[Show abstract][Hide abstract] ABSTRACT: The present invention relates to poly(4-hydroxybutyrate)-b-monomethoxy(polyethylene glycol) copolymer nanoparticles, a method for preparing the same, and a pharmaceutical composition including the same as an active ingredient for treating brain disorders, in which the P(4HB)-b-mPEG nanoparticles according to the present invention break down slowly in brain cells such that gamma-hydroxybutyric acid (GHB) is released and so can sustainedly release a therapeutic level of concentration of GHB which is used as an agent for treating brain disorders such as epilepsy, and the nanoparticles are in the form of a hydrophobic polymer and so cannot easily be used as a hallucinogenic agent or narcotic agent due to the hydrophobic property whereby said nanoparticles do not dissolve in water or alcohol, and so can be used more safely than existing GHB, and hence, can advantageously be used instead of prior-art GHB therapeutic agents.
[Show abstract][Hide abstract] ABSTRACT: Despite several advancements in chemotherapy, cancer is still the second most frequent cause of mortality worldwide. Drug delivery to solid tumors is one of the most challenging aspects in cancer therapy. In pharmaceutical industries biodegradable polymeric nanoparticles as drug carriers
have attracted great research interest because of their biocompatibility, biodegradability and sustained release of drugs. In our study we prepared poly(4-hydroxybutyrate)-mPEG (P(4HB)-mPEG) nanocarriers for the delivery of cisplatin as anticancer drug to mouse hippocampal HT22 cells. P(4HB)
is more suitable candidate to be utilized in pharmaceutical industries due to its wide medical applications. P(4HB) is a homopolymer of 4-hydroxybutyrate (4HB), and belongs to a diverse class of materials called polyhydroxyalkanoates (PHA) produced by microorganisms inside the cells as energy
storage materials. P(4HB) has certain unique properties such as biocompatibility and rapid in vivo degradation, which differentiate it from others PHA based polymers. Novel amorphous amphiphilic block copolymer P(4HB)-mPEG nanocarriers were prepared and characterized. Flow cytometry,
and confocal microscopy revealed a suppression effect by the cisplatin loaded nanocarriers on HT22 cell growth, and enhancement of apoptotic process of the cells compared to free drug treated cells. The amorphous polymeric nanocarriers could be effective vehicles for the sustained delivery
of toxic anticancer drugs for the therapy of cancer.
Journal of Nanoscience and Nanotechnology 11/2014; 14(11). DOI:10.1166/jnn.2014.9924 · 1.56 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Provided is a method for enhancing the production of polyhydroxyalkanoic acid (PHA) from microorganism strains by disrupting a gene associated with the production of an exobiopolymer (EBP) in the Pseudomonas strain to redirect the carbon flux toward the production of the polyhydroxyalkanoic acid, thereby enhancing the production of the polyhydroxyalkanoic acid.
[Show abstract][Hide abstract] ABSTRACT: The present invention relates to poly(4-hydroxybutyrate)-b-monomethoxy
(polyethylene glycol) copolymer nanoparticles, to a production method for same
and to a pharmaceutical composition for brain disorder treatment containing
same as an active ingredient; and the P(4HB)-b-mPEG nanoparticles according
to the present invention break down slowly in brain cells such that gammahydroxybutyric
acid (GHB) is released and so can sustainedly release a therapeutic level of concentration of GHB which is used as an agent for treating brain
disorders such as epilepsy, and the nanoparticles are in the form of a hydrophobic polymer and so can easily be used as a hallucinogenic agent or narcotic agent
due to the hydrophobic property whereby said nanoparticles do not dissolve in water or alcohol, and so can be used more safely than existing GHB, and hence can
advantageously be used instead of prior-art GHB therapeutic agents.
