795

The Journal of Toxicological Sciences (J. Toxicol. Sci.)

Vol.39, No.5, 795-802, 2014

Original Article

Volatile organic compounds of polyethylene vinyl acetate

plastic are toxic to living organisms
Tingzhu Teresa Meng
North Carolina School of Science and Mathematics, 1219 Broad Street, Durham, North Carolina, USA 27705
(Received May 29, 2014; Accepted August 20, 2014)

ABSTRACT Volatile organic compounds (VOCs) in polyvinyl chloride (PVC) plastic products readily evaporate; as a result, hazardous gases enter the ecosystem, and cause cancer in humans and other animals. Polyethylene vinyl acetate (PEVA) plastic has recently become a popular alternative to PVC since
it is chlorine-free. In order to determine whether PEVA is harmful to humans, this research employed the
freshwater oligochaete Lumbriculus variegatus as a model to compare their oxygen intakes while they
were exposed to the original stock solutions of PEVA, PVC or distilled water at a different length of time
for one day, four days or eight days. During the exposure periods, the oxygen intakes in both PEVA and
PVC groups were much higher than in the distilled water group, indicating that VOCs in both PEVA and
PVC were toxins that stressed L. variegatus. Furthermore, none of the worms fully recovered during the
24-hr recovery period. Additionally, the L. variegatus did not clump together tightly after four or eight
days exposure to either of the two types of plastic solutions, which meant that both PEVA and PVC negatively affected the social behaviors of these blackworms. The LD50 tests also supported the observations
above. For the first time, our results have shown that PEVA plastic has adverse effects on living organisms, and therefore it is not a safe alternative to PVC. Further studies should identify specific compounds
causing the adverse effects, and determine whether toxic effect occurs in more complex organisms, especially humans.
Key words: Volatile organic compound, Polyethylene vinyl acetate, Polyvinyl chloride,
Environmental toxin, Carcinogen, Lumbriculus variegatus
INTRODUCTION
Volatile organic compounds (VOCs) are chemicals containing carbon that have high vapor pressures at standard
condition. VOCs are used as ingredients in a wide array
of household products such as shower curtains, plastic
cups, lunch boxes, etc. Their high vapor pressures result
from low boiling points, which allow gases to be easily
emitted while the products are in storage or in use. Thus,
concentrations of many VOCs are up to ten times higher
indoors than outdoors (Hester et al., 1995).
VOCs can cause eye, nose, and throat irritation, headaches, loss of coordination, nausea, developmental damage as well as damage to the liver and central nervous,
respiratory, and reproductive systems (Lester et al., 2008).
Some VOCs like benzene and formaldehyde are known to
cause cancer in animals and are suspected to cause cancer
in humans (Huff, 2007; Hauptmann et al., 2009). However, at present, not much is known about the health effects

of these organic compounds in homes or their ecological

effects (Hester et al., 1995).
Polymers derived from vinyl chloride monomer are
called polyvinyl chloride (PVC) or vinyl. Polyvinyl
chloride is used in a plethora of household products from
flooring to wall covering.
The addition of a chlorine molecule differentiates PVC
from other vinyls. However, during combustion, chlorine produces a carcinogen called dioxin that can cause
reproductive and developmental problems, damage the
immune system, and interfere with hormones. Low exposure to dioxin is a health concern because the chemical
first accumulates in the environment, then in plants, in
animals, and finally in humans through food consumption.
In fact, food accounts for 95 percent of human exposure
to dioxin (Viola, 1969; Struciski et al., 2011). Therefore,
the World Health Organizations International Agency for
Research on Cancer (IARC) established vinyl chloride as
a known human carcinogen (Aitio, 1999).

