# Insight on Extraction and Preservation of Biological Activity of Cereal \(\beta\)-D-Glucans
## 4 Methods of Extraction and Purification of Cereal \(\beta\)-D-Glucans
### Dry Technologies
However, the concentrated fractions with this approach are not suitable to be integrated with all foods, such as liquid matrices, because high quantities would be needed to obtain significant health benefits, and this would result in the impairment of the organoleptic qualities of the product or that it does not contain enough \(\beta\)-D-glucans to qualify as a functional food [29]. The extractability of \(\beta\)-D-glucans depends on particle size, pH, temperature, extraction time, and solvent and solute ratio. Generally, in wet extraction, the \(\beta\)-D-glucans present in barley/oat/wheat flour are solubilized in water, acidified water, or an alkaline solution. As highlighted in the following sections, the most used method involves extraction in an alkaline solution, as it is the best method to preserve the structure of \(\beta\)-D-glucans, as well as allow the removal of starch and other insoluble fibrous fractions. Proteins can be removed by isoelectric precipitation and subsequent centrifugation. Enzymatic treatments with thermostable protease, \(\alpha\)-amylase, are generally performed to hydrolyze proteins and starch bound to fibrous particles to maximize the purity of the extract. Subsequently, fibrous particulates rich in starch and protein-deprived \(\beta\)-D-glucans are recovered by precipitation with ethanol and centrifugation. Ethanol also inhibits some indigenous enzymes and removes dissociated sugars, proteins, and some nonpolar compounds [4,9]. Following this crude extraction, \(\beta\)-D-glucans can be purified further using freezing-thawing methods and alcohol in order to remove starch, protein, fat, and other residual unwanted species [9]. During extraction and purification, an additional phase of \(\beta\)-D-glucanase inactivation should be employed. This enzyme is responsible for the hydrolysis of \(\beta\)-D-glucans, which causes an alteration of the structure, MW, and viscosity of \(\beta\)-D-glucans; consequently, their cholesterol-lowering activity and glucose modulation is reduced. Enzymatic inactivation  
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Figure 3: Scheme for the extraction and purification of \(\beta\)-D-glucans, adapted from [38]. may be possible by autoclave, HCl, trichloroacetic acid, ethanol precipitation, or heating. With alkaline extraction with NaOH, the fraction of arabinoxins (AXs) is also coextracted with \(\beta\)-D-glucan. The percentage of AXs and \(\beta\)-D-glucans coextracted by alkaline extraction with NaOH depends on the cereal source and fraction considered [37]. For example, in the starch endosperm of barley, these two fibers are present in cell walls in these percentages: 70% (1\(\rightarrow\)3, 1\(\rightarrow\)4)-\(\beta\)-D-glucans and 20% AXs. However, in the aleurone cell walls of barley, the situation is the opposite [38]. In contrast, wheat endosperm cell wall polysaccharides comprise about 70% AXs and 20% (1\(\rightarrow\)3, 1\(\rightarrow\)4)-\(\beta\)-D-glucans [39]. Therefore, selecting the fractions and origin of cereal byproducts to extract and isolate \(\beta\)-D-glucans is crucial. Table 2 shows the amount of AXs and glucans (mainly \(\beta\)-D-glucans) in cereal residues and byproducts [38]. Studies on possible molecular interactions between AXs and \(\beta\)-D-glucans indicate a spontaneous and intermolecular solid association between unsubstituted regions of xylan chains and cellulose-like 1,4-\(\beta\)-linked fragments from the \(\beta\)-D-glucan chains. These associations are thought to be based on hydrogen bonds. These noncovalent associations, as well as the extent of cross-interactions between \(\beta\)-D-glucans and AXs in plant cell walls, could contribute to the poor water extractability, solubility, and enzymatic indigestibility of these polysaccharides [37; 40; 41]. Since solubility, which is linked to viscosity, is the main functional and technological characteristic of cereal \(\beta\)-D-glucans, this is a problem.