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Indigo carmine dye is used in many industries like textile, paper, cosmetics, etc. It is specially used in textile industries for dyeing denim. The untreated water discharged from these industries, especially the textile industry, poses a significant environmental threat. The conventional physicochemical methods used in treating industrial effluent are very expensive and cause secondary pollution. In the present study, an extracellular laccase-producing Bacillus licheniformis NS2324 was isolated, and a crude enzyme was used to degrade indigo carmine dye. The conditions needed for decolorisation were standardised. Optimisation studies for dye decolorisation by NS2324 laccase revealed that the enzyme was able to decolorize dye efficiently at 50 oC temperature, pH 8, with an enzyme dose of 10 IU/ml after 6 hours. Up to 96.79±1.06 per cent decolorisation was achieved under optimised conditions without the use of any mediator of laccase. Complete degradation of indigo dye by laccase in the absence of mediator makes the present study very useful for treating textile effluent from the denim industry.

Amylases have discovered applications in various industries including juice preparing, processing of starch, bread improvement and other maturation forms including malting grain and in baking ventures. In this investigation, amylase extracted from Bacillus licheniformis MTCC 1483 was applied for the preparation of buns and for improvement of bread quality. It was evaluated that maximum loaf volume was obtained by an application of 3.5 unit/Kg flour of amylase at 37°C, pH 7.0 for 30min. Tactile attributes of bread viz. color, taste, flavor and acceptability were additionally upgraded. Thus, the bread prepared by using alpha amylase showed an increase in loaf volume and bread height with better taste and flavor. Overall results showed applicability of alpha amylase in baking industry.

Malachite green, a triphenylmethane dye, is used in aquaculture to limit the growth of protozoans and fungi in fish. It is also employed in the food, medicinal, and textile sectors. MG has high toxicity covering micro organisms and higher eukaryotes. Its toxic effects include organ damage, developmental abnormalities and mutagenic/carcinogenic potentials. Hence it is of utmost importance to degrade MG. Therefore, in this study with the help of catalytic properties of laccase, biodegradation of dye-malachite green was studied with respect to different factors like temperature, pH, enzyme dose and treatment time. The maximum of 99.31±0.97% MG decoloration was obtained under the conditions of 10 U ml-1 enzyme, pH 8.0 incubation time 1h and a temperature of 50°C. MG was degraded without any mediator of laccase in the current study.

Amylases are starch-degrading enzymes produced by many bacteria, plants, fungi and animals. It has found tremendous application in industry. One of its applications is in fruit industry wherein amylases are used to clarify fruit juices and reduce its viscosity. In fruits, high amount of starch is present which resulted in increased viscosity of the prepared juices. This may also lead to settling of juice at bottom. Amylases act on starch component and degrade it. In the current study, amylase from B. licheniformis was used for the clarification of apple, orange and grape juices. Total suspended solids, viscosity and total acidity was found to decrease with increasing amylase concentration. The results revealed that after amylase treatment the color, texture and flavor of juices were also improved.

Laccases are multicoopper oxidases catalyzing the oxidation of phenolic as well as non-phenolic compounds. Laccases show typical blue color due to the presence of covalent Type 1 Cu-Cys bond which absorbs at 600 nm. However, recently some white laccases have also been identified which lacks typical spectra of blue laccases and do not show peak at 600 nm. In the present study, a novel white laccase was isolated from Bacillus sp. MSK-01. MSK laccase was purified and characterized in detail and the purified laccase was referred to MSKLAC. It has a molecular weight of 32 KDa. UV–visible spectrum of purified MSKLAC do not show characteristic peak at 600 nm and bend at 330 nm. The enzyme was repressed by conventional inhibitors of laccase like sodium azide, cysteine, dithiothreitol and β-mercaptoethanol. The laccase was highly thermo-stable enzyme having optimum temperature of 75 °C and could treasure more than 50% activity even at 100 °C. The optimum pH for ABTS and guaiacol was 4.5 and 8.0 respectively. MSKLAC was stable in the presence of most of the metal ions and surfactants. The effect of MSKLAC on lung cancer cell line was also assessed. It was observed that MSKLAC is inhibitory to lung cell cancer line. Thus, MSKLAC has potential to be used as an anti-proliferative agent to cancer cells.

