提高微生物油脂生产能力的研究进展

微生物油脂是生物柴油生产领域具有广阔前景的新油脂资源。然而, 利用产油微生物进行油脂的工业化生产仍存在限氮条件下油脂生产强度不够高、对廉价高氮生物质原料的利用效率低等瓶颈问题。随着近年来发酵工程、生物信息学及分子生物学技术的发展, 国内外研究者利用不同策略优化微生物油脂的生产条件, 并对其油脂积累代谢途径进行改造, 旨在获得适用于工业化生产的产油性能优良的油脂菌。本综述总结了国内外利用生化工程、基因工程以及新兴的转录因子工程策略提高产油微生物油脂生产强度和扩大产油微生物廉价底物利用范围方面的研究进展, 并展望了基于组学研究、模块途径工程以及反向代谢工程的综合策略在理性改造产油微生物以提高其油脂发酵性能中的应用。     

加快微生物油脂研究为生物柴油产业提供廉价原料

当前国内外致力于发展生物柴油,因其性能优良,成为化石柴油的替代品。由于以植物油脂生产生物柴油原料成本占总成本的70％-85％,所以亟待开发廉价油脂资源。微生物油脂主要是微生物利用碳水化合物合成的甘油脂,其脂肪酸组成和植物油相近。产油微生物具有资源丰富、油脂含量高、碳源利用谱广等特点,开发潜力大。然而,目前微生物油脂生产成本偏高,研究工作仍以富含多不饱和脂肪酸的高附加值菌油为目标。随着现代分子生物学和生物化工技术的发展,对产油微生物菌种筛选、改良、代谢调控和发酵工程的研究日趋深入,将降低微生物油脂生产成本,为未来生物柴油产业提供廉价原料。     

组学技术在产油微生物中的应用

微生物油脂是未来燃料和食品用油的重要潜在资源。近年来,随着系统生物学技术的快速发展,从全局角度理解产油微生物生理代谢及脂质积累的特征成为研究热点。组学技术作为系统生物学研究的重要工具被广泛用于揭示产油微生物脂质高效生产的机制研究中,这为产油微生物理性遗传改造和发酵过程控制提供了基础。文中对组学技术在产油微生物中的应用概况进行了综述,介绍了产油微生物组学分析常用的样品前处理及数据分析方法,综述了包括基因组、转录组、蛋白(修饰)组及代谢(脂质)组等在内的多种组学技术,以及组学数据基础上的数学模型在揭示产油微生物脂质高效生产机制中的研究,并对未来发展和应用进行了展望。     

产油微生物利用餐厨垃圾生产油脂的研究进展

餐厨垃圾中含有丰富的营养物质,经生物转化过程可以合成对人类有用的化学品。某些产油微生物可以处理餐厨垃圾生产油脂,同时合成高附加值代谢产物如多不饱和脂肪酸、角鲨烯和类胡萝卜素等。这不仅能够降低生产成本,而且提高了产物的经济价值,具有极大的工业化应用潜力。文中主要概括了目前餐厨垃圾的处理研究现状,综述了产油微生物发酵餐厨垃圾生产油脂的研究进展,并结合笔者的工作对未来该领域的发展进行了总结与展望,以期为今后的相关研究提供有益借鉴。     

微生物油脂的研究进展及展望

综述了培养微生物生产油脂的发展历史及研究现状,讨论了产油影响因素,对于其瓶颈因素碳源和氮源的影响。可采用培养后期限制氮源并补加碳源的措施,从而解决增加微生物油脂的含量。展望了采取微生物混合培养方法生产油脂技术研究的发展前景。     

微生物油脂及其生产工艺的研究进展

微生物油脂是一种应用前景广阔的新型油脂资源,正越来越受到人们的重视,尤其在生产富含不饱和脂肪酸的功能性油脂方面已成为研究热点。该文对微生物油脂的特点及组成、产油微生物必备条件及常见种类、微生物油脂的生产工艺等方面进行了综述,展望了其研究的发展前景。     

