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Low Temperature Thermo - chemical Pretreatment of Dairy Waste Activated Sludge for Anaerobic Digestion Process
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Abstract

An investigation into the influence of low temperature thermo-chemical pretreatment on sludge reduction in a semi-continuous anaerobic reactor was performed. Firstly, effect of sludge pretreatment was evaluated by COD solubilization, suspended solids reduction and biogas production. At optimized condition (60C with pH 12), COD solubilization, suspended solids reduction and biogas production was 23%, 22% and 51% higher than the control, respectively. Secondly, semi - continuous process performance was studied in a lab-scale semi-continuous anaerobic reactor (5L), with 4L working volume. With three operated SRTs, the SRT of 15 days was found to be most appropriate for economic operation of the reactor. Combining pretreatment with anaerobic digestion led to 80.5%, 117% and 90.4% of TS, SS and VS reduction respectively, with an improvement of 103% in biogas production. Thus, low temperature thermochemical can play an important role in reducing sludge production.

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*Corresponding author:

Dr. J. Rajesh Banu, Department of Civil Engineering, Anna

University of Technology Tirunelveli, Tirunelveli - 627007, India. Tel: 9444215544, Email id:rajeshces@gmail.com.

Keywords: Waste Activated Sludge, Solubilization, Thermo-chemical pretreatment, Biogas, Biodegradability, Anaerobic Digestion.

1. Introduction
Dairy industry is one of the prime sectors in India. Studies say that Indian dairy industries have a growth at more than 15% and are estimated to cross 150 million tons per annum. Water management in the dairy industry is well documented, but effluent production and disposal remain a problematic issue for the dairy industry. Proper management of excess sludge is a big challenge to wastewater treatment operators because sludge handling and disposal accounts for up to 60% of total treatment plant operating costs (Neyens and Baeyens, 2003). Anaerobic digestion is of particular interest in sludge treatment since it can reduce the overall amount of sludge to be disposed, while producing an energy-rich biogas that can be valorized energetically (Appels et al., 2010). Anaerobic digestion thus optimizes wastewater treatment costs, its environmental footprint and is considered a major and essential part of a modern wastewater treatment plant. Anaerobic degradation can be achieved through several stages: hydrolysis,

acidogenesis, acetogenesis and methanogenesis. Anaerobic digestion of sludge is hampered due to the rigid structure of the microbial cell walls, protecting the inner cell products. Hence, hydrolysis of sludge requires longer retention time and has been pointed as the rate limiting step. In order to improve the rate of hydrolysis and the anaerobic digestion performance, sludge disintegration was developed as a pre-treatment process to accelerate the anaerobic digestion and to increase the degree of stabilization. Increasing the degree of sludge stabilization using a disintegration process provides less sludge production, more stable sludge and more biogas production compared with classical anaerobic digestion. Various methods like Ultrasonic treatment (Pham et al., 2009), Ozone oxidation (Ahn et al., 2002), alkaline treatment (Lin et al.,

2007), thermal treatment (Barjenbruch and Kopplow, 2003), Fenton process (Kaynak and Filibeli, 2008) and biological hydrolysis with enzymes (Ayol et al., 2008) were investigated for sludge disintegration by several researchers in half-scale and lab-scale plants to bypass the rate limiting stage of hydrolysis. Mechanical treatment employs several strategies for physically disintegrating the cells and partly solubilizing their content. Although less results are available for the other pretreatment methods, it is seen that their efficiency of improving anaerobic digestion of sludge is rather low, compared to the other methods. Ultrasonic pretreatment is the alternate method to disrupt sludge cells without doubt. Although cell disintegrations of 100% can be obtained at high-power levels, power consumption thus becomes a serious problem, and the ultrasound probes need replacement every 1.5 to 2 yrs, which causes great concerns on its practical application (Zhang et al., 2007). However its application as a sludge pretreatment process in anaerobic digestion is scarcely mentioned. The heat treatment of waste activated sludge was shown to be an effective pretreatment method for anaerobic digestion. The optimum treatment conditions and digestion improvement are largely depending on the nature of the sludge. Various temperatures, ranging from 60 to 270C have been studied in literature. However, temperatures higher than 180C lead to the production of recalcitrant soluble organics or toxic/inhibitory intermediates, hence reducing the biodegradability (Wilson and Novak, 2009). The only alternative to overcome this drawback is the application of low temperature treatment, and it has been pointed out as an effective treatment for increasing biogas production (Climent et al., 2007). In thermo-chemical methods, an acid or base is added to solubilize the sludge. The addition of acid or base avoids the necessity of high temperature and these methods are carried out at ambient or moderate temperatures. These methods are shown to be effective and cumbersome for sludge solubilization. The

objective of the current work is to study the solubilization of organic and inorganic compounds during thermo-chemical treatment and the effects on anaerobic biodegradability. Finally, this thermo-chemical pretreatment was combined with semi continuous anaerobic digesters in order to evaluate their effects in terms of solids reduction and of biogas production.

