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Lab 1 Water, pH, Buffers, and Amino Acids

Name: Danye Jiang Lab Partner: Marrisa Jarosinsici TA's Name: Selena Rice Date: Sept 4, 2011

Introduction Water is a major component of many bodily fluids including blood, urine, and saliva. Thus, it's important to fully comprehend the unique properties water possesses that makes it well suited for biological systems. The dissociation of water with biological molecules affects the pH of cells and extracellular fluids. This experiment investigates the dissociation of water with both a simple buffer of acetic acid and a solution of an unknown amino acid. Quantitatively, the experiment attempts to identify the pK' of acetic acid and to identify the unknown amino acid by interpreting the titration curve.

Materials and Methods In experiment 1, the pH meter was standardized for the pH 7-12 range. 0.0200L of deionized water was added to the titration vessel. Using a P-1000 Pipetman pipettor, 8 additions of 0.500M KOH at 0.0001L interval were made to the titration vessel until the pH reached 12. Experiment 2 and 3 also involved the standardization of pH meter and titration with 0.500M KOH. In experiment 2, the pH meter was standardized for the pH 1-7 range and acetic acid was titrated until the pH meter read beyond 10. In experiment 3, the pH meter was standardized first for pH 1-7 range and was re-standardized for pH 7-12 range. An unknown amino acid was titrated until the pH meter reached 12.

Graphs and Tables of Results

(1) Experiment 1: Titration of Pure Water Titration of Pure Water with 0.500M KOH Volume of KOH Added(L) 0 0.00010 0.00020 0.00030 0.00040 0.00050 0.00060 0.00070 0.00080 pH 8.71 11.05 11.30 11.55 11.62 11.65 11.85 11.95 12.05 Titration of Pure Water with 0.500M KOH
13.00 12.50 12.00 11.50 11.00

pH

10.50 10.00 9.50 9.00 8.50 0.00000.00010.00020.00030.00040.00050.00060.0007 0.00080.0009

pH

Volum e of KOH Added(L)

(2)

The calculated pH is 11.4 and the observed pH is 11.05. The pH calculated is slightly higher than the pH observed with a deviation of (11.4-11.05)/(11.4)=4%. Water has low buffer capacity.

(3) Experiment 2: Titration of a Buffer, Acetic Acid Titration of Acetic Acid with 0.500M KOH
12.00 11.00 10.00 9.00 8.00

pH

7.00 6.00 5.00 4.00 3.00 2.00 0.0000 0.0005 0.00100.00150.00200.0025 0.0030 0.0035 0.00400.0045

pH

Volum e of 0.500M KOH Added(L)

(4) Experiment 3: Titration of Unknown Amino Acid Titration of Amino Acid with 0.500M KOH Volume of KOH Added(L) 0 0.00040 0.00080 0.00120 0.00160 0.00200 4 pH 1.61 1.81 1.92 2.05 2.20 2.35

0.00240 0.00280 0.00320 0.00360 0.00400 0.00440 0.00480 0.00520 0.00560 0.00600 0.00640 0.00680 0.00720 0.00760 0.00800 0.00840

2.51 2.71 3.01 3.40 7.90 8.69 9.15 9.41 9.62 9.81 10.01 10.20 10.51 10.95 11.70 12.15 Titration of Amino Acid with 0.500M KOH
14.00

12.00

10.00

8.00

pH

pH
6.00

4.00

2.00

0.00 0.0000 0.0010 0.0020 0.0030 0.0040 0.0050 0.0060 0.0070 0.0080

Volum e of 0.500M KOH Added(L)

The unknown amino acid is graphically determined to be glycine. The curve shows that there

are 2 dissociable groups. At the pHI, no buffering power exists as indicated by the steep slope. Note that the first slope spans pH 1 to pH 5 and requires 1 mole of KOH per mole of the unknown amino acid; the second slope spans pH 8 to pH 11 and also requires 1 mole of KOH per mole of the unknown amino acid. These characteristics indicate the unknown amino acid to be glycine. (5.a) Titration of 0.100M Acetic Acid with 0.500M KOH KOH vol. (L) 0.00040 0.00080 0.00120 0.00160 0.00200 0.00240 0.00280 0.00320 0.00360 Sample Calculation: pH observed 3.69 4.00 4.21 4.22 4.45 4.62 4.81 5.05 5.49 [HA] 0.0882 0.0769 0.0660 0.0556 0.0455 0.0357 0.0263 0.0172 0.085 [A-] 0.00980 0.0192 0.0283 0.0370 0.0455 0.0536 0.0614 0.0690 0.0763 PK' calculated 4.64 4.60 4.58 4.40 4.45 4.44 4.44 4.45 4.54

(5.b) The mean of the calculated pK' is 4.50. The standard deviation of the calculated pK' is 0.09.

(5.c) The graphically determined pK' of acetic acid is 4.60. The mean agrees well with the value determined graphically with a deviation of (4.60-4.50)/4.50=2.2%.

(5.d) The variation of the pK' values appears to be mainly due to systematic errors. In experiment 2, when adding 0.500M KOH at 0.0001L interval to the titration vessel, the volume indicator of the P-1000 Pipetman pipettor was not set up correctly that the indicator digits did not line up perfectly. Due to the inconsistent additions of base, the observed pH fluctuated and eventually led to variation in pK'.

(6)

Conclusion The experimentally determined pK' of acetic acid was found to be 4.60, a 2.2% deviation from the calculated value. The small deviation indicated that the pK' of a weak acid could be reliably determined by titration in a simple buffer and a strong base. However, the experimentally determined pK' was compromised by the 2.2% variation due to systematic errors. The unknown amino acid was identified as glycine by interpreting the titration curve. The curve clearly indicated 2 inflection points (locations of the pK's). It's noted that the first dissociable group 7

spanned pH 1 to 5 and required 1 mole of KOH per mole of the unknown amino acid; the second slope spanned pH 8 to 11 and required also 1 mole of KOH per mole of the unknown amino acid. These characteristics thus eliminated the choice of lysine, aspartic acid, and histidine. The three experiments in this lab were able to justify the established hypothesis in the introduction. Overall, this lab successfully demonstrated the important roles of weak acids and bases as buffers in controlling pH and that of water in biological system.