Effect of T3 hormone on neural differentiation of human adipose derived stem cells.

Human adult stem cells, which are capable of self-renewal and differentiation into other cell types, can be isolated from various tissues. There are no ethical and rejection problems as in the case of embryonic stem cells, so they are a promising source for cell therapy. The human body contains a great amount of adipose tissue that contains high numbers of mesenchymal stem cells. Human adipose-derived stem cells (hADSCs) could be easily induced to form neuron-like cells, and because of its availability and abundance, we can use it for clinical cell therapy. On the other hand, T3 hormone as a known neurotropic factor has important impressions on the nervous system. The aim of this study was to explore the effects of T3 treatment on neural differentiation of hADSCs. ADSCs were harvested from human adipose tissue, after neurosphere formation, and during final differentiation, treatment with T3 was performed. Immunocytochemistry, real-time RT-PCR, Western blotting techniques were used for detection of nestin, MAP2, and GFAP markers in order to confirm the effects of T3 on neural differentiation of hADSCs. Our results showed an increase in the number of glial cells but reduction in neuronal cells number fallowing T3 treatment.

Insulin and leptin levels in overweight and normal-weight Iranian adolescents: The CASPIAN-III study.

BACKGROUND: In this study, we aim to compare insulin and leptin levels in adolescents with or without excess weight and in those with or without abdominal obesity. MATERIALS AND METHODS: This case-control study was conducted among 486 samples. We randomly selected 243 overweight and an equal number of normal-weight adolescents from among participants of the third survey of a national surveillance program entitled Childhood and Adolescence Surveillance and PreventIon of Adult Non-communicable diseases study. Serum insulin and leptin were compared between two groups and their correlation was determined with other variables. RESULTS: The mean age and body mass index (BMI) of participants were 14.10 ± 2.82 years and 22.12 ± 6.49 kg/m(2), respectively. Leptin and insulin levels were higher in overweight than in normal-weight adolescents (P < 0.05). Leptin level was higher in children with abdominal obesity than in their other counterparts (P < 0.001). Leptin level was correlated with age, fasting blood glucose, BMI, and insulin level. CONCLUSION: Insulin and leptin levels were higher among overweight and obese children, which may reflect insulin and leptin-resistance. Given the complications of excess weight from early life, prevention and controlling childhood obesity should be considered as a health priority.

Comparing brain-derived neurotrophic factor and ciliary neurotrophic factor secretion of induced neurotrophic factor secreting cells from human adipose and bone marrow-derived stem cells.

Adipose derived stem cells (ADSCs) and bone marrow stem cells (BMSCs) may be equally beneficial in treating neurodegenerative diseases. However, ADSCs have practical advantages. In this study, we aimed to induce neurotrophic factors secreting cells in human ADSCs. Then, we compared the level of brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) secretion in neurotrophic factors secreting cells from human adipose and bone marrow-derived stem cells. Isolated human ADSCs and BMSCs were induced to neurotrophic factor (NTF)-secreting cells. The levels of expression and secretion of BDNF and CTNF of induced cells were assessed using immunocytochemical, Real-Time polymerase chain reaction, and enzyme linked immunosorbent assay (ELISA). The level of BDNF significantly increased in both the induced mesenchymal stem cells (MSCs) relative to ADSCs and the BMSCs (P < 0.01). Moreover, ELISA analysis showed that the release of BDNF in the induced BMSCs was almost twofold more than the induced ADSCs. Overall, NTF-secreting factor cells derived BMSCs and ADSCs could secret a range of different growth factors. Therefore, the variation in neurotrophic factors of different induced MSC populations suggest the possible beneficial effect of each specific kind of neurotrophic factor secreting cells for the treatment of a particular neurodegenerative disease.

Assessment of cagE and babA mRNA expression during morphological conversion of Helicobacter pylori from spiral to coccoid.

Helicobacter pylori (H. Pylori) is an actively dividing spiral bacterium that changes to coccoid morphology under stressful environments. The infectivity of the coccoids is still controversial. The aim of this study was to determine the viability and expression of two important virulence genes (babA and cagE), in antibiotic-induced coccoid forms. Three strains of H. pylori, the standard 26695 and two clinical isolates (p1, p2) were converted to coccoid form by amoxicillin. Coccoids were identified according to Gram-staining and microscopic morphology. The viability of the cells was analyzed by flow cytometry. The expression of cagE and babA in coccoid forms were evaluated and compared to the spirals by quantitative PCR assay. The coccoid forms were developed after 72 h exposure of H. pylori to ½ MIC of amoxicillin, and the conversion form was completed (100 %) at 144 h in all of three isolates. Flow cytometry analyses showed that the majority of the induced coccoids (90-99.9 %) were viable. Expression of cagE and babA was seen in coccoids; however, in lower rate (cagE, ~3-fold and babA, ~10-fold) than these in spiral forms. Coccoid forms of two clinical isolates significantly expressed higher rate of cagE and babA than standard 26695 strain (P = 0.01). These results suggest that the induced coccoid form of H. pylori is not a passive entity but can actively infect the human by expression of the virulence genes for long time in stomach and probably play a role in chronic and severe disease.