Laccase gene from Bacillus licheniformis NS2324 was cloned and expressed in E. coli by using pUC 18 as cloning vector and pet 15b as expression vector. The purified recombinant laccase (rLacNS2324) showed a molecular mass of 66 KDa. The optimum pH and temperature for rLacNS2324 was found to be pH 8 and 40 °C respectively. The half life of rLacNS2324 at pH 7, 8 and 9 is 24 h. The half life of laccase at 45 °C is 8 h. Laccase activity was increased in the presence of Cu

Cost-effective production of bioethanol from waste material is becoming the need of the hour to combat the exhaustive nature of fossil fuel. In this study, bioethanol was produced from microwave-pretreated kitchen waste at high dry material consistency. Pretreatment was performed for 30 min at a constant power of 90 W. Liquefaction/saccharification was done with in-house produced amylase from Bacillus licheniformis MTCC 1483. The liquefaction step was optimized using response surface modeling. Three factors, viz. pH, concentration of dry substrate and amylase, were optimized by using reducing sugar and ethanol yield as response. The optimum conditions of input parameters obtained were pH 7.5, dry material 40% (w/v) and amylase 15 IU g−1. The process developed in the present study leads to 0.129 g ml−1, i.e., 0.32 g per g biomass ethanol production. The novelty of the manuscript lies in the fact that no acid/alkali hydrolysis was carried out for the release of reducing sugar. Instead, microwave treatment was carried out at low power for longer time so as to release maximum sugar. The cost incurred in bioethanol production was also estimated by taking cost of chemicals, instruments and operating cost in account. The total cost of bioethanol produced in the present study was calculated as 0.143 $/l of ethanol. A 8.32-fold decrease in price of ethanol produced in the present study was observed when compared to the market selling price of ethanol. This makes the developed process economically and industrially feasible.

Bioprocessing of pulp requires lignolytic as well as hemicellulolytic enzymes. The present study is the first report of a cocktail of laccase (L), xylanase (X), and mannanase (M), from a single bacterium for pulp biobleaching. A novel strain Bacillus tequilensis LXM 55 produced thermo-alkali stable L + X + M. On optimization higher enzyme yield (IUml−1/fold increase) of laccase (396.35/24.16), xylanase (212.95/81.90) and mannanase (153.33/102.90) were achieved in the cocktail. Treatment of pulp with cocktail of enzymes led to 49.35% reduction in kappa number and considerable enhancement in the brightness (11.59%), whiteness (4.11%), and other pulp properties. Most importantly, no mediator system was required for the application of laccase. 40% less chlorine consumption was required to obtain the paper of the same quality as that of pulp treated without enzyme but with 100% chlorine. Therefore, this cocktail of enzymes is highly suitable for pulp biobleaching in the paper mill.

Decolourization of textile effluent is always being a problem of major concern for the safe disposal of effluent water in river stream. Due to intense colour of the effluent water, even after the treatment, disposal of the water in river stream is under strict environmental regulation. Laccase, a natural oxidase, have the miraculous power to decolourize different dyes with or without mediators. However, stability and cost of implementation of the enzyme at industrial conditions make the entire process unfeasible at industrial scale. In the present study, a highly thermo-alkali-stable, metal, surfactant and organic solvent tolerant laccase from Bacillus sp. MSK-01 was immobilized in Cu-alginate bead with ABTS mediator under standardized conditions. A continuous flow packed bed bioreactor was formulated to develop a continuous method of the treatment of effluent water. Results showed 66% reduction in the colour of the effluent. UV-VIS spectrum analysis of the treated and untreated samples showed the formation of the degradation products of dyes due to the action of laccase. The developed process can be made useful for on site industrial application.

