ochem

Liquid-Liquid and Solid-Liquid Extraction

Jennifer Driskell

Department of Chemistry, The Pennsylvania State University, University Park, PA 16802

CHEM 213B.102 | TA: Omair Khan

Original Submission: February 19, 2011

Resubmission: March 24, 2011

Abstract

Liquid-liquid extraction techniques were used to extract an acid, base and neutral from an

unknown mixture. The technique was done on a microscale level using test tubes and pipets to

separate and extract the layers of interest. Solid-liquid and liquid-liquid extraction were used to

extract caffeine from tea leaves. The caffeine was extracted on a macroscale level using beakers

and separatory funnels. The crude caffeine was sublimed to obtain a pure caffeine sample. For

identification of the compounds extracted (unknowns and caffeine), melting point values were

obtained using a Mel-Temp apparatus. Two of the compounds isolated from the unknown

mixture were also analyzed using a 60 MHz 1H NMR.

Introduction

Whether making coffee, isolating a natural product or separating compounds in solution, the

underlying technique being applied is extraction. Extraction is the process of pulling one

compound away from or out of another compound.1 It is widely used both in everyday life and

the laboratory. Two common examples of extraction are liquid-liquid extraction and solid-liquid

extraction. A Pittsburg laboratory demonstrated liquid-liquid extraction when the scientists were

able to isolate urea from a fluorous-amine scavenger.2 In their reaction, they prepared urea by

mixing an amine (limiting reagent) with an excess of isocyanate. Once the reaction was

Experiment 5 Extraction Driskell / 2

complete they added an excess of fluorous amine quenching reagent to change the un-reacted

isocyanate into a fluorous urea. The liquid-liquid extraction was then used to separate the

organic urea product from the fluorous urea and amine.2 An example of solid-liquid extraction is

brewing coffee. Solid-liquid extraction has two phases a solid phase, the coffee beans, and a

liquid phase, the water. The water is able to dissolve small, polar molecules, like caffeine, away

from the coffee beans due to similar molecular interactions.1 Liquid-liquid extraction works

using solubility differences of the compounds being separated.2 Manipulation of a compound

can cause it to move into a desired layer allowing it to be extracted. Determining which layer the

compound resides or how much of it will be there can be calculated using, the distribution

coefficient, K.1 The distribution coefficient is the distribution of the solute, in the solvent, at

equilibrium. A small distribution coefficient means that not all of the solute resides in the

organic layer. Multiple extractions might be necessary to completely extract the entire

compound.1 The distribution coefficient is used in choosing acid-base reactions to manipulate

compounds into the desired layer.

Acid-base reactions use functional groups to manipulate the solubility of the molecule.

By deprotonating or protonating a functional group we can affect the pH, which in turn will

change the distribution coefficient, pulling molecules into the water layer. An acid is a

compound that will donate a hydrogen ion to a compound capable of accepting a hydrogen ion.

A base is a compound that has the ability to accept a hydrogen ion from a hydrogen donor.3

Compounds with similar functional groups, such as carboxylic acids, will not be separated by

this kind of manipulation because they will both move into the same layer. Neutral compounds

are unique in that they will not be affected by the acid-base reactions because they do not lose or

gain protons.1 Scheme 1 provides a flow chart to give two examples of acid-base reactions.


Hydrochloric acid, a strong acid, donates its hydrogen to the base, 3’-aminoacetophenone. The

conjugate acid of the base now has a positive charge allowing it to be pulled into the water layer.

After extraction of the water layer, containing the conjugative acid, the addition of a strong base,

K2CO3, will deprotonate the conjugative acid returning it to the original base. By deprotonating

the hydroxyl group on the benzoic acid with a weak base, NaHCO3, a negative charge is created

on the oxygen atom pulling it into the water layer. After extraction of the water layer, with the

conjugative base, the addition of a strong acid, HCl, will protonate the oxygen atom leaving the

original uncharged acid.3 The addition of acids and bases can help to extract one compound from

another, however, it does not guarantee purity of the compound.

