2,6-diisopropylnaphthalene

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 Oct. - 19.11.2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 310 (Ready Biodegradability - CO2 in Sealed Vessels (Headspace Test)

Deviations:

yes

Remarks:

low initial concentration (range of water solubility); reduction of the ratio headspace to water volume (50 : 200 mL = 1 : 4); GC analysis

Principles of method if other than guideline:

Primary biodegradation/elimination of single components of diisopropylnaphthalene by analysing the GC profile over time.

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): activated sludge from the activation bassin of a predominantly domestic biological sewage treatment plant (community STP, Stadtentsorgung Neustadt, Im Altenschemel, Lachen-Speyerdorf)
- Method of cultivation: freshly prepared
- Storage conditions: no storage
- Preparation of inoculum for exposure: The sludge was washed with tap water twice, then filtrated through a cloth, resuspended in test medium and aerated. An aliquot was added to the test medium
- Pretreatment: no pretreatment
- Concentration of sludge: 3.36 g dw suspended solid/L

Duration of test (contact time):

56 d

Initial conc.:

ca. 0.2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

test mat. analysis

Remarks:

; GC-analysis of diisopropylnaphthalene isomers

Details on study design:

TEST CONDITIONS
- Composition of medium: standard medium according to test guideline
- Additional substrate: no
- Solubilising agent (type and concentration if used): no
- Test temperature: 22 ± 2 °C
- pH: no data
- pH adjusted: no
- Aeration of dilution water: yes
- Suspended solids concentration: ca. 10 ± 1 mg/L
- Continuous darkness: no, no direct light

TEST SYSTEM
- Culturing apparatus: size of test flasks was 250 mL (fill volume 200 mL). The flasks were made of DURAN 3.3 and closed with red lids and silicon septa. Flasks were set up in a rotation shaker in an air-conditioned room.
- Number of culture flasks/concentration: test vessels, negative and positive controls: 2; medium blank and abiotic control: 1
- Method used to create aerobic conditions: prior to test aeration of dilution water, during test air supply of the head space
- Measuring equipment: carbon analyzer TOC multi N/C 2100S; HP Model 6890N GC
- Test performed in closed vessels due to significant volatility of test substance: yes
- Details of trap for CO2 and volatile organics if used: positive controls: CO2 was measured as carbonate after adjustment of test vessels pH with 1 mL 7M NaOH in closed vessel. Test vessels: no carbon dioxide measurementsTest substance was extracted in 10 mL iso-hexane in closed vessel.

SAMPLING
- Sampling frequency: day 0, 4, 8, 14, 21, 28, 40, and 56
- Sampling method: for CO2 measurement, 1 mL sample was taken directly from test medium 1 hour after addition of NaOH solution.
for analysis of test substance,
for test substance measurements, iso-hexane extracts of test medium (see above "Details on analytical method") were concentrated and aliqots were injected into GC.
- Sample storage before analysis: no

CONTROL AND BLANK SYSTEM (to be checked)
- Inoculum blank: 2
- Abiotic sterile control: 1, poisoned with 50 mg HgCl2/L
- Toxicity control: no (not relevant due to low test concentration)
- Other: 1 medium blank without test substance and inoculum

STATISTICAL METHODS: no statistics

Reference substance:

aniline

Key result

Parameter:

% degradation (test mat. analysis)

Value:

21 - 30

Sampling time:

56 d

Remarks on result:

other: primary biodegradation; range of results from day 21 to 56

Results with reference substance:

Reference substance (aniline, 20 mg C/mL) performed as expected. Biodegradation was ca. 99% after 14 days.
day 4 8 14 21 28 40 56
% degradation 76.2 73.1 98.5 100.5 95.4 101.7 98.0
(mean of 2 replicates)

Within the test period of 56 days, primary degradation of ca. 50% was observed. Abiotic degradation contributed ca. 20 - 30%.

Average biodegradation was approx. 25%. The time course of primary degradation is presented in the following table.

After a test period of ca. 21 days, there was no more substantial increase in total biodegradation.

