Anaerobic digestion of lignocellulosic waste usingphysico-chemical pretreatment methods interms of performance, microbial community, and cost analysis

Abstract

Increasing population and energy demand emphasize the global need to use energy resources more effectively and responsibly. Specifically, reliance on fossil fuels and the repercussions of climate change have hastened the hunt for renewable energy. However, current sustainable energy supplies are insufficient to fulfill rising demand alone. In this context, transforming organic solid waste into a sustainable resource and energy solves the environmental pollution problem while also creating new resource prospects. Although anaerobic treatment is a well-known and widely utilized method, it remains a field for further improvement. However, the lignocellulosic structure of organic solid wastes makes hydrolysis, the first stage in anaerobic digestion, more difficult than with other forms of waste. The use of lignocellulosic wastes in anaerobic digestion systems offers significant waste management potential. However, biodegradation of such wastes is challenging due to their complicated structure, hence several pre-treatment procedures have been devised. Physicochemical pretreatment procedures, particularly microwave and acid treatments, speed up the breakdown of lignin and cellulose structures, allowing microorganisms to operate more efficiently with the waste. As a result, the anaerobic process becomes more efficient, and important chemicals like methane and volatile fatty acids are produced in greater quantities. Sunflower waste is a major raw material source for anaerobic treatment procedures in Turkey and across the world. Turkey is one of the world's largest sunflower producers, with sunflowers growing across vast agricultural regions, particularly in the Thrace region. Stem, head, and other biomass wastes from sunflower cultivation are typically left on agricultural areas or disposed of using inefficient ways. However, these wastes have a significant potential in biogas production thanks to their high cellulose and lignin content. Integrating these wastes into the circular economy makes significant contributions to both environmental sustainability and economic gains. Agricultural wastes have great potential for biomass energy production and can be converted into various products such as biogas, bioethanol, compost. In addition, by utilizing agricultural waste, farmers' income sources diversify and contribute to the strengthening of the rural economy. Products obtained by recycling agricultural wastes both ensure efficient use of resources within the scope of the circular economy model and contribute to meeting Turkey's energy needs with sustainable resources. As a result of these reasons, sunflower was chosen as organic waste in the study. Even though pretreatment methods are basically divided into physical, chemical, biological and combined pretreatment, research has shown that combined pretreatment can be more successful. Combined pretreatments are preferred to ensure more effective breakdown of lignocellulosic wastes, because methods used alone often cannot adequately break down the complex structure of the waste. Combined pretreatments are generally applied by combining physical, chemical or biological methods, whereby the advantages of each method create a synergistic effect, providing more efficient results. For example, the combination of microwave and acid-based pretreatment disrupts the structure of biomass both thermally and chemically, resulting in higher biogas and volatile fatty acid production. While these methods increase energy efficiency, they also provide sustainable solutions in terms of cost effectiveness. In this study, physico-chemical pretreatment was chosen to break down the lignocellulosic structure most successfully. For a physicochemical process, microwave was used for physical pretreatment and hydrochloric acid was used for chemical pretreatment. In this way, a more effective hydrolysis process was achieved with combined pretreatment. The substrate was microwave pretreated in 0.8%, 1.2% and 1.6% HCl solutions for 30 minutes at 120oC and 140oC. After pre-treatment, sCOD values were examined to understand under which condition the efficiency was more successful. Compared to the control sample without pretreatment, it was observed that 47% higher sCOD was obtained in the sample that underwent combined pretreatment in 1.2% HCl solution at 120oC for 30 minutes. The sCOD value measured without pretreatment is 22395 mg/L sCOD, and with 1.2% HCl 120oC pretreatment, the sCOD value is 32908 mg/L. In this way, the most effective pretreatment method was obtained in the study. At the same time, TS and VS values for seed sludge, control sample and the most efficient pretreatment were determined by standard methods. When comparing control samples and pretreated samples, TS degradation increased by 45% and VS by 52%. An increase in the TS and VS values of the solid material is a sign of effective deterioration. Thus, lignocellulosic wastes can be shown