39 Acid base titration: Alanine (HA2) (2- aminopropanoic acid) with sodium hydroxide

Transcription

1 Chemistry Sensors: Loggers: ph, Drop / Bubble counter Any EASYSENSE Logging time: EasyLog Teacher s notes 39 Acid base titration: Alanine (HA2) (2- aminopropanoic acid) with sodium hydroxide Read Amino acids, with certain exceptions, are soluble in water and insoluble in non polar organic solvents such as ether, chloroform and propanol. This is not in keeping with known observations of the properties of carboxylic acids and organic amines. The solubility information on amino acids suggests structures which have charged highly polar structures. Amino acids contain both amino and carboxyl groups and should react with both acids and alkalis. Substances that react to both alkalis and acids are known as amphoteric substances In this investigation the amino acid alanine is used. Alanine has been chosen as it is a low cost, simple to obtain amino acid with data for alanine titrations widely available for comparison. Other amino acids can be used. When alanine is dissolved into water it will give a solution of neutral ph. The solution of alanine in water does not show migration of ions to electrodes if an electric current is passed through it. The same behaviour will also be shown if the molecule exists as the zwitterion, a condition in which negative and positive charges exist at the same time. In this investigation a solution of acidified alanine will have sodium hydroxide added to it to produce a titration curve, from which the isoelectric point, acidic and base dissociation ph values can be determined. Apparatus 1. An EASYSENSE logger. 2. A Smart Q ph sensor. 3. A Smart Q Drop / Bubble counter set to the correct volume range (see calibrating the Drop counter) with alignment adapter fitted. 4. The Drop counter reagent reservoir fitted with two stopcocks and tip x 200 ml beakers. 6. Magnetic stirrer and stir bar cm 3 of 0.1 mol dm -3 Sodium hydroxide (NaOH) cm 3 of acidified Alanine solution ph 1 (see Notes). 9. Measuring cylinders (50 ml capacity). T39-1(V2)

2 Set up of the software and logger The investigation uses EasyLog. Notes Published data for alanine Isoelectric point = ph 6.02 Pka 1 (Acidic dissociation) = ph 2.34 Pka 2 (Base dissociation) = ph 9.69 Using the Drop Counter to measure the volume of titrated fluid, and the standard ph Sensor we have obtained values of ph 6.58, 2.3 and 9.5 (respectively) with good repeatability of results. The sample investigations that lead to this published investigation used, Alanine 0.5 g in 30 cm 3 of analytical water (0.18 mol dm -3 ) 0.1 mol dm -3 reagent quality Sodium Hydroxide 2.0 mol dm -3 reagent quality Hydrochloric acid The alanine solution was acidified to ph 1.0 using drops of the 2.0 mol dm -3 hydrochloric acid. Most published information starts with the acidified alanine and titrate alkali into it. To make extension work / study easier the same method was used. Acid titration into the alkali alanine is possible, again use 1.0 mol dm -3 NaOH to make the solution alkaline (ph 12 will be sufficient). Calibrating the Drop counter This investigation uses the Drop counter with one of its preset calibrated ranges e.g. 27 drops/cm 3 so the drops counted are automatically converted and displayed as volume in cm 3. If accuracy is not critical and you are using the reagent reservoir and tip supplied with a low viscosity liquid (like water) and the flow rate set to: - Fast e.g. 10 plus drops per second, use the 24 drops/cm 3 range. Medium e.g. between 5 10 drops per second, use the 25 drops/cm 3 range. Slow e.g. between drops per second, use the 26 drops/cm 3 range. Very Slow e.g. less than 1.5 drops per second, use the 27 drops/cm 3 range When used with a ph Sensor, the flow rate is best set very slow (less than 1.5 drops per second) to allow the ph Sensor time to settle to a new reading after addition of the titrant. To calculate the number of drops in a cm 3 1. Set up the reagent reservoir in the alignment adapter of the Drop / Bubble Counter. Close both stopcocks and fill the reservoir with the type of solution being used. 2. The first step is to adjust the flow rate. Place a beaker under the stopcock to catch the drops. Fully open the lower stopcock. Slowly turn the top stopcock round until it begins to produce drops and then finely adjust the drop rate. When the correct flow rate of drops is achieved close the lower stopcock to stop the flow. Now the flow rate is set, do not adjust the top stopcock leave in this position. Use the lower stopcock to turn the drops on and off. 3. Top up the reservoir. Place an accurate measuring container e.g. volumetric flask (10 ml or less) under the dropping tip. Open the lower stopcock fully and count the number of drops required to fill up to the volume mark on the measuring container. You can use the Drop / Bubble Counter to count the total number of drops (set to the Drop / bubble count range). Make sure you zero the Sensor before each run. Close the lower stopcock to stop the drops. T39-2(V2)

