Chemex Environmental International Ltd

Chemex Environmental International Ltd

Environmental Fate Services

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An extensive range of studies to help assess the fate of a chemical in the environment. Assessing the degradation and accumulation potential of products plays an important role in measuring their ability to persist in and damage the environment. The degradation of a test material by micro-organisms or by physico-chemical means can be measured. Bioaccumulation can be assessed by analytical techniques or by measuring the body burden following exposure of an organism to a chemical.

Correctly pairing a sample with a test method can mean the difference between a pass and a fail in a biodegradation test.  Chemex currently offer a range of Biodegradation test methods covering, but not restricted to, Ready Biodegradation (OECD 301A, B, D, E and F, 310 and Freshwater BODIS), Inherent Biodegradation (OECD 302B and CONCAWE), Anaerobic (ISO 11734) and Marine Biodegradation (OECD 306 and Marine BODIS).  All studies are conducted with GLP compliance.

Bioaccumulation potential forms part of a HOCNF submission.  Chemex has analysts who have been performing this essential test routinely for many years, in addition, we play an active role in EOSCA as an Associate Member, ensuring that our analytical techniques are current and fit for purpose.

Our HPLC capability and range of detectors (UV/Vis, RI, Fluorescence) enable us to give you comprehensive data for your material(s) and we have years of experience in dealing with ‘difficult’ samples.

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Abiotic Degradation (OECD 111)

Abiotic degradation - Hydrolysis as a function of pH
Hydrolysis is an important reaction controlling degradation. The reaction is particularly relevant for substances with low biodegradability and can influence the persistence of a substance in the environment. This method is only applicable to water soluble substances.

The test material is dissolved in water at low concentration with pH and temperature control. The decrease in the concentration over time is measured by a suitable analytical technique. The test is performed at three pH values (normally 4.0, 7.0 and 9.0). The initial testing is performed at 50oC with two further analyses performed at a temperature below this (typically 40oC).

The rate constants for each condition are derived from plots of the log of concentration against time.

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Anaerobic Biodegradation (ISO/DIS 11734 / OECD 311)

Freshwater Anaerobic Biodegradation Study
This method is intended to provide a simple and reliable screening test for the ultimate biodegradability of chemicals under anaerobic conditions. The method is applicable to soluble and poorly soluble organic chemicals.

A known volume of anaerobic sludge (corresponding to 10% of the sludge concentration in a real digester) suspended in an oxygen free medium is placed in a suitable vessel with a headspace into which any gases produced may be evolved. Prior to sealing a small amount of test compound is added. The vessels are incubated at constant temperature (35 ± 1°C) and pH for a period of 8 weeks. The headspace pressure, resulting from the production of gas, is measured.

From the measured values of net gas production the extent of biodegradation is calculated. The kinetics of the degradation are followed by intermediate measurements at suitable intervals during the course of the test.

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Bioaccummulation Potential (OECD 107)

Bioaccummulation Potential - Shake Flask Method (OECD 107)
This guideline determines experimentally n-octanol/water partition coefficient values in the range log Pow between -2 and 4 (occasionally up to 5). This particular method is not suitable for surface-active materials. The partition coefficient is defined as the ratio of the equilibrium concentrations of a dissolved substance in a two-phase system consisting of two largely immiscible solvents.

The test should be carried out at a temperature in the range of 20 to 25°C and kept constant at ± 1°. There are three runs with different volume ratios of n-octanol to water. Duplicate vessels containing accurately measured amounts of the two solvents and stock solution are used in all three runs. After agitation the separation of the two phases, in general, is achieved by centrifugation. It is necessary to determine the concentrations of the test substance in both phases. Analytical methods which may be appropriate are: photometry, gas chromatography and high performance liquid chromatography. The total quantity of substance present in both phases should be calculated and compared with the quantity originally introduced. A Pow value is calculated from the data of each run. The six log Pow values should fall within a range of ± 0.3 units.

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Bioaccummulation Potential (OECD 117)

Bioaccummulation Potential - HPLC Method (OECD 117)
This Test Guideline describes the HPLC method, which is used to determine the partition coefficient (n-octanol/water) (Pow).

