193 research outputs found
Optimization of a fed-batch fermentation process for production of bleomycin by Streptomyces mobaraensis ATCC 15003
The objective of this work was to optimize inoculum size and pH for rapid production of bleomycin by Streptomyces mobaraensis ATCC 15003 by fed-batch fermentation. In both batch- and fed-batch fermentations, neither production period for bleomycin by S. mobaraensis nor the amount of BLM were affected by increasing the inoculum size from regular 10 to 30% (v/v) level. A fed-batch bioreactor not only shortened the lag phase of BLM production from 114 to 60 h, but also fed-batch fermentationenhanced BLM production when combined with an appropriate pH profile with no effect on the amount produced. Due to the substrate inhibition that takes place at high levels of carbon source, fed-batch fermentation was proposed as a better alternative for BLM production. The combined effects of batch and fed-batch fermentation and various pH profiles on BLM production in a bioreactor were evaluated. The tested pH profiles included; (1) a constant pH profile at 6.8 (profile1); (2) a constant pH profile witha period of auto-acidification for 72 h (profile2) and (3) a step-wise pH profile with pH adjustment every 24 h (profile3). When profile 3 was applied, fed-batch fermentation enhanced BLM production in the bioreactor and yielded about two-fold higher BLM concentration than the irrespective batch fermentation. On the other hand, constant pH profile or that which included one period of autoacidification (profiles 1 and 2) resulted in an insignificantly difference BLM production in fed-batch fermentation. Overall, this study suggested that fed-batch fermentation can be successfully used to enhance BLM production in bioreactor especially with fluctuated pH-profile.Key words: Bleomycin, Streptomyces mobaraensis, rapid production, fed-batch fermentation
Improvement and enhancement of clavulanic acid production in Streptomyces clavuligerus using vegetable oils
Clavulanic acid (CA) is a potent inhibitor of β-lactamases. Oil can be used as a source of carbon and energy in CA production. To develop a policy for enhanced production of CA, the selection of a production medium and an optimum pH, different vegetable oils as carbon source have been used. The effects of different medium containing vegetable oil on cell growth and CA yield production during the fermentation of Streptomyces clavuligerus ATCC 27064 were demonstrated. In this study, three out of eight tested oils supported CA production. Medium containing olive oil showed two-fold higher CA yield than glycerol containing medium. The highest productivity was obtained at initial pH 7. We concluded that using olive oil as a sole source of carbon and energy for cultivation of S. clavuligerus is a promising strategy for CA production. It has several scientific advantages and economic benefits that lead to increased antibiotic titre and can be considered as a cheaper alternative compared to carbohydrates. The results of this study can be applied for the efficient production of β-lactamase inhibitory antibiotics.Key words: Clavulanic acid, enzymes, β-lactamases, vegetable oils, Streptomyces clavuligerus
Solubility of Gliclazide in Transcutol + Water Co-solvent Mixtures at (298.15 to 333.15) K
The aim of present investigation was to determine the mole fraction solubility of a poorly water soluble antidiabetic drug gliclazide (GLZ) in mono-solvents and various Transcutol + water co-solvent mixtures at (298.15 to 333.15) K. The experimental solubility of GLZ was measured by shake flask method and resulting data was correlated with the modified Apelblat model at each temperature studied. Good correlation was observed between the experimental data of GLZ and calculated one with absolute relative deviation in the range of (0.050 to 5.680) %. The correlation coefficients were observed in the range of 0.9966 to 0.9995 which indicated good fitting of experimental solubility data. The lowest mole fraction solubility of GLZ was observed in pure water (1.9 × 10–6 at 298.15 K) whereas the highest one was observed in pure Transcutol (11.9 × 10–3 at 298.15 K). The enthalpies and entropies for GLZ dissolution were observed as positive values in the range of (15.742 to 40.551) kJ mol–1 and (52.801 to 121.721) J mol–1 K–1, respectively in all sample matrices. These results of thermodynamic parameters indicated that the dissolution of GLZ is endothermic and an entropy-driven process. Based on current solubility data, GLZ was considered as practically insoluble (poorly soluble) in pure water and soluble in Transcutol. These preliminary studies indicated that Transcutol could be used as a co-solvent for solubility enhancement of GLZ which could help in preformulation studies and formulation development of GLZ
Hepatocellular proliferation in response to agonists of peroxisome proliferator-activated receptor alpha: a role for kupffer cells?
