Cam Sep Lect 1 Chromatografia Bronislaw K. Glod

Photometric Detection
According to the Lambert–Beer law, the background absorbance, A b
, of the acid, HB, used
as buffer can be expressed as:
A b
= lε B–
[B – ] + lε HB
[HB] (9)
where l denotes the length of detector cell and ε B–
and ε HB
are, respectively, the molar absorption
coefficients of the dissociated and undissociated forms of acid. Absorption at the peak
maximum, on the basis of eqs (1) and (9) is described by:
A max = lε B–
[B – ] max + lε HB
[HB] max (10)
The height of the chromatographic peak can be obtained from eqs (9) and (10):
ΔA = ε B–
[B – ] max + lε HB
[HB] max – lε B–
[B – ] – lε HB
[HB] (11)
By combining eq. (11) with eq. (3), using the definition of the dissociation constant, and assuming
that the concentration of the undissociated form of the solute acid is constant throughout the
chromatographic peak
([HB] max = [HB]), we obtain:
ΔA = –lε B–
[R – ] (12)