Vanillin-hydrochloric acid as a histochemical test for tannin.

Condensed tannin (leucoanthocyanins and catechins) can be demonstrated in fresh plant sections with saturated alcoholic vanillin followed by addition of concentrated HCl. Bright red vanillin-tannin condensates are formed immediately. Preparations may then be made permanent by mounting in a 1:1 mixture of Hoyer's medium and concentrated HCl. Some fading and loss of tannin occurs. The phloroglucinol-HCl test for lignin can also be made permanent with this acidic mountant.     

A Modification of the Cresyl Violet Technic for Staining Nerve Cells

Kill root tips in 1 part glacial acetic acid to 3 parba RB Solute alcohol for 12 or more hours. Remove from king fluid a d place for 5 to 10 minutes in a solution consisting of 1 part 95% alcohol to 1 part concentrated HC1. Transfer to Carnoy's fluid for 5 minutes or longer. Cut a small piece (0.5 mm. or less) off the tip of the root Press directly on the piece of root with a small fiat scalpel; the cells will now separate and float free in the stain. Place cover slip over the drop of stain and apply gentle pressure. Heat carefully by paseing the slide 3 or 4 times thru the flame of an alcohol lamp. Seal with heated mixture of 1 part Parowax to 1 part gum mastie. Make permanent by the McClintock permanent method. and place on a clean slide in a small drop of iron-ace-sinin.     

A Rapid HCl/Toluidine Blue Squash Technic for Plant Chromosomes

Fresh or pretreated root tips are simultaneously fixed and hydrolysed in 5 N HC1 for 15 min at room temperature. After washing they are macerated on a slide in a drop of 0.05% toluidine blue made up in McIlvaine citric acid-Na₂HPO, buffer at pH 4.0. Pressure on the cover slip completes the squash preparation. It is made permanent by removing the cover slip on dry ice, air drying and mounting in Euparal.     

Dinucleotide repeat (GT)n markers on chromosome 21.

To further develop the linkage map of human chromosome 21 (HC21), we have concentrated on identifying highly polymorphic markers based on dinucleotide repeat sequences such as (GT)n, as these are often highly polymorphic, are widespread throughout the human genome, and can be rapidly analyzed by the polymerase chain reaction. We report here nine (GT)n polymorphic markers from HC21.     

单宁细胞形态与部分柿属种及品种相关性研究

采用软柿果肉直接压涂法,在光学显微镜下对204个柿属种及品种果实中的单宁细胞形态特征进行观察分析.结果显示:(1)在6个种柿属植物果实中,均含有单宁细胞,其外形大多属于短形和近圆形,但在数量、大小和颜色上存在差异,其中,油柿(Diospyros oleifera Cheng.)、君迁子(D.lotus Linn.)、柿(D.kaki Thunb.)、浙江柿(D.glaucifolia Metc.)的单宁细胞通常无色,而黑柿(D.nitida Mcrr.)为黄绿色,乌材(D.eriantha Champ.)为紫红色,乌柿(D.eathayensis Stheward.)为淡紫色;单宁细胞从大到小依次为油柿>君迁子>浙江柿>乌材>黑柿>乌柿.(2)单宁细胞在不同品种类型间差异明显,其中涩柿单宁细胞多为无色,单宁细胞比较宽大;甜柿品种均会出现褐变的单宁细胞,单宁细胞较小或瘦长;完全甜柿品种大多存在着凝固型褐变单宁细胞,仅少数凝聚呈球形,且单宁细胞分散存在于果肉中,果肉中的褐斑较细小;不完全甜柿在种子周围的褐斑处,可以看到大量的表面凹形且褐变的收缩型单宁细胞,且常以单宁细胞束的形态存在于果肉中,使果肉中的褐斑大而密;原产我国的完全甜柿中不存在凝聚型的单宁细胞,只有凝固型的单宁细胞.(3)聚类分析结果表明,单宁细胞的特征可以作为不同类型柿属种的分类依据.     

Hydrochloric acid as a Fixative for root Tip Chromosomes

After pretreatment with 0.2 to 0.3% colchicine at 26° C. for 2 hours root tips are fixed and macerated in a 1/10 dilution of concentrated hydrochloric acid at 60° C. for 10 to 14 minutes, washed, transferred to a staining dish with aceto-orcein or another aceto-stain for about 10 minutes, immersed in a drop of the stain, covered and squashed. The preparations may subsqeuently be made permanent.     

