Phenol, paraalkylation products with C12-rich branched olefins derived from propene oligomerisation, ethane-1,2-diol, carbonate, sulfurized, calcium salts, overbased, reacted with alkylene carbonate, including distillates (petroleum), heavy paraffinic C10-C50

Toxicity to terrestrial plants

Link to relevant study record(s)

Reference

Endpoint:

toxicity to terrestrial plants: short-term (with study design considered suitable for long-term assessment)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2014 January 7 to 2014 November 20

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Remarks:

Adjustments had to be made to the protocol to take equipment availability and substitutions necessary to carry out the work as outlined in the guideline.

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.4200 (Seed Germination/Root Elongation Toxicity Test)

Version / remarks:

1996 / Last know version prior to replacement; Required study for submission to China.

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Identifier: EXP1200078
Batch number: E00275-350
Expiration date: end of 2015
Very dark brown (almost black) viscous liquid.
Purity: 100%

Analytical monitoring:

no

Remarks:

Material is UVCB, and there is no specific/appropriate analytical method.

Vehicle:

no

Remarks:

Test material was dissolved in tetrahydrofuran (THF, CAS 109-99-9) prior to preparing samples for use in the study.

Details on preparation and application of test substrate:

A stock solution was prepared by disolving 5.0000 g of EXP1200078 in 500 mL of THF to provide a nominal concentration of 10 mg/mL. The stock was mixed by inversion, and it appeared translucent and brown in color after mixing, with no particulates observed. Test substrate was prepared by mixing 500 mL of the 10 mg/mL stock, with 5000 g of substrate (80 grit glass beads), stirring to mix, then dried to allow the solvent to evaporate. The solvent control soil was prepared in a similar manner as the treated substrate but without the addition of test substance (THF only). Negative control soil consisted of the 80 grit glass beads without solvent or test substance added. The substrate was mixed with soil after drying, and sufficient reverse osmosis water to hydrate the soil was added, and the substrate was mixed again. Prepared substrate was added to each of three test chambers for each of the treatment and control groups. The test concentrations are reported as milligrams of test substance per kilogram of test substrate on a dry weight basis (mg/kg).

Species:

Allium cepa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

Allium cepa (Onion) / Granix Yellow PRR Hybrid; 96% germination; Liliceae family; supplier - Park Seed Co., Greenwood, SC, USA

Species:

Lolium perenne

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

Lolium perenne (Ryegrass) / Gator 3; 90% germination; Poaceae family; Meyer Seed Co., Baltimore, MD, USA

Species:

Triticum aestivum

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

Triticum aestivum (Wheat) / Glenn Hard Red; 94% germination; Poaceae family; Johnny’s Selected Seeds, Winslow, ME, USA

Species:

Zea mays

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

Zea mays (Corn) / Nothstine Dent OG; 87% germination; Poaceae family; Johnny’s Selected Seeds, Winslow, ME, USA

Species:

Beta vulgaris

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Beta vulgaris (Sugarbeet)/ 38WVR0852; 98% germination; Chenopodiaceae family; Betaseed, Inc., Moorhead, MN, USA

Species:

Brassica napus

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Brassica napus (Oilseed Rape) / Dwarf Essex; 96% germination; Brassicaceae family; Johnny’s Selected Seeds, Winslow, ME, USA

Species:

Brassica oleracea var. capitata

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Brassica oleracea (Cabbage) / Late Flat Dutch; 99% germination; Brassicaceae; Sustainable Seed Co., Covelo, CA, USA

Species:

Glycine max (G. soja)

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Glycine max (Soybean) / Williams 82% viability; 85% viability; Fabaceae; Missouri Foundation Seeds, Columbia, MO, USA

Species:

Lactuca sativa

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Lactuca sativa (Lettuce) / Summertime; 98% germination; Asteraceae; Sustainable Seed Co., Covelo, CA, USA

Species:

Lycopersicon esculentum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

Lycopersicon esculentum (Tomato) / Rutgers; 90% germination; Solanaceae; Meyer Seed Co., Baltimore, MD, USA

Test type:

seed germination/root elongation toxicity test

Study type:

laboratory study

Substrate type:

other: 80 grit glass beads

Limit test:

yes

Total exposure duration:

13 d

Remarks:

Dependent upon species. The range was 3 to 13 days.

Post exposure observation period:

Exposure was for duration of study for each, individual seed type.

Justification for exposure duration:

All exposures were less than two (2) weeks, and it did not make sense to disturb the seeds and/or seedlings during the study period.

