Sporidiobolus pararoseus WZ012产类胡萝卜素发酵条件的优化

以响应面方法对Sporidiobolus pararoseus WZ012产类胡萝卜素的摇瓶发酵条件进行了优化。在Plackett-Burman实验的基础上,利用中心组合设计研究了4个主要因素(葡萄糖、酵母膏、KH2PO4、接种量)对产类胡萝卜素的影响。结果表明,较高浓度的酵母膏和较大的接种量有助于类胡萝卜素的生产,而较高浓度的葡萄糖和KH2PO4有助于生物量的提高,生物量与类胡萝卜素产量之间没有必然的联系。优化得到的发酵培养基组成(g/L)为:葡萄糖60.23、酵母膏12.21、KH2PO41.68、接种量为8.38%。在此条件下红酵母产类胡萝卜素的最大产量为29.3mg/L,较优化前提高了54.1%。类胡萝卜素成分分析表明主要含有β-胡萝卜素、圆酵母素和红酵母红素这3种类胡萝卜素物质,其中β-胡萝卜素占62.5%。     

不同碳源对粘性红酵母WP3生长及类胡萝卜素产量的影响

研究了不同碳源及含量对粘性红酵母（Rhodotorula mucilaginosa）WP3生长及类胡萝卜素积累的影响。结果表明,在所研究的碳源中,葡萄糖有利于生长,而果糖有利于类胡萝卜素积累。在碳源含量为40 g/L,果糖和葡萄糖比例为7∶1,果糖含量为35 g/L,葡萄糖含量为5 g/L时,粘性红酵母WP3的生物量为7.94,类胡萝卜素产量达到556.84 μg/L,对粘性红酵母生长及类胡萝卜素合成有利。     

Extraction of carotenoids from Phaffia rhodozyma: A comparison between different techniques of cell disruption

The yeast Phaffia rhodozyma is known for producing carotenogenic pigments, commonly used in aquaculture feed formulation as well as in cosmetic, pharmaceutical, and food industries. Despite the high production of carotenoids from microorganisms by biotechnology, their use has limitation due to the cell wall resistance, which constitutes a barrier to the bioavailability of carotenoids. Therefore, there is a need to improve carotenoids recovering technique from microorganisms for the application of food industries. This study aimed to compare mechanical, chemical, and enzymatic techniques of cell disruption for extracting carotenoids produced by P. rhodozyma NRRL Y-17268. Among the techniques studied, the highest specific concentration of carotenoids (190.35 μg/g) resulted from the combined techniques of frozen biomass maceration using diatomaceous earth and enzymatic lysis at pH of the reaction medium of 4.5 at 55°C, with initial activity of β-1,3 glucanase of 0.6 U/mL for 30 min.     

细胞破壁对螺旋藻藻蓝蛋白提取效果的影响

螺旋藻细胞破壁是藻蓝蛋白提取的关键工序。采用溶胀法、超细剪切法、超声波法、反复冻融法、溶胀—超细剪切法和溶胀—超细剪切—超声波法对螺旋藻细胞进行破壁处理;通过对比分析破壁后提取液中的藻蓝蛋白得率来评价破壁方法的优劣。结果表明,溶胀法、超细剪切法、超声波法、反复冻融法、溶胀—超细剪切法、溶胀—超细剪切—超声波法得到的最高藻蓝蛋白得率分别为8.90%,7.38%,8.00%,8.26%,9.22%,8.88%。综合考虑,溶胀—超细剪切法相对其它5种破壁方法而言,其操作简单便捷,破壁效果更加显著,藻蓝蛋白得率高;溶胀—超细剪切法最佳的提取工艺为:溶胀时间12h,剪切时间5min。     

研磨法破碎小球藻细胞工艺优化

选取破碎过程中的关键因素,以油脂得率为评价指标,通过进行单因素试验和正交试验,对研磨法破碎小球藻细胞的工艺条件进行了研究。结果表明,小球藻细胞破碎最佳工艺条件为藻液质量浓度150 g/L,破碎时间2.5 h,球磨机转速400 r/min,藻液与钢珠的质量比为3:4。在此工艺条件下,能达到较好的破碎效果,油脂得率为46.89%。     

