Efficiency of repeated batch penicillin fermentation using two fermentors. Computer simulation and optimisation

Repeated batch operation using two fermentors (RBTF) to penicillin fermentation was demonstrated by computer simulation to improve productivity. Three operation modes were compared: chemostat, repeated batch operation using a single fermentor (RBSF) and RBTF; in each case account was made of the lag period before growth. The simulated fermentor performances were assessed on the basis of the penicillin productivity and concentration; the simulation was based on published batch fermentation data. It was shown that RBTF was superior to RBSF and chemostat. The advantage of RBTF increased as the lag period became greater.     

A method for multiphase batch process monitoring based on auto phase identification

Batch processes with multiple phases are commonly found in process industries. Process dynamics and correlations among variables also tend to change with the transitions across such phases. Traditional approaches where the model is constructed from data representing the whole batch process would not be sufficient to capture the varying process dynamics and correlation structure. Different ways of phase segmentation and modeling strategies have been previously reported that account for the multi-phase characteristics of such processes. For a given process dynamic or a particular phase in a multi-phase process, a Principal Component Analysis (PCA) model can project the maximum deviation with small number of principal components, and represent a certain percentage of the deviation with fixed number of principal components. As the process dynamics change, the percentage represented by the fixed number of PCs also changes if a single PCA model is applied. In this paper, a new phase identification method is proposed based on the change of the first cumulative contribution between different PCA models. Every phase is modeled separately based on the phase identification. The method is applied to fault detection in the fed-batch penicillin cultivation process. The results show that the method can better capture the process dynamics in different phases and detect process upset in an early stage.     

青霉素酰化酶基因工程菌 E.coli A_(56)(pPA22)水解青霉素 G 的动力学

用初速度法测定了青霉素酰化酶基因工程菌 E.coli A_(56)(pPA22)游离细胞和固定化细胞水解青霉素 G 的动力学常数,并进行了动力学模型的检验。该菌体酶活高,米氏常数小,固定化后米氏常数和苯乙酸抑制常数增大,底物抑制常数和6-APA 抑制常数没有变化。通过青霉素 G 裂解过程的考察,验证了青霉裂解动力学模型的适用性及动力学常数测定的可靠性。     

蜡状芽孢杆菌ZJB-07112酰胺酶的分离纯化及其酶学性质

用一株蜡状芽孢杆菌新菌株ZJB-07112 (Bacillus cereus ZJB-07112)发酵生产酰胺酶,经超声波细胞破碎、High Q阴离子色谱、Phenyl-Sepharose疏水色谱等步骤获得了凝胶电泳均一的酰胺酶。用12.5% SDS-PAGE测得该酶的分子量约为60.6 kD。其N端氨基酸序列为ATIRPDDKAI。该酶水解反应的最适pH和最适温度分别为7.5和35 ℃。在pH 7.5条件下,该酶在50 ℃以上容易失活,60 ℃保温30 min后,仅保留10.8%的酶活。除了Hg⁺ 、Ag⁺等重金属离子和尿素外,其他金属离子和EDTA对该酶的活性影响不大。以丙烯酰胺为底物时,该酶的K_m和V_max值分别为2.64 mmol·L^-1和0.6 μmol·min^-1·ml^-1。     

青霉素酰化酶的分离纯化和固定化研究新进展

青霉素酰化酶又称为青霉素酰胺酶或青霉素氨基水解酶,主要从大肠埃希菌胞内酶和巨大芽孢杆菌胞外酶获得,该酶已大规模应用于工业生产β-内酰胺类抗生素的关键中间体和半合成β-内酰胺类抗生素。主要介绍了青霉素酰化酶固定化技术的进展,讨论了不同固定化技术的特点,并展望了固定化青霉素酰化酶的发展前景。     

Nanocrystalline-diamond thin films with high pH and penicillin sensitivity prepared on a capacitive Si-SiO2 structure

