Science of the Total Environment 630 (2018) 302–307

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Science of the Total Environment

journal homepage: www.elsevier.com/locate/scitotenv

Transfer of arsenic from poultry feed to poultry litter: A mass

balance study
Sanjay K. Gupta a, X. Chris Le c, Gary Kachanosky a, Martin J. Zuidhof b, Tariq Siddique a,⁎
a
Department of Renewable Resources, University of Alberta, Edmonton, AB T6G2G7, Canada
b
Department of Agricultural Food and Nutritional Sciences, University of Alberta, Edmonton, AB T6G2P5, Canada
c
Division of Analytical and Environmental Toxicology, University of Alberta, Edmonton, AB T6G2G3, Canada

H I G H L I G H T S G R A P H I C A L A B S T R A C T

• First study that describes mass balance of

arsenic intake and excretion by chickens
• Two strains Ross and Cobb exhibited
similar response to roxarsone-amended
feed.
• Arsenic did not retain in chickens during
their growth on arsenic containing feed.
• Poultry litter enriched in arsenic war-
rants investigations if used in crop
production.

a r t i c l e i n f o a b s t r a c t

Article history: Roxarsone (rox), an arsenic (As) containing organic compound, is a common feed additive used in poultry pro-
Received 1 January 2018 duction. To determine if As present in rox is excreted into the poultry litter without any retention in chicken meat
Received in revised form 8 February 2018 for safe human consumption, the transference of As from the feed to poultry excreta was assessed using two com-
Accepted 10 February 2018
mercial chicken strains fed with and without dietary rox. The results revealed that both the strains had similar
Available online xxxx
behaviour in growth (chicken weight; 2.17–2.25 kg), feed consumption (282–300 kg pen−1 initially containing
Editor: Jay Gan 102 chicken) and poultry litter production (73–81 kg pen−1) during the growth phase of 35 days. Our mass bal-
ance calculations showed that chickens ingested 2669–2730 mg As with the feed and excreted out
Keywords: 2362–2896 mg As in poultry litter during the growth period of 28 days when As containing feed was used, yield-
Roxarsone ing As recovery between 86 and 108%. Though our complementary studies show that residual arsenic species in
Arsenic rox-fed chicken meat may have relevance to human exposure, insignificant retention of total As in the chicken
Poultry feed meat substantiates our mass balance results. The results are important in evaluating the fate of feed additive
Arsenic intake used in poultry production and its potential environmental implications if As containing poultry litter is applied
Arsenic excretion
to soil for crop production.
Poultry litter
© 2018 Elsevier B.V. All rights reserved.
Arsenic recovery

1. Introduction

⁎ Corresponding author.
E-mail addresses: xc.le@ualberta.ca (X.C. Le), gary.kachanoski@ualberta.ca
Arsenic (As) is a toxic element in the environment and a known car-
(G. Kachanosky), martin.zuidhof@ualberta.ca (M.J. Zuidhof), tariq.siddique@ualberta.ca cinogen that consistently ranks first on the Agency for Toxic Substances
(T. Siddique). and Disease Registry (ASTDR) Substance Priority List (https://www.

https://doi.org/10.1016/j.scitotenv.2018.02.123
0048-9697/© 2018 Elsevier B.V. All rights reserved.
 S.K. Gupta et al. / Science of the Total Environment 630 (2018) 302–307 303

