carry out a simple distillation of 2-propanol (isopropyl alcohol), which has been contaminated with a non-volatile impurity (a dye). In Part B, you will carry out a fractional distillation of a 50:50 (v/v) mixture of ethanol and water.  

During each of these experiments, you will determine the boiling point of the volatile organic compound during distillation. You will monitor the temperature that the volatile liquid has, in the gas phase using a thermometer.  This temperature should remain constant, and should reflect the actual boiling point of the chemical under the atmospheric conditions of the day. Report this boiling point as part of your data.


Part A: Simple Distillation of 2-propanol

Safety:2-propanol is a highly flammable liquid and a severe eye irritant -- no flames will be allowed in lab while it is in use. As for every experiment, goggles must be worn, even though you may not actually be  working the chemicals, if there is anyone using 2-propanol in the lab.

Follow the instructions in chapters 19 and 20 of Zubrick for setting up a simple distillation apparatus, although the instructor will go through the setup, and the use of your organic kit.  You will use a 50-mL round bottom flask as the distillation pot, and a 100-mL round bottom flask as the receiver.  As a standard rule, anytime you are boiling an organic compound, you will always include a few (not a handful!) boiling stones to keep the solution boiling smoothly!

Using a beaker (not a graduated cylinder) obtain about 30-40 mL of "impure" 2-propanol (this sample of 2-propanol has had a small amount of a soluble, non-volatile dye added to it as an impurity). Add the 2-propanol to the distillation pot (never pour anything through a ground-glass opening without using a funnel), add the boiling stones, and begin the distillation (remember to turn on the cooling water before you turn on the heat). Collect your distillate in a pre-weighed graduated cyclinder (10-mL or 25-mL or 50-mL cylinder).

Once the temperature starts to rise above room temperature, you should start to record the thermometer reading every minute.  

You should start to record the temperature at any time up to when the solution starts to boil. Record every minute the temperature you read. Continue to do your distillation until you have collected about 20 mL of distillate. Be sure that the distillation pot never goes dry (never let a heated flask go dry!).

You must plot your data by hand using graph paper, or you can use Excel or another graphing program for a graph for inclusion in your lab notebook and written report.

Measure the volume of the distillate collected.  Using a pre-weighed graduated cylinder (10-mL or 25-mL or 50-mL cylinder), you should determine the density of the collected distillate. You will also determine the refractive index of your distilled liquid.  Your instructor will describe how a refractive index is determined.

Dispose of your liquid, and any liquid remaining in the distillation pot, in the liquid waster.  Be sure to make certain that no boiling stones are deposited into the liquid waste.  Put the boiling stones in the solid waste container.

Part B: Fractional Distillation of an Ethanol/Water Mixture

Safety: Ethanol is a flammable liquid and an irritant; avoid contact and inhalation -- wear gloves while handling it. No flames will be allowed in lab while ethanol is in use.  Goggles must be worn whenever anyone is using chemicals.

Set up a fractional distillation apparatus as demonstrated by your instructor.  Use glass beads to pack the fractionating column (your instructor will demonstrate how to pack the column). Try adding some glass beads directly to your fractionating column. If the glass beads stay in the column, there is no problem, but if any beads go through, try adding a larger amount of glass bead, and their packing inside the fractionating column should allow them to stay in place. Do not ever use glass wool or anything besides beads in the fractionating columns.

To do this part of the experiment, you will use a 100-mL round bottom flask as the distillation pot. You will need a number of receivers; it is best to use test tubes. Measure into one test tube about 4 mL of water. Use this sample to know how much liquid you need to obtain about 4 mL of distillate during this part of the experiment. Continue collecting 4-mL samples until you have collected about 30 mL of distillate. Determine the refractive index of each collected sample, as well as determining the refractive index for pure ethanol and pure water.

Start recording the temperature as soon as your sample begins to boil. Record the temperature every 30 sec. Collect your distillate into test tubes. You should collect about 4 mL in each test tube, but it is not necessary to measure each tube. As a comparison, add about 4 mL of water into a test tube. Collect about the same amount of liquid into each of the tubes during the distillation process. Continue recording the temperature until you stop collecting your samples. Collect about 30 mL of distillate.

You must plot your data by hand using graph paper, or you can use ChemWorks™ on the computers for inclusion in your lab notebook and written report. You should have two plateaus, one for the boiling point of the ethanol and the other for the boiling point of the water. Your graph for your lab report and for your notebook must show these two plateaus.

After the distillation has finished, you will have a good determination for the boiling point of ethanol.  Determine the refractive index for each of your samples.

Do not ever throw any glass beads away.  At an expense of about 25 cents ($0.25) per glass bead, they are very expensive.  Keep your glass beads in your fractionating column (add some tissue to the top to prevent spillage) until the next lab period. Never throw away any glass beads.

