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INSTRUMENTATION PART 1

LIGHT

Type of electromagnetic radiation (EMR)

Photons of energy/packets of energy travelling in waves

WAVELENGTH ( ) trough to trough, crest to crest

Wavelength = Energy

Wavelength = Energy

E= 1/

=lambda

V = 1/

Frequency
# of cycle/second

Frequency of EM wave = V
-INVERSELY PROPORTIONAL to WAVELENGTH
Energy
- INVERSELY PROPORTIONAL to WAVELENGTH of light
Ultraviolet
-light whose WL < the visible region.
- 190-340nm
Visible
-light whose WL falls at 340-700nm
Infrared
-light whose WL > 700 nm
Wavelength Color absorbed Color emitted
350-400 Violet Yellow
400-450 Indigo Yellow
450-500 Blue Orange
500-550 Green Red
550-600 Yellow Indigo
600-650 Orange Blue
650-700 Red Green
A red solution containing hemoglobin will absorb green light at 540nm.
A yellow solution containing bilirubin will absorb indigo at 400-430nm.

SPECTROPHOTOMETRY-measuremnt of the light transmitted by a solution to determine the


concentration of the light-absorbing substance in the solution.
*darker color = greater concentration

Schematic diagram
LIGHT SOURCE
Light source
TYPE DECRIPTION
Monochromator Tungsten/incandescent Most common LS at visible,
tungsten/ Tungsten-iodide near UV and near IR
Sample holder: CUVETTE lamp/Halogen Quartz lamp
Mercury/ hydrogen Fluorometry
Photodetector lamp/Xenon arc *uneven light distribution
(common only in European
GALVANOMETER spectro)
(read-out device) Hollow cathode lamp AAS (Atomic Absorption
Spectroscopy)
Deuterium At UV range; continuous light
emission down to 165 nm
Merst Glower Uses electrically heated rod of
rare earth element oxides
Globar Uses silicon carbide heated on
1200 degree Celsius
ENTRANCE SLIT
-reduces stray light
-prevents scattered light from entering the
monochromator

MONOCHROMATOR
-device that produces light of specific wavelengths from
a light source
-NOTE: Monochromatic light light radiation of a single
wavelength
Types:
1) Prism
-Wedge-shaped piece of glass pieces of glass, quartz, or
sodium chloride or some other material that allows
transmission of light.

2) Diffraction Gratings
-aluminized surfaces that has been cut into tiny grooves that
can act as a prism and a slit.
3) Interference Gratings
-made by placing silver films on both sides of a dielectric
field (magnesium fluoride)
-when light passes through the silvered surface, it passes
through the dielectric field and is reflected from the second
silvered surface back to the first surface
-enhances desired wavelength by constructive interferences
and eliminates others by destructive interferences
(thickness of magnesium fluoride)

CUVETTE
Types:
1) Borosilicate glass-for alkaline solutions
2) Quartz/Plastic-good for wavelength below 320nm
3) Alumina silica glass-good at visible region
4) Soft glass-preferable for acidic solutions

PHOTODETECTORS
-converts transmitted light energy into an equivalent
amount of electrical energy
Types:
1) Barrier-layer cell
2) Photoemissive/Phototube
3) Photomultiplier
Barrier-Layer Cell (BLC)
-least expensive
-composed of light-sensitive materials
-when exposed to light, electrons are excited and are
released into the highly conductive silver
-no need for power source
Photoemissive/Phototube
-has photosensitive material
-composed of : 1 (+) charged anode
2 (-) charged cathode enclosed in a
glass
-cathode emits electron when exposed to light. The
emitted electrons jump over to the positively
charged anode where they are collected and return
through an external measurable circuit
-requires an external source of energy