Autonomic involvement in the permanent metabolic programming of hyperinsulinemia in the high-carbohydrate rat model

Exposure to a high-carbohydrate (HC) milk formula during the suckling period results in permanent metabolic programming of hyperinsulinemia in HC rats. Previous studies have shown that hyperinsulinemia in HC rats involves a programmed hyperresponsiveness to glucose. However, the immediate onset and persistence of enhanced insulin secretion throughout life suggests a role for numerous factors that control insulin secretion. Present in vivo and in vitro studies have shown a role for altered autonomic activity, including increased parasympathetic and decreased sympathetic activities, in the maintenance of hyperinsulinemia in 100-day-old HC rats. HC rats were shown to be more sensitive to cholinergic-induced potentiation of glucose-stimulated insulin secretion (GSIS) in response to acetylcholine and showed increased sensitivity to blockade of cholinergic-induced insulin secretion by the muscarinic-type 3 receptor-specific antagonist 4-diphenylacetoxy-N-methylpiperidine. In addition, HC rats were less sensitive to adrenergic-induced inhibition of insulin secretion by oxymetazoline, whereas treatment with yohimbine resulted in increased GSIS. Furthermore, HC rats showed greater reductions in plasma insulin levels after vagotomy, as well as an attenuation of yohimbine-induced potentiation of GSIS, suggesting that yohimbine-mediated changes are mediated by parasympathetic activity. Changes in autonomic regulation of GSIS are supported by increased mRNA levels of the parasympathetic signaling molecules muscarinic-type 3 receptor, phospholipase Cbeta1, and protein kinase C-alpha and decreased levels of alpha(2a)-adrenergic receptors in islets from adult HC rats. In conclusion, metabolic programming of hyperinsulinemia throughout adulthood of HC rats involves changes in autonomic activity in response to the HC dietary intervention in the suckling period.     

Ultrastructural and biological characterization of human choroid cell cultures transformed by Simian Virus 40

A human diploid cell line of choroid origin was isolated from the retrouveal portion of an enucleated eye and designated HC. After 10 passages, when the proliferative capacity of HC cells decreased, they were infected and transformed by Simian Virus 40 (SV40). A proliferating long-term cultured cell line designated HC/SV40 was established and it has been maintained as monolayer for more than 100 passages so far. The two cell lines, HC and HC/SV40, were compared for growth characteristics, capacity to form colonies in soft agar, presence of nuclear T-antigen, and ultrastructure. Cytogenetic analysis was also performed to determine the presence of chromosomal aberrations due to the permanent viral transformation of the cell line. The results indicate that HC/SV40 should be considered the transformed counterparts of HC cells because they are morphologically similar to the latter but can grow in soft agar, possess T-antigen, and show a pattern of karyotypic changes similar to that induced by SV40 in human fibroblasts. The choroid origin of HC and HC/SV40 cell lines was confirmed by the presence, in their cytoplasm, of typical electron dense granules. Their neural origin will make these cell lines very useful for neuropharmacological and differentiation studies.     

蒲桃和人面子叶片单宁含量与凋落物分解速率及底栖动物定殖的关系

在广州龙洞水库一条天然2级溪流中,测定了蒲桃和人面子凋落物105 d分解过程中单宁含量的变化.结果表明：蒲桃叶片单宁的初始含量（0.191 g·g^-1DM）高于人面子（0.057 g·g^-1DM）.在最初一周内, 两种树木叶片的单宁含量分别下降了45 %和22 %,其中人面子叶片单宁含量降速比蒲桃快;21 d后,其下降速度减缓,而凋落物分解的速度加快,人面子叶片分解比蒲桃迅速（k值分别为0.038和0.013 d^-1）.定殖在人面子叶片上的底栖动物的平均密度显著高于蒲桃叶片（P<0.05）,分别为每克叶片287.9头和26.2头;底栖动物的数量变化随叶片单宁含量的降低而呈逐渐增加趋势.富含单宁成分的蒲桃叶片分解速率缓慢,可能是凋落物中高浓度缩合单宁抑制了底栖动物,尤其是撕食者的定殖所致.     