Correspondence: Tingzhu Teresa Meng (E-mail: meng15t@ncssm.edu)

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T.T. Meng

In addition to vinyl chloride, PVC has other harmful

additives including phenol, cyclohexane, decane, methyl
isobutyl ketone, xylene, undecane, ethanol, dipropylene
glycol methyl ether, phthalates, organotins, lead, mercury,
as well as the carcinogenic compounds toluene and ethylbenzene (Lester et al., 2008). Therefore, a previous study
suggested an association between PVC flooring and the
development of bronchial obstruction in children (Jaakkola
et al., 1999). Also, Di (2-ethyl hexyl) phthalate (DEHP)
caused irregularities in male sexual differentiation when
given to pregnant rats (Wolf et al., 1999).
Therefore, stores like Target and Wal-Mart have presently replaced PVC shower curtains with chlorinefree vinyls, most commonly polyethylene vinyl acetate
(PEVA) plastic (Lester et al., 2008). PEVA is also phthalate-free and biodegradable, which makes it currently a
popular alternative to PVC. The absence of chlorine has
been shown to considerably reduce harmful off-gassing
(Nou et al., 2011).
Polyethylene vinyl acetate (PEVA) is a copolymer of
polyethylene (PE) and ethylene vinyl acetate (EVA), and
is similar to PVC because it is also made from petrochemicals (petroleum and natural gas). A study conducted by
SAFC Biosciences in 2006 detected VOCs, such as aromatic compounds, alcohols, aldehydes, alkanes, alkenes,
ketones, phenolics, ethers, amines, sulfides, etc., in PE
and EVA bags. Although PEVA is chlorine-free, it is full
of untested chemicals that may not make it a safe alternative to PVC. Therefore, it is necessary to test the health
and environmental effects of those VOCs in PEVA plastic.
A lot of methods are used to measure concentrations
of dissolved organic carbon (DOC) of a water sample, among which measuring UV-visible absorbance and
then calculating concentration is a convenient and effective way because aromatic molecules of DOC produce
vibronic excitations under certain wavelengths of UV
(Khan et al., 2014). For example, naphthalene or anthrathene has a UV absorbance at 270 nm. Additionally, the
absorption of radiation corresponds to functional groups
attached to an aromatic ring, and this can include -OH
group at 270 nm, -OCH3 at 269 nm, -CN at 271 nm,
-CO2 at 268 nm, and -COOH at 273 nm (Williams and
Flemming, 1980). Therefore, UV absorbance measurement at 254 nm is usually used as a substitute to estimate organic carbon content in the water (Najm et al.,
1994; Eaton, 1995). However, the value of absorbance at
254 nm particularly depends on the concentrations of
humic acids in water. When the concentration of humic acids is low, the UV254 may be too low and generate a comparatively high random error (Wang and Hsieh,
Vol. 39 No. 5

2001). Therefore, Khan et al. (2014) developed a correlation with very high coefficient (R2 = 0.98) between UV
272 nm absorbance and concentration of DOC of a sample that had been passed through a filter with 0.45 m
pores (Knap et al., 1996). The DOC includes purgeable
(volatile) organic compound (VOC) and non-purgeable
organic carbon (NPOC).
Lumbriculus variegatus, better known as blackworms
or California blackworms, are freshwater oligochaetes
that live in the sediment and silt of ponds and lakes. A
normal behavior for L. variegatus is clumping if several blackworms are together, they will intertwine their
bodies into a ball. In nature, numerous lumbriculus congregate and form large clumped colonies of up to 2,000
individual worms. This allows for increased metabolism, stabilization of body temperatures, effective gas
exchange, and collaborative food sharing (Leppanan and
Kukkonen, 1998). Moreover, clumped worms burrow
their anterior ends in the sediment and extend their posterior up into the overlaying water. This orientation not only
allows for effective gas exchange via dorsal blood vessels,
but also allows for rapid response upon stimuli, to avoid
dangers such as predation and acute toxicity (Drewes and
Fourtner, 1989; Drewes, 1999; Ding et al., 2001).
L. variegatus is an important component of freshwater food webs in Europe and North America because
they feed on microorganisms and organic material, and
is a major food source for fish. Volatile organic chemicals from deposited plastics end up in sediment of lakes
and rivers through wind and rain run-off. These toxins
diffuse through Lumbriculus skin and bio-accumulates
in the blackworm, then in its predators and eventually in
humans. As a result the health of L. variegatus is a vital
indicator of the current and future health of the ecosystem.
This research was conducted to compare effects of
VOCs in both PEVA and PVC on the oxygen intake,
activity and social behavior of the oligochaete Lumbriculus variegatus to determine whether PEVA is toxic and
whether it is a healthy alternative to PVC, and to provide
further information about the health and ecological risk of
these plastics.
MATERIALS AND METHODS
Preparation of PEVA and PVC solutions
To prepare the solution containing volatile organic compounds from PEVA plastics, twenty squares measuring
2 inches by 2 inches were cut from a 100% PEVA plastic
shower liner. 200 mL of distilled water was poured into
a water bath, and then the pieces of plastic were even-