atsdr.cdc.gov/spl/index.html). Sources of As contamination in the envi-
ronment includes its elevated contents in some earth crust minerals, in-
dustrial activities (mining and smelting) and arsenical compounds used
for different purposes (ATSDR, 2007). Drinking water and ingestion of
contaminated food are primary As exposure pathways to biological re-
ceptors (humans and animals) that pose a significant threat to public
health (Naujokas et al., 2013).
Roxarsone (rox; 3-nitro-4-hydroxyphenylarsonc acid), an organic
compound containing As, was added routinely to the poultry feed for
parasitic disease prevention, growth promotion, enhanced feed utiliza-
tion and improved meat pigmentation for N60 years (Nachman et al.,
2013; Fisher et al., 2015) until its use was discontinued in Europe
(European Commission, 1999), USA (FDA, 2013), and Canada (https://
www.theglobeandmail.com/life/health-and-fitness/sales-halted-after-
arsenic-found-in-chicken-drug/article591962/). When the United
States Food and Drug Administration (FDA) approved rox in 1944 as a
feed additive (Nachman et al., 2013), it was believed that the non-
toxic organic As present in rox would not change into toxic inorganic
As inside the chicken body; rather organic As would be excreted un-
changed into the poultry manure leaving chicken meat safe for human
consumption (Schmidt, 2012). However, higher than tolerable amounts
of total As (N2.0 mg kg−1) established by FDA before 1963 (FDA, 1963)
were found in the livers of rox-fed chickens compared to the ones fed
without rox (FDA, 2011).
Subsequent studies focused on the determination of different
species of As in different chicken body parts as the toxicity of As is highly
dependent on its chemical species (Moe et al., 2016). Higher concentra-
tions of inorganic As were found in the conventional compared to
the anti-biotic free conventional chicken meat samples in the US
(Nachman et al., 2013). Similarly, inorganic forms of As was also de-
tected in the feather meal products from six US states (Nachman et al.,
2012). Yao et al. (2016) revealed that rox in chicken diet was trans-
formed into its metabolites (different As species) in chicken manures
which subsequently increased concentrations of these As species in
rice plants (grain, straw and hull) when grown in soil amended with
the chicken manures. However, no study to date has reported the
mass balance of As from poultry feed to poultry litter. In our study, we
conducted an experiment growing two strains of chicken on rox-
supplemented feed or feed devoid of rox. Different arsenic species
were analyzed in chicken liver (Peng et al., 2014; Peng et al., 2017),
chicken breast (Liu et al., 2016) and chicken litter (Yang et al., 2016)
16 pens with a treatment combination of two types of feed, two poultry
strains and four replications. In each pen (169 cm × 420 cm), 102 chicks
(one-day old, mixed sex, previously weighed) were reared. New soft-
wood shavings of known weight were used as the bedding material in
each pen that yielded a depth of 7.5 cm above the pen floor. For the
rox treatment, 3-NITRO® (Alpharma Canada Corporation) containing
20% rox was added to the feed at 250 g ton−1. Because rox contains
28.48% As, the As content was 14.24 mg kg−1 of the feed prepared for
the rox treatment. The feed used for the control treatment had all the
same ingredients except rox. All chickens received diet according to
standard Poultry Research Centre diet composition (Table 1) - starter
diet (3068 kcal kg−1; 23% crude protein) for two weeks, grower diet
(3152 kcal kg−1; 20% crude protein) from two to four weeks, and fin-
isher diet (3196 kcal kg−1; 19% crude protein) from four to five weeks
of age. Rox was added to the starter and grower diets only. Chickens
had ad libitum access to both feed and water (b1 μg As L−1).
2.2. Poultry litter sampling and analysis
Samples of poultry litter (excreta mixed bedding material) were col-
lected from each pen on 14, 28 and 35th days using a steel core of 10 cm
diameter. Fifteen fresh samples were collected randomly up to the full
depth of litter, and then total amount of litter in each pen was estimated
by extrapolating the measurements to the whole pen area. These 15 lit-
ter samples, on each respective sampling day, were pooled together and
a representative sample (a quarter of the composite sample; ~0.5 kg)
was put into a double Ziplock® polyethylene bag, sealed and frozen at
−20 °C until chemical analysis was performed. The remaining sample
was put back into the pen and mixed well with the litter. The total litter
amount was also determined at 35th day (clean out day) by both esti-
mating through sampling method employed for day 14, 28 and 35,
and physically weighing all the litter collected from each pen. Total litter
weight measured at 35th day was higher than the weight estimated
through sampling method. Therefore, a factor of 1.4 was used, which
represented an average ratio of measured to estimated poultry litter
weight from 16 pens, to calculate the weight of poultry litter produced
Table 1
Composition of different growth stage poultry feeds used in the experiment to feed
chickens for 35 days.

