A. W. Khan

Comparison of natural hemicellulose and chemically acetylated xylan as substrates for the determination of acetyl-xylan esterase activity in aspergilli

Abstract Native hemicellulose obtained as a result of an aqueous phase thermochemical treatment of aspen wood was tested as a substrate for the determination of acetyl-xylan esterase activities of a number of Aspergilli cultures. The native hemicellulose contained about 60% xylose, of which every two out of three xylose residues were acetylated. Results obtained using the native hemicellulose and chemically acetylated xylan were comparable for the majority of Aspergilli tested, except the enzyme preparations obtained from A. japonicus and A. niger ATTC strain 10864, which were significantly more active on native hemicellulose than on chemically acetylated xylan. The results obtained using 4-nitrophenyl acetate as a substrate showed no relation with those obtained using native hemicellulose or chemically acetylated xylan as substrates. Among the various species and strains of Aspergilli tested , A. japonicus, A. niger ATCC no. 10864, and A. versicolor produced the highest amounts of acetyl-xylan esterase, and the production of this enzyme was greatly induced by the presence of cellulose or xylan in the medium.

Relation between filter paper activity and saccharifying ability of a Trichoderma cellulase system

SummaryTests made to study the relation between filter paper activity and actual saccharifying ability of Trichoderma cellulases show that 30 IU/g of cellulose were sufficient to achieve over 80% hydrolysis of a 25 g/L cellulose suspension in 24 h. With the same enzyme/substrate ratio, but double the concentration of substrate, about 60% hydrolysis was achieved. End- product inhibition is one factor which seriously limits the degree of hydrolysis and therefore the concentration of sugars achievable by enzymatic hydrolysis at high levels of substrate concentration or enzyme/substrate ratio.

Single step conversion of cellulose to ethanol by a mesophilic coculture

SummaryA coculture consisting of two mesophilic anaerobes, produced about 0.8 mole of ethanol per mole of cellulose from a variety of cellulosic materials. The non-cellulolytic member of this coculture, Clostridium saccharolyticum sp. nov. converted glucose and xylose to ethanol and acetic acid in ratios over 4 to 1.

Comparative degradation of cellulose and sugar formation by three newly isolated mesophilic anaerobes and Clostridium thermocellum

SummaryTo develop a coculture for a single step conversion of cellulose to fuels and chemicals, the cellulose degradation by three newly isolated mesophilic anaerobes was compared with that of Clostridium thermocellum. Degradation of cellulose obtained from cotton linters, and delignified ball-milled pulp and steam-exploded aspen wood by mesophilic anaerobes was comparable to that by Cl. thermocellum. However mesophilic anaerobes produced larger amounts of sugars. In media containing 50 g/l of cellulose suspensions, the sugars produced by mesophilic anaerobes were 10.5 to 14.5 g/l, about 50% of the cellulose used.

Use of Pretreated Lignocellulose for the Production of Cellulases

SummaryTests made to utilize lignocellulosics as a substrate for the production of cellulases showed that the enzyme production from steam and explosion decompressed aspen wood (SED) by Tricoderma reesei RUT-C30 was low, and the enzyme system produced was deficient in exoglucanase and β-glucosidase activities. Mixing this substrate with 10–20% pure cellulose lessened this deficiency and improved enzyme production. The enzyme system produced from the mixed substrate was rich in xylanase and had saccharifying ability equal to that produced in medium containing pure cellulose.

Use of charcoal to minimize end product inhibition in enzymatic hydrolysis of cellulose

SummaryTests made to improve saccharification of cellulose byTrichoderma cellulases showed that charcoal used as an adsorbent minimized the end product inhibition. Charcoal adsorbed both cellobiose and glucose and did not affect the enzymatic hydrolysis of cellulose. Results showed that charcoal is as effective as β-glucosidase in improving the enzymatic saccharification of cellulose.

Conversion of cellobiose and xylose to ethanol by immobilized growing cells of Clostridium saccharolyticum on charcoal support

SummaryAn immobilization technique has been developed for the conversion of both cellobiose and xylose to ethanol, which may be considered as one stage of a process for the conversion of cellulosic biomass to ethanol. Relatively inexpensive charcoal was used as a support material, with 23 mg dry weight of Clostridium saccharolyticum cells per g dry weight of support. Tests were run for 170 h at 0.15 1/h dilution rate. From a 3% (w/v) sugar mixture, 0.7% (w/v) ethanol was obtained with over 97% cellobiose and 62% xylose utilization.

Factors affecting the utilization of steam- and explosion-decompressed aspen wood by cellulolytic anaerobes

Acetivibrio cellulolyticus and Clostridium thermocellum were unable to grow in medium containing 2 g litre−1 of untreated steam- and explosion-decompressed (SED) aspen wood as a source of carbon. When the medium was concurrently supplemented with 5 g litre−1 of Whatman CF11 cellulose powder, growth of both anaerobes, to levels comparable to media containing only CF11 cellulose, was observed. Treatment of the SED wood with ethanol-water, dioxane-water or sodium hydroxide released adhered lignin or lignin residues and rendered the SED wood suitable for the growth of both anaerobes. The results indicate that at concentrations of 4 g litre−1 or lower, growth on untreated SED wood is probably prevented because of inaccessibility of adherence sites for these bacteria, and at concentration of 4 g litre−1 or more it is also inhibited due to increased concentration of inhibitory materials such as lignin and lignin residues.

Formation of esterase, xylanase and xylosidase by cellulolytic anaerobes☆

Three mesophilic anaerobes, namely Acetivibrio cellulolyticus, Acetivibrio cellulosolvens and Bacteroides cellulosolvens and a thermophilic anaerobe, namely Clostridium thermocellum, produced xylanase, xylosidase and esterase activities when grown on medium containing cellulose. Xylanase and xylosidase activities produced by both the mesophilic and thermophilic anaerobes were stimulated by the presence of ascorbic acid, cysteine or dithiothreitol, but differed in their stability at high temperatures, their response to the presence of calcium and magnesium and they had different temperature optima. Esterase activity from both mesophilic and thermophilic anaerobes had a pH optimum between 5.5 and 6.0 but the temperature optimum of the esterase was 37°C for mesophilic and 50°C for thermophilic anaerobes. The strains used in this study were unable to utilize glucose or xylose and their growth on xylan was limited. These anaerobes appear to produce these enzymes as a part of their cellulolytic enzyme system and degrade xylan during their growth on cellulose.

Properties of a butyrate-forming Clostridium present in cellulose-enriched methanogenic culture.

Abstract Tests were made to determine the cause of butyrate formation in a cellulose-enriched culture from sewage sludge containing cellulolytic anaerobes that do not produce this acid. The results showed the presence of a saccharolytic butyrate-forming Clostridium . In pure culture, this Clostridium produced ethanol, acetate and butyrate from a variety of sugars, and was non-motile, without peritrichous flagella. In coculture with cellulocytic anaerobes, this Clostridium helped in improving cellulysis by removing sugars that are not used by these anaerobes but inhibit their growth. In mixed culture fermentations failing because of poor methanogenesis, the H 2 is not completely removed. The results showed that, unlike cellulytic anaerobes, the growth of this Clostridium was not affected by the presence of H 2 . It appears its growth in the presence of H 2 causes the accumulation of butyrate.