Determination of LD 50 and experimental doses

Transcription

1 Determination of LD 50 and experimental doses

2 Determination of LD 50 of sodium arsenate and sodium selenite: Trevan (1927) first suggested the measurement of toxicity of any toxicant by LD 50 determination. Due to variations in the environment, toxicity studies should not be highly standardized (Warren, 1971). Feeding tends to increase the metabolic activities such as rates of respiration, excretion or production of other waste products, which affect the toxicity. So food supply to the test animals should be totally stopped during the experiment (APHA, 1960). For the present experiment the test animals were starved 24 hours prior to and during the 48 hours test period. APHA (1960) also suggested that the preferable duration for a toxicity test should be at least 48 hours or 96 hours. For the present study, exposure period selected was 48 hours. For the determination of median tolerance limits for sodium arsenate and sodium selenite, a bunch of 10 healthy, laboratory-acclimatized mice were selected. Different concentrations of sodium arsenate injected intraperitoneally and different concentrations of sodium selenite were fed orally. A concurrent control was maintained throughout the experimental period under identical conditions by injecting sterile physiological saline (0.7 ). Any change in behaviour and other responses were carefully noted. The number of dead mice was recorded at 48 hours after treatments. Whether any bioassay animals died due to factors other than exposure to the test chemicals, control was calculated using the Abbott s formula as follows: Corrected () = Where, M obs = Observed treatment M control = Control 52

3 Calculation of LD 50 of sodium arsenate: Estimation of the dose range and percentage of of sodium arsenate An approximate LD 50 value of sodium arsenate was initially determined using a small number of mice (two each dose). Then twelve doses were chosen for determination of LD 50, starting from no death to 100. For estimation of LD 50 of sodium arsenate, these twelve doses were given intraperitoneally to the twelve groups of mice, ten in each group. Conversion of percentage mortalities to probits and calculation of LD 50 The percentage of animals those were died at each dose level was then transferred to probit (Table 3). Table 3. Results of the intraperitoneal doses of sodium arsenate on mice for determination of LD 50. Groups Dose (mg/kg) Log 10 dose Total Dead Corrected Probits The 48 hours median tolerance limits or LD 50 were determined following the method of Miller and Tainter (1944) by straight line graphical interpolation of the data. The median value was derived by calculating the experimental data on semi-logarithmic paper, with test concentration on the logarithmic scale and the percentage of survival on the arithmetic scale. A straight line was drawn 53

4 Probits Chapter IV: Determination of LD 50 between the two points, which are above and below the 50 survival line. The point of intersection with the 50 survival line was considered as the LD 50 concentration. The 48 hours LD 50 was found to lie in the range of mg kg -1 body weight. On a second trial the exact value was determined to be 85 mg kg -1 body weight (Fig. 11) LD50 Graph y = x R 2 = Log10 concentration Fig. 11. Plot of the Log 10 concentration versus probits from table 1 for calculation of LD 50 of sodium arsenite administered intraperitoneally. LD 50 regression: y = x = x x = LD 50 = antilog ( ) = mg kg -1 According to material safety data sheet (MSDS, see web based resources in reference chapter); and Sax and Lewis (1987), LD 50 of sodium arsenate was calculated as mg kg -1 when given intramuscular to the mice, although in our study LD 50 of sodium arsenate was estimated as mg kg -1. The doses those were selected for further studies in the present context were 5 mg SA kg -1 54

5 body weight and 30 mg SA kg -1 body weight, and these were 1/3.41 LD 50 and 1/2.84 LD 50 respectively. Calculation of LD 50 of sodium selenite: Estimation of the dose range and percentage of of sodium selenite An approximate LD 50 value of sodium selenite was initially determined using a small number of mice (two for each dose). Then twelve doses were chosen for determination of LD 50, starting from no death to 100. For estimation of LD 50 of sodium selenite, these twelve doses were given orally to the twelve groups of mice, ten in each group. Conversion of percentage mortalities to probits and calculation of LD 50 The percentage of animals those were died at each dose level was then transferred to probit (Table 4). Table 4. Results of the oral doses of sodium selenite on mice for the determination of LD 50. Groups Dose (mg/kg) Log10 dose Total Dead Corrected Probits

6 Probits Chapter IV: Determination of LD 50 The 48 hours LD 50 was found to lie in the range of 8 12 mg kg -1 body weight. On a second trial the exact value was determined to be 11 mg kg -1 body weight (Fig. 12) LD 50 graph y = x R 2 = Log 10 Concentration Fig. 12. Plot of the Log 10 concentration versus probits from table 2 for calculation of LD 50 of sodium selenite administered orally. LD 50 regression: y = x = x x = 1.03 LD 50 = antilog (1.03) = mg kg -1 According to material safety data sheet (MSDS, see web based resources in reference chapter), LD 50 of sodium selenite was calculates as 7.1 mg kg -1 when given orally to the mice, although in our study it was calculated as mg kg -1. The dose that was selected for further studies in the present context were 0.25 mg SS kg -1 body weight which was 1/42.84 LD

7 Selection of exposure concentration: Perkins (1979) suggested that the measurement based upon acute poisoning are unhelpful since they tell nothing of the impact. The effect of toxicity can only be investigated with organisms not at the point of death i.e. in truly sub-lethal doses. In the present investigation, exposure concentration of the sodium arsenate and sodium selenite was selected on the basis of the observed levels at different concentrations. Only sublethal concentrations of these compounds were selected for individual static bioassay where no of mice was recorded throughout the experimental duration. The concentrations of sodium arsenate used were 1/3.41 LD 50 and 1/2.84 LD 50, and the concentration of sodium selenite used was 1/42.84 LD 50 for the present study. 57