User:Khyra A. Neal/Notebook/Chem 571/2014/09/30

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September 30, 2014 Tasklist 1. Calibration Curve for CaCl2 Prepare 25 mL 50 mM CaCl2 using HPLC water 0.13875 g CaCl2 in 25 mL HPLC water Prepare 25 mL of each using serial dilutions: 5 mM, 500 μM, 50 μM, and 5 μM CaCl2 from 50 mM stock CaCl2 Dilute all solutions to 25 mL with HPLC water. Measure Ca2+ using ISE CaCl2 concentration Potential (mV) 5μM -44.6 50μM 8.2 500μM 39.5 5mM 63.5 50mM 86.6 Measure HPLC water, DI water, tap water, and your lysozyme stock solution. Substance Potential (mV) Tap water 51.3 HPLC water -23.0 DI water -51.5 Lysozyme 0.5g/L -19.1 Measure 50 mM and 50 mM of other salts to check for interferences CaCl2 concentration Cu(II)Cl2 concentration KCl concentration Potential (mV) 50mM 0 50mM 86.4 50mM 50mM 0 80.5 2. Calibration Curve for NaCl Prepare 25 mL of 50 mM NaCl using HPLC water 0.07305 g NaCl in 25 mL HPLC water Prepare 25 mL of each using serial dilutions: 5 mM, 500 μM, 50 μM, and 5 μM NaCl from 50 mM NaCl stock solution Dilute all solutions to 25 mL with HPLC water Measure Cl- using ISE NaCl concentration Potential (mV) 5μM 146.5 50μM 129.6 500μM 81.3 5mM 27.5 50mM -23.9 Measure HPLC water, DI water, tap water, and your lysozyme stock solution. Substance Potential (mV) Tap water 65.5 HPLC water 163.5 DI water 158.8 Lysozyme 0.5g/L 89.4 3. Try measuring 50 mM and 50 mM of other salts to check for interferences NaCl concentration NaSO4 Na2HPO42- Potential (mV) 50mM 50 mM 0 -27.0 50mM 0 50mM -30.8 Dialysis Prepare 10 mL 0.5 g/L Lysozyme 0.005 g Lysozyme in 25 mL HPLC water using volumetric flask Prepare 0.25 mM HCl from 2.5 mM HCl stock solution 10x dilution from stock Cut a piece of 3500 MWCO flat dialysis tubing, wet it, and insert it in the 5-well dialysis chamber and clamp the dialysis chamber shut. Fill one side of each well with 1 mL of 0.5 g/L Lysozyme solution Measure out 1 mL with transfer pipette then add 1mL using Pasteur pipette to the wells Fill the other side of the wells with the following solutions: HPLC water, 0.25 mM HCl, 50 mM CaCl2, 500 μM CaCl2, and 50 mM NaCl Insert screws to prevent leaving/evaporation Place on low speed shaker overnight.