Buffers

Hepes Buffer, Ripa Buffer, Ph Buffer, Hssb Buffer

Buffer solutions are a mixture of a weak acid and its conjugate base, or a weak base and its conjugate acid. Buffer solutions are used to help keep the pH of another solution mixed with them constantly. When a small amount of acid or alkali is introduced into the solution by addition or a chemical reaction in the solution, the buffer resists changes in the pH of the entire solution. Buffer solutions are therefore very useful for many different applications where a relatively constant pH is required. Buffer; It may also be called a pH buffer, hydrogen ion buffer, or buffer solution.

For example, blood contains natural buffers to maintain a constant pH between 7.35 and 7.45, which is necessary for our enzymes to work properly. Since enzyme activity changes according to pH, it is necessary to maintain a constant pH value in order to observe the correct level of activity in biochemical measurements. Buffers in commercial applications; It can be found in shampoos to prevent skin irritation, in baby lotions to prevent bacterial growth, and in contact lens solutions to keep the pH of the liquid compatible with that of the eye.

The preparation of buffers consists of several steps: weighing the components, dissolving the components, adjusting the pH, and making up to the final volume. Since the ratio of acid to base in the buffer is directly related to the final pH, it is very important to weigh the components with a high degree of accuracy. It is therefore important that the equipment used (balance, pipettes, and pH meter) is properly calibrated and has a sufficient level of accuracy.

Preparing a buffer solution takes time and must be done carefully so that the buffer does its job as intended. When the quality of your products or biochemical analyzes depends on the performance of your buffer solutions, you want to make sure you prepare them right the first time.

Watch the video and you can save time and workload while preparing the right buffer solutions with precision balances and pH meters.

How Are Buffer Solutions Prepared? Typical Procedure.

Preparation of buffers consists of several steps: calculating ingredients (concentrations and quantities) based on required use and intended volume, weighing ingredients, dissolving ingredients, adjusting pH, filling to the final volume, labeling, documenting results, and storing for direct or later use.

Select a recipe from the database

Recalculate prescription quantities based on buffer volume required

Weigh the ingredients in the container

Dissolve the components in a suitable solvent (typically water)

Check and adjust the pH value using the pH meter

Make up the solution to the required volume

Transfer to a storage bottle and label

Document the results

How Buffer Solutions Work

Buffers are aqueous systems that resist changes in pH when small amounts of acid or base are added, and consist of a weak acid conjugated to a base. Buffers keep the pH of the solution constant by absorbing protons that are released during reactions or by releasing them when the reactions consume protons. The discovery that partially neutralized solutions of weak acids or bases are resistant to pH changes when small amounts of strong acids or bases are added has led to the concept of "buffer".

A conjugate base is an acid that has lost a proton.

HA ↔ H+ + A-

acid ↔ proton + conjugate base

A conjugate acid is a base that has gained a proton.

A + H+ ↔ H+A

base + proton ↔ conjugate acid

A balance is established between decomposed and undifferentiated forms.

For example, weak acetic acid partially dissociates in water to produce the acetate ion:

CH3COOH ↔ H+ + CH3COO-

Undissociated acetic acid, hydrogen ions, and dissociated ions exist in equilibrium in solution.

Sodium acetate also decomposes in water producing the same acetate ion:

CH3COONa ↔ Na+ + CH3COO-

Undissociated sodium acetate and sodium and acetate ions exist in equilibrium in the solution.

In this case, an aqueous solution of a mixture of acetic acid and sodium acetate absorbs H+ ions when an acid is added to it, by combining hydrogen ions with acetate base to form acetic acid. Also, when OH- ions enter the solution by adding an alkali to the solution, these ions combine with acid molecules (H+) and turn into water. In this way, the system resists changes in solution pH as it tries to restore equilibrium. This is how buffer solutions work.

What happens after adding acid to the buffer?

When some strong acid (more H+) is added to an equilibrium mixture of weak acid and conjugate base, the equilibrium shifts to the left according to Le Chatelier's principle.

What happens after adding a base to the buffer?

Similarly, when a strong base is added to the mixture, the decrease in hydrogen ion concentration is less than would be expected for the amount of base added. This is because the reaction shifts to the right to accommodate the H+ ions lost in the reaction with the base.

Buffer Solution Types

Buffer solutions consisting of a weak acid and a conjugate base are called acidic buffers and have pH values ​​less than 7. A buffer prepared with acetic acid (weak acid) and sodium acetate (conjugate base) is an acidic buffer and has a pH of approximately 4.75.

Buffer solutions consisting of a weak base and its conjugate acid are called basic buffers and have a pH greater than 7. An example of a base buffer is an aqueous solution of ammonium hydroxide (weak base) and ammonium chloride (conjugate acid) with a pH of 9.25.

What Should Be Considered While Preparing Buffer Solution?

