Name: absorbance spectra and beer-lambert analysis
Uploaded: 2020-03-26T07:22:26.000Z
Duration: 2 min 53 s
Channel: JoVE

K12 Standards

Texas Essential Knowledge and Skills

High School

Chemistry and Physics

IPC.7

IPC.7.E

Beer's Law

Muitas reações químicas progridem em duas direções, para frente e para trás. Com o tempo, as reações direta e reversa ocorrerão na mesma taxa e a concentração dos reagentes e produtos não mudará mais. Isso é conhecido como equilíbrio químico.
No equilíbrio químico, as concentrações de cada componente estão relacionadas entre si pela constante de equilíbrio, K, que é a razão entre as concentrações do produto e as concentrações do reagente, cada uma elevada à potência de seus coeficientes estequiométricos.
Mas como você determina as concentrações de equilíbrio? Um método mede a intensidade de um comprimento de onda de luz que o produto absorve antes e depois de passar por uma amostra. A diferença de intensidade é chamada de absorbância e corresponde à quantidade do composto absorvente na amostra.
Você deve se lembrar que os elétrons ocupam principalmente o estado fundamental. Quando eles absorvem uma certa quantidade de energia, eles são excitados para um nível de energia mais alto. Essa energia corresponde a um comprimento de onda específico da luz. Você pode encontrar esse comprimento de onda e medir a absorbância com um espectrofotômetro, que direciona um feixe de luz através da amostra e mede a mudança na intensidade em um ou mais comprimentos de onda.
A absorbância é igual ao log negativo da intensidade da luz atenuada sobre a intensidade da luz incidente. Ao plotar os valores de absorbância de várias soluções com diferentes concentrações de produtos conhecidas, observamos uma relação linear entre absorbância e concentração. Este é um exemplo da Lei de Beer.
A Lei de Beer é expressa matematicamente por esta equação, onde A é a absorbância, épsilon é o coeficiente de atenuação molar, uma constante que varia para cada composto, l é o comprimento do caminho da luz através da amostra e c é a concentração do composto.
Ao identificar a função linear de um determinado composto em um comprimento de onda e comprimento de caminho específicos, você pode usar os dados de absorbância de uma solução em equilíbrio para determinar a concentração de equilíbrio do produto. A partir daí, você pode calcular as concentrações de equilíbrio dos reagentes e resolver a constante de equilíbrio. Neste laboratório, você preparará soluções de isotiocianato de ferro (III) e usará um espectrofotômetro para determinar sua absorbância em várias concentrações.

