2002_k.vorkamp_fate of methidathion residues in biological waste during anaerobic digestion

8/10/2019 2002_K.vorkamp_Fate of Methidathion Residues in Biological Waste During Anaerobic Digestion

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Fate of methidathion residues in biological waste

during anaerobic digestion

Katrin Vorkamp *, Esther Kellner, Joorg Taube, Kai D. Mooller,Reimer Herrmann

Department of Hydrology, University of Bayreuth, 95440 Bayreuth, Germany

Received 15 August 2001; received in revised form 24 January 2002; accepted 14 February 2002

Abstract

The aim of this study was to examine the fate of the organothiophosphorus pesticide methidathion during anaerobic

digestion of biological waste. Three reactor experiments were conducted under various conditions of temperature, pH

and retention time. The influence of pH and temperature as well as the partitioning between solid and aqueous phase

were studied in batch experiments. The mesophilic (25, 35 C) reactor experiments showed a decline to about 10% of the

maximum methidathion concentration within 3080 d. In the thermophilic (55 C) reactor experiment, methidathion

disappeared within 20 d. The batch experiments showed an abiotic hydrolysis of methidathion over the experiment

period of 4 d, accelerated by alkaline conditions (pH 10.5 and 12.8) and high temperatures (55 C). The hydrolysis was

also noticeable at a neutral pH, while methidathion was most stable at weakly acid pH values. Methidathion bonded

strongly to the biological waste, and the amount released into the water phase was below the maximum aqueous

solubility. About 10% of methidathion remained non-extractable. High concentrations of dissolved organic carbon andyeast extract as a model substance for disintegrated cells further reduced the content of methidathion in the water

phase, possibly caused by co-sorption to the solid organic matter. 2002 Elsevier Science Ltd. All rights reserved.

Keywords:Anaerobic digestion; Biological waste; Organothiophosphorus compounds; Pesticide residues; Methidathion; Hydrolysis

1. Introduction

Since enactment of a German law in 1993, waste

disposal has been limited to waste containing less than

5% organic matter (TA Siedlungsabfall, 1993). There-fore, biological waste has been collected separately in

order to be composted or anaerobically digested and to

be turned into reusable compost. Biological waste in-

cludes all organic garden and kitchen wastes, which add

up to an annual amount of 4:9 1067 106 t in Ger-many (Scherer, 1995).

Besides its role in a sustainable waste management,

the anaerobic digestion of organic matter has also been

acknowledged as a source of renewable energy, yieldingbiogas containing 5070% methane (Wellinger et al.,

1991). Depending on the preferred group of microor-

ganisms, anaerobic digestion processes are conducted

mesophilically or thermophilically. Generally, mesophi-

lic microorganisms grow between 8 and 45 C, with an

optimum growth temperature of about 35 C. The

growth of thermophilic organisms requires temperatures

above 42 C, ideally 60 C.

In accordance with agricultural practice, which aims

at maximum crop yield, pesticides are commonly used

for crop protection. The Food and Agricultural Orga-

nization of the United Nations (FAO) estimates the

Chemosphere 48 (2002) 287297

www.elsevier.com/locate/chemosphere

* Corresponding author. Present address: Department of

Environmental Chemistry, National Environmental Research

Institute, Frederiksborgvej 399, 4000 Roskilde, Denmark. Fax:

+45-46-301114.

E-mail address: [email protected] (K. Vorkamp).

0045-6535/02/$ - see front matter 2002 Elsevier Science Ltd. All rights reserved.

PII: S0 0 4 5 -6 5 3 5 (0 2 )0 0 0 9 5 -4


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worldwide amount of pesticides applied to be more

than 3 106 t a1. Previous studies have shown thatvarious components of biological waste, such as citrus

fruit peels, vegetables and flowers, contain high con-

centrations of pesticides, including the insecticide

methidathion (Vorkamp et al., 1997, 1999; Foorster et al.,

1999).Methidathion (S-2,3-dihydro-5-methoxy-2-oxo-1,3,4-

thiadiazol-3-ylmethyl-O,O-dimethyl-phosphorodithioate)

is a thioester of thiophosphoric acid (Fig. 5) and belongs

to the group of organophosphorus pesticides. Its water

solubility at 20 C is 240 mg l1 and the logKOW is 2.22

(Richardson and Gangolli, 1993). The vapor pressure at

20 C is specified as 1:87 101 Pa (Richardson andGangolli, 1993) and 1:33 104 Pa (Hayes and Laws,1991). Methidathion is a non-systemic insecticide and

acaricide which has been in use since 1966 to control

sucking and chewing insects and spider mites on many

crops such as corn, fruits, vegetables, tobacco, cottonand sun flowers, in green houses and rose cultures. Its

toxic effect is based on the inhibition of choline esterase

activity (Hayes and Laws, 1991). Methidathion is con-

sidered very toxic to human beings (Richardson and

Gangolli, 1993) as well as to birds, bees and aquatic

organisms (Hartley and Kidd, 1987; Flammarion et al.,

1996).

