96
stirring. Then the supernatant and residue is separated by an exhaustive centrifugation process. The residue material still contains β-glucan, thus to completely separate β-glucan, it is resuspended twice in the alkaline media under the same conditions
of temperature. All of the alkali extracts is also precipitated using ammonium sulfate by keeping the material overnight at a temperature of 4 °C or below followed
by use of centrifugation at high speed under low temperature to recover β-glucan.
This technique is highly effective to recover higher amounts of β-glucan from the
mushroom cell wall along with high levels of purity can be achieved (Nguyen
et al. 1998).
Using alkali as a solvent, β- glucan extraction from mycelium of P. chrysogenum
has also been reported; the mycelium was first freeze-dried and then ground into
powder form. Then the material was treated with alkaline solution for the extraction
of β-glucan. Maximum recovery can be obtained at a concentration of 1 M NaOH
at 40 °C as NaOH is a more effective alkali in this form. β-glucan can be extracted
by the treatment of freeze-dried powder of mycelia with the said concentration of
alkali. β-glucan contains protein impurities, thus it must be separated by the sevage
method that works at low pH. After protein has been excluded, the pH of the media
is again adjusted toward neutral for separating β-glucan. Further purification is also
obtained through dialysis against distilled water (Wang et al. 2007).
Acidic Extraction
Several reports are available on various extraction methods for β-glucan however;
extraction under acidic conditions has been reported only by a few workers.
Ahluwalia and Ellis (1984) have focussed the determination of β-glucan and starch
in barley and its malt. They adopted a simple and quick method for the extraction of
β-glucan and starch by using perchloric acid (50 mM). Then the extract was directly
used without the drying process to determine the β-glucan and starch contents
within the extract solution. For β-glucan determination, the extract was hydrolyzed
to glucose by incubating with sodium acetate buffer and “treated cellulose.” Glucose
released was measured enzymatically using glucose-6-phosphate dehydrogenase
and results were expressed as β-glucan (%) on dry weight basis. Moreover, Babu
(2015) investigated that with defatted oat flour, a lower yield of β-glucan was
achieved under acidic extraction compared to alkaline and enzymatic extractions.
Enzymatic Extraction
To improve purity and extraction within the extraction media, many enzymes are
used to remove impurities which then separated in a series of steps by using centrifugation process. For that purpose a large number of enzymes like endo-β-(1→3)
(1→4) glucanase, feruloyl esterase, endo-xylanases, xyloacetylesterase, and arabinofuranosidase could also be used. Some indigenous enzymes need to be deactivated as they will decrease the quantity and quality of extracted β-glucan during the
N. Jan et al.
stirring. Then the supernatant and residue is separated by an exhaustive centrifugation process. The residue material still contains β-glucan, thus to completely separate β-glucan, it is resuspended twice in the alkaline media under the same conditions
of temperature. All of the alkali extracts is also precipitated using ammonium sulfate by keeping the material overnight at a temperature of 4 °C or below followed
by use of centrifugation at high speed under low temperature to recover β-glucan.
This technique is highly effective to recover higher amounts of β-glucan from the
mushroom cell wall along with high levels of purity can be achieved (Nguyen
et al. 1998).
Using alkali as a solvent, β- glucan extraction from mycelium of P. chrysogenum
has also been reported; the mycelium was first freeze-dried and then ground into
powder form. Then the material was treated with alkaline solution for the extraction
of β-glucan. Maximum recovery can be obtained at a concentration of 1 M NaOH
at 40 °C as NaOH is a more effective alkali in this form. β-glucan can be extracted
by the treatment of freeze-dried powder of mycelia with the said concentration of
alkali. β-glucan contains protein impurities, thus it must be separated by the sevage
method that works at low pH. After protein has been excluded, the pH of the media
is again adjusted toward neutral for separating β-glucan. Further purification is also
obtained through dialysis against distilled water (Wang et al. 2007).
Acidic Extraction
Several reports are available on various extraction methods for β-glucan however;
extraction under acidic conditions has been reported only by a few workers.
Ahluwalia and Ellis (1984) have focussed the determination of β-glucan and starch
in barley and its malt. They adopted a simple and quick method for the extraction of
β-glucan and starch by using perchloric acid (50 mM). Then the extract was directly
used without the drying process to determine the β-glucan and starch contents
within the extract solution. For β-glucan determination, the extract was hydrolyzed
to glucose by incubating with sodium acetate buffer and “treated cellulose.” Glucose
released was measured enzymatically using glucose-6-phosphate dehydrogenase
and results were expressed as β-glucan (%) on dry weight basis. Moreover, Babu
(2015) investigated that with defatted oat flour, a lower yield of β-glucan was
achieved under acidic extraction compared to alkaline and enzymatic extractions.
Enzymatic Extraction
To improve purity and extraction within the extraction media, many enzymes are
used to remove impurities which then separated in a series of steps by using centrifugation process. For that purpose a large number of enzymes like endo-β-(1→3)
(1→4) glucanase, feruloyl esterase, endo-xylanases, xyloacetylesterase, and arabinofuranosidase could also be used. Some indigenous enzymes need to be deactivated as they will decrease the quantity and quality of extracted β-glucan during the
N. Jan et al.