一株产色素细菌的鉴定及所产色素性质的研究
摘要
色素可分为天然色素和合成色素两类。近几年来,苏丹红、孔雀绿等常用合成色素事件引起了一轮又一轮的食品安全警示,因而人们对合成色素的安全性疑虑也逐渐加深。而天然色素,却不断体现出它自身对人类健康有益的许多独特的优点:大多数天然色素来自可食用的动植物组织和微生物,安全性高,无毒副作用;很多天然色素中含有人体必须的营养物质或其本身就是维生素或具有维生素性质的物质;有些天然色素具有药理作用,对某些疾病具有防治作用;天然色素色调自然,易被消费者接受,有的品种还具有特殊的芳香气味,添加到食品中会给人带来愉快的感觉等。所以,天然色素越来越受到人们的青睐。但天然色素却存在其自身的缺点:一方面是天然色素的稳定性差、价格昂贵;另一方面是目前利用的天然色素大部分是从植物组织中提取出来的,而植物的大面积种植占用大量的土地并且受气候的影响,严重制约了它的广泛利用。开发新的天然色素的来源,尤其是利用微生物来生产天然色素引起了广泛的关注。
本工作对一株由土壤中分离的产绿色素菌株进行了初步的研究。首先,对产色素菌株的培养条件进行了探索,发现选择马铃薯培养基,静置或低速摇床培养,碳源选择葡萄糖或蔗糖,氮源选择酵母粉(或膏),不添加生长因子或适当添加微量的硫酸镁,培养条件为温度为28℃、pH值为6左右、培养时间为72h,可获得最大色素产量。其次,将色素进行初步的提取分离,分别采用了有机溶剂萃取、活性炭吸附和层析等方法,发现色素为水溶性色素,不溶于大部分有机溶剂,对该菌株所产色素最适宜的提取方法是:取接种后培养2天的培养液5000rmp离心20分钟,得上清液和菌体。取上清液,加入等体积的丙酮,静置后5000rmp离心20分钟,可得黄色上清液和蓝色沉淀。对上清液真空减压浓缩,干燥,得粘稠状黄色色素粗提物。将蓝色絮状沉淀溶于水后用乙醇纯化,减压蒸馏,真空干燥,得蓝色色素粗提物。对提取出的两种色素对温度、pH值、光照、氧化剂和还原剂以及金属离子等因素的稳定性进行了考察,发现两种色素对以上各因素的稳定性较好。最后对菌株进行了初步的生理生化鉴定和16SrRNA鉴定。
The pigment includes natural pigment and synthetic pigment. In recent years, Sudan, malachite green, and other commonly used synthetic pigment incidents also caused a lot of food safety alerts, which the people of pigment security concerns have gradually deepened. But natural pigments has continuously demonstrated its own beneficial to human health of the many unique advantages: most natural pigment from edible plants、animals and microorganisms, safe and non-toxic; many natural pigments contain the necessary nutrients to human or itself is a vitamin or the material with vitamin natures; some natural pigments have pharmacological effects and can treat some diseases; natural pigment colors are natural, can be easily accepted, and some also have nice smell, added to the food will bring a happy feeling. Therefore, more and more people like to use the natural pigments. However, the natural pigment also has its own shortcomings: on the one hand, the natural pigment has poor stability and expensive; on the other hand, most of the natural pigment today used is been extracted from plant tissue, at the same time the planting is impacted by the climate and occupies lots of land, which Restrict its Extensive use. Developments of new sources of natural pigments, in particularly, the use of microbes to produce natural pigments have caused widespread concerns.
In this work, a bacteria which can produce green pigment isolated from soil is studied. First of all, conditions of the strain to produce the pigment were explored and found that potato medium, static culture, glucose or sucrose choose carbon, nitrogen choose yeast (or cream), culture conditions for the temperature of 28℃, pH about 6, the culture time about 72 h, the stain will produce the largest pigment. Secondly, The extraction and separation of the pigment were studied, we have used organic solvent extraction、activated carbon adsorption、paper chromatography and other methods, this pigment is water-soluble and can not dissolve in most organic solvents, we extract two kinds of pigments: the blue one and the yellow one. Both of the pigments extracted can not be easily damaged by the high temperature、pH、light、oxidants and reducing agents、metal ions factors. Finally, we do some physiological and biochemical experiments and 16SrRNA identification to this strain .
