Author:
ADAMSON NICHOLAS J.,REYNOLDS ERIC C.
Abstract
In an approach to develop a commercial-scale process for the production
of casein phosphopeptides containing the cluster sequence −SerP−SerP−SerP−Glu−Glu–, we
have
studied the relationship between casein
hydrolysis and phosphopeptide release. The degrees of hydrolysis (DH) of
casein
using Novo trypsin PTN 3.0 S and pancreatin 4NF independently, at enzyme
to
substrate (E[ratio ]S) ratios of 1[ratio ]50–1[ratio ]1600 (by weight),
were determined using the pH-stat method. Casein phosphopeptides (CPP)
were
selectively precipitated using
Ca2+ and ethanol from the acid-clarified hydrolysates.
The precipitates were analysed by
high performance capillary electrophoresis to calculate individual phosphopeptide
yields based on extinction coefficients of the purified peptides.
Individual peptides
were purified by reversed-phase HPLC and anion-exchange FPLC4 and
characterized by MALDI-TOF mass spectrometry and amino acid sequence
analysis. For both
enzymes, lowering the E[ratio ]S ratio resulted in reductions in the DH
and
the release of
the CPP, and an increase in peptide chain length. The longer chain length
offset the
reduction in release such that the gravimetric yields of CPP preparations
remained
relatively constant. For Novo trypsin the highest yields of the
major cluster peptides
(β-casein(CN)f(1–25), αs1-CNf(59–79),
αs2-CNf(1–21), αs2-CNf(46–70)
and related
peptides) in the selective precipitates were obtained at a casein DH of
17%. At lower
DH values (9–15%), there was a decrease in yield of the peptides
derived from
αs1-CN and αs2-CN while the yield of
the
β-CN-derived cluster peptides remained
relatively constant. The CPP produced using pancreatin were found to be
truncated
at all E[ratio ]S ratios, relative to the tryptic CPP, owing to
higher levels of chymotryptic
and carboxypeptidase activities in pancreatin. The highest yields of the
truncated
forms of the major cluster peptides using pancreatin were obtained
at a casein DH of 19–23%.
Publisher
Cambridge University Press (CUP)
Subject
Animal Science and Zoology,General Medicine,Food Science
Cited by
21 articles.
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