BLBS102-c13 BLBS102-Simpson March 21, 2012 13:15 Trim: 276mm X 219mm Printer Name: Yet to Come
250 Part 2: Biotechnology and Enzymologyduring capture is often intensive, the muscular glycogen stores
may be almost depleted even before slaughtering takes place.
The activation of myosin ATPase by spinally mediated reflexes
continues after slaughter, but the extent to which it occurs can be
reduced by employing rested harvest techniques such as anes-
thetization (Jerrett and Holland 1998, Robb et al. 2000); by
destroying the nervous system by use of appropriate methods
such asiki jime,where the brain is destroyed by a spike (Lowe
1993), or by use of still other techniques (Chiba et al. 1991, Berg
et al. 1997).
ATPases hydrolyze the terminal phosphate ester bond, form-
ing ADP. ADP is subsequently degraded by adenylate kinase,
forming AMP. The degradation of ADP is favored by the removal
of AMP by AMP deaminase (EC 3.5.4.6), producing inosine
monophosphate (IMP) and ammonia (NH 3 ). AMP deaminase
is a key regulator of the intracellular adenine nucleotide pool
and as such is a highly regulated enzyme inhibited by inorganic
phosphate, IMP and NH 3 and activated by ATP.
It is widely accepted that IMP contributes to the desirable taste
of fresh seafood (Fluke 1994, Gill 2000, Haard 2002). IMP is
present in small amounts in freshly caught relaxed fish but builds
up during the depletion of ATP (Berg et al. 1997). IMP can be
hydrolyzed to form inosine, yet the rate of hydrolysis in fish is
generally low, following zero-order kinetics, indicating that the
IMP-degrading enzymes are fully substrate saturated (Tomioka
et al. 1987, Gill 2000, Itoh and Kimura 2002). However, a large
species variation in the degradation of IMP has been shown,
and thus IMP may persist during storage for several days oreven weeks (Dingle and Hines 1971, Gill 2000, Haard 2002).
The formation of inosine is often rate limiting for the overall
breakdown of nucleotides and is considered to constitute the last
purely autolytic step in the nucleotide catabolism of chilled fish.
The formation of inosine usually indicates a decline in the prime
quality of seafood (Surette et al. 1988, Gill 2000, Haard 2002).
IMP can be hydrolyzed by various enzymes, 5’-nucleotidase
(EC.3.1.3.5) being regarded as the most important of these in
the case of chilled fish (Gill 2000, Haard 2002). Several forms
of 5’-nucleotidase exist, of which a soluble sarcoplasmic form
has been documented in fish (Itoh and Kimura 2002).
The degradation of inosine continues via hypoxanthine and
its oxidized products xanthine and uric acid. These reactions, all
of them related to spoilage, are catalyzed by both endogenous
and microbial enzymes, the activity of the latter depending upon
the spoilage flora that are present and thus differing much be-
tween species and products. Inosine can be degraded by either
nucleoside phosphorylase (EC 2.4.2.1) or inosine nucleosidase
(EC 3.2.2.2), leading to the formation of different by-products
(see Fig. 13.1). The final two-step oxidation of hypoxanthine is
catalyzed by one and the same enzymes, xanthine oxidase (EC
1.1.3.22). The oxidation requires a reintroduction of oxygen in
the otherwise anaerobic pathway, which often limits the extent
of the xanthine oxidase reaction and causes inosine and hypox-
anthine to accumulate. Hypoxanthine is an indicator of spoilage
and may contribute to bitter taste (Hughes and Jones 1966).
The hydrogen peroxide produced by the oxidation of hypoxan-
thine can be expected to cause oxidation, but the technologicalFigure 13.1.The successive reactions of the main pathway for nucleotide degradation in fish. Substances listed below the dotted line are
generally considered to be indicators of spoilage. Piis inorganic phosphate.