Rapid and accurate identification of Mycobacterium tuberculosis complex and common non-tuberculous mycobacteria by multiplex real-time PCR targeting different housekeeping genes.

Rapid and accurate identification of mycobacteria isolates from primary culture is important due to timely and appropriate antibiotic therapy. Conventional methods for identification of Mycobacterium species based on biochemical tests needs several weeks and may remain inconclusive. In this study, a novel multiplex real-time PCR was developed for rapid identification of Mycobacterium genus, Mycobacterium tuberculosis complex (MTC) and the most common non-tuberculosis mycobacteria species including M. abscessus, M. fortuitum, M. avium complex, M. kansasii, and the M. gordonae in three reaction tubes but under same PCR condition. Genetic targets for primer designing included the 16S rDNA gene, the dnaJ gene, the gyrB gene and internal transcribed spacer (ITS). Multiplex real-time PCR was setup with reference Mycobacterium strains and was subsequently tested with 66 clinical isolates. Results of multiplex real-time PCR were analyzed with melting curves and melting temperature (T (m)) of Mycobacterium genus, MTC, and each of non-tuberculosis Mycobacterium species were determined. Multiplex real-time PCR results were compared with amplification and sequencing of 16S-23S rDNA ITS for identification of Mycobacterium species. Sensitivity and specificity of designed primers were each 100 % for MTC, M. abscessus, M. fortuitum, M. avium complex, M. kansasii, and M. gordonae. Sensitivity and specificity of designed primer for genus Mycobacterium was 96 and 100 %, respectively. According to the obtained results, we conclude that this multiplex real-time PCR with melting curve analysis and these novel primers can be used for rapid and accurate identification of genus Mycobacterium, MTC, and the most common non-tuberculosis Mycobacterium species.

Candida galli strain PGO6: a novel isolated yeast strain capable of transformation of isoeugenol into vanillin and vanillic acid.

Candida galli strain PGO6 isolated from oil-contaminated water is the first isolated yeast strain which is capable to form vanillin and vanillic acid during isoeugenol biotransformation. The products were confirmed by thin-layer chromatography (TLC), changes in the UV absorption pattern and high-performance liquid chromatography (HPLC). The phenotypic and physiochemical characteristics as well as molecular phylogenetic analysis based on amplification the ITS1-5.8S-ITS2 rDNA regions indicated the isolated strain PGO6 was identified as C. galli (GenBank accession number HM641231). Resting cells of C. galli PGO6 from the late-exponential of growth phase were used as biocatalysts for the biotransformation of isoeugenol. The optimal molar conversion of vanillin (48%) and vanillic acid (19%) was obtained after a 30 h incubation using 0.1% (v/v) of isoeugenol and 6 mg of dry weight of cells per ml without further optimization. Under these conditions, the total amount of vanillin and vanillic acid was 583 mg l(-1). Further biotransformation was carried out using 0.5% (v/v) of isoeugenol under the resting cells conditions, yielding a vanillin concentration of 1.12 g l(-1) (molar yield 25.7%) after 60 h incubation. This study brings the first evidence for biotransformation of isoeugenol to vanillin and vanillic acid by a yeast strain.

Torque teno mini virus infection and multiple sclerosis.

Multiple sclerosis (MS) is a disease of young adults which is characterized by autoimmune demyelination of the central nervous system. Interaction of genetics and environmental factors are required to cause MS. Among the proposed environmental factors for MS, viral infections are thought to play a role in the pathogenesis of the disease. Torque teno mini virus (TTMV), which has recently been shown to infect humans, is a member of circoviridae, and has a circular DNA with 2860 nucleotides. Since there are a few data about the pathogenicity of this virus, this study sought to investigate the presence of TTMV in sera from MS patients and healthy individuals. We studied 149 serum samples from MS patients and 150 sera of healthy individuals. Serum DNA was extracted using phenol-chloroform and was subjected to nested polymerase chain reaction. TTMV-DNA was detected in 24 (16%) sera of the healthy blood donors and in 21 (14.1%) samples of the MS patients, where the difference did not reach significance (p > .05). The result of this study could not establish an association between TTMV infection and MS.

