SAM产生菌酿酒酵母HYS98的发酵方式

分别研究了分批发酵、恒速流加发酵、变速流加发酵和分批补料发酵4种操作方式下酿酒酵母HYS98发酵生产SAM的代谢特征,并考察了不同发酵方式对发酵结果的影响. 结果表明,酿酒酵母HYS98发酵生产SAM的最佳发酵模式为分批补料发酵,SAM的平均产率和菌体生长速率分别达0.123和1.030 g/(L×h),比国内外文献报道的最高水平分别提高了68%和73%,为发酵工艺进一步优化奠定了基础.     

重组HIV-1 DNA疫苗工程菌中试发酵工艺的优化

目的优化重组HIV-1DNA疫苗工程菌的中试发酵工艺。方法优化质粒转化条件,制备工程菌Jm109/pVAX1-MEGp24种子液,连续传30代,检测其遗传稳定性;优化培养基成分、补料基质、补料方式和培养时间,确定最佳发酵参数;并应用最佳发酵参数,于50L发酵罐进行3批中试规模的发酵,考察在发酵培养过程中质粒的稳定性和超螺旋质粒的比例。结果工程菌在连续传代30次后,质粒拷贝数基本保持稳定;最佳发酵参数为:采用以甘油为碳源的培养基,以梯度恒速流加方式补料,培养时间13h;通过3批稳定发酵,最终可获得湿菌67.0～68.6g/L,质粒含量可达1.62～1.73mg/g菌,超螺旋质粒的比例达93%以上。结论已建立了稳定的重组HIV-1DNA疫苗工程菌中试发酵工艺,为进一步规模化生产奠定了基础。     

治疗性双质粒HBV DNA疫苗工程菌的中试发酵工艺研究

目的研究双质粒HBV DNA疫苗工程菌的中试发酵工艺。方法首先通过计算质粒拷贝数,检测工程菌DH5α/pS2.S和DH5α/pFP在传代过程中的遗传稳定性,通过摇瓶培养确定培养基组成,再在30L发酵罐内,通过改变培养基成分、培养时间、补料方式,确定最佳发酵参数。并将确定的最佳发酵参数应用于50L发酵罐连续3批中试规模的发酵,同时考察在发酵培养过程中质粒稳定性和超螺旋质粒DNA的比例。结果工程菌DH5α/pS2.S和DH5α/pFP在连续传代30次后,质粒拷贝数保持稳定。确定最佳发酵参数为以甘油为碳源的培养基培养,梯度恒速流加方式补料,培养时间为10h。通过3批稳定发酵,最终可获得湿菌58.0~71.8g/L,质粒含量可达到1.11~1.58mg/g菌,超螺旋质粒DNA的比例达93%以上。结论已建立了稳定的双质粒HBV DNA疫苗工程菌中试发酵工艺,为进一步规模化生产奠定了基础。     

重组大肠杆菌表达双功能谷胱甘肽合成酶的发酵工艺研究

对重组表达双功能谷胱甘肽合成酶（GshF）的大肠杆菌进行了5L罐的发酵工艺探索。分别考察了不同培养基、诱导剂种类、诱导时机、诱导温度、补料流加碳源类型等对菌体生长、GshF蛋白表达及其酶活的影响。结果表明,采用葡萄糖进行补料流加．在优化工艺条件下,GshF表达量达到1．94g·L^-1。     

西索米星发酵工艺条件的优化

考察了氯化钴、蛋氨酸、磷酸盐和可发酵糖对西索米星分批发酵的影响,优化了西索米星分批发酵工艺条件. 研究表明,初始发酵培养基中添加6~10 mg/L氯化钴或1.0~2.0 g/L蛋氨酸有助于西索米星的合成,蛋氨酸在产物合成前期添加的效果最佳. 高浓度磷酸盐会提高发酵液中淀粉水解酶活力和丙酮酸浓度、降低碱性磷酸酯酶活力和抑制西索米星的合成. 应分段控制发酵液中的磷酸盐浓度,在菌体生长期应控制在3.14 mmol/L以内,在产物合成期应控制在0.10 mmol/L以下. 与分批发酵过程相比,当以麦芽糖或淀粉水解液为补料液、将发酵液中可发酵糖浓度控制在8.5~11.5 g/L时,不论采用变流率还是恒流率的流加发酵方式均有助于大幅度提高西索米星发酵水平.     

