N.J. Admin. Code § 7:18-7.6 - Calculating, analyzing, and reporting of quantal test results

(a) A toxicity test is invalid if any of the conditions listed in (a)1 through 5 below occurs. When a toxicity test is invalidated, the laboratory shall clearly mark the test results accordingly. The laboratory shall submit the results to the Department, along with a written explanation as to the reason for the invalidation and the expected date that the test will be repeated. If a toxicity test is invalidated, the laboratory shall repeat the test as soon as possible within the monitoring period specified in the permit. A toxicity test is invalid if:

1. In an N.M.A.T. or N.O.A.E.C. test, mortality of the control test organisms exceeds 10 percent;

2. In tests determining either an EC[50] or an LC[50], greater than 10 percent of the control test organisms either show the effect or exhibit mortality;

3. In definitive tests determining an LC[50] or an EC[50], after pooling replicate test chamber responses two or more test concentrations deviate significantly from the expected trend of increasing effluent concentrations exhibiting increasing levels of toxicity. Deviation in response from the expected trend of greater than 10 percent (mortality for an LC[50], or effect for an EC[50]) shall be considered to be significant;

4. The 95 percent confidence limits cannot be calculated for a test which results in an LC[50] or EC[50] that is either less than the highest effluent concentration tested, or greater than the lowest effluent concentration tested. However, the test need not be invalidated if the calculated LC[50] or EC[50] lower confidence limit is ≥30 percent of the highest effluent concentration tested and the upper confidence limit is a positive infinity; or

5. The laboratory is unable to calculate 95 percent confidence limits when required under (b)4ii below.

(b) A laboratory shall satisfy the requirements of (b)1 through 6 below when calculating toxicity test results.

1. The laboratory shall not use biological test results from N.M.A.T. definitive acute toxicity tests to calculate an LC[50] or EC[50]. For all effluent concentrations and the control, the laboratory shall report the number and percentage of dead test organisms or those showing the effect.

2. The laboratory shall express the biological test results from range-finding acute toxicity tests in terms of an LC[50] or EC[50] whenever the data support the estimation. The laboratory shall estimate the LC[50] or EC[50] by the graphical method described in EPA Acute Methods #027F-1993.

i. When the highest effluent concentration kills or affects less than 50 percent of the test organisms, the laboratory shall record the number of organisms showing the response in each treatment; and

ii. When the lowest concentration kills or affects more than 80 percent of the test organisms, the laboratory shall repeat the range-finding test with a lower set of effluent concentrations, before conducting the definitive toxicity test.

3. The laboratory shall express the biological test results from definitive acute toxicity tests in terms of an estimated LC[50] or EC[50], except when no toxicity is observed. The laboratory shall report the numbers and percentages of test organisms dying (for LC[50]) or showing the effect (for EC[50]) for each test chamber.

i. When more than 50 percent of the test organisms die (for LC[50]) or show the effect (for EC[50]) in the lowest effluent concentration, the laboratory shall report an LC[50] or EC[50] of less than the lowest concentration;

ii. When less than 50 percent of the test organisms die (for LC[50]) or show the effect (for EC[50]) in the highest effluent concentration, the laboratory shall report an LC[50] or EC[50] of greater than the highest concentration;

4. When the results of a definitive acute toxicity test falls between the two extremes given in (b)3i and ii above, the laboratory shall calculate LC[50] or EC[50] values using the following methods and procedures.

i. The laboratory shall calculate LC[50] or EC[50] for at least the final exposure time and each 24 hour exposure time between the beginning and the end of the test. For example, in a test with a 96-hour duration, the laboratory would calculate LC[50] or EC[50] 24, 48, 72 and 96 hours after the beginning of the test;

ii. The laboratory shall calculate 95 percent confidence limits for each definitive acute toxicity test LC[50] or EC[50]. If the laboratory cannot do so, the test shall be invalid in accordance with (a)5 above. This requirement does not apply to any toxicity test that results in an LC[50] or EC[50] that has:

(1) Greater than or equal to the highest effluent concentration tested; or

(2) Less than or equal to the lowest effluent concentration tested; or

(3) No partial mortalities.

iii. If any acute toxicity test results in no partial mortalities, the LC[50] or EC[50] is determined using the Graphical Method as described in EPA Acute Methods #027F-1993;

iv. If any acute toxicity test results in two or more partial mortalities and a non-significant Chi Square Test, the LC[50] or EC[50] and corresponding 95% confidence intervals are determined using the Probit Method as described in EPA Acute Methods #027F-1993.

v. The Spearman-Karber Method, as described in EPA Acute Methods #027F-1993, is used to determine the LC[50] or EC[50] and corresponding 95 percent confidence intervals for any acute toxicity test if the following conditions apply:

(1) Zero mortality in the lowest effluent concentration and 100 percent mortality in the highest effluent concentration, and;

(2) One partial mortality, or;

(3) Two or more partial mortalities and a significant Chi Square Test;

vi. If the conditions in (b)4iii, iv, or v do not apply, the LC[50] or EC[50] and corresponding 95 percent confidence intervals for any acute toxicity test are determined using the Trimmed Spearman-Karber Method as described in EPA Acute Methods #027F-1993.

