Mycophenolic acid

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

April 5th, 1993 to September 28th, 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study conducted under recognised test methods.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other:

Deviations:

yes

Remarks:

See "Principles of method if other than guideline"

Principles of method if other than guideline:

The study was conducted in compliance with the U.S. Food and Drug Administration (FDA) Good Laboratory Practice regulations (21 CFR, Part 58). Deviations are as follows: a) Protocol Section: 4.5 Test Procedure - Biological: the definitive test concentrations selected for this study (400, 600, 800, 1000, and 1200 mg/L did not follow the 1.5 to 2.0 ratio specified in the protocol. reason: the range selected bracketed the suspected EC50 more closely than a 1.5 to 2.0 ratio series would have. b) Protocol Section: 4.6 Test Procedure - Chemical and physical: The dissolved oxygen in the test solutions was below the 60% limit by the 24 hour observation period. the control dissolved oxygens were at least 845 saturation for the duration of the study. Reason: mycophenolic acid impacted the dissolved oxygen content in the test water. Effect on the study: None. Daphnids are able to suvive and reproduce in water with very low dissolved oxygen content. The specification for 60% saturation is derived from the DO tolerance level for fish.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Mycophenolic acid

EC Number:

246-119-3

EC Name:

Mycophenolic acid

Cas Number:

24280-93-1

Molecular formula:

C17H20O6

IUPAC Name:

6-(4-hydroxy-6-methoxy-7-methyl-3-oxo-1,3-dihydro-2-benzofuran-5-yl)-4-methylhex-4-enoic acid

Details on test material:

- Name of test material (as cited in study report): Mycophenolic Acid
- Substance type: Monoconstituent
- Physical state: brown powder
- Impurities (identity and concentrations): Phthalide 0.2 %; 1-Hydroxy PMPA 0.21 %
- Storage condition of test material: was stored in a refrigerator

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations:0-, 400-, 600-, 800-, 1000-, and 1200-mg/L nominal test concentrations
- Sampling method: samples were taken at 0, 24, and 48 hours during the study to determine the measured concentration of the test substance for each test level
- Sample storage conditions before analysis:

Test solutions

Vehicle:

yes

Details on test solutions:

Hard Blended Dilution Water: 146 mg/L as CaCO3 /total hardness), 166 mg/L as CaCO3 (total alkalinity), 8.2 (pH), 21.1 degree Celsius (temperature) 300 microMhos/cm (conductivity), and 9.1 mg/L (dissolved oxygen)

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Source: ABC Laboratories in-house daphnid culture. The primary culture was obtained from the Columbia National Researsch Laboratory (CNFRL), Columbia, Missouri, in 1977
- Age at study initiation (mean and range, SD): Only neonates (less than 24 hours old) were selected for testing.
- Method of breeding: in a temperature-controlled area at 20-21 degree Celsius.Lighting at 60-65 footcandles on a 16-hours daylight photoperiod. Daphnia were fed every day a suspension of at least one algae species, a supplement of trout chow, active dry yeast
- Feeding during test: the neonates were not fed during the study.

ACCLIMATION
- Acclimation period: since culturing and testing conditions were the same no acclimatation period was necessary
- Acclimation conditions (same as test or not): same as test

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

Does not apply

Test conditions

Hardness:

146 mg/L as CaC03

Test temperature:

21.1 degree Celsius

pH:

8.2

Dissolved oxygen:

9.1 mg/L

Nominal and measured concentrations:

Nominal test concentrations:0-, 400-, 600-, 800-, 1000-, and 1200-mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:250-ml glass beakers
- Type (delete if not applicable): covered with loose-fitting petri dish covers

- No. of organisms per vessel: five
- No. of vessels per concentration (replicates): four
- No. of vessels per control (replicates): four


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: hard blended water
- Total hardness of 130-160 mg/L as CaCO3
- Culture medium different from test medium: no


OTHER TEST CONDITIONS
- Photoperiod: 16-hours daylight photoperiod with 30-minute simulated dawn and dusk periods
- Light intensity: 650 to 664 lux 16-hours daylight photoperiod with 30-minute simulated dawn and dusk periods
- Temperature (waterbath) range from 19 to 21 degree Celsius


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Not applicable


TEST CONCENTRATIONS
Nominal test concentrations:0-, 400-, 600-, 800-, 1000-, and 1200-mg/L

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

EC50

Remarks:

50% immobilization or mortality

Effect conc.:

755 mg/L

Nominal / measured:

meas. (not specified)

Basis for effect:

mobility

Remarks:

behaviour

Effect concentrations 2

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

440 mg/L

Nominal / measured:

meas. (not specified)

Basis for effect:

mobility

Remarks:

behaviour

Details on results:

Based on the mean measured concentrations of mycophenolic acid the 48-hour EC50 and NOEC for Daphnia magna exposed to mycophenolic acid were 755 and 440 mg/L, respectively.

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

Statistical analysis of the concentration vs. effect data (immobility) was conducted by using a computerised EC50 program. This program calculates the EC50 and its 95% confidence limits using the binimial, the moving average, and the probits tests. The program uses different methods of analyzing the data since no one method of analysis is appropriate for all possible sets of data that may be obtained. The method(s) of calculation selected for presentation in this report was that which gave the narrowest confidence limits for the EC50. However, if no immobility occured or if a dose response could not be demonstrated over a reasonable range (<37 to >63%), the EC50 and /or its 95% confidence limits could not be calculated. A 48-hour dose-response slope was calculated by transforming percent immobile to probit values and then calculation via linear regression.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

This study was designed to determine the 48-hour EC50 value and NOEC for Daphnia maga exposed to mycophenolic acid in a static system. The mean measured concentrations for the study were <3.00, 440, 657, 868, 1080, and 1320 mg/L mycophenolic acid. The abnormal effects of daphnid immobility, surfacing, and daphnids tending to the bottom of the vessel were observed in the 657, 868, 1080, and 1320 mg/L test solutions. Based onthe mean measuredconcentrations, the 48-hour EC50 and NOEC were 775 and 440 mh/L mycophenolic acid respectively.

Executive summary:

The primary objective of this test with Daphnia magna was to evaluate the acute toxicity of mycophenolic acid, CAS No. 24280-93-1, to Daphnia magna under static conditions. The test was designed to yield EC₅₀values following 24 and 48 hours of exposure.

Samples were. taken from each test solution at 0, 24, and 48 hours of the definitive test to determine the measured concentration for each test level. The test concentrations were determined by using high-performance liquid chromatography. The measured concentrations were determined to be 440, 657, 868, 1080, and 1320 mg/L for the nominal concentrtions of 400, 600, 800, 1000, and 1200 mg/L, respectively.

 

Based on the mean measured concentrations of mycophenolic acid the 48-hour EC,, and NOEC forDaphnia magnaexposed to mycophenolic acid were 755 and 440 mg/L, respectively.