In order to achieve high quality compost products,

pesticides in biological waste should be degraded during

the treatment process. No residues should remain in the

digested waste which will be used as a fertilizer in agri-

culture. Chemical hydrolysis and microbial degradation

are two important degradation pathways for organo-

phosphorus pesticides (Eto, 1977; Matsumara, 1982).

Hydrolysis pathways have been proposed for phospho-

ric and thiophosphoric esters, with acid, base or neutral

species serving as catalysts (Schwarzenbach et al., 1993).

According to Hartley and Kidd (1987), methidathion is

rapidly hydrolyzed under alkaline and extremely acidic

conditions, but rather stable in weakly acid and neutral

surroundings. Microbial degradation by the soil bacte-

riumBacillus coagulanswas described by Gauthier et al.

(1988).

The aim of this study was to examine and to evaluate

the fate of methidathion in biological waste during bothmesophilic and thermophilic anaerobic digestion pro-

cesses. The effects of pH and temperature and the par-

titioning between solid and aqueous phase were further

studied in batch assays.

2. Materials and methods

2.1. Bioreactor

The fate of methidathion during anaerobic digestion

was investigated in a stainless steel bioreactor (Fig. 1).

This bioreactor had a total volume of 300 l and an op-

erating volume of 240 l. It was completely sealed by

o-rings and shaft seals to maintain anaerobic conditions.

A content of 5% dry matter was chosen for the experi-

ments. The waste pulp was mixed by a propeller driven

by a transmission engine (Fig. 1). In order to investigate

aerobic conditions, air could be introduced into the bio-reactor. A heating system was used for temperature

control.

2.2. Synthetic biological waste matrix

Since the composition of biological waste differs both

temporally and spatially (Oetjen-Dehne and Tidden,

1991), an artificial biological waste matrix was used as

described by Vorkamp et al. (1997). This matrix con-

tained 82.1% laboratory grade rabbit food, 14.6% cel-

lulose powder and 3.3% paper. The rabbit food andcellulose powder represented kitchen waste and wooden

waste, respectively. The ingredients were ground and

after that mixed according to their percentages. Ap-

proximately 12 kg of the mixed material was added to

the bioreactor.

2.3. Preparatory work

Methidathion was extracted from the commercial

product Ultracid 40 (40% methidathion, Ciba-Geigy,

Frankfurt/Main, Germany). Twenty-five g of Ultracid

40 was mixed with about 500 ml of purified water

(Millipore, Eschborn, Germany). To this mixture, 500

ml cyclohexane (nanograde, Geyer, Renningen, Ger-

many) were added and stirred for 24 h at room tem-

perature. After separation of the water and the solvent

phase, 200 ml cyclohexane were added to the water. The

mixture was extracted ultrasonically for 15 min. The

water and the solvent phase were separated, and the ul-

trasonic extraction was repeated. Finally, the solvent

phase was dried over anhydrous Na2SO4 and concen-

trated by rotary evaporation (40 C, 270 mbar). The

extracts purity was determined by gas chromato-

graphicmass selective analysis (GCMS) in comparisonto a standard substance and found to be above 97%.

The concentrated extract was dissolved in acetone

(nanograde, Geyer, Renningen, Germany). For the three

reactor experiments, 2.035, 2.086 and 2.640 g of meth-

idathion were added to about 500 g of the synthetic

biological waste matrix, which was constantly stirred.

After the solvent had vaporized, the biological waste

was mixed with the remaining 11.5 kg and filled into the

bioreactor, which was then filled up with tap water to a

volume of 240 l. The sealing of the reactor and the si-

multaneous start of the transmission engine were con-

sidered the beginning of the experiment.

288 K. Vorkamp et al. / Chemosphere 48 (2002) 287297


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2.4. Reactor experiments

The fate of methidathion was examined during three

anaerobic digestion experiments, followed by a short

aerobic stabilization. The three experiments differed in

temperature, pH and retention time (Table 1). The

temperature for the first, second, and third experiment

was maintained at 25, 35, and 55 C, respectively.