Buffer solutions work best when their pH values ​​are similar to the pH of the system or the solution under consideration. In the study of enzymes in human biology, a system suitable for the pH of the blood, in the range of 7.35-7.45, is required. Otherwise, the enzymes will not function properly. If the pH of the buffer system is outside of the desired range, this will adversely affect the analysis.

Therefore, it is necessary to be able to prepare buffer solutions with a certain pH and this can be done in different ways:

pH adjustment

The crystalline acid or base is first dissolved in water in a volume of 60-70% of the volume of the buffer solution to be obtained. The pH is tested and then adjusted. If a crystalline acid is used, the pH is adjusted using a base that will not produce ions that might interfere with the system under investigation. If a crystalline base is used, the pH is adjusted using a suitable acid. When the desired pH is reached, the volume of the buffer solution is made up of water to obtain the required volume.

Mixing acid and base solutions

This method mixes an acid or base solution with the corresponding salt solution. The concentrations of the source solutions must be the same as the buffer solution required. The solutions can be mixed in varying proportions to achieve different pH values ​​in the resulting buffer solution. Alternatively, pH can be monitored while adding one solution to another.

Using the Henderson-Hasselbach equation

The Henderson-Hasselbach equation can be used to estimate the pH of the buffer solution, using the dissociation constant pKa. When a weak acid (HA) is in solution, the hydrogen ions it dissociates (H+), and the conjugate base of the acid are in equilibrium (A-), the dissociation constant is a strong measure of the acid at this equilibrium point.

Benefits of Universal Buffers

Universal buffer solutions are made up of various acid-base pairs. This allows universal buffers to maintain the pH of a wide variety of solutions, making them suitable for a wider range of applications.

Additional Tips on Preparing and Using Buffers

Creating a buffer preparation SOP (Standard Operating Procedure)

It's a good idea to have everyone follow the same procedure to document the buffer preparation process and ensure consistency and reproducibility. SOP; It should include details of the materials used, the precise steps of when and how to add the ingredients and measure the pH. The SOP may also include many of the points outlined below.

Wear protective clothing

Appropriate personal protective equipment (PPE), such as protective eyewear and clothing, should be worn, especially when working with strong acids or bases.

Control microbial contamination (especially in biology applications)

Before using any buffer, check the container for microbial contamination. Buffer solutions with pH close to neutral are particularly susceptible to contamination. You may see some turbidity in the solution or impurities deposited on the bottom.

Use your pH meter correctly

To ensure accurate pH measurements, your pH meter should be calibrated and maintained regularly. Before use, the electrode must be properly prepared, using a sufficient amount of buffer solution to ensure proper immersion of the electrode connection. Wait for the pH to stabilize before recording the measurement and rinse the electrode with distilled water after the procedure. Be sure to use your pH meter at ambient temperature or use an electrode with an integrated temperature probe.

Pay attention to the temperature

The amount of decomposition may vary with temperature. Therefore, your buffer solution should be prepared at the temperature at which you will perform your analysis. Also, make sure that the temperature at which you calibrate the electrode is the same as the temperature at which you are measuring.

Pay attention to the concentration

It is common practice to dilute ready-made buffer solutions to the required concentration for analysis. However, changing the concentration can affect the amount of dissociation. Since pH is a measure of hydrogen ions (H+), a change in dissociation can cause a change in pH. After reconstitution, the pH should be rechecked before using the buffer.

Challenges of Buffer Solution Preparation

While the simplest buffer solution consists of just an acid, a base, and water, lab technicians often have to prepare a variety of buffer solutions each day, typically consisting of 2-5 components but can number up to 20. A typical laboratory may have more than 20 buffer solution recipes prepared to produce 1 liter of the buffer.

Buffer solution calculations

In a database of more than 20 prescriptions, the lab technician must first ensure that the correct solution recipe is selected. For buffer solution volumes other than 1 liter, all component amounts must be recalculated to match the new volume and the new amounts recorded. Errors in calculations or when recording new amounts can lead to incorrect pH values ​​in the buffer solution. There is a higher risk of error in systems with manual data recording.

Weighing and recording of weighing results

When weighing all these different buffer solution components, care must be taken to use the correct amount of the correct component. The actual weight of each component should be recorded and care should be taken to avoid recording errors if results are recorded manually or by entering them into a computer.

Depending on how much of each buffer solution component is required and the minimum weight of the balance, it may be necessary to use two different balances. 

Accurate performance of pH meters

Controlling the pH level of the buffer solution is vital. However, pH measurements can be inaccurate if the pH meter used is not calibrated or properly maintained. Using a buffer solution with the wrong pH can seriously affect subsequent analysis or product quality.

Documentation

The final buffer solution should be carefully labeled with all the information to avoid confusion. The expiration date is important to ensure that the buffer solution is still active when it is time to use it. All data on the buffer solution prepared should be recorded and stored securely for future reference and traceability.

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