Evaluation of the leptin receptor in human spermatozoa.

BACKGROUND: Leptin, a 167 amino acid peptide hormone, profoundly effects reproduction exerting its biological effects via interaction with the leptin receptor (ObR) which is widely expressed on peripheral tissues. In this study, we have attempted to assess leptin receptor expression in the spermatozoa of fertile males and those diagnosed with male factor infertility; both at the mRNA or protein levels. METHODS: Semen samples were collected from fertile males and individuals with male factor infertility. In order to evaluate leptin receptor expression several techniques were utilized, including: reverse transcriptase-polymerase chain reaction (RT-PCR), immunostaining, flow cytometry, and western blotting. Mononuclear cells isolated from volunteers' peripheral blood were used as positive controls for leptin receptor expression. RESULTS: leptin receptor was noted on mononuclear cells but we were unable to detect this receptor on spermatozoa at the protein level. Leptin receptor expression was detected on peripheral blood mononuclear cells (PBMCs) as positive controls; however it was not detectable on the spermatozoa of both groups by immunofluorescence microscopy or flow cytometry. Furthermore, positive expression of the ObR long isoform as assessed by RT-PCR was observed in the sperm of only four cases, whereas expression of beta-Actin, a house keeping gene, and HspA2, a testis specific gene, was present in all cases. CONCLUSION: The long isoform of leptin receptor may not be present on human sperm. Species difference may be accounted for diverse reproductive physiology which depends on metabolic requirement. Leptin receptor expression at the mRNA level in some individuals may be related to contamination by other cells in semen.

Effect of plant oils upon lipase and citric acid production in Yarrowia lipolytica yeast.

The nonconventional yeast Yarrowia lipolytica degrades very efficiently hydrophobic substrates to produce organic acids, single-cell oil, lipases, and so forth. The aim of this study was to investigate the biochemical behavior and simultaneous production of valuable metabolites such as lipase, citric acid (CA), and single-cell protein (SCP) by Yarrowia lipolytica DSM 3286 grown on various plant oils as sole carbon source. Among tested plant oils, olive oil proved to be the best medium for lipase and CA production. The Y. lipolytica DSM 3286 produced 34.6 +/- 0.1 U/mL of lipase and also CA and SCP as by-product on olive oil medium supplemented with yeast extract. Urea, as organic nitrogen, was the best nitrogen source for CA production. The results of this study suggest that the two biotechnologically valuable products, lipase and CA, could be produced simultaneously by this strain using renewable low-cost substrates such as plant oils in one procedure.

Isolation of a novel mutant strain of Saccharomyces cerevisiae by an ethyl methane sulfonate-induced mutagenesis approach as a high producer of bioethanol.

In order to obtain mutant strains showing higher bioethanol production than wild-type strains, a commercial Saccharomyces cerevisiae type was subjected to mutagenesis using ethyl methane sulfonate (EMS). After adding EMS to a shaken yeast suspension, the viability of yeast cells was assessed by diluted sample inoculation to solid yeast-extract peptone glucose (YEPG) medium at 15-min intervals. At 45 min, the viability of yeast cells was estimated to be about 40%. Mutagenized cells were recovered from YEPG broth after incubation at 30 degrees C for 18 h. After this period, EMS-treated yeast cells were grown on solid aerobic low-peptone (ALP) medium containing 2-12% (v/v) ethanol. All plates were incubated at 30 degrees C for 2-6 d in order to form colonies. The mutant strains that tolerated high concentrations of ethanol were selected for bioethanol production in microfuge tubes containing fermentation medium. Formation of bioethanol in small tubes was detected by the distillation-colorimetric method. In addition, trehalose content and invertase activity were determined in each mutant strain. Among many isolated mutant strains, there were six isolated colonies that grew on ALP medium supplemented with 10% (v/v) ethanol and one of them produced bioethanol 17.3% more than the wild type.

Identification of a monoclonal antibody against the leptin receptor that acts as an antagonist and blocks human monocyte and T cell activation.

