Toxicity Evaluations of Mixed Chemicals

Detection of EC50 values of mixed chemicals

To clarify whether the toxicities of mixed chemicals were enhanced or reduced in comparison with the cases of single chemicals, each combination of two chemicals was evaluated in the present system. As shown in Figure 3 a, BLI changes of V. fischeri were followed for the mixture of SEA-NINE 211 and TPBP, where they were reduced when the concentrations of two kinds of chemicals as well as incubation times increased. The inhibitory effects became clear at 20-30 min of incubations when the mixture concentrations of SEA-NINE 211 and TPBP were higher than 0.21 and 0.56 mgl _1, respectively. The values of INH (%) were enhanced by an increase in the concentration of both SEA-NINE 211 and TPBP in combination, as shown in Figure 3b. From the crossing point of 50% INH value, the authors determined the EC50 value for the mixture of SEA-NINE 211 and TPBP to be 0.19 and 0.47 mgl_1, respectively. EC50 values for other mixtures (A, B) were also calculated in the same way described above, which are summarized in Figure 4.

The toxicities of samples can be shown based on the EC50 values, in which lower values reflect the higher toxicity, since 50% BLI values in the cell suspension of V. fischeri are inhibited by relatively low concentration of test samples. The EC50 values in Table 1 and Figure 4 show the toxicities of single and their mixed chemicals, respectively. To determine the interaction

500 400

5 10 15 20 25 Incubation time (min)

5 10 15 20 25 Incubation time (min)

5 10 15 20 25 Incubation time (min)

5 10 15 20 25 Incubation time (min)

Figure 2 Typical inhibition patterns of the BLI of V. fischeri in the presence of CuSO4 or Ziram. BLI changes in V. fischeri were followed in the presence of (a) CuSO4 and (b) Ziram as a function of the incubation time. Different concentrations of CuSO4 were added by 0.06 (closed circles), 0.09 (open triangles), 0.13 (closed triangles), 0.20 (open squares), and 0.30 (closed squares) mgl"1. For Ziram, the following concentrations were added: 0.08 (closed circles), 0.12 (open triangles), 0.18 (closed triangles), 0.27 (open squares), and 0.42 (closed squares) mg r1. Control samples containing ASW with 1% (v/v) DMSO are shown as open circles. All the data were obtained from triplicate experiments, averages of which were described. RLU, Relative Luminescence Unit.

Table 1 EC50 values of single chemicals examined (mg l 1)a

Chemical

ec50 value

TBT-Cl

0.02 ± 0.00

Zn-pt

0.08 ± 0.01

Cu-pt

0.12 ± 0.01

CuSO4

0.22 ± 0.02

Ziram

0.31 ± 0.02

SEA-NINE

0.35 ± 0.02

211

TPBP

0.75 ± 0.05

IPBC

8.49 ± 0.79

Diuron

12.74 ± 1.21

DCF

39.02 ± 2.18

Irgarol 1051

>40.00b

aThe EC50 values were calculated at 30 min of incubation from triplicate experiments.

bThe EC50 value of Irgarol 1051 was undetectable due to its solubility limitation (about 40 mg r1).

aThe EC50 values were calculated at 30 min of incubation from triplicate experiments.

bThe EC50 value of Irgarol 1051 was undetectable due to its solubility limitation (about 40 mg r1).

effects in each combination of two chemicals, their toxicities should be compared with those of respective single chemicals.

Comparison of EC50 values of single and mixed chemicals

BLI changes in cell suspension of V. fischeri were followed as a function of the incubation time and different concentrations consisting of CuSO4 and Ziram. As shown in Figure 5, the mixture consisting of 0.05 mglof CuSO4 and 0.06 mgl of Ziram resulted in a significant reduction of BLI value in comparison with the control. In the presence of the same level of concentrations in CuSO4

or Ziram of single form, however, BLI values were almost the same as control, as shown in Figures 2a and 2b. The EC50 values of this mixture were calculated from the BLI changes in the presence of various concentrations of CuSO4 and Ziram (Figure 5). The EC50 values in the single form of CuSO4 and Ziram were sufficiently high, 0.22 and 0.31 mgl _1, respectively, but they became markedly low when they were mixed, as shown in Figure 4. That is to say, the mixture became highly toxic, although the respective single chemical showed low level of toxi-city. Thus, the concentration to reveal the same level of toxicity in the mixture as cases of respective single chemicals resulted to less than one-fifth. It might be of value to point out that the EC50 values for the combinations of CuSO4 together with Cu-pt, Zn-pt, or Irgarol 1051 became lower in comparison with those of single chemicals, as shown in Table 1 and Figure 4. The same concentrations of their combinations containing CuSO4 showed much higher inhibitory activities than respective single chemicals, probably owing to the synergistic effect in combinations with CuSO4. All of the EC50 values in CuSO4-containing combinations mentioned above were reduced to one-fourth or one-fifth of those of each single chemical. The EC50 values for the other combinations were half as those of their single chemicals.

