Characterization of medium chain length (R)-3-hydroxycarboxylic acids produced by Streptomyces sp. JM3 and the evaluation of their antimicrobial properties.

(R)-3-Hydroxycarboxylic acids, chiral enantiomers of bacterial polyhydroxyalkanoates (PHA), may be valuable synthons for the production of numerous industrial materials such as ß-lactams, fungicides, flavors, pheromones and vitamins. In this study, (R)-3-hydroxycarboxylic acid [(R)-3HAs)] synthons were produced by Streptomyces sp. JM3 (JN166713) under batch fermentation. Initial confirmation of PHA production was achieved by matrix assisted laser desorption ionization-time of flight mass spectroscopy and gas chromatography/mass spectroscopy (GC/MS). Subsequently, (R)-3HAs were produced by in vivo depolymerization and the monomers were separated using acid precipitation and anion exchange chromatography. The (R)-3HAs were identified by GC/MS as 3-trimethylsiloxy esters of decanoic, octanoic and butanoic acids. This was further supported by (13)C nuclear magnetic resonance spectrometry. The (R)-3HAs exhibited antimicrobial activity against Escherichia coli O157:H7, Listeria monocytogenes (ATCC 7644) and Salmonella typhimurium (ATCC 14028) with minimum inhibitory concentration ranging from 12.5 to 25 mg ml(-1). However, the minimum bactericidal concentration data suggest that the (R)-3HAs may be bactericidal for E. coli O157:H7 and bacteriostatic for S. typhimurium and L. monocytogenes. Furthermore, the major purified synthon was shown to minimize the invasion of fibroblasts by S. typhimurium (ATCC 14028) [p < 0.05], using the MTT assay [(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)].

Influence of iron-chelated growth conditions on outer membrane protein production and virulence of Vibrio tubiashii.

Growth of two Vibrio tubiashii strains under iron-chelated conditions resulted in the production of a hydroxymate-like siderophore, and expression of outer membrane proteins with homologies to proteins in Vibrio cholerae and Vibrio vulnificus which were not seen in cells grown under non-chelated growth conditions. PCR analysis using primers based on Listonella anguillarum's ferric uptake Repressor protein (fur) gene detected a 316 bp fur gene homolog which also had sequence homology to the fur genes of V. cholerae and V. vulnificus. V. tubiashii cultured under iron-chelated growth conditions induced a greater fluid accumulation (FA) response in suckling mice than cells which were cultured under iron non-chelated growth conditions. Our observations that V. tubiashii possesses a fur-like gene homolog and expresses unique OMPs, a hydroxymate-like siderophore, and produces an increased fluid accumulation response in suckling mice when grown under iron-chelated condition support previous findings that V. tubiashii may have the essential components for the survival and establishment of infections and this report represents the first observations of competent iron acquisition system in V. tubiashii which is similar to those produced by other marine vibrios, many of which are pathogenic for humans.

Biosynthesis and characterization of copolymer poly(3HB-co-3HV) from saponified Jatropha curcas oil by Pseudomonas oleovorans.

Polyhydroxyalkanoates (PHAs) are naturally occurring biodegradable polymers with promising application in the formulation of plastic materials. PHAs are produced by numerous bacteria as energy/carbon storage materials from various substrates, including sugars and plant oils. Since these substrates compete as food sources, their use as raw material for industrial-scale production of PHA is limited. Therefore, efforts have been focused on seeking alternative sources for bacterial production of PHA. One substrate that seems to have great potential is the seed oil of Jatropha curcas plant. Among other favorable properties, J. curcas seed oil is non-edible, widely available, and can be cheaply produced. In this study, Pseudomonas oleovorans (ATCC 29347) was grown in a mineral salt medium supplemented with saponified J. curcas seed oil as the only carbon source under batch fermentation. Optimum PHA yield of 26.06% cell dry weight was achieved after 72 h. The PHA had a melting point (T(m)) between 150 and 160 degrees C. Results of polymer analyses by gas chromatography/mass spectrometry (GC/MS) identified only the methyl 3-hydroxybutanoate monomeric unit. However, electrospray ionization-time of flight mass spectroscopy (ESI-TOF MS) confirmed that the PHA was a copolymer with the characteristic HB/HV peaks at m/z 1155.49 (HB) and 1,169, 1,184-1,194 (HV). The data were further supported by 1H and 13C NMR analysis. Polymer analysis by gel permeation chromatography (GPC) indicated a peak molecular weight (MP) of 179,797, molecular weight (M(W)) of 166,838, weight number average mass (M(n)) of 131,847, and polydispersity (M(w)/M (n)) of 1.3. The data from this study indicate that J. curcas seed oil can be used as a substrate to produce the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3HB-co-3HV).

Characterization of copolymer hydroxybutyrate/hydroxyvalerate from saponified vernonia, soybean, and "spent" frying oils.

Poly(beta-hydroxyalkanoate)s (PHAs) were biosynthesized by Ralstonia eutropha (formerly known as Alcaligenes eutrophus) by using saponified soybean, vernonia, and "spent" frying oils. These PHAs were isolated and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), gas chromatography/mass spectrometry (GC/MS), proton nuclear magnetic resonance spectrometry (1H NMR), and 2-dimensional homonuclear (1H-1H) correlation spectroscopy (COSY). The analytical results revealed that the PHAs produced from saponified vernonia and soybean oils were copolymers of hydroxybutyrate (HB) and hydroxyvalerate (HV), that is, P(HB/HV)s, whereas the saponified "spent" frying oil produced only poly(beta- hydroxybutyrate) (PHB) homopolymer. MALDI-MS, GC/MS, and NMR independently confirmed the composition of the PHAs. Saponified soybean oil and vernonia oil PHAs contained approximately 4 and 1% HV units, respectively. For comparison, commercial PHB and P(HB/HV), produced by R. eutropha by using glucose and a cosubstrate of glucose and propionic acid, respectively, as carbon sources, were similarly characterized.