Degradability in the environment

＊Biodegradability (Law Concerning the Examination and Regulation of Manufacture, etc. of Chemical Substances, and OECD TG301C) Biodegradability tests stipulated by the OECD test guideline consist of OECD TG301A through TG301F, and TG302A through TG302C. The major test procedure in Japan is 301C, which is stipulated by the Law Concerning the Examination and Regulation of Manufacture, etc. of Chemical Substances.
After 100mg/L of a chemical substance is exposed to 30mg/L of activated sludge at 25℃ for 28 days, biodegradability is measured. If a BOD degradation rate is 60 percent or higher, and no decomposition products are observed by direct determination, the Japanese law designates the chemical substance as a readily degradable substance.

Bioconcentration test

＊Bioconcentration test using fish (Law Concerning the Examination and Regulation of Manufacture, etc. of Chemical Substances) Test fish are exposed to fixed concentrations (typically, two concentrations, or not less than one hundred times lower than LC50, and as low as possible) of a test chemical substance in water under flow-through conditions for 28 days. A concentration rate is obtained by calculating a ratio of the concentration of the test substance in water to that in the test fish.

The Japanese law designates a bioconcentration factor of lower than 1,000 as not high bioconcentrative, and a factor of 5,000 or higher as high bioconcentrative.

＊n-Octanol / water distribution factor (Law Concerning the Examination and Regulation of Manufacture, etc. of Chemical Substances, and OECD TG107 and TG117) Two types of test methods, the shake-flask method (TG107) and HPLC (liquid chromatography) (TG117), are stipulated.

If an n-octanol / water partition coefficient (logPow) is lower than 3.5, the chemical is designated as not high concentrative, and in this case, the test is accepted as a substitute for the bioconcentration test with fish. If the value is 3.5 or higher, it is necessary to perform a bioaccumulation test with fish.

Eco-toxicity Tests

Recently, a high value has been put on effects of emission chemical substances on the natural environment. Although until now, the fish acute toxicity test has been mainly performed to study such effects, a recent trend is a comprehensive assessment using results of three tests: the fish acute toxicity test, algal growth inhibition test and daphnids acute immobilization test. The Law Concerning the Examination and Regulation of Manufacture, etc. of Chemical Substance Control Law (Kashinho), requires performance of these three tests for the study of environmental impact. A GHS advisory of the UN, which was adopted in July of 2007, classifies hazardous properties of aquatic life according to results of the three tests.

＊Algal Growth Inhibition Test (OECD TG201)

A test alga (Pseudokirchneriella subcapitata), a producer in the aquatic food chain, is exposed to a chemical substance for 72 hours. The number of cells is measured 24, 48 and 72 hours after the beginning of the exposure. EC50 (effect concentration50) and NOEC (no observation effect concentration) are calculated to check for any impact on algal growth.

＊Daphnids Acute Immobilization Test (OECD TG202)

Daphnia magna, a primary consumer on the aquatic food chain, is exposed to a chemical substance for 48 hours. To check for any impact on the mobility of Daphnia magna, the number of immobile organisms or deaths are observed 24 and 48 hours after the beginning of the exposure to calculate the effect concentration 50 (EC50).

The test can be a preliminary test for Daphnia magna reproduction test (OECD TG211).

＊Fish Acute Toxicity Test (OECD TG203)

A test species of fish, a high-order consumer in the aquatic food chain, is exposed to a test chemical substance for 96 hours. The number of dead fish is observed to calculate the lethal concentration 50 (LC50).
Although general test species are killfish and carp in Japan, bluegill, rainbow trout and guppy are common outside Japan.

In all three tests, Chemical Substance Control Law (Kashinho) , sets the upper limit to 100mg/L while the GHS advisory doesn't require classification of hazardous properties for LC50 or EC50 of higher than 100mg/L.