Nutritional status has a major impact on the immune response and this is in part mediated by leptin, a pro-inflammatory cytokine. Preliminary data suggest that antagonism of leptin may offer a therapeutic approach for the treatment of some inflammatory disorders. We have tested monoclonal antibodies (mAbs) to the human leptin receptor (ObR) for antagonist activity using a leptin signalling bioassay. We identified a mAb, 9F8, which demonstrated dose-dependent antagonist activity in the leptin bioassay. Specificity of the mAb for ObR was confirmed using a plate binding assay. The 9F8 mAb displaced leptin binding to human ObR and enzymatically generated Fab fragments of 9F8 retained antagonist activity. Therefore the Fab fragment of 9F8 was cloned and recombinant 9F8 Fab (rFab) was purified from E. coli periplasmic fraction using a C-terminal His tag. Purified 9F8 rFab bound to human ObR and exhibited leptin antagonist activity. In vitro studies demonstrated that the 9F8 mAb inhibited leptin induced TNF-alpha production from human monocytes and anti-CD3 mAb induced proliferation of human T cells in PBMC culture. In conclusion, this study has identified a mAb to the human leptin receptor which inhibits leptin signalling and acts as a leptin antagonist in vitro.

Leptin serves as angiogenic/mitogenic factor in melanoma tumor growth.

BACKGROUND: Tumor development is angiogenesis dependent. There is evidence that leptin contributes to tumor growth. However, all the mechanisms by which leptin does this has not been clearly established. The objective of the present study was to test the hypothesis that leptin enhances melanoma tumor growth through inducing angiogenesis and cell proliferation. MATERIALS AND METHODS: We injected 2 × 10(6) B16F10 melanoma cells subcutaneously to 32 C57BL6 mice. The mice were randomly divided into four groups of eight animals, on day 8. Two groups received twice daily intraperitoneal (i.p.) injections of either phosphate buffered saline or recombinant murine leptin (1 µg/g initial body weight). Two groups received i.p. injections of either 9F8 an anti leptin receptor antibody or the control mouse IgG at 50 µg/injection every 3 consecutive days. By the end of the 2(nd) week, the animals were euthanized and blood samples and tumors were analyzed. Angiogenesis and proliferation were assessed by immunohistochemical staining for CD31 and Ki-67 respectively. RESULTS: Tumors size, capillary density, plasma levels of vascular endothelial growth factor, and the number of Ki-67-positive stained cells were significantly more in the leptin than 9F8 and both control groups (P < 0.05). CONCLUSION: Taken together, our findings reinforce the idea that leptin acts as an angiogenic and mitogenic factor to promote melanoma growth.

Nanobiocomposite of poly(lactide-co-glycolide)/chitosan electrospun scaffold can promote proliferation and transdifferentiation of Schwann-like cells from human adipose-derived stem cells.

The transdifferentiation of human adipose-derived stem cells (ADSCs) into Schwann-like cells on biocomposite scaffolds may be a critical issue in nerve regeneration medicine. In this study, tissue-engineered scaffold with chitosan (CS) nanopowders and poly(lactide-co-glycolide) (PLGA) was investigated for its potential Schwann cells (SCs) transdifferentiation. The differentiation of human ADSCs into S-like cells was induced with different CS content and direction of nanofibers on PLGA/CS scaffolds. Cell morphology and proliferation of differentiated cells were investigated by scanning electron microscopy and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay respectively. For assessment efficiency of transdifferentiation, the expression of SC markers (glial fibrillary acidic protein and S100), and myelinogenic marker (myelin basic protein) was investigated in different nanochitosan content and direction of nanofibers scaffolds, using immunocytochemistry technique. The nanochitosan can significantly promote cell proliferation of differentiated cells (p < 0.05). The mean percentage of S-like cells on greater CS content nanofibers scaffold was significantly higher than others (p < 0.05). In addition, the align orientation of nanofibers in scaffolds guided the differentiation of ADSCs toward myelinating S-like cells on the constructs. Overall, we found that high CS content and aligned-orientation of nanofibers in biocomposite scaffold (70/30A) can promote differentiation and myelinogenic capacity of S-like cells induced from human ADSCs.

Cell Attachment and Proliferation of Human Adipose-Derived Stem Cells on PLGA/Chitosan Electrospun Nano-Biocomposite.

