Search

A survey of the inhibitory effects of various plant polysaccharides on DNA restrictions (HindIII and EcoRI) revealed that neutral polysaccharides (arabino-galactan, dextran, gum guar, gum locust bean, beta-glucan, inulin, laminaran, mannan and starch) were not very inhibitory. In contrast, acidic polysaccharides (carrageenan, dextran sulfate, gum ghatti, gum karaya, pectin and xylan) were very inhibitory, even at low concentrations. The Elutip-d (RPC-5 type resin) was evaluated for removal of the inhibitory polysaccharides. Used alone or in combination with a phenol/chloroform wash, it proved effective in removing the polysaccharide so that HindIII digestion was possible, except in the cases of carrageenan and dextran sulfate. In addition, the genomic DNA extracts from live oak (Quercus virginiana) and magnolia (Magnolia grandiflora) were sufficiently purified so that the DNAs could be restricted with both EcoRI and HindIII.

The water-soluble polysaccharides from dried Ginkgo biloba leaves were isolated after exhaustive extraction with organic solvents. The polysaccharide mixture could be separated into a neutral (GF1) and two acidic (GF2 and GF3) polysaccharide fractions by ion exchange chromatography. According to the Mr distribution GF1 and GF3 seemed to be homogenous, whereas GF2 could be further fractionated into two subfractions (GF2a and GF2b) by gel permeation chromatography. GF1 (Mr 23,000) showed the structural features of a branched arabinan. The main chain was composed of 1,5-linked arabinose residues and three in 12 arabinose molecules were branched via C-2 or C-3. GF2a (Mr 500,000) consisted mainly of 1,2,4-branched mannose (29%), 1,4-linked glucuronic (32%) and galacturonic (8%) acid as well as terminal rhamnose (25%). After removal of ca 70% of the terminal rhamnose the remaining polysaccharide showed a decrease in 1,2,4-branched mannose and an increase in 1,2-linked mannose indicating that at least half of the rhamnose residues were linked to mannose via C-4. GF3 (Mr 40,000) consisted of 1,4-linked galacturonic (30%) and glucuronic (16) acid, 1,3,6-branched galactose (15%), 1,2-linked (5%) and 1,2,4-branched (3.5%) rhamnose as well as 1,5-linked arabinose (11%). Rhamnose (5%) and arabinose (10%) were present as terminal groups. Mild acid hydrolysis selectively cleaved arabinose and the remaining polysaccharide showed an increased amount of 1,6-linked and terminal galactose and a decreased quantity of 1,3,6-branched galactose. These results indicated that the terminal as well as the 1,5-linked arabinose were mainly connected to galactose via C-3. The GF3 polysaccharide appeared to be a rhamnogalacturonan with arabinogalactan side chains.

Purified polysaccharides from cell cultures of the plant Echinacea purpurea were investigated for their ability to enhance phagocytes' activities regarding nonspecific immunity in vitro and in vivo. Macrophages (M phi) from different organ origin could be activated to produce IL-1, TNF alpha and IL-6, to produce elevated amounts of reactive oxygen intermediates and to inhibit growth of Candida albicans in vitro. Furthermore, in vivo the substances could induce increased proliferation of phagocytes in spleen and bone marrow and migration of granulocytes to the peripheral blood. These effects indeed resulted in excellent protection of mice against the consequences of lethal infections with one predominantly M phi dependent and one predominantly granulocyte dependent pathogen, Listeria monocytogenes and C. albicans, respectively. Specific immune responses to sheep red blood cells (antibody production) and to listeria (DTH) were not affected by the polysaccharides. The possibility of clinical use is discussed.

From the nutrition medium of Arnica montana cell cultures two homogeneous polysaccharides, an acidic arabino-3,6-galactan-protein with mean Mr of 100,000 and a neutral fucogalactoxyloglucan with mean Mr of 22,500 have been isolated by DEAE-Sepharose CL-6B and Sephacryl S-400 column chromatography. Their structures were elucidated mainly by methylation analysis, partial acidic and enzymatic hydrolysis and 13C NMR spectroscopy. The fucogalactoxyloglucan shows a pronounced enhancement of phagocytosis in vivo. The arabino-3,6-galactan-protein displays a strong anticomplementary effect and stimulates macrophages to excrete the tumour necrosis factor (TNF alpha).

