Sunday, May 31, 2020

The main objective of this laboratory report is to identify analgesics in a compound containing a mixture of known analgesics by separating the mixtures using Thin Layer Chromatography (TLC) and determining the individual R_f values of the separated mixtures. The individual R_f values of the mixtures are then compared with the Rf values of known analgesics. Besides, the industrial purpose of the laboratory report is to understand the application of the procedure in the different contemporary fields of professions such as colour separation in textile industries and forensic criminal investigations.

Introduction

Chromatography is routinely used in organic chemistry laboratories as a technique of separation of molecules and subsequent analysis. For instance, TLC is usually used to find the degree of purity of molecules or compounds and to analyze the components of a compound or mixture through exposing them to both stationary (Mohrig et al. 2014). In principle, the identity of molecules is made possible due to the different solubility capacity of molecules and their strength of attraction to the stationary phase.

In this laboratory experiment, TLC was applied in the determination of unknown components in a mixture by comparing their R_fvalues with known analgesic compounds. The analgesics used in this experiment were; Acetaminophen, Asprin, Caffeine, Phenacetin, and salicylamide which are compounds used for pain-relieving processes. This TLC technique is applied in several professional fields of study. For instance, forensic detectives use TLC in matching known illegal drugs with blood samples of criminal suspects.

Procedure

Preparation of TLC Plastic Plate

Using a clean pair of forceps, two separate TLC plates (Silica gel) were placed on a clean surface and marked using a pencil as TLC plate 1 and TLC plate 2. Using a clear pencil marker, baseline and finishing line were drawn 1cm from each edge of both bottom and top of the TLC plates (Silica gel) respectively. On the first plate, spots were drawn and labeled as Ace, Asp, Caf, and Unkn for the first plate representing Acetaminophen, Asprin, Caffeine and Unknown mixture. On the second plate, spots were drawn and labeled as Phen, Sal and Unkn to represent Phenacetin, Salycylamide and unknown mixture. Simultaneously, with intermittent cleaning of TLC capillary tube, 0.25ml of each molecule was measured and transferred to its respective spots and the resulting TLC plates transferred to the developing chamber using clean forceps.

Developing Chamber

Before the transfer of the TLC Silica gel plates using clean forceps into the developing chambers, the ethyl acetate solvent was poured into the chamber at the level of 0.8cm just below the 1cm mark measured using a ruler. The reason for this was to reduce the chances of diffusion of the molecule with solvent, creating an unmeasurable movement due to scattering of diffused molecules. The developing chamber was enclosed with a clear watch glass to allow the environment within the chamber to be saturated. The experiment was then left to run and after 25 minutes, the plates were removed using forceps for visualization. This time was used to ensure that the solvent had traveled the furthest distance to TLC plates

Visualization

The plates could be seen immediately using UV light and each spot from both plates circled using a pencil to avoid fading of the colours due to UV light fluorescence of the spots.

Analysis

The distance in millimeters (mm) from the baseline to the center of each circle for each molecule movement was measured using a ruler and recorded. Additionally, the distance of the movement of the solvent was measured known as the solvent front as shown in table 1.

Molecule	Distance Travelled (mm)
Acetaminophen	30
Asprin	45
Caffeine	17
Phenacetin	40
Salicylamide	46
Unknown spot 1(Both plates)	17
Unknown spot 2 (Both plates)	30
Unknown spot 3 (Both plates)	45
Solvent	65

Table 1. Distance covered by each molecule on the First and Second TLC plates from the baseline to the middle of the spots circled.

Results and Discussion

The results recorded were used to calculate individual R_f values. In TLC plate analysis, R_f value could be determined by identifying the ratio spot to solvent travel (Balshan, 2017; Singh et al. 2016). The formula is as represented below.

Molecule/Spot	R_f Values
Acetaminophen	0.46154
Asprin	0.69230
Caffeine	0.26154
Phenacetin	0.61538
Salicylamide	0.70769
Unknown spot 1(Both plates)	0.26154
Unknown spot 2 (Both plates)	0.46154
Unknown spot 3 (Both plates)	0.69230
Solvent	N/A

Table 2: R_fvalues of different molecules/spots as recorded from TLC plates in the experiment.

This technique that uses the concepts of stationary and mobile phase’s components based on polarity differences to separate and identify mixture in compounds is known as chromatography. A good example used in this experiment is TLC. It is essential in the separation and identification of unknown molecules in a mixture. More polar compounds have strong intermolecular forces resulting in strong affinity to the polar phase of TLC thus will move a shorter distance up the plate.

