SCIENCE EDUCATION > Chemistry

Analytical Chemistry

This collection takes a broad look at quantitative analysis and instrumentation including electrochemistry, spectroscopy, chromatography, and mass spectrometry.

  • Analytical Chemistry

    09:51
    Sample Preparation for Analytical Characterization

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Sample preparation is the way in which a sample is treated to prepare for analysis. Careful sample preparation is critical in analytical chemistry to accurately generate either a standard or unknown sample for a chemical measurement. Errors in analytical chemistry methods are categorized as random or systematic. Random errors are errors due to change and are often due to noise in instrument. Systematic errors are due to investigator or instrumental bias, which introduces an offset in the measured value. Errors in sample preparation are systematic errors, which will propagate through analysis, causing uncertainty or inaccuracies through improper calibration curves. Systematic errors can be eliminated through correct sample preparation and proper use of the instrument. Poor sample preparation can also sometimes cause harm to the instrument.

  • Analytical Chemistry

    09:17
    Internal Standards

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    The goal of many chemical analyses is a quantitative analysis, where the amount of a substance in a sample is determined. In order to accurately calculate the concentration of an unknown from a sample, careful sample preparation is key. Every time a sample is handled or transferred, some of the sample can be lost. There are strategies however, for minimizing sample loss. There are also strategies for coping with sample loss and still making accurate measurements of concentration. To minimize sample loss, the ideal is to minimize the number of sample handling and transfer steps. For example, massing a solid sample directly into a flask that a solution will be made in reduces a transfer step. If it's necessary to transfer from one flask to another and a dilution is being made, then triple rinsing the glassware helps ensure all the sample is transferred. Other strategies are more specific to the sample. For example, samples that adsorb to glass, such as proteins, might better be handled in polypropylene disposable tubes. The tubes are not hydrophilic, so if a small amount of sample is to be pipetted in water, it is best to have already added the water to the tube, so the sample can be pipetted directly into the solvent. It may be better to concentrate, rather than completely dry a sample, due to losses from insolubilities aft

  • Analytical Chemistry

    11:27
    Method of Standard Addition

    Source: Laboratory of Dr. Paul Bower - Purdue University

    The method of standard additions is a quantitative analysis method, which is often used when the sample of interest has multiple components that result in matrix effects, where the additional components may either reduce or enhance the analyte absorbance signal. That results in significant errors in the analysis results.

    Standard additions are commonly used to eliminate matrix effects from a measurement, since it is assumed that the matrix affects all of the solutions equally. Additionally, it is used to correct for the chemical phase separations performed in the extraction process. The method is performed by reading the experimental (in this case fluorescent) intensity of the unknown solution and then by measuring the intensity of the unknown with varying amounts of known standard added. The data are plotted as fluorescence intensity vs. the amount of the standard added (the unknown itself, with no standard added, is plotted ON the y-axis). The least squares line intersects the x-axis at the negative of the concentration of the unknown, as shown in Figure 1. Figure 1. Graphic representation of method of standard addition.

  • Analytical Chemistry

    07:42
    Calibration Curves

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Calibration curves are used to understand the instrumental response to an analyte and predict the concentration in an unknown sample. Generally, a set of standard samples are made at various concentrations with a range than includes the unknown of interest and the instrumental response at each concentration is recorded. For more accuracy and to understand the error, the response at each concentration can be repeated so an error bar is obtained. The data are then fit with a function so that unknown concentrations can be predicted. Typically the response is linear, however, a curve can be made with other functions as long as the function is known. The calibration curve can be used to calculate the limit of detection and limit of quantitation. When making solutions for a calibration curve, each solution can be made separately. However, that can take a lot of starting material and be time consuming. Another method for making many different concentrations of a solution is to use serial dilutions. With serial dilutions, a concentrated sample is diluted down in a stepwise manner to make lower concentrations. The next sample is made from the previous dilution, and the dilution factor is often kept constant. The advantage is that only one initial solution is needed. The disadvantage is that any errors in solution making—pipetting, massing, etc.&#

