Antioxidant compounds produced by Pseudocercospora sp. ESL 02, an endophytic fungus isolated from Elaeocarpus sylvestris

Objective: To isolate endophytic fungi from Elaeocarpus sylvestris (E. sylvestris) and to isolate antioxidant compounds from a potential source fungus. Methods: Endophytic fungi were isolated from fresh leaves and stems of E. sylvestris and identified based on DNA analysis. 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity was used to evaluate the antioxidant activity of the fungi. The po-tential antioxidant fungus was further studied to isolate antioxidant compounds. The isolated compounds were identified by melting point analysis, optical rotation, spectral analysis using a UV spectrophotometer, high resolution fast atom bombardment mass spectrometry, X-ray crystallography analysis, 1H nuclear magnetic resonance analysis and 13C nuclear magnetic resonance analysis. The isolated compounds were evaluated with DPPH radical scavenging, reducing power, and b-carotene bleaching assays. Results: Seven endophytic fungi were successfully isolated from E. sylvestris and identified as Pestalotiopsis sp. EST 01, Pestalotiopsis sp. EST 02, Diaporthales sp. EST 03, Meyerozyma sp. EST 04, Diaporthales sp. EST 05, Pestalotiopsis sp. ESL 01, and Pseudocercospora sp. ESL 02. Of the seven fungi, Pseudocercospora sp. ESL 02 had the highest antioxidant activity [IC50 = (30.54 ± 0.88) mg/mL]. From that fungus, two compounds identified as terreic acid (1) and 6-methylsalicylic acid (2) were isolated with an IC50 of DPPH radical scavenging activity of (0.22 ± 0.02) mmol/L and (3.87 ± 0.27) mmol/L, respectively. The compounds also had good activities from the reducing power and b-carotene bleaching assays. Conclusions: The Pseudocercospora sp. ESL 02 fungus isolated from E. sylvestris looks promising as a novel source of terreic acid.

Effects of saline-alkaline stress on benzo[a]pyrene biotransformation and ligninolytic enzyme expression by Bjerkandera adusta SM46.

Benzo[a]pyrene (BaP) accumulates in marine organisms and contaminated coastal areas. The biotreatment of waste water using saline-alkaline-tolerant white rot fungi (WRF) represents a promising method for removing BaP under saline-alkaline conditions based on WRF's ability to produce ligninolytic enzymes. In a pre-screening for degradation of polycyclic aromatic hydrocarbons of 82 fungal strains using Remazol brilliant blue R, Bjerkandera adusta SM46 exhibited the highest tolerance to saline-alkaline stress. Moreover, a B. adusta culture grown in BaP-containing liquid medium exhibited resistance to salinities up to 20 g l(-1). These conditions did not inhibit fungal growth or the expression of manganese peroxidase (MnP) or lignin peroxidase (LiP). The degradation rate also became higher as salinity increased to 20 g l(-1). Fungal growth and enzyme expression were inhibited at a salinity of 35 g l(-1). These inhibitory effects directly decreased the degradation rate (>24%). The presence of MnSO4 as an inducer improved the degradation rate and enzyme expression. MnP and LiP activity also increased by seven- and fivefold, respectively. SM46 degraded BaP (38-89% over 30 days) in an acidic environment (pH 4.5) and under saline-alkaline stress conditions (pH 8.2). Investigating the metabolites produced revealed BaP-1,6-dione as the main product, indicating the important role of ligninolytic enzymes in initializing BaP cleavage. The other metabolites detected, naphthalene acetic acid, hydroxybenzoic acid, benzoic acid, and catechol, may have been ring fission products. The wide range of activities observed suggests that B. adusta SM46 is a potential agent for biodegrading BaP under saline conditions.