 

Mean percentage of degradation as sum of all peaks (total of diisopropylnaphthalene) [Report, Table 9.2 -d]

Day	Primary degradation total	Primary degradation abiotic	Primary degradation biodegradation
0	0	0	0
4	5	6	-1
8	19	7	12
14	24	16	8
21	32	11	21
28	43	20	23
40	47	17	30
56	53	32	21

 

 

Individual isomers of the diisopropylnaphthalene technical product mixture showed different susceptibility to degradation as is illustrated in the following table.

 

 

Percentage of degradation for individual diisopropylnaphthalene (DIPN) isomers as mean of replicate determinations calculated from peak areas (isomers are characterised by substituent position numbers) [from Report, Table 9.2 -b and 9.2 -c]

Day	Peak 1 (1,3-DIPN)		Peak 2 (1,7- DIPN)		Peak 3 (2,6- DIPN)		Peak 4 (2,7- DIPN)		Peak 5 (1,6- DIPN)		Peak 6 (1,4- DIPN)		Peak 7 (1,5- DIPN)
	total	biotic	total	biotic	total	biotic	total	biotic	total	biotic	total	biotic	total	biotic
0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
4	-3	-10	0	-8	18	12	11	5	7	2	-3	-7	7	0
8	7	2	11	3	40	33	28	20	24	17	7	2	15	7
14	3	-10	9	-8	59	40	37	18	41	24	5	-9	15	1
21	8	-1	13	3	82	70	45	33	38	27	7	-1	30	9
28	8	-10	14	-5	100	76	59	35	78	59	7	-6	33	14
40	12	-3	19	3	100	81	68	49	87	69	13	-1	34	17
56	17	-12	28	-6	100	>81	78	36	92	56	21	-9	52	15

  Negative percentages indicate that the abiotic value was determined to be higher than the biotic portion.

 

Total primary degradation was found for all isomers (between 17 and 100%). But biodegradation was only observed for 4 of 7 isomers (between ca. 15% - 1,5 -DIPN to >81% - 2,6 -DIPN). For 1,3 -DIPN, 1,4-DIPN, and 1,7 -DIPN, no primary biodegradation could be demonstrated in the present experiment.

 

Capability of biodegradation appears to be dependent on isomeric structure. In general, biodegradation appeares to proceed the better if the two isopropyl-substituents are attached to different rings of naphthalene and the further apart the two substituents are from each other (2,6- > 1,6- > 2,7- > 1,5-).

 

Obviously there was no relevant increase in primary biodegradation after day 28 of the experiment. Variable results do not show a clear trend and are considered to be caused by variances of individual test vessels and by measurement inaccuracy.

 

 

Averaged peak areas from single runs (corrected by ISTD) (DIPN isomers are characterised by substituent position numbers)

 

