Classifying Compounds containing Hydroxyl- and Carbonyl groups using specified tests

Abraham, K.D., Acebedo, D.M.A., Alap-ap, M.L.M., Asprec, W.A.A., Balaaldia, K.C.A. and Basilio, Z.A.S. 2B-PH, Group No. 1, Department of Pharmacy, Faculty of Pharmacy, University of Santo Tomas, España Boulevard, 1015 Manila, Philippines

ABSTRACT

Hydroxyl- or carbonyl- containing samples was given for this experiment for analysis. Hydroxyl group refers to a functional group containing OH- when it is a substituent in an whereas carbonyl group refers to a divalent chemical unit consisting of a carbon and an oxygen atom connected by a double bond [1]. Hydroxyl group is the characteristic functional group of and while carbonyl group is the characteristic functional group of aldehydes and [2]. In this experiment, several differentiating tests were conducted with samples ethanol, n-butyl , sec-butyl alcohol, tert-butyl alcohol, benzyl alcohol, n- butyraldehyde, benzaldehyde, , acetophenone, , and acetaldehyde. These tests include solubility of alcohols in water, Lucas test, Chromic Acid test, 2,4- Dinitrophenylhydrazone test, Fehling’s test, Tollens’ Silver Mirror test, and Iodoform test. The first three test done helped in the identification of the structure of the alcohols and tell if they are Primary (the alpha carbon atom where OH group is attach is attached to only one alkyl group), secondary (the alpha carbon is attached to two alkyl group), or tertiary (the alpha carbon is attached to three alkyl groups). The fourth test, The 2,4-Dinitrophenylhydrazon test was used to identify the presence of carbonyl groups-, aldehydes and ketones. 2,4-Dinitrophenylhydrazone was also used to distinguish its aromaticity. A positive result of red-orange precipitate indicates a carbonyl group, while a yellow precipitate indicates a presence of aldehydes and ketones. The fifth and sixth test, the Fehling’s and Tollens’ Silver Mirror test were used to identify the presence of an aldehyde and distinguishing an aldehyde from a . In this experiment, only an aldehyde would yield a positive result while ketones won’t. And lastly, the Iodoform test was used to ensure the presence of CH3CO group or methyl carbonyl group in the samples given.

INTRODUCTION group –OH when it is a substituent in an An alcohol is a compound that has a organic compound. Hydroxyl groups are hydroxyl group bonded to a sp3-hybridized known for their tendency to form hydrogen carbon atom, R-OH. They are also classified bonds either as a donor or as an acceptor. as primary (1”), secondary (2”) or tertiary This is also related to their ability to increase (3”), depending on the number of carbon hydrophilicity and water solubility [3]. substituents bonded to the hydroxyl-bearing The functional group of an aldehyde is a carbon. Their most important physical carbonyl group bonded to a hydrogen atom. property is the polarity of their –OH groups. Another carbonyl group is a ketone bonded Due to the large difference in the to two carbon atoms. Due to the polarity of a electronegativity of C-O and O-H, both of carbonyl group, aldehydes and ketones are their bond of an alcohol are polar covalent polar compounds and interact in the liquid and alcohols are polar molecules. Hydroxyl state by dipole-dipole interaction. The result group is used to describe the functional is that aldehyde and ketones have higher boiling points than those of nonpolar primary and secondary resulting in the compounds with comparable molecular reduction of the orange chromium Cr6+ ion weight [4]. to a blue-green Cr3+ ion. The Lucas test is performed with the use of Carboxylic acids and their derivatives are the Lucas Reagent, where Lucas reagent is a the most abundant of all organic compounds solution of anhydrous zinc chloride in in living organisms and in the laboratory. concentrated hydrochloric acid. This reagent The most common derivatives of carboxylic is used to classify alcohols of low molecular acids are acyl halides, acid anhydrides, weights. This reaction shows a substitution, esters, and amides. They contain acyl group which the chloride replaces a hydroxyl attached to a nucleophilic molecule that group. A positive result indicated the replaced the –OH group of a carboxylic formation of a chloroalkane which is based acid. Meaning that Jones test is used to on the difference in reactivity of the three check for the presence of a primary and classes of alcohols with hydrogen halides. secondary alcohol both of which shows a The difference in reactivity shows the positive result of the formation of a green different ease of formation of the color. Here is the reaction mechanism in carbocations. Tertiary carbons are the most Chromic Acid Test [6]. stable and Primary carbons are the least stable. Tertiary alcohols react immediately to with the Lucas Reagent, Secondary alcohols react within a few minutes while Primary alcohols do not react with Lucas Reagent at room temperature. Meaning that the time taken for a positive effect to be visible is the measure of the reactivity of the class of alcohol, and it is also the way of determining the class of a alcohol the solution is. Here is the reaction mechanism in Lucas Test [5].

