UNIVERSIDADE TIRADENTES UNIT

DIRETORIA DE PESQUISA - DPE

PROGRAMA DE PS-GRADUAO EM ENGENHARIA DE PROCESSOS PEP

RESUMO ARTIGO DE REVISTA

DIEGO LOPES CORIOLANO

Prof. Dr Manuela Leite

Disciplina: Modelagem e Otimizao de Processos

Aracaju
Agosto/2016

Objetivo do trabalho:
Realizar pesquisa de um artigo publicado em revista cientfica. O artigo de tema livre,
porm deve conter aplicaes de balano de massa ministradas na disciplina Modelagem e
Otimizao de Processos do Programa de Ps-Graduao em Engenharia de Processos PEP.
O resumo dever ser dividido em:

Ttulo de artigo;
Autores do artigo;
Nome da Revista Publicada;
Objetivos do artigo; e
Aplicaes de balano de massa.

1. Ttulo de Artigo
Mass balances and distributions of C, N, and P in the anaerobic digestion of different
substrates and relationships between products and substrates

2. Autores do artigo
Heng Li;
Fen Tan;
Lanting Ke;
Dong Xia;
Yuanpeng Wang;
Ning He;
Yanmei Zheng; e
Qingbiao Li.

3. Nome da Revista Publicada

O artigo em questo foi publicado pela Chemical Engineering Journal no ano de
2015.

4. Resumo do artigo
Como resumo o artigo destaca conhecer as distribuies e balanos de massa de C,
N e P durante o processo de digesto anaerbica (AD); Por exemplo, estes valores
podem ser usados para obter informao quantitativa sobre o destino de C, N, P e
tambm de digesto anaerbica, compreender saldos de carbono e de nutrientes, e
para avaliar a contribuio dos processos AD para ciclos elementares. Neste trabalho,
o contedo C, N e P no gs, lquidos e slidos fases foram investigadas por sete
diferentes substratos: palha de milho, palha de arroz, palha de trigo, dejetos de sunos
a partir de sunos alimentados com rao ou do gnero alimentcio, esterco bovino e
esterco de galinha.
Como foco na disciplina em questo, destaca-se a seguinte informao do artigo:
Os balanos de massa de C, N e P para cada substrato diferente eram distintos,
mas todos os saldos foram perto de 100% neste estudo.

5. Introduo
Os autores iniciam o trabalho definindo a digesto anaerbica como um processo
no qual microrganismos anaerbios converter matria orgnica em biogs em um
ambiente livre de oxignio. Muitos estudos tm demonstrado que a tecnologia de

digesto anaerbia pode ser empregada para processar com sucesso muitos tipos de
resduos de biomassa slida, como palha de culturas, aves e estrume animal.
Destacam tambm o cultivo de arroz, milho e trigo e o descarte das palhas dessas
plantaes e tambm a quantidade de estrume animal, principalmente de sunos,
bovinos e galinhas na China. Com isso, os autores sugerem que a AD um mtodo
adequado para converter as palhas da colheita e o esterco em biogs e com isso
haveria produo de energia renovvel e reduo de desperdcio.
Foi realizada uma reviso bibliogrfica sobre o balano de massa e os autores
afirmaram que o balano de massa crucial para a quantificao da produo de
bioenergia e potencial de recuperao de nutrientes a partir de AD, que necessrio
para avaliar a contribuio dos processos AD aos ciclos elementares. Alm disso,
balanos de massa so muitas vezes identificados com referncia a parmetros
qumicos e fsicos convencionais que fornecem uma indicao sobre o destino de
materiais compostveis em todo o AD.
Por fim destacaram revises anteriores que abordaram a distribuies e equilbrio
de massa do C, N e P na AD, incluindo o de nutrientes (N, P e K) do balano de massa
de resduos alimentares domsticos fonte segregada em AD; e distribuio de
tamanho de partculas, padres de elementos traos (N, P, K, Ca, Fe, Mg, S, Al, Cu, Mn
e Zn) e balanos de massa em uma planta de digesto anaerbia em grande escala
tratamento de dejetos de sunos. No entanto, esses estudos raramente consideravam
o balano de massa de C que mais crucial para o processo AD, segundo os autores.
O objetivo do trabalho foi determinar as distribuies de C, N e P e balanos de
massa de adubos e palhas para fornecer uma base terica para otimizar produo de
metano. Por outro lado, a anlise da relao entre os produtos e substratos crucial
para o desenvolvimento de parmetros chave de processo e um modelo de previso
de produtos relacionados com a gerao de energia renovvel atravs de AD.

