Appl Microbiol Biotechnol (2006) 73: 60–66
DOI 10.1007/s00253-006-0454-y
BIOTECHNOLOGICAL PRODUCTS AND PROCESS ENGINEERING
Khaw Teik Seong
.
Yoshio Katakura
.
Kazuaki Ninomiya
.
Yohei Bito
.
Satoshi Katahira
.
Akihiko Kondo
.
Mitsuyoshi Ueda
.
Suteaki Shioya
Effect of flocculation on performance of arming yeast in direct
ethanol fermentation
Received: 15 February 2006 / Revised: 29 March 2006 / Accepted: 31 March 2006 / Published online: 13 May 2006
# Springer-Verlag 2006
Abstract In the direct ethanol fermentation of raw starch
by arming yeast with α-amylase and glucoamylase, it is
preferable to use a flocculent yeast because it can be
recovered without centrifugation. Three types of arming
yeast system, I (nonflocculent), II (mildly flocculent), and
III (heavily flocculent), were constructed and their
fermentation performances were compared. With an
increase in the degree of flocculation, specific ethanol
production rate for soluble starch decreased (0.19, 0.17,
and 0.12 g g-dry-cell
−1
h
−1
for systems I, II, and III,
respectively), but that for raw starch did not decrease as
much as expected (0.06, 0.06, and 0.04 g g-dry-cell
−1
h
−1
for systems I, II and III, respectively). Microscopic
observation revealed that many starch granules were
captured in the yeast flocs in system III during the direct
ethanol fermentation of raw starch. It was suggested that
the capture of starch granules increases apparent substrate
concentration for amylolytic enzymes in arming yeast cell
flocs; thus, the specific ethanol production rate of system
III was kept at a level comparable to those of the other
systems.
Introduction
The exhaustion of fossil fuels in the foreseeable future has
sparked a global effort to develop alternative renewable
energy sources. A number of renewable energy platforms
have been proposed with bioethanol production from
biomass being one of the potential candidates. Bioethanol
production from starchy materials by microorganisms has
been studied extensively over the past few decades.
Bioethanol production by yeast consists of three major
processes: the preparation of starchy materials from
agricultural crops, the bioconversion of starchy materials
to ethanol by yeast, and the recovery of ethanol from the
fermentation broth by distillation. The bioconversion of
starchy materials to ethanol is a critical step in the overall
process because Saccharomyces cerevisiae, which is one of
the best microorganisms for producing ethanol, cannot
directly utilize starchy materials. Although a straightfor-
ward approach to this problem is to add α-amylase and
glucoamylase exogenously to convert starchy materials to
glucose, this method will eventually increase the produc-
tion cost of ethanol. One promising strategy is to develop
an inexpensive single-step fermentation using a recombi-
nant yeast that secretes amylolytic enzymes (Ashikari et al.
1989; Birol et al. 1998; de Moraes et al. 1995; Eksteen et al.
2003). However, the productivity of amylolytic enzymes is
reduced by the successive reuse of yeast cells (Khaw et al.
2005). Because the recovery of secreted enzymes from the
fermentation broth is impractical and the addition of
exogenous enzymes to compensate for the reduction in
productivity is costly, we have constructed a series of
arming yeasts displaying amylolytic enzymes on their cell
surface for the direct ethanol fermentation of starchy
materials (Murai et al. 1999; Shigechi et al. 2002, 2004a,b).
The advantages of the display system constructed are the
high stabilities of amylolytic enzymes and the ease of
recovery of such enzymes together with arming yeast cells.
To recover yeast cells without centrifugation, we have
examined the use of a flocculent strain that sediments
rapidly (Khaw et al. 2005). However, the formation of cell
flocs seems to prevent contact between starch granules and
K. Seong
.
Y. Katakura
.
K. Ninomiya
.
S. Shioya (*)
Department of Biotechnology, Graduate School of Engineering,
Osaka University, 2-1 Yamadaoka,
Suita, Osaka 565-0871, Japan
e-mail: shioya@bio.eng.osaka-u.ac.jp
Tel.: +81-6-68797444
Fax: +81-6-68797444
Y. Bito
.
S. Katahira
.
A. Kondo
Department of Chemical Science and Engineering,
Faculty of Engineering, Kobe University,
1-1 Rokkodaicho, Nada-ku,
Kobe 657-8501, Japan
M. Ueda
Department of Applied Biochemistry,
Division of Applied Life Sciences,
Graduate School of Agriculture, Kyoto University,
Kitashirakawa-Oiwake-cho, Sakyo-ku,
Kyoto 606-8502, Japan