Part C: Fermentation and Distillation of Ethanol

Yeast ferment sugars to produce ethanol. You will use glucose (dextrose) as the sugar the yeast will use for alcohol production. Glucose has a molar mass of 180 g/mol. The molar mass of ethanol is 46 g/mol. The other product of fermentation is carbon dioxide, which we will not consider in the current experiment. The equation for this reaction is as follows:

Based on the balanced equation, one (1) mole of glucose will yield two (2) moles of ethanol. Based on these molar ratios, one mole of glucose is 180 g, and two moles of ethanol is 92 grams. On the other hand, one mole of sucrose will yield four moles of ethanol. Your Instructor will inform you which carbohydrate you will be using.

Based on the amount of carbohydrate (glucose or sucrose) you actually started with (based on the amount you weighed out), what is the theoretical yield of alcohol that you should produce? Use this value to determine your percent yield after you have finished this part of the experiment.

Grams of glucose (or sucrose) fermented: _________________ g

Theoretical yield of ethanol: __________________ g

Fermentation Setup

Set up the fermentation container in the following manner:

Allow the fermentation mixture to stand at a temperature of 25-35oC until fermentation is complete (about a week is required).

First Distillation of Fermentation Mixture

After the fermentation is complete, you will distill the alcohol from the liquid mixture, including the solid, particulate material that is not soluble. Gently pour all of the liquid into a 500-mL round-bottomed flask (checked out from the stockroom).  The liquid will be somewhat cloudy, but this will not interfere with the first distillation (do not ever let a distillation flask go dry, but this should be of little concern here since we will only collect a fraction of the liquid via distillation).  

Add a few (6-8) boiling stones to the flask.  Attach a fractional distillation column (filled with glass beads, as described in Part B) to the top of the distillation flask, and attach the still head and condenser unit. You do not need to monitor the temperature as the first distillate is mostly water, so the temperature will be about 100oC.  Collect about 50 mL of distillate into a pre-weighed graduated cylinder (a 50- or 100-mL graduated cylinder can be used).

After the first distillate has been collected, observe and record the collected volume, and weigh the graduated cylinder to determine the mass of collected distillate (total mass of cylinder containing the distillate minus the mass of the empty graduated cylinder).  You should now calculate the density of the distillate, based on the mass of the distillate you just determined divided by the volume collected.  From the Table of Densities of aqueous ethanol solutions, given in Footnote 2, calculate the mass of alcohol collected in this first distillate.

To make your determination of the amount of ethanol isolated, you can estimate your percent alcohol using the table at the end of this experiment. This table shows you several ways to determin the amount of ethanol in your sample based on density. The easiest column to use is the first column which shows the mass percent of alcohol (% by mass) as follows:

Using the density of your distillate from the above calculations, determine the percent alcohol ( you may have to extrapolate (e.g., Calculate the percent yield of ethanol produced in the fermentation, based on the theoretical amount of ethanol that would be produced from the starting amount of glucose (each glucose molecule produces two ethanol molecules).  This "crude" distillate will be used in a second fractional distillation.

An additional table is available, using an interpolative procedure to determin percentages which do not fall within the larger percent range shown in the table below (percentages are in 5% increments). The new table (which is an Adobe pdf document) shows you how to get to the percent values, based on density, in unit values, e.g., 45%, or 46%, or 47%, etc. The highlighted values in this pdf document shown how to determine percent concentration values between 45% and 50% ethanol. You can download and print this document at this URL of a pdf document.

Discard the residue left in the distillation pot, which contains mostly water.

One can obtain 95% ethanol (but not 100% ethanol; do you know why? What is an azeotrope?) from the dilute alcohol mixture obtained during the first distillation.  For the second distillation, monitor the boiling point carefully, as you should collect the material that distills at a temperature of 78-82oC; if too little distillate is obtained in this range, continue the distillation and collect the fraction boiling at 82-88oC.  

Prior to doing your second distillation, empty the glass beads from the fractioning column into a beaker you will wash them yourself with soap and water, and then reuse them. Also, you need to wash with soap and water each of the following pieces of glassware:

You can now re-assemble your distillation setup for a fractional distillation, and then follow the procedure for collecting and analyzing your second-distillate liquid. At the end of the experiment, put the used glass beads into a beaker on the Instructor's bench to be washed by the Stockroom Personnel. Do not throw the glass bead away.

Procedure for collection of ethanol via a second distillation:

  1. Add the alcohol-water mixture from the first distillation (after weighing and determining its density) to a 100-mL round bottom flask.
  2. Add a few boiling stones to your round-bottom flask (distillation pot) to maintain a slow, steady boiling.
  3. Start monitoring the temperature as soon as you turn on the heat, or at least prior to the solution boils. Monitor the temperature at regular intervals, usually every minute, until you stop collecting your samples.
  4. Start to collect 4-mL samples (using you conical vial in the organic kits or a test tube) until the temperature rises significantly above the normal 78-82oC temperature, as described above. It is still permissible to collect above the 82oC degree range, but your samples contain more and more water. However, if you have not collected at least 4-5 samples, each of about 4-5 mL volumes, continue to collect samples (regardless of temperature) until at least 20 mL of solution is collected. Based on theoretical yeields, and actual lab experience, you could have produced 30-36 mL of ethanol during your fermentation, so collectin at least 20 mL is not out of reason.
  5. Do not determine a refractive, but you must determine a density for each sample. To do this, collect about 4-5 mL in a conical vial (as mentioned above), and then immediate transfer the contents of the vial into a pre-weighed 10-mL graduated cylinder. After transfer of collected material into the 10-mL graduated cylinder, reattach the vial to the vacuum adapter, and collect another 4-5 mL sample.
  6. Determine the mass and volume of the sample in the 10-mL cylinder to calculate a density of the sample. After determining the mass and volume, transfer its contents into a pre-weighed 50-mL graduate cylinder to collect all samples for an overall density and yield of ethanol for the entire experiment.
  7. After you have analyzed individually the 4-mL samples, combine your 4-5-mL collected distillate samples into the 50-mL graduated cylinder, the total volume, the mass of the sample in the 50-mL cylinder, you can determine the density.
  8. From the density just determined (step #7), determine the total mass of ethanol collected during the second fractional distillation. No need to determine a refractive index as density (and total mass of solution) will be our criterian for yield.
  9. Determine percent yield based on the overall yield from step #8, by dividing the actual yield by the theoretical yield times 100 to get a percent yield.

