Final answer:
Binding detection in equilibrium dialysis involves measuring drug concentrations post-equilibrium; the difference aids in calculating drug-protein binding. The unbound drug is quantified via LC-MS/MS, whereas fluorescent quenching monitors binding to proteins like albumin.
Step-by-step explanation:
In equilibrium dialysis, binding can be detected by measuring the concentration of a drug in both the plasma compartment and the buffer compartment after equilibrium has been reached across a semi-permeable membrane. The total drug concentration is determined within the plasma compartment, while the free drug concentration is measured within the buffer compartment. Using this data, the percentage of drug bound to plasma proteins can be calculated. To accurately quantify the drugs, especially due to low levels of free drug in the buffer, LC-MS/MS methods are applied. Sample preparation typically includes protein precipitation followed by centrifugation before analysis by LC-MS/MS.
For drugs like levofloxacin, the binding to albumin can be monitored via a decrease in fluorescence, known as fluorescence quenching. This method helps in determining the binding affinity, which is crucial for understanding the pharmacodynamics and pharmacokinetics of a drug.