Kinetics of protein extraction from excess sludge by thermal alkaline treatment

doi:10.21203/rs.3.rs-1974348/v1

Download PDF

Research Article

Kinetics of protein extraction from excess sludge by thermal alkaline treatment

https://doi.org/10.21203/rs.3.rs-1974348/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

Protein can be extracted effectively from excess sludge by thermal alkaline process. During the extraction process, insoluble protein in sludge was turned into soluble protein, and part of the soluble protein was hydrolyzed into peptide. Then part of the soluble protein and peptide were degraded into amino acids, generating by-products: water, carbon dioxide and ammonium. Therefore, to obtain the maximum concentration of soluble protein, it is necessary to establish a kinetic model describing the process of sludge protein extraction. The report indicated that the extraction mechanism of sludge protein was according to first-order continuous reaction kinetic. A kinetic model of the reaction was established to explain the relationship between the rate constant, temperature and hydrolysis time of the first-order reaction. The optimum conditions were as follows: 0.1 M NaOH, reaction temperature 70°C, hydrolysis time 4 h. Protein separated and obtained from sludge hydrolysate by centrifugation and filtration, which could be used as a feed additive or foaming agent.

Excess sludge

Thermal alkaline process

Protein

Kinetic model

Wastewater treatment plants produce a large number of excess sludge in sewage treatment every year. Its treatment and disposal pose a significant challenge to the operation of wastewater treatment plants (Bou-Sarkis et al., 2022). The composition of residual sludge is very complex, including microorganisms, organic matter, extracellular polymers (EPS) and inorganic salts (Christensen et al., 2015; Zhang et al., 2016). Sludge contains rich organic matter (protein, polysaccharide, nucleic acid and organic acid), which could be used to obtain high calorific value fuels such as methane (biogas), hydrogen (H₂) and volatile fatty acids (VFA) through anaerobic digestion (Geng et al., 2022; He et al., 2022; Li et al., 2016; Wu et al., 2022). In addition, abundant resources in sludge could be recovered through phosphorus separation and protein extraction technologies. Extracellular polymeric substances (EPS) contribute significantly to sludge quality compared to intracellular substances. They account for 60–80% of sludge biomass and are an essential part of microbial aggregates used to hold them in a three-dimensional matrix (Bou-Sarkis et al., 2022; Sheng et al., 2010). EPS can be divided into soluble EPS (S-EPS), loosely bound EPS (LB-EPS) and tightly bound EPS (TB-EPS), composed of proteins, carbohydrates, humic acid-like substances, uronic acids, nucleic acids and biopolymers (Wang et al., 2018).

In recent years, protein extraction from sludge has attracted much attention. Protein accounts for about 40 ~ 60% of the remaining sludge dry weight (Wang et al., 2019). The steps for protein recovery from excess sludge usually include screening, treatment, filtration, precipitation, drying, recovery, characterization and identification (Boleij et al., 2018; Lotti et al., 2019; Seviour et al., 2019; Xiao et al., 2017). Recycled protein could be used as animal feed (Hwang et al., 2008), foam extinguisher (Gao et al., 2020), wood adhesive (Pervaiz and Sain, 2011), compound fertilizer (Liu et al., 2009) and corrosion inhibitor (Su et al., 2014). This would help alleviate global energy shortages. Sludge dissolution is necessary before protein recovery. Sludge hydrolysis methods have been studied for many years, including physical, chemical and biological methods, as well as including some combined treatment process (Gao et al., 2020; Park et al., 2019; Wong et al., 2020; Zou et al., 2022). The thermal alkaline hydrolysis has a good effect on sludge cracking. Alkali could cause the dissolution of membrane proteins and saponification of membrane lipids, thus destroying microbial cells.

