Real-time analyte monitoring of a fungal fermentation, at pilot scale, using in situ mid-infrared spectroscopy

Robust in situ biochemical monitoring is essential for the development of substrate feed control to optimize fermentation processes. The scale up of the fermentation for the fungus Glarea lozoyensis can benefit from such technology to improve the yield of the pharmaceutically important pneumocandin of interest and control the levels of unwanted analogues. A new in situ probe, using a diamond attenuated total reflection element, was evaluated at pilot scale for the quantitative measurement of fermentation analytes using Fourier transform mid-IR spectrometry. The new technology was shown to be stable, unaffected by reactor operation conditions of agitation, airflow, and backpressure, but sensitive to temperature control. Both glucose and phosphate were simultaneously monitored during a seed fermentation at 280 L pilot scale using complex medium with detection to 0.1 g/L for both analytes. Fructose, glutamate, and proline were monitored at 75 L scale using production media with detection limits of 0.1, 0.5, and 0.5 g/L respectively. Partial least squares calibration/prediction models were created for analytes of interest using off-line reference measurements and specific spectral regions. Good fits were obtained between off-line measurements and those predicted by in situ mid-IR. Standard errors of prediction (SEP) for glucose (range 18-0.1 g/L) and phosphate (range 11-7.5 g/L) were 0.16 and 1.8 g/L respectively with mean percentage errors (MPEs) around 2.5%. SEP values for the production process: fructose (range 20-0.1 g/L), glutamate (8-0.5 g/L), and proline (12-0.5 g/L) were 0.44, 0.6, and 0.5 g/L respectively with MPEs of 2.2, 5.3, and 10.1%. The technology effectively demonstrates quantitative multicomponent analysis of fermentation processes using in situ monitoring.     

矿质元素对毛头鬼伞菌丝体生长的影响

研究了4种矿质元素对栽培食用菌毛头鬼伞菌丝体生长的影响。4种矿质单元素添加试验结果表明,能使毛头鬼伞菌丝体旺盛生长的最佳浓度分别是CaCl20.1 mmol/L、MgSO48 mmol/L、KCl 1.22 mmol/L、NaH2PO413 mmol/L。4种元素协同作用试验结果表明,单元素浓度的最佳生长量与混合施用时的最佳生长量一致。以毛头鬼伞的主要栽培料之一的棉子壳估算,原料中的4种矿质元素浓度均偏低,必须补充矿质元素才能满足毛头鬼伞菌丝体的旺盛生长。     

高产γ-氨基丁酸的谷氨酸棒杆菌工程菌的构建和发酵条件优化

为了实现γ-氨基丁酸(GABA)在微生物中"一步法"高效生产,本研究构建出一株高产GABA的谷氨酸棒杆茵工程菌ATCC 13032/pDXW10-gadB1-gadB2,可以直接将自身合成的L-谷氨酸转变成GABA,并对该工程茵的发酵培养基和发酵条件进行了初步优化。结果表明:培养7 h~8 h的种子液按发酵起始OD_(562)=1.6转接至发酵培养基(葡萄糖、玉米浆分别100 g/L、4 g/L),10 h添加PLP至O.1 mmol/L,发酵结束后胞外GABA可达(26.39±1.68)g/L。为工业化"一步法"生产GABA提供理论和实验基础。     

太子参‘柘参1号’离体快繁试验

以太子参‘柘参1号’叶片、茎段为外植体进行离体快繁试验,结果表明：带腋芽的茎段能够诱导出丛生芽,且诱导率较高,最佳诱导培养基为MS + 6-BA 1.0 mg/L + NAA 0.2 mg/L +蔗糖30 g/L;增殖培养基为MS + 6-BA 1.0 mg/L + NAA 0.2 mg/L +蔗糖30 g/L;生根培养基为MS + NAA 0.2 mg/L +蔗糖30 g/L。     

