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Environmental fate & pathways

Biodegradation in water: screening tests

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Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-07-12 to 2012-10-12
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
This study has been carried out according to appropriate OECD guidelines and GLP standards.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
yes
Remarks:
Tested concentrations are lower than recommended because biphenyl-4,4'-diol was suspected to be toxic to the microbial inoculum, and because of its low water solubility. This modification did not compromise the scientific validity of the study.
GLP compliance:
yes
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
The activated sludge used in this study was obtained from a municipal sewage treatment plant (Hildesheim, DE). Prior to use in the definitive CO2 Evolution Test, the sludge solids were washed twice with autoclaved tap water, resuspended in mineral salts medium and maintained under aerobic conditions for 4 hours. The sludge was then homogenised and the supernatant decanted and aerated with CO2 free air. After 4 days the inoculum was centrifuged and the solids resuspended in 1 L tap water. The bacterial suspension was maintained in an aerobic condition by aeration with CO2 free air for two further days. This mixture, added to the vessels at a rate of 10 mL/L medium, was used as the test inoculum. The microbial density of the inoculated test medium in the test vessels at the start of the test was 6.7E007 colony forming units (CFU)/L, determined by standard plate count.
Duration of test (contact time):
60 d
Initial conc.:
7 mg/L
Based on:
test mat.
Initial conc.:
5.4 mg/L
Based on:
TOC
Remarks:
organic carbon content
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
The ready biodegradability of biphenyl-4,4-diol was assessed in an extended (60 day) OECD 301B CO2 evolution test . The test item concentration was deliberately lowered below the recommended range as biphenyl-4,4-diol was considered capable of inhibiting the activity of the microbial inoculum and hence limiting the biodegradation observed in the test. An initial non-GLP preliminary test was first used to establish the viability of the test conditions, followed by the fully GLP-compliant definitive test. To facilitate good discrimination between the CO2 generated by the low concentration of test substance and that formed by background respiration, the test inoculum was rigorously washed by re-suspension and pre-conditioned by prolonged aeration under test conditions to minimise CO2 production in the blank controls during the test.

The CO2 evolution test design comprised a pair of blank control vessels, a reference vessel (definitive test only) with sodium benzoate dosed at 20 mg/L and replicate vessels (2 in the prelim, 4 in the definitive test) dosed with biphenyl-4,4-diol (at 10 mg/L in the prelim, 7 mg/L in the definitive test). A further vessel containing a mixture of the reference and test substance (each component at the same concentration as that at which they were tested individually) served as a toxicity control. The vessels were continually purged with CO2-free air and the exhaust air passed through a train of 3 gas-scrubbers containing Ba(OH)2 solution to trap CO2 formed as a respiration/degradation product. CO2 yields were determined at intervals by detaching the first-in-line scrubbers and titrating their contents with acid/phenolphthalein indicator. Degradation was calculated by expressing measured, blank-corrected CO2 yield as a % of theoretical. With the exception of replicate(s) that served as the source of adapted inocula for the next phase of testing, incubations were terminated by adding HCl (34 days in the prelim, 60 days in the definitive test). Residual CO2 in the test media (formed before acidification, released after lowering pH) continued to be purged and trapped for a further day.

In both the preliminary and definitive stages, the initial test based on measurement of CO2 production was followed by a further, confirmatory test that employed different methodology, based on the measurement of cumulative oxygen uptake (see description in the following data "Part 2_modified OECD301F").
Reference substance:
benzoic acid, sodium salt
Preliminary study:
Two replicate vessels containing biphenyl-4,4-diol (10 mg/L or 7.7 mg organic C-equivalent/L) were used in the preliminary CO2 evolution test. In one replicate, 10% degradation was reached after an adaption phase of 15 days. Rapid degradation followed and the 60% pass level was reached after 25 days - within 28 days and within the 10-day window, i.e. fulfilling the criteria for classification as readily biodegradable. The second replicate reached the 10% level after an adaption phase of 27 days. Thereafter the course of biodegradation was similar to the first replicate and 35% degradation was reached (CO2 production still in progress) when the test was terminated on day 34.

