Spectroscopy

Summary

Spectroscopy is the study of the interactions of electromagnetic radiation, or light, with matter to gain information about the atoms or bonds present within the system. There are many types of spectroscopic techniques, but most are based on the absorption or emission of photons from the material being studied. The applications of spectroscopy span a variety of disciplines and allow scientists to, among countless other things, determine the elemental composition of a nearby dwarf star, the chemical identity of an unknown white powder sample, whether a transfected gene has been expressed, or the types of individual bonds within a molecule.

Definition and Basic Principles

Spectroscopy is the study of how light interacts with matter. It allows scientists across fields to study the composition of extremely large and extremely small systems. Each spectroscopic technique is unique, but the most widely used techniques are based on one of three phenomena—the absorption of light by matter, the emission of light by matter, or the scattering of light by matter. A photon can behave as both a particle and a wave. For most spectroscopic techniques, the wave nature of the photon is the most critical because the wavelength of light being emitted, absorbed, or scattered is where the information about the sample is contained.

Absorption spectroscopy involves the absorption of photons by matter and can give information about the types of atoms or bonds in a molecule. Typically, a given material will absorb specific wavelengths of light and will reflect or transmit all the other wavelengths. Emission spectroscopy involves the emission of photons from a sample on excitation. Scattering deals with light that is inelastically scattered from a sample, meaning the wavelength of light bouncing off the sample is not the same as the wavelength of light that was shined on the sample.

Related disciplines include electron spectroscopy and mass spectroscopy. Electron spectroscopic techniques generally involve either irradiating a sample with light and causing the emission of photoelectrons or bombarding a sample with electrons and causing the emission of X-rays. Either the photoelectrons or X-rays can be detected to gain chemical information about a sample. Mass spectroscopy is unique in that various methods can be employed to liberate ions from a sample, the masses of which are then detected to gain a chemical fingerprint of the present molecules.

Background and History

In 1666, Isaac Newton became the first person to discover that ambient light could be separated into a continuous band of varying colors using a prism. This discovery was the beginning of all spectroscopic research. In 1803, Thomas Young performed his famous slit experiment demonstrating the wave nature of light. William Hyde Wollaston observed the first absorption bands in solar radiation in 1802, and Joseph von Fraunhofer assigned these lines (called Fraunhofer lines) significance in 1817, paving the way for the fields of spectroscopy and astrophysics. In 1848, French physicist Jean Bernard Léon Foucault determined the locations of the lines were characteristic of the elements that were present. Lord Rayleigh (John William Strutt) investigated the elastic scattering of light in 1871, and Albert Einstein followed in his footsteps in 1910. However, a wavelength shift caused by inelastic scattering of light was not observed until 1928, a discovery for which Sir Chandrasekhara Venkata Raman won the Nobel Prize in Physics. In 1960, Theodore Harold Maiman built the first successful optical laser, which would later prove essential for modern spectroscopic instruments.

How It Works

Absorption Techniques. Fourier transform infrared spectroscopy (FTIR) is a vibrational spectroscopy technique that has the potential to elucidate what types of bonds are present within a sample. In transmittance mode, the sample is placed between an infrared light source and a detector and is irradiated with infrared light. Attenuated total reflectance (ATR) is a closely related technique that has minimal sample preparation. A sample is placed in contact with a crystal, such as diamond or germanium, and an evanescent infrared wave is sent through the crystal. Most bonds will absorb a specific wavelength of infrared light that causes them to vibrate or bend, and these wavelengths of light are absorbed in these processes and do not reach the detector. Polar bonds, such as carbonyls and ethers, are the bonds most easily detected using FTIR. The output of both FTIR and ATR is a spectrum with bands showing which wavelengths of light were absorbed by the sample. These wavelengths can then be interpreted to determine what types of bonds or functional groups were present in the sample.

Ultraviolet-visible (UV-Vis) spectroscopy is similar to FTIR in that it involves shining light of a known wavelength range, in this case visible and ultraviolet, at a sample and determining which wavelengths are absorbed by the sample. UV-Vis does not provide as much specific information about bond character as FTIR; however, it is a very useful and fast technique for quantifying the concentration of a known analyte in solution according to Beer's law.

Atomic absorption spectroscopy (AAS) is a quantitative elemental analysis technique that relies on the reproducible absorption of specific wavelengths of light by a given element on excitation. A sample in solution is atomized by drawing the solution into a heat source and then irradiated with a specific wavelength of light selected to excite a particular element. The amount of light absorbed by the sample at that wavelength is used to calculate the concentration of the analyte in the solution. AAS is very sensitive and is often used for detection of trace elements. Several heat sources, including an open flame, a graphite furnace, or an acetylene torch, can be employed to atomize the sample.

Astronomical spectroscopy is a method that astronomers use to determine the elemental composition of far-away celestial bodies that cannot be sampled in a laboratory. These bodies emit electromagnetic radiation across the entire electromagnetic spectrum; however, each element absorbs several characteristic wavelengths of light. So, by measuring which wavelengths of light are not being emitted by the very large sample in question, a fingerprint of all the elements present can be obtained.