补料—批式发酵生产青霉素G的最优控制策略比较

补料－批式发酵通常有两种最优控制策略：以底物的流加速率和流加底浓度作为控制和调节变量。本文以补料－批式发酵生产青霉素Ｇ为模型体系，运用敏感性分析模型比较了种最优控制策略。从系统的易控性、稳定性和最优解的可得性角度研究指出：以流加底物浓度作为最优控制和调节变量具有显著的优越性。     

产放线酮链霉菌30L发酵罐葡萄糖流加发酵实验研究

在前期工作中我们采用了正交实验设计法、均匀实验设计法和葡萄糖流加发酵对一株产放线酮链霉菌发酵培养基及发酵方式进行了优化,并采用化学及物理诱变方法选育出了遗传型比较稳定的生产菌株,在以上工作的基础上摇瓶条件下放线酮的平均产量已经达到了621μg/mL比优化前提高了38%。本实验对放线酮30升罐放大发酵及30 L罐葡萄糖以流加的方式发酵进行初步探索,为日后放线酮的规模化生产提供依据。     

营养物及补料方式对重组酵母产α-淀粉酶的影响

研究了两段培养中基质浓度、配比及天然氮源添加方式对重组酵母发酵生产(-淀粉酶的影响。结果表明,重组酵母发酵适宜的初糖浓度为7g·L-1,葡萄糖和基础氮配比为7:4,生长期补料碳氮比为5:1,且产酶期供给天然氮源以少量多次脉冲补加酵母膏为宜。于2L发酵罐中培养,证明脉冲补加酵母膏、流动添加基本营养物的补料分批培养方式是有效的,指数流加基本营养物较之恒流量流加更能提高菌体浓度和(-淀粉酶表达水平。培养60小时细胞量达24.5g·L-1,酶活力达97.3U·mL-1。     

用于谷胱甘肽合成的重组大肠杆菌的培养和诱导表达

研究了用于谷胱甘肽合成的温度诱导型重组大肠杆菌的补料分批发酵,通过葡萄糖的脉冲补入,结合溶氧反馈控制技术,实时控制发酵过程中乙酸的产生. 在此基础上,进一步考察了在谷胱甘肽合成酶系诱导表达阶段,流加酵母粉和蛋白胨时重组大肠杆菌的补料分批发酵过程. 结果发现,脉冲补料-溶氧反馈控制技术在流加酵母粉和蛋白胨的补料分批发酵中仍能得到很好的应用,乙酸浓度可被有效地控制在2.5 g/L以下. 通过比较诱导表达阶段流加不同组成的补料液成分,发现添加适量酵母粉和蛋白胨可促进谷胱甘肽合成酶系的表达,在42℃诱导4 h收获的重组大肠杆菌酶法合成谷胱甘肽达到2.46 g/L.     

根癌土壤杆菌产辅酶Q10的补料分批发酵工艺

以根癌土壤杆菌(Agrobacterium tumefaciens Q14)为生产菌株,在15 L发酵罐内对补料分批发酵生产辅酶Q10的工艺进行优化.通过逐步添加限制性营养物质,考察其对发酵的影响,获得了较优的补料分批发酵工艺,即流加葡萄糖和KH2PO4溶液,将葡萄糖和磷浓度分别控制在5～15 g/L和50～120 m...     

