测定浮游植物中色素的高效液相色谱与二极管阵列分光光度计联用的方法

建立了高效液相色谱与二极管阵列分光光度计联用分析浮游植物中色素的方法.采用90%的丙酮提取藻类中的类胡萝卜素和叶绿素后在反相C18柱上分离,流动相选用甲醇/醋酸铵/丙酮,流速为2.0 mL*min-1.用可见光检测器检测,波长为440 nm.在线的二极管阵列检测器对色素峰的扫描(400～700 nm)得到各色素峰的吸收光谱的结果显示,保留时间相同的色素峰,吸收光谱可能不同.根据色素峰吸收的光谱特性确定色素的种类,可以避免色素峰保留时间相同而造成的判断误差,从而提高了分辨的准确性.     

不同北虫草菌株产类胡萝卜素分析

通过对不同来源北虫草菌株子实体中提取的色素进行定性分析并对其含量进行测定,获得类胡萝卜素含量高的北虫草菌株。结果表明,浓硫酸反应中溶液在两相交界处呈现特征性的蓝绿色,色素提取液在400～600 nm范围内有波长分别为415、440和460 nm的3个吸收峰与类胡萝卜素标准的吸收光谱一致,证明子实体中提取的色素为类胡萝卜素;6个菌株子实体类胡萝卜素的含量差异较大,且J菌株的类胡萝卜素的含量最高,达到5.021 mg/g。     

几种绿藻、褐藻和红藻的吸收光谱的比较研究

1.用乳白玻璃法测定了我国黄海常见的属于绿藻门、褐藻门和红藻门的十种海藻的吸收光谱。讨论了这三门海藻的吸收光谱的特点。通过对吸收光谱及红藻、褐藻相对于绿藻的差异光谱的分析,讨论了各种色素在活体中的吸收峰和吸收带的位置。2.根据褐藻与绿藻的吸收光谱的差异及差异光谱的比较分析,认为绿藻吸收光谱中位于485(或480)毫微米的吸收峰是叶绿素 b 的蓝光吸收峰。叶绿素 b 的红光吸收峰位于655毫微米附近。岩藻黄素的吸收峰位于525毫微米附近。根据红藻吸收光谱及其同绿藻的差异光谱中的红光吸收峰和负峰的特点,初步认为红藻中的叶绿素 d 的红光吸收峰比叶绿素 a 的红光吸收峰位于较长波长处。3.对几种红藻的吸收光谱进行了比较。观察到原始红藻纲的条斑紫菜(Porphyra yezoen-sis)的藻胆素与真红藻纲的藻类有显著的差异。这种差异及条斑紫菜的藻胆素的含量和组成上的易变性值得进一步研究。     

VB1、VB12和VH对雨生红球藻生长及类胡萝卜素含量的影响

以淡水经济微藻雨生红球藻(Haematococcus pluvialis CH-1)为试验藻种,分别添加维生素B₁(VB₁)、B₁₂(VB₁₂)和维生素H(VH),每种维生素各分6个浓度梯度,测定了雨生红球藻的细胞密度、生物量、叶绿素a和类胡萝卜素含量等指标。结果表明,分别添加维生素B₁、B₁₂和H均显著促进了雨生红球藻细胞的生长。维生素B₁、B₁₂和H对雨生红球藻的最佳添加浓度分别为10mg·L^-1、50μg·L^-1和500μg·L^-1。在维生素B₁、B₁₂和H各自的最佳浓度处理下,雨生红球藻的细胞密度、生物量、叶绿素a和类胡萝卜素含量等4项指标均比对照有显著提高:维生素B₁处理分别提高了20.1%、14.6%、23.2%和21.3%;维生素B₁₂处理分别提高了29.5%、30.0%、28.0%和24.4%;维生素H处理分别提高了17.1%、29.2%、21.8%和10.1%。在雨生红球藻规模化生产的游动细胞培养阶段,适当地添加维生素B₁、B₁₂和H均能够有效地提高藻细胞密度、生物量、叶绿素a和类胡萝卜素含量。     