Antioxidant and α-glucosidase inhibitor activities of natural compounds isolated from Quercus gilva Blume leaves

To isolate and investigate antioxidant and α-glucosidase inhibitor compounds in the leaves of Quercus gilva Blume (Q. gilva). Methods: Dry leaves of Q. gilva were extracted with methanol and the methanolic extract was further separated by silica gel column chromatography using several solvents with increasing polarity. The antioxidant activities of the isolated compounds were evaluated using various in vitro assays: 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity, hydrogen peroxide radical scavenging activity, β-carotene bleaching assay, and reducing power assay. The α-glucosidase inhibitory assay was conducted against α-glucosidase from Saccharomyces cerevisiae. Results: Three compounds were isolated and their structures were identified as catechin (1), epicatechin (2), and tiliroside (3) using an instrumental analysis. Compound 2 had higher antioxidant activity with inhibitory concentrations (IC50) of (22.55 ± 2.23) μmol/L than that of quercetin, which was used as the standard, with an IC50 of (28.08 ± 2.39) μmol/L, followed by compound 1 with IC50 of (40.86 ± 3.45) μmol/L. On the other hand, compound 3 had the lowest antioxidant activity with an IC50 of (160.24 ± 8.15) μmol/L. However, compound 3 had the highest α-glucosidase inhibitory activity with an IC50 of (28.36 ± 0.11) μmol/L, followed by compounds 1 and 2 with (168.60 ± 5.15) and (920.60 ± 10.10) μmol/L, respectively. Conclusions: The results obtained for the antioxidant activities and α-glucosidase inhibitory activities in a methanolic extract from the leaves of Q. gilva confirmed the potential of this plant as a source of natural antioxidants and antidiabetic medicine.

Potential of fungal co-culturing for accelerated biodegradation of petroleum hydrocarbons in soil.

The potential of fungal co-culture of the filamentous Pestalotiopsis sp. NG007 with four different basidiomycetes--Trametes versicolor U97, Pleurotus ostreatus PL1, Cerena sp. F0607, and Polyporus sp. S133--for accelerating biodegradation of petroleum hydrocarbons (PHCs) was studied using three different physicochemical characteristic PHCs in soil. All the combinations showed a mutual intermingling mycelial interaction on the agar plates. However, only NG007/S133 (50/50) exhibited an optimum growth rate and enzymatic activities that supported the degradation of asphalt in soil. The co-culture also degraded all fractions at even higher concentrations of the different PHCs. In addition, asphaltene, which is a difficult fraction for a single microorganism to degrade, was markedly degraded by the co-culture, which indicated that the simultaneous biodegradation of aliphatic, aromatic, resin, and asphaltene fractions had occurred in the co-culture. An examination of in-vitro degradation by the crude enzymes and the retrieval fungal culture from the soil after the experiment confirmed the accelerated biodegradation due to enhanced enzyme activities in the co-culture. The addition of piperonyl butoxide or AgNO3 inhibited biodegradation by 81-99%, which demonstrated the important role of P450 monooxygenases and/or dioxygenases in the initial degradation of the aliphatic and aromatic fractions in PHCs.

Enhanced biodegradation of asphalt in the presence of Tween surfactants, Mn(2+) and H2O2 by Pestalotiopsis sp. in liquid medium and soil.

Asphalt and fractions thereof can contaminate water and soil environments. Forming as residues in distillation products in crude oil refineries, asphalts consist mostly of asphaltene instead of aliphatics, aromatics, and resins. The high asphaltene content might be responsible for the decrease in bioavailability to microorganisms and therefore reduce the biodegradability of asphalt in the environment. In this study, the effect on asphalt biodegradation by Pestalotiopsis sp. in liquid medium and soil of nonionic Tween surfactants in the presence of Mn2+ and H2O2 was examined. The degradation was enhanced by Tween 40 or Tween 80 (0.1%) in the presence of Mn2+ (1 mM) and H2O2 (0.05 mM). A Tween surfactant, Mn2+, and H2O2 can overcome bioavailability-mediated constraints and increase ligninolytic activities, particularly manganese peroxidase and laccase activities. The study is significant for the bioremediation of asphalt and/or viscous-crude oil-contaminated environments.

