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

Molte reazioni chimiche progrediscono in due direzioni, avanti e indietro. Nel tempo, le reazioni dirette e inverse si verificheranno alla stessa velocità e la concentrazione dei reagenti e dei prodotti non cambierà più. Questo è noto come equilibrio chimico.
All'equilibrio chimico, le concentrazioni di ciascun componente sono correlate tra loro dalla costante di equilibrio, K, che è il rapporto tra le concentrazioni del prodotto e le concentrazioni dei reagenti, ciascuna elevata alla potenza dei propri coefficienti stechiometrici.
Ma come si determinano le concentrazioni di equilibrio? Un metodo misura l'intensità di una lunghezza d'onda della luce che il prodotto assorbe prima e dopo il passaggio attraverso un campione. La differenza di intensità è chiamata assorbanza e corrisponde alla quantità di composto assorbente nel campione.
Potresti ricordare che gli elettroni occupano principalmente lo stato fondamentale. Quando assorbono una certa quantità di energia, vengono eccitati a un livello di energia più elevato. Quell'energia corrisponde a una specifica lunghezza d'onda della luce. È possibile trovare questa lunghezza d'onda e misurare l'assorbanza con uno spettrofotometro, che dirige un raggio di luce attraverso il campione e misura la variazione di intensità a una o più lunghezze d'onda.
L'assorbanza è uguale al logaritmo negativo dell'intensità della luce attenuata rispetto all'intensità della luce incidente. Tracciando i valori di assorbanza di più soluzioni con diverse concentrazioni di prodotto note, osserviamo una relazione lineare tra assorbanza e concentrazione. Questo è un esempio della legge della birra.
La legge di Beer è espressa matematicamente da questa equazione, dove A è l'assorbanza, epsilon è il coefficiente di attenuazione molare, una costante che varia per ogni composto, l è la lunghezza del percorso della luce attraverso il campione e c è la concentrazione del composto.
Identificando la funzione lineare per un dato composto a una lunghezza d'onda e a una lunghezza del percorso specifiche, è possibile utilizzare i dati di assorbanza di una soluzione all'equilibrio per determinare la concentrazione di equilibrio del prodotto. Da lì, è possibile calcolare le concentrazioni di equilibrio dei reagenti e risolvere la costante di equilibrio. In questo laboratorio, preparerai soluzioni di isotiocianato di ferro (III) e utilizzerai uno spettrofotometro per determinarne l'assorbanza a varie concentrazioni.

