Isotopic labeling (or isotopic labelling) is a technique used to track the passage of an isotope, or an atom with a variation, through a reaction, metabolic pathway, or cell. The reactant is 'labeled' by replacing specific atoms by their isotope. The reactant is then allowed to undergo the reaction. The position of the isotopes in the products is measured to determine the sequence the isotopic atom followed in the reaction or the cell's metabolic pathway.
In isotopic labeling, there are multiple ways to detect the presence of labeling isotopes; through their mass, vibrational mode, or radioactive decay. Mass spectrometry or nuclear magnetic resonance detects the difference in an isotope's mass, while Infrared spectroscopy detects the difference in the isotope's vibrational modes. The radioactive decay can be detected through an ionization chamber or autoradiographs of gels.
An example of the use of isotopic labeling is the study of phenol (C6H5OH) in water by replacing common hydrogen (protium) with deuterium (deuterium labeling). Upon adding phenol to deuterated water (water containing D2O in addition to the usual H2O), the substitution of deuterium for the hydrogen is observed in phenol's hydroxyl group (resulting in C6H5OD), indicating that phenol readily undergoes hydrogen-exchange reactions with water. Only the hydroxyl group was affected, indicating that the other 5 hydrogen atoms did not participate in these exchange reactions.
[edit] Isotopic tracer
An isotopic tracer, (also "isotopic marker" or "isotopic label"), is used in chemistry and biochemistry to help understand chemical reactions and interactions. In this technique, one or more of the atoms of the molecule of interest is substituted for an atom of the same chemical element, but of a different isotope (like a radioactive isotope used in radioactive tracing). Because the labeled atom has the same number of protons, it will behave in almost exactly the same way as its unlabeled counterpart and, with few exceptions, will not interfere with the reaction under investigation. The difference in the number of neutrons, however, means that it can be detected separately from the other atoms of the same element.
Nuclear magnetic resonance (NMR) and mass spectrometry (MS) are used to investigate the mechanisms of chemical reactions. NMR and MS detects isotopic differences, which allows information about the position of the labeled atoms in the products' structure to be determined. With information on the positioning of the isotopic atoms in the products, the reaction pathway the initial metabolites utilize to convert into the products can be determined. Radioactive isotopes can be tested using the autoradiographs of gels in gel electrophoresis. The radiation emitted by compounds containing the radioactive isotopes darkens a piece of photographic film, recording the position of the labeled compounds relative to one another in the gel.
Isotope tracers are commonly used in the form of isotope ratios. By studying the ratio between two isotopes of the same element, we avoid effects involving the overall abundance of the element, which usually swamp the much smaller variations in isotopic abundances. Isotopic tracers are some of the most important tools in geology because they can be used to understand complex mixing processes in earth systems. Further discussion of the application of isotopic tracers in geology is covered under the heading of isotope geochemistry.
Isotopic tracers are usually subdivided into two categories: stable isotope tracers and radiogenic isotope tracers. Stable isotope tracers involve only non-radiogenic isotopes and usually are mass-dependent. In theory, any element with two stable isotopes can be used as an isotopic tracer. However, the most commonly used stable isotope tracers involve relatively light isotopes, which readily undergo fractionation in natural systems. See also isotopic signature. A radiogenic isotope tracer [1] involves an isotope produced by radioactive decay, which is usually in a ratio with a non-radiogenic isotope (whose abundance in the earth does not vary due to radioactive decay).
[edit] Stable isotope labeling
Stable isotope labeling involves the use of non-radioactive isotopes that can act as a tracers used to model several chemical and biochemical systems. The chosen isotope can act as a label on that compound that can be identified through nuclear magnetic resonance(NMR) and mass spectroscopy(MS). Some of the most common stable isotopes are 2H, 13C, and 15N, which can further be produced into NMR solvents, amino acids, nucleic acids, lipids, common metabolites and cell growth media.[3] The compounds produced using stable isotopes are either specified by the percentage of labeled isotopes (i.e. 30% uniformly labeled 13C glucose contains a mixture that is 30% labeled with 13 carbon isotope and 70% naturally labeled carbon) or by the specifically labeled carbon positions on the compound (i.e. 1-13C glucose which is labeled at the first carbon position of glucose).
A network of reactions adopted from the glycolysis pathway and the pentose phosphate pathway is shown in which the labeled carbon isotope rearranges to different carbon positions throughout the network of reactions. The network starts with fructose 6-phosphate (F6P), which has 6 carbon atoms with a label 13C at carbon position 1 and 2. 1,2-13C F6P becomes two glyceraldehyde 3-phosphate (T3P), one 2,3-13C T3P and one unlabeled T3P. The 2,3-13C T3P can now be reacted with sedoheptulose 7-phosphate (S7P) to form an unlabeled erythrose 4-phosphate(E4P) and a 5,6-13C F6P. The unlabeled T3P will react with the S7P to synthesize unlabeled products. [2] The figure demonstrates the use of stable isotope labeling to discover the carbon atom rearrangement through reactions using position specific labeled compounds.
