5

RATE OF OXYGEN

CONSUMPTION
 40

INTRODUCTION

The rate of oxygen consumption which is the basic physiological pa­

rameters serves as an indicator of physiological stress and changes in the

oxygen consumption could be used to evaluate any change that would occur in

the metabolism due to alteration in the environment. Such stress as pesticides

(Roberts ,1975} heavy metal (Calabrease et. al., 1977) as well as natural stresses

as temperature and salinity (Kinne, 1964b; Precht et. al., 1973; Vernberg, 1972,

Basha Mohideen, 1979; Basha Mohideen and Parvatheswara Rao, 1976) In­

secticides gain entery through the gills of fishes (Holden, 1962, Premdas and

Anderson, 1963; Ferugson and Good year, 1967) and thus it results in the di­

rect entry into the vascular system and affecting various tissues and organs

consequently, various physiological processes would be affected in varying de­

grees (Ashley, 1972) The foremost physiological function to be affected is oxy­

gen consumption of the aquatic animals. Pesticidal compounds like triterpenic

glycoside initially affected the respiratory pump and correspondingly oxygen

consumption (Bhattand Farswan, 1991). Malta Reddy and Basha Mohideen

(1989) reported little drop in the rate of oxygen consumption in Cyprinus carpio

exposed to synthetic pesticide fenvalerate and cypermethrin.

All the above investigations indicate that the sensitive indicator of stress

namely oxygen consumption in fish exposed to pollutants in general is sup­

pressed considerably. Studies involving the oxygen consumption of the whole

animal with reference to the sublethal effects of natural pesticide Azadirachtin

are very few and very little attempts has been made with regard to the food fish

of economic importance that have commercial value to a very great extent.

 *■ K- U. LIBMir }
METHOD ACC. Ni.OJLfl2JL5. t
Call. No.„_ _ _ _ _ _ _
The rate of oxygen consumption was inceasured at differenfsele'CtSC

sublethal exposure periods like 24hrs, 7 days, 15days, 20 days and 30 days

and in fresh water without Azadirachtin serving as control by the improved

Winkler’s method as developed by Basha Mohideen and Kunnemann (1978).

In this method the experimental fish Catla catla was introduced into the

respiratory chamber covered with a black foil. The fish was allowed to settle in

the chamber for a period of five minutes. The initial water samples were col­

lected in the winklers bottles of 50ml capacity half an hour after the introduction

of the fish the final samples were collected into the winklers bottles. The dis­

solved oxygen contents in these samples were estimated as given below with

the help of micro syringe 0.25ml of 40% Mncl, 0.25ml of 10% KOH and 0.25ml

of 3% dye (Leukobarbeline Blue -1) were added respectively into the bottles

containing initial and final samples separately which were then closed and

shaken well and the precipitate was allowed to settle. After 3minutes 1m of

40% citric acid was added and the bottles were again shaken for a minute in

order to dissolve the precipitate. After five minutes a deep blue colour was

developed and 5ml of this blue colour solution was pipetted out into a 50ml

standard flask and was made up to the mark with distilled water. The optical

density of the blue colour was read in spectronic - 20 (Spectrophotometer) at

a wave length of 578 nm using glass cuvettes of 1 cm light path. The intensity of

the blue colour of the solution is directly proportional to the amount of oxygen

present in the sample. The calibration curve was prepared by using water
 42

samples of different oxygen concentration (by bubbling N 2 into tap water) The

equation of calibration and optical density at 578 nm was calculated by linea

regression.

A calibration factor 6.607 was obtained in the present method. The dif­

ference in oxygen concentration between the initial and final sample gives the

rate of oxygen consumption by the fish the rate oxygen consumption is calcu­

lated by using the formula.

Initial O.D - Final O.D x 6.607 x 2

-------------------------------------------------------- = Oxygen consumption (02 ml/gm/hr)
Body weight

The reagents used for this experiment were purchased from Merk

(Darmstadt) and the dye from the Altman (Berlin). The solutions were prepared

with oxygen free distilled water. In order to present the decomposition of the

dye amber coloured bottles were used and 0.3 ml of 25% ammonium hydrox­

ide per 100 ml solution of the dye was used.

RESULTS

The data for the rate of oxygen consumption (02 ml/g/hr) in two different

groups of experimental fish, namely small and large individuals of Catla catla

at different sublethal exposure periods of Azadirachtin like 24hrs. 7 day, 15

day, 20day and 30 day including control medium (freshwater without Azadirachtin)

) and initial % elevation and later % suppression in 02 consumption at different

sublethal exposure periods, calculated in relation to the rate of oxygen con-

 43

sumption in the control as represented in (fig, 3,4 and Table 4). In both the

stages of fish Catla catla in relation to control, the rate of oxygen consumption

increased at 24 hrs. and later decreased at different sublethal exposure peri­

ods and the decrease in oxygen consumption and hence the % suppression of

oxygen consumption, was progressive from 7-day period and reached a maxi­

mal % suppression of oxygen consumption at the middle (15-day period). Fur­

ther towards the end of 30-day exposure there was a rise in oxygen consump­

tion from its maximum suppression and reached nearer to the control, thus this

fish exhibited a fairly good amount of recovery in its oxygen consumption at 30-

day.

