产氢红杆菌类胡萝卜素含量和组分分析

类胡萝卜素在调节光合细菌产氢中具有重要作用。采用丙酮－甲醇有机溶剂法和KOH甲醇皂化法对产氢红杆菌（Rhodobacter sp.）R7菌株类胡萝卜素进行了提取纯化,并进一步采用硅胶G薄板层析法对提取的类胡萝卜素进行了分离,并结合光谱法对分离的类胡萝卜素进行了定性和定量分析。结果表明,丙酮－甲醇（7∶2,V/V）提取3次可将色素提取完全;最佳提取时间为2h;超声波处理与否对提取率影响不明显;该工艺提取类胡萝卜素产率为2.81mg/g湿菌体。硅胶G薄板层析表明该菌株类胡萝卜素有4个主要组分：黄色、红色、浅红色和浅黄色,黄色和红色为主要成分,光谱学数据显示黄色组分为球形烯,红色组分为螺菌黄质系类胡萝卜素,表明产氢红杆菌类胡萝卜素代谢途径独特。     

北萱草与大苞萱草区分为不同物种的核型证据

在文献中,小萱草Hemerocallis dumortierii Morr、北萱草H．esculenta Koidz．和大苞萱草H．mid- dendorfii Trautv et Mey被认为属于同一生物学复合体,并被处理为同一物种的不同变种。本文比较分 析了北萱草及大苞萱草的核型。北萱草和大苞萱草的核型分别为:2n=2x=22=12m+8sm+2T和2n =2x=22=10m+6sm+4st+2T。两个类群的核型差异水平已超出种内变异水。另外,大苞萱草以其 具有宽大苞片包裹的类头状花序,并且产黑龙江和吉林东部,南达辽宁干山地区,但绝不分布到大陆40° N以南地区而显著不同于分布华中、华北的北萱草。根据核型、外部形态及地理分布资料的综合分析,北萱草与大苞萱草应区分为不同物种,而不是同一物种的不同变种;同时,在系统发育上大苞萱草比北萱草进化。     

马蔺花器官表型特征及色素分析

为分析马蔺（Iris lactea var. chinensis）种质花器官表型性状，明确马蔺花瓣的色素成分，以中国6个省市不同生境条件下22份马蔺种质资源为试验材料，对花冠这一重要观赏部位的表型特征及花色素进行系统研究。采用RHSCC比色和色差仪测色方法描述了马蔺种质花器官表型性状，并通过亚硝酸钠-硝酸铝显色法和pH示差法等方法测定了马蔺花色素质量分数，分析不同色系马蔺花色素表达差异。结果表明：22份马蔺种质花瓣花色可分为浅蓝色、浅蓝紫色、深蓝紫色和紫罗兰色4大色系，垂瓣和旗瓣明度（L*）与a*呈负相关，与b*呈正相关，与彩度（c*）呈负相关。不同色系马蔺花器官表型特征不同，4大色系中紫罗兰色花瓣最大、花葶最高、垂瓣花斑最小，浅蓝色花瓣最小、花葶最低、垂瓣花斑最大，说明花瓣颜色越深，花瓣越大，垂瓣花斑则越小；不同色系马蔺花瓣中色素质量分数差异显著，浅蓝色花瓣中类胡萝卜素质量分数显著高于紫罗兰色，而紫罗兰色花瓣中的类黄酮质量分数和花色苷质量分数显著高于浅蓝色。随花瓣颜色加深，类胡萝卜素质量分数降低，类黄酮和花色苷质量分数相应增加。相关性分析表明，类胡萝卜素质量分数与垂旗瓣L*均呈显著正...     