Yu Lin et al. get the protein extraction rate of 69% using calcium oxide as a hydrolytic agent under the conditions of extraction time 2 h, pH 12, and temperature 130°C (Xiang et al., 2017). Additionally, the foamability and foam stability were close to 450% and 88.8%, and the dewatering performance of the hydrolysis sludge improved by 93.1% (Gao et al., 2020). And for thermal treatment, medium temperature treatment has attracted more attention compared with high temperature (> 100°C) and high pressure (> 10 MPa) treatment (Xue et al., 2015). Because the latters require high energy input and special equipment (Suarez-Iglesias et al., 2017; Xiao and Zhou, 2020). However, most of these studies only consider how to increase the soluble protein content, ignoring the ensuing energy consumption and costs. Therefore, costs and energy inputs need to be carefully analyzed. In addition, in the process of protein dissolution, the conversion of proteins to peptides and degradation of proteins and peptides to amino acids and ammonia are ignored. The determination of protein concentration in sludge and solution is usually by Kjeldahl method (Academy, 2018), Dumas method (Salette and Dumas, 1970), Coomash bright blue method (Bradford and Williams, 1976), BCA method (Akins and Tuan, 1992) and Lowry method (Lowry and Hunter, 1945). These methods can not accurately measure the protein concentration because the nitrogen of nucleic acids, amino acids and ammonia generated by protein hydrolysis will interfere with the determination of protein (Lamp et al., 2020). A more systematic and detailed study of these treatments is necessary to obtain more appropriate and economically viable therapies.

The thermal alkaline treatment is the employed sludge hydrolysis technology previously (Hui et al., 2022). Here, we analyzed the kinetics of sludge protein extraction to obtain the maximum content protein. The recovered protein molecular weight distribution, functional groups and the secondary structure were also researched. Mass spectrometry (MS) was used to identify the extracted protein, which was conducive to understand the underlying extraction mechanism. In addition, we aimed to find out its structural components by combining the characterization information of the extracted protein.

2.1. Materials and feedstock analysis

The excess sludge was extracted from the second pond of Dalian Waste Water Plant, which was dried in an oven at 50°C over 24 h before being analyzed and used as a feedstock in the thermal alkaline hydrolysis experiments. The dried sludge was first broken and ground and then sieved through a 100-mesh sieve to prepare the sample.

Table 1 lists the chemical characteristics of the excess sludge. The VSS (volatile suspended solids) content of the dried sludge was 48.56% according to the standard method (Eaton et al., 1966). Protein content in raw sludge was measured by the hydrolyzed amino acid content with the automatic amino acid analyzer. Carbohydrate was determined by the Dubois procedure using glucose as equivalent (Dubois et al., 1956). The element of carbon (C), hydrogen (H), and nitrogen (N) were determined by an elemental analyzer (VARIO MACRO, Germany). The data was calibrated to 2 mg of Elmer Organic Analytical Standard of Acetanilide (formula:C₈H₉NO) with known abundances of carbon (71.11%), hydrogen (6.67%), nitrogen (10.37%) and oxygen 11.85%) (Chacón-Parra et al., 2021). The margin of error was ± 0.1% for carbon, hydrogen and nitrogen calculated against 3 replicates. Amino acids were stained with ninhydrin (Smith and Agiza, 1951). Ammonium concentration were determined by Nessler method using ammonium chloride as standard (Prince, 1945).

2.2. Thermal alkaline hydrolysis experiment

Dried excess sludge powder was incubated in 0.1 M NaOH (40 g/L) for 1–5 h, stirring with a magnetic stirrer at 180 rpm at temperature (30, 50, 70 and 90°C). After centrifugation at 4000 rpm for 30 min at 4°C, the sludge residue was collected and dried. The protein in the supernatant was precipitated out by adding an equal amount of trichloroacetic acid (C₂HCl₃O₂, 10% wt). The precipitated protein was collected by centrifugation at 4000 rpm for 15 min at 4°C and lyophilized.

2.3. Analysis of products from thermal alkaline hydrolysis

SDS-PAGE was performed using BeyoGel™ Plus Precast PAGE Gel (Hepes, 4–20%). The protein samples were prepared in Loading Buffer (Bromophenol blue, 5X). The protein was denatured by incubation at 100°C for 5 min. Subsequently, a 10 µL sample was loaded per well. The Prestained Color Broad Range Protein Ladder was used as a molecular weight marker. The gel electrophoresis was performed at 150 V for 40 min. For visualization of the protein, the operational method of the BeyoBlue™ Coomassie Blue Super Fast Staining Solution (Beyotime) was according to the manufacturer’s instructions. The protein was analyzed by High-Performance Liquid Chromatography coupled with a LTQ-Orbitrap mass spectrometer (Thermo Fisher, Bremen, Germany).

