 Dear students, today we will learn about preparation of buffer. Buffer is a solution that can resist the change in pH upon the addition of acidic or basic component. Buffers are usually able to neutralize the amount of acid or base when added to the solution. Thus it neutralizes the pH of the solution. It is very important to maintain the pH of the solution while working in the laboratory. So buffers are usually used in most of the wet lab experiments. Let's have a look on the types of buffer. Buffers are usually classified into two main types, acidic and basic. Acidic buffers usually have less than 7 pH. Acidic buffers are usually formed by the addition of weak acid plus salt and strong base. For example, for the preparation of an acidic buffer, we will take an example of acetate buffer. How acetate buffer will be formed? First of all, we will take a weak acid and we all know that acidic acid is a weak acid. When it will combine with strong base, let's see what happens. This hydroxyl group will form a bond with hydrogen and water molecule will be released. And the remaining will be the salt of this weak acid, which will be sodium citrate. Now by combining this weak acid with this salt, we can successfully make the acetate. The second type of the buffer is basic buffer. So usually basic buffers have greater than 7 pH. Now let's see how basic buffers are formed. Like acidic buffer, basic buffers are formed by adding weak base, its salt and with its conjugate strong acid. Let's have a look on the example of the basic buffer. We know that ammonium hydroxide is a weak base and hydrochloric acid is a strong acid. When a weak base will be combined with the strong base, what will be formed? This hydrogen will form a bond with this hydroxyl and release the water molecule. The remaining product will be the salt of this weak base, which will be ammonium chloride. Now combining this weak base with its salt, we can make ammonium buffer. Which is an example of basic buffer. In order to make buffer, either it is acidic buffer or basic buffer, we should know the value of dissociation constant for acid which is pKa and this is dissociation value for the base which is pKb. Let's have a look on the preparation of phosphate buffer that how to calculate, how to done calculations before going to the experiment. So for phosphate buffer, first of all, we have sodium dihydrogen phosphate and its molecular weight is 119.98 gram. Similarly for its salt, disodium hydrogen phosphate, the molecular weight is 141.96 gram. We know the dissociation constant value which is 7.21. Now for what strength we should make the buffer? Today we will make 0.1 molar phosphate buffer and our desired pH is 7.4. So the equation will be pH which is 7.4 is equal to the value of pKa which is 7.21 plus log into B over A, which is the concentration of the moles that how much moles of base and how much moles of acid will be required to calculate the mass. So by solving this equation, here we will have this result and after that to evaluate the moles of base and moles of acid, we will calculate, we will solve this equation by giving this value of moles that we want to create for this buffer. For example, we want to make 0.1 molar phosphate buffer. So what we will do? We will kept here 0.1 and solve it with this value. From this equation, we can evaluate the moles of acid. So by solving this, the moles of acid will be 0.059 moles and from the value of moles of acid, we can calculate the moles of base which will just subtracting the required concentration of the phosphate buffer with the moles of acid. So 0.1 minus 0.059 moles which was the value for the acid, we can calculate the moles of base and the value will come 0.091 moles. Now once we have the values for the moles of base and the moles of acid, we can calculate the mass in gram that we will weigh in the lab to make the buffer. So how we will do the calculation? For the acid calculation, we will take the moles of acid multiplied by its molecular weight. The value will be 7.08. Likewise for base, take the value for the moles of base and multiply it molecular weight for the base. By solving this equation, we will have the value for the mass that we are going to weigh for acid and vein. Once we know the mass to be weigh, we can just measure and weigh this quantity of acid and base and dissolve it 1000 ml of distilled water to make the buffer. So this was pre-calculation or you can say this was the calculation before making the buffer in the lab. You have to do the calculation for all type of the buffer that you are going to make. Now let's perform the experiment. To prepare phosphate buffer in the lab, we will weigh the measured quantity of sodium dihydrogen phosphate and disodium hydrogen phosphate. So let's weigh the sodium dihydrogen phosphate. We will weigh 7.08 gram of sodium dihydrogen phosphate. With the help of spatula, first of all, we will weigh the vein balance. By measuring the desired quantity of weak acid, we will add this reagent to beaker. Afterwards, we will again weigh the salt of this buffer, which is disodium hydrogen phosphate. By measuring the quantity of base, we will also add this into the beaker. Once you have measured the quantity of weak acid and its salt or base, you will then take the quantity of distilled water in measuring slender to make the solution. We have done the calculations for 1 liter preparation of phosphate buffer. So we will take 950 ml of distilled water. First, we will take the half quantity of water in order to mix the reagents. After mixing, we will add the remaining quantity of distilled water. We will stir the reagents and water to have a colorless, clear solution. Now we will check the pH of the solution. Through pH meter, we will assess the pH of the solution. This is a pH sensor which is dipped into our neutral buffer solution. It should be noted that this pH sensor should be handled carefully in order to avoid any cooling. Just dip the sensor into your solution and observe the pH on the pH meter. The pH of the solution is 7.1, but we need buffer of 7.4 pH. So what we will do? We will, in order to increase the value of pH, we will add some drops of diluted base. So here you can see we have kept the diluted acid and diluted base. So to maintain our desired pH, we will add some drops of diluted base. After the addition of base into the buffer solution, we will again check the pH of the solution. By adding the base into the solution, the pH of the solution has been dried up. After the pH maintenance, our buffer solution is ready and we can store this buffer in reagent bottle for our future experiment. Now we will proceed towards the preparation of citrate buffer. As I already told you that citrate buffer is a type of acidic buffer. So after doing the calculation, what we will do? We will measure the desired quantity of citric acid and its salt and then we will mix with the distilled water to make the buffer solution. Let's perform the experiment. First of all, we will weigh the quantity of citric acid and its salt. Whenever you want to weigh something in a weighing balance, you must tear the weighing balance before. After the addition of both reagents, we will measure the distilled water in measuring cylinder and add hinda. First we will take the half quantity of water, dissolve it to make a colorless solution. You can also mix the reagents with distilled water with the help of magnetic stirrer on the hot plate. Now, measure the remaining quantity of distilled water. Add the remaining distilled water to raise the volume till 1000 ml or 1 liter. After mixing the reagents of citrate buffer with distilled water, we will check the pH of the buffer solution. You can simply dip the pH sensor in the desired buffer solution to evaluate the pH. In order to reduce the pH, we will add some drops of weak diluted acid. Carefully add some drops of diluted acid to the buffer solution. After the addition of some drops of diluted acid, we will again measure the pH of the buffer solution. Once you have prepared the buffer, you can store it in the reagent bottle for your future experiment. So in this practical, we have analyzed that how to make buffer and we have made phosphate and citrate buffer. In order to make any buffer in the lab, you have to do some calculations and then after measuring the desired quantity, you can easily adjust the pH through pH meter and can make buffer solution.