Column and Thin Layer Chromatography Essay Example for Free

Column and Thin Layer Chromatography Essay sn atomic number 18Spinach extract was divide into dissevers containing mixs of similar diametricality by mainstay chromatography. Based on solid-liquid physique separate, this separation technique exploited the polar polarity of the complicateds in the spinach plant extract. Three elements with different color were obtained. The extract and its splits were analyzed using prune class chromatography (TLC). The TLC results showed that in that respect was matchless commingle (Rf=0. 979) in the low fraction there were three compounds (Rf1=0.839, Rf2= 0.691, Rf3=0.149) in the act fraction there was one compound (Rf=0.017) in the ordinal fraction. The separations of compounds which went to the offset printing and third fraction were relatively satisfying, while the number fraction had some(prenominal) kinds of compounds.IntroductionChromatography is the separation of compounds or ions by distribution amid two microsco pe stagesa runny chassis and a stationary signifier. The technique is based on the differential absorptivities of the constituents between these two phases, due to different properties of the compounds to be separated and the nature of the two phases involved. If one constituent adheres more to the stationary phase than the liquid one, separation will be achieved.There are several different types of chromatography, such as thin layer chromatography (TLC), gas chromatography (GC) and column chromatography. All the chromatographic methods are based on partitioning of molecules between a stationary phase and a mobile phase. In order to measure the amount of partitioning between the two phases, each compound has an unique Partition Coefficient (Kp), which is defined as the ratio of concentrations of the compound between the stationary phase and the mobile phase Kp=xsp/xmp. Therefore, if Kp is great than 1, the substance adheres more to the stationary phase if Kp is smaller than 1, the substance adheres more to the mobile phase if the Kp is equal to 1, the probabilities of the substance to adhere to the two phases are roughly the same. The Kp value is depended on a multitude of factors polarity, solubility in the solvent, hydrogen bonding, volatility in the case of gas chromatography.After the injection, the sample molecules will either stay in the mobile phase or adhere to the stationary phase. When the mobile phase is pushed through the stationary phase by the eluent, the mobile phase, the molecules will move through the column at a rate that depends on their different Kp value. For example, if one constituent is more polar than other and adheres more to the stationary phase when the stationary phase is more polar than the mobile phase, this constituent will lag behind while the constituent that is less polar and adheres less to the stationary phase will move ahead. In this way, the sample can be separated into fractions containing compounds of similar pola rity.The differences among the various types of chromatography are mostly depended on the nature of the two phases involved. Column and thin layer chromatography use solid stationary phase and liquid mobile phase while GC uses a gas as mobile phase and a liquid as stationary phase. Compared with TLC and column chromatography, GC is a more sophisticated method.DiscussionThe intent of this experiment was to successfully separate the constituents of spinach extract using column chromatography. The results of the separation were analyzed by TLC to reveal in which fraction the compounds of the mixture were. Since both of the samples of the first fraction and the third had only one developed location on the TLC plate, the separation of the compounds in the first and third fractions were relatively satisfying. However, there were 3 different compounds reveled for the second fraction. Since the color of the first fraction was yellow and the color of the second and third fractions were gree n, the major compound in the first fraction should be Carotenes, the yellow-orange pigment while the major compounds in the second fraction and the third fraction should be chlorophyl. During the process of column chromatography, the first band gained in the column was very perish and the color was yellow. The second band gained in the column was green and the thickness of the second band was greater than the first. Also, the green color of the second band varied, and colorless area was observed in the second band. Compared with the first band, the second band showed an unsatisfying separation result.Since there was colorless area and various green colourize in the second band, the second fraction must contained several constituents, which was examined by the TLC resultsthere were 3 developed spot of the second fraction sample on the TLC plate. As for the third fraction, there was no clear band in the column while adding the last eluent. The concentration of the compound in thir d fraction was relatively small, resulting in a very small Rf value, 0.017. Compared with the floating policy of the second fraction sample, which had a much clearer spot with similar Rf value, 0.839, the compound in the third fraction must existed in a large amount in the second fraction. In addition, these 2 spots had the same light green color, which can besides examine this conclusion. The lower concentration can also due to the unsatisfying spotting. The diameter of the undeveloped spot in the third fraction was larger than the first and second one, because too much solvent was used.Also, multiple times of spotting was needed. Observing the spots color, the clearest spot in the second fraction had a more intense green color, which showed that the compound in this fraction should be Chlorophyll a. While the major compound in the third fraction should be Chlorophyll a, with a lighter color and lower Rf value. The ideal solvent system should result in Rf values ranging from 0.24 to 0.54. However, in this experiment, none of the Rf value was in this range. Therefore, the polarity gradient of the eluents should be narrowed, which can result in more spots in the ideal range. Since there were 3 spots in the second fraction sample, over 4 eluents are needed to give a come apart separation, assuming the spot with lowest Rf value in the second fraction had the same compound as the spot in the third fraction sample.