Chromatography Assignment 1 Normal phase chromatography uses a polar stationary phase and a non polar or sometimes slightly polar mobile phase 2 phenol acetophenone naphthalene 3 The order of the peaks does not change when the mobile phase composition of B is set to 50 The chromatogram shifts to the left and the signal increases as the nonpolar stationary phase has a higher concentration The same happens for the shift 70 B k tr and decreased for all three of the molecules This is likely due to the changing affinity for the mobile phase as acetonitrile is less polar than water so a greater percentage would make the entire mobile phase less polar When acetonitrile is set to 5 the chromatogram is shifted to the right as the retention factor increases The elution order remains the same but the retention times and k values increased for each compound At 5 acetonitrile the mobile phase is very polar so it does not dissolve the non polar compound effectively At 95 acetonitrile the graph only shows two peaks rather than three The acetophenone and phenol have combined because their k values are practically identical 0 1102 and 0 1199 respectively The retention times of each molecule also decreased substantially Increasing acetonitrile concentrations helps elute polar substances and improve the separation and retention times of each molecule It is practically beneficial as well as a higher percentage of acetonitrile makes the mobile phase less polar which decreases the length of the trials Increasing the water concentration allows the substances to stay in the stationary phase longer 4 The chromatogram shifts to the right as each compound shows an increase in k and retention time As methanol is more polar than acetonitrile there is relatively more interaction of the compounds with the stationary phase due to an increase in relative affinity compared to the mobile phase 5 The total retention time k value and tr for the gradient elution is significantly less than that of the 5 acetonitrile isocratic elution The variance for each compound also decreased drastically suggesting that the results are more precise In the gradient elution there were three distinct peaks compared to only two in the 95 acetonitrile isocratic elution The tR and k values were greater for the gradient elution but not by a very significant margin The benefit of using gradient elution is that the peaks come out more defined and with better resolution and detectability It allows the advantage of the favorable interactions at both high and low polarities within one trial 6 The benefit to a long column in chromatography is that as the length increases the interaction time between the eluting compound and the stationary phase increases increasing the column s efficiency and resolution resulting in well separated peaks The advantage of a shorter column is that the time for separations is reduced because there is a shorter distance for the compounds to travel 7 Based on my observations smaller particle sizes create better efficiencies and less peak dispersion or narrower peaks suggesting less variance associated with a higher level of precision Retention Time min Variance sec 1 2946 2 4553 12 2052 0 6162 1 1554 5 7187 W1 2 min 0 0241 0 0453 0 2240 8 Compound 1 Phenol 2 acetophenone 3 naphthalene 9 Resolution 0 589 t r w 1 2 ave t r 2 4553 1 2946 1 1607 w 1 2 ave 0 0241 0 0453 2 0 0347 resolution 0 589 1 1607 0 0347 19 705 10 N 2 5 55 t r 2 w1 2 H L N phenol acetophenone naphthalene Compound Number of Theoretical Plates Theoretical Plate Height mm N 15969 4 16338 2 16479 9 0 00626 0 00612 0 00607