生物表面活性剂生产Bacillus subtilis-1101发酵过程优化

应用中心组合试验设计和响应面分析方法对影响枯草芽孢杆菌Bacillus subtilis-1101产生表面活性剂的发酵过程进行优化。结果表明,枯草芽孢杆菌Bacillus subtilis-1101产生表面活性剂的最佳发酵条件为发酵温度29.1℃,初始pH值为4.9,装液量为56mL。在此条件下进行实验,结果最大排油圈为7.08cm,与模型预测值接近。说明响应面分析方法是优化表面活性剂生产的有力工具。     

Structural characterization of a biosurfactant produced by Bacillus subtilis at 45°C

Structural and biochemical characterization of a biosurfactant produced by Bacillus subtilis under thermophilic conditions was performed. Preliminary structural determination of CHCl₃/CH₃OH (65∶15) extracts by thin-layer chromatographic reagents showed it to be identical to surfactin. Also, the infrared, ¹H nuclear magnetic resonance, and mass spectroscopy analysis confirmed it to be identical to surfactin. Biochemically, the surfactant was a lipopeptide-containing lipid (17.05%) and protein (13.2%). The surfactant yielded a minimal aqueous surface-tension value of 28 dyne/cm and an interfacial tension value at 0.1% concentration of 0.2 dyne/cm against diesel oil. The critical micelle concentration of the surfactant was 35 mg/L. The biosurfactant exhibited an emulsification value (E ₂₄) of 90 against diesel oil and a sand-pack oil recovery of 62%. It has potential application in microbial-enhanced oil recovery in thermophilic, alkaline, acidic, and halophilic environments.     

Biosurfactant production by microorganisms on unconventional carbon sources

In recent years natural biosurfactants have attracted attention because of their low toxicity, biodegradability, and ecological acceptability. However, for reasons of functionality and production cost, they are not competitive with chemical surfactants. Use of inexpensive substrates can drastically decrease the production cost of biosurfactants. This review describes the use of unconventional carbon sources for biosurfactant production. These sources include urban as well as agroindustrial wastes. With suitable engineering and microbiological modifications, these wastes can be used as substrates for large-scale production of biosurfactants.     

Synthesis of enhanced biosurfactant by Bacillus subtilis MTCC 2423 at 45°C by foam fractionation

Production of a lipopeptide surfactant in a 6.5-L batch fermentor was carried out using Bacillus subtilis MTCC 2423 at 45°C. A good yield was obtained from sucrose (2%) substrate fermentation by continuous removal of the product by foam fractionation. The biosurfactant was recovered from collapsed foam by acid precipitation. The biosurfactant yield (4.5 g/L) was about 4.5 times higher than the yield (ca. 1 g/L) obtained by shake-flask fermentation. Surface activity of the collapsed foam was very high, and total surface activity was observed in the collapsed foam. The structural characterization of this biosurfactant produced at 45°C by the strain used in this study was recently reported. The biosurfactant was analyzed by thin-layer chromatography, infrared, ¹H nuclear magnetic resonance, and mass spectroscopy and was found to be identical to surfactin, a lipopeptide surfactant.     

Biomonitoring of biosurfactant production by green fluorescent protein‐marked Bacillus subtilis W1012

BACKGROUND: Biosurfactant production was investigated using two strains of Bacillus subtilis, one being a reference strain (B. subtilis 1012) and the other a recombinant of this (B. subtilis W1012) made able to produce the green fluorescent protein (GFP). RESULTS: Batch cultivations carried out at different initial levels of glucose (G₀) in the presence of 10 g L^?1 casein demonstrated that the reference strain was able to release higher levels of biosurfactants in the medium at 5.0≤G₀≤10 g L^?1 (B_max = 104–110 mg L^?1). The recombinant strain exhibited slightly lower levels of biosurfactants (B_max = 90–104 mg L^?1) but only at higher glucose concentrations (G₀ ≥ 20 g L^?1). Under these nutritional conditions, the fluorescence intensity linked to the production of GFP was shown to be associated with the cell concentration even after achievement of the stationary phase. CONCLUSION: The ability of the genetically‐modified strain to simultaneously overproduce biosurfactant and GFP even at low biomass concentration makes it an interesting candidate for use as a biological indicator to monitor indirectly the biosurfactant production in bioremediation treatments. Copyright © 2008 Society of Chemical Industry     

The use of agroindustrial by-products for biosurfactant production

Traditionally, hydrocarbons have been used for biosurfactant production. However, urban waste, peat pressate and agroindustrial by-products, such as olive oil mill effluent and acid whey, are possible substitutes for microbial growth and biosurfactant production. The state of the art has been reviewed, augmented by some new information onPseudomonas fermentation of olive oil mill effluent. More research is needed to improve yields and production economics.     