Reverse phase HPLC is performed on analytical columns packed with a solid phase containing long hydrocarbon chains chemically bound onto silica. The chemicals are retained in the column in proportion to their hydrocarbon-water partition coefficient, with hydrophilic chemicals eluted first and lipophilic chemicals last. The HPLC method covers log Pow in the range of 0 to 6, but it can be expanded to cover the log Pow range between 6 and 10 in exceptional cases. The HPLC operation mode is isocratic. The retention time is determined in duplicate. The partition coefficient of the test substance is obtained by interpolation of the calculated capacity factor on the calibration graph. For very low and very high partition coefficients extrapolation is necessary.retained

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Biodegradability in Seawater (OECD 306)

Marine Biodegradability in Seawater – Closed Bottle Method (OECD 306)
This method is appropriate for volatile materials and although good water solubility is desired, poorly soluble materials can be tested. The test vessels are closed filled bottles containing the test substance dissolved in the natural seawater.  The bottles are incubated in the dark at 20°C and the degradation is followed by analysis of the dissolved oxygen over a 28 day period.

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Biodegradation - Closed Bottle Test (OECD 301D)

Freshwater Closed Bottle Test (OECD 301D, EC C.4.4)
This Freshwater method is appropriate for volatile materials and although good water solubility is desired, poorly soluble materials can be tested. As implied by the title, this is conducted in a closed system and oxygen is consumption measured by electrochemical probes or through chemical analysis. The principle is similar to the Modified MITI test and suitable controls are used.

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BODIS in Seawater (PARCOM)

Biodegradability of Insoluble Substances (BODIS) in Seawater (PARCOM)
This method was developed for the assessment of degradation of poorly soluble materials in the marine environment, however, it can also be adpated for Freshwater purposes. The test vessels are closed glass bottles with a known volume of natural seawater (2/3) and air (1/3). They are shaken continuously to ensure steady state oxygen partitioning between liquid and gas phases. The degradation is followed by weekly measurements of the biochemical oxygen demand (BOD) in the aqueous phase for a 28 day period.

The BOD is calculated from the cumulative oxygen uptake.

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DOC Die-Away Test (OECD 301A / EC C.4.3)

Freshwater Biodegradation - DOC Die-Away Test
The loss of dissolved organic carbon (DOC) is followed by analysis every few days during the 28 day study period and is compared with the total organic carbon for the known weight of test substance. A sodium acetate reference is used and greater than 60% loss is expected.

This method is similar to the Modified OECD Screening Test (OECD 301E) but uses a higher concentration of micro-organisms.

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Estimation of the Adsorption Coefficient on Soil and Sewage Sludge (OECD 121)

Estimation of the Adsorption Coefficient (Koc) on Soil and Sewage Sludge using HPLC (OECD 121)An important parameter in the sorption behaviour of substances is the adsorption coefficient (Koc). This is defined as the ratio between the concentration of the substance in the soil/sludge and the concentration of the substance in the aqueous phase at adsorption equilibrium. The adsorption coefficient normalised to the organic content of the soil Koc is a useful indicator of the binding capacity of a chemical on organic matter of soil and sewage sludge and allows comparisons to be made between different chemicals.

The experimental method uses High Performance Liquid Chromatography (HPLC) for the estimation of the adsorption coefficient (Koc) in soil and sewage sludge. While passing through the column in the mobile phase the test substance interacts with the stationary phase. As a result of partitioning between mobile and stationary phases the test substance is retarded.  The dual-composition of the stationary phase having polar and non-polar sites allows for the interaction of polar and non-polar groups of a molecule in a similar way as is the case for organic matter in soil or sewage sludge matrices. This enables the relationship between the retention time on the column and the adsorption coefficient on organic matter to be established.

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Manometric Respirometry (OECD 301F)

Freshwater Manometric Respirometry Test (Oxitop) - Ready Biodegradability
This freshwater ready biodegradibility test uses Oxitop apparatus and is part of the OECD 301 series of methods.

A measured volume of inoculated mineral medium, containing a known concentration of the test substance (100 mg/l giving at least 50-100mg ThOD/l) as the source of organic carbon, is stirred in a closed flask at a constant temperature for up to 28 days. During the course of the test oxygen is consumed and carbon dioxide evolved. The carbon dioxide is absorbed in sodium hydroxide and the drop in pressure in the test vessels therefore determines the oxygen consumption. The caps of the test vessels contain a pressure transducer and microprocessor to measure and calculate BOD.

The amount of oxygen taken up by the microbial population during biodegradation of the test substance is expressed as a percentage of ThOD or, less satisfactorily, COD.

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Modified Sturm Test (OECD 301B)

Freshwater Modified Sturm Test (OECD 301B / EC C.4.3)
This method is suitable for non-volatile substances that may be poorly soluble in water.