Day	Sample	Area Peak 1 (1,3-)	Area Peak 2 (1,7-)	Area Peak 3 (2,6-)	Area Peak 4 (2,7-)	Area Peak 5 (1,6-)	Area Peak 6 (1,4-)	Area Peak 7 (1,5-)
0	Replic. 1	44.42	54.15	48.65	40.45	43.28	47.97	27.44
	Replic. 2	47.62	57.97	52.73	43.74	46.36	50.93	27.28
	Abiotic	56.27	69.27	64.04	52.82	55.03	60.07	29.42
4	Replic. 1	45.88	54.76	40.14	36.11	39.97	48.98	20.77
	Replic. 2	48.56	57.53	42.54	38.57	43.59	53.15	29.87
	Abiotic	52.40	64.14	59.67	49.60	52.18	58.05	27.15
8	Replic. 1	43.29	50.12	29.94	30.15	33.76	46.16	20.34
	Replic. 2	42.22	49.45	30.62	30.71	34.05	46.19	26.06
	Abiotic	53.48	63.84	59.16	48.93	51.23	57.36	27.02
14	Replic. 1	44.70	50.82	20.17	26.60	25.50	46.81	21.10
	Replic. 2	44.84	51.47	21.34	26.41	27.37	47.25	25.31
	Abiotic	49.07	57.84	51.63	42.63	45.47	51.80	25.31
21	Replic. 1	42.34	48.22	6.86	22.15	26.00	45.39	18.01
	Replic. 2	42.70	48.84	11.12	24.28	29.69	46.93	20.40
	Abiotic	51.27	61.91	56.37	46.35	49.16	55.66	23.39
28	Replic. 1	41.66	47.07	< LOD	17.27	8.82	44.29	17.09
	Replic. 2	42.80	48.86	< LOD	17.57	10.63	47.41	19.34
	Abiotic	46.30	55.78	48.83	40.57	44.23	51.92	23.70
40	Replic. 1	36.23	39.75	< LOD	7.48	4.82	38.54	14.78
	Replic. 2	45.35	51.53	< LOD	19.53	7.28	48.17	21.55
	Abiotic	48.27	58.15	51.77	42.62	45.46	51.84	24.56
56	Replic. 1	37.08	41.15	< LOD	12.75	4.75	40.09	15.55
	Replic. 2	37.31	42.49	< LOD	12.92	4.65	41.34	16.81
	Replic. 3	38.67	35.91	< LOD	8.04	3.58	34.65	14.25
	Replic. 4	36.34	45.57	< LOD	-	-	44.32	2.75
	Replic. 5	38.71	41.23	< LOD	10.80	4.14	39.11	15.09
	Replic. 6	39.95	38.02	< LOD	14.11	4.92	37.20	15.71
	Abiotic 1	43.76	51.16	< LOD	35.87	39.18	46.26	19.66
	Abiotic 2	40.10	42.56	< LOD	29.46	32.81	39.42	18.54
	Abiotic 3	38.48	47.00	< LOD	30.99	35.72	44.37	17.93

 

 

In diisopropylnaphthalene test vessels, CO2 development (ultimate biodegradation) was not measured. Ultimate biodegradation was tested in a separate test using 14C radiolabelled test substance (LAUS GmbH 2011a).

 

Procedure controls performed as expected.

Validity criteria fulfilled:

yes

Interpretation of results:

other: not readily biodegradable; only partial primary biodegradation

Conclusions:

In a sealed vessels ready biodegradability test according to OECD test guideline 310, diisopropylnaphthalene (200 µg/L) was incubated for 56 days with activated sludge (10 mg/L SS) from a predominantly domestic sewage treatment plant. Primary degradation was followed by analysing the decrease of diisopropylnaphthalene levels in test vessels. Diisopropylnaphthalene was analysed by a GC-method measuring seven individual diisopropylnaphthalene isomers (individual GC peaks). Degradation was measured on day 4, 8, 14, 21, 28, 40, and 56.
Total primary degradation of diisopropylnaphthalene was ca. 50% within the test period. 20 to 30% was caused by abiotic degradation. Primary biodegradation was found to be about 20 to 30% for all isomers. There appeared no relevant increase in overall biodegradation from day 21 to day 56 (varying levels of biodegradation between 20 to 30%)
Distinct diisopropylnaphthalene isomers showed variable biodegradation. For three isomers, no primary biodegradation could be demonstrated. For four isomers, 15 to 100% primary biodegradation was noted at the end of the test period. Abiotic degradation was similar for all isomers within certain variations. Between day 14 and 40, abiotic biodegradation seemed to level off at around 15 to 20%. On day 56, rates of around 30 to 35% abiotic primary degradation was observed.

Executive summary:

In a sealed vessels ready biodegradability test according to OECD test guideline 310, diisopropylnaphthalene (200 µg/L) was incubated for 56 days with activated sludge (10 mg/L SS) from a predominantly domestic sewage treatment plant. Primary degradation was followed by analysing the decrease of diisopropylnaphthalene levels in test vessels.

Diisopropylnaphthalene was analysed by a GC-method measuring seven individual diisopropylnaphthalene isomers (individual GC peaks). Degradation was measured on day 4, 8, 14, 21, 28, 40, and 56.

Total primary degradation of diisopropylnaphthalene was ca. 50% within the test period. 20 to 30% was caused by abiotic degradation. Primary biodegradation was found to be about 20 to 30% for all isomers. There appeared no relevant increase in overall biodegradation from day 21 to day 56 (varying levels of biodegradation between 20 to 30%).