对苞萱草的核型研究

本文报道用光学显微镜观察对苞萱草Ｈｅｍｅｒｏｃａｌｉｓｆｕｌｖａ（Ｌ．）Ｌ．ｖａｒ．ｏｐ ｐｏｓｉｔｉｂｒａｃｔｅａｔａＨ．ＫｏｎｇｅｔＣ．Ｊ．Ｗａｎｇ,ｖａｒ．ｎｏｖ．的染色体,按全国第一次植物染色体学术讨论会建议的标准进行核型分析。研究结果表明,对苞萱草体细胞染色体数目２ｎ＝２２,核型公式ｋ（２ｎ）＝２２＝１２ｍ＋６ｓｍ＋４ｓｔ,属于Ｓｔｅｂｂｉｎｓ核型的２Ｂ型。甘肃萱草属植物的核型可分为两大类群,对苞萱草的核型属于第二类群。对苞萱草与萱草Ｈ．ｆｕｌｖａ（Ｌ．）Ｌ．的核型较接近,为新变种的确立提供了细胞学方面的依据。     

产氢红杆菌类胡萝卜素含量和组分分析

类胡萝卜素在调节光合细菌产氢中具有重要作用。采用丙酮-甲醇有机溶剂法和KOH甲醇皂化法对产氢红杆菌(Rhodobacter sp.)R7菌株类胡萝卜素进行了提取纯化,并进一步采用硅胶G薄板层析法对提取的类胡萝卜素进行了分离,并结合光谱法对分离的类胡萝卜素进行了定性和定量分析。结果表明,丙酮-甲醇(7∶2,V/V)提取3次可将色素提取完全;最佳提取时间为2h;超声波处理与否对提取率影响不明显;该工艺提取类胡萝卜素产率为2.81mg/g湿菌体。硅胶G薄板层析表明该菌株类胡萝卜素有4个主要组分:黄色、红色、浅红色和浅黄色,黄色和红色为主要成分,光谱学数据显示黄色组分为球形烯,红色组分为螺菌黄质系类胡萝卜素,表明产氢红杆菌类胡萝卜素代谢途径独特。     

类芽孢杆菌(Paenibacillus sp.)蛋白质组学研究中三种蛋白提取方法的比较分析

【目的】革兰氏阳性类芽孢杆菌（Paenibacillus sp.）本身细胞壁的结构特点导致其菌体全蛋白不易获得。本研究选取了3种破碎方法——溶菌酶联合超声破碎法（方法一）、溶菌酶联合SDS热处理破碎法（方法二）、液氮联合超声破碎法（方法三）进行革兰氏阳性菌的细胞破碎,以期获得适于样品菌株基于质谱技术进行蛋白质组学研究的制备方法。【方法】在蛋白样品的制备过程中,对3种不同破碎方法的蛋白提取得率和SDS-PAGE检测分析结果进行比较;随后将3种蛋白样品制备方法的样品用质谱技术进行鉴定,分析不同蛋白样品基于质谱技术鉴定蛋白的差异。【结果】在蛋白样品的制备提取过程中,不同破碎方法的蛋白提取率大致相同。用单因素方差比较3种提取方法质谱鉴定蛋白数的差异性,方法三鉴定的蛋白数最多（2 638个）,其次是方法一（2 452个）,方法二鉴定的蛋白数最少（2 003个）。进一步用韦恩图分析比较不同提取方法的蛋白鉴定通量差异,综合考虑蛋白提取效率的结果以及液氮研磨法提取蛋白的缺点,最终选取溶菌酶联合超声破碎法（方法一）提取菌株全蛋白作为该菌基于质谱分析其蛋白质组学研究中最适合的方法。最后,对质谱鉴定菌株蛋白包括分子量、等电点、疏水性的基本性质进行分析,发现3种破碎方法质谱鉴定的蛋白与模式菌株多黏类芽孢杆菌（Paenibacillus polymyxa）基因组中预测蛋白的各个组分分布占比基本一致,都保证了菌株蛋白质组数据信息的完整性。【结论】基于质谱技术开展革兰氏阳性类芽孢杆菌（Paenibacillus sp.）的蛋白质组学研究,溶菌酶联合超声破碎法是提取该菌株全蛋白最适合的方法。     

甘肃萱草属─新变种

甘肃萱草属─新变种孔红王庆瑞（甘肃天水师范高等专科学校生物系,天水７４１０００）（西北师范大学植物研究所,兰州７３００７０）关键词萱草属;对苞萱草;甘肃ＡＮＥＷＶＡＲＩＥＴＹＯＦＨＥＭＥＲＯＣＡＬＬＩＳＦＲＯＭＧＡＮＳＵ￥ＫｏｎｇＨｏｎｇ（Ｄｅｐａｒ...     