lei da cerveja

Absorbância e Fluorescência
Quando a luz atinge uma substância, ela é absorvida, transmitida ou refletida. Normalmente, uma substância interage com uma faixa de comprimentos de onda de luz, cada um interagindo com as moléculas ou átomos de maneira diferente. Uma substância pode absorver uma faixa específica de comprimentos de onda, refletir outra faixa de comprimentos de onda e transmitir os outros comprimentos de onda da luz.
Quando uma molécula absorve luz, a energia é usada de quatro maneiras diferentes: (1) translação, que faz com que a molécula mude sua velocidade molecular; (2) vibração, que faz com que a distância entre as moléculas mude rapidamente; (3) rotação, que faz com que os átomos girem em torno das ligações em uma molécula; e (4) excitação de elétrons, que faz com que os elétrons façam a transição para níveis de energia mais altos.
<h4>Níveis de energia</h4>
Em 1913, Niels Bohr propôs um modelo para o átomo de hidrogênio no qual os elétrons viajam ao redor do núcleo em órbitas fixas e circulares, chamadas de estados estacionários. A energia associada a cada órbita, ou estado estacionário, existe apenas em energias fixas e discretas. Somente quando um elétron se move para outra órbita é que a energia é absorvida ou emitida. O elétron nunca está entre estados. Essa mudança só ocorre se a energia absorvida ou emitida for igual à diferença entre os dois estados de energia.
No modelo de Bohr, o número quântico n representa a energia do elétron. Quando um elétron ocupa o estado de energia mais baixo possível, diz-se que ocupa o estado fundamental, que é n = 1. Quando um elétron absorve um fóton, cuja energia é igual à diferença entre o primeiro e o segundo estados, o elétron fica excitado e faz a transição do estado fundamental para o estado excitado, onde n = 2. Se a energia do fóton for igual à diferença entre o primeiro e o terceiro estados, o elétron faz a transição para o terceiro estado, ou n = 3, e assim por diante.
Os elétrons podem retornar espontaneamente ao estado fundamental ou a qualquer outro estado excitado inferior. Quando isso acontece, o excesso de energia obtido com a excitação é liberado na forma de um fóton emitido. A energia do fóton é igual à diferença entre os dois estados de energia e corresponde a diferentes comprimentos de onda da luz.
<h4>Espectros de Absorção e Emissão</h4>
Embora a maioria das substâncias absorva ou emita a quantidade máxima de luz em um comprimento de onda, elas também tendem a absorver ou emitir luz em uma variedade de comprimentos de onda. Essa faixa de comprimentos de onda é chamada de espectro. A energia da luz absorvida é quantificada e visualizada usando um espectro de absorção, enquanto a energia da luz emitida é quantificada e visualizada usando um espectro de emissão.
Os espectros de absorção e emissão são medidos usando um espectrofotômetro, que é um dispositivo que transmite luz através de uma amostra e, em seguida, mede o comprimento de onda e a intensidade da luz que passa por ela. Dentro do espectrofotômetro há uma grade de difração ou um prisma, que separa a luz recebida em seus comprimentos de onda componentes. Os diferentes comprimentos de onda são então transmitidos através da amostra e a intensidade é registrada em um detector de dispositivo de carga acoplada linear (CCD). O CCD é um circuito integrado gravado em uma superfície de silício que forma elementos sensíveis à luz chamados pixels. O CCD coleta e classifica a luz difratada e a lê de volta em um comprimento de onda de absorção.
Ao medir a absorbância de uma amostra, o soluto é geralmente dissolvido em um solvente e colocado em um recipiente conhecido como cubeta. Em seguida, a amostra é colocada dentro do espectrofotômetro e a intensidade da luz transmitida é medida junto com os comprimentos de onda da luz para obter um espectro de absorbância. Como esperado, a intensidade da luz transmitida é menor do que quando não há amostra presente dentro do espectrofotômetro.
Isso ocorre porque a luz transmitida é absorvida pela amostra, pela cubeta e pelo solvente. Antes de medir as amostras, o espectrofotómetro deve ser calibrado com um «branco». Um branco é uma cubeta que contém apenas o solvente usado para dissolver o soluto. O espectrofotômetro é calibrado de modo que a absorbância total devida à cubeta e ao solvente seja subtraída da absorbância medida da amostra. Isso nos permite registrar a absorbância que é atribuída apenas às espécies de interesse.
