Decolorization and bioremediation of molasses wastewater by white-rot fungi in a semi-solid-state condition

Molasses wastewater (vinasse; the by-product of distillation of fermented sugar) was decolorized and its chemical oxygen demand (COD) was reduced in static cultivation using the fungi Coriolus versicolor, Funalia trogii, Phanerochaete chrysosporium and Pleurotus pulmonarius ('Pleurotus sajorcaju'). The effect of cotton stalk on decolorizing and COD removing capability of four fungi was determined. In the entire concentration range tested (10-30%), wastewater was effectively decolorized by C. versicolor and F. trogii. Cotton stalk addition stimulated the decolorization activity of all fungi. The utilization of cotton stalk represents several advantages due to its function as an attachment place and as a source of nutrients; its use also reduces process costs.     

Effects of various media and supplements on laccase production by some white rot fungi

White rot fungi produce three main extracellular enzymes involved in ligninolysis; laccase, lignin peroxidase and manganese peroxidase. Though all white rot fungi do not produce all three enzymes, laccase occupies an important place in ligninolysis. The present paper reports its production by some white rot fungi; Daedalea flavida, Phlebia brevispora, Phlebia radiata and Polyporus sanguineus under different nutritional conditions. Of the various basal media tested, mineral salts malt extract broth proved to be the best medium for laccase production. Sugarcane bagasse proved to be the best laccase inducer among the various supplements added to different media.     

Taxonomic identification of Streptomyces exfoliatus K10 and characterization of its poly(3-hydroxybutyrate) depolymerase gene

Abstract Using fungi grown on synthetic agar medium, we evaluated and compared the concentration of various H₂O₂-producing enzymes. Our results showed that oxidase production in solid medium was better than that found in liquid medium and as high as that detected in wood samples. High yields of oxidases made it possible to compare different oxidases in the same culture extracts and under different conditions. Our results also indicated that H₂O₂ production is ubiquitous in the white rot fungi tested and that enzyme levels are influenced by the substrate composition.     

A Comparison between the Cellulolytic Activity of White and Brown Rot Fungi. I. The Activity on Insoluble Cellulose

About 70 strains of white and brown rot fungi were cultivated on media, containing filter paper cellulose as the main carbon source. The cellulolytic activity of the culture filtrates was measured after different periods of growth by means of the turbidimetric method. The results obtained indicate a difference between the two types of wood decay fungi as to the capacity of attacking the cellulose used in the medium and in the cellulase test. No significant C₁activity was found in any of the brown rot cultures whereas all white rot fungi tested exerted a measurable activity on the test substrate. The effect of various carbohydrates and some proteins as inducers of cellulase activity was studied. Especially cellobiose and lactose were active on white rot fungi in this respect, particularly in the presence of yeast extract. Also some brown rot fungi exerted C₁-activity after incubation on glucose or cellobiose.     

Industrial and agricultural wastes as substrates for laccase production by white-rot fungi

White-rot fungi,Coriolus versicolor andFunalia trogii, produced laccase on media with diluted olive-oil mill wastewater and vinasse. Addition of spent cotton stalks enhanced the laccase activity with a maximum after 12 d of cultivation.     

A cost-effective cane molasses medium for enhanced cell-bound phytase production by Pichia anomala

AIM: Formulation of an inexpensive cane molasses medium for improved cell-bound phytase production by Pichia anomala. METHODS AND RESULTS: Cell-bound phytase production by Pichia anomala was compared in synthetic glucose-beef extract and cane molasses media. The yeast was cultivated in 250 ml flasks containing 50 ml of the medium, inoculated with a 12 h-old inoculum (3 x 10(6) CFU ml(-1)) and incubated at 25 degrees C for 24 h at 250 rev min(-1). Different cultural parameters were optimized in cane molasses medium in batch fermentation. The cell-bound phytase content increased significantly in cane molasses medium (176 U g(-1) dry biomass) when compared with the synthetic medium (100 U g(-1) dry biomass). In fed-batch fermentation, a marked increase in biomass (20 g l(-1)) and the phytase yield (3000 U l(-1)) were recorded in cane molasses medium. The cost of production in cane molasses medium was pound 0.006 per 1000 U, which is much lower when compared with that in synthetic medium (pound 0.25 per 1000 U). CONCLUSIONS: An overall 86.6% enhancement in phytase yield was attained in optimized cane molasses medium using fed-batch fermentation when compared with that in synthetic medium. Furthermore, the production in cane molasses medium is cost-effective. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytase yield was improved in cane molasses when compared with the synthetic medium, and the cost of production was also significantly reduced. This enzyme can find application in the animal feed industry for improving the nutritional status of feed and combating environmental pollution.     

