Biol. Rev. (2010), pp. 000–000. 1 doi: 10.1111/j.1469-185X.2010.00124.x Molluscan biological and chemical diversity: secondary metabolites and medicinal resources produced by marine molluscs

Kirsten Benkendorff∗ School of Biological Sciences, Flinders University, GPO Box 2100 Adeliade, 5001, SA, Australia

(Received 4 March 2009; revised 10 December 2009; accepted 17 December 2009)

ABSTRACT

The phylum Mollusca represents an enormous diversity of species with eight distinct classes. This review provides a taxonomic breakdown of the published research on marine molluscan natural products and the medicinal products currently derived from molluscs, in order to identify priority targets and strategies for future research. Some marine gastropods and bivalves have been of great interest to natural products chemists, yielding a diversity of chemical classes and several drug leads currently in clinical trials. Molluscs also feature prominently in a broad range of traditional natural medicines, although the active ingredients in the taxa involved are typically unknown. Overall secondary metabolites have only been investigated from a tiny proportion (<1%) of molluscan species. At the class level, the number of species subject to chemical studies mirrors species richness and our relative knowledge of the biology of different taxa. The majority of molluscan natural products research is focused within one of the major groups of gastropods, the opisthobranchs (a subgroup of Heterobranchia), which are primarily comprised of soft-bodied marine molluscs. Conversely, most molluscan medicines are derived from shelled gastropods and bivalves. The complete disregard for several minor classes of molluscs is unjustified based on their evolutionary history and unique life styles, which may have led to novel pathways for secondary metabolism. The Polyplacophora, in particular, have been identified as worthy of future investigation given their use in traditional South African medicines and their abundance in littoral ecosystems. As bioactive compounds are not always constitutively expressed in molluscs, future research should be targeted towards biosynthetic organs and inducible defence reactions for specific medicinal applications. Given the lack of an acquired immune system, the use of bioactive secondary metabolites is likely to be ubiquitous throughout the Mollusca and broadening the search field may uncover interesting novel chemistry.

Key words: bioactivity, biodiversity, chemical defence, molluscan evolution, marine natural products, natural remedies, secondary metabolism, traditional medicine.

CONTENTS I. Introduction ...... 2 (1) Molluscan biological diversity ...... 2 (2) Molluscan chemical diversity ...... 4 II. Taxonomic Distribution of Molluscan Metabolites ...... 6 (1) Minor classes ...... 6 (2) Cephalopoda ...... 8 (3) Bivalvia ...... 10 (4) Gastropoda ...... 10 (a) Eogastropoda and non-heterobranch orthogastropods ...... 10 (b) Heterobranch gastropods ...... 11

* Address for correspondence: Tel: +61 8 8201 3959; Fax: +61 8 8201 3015; Email: Kirsten.benkendorff@flinders.edu.au

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III. Molluscan Medicines ...... 12 IV. Conclusions ...... 15 V. Acknowledgements ...... 15 VI. References ...... 15

I. INTRODUCTION et al., 2002; Cummins et al., 2006), predatory behaviour (e.g. Roseghini et al., 1996; Craig, 2000; Kanda et al., 2003) and Throughout history, molluscs have provided a wide range of defensive secretions (e.g. Ireland & Faulkner, 1978; Pawlik, human resources, including food, shells, dyes and medicines Albizati & Faulkner, 1986; Marin et al., 1999; Kelly et al., (e.g. Fig. 1). In many cultures shelled gastropods and bivalves 2003; Derby et al., 2007). Consequently, there should be are regarded as a delicacy or healthy food and they also much scope for future drug discovery within this phylum. feature in a range of traditional natural remedies (e.g. Hu, The continual discovery of novel drug leads from the 1980; Herbert et al., 2003; Prabhakar & Roy, 2009). In most enormous pool of available species requires a strategic cases there has been no scientific research undertaken to approach, such as the investigation of traditional medicines substantiate the health benefits of molluscs. However, there and/or previously unstudied sources that are likely to is increasing interest in the bioactivity of mollusc extracts have independently evolved novel pathways for secondary and secondary metabolites (see Cimino & Gavagnin, 2006). metabolism. As outlined by Cimino & Ghiselin (2001) Currently, natural products isolated from molluscs and their chemical defence appears to have evolved differently structural analogues are particularly well represented in the in different types of organisms. Consequently, it could anticancer compounds in clinical trials (Simmons et al., 2005). be predicted that distinct chemical structures will occur Nevertheless, it is presently unclear whether the production within molluscan groups that have evolved under different of bioactive secondary metabolites is ubiquitous within the environmental and biological pressures. The purpose of this Phylum Mollusca. review is to examine the current literature on molluscan The term mollusc is derived from the Latin word molluscus secondary metabolites to identify major gaps in our meaning ‘soft’. Despite the presence of a shell in some knowledge of molluscan chemistry. Combined with an molluscan groups, all molluscs are essentially soft-bodied, assessment of molluscan evolution and medicinal resources, making them vulnerable to predators and pathogens. Even this could help refine future targets for natural products those with a shell must regularly open the shell, or extend research. The approach used here primarily involves a their muscular foot beyond it, for the purposes of feeding taxonomic classification of molluscan secondary metabolites and locomotion. Thus the shell does not present a true to highlight under-represented taxa. The bias towards certain physical barrier to microbial infection. However, molluscs taxa is also compared to the distribution of species used in often live in microbially rich habitats, such as soil and medicinal remedies, to establish further limitations in our leaf litter on land and amongst marine benthic sediments knowledge on bioactive compounds from molluscs. and hard reef communities. The majority of molluscan diversity occurs in the sea, where even in the water (1) Molluscan biological diversity column there is an estimated 105 –106 microbial cells ml−1 (Whitman, Coleman & Wiebe, 1998). When any natural Molluscs are the second largest animal phylum on earth, or artificial surface is placed in the marine environment, constituting approximately 7% of living animals. There bacteria rapidly settle, attach and form biofilms (Davis et al., are currently around 52,000 named species of marine 1989), which can facilitate pathogenic invasion. However, molluscs (Bouchet, 2006) and an estimated diversity of like all invertebrates, molluscs do not have an acquired 100,000–200,000 species (Pechenik, 2000). Molluscs are immunological memory (Sminia & Van Der Knaap, 1986; relatively well described compared to many other inver- Hooper et al., 2007). This suggests that molluscs must have tebrate phyla and the taxonomy is fairly well resolved evolved alternative defence strategies to protect themselves for an enormous diversity of species from many regions against the onslaught of microbial invasion. Indeed, their around the world, despite some remaining unresolved innate immune system does appear to have a well-developed phylogenetic disputes (e.g. Haszprunar, 1996; Ponder & humoral component with the biosynthesis of antimicrobial Lindberg, 1997; Colgan, Ponder & Eggler, 2000; reviewed defence factors (e.g. Tripp, 1975; Mitta et al., 2000b; Mitta, by Ponder & Lindberg, 2008). Molluscs are diverse not Vandenbulcke & Roch, 2000; Cellura et al., 2007; Li, Zhao only in terms of their species richness, but also encom- & Song, 2009). Under the pressure of natural selection, pass a wide range of morphologies and ecological niches. a range of different antibacterial, antifungal, antiparasitic Their habitats range from the highest alpine regions to and antiviral secondary metabolites may have evolved in the deepest sea vents and they have adapted to a range molluscs, for circulation in the haemolymph, as well as for of different life styles, including trophic niches encom- inclusion in mucus secretions on body surfaces. Molluscs have passing predatory, herbivorous, scavenging, detritivorous, also been shown to use secondary metabolites as part of their filter-feeding and even some symbiotic photo- and chemo- communication systems (e.g. Cimino et al., 1991; Zatylny autotrophs. Phylogenetically, molluscs have diversified into