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VOCs of PEVA plastic are toxic

ly spread out in the water. The cover of the water bath

was sealed with tape to prevent gases from escaping. The
water bath was heated to 66 degrees Celsius, and kept at
this temperature for two hours, and then was turned off
and left overnight to allow the VOCs condense. The same
procedure was repeated to obtain the solution containing
VOCs from a 100% PVC plastic shower liner.
Measuring concentrations of DOC in six
solutions
Six solutions, including one distilled water, one tap
water, the PEVA solution, the PVC solution, one lake
water (from North Carolina, USA), and one pond water
(from North Carolina, USA), were separately filtered
with sterile and endotoxin-free polyethersulfone filters
with 0.45 m pores (Whatman TM), and then read UV
absorbance at 272 nm with NanoDrop ND-1000 V3.3.0
(Thermo Scientific). The settings were as follows: UVVis, 1.0 Max absorbance, Normalize ON, and HiAbs ON.
Next, the concentrations were calculated with the following equation:
DOC (mg/L) = 518.93 Absorbance (272 nm) + 1.065
(Khan et al., 2014)
Obtaining and maintaining L. variegatus
L. variegatus from the same source was purchased
from Carolina Biological Supply Company. Each of
these worms was at a young age just two weeks old
and measured 1 to 2 inches long. It was not necessary to
account for gender in the experiment because each worm
has both male and female sex organs, and reproduces
through asexual fragmentation. Also, it was not necessary
to feed the worms because the blackworms are advantageous at long-term exposure without feeding (Sardo
and Soares, 2010) and this experiment did not require
the worms to be maintained for an extended period. The
experimental condition for these worms was at room temperature (25C) in PEVA, PVC or distilled water.
Determining what concentrations of solutions
are used
A LD50 test was performed prior to the experiment to
determine the toxicity of both PEVA and PVC plastics as
well as the appropriate concentrations to use in the actual
experiment. When establishing a LD50 test, a serial dilution is used to drop the concentration of the substance to
levels of interest. In this experiment, the concentrations
100%, 10%, 1%, 0.01% and 0.001% of the stock solution
were desired. Thus, the five desired concentrations were
diluted and poured into five dishes, and then ten blackworms were put in each dish.

After 24-hr, a blackworm died in the 100% PEVA

solution and another blackworm died in the 100% PVC
solution. The blackworms in the 100% solutions did not
clump and tended to be less active than the ones in the
diluted solutions. Although the mortality rates of Lumbriculus during the LD50 were very low, the VOCs in
the 100% solutions did prove to have adverse effect, so
the 100% stock solutions from both PEVA and PVC were
chosen to be used in the experiment.
Measuring oxygen intakes
Since the goal of this experiment was to test consequence when Lumbriculus variegatus was exposed to volatile organic compounds in both PVC and PEVA, three
exposures in different lengths were tested: one day, four
days, and eight days. Each length of exposure had three
petri dishes: PEVA (that is, in PEVA solution), PVC (in
PVC solution), and the control (in distilled water). Using
a graduated cylinder, each petri dish of one exposure was
filled with 15 mL of solution of PEVA, PVC or distilled
water, and then 10 blackworms were transferred into it.
The blackworms in each petri dish were transferred to
an appropriate container using a funnel 24-hr before the
due time of an exposure. Then the opening of the container was sealed with an oxygen probe. The amount of oxygen in the container was measured for 24-hr with a Vernier oxygen gas sensor. After measuring the oxygen, the
blackworms in each container were transferred back into
their original petri dish, and then the solution was sucked
out with a pipette and replaced with 15 mL of distilled
water, and then transferred back into the container again.
The oxygen during blackworms recovery was monitored
for 24-hr to see whether the effects of the exposure to
PEVA or PVC were reversible.
The oxygen intakes were calculated after the amounts
of remaining oxygen (in ppm) in the container were measured with the Vernier oxygen gas sensor. That is, an oxygen intake at a specific time was determined to be equal
to 210,000 ppm (the oxygen concentration in the empty
container) minus the amount of remaining oxygen at that
time. The ppm was then converted to mg by multiplying
the containers total volume (317 mL).
Observations of activity, mortality, and social
behavior
Observations of activity, mortality and clumping were
recorded after each exposure and each recovery period. Activity was measured as follows: Blackworms in
a control group were gently probed with a pipette and
their reaction was recorded as normal. Activity rating
ranged from 1 to 3, where 1 = inactive (the worm did
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T.T. Meng

