Analytical techniques and instrumentation Flashcards
It is a method of determining the chemical composition of sample (specimen) qualitatively and quantitatively
Analytic techniques
It pertains to instrument, system or device used for the measurement of an analyte in a solution.
Analytic techniques
Photometry
analytical techniques
Photoelectric colorimetry
Spectrophotometry
Atomic absorption spectrophotometry (AAS)
Flame emission spectrophotometry (FES)
Infrared spectrophotometry
Mass spectrometry (MS)
Electro chemistry
analytical techniques
Electrophoresis
Ion selective electrode (ISE)
Luminescence spectrometry
analytical techniques
Fluorometry
Chemiluminescence
Flourescence
most commonly used electromagnetic radiation in measuring different analyte in the sample.
visible light
the number of waves produce in a specific time (1 second).
frequency
Can see by the eye, made up of colors.
From 380nm to 780nm.
Visible light (narrow region)
The study of interaction between light and matter.
The study of electromagnetic radiation (light) emitted or absorbed by a chemical in a solution.
All molecules absorbed light.
Spectrometry
Chromatography
analytical techniques
GC-gas chromatography
HPLC- high pressure liquid chromatogaphy
Below 380 nm. Short wavelengths
Harmful to body due to extreme high energy which causes ionization.
Example gamma rays and X rays.
. UV light
Electromagnetic radiation (light) is a combination of electronic and magnetic vibration that travels in a wave-like manner (oscillation).
Electromagnetic radiation
the distance between 2 peaks of wave, measured in nanometer (nm)
wavelength
States that the intensity of transmitted light is inversely proportional to concentration of the solution while absorbed light is directly proportional concentration of the solution.
Beer’s law
Above 780 nm. Long wavelengths
Low frequency waves, not harmful. Non-ionizing
Examples radio waves, microwaves.
Infrared light
Transmitted light or Transmittance (T) can be calculated as follows:
T =I/Io
I= the intensity of light (transmitted light) after it passed through the sample.
Io= the intensity of light (incident light) striking the sample.
Transmitted light is inversely proportional to the concentration of solution.
Absorbed light is directly proportional to the concentration of solution
Beer’s law
Equation: A= abc
A- The absorbance of the solution ( no unit)
a- Molar absorptivity- a measure of how well the molecule absorbs the particular wavelength of radiation
b- path length -the distance travel by light through the solution ( in cm)
c- The concentration of the solution (mol/L)
Absorbance (Abs) is also called
optical density (OD).
The absorbance is directly proportional to the concentration of the solution and the diameter of the test tube or path length.
Beer-Lambert’s Law
States that the amount of absorbance by a color solution is directly proportional to the concentration of the solution and the length of a light path through the solution.
Beer- Lambert’s Law
The intensity of transmitted light decreases as the thickness or path length (diameter of test tube) increases through which the light travels.
Lambert’s law
REMEDIES for beer’s law
use of blank solutions
water blank
reagent blank
serum blank
use of allen correction
Transmitted light is inversely proportional to the path length (the diameter of the tube).
The wider the diameter of the test tube the lower the transmitted light and vice versa.
Lambert’s law
uses only visible light and measures only colored solution
PHOTOELECTRIC COLORIMETRY
White light passes through an appropriate filter which absorbs light of all wavelength except the wavelength denoted by the particular filter.
Light passes through the colored solution (unknown or standard) and this colored solution absorbs some of the light depending on the intensity of the color.
The transmitted light impinges on a photoelectric cell which converts the light energy to electrical energy and this is recorded on a galvanometer as either percent transmittance or absorbance (optical density).
PHOTOELECTRIC COLORIMETRY
A combination of spectrometer and photometer that measures the amount of light absorbed through light transmittance to determine concentration in the solution. It can measures concentration of solution in visible or UV lights.
Spectrophotometer
much like photoelectric colorimetry except the use of monochromator like diffraction grating or quartz prism to disperse white light into a systemic array of its component colors or wavelengths while colorimeter uses filters of specific wavelength.
Spectrophotometer
Light passes through a monochromator and is spread into a spectrum of colors. Slit blocks off all but a narrow band of the light. Sample absorbs some of this light and transmits the rest. The transmitted light strikes a detector which sends a signal to an amplifier. This small signal is amplified and presented as transmittance, absorbance or concentration units by a read-out devices like meters, digital displays, printed read-outs and recorders.
Spectrophotometer
used visible, UV and infrared light for measurement of analyte. It can analyzed both colored and non-colored solution.
Spectrophotometer
Light source
UV
visible light
Infrared light
UV
Hydrogen lamp
Deuterium lamp
Mercury arc lamp
Xenon lamp
Hollow cathode
Visible
Tungsten-iodine lamp
Mercury/sodium vapor lamp
Hollow cathode
Infrared
Merst glower
Globar (silicone carbide)
Basic components of spectrophotometer
Light source
Monochromator (with entrance and exit slits)
Sample cells (cuvettes)
Photodetector
Readout device
made up of parallel groves into aluminized surface of a flat piece of crown glass.
Wavelength is bent when it strikes the diffraction gratings.
Diffraction gratings
commonly used light source in visible and near infrared region
tungsten light bulb
light source used in UV region.
deuterium lamp
isolates the specific wavelength or band of light from the light source.
Monochromator
It has entrance and exit slits which regulates the degree of light the passes through.
Monochromator
Types of monochromator
Glass filter
Prism
Diffraction gratings
less expensive, but not precise. Made up of semi transparent silver film used in photoelectric colorimetry.
Glass filter
it disperse the white light into several spectrum. It is rotated to isolate desired
wavelength.
prism
the most commonly used because it isolates a linear and widely
distributed wavelength. It can operates in visible and UV light.
diffraction gratings
made up of glass, quartz or sodium chloride (wedge shape).
A narrow light focus on a prism is refracted as it enters the more dense glass.
Rotated to a specific wavelength.
prism
concentrate the white light towards the monochromator. Minimizes the entrance of stray light and scattered light into monochromator
entrance slit
concentrates the specific wavelength towards the sample cell. Allows a narrow of light beam to reach the cuvette or sample cell.
exit slit
is the most common cause of loss of linearity at high analyte concentration.
stray light
use to check the accuracy of wavelength setting in spectrophotometer.
holmium oxide
wavelength of
colors
RED
ORANGE
YELLOW
YELLOW GREEN
GREEN
BLUE GREEN
GREEN BLUE
BLUE
VIOLET
651-700
601-650
576-600
556-575
5506-555
496-505
476-495
431-475
350-430
The wavelength at which a substance has its maximum absorbance of light
Lambda max (λmax)
Must be free from scratches to prevent the bouncing of lights that passes through.
Sample cells-
It holds the colored solution or test sample.
Sample cells-