Environmental Toxicology and Chemistry, Vol. 7 , pp. 241-243, 1988 0730-7268/88 $3.00 + .

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Printed in the USA. Pergamon Journals Ltd.

Short Communication
AVOIDANCE OF BRASS POWDER-CONTAMINATED
SOIL BY THE EARTHWORM, LUMBRICUS TERRESTRIS
RANDALL S . WENTSEL* and MARKA. GUELTA
U.S.Army Chemical Research, Development and Engineering Center,
Aberdeen Proving Ground, Aberdeen, Maryland 21010-5423
(Received 13 April 1987; Accepted 15 September 1987)

Abstract-Tests of the avoidance of brass powder (70% Cu, 30% Zn) contaminated soils by the
earthworm Lumbricus terrestris at concentrations ranging from 0 to 200 w g / g were conducted.
Avoidance was significant ( p < 0.001) at brass concentrations as low as 38 pg/g. Avoidance was
found to be a more sensitive indicator of the impact of brass powder on the worms than sublethal
effects measured by weight loss.
Keywords- Toxicity Avoidance Brass powder Earthworms

INTRODUCTION to soils. Bhat et al. [12] found earthworm migra-

The use of earthworms as test organisms for
assessing the impact of chemicals o n terrestrial
ecosystems has been reviewed by several authors
[l-31. Earthworms have been found to be mobile
in soil vertically to a depth of 2.5 m and horizon-
tally up to 10 m/year [4]. Measurements of the
impact of smelting operations [5,6], mining [ 7 ] ,
and the application to land of sludge [8] have
shown that earthworms are sensitive to metals con-
tamination. Gish and Christensen [9] determined
that earthworms could be used to monitor heavy
metal contamination because they accumulate met-
als. Bengtsson et al. [ 5 ] studied the impact on
earthworm populations of atmospherically depos-
ited metals from a brass mill. They evaluated the
relationship between soil metal contamination and
earthworm density.
Laboratory tests on the toxicity of metals to
. earthworms have generated LC5Os and have
recorded sublethal effects such as weight loss and
reduction in cocoon production. A six-week LC50
for Lumbricus rubellus for copper (as CuCl,) in
sandy-loam soils was near 1,000 pg/g [lo]. A sig-
nificant reduction in L. rubellus cocoon produc-
tion was observed at 150 pg/g copper, and body
weight reductions were observed at 372 pg/g cop-
per [ l l ] .
Another indicator of the impact of chemicals
on earthworms may be their avoidance of contam-
inated soils. Avoidance could be a sensitive mea-
sure of the sublethal effects of chemicals applied
~
*To whom correspondence may be addressed.
24 1
tion to be a sensitive test of food preference.
The purpose of this experiment was to deter-
mine if the earthworm Lumbricus terrestris would
avoid soil contaminated with brass powder.
Elemental analysis of the brass powder showed
that it was approximately 70% copper and 30%
zinc, with metal impurities totaling less than 170.
The brass powder (Atlantic Powder Metals) parti-
cles have diameters of 1.7 pm, with a thickness of
between 0.08 and 0.32 fim.
METHODS
To test avoidance, 15 cm x 50 cm chambers
were constructed from Styrofoam. Four chambers
were used at each concentration. A common test
species, Lumbricus terrestris, was purchased from
a local supplier and held for two weeks before the
initiation of the experiments.
The avoidance tests were conducted using non-
sterile loam soils. The soil parameters were ana-
lyzed by the Soil Testing Laboratory, University of
Maryland, College Park, Maryland [13] (Table 1).
The soils were air-dried and sieved through a 2-mm
mesh screen before use.
The test medium consisted of a nonsterile soil
(600 g), distilled water (150 ml) and the brass pow-
der spike. The brass powder concentrations ranged
from 17 to 200 pg/g (Table 2). The corresponding
copper and zinc concentrations ranged from 12 to
140 pg/g and from 12 to 67 pg/g. The method of
metal analysis in soils and the quality control pro-
cedures are described elsewhere [ 141.
242 R. S. WENTSELAND M. A. GUELTA