The hydrolytic and the acidogenic phases of the an-

aerobic digestion process began spontaneously. The

methanogenic phase was deliberately induced by an in-

crease in pH and the addition of methanogenic sewage

sludge as an inoculum. In the thermophilic experi-

ment, the inoculum consisted of a mixture of sewage

sludge and the synthetic biological waste incubated at

55 C until methane was produced. The NaOH and

Table 1

Characterization of the reactor experiments

Experiment Temperature (C) Duration (d) pH-stabilization

R1 25 33 12.5 d: 800 g NaOH (solid)

27 d: 420 g NaOH (solid)

R2 35 121 12.5 d: 3.4 l NaOH (dissolved, 12.5 M)

76 d: 500 g NaOH (solid)

R3 55 71 8.5 d: 1 l NaOH (dissolved, 12.5 M)

28 d: 200 ml NaOH (dissolved, 12.5 M)

40 d: 250 ml NaOH (dissolved, 12.5 M)

50 d: 25 g NH42CO3 (solid)53 d: 500 ml NH42CO3 (dissolved, 5.2 M)

Fig. 1. Construction of the bioreactor for anaerobic digestion experiments.

K. Vorkamp et al. / Chemosphere 48 (2002) 287297 289


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NH42CO3 solutions (Table 1) were added at a rate of70 mlh1. When the methane production declined, air

was introduced into the reactor to convert to the aerobic

phase. The aerobic phases started after 29, 111 and 64 d,

respectively. A suspension of 10 g compost in 500 ml

water was used as an aerobic inoculum.

The process was monitored by continuously mea-suring pH value, redox potential, temperature and the

amount of gas produced. The composition of the reactor

gas and the concentrations of short-chained volatile

fatty acids (VFA) (acetic acid, propionic acid, n- and

iso-butyric acid, n- and iso-valeric acid and capronic

acid) were analyzed daily. The gas concentrations were

analyzed using gas chromatographic separation and

detection by thermal conductivity (GCTCD). The con-

centrations of the gases were determined from an ex-

ternal calibration using different injection volumes of a

gas standard (AGA, Meitingen-Herbertshofen, Ger-

many), which was comprised of 60% CH4, 30% CO2, 5%N2 and 5% O2. Gas samples were taken through a sep-

tum in the top of the bioreactor using a 1 ml-syringe.

VFA were determined using gas chromatographic sep-

aration and flame ionization detection (GC-FID). They

were identified according to their retention times and

quantified on the basis of an internal standard (1-

methylvaleric acid). A detailed description of the ana-

lytical method for VFA is given by Vorkamp et al.

(2001).

Samples for the analysis of VFA and methidathion

were taken daily from the valve at the bottom of the

bioreactor. Prior to sampling, the stirrer was set to

maximum propelling for 5 min to ensure full homoge-

neity and thus representative sampling. The volume

of the valve was accounted for by discarding the first

150 ml of the sample. The following 350 ml were mixed

with 1 ml of a biocide-mixture (actidione, chloramphe-

nicol, streptomycin, tetracycline, 5 mg ml1 each) (Merck,

Darmstadt, Germany), dissolved in water:methanol

(nanograde, Geyer, Renningen, Germany, 10:1, v:v). The

samples were frozen at 20 C until processing.

2.5. Batch experiments

2.5.1. Effects of pH

The effects of pH on the fate of methidathion were

examined at six different pH values in assays with and

without biological waste (Table 2). The assays without

biological waste contained pure buffer solutions. In the

assays with biological waste, the waste was mixed with

buffer solutions. The pH values mainly occurring in the

anaerobic digestion experiments are between 4.1 and 8.5.

However, the addition of NaOH can lead to a short raise

in pH until the reactor is fully mixed. Therefore, the pH

range was extended to pH 10.5 and 12.8. The experi-

ments were carried out at 25

C in triplicate with three

blanks, which contained the corresponding amounts of

solvent.

For the assay without biological waste, 200 lg of

methidathion in acetone were added to 20 ml of each

buffer solution. In order to inhibit microbial degradation

of the insecticide, the buffer solutions also contained

0.05 ml of the biocide-solution. Half of the assays wereanalyzed after short, manual shaking (t0 30 min). Theother half were analyzed after four days (t1 97 h) ofreciprocal shaking at 150 rpm.

For the assay with biological waste, 500 lg of

methidathion in an equal volume mixture of cyclohex-

ane and ethylacetate (nanograde, Geyer, Renningen,

Germany) were added to 2 g of the synthetic waste.

