Waters 2998, Обзор и руководство по обслуживанию

Страница: 16 / 94

December 5, 2017, 715004753 Rev. B
Page 16
1.1.1 Calculating absorbance
The detector computes absorbance by subtracting the dark current (see “Dark current” on
page 23 ) from the reference spectrum (reference energy) and the acquired spectrum (sample
energy). Absorbance is based on the principles of Beer’s law.
1.1.1.1 Beer’s law
The Beer-Lambert law (commonly called Beer’s law) describes the relationship between the
quantity of light of a particular wavelength arriving at the photodiode and the concentration of the
sample passing through the flow cell. Beer’s law is expressed as
A
=

lc
where
A
= dimensionless quantity measured in absorbance units

= constant of proportionality known as the molar absorptivity
l
= path length in centimeters (1.0 cm in the detector’s normal flow cell)
c
= concentration in moles per liter
Beer’s law applies only to well-equilibrated dilute solutions. It assumes that the refractive index of
the sample remains constant, that the light is monochromatic, and that no stray light reaches the
detector element. As concentration increases, the chemical and instrumental requirements of
Beer’s law are sometimes violated, resulting in a deviation from (absorbance versus
concentration) linearity. The absorbance of mobile phase can reduce the linear range.
Table 1–1: Optics assembly components
Component Function
Lamp Deuterium source lamp.
M1 mirror Focuses light from the deuterium source lamp.
Window Used to help minimize air infiltration into the lamp housing.
Filter flag/shutter Flag positions for measuring open (sample) and blocked (dark) beam
energies and a third for wavelength verification.
Flow cell Houses the segment of the flow path (containing eluent and sample)
through which the polychromatic light beam passes.
Spectrograph mirror and
mask The mirror focuses light transmitted through the flow cell onto the slit
at the entrance to the spectrographic portion of the optics. The mirror
mask defines the size of the beam at the grating.
Slit Determines wavelength resolution and intensity of light striking the
photodiodes. The width of the slit is 50 µm.
Grating Disperses light into bands of wavelengths and focuses them onto the
plane of the photodiode array.
Order filter Reduces the contribution of second-order diffraction of UV light (less
than 370 nm) to the light intensity observed at visible wavelengths
(greater than 370).
Photodiode array A linear array of 512 photodiodes. The diode width (50 µm), together
with a 50-µm slit, yields single wavelength resolution of 1.2 nm.