to decompose more efficiently, meaning that a more efficient hydrolysis will result in a larger VFA output. In recent years, it has become quite common to suppress the presence of archaea in the anaerobic digestion process and to ensure the formation of volatile fatty acids. The main reason for this is that although methane is used as an energy source, it is an important source of greenhouse gases. At the same time, CO2 is also formed as a byproduct in addition to methane (CH4) in semi-anaerobic digestion. The most important motivation of this study is to obtain volatile fatty acids. In addition to the development of pre-treatment technologies, the production of volatile fatty acids, which have an important place in the circular economy and are used in many different sectors, has become very popular. In this study, sunflower used as lignocellulosic waste was kept at pH 5.5 after being exposed to combined pretreatment. While archaea, that is, microorganisms that provide methane formation, cannot survive in this pH range, only the formation of volatile fatty acids is possible with acidogenesis microorganisms. At this pH value, microorganisms produce volatile fatty acids, but these volatile fatty acids do not turn into methane because the archaea cannot survive. Once the archaea were inhibited and the pH was checked every day, samples were taken on certain days. The formation amount of volatile fatty acid types, which are used in various sectors and have different usage purposes, on determined days was measured in mg/L sCOD. Cumulative VFA formation was determined for acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid and isovaleric acid. The 1st day sCOD value for the pretreated sample was 32908 mg/L; At the end of the 30th day, it was observed to be 19324 mg/L. It is showed that a success rate of volatile fatty acid formation of approximately 40%. In summary, the acid types, from most produced to least produced, are as follows: Propionic Acid: 3089 mg/L, Isovaleric Acid: 2993 mg/L, Isobutyric Acid: 2700 mg/L, Butyric Acid: 2137 mg/L, Isocaproic Acid: 2421 mg/L, Caproic Acid: 760 mg/L, Acetic Acid: 539 mg/L, Valeric Acid: 245 mg/L. Another important aspect of the study is the generation of genomic sequences of microbial communities for taxonomy categorization. The aim is to fully characterize microbial species using Oxford Nanopore MinION technology in combination with fast and long-read approaches. After DNA isolation, PCR and sequencing were performed under appropriate conditions. 16S, 18S and archaeal microorganism communities were classified and compared as phylum, class and species. Genetic sequencing of microbial communities has revealed a diverse spectrum of microorganisms involved in the anaerobic digestion process, including many species of bacteria and archaea. The most dominant species in microbial communities were observed as follows: Armatimonadota, Caloramator sp. E03, Dysgonomonadaceae:, Stephanoeca, Prototheca ciferrii, Prototheca wickerhamii, Tetramitus dokdoensis, Methanosarcina vacuolata, Methanothrix soehngenii, Methanosarcina barkeri. These findings will provide resources for future studies on understanding the microbial community of the anaerobic digestion process and improving system efficiency. Finally, a cost analysis was performed using all collected data. The economic values according to the types of volatile fatty acids obtained from 1.5 grams of substrate are as follows: Acetic Acid: $17, Propionic Acid: $250, Isobutyric Acid: $476, Butyric Acid: $247, Isovaleric Acid: $673, Valeric Acid: $24, Isocaproic Acid: $385, Caproic Acid: $122. Microbial analysis is very important to check system success efficiency. Archaea were shown to persist in this manner. Anaerobic digestion is an extremely sensitive process. Although successful pretreatment was used and archaea were predicted to be absent, their continued presence indicated that a more precise process application could yield more efficient VFA synthesis. At the end of this entire study, the following conclusions can be made: Increasing population and energy demand increases the need to replace traditional energy sources with sustainable alternatives. In this context, the use of organic solid wastes, especially those with a lignocellulosic structure, in the production of biogas and volatile fatty acids is important. However, the complexity of the lignocellulosic structure makes biodegradation difficult. Therefore, physicochemical pretreatment methods such as microwave and HCl enable waste to be hydrolyzed more easily and increase biogas yield. Studies show that combined pretreatment methods disrupt the structural integrity of wastes, leading to higher methane and volatile fatty acid production. In countries where sunflower production is intense, such as Türkiye, these wastes offer a great energy potential. While volatile fatty acid production in anaerobic treatment stands out as a more environmentally friendly and economical option, effective taxonomic analysis of microbial communities has the potential to further increase process efficiency.