  • Analytical Chemistry

    09:20
    Ultraviolet-Visible (UV-Vis) Spectroscopy

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Ultraviolet-visible (UV-Vis) spectroscopy is one of the most popular analytical techniques because it is very versatile and able to detect nearly every molecule. With UV-Vis spectroscopy, the UV-Vis light is passed through a sample and the transmittance of light by a sample is measured. From the transmittance (T), the absorbance can be calculated as A=-log (T). An absorbance spectrum is obtained that shows the absorbance of a compound at different wavelengths. The amount of absorbance at any wavelength is due to the chemical structure of the molecule. UV-Vis can be used in a qualitative manner, to identify functional groups or confirm the identity of a compound by matching the absorbance spectrum. It can also be used in a quantitative manner, as concentration of the analyte is related to the absorbance using Beer's Law. UV-Vis spectroscopy is used to quantify the amount of DNA or protein in a sample, for water analysis, and as a detector for many types of chromatography. Kinetics of chemical reactions are also measured with UV-Vis spectroscopy by taking repeated UV-Vis measurements over time. UV-Vis measurements are generally taken with a spectrophotometer. UV-Vis is also a very popular detector for other analytical techniques, such as chromatography, because it can detect many compounds. Typically, UV-Vis is not the most

  • Analytical Chemistry

    09:25
    Raman Spectroscopy for Chemical Analysis

    Source: Laboratory of Dr. Ryoichi Ishihara — Delft University of Technology

    Raman spectroscopy is a technique for analyzing vibrational and other low frequency modes in a system. In chemistry it is used to identify molecules by their Raman fingerprint. In solid-state physics it is used to characterize materials, and more specifically to investigate their crystal structure or crystallinity. Compared to other techniques for investigating the crystal structure (e.g. transmission electron microscope and x-ray diffraction) Raman micro-spectroscopy is non-destructive, generally requires no sample preparation, and can be performed on small sample volumes. For performing Raman spectroscopy a monochromatic laser is shone on a sample. If required the sample can be coated by a transparent layer which is not Raman active (e.g., SiO2) or placed in DI water. The electromagnetic radiation (typically in the near infrared, visible, or near ultraviolet range) emitted from the sample is collected, the laser wavelength is filtered out (e.g., by a notch or bandpass filter), and the resulting light is sent through a monochromator (e.g., a grating) to a CCD detector. Using this, the inelastic scattered light, originating from Raman scattering, can be captured and used to construct the Raman spectrum of the sample. In the case of Raman micro-spectroscopy the light passes through a microscope before reaching the

  • Analytical Chemistry

    07:45
    X-ray Fluorescence (XRF)

    Source: Laboratory of Dr. Lydia Finney — Argonne National Laboratory

    X-ray fluorescence is an induced, emitted radiation that can be used to generate spectroscopic information. X-ray fluorescence microscopy is a non-destructive imaging technique that uses the induced fluorescence emission of metals to identify and quantify their spatial distribution.

  • Analytical Chemistry

    09:21
    Gas Chromatography (GC) with Flame-Ionization Detection

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Gas chromatography (GC) is used to separate and detect small molecular weight compounds in the gas phase. The sample is either a gas or a liquid that is vaporized in the injection port. Typically, the compounds analyzed are less than 1,000 Da, because it is difficult to vaporize larger compounds. GC is popular for environmental monitoring and industrial applications because it is very reliable and can be run nearly continuously. GC is typically used in applications where small, volatile molecules are detected and with non-aqueous solutions. Liquid chromatography is more popular for measurements in aqueous samples and can be used to study larger molecules, because the molecules do not need to vaporize. GC is favored for nonpolar molecules while LC is more common for separating polar analytes. The mobile phase for gas chromatography is a carrier gas, typically helium because of its low molecular weight and being chemically inert. Pressure is applied and the mobile phase moves the analyte through the column. The separation is accomplished using a column coated with a stationary phase. Open tubular capillary columns are the most popular columns and have the stationary phase coated on the walls of the capillary. Stationary phases are often derivatives of polydimethylsiloxane, with 5–10% of the groups functionalized to tune the separa