Foaming properties of surfactin,a lipopeptide biosurfactant fromBacillus subtilis

Foaming properties of surfactin were investigated and compared to those of sodium dodecyl sulfate (SDS) and bovine serum albumin (BSA). Foams were formed by a bubbling technique. Evolution of the foam volume and the liquid in the foam was monitored with optical and conductimetric methods to characterize foam formation and stability. Excellent foaming properties of surfactin were shown by its higher ability to form and stabilize the foam at a concentration as low as 0.05 mg/mL, in comparison with SDS and BSA. Surfactin produced a foam with intermediate maximum density and stabilized the liquid in foam, as well as BSA.     

乳酸菌发酵甘蔗糖蜜产乳酸的培养条件优化

乳酸是有广泛应用的有机酸,廉价原料是降低乳酸生产成本的重要因素。以实验室前期筛选的植物乳杆菌sy4为出发菌株研究了利用甘蔗糖蜜生产乳酸的培养条件。在确定发酵温度和初始pH后,通过Plackett-Burman实验和中心复合实验优化乳酸发酵条件。结果表明：植物乳杆菌sy4乳酸发酵的最适条件为温度32℃、60h、初始pH 6.5;Plackett-Burman实验表明酵母提取物、糖蜜和碳酸钙是影响乳酸产量的主要因素;中心复合实验得到三因素的最优组合为：酵母提取物13.19g/L、糖蜜476.63g/L和碳酸钙134.82g/L,确定了最适发酵培养基。在此条件下乳酸产量为（145.53±1.24） g/L,与模型预测值147.23g/L接近。本研究为以甘蔗糖蜜为原料生产乳酸提供了技术支持。     

Purification and characterization of thermophilic and alkalophilic tributyrin esterase fromBacillus strain A30-1 (ATCC 53841)

An extracellular esterase (EC 3.1.1.1) from a thermophilicBacillus A30-1 (ATCC 53841) was purified 139-fold to homogeneity by sodium chloride (6 M) treatment, ammonium sulfate fractionation (30–80%) and phenyl-Sepharose CL-6B column chromatography. The native enzyme was a single polypeptide chain with a molecular weight of about 65,000 and an isoelectric point at pH 4.8. The optimum pH for esterase activity was 9.0, and its pH stability range was 5.0–10.5. The optimum temperature for its activity was 60°C. The esterase had a half-life of 28 h at 50°C, 20 h at 60°C and 16 h at 65°C. It showed the highest activity on tributyrin, with little or no activity toward long-chain (12–20 carbon) fatty acid esters. The enzyme displayed K_m and K_cat values of 0.357 mM and 8365/min, respectively, for tributyrin hydrolysis at pH 9.0 and 60°C. Cyclodextrin (α, β, and γ), Ca²⁺, Co²⁺, Mg²⁺ and Mn²⁺ enhanced the esterase activity, and Zn²⁺ and Fe²⁺ acted as inhibitors of the enzyme activity. The enzyme activity was not affected by ethylenediaminetetraacetic acid, p-chloromercuribenzoate andN-bromosuccinimide. This paper was presented in part at the 82nd Annual Meeting and Exposition of the American Oil Chemists’ Society, held May 12–15, 1991, in Chicago, Illinois.     