A simple and inexpensive purification technique that can be used to remove impurities in a

compound is sublimation.1 Sublimation is the direct phase change from a solid to a gas, without

passing through a liquid phase, at a particular pressure. This experiment sublimed caffeine to

obtain pure crystals from the crude caffeine extracted from tea leaves using solid-liquid and

liquid-liquid extraction techniques. At atmospheric pressure, the impurities in the crude product

had lower sublimation points than caffeine meaning they would have to pass through the liquid

phase before becoming gaseous.1 In order for the impurities to sublime they would need a lower

pressure. Caffeine on the other hand had a higher sublimation point and was able to become

gaseous instead of liquid at atmospheric pressure. When the gas vapors touched the walls of the

test tube they were cooled. The cooler temperature caused the phase change from the gas back

into a solid this time without the impurities.1

In this experiment, three compounds were isolated from a mixture using acid-base

reactions and liquid-liquid extraction techniques. The compounds were then identified using a

60 MHz 1H Nuclear Magnetic Resonance Analysis (NMR) and melting points were obtained


using a Mel-temp apparatus. The second part to this experiment was the extraction of caffeine

from tea leaves, this involved solid-liquid extraction followed by liquid-liquid extraction as the

first step of purification. The caffeine was finally purified using sublimation and the crystals

were identified using melting points obtained from a Mel-temp apparatus.

Results and Discussion

Isolation of Compounds from Unknown 34

The isolation of Unknown 34 can be easily followed using Scheme 1 to keep track of

which acids and bases are being added and which layer the compound resides after the reaction.

The first step was to isolate the three unknowns in the mixture this was done according to the

procedure found in the experimental section. After isolation two 60 MHz 1H NMR analyses

were preformed, one on the isolated amine and one on the isolated neutral. NMR number one

was used for both analyses using d-chloroform as the solvent.


Scheme 1 Flow Chart for Isolation of Unknown 34

This is a visual diagram that will assist following the acid and base reactions that occurred when isolating compounds from the Unknown 34 mixture. The mixture came with a base, an acid and a neutral compound that had to first be isolated

before they could be identified.

The NMR analysis of the neutral compound, Figure 1, assisted in determining that the

neutral compound was fluorenone. This spectral analysis contains a lot of contamination which

can be contributed to the other compounds not being completely extracted. Two of the

contamination peaks can be attributed to the base and one attributed to the acid. There was one

contamination peak that could not be accounted for as coming from the acid or base in particular.

The theoretical shift and splitting was unknown for this particular contamination. The NMR

disproved the compound 1, 4-Dimethoxybenzene because it did not display a methyl hydrogen


peak that would be apparent around 3.5 ppm. All of the peaks of importance are jumbled closely

together, so specific splitting patterns were not determined. The peaks can all be found in the 7-

8 ppm range which was the appropriate range for aromatic hydrogens. This information can be

viewed in Table 1. The NMR obtained for the amine compound, Figure 2, assisted in

determining that the compound isolated was 3’-Aminoacetophenone. All six peaks of interest

are accounted for on the spectrum and displayed in Table 2. The spectrum disproves 4’-

Aminoacetophenone because there was no “Para” splitting of the aromatic hydrogens. The only

problem with the NMR of the base was incorrect integration ratios. A possible explanation for

the incorrect ratios could be when the spectrum was normalized to account for the hydrogens in

the amino group, Hf. It is possible that the machine did not record two hydrogens instead it

viewed them as one. When that peak was normalized to two it added an additional hydrogen to

the other integrated peaks. After subtracting one hydrogen from the other integrated peaks the

numbers produced were expected values for 3’-Aminoacetophenone. The actual shift for

hydrogen A was higher than the theoretical shift. This is possibly due to the electronegative

oxygen atom interacting with the methyl hydrogens. Hydrogen E did not have the theoretical

splitting of a triplet. This hydrogen might have been able to interact with the amino group

resulting in more splitting than what would normally be expected. Peaks pertaining to hydrogen

B, C, and D are all within the circle and although not easy to determine they did in-fact have

correct splitting.


