Journal of Applied Sciences & Environmental Management, Vol. 5, No.
1, June, 2001, pp. 93-96
Aqua-toxicological Effects of Water Soluble Fractions
(WSF) Of Diesel Fuel On O. Niloticus Fingerlings
*DEDE, E.B. KAGLO, H.D.
Department of Pharmacology and Toxicology, College of Health Sciences, University
of Port Harcourt, Port Harcourt, Nigeria.
*Corresponding author
Code Number: ja01016
ABSTRACT
Five concentrations of water
soluble fraction (WSF) of diesel fuel (1.6 ppm, 3.2ppm, 6.4ppm, 12.3ppm and
19.2ppm) were made. To each of the concentrations of the WSF, ten fingerlings
of O. niloticus with an average weight of 3g were incubated for 96hours. The
96hour LC50 was determined (8.08ppm). Histopathological examination
of the fish gills following exposure to the LC50 and LC100 concentrations
of the WSF showed gill elongation and lamellar hyperplasia respectively. There
was reduction of dissolved oxygen content (from 7.2 to 4.5 mg/l) caused by
diesel that dissolved in water. This was below acceptable levels for the
sustenance of aquatic life (i.e. 6.0 8.0 mg/L). However, the pH reading
was not adversely affected. The result tended to suggest that the death
of the fingerlings might be related to the decreased dissolved oxygen content
of the water due to the presence of diesel. The structural changes of the
gills observed may be an adaptation by the fingerlings to oxygen stress.
@JASEM
A recent study on the contaminating
effect of petroleum hydrocarbons on fish species showed that minnows and
mullets were found to accumulated aromatic hydrocarbons to as much as 3 -
4mg/gm wet weight of the fish. The mullet absorbed
the hydrocarbons slowly and depurated them more readily, but the minnows
absorbed them rapidly and retained the compounds for as much as two weeks
after being transferred to uncontaminated sea water (Khan et al 1995). The
current study highlights the possibility that some fishes 9such as minnows)
can store aromatic petroleum hydrocarbons from contaminated waters and pass
them on to higher trophic levels.
Furthermore, two years after the release of 600,000 litres of diesel fuel
into Arthur Harbour in the Antarctic peninsula, spill related contamination
was still detected in the intertidal limpet Nacella concinna (Kennicutt
and Sweet, 1992), and the fish, Fundulus heteroclitus collected from
the Wild Harbour marsh, where a major diesel fuel spill occurred in 1969,
was found to contain up to 75ppm of petroleum hydrocarbon in its tissues
several year later (Sabo & Stegeman, 1977)
Contamination of water bodies by hydrocarbons has been shown to produce
subtle changes in fish that are both chronically or briefly exposed (Sabo & Stegeman,
1977) The toxic effects on fish depends on the amount of oil spilled, the
area covered by the oil and the chemical characteristics of the oil (Kiihnhold,
1980); the fish species and the duration of influence of the oil one the
fish are also important (Onuoha & Nwachukwe, 1990).
The current study is aimed at
investigating the acute toxicological effect of the water soluble fraction
of diesel fuel on tilapia (Oreochromis niloticus) fingerlings. It
seeks to provide some needed toxicological data on the diesel fuel that is
in commercial use in Nigeria, especially on a common tropical fish, O.niloticus; This
will include examination of the effect of changes in physicochemical parameters
of the water, ie. pH and dissolved oxygen content, habitant on the viability
of the fish.
MATERIALS AND METHODS
Eighty fingerlings of tilapia
(Oreochromis niloticus) were obtained from the African Regional Aquaculture
Centre (ARAC), Aluu Port Harcourt Nigeria the fingerlings had an average
weight of 3g. They were conditioned to depended on natural foods for about
14 days, this was intended to simulate conditions obtainable in the natural
habitat.
PREPARATION OF WATER SOLUBLE
FRACTION (WSF)
One part (IL) of fresh diesel fuel obtained from a filling
station, was diluted with four parts (4L) of the water with which the fingerlings
were cultured (the control water), in a 6L flask in accordance with Baden
(1982). The diesel water mixture was stirred slowly for 24 hours with a
Gallenkamp magnetic stirrer. This was to enhance the dissolution in the
water of the water-soluble components of the fuel. The mixture was then
made to stand for 3 hours before it was poured into separating funnels and
allowed to stand overnight, so as to obtain a clear oil-water interphase. The
lower layer of water, containing the WSF of diesel was decanted into containers
(Afolabi et al, 1985). This process was repeated several times until sufficient
quantity of the WSF was obtained to carry out the study.
