Process analytical technology (PAT) tools for the cultivation step in biopharmaceutical production

The process analytical technology (PAT) initiative is now 10 years old. This has resulted in the development of many tools and software packages dedicated to PAT application on pharmaceutical processes. However, most applications are restricted to small molecule drugs, mainly for the relatively simple process steps like drying or tableting where only a limited number of parameters need to be controlled. A big challenge for PAT still lies in applications for biopharmaceuticals and then especially in the cultivation process step, where the quality of a biopharmaceutical product is largely determined. This review gives an overview of the currently available tools for monitoring and controlling the biopharmaceutical cultivation step and of the main challenges for the most common cell platforms (i.e. Escherichia coli, yeast, and mammalian cells) used in biopharmaceutical manufacturing. The real challenge is to understand how intracellular mechanisms (from synthesis to excretion) influence the quality of biopharmaceuticals and how these mechanisms can be monitored and controlled to yield the desired end product quality. Modern “omics” tools and advanced process analyzers have opened up the way for PAT applications for the biopharmaceutical cultivation process step.     

Evaluation of Electron Donor Materials for Solution‐Processed Organic Solar Cells via a Novel Figure of Merit

Organic photovoltaic (OPV) technology offers many advantages, although no commercial applications have been achieved after more than a decade of intensive research and development. Several challenges have yet to be overcome including high power conversion efficiency (PCE), good processability, low cost, and excellent long‐term stability, and so on. In this article, these fundamental challenges are significantly addressed by surveying and analyzing a new merit factor for material applied accessibility containing three parameters: synthetic complexity, device efficiency, and photostability. Thirty‐five donor small molecules are introduced to assess their synthetic accessibility. Furthermore, the PCEs and device photostability of these molecules are carried out, and further measured under one sun illumination within 200 h, respectively. Combining with the characteristics of these three factors, investigated molecules are ranked according to an industrial figure of merit (i‐FOM), while some guidelines for the material design and synthesis are given. It is suggested that a PCE of >14% and an i‐FOM of >20% via active material engineering are realistic for possible industry future of OPV. Along with the systematic study, it is believed that this i‐FOM can be taken into consideration at an early stage of molecular design and provides valuable insight for efficient evaluation of photovoltaic materials for possible commercial applications.     

Status of protein engineering for biocatalysts: how to design an industrially useful biocatalyst

Recent advances in the development of both experimental and computational protein engineering tools have enabled a number of further successes in the development of biocatalysts ready for large-scale applications. Key tools are first, the targeting of libraries, leading to far smaller but more useful libraries than in the past, second, the combination of structural, mechanistic, and sequence-based knowledge often based on prior successful cases, and third, the advent of structurally based algorithms allowing the design of novel functions. Based on these tools, a number of improved biocatalysts for pharmaceutical applications have been presented, such as an (R)-transaminase for the synthesis of active pharmaceutical ingredients (APIs) of sitagliptin (Januvia?) and ketoreductases, glucose dehydrogenases, and haloalkane dehalogenases for the API synthesis toward atorvastatin (Lipitor?) and montelukast (Singulair?).     

Acoustic-resonance spectrometry as a process analytical technology for the quantification of active pharmaceutical ingredient in semi-solids

The purpose of this study was to demonstrate acoustic resonance spectrometry (ARS) as an alternative process analytical technology to near infrared (NIR) spectroscopy for the quantification of active pharmaceutical ingradient (API) in semi-solids such as creams, gels, ointments, and lotions. The ARS used for this research was an inexpensive instrument constructed from readily available parts. Acoustic-resonance spectra were collected with a frequency spectrum from 0 to 22.05 KHz. NIR data were collected from 1100 to 2500 nm. Using 1-point net analyte signal (NAS) calibration, NIR for the API (colloidal oatmeal [CO]) gave anr ² prediction accuracy of 0.971, and a standard error of performance (SEP) of 0.517%CO. ARS for the API resulted in anr ² of 0.983 and SEP of 0.317%CO. NAS calibration is compared with principal component regression. This research demonstrates that ARS can sometimes outperform NIR spectrometry and can be an effective analytical method for the quantification of API in semi-solids. ARS requires no sample preparation, provides larger penetration depths into lotions than optical techniques, and measures API concentrations faster and more accurately. These results suggest that ARS is a useful process analytical technology (PAT). Published: July 14, 2006     

The NYC native air sampling pilot project: using HVAC filter data for urban biological incident characterization