Evaluation of antioxidant and α-glucosidase inhibitory activities of some subtropical plants.

The antioxidant and α-glucosidase inhibitory activities of the methanolic leaf extracts of some subtropical plants were evaluated in the present study. Antioxidant activity was evaluated based on 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity, reducing power, hydrogen peroxide and ß-carotene bleaching assays. α-Glucosidase inhibitory activity and enzyme kinetics as well as the total phenolic content of the extracts were also investigated. Elaeocarpus sylvestris extract had the highest activities on all the antioxidant assays performed such as DPPH scavenging activity (IC50 12.7 ± 0.5 µg mL(-1)), reducing power (491.1 ± 6.3 mg QE g(-1) dry extract), hydrogen peroxide (IC50 65.6 ± 0.4 µg mL(-1)) and ß-carotene bleaching assays (IC50 5.1 ± 1.9 µg mL(-1)). The total phenolic content of the E. sylvestris extract also had the highest values for gallic acid, quercetin and rutin equivalents (353.8 ± 28.6 mg GAE g(-1) dry extract; 294.9 ± 24.5 mg QE g(-1) dry extract; 663.0 ± 52.3 mg RE g(-1) dry extract, respectively). α-Glucosidase inhibition assay revealed that Distylium racemosum had the highest activity with an IC50 value of 22.6 ± 1.9 µg mL(-1). The results of the present study revealed the potencies of E. sylvestris, D. racemosum, Acer mono Maxim and Liquidambar styraciflua as alternative sources for antioxidants and α-glucosidase inhibitors.

Potent α-glucosidase inhibitors isolated from Ginkgo biloba leaves.

In vitro α-glucosidase inhibitory activity of Ginkgo biloba leaves was investigated. The inhibitory activity of methanol extracts from yellow and green leaves was 13.8 and 40.1 µg mL(-1), respectively. Each methanol extract was separated into its respective fraction by solvent-solvent extraction with n-hexane, chloroform, ethyl acetate and n-butanol. The n-hexane fractions (in both methanol extracts from green and yellow leaves) exhibited high α-glucosidase inhibitory activity with IC50 values of 13.6 and 13.4 µg mL(-1), respectively. Further fractionation of the n-hexane fractions by silica gel column chromatography gave the most active fraction which was identified as ginkgolic acid (C13:0) and a mixture (C13:0, C15:0, C15: 1, C17:1 and C17:2). Ginkgolic acid (C13:0) exhibited the highest α-glucosidase inhibitory activity. This is the first study to successfully isolate ginkgolic acids as α-glucosidase inhibitors.

Isolation of hyperoside and isoquercitrin from Camellia sasanqua as antioxidant agents.

Two antioxidant active compounds were isolated from the methanol extract of Camellia sasanqua using various in vitro assays: 1,1-diphenyl-2-picrylhydrazyl (DPPH), ß-carotene bleaching and reducing power assays. The ethyl acetate (EtOAc) fraction of the methanol extract had the highest DPPH radical-scavenging activity with an Inhibition Concentration (IC50) value of 18.3 ± 1.63 µg mL(-1). Sephadex LH-20 column chromatography was used to separate the EtOAc fraction into eight fractions (F1-F8). Antioxidant activity was significantly higher in fraction 5 with an IC50 value 14.61 ± 0.02 µg mL(-1). Fraction 5 was further separated by HPLC preparative with Capcellpak C18 MG followed by the Cosmosil 5C18-AR-II column, using a guided DPPH radical-scavenging assay. The compounds isolated were identified as: Hyperoside (1) and isoquercitrin (2) after recrystallization from ethanol, based on Mass Spectrum (MS) and Nuclear Magnetic Resonance (NMR) analyses. Their DPPH radical-scavenging activities based on the 50% scavenging concentration decreased in the following order: Isoquercitrin (21.6 mM) > hyperoside (27.5 mM). The antioxidant activities of hyperoside and isoquercitrin were 67.52 ± 0.64 and 64.33 ± 0.51%, respectively, in the ß-carotene bleaching assay. These compounds were found to have good reducing powers (OD value: 2.5-3.8) at concentrations of 50-140 µg mL(-1), using the potassium ferricyanide reduction method. Although, these compounds are well-known, hyperoside (1) was isolated from this herb for the first time.

