Tailoring the supramolecular structure of guanidinylated pullulan toward enhanced genetic photodynamic therapy

In the progress of designing a gene carrier system, what is urgently needed is a balance of excellent safety and satisfactory efficiency. Herein, a straightforward and versatile synthesis of a cationic guanidine-decorated dendronized pullulan (OGG3P) for efficient genetic photodynamic therapy was proposed. OGG3P was able to block the mobility of DNA from a weight ratio of 2. However, G3P lacking guanidine residues could not block DNA migration until at a weight ratio of 15, revealing guanidination could facilitate DNA condensation via specific guanidinium-phosphate interactions. A zeta potential plateau (∼+23 mV) of OGG3P complexes indicated the nonionic hydrophilic hydroxyl groups in pullulan might neutralize the excessive detrimental cationic charges. There was no obvious cytotoxicity and hemolysis, but also enhancement of transfection efficiency with regard to OGG3P in comparison with that of native G3P in Hela and HEK293T cells. More importantly, we found that the uptake efficiency in Hela cells between OGG3P and G3P complexes was not markedly different. However, guanidination caused changes in uptake pathway and led to macropinocytosis pathway, which may be a crucial reason for improved transfection efficiency. After introducing a therapeutic pKillerRed-mem plasmid, OGG3P complexes achieved significantly enhanced KillerRed protein expression and ROS production under irradiation. ROS-induced cancer cells proliferation suppression was also confirmed. This study highlights the guanidine-decorated dendronized pullulan could emerge as a reliable nonviral gene carrier to specifically deliver therapeutic genes

Biosynthesis and Thermal Properties of PHBV Produced from Levulinic Acid by <i>Ralstonia eutropha</i>

<div><p>Levulinic acid (LA) can be cost-effectively produced from a vast array of renewable carbohydrate-containing biomaterials. LA could facilitate the commercialization of the polymer poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and PHBV-based products as carbon substrates. Therefore, this paper focused on the production of PHBV by <i>Ralstonia eutropha</i> with LA for hydroxyvalerate (HV) production, which plays an important role in enhancing the thermal properties of PHBV. Accordingly, the HV content of PHBV varied from 0–40.9% at different concentrations of LA. Stimulation of cell growth and PHBV accumulation were observed when 2–6 g L⁻¹ LA was supplied to the culture. The optimal nitrogen sources were determined to be 0.5 g L⁻¹ ammonium chloride and 2 g L⁻¹ casein peptone. It was determined that the optimal pH for cell growth and PHBV accumulation was 7.0. When the cultivation was performed in large scale (2 L fermenter) with a low DO concentration of 30% and a pH of 7.0, a high maximum dry cell weight of 15.53 g L⁻¹ with a PHBV concentration of 12.61 g L⁻¹ (53.9% HV), up to 81.2% of the dry cell weight, was obtained. The melting point of PHBV found to be decreased as the fraction of HV present in the polymer increased, which resulted in an improvement in the ductility and flexibility of the polymer. The results of this study will improve the understanding of the PHBV accumulation and production by <i>R. eutropha</i> and will be valuable for the industrial production of biosynthesized polymers.</p> </div

Biosynthesis of poly(hydroxybutyrate-hydroxyvalerate) from the acclimated activated sludge and microbial characterization in this process

National Basic Research Program of China [2013CB733505]; National Natural Science Foundation of China [41271260, 41071302]; Program for New Century Excellent Talents in University [NCET-12-0326]; Development and Reform Commission of Fujian Province, China [2011-1598]; Science and Technology Program of Xiamen of Fujian Province, China [3502Z20126005]This study investigated the effects of substrate composition (acetate and propionate) on synthesis of various mix of poly(hydroxybutyrate-hydroxyvalerate) (P(HB/HV)) from activated sludge, which was acclimated using a single carbon (acetate) and mixed carbons (acetate and propionate). Results of batch P(HB/HV) production tests indicated that the yield and synthesis rate of P(HB/HV) decreased as the proportion of propionate in the substrate increased. However, mixed-carbon-acclimated sludge with acetate and propionate exhibited better P(HB/HV) production performance than with acetate-acclimated sludge in terms of substrate utilization, yield of P(HB/HV) and HV fraction in P(HB/HV). The desired hydroxyvalerate (HV) fraction (0-74%) of the P(HB/HV) could be obtained based on the substrate composition and sludge acclimation. Furthermore, PCR-DGGE analysis indicated that specific species dominated the activated sludge used for P(HB/HV) production. Acidobacteria and Burkholderiales were the dominant bacterial populations and played an important role in HV synthesis. (C) 2013 Elsevier Ltd. All rights reserved