Hydrolysis of bamboo shoot cell walls with Driselase (a fungal enzyme preparation) gave xyloglucan and arabinoxylan oligosaccharides containing ferulic and p-coumaric acids, respectively. The structures of two oligosaccharides containing phenolic acids are here determined to be O-(4-O-trans-feruloyl-alpha-D-xylopyranosyl)-(1----6)-D-glucopy rano se and O-[5-O-(trans-p-coumaroyl(-alpha-L-arabinofuranosyl]-(1----3)-O-be ta-D-xylopyranosyl-(1----4)-D-xylopyranose, on the basis of n.m.r. spectroscopy, methylation analysis, and f.a.b.-m.s. The possible role of phenolic acid substituents in cell-wall architecture is discussed.

1. Polymers were solubilized from the cell walls of parenchyma from mature runner-bean pods with minimum degradation by successive extractions with cyclohexane-trans-1,2-diamine-NNN'N'-tetra-acetate (CDTA), Na2CO3 and KOH to leave the alpha-cellulose residue, which contained cross-linked pectic polysaccharides and Hyp-rich glycoproteins. These were solubilized with chlorite/acetic acid and cellulase. The polymers were fractionated by anion-exchange chromatography, and fractions were subjected to methylation analysis. 2. The pectic polysaccharides differed in their ease of extraction, and a small proportion were highly cross-linked. The bulk of the pectic polysaccharides solubilized by CDTA and Na2CO3 were less branched than those solubilized by KOH. There was good evidence that most of the pectic polysaccharides were not degraded during extraction. 3. The protein-containing fractions included Hyp-rich and Hyp-poor glycoproteins associated with easily extractable pectic polysaccharides, Hyp-rich glycoproteins solubilized with 4M-KOH+borate, the bulk of which were not associated with pectic polysaccharides, and highly cross-linked Hyp-rich glycoproteins. 4. Isodityrosine was not detected, suggesting that it does not have a (major) cross-linking role in these walls. Instead, it is suggested that phenolics, presumably linked to C-5 of 3,5-linked Araf residues of Hyp-rich glycoproteins, serve to cross-link some of the polymers. 5. There were two main types of xyloglucan, with different degrees of branching. The bulk of the less branched xyloglucans were solubilized by more-concentrated alkali. The anomeric configurations of the sugars in one of the highly branched xyloglucans were determined by 13C-n.m.r. spectroscopy. 6. The structural features of the cell-wall polymers and complexes are discussed in relation to the structure of the cell walls of parenchyma tissues.

A main acidic polysaccharide preparation was isolated from the leaves of Malva sylvestris L. var mauritiana Mill. It is composed of L-rhamnose, D-galactose, D-galacturonic acid, and D-glucuronic acid in the molar ratio of 22:6:22:11, and it contains 7.7% peptide. It was homogeneous by electrophoresis and gel chromatography, which gave a value of 11,000 as molecular weight. The structure of the polysaccharide component was elucidated by methylation analysis and partial hydrolysis. The substance showed considerable anticomplementary activity.

A method to determine rapidly the identities and proportions of the oligosaccharide repeating-units in plant cell-wall xyloglucans by 1D 1H-n.m.r. spectroscopy was developed. Six of the most commonly found xyloglucan oligosaccharide subunits (including three subunits that had not been fully characterized previously) were prepared by endo-(1----4)-beta-D-glucanase digestion of xyloglucans from various plant species. The oligosaccharides were reduced to the corresponding oligoglycosyl-alditols, purified, and characterized by glycosyl composition and linkage analysis, 1H-n.m.r. spectroscopy, and f.a.b.-mass spectrometry. Correlations between the 1H-n.m.r. spectra and the structures of the oligoglycosyl-alditols can be used to identify oligoglycosyl-alditols derived from xyloglucans of unknown structure. The identities and relative amounts of the oligosaccharide subunits of xyloglucans isolated from tamarind seed and rapeseed hulls were determined on this basis.