2. Materials and methods

2.1 Sludge sampling and characterization The Waste activated sludge was obtained from a dairy plant at Tirunelveli in Tamil Nadu (India). Samples were collected and stored at 4C. The Characteristics of the raw sludge were as follows: pH was 6.96, CODS (SCOD) was 420 mg/L, Total COD (TCOD) was 22000 mg/L, Total solids (TS) content was 8513 mg/L, Volatile solids (VS) content was 5160 mg/L, Suspended solids (SS) content was 4700 mg/L, Proteins was 780 mg/L and Carbohydrates were 320 mg/L. 2.2 Thermo-chemical Pretreatment In this study, the effect of low temperature thermo-chemical pretreatment was investigated. The low temperature thermo-chemical pretreatment was carried out at 50, 60, 70 and 80C in order to enhance solubilization of particulate material, as well as enzymatic hydrolysis. The effect of thermo-chemical treatment depends on both temperature and time. In this work, the effect of pretreatment time was evaluated by taking samples at different pretreatment times (6, 9, 12, 24, 36 and 48 h) in order to study the combined effect. Batch reactors containing 1L of sludge were submerged in a thermostatic bath at various temperatures (50, 60, 70 and 80C) during 6, 9, 12, 24, 36 and 48 h. The reactors were covered with an aluminium foil, to avoid water evaporation. The sludge in the reactor was kept in suspension by a slow-speed stirrer (Digital Overhead IKA RW 20), to ensure temperature

homogeneity. For the waste activated sludge studied, different concentrations of NaOH were added to reach different pH values of 10, 11 and 12. Even though, sodium decrease the dewaterability of the sludge, the choice of this alkaline agent was made from different studies indicated that, for anaerobic digestion, pretreatment with NaOH was more efficient than using other alkaline agents (Kim et al., 2003, Lin et al., 1997). The negative influence of sodium over sludge dewaterability is reduced when NaOH is combined with other treatment methods such as microwave and thermal (Jin et al., 2009; Ilgin and Dilek., 2009). Dewaterability of the sodium pretreated sludge can be improved by the subsequent sludge management using lime. Banu et al., (in press) have used lime to decrease the capillary suction time of sodium pretreated sludge. 2.3 Biochemical methane potential assay Biogas production of raw and pretreated sludge samples at various temperatures (50 to 80C) for 6, 9, 12, 24 h was initially determined by batch tests at mesophilic conditions. After pretreatment applications and before the reactor setup, pH of all pretreated samples was neutralized to 7. The biodegradability assays were conducted in batch reactors with 300 mL serum bottles. The rumen micro-organisms of cattle dung were used as an inoculum. Moreover in case of anaerobic biodegradability, the use of a highly active animal inoculum waste will reduce the experimental time significantly, or reduced the amount of inoculum required in full-scale batch digesters, and consequently, the corresponding digester volume (Borja et al., 2003). Each serum bottle was filled with 100 g of inoculum and 50 g of substrate. A blank treatment with only 150 g of inoculum was used to determine biogas production due to endogenous respiration. After adding the substrates and inoculum, the reactors were closed with a rubber septum and an aluminium seal to make them air tight and was subsequently purged with nitrogen gas at the rate of 10 mL per second for 20 minutes into the reactors to maintain anaerobic conditions. Bottles were

