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PART THREE ORGANIC CHEMISTRY (HCT403) PRACTICALS
PRACTICAL 1:Isolation of caffein from tea
PROCEDURE
Place 125ml of distilled water in to 500ml Erlenmeyer flask.Add 12,5g of tea and 12,5g of powdered calcium carbonate.Boil the contents of the flask with constant stirring for about 20 minutes. At this time, filter the hot mixture with suction, and press out theliquid from the tea leaves with a large cork. Transfer the filtrate to a 250ml Erlenmeyer flask and add 100ml of chloroform. Gently stir the two layers together for about 10 minutes and then separate them by means of a separatory funnel. Distil the chloroform layer until only about 10ml remains, transfer the remaining solution to a tared (weighed) 25ml Erlenmeyer flask (filtering if necessary), and evaporate to dryness on the steambath. Recrystallise the solid residue of caffeine from 95% ethanol, using 5 ml per gram. Record your yeild.
- Make an infrared spectrum of caffeine.
- Draw the chemical structure of caffeine.
- Name two other natural products that contain caffeine.
- Determine the melting point of caffeine.
PRACTICAL 2: Carbohydrates
In this experiment, tests that distinguish among various carbohydrates will be performed.
The carbohydrates include:
(i)Aldopentoses:xylose and arabinose
(ii)ii) Aldohexoses: glucose and galactose
(iii)(iii) Ketohexase: fructose
(iv)(iv) Disaccharides: lactose and sucrose
(v)(v) Polysaccharides:starch and glycogen
The structures and reactions of these are expected to be known. The tests are classified as follows:
1)Test based on the reducing property of a carbohydrate(sugar)
(a) Benedict ‘ s test
(b) Barfoed`s test
2) Osazone formation
3) Iodine test for starch
4) Hydrolysis of sucrose
5) Mucic acid test for galactose and lactose
Special Instructions
All procedures in this experiment involves simple test- tube reactions. Most of these are short;osazone formation and the mucic acid test, take relatively longer to complete.You will need a minimum of 10 test tubes, numbered in order. Clean them carefullyeach time they are used. The 1% solutions of carbohydrates and reagents needed for tests will hopefully have been prepared in advance.Be sure to shake the starch solution before using it.Record results and observations and where reaction equations are involved present these concisely and clearly.
1.Test based on Reducing Properties of sugars
(a)Benedict`s Test :
This test is performed under mildly basic conditions, to produce the red precipitate, cuprousoxide. The reagent also reacts with water-soluble aldehydes that are not sugars. Benedict`s test is considered one of the classical tests for determining the presence of an aldehyde functional group.
Procedure
Prepare a boiling water bath for this experiment. Place 1ml of each of the following 1% carbohydrates solutions in separate test tubes: xylose,arabinose,glucose,galactose,fructose,lactose, sucrose,starch(shake it) and glycogen. Add 1ml of distilled water to a reverse test tube (control).Add 5ml of Benedict`s solution to each test tube. Place the test tube in a boiling-water bath for 2-3 minutes. Remove the test tube and note the results.A red , brown or yellow precipitate indicates a positive for a reducing sugar.
(b)Barfoed`s test:
Barfoed`s test distinguishes reducing monosaccharides and reducing disaccharides by the difference in the rate of reaction .The reagents consists of copper ions like Benedict`s reagent. In this test, however, Barfoed’s reagent reacts with reducing monosaccharides, to produce cuprous oxide,
faster than with reducing disaccharides.
Procedure
Place 1ml each of the above carbohydrates.[Procedure reagent to each test tube .Place the tubes in a boiling-bath for 10mins. Remove the tubes and note the results.
2.Osazone formation
A good experimental use for the osazone is to observe the rate of its formation .The reaction vary greatly even though the same osazone may be produced from different sugars.The crystal structure of the osazone is also distinctive e.g arabinose(fine precipitate), glucose(course precipitate).
Procedure
Place 0.1g of each of the carbohydrates in 1(a) in separate test tubes .Then add 1ml of phenylhydrazine
reagent (CARE) to each test tube. Place the tubes in a boiling water-bath simultaneously .Watch for a precipitate and record results (observation and time).After 30mins , cool the tubes and record crystalline form of the precipitate (microscope may help).Reducing disaccharides will not precipitate until the tubes are cooled. Non –reducing disaccharides will hydrolyse first before osazone precipitation.
3.Iodine test:
Check-up the theory surrounding the iodine test.
Procedure
Place 2ml of each of the 1% carbohydrate solutions in three separate test tubes: glucose, starch and glycogen.Add 2ml of distilled water to another test tube to act as a control.Add 1 drop of iodine solution to each test tube and observe .Add a few drops of sodium thiosulphate to the solutions and record the results.
4,Hydrolysis of sucrose:
Read the underlying theory of hydrolysis.
Procedure
Place 5ml of 1% solution of sucrose in a test.Add 2 drops of conc HCl and heat the tube in a boiling water-bath for 10mins.Cool and neutralize the contents with 10% NaOH until the mixture is just basic to litmus.Test the mixture with Benedict`s reagent.Note theresults and compare these with the results on unhydrolysed sucrose[1(a)].