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Fig. 1. Illustration of the diverse resources obtained from molluscs, such as edible whelks in the family Muricidae (Gastropoda); (A) The Australian muricid Dicathais orbita; (B) the Murex homeopathic remedy; (C) dyes and pigmented compounds isolated from muricids by (D) silica chromatography. Chemical structures are shown for the secondary metabolites (E) murexine, a muscle-relaxing choline ester; and the anticancer brominated indoles (F) tyrindoleninone; (G) 6-bromoisatin and (H) 6,6’-dibromoindirubin, which is a minor pigment and structural isomer of (I) 6,6’-dibromoindigo, the well-known historical dye Tyrian purple. seven or eight classes: Gastropoda, Bivalvia, Scaphopoda, Table 1. Published research on secondary metabolites from the Cephalopoda, Polyplacophora, Monoplacophora, Caud- different classes of Mollusca, complied from Faulkner (1984a, b, ofoveata and Solenogastres. The Caudofoveata and 1986, 1987, 1988, 1990, 1991, 1992b, 1993, 1994, 1995, 1996, Solenogastres are often combined into one paraphyletic 1997, 1998, 1999, 2000, 2001, 2002), Blunt et al., (2003, 2004, 2005, 2006, 2007, 2008, 2009), Alam & Thomson (1998), Baker group the Aplacophora (Haszprunar, 2000; Haszprunar, & Murphy (1976, 1981) and additional searches using Chemical Schander & Halanych, 2008) and as these are both minor Abstracts, Biological Abstracts and Medline. The number of species groups they are treated together herein (Table 1, Fig. 2). that have been subject to natural products studies is provided, in About 90% of molluscan diversity is found in one class, addition to the number of compounds isolated and the number the Gastropoda, with an estimated 75,000 –150,000 species of research papers, accounting for overlap between the different (Fig. 2). The Gastropoda have diversified into the widest literature sources. The per cent of species investigated in each range of ecological niches, including all trophic niches and class is calculated based on Avila’s (2006) estimate of the number terrestrial, freshwater, marine benthic, pelagic and infaunal of species in each class (see Fig. 2) habitats. The bivalves are the next most diverse class, with approximately 10,000–20,000 species (Fig. 2), found Taxa Compounds Papers Species Per cent species in marine and freshwater habitats. These are primarily filter- Aplacophora 0 0 0 0 feeding organisms, although some are deposit feeders and a Monoplacophora 0 0 0 0 few are carnivorous (the septibranchs). Some bivalve families Polyplacophora 49 11 10 1.5 Scaphopoda <10 2 4 1.1 have specifically adapted to house microbial symbionts Cephalopoda 24 11 13 1.3 within their gills or mantle tissue to allow autotrophic Bivalvia 190 146 61 0.3 nutrition. The remaining five classes of molluscs are all Gastropoda 948 878 817 1.0 marine and relatively minor in terms of their species diversity (Fig. 2). Most cephalopods are highly adapted to an active predatory lifestyle in benthic and pelagic habitats. The infaunal microcarnivores occurring in soft sediment habitats. polyplacophorans (chitons) are essentially algal grazers on The Aplacophora have mostly been found from deep sea shallow rocky reefs and the Scaphopoda (tusk shells) are and continental shelf samples and are either carnivores or

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Fig. 2. The estimated number of extant species in each class of the phylum Mollusca, from three different sources. The estimated number of species from Ponder & Lindberg (2008) is extracted from relevant chapters, with the exception of bivalves, for which no species richness estimate was available. detritivores (Beesley, Ross & Wells, 1998). The enormous of marine molluscs (Table 1, Figs 3, 4), with respect to adaptive radiation in the biology of molluscs suggests that structural diversity (Fig. 5) and the taxonomic distribution of they are likely also to be diverse in terms of their chemistry the metabolites (Figs 6, 7). (secondary metabolism). Taking into consideration the chemical redundancy between species, at least 977 distinct compounds have been isolated from just 251 species in the annual reviews (2) Molluscan chemical diversity of marine natural products (Faulkner 1984a, b, 1986, Marine molluscs have become the focus of many chemical 1987, 1988, 1990, 1991, 1992b, 1993, 1994, 1995, 1996, studies aimed at isolating and identifying novel natural 1997, 1998, 1999, 2000, 2001, 2002; Blunt et al., 2003, products. As scant information is available on the chemistry 2004, 2005, 2006, 2007, 2008, 2009). Similarly, Alam & of terrestrial and freshwater molluscs, this review focuses Thomson (1998) report 585 compounds from just 135 species on marine species. In the last three decades over 1000 of marine molluscs. This implies that chemical diversity research papers have been published on molluscan secondary in molluscs exceeds their species diversity, although this metabolites (Table 1, Avila, 2006). Alam & Thomson (1998) chemical diversity encompasses many structurally related compiled a valuable reference book detailing 585 metabolites compounds derived from the same biochemical pathways. isolated from marine molluscs. Prior to this, Baker & Murphy The majority of compounds have only been isolated from (1976, 1981) compiled information on 148 molluscan a single species, although the same group of compounds metabolites in their two-volume book ‘Compounds from is often found in molluscs from the same family or genus, Marine Organisms’. Faulkner (1984a, b, 1986, 1987, 1988, when related species are investigated. Infrequently, the same 1990, 1991, 1992, 1993, 1994b, 1995, 1996, 1997, 1998, secondary metabolites converge across diverse classes of 1999, 2000, 2001, 2002) included molluscs in his annual molluscs (e.g. Roseghini et al., 1996; Derby et al., 2007). The review of marine natural products, reporting a total of 729 distribution of compounds across taxonomic groups and compounds from 199 species. These reviews have been species (Fig. 3 & 4) is primarily due to the fact that the continued in recent years by Blunt et al. (2003, 2004, 2005, associated research effort is far from equal. The frequency 2006, 2007, 2008, 2009) reporting a further 190 distinct distribution histogram of the number of species reported compounds from 75 species. Cimino & Gavagnin (2006) to contain multiple metabolites complied from Faulkner provide a modern review of marine molluscan chemistry and (1984a, b, 1986, 1987, 1988, 1990, 1991, 1992b, 1993, biotechnology, including chapters on secondary metabolites 1994, 1995, 1996, 1997, 1998, 1999, 2000, 2001, 2002) from marine gastropods from Antarctica, Southern Africa and Blunt et al. (2003–2007) reveals a strong right-hand and South America (Davies-Coleman, 2006); Australia and skew, with a median number of two and a maximum of New Zealand (Garson, 2006), Indo-Pacific (Wahidullah et al., 58 compounds isolated from a single species of mollusc 2006) and Japan (Miyamoto, 2006). These references offer (Fig. 4). The majority of species have only been subject to a excellent opportunities to examine the chemical diversity single study aimed at the isolation of novel natural products.

