Introduction to Spectrophotometry
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Questions and Answers

What is the primary function of UV-Vis spectroscopy?

  • To measure the amount of light absorbed by or transmitted through a sample (correct)
  • To analyze the thermal properties of materials
  • To determine the density of a sample
  • To measure the amount of light refracted by a sample

Which application is NOT typically associated with UV-Vis spectroscopy?

  • Quality assurance
  • Pharmaceutical analysis
  • DNA & RNA analysis
  • Thermal conductivity measurement (correct)

For how long has UV-Vis spectrophotometry been in general use?

  • For the last 20 years
  • For the last 50 years
  • For the last 80 years (correct)
  • For the last 100 years

What is a common purpose of using a blank sample in UV-Vis spectroscopy?

<p>To compare the absorbance of the sample (D)</p> Signup and view all the answers

Which of the following is NOT a type of analysis conducted using UV-Vis spectroscopy?

<p>Acoustic wave measurement (C)</p> Signup and view all the answers

What is the role of a chromophore in a molecule?

<p>It is responsible for light absorption. (B)</p> Signup and view all the answers

What occurs to a molecule when it absorbs light?

<p>The energy of the molecule increases. (D)</p> Signup and view all the answers

How is the observed color of a substance determined?

<p>By the color that is absorbed by the substance. (B)</p> Signup and view all the answers

Which type of radiation is mentioned as promoting electrons to higher energy orbitals?

<p>Visible and ultraviolet radiation (A)</p> Signup and view all the answers

What happens to a molecule when it emits light?

<p>Its energy level decreases. (D)</p> Signup and view all the answers

What is the primary purpose of spectrophotometry?

<p>To measure chemical concentrations using light (D)</p> Signup and view all the answers

What are the components of light waves in spectrophotometry?

<p>Electric and magnetic fields that oscillate perpendicular to each other (A)</p> Signup and view all the answers

What does the symbol $ u$ represent in the context of light waves?

<p>Frequency of oscillations (D)</p> Signup and view all the answers

What is the unit of frequency in spectrophotometry?

<p>Hertz (D)</p> Signup and view all the answers

Which range of wavelengths defines the visible spectrum?

<p>380 to 780 nanometers (B)</p> Signup and view all the answers

Which of the following best describes the electromagnetic spectrum in relation to spectrophotometry?

<p>It represents molecular processes occurring when different light regions are absorbed (D)</p> Signup and view all the answers

What materials are commonly used for cuvettes in spectrophotometry?

<p>Glass or quartz (C)</p> Signup and view all the answers

Which statement regarding the properties of light is true?

<p>Light can behave both as a particle and a wave (C)</p> Signup and view all the answers

What is the relationship between the energy increase of a molecule and the wavelength of ultraviolet radiation absorbed?

<p>Shorter wavelengths correspond to higher energy. (D)</p> Signup and view all the answers

What does transmittance (T) quantify in the context of UV-Vis Spectrophotometry?

<p>The fraction of original light that passes through the sample. (D)</p> Signup and view all the answers

In Beer-Lambert’s Law, which variable represents the concentration of the sample?

<p>c (C)</p> Signup and view all the answers

How does the quality of a monochromator affect the light beam in a UV-Vis spectrophotometer?

<p>It narrows the range of wavelengths in the emerging beam. (B)</p> Signup and view all the answers

Which of the following factors influences absorbance in a sample according to Beer-Lambert’s Law?

<p>Molar absorptivity (A)</p> Signup and view all the answers

What is the unit of measurement for molar absorptivity in Beer-Lambert's Law?

<p>M^-1 cm^-1 (A)</p> Signup and view all the answers

When light is absorbed by a sample in a UV-Vis spectrophotometer, what happens to the irradiance of the beam?

<p>It decreases. (B)</p> Signup and view all the answers

If a 0.00240 M solution has a molar absorptivity of 313 M^-1 cm^-1 in a 2.00-cm cell, which mathematical relationship would you use to find the absorbance?

<p>Absorbance = c × ε × b (B)</p> Signup and view all the answers

Which of the following is a limitation of Beer’s Law?

<p>It is not valid for solutions with high concentrations of electrolytes. (C)</p> Signup and view all the answers

What characterizes a standard calibration curve?

<p>It shows the relationship between an analytical signal and known concentrations. (B)</p> Signup and view all the answers

What are standard solutions used for in analytical procedures?

<p>To establish a correlation between concentration and analytical signal. (D)</p> Signup and view all the answers

Which statement regarding blank solutions is true?

<p>They assist in measuring the effects of impurities or interference. (C)</p> Signup and view all the answers

What effect does high absorbance have on light measurement in samples?

<p>It limits the effective measurement of light intensity. (A)</p> Signup and view all the answers

What is a common characteristic of solutions that are too dark in color related to Beer’s Law?

<p>They result in low absorbance causing indistinguishable readings. (C)</p> Signup and view all the answers

Which of the following factors is critical for ensuring accurate readings in spectrophotometric measurements?

<p>Using compartments that are tightly closed to prevent stray light. (B)</p> Signup and view all the answers

How does scattering of light due to particulates in a sample affect analytical readings?

<p>It causes variations that may lead to inaccurate absorbance readings. (D)</p> Signup and view all the answers

Flashcards

Beer's Law Limitations

Beer's Law, relating absorbance to concentration, doesn't hold true for high electrolyte/analyte concentration, non-dilute solutions, or samples with particulates. Stray light and high/low absorbance levels can also affect accuracy.

