Function of silica bodies in the epidermal system of rice (Oryza sativa L.): testing the window hypothesis

Silicon has been considered to be important for normal growth and development of the rice plant (Oryza sativa L.). To investigate the physiological function of deposited silica in rice leaves, the hypothesis that silica bodies in the leaf epidermal system might act as a 'window' to facilitate the transmission of light to photosynthetic mesophyll tissue was tested. The silica content of leaves increased with supplied silicon and was closely correlated with the number of silica bodies per unit leaf area in the epidermal system. There was a significant difference in silica deposition and formation of silica bodies between Si-treated and non-treated leaves; silicon was polymerized inside the silica cells and bulliform cells of the epidermis, in Si-treated leaves. Although the 'windows' were only formed in leaves with applied silicon, optical properties of leaf transmittance, reflectance and absorptance spectra in Si-treated and non-treated leaves were almost equal. Furthermore, light energy use efficiency and quantum yield of Si-treated leaves were less than in leaves not containing silica bodies. Thus, silica bodies, at least based on the data, do not function as windows in rice leaves.     

Application of supercritical CO2 for extraction of polyisoprenoid alcohols and their esters from plant tissues

In this study, a method of supercritical fluid extraction (SFE) with carbon dioxide of polyisoprenoids from plant photosynthetic tissues is described. SFE was an effective extraction method for short- and medium-chain compounds with even higher yield than that observed for the “classical extraction” method with organic solvents. Moreover, SFE-derived extracts contained lower amounts of impurities (e.g., chlorophylls) than those obtained by extraction of the same tissue with organic solvents. Elevated temperature and extended extraction time of SFE resulted in a higher rate of extraction of long-chain polyisoprenoids. Ethanol cofeeding did not increase the extraction efficiency of polyisoprenoids; instead, it increased the content of impurities in the lipid extract. Optimization of SFE time and temperature gives the opportunity of prefractionation of complex polyisoprenoid mixtures accumulated in plant tissues. Extracts obtained with application of SFE are very stable and free from organic solvents and can further be used directly in experimental diet supplementation or as starting material for preparation of semisynthetic polyisoprenoid derivatives, e.g., polyisoprenoid phosphates.     

Function of silica bodies in the epidermal system of rice (Oryza sativa L.): testing the window hypothesis

Silicon has been considered to be important for normal growthand development of the rice plant (Oryza sativa L.). To investigatethe physiological function of deposited silica in rice leaves,the hypothesis that silica bodies in the leaf epidermal systemmight act as a ‘window’ to facilitate the transmissionof light to photosynthetic mesophyll tissue was tested. Thesilica content of leaves increased with supplied silicon andwas closely correlated with the number of silica bodies perunit leaf area in the epidermal system. There was a significantdifference in silica deposition and formation of silica bodiesbetween Si-treated and non-treated leaves; silicon was polymerizedinside the silica cells and bulliform cells of the epidermis,in Si-treated leaves. Although the ‘windows’ wereonly formed in leaves with applied silicon, optical propertiesof leaf transmittance, reflectance and absorptance spectra inSi-treated and non-treated leaves were almost equal. Furthermore,light energy use efficiency and quantum yield of Si-treatedleaves were less than in leaves not containing silica bodies.Thus, silica bodies, at least based on the data, do not functionas windows in rice leaves. Key words: Silicon, window hypothesis, rice, optical property, quantum yield     

Isolation of a wide range of minerals from a thermally treated plant: <Emphasis Type="Italic">Equisetum arvense</Emphasis>, a Mare’s tale

Silica is the second most abundant biomineral being exceeded in nature only by biogenic CaCO₃. Many land plants (such as rice, cereals, cucumber, etc.) deposit silica in significant amounts to reinforce their tissues and as a systematic response to pathogen attack. One of the most ancient species of living vascular plants, Equisetum arvense is also able to take up and accumulate silica in all parts of the plant. Numerous methods have been developed for elimination of the organic material and/or metal ions present in plant material to isolate biogenic silica. However, depending on the chemical and/or physical treatment applied to branch or stem from Equisetum arvense; other mineral forms such glass-type materials (i.e. CaSiO₃), salts (i.e. KCl) or luminescent materials can also be isolated from the plant material. In the current contribution, we show the chemical and/or thermal routes that lead to the formation of a number of different mineral types in addition to biogenic silica.     

