Complete the following purification table: Protein (mg) Volume Units / (ml) ml Total Units % Recovery Protein concentration (mg/ml) Units/mg Fold Purification Crude homogenate 382.5 45 45 2025 100 8.5 5.3 40% Ammonium sulphate supernatant 280 40 47 1880 6.9 65% Ammonium sulphate pellet 75.6 6 212 1272 12.6 16.8 3.2
Calculate the final percantage of saturated Ammonium sulphate if 468ml of saturated Ammonium sulphate was added to 410ml of a 2% saturated protein solution
The Correct Answer and Explanation is :
To complete the purification table and calculate the final percentage of saturated ammonium sulphate, let’s break the problem into two parts.
Part 1: Completing the Purification Table
Given Information:
| Protein (mg) | Volume (ml) | Units/(ml) | ml | Total Units | % Recovery | Protein concentration (mg/ml) | Units/mg | Fold Purification |
|---|---|---|---|---|---|---|---|---|
| Crude homogenate | 382.5 | 45 | 45 | 2025 | 100 | 8.5 | 5.3 | 40% |
| Ammonium sulphate supernatant | 280 | 40 | 47 | 1880 | 6.9 | 65% | ||
| Ammonium sulphate pellet | 75.6 | 6 | 212 | 1272 | 12.6 | 16.8 | 3.2 |
Protein concentration (mg/ml) = Total Protein (mg) / Volume (ml)
Units/mg = Total Units / Total Protein (mg)
Fold Purification = Protein concentration in the fraction / Protein concentration in the crude homogenate
Let’s calculate the missing values.
- For the Ammonium sulphate supernatant:
- Protein concentration (mg/ml) = 280 mg / 40 ml = 7 mg/ml
- Units/mg = 1880 units / 280 mg = 6.71 units/mg
- Fold Purification = 7 mg/ml / 8.5 mg/ml = 0.82 fold
- For the Ammonium sulphate pellet:
- Protein concentration (mg/ml) = 75.6 mg / 6 ml = 12.6 mg/ml
- Units/mg = 1272 units / 75.6 mg = 16.8 units/mg
- Fold Purification = 12.6 mg/ml / 8.5 mg/ml = 1.48 fold
Completed Purification Table:
| Protein (mg) | Volume (ml) | Units/(ml) | ml | Total Units | % Recovery | Protein concentration (mg/ml) | Units/mg | Fold Purification |
|---|---|---|---|---|---|---|---|---|
| Crude homogenate | 382.5 | 45 | 45 | 2025 | 100 | 8.5 | 5.3 | 40% |
| Ammonium sulphate supernatant | 280 | 40 | 47 | 1880 | 6.9 | 65% | 7 | 6.71 |
| Ammonium sulphate pellet | 75.6 | 6 | 212 | 1272 | 12.6 | 16.8 | 1.48 |
Part 2: Final Percentage of Saturated Ammonium Sulphate
To calculate the final percentage of saturated ammonium sulphate, we will use the formula:
[
\text{Final %} = \frac{\text{Amount of saturated ammonium sulphate added (mg)}}{\text{Total volume after mixing}} \times 100
]
Step-by-Step Calculation:
- Amount of saturated ammonium sulphate added:
- Ammonium sulphate is 100% saturated at 468 ml.
- 100% saturation means the amount of ammonium sulphate in 1 ml of solution is 100% of the solubility limit.
- So, the total amount of ammonium sulphate in 468 ml of 100% saturated solution is 468 ml (as it is fully saturated).
- Total volume after mixing:
- Volume of protein solution = 410 ml
- Volume of ammonium sulphate solution = 468 ml
- Total volume = 410 ml + 468 ml = 878 ml
- Final % saturation:
- (\text{Final % saturation} = \frac{468}{878} \times 100 = 53.3%)
So, the final percentage of saturated ammonium sulphate is 53.3%.
Explanation:
When we add the ammonium sulphate solution to the protein solution, the amount of ammonium sulphate that is mixed (468 ml of a saturated solution) is diluted by the total volume of the combined mixture (878 ml). Thus, the final concentration of ammonium sulphate in the mixture is reduced, and the percentage of saturation reflects how concentrated the ammonium sulphate is relative to its solubility limit at the final volume.