Only three gum exudates are permitted for pharmaceutical and food use by international regulatory authorities, viz. gum tragacanth (Asiatic Astragalus spp.), gum karaya (Sterculia spp.) and gum arabic [Acacia senegal (L.) Willd.], but a wide range of other tree exudates is used for a variety of uses in their countries of origin. This paper presents analytical data for the gum exudates from Atalaya hemiglauca, Cassine aethiopica, Combretum paniculatum, Sclerocarya birrea, and Pseudocedrela kotschyi. These gums may have local technological applications, but are not recommended for addition to foodstuffs.

alpha-Linked D-xylosyl side chains represent the typical feature common to all xyloglucans not shared by other cell wall polysaccharides. Since no easily available alpha-D-xyloxidase is known, advantage was taken of the conformational and configurational homologies between alpha-D-xylopyranose and alpha-D-glucopyranose to make an alpha-D-glucosidase-gold complex which was able to recognize alpha-D-xylosyl terminal residues of xyloglucans. This marker was used together with alpha-L-fucosidase gold complex for the double labeling on two different structural features of the same macromolecule in plant primary cell wall.

Polysaccharides were isolated from Dragon's Blood (DB), a red resinous secretion from the fruits of Daemonorops species and from pollens of Typha Augustata (TA) by alcohol precipitation followed by deaminoethylaminoethyl (DEAE) cellulose chromatography. The purified polysaccharides of DB and TA gave specific rotations of [alpha]D20 64 and [alpha]D20 of 80 and their molecular weights by membrane osmometry were approximately 25000 and approximately 30000 respectively. The effect of addition of the polysaccharides on coagulation of pooled normal human plasma was investigated. They were found to accelerate the recalcified plasma times at concentrations below 100 micrograms/ml while inhibiting them at higher concentrations. Their procoagulant properties were due to their effects on the activation of Hageman factor (XII) while the anticoagulant effect was mainly directed towards fibrinogen. Studies of the mechanism of the anticoagulant effect showed that addition of TA polysaccharide inhibited the rate of release of fibrinopeptides by thrombin and also the aggregation of fibrin monomers.

The 1 M-KOH extract from the depectinated cell walls of parenchymatous tissues of mature runner bean (Phaseolus coccineus) on neutralization, dialysis and concentration gave insoluble (hemicellulose A) and soluble (hemicellulose B) carbohydrate complexes in the weight ratio 2:1. Both fractions contained polysaccharide, protein and polyphenolic material. The structural features of the carbohydrates were examined by methylation analysis. Hemicellulose A contained mainly pectic arabinogalactan, with lesser amounts of arabinoxylan and glucan. Sequential fractionation of hemicellulose B by anion-exchange and hydroxyapatite chromatography gave a range of polysaccharide-protein-polyphenolic complexes. The main polysaccharides in these complexes were (acidic) arabinoxylans, galactans, arabinogalactans 1 and 2 and xyloglucans. The proteins contained small amounts of hydroxyproline, but were rich in aspartic acid and glutamic acid. Attempts to determine the nature of the polyphenolic material were unsuccessful. The structural features of the polysaccharide-protein-polyphenolic complexes are discussed in relation to the structure of the cell walls of parenchymatous tissues.

Hot-water extraction of defatted garlic-bulbs yielded a mixture of polysaccharides containing a D-galactan, a D-galacturonan, an L-arabinan, a D-glucan, and a D-fructan. A trace of L-rhamnose was also detected in the polysaccharide hydrolyzate. The pectic acid was partially removed by precipitation with aqueous calcium chloride; from the remaining polysaccharide mixture, a pure D-galactan containing 97.3% of D-galactose was isolated by fractional precipitation and repeated chromatography through a column of DEAE-cellulose. Methanolysis and hydrolysis of the permethylated D-galactan yielded 2,3,4,6-tetra-, 2,3,6-tri-, and 2,3,di-O-methyl-D-galactose in the molar proportions of 1:2:1. On periodate oxidation, the D-galactan reduced 1.18 molar equivalents of the oxidant per D-galactosyl residue, and liberated one molar equivalent of formic acid per 4.13 D-galactosyl residues. Smith degradation of the D-galactan was also conducted. From these results, a structure has been assigned to the repeating unit of the D-galactan.

