Spectrophotometry Chapter 2

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Questions and Answers

What is the primary function of a monochromator?

To filter out specific wavelengths of light

To amplify light intensity

To convert monochromatic light to polychromatic light

To separate polychromatic light into individual wavelengths (correct)

Which component of a monochromator creates a parallel beam after the entrance slit?

Camera mirror

Diffraction grating

Collimating mirror (correct)

Exit slit

Which of the following is NOT one of the basic components of a monochromator?

Exit slit

Diffraction grating

Entrance slit

Prism (correct)

What is the role of the diffraction grating in a monochromator?

To disperse light into its individual wavelengths Signup and view all the answers

How many main types of monochromators are there?

Two Signup and view all the answers

What is the function of a cuvette in the given setup?

Hold liquid analyte Signup and view all the answers

Which type of lamp provides power in the 320 to 800 nm range?

Tungsten filaments lamp Signup and view all the answers

What is the main function of a photodetector in the system?

Amplification of the signal Signup and view all the answers

Which of the following is a type of wavelength selector?

Monochromators Signup and view all the answers

What is one of the issues associated with using a tungsten filaments lamp?

Condensation on the glass envelope Signup and view all the answers

What is the role of a microprocessor in the given system setup?

Calculate concentration Signup and view all the answers

What does the Beer-Lambert law relate to?

The attenuation of light through a material Signup and view all the answers

What symbol is used to denote the intensity of light passing through the reference cell?

I0 Signup and view all the answers

If the intensity I is less than I0, what does it indicate?

The sample has absorbed some of the light Signup and view all the answers

Which value in the Beer-Lambert law equation represents the absorptivity?

a Signup and view all the answers

According to the Beer-Lambert law, what is absorbance (A) proportional to?

a, L, and C Signup and view all the answers

Which equation correctly represents the relationship between absorbance (A) and transmittance (%T)?

A = log(100/%T) Signup and view all the answers

What is a major advantage of using a grating over a prism in spectroscopy?

Higher efficiency Signup and view all the answers

Which of the following best describes a cuvette's purpose?

Holds the sample being analyzed Signup and view all the answers

Which law combines the principles of Lambert, Bouguer, Bunsen, Roscoe, and Beer?

Beer's Law Signup and view all the answers

According to Beer-Lambert law, what is the relationship between the concentration of a solution and its absorbance?

Linear Signup and view all the answers

What elements are involved in a grating spectrometer?

Source, grating, detector Signup and view all the answers

What happens to the intensity of light as it passes through a sample with higher absorbance?

It decreases Signup and view all the answers

Which factor does not contribute to the efficiency of gratings over prisms?

Absorption effects Signup and view all the answers

What is the result of the interaction between a patient specimen and reagent in a sample?

Absorption of light selectively Signup and view all the answers

What is the main purpose of wavelength selectors?

To select portions of the power spectrum produced by the power source Signup and view all the answers

Which of the following filters absorb in the higher wavelength visible range?

Blue color filters Signup and view all the answers

What is the primary function of a collimator lens in a spectrometer?

To make sure all light rays are parallel to each other Signup and view all the answers

Which element of a spectrometer is responsible for separating light based on its wavelengths?

Diffraction grating or prism Signup and view all the answers

What type of filter is designed to be low-pass, high-pass, or band-pass?

Glass filters Signup and view all the answers

Which component of a spectrometer focuses the spectrum onto a detector?

Camera lens Signup and view all the answers

What is the function of the exit slit in a spectrometer?

Select a narrow band of wavelengths Signup and view all the answers

Interference filters are named after which aspect?

Designations based on the principle of operation Signup and view all the answers

What component of a UV-visible spectrophotometer selects a specific wavelength of light?

Monochromator Signup and view all the answers

Which light source is commonly used in the visible region (400-800 nm) for spectrophotometry?

Tungsten halogen lamp Signup and view all the answers

Which component in the UV-visible spectrophotometer holds the sample being analyzed?

Cuvette Signup and view all the answers

Which of the following ranges is covered by the ultraviolet light in spectrophotometry?

200-400 nm Signup and view all the answers

What is the function of the detector in the spectrophotometer?

Measures the amount of light that passes through the sample Signup and view all the answers

Which is NOT a component of the basic UV-visible spectrophotometer?

Fiber Signup and view all the answers

Why are LEDs becoming popular in spectrophotometry?

They are energy-efficient and have a long lifespan Signup and view all the answers

In which application area of spectrophotometry would one study the rate of a chemical reaction?

Kinetics Signup and view all the answers

What role does the monochromator play in a UV-visible spectrophotometer?

Selects a specific wavelength of light Signup and view all the answers

Which substance is most likely to emit light in the infrared region, often used in spectrophotometers?

Hydrogen lamp Signup and view all the answers

According to the Beer-Lambert Law, which variable represents the molar absorptivity?

ε Signup and view all the answers

What happens to absorbance if the concentration of the solution is doubled?

Absorbance is doubled Signup and view all the answers

Which of the following describes a major assumption of Beer-Lambert Law?