Textile and clothing industry, besides being a growing market worldwide, is also a big contributor to environmental pollution due to the generation of highly polluted wastewater. Textile dyeing and processing utilized large amount of water and in turn generates huge amount of polluted water. Because of the serious concern of environmentalist, the discard of this water to the river stream without treatment is banned. Therefore, the industries have adopted the various strategies to deal with the effluent before its discard to stream. Moreover, because of scarcity of water in near future, the reutilization of treated water can also be a useful approach. In this chapter, various physical, chemical, biological, and hybrid methods are discussed to deal with the effluent water. Physical methods such as filtration, coagulation, chemical methods such as oxidation, and biological methods such as aerobic and anaerobic oxidation are discussed in respect of the development done in past years with their loopholes. A hybrid approach utilizing the collective effort of all these methods is also discussed.

Laccases are multicopper oxidases containing four copper atoms per monomer distributed in three redox sites. Because of its tremendous applications in different areas, isolation of new laccases with wide range of industrial implementation. The present study focuses on the optimization of laccase production from Bacillus sp. MSK-01 under solid state fermentation conditions using fruit juice waste as the substrate. MSKLAC was produced extracellularly by the bacteria. This laccase was able to oxidize ABTS and syringaldazine. Various nutritional and environmental factors were utilized for increasing the enzyme yield. Plackett Burman was used to study the influence of input parameters on laccase yield. Tween-80, initial moisture ratio and magnesium sulphate were the major influencing factor affecting laccase yield. Central composite design of RSM was used for the modelling of experiment. Desirability approach was used to optimize laccase yield. Maximum laccase yield of 1645 IUg−1 was obtained when 0.55% of tween -80, 1:2.34 initial moisture ratio and 300μM magnesium sulphate was used. A 470 fold increase in the yield of laccase from unoptimized condition was obtained.

Treatments of industrial effluent/s by enzymes offer significant advantage over the use of microorganisms. However they have not been explored for the bioremediation of effluent from pulp and paper industry. This may be because of the less availability of lignolytic enzymes which can be active/stable under extreme environmental conditions that are present in effluent treatment plant of pulp and paper mills. In present study use of thermoalkali stable laccase from Bacillus tequilensis SN4 has been explored for the bioremediation of paper mill effluent. Under standardized conditions significant reduction in various parameters , color 83%, BOD 82%, COD 77%, phenol content 62%, lignin content 74%, TDS 28% and TSS 34%, was achieved. Major advantage was that treatment time was only 4h, much less than the bioremediation reported with the use of microorganisms, which is usually four to five days. Results indicate that SN4 laccase is very suitable to develop a process for the effective bioremediation of effluent from pulp and paper industry.

Protease has been in increasing demand in industries due to its hydrolytic nature. In industries, high yield of enzyme is required to meet the industrial need at a relatively cheaper cost. In the present study, the protease from Bacillus sp. MSK-01 was produced in large quantity by submerged fermentation. Statistical techniques including Plackett-Burman and Response surface methodology are useful tools for optimizing many parameters at a time and are used for increasing the protease production from Bacillus sp. MSK-01. 19 different parameters were chosen, out of which 15 factors had positive effect on protease yield. Four maximum influencing factors were peptone, magnesium sulphate, skim milk powder and casein were chosen to further increase the protease yield. 397.3 IU ml-1 of enzyme yield was obtained under optimized conditions which lead to 198 fold increase in the yield of protease from unoptimized condition.

Amylase has been in increasing demand in industries due to its hydrolytic nature. Solid state fermentation (SSF) is a cost effective method for increasing the enzyme production. In the present study, amylase from Bacillus licheniformis MTCC 1483 was produced in large quantity by solid state fermentation using paddy straw as substrate. Response surface methodology is a useful tool for optimizing many parameters at a time and is used for increasing the amylase production. 8523 IUg-1 of enzyme activity was obtained under optimized conditions which lead to 35 fold increase in the yield of amylase from unoptimized condition.