外源ABA和GA3对红肉脐橙果皮主要色素含量变化的影响

测定了红肉脐橙果实发育期间果皮叶绿素和总类胡萝卜素含量的变化动态,并在果实转色前用不同浓度的外源ABA和GA₃处理后测定果皮色素含量的变化动态。结果表明,红肉脐橙果皮叶绿素总量于9月20日出现最大值(0.1469mg/gfw),总类胡萝卜素含量于12月20日达到最大值(0.0321mg/gfw);转色前用ABA处理后加速果皮叶绿素的降解,但抑制果皮类胡萝卜素的积累;转色前用GA₃处理后延缓果皮叶绿素的降解,并同样抑制果皮类胡萝卜素的积累,严重阻碍了果皮类胡萝卜素的合成。     

粳稻生育后期剑叶光合日变化和光合色素对大气CO2浓度和温度升高的响应——FACE研究

为研究水稻叶片光合色素和光合日变化对大气CO₂浓度和气温升高的响应,我们采用在开放空气中控制升高CO₂浓度和温度的方法,以常规粳稻南粳9108为试验材料,设置了环境CO₂和高大气CO₂浓度(增加200 μmol·mol^-1)、环境温度和增温(增加1～2 ℃)交互的4个处理,测定了灌浆中期和后期水稻剑叶的光合日变化特征和光合色素含量.结果表明: 水稻剑叶净光合速率(P_n)为双峰曲线,发生了光合“午休”现象;大气CO₂浓度升高提高了剑叶P_n,灌浆中期和后期平均分别增加了47.6%和39.1%;高温有降低P_n的趋势,但相关性未达到显著水平.大气CO₂浓度和温度升高导致水稻剑叶生育后期气孔导度(g_s)平均分别降低了17.0%和11.8%.高CO₂浓度水稻剑叶生育后期蒸腾速率(T_r)、叶绿素a、叶绿素b、类胡萝卜素、总叶绿素和叶绿素a/b值显著降低,平均降幅分别为5.9%、50.4%、21.3%、41.4%、39.4%和21.4%,明显增加了剑叶水分利用率(WUE),平均增幅达47.9%.与之相反,生育后期增温使水稻剑叶T_r增加了10.2%,使WUE平均降低了20.4%.综上所述,大气CO₂浓度升高对粳稻生育后期剑叶P_n、g_s和光合色素含量的影响明显大于增温效应.因此,应重视大气CO₂浓度和温度对水稻光合作用和光合色素的综合效应,减弱增温的负效应.     

粳稻生育后期剑叶光合日变化和光合色素对大气CO2浓度和温度升高的响应——FACE研究

为研究水稻叶片光合色素和光合日变化对大气CO₂浓度和气温升高的响应,我们采用在开放空气中控制升高CO₂浓度和温度的方法,以常规粳稻南粳9108为试验材料,设置了环境CO₂和高大气CO₂浓度(增加200 μmol·mol^-1)、环境温度和增温(增加1～2 ℃)交互的4个处理,测定了灌浆中期和后期水稻剑叶的光合日变化特征和光合色素含量.结果表明: 水稻剑叶净光合速率(P_n)为双峰曲线,发生了光合“午休”现象;大气CO₂浓度升高提高了剑叶P_n,灌浆中期和后期平均分别增加了47.6%和39.1%;高温有降低P_n的趋势,但相关性未达到显著水平.大气CO₂浓度和温度升高导致水稻剑叶生育后期气孔导度(g_s)平均分别降低了17.0%和11.8%.高CO₂浓度水稻剑叶生育后期蒸腾速率(T_r)、叶绿素a、叶绿素b、类胡萝卜素、总叶绿素和叶绿素a/b值显著降低,平均降幅分别为5.9%、50.4%、21.3%、41.4%、39.4%和21.4%,明显增加了剑叶水分利用率(WUE),平均增幅达47.9%.与之相反,生育后期增温使水稻剑叶T_r增加了10.2%,使WUE平均降低了20.4%.综上所述,大气CO₂浓度升高对粳稻生育后期剑叶P_n、g_s和光合色素含量的影响明显大于增温效应.因此,应重视大气CO₂浓度和温度对水稻光合作用和光合色素的综合效应,减弱增温的负效应.     

