BCHM 3100K-01

Lab #1: Solutions and pH

Spring 2013

Lab #1: Solution Preparation and pH Determination Purpose: The purpose of this laboratory exercise is to prepare various solutions, including two buffer solutions, and to use a pH meter to adjust the pH of the buffers you prepare. Rationale: No matter where you go in your science career, you will have to make solutions. The ability to make solutions accurately is therefore a critical skill. In addition, these solutions will be used later in the semester for a protein purification project, so it is important that you prepare them carefully. Procedure: 1) You and your lab group will make the following solutions (refer to the chart below): 0.045 M aluminum sulfate 4.5 mM aluminum sulfate 25mM phosphate buffer High salt phosphate buffer Either of the following solutions: o 2M ammonium hydroxide o 2M acetic acid These solutions will be shared among the lab, so we need three groups to make each of them. 2) Before coming to lab, calculate the amount of each substance that should be used to make each solution in the table below. Check your calculations with your lab partners to ensure your math is correct before you make your solutions. 3) Weigh out and add the dry chemical(s) to a labeled beaker or flask. Scales, weigh boats, spatulas, stir plates and stir bars are on your benches. The chemicals are on the prep bench. 4) Add distilled water (dH2O) and a magnetic stir bar. Measure volumes in graduated cylinders; volumes marked on beakers and bottles are inaccurate. Prepare the solution to ~25-50 mL less than the final volume required, then adjust the pH by adding acid or base (if necessary; see below). Once the pH is correct, you will add more water to bring the solution up to the final volume. 5) Place the beaker on a stir plate and start stirring slowly. Gradually increase the speed until adequate mixing is achieved. 6) Your instructor will demonstrate how to use the pH meter. Remember to rinse the electrode after use.

BCHM 3100K-01

Lab #1: Solutions and pH

Spring 2013

7) Adjust the pH of your solution (if required) by adding acid or base as appropriate. If you overshoot the pH of your solution by more than 0.5 pH units, remake the entire solutionDO NOT add more acid or base to bring it back to the desired pH. Doing so will excessively increase the salt content of the solution. 8) Bring your solution to its final volume with dH2O. 9) Pour solutions into bottles and label the bottles properly with the name of the solution, the date and your initials. 10) Rinse all glassware and clean up the lab benches. Stir bars are valuable and love to go down the drain. Remove your stir bar and rinse it separately from your beaker. 11) Turn in your lab notebook sheets (duplicates only) to your instructor.

Solutions to Be Made 0.045 M Al2(SO4)3 18H2O (store at 4C) (3%) 4.5 mM Al2(SO4)3 18H2O (store at 4C) (0.3%) 2M NH4OH 2M acetic acid 25 mM sodium phosphate buffer: 0.1407g NaH2PO4- (acid) 1.7371g Na2HPO4- (conjugate base) Adjust pH to 8.0 0.5M NaCl in 25 mM sodium phosphate, pH 8.0 (= high salt phosphate buffer)

Volumes 50 mL 250 mL 50 mL 25 ml 300 mL 100 mL

Molecular Weight Table Chemical Al2(SO4)3 18H2O NH4OH Glacial Acetic Acid NaH2PO4 H2O (monosodium phosphate, monohydrate) Na2HPO4 7H2O (disodium phosphate, heptahydrate) NaCl

Formula Weight or Stock Concentration 666.42 g/mol 35.04 g/mol 17 M 137.99 g/mol 268.07 g/mol 58.44 g/mol