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Unit 33

EXPERIMENT 33
Identification of an Unknown ___________________________________________
E33.1 OVERVIEW In this experiment you will identify an unknown organic liquid by a combination of analytical tools: IR spectrum, chemical tests, refractive index and 1H-NMR. Your unknown will be a liquid with no more than nine carbon atoms and will have only one of the following functional groups: alcohol, alkyl halide, aldehyde, ketone, carboxylic acid or ester. In addition, your unknown may have an ether group. This experiment is different from other experiments you have performed so far. The amount of experimental work is minimal, but you will be required to fully interpret your data and solve the unknown before leaving the lab. As always, the TA will be there to assist you and to clarify any questions, especially about 1H-NMR. You should come to the lab very well prepared with a good working knowledge of IR and, above all, 1HNMR. E33.2 EXPERIMENTAL PROCEDURE
Background reading: 11.1-11.3; Unit 31; Unit 33 Estimated time: 3-3.5 hr

Obtain an unknown liquid sample from the stockroom. Write down the unknowns number in your lab notebook. Obtain the IR of your unknown. Your IR should be nice looking with the bands ending in sharp peaks rather than being flat at the bottom. These flat signals result when too much sample is used. To avoid this, use a very small drop of liquid: touch your liquid with a Pasteur pipet and then just touch the IR plate with the pipet. This will deposit a very small amount of liquid on the plate, enough to obtain a good IR. Cover the plate with the other half and spread the liquid by rotating the plates. By analyzing the IR spectrum you should be able to tell if your unknown contains any of the following functional groups: alcohol, carboxylic acid, aldehyde, ketone, ester. You should also be able to determine if your compound is aliphatic or aromatic. The lack of IR bands for OH and C=O will mean that your compound is an alkyl halide. If you suspect a carboxylic acid, you should further confirm your suspicion by checking the pH of a solution of your unknown in water (one drop of unknown in 1 mL of water). If the pH is clearly acidic (pH < 3) your unknown is likely to be a carboxylic acid. You should also perform the silver-nitrate-in-ethanol test. Most carboxylic acids produce a precipitate of a silver carboxylate when treated with silver nitrate.

Unit 33

If you suspect an alcohol, you should perform the Lucas test. Tertiary alcohols soluble in the Lucas reagent (ZnCl2 in HCl(c)) react immediately and give an insoluble tertiary alkyl chloride that separates as an immiscible layer or as an emulsion. Secondary alcohols soluble in the Lucas reagent react more slowly than tertiary alcohols and cloudiness or an immiscible layer forms within 5-10 minutes. Primary alcohols do not react with the Lucas reagent at room temperature. The Lucas test is limited to alcohols soluble in the Lucas reagent (monofunctional alcohols with fewer than six carbon atoms). If you suspect an aldehyde, a ketone or an ester perform the 2,4dinitrophenyldrazine test. Aldehydes and ketones react almost immediately and give a yellow-orange (sometimes red) precipitate when treated with 2,4-dinitrophenylhydrazine. Esters do not react; they give a negative test as evidenced by the lack of a precipitate. If your 2,4-nitrophenylhydrazine test was positive, perform the Fehlings test as well. Aliphatic aldehydes give a positive test (the formation of a brown-reddish precipitate of cuprous oxide) with the Fehlings reagent. Aromatic aldehydes and ketones do not react. If you suspect an alkyl halide, perform the sodium-iodide-in-acetone test and the silver-nitrate-in-ethanol test. These tests will allow you to distinguish between primary and tertiary alkyl halides. Read sections 11.1-11.3 and measure the index of refraction of your unknown liquid. The TA will show you how to use the refractometer. The index of refraction of your unknown will be in the range 1.3000-1.5500. Make sure that you estimate the fourth decimal place. After you have identified your unknown, compare the index of refraction measured with the literature value as further confirmation of its identity. The Merck Index, the CRC and any other reliable handbook or online source can be used for this. After you have run the IR spectrum, performed the chemical tests and measured the index of refraction, check your results with your TA. If you are on the right track the TA will give you the 1H-NMR to interpret, otherwise he/she will help you reorient. The interpretation of the NMR should be done in the lab. These are the steps to follow: Notice the solvent used to obtain the NMR and make sure that you do not assign solvent or impurity signals, such as water, to your sample. Check Table 33.14 for a list of commonly used solvents and their NMR signals, including water. Integrate the 1H-NMR following the steps discussed in lab lecture (section 33.9; p. 732). Since you do not know the molecular formula, your integration will be in relative terms. However, you should try to anchor the integration by making a reasonable assumption about the number of hydrogens under a certain peak. Your assumption should be based on the functional group present in your molecule and knowing that the unknown is a relative small molecule with 9 or fewer carbons. Example, if you suspect an aldehyde, locate the aldehyde hydrogen. That peak should integrate for only 1H. Interpret the multiplicity of the peaks following the n+1 rule. This will tell you not only the number of neighbors for each type of hydrogen, but it may also be helpful in anchoring the integration. For example if your spectrum shows two triplets and no other multiplet (doublet, triplet, quartet, etc.), its very likely that