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Fig. 3. The number of (A) species subject to chemical investigation and (B) distinct secondary metabolites isolated from each class in the phylum Mollusca, sourced from the indicated reviews and syntheses of marine natural products.

These studies typically yield a small group of structurally the algal diets of these cosmopolitan grazing seahares. The related compounds (analogues). However, some species have bivalve Patinopectin yessoensis contained the second highest been intensively studied, often by several different research number of compounds (33, Fig. 4) with a range of sterols and groups working on collections from different biogeographic algal toxins. Clearly dietary sources contribute significantly regions. These collections may yield distinct compounds, to the chemical diversity found in molluscs. Nevertheless, according to the isolation procedures and focus of the specific evidence for de novo biosynthesis has been reported in several researchers. For example, 25 compounds have been isolated molluscan taxa (see reviews by Garson, 1993; Cimino & from the seahare Aplysia kurodai including terpenes, nitrogen- Ghiselin, 2001; Moore, 2006; Fontana, 2006). Evidence for containing aliphatic compounds, macrolides and fatty acid the biogenesis of secondary metabolites mostly stems from derivatives. In 2008, a further eight novel metabolites were feeding experiments, which demonstrate the incorporation isolated from this species (Blunt et al., 2009) collected from of radio-labeled precursors in certain groups of heterobranch new locations. molluscs (reviewed by Cimino et al., 2004; Fontana, 2006). The 58 compounds reported from the related species More recently, these experiments have been complemented A. dactylomela (Fig. 4) are primarily terpenes derived from by genetic studies (Fontana, 2006) or histochemistry (Westley

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Fig. 4. Frequency distribution of the number of compounds found in molluscan species complied from Faulkner (1984a,b–2002) and Blunt et al.’s, (2003, 2004, 2005, 2006, 2007) annual reviews of marine natural products. Fifty-eight compounds have been isolated from Aplysia dactylomela (inset with egg masses).

& Benkendorff, 2009) aimed at identifying the biosynthetic II. TAXONOMIC DISTRIBUTION OF processes. MOLLUSCAN METABOLITES The secondary metabolites isolated from molluscs fall into a wide range of structural classes, with some compounds The number of species subject to natural products research being more dominant in certain taxa. Fig. 5 shows the in the different molluscan taxa (Figs 3A, 6A) is likely to relative proportions of different compounds in the two major be influenced by their relative diversity (Fig. 2), as well as molluscan classes compiled from Alam & Thomson (1998), their accessibility for research and how well their biology is Faulkner (1984a, b, 1986, 1987, 1988, 1990, 1991, 1992b, currently understood. In general, the diversity of compounds 1993, 1994, 1995, 1996, 1997, 1998, 1999, 2000, 2001, isolated from the different molluscan classes (Figs 3B, 6B) 2002) and Blunt et al. (2003, 2004, 2005, 2006, 2007). All does mirror their relative species richness (Fig. 2), but even the major types of secondary metabolites are represented the two best studied classes (gastropods and bivalves) are in both classes of mollusc (Fig. 5). In the Gastropoda only represented by a tiny proportion of the available species (Fig 5A) terpenes dominate, whereas only three terpenes (Table 1). Overall, less than 1% of named molluscan species were reported from bivalves (Fig. 5B). Terpenes have been have been investigated for their secondary metabolites. a major focus of research in soft-bodied grazing gastropods, which may acquire these compounds from their diet for (1) Minor classes use in their own defence (e.g. Faulkner, 1984a, 1992a). The Monoplacophorans and Aplacophorans have not been Sterols are the dominants compounds reported from bivalves subject to any chemical studies to date (Fig. 3, Table 1). (Fig. 5B), but are the least frequently reported in gastropods This is not surprising considering the dearth of biological (Fig. 5A). The relatively large research effort on sterols in knowledge on these groups and the fact that they are all deep- bivalves is probably due to their importance in fisheries sea organisms with relatively small body sizes (Beesley et al., and aquaculture, with interest focusing on biochemical 1998). The Monoplacophora, with few living representatives, changes over the reproductive cycle. Polyproprionates and triggered much excitement when extant specimens were first alkaloids have been isolated in reasonably large numbers discovered, as it was suggested that they are the closest to the from both classes of molluscs, whereas aliphatic nitrogen- hypothetical ancestral mollusc (e.g. Morton, 1979). However, containing compounds are relatively uncommon (Fig. 5). more recent research indicates that the ancestral mollusc is Further discussion on the types of compounds found within more aplacophoran-like (Haszprunar et al., 2008), although certain molluscan taxa is given below for each group. This this is not universally accepted (e.g. Ponder and Lindberg, includes additional compounds found by extensive database 1997). The Aplacophora are worm-shaped molluscs that do searching that are not represented in general reviews on not have a shell, but rather are covered by a cuticle embedded marine natural products. with calcareous spicules (Beesley et al., 1998). Consequently,

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Fig. 5. The relative proportion of secondary metabolites in different structural classes for (A) Gastropoda (N = 661) and (B) Bivalvia (N = 92) complied from complied from Alam & Thomson (1998), Faulkner (1984a, b, 1986, 1987, 1988, 1990, 1991, 1992b, 1993, 1994, 1995, 1996, 1997, 1998, 1999, 2000, 2001, 2002) and Blunt et al.’s, (2003, 2004, 2005, 2006, 2007) annual reviews of marine natural products. Miscellaneous refers to compounds of mixed biosynthetic origin. they could be of great interest for investigations into the Otway, 1997). No interesting secondary metabolites from evolution of chemical defence mechanisms and may provide these molluscan classes have been reported in the various interesting insights into the early evolution of biosynthetic reviews of marine natural products (Fig. 3B), although further pathways in the Mollusca. However, further research on database searching revealed a handful of chemical studies their biology should ensure the collection of specimens is (Table 1) on the sterols (Kind & Meigs, 1955; Teshima environmentally sustainable (Benkendorff, 2002). Additional & Kanazawa, 1973, 1978; Voogt & Van Rheenen, 1974; information on their biological attributes would further Teshima et al., 1982), lipids (Lawrence, 1970; Hayashi contribute to effective targeting of species for biodiscovery et al., 1991), hydrocarbons (Yasuda & Fukamiya, 1977) and research. carotene derivatives (Tsushima, Maoka & Matsuno, 1989) of Two other minor classes of molluscs, the Polyplacophora chitons. One study has also investigated the sterols and lipids and Scaphopoda, have also been neglected in the natural of scaphopods (Kandaswarmy & Rajulu, 1978) and at least products literature (Fig. 3, Table 1). The Scaphopoda are two further investigations have incorporated a scaphopod difficult subjects for research because they live completely and/or chitons in comparative studies of sterols in molluscs buried in sediment (Beesley et al., 1998). On the other hand, (Teshima & Kanazawa, 1972; Idler et al., 1978). some species of Polyplacophora (chitons) are common on Future studies aimed at isolating novel secondary intertidal and shallow subtidal reefs. They can reach large metabolites from these two molluscan classes would be sizes (>10 cm) and their biology is relatively well understood strongly justified based on their distinct biological attributes. (e.g. Boyle, 1977; Otaiza & Santelices, 1985; Smith & The Scaphopoda are detritivores and microcarnivores with