本工作对一株由土壤中分离的产绿色素菌株进行了初步的研究。首先,对产色素菌株的培养条件进行了探索,发现选择马铃薯培养基,静置或低速摇床培养,碳源选择葡萄糖或蔗糖,氮源选择酵母粉(或膏),不添加生长因子或适当添加微量的硫酸镁,培养条件为温度为28℃、pH值为6左右、培养时间为72h,可获得最大色素产量。其次,将色素进行初步的提取分离,分别采用了有机溶剂萃取、活性炭吸附和层析等方法,发现色素为水溶性色素,不溶于大部分有机溶剂,对该菌株所产色素最适宜的提取方法是:取接种后培养2天的培养液5000rmp离心20分钟,得上清液和菌体。取上清液,加入等体积的丙酮,静置后5000rmp离心20分钟,可得黄色上清液和蓝色沉淀。对上清液真空减压浓缩,干燥,得粘稠状黄色色素粗提物。将蓝色絮状沉淀溶于水后用乙醇纯化,减压蒸馏,真空干燥,得蓝色色素粗提物。对提取出的两种色素对温度、pH值、光照、氧化剂和还原剂以及金属离子等因素的稳定性进行了考察,发现两种色素对以上各因素的稳定性较好。最后对菌株进行了初步的生理生化鉴定和16SrRNA鉴定。
The pigment includes natural pigment and synthetic pigment. In recent years, Sudan, malachite green, and other commonly used synthetic pigment incidents also caused a lot of food safety alerts, which the people of pigment security concerns have gradually deepened. But natural pigments has continuously demonstrated its own beneficial to human health of the many unique advantages: most natural pigment from edible plants、animals and microorganisms, safe and non-toxic; many natural pigments contain the necessary nutrients to human or itself is a vitamin or the material with vitamin natures; some natural pigments have pharmacological effects and can treat some diseases; natural pigment colors are natural, can be easily accepted, and some also have nice smell, added to the food will bring a happy feeling. Therefore, more and more people like to use the natural pigments. However, the natural pigment also has its own shortcomings: on the one hand, the natural pigment has poor stability and expensive; on the other hand, most of the natural pigment today used is been extracted from plant tissue, at the same time the planting is impacted by the climate and occupies lots of land, which Restrict its Extensive use. Developments of new sources of natural pigments, in particularly, the use of microbes to produce natural pigments have caused widespread concerns.
In this work, a bacteria which can produce green pigment isolated from soil is studied. First of all, conditions of the strain to produce the pigment were explored and found that potato medium, static culture, glucose or sucrose choose carbon, nitrogen choose yeast (or cream), culture conditions for the temperature of 28℃, pH about 6, the culture time about 72 h, the stain will produce the largest pigment. Secondly, The extraction and separation of the pigment were studied, we have used organic solvent extraction、activated carbon adsorption、paper chromatography and other methods, this pigment is water-soluble and can not dissolve in most organic solvents, we extract two kinds of pigments: the blue one and the yellow one. Both of the pigments extracted can not be easily damaged by the high temperature、pH、light、oxidants and reducing agents、metal ions factors. Finally, we do some physiological and biochemical experiments and 16SrRNA identification to this strain .
引文
[1] 阎炳宗. 我国天然色素的现状及发展方针[J]. 中国食品添加剂, 1999, (2): 49-51
[2] 聂晶, 齐兴娟. 食用合成色素研究动态[J]. 中国食品卫生杂志, 2002, 14(1) :58-60
[3] 郭承育. 天然食用色素的开发应用[J]. 青海大学学报(自然科学版), 2001, 19(3): 45-46
[4] 乔华. 天然食用色素的稳定性的研究及应用[D]. 山西: 山西大学, 2006
[5] 冯定远. 天然植物色素及其利用[J]. 广东饲料, 1996, (6): 6-9
[6] 杨革李亚.类胡萝卜素发酵生产条件的研究[J]. 食品科学, 1998,(8):20.