中国萱草属数量分类研究

用聚类分析和主成分分析研究了国产萱草属11个类群的分类。结果发现这些类群形成4簇。第一簇：北黄花菜、黄花菜、小黄花菜和多花萱草。第二簇：小萱草和大苞萱草。第三簇：折叶萱草,西南萱草和矮萱草。第四簇：萱草及其三倍体类型。各簇都有其区别特征。讨论了簇内各类群之间的亲缘关系及属下分组问题。     

秦岭南麓萱草属植物花蕾中酚类物质比较分析

目的：为了对秦岭南麓野生萱草资源进行保护和开发利用。方法：从已收集的野生萱草属植物中选取具有代表性的种质材料，以黄花菜栽培种‘沙苑花’为对照，利用高效液相色谱法和紫外分光光度计对萱草属植物花蕾中酚酸类和黄酮类化合物进行检测和分析。结果：对7种酚酸类物质检测表明，野生材料的绿原酸、丁香酸、香草酸、对香豆酸、阿魏酸含量较栽培种低；但没食子酸和香豆酸的含量较栽培种高，其中野生‘萱草’的没食子酸和香豆酸含量分别是栽培种的1.57倍和1.59倍。在黄酮类物质中，野生材料的儿茶素含量低于栽培种；野生‘大苞萱草’的总黄酮和芦丁含量较栽培种低；但野生材料的槲皮素含量均高于栽培种，其中野生‘萱草’和‘大苞萱草’的平均含量分别是栽培种的1.99倍和2.56倍。结论：在秦岭南麓萱草属植物中，野生材料的没食子酸、香豆酸、总黄酮、槲皮素含量高于栽培种，且绿原酸、总黄酮和儿茶素含量明显高于其他地区的萱草材料，故应加强对该地区野生萱草属植物资源的调查收集和开发利用，这将对丰富萱草资源多样性和营养品质改良具有重要意义。     

丙酮和二甲基亚砜法测定植物叶绿素和类胡萝卜素的方法学比较

丙酮和二甲基亚砜作为提取剂测定植物光合色素含量是最为普遍的两种方法,但是对于二者之间的可比性问题探讨很少。本文基于对19中木本植物叶片和树枝内叶绿素含量的测定结果发现如下结果：（1）二甲基亚砜测定叶绿素a、叶绿素b和叶绿素总量的结果均显著高于丙酮测定结果（而对叶绿素a/b比值的影响较小）,与此相反,丙酮对类胡萝卜素提取测定结果要远高于二甲基亚砜的测定结果。（2）两种方法对叶绿素和类胡萝卜素测定结果之间均存在显著线性相关,可以应用相关线性拟合方程进行不同方法测定结果之间的转换,以减少叶绿素含量、类胡萝卜素含量比较过程中产生的测定方法误差。（3）不同单位表示方法,对两种方法测定结果的相关性存在影响,使用鲜重单位表示的相关程度普遍高于以表面积为单位的相关程度。（4）两种方法对树枝和叶片测定结果的影响存在差异：对于叶绿素含量低的树枝,两种方法测定结果的差异较小,而对于叶绿素含量高的叶片,二甲基亚砜测定结果则远高于丙酮的测定结果,可能原因在于丙酮对于叶绿素的提取效果远低于二甲基亚砜。（5）以二甲基亚砜为浸提试剂,通过不同精确度仪器（0.1和1 nm）检测发现尽管不同仪器之间相关关系达到0.99以上,但二者相比精确度为0.1 nm检测叶绿素a、总叶绿素浓度高出15%~33%,而叶绿素b浓度低4%左右。这些研究发现为不同方法之间的比较提供了依据。     

Karyotype Evidence for Distinguishing Between Hemerocallis esculenta and H. middendorfii

