ACUTE TOXICITY OF PYRITHIONE ACUTE TOXICITY OF PYRITHIONE PHOTODEGRADATION PRODUCTS PHOTODEGRADATION PRODUCTS

TO SOME MARINE ORGANISMSTO SOME MARINE ORGANISMS

T OndukaT Onduka, K Mochida, K Ito, A Kakuno and K Fujii, , K Mochida, K Ito, A Kakuno and K Fujii, National Research Institute of Fisheries and Environment of InlaNational Research Institute of Fisheries and Environment of Inland nd

Sea, Fisheries Research Agency, Japan Sea, Fisheries Research Agency, Japan H Harino, Osaka City Institute of Public Health and EnvironmentaH Harino, Osaka City Institute of Public Health and Environmental l

Sciences, JapanSciences, Japan

International Symposium ON SHIPBUILDING TECHNOLOGY (ISST 2007)

- Fabrication and Coatings –6-7 September 2007

Osaka University, Japan

ContentsContentsIntroduction Introduction Antifouling paint biocide Antifouling paint biocide inin JapanJapanCharacteristic of Characteristic of ppyrithiones (PTs)yrithiones (PTs)

ExperimentalExperimentalTest chemicals, organisms, and methodsTest chemicals, organisms, and methods

Result and DiscussionResult and DiscussionAcute toxicity of PTs and photodegradation productsAcute toxicity of PTs and photodegradation productsComparison of toxicity of PTs Comparison of toxicity of PTs

and tributyltin (TBT) in previous studyand tributyltin (TBT) in previous study

ConclusionConclusionSummary and future taskSummary and future task

AntifouAntifoulling ing paintpaint biocidebiocideOrganotin compounds (OTs) such as TBT have Organotin compounds (OTs) such as TBT have been used as antifouling biocides been used as antifouling biocides

The toxicity of OTs have led to a worldwide ban The toxicity of OTs have led to a worldwide ban by the International Maritime Organizationby the International Maritime Organization

Candidate marine antifouling compounds Candidate marine antifouling compounds developed as alternatives to OTs developed as alternatives to OTs

→→Irgarol 1051, diuron, SeaIrgarol 1051, diuron, Sea--Nine 211, Nine 211, zinc pyrithione (ZnPT)zinc pyrithione (ZnPT), , copper pyrithione (CuPT)copper pyrithione (CuPT)

AAntifountifoulling ing system in Japansystem in Japan

Frequency of Frequency of using PTsusing PTs

: : 61%61%

Diuron

Pyridine-triphenylborane

ZnPT

CuPT

Dicopperoxide

(%)The frequency of using antifouling biocide in Japan

Japan Paint Manufactures‘Association (2006)

0 20 40 60 80

PTs are very PTs are very frequently used as frequently used as antifouling booster antifouling booster

biocidesbiocides

Characteristic of PyrithionesCharacteristic of PyrithionesPTs have toxic effects on fresh water organismsPTs have toxic effects on fresh water organisms

Median lethal concentration (Median lethal concentration (LLC50)C50), , Median effective concentration (Median effective concentration (EC50)EC50)

:: 0.440.44––462< 462< µµgg LL--11

PTs degrade rapidly in the water column PTs degrade rapidly in the water column Photolytic halfPhotolytic half--lives: lives: 7.17.1––29 minutes29 minutesHHydrolytic halfydrolytic half--lives: lives: 12.912.9––990<0< daydayss

→→ PTs PTs areare more stable under dark conditionmore stable under dark condition

The purpose of this study was to elucidate The purpose of this study was to elucidate the effect of the degradation products of PTsthe effect of the degradation products of PTs on on

marine organismsmarine organisms

Test organismsTest organismsIn the functioning of ecosystem, In the functioning of ecosystem,

trophic relations are of prime importance trophic relations are of prime importance

Three marine organisms, Three marine organisms, representing three trophic levels representing three trophic levels

Alga: Skeletonema costatum

Crustacea: Tigriopus japonicus(nauplii)

Fish: Pagrus major(weight: 0.2-0.3g))

Test ChemicalsTest Chemicals

N SS N

N

SO3H

NO

N SS N

O O

N

SH

N

SH

O

Pyridine-2-sulfonic-acid

(PSA)

2-mercaptopyridine-N-Oxide(POS)

2,2‘-dithio-bis-pyridine-N-Oxide

((POS)2)

Pyridine-N-Oxide(PO)

2-mercaptopyridine(PS)