not react with one touch), 2 = normal activity level (the

worm reacted like the control group with one touch), and
3 = very active (one touch triggered others to react, too).
Interpreting data
After the oxygen intakes were calculated, the trends of
the oxygen intakes in each case and their lines of best fit
were graphed with Microsoft Excel. The slope of the best
fit line was the oxygen change rate per hour (mg/hr),
which reflected the health effects when the worms were
exposed to VOCs. The similar analysis was done on the
data from each of the recovery periods.
RESULTS
The UV absorbance of the six solutions (one distilled
water, one tap water, the PEVA solution, the PVC solution, one lake water, and one pond water) at 272 nm were
as follows: 0.005, 0.009, 0.012, 0.021, 0.023, and 0.015,
and their estimated concentrations (mg/L) were as follows: 3.66, 5.74, 7.29, 11.96, 13, and 8.85.
For the one day exposure, the oxygen intakes of both
PEVA (average 9,250 mg) and PVC (23,529 mg) groups
were much higher than the distilled water group (3,041
mg) (see Fig. 1). Furthermore, the high oxygen intakes of
both PEVA (average 11,729 mg) and PVC (17,891 mg)
groups did not fully recover after a 24-hr healing period
in distilled water (3,205 mg) (see Fig. 2). The same was
observed from oxygen intake data in four day exposure,
eight day exposure and their 24-hr recovery periods (see
Table 1).
Generally, the PEVA group had more fluctuations
than the control and PVC groups (see Figs. 2 and 3). The

control and PVC groups had relatively constant oxygen

intakes.
The change in oxygen intake (mg/hr) per hour is
the slope of the best fit line. During the exposures (see
Figs. 1, 3, 5), the slope of the control (distilled water) first
decreased from one day exposure 29.06 to four day exposure -19.74 then remained constant from four day to eight
day exposure. The mg/hr of PVC first decreased from
27.91 to 2.87 then increased from 2.87 to 47.59. Meanwhile, the mg/hr of PEVA increased from 21.38 to 96.94
then decreased from 96.94 to 9.99 (see Table 2).
When comparing the mg/hr of exposure to recovery, PEVAs rate of oxygen intake first increased in one
day exposure and subsequent recovery (from 21.38 to
106.90), then decreased in the four day set (96.94 to 1.36)
and the eight day set (9.99 to 2.69). PVCs mg/hr (27.91
to 19.47, 2.87 to -38.26, 47.59 to 25.23) decreased from
exposure to recovery. The control (29.06 to -12.72, -19.74
to 14.05, -19.92 to 4.57) decreased then increased (see
Table 2).
Additionally, the blackworms did not clump together tightly after four or eight day exposure to either of the
two plastic solutions (Table 3). The number of worms
clumped together mainly decreased with greater time
of exposure. Recovery periods generally had greater
worms clumping together than the exposure period, but
the worms in the PEVA and PVC solutions did not fully
recovery (see Table 3).
The control groups during all periods of exposure and
recovery had normal activity. The activity level of PEVA
and PVC was initially higher than normal and decreased to
below normal by the eight day of exposure (see Table 4).
During all the exposure and recovery periods, all of