  • Analytical Chemistry

    12:56
    High-Performance Liquid Chromatography (HPLC)

    Source: Dr. Paul Bower - Purdue University

    High-performance liquid chromatography (HPLC) is an important analytical method commonly used to separate and quantify components of liquid samples. In this technique, a solution (first phase) is pumped through a column that contains a packing of small porous particles with a second phase bound to the surface. The different solubilities of the sample components in the two phases cause the components to move through the column with different average velocities, thus creating a separation of these components. The pumped solution is called the mobile phase, while the phase in the column is called the stationary phase. There are several modes of liquid chromatography, depending upon the type of stationary and/or mobile phase employed. This experiment uses reversed-phase chromatography, where the stationary phase is non-polar, and the mobile phase is polar. The stationary phase to be employed is C18 hydrocarbon groups bonded to 3-µm silica particles, while the mobile phase is an aqueous buffer with a polar organic modifier (acetonitrile) added to vary its eluting strength. In this form, the silica can be used for samples that are water-soluble, providing a broad range of applications. In this experiment, the mixtures of three components frequently found in diet soft drinks (namely caffeine, benzoate, and aspartame) are separated. Seven prepared solutions contain

  • Analytical Chemistry

    08:51
    Ion-Exchange Chromatography

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Ion-exchange chromatography is a type of chromatography that separates analytes based on charge. A column is used that is filled with a charged stationary phase on a solid support, called an ion-exchange resin. Strong cation-exchange chromatography preferentially separates out cations by using a negatively-charged resin while strong anion-exchange chromatography preferentially selects out anions by using a positively-charged resin. This type of chromatography is popular for sample preparation, for example in the cleanup of proteins or nucleic acid samples. Ion-exchange chromatography is a two-step process. In the first step, the sample is loaded onto the column in a loading buffer. The binding of the charged sample to the column resin is based on ionic interactions of the resin to attract the sample of the opposite charge. Thus, charged samples of opposite polarity to the resin are strongly bound. Other molecules that are not charged or are of the opposite charge are not bound and are washed through the column. The second step is to elute the analyte that is bound to the resin. This is accomplished with a salt gradient, where the amount of salt in the buffer is slowly increased. Fractions are collected at the end of the column as the elution occurs and the purified sample of interest can be recovered in one of these fractions. Another

  • Analytical Chemistry

    08:49
    Capillary Electrophoresis (CE)

    Source: Laboratory of Dr. B. Jill Venton - University of Virginia

    Capillary electrophoresis (CE) is a separation technique that separates molecules in an electric field according to size and charge. CE is performed in a small glass tube called a capillary that is filled with an electrolyte solution. Analytes are separated due to differences in electrophoretic mobility, which varies with charge, solvent viscosity, and size. Traditional electrophoresis in gels is limited in the amount of voltage that can be applied because Joule heating effects will ruin the gel and the separation. Capillaries have a large surface area-to-volume ratio and thus dissipate heat better. Therefore, the voltages applied for a capillary electrophoresis experiment are quite large, often 10,000–20,000 V. Capillary electrophoresis is useful for high-performance separations. Compared to liquid chromatography, CE separations are often faster and more efficient. However, capillary electrophoresis works best to separate charged molecules, which is not a limitation of liquid chromatography. CE has a greater peak capacity than high-performance liquid chromatography (HPLC), meaning the separations are more efficient and more peaks can be detected. The instrumentation can be very simple. However, HPLC is more versatile and many stationary and mobile phases have been developed for different types of molecules.

  • Analytical Chemistry

    10:28
    Introduction to Mass Spectrometry

    Source: Laboratory of Dr. Khuloud Al-Jamal - King's College London

    Mass spectrometry is an analytical chemistry technique that enables the identification of unknown compounds within a sample, the quantification of known materials, the determination of the structure, and chemical properties of different molecules.

