Handbook of Solvents - George Wypych - ChemTech - Ventech!

15.2.2 A simple test to determine toxicity using bacteria 1109
cium. The toxicity of 4 chemicals was enhanced, was greater, when the calcium and EDTA
were present.
The addition of calcium and EDTA at these concentrations had no effect on the apparent
toxicity of: 1,4-dinitrobenzene, the herbicide 2,4-D, 2,4-dinitrotoluene, 4-chloro-benzoate,
carbon tetrachloride, chloroform, cynazine, hexachlorophene, isonicotinic acid the
insecticide Lindane, o-nitrophenol, p-toluidine, the antibiotic Streptomycin,
tetrachloroethylene and trichlorophenol.
The addition of calcium and EDTA reduced the toxicity at least 10% of
2,6-dinitrocresol, 2,4-dinitrophenol, 2,6-dinitrophenol, 2,6-dinitrotoluene, 2-methyl
resourcinol, 3-phenoxybenzoate, 2,4-dinitrocresol, the detergent CTAB, the antibiotic
Neomycin, p-hydroxybenzoate, pentachlorophenol, salicylic acid, and trichloroethylene.
p-amino benzoic acid and p-hydroxy benzoate were no longer toxic with these levels
of calcium and EDTA.
3-chlorobenzoate, 3-methyl-4-nitro-phenol, and the detergents sodium lauryl sulfate
and Tween 80 had increased toxicity with the addition of calcium and EDTA. No correlation
between the structure of the chemical and the effect of EDTA and calcium could be ascertained.
For example, streptomycin and neomycin are both amino glycoside antibiotics.
Calcium and EDTA did not affect the toxicity of streptomycin but nearly eliminated the toxicity
of neomycin.
Several chemicals were tested with 25 μmoles EDTA and 7.4 μmoles of calcium, 10
times the amount used in the experiments reported. The results of this experiment are shown
in Table 15.2.2.6. In the presence of high levels of EDTA and calcium, isonicotinic acid is
no longer toxic. Dinitrophenol and 3-phenoxy benzoate, in the presence of low levels of
EDTA and calcium were not as toxic, in the presence of high levels, it had the same toxicity.
In presence of low levels of EDTA and calcium, Streptomycin was comparably toxic, the
calcium and EDTA did not affect the toxicity. In the presence of high levels, Streptomycin
was extremely toxic. Pentachlorophenol, a common soil contaminant, had reduced toxicity
with low levels of EDTA and calcium, had elevated levels of toxicity with high levels of
EDTA and calcium. This work shows that the effect of EDTA and specific toxins must be
worked out before any conclusions as to the toxicity of the compound in the presence of
EDTA and calcium can be established.
Calcium and magnesium are commonly found in water. Obviously if this assay is to be
used with water samples, EDTA must be added. It must be determined using water uncontaminated
with organic toxins how much EDTA must be used to compensate for the divalent
cations. Often the concentration of divalent cations is determined by atomic absorption
spectroscopy. However, these values do not agree with the toxicity relieved by EDTA. Soil
samples with as much as 5 gm calcium (45 mM) per kg soil have been assayed using 2.5
μmoles EDTA in each sample (Hillaker, 1996). The calcium is complexed with sulfate and
phosphate ions and the calcium is not available to the cell, is not seen by the mechanism that
reduces the dye. Levels of soluble calcium and magnesium in water are very low. We have
found that 2.5 μmoles of EDTA relieves the inhibition caused by divalent cations in all water
and soil samples tested thus far (Botsford, 2000b).