One
Day
Exposure
One
Day
Exposure

One Day Recovery

32000

32000
Oxygen intakes (mg)

y = 27.91x + 23194

24000
20000
Control

16000
y = 21.38x + 8993.5

12000

PEVA
PVC

8000
y = 29.06x + 2692.2

4000

Oxygen intakes (mg)

28000

28000

24000
20000

y = 19.47x + 17657
Contol

16000

y = 106.9x + 10445

PVC

8000
y = -12.72x + 3357.8

4000
0

0
0

10 12 14 16 18 20 22 24

One day exposure. The equation for the trend line is

above each curve. The trend line does not show up if it
completely overlaps with its corresponding curve. The
same rules apply thereafter.

Vol. 39 No. 5

10

12

14

16

18

20

22

24

Time (hr)
Time (hr)

Time
(hr)
Time
(hr)

Fig. 1.

PEVA

12000

Fig. 2.

One day recovery. The dash line crossing the PEVA

curve is its trend line.

799
VOCs of PEVA plastic are toxic

Table 1. Average Oxygen Intake (mg)

Table 2. Change in Oxygen Intake (mg/hr)

Exposure Avg. Oxygen Intake (mg)

Exposure (mg/hr)

Length of Exposure

Control

PEVA

PVC

3,041

9,250

23,529

One Day

Four Days

2,719

10,322

23,271

Eight Days

2,420

8,056

16,823

One Day

Recovery Avg. Oxygen Intake (mg)

Length of Exposure

Length of Exposure

Control

PEVA

PVC

29.06

21.38

27.91

Four Days

-19.74

96.94

2.87

Eight Days

-19.92

9.99

47.59

Length of Exposure

Control

PEVA

PVC

Recovery (mg/hr)

Control

PEVA

PVC

One Day

3,205

11,729

17,891

One Day

-12.72

106.9

19.47

Four Days

3,404

8,192

22,893

Four Days

14.05

1.36

-38.26

Eight Days

2,448

8,213

17,513

Eight Days

4.57

2.69

25.23

Table 3. The Amount of Clumping Worms

Table 4. Activity Level

Exposure (# Clumping)

Exposure (Activity Level)

Length of Exposure

Control

PEVA

PVC

Length of Exposure

Control

PEVA

PVC

One Day

10/10

8/10

9/10

One Day

2.5

Four Days

9/10

5/10

6/10

Four Days

2.5

Eight Days

8/10

2/10

5/10

Eight Days

0.5

Control

PEVA

PVC

Control

PEVA

PVC

Recovery (# Clumping)
Length of Exposure

Recovery (Activity Level)

Length of Exposure

One Day

10/10

9/10

10/10

One Day

1.5

1.5

Four Days

9/10

7/10

6/10

Four Days

1.5

Eight Days

9/10

6/10

4/10

Eight Days

0.5

Note: 1 = inactive, 2 = normal activity, and 3 = very active

the blackworms were respiratory (that is, their tails curled

and were out of the water) and none of the blackworms
died under observation.
DISCUSSION
Oxygen consumption is an effective parameter
of physical stress
Measure of oxygen consumption is an indirect but
common technique to measure energy expenditure
(Levine, 2005) because most of energy in the body of an
organism is produced aerobically, and monitoring energy
expenditure is a standard method to measure the effects of
stressors in aquatic organisms (Giesy, 1988). Therefore, it
is reasonable to measure oxygen intakes of the worms to
indicate the stress that they endure. Actually, the rate of
respiration/intake of oxygen of earth-dwelling worms has
been shown to increase by introducing stress such as light
stimuli (Billstein et al., 2002). Moreover, direct measure-

ment of exercise respiratory gas exchange is a valuable

and standard testing of cardiopulmonary exercise stress in
patients (Crespo et al., 2009).
As Giesy (1988) stated, the methods to measure the
effects of stressors in aquatic organisms can be placed into
six categories: 1) Hormones; 2) Energetics; 3) Enzyme
activities; 4) Osmoregulatory electrolytes; 5) RNA/DNA
and protein content; and 6) other substrates. Obviously, measuring oxygen consumption belongs to the second category, and measuring heat shock protein or immediate-early gene products falls into the fifth category. It
is no doubt that all of them, including oxygen consumption, induction of heat shock proteins, induction of immediate-early gene products, hormones, and osmoregulatory
eletrolytes, are effective parameters to measure stress.