Batch cultivation of <i>R. eutropha</i> for PHBV accumulation under the condition of stable pH and DO, which was automatically kept at 7.0 and 30%, respectively.

<p>1.2 L (pH 7.0) of media was in 2 L fermenter with 5% (v/v) inoculum at 30°C and aeration.</p

DSC thermograms of PHB (a), PHBV₁ (b) and PHBV₂ (c) samples.

<p>DSC thermograms of PHB (a), PHBV₁ (b) and PHBV₂ (c) samples.</p

TG-DTG curve of PHB (a), PHBV1 (b) and PHBV2 (c) samples.

<p>TG-DTG curve of PHB (a), PHBV1 (b) and PHBV2 (c) samples.</p

Batch cultivation of <i>R. eutropha</i> for PHBV accumulation under the condition of stable pH, which was automatically fed to keep the pH of the culture at 7.0. 1.2 L (pH 7.0) of media was in 2 L fermenter with 5% (v/v) inoculum at 30°C and aeration/agitation (500 rpm and 2 vvm air volume/culture volume/min).

<p>Batch cultivation of <i>R. eutropha</i> for PHBV accumulation under the condition of stable pH, which was automatically fed to keep the pH of the culture at 7.0. 1.2 L (pH 7.0) of media was in 2 L fermenter with 5% (v/v) inoculum at 30°C and aeration/agitation (500 rpm and 2 vvm air volume/culture volume/min).</p

Optimization of polyhydroxybutyrate (PHB) production by excess activated sludge and microbial community analysis

In this study, a high value-added and biodegradable thermoplastic, polyhydroxybutyrate (PHB), was produced by excess activated sludge. The effects of the nutritional condition, aeration mode, sodium acetate concentration and initial pH value on PHB accumulation in the activated sludge were investigated. The maximum PHB content and PHB yield of 67.0% (dry cell weight) and 0.740 gCOD gCOD(-1) (COD: chemical oxygen demand), respectively, were attained by the sludge in the presence of 6.0 g L(-1) sodium acetate, with an initial pH value of 7.0 and intermittent aeration. The analysis of the polymerase chain reaction (PCR)-denaturing gradient-gel-electrophoresis (DGGE) sequencing indicated that the microbial community of the sludge was significantly different during the process of PHB accumulation. Three PHB-accumulating microorganisms, which were affiliated with the Thauera, Dechloromonas and Competibacter lineages, were found in the excess activated sludge under different operating conditions for PHB accumulation. (C) 2010 Elsevier B.V. All rights reserved.National Natural Science Foundation of China[30700020]; Science and Technology Program of Fujian Province, China[2008Y0060]; Science and Technology Program of Xiamen of Fujian Province, China[3502Z20073005]; Natural Science Foundation for Youths of Fujian Province, China[2010J05121

Effect of different glucose concentrations on DCW, PHBV and HV production.

<p>Sharing a common lowercase are not significantly different in the HV content and the same capital are not significantly different in the concentration of DCW and PHBV (<i>P</i><0.05). 50 mL (pH 7.0) of media was in 250 mL flask with 5% (v/v) inoculum under shaking conditions at 200 rpm for 72 h at 30°C.</p

Thermal properties of PHBV samples with different fractions of HV.

<p>Xc: crystallinity, Tm: melting point, ▵H_f: melting enthalpy, T_eoi: initial thermal decomposition temperature, T_em: onset temperature, T_eof: finished thermal decomposition temperature, Wt-loss: thermal weight loss.</p