Figure 1 1H NMR Spectral of Neutral Compound

This was the compound remaining after the extraction of both the acid and base. The likelihood of contamination would be greatest since the other compounds were extracted, leaving this one in solution. The spectrum has been annotated

for fluorenone and contamination has been marked and identified in Table 1.

Table 1 1H NMR Spectral Data for the Neutral Compound

This is the tabulated description of results from the neutral NRM analysis, Figure 1. The peaks of interest are closely

packed around 7.5 ppm; the other peaks listed are noted as contamination. This contamination was contributed to the base

and acid not being completely extracted from the neutral.

Hydrogen Theoretical Shift

(ppm) Actual Shift (ppm) Theoretical Splitting Actual Splitting

A 6.5-8.5 ~ 7.5 doublet unknown

B 6.5-8.5 ~7.5 triplet unknown

C 6.5-8.5 ~7.5 triplet unknown

D 6.5-8.5 ~ 7.5 doublet unknown

Contamination Methyl from Base 0.7-1.3 2.5 singlet singlet

Contamination unknown Unknown 5.9 unknown singlet

Contamination Aromatic from Base 6.5-8.5 ~6.8 triplet multiplet

Contamination Aromatic from Acid 6.5-8.5 ~8.1 doublet multiplet


Figure 2 NMR Spectral of Amine Compound

This was the first compound extracted after isolation involving HCl. The chance of contamination for this molecule was low and no contamination peaks were observed on the spectrum. The printout has been annotated for 3’-

Aminoacetophenone with all hydrogens accounted for and displayed in Table 2.

Table 2 NMR Spectral Data for the Amine Compound

This is the tabulated description of results from the amine NRM analysis. The peaks of interest are labeled with letters

corresponding to Figure 2. All of the expected peaks are visible and most theoretical splitting was observed. The biggest

problem was the incorrect integration ratios.

Hydrogen Theoretical Shift (ppm) Actual Shift (ppm) Theoretical Splitting Actual Splitting

A 0.7-1.3 2.5 singlet singlet

B 6.5-8.5 ~6.860 singlet singlet

C 6.5-8.5 ~7.26-7.31 doublet doublet

D 6.5-8.5 ~6.90-7.22 triplet triplet

E 6.5-8.5 ~ 6.80 triplet multiplet

F 0.5-4.0 3.81 Singlet Singlet


Melting points were obtained for all three compounds isolated from Unknown 34 and

compared to literature values from Sigma-Aldrich. Melting points help determine a compound

based on the range and comparison to literature values. The range will be considerably lower

and broad due to impurities in the sample. The melting point data for Unknown 34 can be found

in Table 3. The melting point range of fluorenone was considerably off compared to the Sigma-

Aldrich values obtained. The NMR had contamination peaks which indicated impurities helping

to explain the low melting point. The amine had a melting point range close to the expected

value of 3’-Aminoacetophenone helping to verify the NMR results. The melting point was

slightly lower than normal, this could be contributed to the Mel-temp used not being completely

accurate. The acid was the only compound isolated that did not have an NMR analysis

preformed. The identity of this compound was completely derived from the melting point value

obtained from the Mel-temp apparatus. The melting point range was higher than typically

associated with benzoic acid. However, 3-toluic acid melts lower than benzoic acid therefore it

would be safe to conclude that the isolated acid compound from Unknown 34 was benzoic acid.

The higher melting point range could be associated with the sample not being completely dry

allowing the water to interact with the molecule affecting the melting point. Melting points

assist in identification of a compound but percent recoveries are calculated to determine how

much the compound was obtained after the extraction.