samples taken from this study. In the current manuscript, we report Ingredient (%) Control Rox
mass balance that determines the amount of total As taken in by
Startera Growerb Finisherc Starter Grower Finisher
chickens with feed consumption and the amount of total As excreted
Corn, yellow grain 18 18 15 18 18 15
in poultry litter to assess the possible retention of As inside the chicken
Fat, vegetable 3.8 3.4 4.1 3.8 3.4 4.1
body. Fish meal, menhaden 3.0 5.0 3.5 3.0 5.0 3.5
Arsenic utilization in poultry industry and its subsequent transport Soybean meal 26.9 16.2 15.1 26.9 16.2 15.1
to different components of biosphere may have far-reaching conse- Wheat, hard grain 43 53 58 43 53 58
quences related to human health and the environment. Though rox con- Calcium carbonate 1.5 1.0 1.0 1.5 1.0 1.0
Dicalcium phosphate 1.5 1.0 1.1 1.5 1.0 1.1
taining poultry feed has been discontinued in Europe and North Sodium chloride 0.43 0.34 0.36 0.43 0.34 0.36
America, many other countries continue to use phenylarsenicals in the L-Lysine 0.23 0.15 0.15 0.23 0.15 0.15
poultry industry (Nachman et al., 2012; Yao et al., 2016). DL-Methionine 0.23 0.10 0.09 0.23 0.10 0.09
L-Threonine 0.05 0.10 0.03 0.05 0.10 0.03
2. Methodology Broiler vitamin 0.50 0.50 0.50 0.50 0.50 0.50
premix (0.5%
2.1. Poultry feed and chicken growth experiment inclusion)
Choline chloride 0.50 0.50 0.50 0.50 0.50 0.50
premix (0.5%
Effect of two types of feed (rox containing feed: rox, and feed with- inclusion)
out rox: control) was studied on two commercial strains of poultry Vitamin E 5000 IU kg−1 0.30 0.30 0.30 0.30 0.30 0.30
(Cobb 500 and Ross 308) grown in an environmentally controlled Generic enzyme 0.05 0.05 0.05 0.05 0.05 0.05
(0.5% inclusion)
barn as described previously by Liu et al. (2016). Briefly, in the barn,
Coccidiostat (Amprol) 0.05 0.05 0.05 0.05 0.05 0.05
age related temperature and ventilation protocols were followed as Growth promoter 0.000 0.000 0.000 0.005 0.005 0.000
per the Strain Management Guides of Cobb (Cobb-Vantress, 2008) and (Roxarsone)
Ross (Aviagen, 2009). The lighting program followed a photoperiod of a
0–14 days.
23 h for the first three days, and then 20 h' light and 4 h' dark from b
15–28 days.
fourth day to the end of the experiment (35 days). In total, there were c
29–35 days.
304 S.K. Gupta et al. / Science of the Total Environment 630 (2018) 302–307