maintained at 35C under shaking (220 rpm). Batch reactors were operated with a residence time of 50 days. Samples were analyzed in duplicate. Enhancement of biodegradability was evaluated by comparison of biogas volumes produced by treated and untreated samples.The anaerobic biodegradability tests performance was evaluated by fitting the cumulative biogas production data to the modified Gompertz equation. The Gompertz equation describes cumulative biogas production assuming that, biogas production is a function of bacterial growth (Redzwan and Banks, 2004). The modified Gompertz equation is presented below: B = P {1-exp [-Rm (t-) / P]} Where B is the cumulative biogas production (mL), Rm is the maximum biogas production rate, P is the biogas yield potential (mL), and is the duration of lag phase, days. Using Matlab 7.0, the unknown parameters were calculated in order to measure the difference between the experimental measurement and the corresponding stimulated value. 2.4 Anaerobic digestion reactors Two identical laboratory scale semi continuous reactors with a working volume of 4L were used as anaerobic digesters at mesophilic temperature (35C). Among the two, one is designated as Control Semi Continuous Anaerobic Reactor (CSCAR) which acts as control and another is designated as Experimental Semi Continuous Anaerobic Reactor (ESCAR), where sludge reduction was carried out. Sludge retention times (SRT) of 20, 15 and 12 days were sequentially tried to investigate the performance of the anaerobic digestion of the pretreated sludge. A control reactor fed with untreated sludge was operated at same SRTs. Feeding and withdrawals were carried out each day by peristaltic pumps in a semi continuous mode. Biogas production was measured by water moving in graduated test tubes linked to the reactors.

2.5 Analytical parameters The following parameters were analyzed before and after thermo-chemical treatment: Total Solids (TS), Suspended Solids (SS), Chemical Oxygen Demand (COD), Carbohydrate concentration, Protein concentration and pH as per Standard Methods for the Examination of Water and Wastewater (American Public Health Association, 2005). The analyses were performed on both the sludge and the supernatant to identify the total and soluble fractions of the specific component. TS, VS and SS were measured in the whole sludge (Total TS, VS and SS) and in the supernatant after centrifugation at 30,130 x g, for 15 min and subsequent filtration through 0.45 m microfiber filter paper. VFA was analyzed by distillation-titration method, and the result was expressed in acetic acid. The methane content in the biogas was analyzed using a Baroda gas chromatograph equipped with a thermal conductivity detector and porapack Q column with hydrogen as carrier gas at a flow rate of 40 mL/min. Protein concentration was determined on total sludge and on the supernatant using the Lowry method (Takahashi et al., 2009). After reactions with salts and Folin's phenol reagent, absorbance of samples was determined at 620 nm, using a spectrophotometer. Folin's phenol reagent is a mixture of phosphomolybdate and phosphotungstate used for the

colorimetric assay of phenolic and polyphenolic antioxidants. It works by measuring the amount of the substance being tested needed to inhibit the oxidation of the reagent. Carbohydrate concentration was determined on total sludge and on the supernatant using the anthrone method (Tapia et al., 2009). After reactions with anthrone and sulphuric acid, absorbance of samples was determined at 625 nm, using a spectrophotometer. The increase in SCOD was calculated as given below: (SCOD after pretreatment SCOD before pretreatment) SCOD (%) = X 100

SCOD before pretreatment

3. Results and discussion 3.1 Thermo-chemical pretreatment In this study, thermo-chemical pretreatment of waste activated sludge was performed to improve the treatment efficiency. The control was performed using non-pretreated WAS. The treatment was performed for different temperature and treatment time. The expected effect of the thermochemical treatment of sludge was an increase in soluble materials, with interest focused on SCOD solubilization, suspended solids reduction and biogas production, thus enhancing hydrolysis. 3.1.1. COD solubilization and SCOD release The pretreatment was done to improve the bioavailability of sludge particulate material. SCOD calculations were considered the main parameter for evaluation of sludge particulate material, and it enables an evaluation of the maximum level of sludge solubilization. Increased SCOD is determined as the substance that can be readily used to produce methane during anaerobic digestion (Wang et al., 2005). The SCOD of pretreated sludge increased with increasing temperature. Fig.1a. shows the optimization of time and pH for COD solubilization during low temperature thermo-chemical pretreatment. From the figure, it is evident that, as the treatment time was increased from 6 to 24 h, an increase in COD solubilization was observed. This may be due to the disruption of chemical bonds in cell walls and membranes by thermo-chemical treatment. Therefore intracellular organic material is released to the liquid phase and increases the SCOD (Appels et al., 2010, Banu et al., 2009). As reaction time increases from 24 to 48 h SCOD was found to be decreased. The sludge is subjected to low temperature thermo-chemical

