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This report shall not be reproduced except in full, without the approval of BNM Environmental. All queries concerning the report or its contents
should be forwarded to the Monitoring Team Leader. ______________________________________________________________________________________________________
Bord na Móna Environmental Ltd
Registered Office: Main Street, Newbridge, Co Kildare, Ireland. Registered No: 303312, VAT No: IE6323313B
For the Attention of: Mark Corrigan
Henkel (Ireland) Ltd.
Tallaght Business Park
Whitestown, Tallaght
Dublin 24
Field work completed by: Mr. Stephen Stapleton
Prepared by: Mr. Stephen Stapleton
Environmental Scientist
Reviewed by: Mr. Peter Coogan
Environmental Team Leader
BNM File Ref: ECS5071-BF-Tox
Monitoring Date: 17th December- Sixth Bi-Monthly Event 2015
Monitoring Period: Toxicity 6 of 6 2015
Reporting Date: 19th January 2016
BI-MONTHLY TOXICITY ANALYSIS OF SEWER
EMISSIONS AT THE HENKEL (IRELAND) LTD.
FACILITY IN KYLEMORE PARK NORTH,
BALLYFERMOT, DUBLIN 10 IN COMPLIANCE
WITH THE REQUIREMENTS OF INTEGRATED
POLLUTION PREVENTION AND CONTROL
LICENCE REGISTER: NO. P0078-01.
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Report No. ECS5071-BF-Tox
Henkel (Ireland) Ltd., Ballyfermot Page 2 P0078-01
Executive Summary
Henkel (Ireland) Ltd. is required to assess the toxicity of emissions to water (wastewater discharges)
from the Ballyfermot facility on a bi-monthly basis, to comply with the requirements of the Integrated
Pollution Prevention and Control Licence, Register No. P0078-01. BNM Environmental was contracted
to perform the sampling. A BNM Environmental Scientist visited the Ballyfermot site on the 17th of
December 2015 to carry out the sixth bi-monthly monitoring event of 2015.
A sample was obtained from emission reference point BFW-1A for monitoring of the bi-monthly
parameters. The wastewater sample was analysed by Aquatic Services Unit for 48 hour LC50 (Tisbe
battagliai - Marine copepod) and 30 min EC50 (Vibrio fischeri - Microtox).
The 48h LC50 to Tisbe battagliai test shows a toxicity value of 8.2 Toxic Units. The 30min EC50 to Vibrio
fischeri test shows a toxicity value of 8 Toxic Units.
This would indicate that the BFW-1A wastewater emission point did register a minor acutely toxic effect
on the test species.
This report is certified as accurate and representative of the sampling and associated analysis carried out.
Respectively Submitted,
__________________________
Mr. Stephen Stapleton Mr. Peter Coogan
Environmental Scientist Environmental Team Leader
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Report No. ECS5071-BF-Tox
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1.0 METHODOLOGIES
1.1 Sampling
Representative grab samples of wastewater were extracted from Hach Lange autosampler in
accordance with the following standards;
TABLE 1.1: STANDARDS
ISO Standard
Description
ISO 5667-1-2006
Guidance on the design of sampling programmes and sampling
techniques
ISO 5667-3-2012
Guidance on sample preservation and handling
ISO 5667-14-2014
Guidance on quality assurance of environmental sampling & handling
ISO 5667-10-1992
Guidance on sampling wastewaters
Sampling was conducted in strict accordance with BNM Environmental recognised Standard
Operations Procedures (SOP) TS-W-02. All samples were returned to the laboratory, and stored
between 1-8°C. The sample was subsequently transported to the Aquatic services unit
Laboratory in Cork, Co. Cork.
1.2 Analysis
Analysis was conducted by Aquatic Services Unit. The wastewater sample was analysed for 48
hour LC50 (Tisbe battagliai - Marine copepod) and 30 min EC50 (Vibrio fischeri - Microtox). Table
1.2 presents the methodology used for the toxicity analysis of the wastewater sample.
