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Short-term toxicity of NExBTL (WAF) toward juvenile rainbow trout (Oncorhynchus mykiss) was determined in GLP-compliant study according to OECD Guidelines for Testing Chemicals (1992) no 203 "Fish, acute toxicity tests". No mortality was observed during the test. Sub lethal effects were not detected either. LL50 > 1000 mg/l.

No long-term aquatic toxicity data for fish are available. The modelled NOELR value (long-term), using PetroTox is > 1000 mg/L

Short-term toxicity of the test material (WAF) toward a water flea Daphnia magna was determined in GLP compliant laboratory according to OECD Guidelines for Testing Chemicals (2002) no 202 "Daphnia sp, Acute immobilisation test". No adverse effects were detected in the test. EL50 > 100 mg/l.

Long-term toxicity of the test material toward an aquatic invertebrate was determined in GLP compliant laboratory according to OECD Guidelines for Testing Chemicals (1998) no 211 "Daphnia magna, Reproduction test". The 14 and 21 Day EL50* (immobilisation) values for parental generation were estimated to be greater than 100 mg/l WAF. The 21 Day EL50* (reproduction) value was estimated to be greater than 100 m/l WAF. The LOEC was considered to be 3.2 mg/l WAF on the basis that at this loading rate significantly (p < 0.05) fewer youngs per adult were produced when compared to the control. The NOEC was considered to be 1 mg/l WAF on the basis that at this loading rate there were no significant mortalities (immobilisation) observed in the parental generation and that there were no significant (p > 0.05) differences between the control and the 1.0 mg/l loading rate WAF test group in terms of numbers of live young produced per adult by Day 21.

The aquatic toxicity of NExBTL renewable diesel was tested using three aquatic organisms from different trophic levels (fish, crustancea and algae). Due to the poor water solubility of the substance, the organisms were exposed to a Water Accommodated Fraction (WAF) of the test substance which is a commonly accepted procedure for regulatory studies on poorly soluble substances. The loading rates used in these studies were 1000 mg/l for fish and 100 mg/l for crustancea and algae. In the experimental toxicity tests, no adverse effects were observed. In an acute test with sediment-dwelling amphipod, the LC50 value was 1200 mg/kg dry weight (d. w.), NOEC was 373 mg/kg d. w. and LOEC was 1165 mg/kg d. w.

Short-term toxicity of NExBTL renewable diesel (WAF) toward an aquatic algae Scenedesmus subspicatus was determined in a GLP compliant laboratory according to OECD Guideline 201 (Alga, Growth Inhibition Test)". There was no statistical difference (Student's t-test, p < 0.05) in the growth between the control and the WAF test group. E*L50 > 100 mg/l.

Toxicity of the test material toward micro-organisms (activated sewage sludge) was determined in a GLP compliant laboratory study according to OECD Guideline 209 (Activated sludge, respiration inhibition test). No effects on respiration were observed at the test concentration of 1000 mg/l.

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