CH 242 EXPERIMENT #4 WEEK OF March 30, 2009 NATURAL PRODUCT EXTRACTION: THE ISOLATION AND CHARACTERIZATION OF EUGENOL FROM

CLOVES Introduction
Many plants contain essential oils which are typically high boiling liquids rich in organic compounds. Indeed, some of these compounds have important uses as medicines, perfumes, flavoring agents etc. Examples are the antimalarial agent quinine obtained from the bark of the cinchona tree, the pleasant smelling geraniol from roses, and vanillin from vanilla pods. In this lab, you will extract the essential oil present in cloves by a technique known as steam distillation. The clove oil obtained in this manner is a rich source of eugenol, a compound that has antibacterial properties. Also present in the oil of cloves, albeit in smaller amounts, are eugenol acetate, caryophyllene, and other minor compounds.
OH OCH3 OCOCH3 OCH3 H3C H3C CH3

CH2 CH=CH2

CH2CH=CH2

H2C

Eugenol

Eugenol Acetate

Caryophyllene

In steam distillation, a mixture of immiscible liquids, one component of which is water, is co-distilled at a temperature that is lower than the boiling points of the individual components. This is because the total vapor pressure Ptot of an immiscible liquid mixture is the sum of the individual vapor pressures that each component would exert if it were a pure liquid. When Ptot equals the atmospheric pressure, the mixture boils. Since many essential oils contain liquids that may not survive heating at higher temperatures, steam distillation is an especially advantageous technique for isolating them because their immiscible mixture with water boils below 100 C. After obtaining the oil of cloves by steam distillation, you will need to isolate eugenol by another useful technique known as chemically active extraction. In this process you take advantage of the fact that eugenol, because it is a phenolic compound, is weakly acidic whereas both eugenol acetate and caryophllene are neutral. Thus, treating the clove oil with a strong base such as sodium hydroxide will result in the selective formation of the sodium salt of eugenol which will be soluble in the aqueous phase. The other neutral compounds will not react with sodium hydroxide and therefore will remain

in the organic phase. Simply by separating the aqueous phase and acidifying it you can isolate eugenol. Subsequently, you will need to characterize eugenol by IR spectroscopy. You may work in pairs on this experiment. You must, however, submit individual lab reports that must be developed and written independently.

Procedure for Steam Distillation:

Load a 100 mL round-bottomed flask with 5 g of freshly ground cloves, 40 mL of deionized water, and a boiling chip. Add three drops of antifoam to the flask and set up an apparatus for steam distillation as instructed in lab. Collect about 35 mL of distillate in a graduated cylinder. Pour the distillate into a 125 mL Erlenmeyer flask, add 2 g of NaCl, and gently swirl to dissolve the salt. Cool the solution in an ice bath for a few minutes and pour into a 125 mL separatory funnel. Add 50 mL of hexanes to the separatory funnel in order to extract the clove oil from the aqueous phase. Venting frequently, gently shake the funnel for about a minute. (Remember: vent often to release the pressure from within the funnel.) Drain the aqueous layer into the original Erlenmeyer flask and leave the organic layer containing clove oil in the separatory funnel.

Procedure for Chemically Active Extraction:

Add 25 mL of 5% aqueous sodium hydroxide to the organic layer in the separatory funnel. Shake the funnel for about a minute and be sure to vent periodically. Drain the aqueous layer into a clean 100 mL beaker. Extract the organic layer with another 25 mL of the sodium hydroxide solution and drain the aqueous layer into the same beaker. Collect the organic layer in a separate 125 mL Erlenmeyer flask. Cool the aqueous layer in an ice-water bath and acidify with 6M aqueous hydrochloric acid. Return the acidified solution to the separatory funnel and extract with 15 mL of hexanes. Shake the funnel for about a minute and with frequent venting. Drain the organic layer into a clean and dry 50 mL Erlenmeyer flask. Repeat the extraction with another 15 mL portion of hexanes and combine the organic layers. Add a small amount of anhydrous sodium sulfate to the organic layer and let sit for 5 minutes. Gravity filter the organic layer into a clean, dry, and pre-weighed 50 mL round-bottomed flask. Remove the solvent using a rotary evaporator and weigh the flask again. Calculate the weight of eugenol, estimate the percent yield, and record its IR spectrum. PRELAB (Due in class on Monday March 30, 2009) (a) Identify the major functional groups present in eugenol, eugenol acetate, and caryophyllene. For each compound list two or three prominent bands that you might expect to see in the IR spectrum of that compound. (b) Why are phenols more acidic than simple alcohols? Identify the product(s) of the reaction of eugenol with sodium hydroxide. WHAT SHOULD BE IN YOUR NOTEBOOK? 1. An entry of the title, date, and page number in the table of contents. 2. An entry of the title, date, and partners name on the first page of the experiment (this should be done each week). 2

3. Masses and volumes of materials that were used in the experiment, and the mass of eugenol isolated. 4. Brief descriptions of the procedures that were followed with deviations, if any. 5. A sketch of the steam distillation apparatus. 6. A boiling point range for the steam distillation of clove oil. WHAT SHOULD BE IN YOUR LABORATORY REPORT? (Do not exceed a total of two pages for items 1-5 below. Please type your report, and print double-sided.) 1. Introduction (1 paragraph) 2. ChemBioDraw structures for eugenol, eugenol acetate, and caryophyllene. 3. A flow chart to describe the chemically active extraction scheme that was employed to isolate eugenol from clove oil. 4. Results and discussion including the principles of steam distillation, calculation of the mass % of eugenol present in the clove sample, and analysis of IR data. 5. Conclusion (1 paragraph) 6. Attach a copy of the IR spectrum, and carbon copies from the lab notebook.