6. Clculos de balano de massa

Os balanos de massas de C, N e P foram calculados antes e depois de AD. A massa
elementar inicial foi obtida multiplicando as fraces de C, N e P pela massa do
substrato. A massa final de C foi calculada utilizando as Equaes (1) e (2).

Onde: W a massa final, mg; Cliquid a concentrao de C lquido, mg / L; Csolid a

quantidade de C slido, mg / g; a densidade do lquido; V o volume total efetivo
dos digestores; V3 o volume de lquido perdido no processo de congelao-secagem,
e V1, V2, W1, W2 e W3 so como descritos acima.
As massas finais de N e P foram obtidas pela combinao das massas lquidas e
slidas destes dois elementos. As massas finais foram corrigidas utilizando a massa
final correspondente do digestor em branco.
Os dados dos digestores foram submetidos a uma anlise de varincia (ANOVA)
para determine se as diferenas observadas entre os substratos foram
significativamente diferente. As diferenas foram consideradas significativas com uma
probabilidade de 0,05 (p <0,05). Os autores analisaram a de correlao entre os
produtos e composies de substratos foram realizadas usando SPSS 19.0. Grficos e
processamento de dados foram concludas usando OriginPro 8.0.
Os balanos de massas de diferentes substratos em AD foram calculados como a
massa de gs, lquido e slido de C, N, e P, antes e depois de AD. Os valores de C (Fig. 4
a) so aceitveis considerando as perdas lquidas de 4-10%. Amostragem, erros
analticos e produo de biogs em condies anormais de temperatura e presso
constituem potenciais fontes de erro. Compreendendo o equilbrio C pode ajudar os
operadores de centrais avaliarem a eficincia do processo de AD e modificar os
parmetros de processo para melhorar a eficincia da degradao da AD. Alm disso, a
Fig. 4b mostra perdas de N de 3-15% para vrios substratos, onde a 3-7% para adubos
e 9-15% para palhetas. Tambm houve perdas no teor de P (3-9%) semelhantes s de
N seguindo os processos AD, de acordo com a nossa anlise.

ANEXO - Artigo Original

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Mass balances and distributions of C, N, and P in the anaerobic

digestion of different substrates and relationships between products
and substrates
Heng Lia, Fen Tanb, Lanting Keb, Dong Xiab, Yuanpeng Wangb, Ning He b,Yanmei
Zhengb*, Qingbiao Lia,b,c*
a

College of the Environment and Ecology, Xiamen University, Xiamen 361005,

China
b

Department of Chemical and Biochemical Engineering, College of Chemistry and

Chemical Engineering, Xiamen University, Xiamen 361005, China

c

College of Chemistry and Life Science, Quanzhou Normal University, Quanzhou

362000, China

*Corresponding author: Department of Chemical and Biochemical Engineering,

College of Chemistry and Chemical Engineering, Xiamen University, No. 422,
Southern Siming Road, Xiamen 361005, P.R. China; phone and fax numbers: +86 592
2189595 & +86 592 2183088; E-mail: zym@xmu.edu.cn; kelqb@xmu.edu.cn

Abstract
Knowledge of the distributions and mass balances of C, N, and P during anaerobic
digestion (AD) processes serves several purposes; for instance, these values can be
used to obtain reliable quantitative information on the fate of C, N, and P after AD, to
understand carbon and nutrient balances, and to evaluate the contribution of AD
processes to elemental cycles. In this paper, C, N, and P contents in the gas, liquid and
solid phases were investigated for seven different substrates: corn straw, rice straw,
wheat straw, swine manure from pigs fed feedstuff or foodstuff, cattle manure, and
chicken manure. The conversion efficiency of C in a substrate into methane was
found to be highest in swine manure from pigs fed foodstuff (15.8%), and greater
liquid and solid stream nutrient levels were found in the swine manures and chicken
manure than in the cattle manure and straws. The mass balances of C, N and P for
each different substrate were distinct, but all balances were close to 100% in this

study. Further study revealed that CH4 yields and substrate lignin fitted a linear
negative relationship well, and the COD/TOC, NH4 +-N and TP concentrations in the
supernatant had positive linear relationships with the proteins, TAN/proteins and
potassium of substrates, respectively.
Keywords: Anaerobic digestion, Methane, Nutrients, Organic waste, Distribution.

