Identification and reduction of juvabione, a main constituent of inhibitory effects on Ceriodaphnia dubia reproduction, in a pulp and paper mill effluent using Todomatsu, Abies sachalinesis, mast

We have evaluated and reduced the environmental impact of pulp and paper mill effluents in Japan using a battery of bioassays combined with chemical analyses. Japanese pulp and paper mill effluents examined had consequently no acute toxicity to zebra fish and Daphnia magna whereas some of the effluents exhibited acute toxicity to luminescent bacteria. Dramatic inhibitory effects on Ceriodaphnia dubia reproduction were continuously observed in whole mill effluents (WMEs) of newsprint mills using Todomatsu, Abies sachalinensis Mast. The seven days 25% effective concentrations (7d-EC25s) for C. dubia reproduction were in the range of 0.1 to 0.3% of the WME in one mill(J1), whereas no lethal effect on C. dubia was observed for the WME. As WMEs from Japanese pulp and paper mills are usually integrated a mixture of effluents derived from pulp and paper manufacture processes, it is often difficult to identify the particular constituents responsible for the toxicity of WMEs. The toxicity of the WME was characterized by a simplified toxicity identification evaluation (TIE) methodology using a combination of metal-chelating and activated-charcoal treatment. The effluent exhibited no toxic effects after activated-charcoal treatment, indicating that the toxicity of the effluent was attributed to organic constituents. We then specified the origin of the toxicity by application of the TIE method to various in-mill process effluents (IMPEs). The C. dubia toxicity tests on the IMPEs suggested that noxious substance(s) was mainly derived from the IMPE of mechanical pulping process. Fractionation of this IMPE using HPLC followed by LC-MS analysis revealed that the compound primarily responsible for the WME toxicity to C. dubia was 150 to 500 μ g/l of juvabione, a naturally-occurring sesquieterpenoid ester found in Todomatsu, Abies sachalinensis Mast. The EC25 value of the authentic juvabione was 5 μ g/l, accounting for the strong effect of both the WME and the IMPE. Further investigation revealed that either chemical or biological treatments could hydrolyze juvabione to todomatsuic acid, non-toxic acid. Activated sludge treatment completely removed the toxicities to C. dubia from the effluents of deinking,(DIP) mechanical(TMP) pulping and newsprint machine processes, indicating that juvabione in the effluents was in fact converted to todomatsuic acid. Consequently, the WME discharged by pulp and paper mills of Oji Paper and Nippon Paper, no longer caused any toxicity to C. dubia reproduction.