Screening and HPLC methods for carotenoids in sweetpotato, cassava and maize for plant breeding trials

Analytical methods for sweetpotato, cassava and maize were developed. In orange and salmon-fleshed sweetpotatoes, (all-E)-β-carotene predominated and results of spectrophotometric screening and HPLC quantification did not differ significantly. In yellow-fleshed sweetpotato and cassava, however, spectrophotometric screening overestimated the HPLC values because of the presence of several minor carotenoids. Aside from (all-E)-β-carotene, Z-isomers were present in cassava in appreciable amounts. For both crops, extraction with acetone or tetrahydrofuran:methanol (1:1), using a mortar and pestle or a Polytron homogenizer, gave equivalent results. Rehydration of dry maize at room temperature for 30 min or at 85 °C for 5, 10 or 15 min gave equivalent results. Concentrations obtained with the C18 and C30 columns did not differ significantly for zeaxanthin, lutein, β-cryptoxanthin and β-carotene in the all-E-configuration, but their Z-isomers were difficult to locate in the chromatogram obtained with the C30 column. Extraction with tetrahydrofuran:methanol (1:1) gave significantly lower results for zeaxanthin and lutein.     

蜂花粉破壁的研究进展

以油菜蜂花粉为主要原料,以蜂蜜中选育出的蜜生假丝酵母LGL-2为试验菌株,研究不同发酵时间、发酵温度、接种量和底物含量对油菜蜂花粉酵素总酚含量的影响。以总酚含量为指标,采用响应面法优化油菜蜂花粉酵素发酵工艺条件。在此条件下,进一步比较分析油菜蜂花粉及其酵素的酚类化合物、挥发性风味成分和抗氧化活性。结果表明：油菜蜂花粉酵素的最佳发酵工艺参数为发酵时间48 h、发酵温度30 ℃、接种量20%、底物含量30%,此时油菜蜂花粉酵素总酚含量可达11.48 mg GAE/g。采用高效液相色谱（HPLC）分析酚类化合物,发现其内包含没食子酸、对香豆酸、芦丁、鞣花酸和槲皮素,各酚类化合物含量经蜜生假丝酵母LGL-2发酵后均有所提高。采用相对气味活度值法（ROAV）结合顶空固相微萃取-气相色谱-质谱技术（HS-SPME-GC-MS）对油菜蜂花粉及其酵素的挥发性香气成分进行测定分析,共检测出26种挥发性香气成分,其中7种被鉴定为油菜蜂花粉酵素关键性挥发性香气成分,部分醇类和酯类物质含量提升为产品带来优良风味。抗氧化活性试验表明,相比于发酵前,油菜蜂花粉酵素具有更强的DPPH自由基清除能力（DPPH·）、ABTS⁺自由基清除能力（ABTS⁺·）以及铁离子还原能力（FRAP）。综上,经蜜生假丝酵母LGL-2发酵能有效提高油菜蜂花粉的酚类物质含量、风味品质及抗氧化活性。

     

超声波法提取沙棘中类胡萝卜素条件的优化

在超声波单因素试验结果基础上,进行L9(33)正交试验,研究了超声波法提取沙棘类胡萝卜素最佳工艺条件.结果表明,以丙酮/石油醚(2∶1,v/v)作提取剂,在超声波功率为300W,料液比1∶5(g/mL),室温下提取20 min能较充分提取沙棘中类胡萝卜素,相对提取率可以达到96.5%.     

Integration of peroxide delignification and sulfamic acid sulfation methods for obtaining cellulose sulfates from aspen wood

A new method to obtain cellulose sulfates from available and inexpensive raw material—aspen wood was developed. This method integrates catalytic peroxide delignification and sulfamic acid sulfation stages. Solvents such as acetic acid and water were used for isolation of pure cellulose by wood peroxide delignification with TiO₂ catalyst. Low-aggressive and less-toxic sulfating agent—sulfamic acid–urea mixture was used to obtain cellulose sulfates.     

