STUDIES ON ALKALI-TOLERANT WHITE ROT FUNGI FOR BIOBLEACHING OF KRAFT PULP OF A. CADAMBA

Abstract

The pulp and paper industry is the sixth largest energy consumer in the industrial sector and intensive in terms of consumption of raw materials, energy, water and capital requirement. The pulp and paper industry is also facing the problem of an acute shortage of cellulosic fibrous raw materials and soaring environmental cost with other associated issues and challenges like, tough competition from import, obsolete technology, minimum profit level and high energy cost. Due to constant pressure from government statutory bodies and public awareness to mitigate emissions to air and water and decrease in energy cost keeping in view the economic viability of the mill, it is necessary to develop environmental benign technology with energy conservation. Application of biotechnology in pulp and paper industry is helpful to solve the above mentioned purposes to some extent. The use of microbial enzymes for prebleaching of pulp is effective to reduce kappa number before bleaching and thereby saving bleaching chemicals and reducing pollution load. Most of the enzymes available in market are active at slightly acidic conditions and temperature below 70 °C and are cost effective. The pulp produced after brown stock washing is having high temperature and alkaline in nature. Aiming this, the present investigation focuses at developing thermophilic and alkali tolerant enzymes from white rot fungi (Coperinellus dissiminatus) for biobleaching of A. cadamba and their impact on pulp and paper properties and pollution load generated during bleaching The main objectives of the present study are here as under: ➢ To isolate and screen thermo-alkaline xylanases producing white rot fungi. ➢ To produce xylanases from screened isolates under optimum condition of liquid state fermentation using low cost substrate for subsequent application in biobleaching. ➢ To study anatomical, morphological and chemical characterization of A. cadamba. Variation in basic density and chemical composition of A. cadamba (12 years old) ii along the height on a-cellulose, lignin and alcohol-benzene soluble has also been studied. A Optimization of various operating parameters of kraft pulping of A. cadamba and the effect of anthraquinone at optimum pulping condition on pulp yield and kappa number has also been studied. A To study the impact of enzymatic bleaching on different bleaching sequences with regard to pulp viscosity, optical properties and mechanical strength properties and pollution load generated during bleaching 15 strains of white rot fungi were isolated by enriched culture technique using wheat bran as a sole substrate from 30 decaying wood samples. The isolates were purified on wheat bran agar medium (2% wheat bran + 2% agar). For the screening of alkali-tolerant xylanases, the enzymes were produced under liquid state fermentation conditions at 40 °C and pH levels varying from 4.0 to 11.0 and incubation period 8°1 day using 2% wheat bran as a core substrate. Out of 15, only two were found active at pH above 9.0. Based on spore print, fruiting bodies and microscopic examination both isolates were identified as different strains of Coprinellus disseminates. They were designated as fungal strains MLKO1 and MLK07. Both of the fungal strains were screened for extracellular enzymes and found positive for xylanase, cellulase, laccase, lignin peroxidase, amylase, mannanase and protease. The effect of fermentation conditions (incubation period and pH) on enzyme production by the test strains C. disseminates MLKO1 and MLKO7 under L.S.F. were studied. The effect of different sugars (glucose, xylose, galactose and xylan), urea, lignocelluloses as substrates (wheat bran, sugarcane bagasse, wheat straw, saw dust of wood and ground nut shell) were also studied. The crude xylanases from both of strains were analyzed for pH and temperature optima. The effect of different metal ions at 1.0 mM concentration was also studied. Both of the fungal strains MLKO1 and MLKO7 produced maximum xylanases between 8th to 10th days of incubation. Cellulases activity was found to decrease after 7th day of incubation while lignin peroxidase activity increased with increasing the incubation days up to 12 days for strain MLKO I . Cellulases and lignin peroxidase activities were observed maximum at 11th and 13th day of incubation for strain MLK07. All the sugars, repressed the xylanase as well as cellulases activity while lignin peroxidase activity and fungal growth was increased. Wheat bran was found to be the best and cheaper substrate for xylanages production. Additional nitrogen source (urea) found represses the xylanase and lignin peroxidase induction on the other hand, it enhanced the fungal growth and cellulase secretion for both of fungal strains. Xylanase from MLKO1 showed the maximum xylanase activity at pH 7.5 and temperature 75 °C, while xylanase from MLKO7 showed maximum xylanase activity at pH 8.0 and temperature 65 °C. Both the xylanase activity was inhibited by HgC12 and CuSO4 at 1.0 mM concentration whereas; ZnSO4 and FeSO4 found to be enhanced xylanases activity. The enzymes extracted from fungal strains MLKO I and MLKO7 were designated as enzyme-A and enzyme-B respectively. In order to assess the