legge della birra

Assorbanza e fluorescenza
Quando la luce colpisce una sostanza, viene assorbita, trasmessa o riflessa. Tipicamente, una sostanza interagisce con una gamma di lunghezze d'onda della luce, ognuna delle quali interagisce con le molecole o gli atomi in modo diverso. Una sostanza può assorbire una specifica gamma di lunghezze d'onda, riflettere un'altra gamma di lunghezze d'onda e trasmettere le altre lunghezze d'onda della luce.
Quando una molecola assorbe la luce, l'energia viene utilizzata in quattro modi diversi: (1) traslazione, che fa sì che la molecola cambi la sua velocità molecolare; (2) vibrazione, che fa cambiare rapidamente la distanza tra le molecole; (3) rotazione, che fa ruotare gli atomi attorno ai legami di una molecola; e (4) eccitazione degli elettroni, che provoca la transizione degli elettroni a livelli di energia più elevati.
<h4>Livelli di energia</h4>
Nel 1913, Niels Bohr propose un modello per l'atomo di idrogeno in cui gli elettroni viaggiano intorno al nucleo in orbite fisse e circolari, chiamate stati stazionari. L'energia associata a ciascuna orbita, o stato stazionario, esiste solo a energie fisse e discrete. Solo quando un elettrone si sposta su un'altra orbita viene assorbita o emessa energia. L'elettrone non è mai tra gli stati. Questa variazione si verifica solo se l'energia assorbita o emessa è uguale alla differenza tra i due stati energetici.
Nel modello di Bohr, il numero quantico n rappresenta l'energia dell'elettrone. Quando un elettrone occupa lo stato energetico più basso possibile, si dice che occupa lo stato fondamentale, che è n = 1. Quando un elettrone assorbe un fotone, la cui energia è uguale alla differenza tra il primo e il secondo stato, l'elettrone si eccita e passa dallo stato fondamentale allo stato eccitato, dove n = 2. Se l'energia del fotone è uguale alla differenza tra il primo e il terzo stato, l'elettrone passa al terzo stato, o n = 3, e così via.
Gli elettroni possono tornare spontaneamente allo stato fondamentale o a qualsiasi altro stato inferiore ed eccitato. Quando ciò accade, l'energia in eccesso che è stata guadagnata dall'eccitazione viene rilasciata sotto forma di un fotone emesso. L'energia del fotone è uguale alla differenza tra i due stati energetici e corrisponde a diverse lunghezze d'onda della luce.
<h4>Spettri di assorbimento ed emissione</h4>
Sebbene la maggior parte delle sostanze assorba o emetta la quantità massima di luce a una lunghezza d'onda, tende anche ad assorbire o emettere luce a una gamma di lunghezze d'onda. Questa gamma di lunghezze d'onda è chiamata spettro. L'energia della luce assorbita viene quantificata e visualizzata utilizzando uno spettro di assorbimento, mentre l'energia della luce emessa viene quantificata e visualizzata utilizzando uno spettro di emissione.
Gli spettri di assorbimento e di emissione vengono misurati utilizzando uno spettrofotometro, che è un dispositivo che trasmette la luce attraverso un campione e quindi misura sia la lunghezza d'onda che l'intensità della luce che lo attraversa. All'interno dello spettrofotometro c'è un reticolo di diffrazione o un prisma, che separa la luce in arrivo nelle sue lunghezze d'onda componenti. Le diverse lunghezze d'onda vengono quindi trasmesse attraverso il campione e l'intensità viene registrata su un rivelatore CCD (Line-Charge-Coupled Device). Il CCD è un circuito integrato inciso su una superficie di silicio che forma elementi sensibili alla luce chiamati pixel. Il CCD raccoglie e ordina la luce diffratta e la rilegge a una lunghezza d'onda di assorbimento.
Quando si misura l'assorbanza di un campione, il soluto viene solitamente disciolto in un solvente e posto in un contenitore noto come cuvetta. Quindi, il campione viene posizionato all'interno dello spettrofotometro e l'intensità della luce trasmessa viene misurata insieme alle lunghezze d'onda della luce per ottenere uno spettro di assorbanza. Come previsto, l'intensità della luce trasmessa è inferiore rispetto a quando non è presente alcun campione all'interno dello spettrofotometro.
Questo perché la luce trasmessa viene assorbita dal campione, dalla cuvetta e dal solvente. Prima di misurare i campioni, lo spettrofotometro deve essere calibrato con un "bianco". Un bianco è una cuvetta che contiene solo il solvente utilizzato per sciogliere il soluto. Lo spettrofotometro è calibrato in modo che l'assorbanza totale dovuta alla cuvetta e al solvente venga sottratta dall'assorbanza misurata dal campione. Questo ci permette di registrare l'assorbanza che viene attribuita solo alle specie di interesse.
L'assorbanza è spesso misurata a una lunghezza d'onda, la lunghezza d'onda massima dell'assorbanza. Tuttavia, l'assorbimento può anche essere misurato a una gamma di lunghezze d'onda per acquisire lo spettro di assorbimento. Per questo, il campione viene esposto a una gamma di lunghezze d'onda della luce incidente e l'assorbimento viene registrato a ciascuna lunghezza d'onda. Se il campione emette luce, lo spettro di emissione viene misurato in modo simile, tranne per il fatto che la lunghezza d'onda incidente è fissata alla lunghezza d'onda della massima assorbanza. Lo strumento misura quindi l'intensità della luce emessa su una gamma di lunghezze d'onda.
<h4>Legge di Beer-Lambert</h4>
L'assorbanza di un campione alla lunghezza d'onda della massima assorbanza fornisce informazioni sul campione, ovvero sulla sua concentrazione. La legge di Beer-Lambert è un'equazione che mette in relazione la trasmittanza con la concentrazione del campione. La trasmittanza, o intensità della luce trasmessa, è la frazione di luce originale che passa attraverso il campione, I, divisa per l'intensità della luce incidente, I0.
<img src="/CDNSource/lm/labs/49/49_Concepts_1.jpg" alt="Equazione della trasmittanza \(T = \frac{I}{I_0}\), diagramma per l'analisi della densità ottica." data-altupdatedat="1754907201000">
La legge di Beer-Lambert afferma che l'assorbanza ottica, A, di una specie in soluzione è correlata al log negativo della trasmittanza.
<img src="/CDNSource/lm/labs/49/49_Concepts_2.jpg" alt="Formula dell'assorbanza A=-log(T), equazione per l'analisi dell'assorbimento ottico negli studi di spettroscopia." data-altupdatedat="1754907201000">
Una versione alternativa della legge di Beer-Lambert afferma che l'assorbanza ottica, A, di una specie in soluzione è linearmente proporzionale alla concentrazione, c, di quella specie quando la lunghezza d'onda, λ, e la lunghezza del cammino, l, sono mantenute costanti.
<img src="/CDNSource/lm/labs/49/49_Concepts_3.jpg" alt="Equazione della legge di Beer-Lambert, A=εcl, concetto di assorbimento, analisi spettroscopica dei dati, diagramma fisico." data-altupdatedat="1754907201000">
Il coefficiente di attenuazione molare, ε, è una misura della forza con cui una specie assorbe la luce a una data lunghezza d'onda. Maggiore è il coefficiente di attenuazione molare, maggiore è l'assorbanza. La lunghezza del percorso, l, è la distanza percorsa dalla luce attraverso il campione, che è la larghezza della cuvetta. Le cuvette standard hanno una lunghezza del percorso di 1 cm.
Questa relazione lineare tra assorbanza e concentrazione è un potente strumento che viene utilizzato per determinare la concentrazione di un campione sconosciuto in base alla sua assorbanza. Per fare ciò, viene generata una curva standard utilizzando un gradiente di concentrazioni note del soluto. L'assorbanza alla lunghezza d'onda di assorbanza di picco, λmax, viene misurata per ogni concentrazione.
Tracciando la concentrazione rispetto all'assorbanza, si osserva una relazione lineare che corrisponde all'equazione di Beer-Lambert. La pendenza di questa linea è uguale al prodotto della lunghezza del percorso e del coefficiente di attenuazione molare. Utilizzando questa funzione lineare calcolata, se l'assorbanza del campione sconosciuto è nota, la concentrazione può essere facilmente determinata.
Se il campione analizzato è una reazione all'equilibrio, la legge di Beer può essere utilizzata per determinare la concentrazione di equilibrio di un prodotto o di un reagente se l'assorbanza è misurata a λmax specifico per quel prodotto o reagente. Una volta nota la concentrazione, è possibile determinare le concentrazioni di equilibrio dei reagenti e dei prodotti rimanenti e quindi risolvere la costante di equilibrio Keq.
<h4>referenze</h4>
<ol>
	<li style="font-size:17px">Kotz, J.C., Treichel Jr, P.M., Townsend, J.R. (2012). Chimica e reattività chimica. Belmont, CA: Brooks/Cole, Cengage Learning.</li>
	<li style="font-size:17px">Silderberg, M.S. (2009). Chimica: la natura molecolare della materia e il cambiamento. Boston, MA: McGraw Hill, Boston.</li>
	<li style="dimensione del carattere:17px">Harris, D.C. (2015). Analisi chimica quantitativa. New York, NY: W.H. Freeman e compagnia.</li>
</ol>