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Fig. 6. The number of (A) species subject to chemical investigation and (B) distinct secondary metabolites isolated from Eogastropoda and major groups within the Orthogastropoda, sourced from the indicated reviews and syntheses of marine natural products. a tubular shell open at both ends (Poon, 1987). Their reduced or internal shell plates. Given the early divergence feeding habits involve extending cilia outside their shells of chitons from other molluscan classes, there is potential for to probe the sediment, which could make them vulnerable the discovery of novel secondary metabolites. to predators and pathogens. The Polyplacophora are algal grazers protected by an eight-plate shell on their dorsal surface, which is surrounded by a leathery girdle. Although (2) Cephalopoda their muscular foot on the ventral surface typically clings Despite also being one of the minor classes in terms of very tightly to the rocky substratum, they may be vulnerable species richness (Fig. 2), the Cephalopoda have been subject to pathogens in biofilms and some predators, particularly to several interesting chemical studies (Table 1, Fig. 3). With when moving or detached. Such potential vulnerability could the exception of Nautilus spp., these predatory species are soft- have led to the evolution of chemical defence strategies, bodied organisms, which have secondarily lost the protection particularly in some large slug-like species that have very of the shell as an adaptation to an active pelagic life style.

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Fig. 7. The number of (A) species subject to chemical investigation and (B) distinct secondary metabolites isolated from different groups within the Heterobranchia, sourced from the indicated reviews and syntheses of marine natural products.

Their highly evolved behaviour, camouflage and ability to Myers et al., 1993), more research effort into the venoms move fast was assumed to reduce the need for chemical and secretory glands of cephalopods is likely to be highly defence against predators (e.g. Faulkner, 1992a), although rewarding. the secretion of ink is a cephalopod chemical defence In addition to the venoms, a range of other natural mechanism (Derby et al., 2007). They also remain vulnerable products have been isolated from the Cephalopoda. These to pathogens in the microbially rich marine environment. include cytotoxic tyrosinase from the ink of the cuttlefish Sepia Furthermore, as active predators with venom glands to officinalis (Russo et al., 2003), as well as ovarian jelly peptides subdue their prey, there is much scope for investigation (Bernay et al., 2006), a sperm-attracting peptide (Zatylny et al., into the chemical composition of their venom secretions. 2002) and novel cardioactive peptide isolated from the brain Secondary metabolites isolated from cephalopod venom, to of the common octopus Octopus vulgaris (Kanda & Minakata, date, include tetrodotoxin (Scheumack et al., 1978), peptides 2006). The compounds reported from cephalopods in Baker (e.g. Songdahl & Shapiro, 1974; Kanda et al., 2003) and & Murphy (1981) include fairly simple, widely distributed biogenic amines (e.g. Roseghini et al., 1996). Given the compounds, such as aromatic amino acids, noradrenaline high diversity of bioactive peptides found in the predatory and some benzoquinols. Overall, despite the relatively high caenogastropods in the family Conidae (Olivera et al. 1990, representation of species studied in this class (Table 1), there

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society 10 Kirsten Benkendorff has been fairly limited research into their biologically active products studies (Figs 3, 6, Table 1). Although a relatively organic molecules. There remains good potential for novel large number of compounds have been isolated, the number discoveries within cephalopods, and this is in part being of species included in these studies still represents only addressed by a current Australian Research Council funded a tiny proportion of the possible pool of extant marine collaborative research project into the evolution of their species in what is by far the largest class of molluscs toxicity (Norman, 2008). (∼1%, Table 1). Given the large diversity of species, it is appropriate to break the gastropods into major groups (3) Bivalvia (Fig. 6). The gastropods were traditionally divided into three subclasses: Prosobranchia, Pulmonata and Opisthobranchia, The second largest molluscan class, the Bivalvia (Fig. 2), but are now considered to fall into two major subclasses: is relatively well represented in chemical studies (Fig. 3, Eogastropoda (true limpets- Patellogastropoda being the Table 1). However, much of this research is associated only living representatives) and Orthogastropoda (Ponder & with sterols (Fig. 5B) and bioaccumulated toxins responsible Lindberg, 1997; 2008), encompassing all other gastropods for paralytic shellfish poisoning (see reviews by Bricelj & (recognized groups Vetigastropoda, Caenogastropoda and Shumway, 1998; Llewellyn, Negri & Robertsin, 2006; Kita Heterobranchia treated as superorders). The pulmonates & Uemura, 2006; Ciminiello & Fattorusso, 2006), as opposed and opisthobranchs share a common ancestor and are to defensive secondary metabolites. Some bivalves have now combined under one monophyletic group Euthyneura, adapted to use their dietary toxins in their own defence which is included in the Heterobranchia (Thollesson, against predators (e.g. Kvitek & Bretz, 2005). Most bivalves 1999; Wagele¨ et al., 2008). can also clamp their shells tightly shut to enclose the entire animal, thus providing them with some physical protection (a) Eogastropoda and non-heterobranch orthogastropods from predators. Nevertheless, as sedentary suspension- feeding organisms that pump large volumes of sea water and Taxonomic breakdown of the gastropod metabolites reveals suspended microorganisms through their gills, they are quite that only three species of patellogastropod limpets have been vulnerable to microbial infection, thus requiring chemical studied (Fig. 6A). Twenty-three structurally related fatty acid defence against pathogens. This has been demonstrated derivatives have been isolated from two of these species in recent research into the involvement of antimicrobial (Kawashima, 2005; Kawashima & Ohnishi, 2006), whilst the peptides and proteins in the humoral immunity of bivalves third species yielded a defensive terpene (Pawlik et al., 1986). (e.g. Mitta et al., 2000a, b; Cellura et al., 2007; Zhao et al., Further research into the chemistry of this ancestral class of 2007; Li et al., 2009). Some interesting polyproprionates gastropods is justified, especially considering that limpets are and alkaloids (aromatic nitrogenous compounds) have also only protected by a shell on their top surface and are found been isolated from bivalves (Fig. 5B), although it is presently in very large numbers on most rocky intertidal shores. unclear if these play any defensive role. In the Orthogastropoda, over 91% of the compounds The detection of antimicrobial defence factors in have been isolated from the marine heterobranchs shelled molluscs such as bivalves has been influenced by (Fig. 6B). Only a single species has been studied from recent advances in our understanding of the biosynthesis the Neritimorpha (=Neritopsina, Fig. 6A). Notably, five and regulated expression of these bioactive compounds. antimicrobial isoflavones have been isolated from this nerite Research on a mussel, Mytilus galloprovincialis, revealed a (Sanduja et al., 1985), but it is uncertain whether these are diversity of antimicrobial peptides (reviewed by Mitta et al., de novo synthesised or acquired from their diet. As Nerita 2000b). Interestingly, some of these peptides are expressed spp. are microalgal grazers on rocky intertidal platforms, constitutively, whilst some are only induced after infection comparisons of the chemical composition of these herbivores with bacteria (Mitta et al., 2000a; Cellura et al., 2007). to their local diet would help resolve the origin of these Similarly, an antimicrobial protein from the scallop Argopecten bioactive isoflavones. Nerites have a broad global distribution irradians is rapidly upregulated after bacterial challenge and some neritimorphs can be found in freshwater and (Zhao et al., 2007). The possibility that bioactive secondary terrestrial habitats. Studies on the chemical consistency of metabolites are not always present in the humoral defence extracts from species across different habitats and tropical system in molluscs means that their bioactivity may be and temperate locations could add to the diversity of overlooked in general screening programs. Strategic research secondary metabolites recorded for this abundant group. into regulated defence mechanisms may provide novel In early studies, Baker & Murphy (1976, 1981) report seven discoveries in even the most well-studied molluscan taxa, compounds isolated from seven different vetigastropods. including commercially important bivalves, which are in However, only an additional 12 compounds from three relatively abundant supply for future drug development species have been reported in more recent annual reviews of (Benkendorff, 2009). marine natural products (Fig. 6B). Consequently, this species- rich group is hugely under-represented in chemical studies. Vetigastropoda contains extremely diverse families, such as (4) Gastropoda the Trochidae (topshells) and many large common species Consistent with their high species diversity, the gastropods in the families Haliotidae, Turbinidae and Fissurellidae are the most highly represented molluscan class in natural (see Geiger, N¨utzel & Sasaki, 2008, for a comprehensive