Optimization of lipase production by Penicillium simplicissimum in soybean meal

BACKGROUND: Lipase production by Penicillium simplicissimum using soybean meal as substrate has been investigated. A factorial design technique was used to evaluate the effects of incubation temperature, initial moisture of the meal and substrate supplementation with low cost supplements, on lipase production. Soybean oil and wastewater from a slaughterhouse, which is rich on oil and fat, corn steep liquor and yeast hydrolysate, were tested as supplementary carbon and nitrogen sources. RESULTS Cultivation conditions were optimized for the production of lipase by factorial design and response surface methodology. Results show that the microorganism produces very low protease activity (0.21 U gds^?1 dry substrate), which helps to maximize lipase production. Soybean meal without supplements appears to be the best medium of those tested for lipase production by P. simplicissimum. CONCLUSION: This work showed that temperature and moisture are the factors that most strongly influence lipase production by P. simplicissimum using soybean meal as substrate. The growth conditions that optimize lipase production are 27.5 °C using substrate with 550 g kg^?1 of initial moisture. In optimum conditions lipase activity of 30 U gds^?1 dry substrate was obtained. Copyright © 2007 Society of Chemical Industry     

Transfer of Lauric and Myristic Acid from Black Soldier Fly Larval Lipids to Egg Yolk Lipids of Hens Is Low

Implementing insects, such as the black soldier fly larvae (BSFL), as animal feed commonly includes the previous removal of substantial amounts of fat. This fat may represent an as yet underutilized energy source for livestock. However, transfer of lauric and myristic acid, prevalent in BSFL fat and undesired in human nutrition, into animal-source foods like eggs may limit its implementation. To quantify this, a laying hen experiment was performed comprising five different diets (10 hens/diet). These were a control diet with soybean oil and meal and a second diet with soybean oil but with partially defatted BSFL meal as protein source. The other three diets were based on different combinations of partially defatted BSFL meal and fat obtained by two different production methods. Lauric acid made up half of the BSFL fat from both origins. Both BSFL fats also contained substantial amounts of myristic and palmitic acid. However, in the insect-based diets, the net transfer from diet to egg yolk was less than 1% for lauric acid, whereas the net transfer for myristic and palmitic acid was about 30% and 100%, respectively. The net transfer did not vary between BSFL originating from production on different larval feeding substrates. The results illustrate that hens are able to metabolize or elongate very large proportions of ingested lauric acid and myristic acid, which are predominant in the BSFL lipids (together accounting for as much as 37 mol%), such that they collectively account for less than 3.5 mol% of egg yolk fatty acids.     

脱脂豆粉催化合成大豆油氢过氧化物

研究了脱脂豆粉催化合成大豆油氢过氧化物的反应。考察了豆粉过筛目数、温度、豆粉浓度、pH值对大豆油氧化的影响。最佳反应条件为豆粉过筛目数80目,豆粉浓度180 g/L,氧化温度10℃,缓冲液pH=9,反应时间3 h,大豆油氢过氧化物产率61.9%。在相同条件下(除反应时间),精酶液催化反应1.5 h达平衡,大豆油氢过氧化物产率54.8%。比豆粉催化最佳产率降低了7.1%。     

脱脂豆粉催化合成氢过氧化玉米胚芽油

以植物催化剂脱脂豆粉替代大豆氧合酶催化合成了氢过氧化玉米胚芽油。考察了豆粉粒度、豆粉浓度、温度以及pH值对氢过氧化玉米胚芽油得率的影响。所得最佳反应条件为:80目豆粉,豆粉浓度200 g/L,反应温度10℃,缓冲液pH为9,反应时间3 h,氢过氧化玉米胚芽油得率为61.9%。脱脂豆粉重复利用3次后,催化反应产物得率下降为32.1%,此后呈缓慢下降趋势。     

Exploring seed and meal composition and protein solubility in soybean genotypes from different domestication periods