TABLE 1.2: TOXICITY TESTING OF WASTEWATER SAMPLE
Test Species Method
Tisbe battagliai The direct toxicity assessment of aqueous environmental samples using
the marine copepod Tisbe battagliai lethality test based on the Environmental Agency 2007 test method
Vibrio fischeri Method 6.2 “Determination of the inhibitory effect of water samples on the
light emission of Vibrio fischeri (Luminescent bacteria test) – Part 3: Method using freeze-dried bacteria” based on ISO 11348-3: 2007
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2.0 RESULTS
The results of the toxicity analysis carried out by Aquatic Services Unit are presented in Table
2.1 as follows:
TABLE 2.1: TOXICITY RESULTS FOR EMISSIONS TO SEWER (BFW-1A)
Test Species Test Parameter Test ResultNote1 No. of Toxic Units
Vibrio fischeri 30 min. EC50 12.5 8
Tisbe battagliai 48 hr. LC50 12.2 8.2
Note 1: Effluent Concentration
3.0 COMMENT
The results of the bi-monthly toxicity testing of the wastewater sample extracted from the
emission reference point BFW-1A at the Henkel (Ireland) Ltd. Ballyfermot facility on the 17th of
December 2015 are presented in Table 2.1 above.
The toxicological effect of BFW-1A on the test species has decreased when compared with
previous results obtained in November (ECS5071).The 30 min EC50 to Vibrio fischeri has
decreased since the previous monitoring event with a toxicity value 145→8.0 Toxic Units at a
concentration of 12.5%vol/vol.
The 48h LC50 to Tisbe battagliai test has decreased since the previous monitoring event with a
toxicity value of 19→8.2 Toxic Units at a concentration of 12.2%vol/vol.
Based on the results from testing carried out on BFW-1A it would indicated that the samples is
minor acutely toxic.
Please see Appendix 1 for map of monitoring locations
Please see Appendix 2 for Aquatic services unit toxicity report
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Appendix 1: Site Location Map
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Appendix 2: Aquatic services unit toxicity report
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Report on the IE Toxicity Testing of the
Henkel BFW-1A final effluent
on behalf of
Bord Na Mona Environmental Ltd.
January 2016
Report prepared by
Aquatic Services Unit,
Environmental Research Institute.
Lee Road.
Cork
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2
Introduction
Toxicity testing was requested by for the final effluent sample at the Henkel
facility as part of their IE licensing requirements. The sample code for the Henkel
effluent is BFW-1A and the EPA IPC License number for the effluent is P0078-1. Two
aquatic species from different trophic levels were selected for the testing namely the
marine copepod (Tisbe battagliai) and the bacteria (Vibrio fischeri). Testing
commenced on December 16th, 2015 and finished on December 18th, 2015.
Methodology
The effluent sample was collected on December 15th, 2015 by Bord na Mona
Environmental personnel and then transported by courier service to the Aquatic
Services Unit (ASU) laboratory. The effluent sample arrived to the ASU laboratory on
December 16th, 2015 and had a temperature of 16.0ºC on arrival; the sample was then
immediately placed in a refrigerator at 4 ºC until required for testing.
Tisbe battagliai Bioassay
The Tisbe bioassay was carried out following standard methods as described in
Environment Agency’s Methods for the Examination of Waters and Associated
Materials (Environment Agency, 2007). The effluent was tested for toxicity at the
following concentrations 4, 8, 16, 32, and 64% effluent. The salinity of the effluent
was adjusted to 30 parts per thousand (ppt) using hypersaline brine to make the sample
suitable for testing. Twenty Tisbe isolates (animals 6 days old) were tested for each
concentration. These animals were added into 4 replicates of five animals per test
chamber for each concentration. Testing was carried out in a constant temperature room
at a temperature of 20 ºC ± 2 throughout the test. A light regime of 16 hours light/8
hours dark was used throughout the testing period. The test duration was 48 hours.
Filtered seawater adjusted to 30 ppt by the addition of distilled water was used as
control and dilution water. An additional brine control comprising of distilled water and
hypersaline brine was also tested. A concurrent reference toxicant using zinc sulphate
was conducted to assess the sensitivity of the test organisms. The T. battagliai were
obtained from in house cultures at the ASU.
Vibrio fischeri bacteria bioassay using Microtox system.
The luminescent bacteria Vibrio fischeri are used exclusively in the Microtox
system. Testing was carried out following the ISO 11348-3 guidelines (ISO, 2007).
Two replicates were used for each concentration tested. The effluent sample was
diluted by a factor of 4 using distilled water prior to testing based on the previous
evidence of toxicity associated with the effluent. The concentrations of effluent tested
ranged from 1.56 % to 20 %. A concurrent reference toxicant bioassay was also carried
out to determine the health and suitability of the bacteria.
