一株具有工业化生产DHA前景的破囊壶菌

多不饱和脂肪酸是保持人体健康不可缺少的营养成分之一,尤其是二十二碳六烯酸(DHA)作为细胞膜磷脂的重要组分,具有非常重要的医药应用和营养价值。目前,在食品工业中,DHA已经添加至牛奶或奶粉中,用作功能性营养强化剂。20世纪80年代,DHA的唯一来源是鱼油,但鱼油的腥味、重金属污染等问题,促使人们探索生产DHA的其他途径如微生物发酵。诱变和筛选是微生物选育过程中比较重要的手段,可以快速使菌株朝着人类所需要的方向突变。UV诱变和化学药物胁迫筛选是使野生株定向突变的一种很好的办法。目前国内外研究主要针对裂殖壶菌属菌株进行诱变育种,许永利[1]用紫外线诱变和喹禾灵筛选方法对裂殖壶菌(Schizochytrium limacinu)进行诱变选育,突变菌株生物量和DHA含量比对照菌株均有提高。吴克刚等[2]利用添加植物激素对Thraustochytriu roseum MF2进行培育诱变,从而获得更高的DHA量,但在脂质含量和DHA在脂质占比率上都不存在明显变化。本期介绍梁园梅、成家杨等[3]发表的论文《高产DHA破囊壶菌Aurantiochytrium sp.PKU#SW7诱变株的筛选》,作者采用紫外线和药物双重诱变胁迫破囊壶菌获得一株突变株,其在生物量、脂质含量和DHA在脂质占比率上都有显著性提高,相比前人的研究,作者获得的突变株有显著优越性,且突变株DHA生产能力传代4次后仍然保持稳定,具有较高的工业价值。在后续的研究中作者若能对筛选条件、培养基(如利用廉价碳源)和培养条件等方面实行进一步优化,提高突变株生物量,降低突变株发酵生产DHA生产成本,将能获得更高的商业价值。     

深圳海域6株破囊壶菌的生长特性及油脂成分分析

【目的】从深圳海域分离得到6株破囊壶菌,对其基本形态特征、生活史和油脂含量等进行研究,开发其应用潜力。【方法】使用松花粉垂钓法对破囊壶菌进行分离,通过18S r RNA基因测序的方法对破囊壶菌进行鉴定,用显微镜观察其基本形态特征,通过使用尼罗红(Nile Red)染色法对油脂含量进行定性检测,并用GC-MS分析菌株的油脂含量和组成情况。【结果】18S r RNA基因鉴定其属于Aurantiochytrium sp.、Schizochytrium sp.和Thraustochytrium sp.三个属。破囊壶菌的脂肪酸主要成分为十六碳饱和脂肪酸和二十二碳六烯酸(DHA),其中Mn11和Mn15的饱和脂肪酸含量达到总脂肪酸含量的70%以上,Mn16和Sw7的DHA产量分别达到1.29 g/L和1.26 g/L。【结论】Mn11和Mn15菌株适合用于生物柴油的生产,Mn16和Sw7是DHA发酵生产的潜力菌株。     

生物转化高浓度甘油生产1,3-二羟基丙酮及分批发酵动力学

以生物柴油生产的高浓度副产物甘油为唯一碳源筛选甘油高耐受性1,3-二羟基丙酮(DHA)高产菌株,运用响应面与正交试验优化菌株产DHA条件,提高DHA产量。分子生物学鉴定表明:筛选的高产DHA菌种G40为芽胞杆菌属(Bacillus)菌株,DHA产量为29.46g/L。响应面分析和正交试验优化后,在甘油224.22g/L、K_2HPO_41.60g/L、NaCl0.5g/L、KH_2PO_40.5g/L、(NH_4)_2SO_40.5g/L、酵母膏1.60g/L和pH7.2、35℃、200r/min的条件下,G40菌株发酵60h产生DHA86.84g/L,比优化前提高了194.8%。实验建立了一种利用高浓度甘油高效率发酵生产DHA的方法。     

高产DHA破囊壶菌Aurantiochytrium sp. PKU#SW7诱变株的筛选

【目的】对野生菌株Aurantiochytrium sp.PKU#SW7诱变育种,筛选高产DHA突变株。【方法】采用UV诱变和化学药物胁迫筛选方式,以菌株的生物量、油脂产量、DHA产量作为筛选指标,获得高产DHA突变株。【结果】经鉴定获得一株DHA高产突变株PKU#PM003,该菌株传代4次后仍保持较好的遗传稳定性。摇瓶发酵后,PKU#PM003生物量产量高达6.62 g/L,比原始菌株5.95 g/L提高了11.26%,脂肪酸含量高达4.01 g/L,比原始菌株3.18 g/L提高了26.1%,DHA在脂肪酸中所占比例由29.97%增加到33.43%,产量提高了41.01%,油脂突变效果显著。【结论】突变株PKU#PM003可作为性状优良的工业化发酵生产菌种,并在DHA产量提升上仍具有巨大的空间。     