[7] 赵军. 黄酮类化合物的抗氧作用机制[J]. 华北煤炭医学院学报,2003,5(3): 306-307
[8] 龚盛昭. 黄酮类化合物保健食品大有开发价值[J]. 广州食品工业科技,2002, 18(1): 63-64
[9] 裴凌鹏, 惠伯棣, 金宗濂等. 黄酮类化合物的生理活性及其制备技术研究进展[J]. 食品科学, 2004, (2): 203-207.
[10] 王立, 姚惠源, 陶冠军等. 乌饭树树叶中黄酮类色素的抗氧化活性[J]. 食品与生物技术学报,2006,25(4):81-88.
[11] 刘莉华, 宛晓春. 黄酮类化合物抗氧化活性构关系的研究进展[J]. 安徽农业大学学报,2002,29(3): 265-270
[12] 尤新. 天然食用色素和功能[J]. 中国食品添加剂,2002, (5): 1-3
[13] 方忠祥, 倪元颖. 花青素生理功能研究进展[J]. 广州食品工业科技,2001,17(3): 60-62
[14] 黄梅丽, 江小梅. 食品工艺学[M]. 北京: 中国人民大学出版社,1986
[15] Merken H M,Beehcer G R.Measuerment of Food Flvonoids by High-Performance Liquid ChromatograPhy A Review[J]. J Agric Food Chem, 2000, 48(3): 577-599
[16] Pourat H. Drogues a antho cyaneset maladies vassculaires[J]. Plant Med Phytother, 1997, 11(9): 143-147
[17] Betti.V. Study of Machanism whereby anthocyanosides potentiate the effect of catecholamines on coronary vessels[J]. Fitoterapia, 1985, 54(2): 67-72
[18] Vincieri. Analysis HPLC of anthocyanines present in fluid extracts from Malyasylvestris L.flowers and leaves[J]. Bull Liaison-Groupe Polyphynols, 1992, 16(1): 339-342
[19] 黄自然. 蚕粪制取叶绿素盐衍生物及其在医药上的应用[J]. 华南农学院学报,1980,l(2):145-149
[20] 潘云良. 发展以蚕沙为原料的叶绿素系列产品[J]. 杭州化工, 1993, (2): 24-28
[21] 王贤纯. 姜黄色素稳定性的研究[J]. 食品与发酵工业, 1994, 20(l): 63 -66
[22] 阎江丽, 王建林. 紫草中天然色素物化特性的研究[J]. 精细化工, 1994, (16): 15-19
[23] 王君, 张宝善. 微生物生产天然色素的进展[J]. 微生物学通报, 2007 , 34(3): 580-583
[24] 田黎, 刘晨临. 海洋黄杆菌 S-9801 菌株色素的提取及色素理化性质的研究[J], 海洋学报, 2003, 25(2): 73-76
[25] 仓一华, 陆玲, 王伟. 一株产类胡萝卜素酵母 Rhodotorula.sp 色素的组分及摇瓶发酵研究[J], 食品科学, 2002, 23(5): 34-38
[26] 彭方等. 高产黑色素微生物资源的研究[J], 氨基酸和生物资源, 1996,18(4): 1-4
[27] 倪丽娜. 一株高产黑色素细菌的分离及鉴定[J], 微生物学通报, 2004, 31(1): 55-59
[28] 赵东红, 陆玲, 秦怀兰. 一种微生物发酵产蓝色素的稳定性及毒性研究[J],食品与发酵工业, 1998, 24(5): 21-24
[29] 李一苇, 陈晓琳等. 一株产蓝色素菌株生物学特性及色素基本性质的研究[J] , 生物学杂志, 2007, 24(1): 41-43
[30] 袁保红, 杜青平, 蔡创华. 海洋细菌 Pseudomonas sp.色素的提取及稳定性的研究[J], 海洋通报, 2005, 24(6): 92-96
[31] 张亮, 朱湘民. 一株产虾青素的黄杆菌 CF—60 的研究[J], 微生物学通报, 1999, 26(5): 332-335
[32] Del Campo J A, Rodríguez H, Moreno J, et al. Accumulation of astaxanthin and lutein in Chlorella zofingiensis (Chlorophyta)[J], Applied Microbiology and Biotechnology, 2004, 64(6): 848-854
[33] Margalith P Z. Production of ketocarotenoids by microalgae[J], Applied Microbiology and Biotechnology, 1999, 51(4): 431-438.
