Some Factors Affecting the Production of Carotenoids by <i>Rhodotorula glutinis</i>var. <i>glutinis</i>

A new yeast strain isolated from pin cushion flower (Scabiosa atropurpura) in our laboratory was selected from 200 yeast isolates as carotenoids producer and identified as Rhodotorula glutinis var. glutinis. The selected isolate was grown in synthetic medium to study the effect of carbon to nitrogen ratio, sources of nitrogen and carbon, mineral salts and incubation temperature on carotenoids production. The results indicated the following optimal conditions: carbon to nitrogen ratio of 5, ammonium sulphate as nitrogen source, sucrose as carbon source, presence of zinc sulphate in the medium and cultivation temperature of 25?C. The studied factors affected the dry biomass as well as the proportion of carotenoids and consequently the colour of pellets of the yeast. The yeast strain was grown under the optimal conditions to study the changes occurring in the medium and the pellets during carotenoids production for 6 days. Carotenoids production started after the first day of incubation and most of the carotenoids content in the yeast cells was produced during stationary phase. The highest cellular (861 μg?g–1) and volumetric (1.9 mg?L–1) carotenoids content were ob- tained after 5 days of growth.

黏红酵母Rhodotorula glutinis微生物油脂提取工艺研究

采用酸热耦合超声波法对黏红酵母Rhodotorula glutinis微生物油脂提取工艺进行研究。选取酸处理时间、沸水浴时间、超声处理时间、超声功率、提取液配比、提取时间、NaCl浓度等7个微生物油脂提取相关因素,首先通过Plackett-Burmen试验设计筛选出3个显著因子:提取液配比(P=0.010 5)、超声处理时间(P=0.010 5)、酸处理时间(P=0.036 4)。然后通过Box-Behnken试验设计,对3个显著因素进行优化组合,得到最佳工艺条件,即酸处理时间34 min,超声处理时间7.5 min,提取液配比1.9∶1。与原始工艺条件相比,1 g湿菌所得油脂重量从42 mg增加到57.33 mg,提高了36.5%。

低能离子注入油脂高产菌株黏红酵母(Rhodotorula glutinis)的选育

本文对油脂高产菌株选育过程中的筛选方法进行研究,采用苏丹黑B染色作为定性分析、酸热耦合超声波作为定量分析高产油脂菌株的筛选方法,通过低能离子注入黏红酵母(Rhodotorula glutinis)进行诱变,筛选油脂高产菌株。结果表明:获得了一株油脂高产菌株D30,产量比出发菌株提高33.05%,传代试验表明了D30的遗传稳定性良好,当发酵96 h,其油脂产量达3.10 g/L。另外,对D30发酵动力学进行了研究,当发酵时间为10 d时,其生物量为47.98 g/L(菌体湿重),油脂产量达到7.81 g/L。

粘红酵母(Rhodotorula glutinis CIBAS A1401)苯丙氨酸解氨酶cDNA核心序列的克隆与分析

L苯丙氨酸因其在工业、医药上的重要作用而日益受到人们的重视.苯丙氨酸解氨酶(PAL)生物转化肉桂酸生产L苯丙氨酸成了该领域研究与开发的热点.本文利用简并寡核苷酸PCR法结合cDNA末端快速扩增PCR法,从粘红酵母中克隆了苯丙氨酸解氨酶cDNA核心序列,为克隆其cDNA全长序列及体外表达其活性蛋白奠定了基础.

Isolation, Identification and Screening of Carotenoid-Producing Strains of <i>Rhodotorula glutinis</i>

An attempt was made to isolate yeast strains from different sources (flowers, trees exudates, fruits and cheese) which have a fast growth rate and high content of carotenoids. Seventy cultures of pigmented yeast were isolated. Forty six yeast isolates were selected and identified by simplified identification method which showed that all isolates belong to Rhodotorula glutinis. These isolates were screened in two steps. Seventeen promising isolates of Rhodotorula glutinis were selected after the primary screening. Then 6 yeast isolates were chosen and passed through the second screening. The most promising isolate (isolated from pin cushion flower, Scabiosa atropurpurea) was selected according to its total carotenoids content expressed as torulene (μg?g–1 and μg?L–1). This isolate was reclassified using a number of morphological, biochemical, and physiological characteristics which revealed that the yeast isolate agree well with those of Rhodotorula glutinis Var. glutinis. Dry biomass, amount and proportion of individual carotenoids and lipid content of the selected identified yeast were determined.

