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In this column, we attempt to explore in depth the theory and methodology of our common laboratory techniques and show how an understanding of interworking of those techniques allow for more flexibility in analysis.
In this column, we attempt to explore in depth the theory and methodology of our common laboratory techniques and show how an understanding of interworking of those techniques allow for more flexibility in analysis. The knowledge of the theories behind the scenes can be used to manipulate the method variables for a more accurate and precise result. Up until this point, we have been mainly focused on the techniques commonly used in molecular spectroscopy and chromatography, which all have organic molecules as their target analytes. In the next few columns, we will look at another critical aspect of analytical chemistry—atomic spectroscopy and elemental analysis.
In the previous columns we have examined spectroscopy as a tool of organic chemical analysis. Spectroscopy is the study or measurement of the interaction of matter and electromagnet radiation resulting in spectra (wavelength or frequency of the radiation). In organic analysis, spectroscopy examines the interaction of energy and molecules, but in inorganic chemistry we focus on atomic spectroscopy or the interaction of atoms and energy. The designation of the type of spectroscopy is categorized by the area of the electromagnetic spectrum targeted (for example, infrared, ultraviolet [UV], or X-ray, and so forth), the atomization source and interaction (such as emission, adsorption, transmittance, and so on), or the type of spectroscopy being employed (optical or mass spectrometry).
Electromagnetic (EM) radiation spectrum encompasses a wide band of energy including radio waves, microwaves, infrared, visible light, ultraviolet, X-rays, and gamma rays (see Figure 1).
Many laboratory analytical techniques are focused on the range of waves associated with light from infrared to the X-ray range of wavelengths.
There are numerous interactions between EM and matter. These include some of the most commonly used interactions in instrumentation including absorption, transmission or refraction, reflection, and emission. All of these processes become the basis for some form of detection or spectroscopy measurement technique (see Figure 2).
Spectroscopy techniques are divided into atomic and molecular spectroscopy depending on the target to be measured and the material being evaluated.Spectrometry instruments are often further divided and defined by the type of interactions they produce and the type of energy that is measured like X-ray fluorescence (XRF), which measures X-rays and the amount of fluorescent energy a material emits during exposure. Some techniques focus on a narrow band of interactions (such as emission only) or a small part of the EM spectrum (like X-rays) while other techniques monitor an array of interactions and wavelengths on the spectrum (see Table I).
In atomic spectroscopy, changes in energy are most commonly measured by the movement of electrons in atomic orbitals when exposed to an energy source such as light. The energy movement of electrons in orbitals use or produce measurable changes in energy usually as photons (Figure 3).
The characteristic movement of electrons when exposed to particular types or wavelengths of energy produces signature energy fingerprints called spectra. Each element of the periodic table has a unique spectrum based on the number of electrons where changes in wavelength and intensity can be calculated as the number of atoms of a particular element in a sample (Figure 4).
In atomic absorption spectroscopy (AAS), light of a set wavelength is passed through a sample of atoms and if that wavelength corresponds to the difference in energy as their electrons move to higher energy states, the light (or a portion of the light) is absorbed, and that difference is calculated as an absorbance value.
The basic components of an atomic spectroscopy system include an energy source, an atomizer or nebulizer to distribute and reduce the sample to its corresponding atoms, a focus or monochromator, and finally a detector (Figure 5).
The energy for most of the atomic absorption instrumentation comes from a variety of lamps that produce metal atoms similar to the sources found in other molecular spectroscopy techniques. Sources for absorption spectroscopy are divided into two groups: continuum and line sources.
Continuous sources provide a wide range of wavelengths of energy and do not require very high sensitivity monochromators to differentiate the wavelengths. These sources include some of the common lamps we discussed when examining molecular spectrometers in previous columns (for example, tungsten, mercury, xenon, and deuterium lamps). The most common use for these types of sources are molecular absorption spectrometers or fluorometers. In atomic absorption, these lamps can be used for background correction.
Line sources produce specific portions or bands of a spectrum and can have large gaps between spectral lines because of the fact that they only contain a few wavelengths. Line sources do require high resolution monochromators and separate lamps for each range of wavelengths required for analysis. Most atomic absorption spectroscopy instruments use some type of line source either as a hollow cathode lamp, electrodeless discharge lamp, sodium, or mercury vapor lamps.