Emission Techniques. Atomic emission spectroscopy (AES) is a technique that provides similar information to the information obtained using AAS but by essentially opposite means. In AES, a sample is atomized and excited by the same heat source, often a flame or a plasma, which is a highly ionized and energized gas. If a plasma is used, the technique is called inductively coupled plasma AES (ICP-AES). The excited analyte will emit light at wavelengths characteristic of the elements that are present. By detecting the emitted wavelengths of light and quantifying them, the elements in the sample can be identified and quantified. Multiple elements can be detected at once in AES.

Fluorescence spectroscopy is a family of techniques that uses higher energy radiation, such as ultraviolet light or X-rays, to excite the atoms of a sample. The excited state is unstable, so atoms must release lower energy photons to relax back to a more energetically favored state. Depending on the type of incident radiation, information about the sample can be obtained to varying degrees of specificity. For some analyses, it is enough to induce fluorescence so that the emitted photons can be imaged, as is the case in many biological applications. In other methods, such as X-ray fluorescence (XRF), specific elemental and chemical information can be obtained.

Scattering Techniques. Raman spectroscopy is a nondestructive vibrational spectroscopic technique based on inelastic light scattering. In Raman spectroscopy, a monochromatic laser is shined on the surface of a sample. Most of the light is elastically scattered, meaning that the light bounces off the sample at the same wavelength at which it entered; however, a very small fraction of the light is inelastically scattered, meaning that the light bounces off the sample at a different wavelength than that at which it entered because of interactions with the bonds in the sample. These interactions cause the bonds to vibrate and bend, similar to what happens in FTIR. The magnitude of the wavelength shift is characteristic of the bonds that caused it, and the resulting spectra are quite similar to FTIR spectra. However, whereas FTIR is most sensitive to polar bonds, Raman spectroscopy is most sensitive to polarizable bonds, such as carbon-carbon double bonds and aromatic rings.

Other Spectroscopic Methods. Many other spectroscopic techniques are across industry and academia. However, most are based on the same principles as the more common types. Examples of techniques include circular dichroism, dynamic light scattering, and spectroscopic ellipsometry. One widely used spectroscopic method not based on conventional absorption, emission, or scattering methods is nuclear magnetic resonance (NMR). In organic chemistry, NMR is used to study the organization of bonds in a molecule and can give information about the number and location of hydrogens, carbons, or other elements being studied in a sample.

Applications and Products

Spectroscopy is widely used in industrial and government-funded scientific endeavors for various purposes, including pharmaceutical analysis, failure analysis, materials science, reverse engineering, and toxicology. Vibrational spectroscopy is widely used to aid in the identification of unknown materials. For example, an analytical chemist may use FTIR to identify foreign material found on a manufacturing line or in a finished product, materials used in a competitor's product, or unknown material recovered from a crime scene. Searchable libraries of vibrational spectra can be consulted in conjunction with spectral interpretation to determine the identity of the material in question. There are many commercially available libraries, but libraries can also be constructed from in-house samples and standard materials.

Raman spectroscopy is a nondestructive, surface-sensitive technique often used to identify pigments in historical texts, art, and textiles to learn more about the people who created them and to verify their authenticity. Glass is largely transparent to Raman spectroscopy, making this technique ideal for identifying unknown materials found in glass containers without opening the container, thereby reducing the risks associated with exposure to an unknown material. This technique is especially useful in the identification of unknown flammable liquids. Thermo Fisher Scientific manufactures a portable Raman spectrometer for use in the field to allow emergency workers to evaluate potentially hazardous materials, such as chemical warfare agents or explosives. Techniques such as FTIR and Raman are used in airport screenings, as they are fast and reliable techniques for quickly identifying potentially harmful compounds.

Spectroscopic methods can also be used to quantify known materials. UV-Vis spectrometers can be used alone or as detectors in a chromatographic system in the pharmaceutical industry to quantify concentrations of drugs, biologics, and excipients in solution, as well as to verify the identity, purity, and stability of such compounds. Toxicologists use methods based on spectroscopy to quantify drugs in biological tissues and fluids.

Production facilities will often place spectroscopic instrumentation in line to continuously monitor products. For example, spectroscopic ellipsometry, which has the potential to measure thin films on the order of nanometers, can be placed in a production setting to monitor the thickness of vapor-deposited thin films.

Synthetic chemists typically use NMR to determine or verify the structure of the compounds they synthesize, and it is often used in polymer science, drug chemistry, and materials science. In the medical community, NMR is used to image various types of soft tissue in situ and is better known as magnetic resonance imaging (MRI). Hospitals also use spectroscopy to measure blood counts and couple spectroscopy with immunoassays to screen for drugs or other compounds in urine.

Various elemental analysis techniques, such as AAS or AES, are often used along with Raman spectroscopy to analyze the composition of geological samples and meteorites. By understanding what elements and compounds are present in these samples, geologists can better understand their formation and hypothesize about the processes that would create such a specimen. For example, researchers at Université de Lyon in France used Raman spectroscopy to identify two new types of ultrahard diamonds, one predicted and one unexpected, in the Havaro meteorite, which indicated that the parent asteroid had undergone an extremely violent collision.