in each pen (Table 2). The loss of litter due to periodical sampling over
the study period was also accounted for during the final calculations.
For As analysis, poultry litter samples were homogenized in an all-
plastic blender. Moist samples, equivalent to 1 g oven-dried sample,
were used for digestion to avoid any As loss through volatilization dur-
ing drying. However, the results were reported on dry-weight basis
after using moisture correction factor for each litter sample used in
the digestion. Litter samples were digested with concentrated nitric
acid (HNO3) (A509P212, trace metal grade, Fisher Scientific) in a micro-
wave oven (Ethos Sel, Milestone) using a modified USEPA method
(USEPA, 1996), wherein temperature was ramped from room tempera-
ture to 180 °C within 10 min and maintained at 180 °C for 10 more mi-
nutes to facilitate the complete digestion of samples. After digestion, the
vials were allowed to cool. The solution was diluted to 100 ml using
nanopure water (Barnstead nanopure, Thermo Scientific), and filtered
through 0.45 μm PTFE syringe filter (033911C, Fisher Scientific). One
milliliter of filtered solution was taken in a 50 ml centrifuge tube and di-
luted to a final volume of 50 ml using 1% HNO3 solution. This acidifica-
tion of samples was done to prevent any precipitation before ICP-MS
(Inductively Coupled Plasma Mass Spectrometer) analysis (Javed et al.,
2013).
Approximately 10 ml of this prepared solution was transferred to a
sampling tube and placed on the auto-sampler of ICP-MS (iCAP Q-
Thermo Scientific). During the analysis of experimental samples, con-
tinuous calibration blanks (CCB) to check any metal carry over, and con-
tinuous calibration verification (CCV) external standards (2 and 20
μg As L−1) were also included after every 10 samples to ensure the va-
lidity of the initial calibration of the instrument. External standards
were prepared in 1% HNO3 using a stock standard (CLMS-2AN; SPEX
Certi Prep). Scandium (Sc) and yttrium (Y) were added to the samples
as internal standards for quantitation and accounting for any instru-
mental drift over time. The reagent and experimental blanks were also
analyzed along with the experimental samples. The As concentration
was lower than the limits of detection (LOD: 0.01 μg As L−1 determined
using 3× standard deviation (σ) of intensities of 7 blanks + average of
the 7 blank intensities) in all reagents and experimental blanks. All
the CCB showed As concentrations lower than the LOD, and As
concentrations in all the CCVs were within ±10% of the actual values
of As (2 and 20 μg L−1) (Javed and Siddique, 2016).
2.3. Mass balance calculations
For mass balance calculations, the following equations were used to
quantify As intake and As excreted in the poultry litter:
I ¼ Wf  Cf ð1Þ
I = Cumulative As taken in by chickens through feed consumption
(mg As pen−1).
Wf = Amount of feed consumed by chicken (kg pen−1).
Cf = Concentration of As in poultry feed (e.g. 14.24 mg As kg−1 rox
feed).
E ¼ ðWl  Cl Þ ð2Þ
E = Cumulative As excreted by chickens in poultry litter
(mg As pen−1).
Wl = Total amount of poultry litter producedby chickens (kg pen−1).
Cl = Concentration of As in poultry litter (mg kg−1).
Recovery of As ð%Þ ¼ E=I  100 ð3Þ
3. Results
3.1. Poultry production with feed consumption
Data pertaining to number of chickens, cumulative feed consump-
tion and poultry litter production per pen, and average body weight
per chicken on different days of observation have been given in
Table 2. The experiment was started with 102 chickens in each pen
but their number decreased with time due to death and removal for
meat analyses. By 35th day, the numbers of Cobb chickens remained
were 69 ± 5 and 67 ± 5 pen−1 in the control and rox treatments,

Table 2
Measurements of different growth parameters and poultry litter production used in the mass balance calculations to determine the fate of feed additive roxarsone.

Measured characteristics Strain Treatment (Days)

0 14 28 35
−1
Number of chickens pen Cobb Control 102 ± 0.6 88 ± 1.5 81 ± 5.3 69 ± 5.5
Cobb Rox 102 ± 0.6 88 ± 0.6 80 ± 5.3 67 ± 4.7
Ross Control 102 ± 0.0 88 ± 1.5 83 ± 1.2 71 ± 2.5
Ross Rox 102 ± 0.6 89 ± 0.6 84 ± 0.6 72 ± 1.5
Average body weight of chicken (kg chicken−1) Cobb Control 0.043 ± 0.0003 0.40 ± 0.005 1.55 ± 0.02 2.20 ± 0.03
Cobb Rox 0.040 ± 0.004 0.41 ± 0.007 1.55 ± 0.06 2.25 ± 0.02
Ross Control 0.042 ± 0.003 0.40 ± 0.022 1.53 ± 0.05 2.17 ± 0.09
Ross Rox 0.040 ± 0.0002 0.41 ± 0.005 1.58 ± 0.03 2.21 ± 0.03
Cumulative feed consumption (kg pen−1) Cobb Control 47 ± 1 193 ± 6 297 ± 20
Cobb Rox 46 ± 1 187 ± 9 282 ± 14
Ross Control 47 ± 2 186 ± 5 300 ± 41
Ross Rox 48 ± 1 192 ± 7 292 ± 12
Estimated weight of poultry litter produced (kg pen−1)a Cobb Control 19 ± 2 30 ± 4 57 ± 4 59 ± 3
Cobb Rox 18 ± 4 30 ± 2 52 ± 2 59 ± 1
Ross Control 20 ± 3 30 ± 6 54 ± 3 59 ± 9
Ross Rox 19 ± 5 29 ± 3 53 ± 1 59 ± 3
Calculated weight of poultry litter produced (kg pen−1)b Cobb Control 19 ± 2 43 ± 6 80 ± 5 82 ± 4
Cobb Rox 18 ± 4 42 ± 3 73 ± 3 83 ± 2
Ross Control 20 ± 3 41 ± 9 76 ± 4 82 ± 12
Ross Rox 19 ± 5 41 ± 4 74 ± 17 83 ± 4