Comparison of isolation methods using commercially available kits for obtaining extracellular vesicles from cow milk

Extracellular vesicles (EV) are important for delivering biologically active substances to facilitate cell-to-cell communication. Milk-derived EV are widely known because of their potential for immune enhancement. However, procedures for isolating milk-derived EV have not been fully established. To obtain pure milk-derived EV and accurately reveal their function, such procedures must be established. The aim of the present study was to compare methods using commercially available kits for isolating milk-derived EV. Initially, we investigated procedures to remove casein, which is the major obstacle in determining milk-derived EV purity. We separated whey using centrifugation only, acetic acid precipitation, and EDTA precipitation. Then, we isolated milk-derived EV by ultracentrifugation, membrane affinity column, size exclusion chromatography (SEC), polymer-based isolation, or phosphatidylserine-affinity isolation. Using EV count per milligram of protein, which is a good indicator of purity, we determined that acetic acid precipitation was the best method for removing casein. Using nanoparticle tracking analysis, protein quantity analysis, and RNA quantity analysis, we comprehensively compared each isolation method for its purity and yield. We found that SEC-based qEV column (Izon Science) could collect purer milk-derived EV at higher quantities. Thus, a combination of acetic acid precipitation and qEV can effectively isolate high amounts of pure extracellular vesicles from bovine milk.     

Assessment of conventional and microwave heating induced degradation of carotenoids in olive oil by VIS Raman spectroscopy and classical methods

Raman spectroscopy, as a fast non-destructive method, has shown its potential in revealing heat-induced degradation of extra virgin olive oil carotenoids during microwave versus conventional heating processes. A progressive degradation in carotenoids was observed, starting at 180 °C and 140 °C in microwave and conventional heating processes respectively; this was followed by a rapid degradation at 180 °C only with conventional heating. As the main difference, the Raman bands due to carotenoids completely disappeared at 203 °C with conventional heating, while these bands could still be observed even up to 225 °C with microwave heating. Furthermore a loss of cis double bonds and slightly changes in free fatty acid was also observed in both heating processes. A precise calibration model was established using partial least squares regression, which can be used for monitoring carotenoids content degradation during heating. The accuracy of the model was estimated using the root mean square errors and the correlation coefficient.     

Comparison of NIR and MIR spectroscopic methods for determination of individual sugars,organic acids and carotenoids in passion fruit

In this work, NIR and MIR spectroscopy was investigated and compared for predicting passion fruit ripening parameters as sugars, organic acids and carotenoids. Spectra of 56 samples of the lyophilized passion fruit were collected using an integrating sphere in NIR range and attenuated total reflectance accessory in MIR range. Individual sugars (sucrose, glucose and fructose), organic acids (malic and citric acids) and carotenoids (β-carotene) contents were determined by reference methods. Spectral and reference data were analyzed by principal component analysis. Partial least square regression (PLSR) was used to establish calibration models. MIR technique was better than the NIR technique for glucose (R²_v = 0.942), fructose (R²_v = 0.855), sucrose (R²_v = 0.818), total sugar (R²_v = 0.914) and citric acid (R²_v = 0.913) content determination. On the other hand, NIR was superior for total acids (R²_v = 0.903) content determination. For malic acid and β-carotene contents both methods were unsatisfactory due to low concentrations of these constituents in the passion fruits.     

