Phuket Marine Biological Center Special Publication no. 19(1): 169-172 (1999) 169

EFFECTS OF EXPOSURE TO WATER-SOLUBLE FRACTIONS OFLUBRICANT OIL (MESRAN@ SAE 40 W) ON ATTACHMENT FORCE

OF THE SNAIL LITTORARIA SCABRA(LITTORINIDAE : GASTROPODA)

Markus T. Lasut & James PaulusSub-Laboratory of Toxicology & Marine Pharmaceutics, Laboratory of Marine Sciences

Faculty of Fisheries & Marine Sciences, Sam Ratulangi UniuersityJl. Kampus (Jnsrat Bahu, Manado 95115,ln'donesia

ABSTRACTAttachment force in gastropods has5 been used to measure responses of marine snailsLittoraria sgabra (shell length of 78-24 mm) to water-soluble fractions (WSF) of lubricantoil (Mesran@ SAE 40 W) in different concentrations. The ability to adhere to the substratumwas ineasured in two ways: 1) the weight required to lift up an animal when it attached to

a substratum (termed tenacity). 2) the percentage of animals falling down from the wall ofthe experimental chamber (termed endurance). Higher concentrations of WSF and longerexposure times reduced the attachment force, both with respect to tenacity and endurance.This was related to the production of mucus as the first self-defence response ofZ. scabra

to the oil. Production of mucus was the sublethal response, which required enerry thatfinally drained the resources available for attachment to the substrate. Statistical tests

showed that the effect of WSF on tenacity and endurance, and their interaction, weresignifrcant (p<0.05) in all experiments.

INTRODUCTIONOiI is important for various refinedpetroleum product. InL992, the Institute ofPetroleum UK calculated that the crude oilentering a refinery is converted primarilyto fuel (c.907o) while ca. 17o is converted tolubricant oils (Betton 1994). Petroleum andits products enter the environment fromseveral man-made sources: offshoreproduction, transportation, municipal andindustrial wastes. It is of global concern thatthe marine environment is vulnerable to oilpollution.

Numerous reports are available, whichdescribe the effects of oil pollution on marineenvironments and their organisms (Ahmad1988, Nybakken 1993). However, little isknown concerning of effects of lubricant oilsincluding their water-soluble fractions(WSF) on marine animals (Saezet al. L992).

The water-soluble fractions may comprise

B0-987o of the lubricants in use today (Betton1994). Saez et al. (1992) showed the negativeeffects of oil-derived hydrocarbons on thedigestive gland of the snail Littorina littorea.

The ability to attach to a substrate isessential for intertidal gastropods in relationto their activities (Imrie 1992) Attachmentcould be used as a way to asses effects ofenvironmental pollution. We haveinvestigated this possibility by studying theattachment capability of the snail Zittorariascabra exposed to water-soluble fractions oflubricant oil. The ability to adhere to thesubstratum was measured in two ways: 1)

the tenacity measured as the weightrequired to lift up an animal attached to asubstratum. 2) endurance measured as thepercentage of animals falling down from thewall of the experimental chamber.

170 Tlopical Marine Mollusc Programme QMMP)

MAIERIALS AND METHODS

L. scabrawas collected at Tongkaina, NorthSulawesi, Indonesia. The snails were stored

in the laboratory fot a 2 days adaptationperiod before the experiment. All sea waterused was taken from the site where thespecimens were collected. The salinity and

temperature were 34 %o and 23-26 'Crespectively. @

A lubricant for car engines (Mesran-

SAE 40 W, produced bY PERTAMINA'Indonesia) was used as the test compound'

The oil had been used in a car for up to 2000

km. The oil was mixed with sea water in achamber and left exposed to air for at least30 days to release the water-soluble fraction'The WSF was separated from the water-insoluble fractions (WIF), which always formthe upper layer of the solution. The WSF

concentration was analYsed on a

spectrophotometer (wave length of 450 A)

using t,t,t-Tbichloroethane as a standard'Three concentrations (0'8' 1.6 and 2'4 ppm')

of WSF were prepared by dilution with sea

water.Chambers without lid containing 15

snails (shell length 18-23 mm) were exposed

for 0, 4 and 8 hours to the threeconcentrations of WSF and a control' Bothconcentration and duration ofthe exposure

were variables of the test. After exposure to

WSF, each animal was rinsed in clean sea

water for 2to 3 minutes before being placed

under a modifred wooden tool with glass as

a substratum.Tenacity was measured in terms of the

pull required to detach the animal from the

substralum. A cotton line was attached to

the snail. A small plastic chamber was fixedat the opposite end of the line running over

a block. Water was added to the chamber

until the animal detached. The volume ofwater was measured (1 mI = 1 g)' The

measurements were done in 1-0 replicates'Endurance was measured as the

percentage of snails, which fell down frgm a

chamber wall when they crawled up' Four

plastic chambers without lid (15 x 17 cm)

contained 0.8, 1.6, 2.4 PPm WSF and acontrol. Atotal of 60 snails (shell length 18-

24 mm) were exposed to WSF fot t44 hours'The number of animals falling down fromthe chamber wall was counted evety 24

hours. The measurements were done in 4replicates.