A absorbância é frequentemente medida em um comprimento de onda, o comprimento de onda máximo de absorbância. No entanto, a absorção também pode ser medida em uma faixa de comprimentos de onda para adquirir o espectro de absorção. Para isso, a amostra é exposta a uma faixa de comprimentos de onda de luz incidente, e a absorção é registrada em cada comprimento de onda. Se a amostra emitir luz, o espectro de emissão é medido de forma semelhante, exceto que o comprimento de onda incidente é fixado no comprimento de onda de absorbância máxima. O instrumento então mede a intensidade da luz emitida em uma faixa de comprimentos de onda.
<h4>Lei Beer-Lambert</h4>
A absorbância de uma amostra no comprimento de onda de absorbância máxima fornece informações sobre a amostra, ou seja, sua concentração. A Lei de Beer-Lambert é uma equação que relaciona a transmitância com a concentração da amostra. A transmitância, ou intensidade da luz transmitida, é a fração da luz original que passa pela amostra, I, dividida pela intensidade da luz incidente, I0.
<img src="/CDNSource/lm/labs/49/49_Concepts_1.jpg" alt="Equação de transmitância \(T = \frac{I}{I_0}\), diagrama para análise de densidade óptica." data-altupdatedat="1754907201000">
A Lei de Beer-Lambert afirma que a absorbância óptica, A, de uma espécie em solução está relacionada ao log negativo da transmitância.
<img src="/CDNSource/lm/labs/49/49_Concepts_2.jpg" alt="Fórmula de absorbância A=-log(T), equação para análise de absorção óptica em estudos de espectroscopia." data-altupdatedat="1754907201000">
Uma versão alternativa da Lei de Beer-Lambert afirma que a absorbância óptica, A, de uma espécie em solução é linearmente proporcional à concentração, c, dessa espécie quando o comprimento de onda, λ, e o comprimento do caminho, l, são mantidos constantes.
<img src="/CDNSource/lm/labs/49/49_Concepts_3.jpg" alt="Equação da lei de Beer-Lambert, A=εcl, conceito de absorção, análise de dados espectroscópicos, diagrama de física." data-altupdatedat="1754907201000">
O coeficiente de atenuação molar, ε, é uma medida de quão fortemente uma espécie absorve luz em um determinado comprimento de onda. Quanto maior o coeficiente de atenuação molar, maior a absorbância. O comprimento do caminho, l, é a distância que a luz percorre através da amostra, que é a largura da cubeta. As cubetas padrão têm um comprimento de caminho de 1 cm.
Essa relação linear entre absorbância e concentração é uma ferramenta poderosa usada para determinar a concentração de uma amostra desconhecida com base em sua absorbância. Para fazer isso, uma curva padrão é gerada usando um gradiente de concentrações conhecidas do soluto. A absorbância no comprimento de onda de absorbância de pico, λmax, é medida para cada concentração.
Ao plotar concentração versus absorbância, observa-se uma relação linear que corresponde à equação de Beer-Lambert. A inclinação desta linha é igual ao produto do comprimento do caminho e do coeficiente de atenuação molar. Usando esta função linear calculada, se a absorbância da amostra desconhecida for conhecida, a concentração pode ser facilmente determinada.
Se a amostra que está sendo analisada for uma reação em equilíbrio, a Lei de Beer pode ser usada para determinar a concentração de equilíbrio de um produto ou reagente se a absorbância for medida em λmax específico para esse produto ou reagente. Uma vez que a concentração é conhecida, você pode determinar as concentrações de equilíbrio dos reagentes e produtos restantes e, em seguida, resolver a constante de equilíbrio Keq.
<h4>Referências</h4>
<ol>
	<li style="font-size:17px">Kotz, J.C., Treichel Jr, P.M., Townsend, J.R. (2012). Química e Reatividade Química. Belmont, CA: Brooks / Cole, Cengage Learning.</li>
	<li style="font-size:17px">Silderberg, M.S. (2009). Química: A Natureza Molecular da Matéria e da Mudança. Boston, MA: McGraw Hill, Boston.</li>
	<li style="font-size:17px">Harris, D.C. (2015). Análise Química Quantitativa. Nova York, NY: W.H. Freeman and Company.</li>
</ol>