Vol. 39 No. 5

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T.T. Meng
Four Day Recovery

Four Day Exposure

32000

32000
28000
y = 2.87x + 23237

24000
20000

Control

16000

y = 96.94x + 9159.1

12000

PEVA
PVC

8000
y = -19.74x + 2955.5

4000

Oxygen intakes (mg)

Oxygen intakes (mg)

28000

y = -38.27x + 23253

24000
20000

Control

16000
12000

y = 14.05x + 3271.9

0
0

10 12 14 16 18 20 22 24

Time
(hr)
Time
(hr)

Four day exposure. The dash line crossing the PEVA

curve is its trend line, and the thin line overlapping the
control curve is its trend line.

Fig. 4.

28000

28000

24000

24000

y = 47.59x + 16252

16000

Control
PEVA

y = 9.99x + 7935.7

PVC

Oxygen intakes (mg)

Oxygen intakes (mg)

14

16

18

20

Eight Day Recovery

32000

12000

12

Four day recovery. The dash line overlapping the

PEVA curve is its trend line, and the thin line overlapping the control curve is its trend line.

32000

20000

10

Time
(hr)(hr)
Time

Eight Day Exposure

8000

20000

y = 25.23x + 17210
Control

16000

PEVA
12000
y = 2.69x + 8180.6

PVC

8000

y = -19.92x + 2659.1

4000

4000

y = 4.57x + 2393.2

10 12 14 16 18 20 22 24

Eight day exposure. The dash line overlapping the

PEVA curve is its trend line, and the thin line overlapping the control curve is its trend line.

Both solutions stressed the worms and

irreversibly increased their oxygen intakes
Immediate responses of the blackworms to VOCs were
observed at the LD50 test. That is, a blackworm died in
both the original PEVA stock solution and the original
PVC stock solution. Furthermore, the blackworms in the
stock solutions did not clump and tended to be less active
than the ones in the diluted solutions. Therefore the LD50
test demonstrated that the greater the concentrations, the
more toxic were the effects of the VOCs, as is consistent with toxicity studies while treated blackworms with
caffeine, nicotine, etc. (Bohrer, 2006), and with extracts
from various hosta varieties (Rivera and Myers, 2012).
And, one day, four day and eight day exposures were
Vol. 39 No. 5

10 12 14 16 18 20 22 24
Time (hr)
(hr)
Time

Time
Time (hr)
(hr)

Fig. 5.

PVC

8000
4000

Fig. 3.

PEVA

y = 1.36x + 8179.7

Fig. 6.

Eight day recovery. The dash line overlapping the

PEVA curve is its trend line.

compared to see the effect of VOCs on the oxygen intake

of L. variegatus. PVC had the highest oxygen intake in
all of the graphs (total average 15,605 mg), however,
PEVA (9,209 mg) still had very high intake compared to
the control (2,726 mg) (see Figs. 1-6). Therefore, these
results clearly indicated that PEVA, like PVC, contained
toxins that stressed L. variegatus and thus increased their
oxygen intakes.
The 24-hr recovery period after the exposure represented detoxification. When comparing exposure to recovery, there was not much difference in the average oxygen
intake. The recovery data showed that complete recovery did not occur within the given time period (as shown
from the avg. oxygen intake and clumping). These results