After the solvent had vaporized at 4 C, 50 ml of each

buffer solution and 0.1 ml of the biocide-mixture were

added. After t0 30 min, half of the samples were cen-trifuged and analyzed. The other half were analyzed at

t1 97 h.

A Student-t-test was used to compare the concen-

trations of methidathion. Whether concentrations were

significantly different was tested between the two sam-

pling points, among the different pH values and for the

results of the two assays. The permitted type I error was

5%. As the concentrations could vary to both sides, a

two-tailed test was applied. The t-test assumes normal

distribution, which was previously tested by a Kol-

mogorowSmirnow-test for n 10 replicates. Accord-ing to Markowski and Markowski (1990), the t-test is

insensitive towards heterogeneity of variances, as long as

the sample sizes are equal, making preliminary tests of

homogenous variances unnecessary.

2.5.2. Effects of temperature

The effects of different temperatures on the fate of

methidathion were examined in a similar manner as the

pH experiments, including three replicates and one

blank. 142 lg of methidathion, dissolved in cyclohexane

and ethylacetate was added to 3 g of the synthetic waste.

After the solvents had vaporized at 4 C, 40 ml of water

and 1 ml of the biocide-solution were added. Half of the

samples were stirred at 55 C, and the other half were

stirred at 25 C. Three samples and one blank from each

treatment were analyzed daily.

Table 2

Selected buffer solutions and resulting pH values

Buffer 1 Buffer 2 Ratio Result-

ing pH

0.1 M citric acid 0.1 M HCl 56:44 4.1

0.1 M K2HPO4 0.1 M KH2PO4 5:95 5.6

0.1 M K2HPO4 0.1 M KH2PO4 61:39 7.0

0.1 M K2HPO4 0.1 M KH2PO4 98:2 8.5

0.1 M glycine 0.1 M NaOH 54:46 10.5

0.1 M glycine 0.1 M NaOH 7.5:92.5 12.8



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2.5.3. Partitioning of methidathion

It was examined whether the composition of the

aqueous phase affected the partitioning of methidathion

between the biological waste and the water phase. Three

assays with different water ingredients were compared:

process water from the methanogenic phase of the an-

aerobic digestion experiments, yeast extract solution andpure water (Millipore) as reference. In each assay, the

concentration of methidathion in the water phase was

determined over a certain period of time. The equilib-

rium concentrations of the single assays were compared

for significant differences. All assays were conducted in

triplicate with one blank.

Twenty-five g of biological waste was spiked with

10.90 mg of methidathion, dissolved in cyclohexane and

ethylacetate. After vaporization of the solvents at 4 C,

495 ml of the respective solution and 5 ml of the bio-

cide-solution were added. The samples were turned

throughout the experiment. After previously determinedperiods of time, samples were taken through the Teflon

septum in the cap of the containers. The samples were

centrifuged at 4000 rpm for 20 min, and 4.5 ml of the

water phase were extracted as described below.

The process water was taken from the methanogenic

phase of an anaerobic reactor experiment, which did not

contain pesticides. The water was centrifuged at 5000

rpm for 20 min. The supernatant was further centrifuged

at 9500 rpm for 25 min and filtered through 0.45 lm

cellulose nitrate filters (Sartorius, Goottingen, Germany).

The pH of the final supernatant was 8.2, the electric

conductivity was 14.8 mS cm1, and the dissolved or-

ganic carbon (DOC) was 1822 mg l1. The yeast extracts

(Fluka, Neu-Ulm, Germany) were a mimic for the high

concentration of proteins likely to occur in the reactor

experiments and mainly consisted of amino acids, pep-

tides, vitamins and carbohydrates. The concentrations

of the yeast extract solutions were 2.02, 5.05 and 8.08

g l1.

The equilibrium concentration in the experiment with

process water was compared to the reference assay by

means of a two-tailed Student-t-test. The H0-hypothesis

was that the mean concentration at the last sampling

point was equal to the mean concentration in the ref-

erence assay. The three different assays of the experi-ment with yeast extracts and the reference assay were

analysed for significant differences between the mean

concentrations of the last sampling point by a one-fac-

torial analysis of variance (ANOVA). If the ANOVA

revealed significant differences, the T-method (Range

STP procedure) was applied to identify the significantly

different means. A minimum significant range (MSR) is

calculated and compared to the differences between

two means of the ANOVA. If the difference is larger

than MSR, the means are significantly different. The test

assumes equal sample sizes and allows a type I error of

5%.

