C12-18-(even numbered, C18 unsaturated)-alkylamines acetates

Endpoint summary

Additional information

BIOACCUMULATION: AQUATIC / SEDIMENT

Introduction

As Primary alkyl amines are readily biodegradable it can be expected that metabolism will occur in aquatic species like fish. This is confirmed by in vitro measurement of the metabolic rate of 1-Hexadecanamine (C16 amine) which was selected as model substance for the Primary alkyl amines (Perdu-Durand, 2006). This means that establishing of the Bioconcentration factor BCF should address Adsorption, Distribution, Metabolism and Excretion (so called ADME process).

Primary alkyl amines and acetates are cationic surfactants which sorb strongly to solid phases by van der Waals and ionic interactions (e.g. ion pair formation, cation exchange etc). This makes it extremely difficult or even impossible to ensure a constant concentration of the test item in a flow through system because of sorption but also by biodegradation as the test system is not sterile. Additionally as fish mucous is negatively charged the cationic surfactant is sorbing on the fish surface as in the test water used for a BCF test no dissolved organic carbon (DOC) or suspended matter is present as it would under environmental conditions.

Usually for the determination of the BCF an OECD 305 BCF test would be carried out as the ADME process would be addressed completely. But due to the inherent properties of cationic surfactants technical issues arise and the measurement is likely to be highly uncertain and the result even not valid. Therefore also estimation procedures for BCF have to be considered and evaluated for use in the context of this assessment under REACH.

 

Attempt to measure the BCF in an OECD 305 Test with 1-Hexadecanamine

Despite a high technical effort a constant flow through with acceptable analytical recoveries could not be established (Akzo Nobel, 2007). In addition due to the negatively charged fish surface continous sorption of C16 amine to the fish mucous took place. Therefore real steady state could not be observed. The calculated BCF taking into account the adsorbed fraction in the mucous gave high values which are unrealistic. Removing the adsorbed C16 amine with methanolic hydrogen chloride resulted in BCF values around 300 L/kg but the validity of this figure cannot be judged. The conclusion is that the conditions proposed in the OECD 305 Guideline do not fit to the intrinsic properties of the n-Primary alkyl amines in general. A test in natural river water which would mimic realistic environmental conditions may be more appropriate but could lead to analytical issues like insufficient sensitivity even when using 14C material.

Conclusion: This preliminary study shows that the conditions described in the OECD 305 Guideline will not be applicable for n-Primary alkyl amines. But alternative conditions e.g. use of river water might create other issues like insufficient analytical sensitivity.

SPECIATION OF PRIMARY ALKYL AMINES

In aqueous medium the unprotonated and the protonated amine are in equilibrium. The percentage of the unprotonated and the protonated amine is determined by the acid constant pKa and the given pH. In the table the percentages are given as function of pH and the pKa of 10.5 for Primary alkyl amines.

pH	Percentage Protonated amine (N+)	Percentage Unprotonated amine (N)
9	97.5%	2.5%
7	99.975%	0.025%
4	99.99997%	0.000003%

Log Kow FOR THE C CHAIN MIXTURE OF THE REGISTRATION SUBSTANCE

In a chemical mixture the extensive properties like volume and mass are additive (Brdicka, 1970). The average Mol mass of a mixture can be described as the sum of the mol fraction times mol mass of the chemical components

M_average = x₁*M₁+ x₂*M₂.. + x_i* M_i

and the average Mol volume of the chemical mixture is the sum of the volume fractions times Mol volume of the chemical components

V_average = x₁*V₁+ x₂*V₂..+ x_i* V_i

Based on the additivity for Mol mass and Mol volume in a mixture it is justified to define an average Log Kow by an additive approach taking into account the molar fraction y of Log Kow of the corresponding C chain homologue. Under environmental conditions (pH 4-9) speciation of the primary alkyl amines is observed which means that the unprotonated and the protonated amine co-exist and their molar ratio is determined by the pKa of the amine. This means that for each C chain homologue both species have to be considered in form of their molar fraction x_i for the unprotonated form and x_i+ for the protonated one e.g.

Log Kow_average= (x_Cn-N*Log Kow_C8-N+ x_C8-N+*Log Kow_Cn-N+)*y_C8-N…+ (x_i*Log Kow_i+
                              x_i+*LogKow_i+)*y_i

or in the general form                   

Log Kow_average=    SUM_j(x_i*Log Kow_i+ x_i+*Log Kow_i+)*y_i

x_i is the molar fraction of the unprotonated C chain homologue i
Log Kow_i is the Log Kow of the unprotonated C chain homologue i
x_i+ is the molar fraction of the protonated C chain homologue i
Log Kow_i+ is the Log Kow of the protonated C chain homologue i
y_i is the molar fraction of the C chain homologue i in the total number j of all C chain homologues

Based on this additivity approach the average Log Kow registration substacne mixture is calculated (see Table below)

The log Kow for the unprotonated amines were calculated with the US EPA KOWWIN Program. The log Kow for the protonated amine was calculated from the unprotonated value deducting a factor of 3.5. This approached is supported by Fu et al (2009) and by measured Log Kow based on measured octanol and water solubility / CMC of the amine species. An example is given for the read across substance C16 -18 -(even numbered) alkyl amines. The registration substance has a higher fraction of shorter chain homologues and therefore an overall lower log Kow than the example substance. A lower log Kow means also a lower BCF but for simplicity reasons the calculated BCF of the example substance will be used as a worst case.