多叶斑叶兰繁殖体系建立及基于转录组的发育调控途径功能基因研究

多叶斑叶兰,是兰科斑叶兰属濒危野生植物、国家二级保护植物,具有极高的观赏和药用价值。多叶斑叶兰由于分布种群小,传播扩散能力弱,自然繁殖受到极大限制。以野生多叶斑叶兰茎段作为外植体,建立高效直接的植株再生体系。结合高通量转录组测序技术与生物信息学分析技术,深入挖掘参与多叶斑叶兰器官发育过程的功能基因。结果表明, Morel+2.0 mg/L 6-BA +0.5 mg/LKT+1.0 mg/LNAA+1g/L蛋白胨+25g/L 蔗糖+7.0 g/L Agar+1.0 g/L活性炭+30 g/L 香蕉+50 g/L土豆为最佳的芽诱导培养基;Morel+3 mg/L 6-BA+0.5mg/L NAA+0.5 mg/LKT+0.01 mg/L TDZ+2g/L蛋白胨+25g/L 蔗糖+7.0 g/L Agar+1.0 g/L活性炭30g/L 香蕉+50g/L 土豆为最佳芽增殖培养基;1/2 Morel+1.0 mg/L IBA+0.1 mg/L NAA+1 g/L+花宝2 号+25g/L 蔗糖+7.0g/L Agar +1.0 g/L活性炭+1g·L-1蛋白胨为最佳的生根培养基。转录组测序分析组装获得170688个Unigene,平均长度为584 bp, N50为833 bp。17352个Unigene比对注释到NR、Swiss-Prot、KOG、GO、KEGG数据库。野生苗与组培苗差异表达Unigene GO与KEGG功能富集分析显示, Unigene主要参与调控植物激素信号转导、植物形态发育、次生代谢过程以及能量代谢过程等生物学功能。进一步分析获得511个参与植物器官发育调控相关的转录因子。成功建立了多叶斑叶兰种质资源保存与高效离体再生体系,结合高通量转录组学技术,获得全面完整的多叶斑叶兰转录组信息特征,为后期多叶斑叶兰快速扩繁、遗传转化以及功能基因鉴定、遗传发育及其调控机制研究奠定基础。     

Bioremediation of synthetic high–chemical oxygen demand wastewater using microalgal species Chlorella pyrenoidosa

The microalgal species Chlorella pyrenoidosa was cultivated in synthetic wastewater of initial chemical oxygen demand (COD), nitrate, and phosphate concentrations of 5000, 100, and 40 mg/L, respectively. The aim of the study was to find out the tolerance of microalgae to different COD concentrations and the extent of COD degradation at those concentrations. Three dilutions of wastewater (initial COD concentrations 5000, 3000, and 1000 mg/L) and three inoculum sizes (0.1, 0.2, and 0.3 g/L) were considered for the study. The experimental parameters such as total organic carbon, total inorganic carbon, COD, optical density, total solids, nitrate, and phosphate were measured on a daily basis. Biodegradation kinetics was determined for all cases using first-order reaction and Monod degradation equations. Optimal results showed that up to 90% reduction in TOC was obtained for 1000 COD wastewater while only 38% reduction in total organic carbon (TOC) was achieved for 5000 COD wastewater. Over 95% reduction in nitrate and nearly 90% removal of phosphate were obtained with the lowest microalgal inoculum concentration (i.e., 0.1 g/L) for all COD dilutions. This study showed that microalgal species C. pyrenoidosa can successfully degrade the organic carbon source (i.e., acetate) with significant removal efficiencies for nitrate and phosphate.     

绿僵菌无纺布菌条的制作(I)液体种子培养条件

研究了金龟子绿僵菌菌株M a83液体培养最佳营养及环境条件,为无纺布培养提供良好的液体种子。以生物量为指标,通过液体摇瓶试验确定了绿僵菌M a83菌株液体种子培养参数,对不同的氮源、碳源及其浓度和初始pH进行单因素分析。结果表明,绿僵菌M a83菌株液体种子培养最佳营养条件为4%黄豆粉,2%白砂糖,0.5%MgSO4.7H2O,0.05g/LKC l,0.1g/L FeSO4.7H2O,1.0g/L KH2PO4,pH6.3~6.5。70L发酵罐培养,干生物量第三天可达35g/L,显著优于筛选前培养基(SDA,28g/L,a=0.05)。     