氮、磷营养对雨生血球藻绿色细胞生长的影响

采用BBM培养基,以雨生血球藻(Haematococcus pluvialis CG-11)为研究材料,通过测定细胞生长速率、叶绿素和类胡萝卜素含量、生物量和虾青素含量,探讨氮、磷营养对雨生血球藻营养细胞生长的影响.结果显示:H.pluvialis CG-11生长的适宜氮源形式是NaNO₃和NH₄NO₃;适宜H.pluvialis CG-11生长的氮浓度为41.2mg·L^-1,磷浓度为5.3～53.3mg·L^-1.     

施氮肥缓解臭氧对小麦光合作用和产量的影响

以小麦(Triticum aestivum)品种‘扬麦16’为试材, 利用开放式空气臭氧(O₃)浓度升高平台, 研究了增施氮(N)肥对O₃对小麦光合作用和产量影响的缓解作用。结果表明, O₃胁迫下灌浆期小麦的净光合速率(P_n)、气孔导度(G_s)、蒸腾速率(T_r)、叶绿素a (Chl a)、叶绿素b (Chl b)、类胡萝卜素(Car)、总叶绿素含量(Chl t)和可溶性蛋白的含量显著降低, 降幅分别为28.95%、31.79%、23.17%、58.89%、68.64%、22.89%、60.31%和32.00%; 胞间CO₂浓度(C_i)变化很小; 成熟期生物量和收获时产量也明显下降, 降幅分别为12.23%和12.63%; 而增施N肥可以增加小麦灌浆期的P_n、Chl a、Chl b、可溶性蛋白的含量, 进而增加小麦生物量和产量, 增幅分别为25.66%、83.05%、121.57%、30.33%、14.94%和10.67%, 而对C_i、G_s、T_r、Car含量无明显影响。O₃和N肥对小麦叶片的P_n、Chl t及可溶性蛋白含量有明显的交互作用。因此, 在大气O₃浓度升高条件下增施N肥对小麦O₃损伤有一定的缓解作用。     

沼泽红假单胞菌中类胡萝卜素的提取与分析

目的:研究沼泽红假单胞菌中类胡萝卜素的提取工艺条件。方法:对细胞收集、前处理、色素初浸、二次萃取,皂化和溶剂回收等方面进行了系统的研究,并通过紫外分光光度法和高效液相色谱法对所提取的类胡萝卜素进行了分析。结果:实验结果表明,类胡萝卜素的最佳提取工艺条件为:碱性氯化钙絮凝法收集细胞,乙醇浸泡法进行前处理,49℃下丙酮初浸提浸提4h(搅拌并加抗氧化剂),丙酮用量为5mL/g菌泥,回收丙酮,乙醚二次萃取,30℃下皂化1.5h,洗涤,浓缩并回收乙醚,在此提取工艺条件下不仅能够有效地提取红螺菌中的类胡萝卜素,而且还能分离细菌叶绿素。结论在本实验条件下,根据标准曲线计算出类胡萝卜素的含量高达6.148mg/L菌泥。通过HPLC分析表明:类胡萝卜素提取液中主要成分至少有10种。     

EVALUATION OF PIGMENT EXTRACTION METHODS AND A RECOMMENDED PROTOCOL FOR PERIPHYTON CHLOROPHYLL a DETERMINATION AND CHEMOTAXONOMIC ASSESSMENT1

Many methods have been proposed to extract and quantify algal pigments. Comparative studies have found that pigment extraction efficiency varies among solvent and mechanical disruption protocols due to differential cellular resistance, thereby, leading to potential misinterpretation of pigment data. When the type or resistance of algae are unknown, a method is required that efficiently extract pigments from all taxonomic groups. The objective of this study was to develop a simple and efficient one stage periphyton pigment extraction protocol by comparing the extractability of four solvents (acetone, methanol, methanol/acetone, and methanol/acetone/N,N‐dimethylformamide), the effects of grinding, and the effects of freeze‐drying. The best overall extraction was obtained using freeze‐dried samples extracted with methanol/acetone/DMF/water (MAD). Eighty‐six percent more chlorophyll was extracted when the sample was freeze‐dried relative to fresh/frozen samples extracted with 90% acetone. Freeze‐drying greatly improved the extraction of both polar and non‐polar (lipophilic/hydrophobic) pigments while MAD increased the extractability of polar pigments and improved peak resolution of all pigments. Chemotaxonomic assessment differed between samples that were fresh/frozen or freeze‐dried before extraction. The relative abundance of cyanobacteria was greater for freeze‐dried material compared with fresh/frozen due to the improved extractability of cyanobacterial pigments. Based on the results of this study, the traditional approach of 90% acetone as a solvent is not recommended for periphyton samples containing cyanobacteria or when the composition of the mat is unknown. The combination of freeze‐drying and MAD was sufficient for the extraction of pigments from a periphyton mat containing filamentous cyanobacteria, green algae, and diatoms.     