    A mass spectrometer is composed of an ionization source, an analyzer, and a detector. The process involves the ionization of chemical compounds to generate ions. When using inductively coupled plasma (ICP), samples containing elements of interest are introduced into argon plasma as aerosol droplets. The plasma dries the aerosol, dissociates the molecules, and then removes an electron from the components to be detected by the mass spectrometer. Other ionization methods such as electrospray ionization (ESI) and matrix assisted laser desorption ionization (MALDI) are used to analyze biological samples. Following the ionization procedure, ions are separated in the mass spectrometer according to their mass-to-charge ratio (m/z), and the relative abundance of each ion type is measured. Finally, the detector commonly consists in an electron multiplier where the collision of ions with a charged anode leads to a cascade of increasing number of electrons, which can be detected by an electrical circuit connected to a computer. In this video, the procedure of ICP-MS analysis will be de

  • Analytical Chemistry

    11:40
    Scanning Electron Microscopy (SEM)

    Source: Laboratory of Dr. Andrew J. Steckl — University of Cincinnati

    A scanning electron microscope, or SEM, is a powerful microscope that uses electrons to form an image. It allows for imaging of conductive samples at magnifications that cannot be achieved using traditional microscopes. Modern light microscopes can achieve a magnification of ~1,000X, while typical SEM can reach magnifications of more than 30,000X. Because the SEM doesn’t use light to create images, the resulting pictures it forms are in black and white.  Conductive samples are loaded onto the SEM’s sample stage. Once the sample chamber reaches vacuum, the user will proceed to align the electron gun in the system to the proper location. The electron gun shoots out a beam of high-energy electrons, which travel through a combination of lenses and apertures and eventually hit the sample. As the electron gun continues to shoot electrons at a precise position on the sample, secondary electrons will bounce off of the sample. These secondary electrons are identified by the detector. The signal found from the secondary electrons is amplified and sent to the monitor, creating a 3D image. This video will demonstrate SEM sample preparation, operation, and imaging capabilities.

  • Analytical Chemistry

    10:37
    Electrochemical Measurements of Supported Catalysts Using a Potentiostat/Galvanostat

    Source: Laboratory of Dr. Yuriy Román — Massachusetts Institute of Technology

    A potentiostat/galvanostat (often referred to as simply a potentiostat) is an instrument that measures current at an applied potential (potentiostatic operation) or measures potential at an applied current (galvanostatic operation) (Figure 1). It is the most commonly used instrument in the electrochemical characterization of anode and cathode materials for fuel cells, electrolyzers, batteries, and supercapacitors. Conventionally, these anode and cathode materials are interfaced with a potentiostat via a three-electrode electrochemical cell. The electrode leads from the potentiostat are connected to the reference electrode, the counter electrode (often called the auxiliary electrode), and the working electrode (which contains the test material of interest). The electrochemical cell is then filled with a high ionic strength electrolyte solution, such as an acidic, alkaline, or salt solution. The media for this high ionic strength solution is typically aqueous; however, for applications necessitating higher operating cell potential windows, such as batteries and supercapacitors, non-aqueous media is often used. The cell media is degassed with an inert gas (to prevent unwanted side reactions) or with a test gas (if the test reaction involves a gas at one of the electrodes). Alternatively, a salt bridge or membran

  • Analytical Chemistry

    08:37
    Cyclic Voltammetry (CV)

    Source: Laboratory of Dr. Kayla Green — Texas Christian University

    A Cyclic Voltammetry (CV) experiment involves the scan of a range of potential voltages while measuring current. In the CV experiment, the potential of an immersed, stationary electrode is scanned from a predetermined starting potential to a final value (called the switching potential) and then the reverse scan is obtained. This gives a 'cyclic' sweep of potentials and the current vs. potential curve derived from the data is called a cyclic voltammogram. The first sweep is called the 'forward scan' and the return wave is called the 'reverse scan'. The potential extremes are termed the 'scan window'. The magnitude of reduction and oxidation currents and the shape of the voltammograms are highly dependent on analyte concentration, scan rates, and experimental conditions. By varying these factors, cyclic voltammetry can yield information regarding the stability of transition metal oxidation state in the complexed form, reversibility of electron transfer reactions, and information regarding reactivity. This video will explain the basic setup for a cyclic voltammetry experiment including analyte preparation and setting up the electrochemical cell. A simple cyclic voltammetry experiment will be presented.

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