Hemerocallis dumortierii, H. esculenta and H. middendorfii were considered to belong to a biological complex, and were treated as different varieties of the same species, H. dumortierii. The present authors observed and compared the karyotypes of H. esculenta and H. middendorfii. Two plant samples for each of the species were studied. One sample of H. esculenta was from Wuán County, Hebei Province, at an altitute of 1000 m, the other from Shennongjia, Hubei Province, altitute 2100 m. The two samples of H. middendorfii were respectively from Antu County, at an altitute of 700 m, and Tonghua, altitute 500 m, in Jilin Province. The karyotypes are 2n＝2x=22=12m+8sm+2T for H. esculenta, while 2n＝2x＝22=10m+6sm+4st+2T for H. middendorfii. The karyotypical difference between the two taxa was obviously not intraspecific but interspecific. Based on the karyotype data as well as the morphological and phytogeographical data available, it is considered that H. esculenta and H. middendorfii should be recognized as different species, instead of different varieties of the same species as in literature, and that H. mid-dendorfii is more advanced than H. esculenta.     

Comparison of extraction methods for quantifying vitamin E from animal tissues

Four extraction methods: (1) solvent (SOL), (2) ultrasound assisted solvent (UA), (3) saponification and solvent (SP), and (4) saponification and ultrasound assisted solvent (SP-UA), were used in sample preparation for quantifying vitamin E (tocopherols) in chicken liver and plasma samples. The extraction yields of SOL, UA, SP, and SP-UA methods obtained by adding delta-tocopherol as internal reference were 95%, 104%, 65%, and 62% for liver and 98%, 103%, 97%, and 94% for plasma, respectively. The methods with saponification significantly affected the stabilities of tocopherols in liver samples. The measured values of alpha- and gamma-tocopherols using the solvent only extraction (SOL) method were much lower than that using any of the other extraction methods. This indicated that less of the tocopherols in those samples were in a form that could be extracted directly by solvent. The measured value of alpha-tocopherol in the liver sample using the ultrasound assisted solvent (UA) method was 1.5-2.5 times of that obtained from the saponification and solvent (SP) method. The differences in measured values of tocopherols in the plasma samples by using the two methods were not significant. However, the measured value of the saponification and ultrasound assisted solvent (SP-UA) method was lower than either the saponification and solvent (SP) or the ultrasound assisted solvent (UA) method. Also, the reproducibility of the ultrasound assisted solvent (UA) method was greater than any of the saponification methods. Compared with the traditional saponification method, the ultrasound assisted solvent method could effectively extract tocopherols from sample matrix without any chemical degradation reactions, especially for complex animal tissue such as liver.     

Ultrasound assisted extraction in quantifying lutein from chicken liver using high-performance liquid chromatography

Four sample preparation methods, (1) solvent (SOL), (2) saponification and solvent (SP), (3) ultrasound assisted solvent (UA), and (4) saponification and ultrasound assisted solvent (SP-UA), were used for quantifying lutein in chicken liver samples by HPLC. The lutein concentrations obtained by using SOL, UA, SP, and SP-UA were significantly different with values from 10.4 microg/g (UA) to undetected (SOL). Efficiency of the four different methods for extracting lutein from high to low were the UA, SP, SP-UA, and SOL method. The measured value of lutein in the liver sample using the UA method was approximately two and three times higher than that obtained from the SP and SP-UA method, respectively. The methods with saponification significantly affected the stabilities of lutein in liver samples. The lutein concentration measured with the solvent only method was either much lower than any of the other extraction methods or undetectable. This indicated that little lutein in those samples was in a form that could be extracted directly by solvent. Compared with the saponification method, the ultrasound assisted solvent method could effectively extract lutein from sample matrix and thus avoid chemical degradation reactions, which would be especially important for complex biological tissue such as liver.     

Carotenoid Formation by Staphylococcus aureus

The carotenoid pigments of Staphylococcus aureus U-71 were identified as phytoene; zeta-carotene; delta-carotene; phytofluenol; a phytofluenol-like carotenoid, rubixanthin; and three rubixanthin-like carotenoids after extraction, saponification, chromatographic separation, and determination of their absorption spectra. There was no evidence of carotenoid esters or glycoside ethers in the extract before saponification. During the aerobic growth cycle the total carotenoids increased from 45 to 1,000 nmoles per g (dry weight), with the greatest increases in the polar, hydroxylated carotenoids. During the anaerobic growth cycle, the total carotenoids increased from 20 nmoles per g (dry weight) to 80 nmoles per g (dry weight), and only traces of the polar carotenoids were formed. Light had no effect on carotenoid synthesis. About 0.14% of the mevalonate-2-(14)C added to the culture was incorporated into the carotenoids during each bacterial doubling. The total carotenoids did not lose radioactivity when grown in the absence of (14)C for 2.5 bacterial doublings. The total carotenoids did not lose radioactivity when grown in the absence of (14)C for 2.5 bacterial doublings. The incorporation and turnover of (14)C indicated the carotenes were sequentially desaturated and hydroxylated to form the polar carotenoids.     

