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Many investigations are available concerning the toxicity of phenol to aquatic organisms from different systematic classes including also several non-standard tests: Short-term and/or long-term effects of phenol were investigated in fish, aquatic invertebrates, aquatic algae and cyanobacteria, aquatic macrophytes, amphibians and aquatic microorganisms.

 

Though the test concentrations were not analytically verified in all static or semi-static tests, the phenol concentration is considered to be stable due to the findings in the Daphnia studies from Colgan et al. (1982) and Lewis et al. (1983), where after a 2 day static exposure period the measured phenol concentrations were above 80 % of the nominal concentrations. These findings were confirmed in a test on the stability of phenol over a period of 5 d via TOC analysis (after 5 d 92.1% of the initial test substance concentration was found; SLOVNAFT 2009).

For assessing short-term effects in fish, 24 reliable studies are available with several fish species and different exposure times. The LC50 values of the key and supporting studies were in the range 8.9 to 67.5 mg/L. The tests were conducted under flow-through conditions or semi-static conditions and the exposure concentrations were analytically verified in most of the studies. The most sensitive endpoint is the LC50 of 8.9 mg/L based on measured concentrations obtained under flow-through conditions (DeGraeve et al., 1980) with the cold water fish rainbow trout (Oncorhynchus mykiss).

For long-term effects in fish reliable study results are available from 8 studies with six fish species. The tests were conducted under flow-through or semi-static conditions and the exposure concentrations were analytically verified in the six flow-through studies. In the semi-static tests (Verma et al. 1981, 1984) the interval for test water renewal was 24 h and the phenol concentrations are regarded as stable, since Colgan et al. (1982) demonstrated the stability of phenol under semi-static conditions (48 h interval).

The most sensitive endpoint regarded as reliable and taken as key value for aquatic toxcitity was the NOEC of 77 µg/L, based on nominal concentrations, obtained in a 60 day study with Cirrhina mrigala under semi-static conditions with a 24 hour renewal period (Verma et al., 1984). At test end survival and wet weight of the surviving larvae was recorded. Based on these results a MATC of 77 - 94 µg/L was found and a NOEC of 77 µg/L was derived. This NOEC obtained with larvae of Cirrhina mrigala is confirmed by the 60 d NOEC of 110 µg/L obtained with Cyprinus carpio from the same authors (Verma et al., 1981).

Embryo-larval tests conducted by Birge et al. (1979) and Black et al. (1982; 1983) showing very low NOEC values were not found to be suited for hazard characterisation, in agreement with the decisions of the EU-member states within the framework of preparing the EU RAR (2006).

Data on short-term effects in invertebrates are available from 20 reliable studies with different aquatic invertebrate species. Nine tests conducted with Daphnia magna or Ceriodaphnia followed the standard test approach of international guidelines with 48 hour exposure. The EC50 values are in the range of 3.1 and 20 mg/L. The most sensitive endpoint is the EC50 of 3.1 mg/L, based on measured initial concentrations, obtained in a static 48-h acute toxicity test with Ceriodaphnia dubia (Oris et al., 1991).

Long-term effects of phenol in aquatic invertebrates are assessed using the results from seven reliable studies with D. magna or C. dubia, conducted under flow-through or semi-static conditions. The NOEC or EC10 values of the seven studies are in the range between 0.05 and 2.38 mg phenol/L.

A EC10 of 0.46 mg phenol/L, based on nominal concentrations, was obtained in a 16 day study with D. magna under semi-static conditions (Deneer et al., 1988). Test parameters were inhibition of length increase and reproduction of the daphnids. The most sensitive parameter is growth. As growth reduction will generally result in a lowered reproductive output, this endpoint is of high relevance. The measured EC10 of 2.38 mg/L for survival of adults and reproduction obtained in a 21 day flow-through study fulfilling the validity criteria of the OECD guideline (1982) performed by Tilser and Zagorc-Koncan (1999) are in the same order of magnitude like the EC10 of Deneer et al. (1988) and confirms their findings.