Other experiments dealt with the transport of meth-

idathion through soil (Mooller et al., 1999). In this con-

text, batch experiments were conducted to examine the

effect of DOC on the equilibrium distribution of meth-

idathion between the aqueous phase and soil. A 103 M

solution of KClO4, buffered at pH 6, was spiked with

methidathion at a concentration of 387 lg l1. Eighty mlof the solution, containing the biocide-mixture, were

added to 10 g of soil and shaken for 10 h, until equi-

librium was reached. In a second assay, 1 ml of DOC

was added to 1 l of the solution. DOC was obtained and

treated as described above. The experiment was con-

ducted in triplicate; analysis followed the description

below.

2.6. Analysis of methidathion

The frozen samples of the reactor experiments werethawed at 4 C overnight. All other samples were pro-

cessed immediately after sampling. Unless otherwise

stated, the samples were centrifuged for 15 min at 4000

rpm (Labofuge Ae, Heraeus, Hanau, Germany) and

filtered (0.7 lm glass fiber filter, Whatman, UK) to

separate water and solid phase. Three aliquots of the

water phase were extracted by an equal volume mixture

of cyclohexane and ethylacetate. To avoid emulsions,

the volume of the organic solvent was about 2.5 times

the volume of water. The extraction was carried out in

three steps. The first extraction took place overnight on

a reciprocal shaker at 150 rpm. After separation of the

solvent and the water phase in a separating funnel, new

solvent was added to the water. In the second and third

extraction, the sample was shaken at 150 rpm for 15 min

each. The solvent phases were dried over anhydrous

Na2SO4 and concentrated by rotary evaporation (40 C,

270 mbar) and under nitrogen (Purity 5, Linde, Ger-

many) to a volume of about 500 ll.

The solid phase was freeze-dried (Modylo-4k Freeze

Dryer, Edwards, Marburg, Germany) and separated into

three samples of defined mass. The samples were ex-

tracted ultrasonically for 15 min using the same solvent

mixture of cyclohexane and ethylacetate. Three extrac-

tion steps were carried out with 50 ml of the solvent-mixture. The extracts were filtered through inactivated

glass wool and further processed as described for the

water phases.

Prior to analysis, a known amount of the internal

standard bromophos-ethyl (Promochem, Wesel, Ger-

many, >99% purity) was added to the samples. Thesamples were analyzed by gas chromatographic separa-

tion (model HP 5890) and mass-spectrometric detection

(GCMSD, model HP 5970). The GC was equipped

with a J&W Scientific DB-17 capillary column (30 m

long, 0.25 mm inner diameter, 0.25 lm film thickness).

Samples (1 ll) were injected in the splitless mode. The



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temperature program was as follows: increase from 70 to

150 C at a rate of 10 C min1, increase to 300 C at a

rate of 8 C min1, 5 min at 300 C. The GC injector

temperature was 250 C and the transfer line was at 290

C. The samples were analyzed in the selected ion

monitoring-mode. The m=z-ratio of the target ion was

145, the m=z-ratios of the qualifier ions were 125 and 58.For quantification, the peak areas of the target ions

of methidathion and bromophos-ethyl (m=z-ratio: 359)were used. Response factors were determined for each

analytical sequence from a series of five standards. The

concentration of methidathion in the water phase was

calculated by relating the amount detected in the sam-

ples to the volume of the aliquot. For the solid phase,

the amount was related to the mass of the sample ex-

tracted. The original water content of the samples was

determined by the weight difference before and after

freeze-drying. The concentration of methidathion in the

solid phase was corrected for the amount transferredfrom the water to the solid phase during freeze-drying.

Relative concentrations were calculated by relating the

actual concentrations to the maximum concentrations

governed by the amount of methidathion added to the

respective experiment. In case of the reactor experiment,

the maximum concentration of methidathion in the solid

phase changed throughout the experiment as the solid

organic matter was microbially degraded and solubilized

due to an increase in pH. This disappearance of the solid

organic matter was accounted for by subtracting the

amount of carbon measured as CO2 and CH4 and DOC

from the original amount of carbon in the reactors.

In order to determine the efficiency of the liquid

liquid-extraction, 22.13 lg of methidathion, dissolved in

acetone, were added to 100 ml of pure water and 20 ml

of process water, respectively. The ultrasonic extraction

was assessed by adding 221.25 lg of methidathion to 2 g

of the dry biological waste. Finally, 441.65 lg of meth-

idathion were added to the 2 g of dry matter, which wasmixed with 50 ml water after vaporization of the solvent.

All aqueous matrices contained the biocide-mixture. The

samples were extracted and analyzed as described above.

