Pathlength normalization

Beer s law, on the other hand, is based on the assumption that there is a single pathlength (normally represented by the variable b in the equation A = abc) for all rays through the sample. In a real, physical, measurement system, this assumption is always false, because of the fact that dew cannot be zero. As Figure 29-1 shows, the actual... 

The absorption of the sample will depend on the pathlength of the cell, /, and the concentration of the sample, c, just as it does in normal absorption spectroscopy. According to Beer s Law,... 

In conclusion, one remark. It Is evident Chat Che calculations describing the ideal case are rather far from reality. Further, it should be noticed, that it is absolutely incorrect, to take as a measure of surface concentration the AES signals (eventually normalized) Chat is to put N] = 1. Even for Che signals most sensitive for the surface, N] - 0.5 It is equally incorrect to say (what is very popular in the literature) that the AES signals characterize the average concentration over the free pathlength X of the electrons monitored the contribution of the deeper layer decreases exponentially and not linearly with the distances from the surface (8). 

The last subject in the discussion of inherently chiral compounds deals with the analysis of the aminoacids, peptides, and proteins. Most all of the remarks that were made about the steroids and carbohydrates regarding CD detection apply equally well to these. The enantiomeric purity of aminoacids is usually determined by their optical rotations at the sodium-D line. Rotations are normally so small that concentrated solutions and long pathlengths are needed. The detection is enhanced a little if laser illumination is used [66] or if ORD detection is done around 230 nm [71]. Without derivatization, only aminoacids with aromatic substituents are CD active in the near UV. Signals are generally weak and enantiomeric purity measurements are not quantitative. 

Steady-State Spectral Convolution. The steady state absorption and emission spectra of dilute dye samples can be measured using standard spectroscopic techniques. Once the extinction coefficient, e( ), and the normalized luminescence spectrum, f(v), are known for a particular dye, the self—absorption probability r over a pathlength L in the sample containing the dye at a concentration C is given by... 

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