4-氯乙酰乙酸乙酯羰基还原酶产生菌的筛选及产酶条件研究

从实验室保藏的菌株中,筛选到一株立体选择性较高的产4-氯乙酰乙酸乙酯(COBE)羰基还原酶的菌株———出芽短梗霉(Aureobasidiumpullulans)SW0202,菌体产酶条件研究表明,最佳的发酵培养基配方为:麦芽糖30.0g/L,酵母膏20.0g/L,蛋白胨3.0g/L,(NH4)2SO45.0g/L,KH2PO42.0g/L,MgSO4.7H2O0.7g/L,最适发酵温度及初始pH分别为:28°C和pH6.0。该菌在此条件下发酵培养24h,产菌丝体生物量16.78g干菌体/L,COBE羰基还原酶酶活力达到1007U/L。在COBE的转化反应中,产物S-CHBE的浓度达到10.12g/L,光学纯度>97%e.e.。     

响应面法优化毛霉菌发酵培养基

采用响应面分析方法优化毛霉菌B的发酵培养基,首先通过单因素试验筛选出葡萄糖为最适碳源,酵母膏和玉米浆为最适氮源,用Plackett—Burman试验对葡萄糖、酵母膏、玉米浆、MgSO4、FeSO4、NILCl/、HPO4进行评估并筛选出具有显著效应的3个因素：葡萄糖、酵母膏、玉米浆,再通过最陡爬坡试验逼近其最大响应区域,最后采用Box—Behnken试验对其用量进行优化,得到毛霉菌最佳发酵培养基（g/L）：葡萄糖51．54,酵母膏5．22,玉米浆14．31,MgSO40．5,FeSO40．1,NH4Cl3,k2HPO43,pH6．0～6．5。培养基优化后,毛霉生物量由23．51g／L提高至31．13g/L,比对照组提高32．41％,腺嘌呤转化率由53．59％提高至59．97％,ATP产率由6．56g／L提高至7．34g／L,比对照组提高11．89％。     

Effects of optimum doses of nitrogen,phosphorus, potassium and calcium on silkworm,Bombyx mori L., growth and yield

Silkworm rearing has been an income source for small-holding farmers in Pakistan. Over the years efforts have been made to improve silkworm quality and quantity and thus develop better prospects for increasing yield and income. Mulberry leaves Morus alba L. in a series of experiments have been supplemented with various nutrients in different doses and combinations. Relative success has been achieved in this context. In our present experiments on silkworm Bombyx mori L., we fed the larvae on mulberry leaves supplemented with different doses of N (0.2%), P (0.1%), K (0.3%) and Ca (0.1%) at 26 ± 3°C with 65 ± 5% relative humidity. The results showed better outputs in terms of food consumption (72.55 g/10 larvae), coefficient of utilization (76.45%), body weight (70.07 g/10 larvae) and body length (8.56 cm/larvae) than where simple mulberry leaves were offered. Our net gains with these production parameters have been in terms of heavier cocoons with and without pupa (2.25 and 0.69 g/cocoon), respectively.     

果糖和前体物质对紫杉醇生物合成的影响

研究了果糖和几种前体物质对东北红豆杉生产紫杉醇的影响,结果表明,在第12d加入6g/L果糖可以使紫杉醇产量增加63.89％,在糖协同的作用下,加入前体（0.05mmol/L乙酸钠,0.05mmol/L苯丙氨酸,0.1mmol/L苯甲酸钠）可显著提高紫杉醇的合成,同对照相比,含量分别增加49.36%、13.18%和64.26%,在第15d向培养基中加入0.05 mmol/L乙酸钠、0.1mmol/L苯甲酸钠、1mmol/L苯丙氨酸和6g/L果糖则使紫杉醇含量提高181.89％。     

