Pairoh Pinphanichakarn

Diversity and Activity of PAH-Degrading Bacteria in the Phyllosphere of Ornamental Plants

Phyllosphere bacteria on ornamental plants were characterized based on their diversity and activity towards the removal of polycyclic aromatic hydrocarbons (PAHs), the major air pollutants in urban area. The amounts of PAH-degrading bacteria were about 1–10% of the total heterotrophic phyllosphere populations and consisted of diverse bacterial species such as Acinetobacter, Pseudomonas, Pseudoxanthomonas, Mycobacterium, and uncultured bacteria. Bacterial community structures analyzed by polymerase chain reaction–denaturing gradient gel electrophoresis from each plant species showed distinct band patterns. The uniqueness of these phyllosphere bacterial communities was partly due to the variation in leaf morphology and chemical properties of ornamental plants. The PAH degradation activity of these bacteria was monitored in gas-tight systems containing sterilized or unsterilized leaves. The results indicated that phyllosphere bacteria on unsterilized leaves were able to enhance the activity of leaves for phenanthrene removal. When compared between plant species, phenanthrene removal efficiency corresponded to the size of phenanthrene-degrading bacteria. In addition, phyllosphere bacteria on Wrightia religiosa were able to reduce other PAHs such as acenaphthylene, acenaphthene, and fluorine in 60-ml glass vials and in a 14-l glass chamber. Thus, phyllosphere bacteria on ornamental plants may play an important role in natural attenuation of airborne PAHs in urban areas.

Novel intermediates of acenaphthylene degradation by Rhizobium sp. strain CU-A1 : Evidence for naphthalene-1,8-dicarboxylic acid metabolism

ABSTRACT The acenaphthylene-degrading bacterium Rhizobium sp. strain CU-A1 was isolated from petroleum-contaminated soil in Thailand. This strain was able to degrade 600 mg/liter acenaphthylene completely within three days. To elucidate the pathway for degradation of acenaphthylene, strain CU-A1 was mutagenized by transposon Tn5 in order to obtain mutant strains deficient in acenaphthylene degradation. Metabolites produced from Tn5-induced mutant strains B1, B5, and A53 were purified by thin-layer chromatography and silica gel column chromatography and characterized by mass spectrometry. The results suggested that this strain cleaved the fused five-membered ring of acenaphthylene to form naphthalene-1,8-dicarboxylic acid via acenaphthenequinone. One carboxyl group of naphthalene-1,8-dicarboxylic acid was removed to form 1-naphthoic acid which was transformed into salicylic acid before metabolization to gentisic acid. This work is the first report of complete acenaphthylene degradation by a bacterial strain.

Production and Characterization of a Biosurfactant from Cyberlindnera samutprakarnensis JP52 T

Cyberlindnera samutprakarnensis JP52T, isolated from cosmetic industrial wastes in Thailand, was found to be an efficient biosurfactant-producing yeast when cultured in a medium containing (2% (w/v) glucose and 2% (v/v) palm oil at 30 °C, 200 rpm for 7 d. The crude biosurfactant had the ability to reduce the surface tension from 55.7 to 30.9 mN/m at 25 °C with a critical micelle concentration (CMC) of 0.046%. Physicochemical analysis of the crude biosurfactant revealed that it had wide ranges of optimum pH and pH stability at 6–9 and 3–10 respectively. It was also thermostable and retained 80% activity even after heat treatment, and it tolerated NaCl at 1.0–10%. Furthermore, it effectively emulsified various vegetable oils with an E24 value of over 80%. A partially purified biosurfactant fraction was analyzed for its structure by MALDI-TOF MS and NMR. This revealed that the biosurfactant mainly contained sophorolipids in C18-(MW 574) and C16-diaceltylated (MW 662) forms.

Process development and simulation of glucose isomerase production from birchwood xylan by a Streptomycetes fusant

Abstract A process for glucose isomerase production from an unutilized substrate, xylan, was investigated, employing strain development, investigation of the culture conditions, and an expedient simulation method. Streptomyces sp. strain D3 obtained through protoplast fusion between Streptomyces cyaneus 190-1 and Streptomyces greiseoruber 42-9 was able to produce a substantial amount of intracellular glucose isomerase from xylan. The induction of glucose isomerase synthesis by xylose was confirmed in shaken flask cultivation. In fermentor cultivation, an initial concentration of 8 g l −1 xylan yielded glucose isomerase, 1100 U l −1 . Taking into account the experimental data, a simplified model for the simulation of glucose isomerase production from xylan substrate was constructed based on a statistical regression method. The data selection for the estimation of the specific rates in the regression model was carried out based on the culture time and statistical distance measurement, such as Euclidean, Mahalanobis and modified distances. The results of the simulation of the growth and enzyme production using the model were close to the experimental results, which indicates that the statistical method can be extended to estimate the time courses of process of glucose isomerase production from xylan reasonably well.

Glucose(xylose) isomerase production by Streptomyces sp. CH7 grown on agricultural residues

Streptomyces sp. CH7 was found to efficiently produce glucose(xylose) isomerase when grown on either xylan or agricultural residues. This strain produced a glucose(xylose) isomerase activity of roughly 1.8 U/mg of protein when it was grown in medium containing 1% xylose as a carbon source. Maximal enzymatic activities of about 5 and 3 U/mg were obtained when 1% xylan and 2.5% corn husks were used, respectively. The enzyme was purified from a mycelial extract to 16-fold purity with only two consecutive column chromatography steps using Macro-prep DEAE and Sephacryl-300, respectively. The approximate molecular weight of the purified enzyme is 170 kDa, and it has four identical subunits of 43.6 kDa as estimated by SDS-PAGE. Its Km values for glucose and xylose were found to be 258.96 and 82.77 mM, respectively, and its Vmax values are 32.42 and 63.64 μM/min/mg, respectively. The purified enzyme is optimally active at 85oC and pH 7.0. It is stable at pH 5.5-8.5 and at temperatures up to 60oC after 30 min. These findings indicate that glucose(xylose) isomerase from Streptomyces sp. CH7 has the potential for industrial applications, especially for high-fructose syrup production and bioethanol fermentation from hemicellulosic hydrolysates by Saccharomyces cerevisiae.

An Ingenious Device for Better Prediction of Bio-Processes Using a Multiple Regression Model : A case Study on Glucose Isomerase Production

An ingenious device in the use of a multiple regression analysis model for better prediction of state variable changes in bio-processes is proposed. A correction factor was introduced in the regression equations for the estimation of specific rate parameters taking into account the data distribution resulting in the improvement of the prediction of glucose isomerase production by a Streptomyces fusant during a batch cultivation with periodical monitoring.