From the above tabulations of the Rf calculations, the unknown mixture spots had Rf values of 0.26154, 0.46154, and 0.69230 correspondings to the R_f values of Caffeine molecule (0.26154), Acetaminophen molecule (0.46154), and Asprin molecule (0.69230). This indicates that the constituents of the mixture were Caffeine, Asprin, and Acetaminophen since they moved an equal distance up the TLC Silica gel plate under the influence of the mobile solvent phase. Besides, they had same Rf values after corresponding calculations. The unknown mixture was also able to separate into 3 spots due to different levels of intermolecular forces of attraction holding the molecules together and the different level of solubility in the mobile phase (DCM). The stationary phase containing silica gel has a hydroxyl group in its structure making a stronger polar compound (Mohrig et al, 2014). Conversely, ethyl acetate is the mobile phase since it is a less polar molecule compared to polar silica gel resulting in movement up the TLC plate noted by the higher solvent front. In this case, its affinity for polar compound (Silica gel) is lower. However, using a more non-polar solvent such as 100% hexane reducing the affinity of other less polar compounds in the mixture to it and hence spots/molecules staying closer to the origin/baseline. This separation based on the affinity or polarity can be understood by the structure of the molecules shown below.

List of Figures

Figure 1: Acetaminophen Figure 2: Asprin

Figure 3: Caffeine Figure 4: Phenacetin

Figure 5: Salicylamide

The structure of the TLC plate is a polar SiO₂making any molecule that is more polar to move a less distance up the plate due to attraction/affinity due to the same polarity strengths (Filipic et al, 2016). From our experiment, salicylamide moved the furthest up the TLC plate because of less affinity to the stationary phase. Salicylamide structure, Figure 5, indicates that the molecule is made up of a carboxyl functional group apart from the hydroxyl group creating weak intermolecular forces. As a result, it becomes less polar and traveling further the TLC plate hence a larger Rf value of 0.70769.

Figure 2 indicates the structure of Asprin. The structure has both a carboxyl group and ester functional groups which are similarly attractive to the stationary phase making it travel up the TLC plate further. However, esters are relatively polar making it travel slightly a lesser distance than Salicylamide, The result is an R_f value of 0.69230. Figures 1 and 4 are structures of Acetaminophen and phenacetin. Looking closely, the hydroxyl group in Acetaminophen is replaced by an ethoxide. Therefore, since Acetaminophen structure has a strong hydrogen bond with SiO2 than phenacetin, creating a high affinity to the TLC plate than phenacetin. This makes it to move a lesser distance than phenacetin and Asprin on the TLC Silica gel plate and hence an Rf value of 0.46154 compared to Rf value of 0.61538. However, Caffeine traveled the list distance of the molecules under study. Based on Figure 3, the structure of caffeine is made up of several amine functional groups with numerous oxygen and Nitrogen lone pairs reactively strongly to the SiO2 of the stationary phase (Maia, Batista, & Fernandes, 2017). This makes it travel the list and hence an Rf value of 0.26154.

Conclusion

In summary, the main objective of this laboratory experiment was to identify the analgesics present in the unknown mixture using TLC. Acetaminophen, Asprin, Caffeine and the unknown mixture were placed using the capillarity tube on the spots drawn using pencil on the first TLC plate and the same was done on the second plate with Phenacetin, Salycylamide and two unknown mixtures. The resulting preparations were then developed and visualized under Uv light. The resulting spotted were used to find the Rf value. The results indicated that the unknown mixture contained Caffeine, Acetaminophen, and Asprin with Rf values of 0.26154, 0.46154, and 0.696230 correspondingly. Besides, the Rf values obtained for each molecule were theoretically close to the standard values, signifying that the experiment was conducted successfully with minimal experimental errors. This technique is an important procedure as it is applied in the field of forensic criminal investigation and in separation of colours in textile industries.

References

Balshan, Y., 2017. Performing 1D Thin Layer Chromatography | Protocol. Retrived from https://www.jove.com/science-education/5499

Filipic, S., Antic, A., Vujovic, M., Nikolic, K. and Agbaba, D., 2016. A comparative study of chromatographic behavior and lipophilicity of selected imidazoline derivatives. Journal of Chromatographic Science, 54(7),1137-1145. Doi: 10.1093/chromsci/bmw081

Maia, J. C., Batista, F. S., & Fernandes, D. G. (2017). Residual TLC silica gel: Recuperation process, characterization and application. Revista Gestão Inovação e Tecnologias, 7(2), 3833-3843. doi:10.7198/geintec.v7.i2.770

Mohrig, J., Alberg, D., Hofmeister, G., Schatz, P. and Hammond, C., 2014. Laboratory Techniques In Organic Chemistry. 4th ed. New York: W.H. Freeman, pp.251-264.

