User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/01/29

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Objective

  • to reallocate ADA into 0.0253mg per tube
  • to prepare phosphate buffer
  • to prepare adenosine stock solution
  • to prepare ADA stock solution for today
  • to run ADA kinetics using UV-vis


Description

1. Reallocating ADA

  • Measure 2.5mg was added to a tube and stored at 4°C
    • 0.5U/(19.8U/mg) = 0.02525mg

2. 0.05M Sodium Phosphate Buffer (pH 7.4)

  • Add 3.1 g of NaH2PO4•H2O and 10.9 g of Na2HPO4 (anhydrous) to distilled H2O to make a volume of 1 L, 1M Sodium Phosphate Buffer.
    • The pH of the solution was 7.03, so 1M NaOH was added drop-wise until the solution reached a pH of 7.4.
    • This buffer was stored at 4°C.
  • 50mL of the 1M Sodium Phosphate Buffer was diluted in 1L H2O.

3. Preparing a stock solution of adenosine

  • Initially, 0.0802g of adenosine was added to 10mL of 0.05M Sodium Phosphate Buffer (to make 30mM solution), however the solution was super saturated.
  • Then, 0.0401g of adenosine was added to 10mL of 0.05M Sodium Phosphate Buffer (to make 15mM solution), but the solution was still saturated.
    • 5mL of 0.05M Sodium Phosphate Buffer was added to this solution in order (to make 10mM solution), but one flake of adenosine remained out of solution.
    • 5mL of 0.05M Sodium Phosphate Buffer was added to the 10mM solution (to make 5mM solution) and the adenosine was fully dissolved in solution.
  • The final concentration of the adenosine stock solution was 5mM.



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