The amount of carbon dioxide (CO2) produced as a percentage of theoretical yield (based on total organic carbon analysis) is used as a basis for assessing whether the material biodegrades. CO2 is measured by way of a sodium hydroxide trap. The study is run for a minimum of 28 days and may be continued if the yield of CO2 is showing signs of increase towards the end of the 28 day period.

A positive control of sodium acetate is run alongside and a degradation of greater than 60% over 28 days should be seen. A blank control containing just the inoculum and nutrient mixture is used to check background levels of activity.

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OCNS Suite

Wherever chemicals are used offshore, the Offshore Chemical Notification Scheme (OCNS) requires a range of ecotoxicological tests to be carried out.   
The OCNS suite is a defined range of ecotoxicological tests that are needed to meet the requirements of the OCNS in order to provide sufficient data to allow assessment of potential environmental risk in the marine environment.

The OCNS requires the collection of bioaccumulation, biodegradation, and aquatic toxicity data from three trophic levels (algae, crustacea and fish). This data is used to predict the potential ecosystem risk. This information can then be used to rank the product by Hazard Quotient (HQ).

Further general information regarding the OCNS can be found in the associated information document 'OCNS - Specifications'.

The ecotoxicology tests that Chemex provide to cover the requirements of the OCNS are:

  • Bioaccumulation Potential (OECD 107-Shake Flask or OECD 117-HPLC)
  • Biodegradation (OECD 306-Closed Bottle or Shake Flask methods or Marine BODIS generally)
  • 72 Hour EC50Alga growth inhibition (Phaeodactylum tricornutum) ISO 10253 2006
  • 48 Hour LC50Marine copepod acute toxicity (Acartia tonsa) – Static ISO 14669 1999
  • Marine Fish (Cyprinodon variegatus or Scophthalmus maximus) – Semi-static  PARCOM
  • 10 Day Sediment Test (Corophium sp) LC50 – Static PARCOM - This is not always required and is dependant on the nature of the sample.

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OECD Biodegradation Screening (OECD 301E)

Freshwater Modified OECD Screening test (OECD 301E / EC C.4.2)
The loss of dissolved organic carbon (DOC) is followed by analysis every few days during the 28 day study period and is compared with the total organic carbon for the known weight of test substance. A sodium acetate reference is used and greater than 60% loss is expected.

This method is similar to the DOC Die-Away Test (OECD 301A) but employs a relatively low concentration of micro-organisms.

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REACH Testing

Chemex can offer a range of studies relevant to your REACH Submissions
Chemex can offer a number of studies that will be relevant to your REACH submissions.  You can rest assured that we have the systems, staff and facilities in place to conduct studies for REACH purposes to a very high standard.

REACH is a significant driving force in the work that Chemex conducts, including Physico-Chemical, Biodegradation, Bioaccumulation, Aquatic Toxicology and Terrestrial Toxicology, all to GLP Compliance.

Chemex remain at the forefront of Ecotoxicology testing, supplying high quality data at competitive prices.

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Ready Biodegradability (ISO 14593 / OECD 310)

Freshwater Ready Biodegradability – CO2 Headspace Test
This method is based on ISO 14593 / OECD 310 : ‘Carbon dioxide (CO2) Headspace Biodegradation Test’ and provides a test for assessing the ultimate, aerobic biodegradability of organic substances in the freshwater environment. It provides similar information to the methods described in OECD 301.

The test is performed in sealed bottles with a headspace of air that provides a reservoir of oxygen (O2) for aerobic biodegradation. CO2 evolution from the ultimate aerobic biodegradation of the test substance is determined by measuring the inorganic carbon (IC) produced in the test bottles over that produced in blanks that contain inoculated medium only. The ultimate aerobic biodegradation is the breakdown of an organic chemical by micro-organisms in the presence of O2 resulting in the production of CO2, water and mineral salts (ie mineralisation) and microbial cellular constituents (biomass).

The extent of biodegradation is then expressed as a percentage of the theoretical maximum IC production (ThIC), based on the quantity of test substance (as total organic carbon) added initially. The test is typically run for 28 days.  Values =60% ThIC in a 10 day window demonstrates ready biodegradability.

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Zahn-Wellens/EMPA Test (OECD 302B)

Freshwater Modified Zahn-Wellens/EMPA Test
This freshwater method is not suitable for volatile materials.

The test material is added to a large volume of activated sludge and nutrient mixture, and the decline in dissolved oxygen concentration measured frequently. The duration of the study is generally 28 days but may be extended if a significant lag period is noted at the start. A result of > 70% degradation means that the substance is ‘ultimately’ biodegradable but < 70% and > 20%, it is ‘inherently’ biodegradable.

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