4.1. Structure and properties

The first report on galactomannan structure was probably the one made by Nadelman in the last century [28]. This author reported the presence of galactose and mannose in several legume species and also noticed the property of galactomannans as viscous substances.

With the development of the techniques of linkage determination by methylation analysis, mainly during the second half of the 20th century, the structures of galactomannans from several sources were determined. It is now clear that the majority of the molecules found in plants are composed of a linear backbone of P-(l —»4)-linked D-mannose residues to which single units of D-galactose residues are attached by a-(l-»6)-linkages.

Depending on the degree of branching with galactose, galactomannans can be easily extracted with hot water for a few hours and then precipitated by addition of alcohol, methanol or ethanol being the most commonly used ones. Upon resuspension in water, it solubilises and after freeze drying, usually a fluffy white cotton-like material can be obtained.

Depending on the source, different degrees of contamination with other cell wall polysaccharides may occur [29] and further purification can be obtained only after addition of Fehling solution, which precipitates galactomannans forming a blue complex. After washing and acidification (usually with HC1), this complex is broken and a polymer composed of mannose and galactose only is solubilised. As galactomannans are typical of the legume seed endosperm (Figure 2), they can also be obtained in a reasonably pure state upon extraction with water after endosperm is separated from the embryo. This can be achieved either by braking the dry seeds and taking advantage of the different degrees of hardness of different seed parts [1,2, 30] or by boiling seeds in water or alcohol until imbibition is sufficient for manual separation of seed parts. In seeds like fenugreek for example, the latter method is a great advantage since endosperm cells have very little cytoplasm left after maturation and separating it from embryo is an important step of purification. However, this should not be taken as a rule, since seed anatomy of legume endosperms appears to be very variable (see below) and it can be a good help checking the anatomy of the seed before extraction (Figure 2D).

Galactomannans are cell wall polysaccharides and as such it is expected that either in muro or during extraction, a certain degree of interaction with other polysaccharides is possible. As a result, alkali extraction of galactomannans usually lead to contamination with other cross linking glycans (hemicelluloses). As mentioned above, Fehling's reagent can be used for purification and another option is reprecipitation with alcohol, but both methods lead to considerable loss of yield. The former possibly by breaking down the polymer into smaller molecules which can not be precipitated with alcohol.

As the number of legume species studied increases and the techniques for analysis of the fine structure of polysaccharides improve, some variations of the general structure of galactomannans were found such as p-(l —>2) or p-(l-»3) linkages in the main chain [31, 32 respectively), oc-(l-»2) linked branches with galactose and short chains with ot-(l->6) linked galactoses. A detailed review on legume seed galactomannan structure and the methods for structural determination (especially Nuclear Magnetic Resonance) have been discussed in a recent work by Scherbukin and Anulov [33].

The structure of the galactomannan of some legume species was studied by McCleary in 1979 [34] who used purified fungal endo-p-mannanases and plant a-galactosidases to

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