[34] Buzzini P. An optimization study of carotenoid production by Rhodotorula glutinis DBVPG 3853 from substrates containing concentrated rectified grape must as the sole carbohydrate source [J], J Ind Microbiol Biotechnol,2000, 24(1): 41-45
[35] 陈建军, 杨济龙. 产番茄红素红酵母培养条件研究[J], 云南农业大学学报, 2005, 20(3): 381-383.
[36] 曾强松, 童海宝, 徐静安. 三孢布拉霉发酵生产 β-胡萝卜素工艺研究[J], 工业微生物, 2002, 32(4): 7-10
[37] 张雪霞, 刘刚叁, 李宁. 中国新药杂志, 2005, 14(7): 880-881
[38] 胡爱红, 方尚玲, 马丽. 一株丝状真菌发酵法产红色素培养基的研究[J],湖北工学院学报, 2003, 18(4): 7-9
[39] Unagul P, Wongsa P, Kittakoop P, et al. J Ind Microbiol Biotechnol, 2005, 32: 135-140
[40] Veiga-Crespo P, Blasco L, Santos F R, et al. International Microbiology, 2005, 8: 55-58
[41] Bhosale P. Environmental and cultural stimulants in the production of carotenoids from microorganisms[J], Applied Microbiology and Biotechnology, 2004, 63(4): 351-361
[42] Salguero A, León R , Mariotti A, et al. Appl Microbiol Biotechnol, 2005, 66: 506-511
[43] Fbregas J, Domnguez A, Maseda A, et al. Interactions between irradiance and nutrient availability during astaxanthin accumulation and degradation in Haematococcus pluvialis[J], Applied Microbiology and Biotechnology , 2003, 61(5-6): 545-551
[44] Simova E D, Frengova G I, Besh D M. Zeitschrift für Naturforsch C. A Journal of Biosciences, 2003, 58:225-229
[45] Squina F M, Mercadante A Z. Journal of Food Biochemistry, 2005, 29(6): 638-652
[46] Kim B K, Park P K, Chae H J, et al. Korean J Chem Eng, 2004, 21(3): 689-692
[47] Orosa M,Franqueira D, Cid A, et al. Bioresource Technology, 2005, 96:373-378
[48] Bhosale P, Gadre R V. β-Carotene production in sugarcane molasses by a Rhodotorula glutinis mutant [J], Journal of Industrial Microbiology and Biotechnology, 2001, 26(6): 327-332
[49] Silva C, Cabral J M S, Isolation of a β-Carotene Over-Producing Soil Bacterium, Sphingomonas sp. [J], Keulen F V. Biotechnology Letters, 2004, 26(3): 257-262
[50] Kang C D, Lee J S, ParkTH, et al. Comparison of heterotrophic and photoautotrophic induction on astaxanthin production by Haematococcus pluvialis [J], Applied Microbiology and Biotechnology, 2005, 68(2): 237-241
[51] 王克明, 庄树宏. PVA 固定化红曲霉发酵生产红曲色素的研究[J], 烟台大学学报(自然科学与工程版), 1998, 11(4): 306-309
[52] 张子儒, 郑巧东, 姚善泾. 红曲霉菌微胶囊化培养[J], 食品发酵与工业, 2003, 29(11): 1-4
[53] Shin C S , Kim H J, Kim M J, et al. Biotechnology and Bioengineering, 1998, 59:576-581
[54] Suh J H, Shin C S. FEMS Microbiology Letters, 2000, 190: 241-245
