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突变增强新型鞘氨醇单胞菌生产凝胶多糖:上游优化、动力学建模以及结构和物理功能评估。

Mutagenesis enhances gellan gum production by a novel Sphingomonas spp.: upstream optimization, kinetic modeling, and structural and physico-functional evaluation.

机构信息

Food Engineering and Technology Department, Institute of Chemical Technology, Mumbai, 400019, India.

Department of Microbiology, HiMedia Laboratories Pvt. Ltd, Mumbai, India.

出版信息

Int Microbiol. 2024 Apr;27(2):459-476. doi: 10.1007/s10123-023-00399-1. Epub 2023 Jul 26.

Abstract

Gellan gum (GG) has gained tremendous attention owing to its diversified applications. However, its high production and hence market cost are still a bottleneck in its widespread utilization. In the present study, high GG producing mutant of Sphingomonas spp. was developed by random mutagenesis using ethyl methylsulphonate (EMS) for industrial fermentation and identified as Sphingomonas trueperi after 16S rRNA and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) analysis. The fermentation conditions such as pH, temperature, and inoculum ratio were optimized by one factor at a time (OFAT) followed by screening of medium components by the Plackett-Burman statistical design. The most critical nutrients were further optimized by response surface methodology for maximizing GG production. The effect of dissolved oxygen tension in bioreactor on cell growth, substrate consumption, GG production, and batch productivity was elucidated. The highest GG titer (23 ± 2.4 g/L) was attained in optimized medium at 10% inoculum (6.45 ± 0.5 log cfu/mL) under controlled fermentation conditions of pH (7), temperature (30 °C), agitation (300-600 rpm), and aeration (0.5-2.0 SLPM) at 22 ± 2% dissolved oxygen tension in a 10-L bioreactor. Kinetic modeling of optimized batch process revealed that logistic growth model could best explain biomass accumulation, while GG formation and substrate consumption were best explained by Luedeking-Piret and exponential decay model, respectively. Structural and physico-functional features of GG produced by mutant Sphingomonas spp. were characterized by HPLC, FTIR, NMR, DSC, TGA, GPC, SEM, and rheological analysis. The higher productivity (0.51 g/L/h) under optimized fermentation conditions suggests potential consideration of mutant and process for commercial utilization.

摘要

黄原胶(GG)由于其多样化的应用而受到极大关注。然而,其高生产成本仍然是其广泛应用的瓶颈。在本研究中,通过乙基甲磺酸(EMS)随机诱变,开发了一种用于工业发酵的高 GG 产生突变体 Sphingomonas spp.,并通过 16S rRNA 和基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF-MS)分析鉴定为 Sphingomonas trueperi。通过单因素实验(OFAT)优化了发酵条件,如 pH、温度和接种比例,然后通过 Plackett-Burman 统计设计筛选培养基成分。通过响应面法进一步优化最关键的营养物质,以最大限度地提高 GG 的产量。阐明了生物反应器中溶解氧张力对细胞生长、底物消耗、GG 生产和分批生产力的影响。在优化的培养基中,在 10%的接种量(6.45±0.5 log cfu/mL)下,在控制发酵条件 pH(7)、温度(30°C)、搅拌(300-600 rpm)和通气(0.5-2.0 SLPM)下,在 22±2%的溶解氧张力下,在 10-L 生物反应器中获得了最高的 GG 浓度(23±2.4 g/L)。优化批处理过程的动力学模型表明,Logistic 生长模型可以最好地解释生物量的积累,而 GG 形成和底物消耗则可以用 Luedeking-Piret 和指数衰减模型来最好地解释。突变的 Sphingomonas spp.产生的 GG 的结构和物理化学功能特征通过 HPLC、FTIR、NMR、DSC、TGA、GPC、SEM 和流变分析进行了表征。在优化的发酵条件下,更高的生产率(0.51 g/L/h)表明突变体和工艺具有商业利用的潜力。

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