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Chromatography is a vital tool used to determine the composition of a mixture. It plays a vital role in different industries: pharmaceuticals, clinical trials, environmental and chemical safety, food and beverage, drug testing, forensics, petroleum creation, and molecular biology.
Chromatography is a biophysical technique that creates separation, identification, and purification of a mixture to then analyze the components of the mixture. To do so, chromatography relies specifically on two different phases, the mobile and stationary phases. To separate the components of the mixture, it is dissolved in a solvent during the mobile phase. The mobile phase then carries the mixture through the stationary phase, where the components of the mixture interact with the stationary phase. The components all interact with the stationary phase at their own pace, creating the separation of the components located within the mixture. The components of the mixture will move differently through this phase due to differences in absorption, partition, and molecular weights.
Chromatography was developed in the early 20th century by Mikhail Tsvet. In 1901, Tsvet introduced the technique while working on separating plant pigments such as chlorophyll and carotenoids. He used a column packed with calcium carbonate as the stationary phase and a solvent as the mobile phase, observing how different pigments separated into distinct colored bands — hence the name "chromatography," which comes from the Greek words chroma (color) and graphein (to write).
Despite its early invention, chromatography did not gain widespread attention until the mid-20th century. During the 1930s and 1940s, advances in paper chromatography by scientists such as Martin and Synge improved the method and demonstrated its usefulness for separating amino acids and other biochemical substances. Their work was so significant that they received the Nobel Prize in Chemistry in 1952.
Later developments introduced new types of chromatography, including gas chromatography (GC) in the 1950s and high-performance liquid chromatography (HPLC) in the 1960s. These advancements allowed for faster, more precise, and more sensitive separations, expanding the technique’s applications across chemistry, biology, medicine, and environmental science.
Today, chromatography remains an essential analytical tool with continuous innovations improving its speed, resolution, and applicability to complex mixtures.
Chromatography can be done in many ways to adhere to the different substances being analyzed. Some of the most common types of chromatography are the following:
Gas Chromatography (GC) is a method of separating and analyzing compounds that can be vaporized without decomposition. It involves injecting a sample into a stream of inert gas (called the carrier gas), which transports the sample through a column containing a stationary phase. As the mixture travels through the column, its components interact differently with the stationary phase, causing them to separate. Each substance exits (or elutes) from the column at a different time, allowing for identification and quantification.
The basic principle of gas chromatography involves the distribution of components between two phases: a mobile phase, which is an inert carrier gas (such as helium, nitrogen, or hydrogen), and a stationary phase, which is a liquid or polymer coated onto the inner wall of a narrow capillary column or packed into a traditional column. When a small sample is injected into the system, it is immediately vaporized and carried by the mobile phase into the column.
Liquid Chromatography (LC) is an analytical technique used to separate, identify, and quantify components in a mixture based on their interactions with two phases: a liquid mobile phase and a solid or liquid stationary phase. Unlike gas chromatography, which is limited to volatile substances, liquid chromatography can be used for a wide range of compounds, including large, polar, or thermally unstable molecules.
The principle of LC is based on the different affinities of substances for the mobile and stationary phases. When a mixture is introduced into the system, each component interacts with the stationary phase differently. Some components move faster through the column, while others are retained longer. This difference in travel time, or retention time, allows for the separation of the mixture into its individual components.
Thin-Layer Chromatography (TLC) is a simple, fast, and inexpensive method used in analytical chemistry to separate and identify components of a mixture. It is particularly useful for monitoring chemical reactions, testing purity, and identifying substances in fields such as organic chemistry, pharmaceuticals, and forensic science.
The technique is based on the principle of adsorption chromatography, where components of a mixture move across a stationary phase at different rates due to differences in their physical and chemical properties. The stationary phase is a thin layer of an adsorbent material—usually silica gel, alumina, or cellulose—coated onto a flat, inert surface such as a glass, plastic, or aluminum plate. The mobile phase is a suitable liquid solvent or a mixture of solvents that travels upward through the stationary phase by capillary action.
Column Chromatography is a widely used liquid chromatography technique that separates and purifies components of a mixture based on their differing interactions with a stationary phase and a mobile phase. It is commonly applied in organic chemistry, biochemistry, and pharmaceutical research to isolate pure compounds from complex mixtures.