In the toxicity control containing both test item and sodium benzoate (reference item), CO2 production reached 42% of the combined theoretical yield after 14 days and 60% after 28 days. The course of the biodegradation curve indicates two biodegradation phases, which represent the biodegradation of the reference item, followed by mineralisation of the test item. Biodegradation of the reference item was not inhibited by the presence of 10 mg biphenyl-4,4-diol in the toxicity control.
Test performance:
The definitive test included four vessels (5 L, brown glass) containing biphenyl-4,4-diol at 7 mg/L (5.4 mg organic C/L), a functional control with the reference substance sodium benzoate at 20 mg/L, two inoculum controls (blanks) and a toxicity control containing the test and reference substances combined.
Appropriate amounts of distilled water, mineral salts solutions and inoculum (total 3 L) were placed in each vessel, then aerated for 24 hours with CO2-free air. Biphenyl-4,4-diol and Na benzoate were weighed out for each of the appropriate vessels, the test substance was dispersed in 2x distilled water using ultrasound prior to dosing. Aeration with CO2 -free air (30 to 100 mL/min) then resumed and the outlets of the test vessels were connected to a dedicated series of 3 gas wash bottles, each containing 100 mL of 0.0125 mol/L Ba(OH)2 solution to trap CO2. Vessel contents were continuously stirred and temperatures during the test ranged from 21.0 to 23.0 degrees C. On Day 60, 1 mL conc. HCl was added to each of the vessels, except for test substance replicate (#2) where the most extensive biodegradation had occurred, and aeration continued for 24h to purge remaining CO2 from the test media. Determination of CO2 was by titration of residual alkalinity in the Ba(OH)2 traps. Adapted microorganisms were recovered from test repilicate #2, to inoculate the subsequent respirometry test (see following summary).

Key result
Parameter:
% degradation (CO2 evolution)
Remarks:
Replicate 1 (definitive study)
Value:
12
Sampling time:
28 d
Remarks on result:
other: Biodegradation: 10% reached on day 20 / 18% reached on day 61
Key result
Parameter:
% degradation (CO2 evolution)
Remarks:
Replicate 2 (definitive study)
Value:
61
Sampling time:
28 d
Remarks on result:
other: Biodegradation: 10% reached on day 15 and 60% reached on day 27 / 67% reached on day 61 / 10-day window achieved in this replicate
Key result
Parameter:
% degradation (CO2 evolution)
Remarks:
Replicate 3 (definitive study)
Value:
5
Sampling time:
28 d
Remarks on result:
other: Biodegradation: 10% reached on day 42 / 14% reached on day 61
Key result
Parameter:
% degradation (CO2 evolution)
Remarks:
Replicate 4 (definitive study)
Value:
7
Sampling time:
28 d
Remarks on result:
other: Biodegradation: 10% not reached until day 60 (only 8% reached on day 61)
Details on results:
In the definitive CO2 evolution test the 10% level was reached in one of four replicates after an adaptation phase of 15 days. Biodegradation of 59% was reached within 6 days. The biodegradation slowed down and the 60% pass level was reached on day 27. After 33 days a maximum biodegradation of 67% was reached. Two of the remaining replicates reached the 10% level between day 20 and day 42, but the biodegradation continued only slowly and after 60 days the mean biodegradation reached was 16%. The course of biodegradation in the fourth replicate was qualitatively similar, but only reached 8% by day 60.

In the toxicity control vessel containing both the test and reference substances, biodegradation based on the combined theoretical CO2 yield of the mixture reached 70% within 28 days and 96% at termination, implying complete degradation of both the sodium benzoate and the biphenyl-4,4-diol components. Biodegradation of the reference item was not inhibited by the presence of 7 mg biphenol-4,4-diol in the toxicity control.

Results with reference substance:
In the definitive CO2 evolution test the sodium benzoate reference substance was mineralised by 60% in 7 days and by 96% after 28 days.

Table 1: Biodegradation of biphenyl-4,4'-diol in comparison to the reference item (sodium benzoate) and toxicity control (CO2 Evolution Test) - Definitive ready biodegradability test, unadapted inoculum.