Table 3 Melting Point Data for Unknown 34

The melting point data was obtained using Mel-temps and are listed along with the expected temperature values gathered from Sigma-Aldrich. Fluorenone was very low most of which can be contributed to contamination. The base and

acid are relatively close to their literature values.

Compound

Melting

Temperature °C

Sigma-Aldrich Temperature

(°C) Values 4

Neutral- Fluorenone 47-51 80-83

Amine- 3'-Aminoacetophenone 94-96 97

Acid- Benzoic Acid 126-128 121


Percent recoveries were calculated for each of the compounds isolated from Unknown 34.

Equation 1 shows the calculation for percent recovery and Table 4 has the calculated values for

each percent recovery of the isolated unknowns. The percent recovery for the amine was above

the possible 100%. The first reason for this high percent recovery would be that the mixture of

Unknown 34 did not contain exactly 100 mg of each of the three compounds. This would result

in the wrong original starting weight of the compound causing the calculation to be incorrect. A

second possibility could be the measurements were inaccurate, resulting in faulty numbers. The

amine was the only compound greater than 100% recovery; the acid had a low percent recovery

at 13%. Reasons for this being so low are numerous; one reason could be that not all of the acid

was extracted from the neutral. A second possibility was that crystals were removed along with

the boiling stick and glass stirring rod that were needed to mix and boil the solution. Crystals

could also have been lost on the final step of removal from the Hirsch funnel. The acid and base

demonstrated the two extremes of percent recovery, however, the neutral, had a good percent

recovery at 81%. This number was not accurate due to the NMR analysis and melting point

values obtained demonstrating contamination. The contamination can be contributed to acid and

base compounds that were not extracted completely remaining in the neutral crystals when dried.

The extra crystals from the acid and base would increase the final weight of the neutral and not

give an accurate value recovered.

Table 4 Percent Recovery Data for Compounds Isolated from the Unknown

Percent recoveries for each of the compounds isolated from Unknown 34 are presented in the table. The theoretical weight was obtained by assuming one third of each compound was used in the mixture since 300 mg was initially weighed

out. The percent recoveries were calculated using Equation 1.

Compound

Theoretical

Weight (g)

Weight of Crystals

(g)

Percent Recovery

%

Neutral- Fluorenone 0.1 0.081 81

Amine- 3'-Aminoacetophenone 0.1 0.116 116

Acid- Benzoic Acid 0.1 0.013 13


Equation 1 Percent Recovery

Isolation of Caffeine

Isolation of caffeine from tea leaves worked using solid-liquid extraction because the tea

leaves are made up of many compounds including cellulose, proteins and lipids that cannot

dissolve into the liquid water phase. When the tea leaves were placed in the boiling water to

soak, the caffeine compound dissolved out of the leaves and entered the liquid which was

decanted and used for the experiment. Although solid-liquid extraction worked to extract

caffeine from the tea leaves, it also extracted any other compound that was able to dissolve in the

water. Liquid-liquid extraction was used as a complementary extraction to obtain only caffeine

from the solution. In the liquid-liquid extraction, dichloromethane (DCM) was used to separate

caffeine from the many other water soluble compounds that also dissolved from the tea leaves.

After isolation the caffeine was sublimed to obtain pure crystals from the crude caffeine

product. The caffeine extracted from tea contained many impurities even after liquid-liquid

extraction. These impurities had lower sublimation points than the pure caffeine. Since caffeine

has a higher sublimation point it was able to change from a solid to a gas before the impurities

had a chance to change phases. The sublimation works by allowing the caffeine to directly

change from a solid to a gas at atmospheric pressure without becoming liquid. Once the gas

molecules interacted with the cool walls of the test tube the lower temperature caused the gas to

solidify forming long, delicate, white crystals.