5.Mucic acid test:
Place 0.2g lactose ,0.1g of glucose and 0.1g of galactose in three separate test-tubes .Add 2ml of water to each tube and dissolve the solids, with heating if necessary .Add 2ml of conc HNO3 to each of the tubes. Heat the tubes and allow them to cool for an hour in a hood.
Remove the tubes and allow them to cool slowly.
Triturate the tubes to induce crystallization .Record your results (ca 1/2 hr afterwards results should be noticeable).
Allow the test tubes to stand until the next laboratory session, to complete the crystallization . Mucic acid is insoluble in water .Confirm this, observation. Account for your observations in this experiment.
6.Tests on Unknown:
Obtain an unknown solid carbohydrate from the technicians .Carefully dissolve part of the unknown (0.25g) in distilled water (25ml) i.e a 1% solution .Save the remainder for the test requiring solid material .Apply whatever
Tests are necessary to identify the unknown.
PRACTICAL 3:ANTHRAQUINONE AND ANTHRACENE
Compound A
Place phthalic anhydride (1g) and thiophene-free benzene (36ml) in a 200ml round bottomed flask . Cool the flask in an ice bath util benzene starts to crystallise and then add anhydrous aluminium chloride (15g).Equip the flask with a condenser connected to HCl gas traand shake the flask well and warm it for a few minutes by hand to start the reaction (if necessary cautiosly warm gently over water bath ).Cool the flask in an ice bath if the reaction becomes too vigorous.When the reaction proceeds smoothly at room temperature(in about 5min) carefully reflux and swirl the mixture on a water bath for about 20 min.Remove the condenser and cool the mixture by swirling in an ice bath .(should ni complex separate ,heat for 10 minutes more before proceeding). Under a fume hood , carefully add ice (50g) in small portions , swirl the flask and cool as necessary .Add conc HCL (10ml), water (50ml) and swirl vigorously and again add water (25ml) and shake vigorously.
`Dislodge solid with a stirring rod from walls of the flask and break up the lumps.Pour the mixture into a separatory funnel, rinse the flask with ether (30ml) and add it into the funnel.Shake gently the funnel and discard the lower phase.To reduce the fluffy precipitate that appears at the interface add conc HCL (5ML) and water (12ml) , shake vigorously and drain off the lower layer (if necessary filter the organic layer).Dry the organic layer with anhydrous sodium sulphate and evaporate until the volume is reduced to about 55ml. Add ligroin (petroleum ether) slowly to slight turbidity and allow product A to crystallise at room temperature and finally at 50C .
Determine m.p. Record IR, UV and NMR spectra of the product in the following synthesis and submit the remainder for marking. Write the structure for A.
Anthraquinone:
Place the intermediate product A (2.0g) from above in a 50ml round bottomed flask , add conc sulphuric avid (12ml) and heat on the steam bath with swirling until the solid is dissolved.Then insert a thermometer into the flask and heat with microburner to 1500C and maintain atb 1500C for 5 min. Let the solution cool to 1000C, remove the themometer and add water (5ml) drop by drop with swirling. Let the mixture cool and dilute with water until the flask is full again, letit cool. Collect the product by filtration and wash with dilute ammonia untilthe filtrate from the funnel is basic. Finally wash the product with distilled water until the washings are neutral .Dry the product in an oven (m.p. 286oC) .
Determine yield (using product A as starting material ).Record IR ,UV, andNMR. Use the product in the next reaction.The remainder submit for marking.
Compound B
Mix anthrraquinone (2.5g),glacial acetic acid (20ml0 and a solution made by warming stannous chloride dihydrate (6mg) in conc hydrochloric acid (6ml0 in 50ml flask equipped with a condenser .Reflux gentle with a burner until anthraquinone is dissolveed (about 8 min) , the refux 15min longer.Disconnect the flask ,heat it on water bath (1000C), and add water(about 6ml) in portions until the solution is saturated.Let the solution cool for crysallisation.Filter and dry the product .Determine the yield andd m.p.Record IR,UVand NMR spectra.Write the structure forB.
Anthracene:
Place zinc dust (5g) in a round botomed flask and activate the dust by adding water (30ml) and cupper sulphate solution (Fehling solution I) and swirl for a minute.Add compound B(2g) , sodium hydroxide (5g) , and water(50ml) and reflux over a burner for for 30mins.Anthracene willhave separated as a white precipitate. Remove the burner rinse down anthracene from the condenser with water filter the still hot mixture by suction. (It may be possible to decant from the residue of zinc mass).Wash the gray solid on the filter funnel with water and transfer it into a beaker .Rinse the funnel and paper with water ,add conc HCL (10ML) , heat on the steam bath with stirring for 20min to remove zinc and zinc oxide.Filter themixture ,wash with water, release the suction and rinse down with methanol(enough to cover the product).Apply suction again and wash with enough methanol once. Recrystallize the product in benzene (in fume hood) and filter the hot solution if necessary. Cool and collect the product.
Determine the yeild and m.p (about 220oC). Record IR, UV, and NMR (CDCl3) spectra.
Questions
1. Give two different reactions of anthracene that demonstrate that this compound has double bond character.
2. What anthraquinone(s) would be expected from a sequence starting with 3-nitrophthalic anhydride and (a) benzene, (b) toluene.
References
1.Organic experiments by Fieser and Williamson
2.Organic textbooks
Compiled by Mambanda Isaac (Chief Technician) & Gonzo Muriel (Snr Laboratory Technician)