Polyhydroxyalkanoate (PHA) is a biodegradable material with many potential biomedical applications, including medical implants and drug delivery. This study developed a system for screening production strains in order to optimize PHA production in Cupriavidus taiwanensis 184, 185, 186, 187, 204, 208, 209 and Pseudomona oleovorans ATCC 29347. In this study, Sudan black B staining, Infrared (IR) and Gas Chromatography (GC) analysis indicated that the best strain for PHA synthesis is C. taiwanensis 184, which obtains polyhydroxybutyrate (PHB). Cultivation of C. taiwanensis 184 under a pH of 7.0, at 30 °C, and at an agitation rate of 200 rpm, obtained a PHB content of 10% and PHB production of 0.14 g/L. The carbon and nitrogen types selected for analysis of PHB production by C. taiwanensis 184 were gluconic acid and NH4Cl, respectively. Optimal carbon/nitrogen ratio for PHB production was also determined. This study demonstrated a PHB content of 58.81% and a PHB production of 2.44 g/L when thecarbon/nitrogen ratio of 8/1 was selected for C. taiwanensis 184. A two‑stage fermentation strategy significantly enhanced PHB content and PHB production. Under a two-stage fermentation strategy with nutrient‑limited conditions, C. taiwanensis 184 obtained a PHB content of 72% and a PHB concentration of 7 g/L. Finally, experimental results confirmed that optimizing the growth medium and fermentation conditions for cultivating the indigenous C. taiwanensis 184 strain substantially elevated PHB content from 10% to 72% and PHB production from 0.14 g/L to 7 g/L, respectively.
Fengycin, a lipopeptide biosurfactant, was produced by indigenous Bacillus subtilis F29-3 isolated from a potato farm. Although inhibiting the growth of filamentous fungi, the fengycin is ineffective against yeast and bacteria. In this study, fengycin was isolated from fermentation broth of B. subtilis F29-3 via acidic precipitation (pH 2.0 with 5 N HCl) followed by purification using ultrafiltration and nanofiltration. The purified fengycin product was characterized qualitatively by using fast atom bombardment-mass spectrometer, Fourier transform infrared spectrometer, ultraviolet-visible spectrophotometer, 13C-nuclear magnetic resonance spectrometer and matrix assisted laser desorption ionization-time of flight, followed by quantitative analysis using reversed-phase HPLC system. This study also attempted to increase fengycin production by B. subtilis F29-3 in order to optimize the fermentation medium constituents. The fermentation medium composition was optimized using response surface methodology (RSM) to increase fengycin production from B. subtilis F29-3. According to results of the five-level four-factor central composite design, the composition of soybean meal, NaNO3, MnSO4·4H2O, mannitol-mannitol, soybean meal-mannitol, soybean meal‑soybean meal, NaNO3-NaNO3 and MnSO4·4H2O-MnSO4·4H2O significantly affected production. The simulation model produced a coefficient of determination (R2) of 0.9043, capable of accounting for 90.43% variability of the data. Results of the steepest ascent and central composite design indicated that 26.2 g/L of mannitol, 21.9 g/L of soybean meal, 3.1 g/L of NaNO3 and 0.2 g/L of MnSO4·4H2O represented the optimal medium composition, leading to the highest production of fengycin. Furthermore, the optimization strategy increased the fengycin production from 1.2 g/L to 3.5 g/L.
Polyhydroxybutyrate (PHB) is one of the polyhydroxyalkanoates (PHAs) which has biodegradable and biocompatible properties. They are adopted in the biomedical field, in, for example, medical implants and drug delivery carriers. This study seeks to promote the production of PHB by Vibrio sp.BM-1, isolated from a marine environment by improving constituents of medium and implementing an appropriate fermentation strategy. This study successfully developed a glycerol-yeast extract-tryptone (GYT) medium that can facilitate the growth of Vibrio sp.BM-1and lead to the production of 1.4 g/L PHB at 20 h cultivation. This study also shows that 1.57 g/L PHB concentration and 16% PHB content were achieved, respectively, when Vibrio sp. BM-1 was cultivated with MS-GYT medium (mineral salts-supplemented GYT medium) for 12 h. Both cell dry weight (CDW) and residual CDW remained constant at around 8.2 g/L and 8.0 g/L after the 12 h of cultivation, until the end of the experiment. However, both 16% of PHB content and 1.57 g/L of PHB production decreased rapidly to 3% and 0.25 g/L, respectively from 12 h of cultivation to 40 h of cultivation. The results suggest that the secretion of PHB depolymerase that might be caused by the addition of mineral salts reduced PHB after 12 h of cultivation. However, work will be done to explain the effect of adding mineral salts on the production of PHB by Vibrio sp.BM-1 in the near future.
Chondral or osteochondral defects are still controversial problems in orthopedics. Here, chondrocytes labeled with magnetic nanoparticles were cultivated on a biphasic, type II collagen–chitosan/poly(lactic-co-glycolic acid) scaffold in an attempt to develop cultures with trackable cells exhibiting growth, differentiation, and regeneration. Rabbit chondrocytes were labeled with magnetic nanoparticles and characterized by scanning electron microscopy (SEM), transmission electron (TEM) microscopy, and gene and protein expression analyses. The experimental results showed that the magnetic nanoparticles did not affect the phenotype of chondrocytes after cell labeling, nor were protein and gene expression affected. The biphasic type II collagen–chitosan/poly(lactic-co-glycolic) acid scaffold was characterized by SEM, and labeled chondrocytes showed a homogeneous distribution throughout the scaffold after cultivation onto the polymer. Cellular phenotype remained unaltered but with increased gene expression of type II collagen and aggrecan, as indicated by cell staining, indicating chondrogenesis. Decreased SRY-related high mobility group-box gene (Sox-9) levels of cultured chondrocytes indicated that differentiation was associated with osteogenesis. These results are encouraging for the development of techniques for trackable cartilage regeneration and osteochondral defect repair which may be applied in vivo and, eventually, in clinical trials.
This article studied the effects of platelet-rich plasma (PRP) on the potential of synovial fluid mesenchymal stem cells (SF-MSCs) to differentiate. The PRP and SF-MSCs were obtained from the blood and knees of pigs, respectively. The identification of SF-MSCs and their ability to differentiate were studied by histological and surface epitopes, respectively. The SF-MSCs can undergotrilineage mesenchymal differentiation under osteogenic, chondrogenic, and adipocyte induction. The effects of various PRP concentrations (0%, 20% and 50% PRP) on differentiation were evaluated using the SF-MSCs-alginate system, such as gene expression and DNA proliferation. A 50% PRP concentration yielded better differentiation than the 20% PRP concentration. PRP favored the chondrogenesis of SF-MSCs over their osteogenesis in a manner that depended on the ratios of type II collagen/type I collagen and aggrecan/osteopontin. Eventually, PRP promoted the proliferation of SF-MSCs and induced chondrogenic differentiation of SF-MSCs in vitro. Both PRP and SF-MSCs could be feasibly used in regenerative medicine and orthopedic surgeries.
1 August 2012 in Journal of Bioscience and Bioengineering