[Show abstract][Hide abstract] ABSTRACT: Provided is a method for producing a polyhydroxyalkanoic acid (PHA). More particularly, a method of preparing PHA containing a high content of long-chain aromatic monomer-units by growing a phaZ mutant of a Pseudomonas strain in a medium containing a sugar, a substituted fatty acid, and salicylic acid.
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas aeruginosa, an opportunistic human pathogen is known to synthesize rhamnolipid and polyhydroxyalkanoic acid (PHA) of which the acyl-group precursors (e.g., (R)-3-hydroxydecanoic acid) are provided through RhlA and PhaG enzyme, respectively, which have 57% gene sequence homology. The inhibitory effect of three 2-bromo-fatty acids of 2-bromohexanoic acid (2-BrHA), 2-bromooctanoic acid (2-BrOA) and 2-bromodecanoic acid (2-BrDA) was compared to get an insight into the biochemical nature of their probable dual inhibition against the two enzymes. The 2-bromo-compounds were found to inhibit rhamnolipid and PHA synthesis simultaneously in alkyl-chain-length dependent manner at several millimolar concentrations. The separate and dual inhibition of the RhlA and PhaG pathway by the 2-bromo-compounds in the wild-type cells was verified by investigating their inhibitory effects on the rhamnolipid and PHA synthesis in P. aeruginosa ΔphaG and ΔrhlA mutants. Unexpectedly, the order of inhibition strength was found 2-BrHA (≥90% at 2 mM) > 2-BrOA > 2-BrDA, equally for all of the rhamnolipids and PHA synthesis, swarming motility and biofilm formation. We suggest that the novel strongest inhibitor 2-BrHA could be potentially exploited to control the rhamnolipid-associated group behaviors of this pathogen as well as for its utilization as a lead compound in screening for antimicrobial agents based on new antimicrobial targets.
PLoS ONE 09/2013; 8(9):e73986. DOI:10.1371/journal.pone.0073986 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cisplatin is a chemotherapeutic agent used against a variety of tumors. We determined the efficacy and bioavailability of cisplatin in the form of cisplatin-loaded self-assembled amphiphilic copolymer nanoparticles (NPs). Non-crystallizing bacterial copolyester was employed as hydrophobic segment to increase drug loading efficiency. Novel amorphous amphiphilic block copolymer P(3HV-co-4HB)-b-mPEG was synthesized from bacterial copolyester poly(3-hydroxyvalerate-co-4-hydroxybutyrate) coupled via transesterification reaction using bis(2-ethylhexanoate) tin catalyst to monomethoxypoly(ethylene glycol). The product was characterized, and core-shell particles with nanometer size range were prepared by emulsification-solvent evaporation method. Transmission electron microscopy (TEM) examination revealed that the NPs took the shape of spheres with inner concealed core of hydrophobic P(3HV-co-4HB) polymer and the outer shell formed by hydrophilic mPEG segment. The in vitro release profile of cisplatin from the core hydrophobic domain showed a sustained release of the drug. TEM and confocal microscopy examination revealed clearly the internalization of cisplatin-loaded NPs into the tumor cells. MTT assay, flow cytometry, western blot and confocal microscopy revealed a suppression effect by the NPs on tumor cell growth, and enhancement of apoptotic process of the tumor cells compared to free drug treated cells. The amorphous polymeric NPs could be effective vehicles for the sustained delivery of toxic anticancer drugs.