基于废物资源化的美极梅奇酵母产油研究现状及应用潜力分析

清洁可再生能源生物柴油的开发利用是对当今能源短缺环境下化石燃料替代物的有益探索。微生物油脂作为一种可能实现生物柴油廉价、高效生产的原料引起了广泛的关注,但由于封闭式培养模式操作复杂、成本高制约了其大规模应用。美极梅奇酵母Metschnikowia pulcherrmia是一种新型产油酵母,具有适应性强、底物利用范围广、可在开放体系培养等特点,很有潜力代替传统产油微生物,实现基于生物柴油的废水及固废能源化工程应用。文中对美极梅奇酵母相关研究开展了全面调研,在分析其产油研究及应用现状的基础上,总结了美极梅奇酵母在油脂生产方面所具有的独特优势和关键影响因素,突出强调了其在开放体系培养及利用有机废弃物生产微生物油脂的可行性。此外,文中还指出了美极梅奇酵母在油脂产量、产油机理等方面存在的问题与不足,为实现生物柴油高效生产提供了新的方向和思路,有利于进一步促进其工业化应用。     

能源微生物油脂技术进展

微生物油脂技术是缓解生物柴油规模化生产原料短缺的有效途径之一。介绍了国内外利用产油真菌生产能源微生物油脂的现状,包括拓展发酵原料、选育优良菌株、建立新型调控策略和不同培养模式以及解析油脂过量积累的分子机制;概括了微生物油脂技术产业化面临的问题及其解决方案;最后指出了能源微生物油脂研究未来发展方向。     

培养条件对产油微生物生长的影响

为了筛选出高产油菌株, 首先采用细胞形态学方法与细胞化学方法(苏丹III染色法)对4株高产油脂菌株进行初筛, 并通过索氏提取法对初筛菌株油脂含量进行分析, 确定M2菌株为实验菌株, 其油脂含量达53.09%。为了增加产油微生物油脂产量, 本试验考察了不同发酵条件对其细胞生长和油脂积累的影响。优化工艺参数为: 10° Bx玉米皮渣水解液为培养基质, 0.2% NaNO3为氮源, pH 6.0、28oC下发酵培养6 d, 微生物油脂含量75.21%, 菌体生物量30.40 g/L, 油脂产量22.86 g/L。气相色谱分析表明该油脂的脂肪酸组成与植物油相似, 主要含有16碳和18碳系脂肪酸, 可作为生物柴油的原料, 不饱和脂肪酸含量达68%, 可应用于医药化工领域。     

Cryptococcus curvatus O3酵母菌培养及产油脂特性

生物柴油的发展, 导致全球油脂供求紧张。微生物油脂的甘三酯组成与植物油类似, 发展微生物油脂可部分缓解植物油脂供应压力。本文研究了Cryptococcus curvatus O3酵母利用葡萄糖为碳源生长和积累油脂的特性。Cryptococcus curvatus O3酵母在培养过程中能适应间歇式碳源流加方式达到高密度培养的目的, 但在相同培养条件下, 不同氮源能影响其代谢过程中糖到油脂转化的脂肪系数。Cryptococcus curvatus O3酵母利用葡萄糖作为碳源在30°C下摇瓶发酵, 菌体生物量为51.8 g/L, 油脂含量达65.1%。脂肪酸组成分析结果表明, 菌油富含饱和及低度不饱和长链脂肪酸, 其中饱和脂肪酸之和占总脂肪酸组成的64%左右, 其脂肪酸组成类似于可可脂, 这些结果对于利用产油微生物转化生物质获取如类可可脂等具有高附加值油脂的研究具有重要意义。     

“Oil hyphae” of endolithic lichens and their fatty acid composition

Summary The structure of medullary oil hyphae of twelve endolithic lichen species, belonging to different taxa and colonizing different habitats, was examined by light and electron microscopy. The chemical composition of lipids isolated from the oil hyphae and from two corresponding mycobionts grown in culture was determined.The oil hyphae of the various species appeared in different forms and contained large amounts of lipid in the form of oil globules. The hyphae of mycobionts isolated from two of the endoliths and grown in culture also contained large amounts of lipids. Triacylglycerol was the predominant lipid component in all the organisms examined. Hexadecanoic acid was the main saturated fatty acid; octadecenoic acid and octadecdienoic acid the predominant unsaturated fatty acids. Tetradecanoic, hexadecenoic, octadecanoic and octadectrienoic acids were also detected.The fatty acid distribution pattern appeared unaffected by the nature of substrate and climatic conditions. There is a certain similarity in the fatty acid composition in related species.This work forms part of a dissertation to be submitted by E.Kushnir to the Department of Botany, Tel-Aviv University, in partial fulfillment of the requirements for the degree of Doctor of Philosophy.     

