As methods for identifying small molecules become more accurate and sensitive, metabolic research has become more global. The metabolome consists of all the small molecules and metabolic intermediates within a system, such as a cell or whole organism, at one particular time. Understanding the metabolome is complex because small metabolites affect many other components of a cell. Metabolites flow in a complex network and form many different transient complexes. The network of metabolites may be compared to city streets. At each corner, a decision on which route to take must be made, and such decisions continue until the person reaches the final destination. Each metabolite molecule follows a specific pathway, often with several potential branches, and at each junction, a decision is made before moving on to the next step. Characterizing the metabolome under particular conditions is known as metabolic fingerprinting.
Several techniques that involve separating and/or identifying small metabolites have made metabolomics possible. Nuclear magnetic resonance (NMR) of extracts from cells grown with 13C-glucose has allowed simultaneous measurement of multiple metabolic intermediates. Metabolites have also been identified by thin layer chromatography after growth in 14C-glucose. These methods are not very sensitive, and some metabolites may not be separated or identified.
Mass spectroscopy offers the best way to analyze whole metabolomes. The technique can identify many different metabolites (even novel ones) and is extremely sensitive. Mass spectroscopy can determine the exact molecular formula for a compound, so every metabolite can be identified. Even if isomers exist, the fragmentation pattern will be different although the molecular formula is the same.
The use of mass spectroscopy is often combined with other methods to simplify analysis. Different types of chromatography can be used to separate the metabolites before analysis by mass spectroscopy. Obviously, using HPLC will separate the complex cellular extract into different fractions, which can then be analyzed by mass spectroscopy.
Metabolomics is especially valuable in studying plants, because metabolites affect the pigments, scents, flavors, and nutrient content. These are all commercially important traits, and using mass spectroscopy to analyze these metabolites will aid in developing better-tasting and fresher produce. For example, in strawberries, there are 7000 metabolites that can be identified by mass spectroscopy (Fig. 9.29). Comparing white and red strawberries has identified which of the metabolite peaks in the mass spectrum corresponds to the intermediates in pigment synthesis.
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