Unit 33

each triplet integrates for 2H (the 2H that split the neighbors signal into a triplet). Pay attention to the chemical shifts to determine the environment of each hydrogen (shielded or deshielded). Use the correlation tables (Tables 33.3-33.8) to calculate the chemical shifts of all the hydrogens. The match between your calculations and the experimental values should be acceptable. Measure the coupling constants (section 33.12) and compare them with the expected values (Table 33.2). Keep in mind that contrary to IR, every peak of the NMR spectrum should be accounted for (except for very small peaks that may belong to impurities). If you leave a peak without interpretation, its very likely that your proposed structure is wrong. Before you leave the lab you must present the name and structure of the unknown to your TA and explain your reasoning.

Chemical tests

Interpret your IR spectrum and decide which one(s) of the following tests you will perform. Chemical tests are based on observable changes such as the formation of a precipitate, separation of an immiscible layer or emulsion, a change in color, etc. Record your observations carefully; describe changes in color, precipitate formation, the time it takes for the change to occur and whether it happens at room temperature or after heating. Safety First
Wear gloves when performing the reactions. Gloves should be changed often and removed immediately after completion of the chemical operations. Perform these tests in the fume hood. 2,4-dinitrophenylhydrazine is a skin irritant and may cause skin allergies. Avoid skin contact and inhalation. Hydrochloric acid is corrosive and may cause severe burns. Avoid skin contact and inhalation. Tartrates are irritant. Silver nitrate produces unsightly stains. Avoid skin contact.

Lucas test If you suspect an alcohol, check first its solubility in water: In a small test tube add 1 mL of water and then add a drop of the unknown; mix well. If the unknown doesnt dissolve, consider it insoluble and do not perform the Lucas test below. Its insolubility is an indication that the molecule likely has six or more carbons. If the unknown dissolves in water, go ahead and perform the Lucas test below. In a small test tube place 0.5-0.7 mL of Lucas reagent (ZnCl2 in concentrated HCl) add about seven drops of your unknown and mix well. Note the time it takes to form an emulsion or an insoluble layer. Tertiary alcohols react very fast and form an immiscible layer almost immediately. Secondary alcohols form an immiscible layer within 5-10 minutes. Primary alcohols do not react. Record your observations.

Unit 33

Cleaning up
With the aid of a Pasteur pipet transfer the contents from the Lucas test tubes to a container labeled Lucas Test Waste. Rinse the test tubes with a little ethanol and dispose of the rinses in the same container.

This test is based on the reaction of alcohols with HCl, a nucleophilic substitution reaction catalyzed by acids, that leads to the formation of alkyl chlorides. The test is useful only for alcohols soluble in the Lucas reagent (small alcohols with fewer than six carbons). The product alkyl chloride is insoluble and separates as an immiscible layer or as an emulsion. Thus, the formation of an emulsion or immiscible layer is considered a positive test. The reaction takes place by an SN1 mechanism and thats why tertiary alcohols are the most reactive, followed by secondary alcohols. Primary alcohols do not react with Lucas reagent at room temperature.
R R R OH

C
R

HCl (c)

ZnCl2

C
R

Cl

+ H2O

soluble 3o > 2o >> 1o

insoluble

The role of ZnCl2 is to act as an acid catalyst (Lewis acid).


R R

Zn2+
OH R

Zn O H

C
R

C
R

R R

C
R

+ (ZnOH)+

HCl
R R

C
R

Cl

+ Zn2+

+ H2O

2,4-Dinitrophenylhydrazine test In a disposable 13x100 mm test tube, place 0.5-0.7 mL of 2,4-dinitrophenylhydrazine reagent (2,4-dinitrophenylhydrazine in solution with H2SO4 and ethanol/water) and add five drops of the unknown and mix thoroughly with the help of a glass rod. Aldehydes react almost immediately giving a yellow-orange precipitate; ketones react more slowly and may take a few minutes before they produce a yellow-orange precipitate. In a negative test, no colored precipitate forms. Esters give a negative test. Record your observations.