Traditionally, commercial soybean breeding has focused on increasing seed yield by crossing elite cultivars and limiting the genetic diversity within commercial germplasm. Wild and ancestral soybean genotypes have higher seed protein concentrations than commercial ones. Different seed protein concentrations and compositions result in diverse functional properties of soybean meal, in particular solubility is important for beverages and protein isolates production. The objectives of our study were (i) to characterize seed protein concentration and composition in genotypes from different soybean domestication periods (types) and (ii) to evaluate the protein concentration and solubility profiles of the defatted meals obtained from these genotypes. Variation within seed and meal protein concentration, composition, and solubility was evident along the domestication process. Wild relative (G. soja) and Elite genotypes had the maximum and minimum seed protein concentrations, respectively (42.9 and 36.3 g 100 g⁻¹). Soybean meal protein concentrations were 55.1, 47.7, 48.4 and 44.1 g 100 g⁻¹ for Wild relative (G. soja), Asian landraces, North American (Nam) ancestors and Elite, respectively. Ample genotypic variation was observed for β-conglycinin components, such as for β, α, and α′ subunits and for total glycinin and its components. Asian landraces had the highest protein solubility. Wild and ancestral germplasm are a reservoir of useful traits to improve soybean seed quality. This study opens the gates to the introduction of ancestral germplasm to breeding programs focused on protein quality and functionality.     

Efficient production of di- and tri-acylated mannosylerythritol lipids as glycolipid biosurfactants by Pseudozyma parantarctica JCM 11752(T)

Mannosylerythritol lipids (MELs) are one of the most promising biosurfactants known, because of their multifunctionality and biocompatibility. In order to attain an efficient production of MELs, Pseudozyma parantarctica JCM 11752(T), which is a newly identified strain of the genus, was examined for the productivity of MELs at different culture conditions. The yeast strain showed significant cell growth and production of di-acylated MELs even at 36 degrees C. In contrast, on conventional high-level MEL producers including P. rugulosa, the MEL yield considerably decreased with an increase of the cultivation temperature at over 30 degrees C. On P. parantarctica, soybean oil and sodium nitrate were the best carbon and nitrogen sources, respectively. Under the optimal conditions on a shake-flask culture at 34 degrees C, the amount of di-acylated MELs reached over 100 g/L by intermittent feeding of only soybean oil. Interestingly, the yeast strain produced tri-acylated MELs as well as di-acylated ones when grown on the medium containing higher soybean oil concentrations than 8% (vol/vol). The production of tri-acylated MELs was significantly accelerated at between 34 and 36 degrees C. With 20 % (vol/vol) of soybean oil at 34 degrees C, the yield of tri-acylated MELs reached 22.7 g/L. The extracellular lipase activity considerably depended on the culture temperature, and became the maximum at 34 degrees C; this would bring the accelerated production of tri-acylated MELs. Accordingly, the present strain of P. parantarctica provided high efficiency in MEL production at elevated temperatures compared to conventional MEL producers, and would thus be highly advantageous for the commercial production of the promising biosurfactants.     

Bland undenatured soybean flakes by alcohol washing and flash desolventizing

The conditions for removing the beany and bitter flavor from defatted soybean meal with ethanol and isopropyl alcohol were investigated. In the presence of alcohols, soybean protein is extremely sensitive to denaturation when temperature, moisture, and residence time are increased. If protein is to be isolated in good yield and quality, retention of the original, high watersolubility is important, and denaturation must be kept to a minimum. Defatted soybean flakes were successfully debittered by countereurrent washing with aqueous alcohols on a pilot-plant scale, and the entrained solvent was recovered by flash desolventizing without excessive denaturation of protein. Effective debittering was obtained with 95 volume percentage ethanol and 91 volume percentage isopropyl alcohol whereas satisfactory flavor was not obtained with absolute ethanol. The solubility of the nitrogenous compounds in the meal product (Nitrogen Solubility Index—NSI=water-soluble nitrogen×100÷ total nitrogen) was maintained at 68 NSI, or higher, regardless of the solvent system or conditions used when starting with 80 NSI defatted flakes. Residual alcohol in the desolventized products was reduced to 1–2% with the aqueous alcohol system and to less than 1% for the absolute alcohol system. Lower residual values can be obtained by recycling the material through the desolventizing unit. The desolventizing system described is simple, low in cost, and should be useful in any process requiring the rapid removal of solvent from residual solids where heat-sensitive constituents are present.     