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society Molluscan biological and chemical diversity 11 review of this group). Chemical studies undertaken on and thus remain under-represented. Myers et al. (1993) the turbinid snails revealed toxins, which are possibly provide sequence data for 37 peptides from eight distinct dietary derived (Kigoshi et al., 2000), as well as four pharmacological and structural classes. There are 500–700 novel cytostatic glycosphingolipids (Pettit, Tang & Knight, species of Conus, and of the species studied so far, each 2005). The only documented chemical study on a trochid appears to produce between 40 and 200 different peptides (Callistoma canaliculatum), yielded a novel toxin 6-bromo-2- with little overlap among species (Myers et al., 1993; Terlau mercaptotryptamine from hypobranchial gland secretions & Olivera, 2004; Livett et al., 2004), suggesting enormous (Kelley et al., 2003). Interestingly, this compound also bears chemical diversity within this one family of molluscs. Indeed a remarkable structural similarity to the bromoindoxyl sulfate complex library of up to 100,000 bioactive peptides has been precursors of Tyrian purple secreted from the hypobranchial predicted, each with specific physiological targets (Buczek glands of the caenogastropod family Muricidae (Fig. 1F- et al., 2005). I; Baker & Sutherland, 1968; Cooksey, 2001; Westley & Benkendorff, 2008). The hypobranchial gland is a uniquely molluscan feature (Beesley et al., 1998) and its functional role (b) Heterobranch gastropods in molluscs is not currently well understood (Westley, Vine & There is a strong bias in the natural product literature Benkendorff, 2006). However, histological investigations in towards research on marine heterobranch gastropods (Figs gastropods have revealed it to be a complex organ containing 6, 7), in which the shell is often reduced or completely absent. a range of secretory cells (e.g. Fretter & Graham, 1994; Several reviews focus on the chemical ecology of these soft- Roller, Rickett & Stickle, 1995; Westely & Benkendorff, bodied opisthobranchs ( Karuso, 1987; Cimino and Sodano, 2009). Consequently, further investigation of the chemical 1989; Faulkner, 1992a; Avila, 1995; 2006; Cimino et al., secretions associated with this gland across a broad range of 2001; Wagele,¨ Ballesteros & Avila, 2006; Wahidullah et al., molluscs would be worthwhile. 2006; Miyamoto, 2006; Fontana, 2006). The taxonomy In total, 40 species of caenogastopod are represented in of Heterobranchia is yet to be fully resolved (Thollesson, the annual reviews of marine natural products (Fig. 6A) 1999; Wagele¨ et al., 2008; Mordan & Wade, 2008), but and these have yielded 49 compounds (Fig. 6B). This is further knowledge of their phylogenetic relationships will be the largest group of gastropods (Aktipis et. al., 2008) but beneficial for understanding the full chemical diversity of this has been largely overlooked by natural products researchers group and the potential for shared biosynthetic pathways due to the presence of a protective shell. However, further among taxa. For example, the pulmonates are sister taxa literature searches revealed that three classes of compounds to the main opisthobranch radiation, which is consistent known to occur in caenogastropods are under-represented with the prevalent biosynthesis of polyproprionates (Garson, in the annual reviews used to generate Fig. 6. Roseghini 1993; Cimino & Ghiselin, 2001; Moore, 2006; Fontana, et al. (1996) examined the distribution of five choline esters 2006) across a range of heterobranch taxa including the (e.g. Fig. 1E) and six biogenic amines in 55 species of the Siphonaridae (basommatophoran pulmonates), Plakobranchus caenogastropod subgroup Neogastropoda. Cooksey (2001) sp. (saccoglosan opisthobranch) and Pleurobranchus sp. reviewed Tyrian purple (Fig. 1I) and related indoles isolated (notospidean opisthobranch). from the hypobranchial glands of 17 species of Muricidae. Within the heterobranchs there is clearly unequal Antimicrobial secondary metabolites from the egg masses of a representation of the different orders in chemical further eight species of Muricidae have also been investigated studies (Fig. 7). Four relatively minor groups, Acochlidea, (Benkendorff, Bremner & Davis, 2000; 2001; Benkendorff, Rhodopemorpha, Thecosomata and Gymnosomata, have Pillai & Bremner, 2004), revealing 16 indole derivatives, not been subject to any chemical studies to date. The many of which are brominated, as well as brominated majority of research on the Heterobranchia has concentrated imidazoles and several small mercaptans. Evidence for on the largest and most charismatic marine group, the the biosynthesis of the brominated indoles is provided by Nudibranchia (Fig. 7), with over 386 compounds isolated bromoperoxidase activity in the hypobranchial glands of from 102 species. The Anaspidea (seahares e.g. Aplysia spp. Trunculariopsis (Murex) trunculus (Jannun & Coe, 1987) and and Dolabella spp.) are also well represented with 18 species Dicathais orbita (Westley & Benkendorff, 2009). yielding an amazing total of 247 compounds documented Numerous reviews have been dedicated to the diversity, in the annual reviews of marine natural products (Fig. 7). structure and pharmaceutical applications of conotoxins A broad range of compounds has been isolated from these from predatory marine cone shells (e.g. Olivera et al., 1990; ‘seahares’ (Kamiya, Sakai & Jimbo, 2006), dominated by Myers et al., 1993; Craig, 2000; Livett, Gayler & Khalil, 2004; terpenes (Fig. 5), but also including sterols (e.g. Miyamoto Terlau & Olivera, 2004; Buczek, Bulaj & Olivera, 2005; et al., 1988, Yamaguchi et al., 1992), fatty acid derivatives Prommer, 2006). Recently identified neurotoxic peptides (e.g. Spinella et al., 1997) and alkaloids (e.g. Kigoshi et al., are included in Blunt et al.’s, (2003, 2004, 2005, 2006, 1990). A range of cytotoxic peptides (e.g. the dolastatins, 2007, 2008, 2009) reviews of marine natural products and reviewed by Poncet, 1999) and glycoproteins (e.g. Iijima, contribute significantly to the number of caenogastropod Kisugi and Yamazaki, 2003; Yamazaki, 1993) have also species and compounds in Fig 6, although they were been isolated from the Anaspidea, as well as several not included in earlier reports by Faulkner (1984b–2002), proteinaceous sex pheromones (Cummins et al., 2006). Some