Effect of media compositions on α-glucosidase inhibitory activity, growth and fatty acid content in mycelium extracts of Colletotrichum sp. TSC13 from Taxus Sumatrana (Miq.) de Laub.

The active α-glucosidase inhibitor compounds in the endophytic fungus Colletotrichum sp. TSC13 were found to be the unsaturated fatty acids (oleic, linoleic and linolenic acids). These compounds have potential as antidiabetic agents. The aim of the present study is to investigate the effects of various media composition on growth (mycelium dry weight) and the fatty acids content (µg mg(-1) mycelium DW) of Colletotrichum sp. TSC13 in relation to its α-glucosidase inhibitory activity. For that purpose, the experiments were set up by varying the carbon and nitrogen sources, metal ions and desaturase and fatty acid synthase inhibitors in the media. Colletotrichum sp. TSC13 grown on potato dextrose broth (PDB) was used as control. The α-glucosidase inhibitory activities were (range from 43.9 ± 2.5 to 88.6 ± 5.2%) at 10 µg mL(-1). This activity seemed to correlate with the unsaturated fatty acids content of the samples. Different sugars as carbon source experiment showed that xylose gave the highest growth (938.7 ± 141.6 mg). However, the highest fatty acids content was obtained from fructose medium which containing linoleic acid (38.8 ± 4.9 µ g mg(-1) DW). Soluble starch gave better growth (672.5 ± 62.3 mg) but very low fatty acids content (2.8 ± 0.1 µg mg(-1) DW) was obtained. Yeast extract was the best nitrogen source. Fatty acids production was better as compared to beef extract and soytone. This is the first report of various media compositions on fatty acids content in Colletotrichum sp. TSC13 in relation to its α-glucosidase inhibitory activity.

Bioassay-guided isolation and identification of antioxidative compounds from the bark of Eugenia polyantha.

Eugenia polyantha bark extracts were found to have potential antioxidative activities. This study is an effort to investigate the antioxidative compounds in E. polyantha. In vitro antioxidatve assay were used as guided tools for the isolation of antioxidative compounds. The methanol-water extracts showed the highest level of free radical-scavenging activity (ED50) = 180 microg mL(-1) and protection from beta-carotene bleaching (8.7 microg mL(-1)). The methanol-water (1:1) extracts exhibited strong DPPH scavenging activity and protection against beta carotene bleaching and was subjected to repeated silica gel column chromatography. The n-butanol, acetone and ethyl acetate solubles exhibited the highest antioxidative activities, derivatization was conducted to the isolated antioxidative compounds prior to identification. Catechin, gallic acid and rutin were isolated from those solubles as active compounds present in the Eugenia polyantha bark.

Determination of chrysene degradation under saline conditions by Fusarium sp. F092, a fungus screened from nature.

Sixty-two rotted wood and soil samples were used to screen for chrysene-degrading fungi. A strain of Fusarium, named F092, was identified as most capable of degrading chrysene. F092 was active under saline and nonsaline conditions, breaking down 48% of the chrysene in 30 d. The percentage of chrysene degraded did not change at 35‰ salinity with pH 8.2 in solid and liquid cultures. The degradation under saline conditions increased about 0.6- and 2.1-fold in cultures with polypeptone and Tween80, and 0.03-fold in agitated cultures. F092 secreted nonligninolytic enzymes named 1,2-dioxygenase and 2,3-dioxygenase. The level of 1,2-dioxygenase activity reached 203.5 U L(-1) at 30 d and that of 2,3-dioxygenase activity, 29.7 U L(-1) at 40 d. The degradation pathway was clarified from the intermediates produced; chrysene 1,2-oxide, chrysene trans-1,2-dihydrodiol, 1-hydroxy 2-naphtoic acid, and catechol. F092 is a potential degrader of chrysene for bioremediation.