European journal of pharmaceutics and biopharmaceutics: official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V 12/2011; 80(3):518-27. DOI:10.1016/j.ejpb.2011.11.014 · 3.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Despite the recent research interest in the field of nanoparticles delivery system, their structure modification and transport behavior of various hydrophobic drugs is poorly developed. In this article the synthesis of novel amphiphilic diblock copolymer poly([R]-3-hydroxyvalerate)-block-monomethoxy poly(ethylene glycol) (PHV-block-mPEG) was undertaken by modifying the structure of biodegradable and hydrophobic poly([R]-3-hydroxyvalerate) (PHV) with hydrophilic monomethoxy poly(ethylene glycol) (mPEG). The chemical combination of the two blocks was carried out in the melt using bis(2-ethylhexanoate) tin as transesterification catalyst. The synthesized product was characterized by gel permeation chromatography (GPC), 1H nuclear magnetic resonance (NMR) and differential scanning calorimetry (DSC) analysis. The block copolymer self-assembled into amphiphilic nanoparticles with a core of hydrophobic PHV and a shell of hydrophilic mPEG in aqueous solution. Characterization of the nanoparticles showed the formation of discrete, spherically shaped nanoparticles with mean particle size of 200 +/- 1 nm and zeta potential of -14 +/- 1 mV. A hydrophobic drug thymoquinone was efficiently incorporated into the core hydrophobic domain of the nanoparticles and its release kinetics was studied in vitro. The amphiphilic PEGylated nanoparticles showed biocompatibility when checked in neuronal hippocampal cells of prenatal rat. Our results suggest that the amphiphilic nanoparticles with core-shell structures are potentially useful to develop novel drug carriers.
Journal of Nanoscience and Nanotechnology 07/2011; 11(7):5702-10. DOI:10.1166/jnn.2011.4493 · 1.56 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Amphiphilic biodegradable core-shell nanoparticles were prepared by emulsification-solvent evaporation technique from diblock copolymers which were synthesized by chemical coupling of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) P(3HB-co-3HV) or poly(3-hydroxybutyrate-co-4-hydroxybutyrate) P(3HB-co-4HB) to monomethoxy poly(ethylene glycol) (mPEG) through transesterification reaction. The nanoparticles were found to be assembled in aqueous solution into an outer hydrophilic shell of mPEG connected to the interior hydrophobic polyhydroxyalkanoate (PHA) copolymer core, which was identified by a comparative analysis of enzymatic degradation of the mPEG-coupled and non-coupled PHA nanoparticles. Morphological examination under atomic force microscope showed the formation of smooth spherically shaped nanoparticles. The average particle sizes and zeta potentials of amphiphilic nanoparticles were in the range of 112-162 nm and -18 to -27 mV, respectively. A hydrophobic drug thymoquinone was encapsulated in the nanoparticles and its release kinetics was studied. The in vitro cytotoxicity evaluation of the nanoparticles on prenatal rat neuronal hippocampal and fibroblast cells revealed that biocompatibility of the amphiphilic nanoparticles was generally independent of the ratio of comonomer units in the PHA block. In conclusion, the amphiphilic nanoparticles contained the hydrophobic PHA segments buried in the core and could thus be used as safe carriers for the controlled release of variety of hydrophobic drugs.
International Journal of Pharmaceutics 11/2010; 400(1-2):165-75. DOI:10.1016/j.ijpharm.2010.08.008 · 3.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Polyhydroxyalkanoic acids (PHAs) and rhamnolipids considered as biotechnologically important compounds are simultaneously produced by Pseudomonas aeruginosa. Both are synthesized from common precursors, (R)-3-hydroxyfatty acids. To find the probable metabolic relationship between their syntheses, we investigated the PHA and rhamnolipids production in four pha (phaC1, phaC2, phaZ, and phaG), four rhl (rhlA, rhlB, rhlR, and rhlI) and rpoS mutant strains of P. aeruginosa PA14 and PAO1 grown in minimal medium containing 70 mM fructose or 30 mM decanoic acid. Higher PHA accumulation was found in the rhamnolipid-negative mutants than in the wild-type strains, suggesting that 3-hydroxyfatty acid precursors become more available for PHA synthesis when rhamnolipids synthesis is absent. However, compared to the wild-type strains, rhamnolipids production was not enhanced in the four pha mutants of P. aeruginosa PA14 and PAO1 which indicates that rhamnolipids production in P. aeruginosa could be tightly regulated at the transcriptional level by a quorum-sensing response. The metabolic pathways for PHA and rhamnolipid synthesis from medium-chain-length fatty acids were also investigated using octanoic-1-¹³C acid. ¹³C NMR analysis revealed that the monomer-unit (R)-3-hydroxyoctanoate-1-¹³C being converted from the octanoic acid substrate was effectively incorporated into PHA. In the rhamnolipid synthesis, the (R)-3-hydroxyoctanoate-1-¹³C is suggested to be firstly converted to (R)-3-hydroxydecanoate-1,3-¹³C via fatty acid de novo biosynthesis pathway and then further processed into (R)-3-((R)-3-hydroxyalkanoyloxy)alkanoic acids (HAAs) via RhlA. The ratio of mono- to dirhamnolipids in the product depended on the type of carbon sources. The rhlB mutant could be exploited as an efficient producer of the important biosurfactant HAAs (e.g., ~700 mg/L HAAs was obtained when grown on 60 mM octanoic acid).