Purification of protease-extracted glycosaminoglycans by short-distance paper chromatography with a zinc acetate solvent

A method for the purification of tissue-extracted glycosaminoglycans, using short-distance (5-cm path) paper chromatography with 0.25 m zinc acetate solvent, pH 5.0, is presented. The method removed 96 to 97% of the impurities present in the original tissue extract, based on absorbance at 260 nm and protein and phosphate contents. Recovery of control samples of dermatan sulfate averaged 88%. In place of zinc, manganous and mercuric acetates were used to test the method; however, zinc was found to be preferable.     

Restoration of a species‐rich flood meadow by topsoil removal and diaspore transfer with plant material

Abstract. In previous studies, limited dispersal was revealed to be the main obstacle to restoration of species‐rich flood‐meadows along the northern Upper Rhine in Germany. To overcome dispersal limitation we transferred freshly mown plant material from species‐rich sources to a restoration site on a former arable field. Before plant material application, topsoil was removed to accelerate nutrient impoverishment and create favourable conditions for seedling recruitment. Topsoil removal led to a drastic reduction in organic matter and essential mineral nutrients to the level of target communities (P) or even below (N, K). At a removal depth of 30 cm content of the soil seed bank that comprised exclusively of annual arable weeds, ruderals and some common grassland species, declined by 60 ‐ 80%, while at a removal depth of 50 cm the seed bank was almost completely eliminated. With few exceptions, all species recorded in source plant material were found established at the restoration site. However, the overall correlation between seed content in plant material and establishment success was not very high. Vegetation development at the restoration site was characterized by a rapid decline in arable weeds and ruderals, while resident grassland species and species transferred with plant material increased rapidly from the third year onwards. After four years as many as 102 species were established that could be exclusively attributed to plant material transfer, among them many rare and highly endangered plants. Establishment of species from plant material was most successful in regularly flooded plots, due to the suppression of competitors as well as the creation of favourable moisture conditions for seedling emergence. Diaspore transfer with plant material proved to be an extremely successful method in restoring species‐rich grassland. However, high quality of plant material and suitable site conditions with low competition in early stages of succession seem to be essential prerequisites.     

Detection of Helicobacter pylori DNA in human feces by PCR: DNA stability and removal of inhibitors

In this study, the stability of Helicobacter pylori DNA in human feces and the effect of a diet lacking in plant material, the suspected source of PCR inhibitors in human feces, were investigated. In addition, a method to remove these inhibitors was developed. Stools inoculated with H. pylori were used as a model. For this purpose, a H. pylori suspension (10(8) CFU/ml) was used to spike stool samples obtained from four healthy adults known to be H. pylori negative. The evaluation of the stability of H. pylori DNA in feces showed that DNA was degraded after 3 days of contact with fecal material at 37 degrees C. A 2-day diet completely free of plant material was sufficient to eliminate PCR inhibitors from human feces. However, inhibitors were detected 48 h after a normal diet was resumed. A new technique consisting of agarose blocks containing embedded DNA as a template for PCR amplification was used for removal of inhibitors, following DNA extraction by a modified QIAamp tissue method (Qiagen, Hilden, Germany). When this method was applied to inhibiting stool samples known to have an inhibitory effect and spiked with H. pylori (5.10(8) CFU/g), a positive PCR was obtained showing that inhibitors present in the original DNA samples were completely removed. The agarose embedded DNA block method is highly efficient and provides clean, high quality template DNA for PCR purposes avoiding long and fastidious conventional extraction methods. In conclusion, this study confirms that H. pylori DNA degrades with time in stools. A diet free of plant material or a special DNA preparation can be used to remove inhibitors and to allow the detection of H. pylori.     

Matric potential evaluations and measurements for gelled substrates

A new method for evaluating the matric potential of gelled media has been developed. The method allows the derivation of the matric potential as a limit of a series of measurements of water potential values from gelled media prepared without added components, from agar powders progressively cleaned of mineral impurities. Three commercial agar brands were tested, and for these the matric potential was found to contribute only between 1 and 2% of their total water potential. Thermodynamic features relating matric and osmotic potentials are described. New hypotheses for understanding the water flux mechanism from gel to tissue cultured explants are discussed. Movement of water along polymeric chains is postulated to be a facilitated step in comparison with bulk movement.     