Biologists use fluorescence spectroscopy to determine whether gene expression has occurred. By labeling the gene they wish to introduce into an organism with green fluorescent protein (GFP), they can visualize everywhere that the gene of interest was expressed. GFP can be visualized in cells, tissues, and organisms by exciting the GFP with blue light, which causes the emission of green light. Real-time Polymerase chain reaction (PCR), a method for making many copies of DNA, often uses fluorescent probes that bind to the new double-stranded DNA. By using fluorescence spectroscopy to quantify the amount of fluorescence, biologists can monitor how much DNA they have synthesized.

Several spectroscopic techniques, such as Raman, FTIR, and NMR, can be used to chemically map out surfaces and volumes of materials. By taking individual spectra at various points across a sample, the intensity of a given signal or combination of signals can be tracked and mapped on an intensity scale. The resolution of these techniques, that is, the smallest features that can be resolved, depends on the wavelength of the light used. Thus, Raman microscopy typically has much better resolution than FTIR because of the shorter wavelengths of light used in Raman spectroscopy. Chemical mapping can reveal details that would not otherwise be seen in a sample, such as where a specific protein may be expressed in a type of tissue or cell.

Careers and Course Work

Students interested in spectroscopy have a wide variety of career paths from which to choose, and their final career goal should determine their coursework. Completing a bachelor of science degree in one of the core sciences such as chemistry or physics serves as preparation for graduate school, which, although not required in every case, greatly improves one's prospects of obtaining a career in spectroscopic research. A bachelor's degree qualifies an individual for a position at the laboratory technician level. These employees are usually trained to operate a spectroscopic instrument and perform routine tasks. A master's degree or doctorate and appropriate research experience qualify the holder for a position involving advanced use and maintenance of spectroscopic instruments; design of spectroscopy experiments; spectroscopic data analysis, interpretation, and communication; or possibly the design of new instruments.

Careers involving spectroscopy are as diverse as the spectroscopic methods themselves. Chemists, physicists, astronomers, materials scientists, chemical engineers, biologists, and geologists can all be spectroscopists. Careers involving spectroscopy can be found in industries such as pharmaceutical companies, colleges, and universities and with the government, perhaps at a national laboratory or a Federal Bureau of Investigation crime laboratory. Despite the diversity of these options, applicants must possess a strong foundation in basic science, including chemistry physics, and mathematics. Coursework in analytical and physical chemistry is highly pertinent to most careers involving spectroscopy.

Social Context and Future Prospects

Spectroscopic techniques are powerful methods for identifying and sometimes quantifying system components, and applications of these techniques are likely to continue to diversify. Handheld spectrometers have become more commonplace in field forensic and military work because they allow personnel to identify the compounds at hand and know which safety precautions to take. Spectrometers will likely find their way into everyday-use products and continue to play a significant role in space missions, such as looking for the presence of water or amino acids on distant planets. In early 2021, the National Aeronautics and Space Administration (NASA) successfully landed an advanced rover, Perseverance, on the surface of Mars. In addition to the many goals of the rover's exploratory mission, scientists hoped Perseverance would detect evidence of past microbial life on the planet. In light of that aim, the rover was equipped with several scientific instruments, including two spectrometers designed to analyze X-rays and ultraviolet light. It also had a SuperCam instrument that combined different technologies, including Raman spectroscopy, to aid in detecting and studying rock and sediment samples on the planet.

An area of spectroscopy of interest is research using synchrotron light. Synchrotrons such as the Canadian Light Source and the European Synchrotron Radiation Facility (ESRF) use particle accelerators to produce ultra-high-intensity light of many wavelengths, which can then be diverted from the particle accelerator down a beam line and into a spectroscopy facility using a set of mirrors, gratings, attenuators, and lenses. Synchrotron light can penetrate deeper into materials and space, allowing information to be gained from farther away than before. In 2020, the ESRF was reopened after it had been closed for upgrade work since late 2018 and introduced its new, even more powerful Extremely Brilliant Source synchrotron, which had already been put to use in analysis of the virus causing the coronavirus disease 2019 (COVID-19) pandemic.

The resolution of spectroscopic chemical maps had been limited by physics, with maximum resolution obtained using synchrotron light and operating right at the limit of diffraction. However, several commercial techniques can image below the diffraction limit of light. One example is the nanoIR by Anasys Instruments, which uses an atomic-force microscope tip to gain infrared information about very small areas of a sample. Typically, ATR resolution is on the order of three to ten microns, but with nanoIR, resolution can be obtained at the submicron level. A great deal of research at universities, government-funded labs, and spectroscopy companies has been focusing on achieving better resolution so that smaller and smaller features can be accurately probed. In 2014, scientists Stefan Hell, Eric Betzig, and William Moerner were recognized for their contributions to the development of super-resolved fluorescence microscopy, receiving the Nobel Prize in Chemistry. Their microscopy and spectroscopy techniques allow scientists to study living cells in real-time at the molecular level, providing the opportunity to more accurately investigate the proteins involved in diseases such as Alzheimer's and Huntington's.

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