treatment for a significantly prolonged time (24 h). Thermal treatments are prone to enhance the formation of refractory substances. The fall in SCOD after 24 h might be due to the formation of refractory substances. Thus, for the waste activated sludge sample, a treatment time of 24 h was found to be the optimum. Similarly, a pH 10 seems to be too low for an effective COD solubilization, since even after a treatment time for 24 h only a very limited amount was set free. Thus, in order to achieve better solubilization, pH 12 was considered to be an optimum condition. Likewise, at 50C for pH 12, the COD solubilization was found only 17%. However, for 60, 70 and 80C irrespective of pH, the temperature plays a major role in enhancing COD solubilization, and it was found to be 23, 24 and 25% respectively. From the above, it is clear that temperature above 60C doesnt help in solubilizing the excess sludge. Thus, comparing cost economics and considering energy generation, low temperature thermo-chemical treatment (60C) was considered to be an optimum condition for rupturing cell membranes. The study is carried out at low temperature range of 50 - 80C which are less intense when compared to high temperatures. Thus, the solubilization doesnt show a rapid rise, but the overall solubilization for the treatment is satisfactory. 3.1.2. SS Reduction SS reduction is an indication of sludge stability, and it is used for assessing the effectiveness of a process in stabilizing sludge (Gholamreza et al., 2008). Fig.1b. shows the optimization of time and pH for SS reduction during low temperature thermo-chemical pretreatment. From the figure, it is evident that, as the treatment time was increased from 6 to 24 h, an increase in SS reduction was observed. The main reason for mass reduction of sludge during the thermo-chemical pretreatment might be to rupture the cell wall and to release of extracellular and intracellular matter. Thus, it is evident that at 60C with pH 12, COD solubilization and SS reduction was

found to be 23 and 22% respectively, which might be regarded as a threshold for the predigestion step. In contrast, pH was found to be decreased during thermo-chemical pretreatment, and it may be explained by acidic compounds formation due to floc disintegration. Lipids were hydrolyzed to volatile fatty acids, and these compounds led to be decreased pH (Bougrier et al., 2005). 3.1.3. Soluble Carbohydrate and Protein release Cell lysis releases protein content into the medium is the first stage of floc disintegration. Proteins are the principal constituents of organisms, and they contain carbon, which is a common organic substance as well as hydrogen, oxygen and nitrogen (Nah et al., 2000). For this reason, it was considered that as the level of soluble protein increased, the efficiency of anaerobic digestion would be improved. Due to thermo-chemical treatment, solids were solubilized, especially organic solids. Fig.2. Presents the solubilization rate calculated for proteins and carbohydrates for the samples. As given in figure, a higher pH value of 12 led to more carbohydrate and protein releases than pH 10. Increase in the carbohydrate and protein releases with the increase in pH was stated by Chen et al (2007). Furthermore, it was found that increase in temperature doesnt play a major role, i.e., the solubilization rate increases linearly with temperature only till 60C, after which the rise is nearly flat. Thus, 60C with pH 12 is significant for protein and carbohydrates release. According to Liu and Fang (2002), during the sludge treatment with NaOH, protein is released more compared to carbohydrate and this result is similar to that obtained in the present study. The protein releases presented in the figure are the sums of protein released from EPS as well as the cell lysis. Hence protein concentration is more than carbohydrate. 3.1.4. Biochemical methane potential assay

Biodegradability batch assay was carried out to assess the feasibility of using thermo-chemical pre-treatment to improve the anaerobic biological degradation of dairy sludge. Biodegradability assays, in which cumulative biogas production was monitored, were assessed to both raw and pre-treated substrates under mesophilic conditions. Cumulative biogas production in serum bottles was maintained for 50 days, and the biogas production was monitored every day. Daily biogas was measured by inserting the needle attached to a syringe (10 and 25 mL). The gas pressure in the bottle displaces the syringe plunger, and the displaced volume was recorded. The results showed that thermo-chemical pretreatment of dairy sludge improved anaerobic degradation. Comparative analysis of biogas generation is depicted in Fig.3a. It was observed that, initial biogas production up to day 5 was similar in all cases, and this may be due to the acclimatization. The increase in biogas production was observed for 60, 70 and 80C pretreated sludge and there is no significant difference between biogas productions. The accumulated biogas production at the end of 50 days of the digestion period, was nearly 415 mL for raw sample, and at 60C with pH 12 it was around 845 mL, nearly 51% higher biogas production was obtained than the raw sludge. These results suggest that a low temperature thermo- chemical pretreatment enhances the biogas production. After 50 days of the digestion period, the biogas production ceased indicating that the digestate did not undergo any further degradation. Fig.3b. shows the model fit with the experimental data (solid line), from each assay (circles) for the samples. Table 1 presents the kinetic parameters obtained in the optimization process. From the experiment conducted, it can be noted that biogas production is enhanced by an increase in temperature from 50C (535 mg/L) to 60C (845 mg/L) by 57%. When further increase up to 80C (890 mg/L) the increase in biogas production is only 5.3%. The trend of biogas production (Rm and P values from Table 1) shows that the solubilization of sludge at 60C with pH 12