Copper induction of lignin-modifying enzymes in the white-rot fungus Trametes trogii

Trametes trogii, a white rot basidiomycete involved in wood decay worldwide, produces several ligninolytic enzymes, laccase being the dominant one, with higher titers than those reported for most other white rot fungi studied up to date. The effect of copper on in vitro production of extracellular ligninolytic activities was studied. CuSO(4)·5H(2)O concentrations from 1.6 μM to 1.5 mM were tested in a synthetic medium with glucose 20 g/L and asparagine 3 g/L. The addition of copper (up to 1 mM) did not affect growth but strongly stimulated ligninolytic enzyme production; faster decolorization of the polymeric dye Poly R-478 was observed as well. Maximal production of manganese peroxidase, laccase, and glyoxal oxidase [1.28 U/mL, 93.8 U/mL (with a specific activity of 720 U/mg protein), and 0.46 U/mL respectively] was attained with 1 mM CuSO(4)·5H(2)O. However, higher copper concentrations inhibited growth and notably decreased manganese peroxidase production, although they did not affect laccase secretion. Laccase activity in the culture filtrate was maximal at 50 C and pH 3.4, and the enzyme was completely stable at pH 4.4 and above, and at 30 C for up to 5 d. Denaturing polyacrylamide gel electrophoresis of extracellular culture fluids showed two laccase activity bands (mol wt 38 and 60 kDa respectively). The pattern of isoenzyme production was not affected by medium composition but differed with culture age.     

Degrading ability of oligocyclic aromates by <Emphasis Type="Italic">Phanerochaete sordida</Emphasis> selected <Emphasis Type="Italic">via</Emphasis> screening of white rot fungi

Seventy-nine white rot strains were screened to determine if they had the potential for use in the degradation of oligocyclic aromates (PAHs) by measuring their dye-decoloration rate. Fourteen strains that were selected based on their dye-decoloration rate were then evaluated for the ability to tolerate various levels of PAHs spiked in agar medium. The ability of white rot fungi to degrade 3- or 4-ring PAHs (anthracene, phenanthrene, fluoranthene, pyrene) was determined. Two strains of Phanerochaete sordida (KUC8369, KUC8370) were possible PAHs degraders, degrading a significantly greater amount of phenanthrene and fluoranthene than the culture collection strain P. chrysosporium (a known PAHs degrader). The production of manganese peroxidase, the only extracellular ligninolytic enzyme detected during the cultivation, was evaluated.     

Decolorization of beet molasses vinasse by Bacillus megaterium ATCC 14581

Beet molasses vinasse is formed in large quantities as a waste product of the distillery industry, where the molasses derived from sugar beet is used as a raw material. This vinasse has a dark color, low pH, and chelating properties. Many microorganisms have the ability to decolorize and biodegrade beet molasses vinasse. In this work, different cultivation parameters (the type of the bacterial culture; static or agitated) and medium components ((NH₄)₂SO₄, KH₂PO₄, yeast extract, glucose, and peptone, and the vinasse concentration) were evaluated utilizing Plackett-Burman design to identify the important factors influencing the vinasse decolorization by Bacillus megaterium ATCC 14581. The significant variables were selected as follows: (NH₄)₂SO₄, KH₂PO₄, glucose, and the concentration of vinasse. These four factors should be chosen as being promising for further optimization studies. The maximum color removal was 38%.     

Comparative studies of lignin peroxidases and manganese-dependent peroxidases produced by selected white rot fungi in solid media

Abstract The effect of various incubation conditions and media composition on ligninolytic activity by selected strains of white-rot fungi was determined in solid media. When compared to conventional methods using liquid media or woody substrates, this method is fast, simple and also quantitative. Manganese-dependent peroxidase was easily detected in all strains studied. However, detection of lignin peroxidase required optimisation of both growth medium and enzyme assay conditions. Using this method, we showed that the role of nitrogen and oxygen in ligninolytic activity varies and that conditions must be optimised for each individual even within the same species. Furthermore, several white rot fungi produced manganesedependent peroxidase during the primary growth phase. Keywords: Manganese-dependent peroxidase; Lignin peroxidase; White rot fungus     

Production of Mushroom Mycelium as a Protein and Fat Source in Submerged Culture in Medium of Vinasse

Of ten mushroom cultures investigated, only Agaricus campestris, Boletus indecisus, and Tricholoma nudum were capable of growing in submerged culture in medium of vinasse with added salts. Higher fermentative efficiencies were found under these conditions than in medium containing molasses or waste sulfite liquor. A. campestris showed a better capacity to produce protein but, since B. indecisus is capable of developing greater mycelium weight, its fermentative efficiencies are comparable. Both microorganisms could be grown in medium of vinasse with greatly varied amounts, producing higher mycelial weight in media with greater vinasse. The capacity of B. indecisus and A. campestris to utilize the noncarbohydrate fraction in total solids, instead of the total carbohydrates when they are in smaller amount, was observed in medium containing vinasse. B. indecisus and A. campestris were easily separated by filtration from the medium, although T. nudum was difficult to separate by this procedure. In experiments with A. campestris, the adaptative capacity of the organism to vinasse was demonstrated.     