DIFFERENCES BETWEEN SMALL AND LARGE FISH

The initial % elevation, the % suppression and the % recovery in 02

consumption of these fishes during sublethal Azadirachtin exposure was found

to be specific with reference to size of the fish. The difference between small

and large fishes was also found to be satistically significant. When compared

maximal % suppression of oxygen consumption of small and large fish at 15-

day period was greater in the small individuals. But the % recovery of oxygen

consumption of the two stages of this fish at 30-days period of Azadirachtin

exposure exhibited a reverse order with % suppression.

DISCUSSION

In aquatic animals the first physiological functions to be affected by

pesticides stress is the oxygen consumption. There is a documentary evidence

that the insecticides gain their entry largely through the gills of the fishes (Holden,
 44

1962; Premdas and Anderson, 1963; Ferguson and Goodyear, 1967) and af­

fect the oxygen consumption in aquatic animals.

It is also know that in fishes the haemoglobin content the whole fish blood

various with the number of erythrocytes present and the RBC count for 99% of

oxygen obtained in fishes (Legler etal., 1977)

In the present experiment rate of 02 consumption in both the small and

large Catla catla is abruptly elevated at 24 hrs exposure periods then sup­

pressed during the other sub lethal exposure periods like 7 days and 15 days

period However the % suppression in 02 consumption is maximum at 15-days

exposure i.e 53.16 for small and 45.75 for large fish but there is gradual rise in

both small and large fishes at 30 days exposure period from the earlier sup­

pression this indicates that Azadirachtin a natural pesticide, toxicity is reduced

as reflected by the removal of inhibitors to extent.

The recovery from 76.19 to 83.15 in both small and large fishes respec­

tively. In the rate of 02 consumptions earlier studies also revealed that pesti­

cide exposure in fishes generally resulted in the suppression of 02 consump­

tions (Hunter et al., 1967) further depressed respiration in fishes (Lee, 1969).

It is due to respiratory distress as a consequence of the impairment of

oxidative metabolism. Disturbance in oxidative metabolism was reported ear­

lier under cypermethrin toxicity in Labeo rohita (Sridevi,1991) and Tilapia

mossambica (Reddy and Yellamma,1991). These pesticides shows a relative

resemblance in their mode of action with that of natural pesticide Azadirachtin.

 45

The decreased oxygen consumption observed in the present study may also

be due to the reduction in the oxygen up take by gill probably due to gill dam­

age. Architechtural Changes in gill structure' are reported in Tilapia mossambica

under fen valerate stress (Radhaih and Jayanth Rao,1988) also support the

present trend of the investigation.

The present finding in 02 consumption indicate that the Catla catla un­

dergoes compensating mechanism leading to homeostasis by varying the meta­

bolic rate during Azadirachtin exposure .Such compensatory mechanism is

demonstrated in few of the other fish species. These compensating mecha­

nism is demonstrated in above studies including the present study could be

possible probably due to activation and enhancement of detoxifying enzymes

which bring about biodegradation of the pesticide to reduce its toxicity in the

way of recovery from the earlier suppression and such detoxification mecha­

nism was reported earliar by Mukhopadhya and Dehadrai (1978) in an breath­

ing Catfish, Clariasbatrachus,

 TABLE- 4)

Rate of Oxygen consumption (02 ml/gm/hr) in small and large individuals

of Catla catla exposed to sub-lethal concentrations at different exposure

periods besides in control medium (freshwater with out Azadirachtin).

Each Value is a Mean of Six individual measurements.

Small size Control 24 hrs 7 days 15 days 20 days 30 days

Mean 0.315 0.490 0.199 0.185 0.180 0.240

S.D ± 0.09 0.078 0.052 0.045 0.068 0.089

% Change 25.56 -45.62 * -53.16 -42.85 -23.80

-

% Recovery - 76.19
- - - -

‘t’ test P < 0.05 P < 0.05 P < 0.05 P<0.01 N.S
-

Large size Control 24 hrs 7 days 15 days 20 days 30 days

Mean 0.546 0.657 0.346 0.29 0.38 0.454

S.D ± 0.02 0.03 0.06 * 0.08 0.07 0.09

% Change - 20.32 -36.66 -45.75 30.42 16.60%

%Recovery - - - - - 83.15

‘t’test - P<0.001 P<0.01 P<0.001 P<0.001 P<0.05

Rate of Oxygen consumption (02 ml/gm/ hr) in small and large individuals

of Catla catla exposed to sub-lethal concentrations at different exposure

periods besides in control medium (freshwater with out Azadirachtin).
Each Value is a Mean of Six individual measurements.
o

Ll
b>
o
Rate of Oxygen Consumption (02

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ml/gm/hr) o
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CM

%
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0.0 -
Control 24Hrs 7 days 15 Days 20 Days 30 Days

Sublethal Exposure Periods

Rate of Oxygen consumption (02 ml/gm/ hr) in small and large individuals

of Catla catla exposed to sub-lethal concentrations at different exposure

periods besides in control medium (freshwater with out Azadirachtin).

Each Value is a Mean of Six individual measurements.

Fig-4
oo mo oo No fof l oi n ot n o c oM o r o
% Change in Oxygen
Consumption

iSmall Fish
i Large fish

7 days 15 Days 20 Days 30 Days Control

Sublethal exposure' periods