All assays were carried out in triplicate with one blank.

3. Results and discussion

3.1. Reactor experiments

The anaerobic digestion process proceeded in its

characteristic phases of hydrolysis, acidogenesis andmethanogenesis, as exemplified for the R3 experiment in

Fig. 2. In the hydrolytic phase, aerobic microorganisms

consumed the remaining oxygen and produced CO2,

while the pH decreased to about 4. The acidogenic phase

began when VFA (acetic acid, propionic acid, n-butyric

acid) were produced and the reactor gas almost exclu-

sively consisted of CO2. The pH would have been acidic

if it had not been raised by adding NaOH solution

(Table 1). CH4 could not be detected in the reactor gas

before NH42CO2 solution was added. As a conse-quence of CH4 production, the acetate concentration

decreased and the pH increased to above 8.

Fig. 2. pH, VFA and gases in the anaerobic digestion experiment R3.Iso-butyric acid, n- andiso-valeric (all


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Principally, the reactor experiments R1 and R2

showed the same course of pH, gas and VFA produc-

tion. However, the percentage of CH4 in the reactor gas

was lower. Consequently, less acetate was consumed and

the concentration did not decrease as clearly as shown

for R3 (Fig. 2). The accumulation of VFA is charac-

teristic for an inhibited methane production (McCartyand McKinney, 1961). The inhibition of methanogenesis

can be caused by a toxic effect of Na, as shown by

McCarty and McKinney (1961).

3.2. Effects of pH

Fig. 3 presents the concentrations of methidathion in

the water and the solid phases of the three reactor ex-

periments. Due to losses when drying the samples, there

are no data available for the first 23 d of the solid phase

of R1 experiment.In all reactor experiments, a decrease of methidathion

concentration was observed. The addition of NaOH led

to a sharp decrease in the concentration. This effect was

most obvious for R1, where the addition of solid NaOH

was accompanied by a steep decrease of the concentra-

tions both in the solid phase and in the water phase. A

decrease was also observed after the addition of a NaOH

solution to R2.

The concentrations of methidathion in the batchexperiments at different pH values are presented in Fig.

4. A sharp reduction in the concentrations occurred al-

ready at t0 30 min at pH 10.5 and 12.8. In solid waste-free buffer solutions, the concentrations at t0 ranged

between 74% and 85% at the pH values from 4.1 to 10.5.

At pH 12.8, only 26% of the initial concentration was

recovered (Fig. 4). The experiments with waste and

buffer solutions yielded total recoveries between 50%

and 71% at t0 30 min and pH 4.110.5 (Fig. 4). At pH12.8, only 1.8% of methidathion was detectable. After 97

h, the concentrations in the water phase of the experi-

ments with biological waste had remained relativelyconstant. A significant decrease was found only at pH

10.5. The remaining concentrations (assay without

Fig. 3. Relative concentrations (ratio of actual concentration to maximum concentration) of methidathion in the water and solid

phases of anaerobic digestion experiments R1 (upper curve), R2 (middle curve) and R3 (lower curve).



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biological waste, solid phase and total recovery in the

assay with biological waste) had decreased at t1 97 h.The decrease was largest at higher pH values.

Thus, both the abiotic batch experiments and the

reactor experiments revealed a base-catalyzed degrada-

tion of methidathion. It is worth noting that immedi-

ately after the addition of NaOH to the reactors, the

local pH is likely to have exceeded the values presented

in Fig. 2, until full mixing of the suspension was reached.

Abiotic hydrolysis of methidathion at alkaline pH

values was also reported by Eto (1977), Hartley and

Kidd (1987) and Richardson and Gangolli (1993). Eto

(1977) described a cleavage of the PS-bond as the main

mechanism of alkaline hydrolysis of methidathion (Fig.

5). The general mechanism for the hydrolysis of thio-

phosphoric acid esters is that the esters react with a

nucleophile by nucleophilic displacement (SN2) at eitherthe phosphorus or the carbon atom bound to the sulfur

of the thiol moiety (Schwarzenbach et al., 1993). For

nucleophilic displacement, OH is a better nucleophile

than H2O (Schwarzenbach et al., 1993). At pH 13 and 25

C, the half-life of methidathion in aqueous solution is

reported to be 30 min (Richardson and Gangolli, 1993).

The results of this study reveal a half-life at pH 12.8 of

even less than 30 min.

