Buffers: Principles and Practice

Biochemical processes can be severely affected by minute changes in hydrogen ion concentrations. At the same time, many protons may be consumed or released during an enzymatic reaction. It has become increasingly important to find buffers to stabilize hydrogen ion concentrations while not interfering with the function of the enzyme being examined. The quality of a buffer is dependent on its buffering capacity and its ability to maintain a stable pH upon dilution or addition of neutral salts. There are many factors that must be considered when choosing a buffer. When studying an enzyme, the pH optimum of the enzyme, nonspecific buffer effects on the enzyme, and interactions with substrates or metals must be considered. When purifying a protein, cost becomes an important consideration, as does the compatibility of the buffer with different purification techniques. The good buffers have been shown to be relatively free of side effects. However, inorganic buffers do have a high potential for specific buffer effects. Many enzymes are inhibited by phosphate buffer, including carboxypeptidase, urease, as well as many kinases and dehydrogenases.