1. Introduction
Anaerobic digestion (AD) is a process in which anaerobic microorganisms convert
organic matter into biogas in an oxygen-free environment. Many studies have proven
that anaerobic digestion technology can be employed to successfully process many
types of solid biomass waste, including crop straw, poultry and livestock manure[1-3].
Approximately 755 million tons of crop straw (of which rice, corn and wheat straw
account for 79.5%) and 39.7 billion tons of poultry and livestock manure(of which pig,
cattle and chicken manure account for 75.6%) are produced every year in China [4].
AD is a suitable method to convert crop straw and manure into biogas and could serve
the dual role of producing renewable energy and reducing waste.
The AD energy production method has rapidly developed; however, it is not yet an
effective renewable energy generator because of the diversity of substrates, low
reactor efficiency and issues related to the treatment of supernatants and residues

(by-products of biogas plants) involved in the process. Empirical studies evaluating

the distribution and mass balances of C, N, and P are essential to understand, predict,
and possibly control bioenergy production through AD. Many attempts to optimize
methane yields are closely associated with the distributions of C, N, and P, including
the magnesium ammonium phosphate (MAP) process [5] and ammonia stripping [6,
7]. The effective reuse of organic carbon and nutrients in supernatants and residues is
another way to improve the total efficiency of AD and is also based on the specific
regulation of the C, N, and P distributions within each substrate. Mass balance is also
crucial to the quantification of bioenergy production and nutrient recovery potential
from AD [8], which is needed to evaluate the contribution of AD processes to
elemental cycles. Furthermore, mass balances are often identified with reference to
conventional chemical and physical parameters that provide an indication of the fate
of compostable materials throughout AD [9, 10].
Several studies have addressed the C, N, and P distributions and mass balance in
AD, including the nutrient (N, P and K) mass balance of source-segregated domestic
food waste in AD [2]; and particle size distribution, trace element patterns (N, P, K,
Ca, Fe, Mg, S, Al, Cu, Mn and Zn) and mass balances in a full-scale anaerobic
digestion plant treating pig slurry [11]. However, these studies have rarely considered
the mass balance of C which is most crucial to the AD process. Although the mass
balance of C, N, and P in an integrated microalgae growth-anaerobic digestion
process [8] has been studied, this study was limited to the substrate of microalgae
which has not been introduced as a substrate in biogas plants, whereas the biogas

plants analyzed relied on various substrates such as manures and straws during
operation [12]. Therefore, it is necessary to determine the C, N, and P distributions
and mass balances of manures and straws to provide a theoretical foundation for
optimizing methane yield. On the other hand, analyzing the relationship between the
products and substrates is crucial to the development of key process parameters and a
predictive model of products related to the generation of renewable energy via AD.
Few studies have shown that lignin content is a significant parameter affecting
methane production potential in the AD of different substrates [13, 14], however,
additional key parameters related to organic carbon and nutrient by-products of AD
must be further investigated to provide a reference for the effective reuse of those
by-products. Furthermore, data summarizing the regulation of C, N, and P
distributions, mass balances and relationships within a range of substrates involved in
AD are scarce in the literature.
Therefore, the objectives of the current work are (1) to evaluate biogas production
and the liquid and solid distributions of C, N, and P after the AD of corn straw, rice
straw, wheat straw, swine manure from pigs fed feedstuff (swine manure 1) and
foodstuff (swine manure 2), cattle manure, and chicken manure and to provide
explanations for the distributions; (2) to describe the mass balances of different
substrates calculated as the mass of C, N, and P in gas, liquid and solid form before
and after AD; and (3) to investigate the relationships between gas and liquid products
and the compositions of substrates.

2. Materials and methods

2.1. Substrates and inoculums

Both animal manure and straws were included in the study. Swine manure1 and
chicken manure were collected from pig and chicken farms located in Putian, Fujian,
China. The animals were fed on feedstuff. Cattle manure was obtained from a cattle
farm close to Xiamen, Fujian, China. Swine manure2 was collected from San Ming
and provided by a local villager. The swine were fed on foodstuff. Manures were
collected as semi-solids and were not mixed with cleaning or drinking water from the
animal houses. They were then frozen until used in the batch experiments. Corn straw,
rice straw and wheat straw were obtained from villagers in San Ming and Mu Dan
Jiang, respectively. Dried straws were chopped using a grinder (Hummer 900) to an
approximate size of 2-3 mm. Inoculum from a laboratory-scale working swine manure
1 digester, which had been maintained at 37C for 2 weeks before the test to remove
most of the remaining methane, was used. The TS and VS/TS of the inoculum used
were 2.93% and 58.8%, respectively.
2.2. Experimental design and set-up
The BMP of animal manures and straws were determined using a batch technique
based on the methods described by Mller et al. [15]. Each substrate was tested in
triplicate using Erlenmeyer flasks with a working volume of 250 mL kept at
mesophilic temperature (370.5C)The initial volatile solid (VS) ratio of substrate to
inoculum (S/I) was maintained at 2:1 for all experimental set-ups. After the inoculum
was mixed with the substrate in the reactors, tap water was added to an effective
liquid volume of 200 mL. All the reactors were tightly closed with rubber septa and