The diisopropylnaphthalene isomers showed variable biodegradation. For three isomers, no primary biodegradation could be demonstrated. For four isomers, 15 to > 81% primary biodegradation was noted at the end of the test period. Abiotic degradation was similar for all isomers within certain variations. Between day 14 and 40, abiotic biodegradation seemed to level off at around 15 to 20%. On day 56, rates of around 30 to 35% abiotic primary degradation were observed.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 Oct. - 01 Dec. 2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 310 (Ready Biodegradability - CO2 in Sealed Vessels (Headspace Test)

Deviations:

yes

Remarks:

use of radiolabelled test substance, low initial concentration (range of water solubility); reduction of the ratio headspace to water volume (50 : 200 mL = 1 : 4)

Qualifier:

according to guideline

Guideline:

other: ISO 14593:2005 "Water quality - Evaluation of ultimate aerobic biodegradability of organic compounds in aqueous medium - Method by analysis of inorganic carbon in sealed vessels (CO2 headspace test)"

Principles of method if other than guideline:

Ultimate biodegradation/mineralisation by measuring the formation of 14CO2 from radiolabelled precursor

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure):
activated sludge from the activation basin of a predominantly domestic biological sewage treatment plant
(community STP, Stadtentsorgung Neustadt, Im Altenschemel, Lachen-Speyerdorf)
- Method of cultivation: freshly prepared
- Storage conditions: no storage
- Preparation of inoculum for exposure: The sludge was washed with tap water twice, then filtrated through a cloth,
resuspended in test medium and aerated. An aliquot was added to the test medium.
- Pretreatment: no pretreatment
- Concentration of sludge: 3.36 g dw suspended solid/L

Duration of test (contact time):

56 d

Initial conc.:

ca. 0.2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: standard medium according to test guideline
- Additional substrate: no
- Solubilising agent (type and concentration if used): no
- Test temperature: 22 ± 2 °C
- pH: no data
- pH adjusted: no
- Aeration of dilution water: yes
- Suspended solids concentration: ca. 10 ± 1 mg/L
- Continuous darkness: no, no direct light, bottles used were made of brown glass

TEST SYSTEM
- Culturing apparatus: size of test flasks was 250 mL (fill volume 200 mL). The flasks were closed with a silicon septum and a lid. Flasks were set up in a rotation shaker in an air-conditioned room.
- Number of culture flasks/concentration: test vessels, negative and positive controls: 2; medium blank and abiotic control: 1
- Method used to create aerobic conditions: prior to test aeration of dilution water, during test air supply of the head space
- Measuring equipment: carbon analyzer TOC multi N/C 2100S; liquid scintillation counter Thriathler, Hidex Oy, Turku, Finland
- Test performed in closed vessels due to significant volatility of test substance: yes
- Details of trap for CO2 and volatile organics if used: 1 M NaOH (2 mL) as absorbing agent for CO2 in head space located in an external micro-scrubber attached to test vessels via a needle passing the cover septum of the flask; activated charcoal in small absorber tubes to collect volatile organics in head space

SAMPLING
- Sampling frequency: day 0, 4, 8, 14, 21, 28, 40, and 56
- Sampling method: see above "Details on analytical methods"
- Sample storage before analysis: only charcoal absorber tubes were stored.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 2
- Abiotic sterile control: 1, poisoned with 5 mg HgCl2/L
- Toxicity control: no (not relevant due to low test concentration)
- Other: 1 medium blank without test substance and inoculum

STATISTICAL METHODS: no statistics

Reference substance:

aniline

Remarks:

initial concentration: 20 mg organic carbon/L

Key result

Parameter:

% degradation (CO2 evolution)

Value:

<= 0.1

Sampling time:

56 d

Remarks on result:

other: based on CO2 formation, no biodegradation was observed

Key result

Parameter:

% degradation (radiochem. meas.)

Remarks:

polar 14C marker in aqueous phase

Value:

ca. 15

Sampling time:

56 d

Key result

Parameter:

% degradation (radiochem. meas.)