Production of β-Carotene by a Newly Isolated Rhodotorula Glutinis UCP1555 Strain and Cytotoxic Effect Evaluation

Carotenoids have attracted greater attention due to the beneficial role on human health. It is an essential nutrient and has some biological functions such as species-specific coloration, photoprotection, light absorbing, which is an important component because of its role as precursor of vitamin A. In this study was performed the production of fl-Carotene by Rhodotorula glutinis UCP/WFCC 1555 in presence and absence of blue and white using LED （light-emitting diodes） and evaluation of the cytotoxic effect. The production was investigated in low cost medium constituted by different concentrations of CG （crude glycerin） and CSL （corn steep liquor） from the CCD （Central Composite Design） and the identification and yield of the β-Carotene was investigated by chromatographic profile （HPLC）. Additionally, the fl-Carotene produced was tested to evaluate its cytotoxic effect in human tumor cells MCF-7 （breast cancer） and HL-60 （promyelocytic leukemia） and healthy cells of macrophages. The results showed that in the medium composed by 6% glycerin and 0.6% corn steep liquor, in the absence of light, occurred the maximum production of total carotenoids with values of 160μg·g-1 and these 100.60μg·g-1 correspond to the β-Carotene that showed ability in inhibit cell growth in several tumor cells such as MCF-7 cells （breast cancer） and HL-60 （promyelocytic leukemia）.

Lipid Production by <i>Rhodotorula glutinis</i>from Pulp and Paper Wastewater for Biodiesel Production

This study investigated the potential of oleaginous yeast Rhodotorula glutinis utilizing pulp and paper wastewater effluents as cultivation media for the sustainable production of microbial lipids as biodiesel feedstock. R. glutinis is oleaginous yeast, which has the ability to produce significant quantities of intercellular lipids in the form of triacylglycerols. Yeast lipids are a promising potential feedstock for biodiesel production due to similar fatty acid composition to plant oils. The effect of various carbon sources on biomass production, lipid accumulation, substrate utilization, and fatty acid composition using R. glutinis in the pulp and paper wastewater media was studied. The pulp and paper wastewater was supplemented with glucose, xylose, and glycerol as carbon sources under nitrogen-limited conditions. The maximum lipid productions of 1.3 - 2.9 g&bull;L–1, which corresponded to the intracellular lipid contents of 8% - 15% cell dry weight (CDW), were obtained under various carbon substrates. A kinetic study of the batch fermentation was performed in a 3 L aerobic batch fermenter to describe the cell growth, lipid accumulation, and substrate utilization process, and the kinetic parameter was estimated. The fatty acid profile of oleaginous yeast was rich in palmitic, oleic, and linoleic acids and comparable to vegetable oils. Thus, the results of this study indicated that pulp and paper wastewater could be used to produce lipids as biodiesel feedstock.

Influence of Agitation and Aeration on Single Cell Oil Production by Rhodotorula glutinis from Glycerol

