Enhancement of Microbial Biosolubilization and Biogenic Methane Potential of Low to High Grade Coal of Pakistan

Abstract

The process of bio solubilization depends on coal ranking and its oxidative state. Lignite coal being less complex in structure is easier to solubilize microbially by the production of extracellular enzymes, but bituminous coal has been less used for bioconversions due to its signature features, that are considered appropriate for thermal power generation resulting negative environmental impacts. Green energy procedures can encourage the efficacy of bituminous coal for multiple applications and sustainable environment. This research was aimed to inquire and reveal the enhanced bio solubilization extent of lignite and bituminous coal designated as low and high rank aerobically and anaerobically that contributes to extract and purify eco-friendly and bio active compounds. In this concern, the bituminous coal sample from Baluchistan coalfield, Duki region of Pakistan and lignite coal samples from thar Sindh region of Pakistan, were subjected for chemical characterization to explore its vulnerability for biological transformation. The native coal solubilizing fungal isolates NF-1 and GB, identified as Debaromyeces hansenii and Aspergillus ochraceus. Respectively based on fungal ITS sequences were isolated. Major conditions were optimized for Debaromyeces hansenii NF-1 and Aspergillus ochraceus GB, 1% and 1.5% glucose with 0.5% of pulverized coal was investigated as an ideal ratio for maximum organic liberation within 7 days respectively for bituminous coal while for lignite coal, 2% glucose was utilized by Debaromyeces hansenii NF-1 and 1% glucose was utilized by Aspergillus ochraceus GB with 1% coal loading ratio and incubation time of day 11. Along bio-treatment, enhancing agents were also used to modify medium (sodium acetate) as well as coal structure (NaOH) that intensify liberation of humic acid, assessed through UV Visible spectrophotometer. Conjugative effect of fungal isolates (NF-1 and GB) on pulverized pre oxidize coal residues was found effective in liberating aromatics. Qualitatively, ethyl acetate soluble extracts of bio-liquified black liquids were investigated through GC-MS. MS spectra showed organic pool, in that pool bio-treated pre oxidized solubilized liquids represented more diversity of organics than virgin. Gravimetrically, it was analysed that Debaromyeces hansenii NF-1 showed 52.4% and Aspergillus ochraceus GB 71% weight loss of pre oxidized coal residues within 2 days by increasing process efficiency. Bio-treatment showed significant differences in elemental composition (O/C, H/C and C/N) of solubilized pre oxidized coal residues. FT-IR spectrum indicated that NF-1 and GB fungal isolates intensify the peaks of phenolic, aldehydic, aromatic, carboxylic, aliphatic and humic acid functionalities in pre oxidized residual coal. For quantitative estimation, gravimetric ratio of extracted humic acid from pre oxidized coal residues by using Aspergillus ochraceus was 78% and Debaromyeces hansenii showed 60% xi while 52% and 42% was extracted from its bio-extracts respectively. Spectroscopic E4/E6 ratio, elemental and FTIR spectrum of biologically (NF-1 and GB) extracted humic acid from pre oxidized coal residues specify formation of condensed aromatics and low molecular weight, enhanced nitrogen/oxygen content and vibrational changes in phenolic, amines, carboxylic, aldehydic and aliphatic compounds of humic molecules respectively. The illumine miseq technique of next generation sequencing had shown similarity in bacterial diversity in 6 samples out of 8 while, 2 samples had shown different behaviour from other samples after determining their relative and read abundance, alpha and beta diversity and heat map. While the fungal alpha diversity of 4 samples had shown distinction from each other in their relative abundance. Another sample from Baluchistan Duki coal mine had shown the presence of more Proteobacteria and Actinobacteria than Firmicutes. After solubilization of coal by fungal isolates different compounds that were produced after degradation of coal were extracted. These compounds were name as Humic and Fulvic acids. About 4.5% concentration of KOH had shown maximum yield of humic acid in both types of coal. The percentage of humic acid using NaOH for native lignite and Bituminous coal was 21.15% and 11.6% and for HNO3 treated coal percentage for lignite and Bituminous coal was 57.8% and 46.9% respectively. The FTIR results of HNO3 treated coal had clearly indicated the presence of N-H group peak at 2923.04cm in case of bituminous coal while introduction of nitro group in lignite coal at 2921.84cm, while both peaks were absent in native coal FTIR spectrum. The elemental analysis of HNO3 treated coal had shown the reduction in carbon content and slight change in sulphur and nitrogen and increase in oxygen content that confirms the oxidation process occurred. The O/C ratio was also increased in case of HNO3 treated coal. 60% and 78% humic acid was extracted from bituminous and lignite coal by potential fungal strains. Both the low and high rank coal were allowed to be treated anaerobically in order to find out the extent of methane production. The raw bituminous (HRC) was capable of producing more methane and carbon dioxide as compared to LRC (Lignite). The co-digestion strategy is productive in stimulation of methane generation but also enhanced its yield. A higher methane potential was observed in the biogasification setup of bituminous coal as compared to lignite coal. The co-substrate Grass peanut shell was able to produce more cumulative methane along with bituminous coal as compared to lignite coal. Grass and peanut shells comprised of lignin and celluloses which are degraded by anaerobic microbes releasing hydrogen which is limiting the reaction rate on the first hand for its deficiency in the reaction mixture. The maximum xii yield of methane and carbon dioxide was obtained at optimum temperature of 35° C and pH 7.0 in both lignite and bituminous coal samples. The whole genome sequencing of potential microbial strains had identified the presence of annotations with their nucleotide base data and gene rank. The circular and sequence view of both the potential fungal isolates Debaromyeces hansenii and Aspergillus ochraceus had shown the particular number of bases present in the nucleotide sequences. The total base pairs of isolate Debaromyeces hansenii were determined as 722,974bp and total number of sequences are 516. The total number of sequences lengths include 1781 sequences in Aspergillus ochraceus.