Preparation of steroid radioligands for ultrafiltration assays by a solid-phase transport globulin method using concanavalin A-Sepharose

Concanavalin A-Sepharose (Con A-Sepharose) was applied to separate non-protein-bound and albumin-bound radioactive impurities from steroid radioligands. Con A-Sepharose gel, plasma, and steroid radioligand were mixed, incubated, and then washed with buffer. This method was compared with an affinity diafiltration method which separates non-protein-bound radioactive impurities with a filter membrane. 3H-Water and 3H-estrone sulfate, chosen to serve as molecules representative of non-protein-bound and albumin-bound impurities, were removed quite effectively by the Con A-Sepharose method, while 85% of 3H-estrone sulfate could not be removed by the diafiltration method. Plasma unbound cortisol (F) and testosterone (T) values determined by ultrafiltration using 3H-F and 3H-T prepared by the Con A-Sepharose method were significantly lower than those determined using the radioligands unprocessed or prepared by the diafiltration method. The whole procedure of the Con A-Sepharose method takes only 3-4 hours. This method is a simple, rapid, and effective technique for preparation of steroid radioligands for plasma protein binding studies.     

Silicon in Imperata cylindrica (L.) P. Beauv: content,distribution, and ultrastructure

Silicon concentration, distribution, and ultrastructure of silicon deposits in the Poaceae Imperata cylindrica (L.) P. Beauv. have been studied. This grass, known for its medicinal uses and also for Fe hyperaccumulation and biomineralization capacities, showed a concentration of silicon of 13,705?±?9,607 mg/kg dry weight. Silicon was found as an important constituent of cell walls of the epidermis of the whole plant. Silica deposits were found in silica bodies, endodermis, and different cells with silicon-collapsed lumen as bulliforms, cortical, and sclerenchyma cells. Transmission electron microscope observations of these deposits revealed an amorphous material of an ultrastructure similar to that previously reported in silica bodies of other Poaceae.     

Nanosilica synthesis mediated by Aspergillus parasiticus strain

Rice husks (RHs) are plant waste materials abundant in phytoliths silica bodies. These were used as starting material for fungal-mediated biotransformation leading to the synthesis of a high-value added product. A strain of Aspergillus parasiticus was capable of transforming the amorphous silica conglomerates into structured nanoparticles (NPs) in the process of RHs biotransformation. Silica NPs were produced extracellularly and their size ranged from 3 to 400 nm depending on the biotransformation conditions and the post-biotransformation supernatant processing. To characterize the NP's structure and dimension, SEM, STEM, EDX and FTIR technics were applied. These demonstrated and confirmed that pyramid (400 nm), cubical (85 nm) and spherical (3 nm and 24 ± 8 nm) forms of silica NPs were obtained.     

Variation of silica bodies in leaf epidermal long cells within and among seventeen species of Oryza (Poaceae)

Morphometric procedures were used with scanning electron microscopy backscattered images to study silica bodies in epidermal long cells of four different leaf veins of 17 of the 20 species of Oryza. The veins studied were midrib, large vein, small vein, and marginal vein. Image analysis was used to study morphological variations among the silica bodies. Statistical analyses were based on 11 variables. Even within a single leaf, silica bodies were not uniform. However, the degree of morphological variation normally showed a distribution of morpholgical types around one modal shape. The most significant differences observed were between silica bodies of the midrib and those of other veins. Bodies varied with respect to both size and shape. Computer-assisted image analysis is an effective tool for categorizing basic data and for statistical analysis of variation among silica bodies. Morphological variation among silica bodies of a single leaf may be related to water-conducting systems and their influence on silica availability and phytolith formation.     

凤尾竹硅酸体形态及其发生初探

利用光学显微镜、扫描电子显微镜对自然生长凤尾竹(Bambusa multiplex vat．nana(Roxb．)Keng f．)营养体各部分及生长发育各时期叶片的硅酸体进行观察研究。凤尾竹营养体各部分硅酸体形态分别为：叶片竹节形;叶鞘鞍形、钝方形;箨叶哑铃形;箨叶鞘鞍形;杆的表皮部位有少量椭圃形硅酸体,而内部来见成形的硅酸体;地下茎及根皆来见成形硅酸体。成熟硅酸体均沿脉横向排列。硅酸体在叶片、叶鞘、箨叶和箨叶鞘中基本上是按生长发育顺序从不成形到成形,由小到大变化的,分布由不规则到沿脉横列。同一叶从叶鞘到叶片呈现出由鞍形→钝方形(椭圆形)→竹节形渐变的趋势。光镜下,硅酸体内一般有折光率不同于其它部位的l到多个细小颗粒聚在一起形成的核状结构。凤尾竹不同生长发育期各部分硅酸体的晶核存在状况、硅酸体的形态和大小的演变趋势体现了硅质在植物器官表皮细胞上的连续沉积过程。     