2.4. Kinetic parameters

Sludge proteins (the molecular weight > 10,000 Da) could be precipitated by trichloroacetic acid (10% wt). The amino acids concentration were detected by an automatic amino acid analyzer, which was stained with ninhydrin. Polypeptide concentration was calculated by the hydrolyzed amino acid content minus free amino acid content in the supernatant. The concentration of ammonium was determined by Nessler reagent colorimetry using ammonium chloride as the standard.

As shown in Fig. 1, through the thermal alkaline process, protein in the aqueous phase was continuously hydrolyzed into peptides, oligopeptides, amino acids, and ammonia. In order to simplify the reaction model, the following assumptions were made: The protein content in raw sludge was identified as the initial reaction concentration. The initial protein concentration in the aqueous phase was zero. The protein, peptides, amino acids, and ammonia concentration in the filtrate were identified as the reaction intermediates concentration. Based on the assumptions above, the sludge protein dissolution was considered as a first-order reaction, and the protein in the aqueous phase transformation and degradation was also regarded as a first-order reaction. Therefore, the overall reaction was seen as a first-order continuous reaction. For the first-order continuous reaction, it could be expressed as equations (1)-(9).

The total volume of protein remained changeless in the experimental process. To obtain the kinetic parameters, the MATLAB function ODE45 was used to solve the ordinary differential equations (1)-(5). Based on the Laplace transformation, the transformed results were equations (6)-(9). In MATLAB, the least square algorithm was used for the nonlinear fitting of experimental data. The bounds of the kinetic parameters were set between 0 and 1. The Arrhenius parameter error was calculated as a 95% confidence interval.

\(\frac{{d{C_0}}}{{dt}}= - {k_1}{C_o}\)

(1)

\(\frac{{d{C_1}}}{{dt}}={k_1}{C_o} - {k_2}{C_1}\)

(2)

\(\frac{{d{C_2}}}{{dt}}={k_2}{C_1} - {k_3}{C_2}\)

(3)

\(\frac{{d{C_3}}}{{dt}}={k_3}{C_2} - {k_4}{C_3}\)

(4)

\(\frac{{d{C_4}}}{{dt}}={k_4}{C_3} - {k_5}{C_4}\)

(5)

\({C_0}={C_{A0}}{e^{ - {k_1}t}}\) (6)

\({C_1}=\frac{{{k_1}}}{{{k_1} - {k_2}}}{C_{A0}}({e^{ - {k_2}t}} - {e^{ - {k_1}t}})\)

(7)

\({C_2}=\frac{{{k_1}{k_2}}}{{{k_1} - {k_2}}}{C_{A0}}(\frac{{{e^{ - {k_2}t}} - 1}}{{{k_3} - {k_2}}} - \frac{{{e^{ - {k_1}t}} - 1}}{{{k_3} - {k_1}}})\)

(8)

\({C_3}=\frac{{{k_1}{k_2}{k_3}}}{{{k_1} - {k_2}}}{C_{A0}}\left[ {\frac{{{e^{ - {k_2}t}} - 1}}{{({k_3} - {k_2})({k_4} - {k_2})}} - \frac{{{e^{ - {k_1}t}} - 1}}{{({k_3} - {k_1})({k_4} - {k_1})}}} \right]\)

(9)

Where C_A0 was the initial protein concentration of raw sludge, mg/L; C₀ was the protein concentration of sludge residue, mg/L; C₁ was the protein concentration in the supernatant, mg/L; C₂ was the peptides concentration in the supernatant, mg/L; C₃ was the amino acids concentration in the supernatant, mg/L; C₄ was the ammonium concentration in the supernatant, mg/L; k_i (i = 1, 2, 3, 4, 5) was the first-order rate constant, h^− 1; and t was the hydrolysis time, h.