Unit 33

Cleaning up
With the aid of a Pasteur pipet transfer the contents from the 2,4-dinitrophenylhydrazine test tubes to a container labeled 2,4-DNPH test- liquid waste. Rinse the test tubes with a little ethanol and dispose of the rinses in the same container. Dispose of the test tubes from the 2,4-dinitrophenylhydrazine test in the container labeled 2,4-DNPH test- solid waste.

This test is based on the nucleophilic addition of ammonia derivatives, such as hydrazines (H2N-NH-R), to the carbonyl group of aldehydes and ketones. The mechanism for this reaction (a type of nucleophilic addition to the carbonyl) is discussed on page 419 in the lab book. The end result is the formation of an insoluble 2,4dinitrophenylhydrazone which has a more extended system of conjugation than the original hydrazine and absorbs visible light at longer wavelengths (blue) than the original 2,4-dinitrophenylhydrazine (absorbs violet) resulting in a product that appears orange (the complementary color of blue).
R

C
R

H2N-NHO 2N

NO2

HA

C
R

N-NHO 2N

NO2

+ H2O

2,4-dinitrophenylhydrazine absorbs violet appears yellow

2,4-dinitrophenylhydrazone insoluble absorbs blue appears orange

Esters do not react with 2,4-dinitrophenylhydrazine. Fehlings test In a small test tube place 0.5 mL of Fehlings reagent A (copper sulfate in water) and 0.5 mL of Fehlings reagent B (sodium hydroxide and sodium potassium tartrate in water). Add two drops of the liquid sample and heat the liquid in a boiling water bath for about 10 minutes. A positive test is the formation of a reddish precipitate of cuprous oxide (Cu2O) or metallic copper deposited on the walls of the tube (copper mirror). Aliphatic aldehydes give a positive test. Aromatic aldehydes and ketones do not react. Record your observations. Cleaning up
With the aid of a Pasteur pipet transfer the contents from the Fehlings test tubes to a container labeled Fehlings Test Waste. Rinse the test tubes with a little ethanol and dispose of the rinses in the same container.

This test is discussed on pages 420 and 429 of the lab book. Aldehydes, especially aliphatic aldehydes, can be easily oxidized with Cu2+. The aldehyde gets oxidized to a carboxylic acid (or a carboxylate under the basic conditions of the test) and the copper gets reduced to Cu+ (in the form of Cu2O) and occasionally to Cu0 (metallic copper). The mechanism for this reaction is complex and not fully understood and we will not discuss it here.

Unit 33

C O

+ 2 Cu2+ blue

+ 5 HO-

tartrate heat

C O

+ 3 H2O + Cu2O reddish precipitate

an aliphatic aldehyde

a carboxylate

The reaction, a redox process, is accompanied by a drastic change in color. Cu2+ under basic conditions is deep blue, and the product of the reaction, usually Cu2O rather than metallic copper, is a reddish precipitate. The reaction is performed in the presence of tartrate to keep the Cu2+ in solution under basic conditions. Tartrate is what we call a chelating agent. A chelating agent is an organic compound that can form two or more coordinate bonds with one central metal ion. Chelating agents are important in industry, medicinal chemistry and biological process for their ability to form stable complexes with metals (as an example, think of the heme group in hemoglobin as a chelating agent for the Fe2+ ion found in its center). In the presence of strong base (NaOH) needed for the redox process, Cu2+ would precipitate out in the form of insoluble Cu(OH)2 and would not react with the aldehyde. To prevent this from happening, tartrate is added along with the NaOH. The tartrate forms a complex with the Cu2+ and keeps it in solution where it can react with the aldehyde. The tartratecopper complex is deep blue.
O
HO O

O
OH

O
O O

Cu2+ +

2
HO O O

2 HO-

Cu
O O O HO O O

+ 2 H2O

tartrate

blue complex

Silver nitrate in ethanol test In a small and perfectly dry test tube place 1 mL of a 0.1 M solution of silver nitrate in ethanol. Add two-three drops of your unknown and mix well. Observe the formation of a precipitate and notice how long it takes to form. If no precipitate is observed within five minutes at room temperature, place the solution in a water bath at approximately 80oC and observe if a precipitate forms. The reaction is considered positive if a precipitate forms. Tertiary alkyl halides and allylic halides react very fast, secondary alkyl halides react slower and primary alkyl halides only after heating. Aryl halides and vinylic halides give a negative test even upon heating. In general, alkyl iodides react faster than alkyl bromides and alkyl bromides faster than alkyl chlorides. Carboxylic acids produce an insoluble silver carboxylate. Carefully record your observations. This test is also discussed on p. 276 and 285 of the lab book. Alkyl halides react with silver nitrate in ethanol and give silver halide which separates as a precipitate. The formation of this precipitate is considered a positive test.