Determination of co-metabolism for 1,1,1-trichloro-2,2-bis(4-chlorophenyl) ethane (DDT) degradation with enzymes from Trametes versicolor U97.

Trametes versicolor U97 isolated from nature degraded 73% of the 1,1,1-trichloro-2,2-bis(4-chlorophenyl) ethane (DDT) in a malt extract liquid medium after a 40-d incubation period. This paper presents a kinetic study of microbial growth using the Monod equation. T. versicolor U97 degraded DDT during an exponential growth phase, using glucose as a carbon source for growth. The growth of T. versicolor U97 was not affected by DDT. DDT was degraded by T. versicolor U97 only when the secondary metabolism coincided with the production of several enzymes. Furthermore, modeling of several inhibitors using the partial least squares function in Minitab 15, revealed lignin peroxidase (98.7 U/l) plays a role in the degradation of DDT. T. versicolor U97 produced several metabolites included a single-ring aromatic compound, 4-chlorobenzoic acid.

Isolation of alpha-glucosidase inhibitors produced by an endophytic fungus, Colletotrichum sp. TSC13 from Taxus sumatrana.

Colletotrichum sp. have potential to act as antidiabetic agent, due to its alpha-glucosidase inhibitory. Therefore, the objective of present study was to isolate and identify the bioactive compounds responsible for the alpha-glucosidase inhibitory activity in Colletotrichum sp. TSC13. The methanol extract of TSC13 mycelia, was partitioned with n-hexane, chloroform and ethyl acetate. The n-hexane fraction exhibited the strongest alpha-glucosidase inhibitory activity. Column chromatography of this fraction resulted in 8 sub-fractions (F1-8). Fraction 3 (F3) which showed 71.4 +/- 2.4% inhibition was analysed further. Analysis using GC-MS after methylation of F3 and comparison to spectra databases and confirmation using authentic sample standards showed that F3 had two saturated fatty acid methyl esters, palmitic acid and stearic acid methyl esters and three unsaturated fatty acid methyl esters, oleic acid, linoleic acid and linoleinic acid methyl esters. Unsaturated fatty acids showed higher activity than the saturated fatty acids and the methyl esters form of unsaturated fatty acids showed slightly less active than the free acids. Further analysis using an ethyl acetate extract, it was confirmed that most of the fatty acids were present in the form of free acids. Therefore, it was concluded that the alpha-glucosidase inhibitor compounds in Colletotrichum sp. TSC13 were unsaturated fatty acids. This is the first report that a Colletotrichum sp. from T. sumatrana has alpha-glucosidase inhibitory activity.

Identification of metabolites from phenanthrene oxidation by phenoloxidases and dioxygenases of Polyporus sp. S133.

Phenanthrene degradation by Polyporus sp. S133, a new phenanthrene-degrading strain, was investigated in this work. The analysis of degradation was performed by calculation of the remaining phenanthrene by gas chromatography-mass spectrometry. When cells were grown in phenanthrene culture after 92 h, all but 200 and 250 mg/l of the phenanthrene had been degraded. New metabolic pathways of phenanthrene and a better understanding of the phenoloxidases and dioxygenase mechanism involved in degradation of phenanthrene were explored in this research. The mechanism of degradation was determined through identification of the several metabolites; 9,10-phenanthrenequinone, 2,2'-diphenic acid, salicylic acid, and catechol. 9,10-Oxidation and ring cleavage to give 9,10-phenanthrenequinone is the major fate of phenanthrene in ligninolytic Polyporus sp. S133. The identification of 2,2'-diphenic acid in culture extracts indicates that phenanthrene was initially attacked through dioxigenation at C9 and C10 to give cis-9,10-dihydrodiol. Dehydrogenation of phenanthrene-cis-9,10-dihydrodiol to produce the corresponding diol, followed by ortho-cleavage of the oxygenated ring, produced 2,2'-diphenic acid. Several enzymes (manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase, and 2,3-dioxygenase) produced by Polyporus sp. S133 was detected during the incubation. The highest level of activity was shown at 92 h of culture.