suitability of A. cadamba for pulp and paper making, anatomical, morphological and chemical characterization of A. cadamba was done. Chemical characterization includes water soluble, 1% NaOH, alcohol benzene soluble, holocellulose, lignin, ash and pentosan. A. cadamba contains 20.6% lignin, 19 %, pentosan 76.20%, holocellulose, 20.56% hemicellulose and 44.3% a- cellulose. The variation in basic density and chemical composition (alcohol-benzene soluble, holocellulose, lignin and a—cellulose) of A. cadamba was done along the height i.e. five feet long cylindrical piece from 5, 25 and 40 ft height from base. Average fiber length 1.43 mm, fiber width 38.12 pm, lumen diameter 26.10 pm and cell wall thickness 5.51 was observed. Screened chips of A. cadamba were digested at different cooking conditions. Based on experimental results, the optimum cooking condition for A. cadamba was found as: active alkali 16% (as Na2O), sulphidity 20%, temperature 165 °C, time at temperature 90 min and liquor to wood ratio of 3.5:1. A. cadamba produced 48.74% screened pulp at kappa number iv 22.5. The addition of 0.1 % AQ increases screened pulp yield by 0.38 % but significantly reduces the kappa number by 6.5 units (28.8%). The effect of aging of A. cadamba (2, 3, 4 and 12 years) on screened pulp yield, kappa number and screening rejects was studied. The Baeur-McNett fiber classification of A. cadamba using mess size +20, +48, +100 and +200 of kraft-AQ pulp at optimum cooking condition was carried out. The unbleached pulp of A. cadamba was beaten in PFI mill at different beating levels to optimize various mechanical strength properties like, tear index, tensile index, burst index, double fold, thickness and stretch. The scanning electron microscopic studies of unbleached kraft-AQ fibres were carried out. The unbleached kraft-AQ pulp of A. cadamba was optimized for various operating parameters, like enzyme doses, consistency and reaction time during prebleaching with enzyme-A (strain MLK01) and enzyme-B (strain MLK07) separately and its impact on release of chromophores and reducing sugars in filtrate were studied. The enzymatically prebleached pulps were subjected to alkali extraction with 2% NaOH separately with enzyme-A and B and evaluated for pulp kappa number and viscosity. The impact of enzymatic prebleaching during different CEHH bleaching sequences, carried out at pulps of different kappa numbers, different chlorine demands and at different pH levels during enzymatic prebleaching, on pulp viscosity, brightness, PFI revolutions to get a beating level of 35 °SR, optical and mechanical strength properties and effluent characteristics like, COD, colour and AOX were studied. The bleach boosting effect of enzymatic treatment on 24 kappa number pulp was found better than that of pulp of kappa number 16. At 2% chlorine demand, viscosity, mechanical strength properties and pollution load were observed to improve while brightness was found to decrease compared to 4% chlorine demand. The kraft-AQ pulp of A. cadamba was prebleached at different pH levels i.e. 5.0, 6.0, 8.0 and 9.0 with 4% chlorine demand. The brightness of bleached pulp, COD and colour of combined effluent generated during AXECEHH and BXECEHH bleaching sequences decreased with increasing the pH. On the other hand, pulp viscosity increased with increasing pH and found maximum at pH 9.0. AOX was also found to decrease with increasing the pH. The kraft-AQ pulp of A. cadamba of kappa number 16 was delignified with 02 at consistency 15%, pH 11.0 maintained with 2% NaOH (as such), oxygen pressure 5 kg/cm2, temperature 110 °C and reaction time 90 min in presence of carbohydrate stabilizer i.e. Epsom salt (0.1% MgSO4) followed by alkali extraction with 2% NaOH at 70 °C and 10% consistency for 90 min. After oxygen delignification, the kappa number and pulp viscosity reduced by 38.12 and 18.70% respectively whereas brightness improved by 22.39%. The oxygen delignified pulp was subjected to bleach by ECF bleaching sequences i.e., ODED, OAXEDED and OBXEDED; ODEDP, OAXEDEDP and OAXEDEDP and ODEP, OAXEDEP and OBXEDEP bleaching sequences. The brightness improvement in various rbleaching sequences in ascending order was as: ODED< ODEDP<ODEP for control; OAXEDEP< OAXEDEDP< OAXEDEP for enzyme A and OBXEDED< OAXEDEDP< OBXEDEP for enzyme B. All the ECF bleaching sequences with enzymatic prebleaching after 02 delignification required more PFI revolutions to get 35 °SR beating level compared to control. In all the ECF bleaching sequences, tensile index, burst index and double fold improved except tear index with enzymatic treatment. Both the Enzyme A and B reduce AOX load and increase COD and colour in the combined effluent of respective bleaching sequences compare to control. The reduction in AOX was found decreases sharply after ODL. The introduction of enzyme-A and B in TCF bleaching sequences of kraft-AQ pulp of A. cadamba i.e. OAXEQPP and OBXEQPP improves the brightness and pulp viscosity and increases PFI revolutions to get a beating level of 35 °SR over OQPP bleaching sequence. All the mechanical strength properties except tear index improve in OAXEQPP and OBXEQPP bleaching sequences while COD shows an increasing trend while colour which decreases in both the bleaching sequences compared to. OQPP bleaching sequence