Preparation of Buffers Claire Anne G.

Punsalan De La Salle University-Dasmarias Dasmarias, Cavite

ABSTRACT In this experiment, the pH of different solutions and water were measured to see how pH was affected when acid or base was added into the solution. A buffer is a mixture of a weak acid [HA] and its conjugate base [A-] or a weak base and its conjugate acid. It is a solution that resists a drastic change in pH when acids or bases are added to it. Phosphate buffer (pH 7.40) was the buffer system used in the experiment. It was prepared using H2PO4--HPO4-2. A pH of 7.29 was prepared and tested. Adding 0.100 M HCl or 0.100 M NaOH to the solution tested the buffer. When 5 ml of 0.100 M HCl was added into the buffer, the pH decreased by 0.14 (7.15). When 5 ml of 0.100 M NaOH was added into the other buffer, the pH increased by 0.25 (7.54). It only shows that the buffer was able to resist drastic change when an acid or base was added into the solution. Unlike the buffer solution, the pH of distilled water was greatly affected when acid or base was added into it.

INTRODUCTION The objective if the experiment was to prepare a buffer system starting from needed calculations up to the actual preparation. Buffers are used in biochemistry to stabilize and modulate the pH of a solution. Creating a mixture of a weak acid and its conjugate base can make a buffer solution in a variety of ways. Buffer solutions are used to make a solution exhibit very little change in its pH when an acid or bases are added to it. Buffer solutions also have many applications in chemical manufacturing for processes that require a specific pH range to work. (1). The pH of a buffer solution is calculated by using Henderson-Hasselbalch equation. The equation has the formula pH= pKa + log [A-]/[HA]. The pH of a solution is equal to acid dissociation constant of the weak acid plus the logarithm of the ratio of conjugate base over weak acid. In biological systems, the buffer system of carbonic acid and sodium bicarbonate used to maintain the pH of blood to a constant value (2). In this experiment, buffer system was used to demonstrate how buffer resists drastic change when an acid or base was added.

MATERIALS AND METHODS The reagents used in the experiment were K2HPO4 (base), KH2PO4 (acid), 0.100 M HCl and 0.100 M NaOH. The needed ingredients in preparing 250 ml of 0.200 M H2PO4-HPO4-2 (pH 7.40) were calculated. The calculated amount of K2HPO4 was 5.3131 g and 2.654 g for the KH2PO4The exact amount of reagents was weighed using an analytical balance. The reagents were transferred into 250-ml volumetric flask. The flask was half-filled with distilled water and swirled until the solid dissolves. The flask was filled-to-mark when the reagents were completely dissolved. The flask was inverted repeatedly to ensure complete mixing. 25 ml of the buffer was transferred into two separate 100-ml Erlenmeyer flask using a volumetric pipette. The pH was measured using the pH meter and the percent error was also determined. After measuring the pH, 5.00 ml of 200 M HCl was added to one flask and 5.00 ml of 0.200 M NaOH was added to another flask. The flasks were swirled and the pH of each sample was measured. The procedures were repeated using distilled water instead of the buffer. The remaining buffer was placed in a clean plastic bottle. All used samples were combined for neutralization.

RESULTS AND DISCUSSIONS A buffer is a mixture of a weak acid [HA] and its conjugate base or a weak base [A-] and its conjugate acid. It is capable of resisting drastic change in pH when acids or bases are added to it. The pH of a buffer can be determined by using Henderson-Hasselbalch equation, which was equal to acid dissociation constant of the weak acid plus the logarithm of the ratio of conjugate base over weak acid. The acid dissociation constant (pKa) is the measure of strength of acids. The phosphate buffer is consists of dihydrogen phosphate ions (H 2PO4-) as hydrogen ion donor (acid) and hydrogen phosphate ions (HPO42-) as hydrogen-ion acceptor (base). Dihydrogen phosphate ion rapidly dissociates to produce hydrogen phosphate ion and H30+ as illustrated in Figure 1.

