Androstenedione production by biotransformation of phytosterols

Androstenedione is a key intermediate of microbial steroid metabolism. It belongs to the 17-keto steroid family and is used as starting material for the preparation of different steroids. Androstenedione can be produced by microbial side chain cleavage of phytosterol, which is an alternative to multi-step chemical synthesis. In this review, various methods of biotransformation of sterols to androstenedione are surveyed. It begins with the history and current research status in this field. The existing methods of chemical and biochemical synthesis are examined. Various issues related to these methods and how researchers have addressed them is presented. Among these, the low solubility of sterols in aqueous systems is a critical problem since it limits the product yield. The main content of this review focuses on new methods of biotransformation that are being investigated. Recent biotechnological advances in this field are presented. The review ends with a note on future perspectives.     

Active extracellular substances of Bacillus thuringiensis ITRI‐G1 induce microalgae self‐disruption for microalgal biofuel

Microalgal cultures are a clean and sustainable means to use solar energy for CO₂ fixation and fuel production. Microalgae grow efficiently and are rich in oil, but recovering that oil is typically expensive and consumes much energy. Therefore, effective and low‐cost techniques for microalgal disruption and oil or lipid extraction are required by the algal biofuel industry. This study introduces a novel technique that uses active extracellular substances to induce microalgal cell disruption. A bacterium indigenous to Taiwan, Bacillus thuringiensis, was used to produce the active extracellular substances, which were volatile compounds with high thermal stability. Approximately 74% of fresh microalgal cells were disrupted after a 12‐h treatment with the active extracellular substances. Algal lipid extraction efficiency was improved and the oil extraction time was decreased by approximately 37.5% compared with the control treatment. The substances effectively disrupted fresh microalgal cells but not dehydrated microalgal cells. An analysis of microalgal DNA from fresh cells after disruption treatment demonstrated typical DNA laddering, indicating that disruption may have resulted from programmed cell death. This study revealed that biological treatments are environmentally friendly methods for increasing microalgal lipid extraction efficiency, and introduced a microalgal cell self‐disruption mechanism.     

Application of multivariate statistical techniques in microbial ecology

Recent advances in high‐throughput methods of molecular analyses have led to an explosion of studies generating large‐scale ecological data sets. In particular, noticeable effect has been attained in the field of microbial ecology, where new experimental approaches provided in‐depth assessments of the composition, functions and dynamic changes of complex microbial communities. Because even a single high‐throughput experiment produces large amount of data, powerful statistical techniques of multivariate analysis are well suited to analyse and interpret these data sets. Many different multivariate techniques are available, and often it is not clear which method should be applied to a particular data set. In this review, we describe and compare the most widely used multivariate statistical techniques including exploratory, interpretive and discriminatory procedures. We consider several important limitations and assumptions of these methods, and we present examples of how these approaches have been utilized in recent studies to provide insight into the ecology of the microbial world. Finally, we offer suggestions for the selection of appropriate methods based on the research question and data set structure.     

Challenges and opportunities for hydrogen production from microalgae

The global population is predicted to increase from ~7.3 billion to over 9 billion people by 2050. Together with rising economic growth, this is forecast to result in a 50% increase in fuel demand, which will have to be met while reducing carbon dioxide (CO₂) emissions by 50–80% to maintain social, political, energy and climate security. This tension between rising fuel demand and the requirement for rapid global decarbonization highlights the need to fast‐track the coordinated development and deployment of efficient cost‐effective renewable technologies for the production of CO₂ neutral energy. Currently, only 20% of global energy is provided as electricity, while 80% is provided as fuel. Hydrogen (H₂) is the most advanced CO₂‐free fuel and provides a ‘common’ energy currency as it can be produced via a range of renewable technologies, including photovoltaic (PV), wind, wave and biological systems such as microalgae, to power the next generation of H₂ fuel cells. Microalgae production systems for carbon‐based fuel (oil and ethanol) are now at the demonstration scale. This review focuses on evaluating the potential of microalgal technologies for the commercial production of solar‐driven H₂ from water. It summarizes key global technology drivers, the potential and theoretical limits of microalgal H₂ production systems, emerging strategies to engineer next‐generation systems and how these fit into an evolving H₂ economy.     