[55] Frengova G I, Emilina S D, Beshkova D M. Synthesis of carotenoids by Rhodotorula rubra GED8 co-cultured with yogurt starter cultures in whey ultrafiltrate[J], Zeitschrift für Naturforschung. C. A Journal of Biosciences,2003, 58: 562-567
[56] 刘海丽,王蓓, 余晓斌. 利用洛伐他汀选育高产 β-胡萝卜素的三孢布拉霉菌株[J], 食品与机械, 2005, 21(5): 10-11
[57] 郝宏蕾, 重组菌株合成类胡萝卜素的研究[D], 浙江: 浙江大学, 2001
[58] 阮丽芳, 王玉沽, 沈萍. 产黑色素 B.thuringiensis 重组菌株的构建及其培养条件的优化武汉大学学报(理学版)[J], 2003, 49(2): 257-260
[59] 张兆俊,肖丽娟.天然色素高粱红开发应用[J],天然色素高粱红开发应用[J]粮食与油脂,2005,(5):40-41
[60] 刘树兴,胡小军.姜黄色素的研究进展[J].陕西科技大学学报,2003.21(4):37-39
[61] 丁纯梅,陶庭先,吴之传.龙虾壳的综合利用(1)虾壳红色素的提取及其性质研究[J].化学世界,1999,(8):444-445,434
[62] Hawthorne SB , Miller DJ . Direct comparisen of soxhlet and low—and high—temperature supercritical CO2 extraction efficiencies of organics from Environmental solids [J]. Anal Chem., 1994, 66 (22) :4005-4012
[63] Rozzi N L.Supercritical fluid extraction of lycopene fromtomato processing byproducts [J]. Journal of Agricultural and Food Chemistry 2002 (9):2638-2643
[64] 黎或,刘敏锐,杜友珍等.微波场协同提取野菊花黄色素的研究[J].广东微量元素科学,2004, 11(9):48-52
[65] 王伟华,于国萍,张立冬.番茄红素微波萃取工艺条件的研究[J].东北农 业大学学报,2004,35(5):564-567
[66] 蔡金星,刘秀凤,李兆蒙,等.以微波-超声波法提取草莓色素及其理化性质的研究[J].生产与科研经验,2003,29(5):69-73
[67] 刘毅,宁正祥.红曲色素及其在肉制品中的应用[J].食品与机械,1999,(1):28
[68] 周煜,16SrRNA 序列分析法在医学微生物鉴定中的应用[J].生物技术通讯,1999,10(4):297-305
[69] 伯杰氏细菌鉴定手册(第八版).1984, 274-312
[2] 聂晶, 齐兴娟. 食用合成色素研究动态[J]. 中国食品卫生杂志, 2002, 14(1) :58-60
[3] 郭承育. 天然食用色素的开发应用[J]. 青海大学学报(自然科学版), 2001, 19(3): 45-46
[4] 乔华. 天然食用色素的稳定性的研究及应用[D]. 山西: 山西大学, 2006
[5] 冯定远. 天然植物色素及其利用[J]. 广东饲料, 1996, (6): 6-9
[6] 杨革李亚.类胡萝卜素发酵生产条件的研究[J]. 食品科学, 1998,(8):20.
[7] 赵军. 黄酮类化合物的抗氧作用机制[J]. 华北煤炭医学院学报,2003,5(3): 306-307
[8] 龚盛昭. 黄酮类化合物保健食品大有开发价值[J]. 广州食品工业科技,2002, 18(1): 63-64
[9] 裴凌鹏, 惠伯棣, 金宗濂等. 黄酮类化合物的生理活性及其制备技术研究进展[J]. 食品科学, 2004, (2): 203-207.
[10] 王立, 姚惠源, 陶冠军等. 乌饭树树叶中黄酮类色素的抗氧化活性[J]. 食品与生物技术学报,2006,25(4):81-88.
[11] 刘莉华, 宛晓春. 黄酮类化合物抗氧化活性构关系的研究进展[J]. 安徽农业大学学报,2002,29(3): 265-270
[12] 尤新. 天然食用色素和功能[J]. 中国食品添加剂,2002, (5): 1-3
[13] 方忠祥, 倪元颖. 花青素生理功能研究进展[J]. 广州食品工业科技,2001,17(3): 60-62
[14] 黄梅丽, 江小梅. 食品工艺学[M]. 北京: 中国人民大学出版社,1986
[15] Merken H M,Beehcer G R.Measuerment of Food Flvonoids by High-Performance Liquid ChromatograPhy A Review[J]. J Agric Food Chem, 2000, 48(3): 577-599
[16] Pourat H. Drogues a antho cyaneset maladies vassculaires[J]. Plant Med Phytother, 1997, 11(9): 143-147
[17] Betti.V. Study of Machanism whereby anthocyanosides potentiate the effect of catecholamines on coronary vessels[J]. Fitoterapia, 1985, 54(2): 67-72
[18] Vincieri. Analysis HPLC of anthocyanines present in fluid extracts from Malyasylvestris L.flowers and leaves[J]. Bull Liaison-Groupe Polyphynols, 1992, 16(1): 339-342
