Biomedical Engineering Reference
In-Depth Information
overnight shipment of large culture volumes can be costly and/or impractical. There also exists
a concern that injecting large volumes of culture that may be contaminated with fermentation
byproducts (e.g., VFAs, as shown in Figure
3.3
) or residual growth substrates (e.g., PCE, TCE,
DCE or VC) could lead to further contamination of the site or, at the very least, legal
implications (see above).
One approach for addressing the issues of culture storage and shipping is to concentrate
the cells for storage and shipping. Cell concentration reduces shipping and storage volumes,
and it removes the bulk of the bacterial culture broth and its potential byproducts or
contaminants. The suitability of cell concentration depends on the robustness of the cultured
cells, however, and the potential for losing an important member of a consortium during the
concentration process should be evaluated.
3.5.1 Concentrating Cultures
Several techniques including vacuum evaporation, spray evaporation, continuous
centrifugation and ultra- or cross-flow filtration have been used in biotechnological applica-
tions to concentrate bacterial cells. Many of these, however, are difficult to apply while
maintaining strict anaerobic conditions (Ljungdahl and Wiegel,
1986
). Therefore the SDC-9
TM
culture is concentrated by cross-flow filtration over a custom-built concentrator constructed
with six Kerasep
TM
KBX tubular ceramic membrane units (Novasep, Inc., Boothwyn, Penn-
sylvania) operated in series (Figure
3.10
). Each filter unit contains seven BX-7c ceramic
elements containing seven flow channels each, all of which are contained within stainless
steel shells. The filters represent 72 ft
2
(6.6 m
2
) of membrane surface area with an effective
pore size of 0.2 micrometers (
m
m). The ceramic membranes are chemically cleaned by circulat-
ing a solution of 0.5% NaOH through the system for 8 h prior to cell concentration activities.
All manipulations are performed under strict anaerobic conditions facilitated by charging
the entire system with N
2
prior to introducing the cells, and by connecting the concentrator
directly to the reactors so that liquid does not have to be removed from the system for
concentration activities.
The culture from the 4,000-L reactor is passed over the membranes at a pressure of 50-55 psi
and returned to the reactor by using a two-pump system. The first pump is the reactor circulation
pump (G&L SSH-S 2
2.5-8; A Gould Pump Co., Seneca, New York) that is capable of
transferring 100 gallons (gal)/min (378 L/min), and the second is a high pressure pump (G&L
NPE 1-1/4
1 - ½-1: A Gould Pump Co., Seneca, New York) with a capacity of 50 gal/min
(189 L/min). The culture from the 750-L reactor is concentrated by using a separate lower
capacity (24 gal/min; 91 L/min) pump (Model CHI-4-50; Grundfos Pump Corp. USA, Olathe,
Kansas). The system is designed to remove ~400-500 L of liquid/h at an initial cell concentration
1.0-1.2 g/L of biomass (DWT), or in the case of the 750-L reactor, to remove 80-85 L/h.
The culture from the 4,000-L reactor can be concentrated to ~120 L within the large reactor
vessel (i.e., ~26 fold), or subsequently transferred to the 750-L vessel and concentrated to ~50 L
(i.e., 64-fold). The culture in the 750-L reactor (550-L of broth) can be concentrated to ~50 L (i.e.,
~10-fold). The concentration process also can be stopped at any time during the process to
generate a culture with a desired
Dhc
concentration. Concentrated cells are transferred to N
2
-
charged 18.5-L stainless steel soda kegs (refer to Figure
3.12
), pressurized to 15 psi with N
2
,and
stored at 4
C.
Figure
3.10
shows a photo of the cell concentration system connected to the 4,000-L reactor,
and Figure
3.11
shows the results of the concentration of a 3,500-L SDC-9
TM
culture in the
ceramic membrane concentrator system. The cell culture is chilled during concentration to
ensure maintenance of cell viability. Analyses of the specific activity of the cells before and
Search WWH ::
Custom Search