Nonlinear Calibration Curve

A calibration curve isn't a straight line if the Beer's law limitations are met.

Standard Calibration Curve

A graph showing the relationship between the concentration of an analyte and its response in an analytical method.

Standard Solutions

Solutions containing known concentrations of the substance being measured.

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Blank Solutions

Solutions with all the reagents and solvents EXCEPT the substance being studied, used to account for any interference from impurities in the process.

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Calibration

Determining the relationship between a measured result and a known standard.

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Dilute Solutions

Solutions with low concentrations of solute.

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Stray Light

Light that reaches the detector that isn't from the sample or reference beam, leading to inaccurate measurements.

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UV-Vis Spectroscopy

A technique that measures the amount of UV or visible light absorbed or transmitted by a sample.

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UV-Vis Spectrophotometer

An analytical instrument used in labs measuring light absorption.

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Applications of UV-Vis Spectroscopy

Used in DNA/RNA analysis, pharmaceuticals, bacterial cultures, beverages, and quality assurance.

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Light Absorption by a Sample

The phenomenon that different specific wavelengths of light are absorbed by a sample, making them useful for analysis.

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Analytical Instrument

A tool that is commonly used to determine the chemical composition of a substance.

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Chromophore

The part of a molecule that absorbs light.

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Absorption of Light

Increases a molecule's energy.

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Emission of Light

Decreases a molecule's energy.

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Color of Substance

The complement of the absorbed wavelengths of light.

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UV/Vis Radiation

Promotes electrons to higher energy levels.

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Spectrophotometer

An instrument used to measure the absorbance and transmittance of light through a substance.

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Cuvettes

Small containers used to hold samples for spectrophotometry.

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Spectrophotometry

Technique using light to measure chemical concentrations.

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Wavelength (λ)

The distance between two corresponding points of a wave.

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Frequency (ν)

Number of oscillations per second of a wave.

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Electromagnetic Spectrum

Range of all types of electromagnetic radiation.

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Visible Spectrum

Part of the electromagnetic spectrum humans can see (380-780 nm).

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Molecular Processes

Reactions or changes within molecules induced by absorption of different wavelengths.

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UV radiation energy increase

The increase in energy (in kilojoules per mole) of an O2 molecule when it absorbs ultraviolet radiation of a specific wavelength.

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UV-Vis Spectrophotometer

A device that measures how much light a substance absorbs.

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Monochromatic light

Light consisting of a single wavelength (color).

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Transmittance (T)

The fraction of initial light that passes through a sample.

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Absorbance

A measure of light absorbed by a sample.

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Beer-Lambert's Law

A law relating absorbance to concentration and path length.

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Molar absorptivity (ε)

A constant that describes how strongly a substance absorbs light at a given wavelength.

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Path Length (b)

The distance light travels through the sample.

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Study Notes

Introduction to Spectrophotometry

  • Spectrophotometry is a technique used to measure chemical concentrations using light.
  • Light is both a particle and a wave, consisting of oscillating electric and magnetic fields.
  • UV-Vis spectroscopy measures the amount of UV or visible light absorbed or transmitted through a sample compared to a reference.

UV-Vis Spectroscopy

  • UV-Vis spectroscopy is an analytical technique.

  • It measures the amount of discrete UV or visible light wavelengths absorbed or transmitted through a sample.

  • Applications include DNA/RNA analysis, pharmaceutical analysis, bacterial culture, beverage analysis, and quality assurance.

  • This technique has been in use for over 80 years and is an important analytical tool.

UV-Vis Spectrophotometer

  • A UV-Vis spectrophotometer measures the amount of light absorbed or transmitted by a sample across different wavelengths typically in the ultraviolet and visible light spectrum range.

  • It is an essential instrument in modern laboratories.

Cuvettes

  • Cuvettes are small containers used to hold the sample.
  • They are usually made of quartz or glass.

Electromagnetic Spectrum

  • Light is part of the electromagnetic spectrum.

  • The visible region of the spectrum spans the wavelength range from 380 to 780 nanometers.

  • Different wavelengths of light in the electromagnetic spectrum result in different molecular processes that can be detected.

Spectrophotometry Principles

  • The interaction between light and matter is crucial to spectrophotometry.
  • Light absorption is related to molecular properties.
  • Light energy is directly proportional to frequency and inversely proportional to wavelength.

Beer-Lambert's Law

  • Absorbance (A) is directly proportional to concentration (c), pathlength (b), and molar absorptivity (É›).

  • A=ebc

  • This law is fundamental to quantitative analysis using spectrophotometry.

  • Transmittance (T) is the fraction of light passing through the sample; A= -logT

Limitations to Beer's Law

  • Beer's Law is not always valid at high concentrations.
  • Stray light, scattering, and high concentrations of electrolytes/analytes can cause deviations from this law.
  • For accurate measurements dilute samples are required when possible.

Standard Calibration Curve

  • Calibration curves are used to determine the relationship between the concentration of an analyte and the measured signal.
  • This is an empirical method.
  • Standard solutions with known analyte concentrations are used to construct the curve.

Reference

  • Harris, D. (2016) Quantitative Chemical Analysis.

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Introduction to Spectrophotometry PDF

Description

This quiz covers the fundamentals of spectrophotometry and its analytical applications, focusing on UV-Vis spectroscopy. You will learn about the principles behind light measurement techniques and their importance in fields such as DNA analysis and pharmaceuticals.

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