Journal of Biotechnology 10/2010; 151(1):30-42. DOI:10.1016/j.jbiotec.2010.10.072 · 2.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This is the first report on the characterization and production optimization of poly-N-acetylglucosamine (PNAG) in Staphylococcus saprophyticus. A strain producing glucosamine exopolysaccharide was isolated and characterized by biochemical test and 16S rRNA gene sequence homology analysis and named as S. saprophyticus BMSZ711. The molecular mass of the purified exopolymer was about 12 kDa. Digestion of the PNAG with DispersinB proved that it has beta-1,6 linkage. BMSZ711 can only produce PNAG when grown in M1 minimal medium but not in nutrient rich medium with optimum temperature of 30 degrees C and pH of 7. Glycerol and ammonium sulfate were found to be the best carbon and nitrogen source, respectively. Maximum PNAG production was obtained when glycerol 100mM, ammonium sulfate 0.3%, yeast extract 1.5 g/L, sodium chloride 10 g/L and valine 2mM were used.
[Show abstract][Hide abstract] ABSTRACT: Pseudomonas fluorescens BM07 is known to produce cold-induced exobiopolymer, which is mainly composed of water-insoluble hydrophobic polypeptides (up to 85%) and saccharides (8%), by decreasing the culture temperature down to as low as 10 degrees C. We screened for transposon insertion mutants of P. fluorescens BM07 that were unable to produce the exobiopolymer. Among the eight mutants that showed the deficiency of exobiopolymer and O-lipopolysaccharide, one mutant BM07-59 that had the highest polyhydroxyalkanoates (PHA) production was selected. The transposon inserted gene in BM07-59 was identified as galU. The disruption of the gene galU coded for the putative product, UDP-glucose pyrophosphorylase (GalU), resulted in 1.5-fold more accumulation of PHA compared with the wild-type strain from 70 mM fructose or galactose at 30 degrees C. Electrophoretic analysis of lipopolysaccharide showed that the mutant lacked the O-antigen lipopolysaccharide bands. The glycosyl composition of the lipopolysaccharide produced by the mutant strain was significantly different from that of the wild-type strain. We suggest that the deletion of galU could be a way to shift carbon flux efficiently from exobiopolymer toward PHA in P. fluorescens BM07.