attributes to the significant biogas production. Increasing the temperature has not influenced the process to a larger extent other than power consumption. At 60C with pH 12 the lag time for biogas production is decreased to 0.99 from 1.4 at 50C. The values were calculated for all experimental conditions and are found to be fit (which are not shown). 3.2 Semi continuous anaerobic digestion reactors During this part of the study, the dairy sludge was thermo-chemically pretreated at 60C with pH 12 was used as a feed. In a semi continuous anaerobic reactors, steady state is assessed by the stability of daily gas production, TS, SS and VS concentrations digested sludge from each reactor. With this assessment after 50 days, the fluctuations in these parameters were less than 10% and it was believed that steady state was achieved. Results obtained after reactor stabilization is reported in Table 2. 3.2.1 SRT Variation During the total period of 200 days, the performance evaluation of Semi continuous anaerobic reactors was carried out at three different SRTs such as 20, 15 and 12 days. The particular SRT was maintained constant by feeding the sludge at a constant rate by the peristaltic pump. The feeding was initialized with low organic loading rate of 3.55 g VS/Ldfor maintaining a long SRT of 20 days. The SRT was shifted to the next lower value and so on, once the reduction efficiencies of TS, SS, and VS were found to be consistent with a particular SRT. At individual SRTs, steady state of operations was retrieved and the results mentioned were average of five consecutive consistent readings. It was observed that the overall performance efficiency in terms of reduction in TS, SS and VS was lowering down as the SRT was lowered. 3.2.2 pH

The digester performance was influenced by pH. The value and stability of the pH in an anaerobic reactor are extremely important because methanogenesis only proceeds at a high rate, when the pH is maintained in the neutral range. pH values in these experimental remained in the range from 7.9 to 7.1 during anaerobic digestion. It was observed that during every shift to the next lower SRT, the pH dropped rapidly. The dropping down of pH at shorter SRT is due to the high organic loading rate, which results in high VFA concentration during the acidogenesis phase. The results reveal that, digester could maintain their pH themselves though higher organic loading was supplied. 3.2.3 Alkalinity pH cannot be an effective measure of the stability of an anaerobic process when there is a high buffering capacity (Bprnsson et al., 2000). Under this condition, alkalinity levels reveal a potential anaerobic process performance directly. The alkalinity of a steady system is between 1000 and 5000 mg CaCO3 L-1 (Ren and Wang, 2004). Lower values of effluent alkalinity warn about the impending reactor failure. The variation of pH and alkalinity with digestion time is presented in Fig.4a. The alkalinity of feed sludge and digested sludge varied significantly as pretreatments were used. The ESCAR digester had higher alkalinity compared to CSCAR due to the addition of NaOH. At 20 and 15 days SRTs, the effluent alkalinity levels were 19% and 34% more than the influent alkalinity. Manariotis and Grigoropoulos (2002) observed the increased levels between 19 and 21%, while Bodkhe (2009) observed the range of 25 to 33%. Similar to pH, the alkalinity reduced at shorter SRT is since a more organic matter was supplied, it eventually increased with acid generation and subsequently neutralizes the alkalinity capacity. 3.2.4 Volatile Fatty Acids