Chromosomes of Sequoia sempervirens; 8-Hydroxy-Quinoline-Castor Oil Pretreatment for Improving Preparation

Living root tips of coast redwood were pretreated for 6 hr at 10-14 C in a homogenized mixture of 0.003 M aqueous solution of about 5 ml of 8-hydroxyquinoline and a drop of castor oil. They were rinsed 2-3 times in Carnoy's alcohol-chloroform-acetic acid (3:2:1), and fixation allowed to continue in this fluid for 1 hr at 60 C. They were then hydrolysed in 1 N HC1 at 60 C for 45 min; washed with distilled water, and squashed in a drop of aceto-carmine or aceto-orcein to make a temporary slide, subsequently made permanent by a quick-freezing method. Our work to date seems to confirm 2n = 66 for S. sempervirens.     

Alcoholic Hydrochloric Acid-Carmine as a Stain for Chromosomes in Squash Preparations

A regressive bulk carmine staining schedule was adapted from a formula proposed by P. Mayer. The stain is made by boiling gently 4 gm of certified carmine in 15 ml of distilled water to which 1 ml of concentrated HC1 has been added. After cooling, 95 ml of 85% alcohol is added, and the solution filtered. Fixed tissue is soaked in the stain until thoroughly penetrated; squashes are then prepared as usual, but plain 45% acetic acid is used as the temporary mounting medium instead of aceto-carmine. The advantages of this method are: intense, precise staining of chromosomes coupled with a lightly stained cytoplasm; consistency and uniformity of results; simplicity of use.     

Ultrastructural and biological characterization of human choroid cell cultures transformed by Simian Virus 40

Summary A human diploid cell line of choroid origin was isolated from the retrouveal portion of an enucleated eye and designated HC. After 10 passages, when the proliferative capacity of HC cells decreased, they were infected and transformed by Simian Virus 40 (SV40). A proliferating long-term cultured cell line designated HC/SV40 was established and it has been maintained as monolayer for more than 100 passages so far. The two cell lines, HC and HC/SV40, were compared for growth characteristics, capacity to form colonies in soft agar, presence of nuclear T-antigen, and ultrastructure. Cytogenetic analysis was also performed to determine the presence of chromosomal aberrations due to the permanent viral transformation of the cell line. The results indicate that HC/SV40 should be considered the transformed counterparts of HC cells because they are morphologically similar to the latter but can grow in soft agar, possess T-antigen, and show a pattern of karyotypic changes similar to that induced by SV40 in human fibroblasts. The choroid origin of HC and HC/SV40 cell lines was confirmed by the presence, in their cytoplasm, of typical electron dense granules. Their neural origin will make these cell lines very useful for neuropharmacological and differentiation studies. This work was supported by Grant 82.00346.96 from the C.N.R. finalized project “Control of Neoplastic Growth”.     

Condensed tannins and sugars in the diet of chimpanzees (Pan troglodytes schweinfurthii) in the Budongo Forest, Uganda

Fruits, leaves and bark forming part of the diet of chimpanzees were collected and it was noted whether samples were of a kind being eaten or not eaten. Samples were dried and analysed for condensed tannin content and for three sugars, glucose, sucrose and fructose. It was found that chimpanzees did not select foods according to the level of tannins but did so according to the levels of sugars, preferring the higher levels. Fig seeds contained higher tannin levels than fig pulp, and the chimpanzees made oral boli (“wadges”) of fig seeds which they spat out. Two fig species were compared: the one with lower tannin and higher sugar content was preferred. The bark of one tree species often eaten contained high levels of tannins but also contained sugars. Young leaves with lower tannin levels were preferred to mature leaves with higher levels. Chimpanzees appear to be able to tolerate higher tannin levels than three monkey species in this forest, and considerably higher levels than marmosets (Callitrichidae). Received: 20 October 1997 / Accepted: 1 March 1998     