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VOCs of PEVA plastic are toxic

led to the conclusion that the adverse effects of exposure

are irreversible in a 24-hr period.
In the exposures (see Figs. 1, 3, 5), the slopes of the
control (distilled water) first decreased then remained
constant, which indicated that the blackworms adjusted to the new environment and then continued to take
in the same amount of oxygen. The mg/hr of PVC first
decreased then increased. Similarly, Billstein et al. (2002)
showed that when earth-dwelling worms were exposed
to stress such as light stimulus, their oxygen intake level increased. Thus, the longer blackworms were exposed
to PVC, the more stressed they became. However, mg/
hr of PEVA first increased then decreased significantly
(96.94 mg to 9.99 mg), perhaps because the harmful compounds of PEVA, which affected the ability of L. variegatus to aspirate oxygen, were different from the toxins of
PVC.
Both solutions adversely affected the worms
behavior
As exposure length increased, the number of worms
clumping together decreased. Activity level of worms in
both PEVA and PVC solutions was initially higher than
normal and then decreased to below normal by the eight
day of exposure, which meant that the stress excited the
blackworms body systems, and then weakened their ability to respond. Therefore, there was no doubt that increases in concentration and exposure length to either PEVA
or PVC solution negatively affected blackworms social
behavior. A similar phenomenon was observed by other
researchers: Bohrer (2006) noticed that the blackworms
were not or less clumping while living in water treated
with caffeine or nicotine.
Experimental conditions were well controlled
Not all of the blackworms at each exposure length
demonstrated the same behavior (see Table 3, Table 4).
This can be explained by variability or differences among
the worms. Both extrinsic and intrinsic factors may affect
results of experiments. However, extrinsic factors, such as
temperature and pressure, were controlled via conducting
the experiment in standard room conditions. Intrinsic factors are within individual organisms, such as age, metabolism, and genetic difference. In the experiment, L. variegatus picked for each group were selected to be all roughly
the same length and coloring, thus age and body size was
controlled. Still, genetic difference between blackworms
could have affected results of exposure to VOCs. However, the genetic difference among the worms should be
small since all of them were fresh young, were from the
same source at the same time, and were almost the same

size.
Toxic organic compounds are potential health
risk to human and other animals
UV absorption of organic solutes is directly proportional to their concentration in aromatic compounds (Traina
et al., 1990; Chin et al., 1994), and thus measuring UVVis absorbance is a convenient and effective way to estimate concentrations of DOC in water (Najm et al., 1994;
Eaton, 1995; Khan et al., 2014).
The DOC of PVC solution is lower than that of the lake
water (11.96 mg/L vs. 13 mg/L), and higher than that of
the pond water (11.96 vs. 8.85). The DOC of PEVA solution is close to that of the pond water (7.29 vs. 8.85), and
higher than the tap water (7.29 vs. 5.74). These data indicate that toxic organic compounds are possibly accumulated in the environment such as lakes, ponds etc., representing a potential health risk to human and other animals.
Further steps will include determining what components
in the two plastics are toxic to the blackworms, and how
much they have accumulated in the ecosystem.
As a side note, the concept Dissolved Organic Matter
(DOM) in the reference Khan et al. (2014) is the same
as the concept Dissolved Organic Carbon (DOC) in this
article because of two reasons: Firstly, samples in the two
studies were passed through filters with 0.45 m pores
and then were measured UV-Vis absorbance. Secondly,
the practical definition of DOC is the "dissolved" fraction of organic carbon below 0.45 m (Knap et al., 1996),
which includes VOC and NPOC.
In summary, this research clearly showed that PEVA
has toxic effects on the oxygen intake, social behavior and
activity of Lumbriculus variegatus, even though it is chlorine-free. And, this research proves that there is a need
for further study on the toxicity of VOCs in PEVA plastic, especially to see whether it causes cancer in humans.
In the near future, further studies should determine what
compounds caused these adverse effects, how much they
have accumulated in the ecosystem, and whether the toxic effect occurs in more complex organisms, especially
humans.
ACKNOWLEDGMENTS
This research was supported by North Carolina
Summer Ventures in Science and Mathematics, and River
Center at Duke University. Many thanks to Josh Cannon
for instruction and discussion, and to Dr. Christine Muth,
Brooke Hassett and Chelsea Clifford for their help.

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802
T.T. Meng

Conflict of interest---- The authors declare that there is

no conflict of interest.
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