Deriving bioconcentration factors of constituents of essential oils - - PowerPoint PPT Presentation

Deriving bioconcentration factors

• f constituents of essential oils

using in-vivo benchmarked dietary exposure studies

Roxana Sühring, Chang’er L. Chen, Gisela Horlitz, Michael McLachlan, Matthew MacLeod

Bioconcentration is important …and difficult to measure

𝑪𝑫𝑮 = 𝑫𝒈𝒋𝒕𝒊 𝑫𝒙𝒃𝒖𝒇𝒔 = 𝒍𝟐 𝒍𝑼

Variability? Water-based exposure? Mixtures?

1Arnot & Gobas 2004 2Arnot & Gobas 2006

kD k1 k2 kE kG kM

𝑒𝐷F 𝑒𝑢 = 𝑙1𝐷W + 𝑙𝐸𝐷𝐸 − 𝑙2 + 𝑙𝐹 + 𝑙𝑁 + 𝑙𝐻 𝐷F

Arnot & Gobas, QSAR, 2003，Chen et. al. ES&T 2018, OECD 305 2012 Revisions

Using dietary exposure

𝑪𝑫𝑮 = 𝒍𝟐 𝒍𝑼

measured estimated

What is benchmarking and how can it help?

Benchmarking with a conservative substance

𝑒𝐷 𝑒𝑢 ⇒ 𝑒𝐷/𝑒𝐷𝐶𝑁 𝑒𝑢

Days Ln(C / ng g-1 ww)

No benchmark

Days

Benchmarkd with HCB 3,4

3Xiao et al. 2013 4Chen et al. 2018

Can we measure the BCF of mixtures?

Benefits

• More representative of the chemical product
• Reducing the number of test animals

Evaluation

• 16 chemicals with published tested and

predicted BCF data for individual substances

• Reported BCFs from < 100 to ~17000

BM-BCF BCF-literature

BM-BCF

BCF-literature Not statistically different (t-test, p = 0.33)

Threshold benchmarking5

Is the depuration rate faster or slower than the benchmark?

5Zou et al. 2015

Ln(C / ng g-1 ww)

No benchmark

Days Days

Benchmarkd with PeCB

Very Bioaccumulative

𝑪𝑫𝑮 = 𝒍𝟐 𝒍𝑼

Mixtures? Uncertainty of k1 estimate

𝑪𝑫𝑮𝑪𝑵 = 𝒍𝟐 𝒍𝑼𝑯

Water-based exposure? Variability?

BCF of Essential oils

Pine oil

• > 60 components
• Analytical standards for compounds

> 1% contribution (n = 9)

• > 88% of the mixture

Cedarwood oil (virginian)

• ~ 60 components
• Analytical standards for compounds > 1%

contribution (n = 7, detected n = 6)

• 75% - 80% of the mixture

(depending on batch)

Image from wikipedia.org Image from wikipedia.org

β-pinene (BPN) Carene (CAN) Terpinolene (TPN) Borneol (BNL) Bornyl Acetate (BAC) β-caryophyllene (BCP)

Measured pine oil constituents

α-pinene (APN) Camphene (CAM) Limonene (LIM)

Results: Pine oil

Log BCFBM = 0.97 x Log KOW - 0.88 R² = 0.95 B limit vB limit

Median BCF for BCP meets B criterion No constituents are B at the 95% confidence level

Thujopsene (Thu) Cuparene (Cup) Cedrol (CDL) α-Funebrene (aFun)

Measured Cedarwood oil constituents

α-Cedrene (aCed) β-Cedrene (bCed)

Biotransformation of Cedarwood oil?

Log BCFBM = 0.70 x Log KOW + 0.16 R² = 0.94 B limit vB limit

Biotransformation of Cedarwood oil constituents exceeds biotransformation of the reference substances BUT: Four out of six constituents are B or vB at the 95% confidence level

Comparison with literature data

Benchmark substances analysed in the Cedarwood oil study Benchmark substances analysed in the Pine oil study

Dichlorobenzene (DiCB) Hexachlorobenzene (HCB) Musk xylene (MX) PCB52 Pentachlorobenzene (PeCB) Trichlorobenzene (TrCB)

What about P and T of Cedarwood oil?

Screening T criterion: EC50 < 0.01 mg/L (potential T), < 0.01 mg/L (T)

• Cedarwood oil has applications as natural biocide/insecticide6.
• α-Cedrene EC50: 0.044 mg/L (Daphnia pulex)7
• Cedrol: indications for endocrine disruption8 and gentoxicity9
• No T results for the remaining Cedarwood oil constituents

Readily biodegradable: ≥ 60% degradation in a ready biodegradation test

• α-Cedrene and Cedrol: > 75% biodeg in 28 days (OECD 301 test)10
• Thujopsene: 36 % biodeg in 28 days test, 56 % in 60 days test (OECD 301) 10
• No biodeg test results for the remaining Cedarwood oil constituents
• α-Cedrene: not P, vB, potential T
• Cedrol: not P, not B, unknown acute T, publications indicating potential

genotox, ED

• Thujopsene: not readily but significant 56% degradation, B, unknown T
• No experimental results for most of the analysed constituents

Thank you for your attention! Take home messages

• Dietary exposure with internal benchmarking provides a robust method for depuration rate

measurements of complex mixtures in fish

• Results for reference substances match published measurements for individual components
• Number of test animals can be reduced
• Higher biotransformation of Pine oil and Cedarwood oil compared to reference substances
• Pine oil likely contains at least one B constituent but most constituents are not B
• Most measure constituents of Cedarwood oil are B or vB
• More information on NCSs and constituents is needed

References

1.

