2. As a biochem TA next year, a student comes to you saying that he heard two-dimensional chromatography is better than conventional chromatography. He has run a chromatogram in 1-butanol:acetic acid:water (10:2:5) and wants more of the solvent to do the second dimension now that the first has dried and been turned. What do you tell the student?     (5 points)

3. Write the reaction between KOH and HClO₄. (4 points)
 
4. State the reason for the following actions (20 points)
     a. Diluting enzyme in 0.1% gelatin for kinetic studies.

     b. neutralizing protein acid hydrosylate with Ba(OH)₂. 

     c. diluting stock enzyme in water to determine protein.

     d. in the determination of Km/Vmax, have a tube containing 0.3 M  sucrose and no enzyme be our blank.

5. Plot the following data and determine the protein concentration of the unknown protein in µg/mL. (5 points)
 

                Contents
       ----------------------
       0.2 mg/mL
Tube   BSA std          water        %T
----   -----            -----       -----
1      0.0 mL           1.0 mL      100.0
2      0.2 mL           0.8 mL       68.2
3      0.4 mL           0.6 mL       45.5
4      0.6 mL           0.4 mL       32.1
5      0.8 mL           0.2 mL       10.5
6      1.0 mL           0.0 mL       14.8
7      1.0 mL unknown   0.0 mL       35.5
8      1.0 mL unknown   0.0 mL       34.6
9      1.0 mL unknown   0.0 mL       12.1