霍氏肠杆菌A20降解类胡萝卜素的条件优化

为了提高类胡萝卜素的降解率,以霍氏肠杆菌（Enterobacter hormaechei）A20菌株为对象,研究其培养条件(培养时间、培养温度、转速、pH值、接种量)对类胡萝卜素降解率的影响,并通过单因素和正交试验,以降解率为评价指标,确定微生物的最佳培养条件。结果表明,培养条件的改变可以影响类胡萝卜素的降解效果,各培养条件对类胡萝卜素的降解率的影响程度不同。菌株A20降解类胡萝卜素的最佳培养条件为培养时间42 h,温度25 ℃,转速150 r/min,pH值7.0,接种量1.0%,在此条件下,类胡萝卜素降解率从最初的70.80%提高至91.53%。     

Yeast cell disruption strategies for recovery of intracellular bio-active compounds — A review

Yeasts are cheap, attractive and easily available residual sources of valuable bio-active compounds. Extraction of these compounds requires to break the yeast cells. So efficient damage of cell wall has become an important issue to be resolved. The aim of this paper is to review the potential of some emerging cell disruption techniques for recovery of intracellular bio-active compounds from Baker's yeast including mechanical (bead mill, high pressure homogenization, ultrasonication), and non-mechanical (electrical, physical, chemical and enzymatic) techniques, as well as some newly developed methods. The advantages and drawbacks of different cell disruption methods were summarized by considering the energy consumption, the interaction of the disruption methods with downstream operations and the process economics of alternative strategies. Finally, some future directions for research areas are proposed.Industrial relevanceWine making process entails the generation of significant amount of waste yeast, which represents an attractive source of valuable compounds that has been relatively unexploited to date. To retain the valuable cell content, effective cell disruption strategies are needed to break the rigid yeast cell walls. This review summarizes the state of the art of some emerging cell disruption techniques for recovery of intracellular bio-active compounds from yeasts including mechanical (bead mill, high pressure homogenizer, ultrasonication), and non-mechanical (electrical, physical, chemical and enzymatic) techniques. Thereby, it identifies the process economics of alternative strategies by considering the interaction of the disruption methods with downstream operations as well as the current situations and future research needs.     

Optimisation of growth parameters for the production of carotenoids by Rhodotorula gracilis

 Response surface methodological studies of growth parameters for carotenoid production by Rhodotorula gracilis (ATCC 90950) have been carried out for three parameters employed, namely glucose concentration (%), incubation period (days) and volume of inoculum (ml/100 ml). The results indicated maximum carotenogenesis (0.054%) at a 10% glucose level with a 2 ml/100 ml volume of inoculum for an incubation period of 9 days, which was also verified by experimental data. Received: 25 May 1998     

反相高效液相色谱法测定烟草中的类胡萝卜素及其异构体

采用反相高效液相色谱−二极管阵列检测器（RP−HPLC−DAD）分离和测定烟草中类胡萝卜素及其异构体的组成和含量。烟草样品经过含有0.1%丁基羟基甲苯（BHT）的冰丙酮溶液萃取,浓缩后,经Zorbax SB C18色谱柱分离。流动相组成:A,乙腈−水（体积比为88:12）;B,乙酸乙酯。梯度洗脱程序:0 − 25min,100%A;25 − 50 min,B由0%线性增加为60%;50 − 55 min, 40%A+ 60% B;55 − 60 min,A由40%线性增加为100%。 检测波长:450 nm。进样量:10 µL。流速:1.0 mL/min。该方法简化了样品的前处理,共分离出烟草中11种类胡萝卜素及其异构体。类胡萝卜素物质的加标回收率为87.7−94.6%,;相对标准偏差为3.01−4.29%。同时研究了新鲜烟叶和烘烤后烟叶中类胡萝卜素的分布和含量,结果显示:烟叶中类胡萝卜素的组成及含量与烟叶品种、部位以及调制有关。      

Influence of different drying methods on carotenoids and capsaicinoids of paprika (Cv., Jalapeno)