Table 1

The elemental composition of the waste ctivated sludge.
Composition	Content (%)
TS	20.47
moisture	79.53
VSS/TS	48.56
C	18.31
H	4.38
N	3.39
O	16.14
S	0.53
Protein	21.19

3.1. Characterization of thermal decomposition of excess sludge

As shown in Fig. 2, the thermal gravity (TG)curve gradually decreased as the temperature increased to 900°C, while the differential thermal gravity (DTG)gradually increased. The weight loss of excess sludge in a nitrogen atmosphere could be roughly divided into three stages. The first stage was the dehydration and drying process, which was mainly involves in the sublimation of free water and combined water. This took place at the temperature ranging from 60 to 180°C. The second stage was the decomposition of excess sludge, which was principally the volatilization of volatile organic compounds and the decomposition of the majority of organic matter in the sludge. The primary decomposition of excess sludge during the pyrolysis process occurred at the temperature ranging from 180 to 600°C with 30% mass loss. The third stage was mainly the decomposition of inorganic substances (especially carbonate) and residual organic substances. Gaseous products (NH₃, CO₂ and H₂O) produced from the decomposition of excess sludge were determined.

3.2. Kinetic model

Results for thermal alkaline treatment of excess sludge with varying time and temperature are shown in Fig. 3. The points in Fig. 3 are measured values, and the curve is the simulation result of first-order dissolution kinetics. After reaction at 30°C for 5 h, 15.98% of the protein in the sludge was dissolved. After reaction at 90°C for 5 h, 54.35% of the protein in the sludge was dissolved. The highest total protein recovery rate in the hydrolysate was 75% at 170°C after reacting 20 min, which has been found previously (Lamp et al., 2020). With the increase of reaction temperature and time, the protein content in sludge residue gradually decreased, indicating that the protein solubility of sludge increased. The bacteria in sludge were mainly Gram-negative bacteria, and the lipid content of the cell wall was high, which was easy to react with alkali solution, leading to the destruction of cell wall structure (Hui et al., 2022). An increase in the temperature facilitates the reaction.

As shown in Fig. 4, the protein concentration in the aqueous phase reached its maximum value, 1626.92 mg/L, after the reaction at 70°C for 4 h. At 90°C for 3 h, the protein concentration in the aqueous phase reached the maximum value, 1712.23 mg/L. Subsequently, with the extension of hydrolysis time, the protein concentration in the aqueous phase began to decrease, indicating that the degradation rate of the protein was higher than the dissolution rate of the sludge protein. At temperatures above 70°C, a darker colour of the hydrolysate was observed, possibly due to the Maillard reaction, accompanied by an unpleasant pungent smell and large amounts of ammonia escaping from the solution. Therefore, the optimal extraction conditions of residual sludge protein were as follows: reaction temperature of 70°C, hydrolysis time of 4 h.

Maillard reaction began with the condensation reaction of a non-dissociation amino group (lysine ε-NH₂, the α-NH₂ of protein molecules or amino compounds) and a reducing carbonyl compounds (mainly glucose, fructose, maltose, lactose and pentose) giving carbonyl amine compounds. Then the product formed Schiff base by taking off a molecule water, which can be further converted to aldose-amines or ketose-amines. Finally, these glycosylamines underwent molecular rearrangement to form amino-ketoses or amino-aldoses, respectively. In the intermediate stage, amino-ketoses or amino-aldoses reacted furtherly to form various carbonyl compounds (hydroxymethyl furaldehyde, maltol, iso-maltol and reduced ketones). In the final stage, polycarbonyl unsaturated derivatives (reduced ketone) generated volatile compounds by cracking reaction and produced brown melanoid substances by condensation and polymerization reaction to complete the whole Maillard reaction.

As shown in Fig. 5, the polypeptide concentration gradually increased with the extension of reaction temperature and time. Below 70°C, only a small number of peptides were produced in the first 1 h. At 30°C for 4 h, the polypeptide concentration was 509.42 ± 53.94 mg/L. At 90°C for 4 h, the polypeptide concentration was 2608.38 ± 286.47 mg/L, which increased by 4 times. This indicated that protein degradation was very difficult at low temperature. The higher reaction temperature and longer time, the more significant decomposition of the protein.

In previous study, the polypeptides and amino acids was also contained when determinating the total protein content, resulting in the recovered protein content much lower than the measured value in the solution. As shown in Fig. 6, the concentration of free amino acids was 85.11 ± 4.26 mg/L at 30°C for 4 h. At 70°C for 4 h, the free amino acids concentration was 242.17 ± 12.11 mg/L, which increased by 2 times. The results showed that the content of free amino acids supplemented with the increase of reaction temperature and hydrolysis time.