                           Biodegradation (%)
   Day 6 Day 14  Day 22   Day 28 Day 35  Day 42   Day 49 Day 56   Day 61
Test item, Rep. 1  11  12  14   14 16  18  18 
Test item, Rep. 2  3 59   61 67  67   67  67  67
Test item, Rep. 3  0  5  10  11  13  14
 Test item, Rep. 4  4  8  8  8  8
 Reference item (Na benzoate) 59 86   91  96  n.a.   n.a. n.a.   n.a. n.a. 
 Toxicity Control: Test + reference item  40  54  61  70  75 81  88  92   96

 

Validity criteria fulfilled:
yes
Interpretation of results:
inherently biodegradable, not fulfilling specific criteria
Conclusions:
The ready biodegradability of biphenyl-4,4'-diol was determined with a non-adapted inoculum according to the CO2 Evolution Test (OECD 301B). The test item concentration was deliberately lowered below the range indicated in the guideline because the test item was considered capable of inhibiting the activity of the microbial inoculum and hence limiting the outcome of the test. Extensive biodegradation of biphenyl-4,4'-diol (with CO2 production >60% of the theoretical yield - the threshold that conventionally represents complete mineralisation) was observed in one replicate in both the preliminary and definitive stages. This result met the 'pass' level for classification as readily biodegradable. Nevertheless, the length of the lag phase between test initiation and the onset of significant degradation was highly variable and in some replicates of the definitive test the rapid degradation phase was not observed at all despite extending the incubation to 60 days. In view of its erratic performance in relation to the pass criteria and the divergence permitted between replicates at test-end or plateau, biphenyl-4,4'-diol does not qualify for classification as readily biodegradable.

In both the preliminary and definitive stages, the initial test based on measurement of CO2 production was followed by a further, confirmatory test that employed different methodology based on the measurement of cumulative oxygen uptake. This additional test was conducted according to the manometric Respirometry Test (OECD 301F) and with a higher concentration of the test item (see data description in the other study presented in this dossier "Part 2_modified OECD301F").
For this purpose, adapted sludge was harvested at the end of the incubation from one or more of the replicate test vessels of the CO2 Evolution Test incubation where >60% theoretical CO2 production had occurred. In each case, complete biodegradation occurred in all replicates. In this additional assay, biphenyl-4,4'-diol has exhibited a potential to undergo rapid and ultimate biodegradation and confirms the finding in the CO2 Evolution test.

In conclusion, Biphenyl-4,4'-diol is non-persistent (not P).

As biphenyl-4,4'-diol does not qualify for classification as readily biodegradable, it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.

The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.
Executive summary:

The ready biodegradability of biphenyl-4,4'-diol was determined with a non-adapted inoculum according to the CO2 Evolution Test (OECD 301B). The test item concentration was deliberately lowered below the range indicated in the guideline because the test item was considered capable of inhibiting the activity of the microbial inoculum and hence limiting the outcome of the test. The study was performed in two stages: an initial, non-GLP preliminary test was first used to establish the viability of the test conditions before conducting the main, fully GLP-compliant definitive test.

Extensive biodegradation of biphenyl-4,4'-diol (with CO2 production >60% of the theoretical yield - the threshold that conventionally represents complete mineralisation) was observed in one replicate in both the preliminary and definitive stages. The degradation observed in this replicate met the 'pass' level for classification as readily biodegradable. Nevertheless, the length of the lag phase between test initiation and the onset of significant degradation was highly variable and in some replicates of the definitive test the rapid degradation phase was not observed at all despite extending the incubation to 60 days. In view of its erratic performance in relation to the pass criteria and the divergence permitted between replicates at test-end or plateau, biphenyl-4,4'-diol does not qualify for classification as readily biodegradable.

In both the preliminary and definitive stages, the initial test based on measurement of CO2 production was followed by a further, confirmatory test that employed different methodology based on the measurement of cumulative oxygen uptake. This additional test was conducted according to the manometric Respirometry Test (OECD 301F) and with a higher concentration of the test itema (see data description in the other study presented in this dossier "Part 2_modified OECD301F").