screw caps after being flushed with N2. Digestion continued until no further gas
production was observed (45 days). All reactors were shaken thoroughly on workdays
to ensure mixing of the reactor contents.
The compositions in the different batch set-ups were as follows. In set-up 1, the
initial volatile solid (VS) loading included 20 g VS/L of corn straw, rice straw, wheat
straw, swine manure1, swine manure2, cattle manure and chicken manure. Set-up 2
was designed to provide insight into the relationships between products and substrates;
as such, the AD processes were based on initial VS loadings of 30, 35, and 40 g VS/L
of all substrates. Blank digesters containing the same amount of inoculums and water
were tested to correct for biogas production and C, N, and P in their liquid and solid
forms.
2.3. Analytical Methods
Total solids (TS), volatile solids (VS), chemical oxygen demand (COD), total
phosphorus (TP), total Kjeldahl nitrogen (TKN) and total ammonium nitrogen (TAN)
analyses of raw manures and straws were performed in accordance with the APHA
Standard Methods [16]. Total carbon (TC) and total organic carbon (TOC) were
analyzed using a CHN Elemental Analyzer (Perkin Elmer, USA). TOC was
determined by adding HCl, following the established method [17]. Protein content
was determined by multiplying the difference between TKN and TAN by a factor of
6.25.
The concentrations of metals present (Ca, Mg, K, and Na) were determined by
inductively coupled plasma-optical emission spectroscopy (ICP-OES). The lipid

content was determined via Soxhlet extraction using diethyl ether as a solvent. The
volatile fatty acid (VFA) concentration was determined using GC-FID (SP-2100A).
The cellulose, hemicellulose, and lignin contents were estimated using a VELP
FIWE6 Fiber analyzer (VELP, Italy).
The biogas volume (V1) was measured by its displacement of water. The methane
content in the produced biogas was determined using GC-TCD (SP-2100A). Digestate
samples from both the reactors, including the blank digesters, were weighed (W1) and
centrifuged in a Rotofix 32 laboratory centrifuge for 10 min at 9,000 g at room
temperature. The volume of liquid (V2) was measured and analyzed in terms of COD,
TP, TKN and ammonium nitrogen (NH4+-N) in accordance with the APHA Standard
Methods [16]. The TOC was also determined using a Shimadzu TOC analyzer. Solid
phase products (residue) were weighed (W2), freeze-dried, weighed again (W3) and
analyzed according the substrate analysis methods mentioned above.
2.4. Mass balance calculations
The mass balances of C, N, and P were calculated prior to and after AD. The initial
elemental mass was obtained by multiplying the fractions of C, N, and P by the mass
of the substrate. The final C mass was calculated using Eqs. (1) and (2); these
equations assume that (1) biogas production occurs under normal temperature and
pressure, (2) the amount of N in the gas phase is neglected given the very low
ammonia concentration in biogas [18, 19], and (3) TKN is used to represent total
nitrogen (TN) which based on the amount of NO3-N and NO2-N only represent a
small percentage of TN in liquid after AD [20, 21].

(1)
(2)
(3)
(4)
where W is the final mass, mg; Cliquid is the concentration of liquid C, mg/L; Csolid is
the amount of solid C, mg/g;

is the density of the liquid; V is the total

effective volume of the digesters; V3 is the volume of the lost liquid in the process of
freeze-drying, and V1, V2, W1, W2 and W3 are as described above. The final N and P
masses were obtained by combining the liquid and solid masses of these two elements.
The final masses were corrected using the corresponding final mass of the blank
digester.
2.5. Data analysis
Data from the digesters were subjected to an analysis of variance (ANOVA) to
determine whether the observed differences among the seven substrates were
significantly different. Differences were considered significant below a probability of
0.05 (p < 0.05). Correlation analyses between products and substrate compositions
were performed using SPSS 19.0. Graphs and data processing were completed using
OriginPro 8.0.

3. Results and discussion

3.1. Characterization of the substrates
The substrate and inoculum characteristics are shown in Table 1. The VS were
primarily comprised of lipids, proteins, cellulose, hemicelluloses and lignin. Chicken