Characterization of phenanthrene degradation by strain polyporus sp. S133.

Polyporus sp. S133, a fungus collected from contaminated soil, was used to degrade phenanthrene, a polycyclic aromatic hydrocarbon, in a mineral salt broth liquid culture. A maximal degradation rate (92%) was obtained when Polyporus sp. S133 was cultured for 30 days with agitation at 120 r/min, as compared to 44% degradation in non-agitated cultures. Furthermore, the degradation was affected by the addition of surfactants. Tween 80 was the most suitable surfactant for the degradation of phenanthrene by Polyporus sp. S133. The degradation rate increased as the amount of Tween 80 added increased. The rate in agitated cultures was about 2 times that in non-agitated cultures. The mechanism of degradation was determined through the identification of metabolites; 9,10-phenanthrenequinone, 2,2'-diphenic acid, phthalic acid, and protocatechuic acid. Several enzymes (manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase and 2,3-dioxygenase) produced by Polyporus sp. S133 were detected during the incubation. The highest level of activity was shown by 1,2-dioxygenase (187.4 U/L) after 20 days of culture.

In vitro antioxidative activities and polyphenol content of Eugenia polyantha Wight grown in Indonesia.

Antioxidative activities of three bark extracts (methanol, methanol-water and water) from Eugenia polyantha Wight grown in Indonesia were evaluated using various in vitro assays; 2, 2-diphenyl-1-picrylhydrazyl radical-scavenging, hydrogen peroxide-scavenging and beta-carotene bleaching assays. From the assays, Eugenia polyantha bark extracts were found to be potential antioxidative activities. The methanol-water extract showed the highest level of free radical-scavenging activity (Effective dose (ED50) = 0.18 mg mL-1) and protection from beta-carotene bleaching (85.7% at 100 microg mL-1). The methanol-water extracts showed the highest total phenolic content (856 mg gallic acid equivalent (GAE)/g and 161 mg catechin equivalent (CE)/g) and total antioxidative capacity (449 mg ascorbic acid equivalent (AAE)/g). Furthermore, it exhibited dose-dependent antioxidative activities. A relationship between total antioxidative capacity and total phenolic content was recognized in the three extracts from Eugenia polyantha bark.

Biodegradation of chrysene, an aromatic hydrocarbon by Polyporus sp. S133 in liquid medium.

Polyporus sp. S133, a fungus collected from contaminated-soil was used to degrade chrysene, a polycyclic aromatic hydrocarbon (PAH) in a mineral salt broth (MSB) liquid culture. Maximal degradation rate of chrysene (65%) was obtained when Polyporus sp. S133 was incubated in the cultures supplemented with polypeptone (10%) for 30 days under agitation of 120 rpm, as compared to just 24% degradation rate in non-agitated culture. Furthermore, the degradation of chrysene was affected by the addition of carbon and nitrogen sources as well as kind of surfactants. The degradation rate was increased with increase in added amount of carbon and nitrogen sources, respectively. The degradation rate in agitated cultures was enhanced about 2 times higher than that in non-agitated cultures. The degradation mechanism of chrysene by Polyporus sp. S133 was determined through identification of several metabolites; chrysenequinone, 1-hydroxy-2-naphthoic acid, phthalic acid, salicylic acid, protocatechuic acid, gentisic acid, and catechol. Several enzymes (manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase and 2,3-dioxygenase) produced by Polyporus sp. S133 were detected during the incubation. The highest enzyme activity was shown by 1,2-dioxygenase (237.5 U l(-1)) after 20 days of incubation.

