Extraction of penicillin G from simulated media by an emulsion liquid membrane process

The extraction of penicillin G from simulated media was performed by water/oil/water (w/o/w) emulsion liquid membranes (ELMs) and studied under various operational conditions in a batch system. The degree of extraction achieved was between 80% and 95% under specific conditions. A concentration of greater than nine times the initial concentration of penicillin G in the external phase was obtained in the internal phase. The pH of the internal aqueous solution, containing a basic salt, was theoretically calculated on the basis of the amount of penicillin G transported into the internal phase. The calculated results agreed with the experimental data well and were used to select a suitable type and concentration of a basic salt in the internal phase to give a pH within the range 5 to 8 where penicillin G was stable after the termination of extraction. The extraction of penicillin G was successfully performed by the ELM process with sodium carbonate in the internal phase.     

Bioprocess applications of model-based estimation techniques

The last decade has seen the development of a number of approaches for estimating those variables which are difficult to measure on-line in industrial process situations. Whilst a range of techniques is available, a common element is the use of process knowledge in the form of a system model. In the case of bioprocess systems, although a large range of models has been presented in the literature, their use in estimation schemes on an industrial scale has been limited. A number of reasons can be identified for their low level of utilisation. Of particular significance is the uncertainty which exists in quantifying system performance and the process-model mismatch which inevitably results. The level of ‘pre-defined model’ uncertainty, together with the knowledge gained during the course of the fermentation, serves to dictate estimator structure. The paper considers a range of estimation strategies and contrasts, through industrial applications, their performance characteristics and utility.     

An integrated downstream processing strategy for the recovery and partial purification of penicillin acylase from crude media

An integrated process strategy for the recovery of penicillin acylase was developed, based on precipitation of non‐enzymatic proteins directly from Escherichia coli homogenates or crude extracts using Rolquat (quaternary ammonium salt) and adsorption of the enzyme on Amp‐Seph (3.8 µmole ampicillin cm^?3) under pseudo‐affinity conditions. The effect of pH, concentrations of ammonium sulfate and Rolquat, and also concentrations of protein and cell debris on the precipitation of non‐enzymatic proteins from homogenates and crude extracts of penicillin acylase were analysed. The method of addition of Rolquat to homogenates and crude extracts significantly influenced the size of the precipitated particles. Improved results on the specific activity of penicillin acylase were obtained for 22% and 1% (w/v) of ammonium sulfate and Rolquat, respectively, added sequentially to enzyme solutions and at room temperature. Under these experimental conditions, the specific activity of penicillin acylase in homogenates and crude extracts was enhanced 2.5–3.0‐fold. Finally, the integrated process strategy was implemented first by precipitation of non‐enzymatic proteins and recovery of penicillin acylase directly from the enzyme solution treated with Rolquat using an adsorption/filtration system with an overall yield of 86%. This system allows simultaneously the filtration of cell debris and fine precipitated particles, in situ recovery of penicillin acylase by its adsorption on Amp‐Seph, and selective desorption of the enzyme with a specific activity of 11 IU (mg prot)^?1 and a desorption yield of 95%. © 2002 Society of Chemical Industry     

Novel thermally and mechanically stable hydrogel for enzyme immobilization of penicillin G acylase via covalent technique

κ‐Carrageenan hydrogel crosslinked with protonated polyethyleneimine (PEI⁺) and glutaraldehyde (GA) was prepared and evaluated as a novel biocatalytic support for covalent immobilization of penicillin G acylase (PGA). The method of modification of the carrageenan biopolymer is clearly illustrated using a schematic diagram and was verified by FTIR, elemental analysis, DSC, and INSTRON using the compression mode. Results showed that the gels' mechanical strength was greatly enhanced from 3.9 kg/cm² to 16.8 kg/cm² with an outstanding improvement in the gels thermal stability. It was proven that, the control gels were completely dissolved at 35°C, whereas the modified gels remained intact at 90°C. The DSC thermogram revealed a shift in the endothermic band of water from 62 to 93°C showing more gel‐crosslinking. FTIR revealed the presence of the new functionality, aldehydic carbonyl group, at 1710 cm^?1 for covalent PGA immobilization. PGA was successfully immobilized as a model industrial enzyme retaining 71% of its activity. The enzyme loading increased from 2.2 U/g (control gel) to 10 U/g using the covalent technique. The operational stability showed no loss of activity after 20 cycles. The present support could be a good candidate for the immobilization of industrial enzymes rich in amino groups, especially the thermophilic ones. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