微生物发酵法生产1,3-丙二醇的研究新进展

1,3-丙二醇(1,3-PD)是一种重要的化工原料,广泛应用于医药、化工、食品及化妆品等行业,同时1,3-PD是合成聚对苯二甲酸丙二酯(PTT)的重要单体,市场需求量逐年增多。基于生态友好型、生产安全和可持续发展的要求,利用微生物转化可再生资源来生产1,3-PD受到了人们的广泛重视。综述了微生物发酵法生产1,3-PD的菌株、代谢途径、发酵和下游分离工艺及其新进展,并对工业生产中利用生物技术生产1,3-PD的未来前景和挑战进行了探讨。     

微生物法生产1,3-二羟基丙酮代谢工程研究进展

1,3-二羟基丙酮是一种重要的化工原料和医药中间体,广泛应用于化妆品、医药、食品等领域。以下综述了微生物法生产1,3-二羟基丙酮的代谢途径和关键酶,以及微生物法生产1,3-二羟基丙酮所涉及的代谢工程技术的研究进展。指出利用基因工程的方法对菌株进行改造,提高甘油脱氢酶催化活性,同时根据菌株的代谢特性,对发酵过程进行调控,提高1,3-二羟基丙酮的得率,是今后的研究方向。     

诱变育种在产油微生物生产DHA中的研究进展

二十二碳六烯酸(DHA)对维持人体正常的生理功能至关重要,具有极高的生物医学价值。产油微生物是指具有很强的脂肪酸合成能力,DHA占总脂肪酸含量相对较高的一类微生物,是获取DHA的新途径。诱变育种作为一种有效的变异手段在产油微生物选育与改良中显示了极为重要的作用和十分诱人的前景。综述了通过物理和化学诱变选育高产DHA菌株的机理及方法,其中物理诱变包括γ射线、紫外线、离子束和常压室温等离子体等;化学诱变包括甲基磺酸乙酯、亚硝基胍和硫酸二乙酯等;主要介绍了不同诱变育种方式在产油微生物生产DHA研究方面取得的成就,探讨了诱变育种的发展方向,提出了不同诱变育种方式相结合的复合育种的思路,以期对今后产油微生物生产DHA的研究有所启发。     

成都普瑞法科技开发有限公司

本公司专业从事高含量提取物,高纯度中药/提取物单体和天然产物医药中间体的生产,定制和生产工艺开发。致力于研究开发高纯度药用植物化学成分的最新分离纯化方法,结合成都在西南地区的资源和科     

成都普瑞法科技开发有限公司

本公司专业从事高含量提取物,高纯度中药/提取物单体和天然产物医药中间体的生产,定制和生产工艺开发。致力于研究开发高纯度药用植物化学成分的最新分离纯化方法,结合成都在西南地区的资源和科研优势,依托四川大学等科研院校,为客户提供全方位的优质服务。     

成都普瑞法科技开发有限公司

本公司专业从事高含量提取物,高纯度中药/提取物单体和天然产物医药中间体的生产,定制和生产工艺开发。致力于研究开发高纯度药用植物化学成分的最新分离纯化方法,结合成都在西南地区的资源和科研优势,依托四川大学等科研院校,为客户提供全方位的优质服务。     

Expression of Vitreoscilla Hemoglobin Enhances Cell Growth and Dihydroxyacetone Production in Gluconobacter oxydans

Dihydroxyacetone (DHA) is an important ketose sugar, which is extensively used in the cosmetic, chemical, and pharmaceutical industries. DHA has been industrially produced by Gluconobacter oxydans with a high demand of oxygen. To improve the production of DHA, the gene vgb encoding Vitreoscilla hemoglobin (VHb) was successfully introduced into G. oxydans, where it was stably maintained, and expressed at about 76.0 nmol/g dry cell weight. Results indicated that the constitutively expressed VHb improved cell growth and DHA production in G. oxydans under different aeration conditions. Especially at low aeration rates, the VHb-expressing strain (VHb⁺) displayed 23.13% more biomass and 37.36% more DHA production than those of VHb-free strain (VHb⁻) after 32 h fermentation in bioreactors. In addition, oxygen uptake rate (OUR) was also increased in VHb⁺ strain relative to the control strain during fermentation processes.     