5. Upon completing a definitive acute toxicity test for which LC[50 ]or EC[50] can be calculated, the laboratory shall plot a toxicity curve. The laboratory shall plot the curve using the LC[50] or EC[50] for each observation time, following the methodology presented in "Standard Methods, 16th Edition" p. 717. The laboratory shall report any threshold or incipient LC[50] that it can estimate from the curve. If the laboratory finds no threshold or incipient LC[50], the laboratory shall report this fact. In either case the laboratory shall include the toxicity curve graph in the report of results whenever the data permit it to be plotted.

6. For all effluent concentrations and the control, the laboratory shall determine the N.O.A.E.C. using the hypothesis testing as described in section 11.3 of EPA Acute Methods #027F-1993.

(c) The laboratory shall follow the following requirements when reporting results of acute toxicity tests:

1. The laboratory shall complete a sample report immediately after collecting either dilution water or effluent samples. The laboratory shall make the report either as a separate form or as an entry in a bound logbook. The laboratory shall include the sample report data in its report of the toxicity test results to the Department. The laboratory shall include the following information in the sample report:

i. The sampling location;

ii. Date and time of collection;

iii. Total residual chlorine level in the sample;

iv. The type of sample (composite or grab);

v. The material sampled;

vi. The collector's name; and

vii. Any pertinent comments.

2. The laboratory shall include the following types of information when reporting results of an acute toxicity test:

i. The name of the test, the investigator(s), and the laboratory;

ii. The date on which the test began;

iii. The name of NJPDES permittee, its location, its NJPDES permit number, and the Discharge Serial Number;

iv. The name of the receiving water body;

v. A detailed description of the effluent, including the sampling point, date and time of collection, known physical and chemical properties, and variability;

vi. A detailed description of the dilution water source, the sample location, the date and time of sample collection, the tide stage (if applicable), the dilution water chemical characteristics, and any pretreatment;

vii. A description of acute toxicity test method(s) used, including:

(1) The test protocol, a definition of the adverse effect (death, immobility, etc.) used in the test, a description of the test chambers used (including the depth and volume of test solution), the number of test organisms per replicate treatment, the number of replicate treatments, and the loading rate of the test organisms. In flow-through toxicity tests the report shall also include the number of water volume exchanges per 24 hours in each test chamber;

(2) Detailed information about the test organisms, including the scientific name, mean and ranges of length and weight (where appropriate), age, life stage, source, previous history (if known), observed disease, treatments (if any), record of acclimation procedure, and observations on behavior during the test;

(3) A description of any aeration or salinity adjustments performed on test solutions before or during the test;

(4) The methods used for all chemical analyses;

(5) The mean and range of the acclimation temperature, test temperature, and salinity; and

(6) Any deviations from the test method(s);

viii. The test results, including the following:

(1) The results of all chemical analyses of the effluent and dilution water;

(2) A daily record of the number and percentages of organisms in each test chamber, including the control treatments, that died or showed the selected effect;

(3) A summary of general observations of other effects of symptoms of toxicity observed during the test;

(4) The LC[50] or EC[50] value with specified exposure time and the 95 percent confidence limits. If the highest effluent concentration did not kill or affect 50 percent or more of the test organisms, report the LC[50] or EC[50] as greater than the highest concentration and the percentage of test organisms killed or affected in each experimental treatment. If the lowest effluent concentration killed or affected more than 50 percent of the test organisms, report the LC[50] or EC[50] as less than the lowest concentration and the percentage of test organisms killed or affected in each experimental treatment. Report the method(s) used to calculate the LC[50] or EC[50], and its 95 percent confidence limits. Include a graph of the toxicity curve;

(5) Any deviations from approved methodology, along with a summary of the reason(s) for the deviation(s); and

(6) Any other relevant information; and

ix. Upon completing the toxicity test, the laboratory shall send the original or true duplicate of the results to the client. The original or true duplicate shall be signed by the laboratory manager or a designee identified under N.J.A.C. 7:18-2.11(a)1 iii.

Notes

N.J. Admin. Code § 7:18-7.6

Administrative change.
See: 28 New Jersey Register 4098(a).
Amended by R.1997 d.192, effective 5/19/1997.
See: 28 New Jersey Register 4149(a), 29 New Jersey Register 2275(a).
In (a)1, inserted "or N.O.A.E.C."; and added (b)6.