聚氨酯泡沫固定化重组枯草杆菌三段培养生产青霉素酰化酶

引　言青霉素G酰化酶 (penicillinGacylase ,EC3 5 1 11,PGA)是 β 内酰胺类抗生素工业中的关键酶之一 .目前主要采用大肠杆菌和巨大芽孢杆菌进行工业化生产 ,但由于产酶水平较低、变温发酵和需要苯乙酸诱导等缺点 ,国内外研究者尝试采用各种基因重组技术 ,以期大幅度提高产酶     

Structural and kinetic studies on ligand binding in wild-type and active-site mutants of penicillin acylase

Penicillin acylase catalyses the condensation of Calpha-substituted phenylacetic acids with beta-lactam nucleophiles, producing semi-synthetic beta-lactam antibiotics. For efficient synthesis a low affinity for phenylacetic acid and a high affinity for Calpha-substituted phenylacetic acid derivatives is desirable. We made three active site mutants, alphaF146Y, betaF24A and alphaF146Y/betaF24A, which all had a 2- to 10-fold higher affinity for Calpha-substituted compounds than wild-type enzyme. In addition, betaF24A had a 20-fold reduced affinity for phenylacetic acid. The molecular basis of the improved properties was investigated by X-ray crystallography. These studies showed that the higher affinity of alphaF146Y for (R)-alpha-methylphenylacetic acid can be explained by van der Waals interactions between alphaY146:OH and the Calpha-substituent. The betaF24A mutation causes an opening of the phenylacetic acid binding site. Only (R)-alpha-methylphenylacetic acid, but not phenylacetic acid, induces a conformation with the ligand tightly bound, explaining the weak binding of phenylacetic acid. A comparison of the betaF24A structure with other open conformations of penicillin acylase showed that betaF24 has a fixed position, whereas alphaF146 acts as a flexible lid on the binding site and reorients its position to achieve optimal substrate binding.     

城市污泥混合青霉素菌渣堆肥实验

为分析抗生素残留对菌渣堆肥过程的影响,以青霉素菌渣、市政脱水污泥和木屑为原料,研究好氧堆肥过程温度、碳素、氮素等理化参数的变化及青霉素的降解情况. 首先通过0.4 m³的大堆体研究菌渣好氧堆肥的可行性,然后通过5个6 L的小堆体详细研究菌渣污泥混合堆肥过程. 结果表明：菌渣污泥混合堆肥升温效果优于对照的单纯污泥. 堆肥菌渣质量分数与堆肥物料总有机碳（TOC）质量分数成正比,堆体的TOC质量分数随堆肥时间不断下降且趋于稳定,菌渣残留的青霉素未影响堆肥过程TOC的变化趋势. 投加菌渣增加了堆体的水溶性有机碳（WSOC）质量分数,生物可利用碳源的增加促进了堆肥过程微生物的转化作用,有助于堆肥过程温度的升高. 添加菌渣有助于减少堆肥原料中的氮素损失,且在第5天已检测不到菌渣中的抗生素残留. 可以通过混合堆肥实现菌渣的资源化利用及药物残留的分解.     

一种基于信噪比确定主元个数的方法

多段主元分析(MPCA)是针对间歇进行故障诊断一种行之有效的方法.在MPCA中主元个数的确定是模型的关键,关系到主元模型的可靠性、准确性、完备性.传统的累积方差贡献率(CPV)方法确定主元个数主观性较大并且没有考虑故障因素.为了提高检测性能,有效的提取主元,文中提出一种信噪比(SNR)与MPCA相结合选取间歇过程主元个数的方法,SNR表明的是故障诊断的灵敏度和主元个数的影响关系,在确保主元信息充分描述数据的基础上,该方法考虑了故障的信息对主元个数的影响来选取主元.将此方法应用于青霉素间歇发酵过程故障诊断中,仿真结果表明T2统计量和SPE统计量的响应曲线对故障更加敏感,有效地提高了故障诊断的准确率.     

混合神经网络建模方法在青霉素发酵过程中的应用

本文采用广义混合神经网络建模方法对青霉素发酵过程进行建模，根据青霉素发酵过程可在线测量过程参数二氧化碳生成率CER，实现青霉素发酵过程生物质浓度的测量。由仿真实验结果可以看出，该方法建模精度高、泛化能力强。     

青霉素发酵液平板膜超滤的操作条件研究

选定了超滤处理青霉素发酵液的流程．通过实验确定了平板膜实验装置超滤过程的操作条件，包括搅拌转速、操作压力、流量比等．测定了膜通量随操作时间和滤液体积的变化情况．