The batch experiments at different pH values also

showed a significant decrease at pH values below 10.5

(Fig. 4). Although the decrease was not as steep as it was

at pH 10.5 or 12.8, a slow hydrolysis at neutral or

weakly acidic pH values does take place. Smolen and

Stone (1997) observed an abiotic hydrolysis under neu-

tral and acid conditions for the phosphoric acid estersmethyl-parathion and methyl-chlorpyrifos. Hong and

Pehkonen (1998) showed both a neutral and an acid-

catalyzed hydrolysis of phorate. The phosphoric acid

esters disulfoton, thiometon, diazinon and demeton S

were shown to be hydrolyzed at pH 5.7 (Dannenberg

and Pehkonen, 1998).

The results of the batch experiments present the

stability of methidathion in the absence of microbial

activity. Whether or not a biological hydrolysis occurs in

the reactor experiments, can be determined by a com-

parison of the respective degradation rates. As the data

for the solid phase of R1 are not complete, this com-

Fig. 4. Recoveries of methidathion in batch assays of different pH values: recoveries in pure buffer solutions (upper diagram) and in

biowaste and buffer solutions (lower three diagrams). Total recovery gives the sum of solid and water phase concentrations. t0 30min, t1 97 h, n.d.: not detectable.

Fig. 5. Mechanism of alkaline hydrolysis of methidathion.



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parison can only be carried out for the reactor experi-

ment at 35 C. During the period of 97 h, the original

concentration was reduced by 53% (Fig. 3). The average

pH value was 5.5. The abiotic experiments at pH 5.6

showed a decrease of the start concentration to 64%.

The difference might be caused by a temperature effect

rather than by significant microbial degradation.Macalady and Wolfe (1983) did not observe a mi-

crobial influence on the degradation rate of chlorpyrifos.

However, biological hydrolysis of methidathion has

been reported by Gauthier et al. (1988). According to

Matsumara (1982), the biotic hydrolysis is either cata-

lyzed by non-specific enzymes with a broad spectrum,

e.g. reductases, oxidases and hydrolases, or by specific

enzymes which are abundant in numerous microorgan-

isms. However, microbial activity might have been low

in the reactor experiments as a consequence of the pH

shifts.

3.3. Effects of temperature

The disappearance rate of methidathion in the reactor

experiments increased with an increase in temperature.

For R1, measurable concentrations of methidathion

were obtained throughout the entire experiment, with

the exception of samples following the addition of solid

NaOH. During the second experiment R2, the concen-

tration in the water phase decreased continuously and

dropped below the limit of detection (36.8 lg l1 on the

basis of a 30 ml sample concentrated to 500 ll) after 54 d

of experiment. The concentration in the solid phase,

however, varied between 5% and 43%. For R3, a rapid

decrease in the concentration of methidathion was ob-

served for both the water and the solid phases. After 21

d of experiment, methidathion was no longer detectable.

The concentrations of methidathion in aqueous and

solid phases of the batch samples incubated at 55 C

indicated that methidathion was hydrolyzed exponen-

tially. The concentrations found in the water and in the

solid phase were added and transformed into logarith-

mic values. The hydrolysis followed pseudo-first-order

kinetics (Eq. (1)) (Cabras et al., 1997).

dC

dt kobsC 1

The observed hydrolysis rate constants kobs are

7:93 104 h1 (R2 0:93) at 25 C and 4:41 102 h1

(R2 0:99) at 55 C. The hydrolysis rate at 55 C thusexceeds the rate at 25 C by two orders of magnitude.

The pH value of both assays was 6.3.

Based on the hydrolysis rate constants, the half-life of

methidathion in biological waste at pH 6.3 is approxi-

mately 100 d. The half-life decreases to 41 d when a

temperature of 55

C is applied.

3.4. Sorption and partitioning of methidathion

The data presented for methidathion in the literature

mainly refer to natural aquatic systems. The complex

waste matrix and the high content of dissolved organic

matter can have various effects on the fate of methida-

thion. Dissolved organic matter can act as a co-solvent,thus increasing the apparent water solubility of a hy-

drophobic organic compound (e.g. Chiou et al., 1986).

On the other hand, non-extractable residues can be

formed in the solid phase, which may reduce the bio-

availability of methidathion.

The extraction of methidathion from pure water and

from process water yielded a recovery of approximately

80% (Table 3). Additional DOC, as present in the pro-

cess water, did not reduce the amount of methidathion

extractable with the mixture of cyclohexane and ethy-

lacetate. The ultrasonic extraction from the dry biolog-

ical waste, however, only gave a recovery of 41%,reflecting strong bonds between methidathion and the

organic matter. An adsorption of the rather polar meth-

idathion to polar groups of the organic matter is pos-

sible. The non-extractable percentage of phosphoric acid

esters in soil was found to be 1880% (Calderbank,

1989).