Figure 2 : Jones’ test Rxn Mechanism

The 2,4-Dinitrophenylhydrazone (2,4-DNP) test serves as to form a derivative. The Figure 1 : Lucas Test Rxn Mechanism aldehydes and ketones react with 2,4-DNP to form a solid 2,4-DNP Derivative. If the The Chromic acid test, also known as Jones solid is yellow, this means that the carbonyl Oxidation test, differentiates primary and group in the unknown is unconjugated while secondary alcohols from tertiary alcohols. A a reddish-orange color most likely means primary alcohol is oxidized to an aldehyde that the carbonyl group is conjugated. or to a carboxylic acid, while a secondary Meaning this test help determine whether alcohol to a ketone. Tertiary alcohols in the the carbonyl group is alipathic or aromatic. other hand do not react. The OH-bearing The reddish-orange color means that the carbon must have a hydrogen atom attached substance is aromatic and the yellow color since that carbon atom is being oxidized in means that the substance is alipathic. Here is The Tollens’ Test, also known as Silver the reaction mechanism in 2,4-DNP [7]. Mirror Test, is used to distinguish between an aldehyde and a ketone. This test also has the same function as the Fehling’s Test. This test uses a reagent known as Tollens’ reagent, which is a colorless, basic, aqueous solution containing solver ions coordinated + to ammonia [Ag(NH3)2 ]. Tollens’ reagent oxidizes an aldehyde into the corresponding carboxylic acid. A positive result would make the reagent metallic silver in color and shows a mirror-like expression in the test tube. Ketones are not reactive with the Tollens’ reagent, so it would not show a mirror-like impression in the test tube. Here is the reaction mechanism in Tollens’ Test Figure 3 : 2,4-DNP Rxn Mechanism [9].

The Fehling’s test is used to determine the presence of an aldehyde in the solution and it doesn’t react with ketones. Aldehydes reduces to make the deep blue solution of copper (II) to muddy green solution, and then forms a brick-red precipitate of insoluble cuprous oxide (Cu2O). Meaning this test is usually used for reducing sugars but us known to be not specific with aldehydes. Here is the reaction mechanism in Fehling’s Test [8]. Figure 5 : Tollens’ Test Rn Mechanism

Lastly, the Iodoform test used to see the presence of CH3CO in aldehydes and ketones. The sample is allowed to react with a mixture of iodine and . The alpha hydrogens of the carbonyl group are acidic and will react with a base to form the anion, which then reacts with iodine and forms triiodo compound which then reacts with another base to form the carboxylic acid salt plus a iodoform. A positive result would produce a yellow precipitate, and if there is no CH3CO group in the sample being tested, then it will get different result. Here is the Figure 4 : Fehling’s Test Rxn Mechanism reaction mechanism in Iodoform Test [10]. alcohol was added. The test tube was covered and shaken vigorously and was allowed to stand at room temperature. The 2 other test tubes were placed with 2-3 drops of sec-butyl alcohol and tert-butyl alcohol. The test tubes were shaken vigorously and the mixtures were allowed to stand. Note the time of the formation of the cloudy suspension or the formation of two layers.