As shown in Fig. 7, the ammonium concentration in the hydrolysate increased rapidly during the first hour at the temperature ranging from 30 to 90°C. And then it stayed almost the same at a specific temperature. At 30°C for 4 h, the ammonium concentration was 126.07 ± 6.30 mg/L. At 90°C for 4 h, the ammonium concentration was 293.15 ± 14.66 mg/L, which increased by 1.3 times. This indicated that the degradation rate of protein, peptide and amino acid into ammonium was almost the same as the rate of ammonium and hydroxide reaction into ammonia after reaction 1 h, that is, the rate constant k₄ equals k₅.

As shown in Fig. 8, the number of molecular weight 1808 Da was the most, and the proportion of molecular weight higher than 10000 Da was 12.35%. This indicated that the macromolecular protein was hydrolyzed to low molecular weight polypeptides and amino acids during the alkaline hydrolysis. As shown in Fig. 9, the hydrolysate contained 17 types of amino acids, which consist of 7 types of hydrophobic amino acid such as alanine (Ala), valine (Val), leucine (Leu), isoleucine (Ile), proline (Pro), phenylalanine (Phe), and methionine (Met), and 10 types of hydrophilic amino acids such as aspartic acid (Asp), threonine (Thr), serine (Ser), glutamic acid (Glu), glycine (Gly), cysteine (Cys), tyrosine (Tyr), histidine (His), lysine (Lys), and arginine (Arg). However, the free amino acids accounted for 15.34% of the total hydrolyzed amino acids, indicating that the thermal alkalin treatment destroyed the structure of the protein, resulting in more conversion of macromolecular proteins into low molecular weight polypeptides, and only a tiny part of them were turned into amino acids.

The thermal alkalin treatment could cause an isomerization reaction of amino acid residues, and part of the L-type structure of natural amino acids was transformed into a D-type structure. This reaction was caused by the dehydrogenation of the chiral carbon atom connected to the carbonyl group to form a negative ion with a planar structure. Since the hydrogen ion has two different attack positions when the amino acid residues were reformed again, the final transformation product of L-type and D-type amino acids reached 100%. Because most D-type amino acids have no nutritional value and cannot be digested and utilized by the human body, the racemization of amino acids would decrease their nutritional value.

As shown in Fig. 10, the molecular weight of proteins in the hydrolysate ranged from 10 to 70 kDa, among which the most notable bands were 30 to 40 kDa. The protein obtained by trichloroacetic acid precipitation had powerful bands of 30 ~ 40 kDa and 10 ~ 15 kDa. Although the total protein content was high, the average protein content was low. Therefore, the whole protein bands were blurred.

As shown in Fig. 11, the functional groups of the raw sludge and sludge residue look similar based on the FTIR. The composition of sludge residue and raw sludge did not change significantly. The FTIR spectra of protein from trichloroacetic acid (TCA) precipitation were shown as an example for the comparison of the protein extracted from hydrochloric acid (HCl) and ethanol (EtOH). The FTIR spectra of protein extracted from TCA, HCl and EtOH were similar. A narrow band with a relatively sharp peak at 3377 cm^− 1 was a typical –NH group in the protein. A peak at 2936 cm^− 1 indicated the presence of aromatic amino acids (tryptophan, tyrosine, and phenylalanine) (Dian et al., 2002). The absorption peak at near 1661 cm^− 1 came from the stretching vibration of C = O and C-N and it was the amide I region spectrum peak of the protein (Hui et al., 2022). A band at 1200 − 950 cm^− 1 with two peaks, one peak at 1122 cm^− 1 was due to the presence of -PO₄³⁻, and the other one at 1057 cm^− 1 was because of carbohydrates (Boleij et al., 2019). The comparison declared that the spectral peak had a red shift. According to the circular dichroism results, as shown in Fig. 12, β-sheet accounted for 62.5% of the protein secondary structure, while others accounted for 37.5%. These results indicated that the protein secondary structure may be changed by thermal alkalin treatment.

As shown in Fig. 13, the molecular weight distribution of the protein ranged from 17991.70 to 138559.62 kDa. The number of protein molecules with molecular weight between 29556.96 and 36419.26 kDa was the largest. The most significant protein molecules were located near 35 kDa in the sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Both of them had almost no difference.