[19] 黄自然. 蚕粪制取叶绿素盐衍生物及其在医药上的应用[J]. 华南农学院学报,1980,l(2):145-149
[20] 潘云良. 发展以蚕沙为原料的叶绿素系列产品[J]. 杭州化工, 1993, (2): 24-28
[21] 王贤纯. 姜黄色素稳定性的研究[J]. 食品与发酵工业, 1994, 20(l): 63 -66
[22] 阎江丽, 王建林. 紫草中天然色素物化特性的研究[J]. 精细化工, 1994, (16): 15-19
[23] 王君, 张宝善. 微生物生产天然色素的进展[J]. 微生物学通报, 2007 , 34(3): 580-583
[24] 田黎, 刘晨临. 海洋黄杆菌 S-9801 菌株色素的提取及色素理化性质的研究[J], 海洋学报, 2003, 25(2): 73-76
[25] 仓一华, 陆玲, 王伟. 一株产类胡萝卜素酵母 Rhodotorula.sp 色素的组分及摇瓶发酵研究[J], 食品科学, 2002, 23(5): 34-38
[26] 彭方等. 高产黑色素微生物资源的研究[J], 氨基酸和生物资源, 1996,18(4): 1-4
[27] 倪丽娜. 一株高产黑色素细菌的分离及鉴定[J], 微生物学通报, 2004, 31(1): 55-59
[28] 赵东红, 陆玲, 秦怀兰. 一种微生物发酵产蓝色素的稳定性及毒性研究[J],食品与发酵工业, 1998, 24(5): 21-24
[29] 李一苇, 陈晓琳等. 一株产蓝色素菌株生物学特性及色素基本性质的研究[J] , 生物学杂志, 2007, 24(1): 41-43
[30] 袁保红, 杜青平, 蔡创华. 海洋细菌 Pseudomonas sp.色素的提取及稳定性的研究[J], 海洋通报, 2005, 24(6): 92-96
[31] 张亮, 朱湘民. 一株产虾青素的黄杆菌 CF—60 的研究[J], 微生物学通报, 1999, 26(5): 332-335
[32] Del Campo J A, Rodríguez H, Moreno J, et al. Accumulation of astaxanthin and lutein in Chlorella zofingiensis (Chlorophyta)[J], Applied Microbiology and Biotechnology, 2004, 64(6): 848-854
[33] Margalith P Z. Production of ketocarotenoids by microalgae[J], Applied Microbiology and Biotechnology, 1999, 51(4): 431-438.
[34] Buzzini P. An optimization study of carotenoid production by Rhodotorula glutinis DBVPG 3853 from substrates containing concentrated rectified grape must as the sole carbohydrate source [J], J Ind Microbiol Biotechnol,2000, 24(1): 41-45
[35] 陈建军, 杨济龙. 产番茄红素红酵母培养条件研究[J], 云南农业大学学报, 2005, 20(3): 381-383.
[36] 曾强松, 童海宝, 徐静安. 三孢布拉霉发酵生产 β-胡萝卜素工艺研究[J], 工业微生物, 2002, 32(4): 7-10
[37] 张雪霞, 刘刚叁, 李宁. 中国新药杂志, 2005, 14(7): 880-881
[38] 胡爱红, 方尚玲, 马丽. 一株丝状真菌发酵法产红色素培养基的研究[J],湖北工学院学报, 2003, 18(4): 7-9
[39] Unagul P, Wongsa P, Kittakoop P, et al. J Ind Microbiol Biotechnol, 2005, 32: 135-140
[40] Veiga-Crespo P, Blasco L, Santos F R, et al. International Microbiology, 2005, 8: 55-58
[41] Bhosale P. Environmental and cultural stimulants in the production of carotenoids from microorganisms[J], Applied Microbiology and Biotechnology, 2004, 63(4): 351-361
[42] Salguero A, León R , Mariotti A, et al. Appl Microbiol Biotechnol, 2005, 66: 506-511
[43] Fbregas J, Domnguez A, Maseda A, et al. Interactions between irradiance and nutrient availability during astaxanthin accumulation and degradation in Haematococcus pluvialis[J], Applied Microbiology and Biotechnology , 2003, 61(5-6): 545-551
[44] Simova E D, Frengova G I, Besh D M. Zeitschrift für Naturforsch C. A Journal of Biosciences, 2003, 58:225-229