Use of soybean vinasses as a germinant medium for a Geobacillus stearothermophilus ATCC 7953 sterilization biological indicator

A novel low-cost medium was developed from by-products and wastes from the ethanol agro-industry to replace commercial media in the production of a steam sterilization biological indicator (BI). Various recovery media were developed using soybean or sugarcane molasses and vinasse to prepare a self-contained BI. Media performance was evaluated by viability and heat resistance (D _121 °C value) according to regulatory standards. A medium produced with a soybean vinasse ratio of 1:70 (1.4%) (w/v) produced the results, with D _121 °C = 2.9 ± 0.5 min and Usk = 12.7 ± 2.1 min. The addition of 0.8% (w/v) yeast extract improved the germination of heat-damaged spores. The pH variation from 6.0 to 7.3 resulted in a gradual increase in the D _121 °C value. The absence of calcium chloride resulted in a decrease in germination, while no significant differences were observed with starch addition. Soybean vinasses may thus be used as the main component of a culture medium to substitute for commercial media in the production of self-contained biological indicators. The use of ethanol production waste in this biotechnological process realized a reliable performance, minimized the environmental impact, and decreased BI production costs while producing a high quality product.     

Physiological studies on Phymatotrichum omnivorum. IX. Synthesis of indole acetic acid in vitro.

The cotton root rot fungus, Phymatotrichum omnivorum, synthesized indole acetic acid (IAA) in culture medium containing tryptophan as the only source of nitrogen. Synthesis of IAA was not affected by illumination, and maximum amounts of the auxin were produced at 30 days. The optimum hydrogen ion concentration and temperature for IAA production were 6.5 and 28 degrees C, respectively. Indole ethanol (IEA) was also detected in tryptophan media, primarily during the active growth of the organism.     

毛云芝菌利用糖蜜酒精废水产漆酶培养基优化

为提高毛云芝菌的漆酶产量,降低生产成本,同时使糖蜜废水中的营养成分得到充分利用,变废为宝,以糖蜜废水作为碳源,对毛云芝菌发酵产漆酶进行研究。在单因素试验基础上,以不同浓度糖蜜废水、尿素含量为自变量,漆酶活力为因变量,采用Box-Behnken设计方法得到最佳的培养基配方为糖蜜废水浓度47%,尿素添加量0.5%,漆酶活力可达1 810.0 U/mL,约为优化前的6倍。说明毛云芝菌利用糖蜜废水作为碳源生产漆酶是可行的,为糖蜜废水资源化利用高效生产有价值产品奠定了理论基础。     

Laccase and Tyrosinase in some Wood-rotting Fungi

Several species of wood-rotting fungi have been examined forphenol oxidases. Polystictus sanguineus, a white rot, produceslaccase, secreting it into the medium. This enzyme has beenstudied and partly purified. Phellinus cryptarum, another whiterot, shows varied production of phenol oxidase according tothe medium used for growth. Laccase only is produced on maltmedium, whereas laccase and tyrosinase result after growth ona glucose and salt medium. Lentinus lepideus, a brown rot, producestyrosinase on several media and this enzyme appears to be entirelyintracellular.     

Comparative fermentation studies of industrial strains belonging to four species of solvent-producing clostridia

Industrial and culture collection strains of solvent-producing clostridia, classified as Clostridium acetobutylicum, Clostridium beijerinckii, Clostridium saccharobutylicum, and Clostridium saccharoperbutylacetonicum were utilised in a comparative study of fermentation performance in a laboratory fermentation medium, a molasses fermentation medium, and a maize fermentation medium under standardised culture conditions. At least one representative strain was selected from each of the sub-groups within the four species. Preliminary evaluations were first undertaken for the three different fermentation media to determine the most appropriate media formulations, carbohydrate concentrations, and culture conditions for comparison of the solvent-producing ability of these strains. Standardised fermentation media and culture conditions were then selected for each of the comparative fermentation studies. These included TYA medium containing 4% glucose, a supplemented molasses medium containing 6% fermentable sugars, and a supplemented maize mash medium containing 8% maize. Additional comparative fermentation studies on industrial strains belonging to two species of solvent-producing clostridia were carried out in molasses containing higher concentrations of fermentable sugars, and the sugar concentrations supporting maximum levels of solvent production were determined. Although all the strains tested grew in the maize fermentation medium and degraded starch, only a few strains produced consistently high solvent levels. Optimum starch utilisation and solvent production was obtained at a maize concentration of 80 g/l. Pretreatment of the maize by milling or saccharification decreased the buffering capacity of the medium and resulted in decreased solvent production. Decreasing the time used to gelatinise the starch had little effect. Solvent yields and concentrations obtained in this study were compared with various published data in the scientific and patent literature and appeared to closely simulate the results obtained in the industrial fermentation process. The fermentation performances of individual strains could provide useful comparative data for the selection and development of strains for use on various commercial fermentation substrates.     