The lowest EC10 value (0.05 mg/L) was reported in the study of SLOVNAFT (2009). In this GLP-study conducted according to OECD Guideline 209 under semi-static conditions and using Daphnia magna as test organism, a 21 d EC10=0.05 mg/L nominal based on reproduction was determined. The stability of the test solutions over a period of 5 days was proven via TOC analysis. This study is regarded as valid with some restrictions: the extrapolated EC10 is below the lowest applied test substance concentration; toxicity results determined for the reference substance are reported to be considerably below values in comperable tests, which may point to an extraordinary sensitivity of the used daphnia strain. Therefore, this EC10 is not regarded as key result for long-term toxicity to aquatic invertebrates.

There are two reports on effects of phenol on amphibians available (Birge et al. 1980, Black et al. 1982). The data was not considered useful for the hazard and risk assessment by the EU member states, due to the reasons already laid down for the long-term fish toxicity results reported by Birge et al. and Black et al. for several substances (see above). However, as there are no other tests with amphibians from other authors available, it cannot be excluded that amphibian species may be more sensitive to phenol than other aquatic species.

 

For the freshwater green algae Pseudokirchneriella subcapitata and Chlorella vulgaris three growth inhibition tests with a standard exposure time of 96 h are available. The EC50 values are 61.1 (cell number), 150 (growth rate), and 370 (growth rate) mg/L. For the blue-green algae Microcystis aeruginosa a TTC (toxic threshold concentration) of 4.6 mg/L was found by Bringmann & Kühn (1978). In this test after 8 days exposure the blue-green algae may not longer be in the exponential growth phase and this may have a negative influence on the test result. However, the 8-day Microcystis test has been accepted within the OECD HPV program and therefore considered reliable with restrictions. The low effect value should be used with care.

For the saltwater benthic diatom Entomoneis cf punctulata a 3-day EC50 of 76 mg/L was found (Adams & Stauber, 2004), based on measured concentrations. The EC50 is in the same order of magnitude as the growth rate EC50 values of the freshwater algae, indicating a possibly comparable sensitivity of the marine and the freshwater algal species.

For assessing the aquatic risk for aquatic algae the lowest EC50 of 61.1 mg/L, based on cell number increase (yield) in green algae, out of the tests with standardised exposure time of 96 hours is used (St. Laurent et al. 1992).

 

The effects of phenol on aquatic plants were investigated with Lemna gibba and four strains of Lemna minor in 7 day growth inhibition tests under static conditions (Cowgill et al., 1991). Measured parameters were number of plants, frond number and dry weight. The EC50 values ranged between 157 and 312 mg/L and the NOEC values between 5 and 108 mg/L. Dry weight was the most sensitive parameter. The EC50 of the most sensitive parameter, dry weight of strain 7136, was 157 mg/L and the NOEC was 5 mg/L for all parameters in Lemna minor strain 7136.

In a report finished recently (SLOVNAFT 2009), the toxicity of phenol towards the duckweed Lemna minor was investigated in a GLP-study conducted according to OECD Guideline 221 (Lemna sp. Growth Inhibition test) for seven days under static conditions. The stability of the test solutions over a period of 5 days was proven via TOC analysis (after 5 d 92.1% of the initial test substance concentration was found). In this study the lowest EC10 value (5.92 mg/L) was determined for the endpoint yield based on number of fronds and the lowest EC50 value (61.82 mg/L) was determined for the endpoint yield based on dry weight. As this EC50 value is the lowest available, this value will be used for assessment.

Manifold test with microorganisms (e.g. waste water sludge, methanogens, activated sludge, several single species) are available. The test methods also vary from short-term tests over 5 min to cell multiplication inhibition test over 48 h. The lowest EC50-value of 21 mg/L for microorganisms relevant for waste water treatment plants was acquired in a 24 h nitrification test with a nitrifying enrichment culture obtained from the mixed liquor of an activated sludge plant treating meat-packing, rendering, and hide-curing waste-water.