Calorimetry Lab
First, the buffer was prepared by using the formula as follows:
Figure 1: Calculation for prepare 0.1 M potassium phosphate buffer at pH 6
3.4007g of potassium phosphate was weighed and placed in 300 mL beaker. Then, 125 mL of water was added into the beaker that contained potassium phosphate. The mixture was dissolved using the stirring rod, and then the magnetic stirring bar was placed in the beaker for further dissolve when measuring the pH. The pH meter was used to measure the solution, and the data was documented at pH 4.6. This was the starting point. Next, 1M NaOH was slowly added into the buffer to make it to pH 6. Then, the buffer was transferred to a 1000 mL graduated cylinder, and 125mL of distill water was added to the buffer. …show more content…
As you can see in table 1, the percent bound of BSA-PR at pH 4 (61.543%) was highest compare to other pH. pH 6 has the percent bound of 3.569%, pH 8 has the percent bound of 2.330%, and pH 10 has the percent bound of 2.309%. pH 6, 8, and 10 do not have an optimal binding for bovine serum albumin and phenol red because they have very low percent bound (refer to table 1), mostly is the free phenol red. Therefore it matches with the prediction. Moreover, there were 2 peaks presented when graphing. Based on the order of elution, the first peak is the bound complex of BSA-PR. According to the data in table 1, the percent bound of BSA-PR was 61.65%. It is not 100% bound, so the second peak is the free phenol red. It is the same with pH6, pH8, and pH 10. Order of elution based on the size of the molecules, the complex of BSA-PR (68,000 Da) eluted first because its size is bigger than the exclusion size of the resin and cannot enter the pores of the bead. Thus, it will travel rapidly and elute first. The BSA is about 66,000 Da, and the phenol red is about 350 Da. The free phenol red will elute last due to its smaller size (350 Da). Small molecules easily enter and fit in the many pores of the beads. Therefore, it takes more time to run down the column and then elute
Figure 1: Calculation for prepare 0.1 M potassium phosphate buffer at pH 6
3.4007g of potassium phosphate was weighed and placed in 300 mL beaker. Then, 125 mL of water was added into the beaker that contained potassium phosphate. The mixture was dissolved using the stirring rod, and then the magnetic stirring bar was placed in the beaker for further dissolve when measuring the pH. The pH meter was used to measure the solution, and the data was documented at pH 4.6. This was the starting point. Next, 1M NaOH was slowly added into the buffer to make it to pH 6. Then, the buffer was transferred to a 1000 mL graduated cylinder, and 125mL of distill water was added to the buffer. …show more content…
As you can see in table 1, the percent bound of BSA-PR at pH 4 (61.543%) was highest compare to other pH. pH 6 has the percent bound of 3.569%, pH 8 has the percent bound of 2.330%, and pH 10 has the percent bound of 2.309%. pH 6, 8, and 10 do not have an optimal binding for bovine serum albumin and phenol red because they have very low percent bound (refer to table 1), mostly is the free phenol red. Therefore it matches with the prediction. Moreover, there were 2 peaks presented when graphing. Based on the order of elution, the first peak is the bound complex of BSA-PR. According to the data in table 1, the percent bound of BSA-PR was 61.65%. It is not 100% bound, so the second peak is the free phenol red. It is the same with pH6, pH8, and pH 10. Order of elution based on the size of the molecules, the complex of BSA-PR (68,000 Da) eluted first because its size is bigger than the exclusion size of the resin and cannot enter the pores of the bead. Thus, it will travel rapidly and elute first. The BSA is about 66,000 Da, and the phenol red is about 350 Da. The free phenol red will elute last due to its smaller size (350 Da). Small molecules easily enter and fit in the many pores of the beads. Therefore, it takes more time to run down the column and then elute