The locations of the O-acetyl substituents on the major nonasaccharide repeating unit of the xyloglucan isolated from sycamore extracellular polysaccharides were determined by a combination of analytical methods, including f.a.b.-m.s. and 1H-n.m.r. spectroscopy. The O-2-linked-beta-D-galactosyl residue of the nonasaccharide was found to be the dominant site of O-acetyl substitution. Both mono-O-acetylated and di-O-acetylated beta-D-galactosyl residues were detected. The degree of O-acetylation of the beta-D-galactosyl residue, was estimated by 1H-n.m.r. spectroscopy to be 55-60% at O-6, 15-20% at O-4, and 20-25% at O-3. 1H-n.m.r. spectroscopy also indicated that approximately 50% of the beta-D-galactosyl residues are mono-O-acetylated, 25-30% are di-O-acetylated, and 20% are not acetylated.

A polysaccharide fraction from Lampteromyces japonicus contained a sugar (4%), which was identified as 3-O-methyl galactose by demethylation with boron trichloride, periodate oxidation of the methyl glycoside derivative and gas chromatography-mass spectrometry of the alditol acetate derivative.

From the water or alcaline-water extracts of Echinacea purpurea (L.) Moench and -angustifolia DC., Eupatorium cannabinum L. and -perfoliatum L., Chamomilla recutita (L.) (Rauscher), Calendula officinalis L., Baptisia tinctoria (L.) R.B., Achyrocline satureoides DC., Arnica montana L., Sabal serrulata Roem et Schult. and Eleutherococcus senticosus Maxim. polysaccharide fractions with molecular weights in the range of 25 000 to 500 000 and higher have been isolated, which, according to the granulocytes- and carbon clearance tests, showed significant immunostimulating activities. The isolated compounds belong to the group of water-soluble, acidic heteroglycanes. The linkages in the different polysaccharides do not represent a uniform structure type.

An aqueous alcoholic extract of fresh flowers of Yucca glauca Nutt. showed striking antitumor activity against B16 melanoma in mice. Systematic fractionation of the extract by means of solvent extraction and gel permeation chromatography led to separation of two galactose containing polysaccharide fractions with marked inhibitory activity against B16 melanoma in mice. The extraction, fractionation, purification and preliminary characterization of the active polysaccharide fractions are described. The materials showed no activity against L1210 or P388 leukemias in mice. The implications of the findings for searches for both natural and synthetic anticancer agents are discussed.

Characterization of the carbohydrates of pea (Pisum sativum) hulls, carrot and cabbage using both colorimetric and gas-liquid chromatographic techniques permitted a detailed investigation into the extent of digestion of differing types of fibre. These digestion studies were greatly aided by the development of a rapid bioassay employing starved adult cockerels. Total collection of undigested residues, uncontaminated by food spillage, could be made from trays placed under the cockerels. Chemical analysis showed that pea hulls consisted mainly of fibre with very little available carbohydrate present, whereas more than half of freeze-dried carrot and cabbage consisted of available carbohydrate (sucrose, glucose, fructose, starch) and consequently considerably less fibre was present. The fibre of carrot and cabbage was similarly composed of nearly equal amounts of neutral and acidic polysaccharides, whereas pea-hull fibre had four times as much neutral as acidic polysaccharides. The digestibility of total neutral polysaccharides from all three foodstuffs was extremely low. However, there appeared to be preferential digestion of polysaccharides composed of rhamnose, arabinose and galactose residues, all associated with pectic material, in contrast to the indigestibility of polysaccharides composed of fucose, xylose and glucose. Acidic polysaccharides were digested to a greater extent than neutral ones, and those of carrot and cabbage more so than pea hulls. The polysaccharides which were the most soluble were also the most digestible, but due to the arbitrariness of polysaccharide solubility, quantification of their total digestibility per se was considered not possible.

This paper illustrates post-embedding immunogold labelling of protein and polysaccharide molecules of plant cells. For EM studies, one is restricted (for most plant cells) to the post-embedding approach because the surrounding cell wall prevents access of antibodies (and secondary gold-tagged markers) to internal sites. The large size of many plant cells also does not lend itself to diffusional entry of antibodies. The molecules localized include seed storage proteins that are large and present in major quantities, a smaller less abundant, water soluble albumin, an oxygen-binding protein, components of the photosynthetic electron transport chain, and complex sugars from the cell wall. A range of preparative procedures and embedding plastics are used.