Purification of transgenic tobacco-derived recombinant human monoclonal antibody

Purification of recombinant monoclonal antibody from transgenic plant extract is technically challenging as it involves the processing of large volume of material, containing low titre of antibody, present along with large quantities of native proteins and other impurities. The conventional approach of capturing antibody from a clarified extract using packed-bed chromatography is therefore not particularly suitable. This study evaluates the suitability of using a combination of ultrafiltration and chromatography for purifying transgenic tobacco-derived human monoclonal antibody. A two-stage cascade ultrafiltration process removed about 97% impurities while ensuring almost complete recovery of antibody, providing 32-fold antibody enrichment in the process. The primary objective of the ultrafiltration step was to reduce the burden on the subsequent chromatographic steps. A two-step chromatographic process was then used to eliminate remaining impurities. Using this approach, recombinant human antibody expressed in tobacco could be purified to greater than 95% purity with 50% overall recovery (ca. 12.5 mg antibody/kg tobacco tissues).     

Salt drying: a low‐cost,simple and efficient method for storing plants in the field and preserving biological repositories for DNA diversity research

Although a variety of methods have been optimized for the collection and storage of plant specimens, most of these are not suited for field expeditions for a variety of logistic reasons. Drying specimens with silica gel in polyethylene bags is currently the standard for field‐sampling methods that are suitable for subsequent DNA extraction. However, silica‐gel repositories are not readily available in remote areas, and its use is not very cost‐effective for the long‐term storage of collections or in developing countries with limited research budgets. Salting is an ancient and traditional drying process that preserves food samples by dehydrating tissues and inhibiting water‐dependent cellular metabolism. We compared salt and silica‐gel drying methods with respect to dehydration rates overtime, DNA quality and polymerase chain reaction(PCR) success to assess whether dry salting can be used as an effective plant preservation method for DNA analysis. Specimens from eleven plant species covering a variety of leaf structures, leaf thicknesses and water contents were analysed. Experimental work indicated that (i) levels of dehydration in sodium chloride were usually comparable to those obtained when silica gel was used, (ii) no spoilage, fungal or bacterial growth was observed for any of the species with all drying treatments and (iii) good yields of quality genomic DNA suitable for PCR applications were obtained in the salt‐drying treatments. The preservation of plant tissues in commercial table salt appears to be a satisfactory, and versatile method that may be suitable in remote areas where cryogenic resources and silica repositories are not available.     

人用精制Vero细胞狂犬病疫苗纯化方法选择

人用精制Ｖｅｒｏ细胞狂犬病疫苗为一种安全、有效的新型疫苗,我们在研制该种疫苗时首先比较了密度梯度离心法和凝胶过滤柱层析法,并发现后者最佳。我们选择了以Ｓｅｐｈａｒｏｓｅ ４ＦＦ为介质的凝胶过滤柱层析的工艺,并对该方法的上样量、流速等指标进行了选择,确定了最佳方法,并在研制中使纯化疫苗的杂蛋白去除率达到９９．８％以上,为大规模生产提供了工艺方法。     

Mercury-biogeochemical exploration for mineral deposits

Biogeochemical prospecting for mercury deposits and deposits of other minerals by the chemical analysis of mercury in plants or plant tissue that accumulate this element in linear (or near linear) proportion to the concentration in the soil is an effective method of exploration, even where allochthonous material as much as 200 to 2000 m thick covers the deposits. Plant tissues with this tendency to accumulate mercury (designated as non-barrier to mercury) comprise only a small fraction of the total of 255 types of plant tissues that were tested. Ten of these were considered to be quantitatively informative, and their mercury concentrations exceeded background values 300 or more times. The remaining types of plant tissues ranged in prospecting value from semi-quantitatively informative to qualitatively informative to uniformative (mercury values at or below background). The failure of some earlier uses of this prospecting method is attributed to the use of inappropriate plant tissues, to the mercury in the particular substrate studied existing in a form of low mobility and availability to plants, or to both causes.Prospecting by examining mercury concentrations in soils and rocks (lithogeochemical prospecting) is more effective than the biogeochemical approach only in prospecting for cinnabar deposits having no allochthonous cover. Mercury-biogeochemical prospecting is most effective for non-mercury mineral deposits and for oil and gas deposits. The types of plant tissues used in these studies are listed and are classified according to their value in prospecting. A case history is given of the Ozernoe pyrite-polymetallic deposit in Siberia.     

