User:Khyra A. Neal/Notebook/Chem 571/2014/10/08

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October 8, 2014

[I-] by Precipitation Titration

1. Prepare 50 mL 50 mM KI

  • 0.415 g solid KI in 50 mL H2O

2. Prepare 50 mL 75 mM AgNO3

  • 0.637 g AgNO3 in 50 mL H2O

3. Prepare 100 mL 10 mM NH4SCN

  • 0.07612 g NH4SCN in 100 mL H2O

4. NH4SCN Standardization

  • Add 3 mL 1 M HNO3 and 2 mL AgNO3 to 10 mL DI H2O
  • Add 200 μL Ferric alum indicator
  • Titrate with your NH4SCN solution
  • Titrated 15.5 mL NH4SCN when orange color change was observed.
  • Second Titration = 15.5 mL
  • AgNO3+NH4SCN→AgSCN+NH4NO3
    • Reaction is 1:1
    • 2mL of AgNO3=1.5×10-4 mol
    • 15.5 mL of NH4SCN contains 1.5×10-4mol
    • [NH4SCN]=9.68 mM

5. KI Standardization

  • Add 3 mL 1 M HNO3 and 1 mL KI to 10 mL DI H2O
  • Add 2 mL AgNO3 and swirl to complete precipitation
  • Add 200 μL Ferric alum indicator
  • Titrate with your NH4SCN solution
  • Observed a yellow solution with a yellow precipitate because of I- ion
  • Titrated 10.3 mL NH4SCNwhen orange color change was observed.
  • Second titration = 10.5 mL

6. Prepare Dialysis for 0.6 g/ L Lysozyme vs KI using MWCO

  • Add 1 mL Lysozyme to 5 wells on one side
  • Make serial dilutions of KI at 50 mM, 25 mM, 10 mM, 5 mM, and 2 mM
  • Add a different concentration to each well on the opposite side.
  • Place on low speed shaker for 1 week

Continuation of Dialysis Analysis from 10/7/14

1. Measure ISE for Ca2+ using 20,000 MWCO tubing

Substance Potential (mV)
50 mM CaCl2 (1) 74.9
Colloid (1) 73.5
500 μM CaCl2 (2) 91.4
Colloid (2) 51.2

2. pH readings for first 5 well dialysis chamber on October 1, 2014

  • Only solutions containing H+ ions were measured
Substance pH)
Lysozyme against 0.25 mM HCl 7.426
0.25 mM HCl 7.857

3. Fluorescence of Protein Solutions from October 1, 2014

  • Dilute each dialyzed lysozyme solution by a factor of 100
  • Transfer 100 mL of dialyzed Lysozyme solution to small volume fluorescence cuvette
  • Measure fluorescence from 300 nm-550 nm and excitation at 280 nm
  • Clean fluorescence cuvette after each use with 1 N HCl, methanol and HPLC water