The purpose of sludge hydrolysis was to obtain more protein and avoid protein decomposition as much as possible. It could be seen from Table 2 that the higher the temperature was increased, the higher the rate constant. Although proteolysis was an endothermic reaction, increasing the reaction temperature of the system was beneficial to the breakdown of sludge cells and the release of intracellular protein. However, high temperature not only destroyed the sludge flocculation and the cell structure, resulting in the insoluble protein turned into a soluble protein in the liquid phase, but also made the dissolved protein continue hydrolyze to small molecular peptides, amino acids and ammonium. The higher the temperature was increased, the more intense the Maillard reaction, which not just reduced the protein recovery rate, also contributed to the recovery liquid colour darker.

Table 2

K-values of sludge protein extraction kinetics.
T(°C)	k₁	R₁²	k₂	R₂²	k₃	R₃²	k₄	R₄²
30	0.038	0.97	0.287	0.98	0.0004	0.99	0.283	0.99
50	0.075	0.97	0.341	0.99	0.0005	0.98	0.335	0.99
70	0.132	0.98	0.415	0.99	0.0009	0.97	0.401	0.98
90	0.173	0.98	0.517	0.99	0.0016	0.98	0.4919	0.98

The purpose of this paper was to obtain a high concentration and high yield protein through the thermal alkalin treatment by establishing a first-order kinetic model. The challenge was to maximize excess sludge protein solubility while minimizing degradation. The results showed that sludge protein of 46.33% could be dissolution, and the maximum concentration of protein in the aqueous phase was 1624.52 ± 81.23 mg/L in the optimum conditions of 0.1 M NaOH, reaction temperature 70°C, hydrolysis time 4 h.

The maximum deviation between the model and experimental data was 10%. Precise dynamics models need to develop over a smaller time range with minor time steps. To further develop the model, a more comprehensive raw materials should be used in the experiments under the identical reaction conditions. Such as sludge collected from different types of sewage treatment plants. The influence of inorganic substances should also be taken into account, because of inorganic components of biomass could catalyze and inhibit protein formation.

Ethical Approval, Consent to Participate and Consent to Publish

Not applicable.

Availability of data and materials

The data sets supporting the results of this article are included within the article.

Author Contributions Statement

Wenlong Hui: Data curation, Writing original draft preparation, Software.

Jiti Zhou: Conceptualization, Methodology.

Ruofei Jin: Visualization, Investigation, Supervision.