Figure 6 : Iodoform Test Rxn Mechanism C. Chromic acid test (Jones’ Oxidation) Six test tubes were prepared as well as six METHODOLOGY samples. The samples are n-butyl alcohol, isopropyl alcohol, tert-butyl alcohol, The materials needed for this experiment in acetaldehyde, benzaldehyde, and acetone. this experiment are the following: Lucas One drop of each sample was placed on Reagent, Chromic Acid reagent, 95% different test tubes and was dissolved in ethanol, Fehling’s A and B, Tollens’ reagent, 1mL of acetone followed by the addition of 5% NaOCl solution, Iodoform test reagent, 5 drops of the reagent dropwise with 2,4-dinitrophenylhydrazine, Pasteur pipette, continuous shaking. Place the test tubes in test tubes and vials and some beakers. While 60oC water bath for 5 minutes and note the the sample compounds needed are the color of each solution. following: ethanol, n-butyl alcohol, sec- butyl alcohol, tert-butyl alcohol, benzyl D. 2,4-Dinitrophenylhydrazone Test alcohol, n-butyraldehyde, benzaldehyde, Four test tubes were labelled with the acetone, acetophenone, isopropyl alcohol, samples to be used. The samples used were acetaldehyde. acetone, acetaldehyde, benzaldehyde, and acetophenone. One drop of each sample was A. Solubility of Alcohols in Water placed in different test tubes followed by the Five test tubes were labelled and ten drops addition of 5 drops of 95% ethanol with each of ethanol, n-butyl alcohol, sec-butyl continuous shaking. Then add 3 drops of alcohol, tert-butyl alcohol, and benzyl 2,4-DNP. If no yellow or orange-red alcohol were placed on the test tubes with precipitation forms, allow the solution to the use of a Pasteur pipette. Followed by stand for 15 minutes. Note the results. addition of 1mL of water and shaking of the test tube. If cloudiness occurs, continue E. Fehling’s Test adding a quarter of an mL of water with Into each test tube, 1mL of freshly prepared vigorous shaking, until a homogenous Fehling’s reagent (made by mixing equal dispersion results. If no cloudiness results amounts of Fehling’s A and Fehling’s B) after the addition of 2mL of water, the was placed. 3 drops of the sample to be alcohol is water soluble in water. tested were added and the test tubes were placed into a beaker of boiling water. B. Lucas Test Changes were observed that occurred within Three test tubes were prepared and 1mL of 10-15 minutes. The samples to be used are Lucas Reagent was dropped in each of them. acetaldehyde, acetone, benzaldehyde, and One the first test tube, 2-3 drops of n-butyl acetophenone. Here 1 mL of benzamide was treated F. Tollens’ Silver Mirror Test with 5mL of NaOH solution and was got Four test tubes were prepared that contains immersed in a boiling water bath. During 1mL of freshly prepared Tollens’ reagent heating, a moist red litmus paper was placed and two drops of each sample was placed over the test tube and any change in the into separate test tubes. The samples were litmus paper was noted. acetaldehyde, benzaldehyde, acetone, acetophenone. Shake the mixture well and H. Alcoholysis: Schotten-Baumann Rxn allow it to stand for 10 minutes. If no Acetic Acid reaction has occurred, place the test tubes in In this procedure, 10 drops of acetic a beaker with warm water for 5 minutes. acid, 1mL ethanol, and 5 drops of Record any observation. concentrated H2SO4 was warmed on a water bath for 2 minutes and then note the odor of I. Iodoform Test the ester formed. 2 drops of each sample (acetaldehyde, acetone, acetophenone, benzaldehyde, and Acyl halides and Acid anhydrides isopropyl alcohol) were placed in different In another test tube, we placed 0.5 test tubes. 10 drops of 10% KI solution were mL ethanol, 1mL water, and 0.2mL of acyl added. Followed by the addition of 20 drops halide or acid anhydride sample. Then add of fresh chlorine bleach (5% sodium 2mL of 20% NaOH solution. Plug the test hypochlorite) were added slowly to each test tube using a cork stopper or parafilm and tube with continuous shaking. The formation shake the mixture for several minutes. of a yellow precipitate was noted. Afterwards, note the odor and the layers of the ester formed. G. Hydrolysis of Acid Derivatives Acid Halides and Acid anhydrides I. Aminolysis: Anilide Formation In a test tube, 1mL of water was Acyl halides and acid anhydrides placed, and then 10 drops of the sample was A few drops of either acetyl chloride cautiously added dropwise. The resulting or were added to 0.5mL mixture was divided into two portions. 1mL aniline. The mixture was then transferred to of 2% AgNO was added to the first test tube a new test tube containing 5mL of water. while 1mL of saturated NaHCO3 was added Formation of precipitation was then to the 2nd portion. observed.