BACKGROUND: It has been proposed that PPARα agonists stimulate Kupffer cells in rodents which in turn, release mitogenic factors leading to hepatic hyperplasia, and eventually cancer. However, Kupffer cells do not express PPARα receptors, and PPARα agonists stimulate hepatocellular proliferation in both TNFα- and TNFα receptor-null mice, casting doubt on the involvement of Kupffer cells in the mitogenic response to PPARα agonists. This study was therefore designed to investigate whether the PPARα agonist PFOA and the Kupffer cell inhibitor methylpalmitate produce opposing effects on hepatocellular proliferation and Kupffer cell activity in vivo, in a manner that would implicate these cells in the mitogenic effects of PPARα agonists. METHODS: Male Sprague-Dawley rats were treated intravenously via the tail vein with methylpalmitate 24 hrs prior to perfluorooctanoic acid (PFOA), and were sacrificed 24 hrs later, one hr after an intraperitoneal injection of bromodeoxyuridine (BrdU). Sera were analyzed for TNFα and IL-1β. Liver sections were stained immunohistochemically and quantified for BrdU incorporated into DNA. RESULTS: Data show that PFOA remarkably stimulated hepatocellular proliferation in the absence of significant changes in the serum levels of either TNFα or IL-1β. In addition, methylpalmitate did not alter the levels of these mitogens in PFOA-treated animals, despite the fact that it significantly blocked the hepatocellular proliferative effect of PFOA. Correlation between hepatocellular proliferation and serum levels of TNFα or IL-1β was extremely poor. CONCLUSION: It is unlikely that mechanisms involving Kupffer cells play an eminent role in the hepatic hyperplasia, and consequently hepatocarcinogenicity attributed to PPARα agonists. This conclusion is based on the above mentioned published data and the current findings showing animals treated with PFOA alone or in combination with methylpalmitate to have similar levels of serum TNFα and IL-1β, which are reliable indicators of Kupffer cell activity, despite a remarkable difference in hepatocellular proliferation
Development of a new medium containing date syrup for production of bleomycin by Streptomyces mobaraensis ATCC 15003 using response surface methodology
A combined statistical approach of orthogonal design and polynomial regression were applied to optimize the composition and concentration of a liquid fermentation medium for the production of bleomycin (BLM) by Streptomyces mobaraensis. Optimal conditions for maximal productivity were determined based on eight parameters at three different levels. The sources of carbon and nitrogen concentration and their interactions with other precursors were found to be statistically significant factors. When date syrup was used as an additional carbon source, higher BLM amount was obtained in comparison to glucose. It was found that the optimum nitrogen source was achieved with the use of soyabean meal. The combined orthogonal design and response surface methodology predicted optimal conditions for production of BLM to be 138 mg dl-1. A confirmatory experiment of the optimal medium composition produced 142 mg dl-1 in the fifth day fermentation at 30°C. The complex medium containing 40 gml-1 date syrup as additional carbon source enhanced the production of BLM by 73%. The combined statistical approach enabled rapid identification and integration of key medium parameters for optimizing secondary metabolite production and could be very useful in pharma-ceutical screening programs.Keywords: Bleomycin, Streptomyces mobaraensis, orthogonal design, medium optimization, date syrupAfrican Journal of Biotechnology Vol. 9(33), pp. 5450-5459, 16 August, 201
Physical pegylation enhances the cytotoxicity of 5-fluorouracil-loaded PLGA And PCL nanoparticles.