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Statistical Analyses
Statistical analyses to generate LC50 (Lethal Concentration to cause 50 percent
mortality) or EC50 (Effective Concentration to cause 50 percent effect) data were
performed using the ToxCalc v5.0.32 Environmental Toxicity Data Analysis System,
(Tidepool Scientific, 2007). Microtox statistical analyses were carried out using the
manufacturers software programme MicrotoxOmni (Azur Environmental Ltd., 1995).
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Results and Discussion
Tisbe battagliai Bioassays
All of the tests were deemed to be valid given that there was greater than 90%
survival in the controls indicating that the animals and testing conditions were
satisfactory (Environment Agency, 2007).
Survival across the concentration series of effluent ranged from 100% at lowest effluent
concentration to 0% at concentrations exceeding 16% effluent. This bioassay predicted
an LC50 value of 12.2 % effluent. This corresponds to a Toxicity Unit (TU) value of
8.2 for the effluent. Results of this bioassay are displayed in Figure 1.
Figure 1 Average survival of Tisbe battagliai after 48 hours in a concentration
series of the Henkel BFW-1A effluent.
A concurrent reference toxicant bioassay using zinc sulphate was carried out
alongside the effluent bioassays. This bioassay produced an EC50 of 0.43 mg/L of Zinc
sulphate, this EC50 value is in agreement with previously published data for Tisbe sp.
(US EPA, 2007) and with ongoing toxicity testing at the ASU. This indicates that the
animals were of suitable sensitivity to be used for toxicity testing. Water quality
measurements in the test chambers remained within normal limits for the duration of
the bioassay. These data are presented in Appendix 1.
0
20
40
60
80
100
Control BrineControl
4 8 16 32 64
Su
rviv
ial
(%)
Effluent Concentration (%)
48 Hr Tisbe Bioassay
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Microtox results.
The Microtox tests carried out on December 17th, 2015 were deemed to be valid. A
reference toxicant test was carried out before actual testing of samples took place to
ensure that the bacterium and reagents were suitable for testing. The reference toxicant
using zinc sulphate determined an EC50 of 1.27 mg/L of (Zinc++) after 15 minutes
exposure. This is within the guideline range of 0.6 -2.2 mg/L (Zinc++) as specified by
the Microtox manufacturer (Azur Environmental, 1995). These data are presented in
Appendix 2.
A dose dependent increase in toxicity was observed between the controls and
throughout the effluent concentration series. Significant toxicity was observed at all
concentrations tested. A GL value of greater 48 was generated for this test. The GL
value is the dilution level at which an effluent causes less than 20% inhibition to the
species. Effluent samples with GL values ranging between 1-10 are considered to have
low toxicity, GL values of 10-100 are considered to be moderate toxicity and GL values
greater than 100 are considered to have high toxicity. This GL ranking system is based
on research carried out on selected effluents. (Wang et al., 2002). This bioassay
produced an EC50 of 12.5%. This would equate to a TU value of 8 for the BFW-1A
effluent. The data for this test are presented in Figure 2.
Figure 2 Plot of the Microtox results for the Henkel BFW-1A effluent sample.
Values represent light production relative to the controls. The highest
concentration of effluent tested was 20%.
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Summary and Conclusions
All of the bioassays performed in the current round of testing were deemed to
be valid as they meet all of the criteria as specified in the guidelines.
Exposure of the marine copepod, Tisbe battagliai, to the Henkel BFW-1A
effluent did produce significant toxicity concentrations exceeding 8% effluent. The
Tisbe bioassay determined an LC50 of 12.2 % effluent or a TU value of 8.2.
In the Microtox bioassay, exposure to the BFW-1A effluent did have a
statistically significant inhibitory effect on light production in the Vibrio fischeri
bacteria. A dose dependent decrease in light production was observed across the
concentration series. The Microtox bioassay determined an EC50 of 12.54% effluent
which corresponds to TU value of 8. Both tested produced similar Toxic Unit values
for the effluent.
