基于傅里叶变换近红外光谱实时分析1,3-丙二醇发酵过程生物量的在线监测方法

生物量是反映生物发酵过程进展的重要参数,对生物量进行实时监测可用于对发酵过程的调控优化。为克服目前主要采用的离线方法检测生物量时间滞后和人工测量误差较大等缺点,本研究针对1,3-丙二醇发酵过程设计了一个基于傅里叶变换近红外光谱实时分析技术的生物量在线监测实验平台,通过对实时采集光谱预处理以及敏感光谱段分析,应用偏最小二乘算法,建立了1,3-丙二醇发酵过程生物量变化的动态预测模型。以底物甘油浓度为60 g/L和40 g/L的发酵过程作为外部验证实验,分析得到模型的预测均方根误差分别为0.341 6和0.274 3,结果表明所建立的模型具有较好的实时预测能力,能够实现对1,3-丙二醇发酵过程中生物量的有效在线监测。     

基于近红外光谱实时监测乙醇发酵过程多组分参数

生物量、葡萄糖浓度和乙醇浓度是乙醇发酵过程的重要参数,传统的方法通常对发酵液取样作离线测量,不仅需要采用多种仪器进行测试分析,而且耗时耗力,成为实时过程调控和优化的障碍。文中针对这些重要过程参数提出了一个基于近红外光谱技术的原位实时检测方法。通过采用浸入式近红外光谱仪对发酵溶液进行原位测量,基于多输出最小二乘支持向量机回归(MLS-SVR)方法建立了利用近红外光谱同时分析葡萄糖浓度、生物量和乙醇浓度的多输出预测模型。实验结果表明,该方法能实时准确地检测乙醇发酵过程中的葡萄糖浓度、生物量和乙醇浓度,而且相对于现有的偏最小二乘法(PLS)分别对各组分建模和预测,能明显提高测量准确性和可靠性。     

原子力显微镜 (AFM) 在细菌生物被膜研究中的应用

原子力显微镜(AFM)作为一项重要的表面可视化技术,以其独特的优势(纳米级的空间分辨率、皮牛级力灵敏度、免标记、可在溶液环境下工作)被广泛应用于生物被膜的研究。AFM不仅可以在近生理环境下对生物被膜表面超微形貌进行可视化表征,同时还可以通过纳米压痕对生物被膜的机械特性(弹性和粘性)进行定量测量,利用AFM单细胞和单分子力谱技术可以获得生物被膜形成过程中细胞-基底以及细胞-细胞之间的相互作用力,为生物被膜的实时原位系统研究提供了可行性。本文简述了AFM的基本操作原理,综述了近年来AFM用于生物被膜表面超微结构成像、机械特性测量以及相互作用力研究方面的进展,并对AFM在生物被膜研究中面临的问题和未来的发展方向进行了讨论。     

荧光定量PCR方法检测白酒发酵过程中Aspergillus tubingensis生物量

【目的】为了更好地分析霉菌在白酒发酵过程中的作用,需要快速准确地测定发酵过程中霉菌生物量的变化,本实验以白酒酿造中常用的塔宾曲霉(Aspergillus tubingensis)为例,建立一套快速准确定量塔宾曲霉生物量的方法。【方法】优化从酒醅中提取基因组的方法,设计和验证专一性引物,建立实时荧光定量PCR(Real-time quantitative PCR)方法,验证方法的有效性并应用于白酒发酵过程中塔宾曲霉生物量的检测。【结果】用原位机械破碎法提取酒醅中总基因组,其DNA的浓度能够达到1.060×105 ng/g酒醅;同时建立了一套快速准确测定固态基质中霉菌生物量的方法,并应用于白酒生产(制曲、堆积发酵和窖池发酵过程)中塔宾曲霉生物量的定量。【结论】实时荧光定量PCR方法能够快速准确地测定固态基质中霉菌的生物量,且检测限较低,对今后的相关研究具有借鉴意义。     

微生物发酵过程生理参数检测传感器技术与过程优化

工业生物发酵过程是一个复杂的代谢表达调控系统。阐述了开发和利用先进的发酵过程传感仪表技术,进行环境参数和细胞生理代谢特性参数的实时精确测定,是系统了解和分析工业微生物的宏观与微观代谢特性的基础。实现生物传感器与信息技术、过程多参数智能化过程分析的软件识别系统联合,是构建新型的智能化工业生物过程控制与生产模式、建立生产过程的绿色智能制造新技术的关键。     

细胞内 NO 的荧光成像

NO是一种具有重要生物学意义的信息分子,在体内具有广泛的生物学特征。但由于NO的自由基性质,使得在活细胞中对低浓度、低寿命的NO实时监测异常困难。为了进一步了解NO在神经、免疫、血管和消化等多种系统中的生理功能,高度专一性的、高灵敏的荧光探针结合激光扫描共聚焦显微镜对活细胞中的NO进行实时、连续的成像已被广泛研究。该综述了近年来NO荧光探针的发展及其在生物成像中的应用。     