Typical patterns of interaction of mixed chemicals

Further evidence of toxicity enhancement by mixing the chemicals was recognized for the combinations such as Ziram + CuSO4, Zn-pt + CuSO4, SEA-NINE 211 + TPBP, and Cu-pt + Diuron. As shown in

700

100

: 1

600

90

-

500

80 70

EC50 = (0.19, 0.47)

□C

)

60 50 40

/

Ij 300 B

IN

200

30

100

20 10

0

0

5 10 15 20 25 30 Incubation time (min)

5 10 15 20 25 30 Incubation time (min)

SEA-NINE 211 concentration (mg l-1)

Figure 3 Inhibition of BLI of V. fischeri in the presence of mixed chemicals, SEA-NINE 211 and TPBP. (a) BLI changes in V. fischeri were followed in the presence of the mixture composed of SEA-NINE 211 and TPBP as a function of the incubation time. Different concentrations of SEA-NINE 211 and TPBP were added by 0.06 and 0.16 (closed triangles), 0.09 and 0.25 (open triangles), 0.14 and 0.38 (closed diamonds), 0.21 and 0.56 (open diamonds), and 0.32 and 0.84mgl_1 (closed squares), respectively. Control samples containing ASW with 1% (v/v) DMSO are shown by open circles. (b) The INH (%) values of BLI forthis mixture were calculated at 30 min of incubation. The EC50 values of mixed chemicals were obtained as the concentration corresponding to 50% of INH value. All the data were obtained from triplicate experiments, averages of which were described.

Irgarol 1051

Diuron

UDa

Ziram

0.268, 19.05s

0.281, 6.434

Cu-pt

0.097, 14.86

0.123, 6.027

0.071, 0.153

Zn-pt

0.063, 17.16

0.071, 4.094

0.042, 0.106

0.058, 0.068

TPBP

UDa

0.714, 5.976

0.589, 0.216

0.553, 0.094

0.443, 0.064

SEA-NINE 211

0.247, 19.29

0.216, 5.442

0.190, 0.209

0.180, 0.092

0.137, 0.060

0.185, 0.465

DCF

22.80, 13.44

32.18, 6.103

25.75, 0.213

21.86, 0.084

18.50, 0.061

21.05, 0.477

26.42, 0.199

CuSO4

0.099, 16.08

0.122, 4.221

0.048, 0.072

0.045, 0.032

0.047, 0.028

0.110, 0.452

0.093, 0.127

0.118, 21.59

IPBC

UDa

4.393, 6.434

3.472, 0.222

3.160, 0.094

2.674, 0.068

2.899, 0.507

3.004, 0.175

4.393, 33.92

4.393, 0.186

Irgarol 1051

Diuron

Ziram

Cu-pt

Zn-pt

TPBP

SEA-NINE 211

DCF

CuSO4

IPBC

Figure 4 EC50 values for the mixed chemicals examined (mg r1). The EC50 values are calculated at 30 min incubations from triplicate experiments. aUD: undetectable due to the low toxicity of Irgarol 1051. bThe values in (A, B) of each cell indicate that 50% inhibition for BLI of V. fischeri was afforded by the mixture composed from the chemicals in row and column titles, the concentrations of which are A and B, respectively.

The relationship between the toxicities of single chemicals and their combinations shown in Figures 6b and 5 c, however, is different from that of Figure 6a. Inhibitory reduction of BLI due to the action of SEA-NINE 211 (0.19 mgl or TPBP (0.47 mgl presented as single form showed approximately 25% value at 30 min of incubation, as shown in Figure 6b, while it increased to 44% by mixing them. In the case of Diuron and Cu-pt, however, the toxicity did not change remarkably by mixing them, that is, the toxicity level of their mixture was almost same as that of Cu-pt alone (Figure 6c). In comparison with two examples of combination shown in Figures 6b and 6c, the mixtures of CuSO4 with either Ziram or Zn-pt showed a significant increase of toxicity in combination (Figure 6a), from which it is concluded that these combinations are classified as synergistic.

In the same way, toxicity analyses for CuSO4 in combination with Cu-pt or Irgarol 1051 were performed, and then marked synergistic effects were also recognized although data was not shown. The interactions of the combinations shown in Figures 6b and 6c are obviously different from the synergism, the details of which are discussed in the next section.

Oplan Termites

Oplan Termites

You Might Start Missing Your Termites After Kickin'em Out. After All, They Have Been Your Roommates For Quite A While. Enraged With How The Termites Have Eaten Up Your Antique Furniture? Can't Wait To Have Them Exterminated Completely From The Face Of The Earth? Fret Not. We Will Tell You How To Get Rid Of Them From Your House At Least. If Not From The Face The Earth.

Get My Free Ebook


Post a comment