[Show abstract][Hide abstract] ABSTRACT: The deletion of the intracellular polyhydroxyalkanoate (PHA) depolymerase gene (phaZ) in Pseudomonas fluorescens BM07 was found to increase more efficiently the levels of longer medium-chain-length (MCL) omega-aromatic monomer-units than in the wild-type strain when the cells were grown with a mixture of fructose and MCL omega-aromatic fatty acid in the presence of salicylic acid that is known as a beta-oxidation inhibitor in BM07 strain. When 11-phenoxyundecanoic acid was used as co-carbon source, the longest monomer-unit 3-hydroxy-11-phenoxyundecanoate, not reported in literature yet, was incorporated into the polymer chain up to approximately 10 mol%. An advantage of salicylic acid inhibition technique is that salicylic acid is not metabolized in BM07 strain, thus, the effective concentration of the inhibitor remaining constant throughout the cultivation. In conclusion, this new technique could be exploited for the enhanced production of side-chain modulated functional MCL-PHA with improved physicochemical properties in P. fluorescens BM07.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study is to prepare amphiphilic biodegradable core-shell nanoparticles using poly(3hydroxybutyrate-co-4hydroxybutyrate) copolymer as hydrophobic core and monomethoxy poly(ethylene glycol) as outer hydrophilic shell. The nanoparticles were prepared by emulsification-solvent evaporation technique using sodium deoxycholate as emulsifier. Particle size, surface morphology and zeta potential were measured by DLS, FE-SEM and ELS, respectively. Characterization of the nanoparticles shows the formation of spherically shaped nanoparticles with mean particle size range of 100 to 200 nm and zeta potential of -22 mV. The modified nanoparticles could be used as better source for drug delivery, ameliorating the toxicity associated with PHA based drug delivery materials.
[Show abstract][Hide abstract] ABSTRACT: Medium-chain-length-polyhydroxyalkanoic acids (MCL-PHAs) formed in Pseudomonas spp. have a rather broad distribution of monomer-units whose precursors are supplied via beta-oxidation degradation of MCL fatty acids fed as the carbon source and/or via PhaG enzyme catalyzing the acyl-group transfer from 3-hydroxyacyl-ACPs derived from acetyl-CoA to coenzyme A. It was found that salicylic acid (SA), in a concentration dependent manner, suppressed the accumulation of PHA in Pseudomonas fluorescens BM07 from fructose as well as shifted the distribution of monomer-units derived from a MCL fatty acid co-added as carbon source (e.g., 11-phenoxyundecanoic acid (11-POU)) to longer monomer-units. Both SA and acrylic acid were found to induce high accumulations of 3-ketohexanoic acid in BM07 wild-type cells grown with n-hexanoic acid as well as to inhibit the formation of acetyl-CoA from acetoacetyl-CoA by BM07 cell extract, suggesting that 3-ketoacyl-CoA thiolase is their common beta-oxidation target. The structural motif of acrylic acid present in the molecular structure of SA may self-explain the similar actions of the two inhibitors. A comparison of monomer modulation between BM07 wild-type and DeltaphaG mutant cells grown on the mixtures of fructose and 11-POU revealed that both PhaG and beta-oxidation inhibitor may play a critical role in the synthesis of PHA with longer side-chain omega-functional substitutions.
Biotechnology and Bioengineering 03/2009; 102(4):1209-21. DOI:10.1002/bit.22149 · 4.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Quantitative Ion Character-Activity Relationship (QICAR) was used for correlating metal ionic properties with maximum biosorption capacity (q(max)). Heat inactivated biomass of Staphylococcus saprophyticus BMSZ711 was studied for biosorption of nine metal ions. Influence of contact time and initial pH was checked. q(max) was determined by Langmuir isotherm and followed a descending sequence (in mmol/g): Pb(2+)>Cd(2+)>Cr(3+)>Zn(2+)>Hg(2+)>Cu(2+)>Co(2+)>Ni(2+)>K(+). q(max) values was modeled with 20 metal ionic characteristics, among these covalent index (X(m)(2)r) was best fitted with q(max) for all metal ions tested, in the following model: q(max)=0.09+0.11(X(m)(2)r) (R(2)=0.73, AIC=-4.14). Classification of metal ions according to valence or soft/hard improved QICARs modeling and more characteristics significantly correlated with q(max) which revealed that covalent bonding played major role in biosorption of soft metal ions and ionic bonding for borderline and hard ions. Biosorption capacity was most effectively predicted (R(2)=0.99, AIC=-8.04) with a two variable model containing electro-negativity (X(m)) and softness index (o(rho)(')).