Compaitive Study on Liposoluble Compoands in Autotrophic and Heterotrophic Chlorella protothecoides

Both autotrophically and heterotrophically grown Chlorella protothecoides cells have been obtained in cell cultures. The content of liposoluble compounds in the cells of heterotrophic algae occupied 72% of the total cells in dry weight, which was more than 4 times as high as that in the autotrophic algal cells. There existed remarkbly different distribution patterns of the hydrocarbons in thesetwo kinds of cells. The hydrocarbons in autotrophic cells were characterised by the predominance of C17 normal alkanes, wheraes the heterotrophic cells were rich in normal alkanes of higher molecular weight or longer carbon chain with C25 as the dominant carbon. The structure of the compounds in benzene fraction is not quite clear, but the compounds in autotrophis sample may be related to the degeneration of the pigments. The compounds in heterotrophic sample probably come from lipid acids. The visible--ultraviolet absorption spectrum of the pigment compounds demonstrated the absorption peaks of the acetone extract from the autotrophic cells at 432.5, 451.5, 472.5 and 661.5 nm, reflecting the existence of chlorophyll and carotenoid, both with a rather high concentration. However, the acetone extract from the hetertrophic algal cells only showed absorption peaks at 427.4, 450.8 and 477.5 nm. The absorption peaks of the original green cells completely disappeared at 432.5 and 661.5 nm, reflecting the disappearance of chlorophyll in cells on the whole; the remaining absorption peaks only reflected the existence of carotenoid, but its concentration had already been greatly reduced. The resuls from comparative experiments were of essential significance on the study of physiological metabolism in heterotrophically grown C. protothecoides and on the exploration and application of the lipid compounds in this kind of algae.     

An efficient method for the extraction of astaxanthin from the red yeast Xanthophyllomyces dendrorhous

This study investigated an efficient method for the extraction of astaxanthin from the red yeast Xanthophyllomyces dendrorhous. The extraction process comprised three steps: (1) cultivating the yeast; (2) treating the yeast culture suspension with microwaves to destroy the cell walls and microbodies; and (3) drying the yeast and extracting the astaxanthin pigment using ethanol, methanol, acetone, or a mixture of the three as the extraction solvent. Ultimately, various treatment tests were performed to determine the conditions for optimal pigment extraction, and the total carotenoid and astaxanthin contents were quantified. A frequency of 2,450 MHz, an output of 500 watts, and irradiation time of 60 s were the most optimum conditions for yeast cell wall destruction. Furthermore, optimal pigment extraction occurred when using a cell density of 10 g/l at 30 C over 24 h, with a 10% volume of ethanol.     

Extraction of chlorophylls a and b from phytoplankton using standard extraction techniques *

SUMMARY. Pigments extracted in methanol, acetone and ethanol from three cultures of green algae and one blue-green alga revealed different extraction efficiencies depending on the species, the extraction solvent used and the extraction time. Chromatographic identification and quantitative measurements of chlorophylls a and b were made from six green algae. When extraction of pigments was incomplete, chlorophyll-b was extracted faster than chlorophyll-a. This effect was more pronounced for acetone extractions, whereas methanol extractions gave the stable ratios of chlorophyll b/a after about 6–10 h. When green algae are frequent, a 6–10 h methanol extraction, without any extra manipulations, is sufficient to ensure reliable ratios of chlorophyll b/a and extraction of the major proportion of the chlorophylls without risk of induced destruction of the chlorophylls.     