For this purpose, adapted sludge was harvested at the end of the incubation from one or more of the replicate test vessels of the CO2 Evolution Test incubation where >60% theoretical CO2 production had occurred. In each case, complete biodegradation occurred in all replicates of the respirometric test based on oxygen uptake. In this additional assay, the test substance biphenyl-4,4'-diol has exhibited a potential to undergo rapid and ultimate biodegradation (i.e. complete mineralisation) and confirms the finding in the CO2 Evolution test.

In conclusion, Biphenyl-4,4'-diol is non-persistent (not P).

As biphenyl-4,4'-diol does not qualify for classification as readily biodegradable, it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.

The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.

Biphenyl-4,4'-diol is classified as "rapidly biodegradable" for CLP purposes.

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-07-12 to 2012-10-12
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
This study has been carried out according to appropriate OECD guidelines and GLP standards.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
yes
Remarks:
The inoculum was pre-adapted to biphenyl-4,4'-diol. This invalidates the test as an investigation of "ready" biodegradability, but does not compromise its validity as a confirmation of ultimate biodegradability.
GLP compliance:
yes
Specific details on test material used for the study:
Biphenyl-4,4'-diol / CAS 92-88-6
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, adapted
Details on inoculum:
In the definitive test of the OECD 301B (see description in the data of the study presented in this dossier, "Part 1_OECD301B"), adapted sludge was harvested at the end of the incubation from the vessel where >60% theoretical CO2 production had occured. It was used to inoculate a test based on the manometric respirometry test (OECD 301 F), with a higher concentration (20 mg/L) of the test item. The inoculum was recovered from the relevant vessel of the prior test by filtration (142 mm diameter membrane filter, 45 micron pore size), re-suspended by shaking the filter end-over-end in 100 mL tapwater for 60 minutes and finally transferred to 2 L test medium. The final inoculum density was not reported, however it would have been lower than that present in the test flask of the CO2 evolution test and uis not likely to have exceeded 30 mg dry weight suspended solids/L.
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
test mat.
Initial conc.:
46.4 mg/L
Based on:
ThOD
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
In both the preliminary and definitive stages of the study, the initial test was based on measurement of CO2 production (see description in the study presented in this dossier "Part 1_OECD301B). Then it was followed by a further, confirmatory test that employed different methodology, based on the measurement of cumulative oxygen uptake according to the OECD 301F Manometric Respirometry test. For these tests, adapted sludge was harvested at the end of the incubation from the replicate vessel(s) of the CO2 evolution test where >60% theoretical CO2 production had occured, and was used to inoculate a test based on the manometric respirometry test (OECD 301 F), with a higher concentration (20 mg/L) of the test item.

The test item concentration was raised to 20 mg/L, equivalent to a theoretical oxygen demand (ThOD) of 46.4 mg O2/L, compared to the concentration of 7 mg biphenyl-4,4'-diol applied in the earlier phase based on measurement of CO2 production. Inoculated test medium (250 mL) was dispensed to brown glass incubation bottles and direct additions of the test or reference substance were made as appropriate to the relevant vessels. A CO2 absorber containing soda lime was suspended in the headspace of each vessel before sealing the bottles with WTW OxiTop caps, each fitted with a pressure-sensitive data-logging device. The vessel contents were continuously stirred and cumulative measurements of headspace pressure reduction made throughout the incubation were automatically converted to O2 consumption readings.