Based on the amount of collected alcohol, its density, what is the percent yield? What is the theoretical yield?. Your instructor will help you determine the percentage of alcohol based on your refractive index and density determinations.

Do not throw any glass beads away.  At an expense of about 25 cents ($0.25) per glass bead, they are very expensive.  After all distillations are completed, pour your glass beads into a large beaker on your instructors bench, so they can be cleaned.  Never throw glass beads away.


1Pasteur salts include:

2Percent yield calculation information is shown below:

To calculate %Yield, you need to know the Theoretical Value you could obtain, based on balanced equations, and if 100% of the reactant(s) is converted to product.  After obtaining your Experimental Value (actual yield), divide it by the Theoretical Value, then multiply by 100 to get %Yield.

3Refractive Index Determination: Refractive Index must be used to help characterize your sample. It can also be used to help determine the percent of a chemical (such as ethanol) in an aqueous solution. Refractive Index is always reported to 4 decimal places. An example of the Refractive Index scale is shown below. The correct reading would be 1.3764.

As an exercise, please give the values for the refractive index for each of the arrows (to 4 decimal places):

#1 _______________  #2 _______________ #3________________

4Aqueous Ethanol Table: Column headings are: density, alcohol % by weight, alcohol % by volume, and g alcohol/100 mL solution.

Aqueous Ethanol
Density (g/ml) % (m/m) % (v/v) g/100 mL Density (g/ml) % (m/m) % (v/v) g/100 mL
0.98938 5 6.2 4.9 0.85564 75 81.3 64.2
0.98187 10 12.4 9.8 0.84344 80 85.5 67.5
0.97514 15 18.5 14.6 0.83095 85 89.5 70.6
0.96864 20 24.5 19.4 0.81797 90 93.3 73.6
0.96168 25 30.5 24.0 0.81529 91 94.0 74.2
0.95382 30 36.2 28.6 0.81257 92 94.7 74.8
0.94494 35 41.8 33.1 0.80983 93 95.4 75.4
0.93518 40 47.3 37.4 0.80705 94 96.1 75.9
0.92472 45 52.7 41.6 0.80424 95 96.8 76.4
0.91384 50 57.8 45.7 0.80138 96 97.5 76.9
0.90258 55 62.8 49.6 0.79846 97 98.1 77.4
0.89113 60 67.7 53.5 0.79547 98 98.8 77.9
0.87948 65 72.4 57.1 0.79243 99 99.4 78.4
0.86766 70 76.9 60.7 0.78934 100 100 78.9

To use the above chart, find the density value that is closest to the density that you determined for your ethanol samples. Then, look at the column that gives g/100mL, which gives the mass of ethanol in 100 mL of solution (alcohol and water together). For example, if your total volume of ethanol from the second distillation was 8.5 mL, and if your mass of aqueous alcohol was 6.88 g, the density of this solution would be 0.80983 g/mL, meaning that 100 mL of this solution would contain 75.4 g of alcohol (see chart). Since you have 8.5 mL, the amount of ethanol would be 6.41 (75.4 x 0.085) grams ethanol (your actual or experimental yield). Use this value to determine percent yield.

For example, if the density was 0.97514 g/ml (from section highlighted in green), the mass percent of alcohol is 15%, meaning that 15% of the total mass would be alcohol.  If it was 18.5% by volume, then based on the volume of liquid, 18.5% would be alcohol.  The last column says that 14.6g of alcohol would be present in a 100 mL volume of collected material

An Adobe Acrobat document is available to correlate density and percent ethanol in one-percent increments here.


Chemicals and Reagents
Compound MW Amount mmol mp bp Density ηD msds
2-propanol (isopropyl alcohol) 60.1 50 mL   -88.5 82.5 0.78505 1.3772 msds
Dextrose (glucose) 180.16 50.0 g 277.53 146 --- 1.54   msds
Sucrose 342.3 50.0 g 146.07 160-186 --- 1.59   msds
Ethanol 46.0414     -114.1 78.5 0.8 1.3614 msds
Compound g/mol grams or mL 10-3 M oC oC g/mL ηD msds

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