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society 12 Kirsten Benkendorff other heterobranch groups, such as the Cephalaspidea and III. MOLLUSCAN MEDICINES Saccoglossa are also reasonably well represented in chemical studies (Fig. 7). In addition to a range of polyproprionates, Molluscs have long provided a source of medicinally the Cephalaspidea produce phenolic and alkaloid alarm useful products for many cultures around the world. pheromones isolated from a range of species including Pliny the Elder described the use of certain molluscs, Navanax inermis (e.g. Sleeper, Paul & Fenical, 1980), Haminoea such as terrestrial pulmonates, in medicinal remedies from navicula (e.g. Cimino et al., 1991) and Bulla spp. (Marin et al., Ancient Rome (Bailey, 1929; Bonnemain, 2005). Bivalve 1999), as well as cytotoxic peptides from Philinopsis spp. (e.g. mussels (Mytillidae) were used as therapy in ancient Crete Nakao et al., 2004). Chemical defence in the saccoglossan (Kamm, 1997) and more recently have been subject to Eylsia spp. is also the source of several cytotoxic peptides, several patents as a source of antimicrobial (de Faire, the kahalalides (e.g. Hamann et al., 1996; Goetz, Nakao & 1999; Roch et al., 2001) and antiviral (Rothman, 1984; Scheuer, 1997, Horgen et al., 2000) and diterpenes (Paul & Bichurina et al., 2001) peptides. Molluscs also feature in Van Alstyne, 1988), which appear to be dietary derived. a number of traditional medicines (Fig. 8) from South Chemical studies on the marine pulmonate gastropods Africa (Herbert et al., 2003), India (Prabhakar & Roy, 2009) have focused largely on the polyproprionates from the limpet- and China (Hu, 1980; Yeung, 1983). Several molluscan- like Siphonaria spp. (Darias, Cueto & D´ıaz-Marrero, 2006), derived therapies are listed on the Homeopathic Materia although some novel fatty acid derivatives have also been Medica (Boericke, 1999, Fig. 8) and extracts from the isolated from species in this genus (Carballeira et al., 2001). New Zealand green-lipped mussel, Perna canaliculus,are Polyproprionates have also been isolated from the intertidal included in the Natural Medicines Comprehensive Database systellommatophoran slug Onchidium sp. (Rodriguez, Riguera (www.naturaldatabase.com). Currently, several bioactive & Debitus,1992), in addition to a mixture of pyrone esters natural products from molluscs are under development (Ireland et al., 1984) and a novel peptide (Fernandez et al., for pharmaceutical drugs (Simmons et al., 2005; Fig. 8). 1996). Earlier studies on the defence secretions of its sister Recently, Ziconotide, derived from the venom of predatory genus Onchidella revealed a lipid-soluble compound onchidal cone snails, was the first marine drug to be approved for (Ireland & Faulkner, 1978). No further studies have been clinical use (Prommer, 2006), as a treatment for chronic pain. undertaken on the chemistry of this genus or any other Dolastatin 10, and synthetic analogues, from the aplysiid systellommatophorans. The much larger pulmonate group Dolabella auriculata are currently in Phase II clinical trials as Eupulmonata is only represented by a few studies on the anticancer agents (Madden et al., 2000). Kahalalide F first sterols and terpenes of Trimusculus sp. (e.g. Manker & identified from the saccoglossan Elysia rufescens (Lopez-Macia Faulkner, 1996; San-Martin et al., 1996). The Eupulmonata et al., 2001), as well as ES-285 from the bivalve Mactromeris includes the large and diverse family Ellobiidae, which polynyma have passed through Phase I clinical trials (Den are mostly marine; eupulmonates are otherwise primarily Brock et al., 2005; Faircloth & Cuevas, 2006). terrestrial and thus are not included in annual reviews of There appears to be no correlation between the number marine natural products. of species from different molluscan taxa that are used There are no equivalent reviews of the literature in medicines and the number of species that have been on terrestrial invertebrate natural products and as a subject to investigation in the natural products literature consequence it is much more difficult to find chemical (r2 = 0.04). The lack of correlation is largely driven by the studies on terrestrial species. Extensive searches of scientific heterobranch molluscs, which rarely feature in molluscan databases reveal very few studies on secondary metabolites medicines (Fig. 8B), despite intensive chemical investigation from terrestrial pulmonates, with a much greater focus (Fig. 7). Conversely, the Cephalopoda are used in a wide on plant metabolites that may be useful as molluscicides. range of traditional medicines (Fig. 8A), but there are However, the mucoproteins produced by garden snails, Helix relatively few chemical investigations (Fig. 3). Listings of spp., have been studied since the early 1900s (Levene, 1925) Cephalopods in the Chinese Materia Medica include squid and there has been considerable interest in the bioactive and cuttlefish (Sepia sp.) ‘‘bone’’, as well as the meat, ink and properties of the mucus-associated lectins (reviewed by eggs from cuttlefish (Hu, 1980). Sepia ink is also listed in the Bonnemain, 2005). Some novel terpenes that inhibit HIV-1 homeopathic Materia Medica for the treatment of depression reverse transcriptase have been isolated from extracts of (Cazalet, 2007). Four cephalopod species were listed in a the giant African snail Achatina fulica (Patil et al., 1993) survey of South African medicines; the internal shells of and cardioexcitatory peptides have been identified from the the species Spirula spirula were the most expensive marine freshwater snail Lymnaea stagnalis (Tensen et al., 1998). There is invertebrate on sale at the traditional medicine market in evidence that some terrestrial pulmonate snails can sequester Durban (Herbert et al., 2003). The medicinal applications bioactive terpenes from their lichen diets (Hesbacher et al., of these species remain unknown and they are yet to be 1995) and terrestrial slugs, such as Arion lusitanicus, have been subjected to any detailed natural product investigations. shown to sequester and detoxify alkaloids from a variety of Several species of chiton (Polyplacophora) were also amongst plants (Aguiar & Wink, 2005). These studies highlight the the most valuable marine invertebrate taxa reported in a potential for further discoveries within the natural products survey of South African traditional medicines (Herbert et al., and chemical ecology of terrestrial pulmonates. 2003). These chitons are reported to cause vaginal spasm in