Advances in field-based studies of primate behavioral endocrinology

The papers in this issue were inspired by a symposium we organized for the 27th Annual Meeting of the American Society of Primatologists, held in June 2004. The purpose of the symposium was to highlight some of the new ways in which noninvasive techniques for measuring steroids are being employed to investigate the relationships between hormones and behavior in wild primates. Endocrinological data from the field provide new insights and more precise interpretations of behavioral data on topics ranging from social and reproductive strategies to seasonality, reproductive suppression, concealed ovulation, and development. Advances in the development of methods for extracting steroids from urine and feces have increased the options available to field workers, who can now select the most appropriate methodology for their study subjects, field conditions, and specific research questions. Field-based studies of behavioral endocrinology have spread to include a wide diversity of primate species. As the scope of these studies continues to grow, we anticipate the emergence of a dynamic new field in which comparative models of primate behavioral endocrinology contribute new perspectives on primates.     

Microalgal transformation of progesterone by the terrestrial-isolated cyanobacterium Microchaete tenera

A large number of microorganisms including various microalgal strains are able to convert steroid compounds into useful metabolites. In the present study, the ability of Microchaete tenera, a rice paddy field-isolated microalga, was investigated for biotransformation of progesterone. The incubation was carried out at 25°C under continuous illumination in the present of 0.25?g?L^?1 of progesterone. After 5?days incubation of the microalga in BG-11 liquid medium, the broth was extracted and the products were purified by the aid of chromatographic methods. Structure elucidation of the metabolites was performed by spectral data (¹³C NMR, ¹H NMR, FTIR, and MS) and physical constants (melting points and optical rotations). Eventually, four major steroids including 20β-hydroxypregn-4-en-3-one, 20α-hydroxypregn-4-en-3-one, 6β-hydroxypregn-4-en-3,20-dione and 6α-hydroxypregn-4-en-3,20-dione were the results of this biotransformation. The study also showed that the best concentration of starting material, temperature, photoregime, and the influence of CO₂ partial pressure on the production of bioconverted metabolites were 0.25?g?L^?1, 25°C, continuous light and 2.0?±?0.1% (v/v), respectively. Highest concentrations of all biotransformed metabolites were obtained in the 5th day.     

Approaches to Repigmentation of Vitiligo Skin: New Treatment with Ultrasonic Abrasion,Seed‐Grafting and Psoralen Plus Ultraviolet A Therapy

Vitiligo vulgaris is a common disease throughout the world although its pathogenesis is not yet known. The most frequent treatment used for vitiligo is PUVA (psoralen plus ultraviolet A) and topical steroids but against stable refractory vitiligo, various other surgical techniques have been developed such as autografting, epidermal grafting with suction blisters, epithelial sheet grafting, and transplantation of cultured melanocytes. We have discovered a new method using ultrasonic abrasion, seed‐grafting and PUVA therapy. The ultrasonic surgical aspirator abrades only the epidermis of recipient sites. This easily and safely removes only the epidermis, even on spotty lesions or intricate regions which are difficult to remove using a conventional motor‐driven grinder or liquid nitrogen. Epidermal seed‐grafting can cover more area than sheet‐grafting, and subsequent PUVA treatment can enlarge the area of pigmentation with coalescence of adjacent grafts. In this article, we provide a general overview of the current surgical therapies including our method for treating stable refractory vitiligo.     

Microalgae as bioreactors

Microalgae already serve as a major natural source of valuable macromolecules including carotenoids, long-chain polyunsaturated fatty acids and phycocolloids. As photoautotrophs, their simple growth requirements make these primitive plants potentially attractive bioreactor systems for the production of high-value heterologous proteins. The difficulty of producing stable transformants has meant that the field of transgenic microalgae is still in its infancy. Nonetheless, several species can now be routinely transformed and algal biotechnology companies have begun to explore the possibilities of synthesizing recombinant therapeutic proteins in microalgae and the engineering of metabolic pathways to produce increased levels of desirable compounds. In this review, we compare the current commercially viable bioreactor systems, outline recent progress in microalgal biotechnology and transformation, and discuss the potential of microalgae as bioreactors for the production of heterologous proteins.     