Detection of soybean antigenicity and reduction by twin-screw extrusion

Antiserum toward soybean meal was prepared by feeding a soybean meal diet containing large amounts of antigens to calves. The antiserum was used to develop a sensitive detection method for soybean meal antigens by competitive inhibition enzyme-linked immunosorbent assay. With this method, the effectiveness of twin-screw extrusion cooking in reducing antigenicity in soybean meal was examined. Antigenicity was reduced to 0.1% of the original activity by extrusion cooking with screws containing kneading-disc elements and die-end temperatures exceeding 66°C. Electrophoretic analysis of the cooked meal indicated that the reduction in antigenicity was due to degradation of protein structures, particularly those with molecular weights exceeding 40 kDa.     

Functional properties of soybean and lupin protein concentrates produced by ultrafiltration-diafiltration

Ultrafiltration followed by diafiltration (UF-DF) was evaluated for the production of protein products from partially defatted soybean meal or undefatted lupin (Lupinus albus L.2043N) meal. This study determined the effects of UF-DF on functional properties of the extracted proteins and compared the results with those of protein prepared by acid-precipitation (AP). UF-DF produced only protein concentrates (73% crude protein, dry basis, db), while AP produced protein isolates (about 90% crude protein, db). Soybean protein produced by UF-DF showed markedly higher values for solubilities up to pH 7.0, surface hydrophobicity index, emulsion activity index, and foaming capacity than did the AP soybean protein. UF-DF soy protein was also the most heat-stable among all protein samples tested. With lupin proteins, only the surface hydrophobicity and emulsion activity indices were significantly improved by using UF-DF. UF-DF generally had no adverse effects on, and in most cases even improved, the functional properties of soy protein concentrate produced by this method. UF-DF did not produce a comparable improvement in functional properties of lupin proteins as it did for soybean protein.     

Pancreatic lipase activity in emulsions containing seed meals: Effect of extrusion

The effect of extrusion of soybean (Glycine max), yellow lupin (Lupinus luteus), and linseed (Linum usitatissimum) on the activity of porcine pancreatic lipase (PPL) was evaluated through pH‐Stat methodology with tributyrin emulsions in which the seed meals [extruded (E) and non‐extruded (NE)] were dispersed. The seed meals affected PPL activity in a concentration‐dependent fashion; at 2.3 mg/mL the meals reduced PPL activity (11.7–45.0% reduction) while the opposite was true (3.1–28.0% increase) at 6.8 mg/mL. PPL activity in the assays containing E‐lupin, E‐soybean and E‐linseed meals at 2.3 mg/mL was 109.7 and 11.3% higher and 25.5% lower than the activity in the emulsions containing the NE‐meals, respectively. The differences between the PPL activity in emulsions containing the extruded and NE‐meals could be due to changes in the emulsifying properties of the meals after extrusion; extrusion increased (10.2%) the emulsification capacity of lupin meal and for linseed meal this property was reduced (9.2%). In spite of the low PPL activity at low meal concentrations, the percentage of free fatty acids released after 30 min was similar to the theoretical maximum (66.6%) with two exceptions – NE‐soybean meal (59.5 ± 2.7%) and E‐linseed meal (57.9 ± 1.2%). Practical applications: Feed and food industries use plant protein sources that contain inhibitors of digestive enzymes. Various proteases inhibitors are inactivated during extrusion being less known the effect of extrusion on lipase inhibitors. Reduction of lypolysis rate, carried out mainly by pancreatic lipase in human and other animals, could influence feed and food consumption because satiety is controlled – at least in part – by undigested lipids. Our results show that extrusion can be used to reduce the inhibitory activity of lupin and soybean on PPL. On the other hand, extrusion of linseed increased the negative effect of this meal on PPL activity effect that could have applications in the development of food and ingredients for weight control.