The method relies on the principle of differential adsorption. The stationary phase, usually a solid adsorbent such as silica gel or alumina, is packed into a vertical glass column. A liquid solvent or mixture of solvents, known as the mobile phase, flows through this stationary phase. When the sample mixture is introduced at the top of the column, the components move through the column at different rates depending on their affinity to the stationary phase. Compounds that interact more strongly with the stationary phase move more slowly, while those with weaker interactions travel faster, resulting in their separation.
Chromatography plays an important role in the safety of pharmaceuticals. Pharmaceutical companies use chromatography to quantify and analyze compounds for contaminants. For example, chiral compounds have two different forms due to their atoms differing slightly in space. One form of chiral compounds is known to be toxic. Chromatography can ensure that the safe form is separate from the dangerous form of the chiral compound.
Vaccination creation is also an application of chromatography. Chromatography can be used to determine which antibodies are the best for fighting and neutralizing certain diseases.
In the fight against the Ebola virus outbreak, which tragically claimed over 11,000 lives, scientists utilized chromatography to develop the experimental immunization Zmapp. This critical process involved identifying the most effective antibodies capable of neutralizing the deadly virus. By isolating and analyzing these antibodies, researchers were able to make significant strides in combating the outbreak.
Through this detailed approach, chromatography not only facilitated the development of vaccines but also provided essential insights into the antibody interactions necessary for effective immunization strategies.
Quality control within the food and beverage industry can be enacted through chromatography.
In the food industry, chromatography is used to separate and analyze additives, vitamins, proteins, amino acids, and other nutritional compounds in food items. Chromatography can also be used to determine expiration dates by distinguishing the number of organic acids present as well as to detect any harmful toxins that may have been added to the food item.
Chromatography, specifically high-performance liquid chromatography combined with mass spectrometry (HPLC-MS), emerged as a crucial tool. This advanced technique provided the precision needed to identify the true composition of meat products. By pinpointing whether the labeled beef was indeed beef, horse, or a mixture of various ingredients, chromatography offered a reliable solution to food testing challenges.
The chemical industry must adhere to numerous environmental safety precautions. Perfluoroalkyl substances, also known as PFAS, have become a persistent threat to the human body and the environment. PFAS can be found in items such as protective coatings on shoes and other fabrics, electronics, and even firefighting foams. While these substances benefit products by making them extremely durable, they pose an environmental concern as they continue to accumulate. PFAS in our drinking water can also lead to damaging health concerns such as reproductive and developmental setbacks. By using solid-phase extraction, liquid chromatography, and mass spectrometry, we can detect PFAS in the environment and our drinking water, even at very low limits.
Similar to security precautions, Forensics is a unique application of chromatography. Gas chromatography can be used for more in-depth forensics procedures, for example, crime scene analysis to test evidence such as blood, hair, and fabric samples to further understand what may have happened at the scene. Chromatography is massively important to forensic pathology work. Gas chromatography is widely used to identify the types of fluids and compounds that exist in a body postmortem. In such cases, a possible cause of death and motive can be determined based on finding drugs, alcohol, or toxic substances in the body. Another unique form of forensics that can be assisted by chromatography is arson verification. By using chromatography in arson verification, it’s possible to identify flammable substances in fire debris to determine the exact substance that created the fire.
One of the most complex uses of chromatography is molecular biology studies. Hybrid techniques between electrochemistry (EC) and mass spectrometry with chromatography are often applied to studies of proteins, peptides, and nucleic acids. This combination is largely used for metabolomics such as biotransformation reactions like oxidative reactions and proteomics such as the purification of plasma proteins, hormones, and antibodies. Chromatography in nucleic acid research plays a role in accelerating the identification process of nucleobases, nucleotides, and nucleosides, as well as identifying their oxidization process.
In essence, chromatography is not just a scientific technique; it's a cornerstone of modern industry practices that ensures the quality and safety of countless products and services integral to everyday life.
From safeguarding pharmaceutical drugs and ensuring food and beverage quality to advancing environmental safety, forensics, and molecular biology, chromatography's versatility and precision make it a cornerstone of modern technology and research. Its ability to separate, identify, and analyze complex mixtures ensures the integrity of countless products and processes that touch our daily lives.
If you're in need of high-quality chromatography supplies and equipment to support your industry or research, we can help you. We provide a wide range of reliable products to help you achieve precision and excellence in your chromatography processes. Explore our catalog to find the tools you need!
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