Effect of green microalgal extracts exhibiting antibacterial activity on viability of human malignant and non‐malignant cells

Microalgae have been investigated for their ability to produce metabolites that exhibit antibacterial activity. However, as research on antibacterial activity progresses, the effect of microalgal extracts on mammalian cells needs to be also assessed. The in vitro effect of microalgal extracts with demonstrated antibacterial activity against the human opportunistic pathogen Staphylococcus aureus was examined on the viability of non‐malignant (MCF10A and 184B5 cells) and malignant human cell lines (A2780 and MCF7). Direct cell counts indicated that the MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide) proliferation assay was found to under/overestimate cell viability when specific microalgal extracts and/or concentrations were tested. From direct cell counts, the viability of non‐malignant cells was not reduced after exposure to the extracts, whereas the viability of malignant cells was affected by specific microalgal extracts and concentrations. The results suggest that green microalgae are worthy of further investigation as potential sources of antibiotics, since they did not show a negative effect on the non‐malignant cell lines, MCF10A and 184B5. Additional studies should evaluate the compounds responsible for the anti‐proliferative activity of specific microalgal extracts observed on the malignant cells A2780 and MCF7.     

Manipulation of the microalgal chloroplast by genetic engineering for biotechnological utilization as a green biofactory

The chloroplast is an essential organelle in microalgae for conducting photosynthesis, thus enabling the photoautotrophic growth of microalgae. In addition to photosynthesis, the chloroplast is capable of various biochemical processes for the synthesis of proteins, lipids, carbohydrates, and terpenoids. Due to these attractive characteristics, there has been increasing interest in the biotechnological utilization of microalgal chloroplast as a sustainable alternative to the conventional production platforms used in industrial biotechnology. Since the first demonstration of microalgal chloroplast transformation, significant development has occurred over recent decades in the manipulation of microalgal chloroplasts through genetic engineering. In the present review, we describe the advantages of the microalgal chloroplast as a production platform for various bioproducts, including recombinant proteins and high-value metabolites, features of chloroplast genetic systems, and the development of transformation methods, which represent important factors for gene expression in the chloroplast. Furthermore, we address the expression of various recombinant proteins in the microalgal chloroplast through genetic engineering, including reporters, biopharmaceutical proteins, and industrial enzymes. Finally, we present many efforts and achievements in the production of high-value metabolites in the microalgal chloroplast through metabolic engineering. Based on these efforts and advances, the microalgal chloroplast represents an economically viable and sustainable platform for biotechnological applications in the near future.     

Imaging Carrier Transport Properties in Halide Perovskites using Time‐Resolved Optical Microscopy

Halide perovskites have remarkable properties for relatively crudely processed semiconductors, including large optical absorption coefficients and long charge carrier lifetimes. Thanks to such properties, these materials are now competing with established technologies for use in cost‐effective and efficient light‐harvesting and light‐emitting devices. Nevertheless, the fundamental understanding of the behavior of charge carriers in these materials—particularly on the nano‐ to microscale—has, on the whole, lagged behind empirical device performance. Such understanding is essential to control charge carriers, exploit new device structures, and push devices to their performance limits. Among other tools, optical microscopy and spectroscopic techniques have revealed rich information about charge carrier recombination and transport on important length scales. In this progress report, the contribution of time‐resolved optical microscopy techniques to the collective understanding of the photophysics of these materials is detailed. The ongoing technical developments in the field that are overcoming traditional experimental limitations in order to visualize transport properties over multiple time and length scales are discussed. Finally, strategies are proposed to combine optical microscopy with complementary techniques in order to obtain a holistic picture of local carrier photophysics in state‐of‐the‐art perovskite devices.     

