Carbohydrates

• Generalities (this section)
  • Role of carbohydrates
  • Classification
  • Sample preparation
• Next sections:
  • Analysis of mono- and oligosaccharides
    • Chromatographic and electrophoretic methods
    • Chemical methods
    • Enzymatic method
    • Physical methods
    • Immuno assays
  • Analysis of polysaccharides and fiber
    • Analysis of starch
    • Analysis of fiber

Role of carbohydrates

The carbohydrates, which include starches, sugars, cellulose and gums containing only the elements carbon, hydrogen and oxygen, are usually the cheapest source of energy in foods and feeds. Fish and shrimp, however, vary in their ability to digest carbohydrate effectively. Many fish appear to be able to utilize simple carbohydrates, such as sugars, more effectively than complex starches; the reverse appears to be true for shrimp and prawns.
This observation may be confused by the beneficial effect that carbohydrates tend to have on the structural integrity of the feed, caused by the binding quality of starches. Carnivorous fish such as salmon and trout and, particularly, marine fish are not efficient converters of carbohydrate. Channel catfish, like shrimp, appear to be able to utilize complex carbohydrates more readily than simple sugars. Channel catfish and carp can utilize quite high levels of dietary carbohydrate. The natural diet of grass carp is very high in this component. The topic of the energy value of dietary carbohydrate is dealt with separately.

Some carbohydrates are normally regarded as indigestible. These are reported separately in the tables of feed composition as 'fibre' or 'crude fibre' . Fibre includes substances such as celluloses (from plants), lignin, chitin, etc. Many fish do not have the enzyme cellulase which is necessary for the digestion of cellulose, and fibre is usually regarded as unavailable as an energy source. At small levels, however, it may aid pelletability. Cellulase however is produced by the gut bacteria of many fish, as is chitinase in crustacea, and herbivorous fish are able to digest fibre.

Classification

Monosaccharides

Monosaccharides are water-soluble crystalline compounds. They are aliphatic aldehydes or ketones which contain one carbonyl group and one or more hydroxyl groups. Most natural monosachharides have either five (pentoses) or six (hexoses) carbon atoms. Commonly occurring hexoses in foods are glucose, fructose and galactose, whilst commonly occurring pentoses are arabinose and xylose. The reactive centers of monosaccharides are the carbonyl and hydroxyl groups.

Oligosaccharides

These are relatively low molecular weight polymers of monosaccharides (< 20) that are covalently bonded through glycosidic linkages. Disaccharides consist of two monomers, whereas trisaccharides consist of three. Oligosaccharides containing glucose, fructose and galactose monomers are the most commonly occurring in foods.

Polysaccharides

The majority of carbohydrates found in nature are present as polysaccharides. Polysaccharides are high molecular weight polymers of monosaccharides (> 20). Polysaccharides containing all the same monosaccharides are called homopolysaccharides (e.g., starch, cellulose and glycogen are formed from only glucose), whereas those which contain more than one type of monomer are known as heteropolysaccharides (e.g., pectin, hemicellulose and gums).

Sample preparation

One of the most commonly used methods of extracting low molecular weight carbohydrates from foods is to boil a defatted sample with an 80% alcohol solution. Monosaccharides and oligosaccharides are soluble in alcoholic solutions, whereas proteins, polysaccharides and dietary fiber are insoluble. The soluble components can be separated from the insoluble components by filtering the boiled solution and collecting the filtrate (the part which passes through the filter) and the retentante (the part retained by the filter). These two fractions can then be dried and weighed to determine their concentrations. In addition, to monosaccharides and oligosaccharides various other small molecules may also be present in the alcoholic extract that could interfere with the subsequent analysis e.g., amino acids, organic acids, pigments, vitamins, minerals etc. It is usually necessary to remove these components prior to carrying out a carbohydrate analysis. This is commonly achieved by treating the solution with clarifying agents or by passing it through one or more ion-exchange resins.

• Clarifying agents: Water extracts of many foods contain substances that are colored or produce turbidity, and thus interfere with spectroscopic analysis or endpoint determinations. For this reason solutions are usually clarified prior to analysis. The most commonly used clarifying agents are heavy metal salts (such as lead acetate) which form insoluble complexes with interfering substances that can be removed by filtration or centrifugation. However, it is important that the clarifying agent does not precipitate any of the carbohydrates from solution as this would cause an underestimation of the carbohydrate content.
• Ion-exchange: Many monosaccharides and oligosaccharides are polar non-charged molecules and can therefore be separated from charged molecules by passing samples through ion-exchange columns. By using a combination of a positively and a negatively charged column it is possible to remove most charged contaminants. Non-polar molecules can be removed by passing a solution through a column with a non-polar stationary phase. Thus proteins, amino acids, organic acids, minerals and hydrophobic compounds can be separated from the carbohydrates prior to analysis

Prior to analysis, the alcohol can be removed from the solutions by evaporation under vacuum so that an aqueous solution of sugars remains.