						Average Log Kow
	Chain length	Weight Percentage	MW (g/mol)	Mol	Mol-%	R-NH2 contrib.	R-NH3 contrib.	pH4	pH7	pH9
C16-18-(even numbered) alkylamines (Hydrogenated tallow alkyl amines)	C12	1%	185.36	0.05	1%	0.05	0.01	0.01	0.01	0.01
	C14	4%	213.41	0.19	5%	0.23	0.09	0.09	0.09	0.10
	C16	32%	241.46	1.33	34%	2.14	1.02	1.02	1.02	1.05
	C18	62%	269.52	2.30	59%	4.77	2.60	2.60	2.60	2.66
	OVERALL	99%		3.87		7.20	3.73	3.73	3.73	3.82

ARNOT & GOBAS BCF FISH ADME MODEL (US EPA BCF BAF Estimation Program)

Under environmental conditions pH 4 -9 most of the registration substance exists in protonated form (see above). The protonated form is a cationic surfactant with properties like strong sorption to negatively charged surfaces e.g. glass or fish, algae which makes classical BCF testing unfeasable. Therefore using the Arnot & Gobas ADME Fish BCF Model is currently the best available method for estimating the BCF Fish of the registration substance mixture taking into account speciation as well.

In the table below the results from the ADME Model for the registration substance mixture is given

					Average Log Kow					Average BCF (BCFBAF Arnot Gobas)
	Chain length	Weight %	MW (g/mol)	Mol-%	R-NH2 contrib.	R-HH3 contrib.	pH4	pH7	pH9	R-NH2 contrib.	R-HH3 contrib.	pH4	pH7	pH9
C16-18-(even numbered) alkylamines (Hydrogenated tallow alkyl amines)	C12	1%	185.36	1%	0.05	0.01	0.01	0.01	0.01	12	0	0.0	0.0	0.3
	C14	4%	213.41	5%	0.23	0.09	0.09	0.09	0.10	81	1	0.9	0.9	2.9
	C16	32%	241.46	34%	2.14	1.02	1.02	1.02	1.05	559	37	37	37	50
	C18	62%	269.52	59%	4.77	2.60	2.60	2.60	2.66	327	263	263	263	264
	OVERALL	99%			7.20	3.73	3.73	3.73	3.82	979	301	301	301	318

CONCLUSION

In the environmental pH range 4 -9 the BCF Fish of the registration substance (C12 -18 -(even numbered, C18 -unsaturated)-alkylamines acetate which is similar to C16-18-(even numbered) alkylamines mixture is in the range 301 to 318 L/kg wwt (worst case) which is far below the 'B' criteria of the PBT / vPvB Assessment. The value of 318 L/kg wwt is used in exposure calculations.

BIOACCUMULATION: TERRESTRIAL

Unfortunately no ADME Model exists for earthworms. But as a worst case the BCF worm may be estimated by a QSAR not taking into account metabolism (see REACH Guidance R16.6.7) .

BCF = (0.84 + 0.012*Kow)/RHO earthworm with Kow = 6606 (=Log Kow 3.82 see above) and RHO earthworm = 1 kg/L wwt

BCF = 80 L/kg wwt which is even lower than the BCF fish estimated before.

CONCLUSION

Based on the highest Log Kow for the example substance mixture of 3.82 (pH 9) a BCF worm of 80 L/kg wwt was calculated which is lower than the BCF fish. The registered substance has a lower log Kow and therefore the BCF worm of 80 L/kg wwt can be regarded as worst case.

Additional Literature

• Albro et. al (1993): The metabolism of di-(2-ethylhexyl) phthalate in the earthworm Lumbricus terrestris,  Comp. Biochem. Physiol. C: Comp. Pharmacol.; 1993, 104 (2), 335 -344 

• Armitage & Gobas (2007): A terrestrial food-chain bioaccumulation model for POPs, Environ. Sci. Technol., 2007, 41(11), 4019-25.

• Perdu-Durand et al (2006): Hexadecylamine biotransformation rates in carp and rainbow trout liver subcellular fractions, ERASM (http://www.erasm.org/Study/INRA_report-subcellular-C16PA-060701.pdf), 2006

• Akzo Nobel (2007): Thomas, PC et al., Status report on the methodology development work to determine the bioconcentration factor (BCF) of 1-Hexadecylamine in the carp Cyprinus carpio under flow through conditions, Final research report, 13.12.2007