A novel downstream process for 1,3-propanediol from glycerol-based fermentation

In this paper, a downstream process for purification of 1,3-propanediol from glycerol-based fermentation broth was investigated. The purification of 1,3-propanediol from fermentation broth was achieved by a process combining microfiltration, charcoal treatment, vacuum distillation, and silica gel chromatography. The broth was first filtered through hollow fiber cartridge, wherein 98.7% of biomass was removed. Soluble proteins and other color impurities in the broth were removed by the use of activated charcoal at optimal concentration of 30 g l⁻¹ where the soluble proteins in the broth decreased to 0.1 g l⁻¹ (96.0% protein loss). The obtained broth when concentrated by vacuum distillation resulted in the crystallization of inorganic salts. Subsequently, 1,3-propanediol was purified by gradient chromatography using silica gel as a stationary phase and mixture of chloroform and methanol as a mobile phase. Finally, with the optimal flow rate of 10 ml min⁻¹ and loading amount of 80 ml, the yield of 1,3-propanediol achieved was 89%. The overall yield of 1,3-propanediol using the proposed procedure was 75.47%. The developed method was found to be a simple, rapid, and efficient procedure for the purification of 1,3-propanediol from fermentation broth.     

Evaluation of adsorbents for separation and purification of paclitaxel from plant cell cultures

In this study, we evaluated the efficiency of different adsorbents for the removal of plant-derived impurities during the pre-purification of paclitaxel from plant cell cultures. Using the synthetic adsorbents sylopute and active clay and their major components SiO₂ and MgO, we performed adsorbent treatment and analyzed the paclitaxel precipitates recovered from hexane precipitation. When SiO₂ was used, the highest purity (～58.1%) and yield (～91.5%) of paclitaxel were obtained. We also determined differences in the effectiveness of the adsorbent treatment according to changes in the surface area, pore volume and pore diameter of SiO₂. Adsorbent treatment was more effective when pore diameter was larger (silica I [2.19 nm] < silica II [4.92 nm] < silica III [9.07 nm]). The highest purity (～74.3%) and yield (～92.9%) of paclitaxel were obtained when silica III was used in the adsorbent treatment. Pore diameter had a greater effect on the removal of plant-derived impurities during the pre-purification of paclitaxel compared with surface area and pore volume. This result could be confirmed by HPLC analysis of the absorbent after treatment and TGA of the organic substances that were bonded to the adsorbent.     

Biomining with bacteriophage: selectivity of displayed peptides for naturally occurring sphalerite and chalcopyrite

During mineral processing, concentrates of sulfide minerals of economic interest are formed by froth flotation of fine ore particles. The method works well but recovery and selectivity can be poor for ores with complex mineralogy. There is considerable interest in methods that improve the selectivity of this process while avoiding the high costs of using flotation chemicals. Here we show the first application of phage biotechnology to the processing of economically important minerals in ore slurries. A random heptapeptide library was screened for peptide sequences that bind selectively to the minerals sphalerite (ZnS) and chalcopyrite (CuFeS2). After several rounds of enrichment, cloned phage containing the surface peptide loops KPLLMGS and QPKGPKQ bound specifically to sphalerite. Phage containing the peptide loop TPTTYKV bound to both sphalerite and chalcopyrite. By using an enzyme-linked immunosorbant assay (ELISA), the phage was characterized as strong binders compared to wild-type phage. Specificity of binding was confirmed by immunochemical visualization of phage bound to mineral particles but not to silica (a waste mineral) or pyrite. The current study focused primarily on the isolation of ZnS-specific phage that could be utilized in the separation of sphalerite from silica. At mining sites where sphalerite and chalcopyrite are not found together in natural ores, the separation of sphalerite from silica would be an appropriate enrichment step. At mining sites where sphalerite and chalcopyrite do occur together, more specific phage would be required. This bacteriophage has the potential to be used in a more selective method of mineral separation and to be the basis for advanced methods of mineral processing.