Native air sampling (NAS) is distinguished from dedicated air sampling (DAS) devices (eg, BioWatch) that are deployed to detect aerosol disseminations of biological threat agents. NAS uses filter samples from heating, ventilation, and air conditioning (HVAC) systems in commercial properties for environmental sampling after DAS detection of biological threat agent incidents. It represents an untapped, scientifically sound, efficient, widely distributed, and comparably inexpensive resource for postevent environmental sampling. Calculations predict that postevent NAS would be more efficient than environmental surface sampling by orders of magnitude. HVAC filter samples could be collected from pre-identified surrounding NAS facilities to corroborate the DAS alarm and delineate the path taken by the bioaerosol plume. The New York City (NYC) Native Air Sampling Pilot Project explored whether native air sampling would be acceptable to private sector stakeholders and could be implemented successfully in NYC. Building trade associations facilitated outreach to and discussions with property owners and managers, who expedited contact with building managers of candidate NAS properties that they managed or owned. Nominal NAS building requirements were determined; procedures to identify and evaluate candidate NAS facilities were developed; data collection tools and other resources were designed and used to expedite candidate NAS building selection and evaluation in Manhattan; and exemplar environmental sampling playbooks for emergency responders were completed. In this sample, modern buildings with single or few corporate tenants were the best NAS candidate facilities. The Pilot Project successfully demonstrated that in one urban setting a native air sampling strategy could be implemented with effective public-private collaboration.     

Automatable production of shippable bilayer chips by pin tool deposition for an ion channel measurement platform

As a step towards an automated and operator-free ion channel measurement platform we have previously demonstrated a solution formulation for artificial lipid bilayers that enabled the indefinite storage and shipping of frozen bilayer precursors. In this work, the solutions were deposited by hand. Here, we have adapted pin tools to deposit the bilayer precursor solutions onto multi-element arrays, a popular method for microarray solution deposition. The pin tools have enabled the deposited volume to be applied highly repeatably and controllably, resulting in reduction of bilayer formation times to <1 h. The pin tools are also compatible with computerized motion control platforms, enabling automated and high throughput production. We discuss these results and the prospects of this technology to produce high density bilayer arrays for high throughput measurement of ion channels incorporated into artificial bilayers.     

Isozyme Analysis and Soluble Mycelial Protein Pattern in Iranian Isolates of Several formae speciales of Fusarium oxysporum

A total of 13 representative isolates of Fusarium oxysporum f. sp. melonis (FOM) from Iran, USA and France, eight isolates of seven formae speciales from Iran and one isolate of F. oxysporum f. sp. niveum from the USA were compared based on isozyme analysis and soluble mycelial protein pattern. Isozyme analyses of alkaline phosphatase (ALP), catalase (CAT), esterase (EST), malate dehydrogenase (MDH), superoxide dismutase (SOD) and xanthine dehydrogenase (XDH) revealed polymorphism among the F. oxysporum isolates in which 22 electrophoretic phenotypes (EP) were determined. At least 10 putative loci for these six enzymes were detected and they were all polymorphic. Maximum genetic diversity was observed in CAT, EST and XDH loci. Using UPGMA, the 22 isolates were separated into three main groups with one of the groups divided into two subgroups. Group I included isolates belonging to five formae speciales from Iran, whereas group II that included FOM isolates from both Iran and the USA was divided into two subgroups each containing the vast majority of the respective isolates from either country. Group III constituted FOM isolates from France and one pathogenic isolate on pepper from Iran. FOM isolates representing five different geographical regions from Iran belonged to two different races of 1 and 1,2Y and one vegetative compatibility group (VCG)0134 and thus were genetically homologous. Isozyme polymorphism in these isolates was highly correlated with VCG and geographical origins and to a lesser extent with races. Variations in soluble protein profile in FOM isolates were correlated with genetic distances determined in isozyme analysis. This study suggests that isozyme analysis could be a useful tool for identifying genetic diversity not only in FOM but also several formae speciales of F. oxysporum.     

Developing transgenic Anopheles mosquitoes for the sterile insect technique

In the last 10 years the availability of the genome sequence of Anopheles gambiae and the development of a transgenic technology for several species of Anopheles mosquitoes have, in combination, helped in enabling us to gain several insights into the biology of these mosquitoes that is relevant to their capacity as vectors of the malaria parasite. While this information is anticipated to inform many novel vector control strategies, the technique most likely to benefit in the near future from the availability of a reliable transgenic technology is the sterile insect technique (SIT), which relies on releasing large numbers of sterile insects to compete for mates in the wild, leading to population suppression. Although SIT has been proven to work reliably for many insects, the construction of suitable strains, and induction of sterility, has until now been a laborious process, combining classical genetics with radiation-induced sterility. Using transgenesis to create strains of Anopheles suitable for SIT could potentially offer several advantages over current approaches, in that the basic design of transgenic constructs designed for other insects should be rapidly transferable to mosquitoes, and induction of sterility as a product of the transgenic modification could obviate the requirement for radiation and its associated deleterious effects. In this paper the progress of different transgenic approaches in constructing tools for SIT will be reviewed.     

