PH meter - Determination and adjustment of medium pH
All microorganisms are affected by the pH of the environment in which they live. Often, the presence of selective species narrows the pH range in which they grow. Sometimes, the acidity or alkalinity of the medium can also act as a growth selection factor. For biochemical test media, however, the pH range in which microorganisms exhibit tk activity in the substrate is narrower than the pH range required for growth: therefore, it is important to adjust the pH of the media to the expected range. When the laboratory prepares the medium according to the medium composition, the pH must be checked and adjusted. It must be pointed out that even for the ready-made synthetic dehydrated medium provided by the manufacturer (especially for some stock dehydrated medium), the final pH needs to be measured after making the experimental medium.
The most accurate way to measure the pH of a medium is with a pH meter. Before use, it must be calibrated with a standard buffer solution of known pH (usually pH is 4 to 7). Rinse the electrode of the pH meter with distilled water and immerse it in the culture medium in the beaker, and then read the pH of the culture medium from the scale of the pH meter. Each manufacturer provides detailed operating instructions for each type of pH meter.
When preparing the culture medium, the PH can be easily adjusted by using a pH meter with mechanical stirring. The culture medium should be prepared in a container (such as a large beaker) that can be stirred continuously. or Teflon-wrapped magnets. The pH of the agar-containing medium needs to be adjusted after thawing, which is easily done with a heatable magnetic stirrer. When adjusting the pH, use a standard buffer solution of known pH to calibrate the pH meter first. Then put the composite electrode and resistance thermometer (if any) in the medium, slowly add appropriate amount of hydrochloric acid and sodium hydroxide from the end of the beaker away from the electrode, so that the medium can reach and maintain the required pH.
The temperature compensation of the pH meter should take into account the effect of temperature on the electrode system: in some dilutions and media, the pH changes considerably with temperature. For liquid media and dilutions, the pH can usually be adjusted and measured either at 25°C or at the temperature at which they are used.
The pH of the agar medium must be adjusted after dissolving, and the temperature of dissolving is above 45°C: therefore, the pH of the solid medium should be determined at its use temperature. At this time, a flat-head composite electrode should be used, and the temperature compensation of the pH meter should be manually calibrated. When preparing the culture medium, if it is determined that the pH of the culture medium is different between the melting state and the solidifying state, it means that there is an error in adjusting the pH.
Likewise, when the prepared medium is heat-sterilized (eg, autoclaved), it may also undergo pH changes. For the dehydrated medium, the manufacturer has taken this into consideration, and usually there is no need to adjust the pH, but the pH of the medium to be used should be carefully checked. If the pH of the dehydrated agar medium must be adjusted during redissolution or preparation, it is best to adjust it with filter-sterilized hydrochloric acid and sodium hydroxide solution after the sterilization step to avoid formation of overheating of the medium. When preparing media with different raw materials or pharmaceuticals, the pH of the media must be checked after autoclaving. In addition, if the pH of the medium is found to change the same after each autoclave, the pH can be adjusted appropriately when preparing the medium. However, it is still necessary to determine the final pH of the medium after sterilization.
