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KeyLabPractical1_2011

KeyLabPractical1_2011 - LAB PRACTICAL 1 18/19 APR 2011 MCB...

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LAB PRACTICAL 1, 18/19 APR 2011 MCB 120L, SPRING 2011 Name:_ ANSWER KEY ______________________ Unknown Buffer #_____ Odd & Even _______ Lab Section (CIRCLE): Lagarias A01 Morand A02 Liu A03 Hilt A04 Lab Practical Problem: Determine the pH of an Unknown Buffer using a pH indicator dye. You have each been given 1.5 ml of an aqueous solution of the pH indicator dye p-nitrophenol (pKa 7.15) and 10 ml of a 0.1 M unknown buffer. Using the microplate reader and 0.1 M stock solutions of HCl and NaOH (available in the front of the lab), design a spectrophotometric assay protocol to determine the pH of the buffer. Please include protocols devised, tables and/or graphs on the accompanying pages and label/append all relevant data. Put your answer in the box below. pH of Unknown Buffer odd = 7.87 (actual = 7.5) even = 7.26 (actual = 7.0) Note 1: No pH meters will be used for this analysis. But, can you assume that the pH of the diluted dye solutions will be the same as those of the buffers/solutions used in their preparation? Note 2: Use 200 μ l final volume in each well of the microplate. You can prepare dilutions in the microplate similar to those used in Exp 0. If you wish to prepare dilutions in microcentrifuge tubes and then transfer to the microplate wells, you will need prepare excess volume of each dilution (eg. 500 μ l), so that you can perform duplicate readings of each dilution. Please use the Microplate template (below) and (if desired) the blank protocol table attached page. 1 2 3 4 5 6 7 8 9 10 11 12 A #1 #9 #17 #25 #33 #41 #49 #57 #65 #73 #81 #89 B #2 #10 #18 #26 #34 #42 #50 #58 #66 #74 #82 #90 C #3 #11 #19 #27 #35 #43 #51 #59 #67 #75 #83 #91 D #4 #12 #20 #28 #36 #44 #52 #60 #68 #76 #84 #92 E #5 #13 #21 #29 #37 #45 #53 #61 #69 #77 #85 #93 F #6 #14 #22 #30 #38 #46 #54 #62 #70 #78 #86 #94 G #7 #15 #23 #31 #39 #47 #55 #63 #71 #79 #87 #95 H #8 #16 #24 #32 #40 #48 #56 #64 #72 #80 #88 #96
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Name:______ ANSWER KEY ___________________ Page #____ 2 ____ S2011 Lab Section (CIRCLE): Lagarias A01 Morand A02 Liu A03 Hilt A04 Table _ 1 _ Sample Mixtures Final vol = 500 μ l Absorbance Values 200 μ l/well Calculations Tube # Dye, μl Unknown Buffer, μl 0.1 N NaOH or 0.1 N HCl Raw Abs λ = 405 Corr Abs λ = 405 relative dye concentration pH A
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