实施药事精准化监管 切实保障医疗质量与安全

精准化管理作为一种行之有效的管理理念与模式,正在被逐步运用于医院管理领域。通过建立药事数据分析平台、利用信息化手段对关键流程进行控制、实施全流程药品质量无缝监管及药物不良反应上报流程标准化,对药事管理信息进行收集、统计、分析,对药学服务流程进行优化整合,并形成戴明环（PDCA循环）管理机制,切实保障医疗质量与安全。     

Calculation of the"Net Additions to Stock" Indicator for the Czech Republic Using a Direct Method

Abstract: Net additions to stock (NAS) are an indicator based on economy-wide material flow accounting and analysis. NAS, a measure of the physical growth rate of an economy, can be used for estimates of future waste flows. It is calculated using two methods: The indirect method of calculation is a simple difference between all input and output flows, whereas the direct method involves measuring the amounts of materials added to particular categories of physical stock and the amounts of waste flows from these stocks.
The study described in this article had one leading objective: to make available direct NAS data for the Czech Republic, which could later be used for predicting future waste flows. Two additional objectives emerged from the first: (1) to develop a method for direct NAS calculation from data availability in the Czech Republic; (2) to calculate NAS directly, compare the results with those achieved in indirect NAS calculation, and discuss the identified differences.
The NAS for the Czech Republic calculated by the direct method is equal to approximately 65 million tonnes on average in 2000–2002 and is approximately 27% lower than the NAS acquired by the indirect method of calculation. The actual values of directly calculated NAS and its uncertainties suggest that the indirect NAS is more likely to be an overestimation than an underestimation. Durables account for about 2% of the total direct NAS, whereas the rest is attributed to infrastructure and buildings. The direct NAS is dominated by nonmetal construction commodities such as building stone and bricks, which equal approximately 89% of the total direct NAS.
Calculation of NAS by the direct method has been proved to be feasible in the Czech Republic. Moreover, uncertainties related to direct NAS are lower than those related to indirectly acquired NAS.     

Factorial and time course designs for cDNA microarray experiments

Microarrays are powerful tools for surveying the expression levels of many thousands of genes simultaneously. They belong to the new genomics technologies which have important applications in the biological, agricultural and pharmaceutical sciences. There are myriad sources of uncertainty in microarray experiments, and rigorous experimental design is essential for fully realizing the potential of these valuable resources. Two questions frequently asked by biologists on the brink of conducting cDNA or two-colour, spotted microarray experiments are 'Which mRNA samples should be competitively hybridized together on the same slide?' and 'How many times should each slide be replicated?' Early experience has shown that whilst the field of classical experimental design has much to offer this emerging multi-disciplinary area, new approaches which accommodate features specific to the microarray context are needed. In this paper, we propose optimal designs for factorial and time course experiments, which are special designs arising quite frequently in microarray experimentation. Our criterion for optimality is statistical efficiency based on a new notion of admissible designs; our approach enables efficient designs to be selected subject to the information available on the effects of most interest to biologists, the number of arrays available for the experiment, and other resource or practical constraints, including limitations on the amount of mRNA probe. We show that our designs are superior to both the popular reference designs, which are highly inefficient, and to designs incorporating all possible direct pairwise comparisons. Moreover, our proposed designs represent a substantial practical improvement over classical experimental designs which work in terms of standard interactions and main effects. The latter do not provide a basis for meaningful inference on the effects of most interest to biologists, nor make the most efficient use of valuable and limited resources.     

A DNA biochip for on-the-spot multiplexed pathogen identification

Miniaturized integrated DNA analysis systems have largely been based on a multi-chamber design with microfluidic control to process the sample sequentially from one module to another. This microchip design in connection with optics involved hinders the deployment of this technology for point-of-care applications. In this work, we demonstrate the implementation of sample preparation, DNA amplification, and electrochemical detection in a single silicon and glass-based microchamber and its application for the multiplexed detection of Escherichia coli and Bacillus subtilis cells. The microdevice has a thin-film heater and temperature sensor patterned on the silicon substrate. An array of indium tin oxide (ITO) electrodes was constructed within the microchamber as the transduction element. Oligonucleotide probes specific to the target amplicons are individually positioned at each ITO surface by electrochemical copolymerization of pyrrole and pyrrole−probe conjugate. These immobilized probes were stable to the thermal cycling process and were highly selective. The DNA-based identification of the two model pathogens involved a number of steps including a thermal lysis step, magnetic particle-based isolation of the target genomes, asymmetric PCR, and electrochemical sequence-specific detection using silver-enhanced gold nanoparticles. The microchamber platform described here offers a cost-effective and sample-to-answer technology for on-site monitoring of multiple pathogens.     