分批补料对木糖发酵生产乙醇的影响

以树干毕赤酵母为发酵菌种,纯木糖为发酵底物,通过分批补料来提高糖利用率以及乙醇得率。结果表明,在24h内,最佳初始木糖浓度为80g/L,在28h的发酵周期中,可以将木糖浓度提高至90g/L,在32h发酵周期内可以将木糖浓度提高至100g/L。通过分批补料,乙醇浓度得到明显提高。当总糖浓度分别为80g/L、90g/L时,24h发酵周期内,分批补料次数以1次为宜,乙醇浓度分别达30.95g/L、32.60g/L,相比于不补料即一次性投料,乙醇浓度分别提高了9.36%、9.18%。总糖浓度100g/L,28h发酵周期内,补料2次效果最佳,乙醇浓度达37.49g/L,比一次性投料下提高了10.36%,较一次性投料达到相同发酵效果缩短了4h。     

Direct hapten-linked multiplexed immunoassays on polycarbonate surface

Direct hapten-linked multiplexed immunoassay is developed on the polycarbonate surface of standard Digital Versatile Discs (DVDs) for six compounds of environmental concern, as proof of concept. Carboxylated haptens are directly linked to the aminated polycarbonate surface through carbodiimide/succinimide coupling. The modified DVD surface maintained its physical and optical properties. Multiplexed assay reached detection limits down to 0.1 μg/L for chlorpyrifos, 2,4,5-trichlorophenoxypropionic acid, sulfathiazole and sulfasalazine and down to 1.0 μg/L for fenthion and malathion. This approach presents advantages such as the improvement in sensitivity in comparison to protein-hapten conjugate format for all the studied analytes and the absence of cross-interference effects, allowing high throughput multianalysis on the same surface. Also, a comparison of the performance of two sensing strategies indicated that DVD disc and drive approach turned out in a simpler mode, the assays being more reproducible and with higher signal to noise ratios.     

培养因子对艾西丝南瓜芽增殖及不定根形成的影响

以艾西丝南瓜带芽茎段为外植体,研究了基本培养基、激素、糖、光照、培养基支持物等因子对芽增殖及不定根形成的影响。结果表明：艾西丝南瓜芽增殖的最佳培养条件为：MS+BA 0.5～1.0 mg/L+IAA 0.1～0.5 mg/L+食用白糖30 g/L,芽的月[KG(*9]增殖系数稳定在10左右;不定根诱导的适宜条件是：[KG)]1/2MS＋食用白糖20 g/L,生根率达86%;且自然散射光条件(1000～5000Lx)优于灯光(1000～2000Lx);以脱脂棉作生根培养基支持物效果优于琼脂,其芽增殖系数和生根率分别提高26%和7%。     

朱顶红鳞茎芽诱导及植株再生体系的建立

以朱顶红杂交品种红孔雀(Amaryllis vittatacv. Red Peacock)鳞茎盘切块为外植体,对影响鳞茎芽直接萌发增殖及生根的主要因素进行了研究,以建立其高效再生体系.结果表明:(1)1/2MS基本培养基有利于鳞茎芽的萌发和生长;切分方式不同对朱顶红鳞茎增殖具显著效应,偏离切分鳞茎可显著提高增殖系数,最高增殖系数可达6,平均增殖系数为3.6;而对半切分或未切分的增殖系数较低.(2)1/2MS 0.1 mg/L NAA 1.5 mg/L 6-BA 2.0 g/L活性炭的培养基利于鳞茎增殖;活性碳可显著提高鳞茎生根率,在1/2MS 0.1 mg/L NAA 2.0 g/L活性炭的培养基上生根率可达100%,而未加活性炭的生根率较低;再生植株经25 d的自然光照驯化后进行移栽,成活率达90%以上.     

Statistical optimization of culture media for growth and lipid production of <Emphasis Type="Italic">Botryococcus braunii</Emphasis> LB572

Botryococcus braunii has an outstanding ability to produce lipid; however, it is a slow-growing green microalgae. Statistical optimization of growth media was performed to faster growth and to increase lipid concentration. The effect of media composition on the growth of B. braunii LB572 was examined using fractional factorial design and central composite design. The media components examined include sodium carbonate, potassium phosphate, calcium chloride, magnesium sulfate, ferric citrate, and sodium nitrate. The results indicated that potassium phosphate and magnesium sulfate were major impact factors. The optimum concentrations of potassium phosphate and magnesium sulphate were found to be 0.058 and 0.09 g/L, respectively, for growth and 0.083 and 0.1 g/L, respectively, for lipid production. These values were validated using bubble column photobioreactors. Lipid productivity increased to 0.19 g/L/day in lipid-optimized media, with an average biomass productivity of 0.296 g/L/day and 64.96% w/w. In growth-optimized media, lipid productivity was 0.18 g/L/day, with an average biomass productivity of 0.304 g/L/day and 59.56% w/w.     