山茱萸果实发育过程中单宁物质的分布与积累特征

选取不同发育时期的山茱萸果实作为研究对象,采用果实形态观察法、显微及超微技术、组织化学定位法以及紫外分光光度计法对山茱萸果实发育过程中单宁物质分布及积累特征进行观察分析,并以单因素ANOVA检验不同发育时期单宁含量的差异,以揭示单宁物质在山茱萸果实发育中的变化规律,为山茱萸果实涩味调控机制研究提供理论依据。结果表明:(1)山茱萸果实发育过程中果皮颜色和果实体积变化明显,可将其发育过程划分为幼果期、中果期、成熟期3个时期;单宁物质主要分布在山茱萸果实中果皮的单宁细胞中。(2)在山茱萸果实发育过程中单宁细胞数目呈先增后减的变化趋势,幼果期单宁细胞从无到有,随着果实发育单宁细胞数目不断增多,至中果期单宁细胞数目开始减少。(3)单宁含量的变化规律与单宁细胞数目的变化一致,单宁含量在花后120 d时达到最多,随后逐渐减少。(4)单宁物质首先在细胞质的小液泡中积累,中央大液泡形成后则为单宁物质积累的主要场所,其积累形态主要有颗粒状、不规则状和板块状3种;单宁细胞中线粒体数目较多,中果期后期及成熟期在中央大液泡液泡膜附近有电子致密物质积累。研究认为,山茱萸果实中中果皮薄壁细胞为单宁物质积累的专属细胞,即单宁细胞,单宁物质的合成运输与液泡、囊泡以及线粒体的作用密切相关;成熟期山茱萸果实总单宁含量降低,涩味降低,表明单宁物质积累的动态变化与植物对环境的适应性和果实涩味息息相关,可结合代谢组和转录组的方法对山茱萸果实中单宁物质的合成机制进行进一步研究。     

Interaction of heparin cofactor II with neutrophil elastase and cathepsin G

We investigated the interaction of the human plasma proteinase inhibitor heparin cofactor II (HC) with human neutrophil elastase and cathepsin G in order to examine 1) proteinase inhibition by HC, 2) inactivation of HC, and 3) the effect of glycosaminoglycans on inhibition and inactivation. We found that HC inhibited cathepsin G, but not elastase, with a rate constant of 6.0 x 10(6) M-1 min-1. Inhibition was stable, with a dissociation rate constant of 1.0 x 10(-3) min-1. Heparin and dermatan sulfate diminished inhibition slightly. Both neutrophil elastase and cathepsin G at catalytic concentrations destroyed the thrombin inhibition activity of HC. Inactivation was accompanied by a dramatic increase in heat stability, as occurs with other serine proteinase inhibitors. Proteolysis of HC (Mr 66,000) produced a species (Mr 58,000) that retained thrombin inhibition activity, and an inactive species of Mr 48,000. Amino acid sequence analysis led to the conclusion that both neutrophil elastase and cathepsin G cleave HC at Ile66, which does not affect HC activity, and at Val439, near the reactive site Leu444, which inactivates HC. Since cathepsin G is inhibited by HC and also inactivates HC, we conclude that cathepsin G participates in both reactions simultaneously so that small amounts of cathepsin G can inactivate a molar excess of HC. High concentrations of heparin and dermatan sulfate accelerated inactivation of HC by neutrophil proteinases, with heparin having a greater effect. Heparin and dermatan sulfate appeared to alter the pattern, and not just the rate, of proteolysis of HC. We conclude that while HC is an effective inhibitor of cathepsin G, it can be proteolyzed by neutrophil proteinases to generate first an active inhibitor and then an inactive molecule. This two-step mechanism might be important in the generation of chemotactic activity from the amino-terminal region of HC.     

四种类型栓皮栎栲胶含量

为了阐明不同类型(薄皮深裂、薄皮浅裂、厚皮深裂、厚皮浅裂)栓皮栎各器官栲胶含量,系统采集了秦岭南坡商州区4种类型的栓皮栎不同径级植株根、主干木材、叶片、枝条和树皮样品,用紫外光分光光度法(UV Spectrophotometry)测定了4种类型栓皮栎不同器官栲胶含量,分析了林木生长与栲胶含量的关系。结果表明,(1)4种类型栓皮栎的根、主干木材、叶片、枝条和树皮单宁含量平均值分别为2.64%—3.02%、1.74%—2.02%、6.63%—7.21%、4.69%—5.10%、8.87%—9.46%,叶片、枝条和树皮均可以作为栲胶生产主要原材料。(2)不同类型各器官单宁含量存在极显著差异(P0.01),4种类型的树皮、叶片、枝条单宁含量均表现为:厚皮深裂型厚皮浅裂型薄皮深裂型薄皮浅裂型;根部和主干木材在4种类型间单宁含量表现为:薄皮深裂型薄皮浅裂型厚皮深裂型厚皮浅裂型;厚皮深裂类型的栓皮栎是提取栲胶的最佳类型。(3)4种类型栓皮栎各器官单宁含量与胸径呈显著正相关,各器官单宁含量随着栓皮栎胸径增大而增多,并最终趋于稳定,在第Ⅳ径级(胸径20.1—25cm)时,栲胶含量均值最大。因此,栲胶生产可以高效利用第Ⅳ径级及其以上径级的栓皮栎,并对第Ⅳ径级以下栓皮栎进行定向培育和重点保护。     