全细胞生物传感器的设计及其在环境监测中的应用

全细胞生物传感器是一种以微生物全细胞为敏感元件,可以快速感应环境中的毒性物质及污染物的装置。因其具有响应快、体积小、成本低、可实现原位监测等优势,在环境监测、药品研发、食品工业等领域显示出巨大潜力。综述了全细胞生物传感器的原理、分类及其在环境污染物监测领域的应用进展等,并对其未来的发展趋势进行了展望,以期为全细胞生物传感器的开发与利用提供参考。     

红外光谱技术在生物过程监测中的应用

在线监测化学组分的浓度对许多生物过程都是十分必要的。然而,探头需耐高温灭菌的要求和生物体系自身的复杂性给许多分析技术的在线监测带来了困难。近几年,随仪器和数据处理技术的迅速发展,应用红外光谱技术对生物过程的原位或在线监测日益广泛。本文对红外过程分析技术进行了较全面的综述,介绍了红外分析的原理、进展及在生物过程监测中的应用。     

大数据-模型混合驱动下生物过程优化与放大的新机遇与挑战

当前,生物制造技术和产业是世界关注的热点。然而,生物过程优化与放大过程中普遍面临以下几个难题,包括:过程检测手段缺乏,难以满足关键指标参数的监控;细胞代谢认知匮乏,无法理性实现过程最优化调控;反应器环境差异大,导致逐级放大效率低下。文中针对以上亟待解决的关键问题,通过案例分析介绍发酵过程实时检测-动态调控-理性放大全链条关键技术创新。在未来,生物过程设计将以集成细胞生理学(时空多尺度细胞代谢模型)和流体动力学(CFD模型)的全生命周期模型为指导,推进计算机辅助设计与开发,加速生物过程实现大规模智能化生产,开启绿色生物制造新时代。     

电化学法快速检测微生物的发展现状及趋势

自1898年Stewart提出利用电化学法检测微生物,电化学法已发展成为一种微生物快速检测的方法。根据检测的参数不同,电化学微生物检测法可以分为阻抗微生物法和介电常数法。阻抗法主要用于食品工业中微生物的快速检测(≤10⁷ cfu/mL),尤其用于易腐食品的微生物快速检测,以期实现在其发生明显腐败之前得到检测结果。而介电常数则用于生物发酵过程中的微生物数量的快速测定,可以实现在线监测微生物数量及生物发酵过程的实时控制。电化学法由于其检测迅速、可以实现自动化检测,在工业化生产中具有广阔的应用前景。     

Online monitoring of Mezcal fermentation based on redox potential measurements

We describe an algorithm for the continuous monitoring of the biomass and ethanol concentrations as well as the growth rate in the Mezcal fermentation process. The algorithm performs its task having available only the online measurements of the redox potential. The procedure combines an artificial neural network (ANN) that relates the redox potential to the ethanol and biomass concentrations with a nonlinear observer-based algorithm that uses the ANN biomass estimations to infer the growth rate of this fermentation process. The results show that the redox potential is a valuable indicator of the metabolic activity of the microorganisms during Mezcal fermentation. In addition, the estimated growth rate can be considered as a direct evidence of the presence of mixed culture growth in the process. Usually, mixtures of microorganisms could be intuitively clear in this kind of processes; however, the total biomass data do not provide definite evidence by themselves. In this paper, the detailed design of the software sensor as well as its experimental application is presented at the laboratory level.     

燃料乙醇发酵技术研究进展

在分析木质纤维素类生物质制备燃料乙醇原理基础上,重点对燃料乙醇转化过程的发酵工艺进行了论述。目前乙醇发酵工艺主要包括直接发酵、分步糖化发酵、同步糖化发酵、同步糖化共发酵和联合生物加工技术等,对这几种技术的研究现状进行了分析并对其发展趋势进行了展望,通过基因工程构建高效发酵菌种的联合生物加工技术将是未来高效发酵工艺的发展趋势,旨在为有效提高发酵菌株的底物代谢能力,获得高的乙醇产量提供重要参考。     

Enhanced production of L-(+)-lactic acid in chemostat by Lactobacillus casei DSM 20011 using ion-exchange resins and cross-flow filtration in a fully automated pilot plant controlled via NIR