manure, cattle manure, and swine manures 1 and 2 all contained higher amounts of
proteins and lipids than did the straws (corn, rice and wheat straws), whereas the
straws all exhibited a higher fiber content (approximately 75% of TS), as represented
by cellulose, hemicelluloses, and lignin. Furthermore the amounts of proteins, lipids
and lignin were considerably different among swine, cattle and chicken manure, even
between swine manures 1 and 2. This result is expected, given that the characteristics
of manures are strongly determined by the type of foods [15, 22]. The proportions of
lignin and cellulose in swine manure 2 were higher than those in swine manure 1,
which can be attributed to the fact that swine are fed on foodstuff, which contains
greater amounts of lignin complexed with cellulose than feedstuff does.
Fig. 1 shows the C, N, and P compositions of the seven substrates. C is the primary
substance found in the substrates, accounting for significant portions (over 82.56% in
manures and over 97.71% in straws) of every substrate. The TKN and TAN values in
the manures were lowest in cattle manure (29.86 and 1.33 g/kg TS, respectively) and
highest in chicken manure (65.78 and 17.58 g/kg TS, respectively). TAN levels fell
below the detection limit of the measuring method in every straw. TP in swine
manures 1 and 2 (34.97 and 30.80 g/kg TS, respectively) was considerably higher
than in other substrates.
3.2. Anaerobic digestion performance
To obtain accurate measurements of liquid and solid C, N, and P, the corresponding
parameter values from the blank digester were used to correct all of the other values;
as a result larger errors in the determination of liquid and solid C, N, and P levels

would arise under lower VS loading conditions. Therefore, the initial VS loadings
designed in this study were higher than that used in other reported works [14, 23].
Consequently, the evaluation of CH4 yields in this study did not occur under optimal
conditions; nevertheless, the CH4 yields obtained for the straws are comparable to
those found in other research [24] and in the ranges of 0.1-0.44, 0.1-0.37, 0.01-0.5
m3/kg VS reported for swine, cattle and chicken manures, respectively [25]. Swine
manure1 produced the highest CH4 yield of 210 mL/g VS, whereas cattle manure
produced the lowest CH4 yield of 124 mL/g VS. Swine manure1 showed a higher CH4
yield than did swine manure2 (164 mL/g VS) due to the different characteristics of the
two manures caused by the different feed types. The CH4 yields from each substrate
with initial VS loadings of 30, 35, and 40 g VS/L are provided in the supplementary
information.
As shown in Table 2, the measured parameters reflect the diversity of each
substrate at an initial VS loading of 20 g VS/L. The ANOVA of the data indicated
that the variations in parameters of different substrates were statistically significant (p
< 0.05). The pH and total VFA are important indicators used for monitoring the AD
process [26]. The pH values of the different digesters all fell within a suitable range of
6.5 to 8.5 [27], and the total concentrations of the seven detected VFA exhibited low
values (total concentrations of 198-425 mg/L) compared with the threshold of VFA
(4000 mg/L) [28]. VS is another important parameter for measuring biodegradation
and is a direct indicator of the metabolic status of some of the most delicate microbial
groups in an anaerobic system [29]. The VS destruction results for the various

substrates indicated that the highest VS removal (60%) occurred in the swine manure1
digester, whereas straws and cattle manure exhibited lower VS removal (below 51%),
with only minor differences observed between the straws and cattle manure (p > 0.05).
Given the substrates characteristics and the poor biodegradability of straws and cattle
manure, the results obtained met with our expectations.
3.3. Liquid and solid C, N, and P
AD leads to the release of organic carbon and nutrients, which generally increase
contamination of the supernatant. In this study, COD/TOC, NH4 +-N, and TP
concentrations represent the liquid phases of C, N, and P, respectively, and the C, N,
and P solid mass fractions represent the solid phases of C, N, and P, respectively. The
COD concentrations in the supernatant reflect both the untreated portion of a
substrates soluble COD and hydrolyzed organic particulate matter [30]. As shown in
Fig. 3a, chicken manure exhibited higher COD concentrations in the supernatant
(1717 mg/L) than any of the other six substrates, which indicates that a higher
proportion of untreated substrate could exist in the chicken manure digester system.
The NH4+-N concentrations in the swine manure1 (415 mg/L), swine manure2 (264
mg/L) and chicken manure (1,574 mg/L) supernatants were likely considerably higher
than the concentrations in the other four substrates (below 57 mg/L) because the
swine and chicken manures had higher TAN levels than the other substrates (Table 1).
Another feasible explanation relates to the anaerobic bioconversion of proteins
contained in animal manure into amino acids and then ammonia [31]. The TP
concentrations in the swine manure1 (7.2 mg/L) and 2 (4.9 mg/L) supernatants were

