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Paper and pulp mills generate substantial quantities of cellulose-rich sludge materials that are disposed in landfills at a large scale. For sustainability purposes, sludge materials can be bioprocessed to produce renewable fuels and useful chemicals. The enzymatic hydrolysis of cellulose is the process bottleneck that affects the conversion economics directly by using zero-cost raw materials. In order to study and optimize the process, the characteristics of the sludge raw materials should be first evaluated. In this work, sludge samples were obtained from paper mills located at different locations in Wisconsin and Minnesota. Part of the sludge samples was washed (de-ashed) with hydrochloric acid while the other part remained unwashed. The samples were subjected to multiple spectroscopic analyses techniques to evaluate the morphological properties of cellulose fibers and to estimate the total structural carbohydrate content. The results showed that the de-ashing process changed some fiber characteristics and cellulose crystallinity structure in all sludge samples. Sludge sample A (obtained from Kraft pulp and recycled paper mill region) showed a high percentage of fiber, with crystalline cellulose, compared to the other two sludge samples suggesting that sludge A is a valuable source to make value-added products. Aspen Plus mass and energy calculations performed in view of the ‘zero’ cost and the reliable supply of sludge raw materials producing 2 mol H2/mol glucose. Moreover, the results showed that extracting crystalline cellulose from these sludge samples is more profitable than crystalline cellulose made from the other lignocellulosic feedstocks. The results reported here showed that the utilization of these sludge materials would be an economically attractive and promising alternative for the production of hydrogen.

The white rot basidiomycete, Phanerochaete chrysosporium, employs an array of extracellular enzymes to completely degrade the major polymers of wood: cellulose, hemicellulose and lignin. Towards the identification of participating enzymes, 268 likely secreted proteins were predicted using SignalP and TargetP algorithms. To assess the reliability of secretome predictions and to evaluate the usefulness of the current database, we performed shotgun LC-MS/MS on cultures grown on standard cellulose-containing medium. A total of 182 unique peptide sequences were matched to 50 specific genes, of which 24 were among the secretome subset. Underscoring the rich genetic diversity of P. chrysosporium, identifications included 32 glycosyl hydrolases. Functionally interconnected enzyme groups were recognized. For example, the multiple endoglucanases and processive exocellobiohydrolases observed quite probably attack cellulose in a synergistic manner. In addition, a hemicellulolytic system included endoxylanases, alpha-galactosidase, acetyl xylan esterase, and alpha-l-arabinofuranosidase. Glucose and cellobiose metabolism likely involves cellobiose dehydrogenase, glucose oxidase, and various inverting glycoside hydrolases, all perhaps enhanced by an epimerase. To evaluate the completeness of the current database, mass spectroscopy analysis was performed on a larger and more inclusive dataset containing all possible ORFs. This allowed identification of a previously undetected hypothetical protein and a putative acid phosphatase. The expression of several genes was supported by RT-PCR amplification of their cDNAs.