Utilization of hydrocarbons by cyanobacteria from microbial mats on oily coasts of the Gulf

Several pieces of evidence indicate that Microcoleu chthonoplastes and Phormidium corium, the predominant cyanobacteria in microbial mats on crude oil polluting the Arabian Gulf coasts, contribute to oil degradation by consuming individual n-alkanes. Both cyanobacteria grew phototrophically better in the presence of crude oil or individual n-alkanes than in their absence, indicating that hydrocarbons may have been utilized. This result was true when growth was measured in terms of dry biomass, as well as in terms of the content of biliprotein, the accessory pigment characteristic of cyanobacteria. The phototrophic biomass production by P. corium was directly proportional to the concentration of n-nonadecance (C₁₉) in the medium. The chlorophyll to carotene ratio of hydrocarbon-grown cyanobacteria did not decrease compared to the ratio in the absence of hydrocarbons, indicating that on hydrocarbons the organisms were not stressed. Comparing the fatty acid patterns of total lipids from hydrocarbon-grown cyanobacteria to those of the same organisms grown without hydrocarbons confirms that n-alkanes were taken up and oxidized to fatty acids by both cyanobacteria.     

Oleaginous yeasts: Biodiversity and cultivation

Microbial lipids produced by oleaginous microorganisms, also called microbial oils and single cell oils (SCOs), are very promising sources for several oil industries. The exploration of efficient oleaginous yeast strains, meant to produce both high-quantity and high-quality lipids for the production of biodiesel, oleochemicals, and the other high value lipid products, have gained much attention. At present, the number of oleaginous yeast species that have been discovered is 8.2% of the total number of known yeast species, most of which have been isolated from their natural habitats. To explore high lipid producing yeasts, different methods, including high-throughput screening methods using colorimetric or fluorometric measures, have been developed. Understanding of the fatty acid composition profiles of lipids produced by oleaginous yeasts would help to define target lipid-related products. For lipid production, the employment of low-cost substrates suitable for yeast growth and lipid accumulation, and efficient cultivation processes are key factors for successfully increasing the amount of the accumulated lipid yield while decreasing the cost of production.     

五种微绿球藻产油和产多不饱和脂肪酸的研究

从5种微绿球藻中鉴别出4个高产油藻种和1个产油量很低的藻种。4种高产油微绿球藻在平台期油脂含量最高,占细胞干重的57%以上,其中三酰基甘油的含量占细胞干重的32.4%-45.2%。分析5种微绿球藻细胞的脂肪酸组成及4种高产油藻三酰基甘油中的脂肪酸组成,发现在高产油藻中,总的饱和脂肪酸和单不饱和脂肪酸的比例达到95%以上,多不饱和脂肪酸在5%以下,而在产油量很低的微绿球藻中多不饱和脂肪酸比例达45%以上。高产油微绿球藻三酰基甘油的多不饱和脂肪酸含量在4%以下,是生物柴油的优质原料,而产油量低的微绿球藻可用于提取C20:5脂肪酸(EPA)。       

Archaeal lipids

The major archaeal membrane glycerolipids are distinguished from those of bacteria and eukaryotes by the contrasting stereochemistry of their glycerol backbones, and by the use of ether-linked isoprenoid-based alkyl chains rather than ester-linked fatty acyl chains for their hydrophobic moieties. These fascinating compounds play important roles in the extremophile lifestyles of many species, but are also present in the growing numbers of recently discovered mesophilic archaea. The past decade has witnessed significant advances in our understanding of archaea in general and their lipids in particular. Much of the new information has come from the ability to screen large microbial populations via environmental metagenomics, which has revolutionised our understanding of the extent of archaeal biodiversity that is coupled with a strict conservation of their membrane lipid compositions. Significant additional progress has come from new culturing and analytical techniques that are gradually enabling archaeal physiology and biochemistry to be studied in real time. These studies are beginning to shed light on the much-discussed and still-controversial process of eukaryogenesis, which probably involved both bacterial and archaeal progenitors. Puzzlingly, although eukaryotes retain many attributes of their putative archaeal ancestors, their lipid compositions only reflect their bacterial progenitors. Finally, elucidation of archaeal lipids and their metabolic pathways have revealed potentially interesting applications that have opened up new frontiers for biotechnological exploitation of these organisms. This review is concerned with the analysis, structure, function, evolution and biotechnology of archaeal lipids and their associated metabolic pathways.     

Identification, isolation and characterization of tetracosapolyenoic acids in lipids of marine coelenterates.