lower than those for the other five substrates. The concentrations of COD/TOC,
NH4 +-N, and TP in the supernatant of each substrate with initial VS loadings of 30, 35,
40 g VS/L are provided in the supplementary information.
The mass fractions of solid (residue) C, N, and P in the different substrates are
shown in Fig. 3b. The C mass fractions (over 478.8 mg/g) in the straw residues are
considerably higher than those in the manures. Higher N and P mass fractions were
observed in swine manure1 (43.3 and 44.4 mg/g, respectively) and swine manure2
(36.9 and 43.7 mg/g, respectively), the recovery of which could improve the total
efficiency of swine manure AD. These results can assist in the development of a
sustainable method to recover C, N, and P from straw and manure residues.
3.4. C, N, and P distributions and mass balances in the AD of different substrates
The C, N, and P distributions were calculated based on the ratios of gas, liquid, and
solid C, N, and P in the seven substrates for an initial VS loading of 20 g VS/L (Table
3). Most C remained in solid form (over 60.3%), whereas some was present in liquid
form (below 7.6%). The efficiency of Cs conversion from the substrate into methane
was highest for swine manure 2 (15.8%), whereas that for rice straw (11.2%) was
higher than that for corn straw (8.2%) and wheat straw (9.5%). Liquid N was more
abundant in chicken manure (81.1%) than in the other six substrates, which can be
attributed to chickens common excretion of urine and feces, as they are monogastric
animals [32]. Liquid P was present in straws, whereas over 80% of the P in manures
existed as precipitate in solid form, particularly in the swine manure (over 97%). This
phenomenon is likely due to the presence of greater concentrations of calcium and

magnesium in pig manure, which are able to re-fix phosphates as chemical

precipitates [33, 34]. Moreover, both solid N and may exist as precipitates, e.g., of
struvite (NH4MgPO4.6H2O), within the digester system [2]. Furthermore, the C, N
and P distributions were slightly different between the two types of swine manure,
which indicates that not only the biogas production but also the C, N and P
distributions were affected by the animals feed type for the manure substrates.
The mass balances of different substrates in AD were calculated as the mass of gas,
liquid and solid C, N, and P before and after AD. The balances of C (Fig. 4 a) are
acceptable considering the net losses of 4-10%. Sampling, analytical errors and biogas
production under abnormal temperature and pressure conditions constitute potential
sources of error. Understanding the C balance may help plant operators evaluate the
efficiency of the AD process and modify the process parameters in situ to improve the
degradation efficiency of AD. In addition, Fig. 4b shows N losses of 3-15% for
various substrates, where 3-7% for manures and 9-15% for straws. There were also
modest losses in P content (3-9%) similar to those of N following the AD processes,
according to our analysis. The relative nutrient balances during AD may influence the
possibilities of reusing digestate considerably.
3.5. Relationships between the products and substrate compositions
To explore possible relationships between the C, N, and P contents in substrates
and their AD products, the ratios of C, N, and P in supernatants and in substrates with
different VS loading were calculated. Fig. 5 shows that the C, N, and P ratios were
similar regardless of the VS loading, reflecting the fact that C, N, and Pare

specifically regulated in the supernatant of different substrates. However, the ratios of

C, N, and P in chicken manure varied with VS loading; in particular, the proportion of
C was inhibited by high NH4+-N concentrations at the higher VS loading but was not
significantly inhibited at the lower VS loading. To investigate the relevant factors
affecting the products, linear regression analyses of CH4 yields and COD, TOC,
NH4 +-N, and TP concentrations in the supernatant of each substrate (Table 1) using
SPSS were performed, and correlations were determined. A significant correlation
was found between the CH4 yield and lignin content of each substrate. With respect to
C, N, and P in the supernatants, significant correlations were discovered between
COD/TOC concentrations and proteins in the substrates, between NH4+-N
concentrations and TAN/proteins in the substrates, and between TP concentrations
and potassium in the substrates.
The statistical analyses of the relationships between products and the factors
mentioned above were performed by linear regression, and the results are displayed in
Fig. 6. A negative correlation was found between CH4 yields and lignin content in the
substrates (Fig. 6 a), which is in agreement with previous studies [13, 14]. Moreover,
the COD and TOC concentrations in the supernatants increased with increases in the
substrate protein levels (Fig. 6 b), suggesting that soluble COD existed relative to
substrate protein concentrations. This trend can be explained by the anaerobic
bioconversion of proteins into VFAs and then into CH4 or CO2 [35], as well as the fact
that VFAs not only are formed through bacterial metabolism but also may already be
present in considerable amounts as organic carbon in influent [36]. Positive

correlations were also found between NH4+-N concentrations in the supernatants and
TAN/proteins in the substrates analyzed (Fig. 6c), as well as between TP in the
supernatants and potassium in the substrates (Fig. 6d). This correlation is excepted
because NH4+-N in the supernatants was derived from TAN in the substrates and the
anaerobic bioconversion of proteins in the substrates [31], whereas TP in the
supernatants occurred in direct proportion to potassium in the substrates [33]. These
results strongly suggest that lignin, TAN, proteins and potassium within substrates
have an important impact on products produced through AD. In addition, derived
linear models may be used to predict CH4 yields and COD, TOC, NH4+-N and TP
concentrations in the supernatants of different substrates.

