Lec 3 Flashcards
measures light and is applied to spectrophotometry
Colorimetry
Law of colorimetry
Involves measurement of the amount of light absorbed by the solution, and that absorption is related to concentration.
2 primary considerations in colorimetric analysis
• quality of light
• intensity of light
A kind of colorimetry that uses the eye in determining the end product/point.
Visual Colorimetry
Duboscq Colorimeter follows _____ Law.
Beer’s Law
A = C x L x K
Beer’s Law formula
The solution’s absorbance is directly proportional to the solution’s concentration
Beer’s Law
The solution’s absorbance is inversely proportional to the transmittance
Beer’s Law
A refined type of visual colorimeter. It is used to determine the concentration of UNKNOWN GLUCOSE solutions
Duboscq Colorimeter
The concentration of the absorbance is equal to the concentration of the substance times the depth
Beer’s Law
Spectrophotometry
Measures light intensity using a device (prism/grating) to disperse the source of light into a continuous spectrum.
Measures light intensity of multiple wavelength using a filter to isolate part of the spectrum
Filter Photometry
Entrance slit is also known as _____
Collimator lens
It narrows down the rays of light coming from the light source
Entrance slit
This separates the white light into different colors, and only one of those colors is allowed to pass through.
Monochromator
It provides radiant energy in the form of visible light that passes through the monochromator.
Light Source
8 types of light source
- Tungsten lamp
- hydrogen lamp
- quartz halide lamp
- deuterium lamp
- Mercury vapor/arc lamp
- Hollow cathode lamp
- LED (light emitting diodes)
- Laser
This light source provided energy wavelength from 340 to 780 nm and wavelength is visible.
Tungsten Lamp
Tungsten lamp that is for visible and UV
Tungsten Iodine Lamp
Tungsten Lamp which is the filament of the lamp.
Tungsten Halide Filaments
TRUE or FALSE:
Tungsten Halide Filaments
- long lasting
- less light at shorter wavelength
- emits a higher intensity radiant energy
FALSE
- MORE light at shorter wavelength
A light source with a wavelength less than 300 nm
Deuterium Lamp
This part of the spectrophotometer minimizes stray light from the light source by preventing scattering of light.
Entrance Slit
Monochromator
A wavelength selector which isolate specific wavelength of light by the use of a prism/grating or both.
4 types of monochromator
Prism, Diffraction grating, Colored filter, Interference filter
A type of monochromator that is a triangular wedge-shaped piece. Made of either glass, quartz, NaCl or KBr.
Prism Monochromator
A prism monochromator for visible spectrum
Glass prism
A prism monochromator used in UV
Quartz / Fused silica prism
A prism monochromator for infrared spectrum
Prism made of NaCl or KBr
A monochromator that has small grooves cut into it. May have >3000 grooves per millimeter
Diffraction Grating
Colored Filter
A type of monochromator that allows a wide range of wavelengths to pass through wide bandwidths of 20-60nm. Less expensive but less sensitive
A type of monochromator that is used to obtain special purity.
Interference Filter
General methods under the classifications of Clinical Chem instrumentation that measure EMR
Spectrophotometry,
Electrophoresis,
Immunoassay,
Nephelometry,
Ion-selective electrodes
Basic Categories of Analytical Chem
Spectrometry,
Luminescence,
Electroanalyctical methods,
Chromatography
The measurement of radiant energy that is absorbed or given off by a molecule after illumination of light source.
Photometry
The radiant energy that includes short gamma rays to long radio waves. They are photons of energy traveling in a wave-like manner.
EMR
Types of EMR (in order)
Cosmic rays,
Gamma rays,
X-rays,
UV rays,
Visible rays,
Infrared,
Radio waves
It is a photon or discrete packet of energy traveling in waves.
Light
Wavelength of light
The distance between 2 successive peaks.
Value of h (Planck’s constant)
6.62 x 10^(-27)
Refers to the wavelength interval in which the lowest type of radiant energy occurs
Planck’s constant
A term used to define a light wave. It is the distance between trough & peak
Amplitude
Nanometer (nm)
• 10^(-9)
• the standard international unit for expressing wavelength of light
• old term for wavelength
• 1 mu = 1 nm
Millimicron (mu)
Angstrom (A)
• smaller compared to Nanometer and Millimicron.
• 10^(-10)
A kind of wavelength with approximately 390 to 750 nm
Visible spectrum/spectra/light/ray
2 rays under Invisible Spectra
UV rays, Infrared ray
A kind of invisible spectrum with a wavelength of 190 to 390 nm
UV rays
A kind if invisible spectrum with a wavelength of more than 800 nm
Infrared rays
Delivers the light desired wavelength by adjusting the wavelength adjustment knob
Exit slit
A certain range of band of wavelengths are allowed to pass into the sample
Bandpass/Bandwidth
An absorption cell used to hold a solution inside the instrument. Absorption is measured.
Analytical Cell / Cuvette / Sample Holder
A kind of cuvette used for solutions that are alkaline and do not etch the glass.
Soft glass cuvette
A kind of cuvette suitable for visible portion of light
Borosilicate glass cuvette
A kind of cuvette that does NOT absorb UV radiation, so it is used to measure UV radiation (below 320 nm)
Quartz / Plastic cuvette
An electron tube amplifying current that can convert transmitted radiant energy into an equal amount of Photoelectric energy
Detector
A type of detector that is less expensive, durable, and temperature sensitive.
Barrier-layer cell / Photocell
A type of detector that gives off electrons when light energy strikes. It requires an outside voltage, thus it’s expensive.
Photoemission Tube
A type of detector that detects and amplifies radiant energy from the solution to the detector, wherein it is already converted to electrical energy.
Photomultiplier
A type of detector which produces a photocurrent that is proportional to the incident radiant energy.
Photodiode
The simplest method of displaying the output of the detection system.
Data read-out device
It is a reference solution when electrical read out of the instrument is arbitrarily at 100%T or 0.0 A
Blanking / Blank (BLK)
4 types of BLK
Distilled water blank,
Reagent blank,
Air blank,
Serum / Sample blank
A type of BLK which contains only distilled H2O. It is used to set Zero Absorbance (0.0 A)
Distilled water blank
A type of BLK that contains all the components of the chromogenic reaction EXCEPT the sample and its analyte.
Reagent Blank
A type of BLK that contains all the reagent and the sample. EXCEPT for the last reagent that will give color to the reaction.
Sample BLK
A measure of the amount of light that passes through the solution
Transmittance
A proportion of light that penetrates the solution
Transmittance
The amount of light being absorbed or stopped by a solution
Absorbance
Formula used in the Spectrophotometer
Cu = Au x Cs / As
5 Calibration procedures for Quality Assurance of Spectrophotometry.
Wavelength calibration,
Absorbance/Photometric Accuracy checks,
Linearity check,
Electronic Stability,
Stray light check
The 3 standards for Photometric Accuracy Checks
Potassium dichromate sol,
Cobalt ammonium sulfate sol,
Potassium nitrate sol
Any wavelength of light outside the band transmitted and selected by the monochromator
Stray light