[45] Squina F M, Mercadante A Z. Journal of Food Biochemistry, 2005, 29(6): 638-652
[46] Kim B K, Park P K, Chae H J, et al. Korean J Chem Eng, 2004, 21(3): 689-692
[47] Orosa M,Franqueira D, Cid A, et al. Bioresource Technology, 2005, 96:373-378
[48] Bhosale P, Gadre R V. β-Carotene production in sugarcane molasses by a Rhodotorula glutinis mutant [J], Journal of Industrial Microbiology and Biotechnology, 2001, 26(6): 327-332
[49] Silva C, Cabral J M S, Isolation of a β-Carotene Over-Producing Soil Bacterium, Sphingomonas sp. [J], Keulen F V. Biotechnology Letters, 2004, 26(3): 257-262
[50] Kang C D, Lee J S, ParkTH, et al. Comparison of heterotrophic and photoautotrophic induction on astaxanthin production by Haematococcus pluvialis [J], Applied Microbiology and Biotechnology, 2005, 68(2): 237-241
[51] 王克明, 庄树宏. PVA 固定化红曲霉发酵生产红曲色素的研究[J], 烟台大学学报(自然科学与工程版), 1998, 11(4): 306-309
[52] 张子儒, 郑巧东, 姚善泾. 红曲霉菌微胶囊化培养[J], 食品发酵与工业, 2003, 29(11): 1-4
[53] Shin C S , Kim H J, Kim M J, et al. Biotechnology and Bioengineering, 1998, 59:576-581
[54] Suh J H, Shin C S. FEMS Microbiology Letters, 2000, 190: 241-245
[55] Frengova G I, Emilina S D, Beshkova D M. Synthesis of carotenoids by Rhodotorula rubra GED8 co-cultured with yogurt starter cultures in whey ultrafiltrate[J], Zeitschrift für Naturforschung. C. A Journal of Biosciences,2003, 58: 562-567
[56] 刘海丽,王蓓, 余晓斌. 利用洛伐他汀选育高产 β-胡萝卜素的三孢布拉霉菌株[J], 食品与机械, 2005, 21(5): 10-11
[57] 郝宏蕾, 重组菌株合成类胡萝卜素的研究[D], 浙江: 浙江大学, 2001
[58] 阮丽芳, 王玉沽, 沈萍. 产黑色素 B.thuringiensis 重组菌株的构建及其培养条件的优化武汉大学学报(理学版)[J], 2003, 49(2): 257-260
[59] 张兆俊,肖丽娟.天然色素高粱红开发应用[J],天然色素高粱红开发应用[J]粮食与油脂,2005,(5):40-41
[60] 刘树兴,胡小军.姜黄色素的研究进展[J].陕西科技大学学报,2003.21(4):37-39
[61] 丁纯梅,陶庭先,吴之传.龙虾壳的综合利用(1)虾壳红色素的提取及其性质研究[J].化学世界,1999,(8):444-445,434
[62] Hawthorne SB , Miller DJ . Direct comparisen of soxhlet and low—and high—temperature supercritical CO2 extraction efficiencies of organics from Environmental solids [J]. Anal Chem., 1994, 66 (22) :4005-4012
[63] Rozzi N L.Supercritical fluid extraction of lycopene fromtomato processing byproducts [J]. Journal of Agricultural and Food Chemistry 2002 (9):2638-2643
[64] 黎或,刘敏锐,杜友珍等.微波场协同提取野菊花黄色素的研究[J].广东微量元素科学,2004, 11(9):48-52
[65] 王伟华,于国萍,张立冬.番茄红素微波萃取工艺条件的研究[J].东北农 业大学学报,2004,35(5):564-567
[66] 蔡金星,刘秀凤,李兆蒙,等.以微波-超声波法提取草莓色素及其理化性质的研究[J].生产与科研经验,2003,29(5):69-73
[67] 刘毅,宁正祥.红曲色素及其在肉制品中的应用[J].食品与机械,1999,(1):28
[68] 周煜,16SrRNA 序列分析法在医学微生物鉴定中的应用[J].生物技术通讯,1999,10(4):297-305
[69] 伯杰氏细菌鉴定手册(第八版).1984, 274-312