4. Conclusions
The approach used to measure C, N, and P distributions and mass balances in this
paper may be useful for evaluating the efficiency of the AD process, optimizing
methane yield, and establishing a mechanism by which digestate can be reused. There
are delicate relationships between products and substrates, as revealed by the common
regulation of different substrates: CH4 yields, COD/TOC, NH4+-N and TP
concentrations in supernatants were largely dependent on the presence of lignin,
proteins, TAN/proteins and potassium in substrates, respectively. Additionally, the
derived linear models could be used to predict the methane production and COD,
TOC, NH4+-N, and TP concentrations in supernatants. Future research into the C, N,
and P distributions in the co-digestion of different substrates and into C, N, and P
distribution predictions would help elucidate these issues further.

Acknowledgements
This work was supported by the National Basic Research Program of China
(2013CB733505), the National Natural Science Foundation of China (41276101), and
Shenzhen

Science

and

Technology

Research

and

Development

Project

(JCYJ20120615161649123).

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Fig. 1 Average C, N, P compositions in different substrates: CS, corn straw; RS, rice straw;
WS, wheat straw; SM1, swine manure1; SM2, swine manure2; CAM, cattle manure; CHM,
chicken manure.

Fig. 2 CH4 yields for seven substrates with an initial VS loading of 20 g VS/L CS, corn straw;
RS, rice straw; WS, wheat straw; SM1, swine manure1; SM2, swine manure2; CAM, cattle
manure; CHM, chicken manure.

Fig. 3 (a) NH4+-N, COD and TP in the liquid and (b) C, N and P in the solid from the
anaerobic digestion of different substrates with initial an VS loading of 20 g VS/L CS, corn
straw; RS, rice straw; WS, wheat straw; SM1, swine manure1; SM2, swine manure2; CAM,
cattle manure; CHM, chicken manure.

Fig. 4 Distribution of C, N, P during gas-liquid-solid threephases in the anaerobic digestion

of different substrates CS, corn straw; RS, rice straw; WS, wheat straw; SM1, swine
manure1; SM2, swine manure2; CAM, cattle manure; CHM, chicken manure.

Fig. 5 C, N and P ratios in the supernatants and substrates with different VS loading during
the anaerobic digestion of different substrates CS, corn straw; RS, rice straw; WS, wheat
straw; SM1, swine manure1; SM2, swine manure2; CAM, cattle manure; CHM, chicken
manure.

Fig. 6 Correlations between supernatant quality and compositions of substrates CS, corn
straw; RS, rice straw; WS, wheat straw; SM1, swine manure1; SM2, swine manure2; CAM,
cattle manure; CHM, chicken manure.

Table 1 Studied substrate compositions

Characteristics

CS

RS

WS

SM1

SM2

CAM

CHM

Inoculum

TS%

91.820.16

92.460.22

92.180.25

24.040.39

27.320.49

21.640.71

24.720.61

4.020.35

VS%

86.300.36

77.400.35

74.950.42

15.560.24

18.660.32

17.310.51

17.430.25

1.860.21

TC(g/kg TS)

465.041.23

398.571.07

401.521.91

351.313.24

299.741.39

384.712.51

354.822.98

ND

TOC(g/kg TS)

416.631.31

367.042.02

369.610.98

276.921.69

233.244.13

334.711.95

295.422.56

ND

CODt (g/kg TS)

1075108

98864

96589

139471

1362148

101252

148096

65875

CODs (g/kg TS)

82.428.61

64.453.82

60.205.26

75.836.85

63.255.85

20.363.25

149.4910.2

35.286.25

TP(g/kg TS)

1.020.08

0.960.04

0.890.26

34.970.54

30.80.82

3.630.06

9.550.19

ND

TKN(g/kg TS)

9.840.78

4.590.49

4.860.58

41.680.49

32.250.93

29.860.83

65.780.96

ND

TAN(g/kg TS)

BD

BD

BD

5.140.04

4.790.14

1.330.12

17.580.07

4.350.56

Calcium(g/kg TS)

3.800.51

1.940.23

1.820.18

58.283.68

61.175.32

23.243.58

27.982.68

ND

Magnesium(g/kgTS)

1.130.15

0.990.10

0.820.09

11.480.15

6.580.35

4.980.20

4.030.18

ND

Potassium(g/kg TS)

17.461.23

25.051.38

201.26

4.680.63

5.920.23

10.230.53

32.391.25

ND

Sodium(g/kg TS)

1.350.12

1.230.08

1.150.10

15.401.02

6.440.86

2.010.15

5.790.23

ND

Protein(g/kg TS)

