Toxicon, Vd . 22, No. S, pp . B21-822, 1984 .

0041-O101/&1 53 .00+ .00Printed in ~reat Brltsio.

p 1984 PerBamon Presa Ltd .

LACK OF TOXINS IN ALCOHOLIC EXTRACTS OF THE SPONGETEDANIA IGNIS

D. MEaSZentrum der Rechtsmedizin, University of Frankfurt, Frankfurt/Main, Federal Republic of Germany

(.9crepted Jor publication 22 February 1984)

D. MBBS. Lack of toxins in alcoholic extracts of the sponge Tedania ignis. Toxicon 22, 821- 822,1984 . - Alcoholic extracts of the sponge Tedania ignis collected onthe coast ofthe Florida Key's(LT.S.A.) were essentially free of toxins which had been found in specimens from the Venezuelancoast, supporting the assumption of planlttonic origin of these toxins .

SEVCIK and BARiiOZA (1983) described the isolation of two fractions from the alcoholicextract of the fire sponge Tedania ignis which exhibited reversible presynaptic effects onthe frog neuromuscular junction blocking the release of acetylcholine. A toxin withsimilar chemical and pharmacological properties (i .e. a polar substance with a mol. wt ofabout 950) was extracted from fish and plankton samples collected in areas (Venezuelancoast) where ichthyotoxism occurred (MIJARES et al ., 1983). The authors concluded thatthe fractions isolated from fish and sponge may have a common planktonic origin .Experiments were, therefore, performed to purify these or similar toxins from sponges ofa distant area .Samples of the sponge Tedania ignis were collected in April 1983 from the shallow

waters around the east (Atlantic) coast of Islamorada (Florida Key's, U.S.A.) . Thesponges were kept frozen at -20°C. Several batches of 50-80 g (fresh weight) fromdifferent locations of the collection area were extracted according to the proceduredescribed by $13VCIK and BARBOZA (1983) . Fifty g sponge tissue was homogenized in500 ml distilled water in a blender. To the homogenate (about 600 ml) 1400 ml ethanolwere added, it was kept at 4°C overnight and centrifuged at 6000 rev/min for 20 min. Thesupernatant was reduced to 200 ml in a rotary evaporator (29°C, 30 kHg), 800 ml of coldacetone (4°C) wasaddedand the small precipitate formed was collected by centrifugation .The supernatant was reduced to 300 ml (rotary evaporator, 30°C), mixed with an equalvolume of acetic acid, the precipitate collected by centrifugation and the supernatantlyophilized . The resulting yellow-brown preparation was soluble in aqueous solutions . Allfractions obtained were bioassayed by i.p . injection into mice using four dilutions (1/1,1/2, 1/4, 1/8) and groups of five animals each .The aqueous extracts of the various batches of sponge produced toxic symptoms in

mice within 6-12 hr when injected i.p . (0 .5 ml of S00 ml extract) . The animals exhibitedsedation, abdominal cramps, slow movement and died within 24 -48 hr, but no signs of

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flaccid paralysis indicative of neurotoxic activity occurred . However, these relativelyunspecific symptoms which are often observed when extracts of marine sponges areinjected into mice (blebs, unpublished results) maysuggest the presence of a toxic agent inlow concentration or of low activity. On the other hand, neither the alcoholic extracts(after evaporation of the alcohol, lyophilization and redissolving in 50 ml physiologicalsaline ; 0.5 ml was injected into mice) where the presynaptic toxins should be concentratednor samples of the fractionation steps had any significant influence on the behaviour ofthe test animals .From these results it is reasonable to conclude that the specimens of Tedania ignis

collected in that specific area (Florida Key's) are essentially free of those small molecularweight, polar substances which may possess neurotoxic (i .e . presynaptic) effects asdescribed by SEVCIK and BARBOZA (1983) . This supports the assumption of MIJARES etal . (1983) that the toxic fractions isolated from specimens from the Venezuelan coast maybe of planktonic origin . It should be interesting to see if other sponge species are also ableto accumulate those toxins .

REFERENCESMunxFS, A. J., $EVCIK, C., BNeHOZn, C. A. and SnnvEnxn, J. A. (1983) Ichthyotoxism by a paralytic

neurotoxin probably produced by the dinoflagellate Ceratium furry (Abstract) . In : Proceedings of the SthEuropean Symparium on Anima/, PlantandMicrobial Toxins, p . 84 (MEHS, D. and HwestuteEn., G. G. Eds) .Hannover: European Section of the International Society on Toxinology .

S$vccx, C. and BARHOZA, C. A. (1983) The presynaptic effect of fractions isolated from the sponge Tedaniaignis. Toxicon 21, 191 .