10 Analytical Protocols in the Measurement …
235
contained SCC homogenized in an applesauce matrix with 10% corn oil (AS). The
final SCC concentrations ranged from 50 to 250 μg/mL for all experimental preparations. Aliquots of each preparation were exposed to a two-phase in vitro digestion
protocol. The gastric phase included acidification of the sample to pH 2.0 with
100 mM HCl and exposure to porcine pepsin (3 mg/mL) with incubation at 37 °C
for 1 h in a shaking water bath (95 rpm). The small intestinal phase was initiated by
neutralizing the gastric phase with NaHCO 3 , adding porcine pancreatin (0.4 mg/mL),
lipase (0.2 mg/mL), and bile extract (2.4 mg/mL), and adjustment of the final pH value
to 7.0 with 1 N NaOH prior to incubation at 37 °C in a shaking water bath (95 rpm).
After 2 h, aliquots of the digestate were centrifuged at 167,000 g at 4 °C for 35 min to
separate the aqueous micellar fraction from residual solids and oil. Isolated aqueous
fractions were filtered (0.2-micron pore size) to remove contaminating aggregates.
Caco-2 human intestinal cells were used to characterize the accumulation of SCC.
The authors found that Cu(II)chlorin e 4 , the major chlorin component of SCC, was
relatively stable during simulated digestion. On the other hand, more than 90% of
Cu(II) chlorin e 6 was degraded to undetermined products during digestion. Accumulation of SCC derivatives was also investigated by using differentiated cultures
of the TC7 clone of the Caco-2 human intestinal cell line. Cellular accumulation in
media containing 0.5 to 60 ppm SCC was linear with intracellular content ranging
between 0.2 and 29.6 μg of total SCC per mg of cellular protein.
The literature data on bioavailability of phycobiliproteins is very scarce, showing
the importance to investigate this field, especially in order to stablish protocols
that allow the exchange and comparison between results obtained worldwide. Most
studies that evaluate the bioavailability related to microalgae involve the availability
and digestibility of some of the constituents, particularly the minerals. However, the
steps to evaluate bioavailability are the same followed to evaluate the absorption of
carotenoids and chlorophyll.
In recent years, there is a growing number of studies on protein hydrolysis as an
effective method to produce bioactive peptides and several works have been pointing
to the possibility that the biological effects that bring health benefits through the intake
of microalgal pigments are related to these peptides. Bioactive peptides are obtained
from enzymatic hydrolysis and exhibit excellent biological effects. Protein hydrolysis is generally performed by enzymes derived from microorganisms or plants, but
digestion by enzymes of the gastrointestinal tract (GIT), such as pepsin or trypsin,
present physiological relevance (Begum et al. 2016).
Ingestion of phycocyanin, resulting from the intake of Spirulina dietary supplements, is a very plausible way to evaluate the bioavailability for these pigments, since
there is a potential susceptibility to GIT proteolysis related to the oral administration
and consumption, to better understand the structure and bioactivities of the released
chromopeptides. There are very few data on the literature about bioactivities of
peptides obtained after C-phycocyanin digestion, as well as bioactivities of peptides
with covalently bound bioactive chromophores (chromopeptides) in general.
Minic and others (2016) examined the digestibility of C-phycocyanin (C-PC)
by pepsin in simulated gastric fluid. The authors used simulated in vitro digestion,
followed by SDS polyacrylamide gel electrophoresis. The pepsin digest of C-PC was
235
contained SCC homogenized in an applesauce matrix with 10% corn oil (AS). The
final SCC concentrations ranged from 50 to 250 μg/mL for all experimental preparations. Aliquots of each preparation were exposed to a two-phase in vitro digestion
protocol. The gastric phase included acidification of the sample to pH 2.0 with
100 mM HCl and exposure to porcine pepsin (3 mg/mL) with incubation at 37 °C
for 1 h in a shaking water bath (95 rpm). The small intestinal phase was initiated by
neutralizing the gastric phase with NaHCO 3 , adding porcine pancreatin (0.4 mg/mL),
lipase (0.2 mg/mL), and bile extract (2.4 mg/mL), and adjustment of the final pH value
to 7.0 with 1 N NaOH prior to incubation at 37 °C in a shaking water bath (95 rpm).
After 2 h, aliquots of the digestate were centrifuged at 167,000 g at 4 °C for 35 min to
separate the aqueous micellar fraction from residual solids and oil. Isolated aqueous
fractions were filtered (0.2-micron pore size) to remove contaminating aggregates.
Caco-2 human intestinal cells were used to characterize the accumulation of SCC.
The authors found that Cu(II)chlorin e 4 , the major chlorin component of SCC, was
relatively stable during simulated digestion. On the other hand, more than 90% of
Cu(II) chlorin e 6 was degraded to undetermined products during digestion. Accumulation of SCC derivatives was also investigated by using differentiated cultures
of the TC7 clone of the Caco-2 human intestinal cell line. Cellular accumulation in
media containing 0.5 to 60 ppm SCC was linear with intracellular content ranging
between 0.2 and 29.6 μg of total SCC per mg of cellular protein.
The literature data on bioavailability of phycobiliproteins is very scarce, showing
the importance to investigate this field, especially in order to stablish protocols
that allow the exchange and comparison between results obtained worldwide. Most
studies that evaluate the bioavailability related to microalgae involve the availability
and digestibility of some of the constituents, particularly the minerals. However, the
steps to evaluate bioavailability are the same followed to evaluate the absorption of
carotenoids and chlorophyll.
In recent years, there is a growing number of studies on protein hydrolysis as an
effective method to produce bioactive peptides and several works have been pointing
to the possibility that the biological effects that bring health benefits through the intake
of microalgal pigments are related to these peptides. Bioactive peptides are obtained
from enzymatic hydrolysis and exhibit excellent biological effects. Protein hydrolysis is generally performed by enzymes derived from microorganisms or plants, but
digestion by enzymes of the gastrointestinal tract (GIT), such as pepsin or trypsin,
present physiological relevance (Begum et al. 2016).
Ingestion of phycocyanin, resulting from the intake of Spirulina dietary supplements, is a very plausible way to evaluate the bioavailability for these pigments, since
there is a potential susceptibility to GIT proteolysis related to the oral administration
and consumption, to better understand the structure and bioactivities of the released
chromopeptides. There are very few data on the literature about bioactivities of
peptides obtained after C-phycocyanin digestion, as well as bioactivities of peptides
with covalently bound bioactive chromophores (chromopeptides) in general.
Minic and others (2016) examined the digestibility of C-phycocyanin (C-PC)
by pepsin in simulated gastric fluid. The authors used simulated in vitro digestion,
followed by SDS polyacrylamide gel electrophoresis. The pepsin digest of C-PC was