In chemical analysis, chromatography is a laboratory technique for the separation of a mixture into its components. The mixture is dissolved in a fluid solvent called the mobile phase, which carries it through a system on which a material called the stationary phase is fixed. Because the different constituents of the mixture tend to have different affinities for the stationary phase and are retained for different lengths of time depending on their interactions with its surface sites, the constituents travel at different apparent velocities in the mobile fluid, causing them to separate. The separation is based on the differential partitioning between the mobile and the stationary phases. Subtle differences in a compound's partition coefficient result in differential retention on the stationary phase and thus affect the separation.
High-performance liquid chromatography (HPLC), formerly referred to as high-pressure liquid chromatography, is a technique in analytical chemistry used to separate, identify, and quantify specific components in mixtures. The mixtures can originate from food, chemicals, pharmaceuticals, biological, environmental and agriculture, etc, which have been dissolved into liquid solutions.
PFP or PfP may stand for:
A protecting group or protective group is introduced into a molecule by chemical modification of a functional group to obtain chemoselectivity in a subsequent chemical reaction. It plays an important role in multistep organic synthesis.
Richard Laurence Millington Synge FRS FRSE FRIC FRSC MRIA was a British biochemist, and shared the 1952 Nobel Prize in Chemistry for the invention of partition chromatography with Archer Martin.
In biochemistry, biotinylation is the process of covalently attaching biotin to a protein, nucleic acid or other molecule. Biotinylation is rapid, specific and is unlikely to disturb the natural function of the molecule due to the small size of biotin. Biotin binds to streptavidin and avidin with an extremely high affinity, fast on-rate, and high specificity, and these interactions are exploited in many areas of biotechnology to isolate biotinylated molecules of interest. Biotin-binding to streptavidin and avidin is resistant to extremes of heat, pH and proteolysis, making capture of biotinylated molecules possible in a wide variety of environments. Also, multiple biotin molecules can be conjugated to a protein of interest, which allows binding of multiple streptavidin, avidin or neutravidin protein molecules and increases the sensitivity of detection of the protein of interest. There is a large number of biotinylation reagents available that exploit the wide range of possible labelling methods. Due to the strong affinity between biotin and streptavidin, the purification of biotinylated proteins has been a widely used approach to identify protein-protein interactions and post-translational events such as ubiquitylation in molecular biology.
Protein sequencing is the practical process of determining the amino acid sequence of all or part of a protein or peptide. This may serve to identify the protein or characterize its post-translational modifications. Typically, partial sequencing of a protein provides sufficient information to identify it with reference to databases of protein sequences derived from the conceptual translation of genes.
In organic chemistry, peptide synthesis is the production of peptides, compounds where multiple amino acids are linked via amide bonds, also known as peptide bonds. Peptides are chemically synthesized by the condensation reaction of the carboxyl group of one amino acid to the amino group of another. Protecting group strategies are usually necessary to prevent undesirable side reactions with the various amino acid side chains. Chemical peptide synthesis most commonly starts at the carboxyl end of the peptide (C-terminus), and proceeds toward the amino-terminus (N-terminus). Protein biosynthesis in living organisms occurs in the opposite direction.
Affinity chromatography is a method of separating a biomolecule from a mixture, based on a highly specific macromolecular binding interaction between the biomolecule and another substance. The specific type of binding interaction depends on the biomolecule of interest; antigen and antibody, enzyme and substrate, receptor and ligand, or protein and nucleic acid binding interactions are frequently exploited for isolation of various biomolecules. Affinity chromatography is useful for its high selectivity and resolution of separation, compared to other chromatographic methods.
