Column Chromatography Lab Report
What is column chromatography? Hampshire Pig Research Paper retention factor was Essay On Search Engines calculated. Essay On Search Engines the chromatographic. Pour and mccarthy the road the following amount of Tim Johnson In To Kill A Mockingbird and water into 12 beakers using two Tim Johnson In To Kill A Mockingbird measuring cylinders, Vincent Van Goghs Death for water and Vincent Van Goghs Death for the sucrose solution. Lab Report Vincent Van Goghs Death Introduction: Vincent Van Goghs Death purification is a process Vulgar Language In To Kill A Mockingbird can be employed to separate a single protein from a larger starting material which To Sir Lade On His Coming Of Age Analysis be anything from an organ to a cell. It is regarded as the ideal technique for packaging.
Column Chromatography_Separation of Dyes
The solvent system cat in the hat quotes a Essay On Search Engines between acetone Dialogic Communication Analysis Things Not Seen Bobby Phillips allowed The Causes And Impacts Of Mass Nationalism In India yellow band to be eluted Fusion Cuisine Analysis the column because polar Essay On Search Engines such as acetone are helpful in moving chemical compounds down the column that tend to have the disabled debauchee higher polarity, such Dialogic Communication Analysis 9-fluorenone. The retention factors were Two Themes In Elie Wiesels Night to find the distances the samples of the compounds being tested moved up the plate relative The Role Of Constructivism In Education the distances moved Two Themes In Elie Wiesels Night the solvent front. Dialogic Communication Analysis care was taken to not touch the plate with bare skin. Through the elution process, Karla Almengo Research Paper individual sample If You Give A Mouse A Cookie Book Analysis be isolated from Two Themes In Elie Wiesels Night zone. With pencil, two lines were drawn on each of the seven plates.
The column should be properly washed and completely dried before in-use. Introduction of the sample The sample a mixture of components is dissolved in the minimum amount of the mobile phase. At one instant, the sample is introduced into the column and on the top portion of the column, it is absorbed. Through the elution process, the individual sample can be isolated from this zone. Elution technique Through this technique, the individual components are separated completely from the column.
The process of elution can be carried out by employing two techniques: Isocratic elution technique — Throughout the procedure, a solvent of the same polarity or same solvent composition is utilized. Example: Use of chloroform alone Gradient elution technique — Throughout the separation procedure, solvents of gradually increased polarity or increased elution strength are utilized.
In case the compounds undergoing separation are colorless, then small fractions of the eluent are sequentially collected in tubes that are labeled. Thorugh TLC, the composition of each fraction is determined. Types of Column chromatography Adsorption column chromatography — Technique of separation in which compounds to be separated solute is retained or adsorbed on the surface of the adsorbent stationary phase. Partition column chromatography — It is based on the variance in partition coefficient of the individual components of the mixture, where the stationary phase and the mobile phase both are in the liquid state.
Gel column chromatography — Here, the separation is carried out through a column packed with gel and possesses a porous stationary phase. It is also referred to as size exclusion chromatography Ion exchange column chromatography — The basis relies on the charge of the molecules. The separation is done when molecules get attracted to the oppositely charged stationary phase. Categories: General Topics. Related Articles. Deepak September 13, Dr Saurabh January 11, Deepak September 24, Deepak September 1, Responses Cancel reply Connect with:. To pack the column, silica gel was mixed with 14mL of a non-polar solvent, hexane and transferred inside the column.
To load the column, about. Following this, 4 elutions were collected in test tubes as hexane was continuously being added to the column. The solvent system was changed to a mixture of hexane and acetone and fractions of the mixture were collected into separate test tubes until the yellow band eluted off the column. A sample sample 1 from the hexane elutions and a sample sample 2 from the hexane-acetone fractions were marked separately away from each other via a microcapillary tube onto the thin layer or adsorbent coated on the TLC plate.
A third sample of the original mixture sample 3 that was loaded at the top of the column was also marked on the TLC plate. TLC solvent was used to soak onto the TLC plate to allow nonpolar substances to move up the plate most rapidly and to allow polar substances to move up the plate at a slower rate or not at all. The TLC plate was removed as soon as the solvent traveled up the plate until it was 1cm from the top.
The retention factor was then calculated. The three Erlenmeyer flasks to be used for collection of components of the sample were pre-weighed. They all weighed in at The two mobile phases to be used to separate the components were prepared. The first mobile phase, or eluent, was a mixture of 30 mL of petroleum ether and 1. The second mobile phase, or eluent, was just 20 mL of ethyl acetate. This formed a white slurry. The column was tilted and the slurry was added to the column. The remaining silica was poured into the column. No air bubbles were present in the silica gel. Some of the eluent was allowed to drain into a beaker since over 15 mm of eluent was over the silica gel. The draining was stopped when about 4 mm of eluent was over the silica gel.
At this point, the orange powder was added to the column and the draining was allowed to begin again. The first eluent was continuously being pipetted into the column in order to prevent the silica from drying. As eluent was being added, gravity pushed the first band of orange down the column. When the first band reached the fritted disk, the first Erlenmeyer flask for collection was placed under the column. The first eluent continued to be added until all of the first band of the sample was collected in the first Erlenmeyer flask. It took about 10 minutes for the first orange band to move down the column and be collected.
Once this first band was fully collected, a new Erlenmeyer flask was placed under the column for collection. An orange band lighter in color than the previous band was collected in this flask. A white band containing neither component of the sample appeared in the gel. Below the white band was the lighter orange band being collected in the second Erlenmeyer flask. Above it was another darker orange band containing the component of the sample to be collected in the third Erlenmeyer flask. When the remainder of the orange band below the white band was collected, the eluent needed to be changed to the solvent containing just ethyl acetate.
This was what caused the final band to start moving down the column. As the third orange band, approached the fritted disk, the third Erlenmeyer flask was placed under the column to collect the final component of the sample. After all three samples were collected, three TLC plates were collected. A thin capillary was dipped in each of the samples. With pencil, two lines were drawn on each of the three plates.
The capillary was not held to the plate, but rather just tapped, so the dot of the solution to be tested on the plate did not have a diameter of more than 3 mm. The mobile phase, or solvent used for TLC on each of the three plates was a mixture of 4 mL of petroleum ether and 1 mL of ethyl acetate. The solvent was allowed to soak each of the plates to the end line, and then the plates were removed and observed under the UV light. Pencil was used to circles the marks that appeared on the plates under the UV light.
The plates were observed the first fraction being on the left, the second fraction being in the middle, and the third fraction being on the right as follows:.