The definitive manometric respirometry test design comprised a pair of blank control vessels, a reference vessel with sodium benzoate dosed at 20 mg/L and triplicate vessels dosed with biphenyl-4,4-diol, also at 20 mg/L. Degradation was calculated by expressing measured, blank-corrected O2 uptake as a % of the theoretical maximum.
Reference substance:
benzoic acid, sodium salt
Preliminary study:
Four replicate vessels containing biphenyl-4,4'-diol (20 mg/L, 46.4 mg ThOD-equivalent/L) were used in the preliminary manometric respirometry test. Rapid and consistent degradation occurred in all four replicates: 12% degradation occurred in all vessles within 2 days and degradation had reached 67, 67, 67 and 64% at termination on Day 28.
Test performance:
The definitive test included three vessels containing biphenyl-4,4-diol at 20 mg/L (46.4 mg ThOD/L), a functional control with the reference substance sodium benzoate at 20 mg/L and two inoculum controls (blanks).
Key result
Parameter:
% degradation (O2 consumption)
Value:
65
Sampling time:
7 d
Remarks on result:
other: Average value from the yhree samples (65%, 62%, 68%) / The maximum mean biodegradation was reached by day 7
Key result
Parameter:
% degradation (O2 consumption)
Value:
63
Sampling time:
28 d
Remarks on result:
other: Average value from the three samples (71%, 56% and 62%)
Details on results:
Biphenyl-4,4-diol was rapidly mineralised in the manometric respirometry test using adapted sludge from the prior definitive CO2 evolution test (see previous summary for details): 10% degradation was reached in all three biphenyl-4,4-diol replicates by day 4. In spite of using a test concentration of 20 mg/L (more than 2x the concentration applied in the preceding definitive CO2 evolution test), the 60% pass level was reached after 6 days. A mean maximum biodegradation of 65% and the degradation plateau were both reached on day 7.
Results with reference substance:
In the definitive manometric respirometry test the biodegradation of sodium benzoate reached >60% on day 3 and a maximum of 76% by day 20.

Table 1: Biodegradation of Biphenyl-4,4'-diol in comparison to the reference item (Manometric Respirometry Test) - Definitive confirmation test, adapted inoculum.

          Biodegradation (%)  
   Day 7 Day 14  Day 21  Day 28 
Test item, Rep. 1  65  65  70 71 
Test item, Rep. 2  62   59  58 56
Test item, Rep. 3 68  62  64 62 
Reference item (Na benzoate)  74  72 75  72 
Validity criteria fulfilled:
not applicable
Interpretation of results:
inherently biodegradable, not fulfilling specific criteria
Conclusions:
In a previous test, Biphenyl-4,4'-diol exhibited a potential to undergo rapid and ultimate biodegradation (i.e. complete mineralisation) under the conditions of the CO2 Evolution Test (OECD 301B). This finding was confirmed by further tests performed according to the Manometric Respirometry Test (OECD 301F), using an inoculum pre-adapted from in the earlier testing phase (from the replicate where > 60% theoritical CO2 production accoured).
In the Manometric Respirometry Test with a pre-adapted inoculum, 10% degradation was reached in all three biphenyl-4,4-diol replicates by day 4. In spite of using a test concentration of 20 mg/L (more than 2x the concentration applied in the preceding definitive CO2 evolution test), the 60% pass level was reached after 6 days. A mean maximum biodegradation of 65% and the degradation plateau were both reached on day 7.

Biphenyl-4,4'-diol is unambiguously non-persistent (not P).

Biphenyl-4,4'-diol did not qualify for classification as readily biodegradable, however it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.

The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.

Biphenyl-4,4'-diol is classified as "rapidly biodegradable" for CLP purposes.
Executive summary:

In a previous test, Biphenyl-4,4'-diol exhibited a potential to undergo rapid and ultimate biodegradation (i.e. complete mineralisation) under the conditions of the CO2 Evolution Test (OECD 301B). This finding was confirmed by a further test performed according to the Manometric Respirometry Test (OECD 301F), using an inoculum pre-adapted from in the earlier testing phase (from the replicate where >60% theoretical CO2 production had occurred.

In this Manometric Respirometry Test, 10% degradation was reached in all three biphenyl-4,4-diol replicates by day 4. In spite of using a test concentration of 20 mg/L (more than 2x the concentration applied in the preceding definitive CO2 evolution test), the 60% pass level was reached after 6 days. A mean maximum biodegradation of 65% and the degradation plateau were both reached on day 7. This complete biodegradation observed in the respirometric test based on oxygen uptake, confirmed the outcome of the earlier phase that relied on CO2 yield measurements. Since this confirmatory test used adapted inocula, it cannot be used to demonstrate "ready" biodegradability, nevertheless they provide unambiguous supporting evidence of the non-persistence of biphenyl-4,4'-diol and its potential to udergo rapid biodegradation.