The toxicity of effluents has been classified based on their TU values using the
following scale TU<1 (relatively not acutely toxic), TU between 1 and 10 (minor
acutely toxic), TU between 10 and 100 (moderately acutely toxic) and TU>100 (very
acutely toxic) (Tonkes et al., 1999). Based on this scale, the results from the bioassays
performed on the BFW-1A final effluent sampled on December 15th, 2015 indicate that
the sample is minor acutely toxic.
References
Azur Environmental, 1995. Microtox Acute Basic Test Procedures. Azur
Environmental ,. Carlsbad, California, USA.
Environment Agency, 2007. The direct toxicity assessment of aqueous environmental
samples using the marine copepod Tisbe battagliai lethality test. Methods for the
Examination of Waters and Associated Materials. http://www.environment-
agency.gov.uk/nls
ISO 11348-3:2007. Water quality-Determination of the inhibitory effect of water
samples on the light emission of Vibrio fischeri (Luminescent bacteria test)—Part 3:
method using freeze dried bacteria. International Standards Organisation.
Tonkes, M. Graaf, P.J.F., and Jannes, G.,1999. Assessment of Complex Industrial
Effluents in the Netherlands using a Whole Effluent Toxicity (or WET) Approach.
Water Science and Technology, Vol. 39, No. 10-11, pp 55-61.
U.S EPA, 2007. ECOTOX User Guide: ECOTOXicology Database System. Version
4. http:/www.epa.gov/ecotox.
Wang, C., Yediler, A., Lienert, D., Wng, Z., and Kettrup, A., 2002. Toxicity
evaluation of reactive dyestuffs , auxiliaries and selected effluents in the textile
finishing industry to luminescent bacteria Vibrio fischeri. Chemosphere, 46, pp 339-
344
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Appendix 1- Water Quality Measurements for Tisbe battagliai Bioassayss
48 Hour Tisbe Bioassay
Bord na Mona BFW-1 Effluent 16/12/15
Concentration Survival D.O (mg/L) pH Salinity (ppt) Temp (ºC)
(%) 0hrs 24hrs 48hrs 0hrs 24hrs 48hrs 0hrs 24hrs 48hrs 0hrs 24hrs 48hrs 0hrs 24hrs 48hrs
Control A 5 5 5 6.9 7.1 7.93 8.10 30 30 21.0 20.8 19.4Control B 5 5 5Control C 5 5 5Control D 5 5 5Control 2A 5 5 5 6.9 7.1 7.93 8.10 30 30 21.0 20.8 19.4Control 2B 5 5 5Control 2C 5 5 5Control 2D 5 5 5
Brine Control A 5 5 5 7.6 7.0 7.98 7.88 29 30 19.2 20.8 19.2Brine Control B 5 5 5Brine Control C 5 5 5Brine Control D 5 5 5Brine Control 2A 5 5 5 7.6 7.0 7.98 7.88 29 30 19.2 20.8 19.2Brine Control 2B 5 5 5Brine Control 2C 5 5 5Brine Control 2D 5 5 5
4A 5 5 5 7.1 5.9 7.97 7.96 30 30 19.2 20.8 19.24B 5 5 54C 5 5 54D 5 5 58A 5 5 5 6.9 5.9 7.95 7.94 29 30 19.2 20.8 19.28B 5 5 48C 5 5 58D 5 5 516A 5 3 0 6.8 5.7 7.87 7.89 29 30 19.2 20.8 19.216B 5 3 216C 5 4 016D 5 3 132A 5 2 0 6.6 5.4 8.32 7.87 29 31 19.2 20.8 19.132B 5 3 032C 5 3 032D 5 0 064A 5 0 0 5.7 3.3 8.08 7.55 29 31 19.1 20.8 19.164B 5 0 064C 5 0 064D 5 0 0
Notes Adjusted pH of 32 and 64% from 7.29 to8.32 and 6.80 to 8.08 respectively.
Day 0 Initiated at 16.00 , Animals 6 days old at initiation
24hrs 16.00
48hrs Terminated at 14.30
Testing performed by Aquatic Services Unit, ERI Building, Lee Rd., Cork
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Appendix 2- Microtox Reference Toxicant Bioassay
Plot of light inhibition relative to the controls versus Zinc++ concentration after 5
and 15 minutes exposure.
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The toxicity testing carried out for this report complies with internationally accepted
guidelines. The results of these toxicity tests contained in this report are deemed valid
under these guidelines.
Signed:
David Gillespie MSc (Toxicology)
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