Purpose :
The main goal of this study is to evaluate the impact of physical incorporation of polyethylene glycol (PEG) into 5-fluorouracil (5-FU)-loaded polymeric nanoparticles (NPs).
METHODS:
The 5-FU-loaded NPs were prepared utilizing a simple double emulsion method using polycaprolactone (PCL) and polylactic-co-glycolic acid (PLGA) with or without PEG 6000. The surface charge, particle size, and shape of NPs were evaluated by standard procedures. Both Fourier Transform Infrared Spectroscopy and X-ray diffraction spectra of the 5-FU loaded NPs were compared against the pure 5-FU. The in vitro release profile of 5-FU from the NPs was monitored by the dialysis tubing method. Cell death and apoptosis induction in response to 5-FU NP exposure were measured by MTT and Annexin-V/7-amino-actinomycin D (7-AAD) assays, respectively, in Daoy, HepG2, and HT-29 cancer cell lines.
RESULTS:
The 5-FU loaded NPs were found to be spherical in shape with size ranging between 176±6.7 and 253.9±8.6 nm. The zeta potential varied between -7.13± 0.13 and -27.06±3.18 mV, and the entrapment efficiency was between 31.96% and 74.09%. The in vitro release of the drug followed a two-phase mode characterized by rapid release in the first 8 hrs followed by a period of slow release up to 72 hrs with composition-based variable extents. Cells exposed to NPs demonstrated a significant cell death which correlated with the ratio of PEG in the formulations in Daoy and HepG2 cells but not in HT-29 cells. Formulations (F1-F3) significantly induced early apoptosis in HT-29 cell lines.
CONCLUSION:
The physical PEGylation significantly enhanced the entrapment and loading efficiencies of 5-FU into NPs formulated with PLGA and PCL. It also fostered the in vitro cytotoxicity of 5-FU-loaded NPs in both Daoy and HepG2 cells. Induction of early apoptosis was confirmed for some of the formulations
PEGylation of PLGA and PCL nanoparticles enhanced the cytotoxicity of loaded 5-flurouracil
Introduction: The high metabolic rate and the short biological half-life of 5-flourouracil (5-FU) required a continuous administration of large doses and consequently resulting in high profile of adverse effects. Incorporation of 5-FU into biodegradable nanoparticles (NPs) would enhance the therapeutic efficacy through prolongation of the biological half-life and the duration of tumour exposure to 5-FU. This study aimed to monitor the effect of physical incorporation of PEG on the attributes of 5-FU-loaded polymeric NPs.
Methods: An emulsification-solvent evaporation technique was employed for the preparation of 5-FU-loaded NPs using polylactic-co-glycolic acid (PLGA) and polycaprolactone (PCL). The effect of incorporating polyethylene glycol (PEG6000) was also investigated. The prepared NPs were evaluated for their particle sizes and morphology and characterized by FTIR and X-ray diffraction. The in vitro drug release profiles were evaluated and the anticancer activity was assessed utilizing MTT assay Daoy, HepG2, and HT29 cancer-cell-lines.
Results: The NPs average sizes were between 176±6.7-253.9±8.6 nm and zeta potential between -7.13± 0.13 and -27.06±3.18 mV. The 5-FU %EE of ranged between 31.96-73.54% and enhanced significantly by PEG incorporation. The SEM images showed spherical particles with smooth surfaces. The in vitro release studies showed an initial rapid 5-FU release up to 8 h followed by a slow release ranging from 36 to 84% after 72 h. The higher was the ratio of PEG, the faster was the drug release rate. Significant % cell death was achieved with all the prepared NPs in the three tested cancer cell lines after 48 and 72 hours incubations. The PEG ratio correlated well with the magnitude of cell death in both Doay and HepG cells only.
Conclusion: The physical PEGylation resulted in significant increase in the entrapment and loading efficiency of 5-FU in both PLGA and PCL NPs. They also improved both the drug release profile and the extent of in vitro cytotoxicity in both Doay and HepG2 cancer cell lines
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