Effect of tamarind (Tamarindus indica) seed husk tannins on in vitro rumen fermentation

This study was conducted to determine the effect of tamarind seed husk (TSH) as a source of tannin on various parameters of rumen fermentation in vitro. The TSH contained 14% tannin (DM basis). The biological interference of TSH tannin on rumen fermentation was assessed using polyethylene glycol (PEG) 6000 as an indicator. Three compound feed mixtures (CFM) were prepared either without TSH (CFM-I), with 2.5% TSH (CFM-II) and with 7.5% TSH (CFM-III). Parameters studied were in vitro gas production with PEG, rate of substrate degradation, and microbial protein synthesis. Addition of PEG to TSH resulted in an increase in gas production from 5.5 to 16.5 ml per 200 mg DM. Presence of TSH tannin depressed cumulative gas production by 16.8% in CFM-II, and by 29.2% in CFM-III during initial stages of fermentation (i.e. at 8 h). Rate of substrate disappearance (T^1/2) was 14.4, 17.6 and 20.5 h in CFM-I, CFM-II and CFM-III, respectively. Irrespective of the carbohydrate source, presence of TSH tannin improved the efficiency of microbial protein synthesis in vitro. Thus, TSH is a natural source of tannin that can be used to beneficially manipulate rumen fermentation.     

Concentration of erythrocyte-based magnetic carriers in the vascular bed

The in vivo and in vitro experiments have shown that magnetic erythrocytes (loaded with ferromagnetic material) can be retained in certain sites of the vascular bed. Magnetic erithrocytes could be concentrated in the abdominal part of the dog aorta using a small permanent magnet fixed on the outer wall of the aorta.     

Aleppo tannin: structural analysis and salivary amylase inhibition

The effectiveness and specificity of a tannin inhibition on human salivary amylase (HSA) catalyzed hydrolysis was studied using 2-chloro-4-nitrophenyl 4-O-beta-D-galactopyranosyl-alpha-maltoside (GalG(2)-CNP) and amylose substrates. Aleppo tannin was isolated from the gall nut of Aleppo oak. This tannin is a gallotannin, in which glucose is esterified with gallic acids. This is the first kinetic report, which details the inhibitory effects of this compound on HSA. A mixed non-competitive type inhibition has been observed on both substrates. The extent of inhibition is markedly dependent on the substrate-type. Kinetic constants were calculated from Lineweaver-Burk secondary plots for GalG(2)-CNP (K(EI) 0.82 microg mL(-1), K(ESI) 3.3 microg mL(-1)). This indicates a 1:1 binding ratio of inhibitor-enzyme and/or inhibitor-enzyme-substrate complex. When amylose was the substrate the binding ratio of inhibitor to enzyme-substrate complex was found to be 2:1, with the binding constants of K(EI) 17.4 microg mL(-1), K(ESI) 14.9 microg mL(-1), K(ESI(2)) 9.6 microg mL(-1). Presumably, the tannin inhibitor can bind not only to HSA, but to the amylose substrate, as well. Kinetic data suggest that Aleppo tannin is a more efficient amylase inhibitor than the recently studied other tannin with quinic acid core (GalG(2)-CNP: K(EI) 9.0 microg mL(-1), K(ESI) 47.9 microg mL(-1)).     

Role of caffeine and tannin in anti-toxigenic properties of coffee and tea

Aflatoxins B₁, B₂, G₁ and G₂ produced by Aspergillus parasiticus var. globosus IMI 120920 in detannin-caffeinated coffee and black tea were five times more concentrated than in normal tea and coffee. Extracts of normal coffee and tea powders significantly reduced aflatoxins production in liquid broth at 1 and 3 % concentrations, with tea extract. having a more pronounced effect than coffee extract Relevant anti-aflatoxigenic properties appear to be due to tannin and caffeine. These induced 95 % inhibition in aflatoxins at 0.3 % and 0.6 %, respectively. Roasting of contaminated coffee beans at 200 °C for 20 min is effective in the reduction of aflatoxins.