Values are the means of the three replications with associated standard deviations (±).
a
Initially (day 0) a known weight of wood chips was spread on each pen floor. Afterwards on 14th, 28th and 35th days, the weight of poultry litter was estimated based on sample
weight and pen area.
b
On 35th day, litter in each pen was also weighed physically. After comparing the estimated and measured weights on 35th day, a factor of 1.4 (average ratio of measured to estimated
poultry litter weight from 16 pens) was used to calculate the poultry litter produced. These values were used in the mass balance to calculate the amount of arsenic excreted by chicken in
each pen.
 S.K. Gupta et al. / Science of the Total Environment 630 (2018) 302–307 305

respectively, while 71 ± 3 and 72 ± 2 Ross chickens pen−1 remained in

each control and rox treated pens, respectively. Cumulative feed con-
sumption increased significantly with age for both the strains under
both the treatments. In 35 days, Cobb chickens consumed 297 ± 20
and 282 ± 14 kg feed pen−1 under the control and rox
treatment, respectively. Similar feed consumption 300 ± 41 and 292
± 12 kg pen−1 was recorded in control and rox fed pens, respectively,
growing the Ross strain. The average body weight of chickens also in-
creased with age for both the strains, and no significant differences
were observed within the chicken strains and the two treatments. The
average body weight ranged between 2.17 ± 0.09 to 2.25 ±
0.02 kg chicken−1 at day 35.

3.2. Arsenic intake

Total As intake was calculated using the amount of feed consumed

and concentration of As present in the feed (Eq. (1)). In control feed,
rox was not added (Table 1), therefore we assumed that no As would
be taken up by the chickens under the control treatment. In the rox
treatment, As was added at 14.24 mg kg−1 of feed. As the chickens
grew over time, they consumed more feed, and consequently, more As
was taken up by chickens while consuming rox-treated starter diet
(0–14 days) and grower diet (14–28 days). Supply of As was stopped
after 28th day as the grower diet was replaced by the finisher diet
which did not contain rox. Because the two strains were not different
in their feed consumption (Table 2), As intake calculated for both strains
was similar. At 14th day, As intake was 661 ± 10 and 680 ± 8 mg pen−1
which increased cumulatively to 2669 ± 128 and 2730 ± 92 mg pen−1
in Cobb and Ross, respectively at 28th day (Table 3).

3.3. Poultry litter production and arsenic concentration in the poultry litter

Initially, a known amount of wood chips was spread on each pen

floor. As the chickens grew in age and size, they consumed feed and
water, and excreted urine and feces on to the bedding material. Conse-
quently, weight of poultry litter (mixture of wood chips and excreta) in-
creased significantly with time. Cobb chickens produced 43 ± 6 and 42
± 3 kg dry poultry litter pen−1 at 14th day and 80 ± 5 and 73 ±
3 kg litter pen−1, in control and rox-fed pens, respectively at 28th day.
Similarly, the poultry litter production by Ross increased from 41 ± 9
and 41 ± 4 kg litter pen−1 at 14th day to 76 ± 4 and 74 ±
17 kg litter pen−1 at 28th day under control and rox treatments, respec-
tively (Table 2). Though higher amounts of poultry feed were consumed
during the last week (29–35 days) of chicken growth, not much corre-
sponding increase in poultry litter was recorded at 35th day that obvi-
ously indicates some discrepancy in poultry litter determination at
35th day.
Regarding As concentrations in poultry litter collected from control
pens, 0.25 ± 0.1 to 1.75 ± 0.4 mg kg−1 As was detected in the litter
from both the strains analyzed during their growth period (Fig. 1)
even though the control feed was devoid of rox. Arsenic concentration
was significantly higher in the poultry litter of rox-fed chickens than
Fig. 1. Arsenic concentration in poultry litter (A) produced by two strains of chicken (Cobb
and Ross) and cumulative arsenic excreted by both the strains (B). Cobb-cont and Ross-
cont are chickens that received poultry feed without roxarsone, whereas Cobb-Rox and
Ross-Rox received poultry feed containing roxarsone. Error bars showed ± standard
deviation from 3 replications.
the poultry litter produced in the control treatment. With time, the con-
centration of As increased significantly in the poultry litter collected
under both the strains. Mean As concentration was 17 ± 3 and 15 ±
4 mg kg−1 when measured at 14th day and 40 ± 7 and 36 ±
4 mg kg−1 at 28th day in the poultry litter of Cobb and Ross, respec-
tively, which decreased to 23 ± 2 and 24 ± 2 mg kg−1, respectively at
35th day as the grower diet was replaced by the finisher diet after
28th day which did not contain rox (Fig. 1A).