Table 1. Soil parameters added at the interface between the two soils. The
chambers then were placed randomly into a pre-
Texture
Type
Loam
Sassafras
cision low-temperature incubator set at 13.0 *
4.6 0.2"C. After 7 d the locations of the earthworms
PH
Cation exchange capacity 3.6 meq in the chamber were noted. The earthworms were
Organic matter 1.O% observed t o be in the control soil, the contami-
Magnesium 15.0 wg/g nated soil or, for scoring purposes, in a neutral
Phosphate 3 pg/g 3- to 4-cm strip at the soil interface. A binomial
Potash 32 clg/g
Boron 0.1 pg/g distribution was used to determine significant dif-
Calcium 40 pg/g ferences in the avoidance experiments.
Nitrate 3.5
Particle size analysis RESULTS
Sand 32%
Silt 50% The results of the avoidance studies are pre-
Clay 18% sented in Table 3. The earthworms' avoidance was
significant ( p c 0.001) at brass powder concen-
trations of 38 to 200 pg/g. No significant avoid-
Table 2. Measured metal concentrations in soil" ance was observed in the earthworms exposed to
17 pg/g brass powder. The threshold avoidance
Calculated concentration is equivalent to elevated copper and
brass zinc soil concentrations of 26 and 20 pg/g,
Copper Zinc concentration respectively.
(Fg/g; fSD) ( p d g ; +sD) (&g)
Control 0" Ob 0' DISCUSSION
Level 1 1 2 + 1.1 *
12 3.8 17
Level 2 26 * 2.3 20 k 0.1 38 Field observations of earthworm population
Level 3 84 * 8.7 47 k 6 120 declines in copper-contaminated soils have been
Level 4 140 + 10 *
67 7 200 thought to be due to toxic effects of the copper.
"Background metal subtracted out. However, avoidance of the area by the worms
bBackground metal levels of copper, 13.4 2.7 pg/g. could also be a contributing factor.
'Background metal levels of zinc, 35 3 pg/g. In a previous study [14], the toxicity of brass to
the earthworm L. terrestris was determined. An
LC50 of 190 p g / g brass powder, with 95% fidu-
Table 3. Avoidance of spiked soil by earthworms cia1 limits of 169 and 220 pg/g, was generated.
Sublethal effects, measured as weight loss, were
Brass powder-spiked soil also observed. Brass concentrations as low as 120
concentration (pg/g) p g / g were found to significantly ( p < 0.05) reduce
200 140 38 17 0
earthworm weight. These results were generated in
Earthworm - - - - - the same soil used in these avoidance studies.
location": C S C S C S C S C Cb The results of this experiment showed t h a t L.
terrestris avoided soil brass concentrations of 38 ,
5 0 5 0 5 0 4 1 2 3 p g / g . Avoidance was a more sensitive indicator of
5 0 5 0 5 0 2 3 2 2
5 0 4 1 5 0 2 3 2 3 the environmental effects of brass powder than
5 0 5 0 4 1 3 2 3 2 LC50 or weight loss. A 17 pg/g brass concentra-
Total 20 0 19 1 19 1 11 9 9 10 tion did not produce significant avoidance by L.
terrestris. The threshold avoidance concentration
"C, control soil side of chamber; S, spiked soil side of lies between 17 and 38 pg/g, or elevated copper
chamber. and zinc concentrations of 12 to 26 pg/g and 12 to
bC, control soil versus control soil.
20 pg/g, respectively.
The characteristics of the soil used in these
experiments tended to make the metals more bio-
The soil spiked with the brass powder was available and therefore the soil was more toxic
placed in one-half of the chamber and uncontami- than most. Other soils that have higher pH, or-
nated soil was placed in the other half. Five earth- ganic matter content and cation exchange capaci-
worms arbitrarily selected from holding trays were ties will complex with metals to a greater degree
 Avoidance of brass powder by earthworms
and will result in higher threshold avoidance con-
centrations. However, the greater sensitivity of
avoidance, as compared with LC50 or weight loss,
should be the same regardless of what soil or
artificial soil mix is used.
Acknowledgement -This work was supported by the
U.S. Army Biomedical Research and Development Lab-
oratory, Ft. Detrick, Maryland, Mr. Gardner, Project
Officer.
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