Exposure of TEST ORGANISMS
The WSF was made into five concentrations;
1.6 ppm, 3.2ppm, 6.4ppm, 12.8ppm and 19.2ppm. The dilutions were made with
the control water (habitat0 with which the fingerlings were cultured at ARAC;
Aluu, near Port Harcourt, Rivers State. Ten fingerlings per group were exposed
to 5 litre each of the five concentrations levels of the water soluble fraction
(WSF) plus the habitat water as control. The fingerlings were observed for
96 hours. Fingerlings were confirmed dead when they no longer responded to
prodding (floating in water) Once death was noticed in each of the five concentrations
of the WSF, the dead fish was removed. The gill of a dead fingerling from
the lowest concentration that killed all the fishes (LG100) was taken out. A
representative of the control group was killed and its gill taken out. These
gills were preserved in formalin.
LC50 DETERMINATION
The number of dead fishes per
group were recorded against the time of their death in a tabular form as
specified by Sprague (1972). The data obtained was used to calculate the
median lethal concentration (LC50) of the WST of the fuel on O.niloticus fingerlings
using Arithmeke method of Karber and Dede (1997). A concentration of the
determined LC50 value Igbigke was made by diluting the stock concentration
of the WSF, and ten fingerlings were exposed to it; the gill of a dead fingerling
from this group was taken out and preserved in formalin.
HISTOPATHOLOGY OF GILL
The preserved gill from the control,
LC50 and LC100 groups were processed, embedded in paraffin
wax and sectioned with a Shandon AS 325 rotary microtome, and slides were
prepared from the sections. Photomicrographs of the sections were made with
a Leitz Camera microscope (Diahu 20 model)
PHYSICOCHEMICAL PARAMETERS
The pH and dissolved oxygen content
of the various WSF concentrations and the control water were the physicochemical
parameters measured in this study.
The pH was measured with a La
Motte Analogy pH meter while the dissolved oxygen content of the water was
measured with a YS1® Dissolved Oxygen meter.
RESULTS
(a) Toxity
testing of Water Soluble fraction (WSF) diesel fuel oil Oreochromis niloticus fingerlings
within 24 hours. Within 24 hours, death was observed with 19.2 ppm WSF diesel
fuel (see table 1).
(b) Toxicity
testing of water soluble fraction (WSF) diesel fuel in Oreochroms niloticus fingerlings
within 48 hours. Within 48 hours death of fingerlings were observed from
12.8 ppm (one death) and with 19.2ppm (two deaths) (see table 2).
(c) Toxicity
testing of water soluble fraction (WSF) diesel fuel or Oreochromis niloticus fingerlings
within 72 hours. Within 72 hours, deaths of oreochrous niloticus observed
with 3.2ppm WSF diesel, 3.2ppm (one death) 6.4 ppm (3 deaths); 12.8 ppm (5deaths)
and 19.2ppm (7 deaths) (table 3).
(d) Toxicity
lasting of Water soluble fraction (WSF) diesel fuel or Oreochromis niloticus fingerlings
within 96 hours fingerlings were observed with 1.6 ppm (2 deaths). 3.2 ppm
(3 death). 6.6ppm (5) deaths). 128 ppm ( 7 deaths) and 19.2ppm (10 deaths all
died). See table 4.