2,2‘-dithio-bis-pyridine(DPS）

NO

Cu

SN

O

SN

O

Zn

SN

O

S

ZnPT CuPT

AlgaAlgaｌｌ growth inhibition testgrowth inhibition test

・ 30ml f/2 medium・ Initial cells concentration: 104cells mL-1

・ Temperature 20℃・ Light source: three ultraviolet screening

fluorescent tubes・ 40-80 µmol m-2 sec-1, 14hL:10hD ・ In vivo fluorescence of the alga

directly with a fluorescence meter

・72-h EC50 value was estimated by probit analysisrate of growth inhibition, nominal concentration

Recommended in the test guidelines of the OECD 201

0h 24h 48h 72h

Monitor in vivo fluorescence

CuPT: mostly stableZnPT:100µg/L→7µg/L

Copepod immobilisation testCopepod immobilisation test

・ 12 wells culture plate・ Filtered seawater・ Five nauplii / well・ 4 wells / concentration・ No feed・ Test period: 24 hours・ Under the dark condition・ Temperature 20℃

・ Inability to swim within 15 s after gentle agitation→ Immobilisation・ 24-h EC50 value was calculated by probit analysis

immobilisation rate, nominal concentration

Recommended in the test guidelines of the OECD 202

Fish acute toxicity testFish acute toxicity testRecommended in the test guidelines of the OECD 203

・ 20L glass aquaria・ 20 fishes / concentration・ No feed, Under the dark condition・ Temperature: 23.3-24 ℃・ Dissolved oxygen: 5.7-5.9 mg L-1

・ pH: 8.1

・ 96-h LC50 value was calculated by trimmed Spearman-Karber technique

fish mortality, nominal or actual concentration

0h 24h 48h 72h 96h

Change test solution, Monitor mortality

Acute toxicity of pyrithionesAcute toxicity of pyrithiones

Alga Crustacea FishEC50

CuPT 1.5 23 9.3*ZnPT 1.6 280 98.2*

EC50 LC50

* Reported by Mochida et al. (2006); the 96-h LC50 was calculated from the actual concentration

Unit : μg L-1

TBT 0.33-1.28 0.6-5.7 3.2-25.9Previous study

・ Alga ： most sensitive・ Toxicity : CuPT ＞ ZnPT to the crustacea and the fish

Discussion of acute toxicity of pyrithiones Discussion of acute toxicity of pyrithiones

Toxicity Alga: CuPT = ZnPTCrustacea and fish : CuPT > ZnPT

ZnPT reacts with the Cu2+ in seawater, and convert to CuPT

f/2 medium: +1.8µg L-1 (7.1nM) of Cu2+

in seawaterZnPT the algal test range: 0.5 – 2.0µg L-1

(1.6-6.5nM) →there was enough Cu2+

to convert ZnPT to CuPT

Acute toxicity of Acute toxicity of the photodegradation products of pyrithionesthe photodegradation products of pyrithiones

Alga Crustacea FishEC50

POS 1.1 >12 500 ―

(POS)2 3.4 >1 250 ―

EC50 LC50

PO >100 000 >100 000 >100 000

PSA >100 000 >100 000 >100 000

DPS 65 550 520

PS 730 76 000 45 000

Unit : μg L-1

Comparison of acute toxicity of Comparison of acute toxicity of photodegradation products and pyrithionesphotodegradation products and pyrithiones

Alga Crustacea FishEC50

POS 1.1 >12 500 ―

(POS)2 3.4 >1 250 ―

EC50 LC50

DPS 65 550 520PS 730 76 000 45 000

Unit : μg L-1

CuPT 1.5 23 9.3*ZnPT 1.6 280 98.2*

Pyrithiones

Kinetics of photodegradation Kinetics of photodegradation products of pyrithionesproducts of pyrithiones

OH－ O

O

O

OH-

H＋

hνRedox

Mn+

nMn+

(Seymour and Bailey, 1981)

(POS)2

POS

ZnPT、CuPT

N

S

O

N

SO3H

O

NSSN

O O

N

SH

O

N

S

O

Test chemicals transfer different products

Analytical methods of PTs transfer products in seawater are developing

POS and (POS）2 can transform to pyrithiones

Summary and future taskSummary and future task

PTs have toxic effects on marine organismsPTs have toxic effects on marine organisms

The toxic influence of PTs and some of their The toxic influence of PTs and some of their photodegradation products on algaphotodegradation products on algaee

Future taskFuture taskThe fate of PTs in seawater The fate of PTs in seawater

has to be elucidatedhas to be elucidated

ThThee environmental PTs and photodegradation environmental PTs and photodegradation products risk assessments are neededproducts risk assessments are needed