The total recovery from a system containing a sus-

pension of biowaste and water was approximetely 70%

(Table 3). The amount of methidathion detected in the

water phase is slightly larger than the amount extracted

from dry waste using cyclohexane and ethylacetate. In

contrast to the solvent extraction of the dry biological

waste, water was added shortly after the addition of

methidathion, when bonds possibly were weaker than

after 24 h. Due to the strong bond to the organic matter,

less than the maximum soluble amount of methidation

was released into the aqueous phase. Correcting the re-

sults of the water phase for the recovery found from

process water analysis, the total recovery increased to

about 90%. Hence, approximately 10% of the methida-

thion added to the experiments could not be extracted

from the suspended biological waste with this analytical

method.

The results of the experiments with different com-

position of the aqueous medium are presented in Fig. 6.

Table 3

Extraction efficiencies (n 3)

Assay Recovery

(%)

Standard de-

viation (%)

Pure water 79.00 2.70

Process water 79.00 3.57

Dry biological waste 40.85 1.68

Suspension: aqueous phase 53.19 0.82

Suspension: solid phase 13.74 0.65

Suspension: total 66.93 1.05



10/11

In all assays, a slight decrease in the concentration of

methidathion was observed, indicating slow hydrolysis

during the course of the experiment. The contents of

methidathion in the water phase of the experiments with

process water as well as yeast extracts were significantlylower than the content in the reference assay. Interest-

ingly, within the three assays with yeast extracts only the

assay with the lowest concentration of yeast was sig-

nificantly different from the others.

Clearly, yeast cells and high concentrations of DOC

do not act as co-solvents, but reduce the release of

methidathion from the solid phase into the water phase.

Experiments with soil confirm this observation (Table

4). The addition of DOC to a system of soil, water and

dissolved methidathion reduced the amount of meth-

idathione in the water phase, thus leading to a shift of

the equilibrium distribution towards the solid phase. An

explanation for this observation might be co-sorption.

Applied dissolved organic matter sorbed to soil may

retard organic pollutants (Koogel-Knabner et al., 1993).

4. Conclusions

The fate of methidathion was determined under

various experimental conditions, including biological

and abiotic processes as well as varying temperature and

pH and different compositions of the aqueous phase. In

a technical anaerobic digestion process, the variety of

process conditions will be smaller. Nevertheless, a par-

tial or complete degradation of methidathion by hy-

drolysis was observed in anaerobic mesophilic and

thermophilic digestions. On the basis of the results of

this study, thermophilic digestion is considered to have a

higher potential for methidathion degradation. Adding

methidathion containing waste to a solution of high

DOC will reduce the release of methidathion into the

water phase and might thus decelerate degradation.

Effective degradation of methidathion can also be

achieved by raising the pH to alkaline values. However,

all means of process control have to ensure that stable

microbial communities are maintained. An inhibition of

microbial activity will disturb the anaerobic digestion

process and might, for example, lead to a reduced biogas

yield. An alkaline pre-treatment of the biological waste

was formerly included in a technical anaerobic diges-tion process, but was dismissed for economic reasons

(Scherer, 1995).

It should be kept in mind, however, that pesticides

belong to a number of different chemical groups, which

are likely to react differently to certain adjustments of

the digestion process. Some pesticides are known to be

persistent (Vorkamp et al., 1999), others might form

toxic metabolites, as was shown for some phosphoric

acid esters (Dannenberg and Pehkonen, 1998; Hong and

Pehkonen, 1998). Even though this study indicated hy-

drolysis of methidathion during mesophilic and therm-

ophilic anaerobic digestion, any extension of these

results to the degradation of other compounds has to be

carefully considered.

Acknowledgements

The work was financed by the Bavarian Network on

Waste Research and Residual Management (Bayerischer

Forschungsverbund fuur Abfallforschung und Restst-

offverwertung). The authors wish to thank Martina

Rohr, Violetta Schittko and Heidi Zier for technical as-

sistance.

Table 4

Recovery of methidathion from a soil and water system, with

additional DOC (n 3)

Assay Recovery

(%)

Standard

deviation (%)

Water phase 22.09 2.60

Soil 0.32 0.046

Water phase DOC 7.78 2.26Soil DOC 7.85 0.60

Fig. 6. Relative concentrations of methidathion in the aqueous phases of the experiments with process water and yeast extracts in

comparison to a reference assay with pure water.



11/11

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