Commercial production of microalgae: ponds, tanks, tubes and fermenters

The commercial culture of microalgae is now over 30 years old with the main microalgal species grown being Chlorella and Spirulina for health food, Dunaliella salina for β-carotene, Haematococcus pluvialis for astaxanthin and several species for aquaculture. The culture systems currently used to grow these algae are generally fairly unsophisticated. For example, Dunaliella salina is cultured in large (up to approx. 250 ha) shallow open-air ponds with no artificial mixing. Similarly, Chlorella and Spirulina also are grown outdoors in either paddle-wheel mixed ponds or circular ponds with a rotating mixing arm of up to about 1 ha in area per pond. The production of microalgae for aquaculture is generally on a much smaller scale, and in many cases is carried out indoors in 20–40 l carboys or in large plastic bags of up to approximately 1000 l in volume. More recently, a helical tubular photobioreactor system, the BIOCOIL™, has been developed which allows these algae to be grown reliably outdoors at high cell densities in semi-continuous culture. Other closed photobioreactors such as flat panels are also being developed. The main problem facing the commercialisation of new microalgae and microalgal products is the need for closed culture systems and the fact that these are very capital intensive. The high cost of microalgal culture systems relates to the need for light and the relatively slow growth rate of the algae. Although this problem has been avoided in some instances by growing the algae heterotrophically, not all algae or algal products can be produced this way.     

Glucose‐6‐Phosphate Dehydrogenase from the Oleaginous Microalga Nannochloropsis Uncovers Its Potential Role in Promoting Lipogenesis

Microalgae have long been considered as potential biological feedstock for the production of wide array of bioproducts, such as biofuel feedstock because of their lipid accumulating capability. However, lipid productivity of microalgae is still far below commercial viability. Here, a glucose‐6‐phosphate dehydrogenase from the oleaginous microalga Nannochloropsis oceanica is identified and heterologously expressed in the green microalga Chlorella pyrenoidosa to characterize its function in the pentose phosphate pathway. It is found that the G6PD enzyme activity toward NADPH production is increased by 2.19‐fold in engineered microalgal strains. Lipidomic analysis reveals up to 3.09‐fold increase of neutral lipid content in the engineered strains, and lipid yield is gradually increased throughout the cultivation phase and saturated at the stationary phase. Moreover, cellular physiological characteristics including photosynthesis and growth rate are not impaired. Collectively, these results reveal the pivotal role of glucose‐6‐phosphate dehydrogenase from N. oceanica in NADPH supply, demonstrating that provision of reducing power is crucial for microalgal lipogenesis and can be a potential target for metabolic engineering.     

The potential of cloud point system as a novel two-phase partitioning system for biotransformation

Although the extractive biotransformation in two-phase partitioning systems have been studied extensively, such as the water–organic solvent two-phase system, the aqueous two-phase system, the reverse micelle system, and the room temperature ionic liquid, etc., this has not yet resulted in a widespread industrial application. Based on the discussion of the main obstacles, an exploitation of a cloud point system, which has already been applied in a separation field known as a cloud point extraction, as a novel two-phase partitioning system for biotransformation, is reviewed by analysis of some topical examples. At the end of the review, the process control and downstream processing in the application of the novel two-phase partitioning system for biotransformation are also briefly discussed.     

Flow‐based analysis using microfluidics–chemiluminescence systems

This review will discuss various approaches and techniques in which analysis using microfluidics–chemiluminescence systems (MF–CL) has been reported. A variety of applications is examined, including environmental, pharmaceutical, biological, food and herbal analysis. Reported uses of CL reagents, sample introduction techniques, sample pretreatment methods, CL signal enhancement and detection systems are discussed. A hydrodynamic pumping system is predominately used for these applications. However, several reports are available in which electro‐osmotic (EO) pumping has been implemented. Various sample pretreatment methods have been used, including liquid–liquid extraction, solid‐phase extraction and molecularly imprinted polymers. A wide range of innovative techniques has been reported for CL signal enhancement. Most of these techniques are based on enhancement of the mixing process in the microfluidics channels, which leads to enhancement of the CL signal. However, other techniques are also reported, such as mirror reaction, liquid core waveguide, on‐line pre‐derivatization and the use of an opaque white chip with a thin transparent seal. Photodetectors are the most commonly used detectors; however, other detection systems have also been used, including integrated electrochemiluminescence (ECL) and organic photodiodes (OPDs). Copyright © 2012 John Wiley & Sons, Ltd.     