Funding information

The authors thank the wastewater treatment plant of Everbright in Lianyungang City for providing the granular sludge. No funding was received.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Academy, A.S., 2018. Determination of crude protein in feeds—Kjeldahl method. State Administration for Market Regulation, p. 8.
Akins, R.E., Tuan, R.S., 1992. Measurement of Protein in 20 Seconds Using a Microwave Bca Assay. Biotechniques 12, 496-&.
Boleij, M., Pabst, M., Neu, T.R., van Loosdrecht, M.C.M., Lin, Y.M., 2018. Identification of Glycoproteins Isolated from Extracellular Polymeric Substances of Full-Scale Anammox Granular Sludge. Environ Sci Technol 52, 13127-13135.
Boleij, M., Seviour, T., Wong, L.L., van Loosdrecht, M.C.M., Lin, Y., 2019. Solubilization and characterization of extracellular proteins from anammox granular sludge. Water Res 164.
Bou-Sarkis, A., Pagliaccia, B., Ric, A., Derlon, N., Paul, E., Bessiere, Y., Girbal-Neuhauser, E., 2022. Effects of alkaline solvents and heating temperatures on the solubilization and degradation of gel-forming Extracellular Polymeric Substances (EPS) extracted from aerobic granular sludge. Biochemical Engineering Journal 185, 108500.
Bradford, M.M., Williams, W.L., 1976. New, Rapid, Sensitive Method for Protein Determination. Fed Proc 35, 274-274.
Chacón-Parra, A., Lewis, D., van Eyk, P., 2021. Elemental nitrogen balance, reaction kinetics and the effect of ethanol on the hydrothermal liquefaction of soy protein. Chem Eng J 425, 130576.
Christensen, M.L., Keiding, K., Nielsen, P.H., Jorgensen, M.K., 2015. Dewatering in biological wastewater treatment: A review. Water Res 82, 14-24.
Dian, B.C., Longarte, A., Mercier, S., Evans, D.A., Wales, D.J., Zwier, T.S., 2002. The infrared and ultraviolet spectra of single conformations of methyl-capped dipeptides: N-acetyl tryptophan amide and N-acetyl tryptophan methyl amide. JOURNAL OF CHEMICAL PHYSICS 117, 10688-10702.
Dubois, M., Gilles, K.A., Hamilton, J.K., Rebers, P.A., Smith, F., 1956. Colorimetric Method for Determination of Sugars and Related Substances. Anal Chem 28, 350-356.
Eaton, A.D., Clesceri, L.S., Greenberg, A.E., Franson, M.A.H., 1966. Standard methods for the examination of water and wastewater. Am J Public Health Nations Health 56, 387-388.
Gao, J.L., Wang, Y.C., Yan, Y.X., Li, Z., Chen, M.L., 2020. Protein extraction from excess sludge by alkali-thermal hydrolysis. Environ Sci Pollut R 27, 8628-8637.
Geng, H., Xu, Y., Zheng, L.K., Liu, H.Y., Dai, X.H., 2022. Cation exchange resin pretreatment enhancing methane production from anaerobic digestion of waste activated sludge. Water Res 212.
He, Z.W., Zou, Z.S., Sun, Q., Jin, H.Y., Yao, X.Y., Yang, W.J., Tang, C.C., Zhou, A.J., Liu, W.Z., Ren, Y.X., Wang, A.J., 2022. Freezing-low temperature treatment facilitates short-chain fatty acids production from waste activated sludge with short-term fermentation. Bioresource Technol 347.
Hui, W., Zhou, J., Jin, R., 2022. Proteins recovery from waste activated sludge by thermal alkaline treatment. J Environ Chem Eng 10, 107311.
Hwang, J., Zhang, L., Seo, S., Lee, Y.W., Jahng, D., 2008. Protein recovery from excess sludge for its use as animal feed. Bioresource Technol 99, 8949-8954.
Lamp, A., Kaltschmitt, M., Ludtke, O., 2020. Protein recovery from bioethanol stillage by liquid hot water treatment. J Supercrit Fluid 155.
Li, X.M., Zhao, J.W., Wang, D.B., Yang, Q., Xu, Q.X., Deng, Y.C., Yang, W.Q., Zeng, G.M., 2016. An efficient and green pretreatment to stimulate short-chain fatty acids production from waste activated sludge anaerobic fermentation using free nitrous acid. Chemosphere 144, 160-167.
Liu, Y.S., Kong, S.F., Li, Y.Q., Zeng, H., 2009. Novel technology for sewage sludge utilization: Preparation of amino acids chelated trace elements (AACTE) fertilizer. J Hazard Mater 171, 1159-1167.
Lotti, T., Carretti, E., Berti, D., Martina, M.R., Lubello, C., Malpei, F., 2019. Extraction, recovery and characterization of structural extracellular polymeric substances from anammox granular sludge. J Environ Manage 236, 649-656.
Lowry, O.H., Hunter, T.H., 1945. The Determination of Serum Protein Concentration with a Gradient Tube. J Biol Chem 159, 465-474.
Park, M., Kim, N., Lee, S., Yeon, S., Seo, J.H., Park, D., 2019. A study of solubilization of sewage sludge by hydrothermal treatment. J Environ Manage 250.
