The heart of an efficient HPLC detector is the cell. Figure above shows the schematic of a modern follow-through cell. The optics provide focusing of the light beam in the center of the cell where it is virtually unaffected by the entry-exit-window-interface disturbance, or drift induced by flow, temperature, or refractive index changes. The short, wide cell assures that maximum energy is transmitted, and a post-cell collecting lens focuses all of the existing light from the cell onto the photodetector.
Modern LC systems provide high resolution in a short time. For example, if we are using a column which provide 10,000 theoretical plates and has 15 cm length, then for the component eluted in 2 min (2 ml at 1 ml/min flow rate) we will ideally have peak width of 80 ml. Thus, havin the flow-cell volume of 20 ml we will have only 4 independent measurements on this peak. This is definetely not enough to correctly describe peak shape and will introduce apparent peak broadening.
Another important feature of the flow-cell is to compensate on the refraction effect. When the components of your mixture pass though the cell they actually change (slightly) the eluent composition which will change the refraction coefficient. If the light beam is not parallel the change in the refraction will change light scattering and will contribute into the apparent adsorption readings. Unfortunately very few cell designs has this compensation.
|Apparent Peak broadening happens if the flow cell volume is more then 1/10 of the peak volume, or if the cell geometry is not optimized.|
|Since the peak area is proportional to the amount of the injected analyte, the peak broadening will cause the reduction of the peak height. In case of apparent peak broadening due to the big cell volume but if the cell geometry is optimized we may not see a significant height reduction.|
|The resolution may be decreased as a result of the peak broadening and peak overlapping occurs.|
|If the cell geometry is not optimized one cen notice an apearance of peak tailing. This indicates the presence of flow stagnant zones.|