不同培养基对富集筛选脱色真菌菌群的效果比较

利用活性黑RB5和活性红M-3BE作为筛选因子,从染料脱色效果、菌群产酶能力以及菌群中的微生物丰富度三方面比较了酵母培养基A、产漆酶真菌培养基B和白腐真菌培养基D在脱色真菌富集筛选方面的效果。富集筛选结果共得到11组具有明显脱色效果的真菌菌群,其中5组来自于D培养基,A和B培养基各获得3组。来自A培养基的3组菌群显示出最好的脱色效果和最大的菌群丰富度,对50mg/L的活性红M-3BE和酸性红A溶液的脱色率最高达到99.53%和97.42%,从中分离到了16株真菌,初步鉴定分属于水霉科、曲霉科(红曲霉属)、节壶菌科和白粉菌科;而B和D培养基中所获得的菌群脱色效果稍差,从中仅得到3株和2株真菌,初步鉴定属于酵母和青霉。A、B两种培养基在各种染料存在下更易产生木质素过氧化物酶,产漆酶能力较弱,而D培养基产漆酶活性较高。     

A critical review of the application of white rot fungus to environmental pollution control

Research on white rot fungi for environmental biotechnology has been conducted for more than 20 years. In this article, we have reviewed processes for cell growth and enzyme production including the factors influencing enzyme productivity and the methods for enhancement of enzyme production. Significant progress has been achieved in molecular biology related to white rot fungi, especially related to the extraction of genetic material (RNA and DNA), gene cloning and the construction of genetically engineered microorganisms. The development of biotechnologies using white rot fungi for environmental pollution control has been implemented to treat various refractory wastes and to bioremediate contaminated soils. The current status and future research needs for fundamentals and application are addressed in this review.     

Extracellular enzyme activities during humic acid degradation by the white rot fungi Phanerochaete chrysosporium and Trametes versicolor

Abstract The relationship between humic acid biodegradation and extracellular lignin peroxidase and Mn-dependent peroxidase activities of two white rot fungi, Phanerochaete chrysosporium and Tranetes versicolor , reported to be lignin degraders, was examined. In experimental conditions promoting culture aeration, particularly with T. versicolor no extracellular peroxidase activity could be detected unless humic acids were included in the culture medium. In the presence of humic acids, appreciable enzymatic activities were determined in the culture filtrate of the two fungi. However, T. versicolor was a more effective degrader than P. chrysosporium , and mineralization assays on synthetic humic acids with culture filtrates showed the important role played by Mn²⁺. The surfactant properties of humic acids are suggested to be responsible for the increase of enzymatic activities.     

ε-PolyLysine production from sugar cane molasses by a new isolates of Bacillus sp. and optimization of the fermentation condition

An attempt was made to use cane molasses as a culture medium for ε-PolyLysine (ε-PL) production by a natural bacterial isolate. The bacterium was identified as Bacillus sp., as confirmed by 16S rDNA sequence analysis. A BLAST result of the sequence indicated that the closest relative of this Bacillus BHU strain was B. thuringiensis, with 97 % homology. The molasses was found to be a better culture medium compared to commonly used culture media comprised of either glucose or glycerol as a carbon source. The various physicochemical parameters were studied for culture growth and polymer production, and were further optimized using response surface methodology (RSM). The correlation coefficient of the resulting model was found to be R ²?=?0.9828. The RSM predicted optimum conditions for ε-PL production (2.46 g/l) by the Bacillus strain was achieved by using molasses, 59.7 g/l; yeast extract, 15.2 mg/l; pH, 6.8 and fermentation time, 42 h at 30 °C. This study represents the first report on the potential application of cane molasses (a byproduct of sugarcane industries) as a culture medium for ε-PL production by Bacillus species. The specific Bacillus strain used in the present study can be exploited for developing a novel technology using inexpensive renewable resources for ε-PL production, a polymer of commercial interest.