Basic compounds overloading

Band broadening and/or unusual retention are also known to occur when specific interactions take place between particular solutes and the stationary phase. The most common cause of such band broadening is a stationary phase overload. It can be also an exclusion phenomenon due to the small size of the stationary phase pores and the large size of the solute. Acid-base interactions between the surface silanols and basic compounds, especially the amine compounds, dramatically broaden the corresponding peaks. Peak deformations, either frontings or tailings, are observed. 

The form of the isotherm need not be Langmuir in nature, but in any event, must be experimentally determined in order to identify the true profile of the overloaded peak. In practice, the determination of the adsorption isotherm of each compound to be separated by a preparative chromatographic procedure can be arduous and time consuming. A better alternative might be to design the fully optimized column from basic principles in the manner previously described. 

Direct, on-line coupling of an NMR spectrometer to an HPLC has required the development of special interfaces called flow probes. These systems can basically work in two ways. One is through an on-flow mode (the solute passes through the system as it passes out of the column and is analyzed), but this process has a low sensitivity even when the HPLC column is overloaded. In another way, which is much more used nowadays, the HPLC control unit is equipped with a valve that invokes a stop flow of the effluent when a compound (detected by a UV detector that is always used in connection with these systems) leaves the column in this case, the flow is stopped in order to have enough time for the acquisition of the NMR spectrum. 

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