Unit 33

R X

+ AgNO3

ethanol

R ONO2 + AgX

The reaction takes place by an SN1 mechanism and therefore tertiary alkyl halides react faster than secondary. Primary alkyl halides react slowly and give a precipitate only upon heating. Carboxylic acids react with silver nitrate in ethanol and produce a precipitate of silver carboxylate.
R-COOH + AgNO3
ethanol

R-COOAg

+ HNO3

Sodium iodide in acetone test In a small and perfectly dry test tube place 1 mL of a 0.1 M solution of a 15% (w/v) solution of sodium iodide in acetone. Add two-three drops of your unknown and mix well. If no precipitate is observed after three minutes at room temperature, place the solution in a water bath at approximately 50oC for about five minutes and observe if a precipitate forms. The reaction is considered positive if a precipitate of sodium chloride or sodium bromide forms. Bromides react faster than chlorides. Primary alkyl bromides react at room temperature; primary and secondary alkyl chlorides and secondary and tertiary alkyl bromides react upon heating at 50oC. Tertiary alkyl chlorides, aryl halides and vinylic halides do not react even after heating. Record your observations. This test is also discussed on p. 277 and 285 of the lab book. Alkyl chlorides and alkyl bromides react with sodium iodide in acetone and give an alkyl iodide and sodium chloride or sodium bromide, respectively, which are insoluble in acetone. The formation of a whitish precipitate of sodium chloride or bromide is considered a positive test.
R X + NaI
acetone

R I

+ NaX

X = Cl, Br

This test is based on the limited solubility of sodium chloride and sodium bromide in acetone; while sodium iodide (the reagent) is soluble in acetone, sodium chloride and sodium bromide (the possible side products) are not. The reaction takes place by an SN2 mechanism and, therefore, primary alkyl bromides and chlorides react faster than secondary ones. Tertiary alkyl chlorides, aryl halides and vinyl halides do not react. Cleaning up
With the aid of a Pasteur pipet transfer the contents from the silver nitrate and sodium iodide test tubes to a container labeled Alkyl Halides Tests Waste. Rinse the test tubes with a little ethanol and dispose of the rinses in the same container. Dispose of the unused unknowns in the bag labeled Unknowns-waste.

EXPERIMENT 33 REPORT

Unit 33

Pre-lab 1) How would you distinguish each compound in the following pairs by IR spectroscopy? Give the bands (with their assignments) that you would use to tell each compound apart. If you do not know their structures check them out in a textbook or online: a) 1-pentanol and 1-pentanal b) benzoic acid and phenol c) cyclohexanone and benzaldehyde d) methyl acetate and acetic acid 2) Write a chemical reaction for 3-methyl-3-pentanol and the Lucas reagent. Show the mechanism for the reaction. What is considered a positive result for the Lucas test? 3) Which alcohols react faster with the Lucas reagent? Why? 4) Write a chemical reaction for acetone and 2,4-dinitrophenylhydrazine. 5) Explain how 2,4-dinitrophenylhydrazones are more conjugated than 2,4dinitrophenylhydrazine. 6) What is considered a positive 2,4-nitrophenylhydrazine test? 7) Write a chemical reaction for formaldehyde and the Fehlings reagent. 8) What is considered a positive Fehlings test? Do aromatic aldehydes react? 9) Using chemical reactions, explain the role of tartrate in the Fehlings test. What is a chelating agent? 10) Write a chemical reaction for 2-methyl-2-chlorobutane and silver nitrate in ethanol. What is considered a positive test? Which alkyl halides react faster? 11) Write a chemical reaction for 1-bromobutane and sodium iodide in acetone. What is considered a positive test? Which alkyl halides react faster? 12) How would you distinguish these two compounds by 1H-NMR? Use chemical shift arguments. Include calculation of chemical shifts using correlation tables.
O O

13) Using the concept of critical angle (show a drawing), give the definition of the index of refraction in the form of an equation. In-lab 1) Identify your unknown. Draw its structure and name it. Briefly explain your reasoning. 2) Interpret the IR of your unknown. Present the important bands in the usual table format. Which band(s) did you use to identify the functional group? 3) Report the results of your chemical tests, including pH determination, if applicable. Interpret and discuss the results of such tests. 4) Report the index of refraction of your unknown. How does it compare with the index of refraction reported in the literature? 5) Interpret the 1H-NMR. Account for integration, coupling and chemical shift. Present the data in a table format. Show your integration calculations. Account for coupling according to the n+1 rule. Calculate the chemical shifts of all the hydrogens using correlation tables. Measure the coupling constants and compare them with table values (Table 33.2). Show all your calculations.

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