Volatile fatty acids are considered as a central parameter that governs the healthiness of the anaerobic treatment (Pind et al., 2002). These are the most important intermediates, where they are degraded by proton-reducing acetogens in association with hydrogen consuming methanogenic bacteria (Mechichi and Sayadi., 2005). Thus, VFA determination in conjunction with pH measurement is an essential prerequisite for maintainance of desired environmental conditions in the anaerobic reactor. Fig.4b. presented the VFA Concentration of anaerobic digester which followed the trend of increasing, firstly, and then decreasing. The VFA concentration varied in a range of 227-790 mg/L and fell well within the recommended value for healthy anaerobic digestion. As a result of increased concentrations of VFA, the acids accumulate and the pH decreases to such a low value that the hydrolysis and acetogenesis can be inhibited (Siegert and Banks, 2005). The average VFA concentration of ESCAR (363 mg/L) was found to be lower than CSCAR (590 mg/L) indicating greater utilization of VFA by methanogens and subsequent biogas production in ESCAR. According to Dearman and Bentham (2006), when VFA concentration started to decrease, biogas production rate increase. 3.2.5 TS removal Fig.5a. shows the variation of TS in digested sludge. For TS removal, CSCAR was in the range of 6.1 to 9.6% and digester ESCAR 13 to 25.1%. TS analysis shows that there was an increase of 161% in TS reduction when compared with CSCAR, since TS reduction was 25.1% in ESCAR at 15 days SRT. The results revealed that thermo-chemical is an effective technique for breaking down the microbial cells or difficult hydrolyzed compounds to easy biodegradable compounds. 3.2.6 SS and VS removal

The SS and VS reductions can be regarded as the reductions that are expected to be achieved in total mass of sludge. The variation of SS and VS removal is graphically presented in Fig.5b. For SS removal, CSCAR was in the range of 17 to 19.5% and digester ESCAR was in the range of 33 to 39.2%. The highest SS removal achieving from ESCAR digester at 15 days SRT was 39.2% with an increase of 101% over CSCAR. Thus, a pretreatment with thermo-chemical gave a great advantage in SS removal improvement compared to non-pretreatment. For VS removal, CSCAR was in the range of 14 to 17% and digester ESCAR 25 to 33%. The VS removal of ESCAR at 20 and 15 days SRT was almost constant and slightly reduced at 12 days SRT. However, it reduced for CSCAR from long to short SRT. This higher performance is due to the combination effects of thermal and chemical, which help to break down the microbial cells for faster subsequent degradation. It eventually facilitates the decomposition reaction which leads to biodegrade more compounds in the digester. 3.2.7 Biogas production Fig.6. shows the comparison of biogas production of the digesters at different SRTs. The first SRT (20 days) was started at digestion run time of 55 days until 115 days, then second SRT (15 days) until 165 days, and finally, the third SRT (12 days) until 200 days. By comparing to control, the methane production of thermo-chemically pretreated sludge increased by 90, 109 and 111% at 20, 15 and 12 days SRT, respectively. It clearly showed that the methane production rate appreciably increased from 20 to 15 days SRT, while minor improvement was observed from 15 to 12 days SRT. The methane content in the biogas varied from 65 to 70%. The higher gas production in thermo-chemically pretreated sludge evidently indicated that it hydrolyzed more organic material solution, which is immediate used by anaerobic bacteria and eventually

facilitates the digestion processes. Furthermore, the biogas production rate was high at short SRT and low at long SRT. Once the SRT was reduced, the organic loading rate automatically increases, which eventually come to over loading phenomenon. It was clearly shown by dropping pH at 12 days SRT. Therefore, by considering both biogas and methane production, SRT of 15 days was found to be the suitable retention time for effective sludge degradation. 3.2.8 Selection of pertinent SRT The minimum SRT at which the reactor yields the designed treatment efficiency highly influences the reactor volume, its compactness and the economy of operation. The economic viability of sludge management is not feasible when the reactor volume is larger. Therefore, the selection of the most suitable minimum possible SRT is very important to ensure the costeffectiveness of the reactor operation. The results obtained at various SRTs identified, the SRT of 15 days to be the most appropriate SRT for economic operation of the reactor.

4. Conclusions
The influence of low temperature thermo-chemical pretreatment of dairy sludge was studied. At 60C with pH 12, COD solubilization and suspended Solids reduction was 23% and 22% higher than that of control. BMP assay results of pretreated sludge confirmed that the observed solubilization led to an increase in sludge biodegradability, nearly 51% higher biogas production than control. Thus, 60C with pH 12 was chosen for semi-continuous digesters and by combining pretreatment with anaerobic digestion led to 80.5%, 117% and 90.4% of TS, SS and VS reduction respectively, with an improvement of 103% in biogas production.

Acknowledgements

Authors are thankful to Department of Biotechnology, India for partial financial assistant to this project (BT/PR13124/GBD/27/192/2009) under their scheme Rapid Grant for Young Investigator (RGYI).

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