Isolation and identification of herbivorous ciliates from contaminated microalgal cultures

Ciliates are a common but understudied group of grazers that can invade microalgal cultures. To estimate the potential impact of ciliates on microalgal culture productivity, the identification of species that can invade these cultures is essential. Furthermore, isolation of these herbivorous ciliates allows to use them in experiments that investigate the impact of ciliate grazing on the productivity of microalgal cultures. The main aims of this study were to isolate and identify ciliates that invade cultures of the freshwater microalgae Chlorella and Chlamydomonas, and to establish a live collection of these ciliates for usage in future experiments. To this end, we optimized a method for isolating ciliates from contaminated microalgal cultures and we developed a new PCR primer set for amplifying the partial 18S rDNA of ciliates belonging to the classes Spirotrichea, Oligohymenophorea and Colpodea. As a result, we isolated 11 ciliates from microalgal enrichment cultures inoculated with non-sterile dust and various freshwater sources. Of these 11 species, 7 were found to be feeding on Chlamydomonas. Ciliate species that fed on Chlorella could not be isolated in this study. Ciliate species feeding on Chlamydomonas were identified based on a combination of morphological observations and molecular analyses of partial 18S rDNA sequences.     

微藻采收方法的研究进展

微藻的生产过程可以实现能源生产、废水净化和CO2减排的高度耦合,在能源危机日益紧张、环境问题日趋严峻的今天,微藻的开发利用具有重要的研究价值和经济、社会效益。制约微藻产业化的瓶颈问题是采收成本过高,一种经济合理的采收方法不但可以大大降低生产成本,还可以奠定微藻产业化发展的基础。本文对目前应用较为普遍的微藻采收方法进行了介绍,重点阐述了絮凝法采收微藻,以期对微藻的低成本高效率采收以及产业化发展提供帮助。     

Methods of downstream processing for the production of biodiesel from microalgae

Despite receiving increasing attention during the last few decades, the production of microalgal biofuels is not yet sufficiently cost-effective to compete with that of petroleum-based conventional fuels. Among the steps required for the production of microalgal biofuels, the harvest of the microalgal biomass and the extraction of lipids from microalgae are two of the most expensive. In this review article, we surveyed a substantial amount of previous work in microalgal harvesting and lipid extraction to highlight recent progress in these areas. We also discuss new developments in the biodiesel conversion technology due to the importance of the connectivity of this step with the lipid extraction process. Furthermore, we propose possible future directions for technological or process improvements that will directly affect the final production costs of microalgal biomass-based biofuels.     

A methodological review on the characterization of microalgal biofilm and its extracellular polymeric substances

Biofilm secreted by microalgae are extracellular polymeric substances (EPSs) composed mainly of polysaccharides, proteins, nucleic acids and lipids. These EPSs immobilize the cells and stabilize biofilm, mediating adhesion towards solid surfaces. The EPSs valorization through industrial exploitations and scientific works is becoming more popular, but the bottleneck of such studies is the lack of consensus among researchers on the selection of detection techniques to be used, especially for novice researchers. It is a daunting task for any inexperienced researcher when they fail to identify the right tools needed for microalgal biofilm studies. In this review, a well-refined analysis protocol about microalgal biofilm and EPSs were prepared including its extraction and characterization. Pros and cons of various detection techniques were addressed and cutting-edge methods to study biofilm EPSs were highlighted. Future perspectives were also presented at the end of this review to bridge research gaps in studying biofilm adhesion via EPSs production. Ultimately, this review aims to assist novice researchers in making the right choices in their research studies on microalgal biofilms in accordance to the available technologies and needs.     

Biotransformations using plant cells, organ cultures and enzyme systems: current trends and future prospects

Plants are valuable sources of a variety of chemicals including drugs, flavours, pigments and agrochemicals. Some of the biochemical reactions occurring in plant cells are complex and cannot be achieved by synthetic routes. In vitro plant cell and organ cultures and plant enzymes act as suitable biocatalysts to perform these complex reactions. A wide variety of chemical compounds including aromatics, steroids, alkaloids, coumarins and terpenoids can undergo biotransformations using plant cells, organ cultures and enzymes. The biocatalyst-mediated reactions are regiospecific and stereospecific. Reaction types include oxidations, reductions, hydroxylations, methylations, acetylations, isomerizations, glycosylations and esterfications. Genetic manipulation approaches to biotransformation offer great potential to express heterologous genes and to clone and overexpress genes for key enzymes. Biotransformation efficiencies can further be improved using molecular techniques involving site-directed mutagenesis and gene manipulation for substrate specificity.