Development and application of microwave-assisted extraction technique in biological sample preparation for small molecule analysis

Microwave-assisted extraction (MAE) has emerged as an efficient extraction technique for various kinds of biological samples due to its low usage of extraction solvents and shorter extraction time. This review will focus on the recent developments and advantages of incorporating MAE in sample preparation protocols for the analysis of small molecules in plant, food and clinical samples in recent years. The operating principles of this technique and the key parameters influencing its extraction efficiency, including the nature of solvent, temperature, power and extraction time and their limitations are first mentioned. This is followed by a discussion on the advantages of applying MAE to extract organic contaminants in food for routine food safety analysis and active ingredients recovery. The successful application of MAE technique to recover bioactive compounds from plants in drug discovery studies and quality control purposes is then described. Additionally, the feasibility of using green solvents such as water, micelle and ionic liquids with MAE for plant metabolite profiling studies is evaluated and the associated challenges discussed. Finally, the application of MAE in clinical samples is highlighted. The use of MAE in this field is currently limited to the targeted detection of small molecules in human samples, due to a lack of knowledge of its effects on thermally labile metabolites. Consequently, the need for additional studies on how MAE impacts the recoveries of different metabolite classes in mammalian samples is discussed. The outcome of these studies can potentially broaden MAE applications in the clinical field.     

A comparison of alternative sample preparation procedures for the analysis of swainsonine using LC‐MS/MS

Introduction – Swainsonine, a polyhydroxy indolizidine alkaloid and known glycosidase inhibitor, is found in a number of different plants that cause a lysosomal storage disease known as locoism in the western USA. Most recently swainsonine has been analysed by LC‐MS/MS after sample extraction and preparation from ion‐exchange resins. Objective – To compare previously published sample preparation procedures with several new alternative procedures to provide methods using either commercially available solid‐phase extraction equipment or procedures which significantly reduce sample preparation time. Methodology – A previously reported and validated sample preparation method using ion‐exchange resin was compared with methods using a commercially available solid‐phase extraction cartridge, a solvent partitioning procedure or a single solvent extraction procedure using one of two solvents. Twenty different plant samples of varying swainsonine concentrations were prepared in triplicate and analysed by LC‐MS/MS. The measured concentration of swainsonine was then statistically compared between methods. Results – There were no statistically significant differences found between four of the five different sample preparation methods tested. Conclusion – A commercially available SPE cartridge can be used to replace the previously used ion‐exchange resin for swainsonine analysis. For very rapid analyses the SPE procedure can be eliminated and a simple, single solvent extraction step used for sample preparation. Published in 2010 by John Wiley & Sons, Ltd.     

The biological interpretation of metabolomic data can be misled by the extraction method used

The field of metabolomics is getting more and more popular and a wide range of different sample preparation procedures are in use by different laboratories. Chemical extraction methods using one or more organic solvents as the extraction agent are the most commonly used approach to extract intracellular metabolites and generate metabolite profiles. Metabolite profiles are the scaffold supporting the biological interpretation in metabolomics. Therefore, we aimed to address the following fundamental question: can we obtain similar metabolomic results and, consequently, reach the same biological interpretation by using different protocols for extraction of intracellular metabolites? We have used four different methods for extraction of intracellular metabolites using four different microbial cell types (Gram negative bacterium, Gram positive bacterium, yeast, and a filamentous fungus). All the quenched samples were pooled together before extraction, and, therefore, they were identical. After extraction and GC?CMS analysis of metabolites, we did not only detect different numbers of compounds depending on the extraction method used and regardless of the cell type tested, but we also obtained distinct metabolite levels for the compounds commonly detected by all methods (P-value?<?0.001). These differences between methods resulted in contradictory biological interpretation regarding the activity of different metabolic pathways. Therefore, our results show that different solvent-based extraction methods can yield significantly different metabolite profiles, which impact substantially in the biological interpretation of metabolomics data. Thus, development of alternative extraction protocols and, most importantly, standardization of sample preparation methods for metabolomics should be seriously pursued by the scientific community.     