Esters J. Hydroxamic Acid Test 2mL of 25% NaOH solution was Preliminary test was done by mixing added to 1mL of ethyl acetate. The mouth of 1mL of 95% ethanol and 1M HCl to the test tube was covered with a marble then separate test tubes that contained drops of the test tube was heated in a boiling water the sample to be used. If upon addition of 1 bath for 5 minutes. Afterwards, the mixture drop of 5% FeCl3, a color other than yellow was neutralized with 10% HCl solution. The will make the test futile. 2mL of alcoholic test tube was wafted to take note of the odor. NH2OH∙HCl and 1mL of 1M KOH was added to separate test tubes of 2 drops ethyl acetate and acetamide. Immersion in a boiling water bath for 2 minutes was done; afterwards the mixture was allowed to cool. Amides 1mL of 5% FeCl3 was introduced in each true for organic compounds that have the test tube. Then the color of the formed same number of carbon atoms present. precipitate was noted. The solubility of alcohols decreases as the length of the hydrocarbon chain of the RESULTS AND DISCUSSION alcohol increases. For the test of the solubility of alcohols in The next test was conducted with the use of water, the turbidity of the solution was the Lucas Reagent and it differentiated 1”, observed and it was the basis if the specific 2” and 3” alcohols. Alkyl chloride formation alcohol is soluble to water. Also, the amount was observed and caused turbidity or of water needed to produce homogenous cloudiness. Also, the rate of the reaction was dispersion was observed. Table 1 shows the observed. Table 2 shows the result of Lucas data gathered from the test. Test. Amount of Sample Observation water needed n-butyl alcohol Homogenous solution Solubility Alcohol to produce Sec-butyl alcohol Homogenous solution to water homogenous Tert-butyl alcohol Formation of 2 layers dispersion Table 2 : Lucas Test results ethanol 1mL Soluble According to the table above, n-butyl n-butyl alcohol 1.50mL Soluble alcohol and sec-butyl alcohol are both sec-butyl 1mL Soluble soluble in the Lucas Reagent while tert- alcohol butyl alcohol formed a cloudy layer. The Tert-butyl 1mL Soluble absence of visible reaction at room alcohol temperature and the appearance of the Benzyl alcohol 2mL Insoluble cloudy layer on the application of heating, represents that it is a primary alcohol. A Table 1 : Solubility of Alcohols in Water delayed formation of a cloudy layer; around Table 1 shows the solubility of alcohols to 3-5 minutes tells us that it is a secondary water and the amount of water that is needed alcohol. And lastly, if the substance turn to produce a homogenous dispersion.Out of cloudy immediately after the addition of the 5 alcohol samples, only Benzyl Alcohol Lucas reagent and the phases separate means is insoluble. The principle behind this is that that it is a tertiary alcohol. Basing on the “like dissolves like”. It can be said that the table above, tert-butyl alcohol is a tertiary other alcohols are soluble to water because alcohol due to the immediate clouding of the they exhibit a polar bond and water is also alcohol and the separation of layers. known to have a polar bond as well. There Chromic Acid test (Jones Oxidation) permits are factors that affect the solubility of the conversion of primary alcohols to alcohols to water. One of these factors is the aldehydes and secondary alcohols to ketones number of carbon atoms present. The lower through oxidation. Table 3 shows the results the number of carbon atoms present, the of Chromic Acid test. more soluble or miscible a substance is. Another factor is the branching of carbon chains. The more branching present, the more soluble a compound is. This is only In the test, acetophenone was the only sample to give an orange-red ppt. while the other samples gave a yellow-orange and Sample Observation canary yellow precipitates. The orange-red n-butyl alcohol Blue-green solution precipitate represents the presence of a Tert-butyl alcohol Dirt yellow solution conjugated carbonyl compound and the Acetaldehyde Blue-green solution appearance of the yellow precipitate Benzaldehyde Blue-green solution indicates the presence of unconjugated Acetone Green solution carbonyl compound. Acetophenone Dirty yellow solution Fehling’s Test is another differentiating test Isopropyl Alcohol Blue-green solution for aldehydes and ketones. In this test aldehydes reacted in this test while ketones Table 3 : Chromic Acid test results did not produce any reaction. In the data gathered, n-butyl alcohol, Sample Observation acetaldehyde, benzaldehyde, and isopropyl alcohol gave a positive result of a blue-green Acetaldehyde Brick-red ppt. solution while tert-butyl alcohol and Benzaldehyde Brick-red ppt. acetophenone both gave a dirty yellow Acetone Dark blue solution solution while acetone gave a green Acetophenone Royal blue solution solution. Chromic test or Jones Oxidation Table 5 : Fehling’s test results involved reduction-oxidation or redox reaction. 