Lei da Cerveja

Absorbância e Fluorescência
Quando a luz atinge uma substância, ela é absorvida, transmitida ou refletida. Normalmente, uma substância interage com um intervalo ...

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absorbance spectra and beer-lambert analysis

Absorbance spectra and the Beer-Lambert Law show how light can be used to study molecules in solution. When light hits a substance, it may be absorbed, transmitted, or reflected. A substance often interacts with a range of wavelengths, and each wavelength can affect the molecules or atoms in a different way.

When a molecule absorbs light, that energy can be used for translation, vibration, rotation, or electron excitement. Electron excitement is especially important in spectroscopy because it moves an electron to a higher energy level. In Niels Bohr’s model, electrons in the hydrogen atom move in fixed circular orbits called stationary states, and each state has a discrete energy. An electron can move only when it absorbs or emits energy equal to the difference between two states.

The lowest energy state is the ground state, or n = 1. If a photon has the right energy, the electron can move to a higher state such as n = 2 or n = 3. When the electron returns to a lower state, it releases the extra energy as an emitted photon. The photon energy matches the energy gap between the two states and corresponds to specific wavelengths of light.

Most substances absorb or emit most strongly at one wavelength, but they usually interact with a range of wavelengths. That range is called a spectrum. An absorption spectrum shows how much light a sample absorbs at different wavelengths, and an emission spectrum shows the light a sample gives off. These spectra are measured with a spectrophotometer, which sends light through a sample and records the wavelength and intensity of the light that passes through.

Inside the spectrophotometer, a diffraction grating or a prism separates incoming light into its component wavelengths. The light then passes through the sample, and a linear charge-coupled device, or CCD, detector records the intensity. A CCD is an integrated circuit on silicon that contains light-sensitive pixels. It collects and sorts the diffracted light and reads it back at an absorption wavelength.

For absorbance measurements, the solute is usually dissolved in a solvent and placed in a cuvette, which is the container used in the instrument. The sample is then placed in the spectrophotometer, and the intensity of transmitted light is measured along with wavelength to produce an absorbance spectrum. The transmitted light is lower than when no sample is present because light is absorbed by the sample, the cuvette, and the solvent. Before samples are measured, the spectrophotometer is calibrated with a blank, which is a cuvette that contains only the solvent. This step removes the absorbance from the cuvette and solvent so the reading reflects only the species of interest.

Absorbance is often measured at the wavelength of maximum absorbance, but it can also be recorded across a range of wavelengths. For emission spectra, the incident wavelength is fixed at the wavelength of maximum absorbance, and the instrument measures emitted light over a range of wavelengths. The Beer-Lambert Law then connects absorbance to sample concentration. Transmittance is the fraction of light that passes through the sample, I divided by I0, and absorbance is related to the negative log of transmittance.

A second form of the Beer-Lambert Law shows that absorbance is directly proportional to concentration when wavelength and pathlength stay constant. The molar attenuation coefficient, ε, describes how strongly a species absorbs light at a given wavelength. The pathlength, l, is the distance the light travels through the sample, which is the width of the cuvette. Standard cuvettes have a 1 cm pathlength. This linear relationship makes it possible to find the concentration of an unknown sample from its absorbance.

To do that, a standard curve is made with known concentrations of the solute. The absorbance at λ max is measured for each standard, and the results are plotted as concentration versus absorbance. The line that appears matches the Beer-Lambert equation, and its slope equals the product of the pathlength and the molar attenuation coefficient. Once that line is known, the concentration of an unknown sample can be calculated from its absorbance.

Beer’s Law can also be used for reactions at equilibrium. If absorbance is measured at the λ max of a specific product or reactant, the equilibrium concentration of that species can be found. The concentrations of the other reactants and products can then be determined, and the equilibrium constant K eq can be solved.

Absorbance Spectra and Beer-Lambert Analysis

absorbance-spectra-and-beer-lambert-analysis

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identification-of-unknown-aldehydes-and-ketones

Carbonyl Tests for Aldehydes and Ketones

Aldehydes and ketones are carbonyl compounds, which means they contain a C=O functional group. A ketone has two alkyl or aryl groups attached to the carbonyl carbon. Acetone is the simplest ketone, with two methyl groups. An aldehyde is similar, but it has at least one hydrogen attached to the carbonyl carbon. Formaldehyde is the simplest aldehyde, and acetaldehyde has one hydrogen and one methyl group. The carbonyl carbon in both aldehydes and ketones is electrophilic because the oxygen atom...