Arnot JA, Gobas FAPC. 2004. A food web bioaccumulation model for organic chemicals in aquatic ecosystems. Environ. Toxicol. Chem. 23 (10): 2343−2355.

2.

Arnot JA, Gobas FAPC. 2006. A review of bioconcentration factor (BCF) and bioaccumulation factor (BAF) assessments for organic chemicals in aquatic organisms. Environ.

• Rev. 14: 254–297.

3.

Xiao RY, Adolfsson-Erici M, Akerman G, McLachlan MS, MacLeod M. 2013. A Benchmarking Method to Measure Dietary Absorption Efficiency of Chemicals by Fish. Environ.

• Toxicol. Chem. 32 (12): 2695−2700.

4.

Chen CL, Löfstrand K, Adolfsson-Erici M, McLachlan MS, MacLeod M. 2018. Deriving in Vivo Bioconcentration Factors of a Mixture of Fragrance Ingredients Using a Single Dietary Exposure and Internal Benchmarking. Environ. Sci. Technol. 2018, 52, 5227−5235.

5.

Zou H, Radke M, Kierkegaard A, MacLeod M, McLachlan MS. 2015. Using Chemical Benchmarking to Determine the Persistence of Chemicals in a Swedish Lake. Environ. Sci.

• Technol. 2015, 49, 1646−1653.

6.

Kramer A, Guggenbichler P, Heldt P, Juergen K, Ladwig A, Thierbach H, Weber U, Daeschlein G. 2006. Hygenic Relevance and Risk Assessment of Antimicrobial-Impregnated

• Textiles. In: Hipler UC & Elsner P. Biofunctional Textiles and the Skin. Current Problems in Dermatology, Vol. 33. ISBN: 3-8055-8121-1. page 94.

7.

Passino-Reader DR, Hickey JP, Ogilvie LM. 1997. Toxicity to Daphnia pulex and QSAR Predictions for Polycyclic Hydrocarbons Representative of Great Lakes Contaminants.

• Bull. Environ. Contam. Toxicol. 59:834-840.

8.

Simon C, Onghena M, Covaci A, VanHoeck E, Van Loco J, Vandermarken T, Van Langenhove K, Demaegdt H, Mertens B, Vandermeiren K, Scippo ML, Elskens M. 2016. Screening of endocrine activity of compounds migrating from plastic baby bottles using a multi-receptor panel of in vitro bioassays. Toxicology in Vitro 37. 121–133.

9.

Mertens B, Simon C, Van Bossuyt M, Onghena M, Vandermarken T, Van Langenhove K, Demaegt H, VanHoeck E, Van Loco J, Vandermeiren K, Covaci A, Scippo ML, Elskens M, Verschaeve L. 2016. Investigation of the genotoxicity of substances migrating from polycarbonate replacement baby bottles to identify chemicals of high concern. Food and Chemical Toxicology 89. 126-137.

10.

Jenner KJ, Kreutzer G, Racine P. 2011. Persistency assessment and aerobic biodegradation of selected cyclic sesquiterpenes present in essential oils. Environ Toxicol Chem30:1096 – 1108.

Hexane phase

ACN

Elute with Hex

Ultrasound assisted extraction - Purge & trap – GC-MS

MQ water hexane

BCF BM-BCF Target 5th Median 95th 5th Median 95th α-Cedrene 4600 138000 infinite 6900 11000 23000 β-Cedrene 4400 69000 infinite 6000 9200 20000 Thujopsene 3000 8600 infinite 3600 4800 6900 Cuparene 1900 2900 6900 1700 2100 2600 Cedrol 570 710 940 520 600 720 α-Funebrene 4100 23000 infinite 4400 6900 15000 BMs TrCB 610 800 1200 650 750 870 PeCB 3500 12000 infinite 3800 5500 9900 HCB 12000

• 10615

infinite n.a. n.a. n.a. PCB3 2800 6900 infinite 2800 4100 7700

Name kT kTG α-Cedrene 0.001 ± 0.029

• 0.012 ± 0.01

β-Cedrene 0.002 ± 0.029

• 0.01 ± 0.009

Thujopsene 0.016 ± 0.03 0.004 ± 0.011 Cuparene 0.047 ± 0.027 0.044 ± 0.012 Cedrol 0.195 ± 0.049 0.203 ± 0.036 α-Funebrene 0.006 ± 0.028 -0.004 ± 0.009 BMs TrCB 0.173 ± 0.053 0.15 ± 0.024 PeCB 0.012 ± 0.027 n/a HCB

• 0.013 ± 0.024 -0.025 ± 0.011

PCB3 0.02 ± 0.03 0.01 ± 0.014

Threshold benchmarking

Composition [%]