1” and 2” alcohols and aldehydes As shown on the table below above, went through oxidation and chromium acetaldehyde and benzaldehyde both underwent reduction from Cr6+ to Cr3+. The produced a brick-red precipitate indicating reduction of the chromium ion which is that they are both aldehydes. Meanwhile colored orange resulted to a blue-green acetone and acetophenone did not produce a solution telling us that the 1” and 2” reaction what so ever. Fehling’s test alcohols were oxidized. Ketones are the involved reduction-oxidation or redox result of the reaction of 2” alcohols with reaction. Aldehydes got oxidized while chromic acid, which do not oxidize further. ketones did not undergo oxidation. Tertiary alcohols are nonreactive and Tollens’ Silver Mirror test differentiated aldehydes got oxidized to carboxylic acids. aldehydes from ketones wherein aldehydes The next test, which is 2,4- were expected to be oxidized while ketones Dinitrophenylhydrazne test / 2,4-DNP is the did not undergo any oxidation. Table 6 test used for distinguishing the presence of shows the results on Tollens’ Silver Mirror an aldehyde or ketone in a compound. Table test. 4 shows the data gathered from the test. Sample Observation Sample Observation Acetaldehyde Silver mirror Acetaldehyde Yellow-orange ppt. Benzaldehyde Gray sol’n w/ globules Benzaldehyde Yellow-orange ppt. Acetone Colorless sol’n Acetone Canary yellow ppt. Acetophenone Turbid gray sol’n Acetophenone Orange-red ppt. Table 7 : Tollens’ Silver Mirror test results Table 4 : 2,4-DNP test results On the table above, only acetaldehyde formed a silver mirror. Benzaldehyde, even though it is an aldehyde did not form any silver mirror. Acetone formed a colorless solution and acetophenone produced a turbid gray solution. This test is similar to Fehling’s test making it undergo redox From the Internet reaction as well. Aldehydes got oxidized while ketones did not. 3. Clark, J. (2003). An Introduction to Alcohols. Retrieved from Iodoform test was used to detect the http://www.chemguide.co.uk/organic presence of an aldehyde or a ketone in a props/alcohols/background.html substance where methyl groups are attached 4. Clark, J. (2003). Aldehydes and directly to the carbonyl carbon. Ketones. Retrieved from Sample Observation http://www.chemguide.co.uk/organic Acetaldehyde Yellow ppt. props/carbonyls/background.html n-butylraldehyde Yellow solution 5. Lucas Reagent (n.d.) Retrieved from Benzaldehyde Red ppt. w/ globules http://www.chemistrylearner.com/luc as-reagent.html Acetone Yellow ppt. 6. Test for Aldehydes and Ketones (n.d.) Acetophenone Yellow ppt. Retrieved from Isopropyl Alcohol Yellow crystal ppt. http://academics.wellesley.edu/Chem Table 7 : Iodoform test results istry/chem211lab/Orgo_Lab_Manual /Appendix/ClassificationTests/aldehy As shown in the table above, acetaldehyde, de_ketone.html acetone, and acetophenone produced a 7. 2,4-Dinitrophenylhydrazine test yellow precipitate. Benzaldehyde resulted (n.d.) Retrieved from produced a red precipitate with globules. http://www.harpercollege.edu/tm- Isopropyl alcohol resulted in a yellow ps/chm/100/dgodambe/thedisk/qual/ crystalline precipitate and n-butyraldehyde dnp.html resulted in a yellow solution. The 8. Fehling’s Test (n.d.) Retrieved from appearance of a yellow precipitate or a yellow crystal indicates a positive result. It https://fenix.tecnico.ulisboa.pt/downl also tells us that the carbonyl group of the oadFile/3779571247498/Testes sample has a methyl group attached directly to it. %20de%20a%C3%A7ucares- REFERENCES alunos.pdf 9. Tollens’ Test (n.d.) Retrieved from From books http://chemwiki.ucdavis.edu/? 1. Brown, W., Poon, T. (2011). title=Organic_Chemistry/Aldehydes Introduction to organic chemistry _and_Ketones/Reactivity_of_Aldehy international student version (5th des_%26_Ketones/Tollens edition). NJ, USA: John Wiley & %E2%80%99_Test Sons, Inc. 10. Clark, J. (2004). Iodoform Reaction 2. Zumdahl, S., Zumdahl, S. (2012). with Aldehydes and Ketones. Chemistry: An Atoms First Approach Retrieved from (International edition). USA: http://www.chemguide.co.uk/organic Brooks/Cole, Cengage Learning. props/carbonyls/iodoform.html