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society Molluscan biological and chemical diversity 13

Fig. 8. The number of taxa in (A) the different classes of Mollusca and (B) the Eogastropoda and major groups from the Orthogastropoda that are used in medicinal remedies. Taxa refers to distinct species, except for the Caenogastropoda, where the number of families is plotted due to related uses and limited information at the species level. Pharmaceuticals refers to compounds derived from molluscs that are commercially available or in clinical trials, whereas all of the natural and traditional remedies are commercially available. Nutraceutical is a modern term derived from the combination of nutritional and pharmaceutical and describes natural extracts or products purified from foods with demonstrated pharmacological activity. women and prevent bed wetting in children (Herbert et al., melancholic disposition (Cazalet, 2007; Boericke, 1995). 2003), suggesting that future chemical investigation into A minor constituent of this dye, 6,6’ dibromoindirubin bioactive metabolites could be worthwhile in this previously (Fig. 1H), has been shown to inhibit cell proliferation neglected class of molluscs (see Fig. 3). with selectivity towards glycogen synthase kinase-3 (GSK-3) In most cases there are no data to support the application α/β receptors (Meijer et al., 2003; Magiatis & Skaltsounis, of traditional and homeopathic remedies from molluscs. 2006). The intermediate dye precursor tyrindoleninone Indeed, there are few data to support the biological activity of (Fig. 1F) also appears to have selective cytotoxicity towards homeopathic remedies in general and few have been tested severalcancercelllines(Vineet al., 2007; Benkendorff, for safety and effectiveness using rigorous methodologies McIver & Abbott, 2009) and the oxidation product 6’ (Straus, 2000). One possible exception is the Murex remedy bromoisatin (Fig. 1G) is generally cytoxtoxic (Westley et al., (Fig. 1B) derived from the purple dye secretion of muricid 2006; Vine et al., 2007). The ultimate dye precursor is whelks (Muridicae; Caenogastropoda), such as Trunculariopsis held as a salt of choline esters. These choline esters (e.g. trunculus, used to treat pain and ‘women’s problems’ including murexine, Fig. 1E) display potent neuromuscular blocking uterine cancer, dysmenorrhoea, chronic endometritis, and muscle-relaxing activity, as well as nicotinic action metrorrhagia, leucorrhoea, nymphomania, anxiety and (Whittaker, 1960; Baker & Duke, 1976; Roseghini et al.,

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society 14 Kirsten Benkendorff

1996). These bioactive compounds could all potentially are common and cowrie shells (Cypraeidae) are incorporated contribute to the homeopathic action of the Murex remedy. into headbands and necklaces for sale at Durban markets However, the composition of dye extracts from different (Herbert et al., 2003). Cowrie shells also feature in Chinese species of Muricidae can vary according to the presence of traditional medicines (Hu, 1980) and have been patented for different ultimate precursors (Cooksey, 2001; Benkendorff use in dental fillings (Weil, 1994). et al., 2001). Neither the source species, nor the chemical Some terrestrial and freshwater caenogastropods in the composition of the homeopathic tincture, are provided basal group Architaenoglossa feature in traditional Indian on the commercially available products. Serial dilution medicine. The eggs of terrestrial apple snails from the is characteristic of homeopathy and even after 100 fold family Ampullariodoidae (Pila spp.) are used to treat rickets, concentration only traces of 6’ bromoisatin could bedetected whilst extracts from freshwater snails (Bellamya spp.) are in some batches of the commercially available Murex remedy used for inflammatory problems including asthma, arthritis and there was no evidence of anticancer effects in either and rheumatism, as well as a treatment for conjunctivitis the dilute or concentrated product (Benkendorff et al., 2009). (Prabhakar & Roy, 2009). As reviewed by Bonnemain (2005), Trace amounts of choline esters were found in some, but not the terrestrial land snail Helix spp. (Stylommatophora) has all, batches of the commercial product suggesting that, with been used in Western health care, from antiquity to the quality control, there could be chemical support for some present. Traditionally extracts and mucus from these snails homeopathic applications. In a different line of research, lipid were used to treat a range of ailments including hernias, extracts from the muricid Rapana venosa have been shown to tuberculosis, resistant colds bronchitis and other chronic be highly efficient in healing induced skin burns in Wistar chest diseases. Studies on the bioactive properties of the rats (Badiu et al., 2008). Overall, there is much potential to mucus from Helix have revealed mucolytic and bacteriolytic derive medicines from the Muricidae. activities, as well as sedative or antispasmodic properties (see Abalone (Haliotidae) is another family of gastropods that Bonnemain, 2005). Analysis of extracts of the mucus have is valued as a healthy delicacy and features prominently in revealed a wide range of enzymes and glycosaminoglycans, natural remedies. In traditional Chinese medicine abalone as well as prostaglandins, which may contribute to its shell and powder is used as a liver tonic, to improve vision and biological effects. Cryptomphalus (Helix) aspersa mucus is to treat cataracts, hypertension, vertigo and limb convulsion incorporated into a commercially available cosmaceutical (Yeung, 1983). Dried abalone power from New Zealand called BIOSKINCARE™, which is claimed to ‘‘repair scars, is sold as a nutraceutical (defined as products purified from prevent and eliminate new stretch marks and firm breasts’’ foods with demonstrated protective or physiological benefits). (BioSkinCare, 2009). Recent pharmacological studies on Abalone powder is promoted for generally enhancing health C. aspersa secretions support the skin-regenerative properties and sexual life, as well as supporting the immune system through evidence of anti-oxidant activity, stimulation of and blood circulation, preventing anaemia and providing fibroblast proliferation and extracellular matrix assembly, minerals, vitamins and omega 3 unsaturated fatty acids as well as regulation of metalloproteinase activities (Brieva (Aroma New Zealand Ltd., 2007). Powder of abalone shell is et al., 2008). also incorporated into a patented formula for treating burns Natural and traditional medicines from bivalve molluscs without pain and scaring (Lee, 1994). The haemolymph of (Fig. 8A) primarily comprise ground shells, whole-body abalone has demonstrated antibacterial (Li, 1960; Vakalia powders and mother of pearl from the subclass & Benkendorff, 2005) and antiviral (Li, Prescott & Jahnes, Pteriomorphia, including oysters (Ostreina), pearl oysters 1962A, Li et al., 1962B) properties against a range of human (Pterioida), clams (Arcoida) and mussels (Mytiloida). Oyster pathogens in vitro. The antiviral fraction was shown to protect shell lysate from Ostrea edulis provides a bioavailable form mice infected with poliovirus and influenza (Li et al., 1962A). of calcium carbonate for osteoporosis patients (Fujita et al., These studies indicate that the active factors are likely to be 1990) and is used in the homeopathic treatment of bone macromolecules such as glycoproteins. However, treatment deficiencies (Cazalet, 2007). Oyster shell has also been shown of the abalone haemolymph with heat or proteases to destroy to prevent gastric ulcers in vivo using a rodent model (Nie the proteins caused no reduction in the antibacterial activity, et al., 1994). Oyster shell (Crassostrea gigas, Ostrea spp.) is whereas the lipophylic compounds extracted on a dianion used as a Chinese remedy to treat headaches, dizziness, resin column were found to be active (Benkendorff, K., palpitations, insomnia, sweating, leucorrhea and uterine unpublished data). Further bioassay-guided fractionation bleeding (Yeung, 1983), whereas oyster extract powder is required to isolate and identify the active factors from (Crassostrea gigas) is promoted as a dietary supplement abalone. containing natural taurine and zinc for cardiovascular health, Shelled gastropods from at least 13 families feature in the liver problems, arthritis, skin problems wound healing and South African traditional medicine market (Herbert et al., resistance to infection (Aroma New Zealand Ltd., 2007). 2003), although it is unclear what these are actually used Several alkaloids have been isolated from Ostrea rivularis,a for. In some cases they may be used as symbols or regalia traditional Chinese medicine used to treat vertigo, tinnitus, by traditional healers to bring good fortune or ward off pulmonary tuberculosis and to reduce phlegm (Ouyang, misfortune, rather than as curative medicines per se.For 2006). Water extracts from oysters (Ostrea gingas syn. Crassostrea example, wrist bands consisting of the shells of Nerita albicilla gigas) were found to inhibit liver damage and stimulate