DNA sequence analysis and gene expression analysis through genomic signal processing played an important role in cancer diagnosis in recent years. Cancer diagnosis through gene expression data, discrete Fourier transform, discrete wavelet transform (DWT) and IIR low pass filter are frequently used but suffer from drawbacks like longer essential time-support. Analytic wavelet transform with optimal filter coefficients for cancer diagnosis using genomic signals is designed in this paper. The proposed technique consists of three modules namely, pre-processing module, optimisation module and transform and cancer diagnosis module. Initially, the filter coefficients are optimally found out using group search optimiser. Then, the optimal coefficients and the pre-processed DNA sequence is applied to analytic wavelet transform and subsequently, diagnosis for the cancer cell is made based on the threshold. DNA sequences obtained from National Centre for Biotechnology Information (NCBI) forms the database for the evaluation. Evaluation metrics parameters employed are sensitivity, specificity and accuracy. Comparison is made to the base method and analytic transform technique for more analysis. From the results, we can observe that the proposed technique has achieved good results attaining accuracy of 91.6% which is better than other compared techniques.

Degradation of residual lignin in kraft pulp by chemical bleaching is implicated in causing environmental pollution. The use of thermo- and alkali-tolerant bacterial laccases is considered to be important biological alternative to chemical processing. Laccases from Bacillus species have shown promise in this respect but their intracellular/spore bound presence make their industrial application economically unfeasible. We report here on a novel extracellular active thermo-alkali-stable laccase (SN4 laccase) which is active at 90 °C and pH 8.0 using 2,6-dimethoxyphenol as substrate from Bacillus tequilensis SN4. SN4 laccase retained 27 % activity for 5 min at 100 °C and more than 80 % activity for 24 h at 70 °C. The enzyme is also stable at a higher pH (9.0–10.0). Enzyme production was optimized by submerged fermentation. Relatively high yields (18,356 nkats ml−1) of SN4 laccase was obtained in a medium containing 650 μM MnSO4, 350 μM FeSO4, and 3.5 % ethanol. A 764-fold increase in laccase activity was observed under optimal conditions. In addition, reduction in kappa number and increase in brightness of softwood pulp by 28 and 7.6 %, respectively, were observed after treatment with SN4 laccase without a mediator. When N-hydroxybenzotriazole was used as a mediator, the kappa number was decreased to 47 % and brightness was increased to 12 %.

A novel extracellular thermo-alkali-stable laccase from Bacillus tequilensis SN4 (SN4LAC) was purified to homogeneity. The laccase was a monomeric protein of molecular weight 32 KDa. UV-visible spectrum and peptide mass fingerprinting results showed that SN4LAC is a multicopper oxidase. Laccase was active in broad range of phenolic and non-phenolic substrates. Catalytic efficiency (k cat/K m) showed that 2, 6-dimethoxyphenol was most efficiently oxidized by the enzyme. The enzyme was inhibited by conventional inhibitors of laccase like sodium azide, cysteine, dithiothreitol and β-mercaptoethanol. SN4LAC was found to be highly thermostable, having temperature optimum at 85°C and could retain more than 80% activity at 70°C for 24 h. The optimum pH of activity for 2, 6-dimethoxyphenol, 2, 2′-azino bis[3-ethylbenzthiazoline-6-sulfonate], syringaldazine and guaiacol was 8.0, 5.5, 6.5 and 8.0 respectively. Enzyme was alkali-stable as it retained more than 75% activity at pH 9.0 for 24 h. Activity of the enzyme was significantly enhanced by Cu2+, Co2+, SDS and CTAB, while it was stable in the presence of halides, most of the other metal ions and surfactants. The extracellular nature and stability of SN4LAC in extreme conditions such as high temperature, pH, heavy metals, halides and detergents makes it a highly suitable candidate for biotechnological and industrial applications.