One of the most commonly used lamps is the hollow cathode lamp, which contains a tungsten anode and a hollow cylindrical cathode made with an element selected for its spectrum. The cathode tube is a sealed glass vessel filled with an inert gas (argon or neon). A high voltage is applied to ionize the gas and as a current flows between the anode and cathode the metal atoms are liberated from the cathode and collide with the gas and are excited producing characteristic radiation. The cathode of the lamp is created using the metals targeted for analysis. Some multielement lamps are created by the fusion of several target metals together producing the ability to measure more than one element at the expense of overall sensitivity for any one element. These multielement lamps can also be a source of spectral interferences in the analysis. Modern atomic absorption spectrometers can contain anywhere from 2 to more than 12 lamps in a system.
In order for the sample to be analyzed, the material must be broken down into atoms before exposure to the energy sources. The first step of many liquid matrix samples is nebulization or the process of creating fine droplet spray from a liquid sample. Nebulizers and the supporting pumps pull samples into the system at a controlled rate and create a fine spray that is then injected into the atomizer along with some type of fuel and oxidizer.
The atomizers that are most commonly found in use are either flame or electrothermal atomizers. The oldest technology still in common use is the flame atomizer. Flames are produced by a mix of air or nitrous oxide with acetylene. The mix of air-acetylene has a temperature of about 2300 °C while the nitrous oxide mix can be up to 2700 °C. Once liquid sample (via a nebulizer) enters the flame it undergoes drying then vaporization to transfer the material to the gas phase. These systems are straightforward and economical, but can be subject to increased levels of interference during the introduction and atomization phases and possible ionization of the sample leading to reduced sensitivity because of ground state depletion. This technique generally has sensitivity in the parts per million range with some elements having lower detection.
In electrothermal atomizers a graphite tube about 20 mm long and 5 mm diameter becomes a heated chamber via an electrical current allowing for precise control of the temperature of the furnace or chamber. The samples (either solid, liquid, or gas) enter the tube and are subjected to a temperature program that dries then pyrolizes the sample before atomization transitions samples to the gas phase. This technique is most commonly referred to as graphite furnace atomic absorption spectroscopy (GFAAS). It has the most flexibility of the atomic absorption techniques and can analyze many different forms of sample and provides part per billion (ppb) and sub-ppb analytical ranges. GFAAS is also known to have less interferences than traditional flame AA, which has made it a popular technique for atomic absorption.
Atomic absorption techniques are limited to a small number of elements in the periodic table (usually about 70). Both techniques require separate lamps for each targeted element. Flame absorption systems require flammable gas mixtures and rely upon the skill of the operator and cleanliness of the samples and system to avoid interferences and ionization. GFAAS has a higher degree of sensitivity but is still limited by the number of elements that can be analyzed. The GFAAS systems do have a wider range of functionality as to sample type and matrix being able to analyze liquids, gases, and solids.
In atomic emission spectroscopy (AES), a high intensity source of light energy such as a spark, arc, flame, or plasma is produced to excite atoms. The sample pathway is similar to atomic absorption where the sample as a gas or a solution is aerosolized in a nebulizer before being vaporized and atomized. The atoms are then excited to a higher energy state by the source before moving back to lower energy and ground states. The movement of the electrons from excited to ground states emits photon energy (such as, light) and is a reason AES is also known as optical emission spectroscopy (OES). The emissions from the element are focused by mirrors and slits to a detector (Figure 6).
The nebulizer or atomizer is the basis of the sample introduction system. Nebulizers by definition create a mist of particles from liquids. There are three basic groups of nebulizers: ultrasonic, electrothermal, and pneumatic nebulizers based on their primary mode of nebulization. Ultrasonic nebulizers, as those found in many medical settings, force liquids onto a surface of vibrating surface. Electrothermal nebulizers use a closed chamber evaporator with a method of heating and cooling to produce nebulized particles. There are spectroscopy systems that do use electrothermal nebulizers. But the most common type of analytical nebulizers are pneumatic nebulizers that use induction to atomize liquids. When a gas of a higher-pressure flows into a lower-pressure gas it creates a jet and pulls in the lower-pressure aerosol. The liquid is pumped into the system and the high-pressure gas draws in and disperses the particles into the jet.
There are many types of modern nebulizer designs that fit into several categories including: concentric, cross flow, Babington or V-Groove, porous disc, parallel path, flow blurring, and vibrating mesh. The first two (concentric and cross flow) are the most used configuration of nebulizer for AES and OES.