3 4. Divide the number of drops by the volume (in cm 3 ) to get the drops per cm 3 value e.g. 272 drops fill a capacity of 10 ml = 27.2 drops/cm 3. Top up the reservoir and repeat three times to get an average value. Results and analysis The data will have been collected as a volume and ph vs. time graph. When the Values tool is used to read ph or Volume values, the data from the graph lines appear in the data boxes in the bar above the graph. The determination of isoelectric point and Pka 2 are relatively straightforward and take advantage of the advanced features of the software. The determination of the Pka 1 value requires a little more work, and it is worth spending time to understand the process involved. Advanced To find the isoelectric point of alanine use the maximum rate of ph change. The graph produced is known as the first derivative and is the method used in automated titrations to determine end / equivalence points. If the process is repeated a second derivative is created which increases the accuracy of finding the end point. The response time of the ph electrode is such that the second derivative should be regarded as an extension point in this investigation. The a dx/dt function can be applied to the ph data to produce a first derivative curve. From the Tools menu select Post-log Function. Choose either the: Formula function and select the function a dx/dt, use ph as the channel. Preset function and select Titration, First derivative of ph, use ph as the channel. If the titration was of alkaline alanine to acidic alanine the value for a =-100, the falling rate of change will give a negative peak, using a negative sign converts the peak back to a positive value. The value of a is set at 100 to make sure the peak on the new plot fits the same axis limits, if a value of 1 is used the sensor axis limits will need to be adjusted for the new rate of change plot. The new plot should reveal 2 sharp peaks. The first peak will indicate the isoelectric point of the alanine molecule. Click on Values and line the cursor to the first peak on the rate of change of ph plot, read off the ph value and volume of titrate from the data boxes at the top of the graph screen, record the information. The ph value is the isoelectric point; the volume value is used to determine Pka 2. The method for determining the two Pka values is outlined in the student s notes. The second peak will represent the point at which the alanine molecule became basic. T39-3(V2)

4 Rate of change (ph) Pka V 1 V Isoelectric point V m Time 1hr 4m To find the Pka 1 Value The Pka 1 value is the halfway point between the initial stable ph and the rapid change of ph as the isoelectric point is reached. It is not easily seen and the rate of change of ph that marks the Pka 1 point is not sufficient for it to be revealed as first or second derivative. If you look carefully at the first part of the graph you will notice that at first the rate of change of ph with each addition of the sodium hydroxide is a linear and very slight. As the volume of sodium hydroxide increases the rate of change of ph also increases. What we are looking for is the point at which the slow change in ph becomes the faster change in ph. To find this you will need to 1. Set the graph to display Volume on the y-axis (vertical) and Time on the x-axis (horizontal). 2. Select Print Graph (File menu). If possible print onto graph paper with a printed grid in landscape mode. Rate of change (ph) Line through ph and Volume at Isoelectric point V Intersect with volume graph V 5 V m Time 1h 4m On the printed graph use a ruler to draw a best fit straight line along the first section of the ph curve (L 1 ). 1. Use a ruler to draw a best fit straight line along the ph curve back from the region just before the ph starts to make a rapid change (L 2 ). T39-4(V2)

5 2. Make sure the two best fit lines intersect. Draw a line from the intersect up to the Volume trace and then across to the y-axis (L 3 ) to estimate the volume (V 4 ) at this point. 3. You will have already found the volume at the first peak (V 1 ) - the isoelectric point. 4. Add V 4 to V 1, and divide by 2. This will give the volume at the Pka 1 point (V 5 ). 5. Go back to the graph on the computer and use Values to find the value for ph at the volume you have just calculated (V 5 ). This ph value is Pka 1. Extension work If you have not seen a titration of sodium hydroxide vs. hydrochloric acid, use 0.1 mol dm -3 solutions of each to conduct a titration. Use 30 cm 3 in the beaker to ensure the tip of the ph electrode is covered. Try a titration of other amino acids, dissolve the amino acid in water and acidify to ph 1 with hydrochloric acid (sample titration graphs and methods are available on the web, try searching for the amino acid name and titration). T39-5(V2)