Test temperature:

25 +/- 1 deg C

pH:

unspecified

Moisture:

not specified

Details on test conditions:

Test chambers were held in a dark environmental chamber. Temperature in the environmental chamber was monitored continuously with a maximum/minimum thermometer.

Seeds were placed on the surface of the substrate in plastic petri dishes (150 mm in diameter) after the substrate was rehydrated with reverse osmosis water. Petri dishes were labeled with the species name, project number, treatment group designation and replicate. Seeds were placed on the surface of the substrate with seeds spaced such that roots had room to grow, with a total of ten seeds in each petri dish. Petri dishes were covered with lids, and the lids were taped into place. The test chambers were placed in an environmental chamber in a randomized block design on shelves that were placed at a slight angle to facilitate linear growth of the plant roots.

Seed germination was observed daily, and the test was terminated for each species when at least 65% of the control seeds had germinated and produced roots or shoots at least 20 mm in length. Observations for germination consisted of noting the number of seeds that had germinated in each replicate. The growth of germinated seeds was evaluated by measuring the length of the roots or shoots. When insufficient root length was apparent after several days of growth, shoot lengths were measured as an indication of plant growth. Root lengths were measured with a ruler from the point between the hypocotyl and the root to the root tip. Shoot lengths were measured with a ruler from the point between the hypocotyl and the root to the hypocotyl tip. The lengths were rounded to the nearest whole millimeter. Seed condition was described by noting the presence or absence of abnormal development. Mean germination and root or shoot lengths were determined for each treatment group.

Nominal and measured concentrations:

A nominal concentration of 1000 mg EXP1200078/kg of substrate was chosen in concentration with the Sponsor based on the results of a non-GLP rangefinding test. Negative control and solvent control groups were maintained concurrently in substrate prepared without the addition of EXP1200078.

Reference substance (positive control):

no

Remarks:

NOMINAL TEST LEVELS: Negative Control, Solvent Control, 1000 mg EXP1200078/kg dry substrate

Key result

Species:

Triticum aestivum

Duration:

5 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 g/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat. (total fraction)

Basis for effect:

germination

Remarks on result:

other: An important monocotolydon was tested and no adverse effects were noted.

Key result

Species:

Brassica napus

Duration:

8 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/kg soil ww

Nominal / measured:

nominal

Conc. based on:

test mat. (total fraction)

Basis for effect:

germination

Remarks on result:

other: An important dicotocotolydon was tested and no adverse effects were noted.

Details on results:

Germination
For all ten species, sufficient germination (at least 65% germinated) was observed in the negative control group, and there were no significant differences between the two control groups for number of seeds germinated. Additionally, when compared to the pooled control, the treatment group germination was not significantly different in any species tested (Dunnett’s test, p < 0.05).

Root/Shoot Elongation
For Z. mays, L. sativa and L. esculentum (Tables 4, 9 and 10, respectively), sufficient root growth and germination (at least 65% seeds germinated with roots of at least 20 mm) were observed in the negative control, control groups were not significantly different, and the treatment group was not significantly different from the pooled control (Dunnett’s test, p < 0.05).
For A. cepa and T. aestivum (Tables 1 and 3, respectively), sufficient root growth and germination (at least 65% seeds germinated with roots of at least 20 mm) were not observed in the negative control, but sufficient growth and germination were observed in the treatment group, indicating that the seeds had sufficient time to germinate and develop roots. Control groups were not significantly different, and the treatment group was not significantly different from the pooled control (Dunnett’s test, p < 0.05).
For L. perenne and B. oleracea (Tables 2 and 7, respectively), minimal root development was observed after 13 days of exposure to substrate, but strong shoot growth was observed. Because it was apparent that the 20 mm root length requirement was not going to be obtained, shoot lengths were instead measured, as an alternate indication of seedling growth. Shoot lengths obtained sufficient growth and germination (at least 65% seeds germinated with shoots of at least 20 mm). There were no significant differences between the control groups, and the treatment group was not significantly different from the pooled control (Dunnett’s test, p < 0.05).
For G. max (Table 8), sufficient root growth and germination (at least 65% seeds germinated with roots of at least 20 mm) were not observed in the negative control or the treatment group, but the test was terminated because it appeared that the root length requirement had been met. Due to the roots being below the substrate, the determination of root length was hindered prior to removing the seeds from the substrate. However, the growth of the roots appeared sufficient to assess whether differeces occurred between the control and treatment group. Control groups were not significantly different, and the treatment group was not significantly different from the pooled control (Dunnett’s test, p < 0.05).
For B. napus (Table 6), sufficient root growth and germination (at least 65% seeds germinated with roots of at least 20 mm) were not observed in the negative control, but sufficient growth and germination were observed in the solvent control and treatment group, indicating that the seeds had sufficient time to germinate and develop roots. The solvent control was significantly stimulated as compared to the negative control. Because there was no negative effect due to the solvent, the control groups were pooled, and there was no significant difference in the treatment group when compared to the pooled control (Dunnett’s test, p < 0.05).
For B. vulgaris (Table 5), sufficient root growth and germination (at least 65% seeds germinated with roots of at least 20 mm) were observed in the negative control. The solvent control was significantly stimulated as compared to the negative control. Because there was no negative effect due to the solvent, the control groups were pooled, and there was no significant difference in the treatment group when compared to the pooled control.