In the concentric nebulizer the gas surrounds the sample introduction or vice versa so that concentric rings of sample and gas are formed producing a plume at the end of nebulized material (Figure 7a). Cross flow nebulizers have a configuration where the gas and the liquid flow are at right angles to each other usually meeting at the tip of the nebulizer to come together to form the plume in a spray chamber (Figure 7b). Cross flow nebulizers allow for more sample to be introduced to the system, but can be prone to clogging.
The next stop for the nebulized sample is the light source which excites the atoms and causes the emission of photons. Again, there are many different types of sources for AES including arc, spark, flame, and plasma.
The earliest types of AES systems used arcs, sparks, and flames, as the source of excitation. Sparks are short bursts of excitation energy generated by an electrode usually composed of graphite. Arcs are sustained AC or DC currents of energy between two electrodes with frequency similar to sparks. DC arcs are considered low-current discharges of about up to 15 A, while AC arcs are higher between 15 to 30 A. The current and the temperature of the arc discharges are controlled by adjusting the applied potential.
These techniques are used in the analysis of metallic solid samples. Materials that are nonconductive are mixed with a conductive matrix such as graphite to facilitate analysis. The sample is heated to a high temperature by the passage of the spark or arc through the sample. The excited atoms emit characteristic wavelength light that is filtered by the monochromator for the detector. Older arc and spark technologies were almost always qualitative or at best semi-quantitative, but newer advances have increased the sensitivity and accuracy of both types of systems. Despite the advances, these systems are not for precise and fine analytical work with low levels of detection.
Another early type of AES was flame emission spectroscopy (FES). FES uses the energy from the flame to excite species of certain metals like Group I alkali metals and Group II alkaline metals. The high temperatures of the flame can cause some instability in some elements.
This type of instrumentation uses a burner and a flammable gas or liquid to produce a flame. A sample is mixed with an oxidant and nebulized in a mixing chamber with the fuel gas. The flame then vaporizes and atomizes the sample. The energy of the change of states as the high temperature changes the energy levels is focused by lenses and filtered by a monochromator before reaching the detector (Figure 8).
There are different types of burners and mixtures of gas that are used in these applications. Gases are chosen regarding temperature and the nature of the target elements; more energy and higher temperatures are needed to excite larger numbers of atoms to higher states. Some metals also require higher temperatures to vaporize. The most common mix of gases for the burner contain a mix of air, oxygen, hydrogen, nitrous oxide, or acetylene depending on the target elements and temperatures that need to be achieved to vaporize those elements without oxidation or instability (Table II).
The most widely used types of atomic emission spectrometers use a plasma source to vaporize and atomize the samples. A plasma is a homogenous mixture a fine mist or fog of ionized gases composed of electrons, ions, and neutral species. The most common gas for spectroscopy plasma is argon. The method in which the plasma is depends on the source of the electric field that produces it. The mostly widely used types of plasma are either by direct current plasma (DCP) or inductively coupled plasma (ICP). Other types of plasma with less applications in the analytical world are laser-induced plasma, laser breakdown, and microwave-induced plasma.
In DCP, ionization occurs in a discharge tube with between two and three electrodes (often graphite anode blocks with a tungsten cathode block). Argon gas flows around the anode blocks and the plasma is produced by the contact of the cathode with the anodes. The temperature at the core can be more than 8000 K.
For ICP, there are three concentric silica quartz tubes (called the torch) in which argon gas moves the sample in its aerosol through an applied Rf power supply. The plasma is ignited by a spark from a Tesla coil. The hottest area of the plasma torch is at the base around 10,000 K while the further away from the tip of the torch the temperature drops to 6000 K (Figure 9).
Plasma sources for atomic emission spectroscopy offer a much broader range of applications than other types of both atomic emission and atomic absorption spectrometry because of their sensitivity and their ability to produce spectra for a wide range of the periodic table of elements. Detection limits can be dropped even further with the coupling of mass spectrometers to ICP systems thereby allowing for sub-ppb levels of routine detection and expanding the analytical reach to ultra-trace level detection. In upcoming columns, we will look deeper into applications and the configurations of ICP-OES and ICP-MS systems and how they have become indispensable tools for the cannabis laboratory.
Patricia Atkins is a Senior Applications Scientist with Spex, an Antylia Scientific company and has been a member of many cannabis advisory committees and working groups for cannabis including NACRW, AOAC and ASTM.
P. Atkins, Cannabis Science and Technology® Vol. 5(7), 16-22 (2022).