In the present study, the yeast Rhodotorula glutinis has been assessed with the aim of producing microbial lipids from glycerol under different aeration conditions. For such a purpose, experiments were carried out in shake flasks, under different conditions of agitation (150 to 250 rpm) and aeration (2.5 to 5.0 of flask volume-to-medium volume ratio). Furthermore, their influence on fermentative parameters (lipid and cell concentration, biomass yield;lipid yield;and lipid volumetric productivity) has been investigated using a 22 full factorial design. The statistical analysis has revealed a strong influence of both variables on substrate consumption, lipid accumulation, cell growth and lipid productivity. As a whole, results suggest that higher aeration levels provide greater cell and lipid concentrations, and lipid volumetric productivity. The best results (4.5 g/L of lipids and QP = 0.95 g/L&sdot;day) were achieved at the highest aeration (5.0 flask volume-to-medium volume ratio) and agitation (250 rpm) levels. Their fatty acid profile showed that oleic acid was produced in greater quantity (53.5%), followed by linoleic acid (18.7%), palmitic acid (6.8%) and stearic acid (9.9%). The microbial oil presented viscosity of 39.3 cP at 50°C and free fatty acid content of 1.93% ± 0.08%. These are significant results and contribute to establishing operational conditions that maximize single-cell oil production from glycerol by Rhodotorula glutinis, i.e. an alternative source as renewable raw material for lipid-based biorefineries.

Utilization of Soluble Starch by Oleaginous Red Yeast Rhodotorula glutinis

Starch containing wastewaters from the food and feed industry have been identified as potential cheap carbon sources for the production of microbial lipids. Due to its high potential lipid content the oleaginous yeast Rhodotorula glutinis is often used for fermentations in this field. Moreover it is investigated in the context of microbial carotenoid production, which also requires a cheap source of carbon. Thus, the ability of R. glutinis (ATCC 15125TM) to degrade and utilize soluble starch for the production of lipids has been assessed in this study. While glucose and fructose were readily consumed from the medium, starch was only slightly reduced in one treatment. The yield of fatty acid methyl esters (FAME) was graduated corresponding to the initial sugar contents, with the highest FAME yield (1.5 g·L-1) at the highest initial sugar content. In the treatment that contained starch as single carbon source, no FAME production was realized. Accordingly, if starchy wastewaters should be used for microbial cultivation with R. glutinis, an enzymatic or chemical pretreatment for starch hydrolysis should be applied, to increase the availability of this carbon source.

Rhodotorula glutinis发酵糖蜜产油脂影响因素研究

微生物油脂生产是解决生物柴油原料问题的重要途径之一。本文以制糖厂废糖蜜为原料,通过间歇发酵试验,考查了装液量、pH、接种量、温度,以及氮素和碳素等因素对黏红酵母(Rhodotorula glutinis)增殖与油脂合成的影响。结果表明,在稀释糖蜜COD浓度11100mg/L、初始pH值6.5、30℃和160r/min条件下发酵132h,黏红酵母的总细胞干重和油脂产量分别达到2.42和0.8g/L,去除单位COD的细胞产量和油脂产量分别为0.35和0.12kg/kg;在黏红酵母发酵系统中,氮源的补充对细胞增殖具有显著刺激作用,但对油脂合成与积累无显著影响;在发酵过程中一次性补加碳源葡萄糖10g/L,可使黏红酵母的细胞和油脂产量分别提高到3.74和1.3g/L。

基于菌藻混合培养合成天然类胡萝卜素的工艺研究及转录组水平分析

为突破单菌培养在微生物类胡萝卜素生产中产率低的问题,采用5 L发酵罐等体积接种高活性粘红酵母和小球藻,研究菌藻混合培养工艺的可行性,进一步从转录水平分析混合体系特定时间的菌藻协同代谢分子机制。结果表明,混合培养可获得最大生物量、残糖利用率、类胡萝卜素产量分别为51.7 g/L、83.3%和29.8 mg/L,实验组类胡萝卜素产量比菌藻纯培养组分别提高了21.5%和77.2%,且混合培养可获得7种类胡萝卜素,提高了玉米黄质和虾青素的占比;发酵36 h时取样测定分析转录组数据发现,实验组与菌藻对照组对比分别获得85个和2858个差异表达基因,GO聚类和KEGG代谢通路分析表明,小球藻可通过强化光合作用和特定类胡萝卜素合成相关基因的上调,实现兼养和改变类胡萝卜素种类的占比。