Table 3
Mass balance of arsenic taken up by chicken fed with roxarsone containing feed and arsenic excreted in the chicken poultry litter.

Parameters Strain Day 14 Day 28 Day 35

Arsenic intake (mg pen−1) Cobb 661 (±10) 2669 (±128)

Ross 680 (±8) 2730 (±92)
Arsenic excreted (mg pen−1) Cobb 703 (±149) 2896 (±414) 1838 (±192)
Ross 625 (±185) 2362 (±228) 2109 (±94)
Arsenic recovered in poultry litter (%) Cobb 106 ± 21 108 ± 11 69 ± 4
Ross 92 ± 26 86 ± 5 77 ± 4

Arsenic recovery for day 35 was calculated using arsenic intake data calculated for day 28 because arsenic containing rox feed was stopped at day 28. Parenthesis shows ± standard de-
viation calculated from three replications.
306 S.K. Gupta et al. / Science of the Total Environment 630 (2018) 302–307
3.4. Mass balance to determine recovery of As in the poultry litter
Excretion of As in litter was calculated using Eq. (2). At 14th day, As
excretions by Cobb and Ross were 703 ± 149 and 625 ± 185 mg pen−1,
which increased to 2896 ± 414 and 2362 ± 228 mg pen−1, respectively,
by 28th day (Table 3). At 35th day, 1838 ± 192 and 2109 ±
94 mg As pen−1 was determined in Cobb and Ross pens as excreted
As (Table 3, Fig. 1). Small amount of As (101 ± 21 to 141 ±
29 mg pen−1) was also excreted in the control treatment (Fig. 1B).
Mass balance calculation (Eq. (3)) was performed to determine the re-
covery of As in the poultry litter produced during the experiment
(Table 3). During the first 14 days, 106 ± 21% and 92 ± 26% of ingested
As was recovered from the poultry litter produced by Cobb and Ross
strains, respectively. Similar recovery of As (86 ± 5 to 108 ± 11%)
was achieved at 28th day when feeding chicken with rox containing
grower diet was stopped and cumulative As excreted in poultry litters
produced by Cobb and Ross was determined. Arsenic recovery de-
creased in the poultry litters of Cobb and Ross when calculated for
35th day due to decreased As concentration in the poultry litter and
some discrepancy in poultry litter measurements at 35th day (Table 3).
4. Discussion
The mass balance of As from poultry feed to poultry litter reported in
this manuscript is a part of a comprehensive study envisaged to track As
from rox containing poultry feed to poultry litter including its metabo-
lism in the chicken meat (Peng et al., 2014; Liu et al., 2016; Peng et al.,
2017). Two strains of chickens (Cobb and Ross) were grown on feed
with or without the addition of rox in separate pens. Quite expectedly,
cumulative feed consumption and body weight of the chickens in-
creased with age. Intake of As by the chickens kept increasing with
time in rox treatment as starter and grower diets contained recom-
mended dose of rox. The concentration of As in the poultry litter of
rox treatment increased till 28th day and then decreased when poultry
litter was analyzed at 35th day. Because as the chickens grew in size and
weight between 28th and 35th day, they consumed rox-free (Finisher)
diet and produced excreta, which led to the decrease in As concentra-
tion when As was measured in litter at 35th day (Fig. 1A). Arsenic con-
centration of 0.25–1.75 mg kg−1 was also detected in the litters of
control treatment chickens that indicated the presence of As in the poul-
try feed even without the addition of rox. Previously, a similar study also
reported 0.6 mg As kg−1 poultry litter where chickens were not fed with
rox (Garbarino et al., 2003). In our study, the source of As in the control
feed was menhaden fishmeal, a protein source in the feed (Table 1) (Liu
et al., 2016). Though the concentrations of As in poultry litter reported
in previous studies vary significantly from 0 to 77 mg kg−1, there is no
mention of recovery or mass balance of As from poultry feed to poultry
litter (Morrison, 1969; Sims and Wolf, 1994; Anderson and Chamblee,
2001; Arai et al., 2003; Toor et al., 2007; Fisher et al., 2015).
For mass balance calculation to determine the recovery of As in poul-
try litter, we used As intake calculated for 14th and 28th days and com-