Maleo- and fumaro-pimaric acids synthesized from Indonesian Pinus merkusii rosin and their sizing properties.

This research is directed at characterizing purified maleopimaric acid (MPA), looking at the amount of MPA and fumaropimaric acid (FPA) made from a large amount of rosin and maleic anhydride (MA) or fumaric acid (FA) with various molar ratios and sources of rosin and evaluating the properties of fortified rosin sizes made from both MPA and FPA. Results showed that identification of the Diels-Alder adduct of abietic acid and maleic anhydride using Mass spectrometry produced a mixture of endo-maleopimaric acid methyl ester with endo-maleopimaric acid tri methyl ester, as indicated by a fragment ion at m/z = 146 with a molecular weight of 414 and fragment ions at m/z = 121, 187, 316 and 386, denoting an endo-maleopimaric acid methyl ester. A fragment ion at m/z = 146 with a molecular weight of 460 and other fragment ions at m/z = 187, 121, 400 and 428 indicated endo-maleopimaric tri methyl ester. Using a large amount of rosin as a raw material to produce MPA, the equation Y = -0.8475 X2 + 10.448X - 9.7125, at a reaction temperature of 200 degrees C is still relevant as it denoted that a molar ratio of 1:6.2 (rosin and MA) is the best. However, the equation Y = -0.46X2 + 5.268X - 4.47 did not apply to FPA. Using a large amount of rosin, an increase in the molar ratio led to an increase in FPA products. In terms of free rosin and pH, the maleo-and fumaro-pimaric rosin sizes have met the requirement of Indonesian national standards for paste rosin size. In terms of free alkali property, the maleo- and fumaro-pimaric rosin sizes were better than the free alkali of the commercial forms.

Biodegradation of phenanthrene by fungi screened from nature.

Microbial degradation of Phenanthrene with several fungi screened from nature was conducted to select fungi for the bioremediation ofPhenanthrene. Thrichoderma sp. S019, a fungus collected from soil, had the highest rate of degradation on the agar medium containing Phenanthrene. Maximal degradation (72%) was obtained when Trichoderma sp. S019 was incubated for 30 days after the addition of 0.1 mM of Phenanthrene to the liquid medium. Furthermore, the degradation of Phenanthrene was affected by the addition of a carbon source, the addition of a nitrogen source and agitation. Also, 1,2-Dioxygenase and 2,3-Dioxygenase were produced by Trichoderma sp. S019 in a liquid medium. These enzymes play an important role in the metabolism of substrates, revealing a high stereoselectivity for initial dioxygenase and enzymatic hydration since the K-region of phenanthrene was the major site of metabolism. Phenanthrene was indeed degraded by Trichoderma sp. S019 because 1-Hydroxy-2-naphthoic acid, Salicyaldehyde, Salicylic acid and Catechol, considered to be the intermediates in the bioremediation of Phenanthrene, were detected among the reaction products.

Reaction of abietic acid with maleic anhydride and fumaric acid and attempts to find the fundamental component of fortified rosin.

Identification of the Diels-Alder adduct of Abietic Acid (AA) and maleic anhydride (MA) or Fumaric Acid (FA) using a Shimadzu QP 5050A Gas chromatograph Mass spectrometer revealed that AA and MA produced endo-maleopimaric acid (MPA) and endo-maleopimaric acid tricarboxylic acid. A reaction product of abietic acid and fumaric acid generated three peaks identified based on their mass spectra as fumaropimaric acid (FPA), FPA adducts and endo-MPA. To maximize the reaction between AA and MA or FA, molar ratio (AA and MA or FA), reaction time and reaction temperature were investigated. The best Diels-Alder reaction between abietic and maleic anhydride was at 125 degrees C for 1 h with a molar ratio of AA to MA of 1: 2. The best Diels-Alder reaction between AA and FA was at 200 degrees C for 1 h with a molar ratio of AA and FA of 1: 2.