Food Biochemistry and Food Processing (2 edition)

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14 Seafood Enzymes: Biochemical Properties and Their Impact on Quality 277

Control SMS 0.05% Catechin 0.1% Catechin

Figure 14.7.Pacific white shrimp without and with different treatments at day 10 of iced storage (Nirmal and Benjakul 2009b). SMS, sodium

metabisulfite.

crustaceans during postmortem storage or handling. Whole Pa-

cific white shrimp treated with catechin solution (0.05% or 0.1%)

kept in ice for 10 days retarded melanosis in comparison with

the control and those treated with 1.25% sodium metabisulfite

(Nirmal and Benjakul 2009b) (Fig. 14.7).

TRIMETHYLAMINE-N-OXIDE

DEMETHYLASE

TMAOase (EC 4.1.2.32) catalyzes the conversion of trimethy-

lamine oxide (TMAO) to dimethylamine (DMA) and formalde-

hyde (FA) (Gill and Paulson 1982). TMAOase is one of the

important enzymes in fish exhibiting an adverse effect on fish

quality. TMAOase has been found at different degrees in various

fish species. The inhibition or lowering of TMAOase activity in

the postharvest animal could be an effective approach to main-

tain fish quality.

Distribution of TMAOase

Although TMAO is distributed among all kinds of marine fish

and invertebrates, and in some fresh water fish, TMAOase

activity has been identified in only 30 species of marine fish

belonging to 10 families and 8 species of invertebrates, mainly

mollusk (Sikorski and Kostuch 1982). Accumulation of DMA

and FA has been described in species belonging to Gadidae,

Merluccidae, and Myctophidae (Sikorski and Kostuch 1982).

Among the fish, Gadiform (families Gadidae and Merluccidae)

is a group of fish with high TMAOase activity (Sotelo and

Rehein 2000).

The level of TMAOase in a particular species and tissue ex-

hibits variations (Rehbein and Schreiber 1984) and seems to be

influenced by factors such as gender, maturation stage, tempera-

ture of the habitat, and feeding status, or size (Sotelo and Rehein

2000).

Depending on the source of the enzyme, the activity may be

located in the soluble fraction or in the particulate matter of ho-

mogenates of organs or tissues (Sotelo and Rehein 2000). The

organs and tissues including viscera and skin contain TMAOase.

Harada (1975) isolated TMAOase from the liver of lizardfish

(Saurida tumbil). Distribution of TMAOase in various inter-

nal organs of lizardfish (Saurida micropectoralis) were studied

(Benjakul et al. 2004b). Rey-Mansilla et al. (2002) studied the

activities of TMAOase from several internal organs of hake

(kidney, spleen, liver, heart, bile, and gallbladder). It was found

that kidney and spleen showed the highest activities while liver,

heart, bile, and gallbladder activities were much lower. More-

over, TMAOase has been found in muscle of some fish species,

such as red hake (Phillipy and Hultin 1993) and walleye pollack

(Kimura et al. 2000).

Purification and Characterization of

TMAOase

Harada (1975) partially purified TMAOase from liver of lizard-

fish (Saurida tumbil) and found that the enzyme had an optimum

pH of 5.0 and required methylene blue for activation. Tomioka

et al. (1974) found that TMAOase activity from pyloric ceca of

Alaska pollack was stimulated by Fe^2 +,l-ascorbic acid, flavin

mononucleotide (FMN) and inhibited by Fe^3 +,Cu^2 +, EDTA,

trimethylamine, and choline. TMAOase associated with Alaska

pollack myofibrils has been characterized (Kimura et al. 2000).

The enzyme requires cofactors for full activity. The system of

nicotinamide adenine dinucleotide and FMN requires anaerobic

conditions while the system of iron and cysteine and/or ascor-

bate functions in the presence or absence of oxygen (Parkin and

Hultin 1986). Benjakul et al. (2003b) purified TMAOase from

lizardfish (Saurida micropectoralis) kidney by acidification fol-

lowed by DEAE–cellulose column chromatography, in which a

purification fold of 82 with a yield of 65.4% were obtained. The

molecular mass of the partially purified enzyme was estimated

to be 128 kDa based on activity staining. Optimum pH and tem-

perature were 7.0 and 50◦C, respectively. The activation energy

was calculated to be 30.5 kJ mol−^1 K−^1. Combined cofactors

(FeCl 2 , ascorbate, and cysteine) were required for full activa-

tion. Fe^2 +exhibited a higher stimulating effect on TMAOase

activity than Fe^3 +. At concentrations less than 2 mM, ascor-

bate was more stimulatory of TMAOase activity than cysteine.

TMAOase was tolerant to salt at concentrations up to 0.5 M.

The enzyme had aKmof 16.24 mM andVmaxof 0.35μmol/min,

and was able to convert TMAO to DMA and FA. Leelapongwat-

tana et al. (2008a) partially purified TMAOase from lizardfish