Crypthecodinium cohnii with emphasis on DHA production: a review

Docosahexaenoic acid (DHA) is a polyunsaturated fatty acid (PUFA) that belongs to the ω-3 group. In recent years, DHA has attracted much attention because of its recognized beneficial effect on human health. At present, fish oil is the major source of DHA, but it may be produced by microorganisms with additional benefits. Marine microorganisms may contain large amounts of DHA and are considered a potential source of this important fatty acid. Some of these organisms can be grown heterotrophically on organic substrates without light, offering the possibility of greatly increasing microalgal cell concentration under controlled and monitored conditions, resulting in a very high quality product. Among the heterotrophic marine dinoflagellates, Crypthecodinium cohnii has been identified as a prolific producer of DHA. The organism is extraordinary in that it produces no other PUFAs than DHA in its cell lipid in any significant amount, which makes the DHA purification process very attractive, particularly for pharmaceutical and nutraceutical applications. This paper reviews recent advances in the biotechnological production of DHA by C. cohnii.     

A HIGH THROUGHPUT SCREENING METHOD FOR 1,3-DIHYDROXYACETONE-PRODUCING BACTERIUM BY CULTIVATION IN A 96-WELL MICROTITER PLATE

1,3-Dihydroxyacetone (DHA) is used extensively in the cosmetic industry, and is the main active ingredient in all sunless tanning skincare preparation. In order to more efficiently and rapidly screen suitable strains or mutants for production of DHA, a high throughput screening method for DHA-producing bacterium by cultivation in a 96-well microtiter plate was developed. With this screening method, more than 100 strains that were able to convert glycerol to DHA were isolated from soil samples, and a mutant of Gluconobacter oxydans ZJB-605 that displayed the highest DHA productivity was obtained.

PRACTICAL APPLICATIONS

The practical application of this work is to promote the microbial process for isolating DHA-producing bacterium and screening DHA-overproducing mutant. With it, DHA manufactory can improve efficiency of strain operation, reduce labor and decrease production costs of DHA. It also can be used for reference about researches of glycerol dehydrogenase, and other alcohol dehydrogenase.     

Efficient production of triacylglycerols rich in docosahexaenoic acid (DHA) by osmo-heterotrophic marine protists

Thraustochytrids have recently emerged as a promising source for docosahexaenoic acid (DHA) production due to their high growth rate and oil content. In this study, two thraustochytrid isolates, Aurantiochytrium sp. PKU#SW7 and Thraustochytriidae sp. PKU#Mn16 were used for DHA production. Following growth parameters were optimized to maximize DHA production: temperature, pH, salinity, and glucose concentration. Both isolates achieved the highest DHA yield at the cultivation temperature of 28 °C, pH 6, 100 % seawater, and 2 % glucose. A DHA yield of 1.395 g/l and 1.426 g/l was achieved under the optimized culture conditions. Further investigation revealed that both isolates possess simple fatty acids profiles with palmitic acid and DHA as their dominant constituents, accounting for ～79 % of total fatty acids. To date, very few studies have focused on the DHA distribution in various lipid fractions which is an important factor for identifying strains with a potential for industrial DHA production. In the present study, the lipids profiles of each strain both revealed that the majority of DHA was distributed in neutral lipids (NLs), and the DHA distribution in NLs of PKU#SW7 was exclusively in the form of triacylglycerols (TAGs) which suggest that PKU#SW7 could be utilized as an alternative source of DHA for dietary supplements. The fermentation process established for both strains also indicating that Aurantiochytrium sp. PKU#SW7 was more suitable for cultivation in fermenter. In addition, the high percentage of saturated fatty acids produced by the two thraustochytrids indicates their potential application in biodiesel production. Overall, our findings suggest that two thraustochytrid isolates are suitable candidates for biotechnological applications.     

Effects of organic and inorganic salts on docosahexaenoic acid (DHA) production by a locally isolated strain of Thraustochytrium sp. T01

Abstract

The traditional source for docosahexaenoic acid (DHA) i.e. fish oil is currently being replaced by microbial sources due to the unpleasant odor and the risk of chemical contamination of fish. Thraustochytrium sp., marine microalgae-like protist is a known source of DHA. In our previous study, we reported a high yielding strain, T01, of Thraustochytrium sp. for DHA production isolated from the mangroves of South India. This strain shows promising yields of biomass and DHA. Shake flask study of T01 yielded 6.17?±?0.04 gL^?1 of DHA. In the present work, we report the effects of organic and inorganic salts on DHA production. Addition of organic salts such as sodium acetate, pyruvate, citrate and malate led to increase in the DHA content in T01 strain. The DHA content increased by 40–46% on addition of sodium salts of organic acids, while inorganic phosphates increased DHA by 33%. The total lipid content also increased (28–33%) with salts of organic acids and 28% with phosphate, but not as much as the increase in DHA. Addition of all the salts together did not show a significant increase in lipid and DHA contents as compared to the addition of individual salts.     