TABLE 1: Toxicity Testing at 24 hours
CONC (ppm)
|
No. Surviving
|
% alive
|
% dead
|
0
|
10
|
100
|
0
|
1.6
|
10
|
100
|
0
|
3.2
|
10
|
100
|
0
|
6.4
|
10
|
100
|
0
|
12.8
|
10
|
100
|
0
|
19.2
|
8
|
90
|
10
|
TABLE 2: Toxicity of Testing at 48 hours
CONC (ppm)
|
No. Surviving
|
% alive
|
% dead
|
0
|
10
|
100
|
0
|
1.6
|
10
|
100
|
0
|
3.2
|
10
|
100
|
0
|
6.4
|
10
|
100
|
0
|
12.8
|
9
|
90
|
10
|
19.2
|
8
|
80
|
20
|
Table 3: Toxicity Test at 72 hours
CONC (ppm)
|
No. Surviving
|
% alive
|
% dead
|
0
|
10
|
100
|
0
|
1.6
|
10
|
100
|
0
|
3.2
|
10
|
90
|
10
|
6.4
|
10
|
70
|
30
|
12.8
|
10
|
50
|
50
|
19.2
|
10
|
30
|
70
|
Table 4: Toxicity Testing at 96 hours
CONC (ppm)
|
No. Surviving
|
% alive
|
% dead
|
0
|
10
|
100
|
0
|
1.6
|
8
|
80
|
20
|
3.2
|
7
|
70
|
30
|
6.4
|
5
|
50
|
50
|
12.8
|
3
|
30
|
70
|
19.2
|
0
|
0
|
100
|
96 HOURS LC50 DETERMINATION: Using Arithmetic method of
Karber the LC50 value was determined as follows:
conc (ppm)
|
conc. difference
|
no. alive
|
no. dead
|
mean death
|
mean death dose diff
|
O (control)
|
-
|
10
|
0
|
-
|
0
|
1.6
|
1.6
|
8
|
2
|
1
|
1.6
|
3.2
|
1.6
|
7
|
3
|
2.5
|
4
|
6.4
|
3.2
|
5
|
5
|
4
|
12.8
|
12.8
|
6.4
|
3
|
7
|
6
|
38.4
|
19.2
|
6.4
|
0
|
10
|
8.5
|
54.4
|
|
|
|
|
|
111.2
|
Table 2: Table for 96 hour LC50 determination based on
Arithmetic method of Karber. (adapted by Dede 1992).
Table 5: Histopathological findings in the gills
of O.niloticus following exposure to the WSF of diesel fuel
Tissue
|
Concentration
|
Effect
|
gill
|
Control
|
Normal gill lamellae
|
LC50
|
Elongation of gill lamellae
|
LC100
|
Lamellar fusion and hyperplasia
|
PHYSICOCHEMICAL PARAMETERS:The average values obtained for the physicochemical
parameters (pH and Dissolved oxygen content) measured for the WSF and control
water as compared with the Federal Environmental Protection Agency, (FEPA)
standard of water quality for aquatic life (Horsfall & Spiff, 1998)
Table 6: Comparison of the WSF diesel with habitat
water based on pH and dissolved oxygen content.
|
pH
|
Dissolved oxygen (mg/L)
|
Water
|
7.3
|
4.5
|
Habitat
|
7.8
|
7.2
|
FEPA
|
6.0-9.0
|
6.0-8.0
|
DISCUSSION
The exposure of O.niloticus fingerlings
to water-soluble fraction of diesel fuel showed mortality even at low concentrations.
From the international classification of toxicity of substances based on
their median lethal concentration (LC50) the water soluble fraction
(WSF) of diesel fuel is slightly toxic to tilapia fingerlings. This corroborated
with previous reports on the effect of water-soluble components of hydrocarbon
on aquatic life (Tatem et al, 1978; Kiihnhold, 1980). Histopathological study
of the gills showed structural abnormalities such as elongation, fusion and
hyperplasia of the lamellae. This agreed with the study of Oladimeji and
Onwumere (1988) that refinery effluents caused gill damage in O. niloticus.
The dissolved oxygen content
of the WSF was found to decrease below the FEPA acceptable level for water
quality for the sustenance of aquatic organism. The dissolved oxygen tension
in aquatic organisms in water contaminated with organic pollutants has been
shown to be due to the diversion of the dissolved oxygen meant for respiration
to the oxidation of organic pollutant. The extant of depletion of oxygen
in the water is often a function of the concentration of the organic pollutants
in it (Horsfall & Spiff, 1998).
The oxygen stress encountered by the fish which is responsible for the respiratory
distress and death, was due to their inability to withstand the oxygen depletion
of the water induced by the organic compounds in the water soluble fraction
of the fuel. Similar oxygen stress imparted by the water soluble fraction
of crude oil had been studied in the shrimp, Palaemon adspersus (Baden, 1982)
and in the catfish Clarias gariepinus (Igloh etal, 2001 Furthermore,
the gill elongation and hyperplasia in the WSF-Cultured fingerlings may be
induced by this oxygen stress.
The pH of the water was within the FEPA acceptable range for the sustenance
of aquatic life (Table 6) and may unlikely contribute to the toxicity of
the water soluble fraction.
Improvement on survival of
some of the fish with duration of incubation could be the result of enzyme
induction which enhanced the rate of breakdown of the pollutants. This confirmed
the report that detoxifying enzymes could be induced in fish with prolonged
exposure to pollutants (Pyne & Penrose, 1975).
REFERENCES