Several tetracosapolyenoic acids (TPA) were detected in lipids of different marine coelenterates. Two of these acids were isolated and their structures were confirmed by chemical and spectral methods as all-cis-6,9,12,15,18-tetracosapentaenoic and all-cis-6,9,12,15,18,21-tetracosahexaenoic acid. Their distribution among lipids of a number of species of different classes of coelenterates from the northern and tropical seas, among neutral and polar lipids of these organisms was investigated. Significant quantities of TPA were found in all of the Octacorallia species studied. In some cases the sum of TPA reaches the level of 20% of total lipid fatty acids. The fatty acid composition of different coelenterates is also discussed.     

Oily yeasts as oleaginous cell factories

Oily yeasts have been described to be able to accumulate lipids up to 20% of their cellular dry weight. These yeasts represent a minor proportion of the total yeast population, and only 5% of them have been reported as able to accumulate more than 25% of lipids. The oily yeast genera include Yarrowia, Candida, Rhodotorula, Rhodosporidium, Cryptococcus, Trichosporon, and Lipomyces. More specifically, examples of oleaginous yeasts include the species: Lipomyces starkeyi, Rhodosporidium toruloides, Rhodotorula glutinis, and Yarrowia lipolytica. Yeast do exhibit advantages for lipid production over other microbial sources, namely, their duplication times are usually lower than 1 h, are much less affected than plants by season or climate conditions, and their cultures are more easily scaled up than those of microalgae. Additionally, some oily yeasts have been reported to accumulate oil up to 80% of their dry weight and can indeed generate different lipids from different carbon sources or from lipids present in the culture media. Thus, they can vary their lipid composition by replacing the fatty acids present in their triglycerides. Due to the diversity of microorganisms and growth conditions, oily yeasts can be useful for the production of triglycerides, surfactants, or polyunsaturated fatty acids.     

Recent advances in the mass spectrometric profiling of bacterial lipids

Exploring the lipids of bacteria presents a predicament that may not be broadly recognized in a field dominated by the biology and biochemistry of eukaryotic — and especially, mammalian — lipids. Bacteria make multifarious metabolites that contain fatty acyl chains of unusual length and unsaturation attached to assorted headgroups, including sugars and fatty alcohols. Lipid profiling approaches developed for eukaryotic lipids often fail to detect, resolve, or identify bacterial lipids due to their wide range of polarities (including very hydrophobic species) and diverse positional and stereochemical variations. Global lipid profiling, or lipidomics, of bacteria has thus developed as a separate mission with methodological and scientific considerations tailored to the biology of these organisms. In this review, we summarize findings primarily from the last three years that exemplify recent advances and continuing challenges to learning about bacterial lipids.     

Composition, accumulation and utilization of yolk lipids in teleost fish

Lipid reserves in teleost eggs are stored in lipoprotein yolk and, in some species, a discrete oil globule. Lipoprotein yolk lipids are primarily polar lipids, especially phosphatidylcholine and phosphatidylethanolamine (PE), and are rich in (n–3) polyunsaturated fatty acids, especially 22:6(n–3) (docosahexaenoic acid, DHA). Oil consists of neutral lipids and is rich in monounsaturated fatty acids (MUFA). Egg lipids are derived from dietary fatty acid, fatty acid mobilized from reserves and possibly fatty acid synthesized de novo. There is selective incorporation of essential fatty acids, particularly DHA, into yolk lipids and discrimination against incorporation of 22:1(n–11). Lipid is delivered to the oocyte by vitellogenin, which is rich in polar lipids, and likely also by other lipoproteins, especially very low density lipoprotein, which is rich in triacylglycerol (TAG). All classes of lipid may be used as fuel during embryonic and larval development and MUFA are preferred fatty acids for catabolism by embryos. Catabolism of oil globules is frequently delayed until latter stages of development. In some species, DHA derived from hydrolysis of phospholipid may be conserved by transfer to the neutral lipid. Recent work has expanded knowledge of the role of DHA in membrane structure, especially in neural tissue, and molecular species analysis has indicated that PE containing sn-1 oleic acid is a prime contributor to membrane fluidity. The results of this type of study provide an explanation for the selection pressures that influence yolk lipid composition. Future work ought to expand knowledge of specific roles of individual fatty acids in embryos along with knowledge of the ecological physiology of ovarian recrudescence, environmental influences on vitellogenin and yolk lipid composition, and the control of yolk lipid accumulation and utilization.