Singh, J., Shukla, S. K., & Sharma, M. (2016). Chromatographic separation studies of carbapenem antibiotics on silica gel G layers with different solvent systems. JPC - Journal of Planar Chromatography - Modern TLC, 29(3), 229-231. doi:10.1556/1006.2016.29.3.11

Distillation of Methanol from Water

Simple and Fractional Distillation

Institution Affiliation

Date

Table of Contents

Preparation for Simple distillation

Observations from simple distillation experiment

Fractional distillation

Apparatus required

Preparation of Fractional distillation

Observations from fractional distillation experiment

Discussion

Graphical illustrations

Distillation Process

Differences between simple distillation and fractional distillation

Explanation of Observations from the Graphs

Boiling point of methanol

Waste Disposal

Safety Protection and Precaution

Distillation of Ethanol

Real-world applications of distillation

The precision of the Thermometer

Errors and Omissions

Conclusion

References

PURPOSE

The objective is to help understand the concept of distillation by separating a mixture of methanol and water using fractional and simple distillation methods. Water and Methanol are miscible. Simple and fractional distillation methods will be used to demonstrate how to separate miscible substances. Although the distillate that is collected from this experiment is not expected to be 100% pure, the effectiveness and efficiency of the process will be discussed with respect to each distillation process to help compare and determine between the two which one is more effective and why. These experiments will help learners understand the complex industrial distillation process done on a large scale.

INTRODUCTION

Miscible liquids are separated through the process of distillation. A solution is heated until it starts to vaporize, then cooled down to back to liquid. Liquids which have different boiling point evaporates at different stages (Nicholas 2017). Simple distillation is used to separate liquids which has a wide gap in boiling point, while fractional distillation is suitable for separating substances which has a closer gap in boiling point. The liquid with a lower boiling point will change to vapor first, passed through the condenser where it turns back to liquid and is collected container as distillate (Nicholas 2017).

EXPERIMENT PROCEDURE

Simple distillation

Apparatus required

Round bottom flask, Vacuum Adapter, Water condenser, Thermometer adapter, Thermometer, ‘Y’Shaped multipurpose adapter, Jack, Woodblock, Clamps, Graduated cylinder, Boiling chip, Water tube and Heating mantle.

Preparation for Simple distillation

I. Set up the apparatus above as shown in fig 1.1

Figure 1.1 a complete set-up of apparatus for simple distillation

II. put 20ml methanol and 20ml water in round bottom flask

III. Add a boiling chip to the mixture

IV. When the set up is complete turn on the water and the heat source.

Observations from simple distillation experiment

I. The table below contains the reading of the Volume of distillate after every 2ml against their respective temperature.

Vol ml	0	2	4	6	8	10	12	14	16	18	20	22	24	25
Temp (°C)	63.4	63.7	65	66.3	67.8	68	70	70.8	72.8	75.5	78.2	83.3	87.8	88.6

Fractional distillation

Apparatus required

Round bottom flask, Vacuum Adapter, Water condenser, Thermometer adapter, Thermometer, ‘Y’ Shaped multipurpose adapter, Clamps, Graduated cylinder, Boiling chip, Water tube, Heating mantle, Fractional column.

Preparation of Fractional distillation

I. Gather all the apparatus required and set-up a fractional distillation as shown in fig 1.2 below.

Figure 2.1 complete set up of apparatus for fractional distillation.

II. Mix water 20ml and Methanol 20ml into a round bottom flask.

III. Add a boiling chip into the mixture.

IV. Switch on the heating mantle to Start heating the mixture.

V. Connect the water source and ensure that cold water is continuously running through the condenser.

VI. Continue heating the solution even after it start boiling.

VII. Once the first drop is collected in the graduated cylinder, read the temperature on the thermometer.

VIII. Take the reading of the thermometer after every increment of the distillate in 2ml.

IX. After all the readings are taken, disassemble the apparatus once they cool down.

Observations from fractional distillation experiment

a) The table below contains the reading of the Volume of distillate after every 2ml against their respective temperature.

Vol ml	0	2	4	6	8	10	12	14	16	18	20	22	24	25
Temp (°C)	58	60	60.7	61.5	62.3	63.1	63.9	65.0	66.7	68.0	73.8	86.2	90.8	91.3

DISCUSSION

Graphical illustrations

Distillation Process

Add 20ml water and 20ml methanol into a round bottom flask, then set-up the apparatus as shown in fig 1.1. Start heating the mixture by adjusting the power knob that supplies power to the heating mantle. Continue heating the mixture until it boils. Ensure a continuous flow of cold water through the condenser. Take note of the temperature when the first drop falls to the graduated cylider. Continue recording the temperature after the distillate increases at an interval of 2ml in the graduated cylinder.