OBJECTIVE: In this study, nano-biocomposite composed of poly (lactide-co-glycolide) (PLGA) and chitosan (CS) were electrospun through a single nozzle by dispersing the CS nano-powders in PLGA solution. The cellular behavior of human adipose derived stem cells (h-ADSCs) on random and aligned scaffolds was then evaluated. MATERIALS AND METHODS: In this experimental study, the PLGA/CS scaffolds were prepared at the different ratios of 90/10, 80/20, and 70/30 (w/w) %. Morphology, cell adhesion and prolif- eration rate of h-ADSCs on the scaffolds were assessed using scanning electron microscope (SEM), 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay and trypan blue staining respectively. RESULTS: H-ADSCs seeded on the matrices indicated that the PLGA/CS composite matrix with aligned nanofibres and higher content of CS nano-powders gave significantly better performance than others in terms of cell adhesion and proliferation rate (P<0.05). CONCLUSION: We found that CS enhanced cell adhesion and proliferation rate, and aligned nanofibers guided cell growth along the longitudinal axis of the nanofibers, which would provide a beneficial approach for tissue engineering.

Morphological and Bactericidal Effects of Different Antibiotics on Helicobacter pylori.

BACKGROUND: Helicobacter pylori (H. pylori) is a spiral Gram negative bacteria that can transform to the coccoid form in adverse conditions. OBJECTIVES: The aim of this study was to determine the in vitro morphological and bactericidal effects of metronidazole, amoxicillin and clarithromycin on H. pylori. MATERIALS AND METHODS: The standard strain 26695 of H. pylori was cultured on Brucella agar (BA) and the minimum inhibitory concentrations (MICs) of three antibiotics were determined by E-test method. The bacteria were exposed to antibiotics at 1/2 MIC, MIC and 2X MIC concentrations in Brucella broth (BB). Induced coccoid forms were confirmed by Gram staining and light microscopy. The viability of cells as well as the susceptibility of viable coccoids to antibiotics were examined using the flow cytometry method. RESULTS: All of the three antibiotics at sub-MIC induced coccoid forms. The highest rates of coccoids (> 90%) were induced at 0.008 µg/mL concentration (1/2 MIC) of amoxicillin, 72 hours postexposure. Metronidazole and clarithromycin with 1/2 MIC (0.5 and 0.125 µg/mL respectively) induced lower rates of coccoid forms (60% and 40% respectively). Potent bactericidal effects on coccoids were observed with Metronidazole at 2X MIC and clarithromycin at MIC (0.25 µg/mL) (80 - 90%). Amoxicillin with MIC and 2X MIC had no bactericidal effect on coccoid forms. CONCLUSIONS: Despite the good in vitro bactericidal effect of amoxicillin on spiral forms of H. pylori, this antibiotic has little effect on induced coccoids that may develop after the inappropriate in vivo antibacterial treatment. Hence, for successful therapy, it is essential not only to eradicate the spiral forms, but to eliminate the viable coccoids.

Preparation and Cytotoxic Evaluation of Magnetite (Fe3O4) Nanoparticles on Breast Cancer Cells and its Combinatory Effects with Doxorubicin used in Hyperthermia.

BACKGROUND: Magnetic nanoparticles in a variable magnetic field are able to produce heat. This heat (42-45°C) has more selective effect on fast dividing cancer cells than normal tissues. METHODS: In this work magnetite nanoparticles have been prepared via co-precipitation and phase identification was performed by powder x-ray diffraction (XRD). Magnetic parameters of the prepared nanoparticles were measured by a Vibrating Sample Magnetometer (VSM). A sensitive thermometer has been used to measure the increase of temperature in the presence of an alternating magnetic field. To evaluate the cytotoxicity of nanoparticles, the suspended magnetite nanoparticles in liquid paraffin, doxorubicin and a mixture of both were added to the MDA-MB-468 cells in separate 15 ml tubes and left either in the RT or in the magnetic field for 30 min. Cell survival was measured by trypan blue exclusion assay and flow cytometer. Particle size distribution of the nanoparticles was homogeneous with a mean particles size of 10 nm. A 15°C temperature increase was achieved in presence of an AC magnetic field after 15 min irradiation. RESULTS: Biological results showed that magnetite nanoparticles alone were not cytotoxic at RT, while in the alternative magnetic filed more than 50% of cells were dead. Doxorubicin alone was not cytotoxic during 30 min, but in combination with magnetite more than 80% of the cells were killed. CONCLUSION: It could be concluded that doxorubicin and magnetite nanoparticles in an AC magnetic field had combinatory effects against cells.

Conversion of isoeugenol to vanillin by Psychrobacter sp. strain CSW4.