4种微生态制剂对对虾育苗水体主要水质指标的影响

研究了由球形红假单胞菌（Rhodopseudomlonas sphaeroides）、噬菌蛭弧菌（Bdellovibrio bacteriovorus）及黏红酵母（Rhodotorula glutinis）制成的4种微生态制剂对中国对虾（Fenneropenaeus chinensis）育苗水质的影响。结果表明，试验组与对照组相比都能明显净化水质，试验组中以添加噬菌蛭弧菌和黏红酵母组的效果最差，添加球形红假单胞菌和噬菌蛭弧菌组的效果最好，此组水质的pH值稳定，NH4^＋N明显下降，而亚硝酸盐、化学需氧量及硫化物等指标也优于其他试验组。统计分析显示，试验组与对照组相比，细菌数降低了3个数量级，添加嗜菌蛭孤菌试验组在减少异养菌（包含有害细菌）方面效果较好。同时用鳗孤菌攻毒，在5d内，对照组的死亡率显著高于试验组（P〈0．05），试验组免疫保护率为29．4％～58．5％，以添加嗜菌蛭弧菌和球形红假单胞菌组最高。建议联合使用噬菌蛭弧和球形红假单胞菌，混合密度初步定为2×10^5个／mL。

粘红酵母与金属离子结合使用对柑橘采后青霉病的抑制效果

研究了粘红酵母(Rhodotorulaglutinis)不同处理方法对柑橘果实采后病害(指状青霉)的抑制效果。结果表明:粘红酵母细胞悬浮液的抑菌效果最好,当浓度为1×109CFU/mL时,几乎完全抑制病害的发生,而滤液则会使病害加重。当粘红酵母分别与0.1mol/L浓度的Ca2+、0.1mol/L的Mg2+结合使用时,可提高抑菌效果。如与Fe2+使用时,高浓度的Fe2+会加重病害的发生,低浓度对抑菌效果不明显,当仅以浓度为2.5μmol/L的添加量抑菌效果较好。

粘红酵母对铀的吸附研究

研究了粘红酵母对水溶液中铀的吸附行为,发现其吸附铀的最佳pH值为6～7,最大吸附量为149.4mgU.g-1,其吸附等温线和Langmuir吸附等温方程符合较好,相关系数R2达到0.99;比较吸附铀前后粘红酵母的红外光谱图发现,吸附过铀的菌体的红外光谱在904cm-1处出现了一个新的峰,此峰为UO2的伸缩振动峰,说明粘红酵母确实对铀发生了吸附作用。此外,氨基或羟基的伸缩振动峰由3309移至3287cm-1,细胞壁中碳水化合物或醇中C—O键伸缩振动发生位移,由1068移至1080cm-1,说明这些基团可能参与了吸附过程;蛋白质的特征吸收峰(1653,1540,1237cm-1)在吸附前后基本无明显变化,表明粘红酵母的主要成分及结构仍保持完整。吸附后的菌体利用0.1mol.L-1的NaHCO3处理后可解吸出其中96%的铀,可见该菌在铀矿废水处理方面具有广阔的应用前景。

拉曼镊子分析红酵母合成类胡萝卜素

利用拉曼镊子对红酵母合成类胡萝卜素进行分析,考查氮源和碳源对类胡萝卜素产量的影响。取发酵终点细胞,一部分用于紫外光谱法测定,另一部分用拉曼镊子检测。原始光谱经过背景扣除、基线校正等方法预处理,定性分析不同培养基培养细胞的平均光谱,类胡萝卜素的拉曼信号强度有明显不同;紫外检测结果和拉曼峰高数据有良好的相关性,拟合参数的相关系数分别达到0.907 8和0.912 1;定量分析1 508cm-1峰高表明适宜红酵母细胞生长和类胡萝卜素合成的氮源和碳源分别是酵母粉+胰蛋白胨、葡萄糖。以上结果说明,拉曼镊子能提供红酵母胞内类胡萝卜素的含量信息,是实时检测红酵母细胞类胡萝卜素合成和优化发酵培养基的有效工具。