Remarks:

non-polar 14C-marker in aqueous phase

Value:

>= 65 - <= 85

Sampling time:

56 d

Results with reference substance:

Reference substance (aniline, 20 mg C/mL) performed as expected. Biodegradation was ca. 99% after 14 days.
day 4 8 14 21 28 40 56
% degradation 76.2 73.1 98.5 100.5 95.4 101.7 98.0
(mean of 2 replicates)

Reference radioactivity for initial test substance concentration (40 µg test substance)

Determination of total radioactivity for initial test substance concentration as described under "Any other information on materials and methods incl. tables" resulted in an activity of 330770 cpm.

 

CO2 formation

In CO2 headspace samples, virtually no radioactivity was detected (< 0.05%) during the total 56 day test period (see table below). There was no ultimate biodegradation of diisopropylnaphthalene.

 

Activated charcoal from absorber

The amount of radioactivity in the charcoal of the absorber from sampling of headspace was ≤ 0.02%. No organic radioactive substances were found in the headspace withdrawn from the test vessels.

 

Residual radioactivity in test vessels

Non polar substances (xylene extractable fraction under acidic condition)

In xylene extracts of the test medium from test vessels, ca. 65 to 85% of radioactivity was detected. Information about the nature of chemical substances present is not available.

Polar substances (fraction remaining in aqueous phase)

Radioactivity in the residual aqueous phase accounted for 10 to 21% of total radioactivity (single values, mean 15 %), indicating to the formation of polar substances from diisopropylnaphthalene, probably by biotic and abiotic transformation of the side chain.

 

Abiotic control

Distribution of radioactivity in abiotic controls was similar to that in test vessels. Depending on the incubation period, xylene extracts of test medium accounted for approx.. 70 to 85% of radioactivity. In the residual aqueous phase ca. 10 to 20% of total radioactivity (single values, not shown) was found, on the average apoprox. 15% from day 28 through 56.

 

 

Distribution of radioactivity (in %) (from Report Table 8.3 -i and 8.2 -j)

 

Day	Headspace			Test medium
	CO2			Xylene extract			Aqueous phase (after extraction)
	Test 1	Test 2	Abiotic control	Test 1	Test 2	Abiotic control	Test 1	Test 2	Abiotic control
4	0.005	0.00	0.002	85.4	84.7	106.3	no data	no data	no data
8	0.005	0.003	0.002	89.7	78.3	86.0	no data	no data	no data
14	0.009	0.006	0.005	73.1	60.8	85.0	no data	no data	no data
21	0.009	0.011	0.008	65.8	80.3	83.8	no data	no data	no data
28	0.004	0.016	-0.003	71.7	67.8	73.7	15.5	15.7	16.8
40	0.014	0.102	0.006	72.4	70.8	76.1	12.8	17.0	19.7
56#	0.032	0.019	0.007	66.6	70.4	72.4	15.6	12.2	11.7

  # Day 56: Average value of three replicates.

 

Ultimate degradation (in %) of test substance as determined by CO2 measurements (radioactive CO2)

 

Day	Test 1	Test 2	Abiotic control
4	0.01	0.00	0.00
8	0.00	0.00	0.01
14	0.01	0.01	0.01
21	0.01	0.01	0.01
28	0.00	0.02	0.00
40	0.01	0.10	0.01
56	0.03	0.02	0.01

 

 

Information on primary biodegradation was investigated in a separate experiment and is reported in LAUS GmbH 2011b.

Validity criteria fulfilled:

yes

Interpretation of results:

other: under test conditions no ultimate biodegradation observed

Executive summary:

In a sealed vessels ready biodegradability test according to OECD test guideline 310, 14C-labelled diisopropylnaphthalene (200 µg/L) was incubated for 56 days with activated sludge (10 mg/L SS) from a predominantly domestic sewage treatment plant. Formation of CO2 was determined on day 4, 8, 14, 21, 28, 40, and 56 by counting of CO2 radioactivity isolated from test vessels.

No radioactive CO2 (ultimate biodegradation) could be detected during the test period. Radioactivity was found prevalently in the organic extract of the test medium (extraction with xylene under acidic condition). Radioactivity in the organic extract decreased over time from ca. 85% at day 4 to ca. 60 to 75% at the end of test period. On the average, the residual aqueous phase accounted for approx. 15% (only examined from day 28).

