Biological processes buffered systems

Since biological systems are dynamic, with many different processes taking place and many different substances present, buffers are necessary to prevent the kind of wide variation of pH that can inhibit proper enzyme catalysis. Thus, a proper pH aids in regulating the reaction rates associated with certain enzymes and maintaining them at levels appropriate for their particular functions. Two important biological buffers are the phosphate buffer system that regulates pH for the fluid inside cells and the carbonic acid buffer system that regulates pH for blood plasma. The chemical equations for these buffers are shown below for an aqueous solution. 

Buffer systems are particularly important in biological systems. Blood and other body fluids are buffer solutions which prevent small additions of acids and bases from causing drastic changes in the pH. The functioning of the enzymes—the biological catalysts—is very sensitive to pH changes, and if biological systems were not buffered there would be serious variations in the rates of metabolic processes. 

Recent studies suggest that many factors may affect hydroxyl radical generation by microsomes. Reinke et al. [34] demonstrated that the hydroxyl radical-mediated oxidation of ethanol in rat liver microsomes depended on phosphate or Tris buffer. Cytochrome bs can also participate in the microsomal production of hydroxyl radicals catalyzed by NADH-cytochrome bs reductase [35,36]. Considering the numerous demonstrations of hydroxyl radical formation in microsomes, it becomes obvious that this is not a genuine enzymatic process because it depends on the presence or absence of free iron. Consequently, in vitro experiments in buffers containing iron ions can significantly differ from real biological systems. 

For the determination of CCA in biological samples, methods not based on LC-MS/MS technology [39, 41-43] and methods that used LC-MS/MS [40, 52] have been reported. Most of the sample extraction methods used liquid-liquid extraction (LLE) technology, since this extraction method is simpler and able to minimize matrix effects. Consequently, LLE methods are considered to provide cleaner samples as compared to solid phase extraction (SPE) methods. Since LC-MS/MS methodology uses nonvolatile solvents or a combination of nonvolatile and volatile solvents, difficulties in the evaporation process and associated interferences when samples are injected onto the system can arise [51]. However, Bahrami as well as Souri [42,43] applied a combination of nonvolatile and volatile solvents in which the nonvolatile solvents were acidic buffers (pH 5 or less). Analytes eluted from SPE prepared samples did not undergo evaporation as applied commonly encountered in extraction procedures [37, 45]. 

---