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of dialysable Fe (Argyri
et al.
, 2009).
Furthermore, this method has also been
used to measure Zn bioavailability (Argyri
et al.
, 2011).
Another method for estimating bio
-
availability is to use Caco-2 cells, which is a
quick and inexpensive method (Glahn
et al
.,
1997). These human colon cells are seeded
at a density of 5 × 104 cells/cm
2
and grown
for 21-25 days to obtain a cell monolayer
(Aherne
et al.
, 2010). The mucus layer cov-
ering the intestinal epithelium has a major
role in the absorption. The mucus is mainly
composed of mucin, which is secreted by
goblet cells in the epithelium. These cells
can be emulated
in vitro
using HT29-MTX
cells, cells that are human colon adenocar-
cinoma resistant to methotrexate. A co-culture
of Caco-2 and HT29-MTX represents two
major types of cells in the epithelium of the
small intestine. When Caco-2 cells and gob-
let cells are cultured together they form a
monolayer, with tight junctions between the
two cell populations. This model has been
used for studies of Fe bioavailability (Mahler
et al.
, 2009).
(2000), with the difference that all steps
occurring in the colon (ascending, trans-
verse and descending colon), were per-
formed in a single reactor. Other
in vitro
colon simulators have long fermentation
times, of between 1 and 14 days (Yoo and
Chen, 2006), which is not suitable to esti-
mate the bioavailability, but this involves
shorter fermentation times in the colon.
Figure 4.1 shows a diagram of the
in vitro
simulation conditions of the entire diges-
tive process.
An example of the use of
in vitro
diges-
tion process in a single batch was the com-
plete digestion process of Gouda cheese,
which contained 15 g linseed meal per kg of
cheese, resulting in the detection of several
lignans (Fig. 4.2). During
in vitro
digestion
of the small intestine secoisolariciresinol
diglucoside (SDG) was identified; this plant
lignan was probably released by pancreatic
enzymes in the intestine. Eeckhaut (2008)
MASTICATION
•
α
-Amylase, mucine
• NaHCO
3
, NaCI, KCI, CaCl
2
2 H
2
O, K
2
PO
4
• pH 7.0 for 20 s
4.9 The
In Vitro
Digestive Process:
Design in a Single Batch
STOMACH
The
in vitro
simulator of the digestive
process in a single batch was designed to
evaluate the bioaccessibility of flaxseed
lignans. The upper gastrointestinal tract
was simulated according to the methodol-
ogy reported by Sumeri
et al
. (2008) with
modifications; these changes altered the
time of passage of the bolus through the
stomach and through the small intestine.
Moreover, instead of directly adding the
sample, a corresponding bolus of food and
saliva mixture was drawn. For this, artifi-
cial saliva was prepared according to
Arvisenet
et al.
(2008). In this
in vitro
sim-
ulation conditions were recreated that
occur during fermentation in the colon,
keeping the whole process in the same bio-
reactor. The large intestine stage was per-
formed according to methodology reported
by Possemiers (2004) and De Boever
• Pepsin
• pH 2.0
• 1 h
SMALL INTESTINE
• Pancreatine
• Biles
• pH 6.0-7.5 for 4 h
LARGE INTESTINE
• Faecal bacteria
• Brain-heart infusion
• pH 5.5-7.0 for 48 h
Fig. 4.1.
A scheme of the digestive process
in vitro
in a single batch.
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