Legge della birra

Assorbanza e fluorescenza
Quando la luce colpisce una sostanza, viene assorbita, trasmessa o riflessa. Tipicamente, una sostanza interagisce con una gamma ...

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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

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esterification

Ester Formation and Everyday Fragrances

Esters are organic compounds with a carbonyl group attached to an oxygen that connects to another alkyl or aryl group. Their general formula is RCOOR'. They are often made by reacting a carboxylic acid with an alcohol in an esterification reaction. In this process, the hydroxyl group of the acid is replaced by an alkyl or aryl group. Many simple esters have familiar fruit and flower smells because their structures are small and less complex. Formic acid and methanol can combine to form methyl...

identification of unknown aldehydes and ketones

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

Video completi e incentrati sul curriculum sia per gli istruttori che per gli studenti nei corsi di laboratorio di Chimica introduttiva.

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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

Preparati a insegnare Legge della birra con questa guida per istruttori di 7 minuti. Copre i passaggi chiave di preparazione per il laboratorio Chemistry su JoVE Lab Manual.

Preparazione dell'istruttore per Legge della birra in Chemistry - Oltre 4.7K visualizzazioni | JoVE Lab Manual

Segui le procedure passo dopo passo di Legge della birra in questo video di protocollo studente di 17 minuti. Parte della collezione Chemistry su JoVE Lab Manual.

Protocollo studente per Legge della birra in Chemistry - Oltre 40K visualizzazioni | 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 ...

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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