Approx. 15% radioactivity was not detected anymore in the fractions examined. There may be some loss of test material due to adsorption to test vessel walls. An abiotic control showed the same pattern. Radioactivity was found in the organic extract and the residual aqueous phase of the test medium. Distribution of radioactivity was similar to experiments with inoculum.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Refer to analogue justification provided in IUCLID section 13

Reason / purpose for cross-reference:

read-across source

Principles of method if other than guideline:

Read-across to preceding entry:
Source substance: Bis(isopropyl)naphthalene [DIPN, mixture of isomers], radiolabeled
Reference: Laus GmbH 2011a

Key result

Parameter:

% degradation (CO2 evolution)

Value:

<= 0.01

Sampling time:

56 d

Remarks on result:

other: based on CO2 formation, no biodegradation was observed, but no definite conclusion can be drawn for 2,6-DIN

Key result

Parameter:

% degradation (radiochem. meas.)

Remarks:

14C-marker in aqueous phase

Value:

ca. 15

Sampling time:

56 d

Key result

Parameter:

% degradation (radiochem. meas.)

Remarks:

non-polar 14C-marker in aqueous phase

Value:

>= 65 - <= 85

Sampling time:

56 d

Details on results:

The 14C-label for 2,6-DIPN was relatively low, only 2.9 % of the total radioactivity, due to disadvantageous chemical-synthetic conditions. 14CO2 formed from 2,6-DIPN, and thus ultimate biodegradability may not have been discriminated from the background variation.

Reference 4

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Refer to analogue justification provided in IUCLID section 13

Reason / purpose for cross-reference:

read-across source

Principles of method if other than guideline:

Read-across to preceding entry:
Source substance: Bis(isopropyl)naphthalene [DIPN, mixture of isomers]
Reference: Laus GmbH 2011b

Parameter:

% degradation (test mat. analysis)

Value:

ca. 40

Sampling time:

8 d

Key result

Parameter:

% degradation (test mat. analysis)

Value:

ca. 59

Sampling time:

14 d

Parameter:

% degradation (test mat. analysis)

Value:

ca. 82

Sampling time:

21 d

Key result

Parameter:

% degradation (test mat. analysis)

Value:

ca. 100

Sampling time:

28 d

Details on results:

Among the consitutents of the source substance, DIPN isomer mixture, the isomer 2,6-DIPN is found to be one of the best biodegradable constituents. But the test conditions do not all to conclude ready biodegradability.

Description of key information

No biodegradation study concerning only 2,6-diisopropylnaphthalene (CAS 24157-81-1; 2,6-DIPN) was available. But two GLP studies according to OECD 210 examining the primary and ultimatie biodegradation of Bis(isopropyl)naphthalene (DIPN), an isomeric mixture which consist of seven isomers (1,3-, 1,4-, 1,5-, 1,6-, 2,6- and 2,7-DIPN) is available and used in a read-across approach.

In a sealed vessel CO2 headspace test using 14C ring labelled diisopropylnaphthalene isomer mixture, no ultimate biodegradation could be demonstrated. Analysing DIPN content in test medium, mean primary biodegradation (ca. 20 - 25%) was observed, while the degree of primary degradation was ~100% after 28 days for 2,6-DIPN. On the other hand, ultimate degradation could not be concluded for the target substance, 2,6-DIPN. This may also be due to the low fraction of 2,6-specific radiolabel, which accounted for not more than 3% in the radioactive test mixture as compared to >13% in the technical mixture.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

No biodegradation study concerning only 2,6-diisopropylnaphthalene (CAS 24157-81-1; 2,6-DIPN) was available. But two GLP studies according to OECD 210 examining the primary and ultimative biodegradation of Bis(isopropyl)naphthalene (DIPN), an isomeric mixture which consist of seven isomers (1,3-, 1,4-, 1,5-, 1,6-, 2,6- and 2,7-DIPN) is available and used in a read-across approach.