Results with reference substance (positive control):

Not available; not conducted.

Reported statistics and error estimates:

Statistical analyses were used to assess the effects of test substance application on seedling germination and root/shoot length. The negative control was compared to the solvent control using a Student’s t-test. A probability of less than 0.05 was considered to be statistically significant. For each endpoint, replicate means were averaged in determination of test group means. The inhibition of treatment group means relative to the control mean was determined with the following equation:

Percent inhibition = [(pooled control mean – treatment mean)/control mean] x 100%

A negative percent inhibition indicated that the treatment mean was greater than the pooled control mean. Because inhibition of at least 25% relative to control means was not observed at the rate tested, the 25% and 50% effect rates (ER25 and ER50, respectively) were not calculated.

Treatment group means were compared to the pooled control mean with a Dunnett’s test. A probability of less than 0.05 was considered to be statistically significant. All computations were made using SAS statistical software

	Content of Tables in Attached PDF
Table	Content
Table 1	Effects of EXP1200078 on Germination and Root Length of Allium cepa (Onion) in a 10-Day Seed Germination Test
Table 2	Effects of EXP1200078 on Germination and Root Length of Lolium perenne (Ryegrass) in a 13-Day Seed Germination Test
Table 3	Effects of EXP1200078 on Germination and Root Length of Triticum aestivum (Wheat) in a 5-Day Seed Germination Test
Table 4	Effects of EXP1200078 on Germination and Root Length of Zea mays (Corn) in a 3-Day Seed Germination Test
Table 5	Effects of EXP1200078 on Germination and Root Length of Beta vulgaris (Sugarbeet) in a 6-Day Seed Germination Test
Table 6	Effects of EXP1200078 on Germination and Root Length of Brassica napus (Oilseed Rape) in a 8-Day Seed Germination Test
Table 7	Effects of EXP1200078 on Germination and Root Length of Brassica oleracea (Cabbage) in a 13-Day Seed Germination Test
Table 8	Effects of EXP1200078 on Germination and Root Length of Glycine max (Soybean) in a 9-Day Seed Germination Test
Table 9	Effects of EXP1200078 on Germination and Root Length of Lactuca sativa (Lettuce) in a 6-Day Seed Germination Test
Table 10	Effects of EXP1200078 on Germination and Root Length of Lycopersicon esculentum (Tomato) in a 9-Day Seed Germination Test

Validity criteria fulfilled:

yes

Conclusions:

An OPPTS Guideline study (USEPA - 850.4200 Seed germination,root elongation toxicity test) was conducted on the test material at the limit concentration. No adverse effects were observed in any of the ten (10) species tested as specified by the Guideline.

Executive summary:

An application of EXP1200078 at rates of 1000 mg EXP1200078/kg dry substrate resulted in no adverse, treatment-related effects on all ten species tested.  The ER25 and ER50 values for all endpoints for the ten species were determined to be greater than 1000 mg EXP1200078/kg, the highest application rate tested.

Description of key information

No effects on any germination or growth parameters per study protocol showed any effects at the highest concentration of 1000 mg/kg soil on a dry weight basis.

No long-term toxicity testing to plants is proposed, as in accordance to the Annex X, Paragraph 9.4, Column 2, exposure to soil is negligible and the CSA does not indicate the need to further investigate, with terrestrial RCRs of <0.02 for all exposure scenarios.

Key value for chemical safety assessment

Short-term EC50 or LC50 for terrestrial plants:

1 000 mg/kg soil dw

Long-term EC10, LC10 or NOEC for terrestrial plants:

1 000 mg/kg soil dw

Additional information

An application of EXP1200078 at rates of 1000 mg EXP1200078/kg dry substrate resulted in no adverse, treatment-related effects on all ten species tested.  The ER25 and ER50 values for all endpoints for the ten species were determined to be greater than 1000 mg EXP1200078/kg, the highest application rate tested.