A capacitive field-effect EDIS (electrolyte-diamond-insulator-semiconductor) sensor with improved pH and penicillin sensitivity has been realised using a nanocrystalline-diamond (NCD) film as sensitive gate material. The NCD growth process on SiO₂ as well as an additional surface treatment in oxidising medium have been optimised to provide high pH-sensitive, non-porous O-terminated films without damage of the underlying SiO₂ layer. The surface morphology of O-terminated NCD thin films and the layer structure of EDIS sensors have been studied by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) methods. To establish the relative coverage of the surface functional groups generated by the oxidation of NCD surfaces, X-ray photoelectron spectroscopy analysis was carried out. The hydrophilicity of NCD thin films has been studied by water contact-angle measurements. A nearly Nernstian pH sensitivity of 54-57 mV/pH has been observed for O-terminated NCD films treated in an oxidising boiling mixture for 80 min and in oxygen plasma. The high pH-sensitive properties of O-terminated NCD have been used to develop an EDIS-based penicillin biosensor. A freshly prepared penicillin biosensor possesses a high sensitivity of 85 mV/decade in the concentration range of 0.1-2.5 mM penicillin G. The lower detection limit is 5 μM.     

超高效液相色谱串联质谱法测定乳制品中 10种青霉素类抗生素残留

目的建立同时测定乳制品中10种青霉素类抗生素残留的超高效液相色谱-串联质谱分析方法。方法样品经乙腈提取后,乙腈饱和正己烷除去脂肪,经0.22μm微孔滤膜过滤,用乙腈-水(含0.1%甲酸5mmol/L乙酸铵)为流动相,C18柱分离,20 min内梯度洗脱分离10种青霉素;电喷雾正离子模式电离(ESI+),多反应监测模式检测(MRM),外标法定量。结果在0.1~20μg/L浓度范围内,10种青霉素类药物在各种乳制品基质中均有良好的线性关系,线性相关系数均0.999;液体乳最低检测限为0.2μg/kg~1.0μg/kg,乳粉的最低检测限1.0μg/kg~5.0μg/kg;方法回收率在70.2%~108.2%,相对标准差为15%。结论该方法测定乳制品中10种青霉素药物的残留量简便、快速、准确,可以满足对青霉素类抗生素的监测要求。     

Extraction equilibria of penicillin G in four different types of organic solvent systems

Equilibrium experiments for physical and reactive extraction of penicillin G were carried out with varying pH, overall penicillin concentration and overall amine concentration in liquid–liquid extraction systems with four different types of organic solvents, i.e. amphiprotic solvent, hydrogen-bond donor solvent, hydrogen-bond acceptor solvent, and nonpolar organic solvent. Also, physical and reactive extraction equilibrium models were suggested to find the appropriate equilibrium model for each organic solvent type. By comparison between the experimental data and the calculated results from the extraction equilibrium models, the most probable physical and reactive extraction equilibrium models were determined. The physical extraction equilibrium models for all types of the organic solvents except for nonpolar organic solvent reflected solute–solvent interaction, while the reactive extraction equilibrium models were dependent on the organic solvent type.     

Improvement of the catalytic performance of immobilized penicillin acylase through assembly of macromolecular reagents in nanopore to create a crowding environment

Macromolecular reagents were co-assembled with penicillin acylase (PA) and immobilized in mesocellular siliceous foams (MCFs) to resemble living cells. Types and concentrations of macromolecules were studied. The catalytic characteristic and stability of PA preparations were also investigated. PA assembled with dextran 10 k in MCFs showed maximum specific activity, 1.32-fold of that of the solely immobilized PA. The optimum pH of dextran and BSA derivatives shifted to neutrality, and the optimum temperature increased by 10 °C. Also, Km of BSA derivative of PA declined 56.7% compared to solely immobilized PA, while the Kcat/Km of PA assembled with BSA was enhanced to 147%. After incubation at 50 °C for 6 h, residual activity of PA assembled with BSA exhibited 53.0%. The ficoll derivative showed 82.8% of its initial activity at 4 °C after 8-week storage. The results indicated that macromolecular reagents assembled with PA in MCFs could dramatically improve the catalytic performance and stability of immobilized enzyme.     

磁性微粒对青霉素G酰化酶的固定化研究

将无机磁性粒子Fe3O4与有机材料海藻酸钠结合起来制成一种复合的磁性微粒,并将其进行表面修饰,通过化学共价法来固定青霉素G酰化酶。通过扫描电镜等对微粒进行形态学观察,并用傅立叶红外图谱表征微粒表面修饰基团。酶学性质研究表明,该微粒固定化的青霉素G酰化酶的最适pH值为7.5,最适温度为40℃。固定化酶与底物的亲和力有所降低,但是稳定性显著提高。重复催化研究结果表明,固定化酶具有比游离酶更广泛的温度及pH值适用范围,并且具有良好的热稳定性、可循环使用性和贮存稳定性。