Pervaiz, M., Sain, M., 2011. Protein Extraction from Secondary Sludge of Paper Mill Wastewater and Its Utilization as a Wood Adhesive. Bioresources 6, 961-970.
Prince, A.L., 1945. Determination of Total Nitrogen, Ammonia, Nitrates, and Nitrites in Soils. Soil Sci 59, 47-52.
Salette, J., Dumas, Y., 1970. Behavior Constant of Digitaria-Decumbens Stent - Relation Between Nitrogen Content and Dry Material - Influence of Different Medium Condition. Cr Acad Sci D Nat 271, 324-&.
Seviour, T., Derlon, N., Dueholm, M.S., Flemming, H.C., Girbal-Neuhauser, E., Horn, H., Kjelleberg, S., van Loosdrecht, M.C.M., Lotti, T., Malpei, M.F., Nerenberg, R., Neu, T.R., Paul, E., Yu, H.Q., Lin, Y.M., 2019. Extracellular polymeric substances of biofilms: Suffering from an identity crisis. Water Res 151, 1-7.
Sheng, G.P., Yu, H.Q., Li, X.Y., 2010. Extracellular polymeric substances (EPS) of microbial aggregates in biological wastewater treatment systems: A review. Biotechnol Adv 28, 882-894.
Smith, A.M., Agiza, A.H., 1951. The Determination of Amino-Acids Colorimetrically by the Ninhydrin Reaction. Analyst 76, 623-627.
Su, W., Tang, B., Fu, F.L., Huang, S.S., Zhao, S.Y., Bin, L.Y., Ding, J.W., Chen, C.Q., 2014. A new insight into resource recovery of excess sewage sludge: Feasibility of extracting mixed amino acids as an environment-friendly corrosion inhibitor for industrial pickling. J Hazard Mater 279, 38-45.
Suarez-Iglesias, O., Urrea, J.L., Oulego, P., Collado, S., Diaz, M., 2017. Valuable compounds from sewage sludge by thermal hydrolysis and wet oxidation. A review. Sci Total Environ 584, 921-934.
Wang, B.B., Liu, X.T., Chen, J.M., Peng, D.C., He, F., 2018. Composition and functional group characterization of extracellular polymeric substances (EPS) in activated sludge: the impacts of polymerization degree of proteinaceous substrates. Water Res 129, 133-142.
Wang, W.Y., Zhang, J., Li, X.S., Zang, Y., Li, R.D., 2019. Effects of typical modified passivators on speciation of heavy metals in protein extracted from sewage sludge. Environ Sci Pollut R 26, 10875-10886.
Wong, L.L., Natarajan, G., Boleij, M., Thi, S.S., Winnerdy, F.R., Mugunthan, S., Lu, Y., Lee, J.M., Lin, Y.M., van Loosdrecht, M., Law, Y., Kjelleberg, S., Seviour, T., 2020. Extracellular protein isolation from the matrix of anammox biofilm using ionic liquid extraction. Appl Microbiol Biot 104, 3643-3654.
Wu, R., Luo, J., Chen, S., Liu, J., Zhang, R., Li, Y.-Y., 2022. Mechanisms of calcium ion inhibiting alkaline fermentation of waste activated sludge for short-chain fatty acids production. Environmental Technology & Innovation 26, 102363.
Xiang, Y.L., Xiang, Y.K., Wang, L.P., 2017. Kinetics of activated sludge protein extraction by thermal alkaline treatment. J Environ Chem Eng 5, 5352-5357.
Xiao, K.K., Chen, Y., Jiang, X., Seow, W.Y., He, C., Yin, Y., Zhou, Y., 2017. Comparison of different treatment methods for protein solubilisation from waste activated sludge. Water Res 122, 492-502.
Xiao, K.K., Zhou, Y., 2020. Protein recovery from sludge: A review. J Clean Prod 249.
Xue, Y.G., Liu, H.J., Chen, S., Dichtl, N., Dai, X.H., Li, N., 2015. Effects of thermal hydrolysis on organic matter solubilization and anaerobic digestion of high solid sludge. Chem Eng J 264, 174-180.
Zhang, J., Zhang, J., Tian, Y., Li, N., Kong, L.C., Sun, L., Yu, M., Zuo, W., 2016. Changes of physicochemical properties of sewage sludge during ozonation treatment: Correlation to sludge dewaterability. Chem Eng J 301, 238-248.
Zou, X., He, J., Zhang, P., Pan, X., Zhong, Y., Zhang, J., Wu, X., Li, B., Tang, X., Xiao, X., Pang, H., 2022. Insights into carbon recovery from excess sludge through enzyme-catalyzing hydrolysis strategy: Environmental benefits and carbon-emission reduction. Bioresource Technol 351, 127006.

Download PDF

Version 1

posted

You are reading this latest preprint version

Kinetics of protein extraction from excess sludge by thermal alkaline treatment

Status:

Version 1

Abstract

Figures

1. Introduction

2. Materials And Methods

2.1. Materials and feedstock analysis

2.2. Thermal alkaline hydrolysis experiment

2.3. Analysis of products from thermal alkaline hydrolysis

2.4. Kinetic parameters

3. Results And Discussion

3.1. Characterization of thermal decomposition of excess sludge

3.2. Kinetic model

4. Conclusions

Declarations

References

Status:

Version 1