61.5

28.7

30.4

227.8

171.6

178.3

207.5

ND

VFA(g/kg TS)

BD

BD

BD

4.320.30

4.15 0.25

2.05 0.15

6.34 0.36

0.020.004

Lipid(g/kg TS)

3.740.35

5.230.40

5.680.51

57.103.89

65.385.32

20.153.20

27.852.98

ND

Cellulose (%)

42.611.12

42.512.45

40.141.59

7.241.15

15.851.64

192.59

20.211.56

ND

Hemicellulose (%)

29.232.11

24.231.59

25.121.48

19.350.84

18.350.92

17.891.16

21.352.15

ND

Lignin (%)

10.150.91

7.540.58

10.30.98

4.250.25

7.640.58

19.50.26

2.250.24

ND

ND, not determined; CS, corn straw; RS, rice straw; WS, wheat straw; SM1, swine manure1;
SM2, swine manure2; CAM, cattle manure; CHM, chicken manure; BD, below the detection
limit of the measuring method.

Table 2 Anaerobic digestion performance

Parameter

CS

RS

WS

SM1

SM2

CAM

CHM

TS initial (g/L)

21.120.8

23.991.2

24.781.1

31.401.5

28.371.2

25.351.0

29.241.3

TS final (g/L)

12.781.6

14.051.5

14.861.6

15.221.8

14.642.0

14.261.8

14.681.8

TS removal (%)

39.482.8

41.433.1

40.033.0

51.533.7

48.403.1

43.753.2

49.793.4

0.002

P1=0.502

P1=0.844

P1=0.016

P1=0.030

P1=0.182

P1=0.019

VS initial (g/L)

19.850.5

20.080.8

20.150.3

20.320.2

19.380.3

20.280.1

20.620.5

VS final (g/L)

10.151.15

9.781.23

10.251.25

8.070.82

8.620.96

10.581.21

8.810.93

VS removal (%)

48.884.6

51.293.7

50.864.4

60.294.6

55.523.4

47.834.7

57.783.8

0.002

P1=0.443

P1=0.692

P1=0.014

P1=0.038

P1=0.808

P1=0.022

28550

32453

27058

35032

28325

23238

35333

511

521

521

601

581.5

551

581

14528

16825

14032

21015

16412

12718

20415

0.002

P1=0.402

P1=0.866

P1=0.021

P1=0.383

P1=0.407

P1=0.029

Biogas yield
(mL/g added VS)
Methane (%)
Methane yield
(mL/g added VS)

pH

7.250.15

6.730.13

6.850.2

7.320.15

7.280.1

6.930.08

8.340.12

VFA (mg/L)

25815

35320

28618

42536

38632

23425

19823

0.000

P1=0.003

P1=0.107

P1=0.002

P1=0.003

P1=0.227

P1=0.019

CS, corn straw; RS, rice straw; WS, wheat straw; SM1, swine manure1; SM2, swine
manure2; CAM, cattle manure; CHM, chicken manure.
p: Indicates a statistical difference between the performances of different substrates.
p1: Indicates a statistical difference between the performance of CS and another substrate
(RS, WS, SM1, SM2, CAM and CHM).

Table 3 C, N and P distributions of seven substrates

CS

RS

WS

SM1

SM2

CAM

CHM

C%
CH4 in gas

8.21.1

11.21.3

9.51.2

14.11.3

15.82.3

8.51.2

12.81.8

CO2 in gas

7.91.2

10.31.1

8.71.2

9.41.8

11.41.8

6.91.1

9.31.4

TOC in liquid

2.10.2

2.10.1

2.20.2

2.60.2

4.20.3

2.80.1

3.80.3

TC in liquid

4.10.3

4.00.2

4.30.4

5.10.5

7.50.5

5.40.3

7.60.6

TOC in solid

57.32.8

53.13.1

56.83.2

44.34.3

39.62.8

60.93.5

47.54.2

TC in solid

76.44.1

69.54.9

72.14.7

64.96.4

60.34.6

74.65

64.15.5

In liquid

25.92.3

38.42.8

36.93.2

18.12.8

21.52.1

5.41.1

81.13.1

In solid

64.23.5

47.63.8

47.33.8

79.34.1

75.73.6

86.84.5

15.11.5

In liquid

82.72.8

84.543.2

80.312.5

0.820.3

0.710.3

16.012.3

9.81.9

In solid

11.381.2

13.041.8

18.722.3

97.012.8

97.182.5

803.5

79.794.8

N%

P%

CS, corn straw; RS, rice straw; WS, wheat straw; SM1, swine manure1; SM2, swine
manure2; CAM, cattle manure; CHM, chicken manure.

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