Biological Reviews (2010) 000–000 © 2010 The Author. Journal compilation © 2010 Cambridge Philosophical Society Molluscan biological and chemical diversity 15 lipogenesis in rats (Kimura, Ohminami & Okuda, 1998). IV. CONCLUSIONS Taurine has been identified as the active ingredient in the Chinese remedy Zhenzhu Jingmu from the serum of (1) Natural products research aimed at the isolation pearl oyster Pinctada martensii gonads, causing constrictions and identification of novel secondary metabolites in guinea pig uterus and reducing bleeding time in mice has only been undertaken on a small proportion of (Xu, 1997). Mother of pearl and pearls (Pinctada spp.) molluscan species to date. This research has been provide another Chinese remedy (Hu, 1980), used as a traditionally biased towards soft-bodied heterobranchs topical eye ointment and to treat various skin conditions. (opisthobranchs) based on their apparent vulnerability A cream containing mother of pearl has been patented for due to the lack of a physically protective shell. This dermatological and cosmetic use (Camprasse & Camprasse, approach has been rewarding, with an extremely 1998). Additional Chinese remedies derived from the pearl diverse range of compounds revealed, including several oyster Pteria margaritifera are used to treat headache, vertigo, drug leads in clinical trials. dizziness, tinnitius and cataracts (Yeung, 1983). The arcid (2) The under-representation of other molluscan taxa in bivale (Arca inflata) is also listed as a Chinese remedy, used for natural products research appears to be unjustified goitre, spleen, liver and gastric problems, as well as duodenal given that most molluscan traditional medicines are cancer (Yeung, 1983). Oyster and clam extracts have been actually derived from shelled molluscs. The bioactivity shown to have antibacterial and antiviral properties similar of many molluscan traditional medicines is yet to be to abalone (Haliotis spp., Li et al., 1962b; Prescott et al., 1966; substantiated, but preliminary data available from Olicard et al., 2005). bivalves, cephalopds and caenogastropods suggests Natural remedies from the New Zealand green-lipped that there is likely to be some chemical basis to mussel (Perna canaliculus) species are generally promoted as their medical applications. Further studies aimed at identifying the bioactive factors in well-known cultured anti-inflammatory agents, effective in halting the progression molluscs, such as abalone, would be valuable given of joint and connective tissue problems and relieving the their wide use for a range of medicinal applications. symptoms of arthritis. Several novel anti-inflammatory (3) Broadening the search field to encompass terrestrial omega 3 polyunsaturated fatty acids have been identified gastropods and some minor molluscan classes could from P. canaliculus extracts (Treschow et al., 2007). A range also be worthwhile given the possibility for divergent of nutraceuticals are produced from this species including evolution of metabolic pathways and distinct selective mussel powder, a mussel extract claimed to have up to five pressures for secondary metabolites. In particular, times greater anti-inflammatory properties than the powder the polyplacophorans would be worthy of further (Aroma New Zealand Ltd., 2007) and a stabilized lipid investigation given their availability, the reasonably extract (Lyprinol), which has been patented and proven large size of some species and their apparent use in effective in a range of clinical trials for arthritis and asthma traditional South African medicines. (Gibson, 2000; Halpern, 2000). The cost of these products (4) Overall, there is a need for more targeted research − increases accordingly; $AUD 150 kg 1 for the powder, based on specific hypotheses related to molluscan − − $1600 kg 1 for the extract and around $16,000 kg 1 for bioactivity and the defence systems of shelled molluscs, Lyprinol. Lyprinol has been shown to be effective in a to maximize the success rates of future investigations. rodent model for prevention of inflammatory bowel disease (Tenikoff et al., 2005) and pharmaceutical preparations containing P. canaliculus extracts have been patented for use V. ACKNOWLEDGEMENTS in treating side effects caused to gastro-intestinal mucosa after oral ingestion of analgesics (McFarlane & Croft, 1984). I would like to thank Winston Ponder (Australian Museum) To date, no species of Perna or other related bivalves and Prof. G. Cimino (Italian National Council of Research) outside New Zealand appear to have been tested for anti- for useful comments and references that have contributed to inflammatory activity. However, a lipid extract from Mytilus this review. I would also like to thank Chantel Westley, Vicki galloprovincialis was found to reduce the healing time for Edwards, Casey Campleman, Warwick Noble and Patrick induced skin burns in a rodent model, similarly to results Laffy from the mollusc lab at Flinders University for useful for the muricid Rapana venosa (Badiu et al., 2008). In India, feedback. shell powder from the freshwater mussels Lamellidens spp. and Parreysia spp. (Paleoheterodonta: Unionoidea) is used to control blood pressure, giddiness and dehydration, whereas a VI. REFERENCES soup prepared from the foot is used to treat cardiac aliments (Prabhakar & Roy, 2009). This is the only study that refers to Aguiar,R.&Wink, M. (2005). 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