Ultimate biodegradation

In the study by LAUS 2011, no ultimate biodegradation of DIPN was observed. Radioactivity of CO2 samples was of the order of background samples. Biodegradation was determined to be < 0.05%. In addition, no radioactivity was observed in samples of absorption material from traps for organic volatiles in the air stream. In acidic organic extracts (xylene) of the test medium, about 85% of total radioactivity was determined at day 4 decreasing to ca. 70% at day 56 (abiotic control 100 % at day 4, 73% at day 56). Following day 28, there was no substantial change/increase in the content of radioactivity in the xylene extracts. In the remaining aqueous phase, the percentage of total radioactivity from day 28 to 56 was between 10 and 21 % for single samples, the mean about 15 % (abiotic control 11 to 20%). There was no substantial change over time in the fraction of radioactivity in the aqueous phase.

No conclusion on ultimate degradation of the target substance 2,6 -DIPN could be drawn. This can also be due to the extremely low fraction of radiolabel of 2,6 -DIPN in the radioactive test mixture: this was surprisingly only approx. 3%, which may have been less than the background variation of the analytical method.

Mean percentage of radioactivity in distinct test fractions:

Day	^14CO2 (%)	Absorbant for volatile organics (%)	Xylene extract (%)	Residual aqueous phase (%)	Balance
4	0.03	0.002	85.1	-	-
21	0.01	0.008	73.1	-	-
40	0.058	0.006	71.5	14.9	86.5
56	0.026	0.007	68.5	13.9	82.4

Overall conclusion:

1. Deficit in the radioactive mass balance: approx. 15% (not found initially)

2. Volatiles (CO2 or organic compounds): approx. 0%.

3. Xylene extractable material (day 28, 40, 56): approx. 70% (apolar under acidic condition)

4. Xylene non-extractable material (day 28, 40, 56): approx. 15% (polar under acidic condition)

It is concluded that of the 100 % of initial radioactivity approx. 15 % were transformed to polar substances.

This appeared to occur in the abiotic control to similar extent.

Primary biodegradation

Primary degradation (total and abiotic) was determined by GC-analysis of DIPN content in test vessels and abiotic controls. Biodegradation was the difference between total and abiotic degradation. Degradation of total DIPN was calculated by summation of the seven DIPN isomers analysed. The seven positional isomers measured in GC-analysis were (in order following peak elution): 1,3-DIPN, 1,7-DIPN, 2,6-DIPN, 2,7-DIPN, 1,6-DIPN 1,4-DIPN, and 1,5-DIPN (for quantitative distribution of isomers see study record).

Total primary degradation was ca. 50% with ca. 20 to 30% biodegradation. Biodegradation of total DIPN seemed to level off after 21 days, while abiotic degradation appeared to increase from day 40 to day 56 after a plateau between day 14 and day 40.

Abiotic primary degradation was similar for all DIPN isomers. Final abiotic degradation was between 28 and 39%, average 32%.

Mean primary degradation of total DIPN

Day	Primary degradation total (%)	Primary degradation abiotic (%)	Primary degradation biodegradation (%)
4	5	6	-1
21	32	11	21
40	47	17	30
56	53	32	21

Regarding primary biodegradation, there were substantial differences between isomers. 1,3- DIPN, 1,4-DIPN and 1,5-DIPN did not show any primary biodegradation. 2,6- DIPN was biodegraded by > 81%, 1,6-DIPN by 56%, 2,7-DIPN by 36% and 1,5-DIPN by 15%. Except for 2,6-DIPN there was no substantial increase in primary biodegradation following day 28 (see table in study record).

The ability of biodegradation appears to be dependent on isomeric structure. In general, biodegradation appears to proceed the better if the two isopropyl-substituents are attached to different rings of naphthalene and the further apart the two substituents are from each other (2,6- > 1,6- > 2,7- > 1,5-).

Mean percentage of primary degradation for DIPN isomers at test end (56 days) (%)

1,3-DIPN

1,4-DIPN

1,5-DIPN

1,6 DIPN

1,7 DIPN

2,6 DIPN

2,7-DIPN

total

biotic

total

biotic

total

biotic

total

biotic

total

biotic

total

biotic

total

biotic

17

-12

21

-9

52

15

92

56

28

-6

100

>81

78

36

Negative percentages indicate that the abiotic degradation was faster than the biotic portion.