矮牵牛新种质的花色表型及花色素分析

以27个上海交通大学自育矮牵牛新种质为研究材料,对花色这一重要观赏性状及其花色素进行了系统研究。用RHSCC比色和色差仪测色方法描述了矮牵牛的花色表型,通过特征显色反应初步判断了矮牵牛的花色素类型,以标准曲线法和pH示差法等方法测定了矮牵牛3类花色素的含量。研究表明：这27个矮牵牛种质的花色可归于5个色系,以紫红色和红色为主;矮牵牛花色在CIELab表色系统中分布较广,而且不同色系花色参数的区分度较大。矮牵牛花瓣中含有类黄酮和花色苷,不含或含少量类胡萝卜素。13个被测种质的花瓣类黄酮含量在2.5～12.2 mg·/g–1 ·FW之间,花色苷含量在0.08～3.88 mg·g–1 FWmg/g·FW之间,而类胡萝卜素在矮牵牛花瓣中含量很低,远远低于类黄酮含量,在7个被测种质中,最高仅为0.216 mg·g–1 FWmg/g·FW,最低为0.004 mg·g–1 FWmg/g·FW。以上结果显示,5个色系矮牵牛所含花色素种类不尽相同,含量也有明显差异,其中紫红色系和红色系花瓣大多不含或含极少量类胡萝卜素,黄色系、白色系和紫色系花瓣的类黄酮含量较高,紫色系和紫红色系花瓣花色苷含量较高。     

A rapid procedure for the separation and analysis of metabolites of the sterol and dolichol pathways

We have previously shown that of several methods examined, the maximum yield of dolichyl phosphate (Dol-P) from rat liver was achieved by saponification of the tissue and subsequent extraction with diethyl ether (R.K. Keller et al. (1985) Anal. Biochem. 147, 166-173). In the present report, we have developed a rapid procedure using non-toxic solvents which resolves the ether extract on a C18 cartridge column into four major fractions: (1) fatty acids; (2) squalene and sterols; (3) dolichol; (4) Dol-P. The utility of the new procedure was demonstrated by preparing the four fractions from liver slices which had been incubated with [3H]acetate. HPLC analysis of the sterol, dolichol and Dol-P fractions yielded well resolved elution profiles, thereby allowing determination of radioactivity incorporated into the major isoprenoids and their metabolites.     

Evaluation of extraction methods for recovery of fatty acids from lipid-producing microheterotrophs

The effect of different extraction techniques on the recovery of fatty acids from freeze-dried biomass of two lipid-producing microheterotrophs was examined. Two procedures were used: the extraction of lipids from biomass followed by transesterification of the fatty acids (extraction-transesterification); and the direct transesterification of biomass to produce fatty acid methyl esters (i.e. without the initial extraction step). Variable factors in the extraction-transesterification experiment were the sequence in which solvents were added to the samples, the relative amount of methanol in the solvent mix, and sonication of biomass while in the solvent mix. Variable factors in the direct transesterification experiment were sample size, and reaction duration. Statistical analysis of data (level of significance P<0.05) showed that: (1) extraction of total fatty acids prior to transesterification was significantly more efficient when solvents were added in the order of increasing polarity; (2) neither sonication nor increasing the proportion of methanol in the extraction solvent significantly affected extraction of fatty acids prior to transesterification; (3) efficiency of direct transesterification of fatty acids increased significantly with reaction time; (4) efficiency of direct transesterification of fatty acids was not significantly affected by sample size; (5) the most efficient method for extraction of fatty acids prior to transesterification yielded significantly less fatty acids than the most effective direct transesterification method. While the study examined only two strains, our results suggest that fatty acid analysis methodology for microheterotrophs under consideration for biotechnological exploitation requires optimisation and validation.