The tenacity was analyzed by means of

two-way ANOVA (Analyses of Variance)while one-way ANOVA was aPPlied inmeasurements of the endurance. A Ttrkey-

test was applied in both measurements(Sokal & Rohlf 1981; Fowler & Cohen 1990)'

RESULTS

Extraordinary amounts of mucus weresecreted by L. scabro exposed to WSF,

indicating a stressed environment compared

with the control. Further, the tenacity ofsnails decreased as the concentrations and

the duration of exposure increased Gig' 1)'

conttol 0,8 1,5 2'4

Concennaion ofWSF ofoil (PPm)

Figure 1. Tenacity of individual L' scobr6-exposed-to

WbF prepared from lubricant oil (Mesran* SAE 40 W)'

The tenacity is expressed in terms of the weight (g) tsd needed to pull the snails offthe substratum after 0,

4, and 8 hrs -of

e*posrrre to WSF at concentrations of

0.8 to 2.4 ppm. The control was clean sea water'

n oto*

I 4ho*t

I gtto*

The effect was statistically significant lubricant had been used in a car engine for

(p<0.05). Interactiorr"r u"lr, conclerrttation up to 2000 km prior to the test on the snails'

and duration was also significantly (P<0.05). However' the composition of the oil was not

ftt" "ffu"t of the WS'F was significant analysed'

(P<0.05)ataconcentrationof0.8ppmandTheWsFwasprobablytakenupbythe4hours of exposure.t/r'1urw'vr

v'v vvu' F---- snails via the gills in connection withrespitation (McMahon L992)' Nelson-Smith

Og72) iru Laws (1993) suggested that the

Phuhet Marine Biotogical center special Publication no' 79(1): 169-172 (1999) 771

looft-'-r-1-+\\-.\ toxic hydrocarbons apparently exert their----. ;i;;;"p.rtbybecomingincorporatedinto

^80s -.-o ppm iftu futtv iuv"t ihtt makesup the interior of

-,-;;;, cell membranes.' As a result' the1Tli,ll:ts l\\ -\ -'_::::: irarropt"aandceasestoproperlyregulateF *l \t^..- \.- ;;:;: tr'" """i'"ge

of substances between the

I I \ \ -\ ' ini""iot utta exterior of the cell''zg I \ tt -:r\

Hvdrocarbons also interact with proteins in

F *t \ \^ -.--....-.-. a ,rariety of plants and animals. Both

3 I \r '\ enzymes and structural proteins upp'"?: t?

E ," I "\- \^--.- be affected. It is jenera'y ug"""d thut--\*--^.-^ aromatic hydrocarbons are the most toxic

| \ iotto*"abycycloalkanes' olefins' andlastly^Ikanes (Laws 1993)''o zo 48 72 90 'l2o '144 a'

The first responses we observed in L'TIME (HouR)

scabra was a high production of mucus

during exposure to ttre WSF Production of

Figure 2. Endurance or L. scabra6::qT*::^HF *rr.oi drains energ'y of the snail (Davies ei

orEou."a from lubricant oil (Mesranw SAE 40 w). Tle ot. tggZl. The decreasing ability to stick toil;;; tt expressed as the percentage of snails' ifr"-r"frir"t"ttt (tft" teriacity test) or stay*hi.h *"r" ableio stay attached to the walls of tan

containing from 08 to 2'4 ppm wsF The "o"ttol

*ul attached to the wall (the endurance test)

clean sea water. The ""a"It"u""t

was estimated every may be the visible signs of a drain of energy'

24 hrs over a period of 144 hrs ioi"rr"r, no relevant comparative studies

rhe endurance decreased over time in all :,il:"lr"Jili?:X1.:ltt}i'r.u'" unable to

concentrations,includingthecontrol(Fig'-z) aLJ" .ffl*s of oil are not observed below

Ho*"-n"" aft,er 144 hrs a total of 87Vo of the ;;;;;;;ions of 10 ppb. However, Gerlach

tttuitt were still attached to the wall in the ;i;;il;;ested thai i pg/'of water-soluble

.o"t"of while onlv 67o marra$ed to do the ;;i;"-;";;;ts in water can harm sensitive

same in the highest concentration of 2'4 ppm ;;;;;i*". as fish larvae hatching from fishWSF The effect of the WSF on the endurance ;; ; healthy. EPA (1976) inLaws (1993)

of the snails was significant from a ";i""""a a reiuction in the chemotactic

concentration of 1.6 ppm (P<0.05). p""."ption of food by the snail Nassarius

DrscussroN * ,n:lfutiT6tTltffii6i:lJi':The toxic effects of oil are believed to be

"ilydrocarbons.on gastropods was 1-100 mg/I

caused by tow-molec;i;r:;;igh Qp-CzD (Betton 1994)' Grazing mollusc are more

compounds, which are soluble in *uiJ" sensitive to oil than filter-feeding mollusc'

(Laws 1gg3). r aru Dvruv

which can separate oil from food particles

The WSF in the present study may have (Cormack 1983)'

n""t tff""ted bV heating because the

t72 Tlopical Marine Mollusc Programme QMMP)

ACKNOWLEDGEMENTSWe are much indebted to the TropicalMarine Mollusc Programme (TMMP)sponsored. by DANIDA for the opportunityto present ttti. paper in the- Ni.nlnConference/ Workshop of TMMP in Lombok,

Indonesia. We wish to thank Ms' NoraTicoalu, SPi. for data collection'

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