The toxins causing DSP: okadaic acid and the dinophysistoxins

Okadaic acid and dinophysistoxins (DTXs), the latter so named because they occur in the dinoflagellate genusDinophysis (see Table 4.1), are acidic compounds (Fig. 4.2b (p. 59)

CH₃

Fig. 4.2 (i) The chemical structures of some of the pectenotoxin group (from van Egmond et al., 2004).

See also Halim and Brimble (2006).

and see Quilliam, 2004a). The most important of this group of toxins as regards shellfish contamination are OA, DTX-1 and DTX-2, which may occur singly or together. The polyether backbone of OA, DTX-1 and DTX-2 may be acylated with a range of saturated and unsaturated fatty acids from C14 to C22 in the digestive glands of various shellfish to produce a mixture of compounds with diarrhetic activity, collectively known as ‘DTX-3’ (Quilliam and Wright, 1995; Wright, 1995; Quilliam, 2004a; van Egmondet al., 2004). In addition to bivalves, there is evidence that fatty acid esters of OA can be produced in the digestive gland of Brown crabs (Cancer pagurus) (Torgersen et al., 2005).

Two OA derivatives, DTX-4 and DTX-5a and -b, which are unusual in being water-soluble, have been isolated from the benthic dinoflagellatesProrocentrum lima and P. maculatum (Hu et al., 1995b,c; Quilliam et al., 1996). They possess an aliphatic side chain containing hydroxyl and sulfate groups and in the case of DTX-5 an amide moiety. Quilliam (2004a) suggests that DTX-4 and -5 have not been detected in shellfish because of the action of esterases.

CH₃

Fig. 4.2 (j) The chemical structures of some of the yessotoxin group and of adriatoxin (from van Egmond et al., 2004). See also Bowden (2006) and Hess and Aasen (2007).

4.5.2 Toxic effects

The symptoms of DSP in humans are summarized in Table 4.1. Gastrointestinal disturbance is the main effect and the symptoms usually disappear within 3 days. Studies on the pathological effects of OA and DTXs as well as the PTXs and YTXs (see Section 4.10) have been confined mainly to suckling and adult mice and to rats – detailed accounts can be found in Teraoet al.

(1990a,b, 1993), Aune and Yndestad (1993) and in van Egmondet al. (2004). The acute toxicities (LD₅₀, μg kg⁻¹ b.w.) to mice of OA, DTX-1 and DTX-3 are 200, 160 and 500 respectively (Fern´andezet al., 2004a). Aune et al. (2007) demonstrated that DTX-2 has about 60% of the toxicity of OA when estimated by the mouse bioassay (i.p. injection) and an enzyme inhibition assay showed good correspondence.

Table 4.2 Sites of action and primary physiological effects of the major classes of phycotoxins subject to governmental regulation in Europe and elsewhere.

Toxin Site of action Primary physiological effects

Saxitoxin group Site 1 on voltage-dependent sodium channels

Block sodium ion influx in neuronal and muscular sodium channel. Prevents propagation of the action potential essential for conduction of nerve impulses and contraction of muscles. Peripheral nervous

Pectenotoxins Not known Hepatotoxic. Damage to cytoskeleton, e.g.

F-actin depolymerization to cells in vitro Yessotoxins Not known. Possible interaction

with calcium channels reported

Cardiotoxic

Azaspiracids Not known Affects liver, pancreas, thymus, spleen and gastrointestinal tract and T and B lymphocytes in rodents. AZA-1 has effects on cytoskeleton (F-actin). Effects on intracellular calcium in cells in vitro. See Tables 4.3 and 4.4 Domoic acid Ionotropic class of glutamate

receptors in central nervous system

Binds to glutamate receptors, i.e. competes with glutamate; receptor-induced

depolarization and excitation NSP toxins Site 5 on voltage-dependent

sodium channels

Induce channel-mediated sodium ion influx;

depolarizes isolated muscle and nerve cells Ciguatera toxins:

Ciguatoxin

Site 5 on voltage-dependent sodium channels

Opening of sodium channels at resting potential and inability of open channels to be inactivated during subsequent depolarization Maitotoxin Calcium channels Calcium ion influx, which may lead to cell

death

Table updated from Leftley and Hannah (1998). See also Hampson and Manalo (1998), Burgess and Shaw (2001), van Egmond et al. (2004), Arias (2006), Bowden (2006) and Llewellyn (2006).

Intraperitoneal and oral doses of OA and DTXs cause marked changes in the small intes-tine, such as fluid accumulation and distension. Ultrastructural changes include degeneration of the intestinal absorptive epithelium. Teraoet al. (1990a,b, 1993) also observed liver dam-age in rats and mice given i.p. and oral doses of OA and DTX-1. The effects of purified OA and DTX-1 on freshly prepared rat hepatocyte cells have been observed using light and electron microscopy (Auneet al., 1991). OA and DTX-1 produced dose-dependent changes including blebbing of the cell surface and overall irregular shape.

Severe injuries to the intestinal mucosa of mice were observed within an hour of oral administration. OA injected into ligated loops of rat intestine produced progressive damage to the villi within 15 minutes and the degree of damage was dose-dependent. OA and DTX-1 induced liver damage in mice and rats after oral as well as i.p. administration. DTX-1 and DTX-3 also induced damage to the epithelium in the small intestine after both oral and i.p.

dosing (van Egmondet al., 2004).

A number ofin vitro cytotoxicity studies have been carried out with various cell lines.

Among the most significant findings were that protein and DNA synthesis in monkey kidney cells (Vero line) were both inhibited by OA in a concentration-dependent manner and it was concluded that the first and main site of action of OA was protein synthesis (van Egmond et al., 2004). OA administered to human intestinal epithelial cells (T84 line) increased the paracellular permeability and it was suggested that this might contribute to the diarrhetic effect of DSP toxins (Tripuraneniet al., 1997).

The main concern about OA and the DTX toxins as regards human health is their role as potent tumour promoters. An effect of OA and DTX-1 on mouse skin was to induce ornithine carboxylase, an enzyme associated with various cancers that is known to be involved in signal transduction pathways involved in cell proliferation. OA has been observed to induce ornithine decarboxylase in rat stomach and also enhanced neoplastic changes in rat stomach after it had been pre-treated with a carcinogen (van Egmondet al., 2004).

The possible effects of chronic exposure to these toxins in humans still have to be estab-lished (Aune and Yndestad, 1993; van Egmondet al., 2004). An epidemiological study was carried out in France to investigate digestive cancer mortality in coastal regions related to consumption of seafood contaminated by DSP toxins. A very tenuous link was established between possible exposure to DSP toxins via seafood and some digestive cancers, mainly in men (Cordieret al., 2000).

At a biochemical level OA and DTXs are powerful inhibitors of protein phosphatases PP1 and PP2 and 2A, which are important in regulating many important metabolic processes in eukaryotic cells, including cell proliferation (Table 4.2).