耐高COD淀粉废水的高产油脂粘红酵母的驯化和筛选

为减轻高COD淀粉废水对产油菌株粘红酵母生长的抑制,提高油脂产量,本文利用淀粉废水对该菌株进行了耐高COD梯度驯化,并采用流式细胞仪对经尼罗红染色的粘红酵母细胞进行了高油脂含量菌株的筛选。结果表明:经多次驯化后,粘红酵母耐受淀粉废水COD高达75000 mg/L;流式细胞仪筛选得到一株油脂含量为25.7%(质量分数)的粘红酵母,比原始菌株油脂提高了140%。400 L发酵罐实验表明,在初始COD 75000 mg/L,葡萄糖质量浓度为36 g/L,pH4.8及30℃条件下,培养33 h后,粘红酵母生物量达25.3 g/L,菌体油脂含量为29.5%,COD降至5600 mg/L,降解率为92.5%。

粘红酵母在味精废水中发酵生产油脂

对粘红酵母菌株进行驯化得到一株优良菌株Rh8,利用其在味精废水中发酵以去除废水中的化学需氧量(COD)并生产油脂;考察废水pH以及添加葡萄糖母液、营养因子等对菌株Rh8在味精废水中生长、产油和COD去除效果的影响,发现将废水稀释4倍、调节pH至5.5时,菌株可以较好地生长;而添加废葡萄糖母液、酵母粉、KH2PO4、MgSO4、MnSO4均能够促进菌体的生长、产油和废水中的COD去除,在250 mL摇瓶中,生物量最高可达15.6 g/L,干菌体中油脂质量分数达到29.61%,COD去除率达到45.1%。

黏红酵母产油脂培养基的响应面优化

利用单因素试验和响应面设计相结合,对黏红酵母产油脂培养基进行了优化。单因素试验得到初步发酵培养基成分为葡萄糖、蛋白胨、KH2PO4。经响应面优化发现,当发酵培养基中葡萄糖含量为73.40g/L,蛋白胨含量为1.06 g/L,KH2PO4含量为3.56 g/L时,油脂产量的理论预测值可达到3.49 g/L,比优化前提高了13%。气相分析其油脂组成,多不饱和脂肪酸质量分数为26.97%。然后又对高产菌株的发酵特性进行研究,在10 d时,生物量和油脂产量达到最高,此时达到发酵终点,生物量为47.98 g/L(菌体湿重),油脂产量达到7.81 g/L。

粘红酵母对水果采后病害生物防治的研究进展

粘红酵母作为一种用于生物防治的拮抗菌,具有拮抗效果好,不产生毒素,可以和化学杀菌剂共同使用等优点。对粘红酵母(Rhodotorula glutinis)对水果采后病害生物防治的研究现状进行了综述,其中包括防治效果、防治机理、防治的优势、存在的问题以及未来的研究方向等。还提出了使用生物技术改造粘红酵母从而提高其生物防治效果,是现在应用粘红酵母对水果采后病害生物防治研究的最新领域。

噬菌蛭弧菌和粘红酵母对中国对虾生长及非特异免疫因子的影响

将噬菌蛭弧菌(Bd)、粘红酵母(Rh)及二者混合物分别以1%的比例添加到中国对虾基础饲料里,同时以基础饲料为对照。采用连续投喂的方法,饲喂1.5cm左右的稚虾25d,分别测定第5、10、15、20和25天的酚氧化酶、超氧化物歧化酶、溶菌活力等免疫因子活性。试验结束后,试验组中Rh组存活率达到63%,Bd组和Bd+Rh组分别为45%和40%,二者间无显著差异(P>0.05)。投喂Rh组的增长率、增重率至少高于其他组10.72%和105.05%,比对照组高出36.37%和320.22%。试验组酚氧化酶活力、超氧化物歧化酶活力、溶菌活力从第10天起均高于对照组两倍左右。试验结果表明,饲养实验结束7d后,用鳗弧菌攻毒,在8d内,对照组的死亡率显著高于试验组(P<0.05),试验组免疫保护率为50%～65%,以Rh组最为显著。试验结果表明,微生态制剂能有效促进对虾的生长及提高免疫因子的活性;从免疫水平和生长水平综合考虑,建议单独使用粘红酵母,添加量为1%左右为宜。