1. PREPARING LABORATORY SOLUTIONS AND REAGENTS II

2. Solutions With More Than One Solute
RECIPE I
Na2HPO g
KH2PO g
0.4% glycerol 10 mL
Dissolve in water.
Bring to a volume of 1 liter.
Recipe lists amounts of solutes.
This is easy to follow

3. Solutions With More Than One Solute Cont…
RECIPE II
1 M MgCl2
0.1 M Tris
Gives the final concentration of each solute.
You must calculate the amount of each to use.
This is harder to interpret

4. A Recipe With Multiple Solutes, SM Buffer
0.1 M NaCl
1 mM MgSO4
0.2 M Tris, pH 7.5
0.01% gelatin
This might be whole recipe as given in a manual

5. Strategy 1: PREPARING SM BUFFER WITHOUT STOCK SOLUTIONS, OVERVIEW:
Prepare a solution of 0.2 M Tris, pH 7.5.
Calculate grams of each of the other solutes required.
Weigh out solutes and dissolve directly in Tris buffer.

6. Strategy 1 Decide how much buffer to make, for example, 1 liter.
1 liter of 0.2 M Tris requires 24.2 g of Tris base. (MW = g/mole.)
Dissolve Tris in about 700 mL of water and bring pH to 7.5.
Do not bring Tris to volume.

7. 4. 1 liter of 0. 1 M NaCl requires 5. 84 g of NaCl
4. 1 liter of 0.1 M NaCl requires 5.84 g of NaCl. Add this to Tris buffer.
5. 1 liter of 1 mM MgSO4 requires 1/1000 of its MW.
MgSO4 comes in more than one hydrated form; read
container to determine MW.
Weigh out correct amount.
Add it to Tris buffer.

8. 0.01% gelatin is 0.1 g in 1 L. Weigh out and add to Tris buffer.
Stir to dissolve
BTV 1 L
Check and record pH.

9. Strategy 2: PREPARING SM BUFFER WITH STOCK SOLUTIONS In overview:
Four solutes are each prepared separately as concentrated stock solutions.

10. When the four stocks are combined, they dilute one another to the proper final concentrations

11. Steps Are: Prepare a stock solution of Tris buffer at pH 7.5.
a. No set rule as to what should be concentration of stock.
b. Example: To make 1 L of 1 M stock, dissolve g of Tris base in about 900 mL of water.

12. c. Bring pH to 7.5.
d. BTV 1L.
2. Prepare a stock solution of magnesium sulfate, for example, 1 M.
a. To make 100 mL, dissolve 0.1 MW of MgSO4 in water, bring to a volume of 100 mL.

13. 3. Prepare a stock solution of NaCl, for example,1 M.
a. To make 100 mL of stock, dissolve 5.84 grams in water and BTV 100 mL.

14. Prepare a stock solution of gelatin, for example, 1%.
a. Dissolve 1 g in a final volume of 100 mL of water.

15. 5. To make the final solution, combine the right amounts of each stock.
a. Since this is a situation where stocks are diluted, use the C1V1 = C2V2 equation
four times, once for each solute.

16. For example, to calculate how much
Tris stock is required:
C1 V1 = C V2
(1 M) (?) (0.2 M) (1000 mL)
? = 200 mL

17. Summary To make 1000 mL of SM buffer, combine: 200 mL Tris stock
1 mL MgSO4 stock
100 mL NaCl stock
10 mL Gelatin stock
BTV 1L; check and record pH.

18. Which Strategy? Both strategy 1 and 2 are correct.
Generally efficient to make stock solutions of frequently used solutes because weighing out chemicals is time-consuming.