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Xue W.-J.,Shaanxi Provincial Institute of Microbiology | Fan D.-D.,Northwest University, China
Huaxue Gongcheng/Chemical Engineering (China) | Year: 2011

To control the acetate production and achieve a high-level expression of the target protein in the different-scale bioreactors during fed-batch production of human-like collagen (HLC) with recombinant Escherichia coli. A probing feeding strategy using feed-up DO-transient control was employed in two different-scale bioreactors and the dissolved oxygen response signal was used to detect the acetate production in pulsed fed-batch cultivation of E. coli. The final dry cell weight (DCW) of 69.1 g/L and HLC mass concentration of 13.1 g/L were obtained on the laboratory scale and the results are similar to that optimized in the previous study. When the process was scaled up to pilot-scale (500 L), the final DCW decreased from 80.3 g/L to 54.1 g/L with non-induced cultivations while acetate mass concentration was similar on the both scales. The difference in DCW was mainly resulted from the difference in oxygen transfer capacity. The final DCW of 51.7 g/L and HLC mass concentration of 9.6 g/L were obtained on the pilot-scale upon optimization of induction timing, and the results are satisfied based on the HLC content. The feed-up DO-transient control strategy can be successfully employed to control the acetate production and achieve a high-level expression of the target protein during the fed-batch production of HLC with recombinant E. coli.

Ma R.,Northwest University, China | Fan D.-D.,Northwest University, China | Fan D.-D.,Xian Giant Biogene Technology Ltd Company | Xue W.-J.,Shaanxi Provincial Institute of Microbiology | And 4 more authors.
Separation Science and Technology | Year: 2010

Interaction between human-like collagen (HLC) and endotoxin makes endotoxin removal from HLC more complicated. In this study, a simple, effective, and low-cost method was developed for endotoxin removal. A 0.4% (volume fraction) TritonX-100 solution was used to dissociate the HLC-endotoxin complexes and Q XL resin was used to remove endotoxin from the HLC solution. Product solution containing 1.2 mg. ml-1 HLC in Tris-HCl buffer (pH 7.5) and 50mM NaCl was added to the Q XL resin; the endotoxin removal efficiency was more than 98% and HLC recovery of up to 95% could be obtained; HLC purity was also enhanced after this process. The residual TritonX-100 was removed by using a superdex-200 column. This method greatly reduced the cost of purification but provided high endotoxin removal efficiency and HLC recovery. © Taylor & Francis Group, LLC.

Li X.,Northwest University, China | Fan D.-D.,Northwest University, China | Deng J.-J.,Northwest University, China | Hui J.-F.,Northwest University, China | And 3 more authors.
Asian Journal of Chemistry | Year: 2013

Injectable in situ forming biodegradable chitosan-human like collagen (CS-HLC) based hydrogels has proved to be a potential candidate as an injectable biomaterial for tissue engineering. With crosslinking agent of carbodiimide, the properties of the novel chitosan-humanlike collagen/b-sodium glycerophosphate-carbodiimide hydrogel were also examined. The gelation time, structure, equilibrium swelling and degradation in vitro and in vivo were dependent upon crosslinking and structure of composite hydrogels. The chitosan-human-like collagen/b-sodium glycerophosphate-carbodiimide hydrogel showed a desirable gelling time, swelling ratio, a smooth surface, regular porous networks and biodegradable. Therefore, the chitosan-human-like collagen/b-sodium glycerophosphate-carbodiimide hydrogels are excellent candidates for use in biomedical fields, such as in soft tissue defect filling and drug delivery.

Li X.,Northwest University, China | Xue W.,Shaanxi Provincial Institute of Microbiology | Liu Y.,Northwest University, China | Li W.,Northwest University, China | And 3 more authors.
Materials Science and Engineering C | Year: 2016

New locally injectable biomaterials that are suitable for use as soft tissue fillers are needed to address a significant unmet medical need. In this study, we used pullulan and human-like collagen (HLC) based hydrogels with various molecular weights (MWs) in combination therapy against tissue defects. Briefly, pullulan was crosslinked with NaIO4 to form a pullulan hydrogel and then may coupled with HLC using the reaction between the -NH2 end-group of HLC and the -CHO group present on the aldehyde pullulan to form the HLC/pullulan hydrogel, wherein the NaIO4 acted as the crosslinking and oxidizing agent. The good miscibility of pullulan and HLC in the hydrogels was confirmed via Fourier transform infrared spectroscopy, scanning electron microscopy, compression testing, enzyme degradation testing, cell adhesions, live/dead staining and subcutaneous filling assays. Here, pullulan hydrogels with various MWs were fabricated and physicochemically characterized. Limitations of the pullulan hydrogels included inflammation, poor mechanical strength, and degradation. By contrast, the properties of the HLC/pullulan hydrogels strongly enhanced. The efficacy of these hydrogels was evaluated both in vitro and in vivo. Our results indicate that HLC/pullulan hydrogels may have therapeutic value as efficient soft tissue fillers, with reduced inflammation, improved cell adhesion and delayed hydrogel degradation. © 2015 Published by Elsevier B.V.

Li X.,Northwest University, China | Xue W.,Shaanxi Provincial Institute of Microbiology | Zhu C.,Northwest University, China | Fan D.,Northwest University, China | And 2 more authors.
Materials Science and Engineering C | Year: 2015

Abstract Novel hydrogels based on carboxyl pullulan (PC) and human-like collagen (HLC) crosslinking with 1,4-butanediol diglycidyl ether (BDDE) are promising soft fillers for tissue engineering due to their highly tunable properties. Recent studies, however, have shown that incorporating hyaluronic acid and BDDE results in hydrogels with a microporous structure, a large pore size and high porosity, which reduce cell adhesion and enhance degradation in vivo. To improve biocompatibility and prevent biodegradation, the use of PC to replace hyaluronic acid in the fabrication of PC/BDDE (PCB) and PC/BDDE/HLC (PCBH) hydrogels was investigated. Preparation of gels with PC is a promising strategy due to the high reactivity, superb selectivity, and mild reaction conditions of PC. In particular, the Schiff base reaction of HLC and PC produces the novel functional group -RCONHR′ in PCBH hydrogels. Twenty-four weeks after subcutaneous injection of either PCB or PCBH hydrogel in mice, the surrounding tissue inflammation, enzymatic response and cell attachment were better compared to hyaluronic acid-based hydrogels. However, the biocompatibility, cytocompatibility and non-biodegradability of PCBH were milder than those of the PCB hydrogels both in vivo and in vitro. These results show that the proposed use of PC and HLC for the fabrication of hydrogels is a promising strategy for generating soft filler for tissue engineering. © 2015 Elsevier B.V.

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