工业乳酸发酵的近期进展

乳酸是一种重要的多用途有机酸。通过菌种改良和发酵工艺技术的改进,可以大大提升微生物发酵技术水平,降低成本。简要综述有关的研究进展。     

Recent advances in microbial production of mannitol: utilization of low-cost substrates,strain development and regulation strategies

Mannitol has been widely used in fine chemicals, pharmaceutical industries, as well as functional foods due to its excellent characteristics, such as antioxidant protecting, regulation of osmotic pressure and non-metabolizable feature. Mannitol can be naturally produced by microorganisms. Compared with chemical manufacturing, microbial production of mannitol provides high yield and convenience in products separation; however the fermentative process has not been widely adopted yet. A major obstacle to microbial production of mannitol under industrial-scale lies in the low economical efficiency, owing to the high cost of fermentation medium, leakage of fructose, low mannitol productivity. In this review, recent advances in improving the economical efficiency of microbial production of mannitol were reviewed, including utilization of low-cost substrates, strain development for high mannitol yield and process regulation strategies for high productivity.     

Fed-batch cultivation of the docosahexaenoic-acid-producing marine alga Crypthecodinium cohnii on ethanol

The heterotrophic marine microalga Crypthecodinium cohnii produces docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications. So far, DHA production has been studied with glucose and acetic acid as carbon sources. This study investigates the potential of ethanol as an alternative carbon source for DHA production by C. cohnii. In shake-flask cultures, the alga was able to grow on ethanol. The specific growth rate was optimal with 5 g l(-1) ethanol and growth did not occur at 0 g l(-1) and above 15 g l(-1). By contrast, in fed-batch cultivations with a controlled feed of pure ethanol, cumulative ethanol addition could be much higher than 15 g l(-1), thus enabling a high final cell density and DHA production. In a representative fed-batch cultivation of C. cohnii with pure ethanol as feed, 83 g dry biomass l(-1), 35 g total lipid l(-1) and 11.7 g DHA l(-1) were produced in 220 h. The overall volumetric productivity of DHA was 53 mg l(-1 )h(-1), which is the highest value reported so far for this alga.     

Scale-up of a new bacterial mannitol production process

D-Mannitol is a sugar alcohol with applications in chemistry, food and pharmaceutical industries, and medicine. Commercially, mannitol is produced by catalytic hydrogenation. Although this process is widely used, it is not optimal for mannitol production. New processes, including chemical, enzymatic, and microbial processes, are frequently developed and evaluated against the existing hydrogenation processes. In earlier papers, we have described the identification of a food-grade lactic acid bacterium strain, Leuconostoc mesenteroides ATCC-9135, with efficient mannitol production capabilities and the development and optimization of a new bioprocess in which the strain was applied. The new bioprocess is simple. It requires a reduced bioreactor with the following features: sterilization, pH and T control (at mild conditions), and slow mixing. The contamination risk of the new bioprocess is low, and the downstream processing protocol comprises simple, widely used unit operations: evaporation, crystallization, crystal separation, and drying. On a 2-L laboratory scale, high mannitol yields from fructose (93-97%) and volumetric mannitol productivities (>20 g L(-1) h(-1)) were achieved. In this paper, the scalability of the new bioprocess was tested on a small pilot scale (100 L). In the pilot plant, production levels were achieved similar to those in the laboratory. Also, high-purity mannitol crystals were obtained at similar yield levels. The results presented in this paper indicate that the new bioprocess can easily be scaled-up to an industrial scale and that the production levels achieved with it are comparable to the catalytic hydrogenation processes.     

微生物发酵产二十二碳六烯酸代谢机理的研究进展

二十二碳六烯酸(简称DHA)是一种重要的长链多不饱和脂肪酸,对人体具有重要的生理功能。微生物发酵生产的DHA与鱼油来源的DHA相比,具有诸多优点,其发展前景广阔。以下从发酵菌株、代谢途径、关键酶以及油脂积累机制等方面进行了综述,为通过代谢工程等技术手段进一步提高DHA产量提供了参考。