利用甘蔗糖蜜半连续发酵生产琥珀酸

为获得较高的琥珀酸发酵产量和生产强度,对Actinobacillus succinogenes CGMCC1593两级双流半连续发酵甘蔗糖蜜生产琥珀酸的工艺过程进行了研究。通过对一级罐初始总糖浓度、补加培养基体积分数和批次发酵时间等发酵条件的优化,琥珀酸产量较分批发酵36 h提高12.9％,与补料分批发酵结果接近;生产强度较分批发酵和补料分批发酵分别提高111％和114%。     

Utilization of canola oil and lactose to produce biosurfactant withCandida bombicola

The prerequisites for a commercial fermentation process of biosurfactants include the use of low- or negative-cost substrates and maximum conversion yields. Under competitive market conditions, the price of canola oil is expected to decrease in response to its increased supply. Lactose, obtained from cheese whey, is a by-product of the dairy industry. In this work, canola oil with glucose or lactose as carbon sources was used as substrates to produce sophorose lipids (SLs) by means of the yeastCandida bombicola. Fermentations were conducted in either shaker flasks or 1-L Bellco (Vineland, NJ) stirred reactors for 5–7 d at 450 rpm and 30°C. The production of SLs reached 150–160 g/L in a medium consisting of 10% glucose, 10.5% canola oil, 0.1% urea and 0.4% yeast extract. When lactose was substituted for glucose, 90–110 g/L SL was obtained. The apolar SL 17-l-([2′-O-β-d-glucopyranosyl-β-d-glucopyransoyl]oxy)-octadecanoic acid 1′-4″-lactone 6′,6″-diacetate (SL-1) was the major one (73%) when canola oil was used instead of safflower oil (SL-1, 50%). Use of canola oil generally resulted in increased yields of SLs comparable to the yields obtained when safflower oil was used in the medium. Other literature reports present yields of 70 g/L and 120 g/L SLs, respectively, with these substrates.     

蕈菌云芝用于酒精废液生物漂白及收获菌丝体的研究

利用蕈菌云芝对不同稀释度酒精废液进行生物漂白及收获菌丝体研究。摇瓶培养结果表明,云芝对20%的酒精废液的脱色率为50%,生物量为8.1g/L。培养过程中培养液pH值呈上升趋势。利用5L发酵罐,通过控制发酵过程中pH值和通气量,培养54h,使脱色率上升到65%,生物量增加到10.9g/L,COD去除率为71%;通过检测发酵液中还原糖变化规律表明云芝对废水中的有色物质有分解利用的过程。与其他生物法处理酒精废液相比,云芝表现出较强的有机负荷承受能力、脱色能力及运行周期短的优势。     

Optimization of the production of chitosan from beet molasses by response surface methodology

Chitosan was produced by Rhizopus oryzae 00.4367 in shake flask culture and a stirred tank fermenter. Synthetic medium, treated and untreated beet molasses were used as cultivation media in shake flask cultures. In the stirred tank fermenter, the cultivation media were synthetic medium and untreated beet molasses. Shake flask culture containing untreated molasses with a sugar concentration of 40 g dm^?3 produced the maximum chitosan yield (961 mg dm^?3). Chitosan concentration reached its maximum value at the late exponential growth phase of R oryzae. In all experiments almost 8–10% of biomass and 32–38% of alkali‐insoluble material was extracted as chitosan. A central composite design was employed to determine the optimum values of process variables (aeration rate, agitation speed and initial sugar concentration) leading to maximum chitosan concentration in the stirred tank fermenter. In all cases, the fit of the model was found to be good. Aeration rate, agitation speed and initial sugar concentration had a strong linear effect on chitosan concentration. Moreover, the concentration of chitosan was significantly influenced by the negative quadratic effects of the given variables and by their positive or negative interactions. A maximum chitosan concentration of 1109.32 mg dm^?3 was obtained in untreated molasses medium containing an initial sugar concentration of 45.37 g dm^?3 with an aeration rate and agitation speed of 2.10 vvm and 338.93 rpm, respectively. Copyright © 2004 Society of Chemical Industry     

Economic production of Bacillus subtilis SPB1 biosurfactant using local agro‐industrial wastes and its application in enhancing solubility of diesel