Influence of Refractance Window™ Drying (RWD), a novel contact drying method, on carotenoids, capsaicinoids, Retinol Activity Equivalent (RAE) and Scoville Heat Unit (SHU) of paprika (Cv., Jalapeno) was investigated in comparison with freeze drying (FD), oven drying (OD), and natural convective drying (NCD) methods. Eight carotenoids (capsanthin, capsorubin, capsolutein, β-carotene, β-cryptoxanthin, mutatoxanthin, violaxanthin and zeaxanthin) and five capsaicinoid analogues (capsaicin, dihydrocapsaicin, homodihydrocapsaicin, isodihydrocapsaicin, nordihydrocapsaicin) were identified in paprika. All these components were significantly (P < 0.05) decreased by the RWD, FD and OD methods. However, due to ongoing synthesis, the NCD method resulted in higher carotenoids, except violaxanthin and mutatoxanthin, and capsaicinoids content than those of the others, even puree. Mutatoxanthin, naturally occurring pigment in red pepper, could only be detected in FD paprika. The highest RAE and SHU values, which were derived from the data of carotenoids and capsaicinoids, respectively, were also determined in NCD paprika.     

Characterization and on-line monitoring of cell disruption and lysis using dielectric measurement

The dielectric measurement of Escherichia coli suspension was carried out between 0.01 and 100 MHz during cell disruption using a physical method or cell lysis induced by chemicals for the purpose of on-line monitoring of cell disruption or lysis of bacterial cells. The dielectric dispersion of relative permittivity centered at about 1.4 MHz and the dielectric parameters based on the fitting calculation well reflected the damage of these cells under physical and chemical stresses. The degree of cell disruption as determined by the dielectric parameters, the amplitude of dispersion and the conductivity, well corresponded to those obtained by other conventional kinetic analyses. This methodology can be utilized for the on-line monitoring of physical cell disruption and cell lysis induced by detergent and could provide direct information to control these processes. Based on these results, the dielectric measurement was successfully applied to monitor the stress-mediated cultivation of E. coli cells.     

Sporidiobolus pararoseus中红酵母红素的柱色谱分离纯化鉴定及其对LNCaP细胞增殖的影响

建立了一种通过柱色谱从Sporidiobolus pararoseus发酵产物中分离纯化红酵母红素的方法。通过高效液相色谙(high performance liquid chromatography,HPLC)鉴定色素的纯度在96%以上。同时通过HPLC-二极管阵列检测器(diode array detector,DAD)、液相(liquid chromalography,LC)-电离子喷雾(electronic spray ion,ESI)-质谱分析(mass spectrometry,MS)、傅立叶变换红外光谱(Fourier-transform infrared spectroscopy,FTIR)和拉曼光谱检测得到了一系列红酵母红素的光谱数据。该研究还发现红酵母红素对于前列腺癌LNCa P细胞的生长具有抑制作用,且该作用与剂量有一定关系。WST-1检测发现,用30μmol/L的β-胡萝卜素、番茄红素和红酵母红素分别处理24 h,细胞的生长率分别降低了(20.5±3.1)%、(27.5±3.7)%和(60.2±1.8)%。。     

N~+注入选育类胡萝卜素高产菌株及其固态发酵条件优化

为获得类胡萝卜素高产菌株,利用N+注入方法对黏性红圆酵母菌RM-1进行诱变处理,结果表明,当注入能量为10keV时,最适诱变注入剂量为2.0×1014ion/cm2,筛选所得突变株RM-21产类胡萝卜素的能力明显提高,较出发菌株增加了60.85%。传代结果表明,传代10次突变株RM-21遗传性状稳定,可作为工业生产菌种。以玉米粉和稻米为原料,对突变株RM-21固态发酵产类胡萝卜素进行条件优化研究,结果表明,当m玉米粉:m稻米=3:1,含水量75%时,固态发酵最适条件为:葡萄糖4.0g/L、NH4NO32.5g/L、初始pH6.0、接种量10%。在此基础上,对固态发酵产类胡萝卜素进行变温培养研究,结果表明,在96h的发酵时段中,0～84h温度控制在33℃,85～96h温度控制在28℃,类胡萝卜素产量达到7.04μg/g(以干基计),比恒温发酵提高了20.96%。