STUDIES ON THE MICROBIAL PRODUCTION OF FUEL GASES FROM APPLE WASTE AND OTHER SUBSTRATES

Abstract

Hydrogen and methane evolving capacities of apple pomace, dumping wheat and vegetable wastes were monitored in both batch culture digestion and continuous daily batch fed culture digestion, in laboratory scale digesters (0.3 -10 Icapacity). Each kg (dry weight) of apple pomace and dumping wheat fed could generate 40 Iand 36 Iof H2 followed by 140 I&218 Iof CH4 respectively in separate stages. In the case of vegetable waste (raddish leaves, cauliflower leaves &stalk and rotten cabbage), subjected to methanogenesis directly, 320 Iof biogas containing 68% of CH4 was generated. Adaptation of methanogens to changing fruit and vegetable feed material was also observed. The direct biomethanation of apple pomace and dumping wheat increased the CH4 yields, to 157 Iand 280 Iof CH4 / kg of total solids fed respectively, whereas recycling of the unutilized solids in the latter case increased the CH4 efficiency by 27.6%. The digestions led to the maximum reduction of Total solids (TS) by 76.4%, Organic solids (OS) by 80.2% and Chemical oxygen demand (COD) by 70%. Similar biodegradable properties were also seen in other crop grains like maize, rice, pure starch and poultry feed &cattle feed wheat qualities of damaged wheat grains. CH4 content was found to be upto 80% in the total biogas evolved, whereas maximum H2% was of the order of 60% in the total biogas-H generated. The mixed microbial population of H2 producers and pure isolated culture of Bacillus licheniformis, produced H2 from glycolytic pathway intermediates, like glucose, glucose-6- phosphate and pyruvate, in approximately proportionate volumes. The mixed and isolated pure culture actively fermented formate to H2 with a conversion efficiency of 68-85%, thereby following the Escherichia coli pathway as one of the biochemical pathways for H2 generation. The mixed population of H2 producers showed a nitrogenase activity of 0.460 nmoles C2H4 / mg protein / h, when the cells were activated by preincubation for 18-24 h with 0.2% glucose supplementation. Significant inhibitory effect on expression of nitrogenase at 02 levels of >0.25% was observed when 02 sensitivity was monitored. The inhibitory effects of C2H2, CO, KNO3 on nitrogenase and hydrogenase activities for H2 production were measured. These revealed 63% of H2 generation through nitrogenase. The culture fluids were tested for fatty acid and alcohol contents. When mixed culture and pure culture of B. licheniformis was grown separately on glucose, dumping wheat and apple pomace, the fermentation products were acetic acid, propionic acid, butyric acid and valeric acid. The concentration of acetic acid produced has been the highest (222.46 mmoles /I) whereas the concentration of other acids has been 1/10 th as high. Maximum ethanol production of 41.75 mmoles / I has been observed in dumping wheat fermentation, whereas methanol production was found only in apple pomace fermentation. pH changes were monitored during H2 production. It was observed that during active H2 producing stages the pH ranged between 4.6-4.8, whereas the pH dropped to 5.3 from an initial pH of 7.0 when H2 production started. S04 "and Fe +in the concentration range of 1-10 mM and 50 mM respectively, in the culture medium increased the H2 production by 15-30%. Ni2+ showed neutral effects in the concentration range of 0-200 nM tested. Activities of the extracellular hydrolytic enzymes were measured during fermentation of dumping wheat and apple pomace on shake and stationary cultures. The maximum activities observed were, 3x 103, 2.85 x 103, 3.2 x 102 & 1.8 x 101 units/I of hemicellulolytic, proteolytic, anxiolytic and cellulolytic respectively. Viable cells of H2 producers (mixed culture and isolated pure cells of B. licheniformis) were immobilized on solid support of baked bricks and in calcium alginate beads and were tested for H2 production using glucose as substrate. In batch culture biotransformations, 2.4 - 4 fold increase in H2 yields was obtained on bricks support generating a maximum of 1.5 mole of H2 / mole of glucose fed. On the other hand calcium alginate immobilized cells showed a maximum of 2 -fold increase in the yield, thereby evolving 0.75 moles of H2 / mole of glucose employed. However, the daily batch fed continuous culture biotransformations of glucose into H2 by these immobilized cells lead to only 39% improvement of yield. The H2 evolving activity of the calcium alginate immobilized cells could be maintained over a period of one year of storage under sterile conditions.