Isolation of activity and partial characterization of large non-proteinaceous lytic allelochemicals produced by the marine dinoflagellate Alexandrium tamarense

Abstract

Certain strains of the toxigenic dinoflagellate Alexandrium tamarense produce potent allelochemicals with lytic activity against a wide variety of marine microorganisms. Our efforts to characterize these allelochemicals from a lytic strain focused on the less polar components because of their higher lytic activity.Fractionation and partial purification after solid phase extraction (SPE) were achieved via alternative chromatographic methods, namely HPLC separation on C8 and HILIC phases. Through MALDI-TOF mass spectrometry we compared the mass differences in SPE, C8 HPLC, and HILIC fractions between a lytic and non-lytic strain of A.tamarense. Several large species with masses between 7 kDa and 15 kDa were found in the HILIC lytic fraction by MALDI-TOF MS. Tryptic digestion and tryptic digestion-coupled size-exclusion chromatography (SEC) suggested that the lytic compounds are large non-proteinaceous molecules (<23.3 kDa, trypsin). Although there is no direct proof that the large molecules found in the lytic HILIC fraction are responsible for the lytic activity of this fraction, the mass range deduced from SEC strongly supports this hypothesis. Total sugar content analysis showed that the lytic HILIC fraction contained two-fold more sugar than the non-lytic one. Nevertheless, the low percentage of saccharide per dry mass equivalent (0.18 ± 0.01 %) indicates that sugar residues are likely not a major component of the lytic compounds. We concluded that at least one group of lytic allelochemicals produced by A. tamarense comprise a suite of large non-proteinaceous and probably non-polysaccharide compounds between 7 kDa and 15 kDa.

Variability in the allelopathic action of the Alexandrium tamarense species complex along the coast of China

“…After standing for 1 h, the precipitate was collected by centrifugation, dissolved in 100 ml of 1 M HCl, and dialyzed in 0.1 mM HCl for 3 days. According to the previous findings (Yamasaki et al, 2008;Ma et al, 2011), some amphipathic extracellular substances with large molecular weights may be responsible for the allelopathic action of A. tamarense. Additionally, Emura et al (2004) isolated a protein-like hemolytic compound with a molecular weight of more than 10 kDa by dialysis from Alexandrium taylori.…”

“…Though many studies have been performed, the nature of allelochemicals involved in the allopathic action of Alexandrium tamarense remains unclear (Emura et al, 2004;Grané li et al, 2008;Yamasaki et al, 2008;Ma et al, 2011). Some large amphipathic extracellular substances, independent of known phycotoxins, PSP toxins and spirolides, have been recommended to be responsible for the action (Tillmann and John, 2002;Emura et al, 2004;Tillmann et al, 2007;Yamasaki et al, 2008;Ma et al, 2011).…”

“…Some large amphipathic extracellular substances, independent of known phycotoxins, PSP toxins and spirolides, have been recommended to be responsible for the action (Tillmann and John, 2002;Emura et al, 2004;Tillmann et al, 2007;Yamasaki et al, 2008;Ma et al, 2011). In our study, to completely remove the algal cells and cell debris that cannot be removed by centrifugation, cell-free supernatants were filtered through 0.22-mm filters.…”

Modelling paralytic shellfish toxins (PST) accumulation in Crassostrea gigas by using Dynamic Energy Budgets (DEB)

“…It has been shown that some Alexandrium spp. strains are able to produce extracellular toxic compounds (Arzul et al, 1999) known to have allelopathic activity on algae (Ma et al, 2011;Lelong et al, 2011) and ichtyotoxic activity on bivalves (Ford et al, 2008;Borcier et al, 2017;Castrec et al, 2018). Recent studies using the AM89BM A. minutum strain (used in the dataset 3) suggested that these compounds might also negatively affect feeding behavior in Pecten maximus (Borcier et al, 2017) and C. gigas (Castrec et al, 2018).…”

Bioactive compounds of marine dinoflagellate isolates from western Greenland and their phylogenetic association within the genus Alexandrium

“…has been shown to affect membrane integrity, mobility via loss of flagella and/or cell survival against a wide variety of other microalgae (Arzul et al, 1999;Tillmann et al, 2008;Tillmann and Hansen, 2009), heterotrophic protists (Hansen et al, 1992;Matsuoka et al, 2000;Tillmann and John, 2002) and diverse members of microbial communities (Weissbach et al, 2011). Although molecular structures and exact mode of action of allelochemicals from Alexandrium still are poorly known (Ma et al, 2009;Ma et al, 2011), all evidence suggest that the lytic activity against target protists is not related to the known toxins, such as saxitoxins and spirolides, produced among members of this genus (Tillmann and John, 2002;Tillmann et al, 2007).…”