To screen strains of halotolerant or halophile bacteria which are able to convert isoeugenol to vanillin, 36 different strains of bacteria isolated from the salty environments in Iran were investigated. During growth on isoeugenol, a moderately halotolerant Gram-negative coccobacil showed capability of converting isoeugenol to vanillin. Based on morphological, physiological, and phylogenetic studies, strain CSW4 was classified as a bacterium belonging to the genus Psychrobacter. The bioconversion products were confirmed by thin-layer chromatography, high-performance liquid chromatography, and spectral data obtained from UV/Vis spectroscopy, FTIR, and mass-spectroscopy. Using growing cells, vanillin reached its maximum level of 88.18 mg L(-1) after 24 h of reaction time in the presence of 1 g L(-1) isoeugenol, resulting in a molar yield of 10.2%. The use of resting cells led to the optimal yield of vanillin (16.4%) which was obtained after 18-h reaction using 1 g L(-1) isoeugenol and 3.1 g of dry weight of cells per liter harvested at the end of the exponential growth phase. To improve vanillin yield, the effect of substrate concentration on vanillin production under resting cells conditions was also investigated. Using 10 g L(-1) isoeugenol, the maximal vanillin concentration (1.28 g L(-1)) was achieved after a 48-h reaction, without further optimization. The present study brings the first evidence for biotransformation of isoeugenol to vanillin in the genus Psychrobacter.

Effect of additives on freeze-drying and storage of Yarrowia lipolytica lipase.

The extracellular lipase of Yarrowia lipolytica presents numerous potentialities for biotechnological applications. This work describes the development and storage of powders obtained from supernatants containing Y. lipolytica lipase by freeze-drying as downstream process that is important in obtaining a stable lipase powder with high enzymatic activity. Lipase was produced by Y. lipolytica U6 mutant strain in 20-L bioreactor. Non-concentrated cell-free culture supernatant samples were supplemented with different concentrations (0.5-1 %) of maltodextrin and glycerol as additives to freeze-drying. Effects of additives, temperature, pH, and storage time on lipase powders were determined. After addition of additives, freeze-drying yield increased 3.5-fold compared to supernatant without additive. Maltodextrin with 0.5 % concentration gave the best protection of lipase during dehydration treatment and its freeze-drying yield (77 %) is better than other formulations. Lipase powders were stored at 4 and 25 °C for 46 weeks without loss of lipase activity. A common impediment to the production of commercial enzyme is their low-stability aqueous solutions. The present study shows that freeze-dried lipase powders of Y. lipolytica have good stability for storage and various applications.

Use of Growing Cells of Pseudomonas aeruginosa for Synthesis of the Natural Vanillin via Conversion of Isoeugenol.

The great demand of people for consumption of natural additives resulted in producing natural vanillin. There are plant sources and chemical procedures for vanillin production but microbial bioconversions are being sought as a suitable alternative. In the present work, the ability to produce vanillin from isoeugenol was screened using growing cultures of various bacteria. Among the 56 strains of bacteria isolated from the soil environments of Iran, a Gram-negative rod designated as strain ISPC2 showed the capability of promoting the formation of high amounts of vanillin when grown in the presence of isoeugenol. On the basis of morphological and physiochemical characteristics and 16S ribosomal ribonucleic acid (rRNA) gene sequence analysis, the isolate was identified as Pseudomonas aeruginosa ISPC2. Vanillin formation was analyzed by GC/FID. In the presence of isoeugenol, a growing culture of P. aeruginosa ISPC2 produced 1.62 g/L vanillin (molar yield of 17.3%) after a 72 h reaction at 30°C and 200 rpm. This proposed procedure is an alternative approach to obtain vanillin in an environmentally friendly way. Further studies for standardization and optimization for higher yield of vanillin production, needs to be investigated.

Pseudomonas resinovorans SPR1, a newly isolated strain with potential of transforming eugenol to vanillin and vanillic acid.

In this study a novel strain was isolated with the capability to grow on eugenol as a source of carbon and energy. This strain was identified as Pseudomonas resinovorans (GenBank accession no. HQ198585) based on phenotypic characterization and phylogenetic analysis of 16S rDNA gene. The intermediates coniferyl alcohol, coniferyl aldehyde, ferulic acid, vanillin and vanillic acid were detected in the culture supernatant during eugenol biotransformation with this strain. The products were confirmed by thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and spectral data achieved from UV-vis, FTIR and mass spectroscopy. Using eugenol as substrate and resting cells of P. resinovorans SPR1, which were harvested at the end of the exponential growth phase, without further optimization 0.24 g/L vanillin (molar yield of 10%) and 1.1g/L vanillic acid (molar yield of 44%) were produced after 30 h and 60 h biotransformation, respectively. The current work gives the first evidence for the eugenol biotransformation by P. resinovorans.