以糙米为原料流加L-蛋氨酸的S-腺苷蛋氨酸发酵优化

研究了S-腺苷甲硫氨酸(SAM)高产菌啤酒酵母S-W55的廉价培养基及分批补料发酵过程优化.对啤酒酵母S-W55生长和SAM产量影响最为重要的糙米水解糖和酵母粉进行了响应面优化,得到了最优化的配方为糙米水解糖51.4g/L、酵母粉4.74g/L,此条件下啤酒酵母S-W55的SAM产量达2.61 g/L.不同分批补料发酵...     

Development of suitable hydroponics system for phytoremediation of arsenic-contaminated water using an arsenic hyperaccumulator plant Pteris vittata

In this study, we found that high-performance hydroponics of arsenic hyperaccumulator fern Pteris vittata is possible without any mechanical aeration system, if rhizomes of the ferns are kept over the water surface level. It was also found that very low-nutrition condition is better for root elongation of P. vittata that is an important factor of the arsenic removal from contaminated water. By the non-aeration and low-nutrition hydroponics for four months, roots of P. vittata were elongated more than 500 mm. The results of arsenate phytofiltration experiments showed that arsenic concentrations in water declined from the initial concentrations (50?μg/L, 500?μg/L, and 1000?μg/L) to lower than the detection limit (0.1?μg/L) and about 80% of arsenic removed was accumulated in the fern fronds. The improved hydroponics method for P. vittata developed in this study enables low-cost phytoremediation of arsenic-contaminated water and high-affinity removal of arsenic from water.     

The Influence of Different Factors on Cell Growth and α-tocopherol Content of Culture from Carthamus tinctorius

Calli were induced from root, hypocotyl, cotyledon and flower bud of Carthamtus tinctorius. All calli had the capability to synthesize α-tocopherol. Among these calli, the hvpocotylcallus was better than others in cell growth rate and α-toeopherol content. Culture conditions could intensively influence the growth rate and α-tocopherol production of callus from Carthamus tinctorius. Sucrose (30g/L) was good for callus growth and glucose (30g/L) was good for α-tocopherol accumulation. High concentration (0.55%) of inositol could obviously stimulate both growth rate and α-tocopherol synthesis of callus. The inoculum quantity for best callus growth was 0.035 to 0.067g dry wt/flask (50ml volume). In addition, α-tocopherol content was effectively increased by culture callus in high CO2 concentration. Studies on optimum cuhure conditions of callus culture showed that the callus growth rate, α-tocopherol content and yield were 1.88, 2.03 and 3.30 times respectively as high as of the control by administration of 0.45%–0.55% inositol, 10% coconut milk, 0.1–0.5% casamino acid, 30g/L sucrose and 10g/L glucose.     

维生素在丙酮酸过量合成中的重要作用

研究了烟酸、硫胺素、吡哆醇、生物素和核黄素对一株光滑球拟酵母(\%Torulopsis glabrata\%) WSH\|IP303以葡萄糖为碳源、以氯化铵为唯一氮源生产丙酮酸的影响。利用正交试验方法,确证了硫胺素是影响WSH\|IP303生产丙酮酸的最重要因素。在硫胺素浓度一定(0.01～0.015mg/L)的前提下,提高烟酸浓度有助于加快耗糖速度。当烟酸、硫胺素、吡哆醇、生物素和核黄素的浓度分别为8、0.015、0.4、0.04和01mg/L时,摇瓶发酵48h,丙酮酸产量和产率可分别达到52.4g/L和0525g/g。采用优化的维生素组合方式,进行2.5L罐分批发酵,在初糖浓度120g/L的条件下发酵57.5h,丙酮酸产量和产率分别达到69.4g/L和0593g/g,分别比摇瓶培养的最好结果提高了32.%和13%。