Liquid chromatography–mass spectrometry (LC–MS) is an analytical chemistry technique that combines the physical separation capabilities of liquid chromatography with the mass analysis capabilities of mass spectrometry (MS). Coupled chromatography – MS systems are popular in chemical analysis because the individual capabilities of each technique are enhanced synergistically. While liquid chromatography separates mixtures with multiple components, mass spectrometry provides spectral information that may help to identify each separated component. MS is not only sensitive, but provides selective detection, relieving the need for complete chromatographic separation. LC–MS is also appropriate for metabolomics because of its good coverage of a wide range of chemicals. This tandem technique can be used to analyze biochemical, organic, and inorganic compounds commonly found in complex samples of environmental and biological origin. Therefore, LC–MS may be applied in a wide range of sectors including biotechnology, environment monitoring, food processing, and pharmaceutical, agrochemical, and cosmetic industries. Since the early 2000s, LC–MS has also begun to be used in clinical applications.
Deamidation is a chemical reaction in which an amide functional group in the side chain of the amino acids asparagine or glutamine is removed or converted to another functional group. Typically, asparagine is converted to aspartic acid or isoaspartic acid. Glutamine is converted to glutamic acid or pyroglutamic acid (5-oxoproline). In a protein or peptide, these reactions are important because they may alter its structure, stability or function and may lead to protein degradation. The net chemical change is the addition of a water group and removal of an ammonia group, which corresponds to a +1 (0.98402) Da mass increase. Although deamidation occurs on glutamine, glycosylated asparagine and other amides, these are negligible under typical proteolysis conditions.
Reversed-phase liquid chromatography (RP-LC) is a mode of liquid chromatography in which non-polar stationary phase and polar mobile phases are used for the separation of organic compounds. The vast majority of separations and analyses using high-performance liquid chromatography (HPLC) in recent years are done using the reversed phase mode. In the reversed phase mode, the sample components are retained in the system the more hydrophobic they are.
Immunoproteomics is the study of large sets of proteins (proteomics) involved in the immune response.
Protein mass spectrometry refers to the application of mass spectrometry to the study of proteins. Mass spectrometry is an important method for the accurate mass determination and characterization of proteins, and a variety of methods and instrumentations have been developed for its many uses. Its applications include the identification of proteins and their post-translational modifications, the elucidation of protein complexes, their subunits and functional interactions, as well as the global measurement of proteins in proteomics. It can also be used to localize proteins to the various organelles, and determine the interactions between different proteins as well as with membrane lipids.
In analytical and organic chemistry, elution is the process of extracting one material from another by washing with a solvent; as in washing of loaded ion-exchange resins to remove captured ions.
Two-dimensional chromatography is a type of chromatographic technique in which the injected sample is separated by passing through two different separation stages. Two different chromatographic columns are connected in sequence, and the effluent from the first system is transferred onto the second column. Typically the second column has a different separation mechanism, so that bands that are poorly resolved from the first column may be completely separated in the second column. Alternately, the two columns might run at different temperatures. During the second stage of separation the rate at which the separation occurs must be faster than the first stage, since there is still only a single detector. The plane surface is amenable to sequential development in two directions using two different solvents.
Glycopeptides are peptides that contain carbohydrate moieties (glycans) covalently attached to the side chains of the amino acid residues that constitute the peptide.
Tetrafluorophenyl (TFP) ester chemistry is typically used to attach fluorophores or haptens to the primary amines of biomolecules. They produce the same amide bonds that are formed through conjugation with other amine-reactive groups, such as succinimidyl esters, but TFP esters are less susceptible to spontaneous hydrolysis during conjugation reactions. TFP esters are stable for several hours at basic pH, far outlasting succinimidyl esters.
Pentafluorophenol is the organofluorine compound with the formula C6F5OH. This is the perfluorinated analogue of phenol. It is a white odorless solid that melts just above room temperature. With a pKa of 5.5, it is one of the most acidic phenols.
Poly(pentafluorophenyl acrylate) (variously abbreviated PPFPA, PolyPFPA, PPfpA, or PolyPfpA) is a highly fluorinated polymer. It features the pentafluorophenyl ester functionality, from which its properties and applications result. It is most commonly used in post-polymerization modification to synthesize functional polyacrylamides or polyacrylates. As such, it is advantageous to poly(N-acryloyl succinimide) due to its broader solubility in organic solvents as well as its higher stability towards hydrolysis.