Absorbance is directly proportional to concentration Signup and view all the answers

What does an absorbance of 0 at a specific wavelength indicate?

No light of that particular wavelength has been absorbed Signup and view all the answers

Why is it important to correct absorbance measurements for the concentration of the sample?

To avoid meaningless absorbance measurements Signup and view all the answers

Which of the following is NOT a limitation of spectrophotometry?

High sensitivity Signup and view all the answers

In spectrophotometry, what would likely happen if a solution is too dilute?

Absorbance will be very low Signup and view all the answers

Which research application is commonly associated with spectrophotometry?

Protein analysis Signup and view all the answers

What is the unit of molar absorptivity?

L mol-1 cm-1 Signup and view all the answers

In Beer's Law, which variable represents concentration?

c Signup and view all the answers

If the path length is doubled and the absorbance is kept constant, what happens to the concentration?

It halves Signup and view all the answers

Which of the following is a typical application of spectrophotometry?

Measurement of nucleic acids Signup and view all the answers

What was the concentration of guanosine found in the example calculation?

8.33 x 10-5 mol/L Signup and view all the answers

Which of the following is not a limitation of spectrophotometry?

It has high sensitivity and selectivity Signup and view all the answers

What is Beer's Law equation?

A = εlc Signup and view all the answers

What factor influences the proportion of light absorbed by a solution?

The number of molecules it interacts with Signup and view all the answers

Study Notes

Definition of Spectrophotometry

A scientific method that measures the absorption of light by a substance at different wavelengths.
Wavelengths range from ultraviolet (200-400 nm), visible (400-700 nm), or near infrared (700-800 nm).

Basic Principle of Spectrophotometry

Measures the concentration of solutes in a solution by measuring the amount of light absorbed.
Uses a cuvette placed in a spectrophotometer.

Basic Components of UV-Visible Spectrophotometer

Light source: Emits light in the desired wavelength range (e.g., deuterium lamp for UV region, tungsten halogen lamp for visible region).
Monochromator: Selects a specific wavelength of light (e.g., filter, prism, gratings).
Entrance slit: Determines the width of the light beam.
Dispersive element: Separates the light into different wavelengths.
Exit slit: Allows only the selected wavelength to pass through.
Sample cell (cuvette): Contains the sample being analyzed (made of glass, plastic, or quartz).
Detector: Measures the amount of light that passes through the sample.

Components of UV-Visible Spectrophotometer

Source: Provides a beam of light.
Filter (optional): Selects a specific wavelength range.
Fiber: Guides the light from the source to the cuvette.
Cuvette: Holds the sample being analyzed.
Photodetector: Detects the light that passes through the sample.
Readout: Displays the data.
Microprocessor: Processes the data.
Recorder: Records the data.

Sources of Light

Hydrogen lamp: Emits light in the infrared region and is often used in spectrophotometers that measure absorbance in the ultraviolet region.
Tungsten filament lamp: Produces visible light and is used in spectrophotometers that measure absorbance in the visible region.
LED: Energy-efficient and has a long lifespan.

Applications of Spectrophotometry

Quantitative analysis: Determines the concentration of a substance in a solution.
Qualitative analysis: Identifies the presence of a substance in a solution.
Kinetics: Studies the rate of a chemical reaction.
Biochemistry: Studies the properties of proteins, enzymes, and other biological molecules.
Environmental monitoring: Measures the levels of pollutants in the environment.

Component Functions

Source of light: Electromagnetic waves.
Filter/prism: Spectral isolation.
Fiber optic: Directional control of the beam.
Cuvette: Holds the liquid analyte.
Photodetector: Amplification of the signal.
Readout device: Digital reading.
Microprocessor: Calculates the concentration.
Recorder: Gives the result.

Wavelength Selectors

Monochromators: Devices that separate light based on its wavelengths.
Filters: Glass or interference filters that absorb power for specific wavelengths.

Monochromators

Prisms: Refraction of sunlight.
Grating spectrometer: Source, grating, mirrors, detector.

Cuvettes

Definition: A small rectangle, round, or square vessel.
Function: Holds the substance being analyzed and maintains the optical characteristic.

Samples

A sample substance results from the interaction between a patient specimen and reagent.
Absorbs light selectively according to the law of Lambert, Bouguer, Bunsen, Roscoe, and Beer's Law.

Beer's Law

States that there is a linear relationship between the concentration of a solution and the absorbance of the solution.
Enables the concentration of a solution to be calculated by measuring its absorbance.

Absorbance of a Sample

The intensity of the light passing through the sample is referred to as the absorbance.
The intensity of light passing through the sample is dependent on the sample's absorbance.
Reflected in the cuvette surface.

The Importance of Concentration

The proportion of the light absorbed depends on how strongly it interacts with the sample.
Concentrated solutions have higher absorbance, while dilute solutions have lower absorbance.

Limitations of Spectrophotometry

Restriction to the solution phase only.
Cost: Spectrophotometers can be expensive.
Low sensitivity and selectivity.

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Description

Learn about the scientific method of spectrophotometry, which measures the absorption of light by substances at different wavelengths. This technique is used to determine the concentration of solutes in a solution.