光合细菌H3菌株色素分析

H3菌株系由盐田微生物层中分离获得的光合细菌株。具有丰富的天然色素。经活细胞色素光谱吸收峰值测定,色素经有机溶剂提取、硅胶薄板层析、SDS－PAGE电泳等,结果表明H3菌株的主要色素包括细菌叶绿素a、细菌脱镁叶绿素（Bacteriophaeophytin）和三种类胡萝卜素。总胡萝卜素含量占细胞于重的0．6％,胡萝卜素蛋白复合体的分子量约11,000.培养条件的差异对色素形成及相对含量有不同程度的影响。     

Extraction of chlorophyll a from freshwater phytoplankton for spectrophotometric analysis

The efficiency of pigment extraction forms the crux of the spectrophotometric analysis of chlorophyll a. The alcoholic solvents, methanol and ethanol, proved to be superior to acetone and acetone with DMSO. Homogenisation and sonication did not improve the extraction in the alcoholic solvents. Boiling at 100°C had an adverse effect whereas complete extraction of the pigments was obtained at the solvents boiling point and allowing the samples to stand for 24 h in the dark.     

The relationship between chlorophyll and the carotenoids in the algal flagellate,Euglena

1. We have obtained an action spectrum for chlorophyll formation in Euglena gracilis. This action spectrum is similar to the absorption spectrum of protochlorophyll. However, efforts to isolate and identify this pigment have been unsuccessful. 2. Porphyrins have been extracted from both the normal and dark-adapted Euglena and a chlorophyll-free mutant. 3. The "action" spectra for chlorophyll and carotenoid synthesis have been found to almost coincide, indicating that the same porphyrin-like molecule may influence the synthesis of both pigments. 4. It is indicated that two porphyrin-like systems are in operation simultaneously, one concerned with carotenoid "removal" and another involved in carotenoid and chlorophyll synthesis.     

A multichannel atomic absorption instrument: simultaneous analysis of zinc, copper, and cadmium in biologic materials

A new method is deseribed for the extraction and determination of chlorophylls a and b. The method is suitable for use with both normal and regreening nitrogen-deficient Chlorella fusca. The assay involves extraction of chlorophylls by an alkaline pyridine reagent which converts the isocyclic ring of the pigment to a cyclic hydroxylactone. Millimolar extinetion coefficients for the hydroxylactone derivatives of both chlorophylls a and b have been determined at 419 and 454 nm. Using these coefficients, equations have been derived for the calculation of chlorophyll a and b concentrations. The new chlorophyll assay has been compared with other assays which involve the extraction of the pigments with 80% acetone or methanol. The new procedure extracts chlorophylls from rormally grown C. fusca more readily than methanol; the chlorophylls of normal Chlorella cannot be extracted with 80% acetone. The new assay is especially useful in the study of chlorophyll synthesis in regreening nitrogen-deficient C. fusca since the chlorophylls present in these deficient cells cannot be completely extracted with acetone, methanol, methanol-dimethylsulphoxide mixtures, or KOH-methanol.     

Detection of chlorophyllide in chlorophyll-free plasma membrane preparations from Anacystis nidulans

Plasma and thylakoid membranes were isolated and purified from the cyanobacterium Anacystis nidulans. Spectrophotometric examination of acetone extracts gave major absorption bands resulting from carotenoids and chlorophyll a in plasma and thylakoid membranes, respectively. Only a very small absorption peak at 663 nm was detected in acetone extracts of plasma membranes which, in contrast to the corresponding peak from thylakoid membranes, could not be extracted into n-hexane; methanol, on the other hand, was effective with both plasma and thylakoid membranes. Aqueous membrane suspensions excited at 435 nm gave strong fluorescence emission at 662 nm for plasma membranes, but only a very small one for thylakoid membranes which had been adjusted to equal absorbance at 678 nm. Excitation spectra of the 668 nm fluorescence emission peak in acetone extracts of plasma and thylakoid membranes were strikingly different from each other. Finally, high performance liquid chromatography afforded clear-cut preparative separation of the two "chlorophyll-like" pigments in plasma and thylakoid membranes, respectively, and identification by comparison with retention characteristics known from the literature, together with a pure chlorophyll a standard. Our results indicate that the highly fluorescent and polar "chlorophyll-like" pigment in plasma membranes of Anacystis is a chlorophyll precursor, viz. chlorophyllide a.