Due to the lack of suitable in-process sensors, on-line monitoring of fermentation processes is restricted almost exclusively to the measurement of physical parameters only indirectly related to key process variables, i.e., substrate, product, and biomass concentration. This obstacle can be overcome by near infrared (NIR) spectroscopy, which allows not only real-time process monitoring, but also automated process control, provided that NIR-generated information is fed to a suitable computerized bioreactor control system. Once the relevant calibrations have been obtained, substrate, biomass and product concentration can be evaluated on-line and used by the bioreactor control system to manage the fermentation. In this work, an NIR-based control system allowed the full automation of a small-scale pilot plant for lactic acid production and provided an excellent tool for process optimization. The growth-inhibiting effect of lactic acid present in the culture broth is enhanced when the growth-limiting substrate, glucose, is also present at relatively high concentrations. Both combined factors can result in a severe reduction of the performance of the lactate production process. A dedicated software enabling on-line NIR data acquisition and reduction, and automated process management through feed addition, culture removal and/or product recovery by microfiltration was developed in order to allow the implementation of continuous fermentation processes with recycling of culture medium and cell recycling. Both operation modes were tested at different dilution rates and the respective cultivation parameters observed were compared with those obtained in a conventional continuous fermentation. Steady states were obtained in both modes with high performance on lactate production. The highest lactate volumetric productivity, 138 g L(-1) h(-1), was obtained in continuous fermentation with cell recycling.     

An on-line approach to monitor ethanol fermentation using FTIR spectroscopy

Fermentation process control is currently limited by its inability to measure parameters such as substrate, product, and biomass concentrations rapidly for consistent on-line feedback. Physical and chemical parameters, such as temperature and pH, currently can be obtained on-line using appropriate sensors. However, to obtain information on the concentration of the substrate, product, and biomass, samples must be taken off-line for measurement. With the use of spectroscopic techniques, real-time monitoring of process constituents such as product and substrate is possible. Spectroscopic techniques are rapid and nondestructive, require minimal or no sample preparation, and can be used to simultaneously assess several constituents in complex matrices. The production of ethanol is the largest fermentation process in terms of production volume and economic value as a result of its prominence in the food, agricultural, and fuel industries. This study attempts to develop an on-line ethanol fermentation monitoring technique using Fourier transform infrared (FTIR) spectroscopy with a flow-through ATR capability. Models developed using multivariate statistics, employed to obtain on-line FTIR measurements, were successfully validated by off-line HPLC analysis and spectrophotometry data. Standard errors of prediction (SEP) values of 0.985 g/L (R2 = 0.996), 1.386 g/L (R2 = 0.998), and 0.546 (R2 = 0.972) were obtained for ethanol, glucose, and OD, respectively. This work demonstrates that FTIR spectroscopy could be used for rapid on-line monitoring of fermentation.     

Multirate state and parameter estimation in an antibiotic fermentation with delayed measurements

This article discusses issues related to estimation and monitoring of fermentation processes that exhibit endogenous metabolism and time-varying maintenance activity. Such culture-related activities hamper the use of traditional, software sensor-based algorithms, such as the extended kalman filter (EKF). In the approach presented here, the individual effects of the endogenous decay and the true maintenance processes have been lumped to represent a modified maintenance coefficient, m(c). Model equations that relate measurable process outputs, such as the carbon dioxide evolution rate (CER) and biomass, to the observable process parameters (such as net specific growth rate and the modified maintenance coefficient) are proposed. These model equations are used in an estimator that can formally accommodate delayed, infrequent measurements of the culture states (such as the biomass) as well as frequent, culture-related secondary measurements (such as the CER). The resulting multirate software sensor-based estimation strategy is used to monitor biomass profiles as well as profiles of critical fermentation parameters, such as the specific growth for a fed-batch fermentation of Streptomyces clavuligerus. (c) 1994 John Wiley & Sons, Inc.     

响应面分析优化L-色氨酸清液发酵培养基

L-色氨酸是八种必需氨基酸之一,随着L-色氨酸应用市场的不断扩大,进行发酵法生产L-色氨酸的研究具有重要的现实意义。为了提高L-色氨酸产量,本文利用响应面分析法对L-色氨酸清液发酵培养基进行优化。利用优化培养基进行发酵,考察清液发酵对L-色氨酸发酵过程中生物量、L-色氨酸产量、副产物生成量的影响。结果表明:在优化条件下利用清液发酵培养基发酵,乙酸含量与原工艺相比降低了(6.75±1.26)%,L-色氨酸产量提高了(16.54±1.15)%,实验值与响应面分析预测值基本相符。     

Rapid estimation of glucose and amylase in fermentation culture broth with test strips by reflectance photometry

Summary The applicability of clinical multilayer test strips based on dry reagent chemistries for the quantitative analysis of fermentation media constituents was evaluated. For glucose and amylase determinations a significant correlation to conventional wet chemical methods was obtained. Bacterial biomass in the range of 0.5–5 g/l has no interfering effect. It is a rapid and cost-effective method for fermentation process monitoring and control.     

Determination of reactor biomass by acoustic resonance densitometry

Acoustic resonance densitometry (ARD) is reported as a method suitable not only for precise investigations into changes of specific gravity in bioreactor media but also as a technique able to provide an accurate wide range and direct determination of cellular mass in fermentation processes. It is further shown that this method can replace present optical procedures, minimizing dilution errors and operator involvement and is suitable for development as an on-line biomass monitoring system.