Figure 1. Acid dissociation of Dihydrogen Phosphate Addition of acid or base can affect the pH of the buffer. The pH increased or decreased depending on the reagent added into the buffer solution. Meanwhile, the table below shows the initial pH of the buffer solution and distilled water and its pH when 5.00 ml of 0.100 M HCl or 5.00 ml of 0.100 M NaOH was added to it. Table 1. Addition of Acid or Base pH Theoretical Initial + 5.00 ml of 0.100 M HCl + 5.00 ml of 0.100 M NaOH 7.40 7.23 7.60 Phosphate Buffer Experimental 7.29 7.15 7.54 Distilled Water % Error 1.49 % 1.11 % 0.79 % 7.24 2.21 11.59

A pH of 7.29 was prepared and tested. When 5 ml of 0.100 M HCl was added into the buffer, the pH decreased by 0.14 (pH=7.15). When 5 ml of 0.100 M NaOH was added into the other buffer, the pH increased by 0.25 (pH=7.54). It only shows that the buffer was able to resist drastic change when an acid or base was added into the solution. The pH of distilled water was greatly affected when acid or base was added into it. The pH of water decreased by 5.03(pH=2.21) when 5 ml of 0.100 M HCl was added. The pH increased by 4.35(pH=11.59) when 5 ml of 0.100 M NaOH was added. In doing the experiment, there are possible errors that could take place that can affect the experimental pH of the buffer. Contamination could be a source of error in this experiment. Different beakers and other equipment were used in the experiment. Unclean equipment could contain contaminants that could affect the pH of the buffer. Adding the wrong amount of acid or base could also affect the buffer. Accuracy of pH meter could also be a source of error. Some pH meter could possibly display inaccurate results. Bicarbonate buffer is an example of a synthetic organic buffer. It was prepared by using carbonic acid (H2CO3) as acid and bicarbonate ion (HCO3 -) as the conjugate base. Carbonic acid rapidly dissociates to produce bicarbonate ion and H30+ as illustrated in Figure 2.

Figure 2. Acid dissociation of Carbonic acid The advantage of using this buffer is it is capable of shifting the chemical equilibrium according to Le Chateliers principle. When the blood gained excess hydrogen ions, some of those hydrogen ions would shift to carbon dioxide, minimizing the increased acidity (3). Blood plasma is a biological fluid that plays a significant role in maintaining the pH. In blood plasma, the carbonic acid and hydrogen carbonate ion buffers the pH. In this buffer, carbonic acid (H2CO3) is the hydrogen-ion donor (acid) and hydrogen carbonate ion (HCO 3-) is the hydrogen-ion acceptor (base). This buffer functions in exactly the same way as the phosphate buffer. Additional H+ is consumed by hydrogen carbonate and additional OH- is consumed by carbonic acid. When carbon dioxide concentration increases, it stimulates increased breathing, and the excess carbon dioxide is released into the air in the lungs. Respiratory acidosis took place when the blood pH falls due to decreased respiration. The concentration of dissolved carbon dioxide in the blood increases when respiration is restricted which makes the blood too acidic. Metabolic acidosis is the decrease in blood pH that results when excessive amounts of acidic substances are released into the blood. Excessive breathing which produces an increase in blood pH results to Respiratory alkalosis. Metabolic alkalosis is due to an increase in blood pH resulting from the release of alkaline materials into the blood (4).

CONCLUSION

Phosphate buffer is capable of resisting drastic change when an acid or base is added to it. This is due to the presence of a weak acid [HA] and its conjugate base [A-] or a weak base and its conjugate acid. When hydrogen ions were added to a buffer, the base in the buffer neutralized them. The acid neutralized hydroxide ions. These neutralization reactions would not have much effect on the overall pH of the buffer solution. REFERENCES (1) Cloe, Adam. What Does a Buffer Do in Chemistry?. Retrieved 30 November 2009 from http://www.ehow.com/about_4609033_what-does-buffer-do-chemistry.html (2) Buffer Solutions. (n.d.). Transtutors. Retrieved November 30, 2009, from http://www.transtutors.com/chemistry-homework-help/ionic-equilibrium/buffersolutions.aspx (3) Numerator, c. t. (n.d.). StateMaster - Encyclopedia: Chemical equilibrium. StateMaster - US Statistics, State Comparisons. Retrieved November 30, 2009, from http://www.statemaster.com/encyclopedia/Chemical-equilibrium (4) Chemical of the Week -- Biological Buffers. (n.d.). Science is Fun in the Lab of Shakhashiri. Retrieved November 30, 2009, from http://scifun.chem.wisc.edu/CHEMWEEK/BioBuff/BioBuffers.html