Developing a Hybrid Programmable Logic Controller Platform for a Flexible Manufacturing System

In this article, we present the design and implementation of a flexible manufacturing system (FMS) control platform based on a programmable logic controller (PLC) and a personal computer (PC)-based visual man-machine interface (MMI) and data acquisition (DAS) unit. The key aspect of an FMS is its flexibility to adapt to changes in a demanding process operation. The PLC provides feasible solutions to FMS applications, using PC-based MMI/DAS, whereby PLCs are optimized for executing rapid sequential control strategies. PCs running MMI/DAS front-ends make intuitive operation interfaces, full of powerful graphics and reporting tools. Information from the PC can be distributed through a company's local area network or web using client-server technologies. Currently, with the convergence of underlying microprocessor technology and software programming techniques, many users find that PLCs provide a cost-effective solution to real-time control in small- to medium-sized process plants, especially when combined with supervisory PCs using hybrid systems. The major work of this article demonstrates that PLCs are responsive to rapid and repetitious control tasks, using PCs that present the flow of information automation and accept operator instructions, thereby providing the user a tool to modify and monitor the process as the requirements change.     

Impact of Fermented Mulberry Leaf and Fish Offal in Diet Formulation of Indian Major Carp (Labeo rohita)

Large quantities of fish offal and mulberry leaf are generated globally. The present study aimed to understand their potential utilization in aqua diet formulation, after proper fermentation, as raw materials to replace fish meal in Indian major carp (Labeo rohita) compounded diet. Fish offal meal (FOM) and mulberry leaf meal (MLM) were used in a 2 × 3 factorial design, to evaluate (i) two different fermented mixtures with the inclusion of both FOM and MLM or only MLM and (ii) to replace three different level of dietary fishmeal: 50, 75 or 80 %. An indoor trial, to evaluate diet intake and digestibility and an outdoor trial to evaluate growth performances were impended in Indian major carp fingerlings. The results showed that FOM and MLM are promising raw materials that can be successfully used in the formulation of diet for the Indian major carp. Specifically, the addition of a proper amount of MLM in the fermentation of FOM produced a fermented mixture that could successfully replace up to 80 % of FM in the diet formulation.     

Alchemical free energy methods for drug discovery: progress and challenges

Improved rational drug design methods are needed to lower the cost and increase the success rate of drug discovery and development. Alchemical binding free energy calculations, one potential tool for rational design, have progressed rapidly over the past decade, but still fall short of providing robust tools for pharmaceutical engineering. Recent studies, especially on model receptor systems, have clarified many of the challenges that must be overcome for robust predictions of binding affinity to be useful in rational design. In this review, inspired by a recent joint academic/industry meeting organized by the authors, we discuss these challenges and suggest a number of promising approaches for overcoming them.     

High throughput proteome screening for biomarker detection

Mass spectrometry-based quantitative proteomics has become an important component of biological and clinical research. Current methods, while highly developed and powerful, are falling short of their goal of routinely analyzing whole proteomes mainly because the wealth of proteomic information accumulated from prior studies is not used for the planning or interpretation of present experiments. The consequence of this situation is that in every proteomic experiment the proteome is rediscovered. In this report we describe an approach for quantitative proteomics that builds on the extensive prior knowledge of proteomes and a platform for the implementation of the method. The method is based on the selection and chemical synthesis of isotopically labeled reference peptides that uniquely identify a particular protein and the addition of a panel of such peptides to the sample mixture consisting of tryptic peptides from the proteome in question. The platform consists of a peptide separation module for the generation of ordered peptide arrays from the combined peptide sample on the sample plate of a MALDI mass spectrometer, a high throughput MALDI-TOF/TOF mass spectrometer, and a suite of software tools for the selective analysis of the targeted peptides and the interpretation of the results. Applying the method to the analysis of the human blood serum proteome we demonstrate the feasibility of using mass spectrometry-based proteomics as a high throughput screening technology for the detection and quantification of targeted proteins in a complex system.     

Consolidated strategy for the analysis of microarray spike-in data

As the number of users of microarray technology continues to grow, so does the importance of platform assessments and comparisons. Spike-in experiments have been successfully used for internal technology assessments by microarray manufacturers and for comparisons of competing data analysis approaches. The microarray literature is saturated with statistical assessments based on spike-in experiment data. Unfortunately, the statistical assessments vary widely and are applicable only in specific cases. This has introduced confusion into the debate over best practices with regards to which platform, protocols and data analysis tools are best. Furthermore, cross-platform comparisons have proven difficult because reported concentrations are not comparable. In this article, we introduce two new spike-in experiments, present a novel statistical solution that enables cross-platform comparisons, and propose a comprehensive procedure for assessments based on spike-in experiments. The ideas are implemented in a user friendly Bioconductor package: spkTools. We demonstrated the utility of our tools by presenting the first spike-in-based comparison of the three major platforms–Affymetrix, Agilent and Illumina.