The pure caffeine crystals were weighed to determine the percent recovery and tested for

purity with a melting point, this data can be found in Table 5. The literature melting point range

for caffeine is 234-236°C, the melting point for the extracted caffeine was 226-233°C.4 The

slight melting point depression could be due in part to the Mel-temp used not being completely


accurate. It could also be that the crystals were still wet and the water interfered with the melting

temperature. Although the melting point was off, it fell into the accepted literature values when

the standard deviation, of +/- 5 was applied. To determine how much caffeine was extracted

from the tea the percent recovery was calculated, using Equation 1, and found to be very low.

This was probably due to the repeated transfer of the compound from the beaker to the flask to

the test tube, along the way product was lost. Another possibility was that during the

sublimation the caffeine could have been evaporated out of the test tube resulting in loss of

product. The data for caffeine can be found in Table 5.

Table 5 Caffeine Data

The theoretical yield for caffeine was determined by multiplying the amount of caffeine in one tea bag by three (55 mg x 3) since the experiment used three tea bags. The percent recovery was calculated using Equation 1. The melting point

values were obtained from the Mel-temp and the expected values were obtained from Sigma-Aldrich.

Compound

Theoretical

Weight (g)

Weight of

crystals (g)

Percent

Recovery %

Melting

Point °C

Sigma-Aldrich

Value °C 4

Caffeine 0.165 0.012 7.27 226-230 234-236

The Utility of the Solid-Liquid and Liquid-Liquid Isolation Methods

To assess the solid-liquid and liquid-liquid isolation methods, it would be best to look at the

percent recoveries that were calculated from the data. For the isolation of the three unknown

compounds from Unknown 34, it can be stated that the liquid-liquid extraction technique was not

great. Table 4 shows the results for percent recoveries and none of the values were good. The

best value, the neutral, was not accurate due to evidence of contamination. For the isolation of

caffeine from tea leaves, the effectiveness of the solid-liquid and liquid-liquid extraction was not

good; the percent recovery was only 7%. These isolation methods worked to isolate the

compounds but need to be improved upon to be able to gather a better recovery of the product.


Conclusion

This experiment used liquid-liquid extraction to isolate a base, acid and neutral from an

unknown. The identities were determined using NMR analyses and melting points. The

compounds were determined to be 3’-Aminoacetophenone, benzoic acid and fluorenone.

Percent recoveries were calculated for each isolated compound and were: 116% (base), 13%

(acid) and 81% (neutral). Solid-liquid and liquid-liquid extraction were both used to isolate

caffeine from tea leaves. The solid-liquid extraction isolated the caffeine from the leaves into the

water and the liquid-liquid extraction isolated the caffeine from the water. The caffeine

sublimed in the test tube forming long, white needles on the inside of the tube which were used

to determine the melting point. The sublimed crystals were determined to be caffeine due to the

melting point of 226-230°C. The percent recovery of caffeine was low at about 7%. The

experiment obtained results that were expected, but can be improved upon.

For future experiments, the isolation of the unknowns should be done on a larger scale so that

the separatory funnel can be used to separate the two layers, this will provide more accurate

extraction compared to pipets. For the caffeine procedure, trying a couple of different

experiments with different types of tea leaves and coffee beans to determine how much caffeine

is extracted from each type would be interesting. This experiment is practical in that most

people use these simple techniques everyday without actually knowing the science behind what

they are drinking.

Experimental Section

Procedure 1: Microscale separation of basic, acidic, and neutral substances

The mixture of Unknown 34 (300 mg) was weighed out and put into a test tube which was

dissolved in ether (2 mL).


Section A Separation of the Basic Component by Extraction with Acid: 5% aqueous HCl (1.0

mL) was added to the test tube containing the unknown and ether. Once layers separated, after

mixing, the bottom layer was extracted into a second test tube. Water (.25 mL) was mixed with

the remaining solution in the original test tube and the bottom layer was extracted into the second

test tube. Ether (6 drops) was added to the second test tube and allowed to evaporate in the

hood.

Section B Separation of the Acid Component by Extraction with Base: 10% aqueous sodium

bicarbonate (1.0 mL) was mixed into the original test tube now containing the acid and neutral.