Differences between simple distillation and fractional distillation

Fractional distillation is a more complex process that uses fractional column while simple distillation is a simpler process and the fractional column is not used. The reason why the fractional column is used in distillation is to separate a mixture of fluids which has a relatively closer boiling point (Mia et al. 2017).

Explanation of Observations from the Graphs

The first drop in the fractional distillation came out at 58⁰C while that of simple distillation came out at 63.4⁰C. Also, both lines increase at almost the same gradient up to at around 20ml after which the gradient for fractional distillation becomes steeper as compared to that of simple distillation. This implies that most of the all methanol had vaporized and the remaining solution has to be heated harder for it to vaporize.

Boiling point of methanol

From the graphical illustrations, both graphs have two different boiling points. The boiling point of methanol is 63.4 ⁰C in simple distillation and 58⁰C in fractional distillation. 64.7⁰C is the accepted boiling point for methanol (Jerry et al. 2014).

Contrary to my expectation, the accepted boiling point at 64.7⁰C is closer to the boiling point of methanol at simple distillation. This should not have been the case if the error discussed hereunder wasn’t there. The boiling point of fractional distillation should have been closer to the accepted boiling point because it does not have the fractional column which contains steel wool or glass beads meant to increase the surface area. This makes the vapor to condense and return back to the boiling mixture many times before vaporizing and reaching the topmost of the column, due to the aspect of condensing and re-vaporizing the boiling point reading becomes more accurate than that of simple distillation method.

Waste Disposal

At the end of our distillate, we need to dispose of the distillate into the non-halogen organic solvent bin. After disposing we should ensure that the bottle where we put the waste is closed always.

Safety Protection and Precaution

Methanol is poisonous and therefore, after the experiment one should always wear lab coats, Protective eyewear, gloves, and closed-toe shoes at all times when experimenting. It is also highly flammable and therefore one should take caution not to sprinkle it so as not to cause fire. We should not eat anything in the laboratory as it might be contaminated and dispose properly laboratory waste (Peter 2016).

1n 1926, the federal government intentionally increased the amount of methanol used in industrial alcohol to poison alcohol in an attempt to curb alcohol consumption.

Distillation of Ethanal

Ethanal can be distilled from a mixture of water using both methods because they have different boiling points (Jerry et al. 2014).

Distillation Real-world applications

a) Separation of oil products

Crude oil comprises different petroleum products, these petroleum products in crude oil have different boiling points and therefore they are separated using fractional distillation (Sukasem, Hareemao, & Sudawong. 2017).

b) Distillation of salty water

A large distillation plant can be used to purify salty water and supply many people with clean drinking water and also be used to cook.

The precision of the Thermometer

The precision of the thermometer is the fraction to which a thermometer can be read, it can also be defined as the nearest whole number or decimal to which a thermometer can be read. In our experiment the precision of the thermometer is 0.1⁰C.

Errors and Omissions in the experiment

There was a critical error made during the transition from simple distillation to fractional distillation. The instructor did not completely change the entire set of apparatus he used for simple distillation. Instead, he only added a fractional column and proceed with the experiment on fractional distillation.

CONCLUSION

Distillation is the most commonly used method of separating a mixture of two or more liquids. The process applies a simple rule of separating mixtures based on their boiling points. It has traditionally been used to perform complex industrial separation of substances in the food industry, chemical, and petroleum industry. It should be understood that distillation is an irreversible process, and is commonly used alongside other procedures to ensure that the distillate is 100% pure.

REFERENCES

Pater, L. (2016). The mimic of fractional distillation technology distillation process modeling. arXiv preprint arXiv:1605.00097. Jerry . M., David G. A., Gretchen E.

Mia, M., Islam, A., Rubel, R. I., & Islam, M. R. (2017). Fractional distillation & of tire- derived pyrolysis oil.

Sukasem, N., Hareemao, T., & Sudawong, C. (2017). The mimic of fractional distillation technology for the development of homegrown pot distillery for ethanol distillation. Energy Procedia, 138, 985-990.

Jerry R., Paul F. S., & Christina N. H. (2014) Laboratory Techniques in Organic Chemistry. Library of Congress Control Number: 2013955847

Dr. Nicholas L., (2017). Fractional Distillation. URL: https://www.jove.com/science- education/5700