uv-vis spectroscopy of dyes

How Dyes Absorb Visible Light

Dyes absorb visible light because their molecular structure lets them interact with specific wavelengths. When light hits a substance, some light is absorbed and the rest is reflected or transmitted. The color we see depends on the wavelength that is reflected. A blue substance reflects blue light, around 430 to 480 nm, and absorbs the complementary orange region, around 590 to 630 nm. Compounds that do not absorb in the visible range appear colorless to the human eye. Light also has energy and...

identifying alcohols

Alcohol Tests for Primary, Secondary, and Tertiary

Alcohol tests can help sort alcohols by structure and reactivity. Alcohols are organic molecules with a hydroxyl group attached to an alkyl or aryl group, written as ROH. If the carbon with the hydroxyl group has one R group, the alcohol is primary. If it has two R groups, it is secondary. If it has three R groups, it is tertiary. Alcohols can also be aromatic when they contain a benzene ring. Phenol is the simplest aromatic alcohol. To identify an alcohol, chemists can use its chemical...

chemistry

Vídeos abrangentes e focados no currículo para instrutores e alunos de cursos introdutórios de laboratório de química

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Explore how spectrophotometers measure absorbance spectra, how blank calibration works, and how Beer-Lambert Law is used to find concentration and equilibrium values.

Absorbance Spectra and Beer-Lambert Analysis | JoVE K12

Louisiana Student Standards for Science

motion-and-stability-forces-and-interactions

Motion And Stability: Forces And Interactions

HS-PS2-6

Prepare-se para ensinar Lei da Cerveja com este guia do instrutor de 7 minutos. Abrange as principais etapas de preparação para o laboratório Chemistry no JoVE Lab Manual.

Preparação do Instrutor para Lei da Cerveja em Chemistry - 4.7K+ visualizações | JoVE Lab Manual

Siga os procedimentos passo a passo de Lei da Cerveja neste vídeo de protocolo estudantil de 17 minutos. Parte da coleção Chemistry no JoVE Lab Manual.

Protocolo do Estudante para Lei da Cerveja em Chemistry - 40K+ visualizações | JoVE Lab Manual

The Atomic Theory of Matter

The earliest recorded discussion of the basic structure of matter comes from ancient Greek philosophers. Leucippus and Democritus argued that all matter ...

Subatomic Particles

Dalton was only partially correct about the particles that make up matter. All matter is composed of atoms, and atoms are composed of three smaller ...

Elements: Chemical Symbols and Isotopes

A chemical symbol is an abbreviation used to indicate an element or an atom of an element. For example, the symbol for mercury is Hg. The same symbol is ...

Ions and Ionic Charges

In ordinary chemical reactions, the nucleus &mdash; which contains the protons and neutrons of each atom and thus identifies the element &mdash; remains ...

The Periodic Table

As early chemists discovered more elements, they realized that various elements could be grouped by their similar chemical behaviors. One such grouping ...

Atomic Mass

Atoms &mdash; and the protons, neutrons, and electrons that compose them &mdash; are extremely small. For example, a carbon atom weighs less than 2 ...

Molar Mass

The identity of a substance is defined not only by the types of atoms or ions it contains but by the quantity of each type of atom or ion. For example, ...

Periodic Classification of the Elements

The periodic table arranges atoms based on increasing atomic number so that elements with the same chemical properties recur periodically. When their ...

Types of Chemical Bonds

Chemical bonding theories were pioneered by American chemist Gilbert&#xA0;N. Lewis. He developed a model called the Lewis model to explain the type and ...

Ionic Bonding and Electron Transfer

Ions are atoms or molecules bearing an electrical charge. A cation (a positive ion) forms when a neutral atom loses one or more electrons from its valence ...

The Born-Haber Cycle

Lattice Energy&#xA0;

An ionic compound is stable because of the electrostatic attraction between its positive and negative ions. The lattice energy of a ...

trends-in-lattice-energy-ion-size-and-charge

Trends in Lattice Energy: Ion Size and Charge

An ionic compound is stable because of the electrostatic attraction between its positive and negative ions. The lattice energy of a compound is a measure ...