pared with the As excreted at both the time points (Table 3). The results
(86–108% As recovery in poultry litter at 14th and 28th days) suggested
that there was no significant retention of As in the body of chickens fed
on rox containing feed. These mass balance results are corroborated by
the results of As in chicken meat which revealed insignificant retention
of As in the meat of chickens sampled from the pens (in the current ex-
periment) on 35th day and taken for the analyses of different As species
(Peng et al., 2014; Liu et al., 2016; Peng et al., 2017; Liu et al., 2018).
Breast meat from the control and rox-fed chickens showed very minute
As concentrations; meat from rox-fed treatment contained ~42 μg kg−1
(rox plus all individual As species) versus ~33 μg kg−1 in the control
treatment chicken breast meat (Liu et al., 2016). The liver samples
from these chickens accumulated relatively higher concentrations of
rox and other individual As species (~10 times higher than the breast
meat); ~400 μg kg−1 in rox-fed chicken compared to ~80 μg kg−1 in
the control chicken (Peng et al., 2014). If we assume and apply this
liver As concentration to whole chicken body, even then it would not
contribute much (0.4 mg kg−1) to our overall mass balance calculations
that deals higher As concentrations in poultry feed and litter, and the
meat As concentrations are far below the tolerable levels (2.0 mg kg−1
previously established by FDA before 1963) (Nachman et al., 2013).
Therefore, our results do not align with the findings of FDA (2011)
where higher concentrations of total As (2.8 ± 1.4 mg kg−1) in liver
samples of rox-fed chicken were reported. However, recent findings
on the identification of additional three methylated phenylarsenical
metabolites in the liver of chickens fed with rox containing feed from
this study revealed that the concentrations of metabolites (~500
μg kg−1) in liver decreased to ~130 μg kg−1 after 5 days of rox feed ces-
sation; these residual As species in chicken liver are relevant to human
exposure if chicken livers are consumed (Peng et al., 2017).
Arsenic recovery for 35th days was also determined by comparing
cumulative As intake at 28th day when chickens stopped receiving rox
containing feed, with excreted As at 35th day. A recovery of 69–77%
was calculated in both Cobb and Ross chicken strains. This lower As re-
covery is attributed to discrepancy in poultry litter determination at
35th day and/or other factors such as environmental losses due volatil-
ization, or loss of As to the pen floor via urine (Nigra et al., 2017). This
study also revealed that the poultry litter from rox-fed chickens
contained higher As (~40 mg kg−1). Because poultry litter is commonly
used as an organic amendment to fertilize soil (Wilkinson, 1979) for
crop production, As and its different metabolites/species can be
translocated to different plant parts that may arise some public health
issues (Rutherford et al., 2003; Yao et al., 2009; Yao et al., 2016).
5. Conclusions
The results of our study show that arsenic from poultry feed ends up
in poultry litter when two poultry stains are fed with rox-amended feed.
Our mass balance calculations showed As recovery between 86 and
108% during the growth period of 28 days which suggested no signifi-
cant retention of As in the chickens. Our complementary studies on
chicken meat analysis also substantiated our mass balance results.
Higher arsenic in poultry litter warrants investigations on its environ-
mental implications if poultry litter enriched in As is applied to agricul-
tural soils for crop production.
Acknowledgements
Authors gratefully acknowledge the financial support provided by
Natural Sciences and Engineering Research Council of Canada (NSERC
DG; 371909-2009), Canada Foundation for Innovation (128377), Cana-
dian Water Network, Poultry Industry Council, and Alberta Livestock
and Meat Agency Ltd. (ALMA).
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