BACKGROUND: The present work aimed to optimize a new economic medium for lipopeptide biosurfactant production by Bacillus subtilis SPB1 for application in the environmental field as an enhancer of diesel solubility. Statistical experimental designs and response surface methodology were employed to optimize the medium components. RESULTS: A central composite design was applied to increase the production yield and predict the optimal values of the selected factors. An optimal medium, for biosurfactant production of about 4.5 g L^?1, was found to be composed of sesame peel flour (33 g L^?1) and diluted tuna fish cooking residue (40%) with an inoculum size of 0.22. Increased inoculum size (final OD₆₀₀) significantly improved the production yield. The emulsifier produced was demonstrated to be an alternative to chemically synthesized surfactants since it shows high solubilization efficiency towards diesel oil in comparison with SDS and Tween 80. CONCLUSION: Optimization studies led to a strong improvement in production yield. The emulsifier produced, owing its high solubilization capacity and its large tolerance to acidic and alkaline pH values and salinity, shows great potential for use in bioremediation processes to enhance the solubility of hydrophobic compounds. © 2012 Society of Chemical Industry     

Novel sucrose lipid produced by Serratia marcescens and its application in enhanced oil recovery

Serratia marcescens MTCC 86 when grown on sucrose produced a novel sucrose lipid that was different from the amino lipid produced from n-alkane by other strains of S. marcescens. The sucrose lipid isolated by acetone precipitation technique was found to be predominantly a mixture of 3-(3′-hydroxytetradecanoyloxy) decanoate and 3-(3′-hydroxyhexadecenoyloxy) decanoate. This sucrose lipid exhibited excellent emulsification activity with a wide variety of hydrocarbons. It was effective in recovering up to 90% residual oil from the oilsaturated sandpack column suggesting its use in enhanced oil recovery. It also showed a strong ability to remove oil from the walls of the containers, indicating its potential use in cleaning operations.     

Effects of molasses grass,Melinis minutiflora volatiles on the foraging behavior of the cereal stemborer parasitoid,Cotesia sesamiae

Olfactory responses of the cereal stemborer parasitoid Cotesia sesamiae to volatiles emitted by gramineous host and nonhost plants of the stemborers were studied in a Y-tube olfactometer. The host plants were maize (Zea mays) and sorghum ( Sorghum bicolor), while the nonhost plant was molasses grass (Melinis minutiflora). In single-choice tests, females of C. sesamiae chose volatiles from infested and uninfested host plants and molasses grass over volatiles from the control (soil). In dual-choice tests, the wasp preferred volatiles from infested host plants to those from uninfested host plants. There was no discrimination between molasses grass volatiles and those of uninfested maize, uninfested sorghum, or infested maize. The wasp preferred sorghum volatiles over maize. Combining uninfested maize or sorghum with molasses grass did not make volatiles from the combination more attractive as compared to only uninfested host plants. Infested maize alone was as attractive as when combined with molasses grass. Infested sorghum was preferred over its combination with molasses grass. Local growth conditions of the molasses grasses influenced attractiveness to the parasitoids. Volatiles from Thika molasses grass were attractive, while those from Mbita molasses grass were not. Growing the Thika molasses grass in Mbita rendered it unattractive and vice versa with the Mbita molasses grass. This is a case of the same genotype expressing different phenotypes due to environmental factors.     

褪黑素诱导糖蜜废醪液培养单针藻Monoraphidium sp.FXY-10 产生物燃料

研究褪黑素对糖蜜酒精废醪液中单针藻Monoraphidium sp. FXY-10生长和油脂积累的影响。结果表明，以糖蜜酒精废醪液为基础培养基，添加10^-3μmol/L的褪黑素，微藻中的油脂含量达到68.69%，是对照组的1.15倍，且最高生物量为1.22g/L，相比对照组提高了41.86 %。与对照组相比，藻细胞内蛋白质和碳水化合物含量分别是对照组的0.61和1.41倍。在微藻培养过程中，糖蜜废醪液中的COD、总氮及总磷的清除率分别达到92.33%、90.07%和86.04%。此外，外源褪黑素的添加提高了乙酰辅酶A羧化酶（acetyl coenzyme A carboxylase，ACCase）、苹果酸酶（malic enzyme，ME）的活性，而降低了磷酸烯醇式丙酮酸羧化酶（phosphoenolpyruvate carboxylase，PEPC）的活性。初步技术经济分析表明，1kg单针藻Monoraphidium sp. FXY-10的干燥粉可以制备535.8g生物柴油。研究表明，褪黑素可以促进糖蜜酒精废醪液中微藻的生长和油脂的积累，降低培养成本，为微藻扩大化生产提供了新的理论基础。