Covalent Bonding and Lewis Structures

Compared to ionic bonds, which results from the transfer of electrons between metallic and nonmetallic atoms, covalent bonds result from the mutual ...

Electronegativity

Whether a bond is nonpolar or polar covalent is determined by a property of the bonding atoms called electronegativity.&#xA0;

Electronegativity values of ...

bond-polarity-dipole-moment-and-percent-ionic-character

Bond Polarity, Dipole Moment, and Percent Ionic Character

Bond Polarity

The absolute value of the difference in electronegativity (&Delta;EN) of two bonded atoms provides a rough measure of the polarity to be ...

lewis-structures-of-molecular-compounds-and-polyatomic-ions

Lewis Structures of Molecular Compounds and Polyatomic Ions

To draw Lewis structures for complicated molecules and molecular ions, it is helpful to follow a step-by-step procedure as outlined:


	Determine the ...

Bond Energies and Bond Lengths

Stable molecules exist because covalent bonds hold the atoms together. The strength of a covalent bond is measured by the energy required to break it, ...

Bonding in Metals

Metallic bonds are formed between two metal atoms. A simplified model to describe metallic bonding has been developed by Paul Dr&#xFC;de called the ...

Model basic atomic structure and relate an element's atomic structure to its bonding, reactivity, and placement on the Periodic Table.

IPC.7.A

physical-and-chemical-properties-of-matter

Physical and Chemical Properties of Matter

The characteristics that enable us to distinguish one substance from another are called properties.

Physical Properties of Matter

A physical property is ...

atomic-radii-and-effective-nuclear-charge

Atomic Radii and Effective Nuclear Charge

The elements in groups of the periodic table exhibit similar chemical behavior. This similarity occurs because the members of a group have the same number ...

Ionic Radii

Ionic radius is the measure used to describe the size of an ion. A cation always has fewer electrons and the same number of protons as the parent atom; it ...

Ionization Energy

The amount of energy required to remove the most loosely bound electron from a gaseous atom in its ground state is called its first ionization energy ...

Electron Affinity

The electron affinity (EA) is the energy change for adding an electron to a gaseous atom to form an anion (negative ion).



This process can be either ...

Alkali Metals

Group 1 elements are soft and shiny metallic solids. They are malleable, ductile, and good conductors of heat and electricity. The melting points of the ...

Halogens

Group 17 elements, known as halogens, are nonmetals. At room temperature, fluorine and chlorine are gases, bromine is a liquid, and iodine a solid. ...

Noble Gases


The elements in group 18 are noble gases (helium, neon, argon, krypton, xenon, and radon). They earned the name &ldquo;noble&rdquo; because they were ...

Use patterns within the Periodic Table to predict the relative physical and chemical properties of elements.

IPC.7.B

Classifying Matter by State

Chemistry is the study of matter and the changes it undergoes. Matter is anything that has mass and occupies space. Matter is all around us; the air, ...

Classifying Matter by Composition

Matter: Pure Substances and Mixtures
According to its composition, the matter can be classified into two broad categories &mdash; pure substances and ...

Phase Transitions

Whether solid, liquid, or gas, a substance&#39;s state depends on the order and arrangement of its particles (atoms, molecules, or ions). Particles in the ...

phase-transitions-vaporization-and-condensation

Phase Transitions: Vaporization and Condensation

The physical form of a substance changes on changing its temperature. For example, raising the temperature of a liquid causes the liquid to vaporize ...

Phase Transitions: Melting and Freezing

Heating a crystalline solid increases the average energy of its atoms, molecules, or ions, and the solid gets hotter. At some point, the added energy ...

Explain how physical and chemical properties of substances are related to their usage in everyday life such as in sunscreen, cookware, industrial applications, and fuels.

IPC.7.C

Science concepts. The student knows that relationships exist between the structure and properties of matter. The student is expected to:

Absorbance Spectra and Beer-Lambert Analysis

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