Once the layers separated, the bottom layer was extracted into a third test tube. Sodium

bicarbonate (6 drops) was mixed to tube one, after separation the bottom layer was extracted into

the third test tube. The third tube was backwashed with ether (6 drops) and allowed to evaporate

in the hood.

Section C Isolation of the Neutral Component: Saturated NaCl (1 mL) was mixed to the

original test tube now containing just the neutral. Once the layers separated, the bottom aqueous

layer was extracted into a beaker. Anhydrous sodium sulfate (about 0.75 mL) was added to the

first test tube, which was corked to shake and vent for 10 minutes. The ether solution was then

decanted from the drying agent into a fourth test tube. After drying, the crystals were weighed

(0.081 g, 81% recovery). A melting point of 47-51°C was obtained and an NMR was conducted.

This data can be found in Figure 1 and Tables 1, 3 and 4.

Section D Isolation of the Amine Component: 50% potassium carbonate (4 drops) was mixed

to tube two and the pH was tested using litmus paper. The tube was corked, to shake and vent

for one minute, and then placed in an ice bath. The cold solution of crystals was added to a clean

100 mL beaker and allowed to dry. Once dry the crystals were weighed (0.116 g, 116%


recovery). A melting point of 94-96°C was obtained and an NMR was conducted. This data can

be found in Figure 2 and Tables 2, 3 and 4.

Section E Isolation of the Carboxylic Acid Component: Concentrated HCl (.128 mL, 0.0015)

was mixed to tube three and the pH was tested using litmus paper. The calculation for the

amount of HCl added can be seen in Equation 2. The tube was placed in an ice bath to form

crystals, after which the liquid was extracted and discarded. Distilled water (about 1.0 mL) was

added to the tube and placed in a sand bath to dissolve the crystals. After cooling in an ice bath,

a Hirsch Funnel was used to collect the crystals. Once dry the crystals weight was obtained

(0.013g, 13% recovery) and a melting point of 126-128°C was recorded. This data can be found

in Tables 3 and 4.

Procedure 2: Isolation of Caffeine from Tea

Three tea bags were allowed to soak in boiling water (75 mL) for 10 minutes and then the

liquid was decanted into a 125 mL Erlenmeyer flask. When the solution cooled it was poured

into a separatory funnel with dichloromethane (15 mL). The mixture was swirled and vented for

two minutes and then allowed to separate. The bottom layer was extracted into a clean flask.

The solution was dried and formed yellowish/green crystals. DCM was added to bring the

crystals into solution to be transferred to a test tube. The test tube was placed in a sand bath and

the DCM was boiled off. The contents of the tube were allowed to sublime for 30 minutes.

Long, white needles formed on the inside of the tube. After cooling and collecting the crystals

they were weighed (0.012 g, 7.27% recovery) and a melting point of 226-230°C was obtained.

This data can be found in Table 5.

Equation 2 Amount of HCl Needed for Acidification


References

1 Linclau, Bruno; Sing, Ashvani K.; Curran, Dennis P. Journal of Organic Chemistry. 1999, 64

(8), 2835-2842.

2 Minard, B.; Masters, K.M.; Halmi, T.O.; Williamson, K.L. Lab Guide for Chemistry 203 and

213B, 2010-2011, 111-138.

3 Brown, Theodore L.; LeMay, H. Eugene Jr.; Bursten, Bruce E.; Murphy, Catherine J.

Chemistry The Central Science, 11ed. 2009: Pearson Prentice Hall, 129-130.

4 Sigma-Aldrich. Sigma-Aldrich Co, 2011. February 1, 2011- February 14, 2011.

http://www.sigmaaldrich.com/united-states.html.

5 https://cms.psu.edu/section/default.asp?id=MRG%2D110110%2D102159%2DSAD270,

“Addendum to the Lab Guide, Chem 213 and Chem 213B, spring 2011, Experiment 5.”

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