3. FOOD ChEMISTRy & bIOTEChNOLOGy 3.1. Lectures

Chem. Listy, 102, s265–s1311 (2008) Food Chemistry & Biotechnology
P39 TriCHOsPOrON CuTANeuM: CELL
ADhESION ON CELLOPhANE SuRFACE
JITKA HRDInOVá, TEREZA KRULIKOVSKá,
VLADIMíR JIRKů, OLGA SCHREIBEROVá, ALEnA
ČEJKOVá and JAn MASáK
Institute of Chemical Technology Prague,
Technická 5, 166 28 Praha 6 – Dejvice, Czech Republic,
jitka.hrdinova@vscht.cz
Introduction
Recently we have faced the problem of increasing
amount of solid cellulose wastes that come mainly from agriculture
activities, food industry as well as from municipal
waste 1,2 .
Cellulose is considered to be a solid nontoxic pollutant;
however, its recalcitrant nature causes many difficulties in
removal of cellulosic waste. The microbial degradation is one
of the possibilities how it could be solved.
Colonization of solid material by microbial cells is
the crucial step in their biodegradation. Biofilm formation
by cellulolytic microorganisms is not investigated enough.
Therefore, establishing more efficient arrangement for technological
solubilization of solid cellulosic wastes could be
achieved using cellulolytic biofilms, formed by direct colonization
of these wastes by cell populations. In this association,
cellophane was chosen as a representative of solid cellulosic
substrates (carrier) and the yeast Trichosporon cutaneum as a
relatively little researched cellulolytic strain.
Experimental
The yeast Trichosporon cutaneum CCY 30-4-5 was
obtained from Department of Genetics and Microbiology,
Faculty of Science, Charles University, Czech Republic.
Inoculum was grown in complex medium and after
2 days it was replaced in minimal medium supplemented
with 1% cellulosic substrates as a sole source of carbon. Sigmacell
Type 101, carboxymethylcellulose – CMC, hydroxyethylcellulose
– HEC, cellophane and filter paper was used
as the representatives of these substrates. Minimal medium
composition in g dm –3 : KH 2 PO 4 – 1.7; na 2 HPO 4 . 7H2 O
– 0.75; (nH 4 ) 2 SO 4 – 5.0; MgSO 4 . 7H2 O – 0.02; CaCl 2 – 0.02;
FeSO 4 . 7H2 O – 0.001; MnSO 4 . H2 O – 0.001, pH 5.8. The
temperature was maintained at 28 °C. Cultures were harvested
during the exponential phase; at 48–72 hr. Separated
and rinsed cells were used for experiments.
F C 8 1 F l o w C e l l D e s c r i p t i o n
The FC 81 Flow Cell in Fig. 1. is a flat plate flow cell
designed for use with transmitted light microscopes and it
was used for observation of biofilm formation by yeast Trichosporon
cutaneum. The capability of the yeast cells to
colonize cellophane as well as the effect of shear stress was
investigated.
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Experiment conditions:
Cell suspension – OD 400 nm 0.1
Temperature – 22 °C
Flow rate – 2-20 ml min –1
Time period – 2 hr.
Fig. 1. Photograph of the FC 81 Flow Cell (biosurface Technologies,
Corp., uSA)
Experiments were carried out as shown by Fig. 2.;
Fig. 3. illustrates an emplacement of cellophane stripe in the
flow cell.
H y d r o p h o b i t y o f Y e a s t C e l l s
The yeast populations were prepared in cultivation
medium supplemented with different types of cellulosic substrates.
Hydrophobity of yeast cell surface was determined
using MATH method 3 .
M i c r o s c o p y a n d I m a g e A n a l y s i s
The colonization of cellophane was observed with
transmitted light microscopy and analysis of the images was
accomplished with Lucia (Laboratory Imaging, Ltd., UK).
The areal parameters of objects (colonies, cells) such as
length, width and colonized area were measured with image
analysis4. The observation area of the flow cell was divided
into three fields (inflow, middle, and outflow). Ten images
were taken in every field.
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Microscope – nIKOn Eclipse E400, Plan Fluor objective,
10 ×/0.30, Ph1 DLL, ∞/0.17 WD 16.0 (Japan)
Filter – 45 mm, nCB11
Camera and software – Canon PowerShot A620, Zoom-
Browser EX 5.5