Mild enzymatic isolation of mannan and glucan from yeast Saccharomyces cerevisiae

Abstract

SUMMARY: An enzymatic procedure to isolate polysaccharides from the yeast saccharomyces cerevisiae was developed which would both be environmentally friendly and preserve the native structure of the polymers. Two proteases and three enzyme cocktails were employed.To isolate the cell wall polysaccharides, the cell walls were first separated from the internal components by mechanical cell disruption. The quality of cell disruption was investigated for four types of glass beads with diameters from 0.25 -1.5 mm and monitored via the inner cellular proteins released. With large glass beads (Ø = 0.75 -1.5 mm) both brewer's and baker's yeast cells exhibited more than 90% cell disruption after as little as 12 min, whereas disruption was still incomplete with smaller beads. The isolated cell wall material was purified on microporous membranes, freeze-dried and then incubated with enzymes. Proteases caused protein lysis in the outer cell wall and released soluble mannan. The insoluble glucan was separated by centrifugation and the mannan was purified by dialysis or chromatography.In the digestion with proteases, complete conversion was achieved at concentrations higher than 240 mg g -1 of cell wall material within 26 h. Cell walls from baker's yeast had to undergo a preliminary extraction with petroleum ether. Complete conversion was only possible with one of the enzyme cocktails investigated, and this was achieved after as little as 6 h for cell walls from both baker's and brewer's yeast. The activity of the two other enzyme cocktails investigated was weaker, with brewer's yeast cell walls generally reacting more readily than those of baker's yeast. ZUSAMMENFASSUNG: Im Hinblick auf die Entwicklung umweltfreundlicher Verfahren und den Schutz der nativen Polymerstruktur wurde ein enzymatisches Verfahren zur Isolierung von Polysacchariden aus derHefe Saccharomyces cerevisiae entwickelt. Eingesetzt wurden zwei Proteasen sowie drei Enzymcocktails.Zur enzymatischen Gewinnung der Zellwand-Polysaccharide wurden zunächst die Zellwände durch einen mechanischen Zellaufschluß vom Zellinneren getrennt. Die Aufschlußgüte wurde für vier Glasperlensorten mit Durchmessern von 0,25 -1,5 mm untersucht und anhand der freigesetzten innerzellulären Proteine kontrolliert. Sowohl für Brauereihefezellen als auch Bäckerhefezellen zeigte sich mit großen Glasperlen (Ø = 0,75 -1,5 mm) bereits nach 12 min ein mehr als 90proz. Zellaufschluß, während bei kleineren Glasperlen der Aufschluß noch unvollständig war. Das gewonnene Zellwandmaterial wurde an mikroporösen Membranen gewaschen, gefriergetrocknet und anschließend mit Enzymen umgesetzt. Proteasen zerstörten in der äußeren Zellwand die Proteinmatrix und setzten lösliches Mannan frei. Das unlösliche Glucan wurde durch Zentrifugieren abgetrennt und das Mannan durch Dialyse oder Chromatographie gereinigt.Bei Umsetzung mit Proteasen war vollständiger Umsatz ab einer Konzentration von 240 mg g -1 Zellwandmaterial in 26 h möglich. Zellwände aus Bäckerhefe mußten zuvor mit Petrolether extrahiert werden. Vo...

Antioxidative activity of (1→3), (1→6)-β-d-glucan from Saccharomyces cerevisiae grown on different media

“…Yeast cells are enclosed by a cell wall containing 29-64% b-glucans, 31% mannans, 13% proteins, 9% lipids and 1-2% chitin (Bacon, Farmer, Jones, & Taylor, 1969;Fleet & Manners, 1976;Kath & Kulicke, 1999).…”

“…further subdivided into an alkali insoluble (80-85% of the entire content) and an alkali soluble part (Kath & Kulicke, 1999).…”

Yeast (Saccharomyces cerevisiae) Glucan Polysaccharides – Occurrence, Separation and Application in Food, Feed and Health Industries

“…Table 1. The cell wall components of Saccharomyces cerevisiae (Klis et al 2002;Kath & Kulicke, 1999;Lessage & Bussey, 2006;Kwiatkowski et al, 2009) The yeast cell wall forms a border that defines the yeast cell's dimensions and separates its organelles from the negative influences of the environment. The individual constituents of the cell wall connect to each other by covalent bonds forming a single supra-molecular biopolymer.…”

“…All of the individual constituents of a yeast cell wall connect to each other by covalent bonds forming a single supra-molecular biopolymer". The (1→3) (1→6)--D-glucan and the (1→2) (1→3) (1→6)--D-mannan/protein complexes (Vinogradov et al, 1998) are the major components of this supra-molecular biopolymer (Lessage & Bussey, 2006;Kath & Kulicke, 1999). They connect to each other by a covalent bond the nature of which is still under investigation (Kaptein et al, 1999;Kollar et al, 1997).…”

“…They connect to each other by a covalent bond the nature of which is still under investigation (Kaptein et al, 1999;Kollar et al, 1997). In the process of yeast cell wall isolation the cell wall is first cracked open, with the help of physical, chemical and/or enzymatic treatments (Kath & Kulicke, 1999) and then separated from the soluble yeast cell components by centrifugation (Sedmak, 2006;Jamas et al, 1998). The concentrated yeast cell wall solids containing ~30% dry weight material are then spray-dried.…”