In conclusion, Biphenyl-4,4'-diol is unambiguously non-persistent (not P).  

Biphenyl-4,4'-diol did not qualify for classification as readily biodegradable, however it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.  

The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.  

Biphenyl-4,4'-diol is classified as "rapidly biodegradable" for CLP purposes.  

Description of key information

Biphenyl-4,4'-diol is non-persistent (not P).
As biphenyl-4,4'-diol does not qualify for classification as readily biodegradable because of divergent performance of replicates under ready biodegradability test conditions, it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.
The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.
Biphenyl-4,4'-diol is classified as "rapidly biodegradable" for CLP purposes.

Key value for chemical safety assessment

Biodegradation in water:
inherently biodegradable, not fulfilling specific criteria

Additional information

The ready biodegradability of biphenyl-4,4'-diol was determined with a non-adapted inoculum according to the CO2 Evolution Test (OECD 301B). The test item concentration was deliberately lowered below the range indicated in the guideline because the test item was considered capable of inhibiting the activity of the microbial inoculum and hence limiting the outcome of the test. The study was performed in two stages: an initial, non-GLP preliminary test was first used to establish the viability of the test conditions before conducting the main, fully GLP-compliant definitive test. Extensive biodegradation of biphenyl-4,4'-diol (with CO2 production >60% of the theoretical yield - the threshold that conventionally represents complete mineralisation) was observed in at least one replicate test vessel of the CO2 Evolution Test in both the preliminary and definitive stages.

In both the preliminary and definitive stages, the initial test based on measurement of CO2 production was followed by a further, confirmatory test that employed different methodology based on the measurement of cumulative oxygen uptake. For this purpose, adapted sludge was harvested at the end of the incubation from one or more of the replicate test vessels of the CO2 Evolution Test incubation where >60% theoretical CO2 production had occurred and was used to inoculate a confirmatory assessment performed according to the Manometric Respirometry Test (OECD 301F), with a higher concentration of the test item. In each case, complete biodegradation occurred in all replicates of the respirometric test based on oxygen uptake, confirming the outcome of the earlier phase that relied on CO2 yield measurements.

Biphenyl-4,4'-diol degraded erratically in the presence of unadapted microorganisms in the CO2 Evolution Tests of both the preliminary and definitive test stages. Degradation was sufficient in some replicates to indicate complete mineralisation and meet the 'pass' level for classification as readily biodegradable, but not in others. The length of the lag phase between test initiation and the onset of significant degradation was highly variable and in some replicates of the definitive test the rapid degradation phase was not observed at all despite extending the incubation to 60 days. In view of its erratic performance in relation to the pass criteria and the divergence permitted between replicates at test-end or plateau, biphenyl-4,4'-diol does not qualify for classification as readily biodegradable. Nevertheless, biphenyl-4,4'-diol has exhibited a potential to undergo rapid and ultimate biodegradation (i.e. complete mineralisation) and this finding in the CO2 Evolution Test was confirmed by further tests performed according to the Manometric Respirometry Test, using an inoculum pre-adapted in the earlier testing phase.

Biphenyl-4,4'-diol is non-persistent (not P).

As biphenyl-4,4'-diol does not qualify for classification as readily biodegradable, it is classified as "inherently biodegradable, not fulfilling specific criteria" (no specific inherent biodegradability tests have been performed) for the purposes of predicting concentrations of biphenyl-4,4'-diol that may enter the aquatic compartment via municipal STP.

The consistently high level of degradation achieved with an adapted inoculum is relevant to situations where waste-waters and process effluents containing biphenyl-4,4'-diol are likely to pass through dedicated industrial STPs before reaching the aquatic environment.

Biphenyl-4,4'-diol is classified as "rapidly biodegradable" for CLP purposes.