The structure of papaya polysaccharide II (PP II) isolated by Chandrasekaran et al. [Carbohydr. Res., 60 (1978) 105-115] has been investigated by methylation analysis of the carboxyl-reduced polymer and by partial hydrolysis of both the intact (arabinose, 31.0; rhamnose, 13.3; galactose, 42.6; glucuronic acid, 10.3; and 4-O-methylglucuronic acid, 2.8%), and carboxyl-reduced polymers. Methylation analysis of carboxyl-reduced PP II indicated a very highly branched structure in which approximately 39% of the galactopyranose units are disubstituted, 24% are monosubstituted, 20% are trisubstituted, and 17% are nonreducing end units. Methylation analysis of products of partial hydrolysis of both intact and carboxyl-reduced polymers indicated that the backbone of the polysaccharide is made up of galactosyl residues substituted at either O-3 or -6, that the principal aldobiouronic acid fragment is 6-O-(glucopyranosyluronic acid)galactose, that the rhamnosyl units are substituted at O-3 with either terminal arabinofuranosyl or galactopyranosyl groups, and that the rhamnosyl residues are themselves linked to glucuronic acid residues through O-4. From this information, a possible statistical fragment with six arabinofuranose and two galactopyranose nonreducing end-groups per 19 sugar units [five units in the main chain of (1----3)-linked galactopyranose units] is proposed.

The polysaccharide from Aloe vahombe (lilaceae) was submitted to partial hydrolyses by sulfuric, oxalic and phosphoric acid. Some oligosaccharides were isolated and investigated by chemical and 13C NMR spectroscopic methods. Their structure was determined. The results prove unambiguously that in the analyzed material the D-mannose is linked to the D-glucose by beta-1----4 linkages to form a carbohydrate heteropolymer.

The arborvitae or Thuja occidentale L., one of the Cupressaceae, has rarely been investigated until now. Several authors have demonstrated that allopathic extracts of this plant could be used as strong antiviral agents directed against plant and animal viruses. Polysaccharide fractions with molecular weights ranging between 20,000 and 1,000,000 and higher have been isolated from the aqueous alkaline extract of the herbal parts of T. occidentale L. by ethanol precipitation and fractionation by ultrafiltration. High molecular subfractions of Thujapolysaccharides (TPS) proved to be highly mitogenic on peripheral blood leukocytes. It was demonstrated by the alcalic immune phosphatase-antiphosphatase and Pappenheim staining methods that the mitogenic and cluster-forming activity of TPS cause T cell induction, in particular, of CD 4-positive T-helper/inducer cells as opposed to B cells. The CD-4+ T-helper/inducer cell induction is connected to an increased production of IL-2. The cluster-forming ability and mitogenity of TPS correlates well with the 3H-thymidine uptake and seems to be IL-1 and IFN-gamma dependent as could be shown by blocking the mitogenic effect using anti-IL-1- and anti-IFN antibodies. Whether it is possible to use these polysaccharide fractions as an adjuvant in the therapy of immune deficiency syndromes and cancer must now be further investigated.

A polysaccharide fraction of Saccharum officinarum was tested for its effects on carbohydrate and lipid metabolism in normal rats and those fed a high sugar diet. Feeding with the high sugar diet induced an elevation of serum glucose and significant accumulation of lipid peroxides in the serum and liver. The accumulation of lipid peroxides was inhibited by combined feeding with the polysaccharide fraction. Pathological examination showed that endothelial cell swelling in ascending aorta was found in one third of rats receiving the high sugar diet control but no pathological change was observed in all of the rats concurrently treated with the polysaccharide fraction.

We have investigated how the stereochemical situation of the dye binding negative side groups on polysaccharides influences metachromatic staining effects. We found that selective binding of toluidine blue (pH = 1.0) in the places of substituted (acidified) vicinal OH groups C2 and C3 resulted in the clearest metachromasia with the strongest anisotropy effects and with a bright clear green polarization color. On the other hand, selective toluidine blue binding in the places of C6 of the carbohydrate rings of the polysaccharide chains resulted in orthochromatic basophilia and lack of birefringence. After transformation of all OH side groups into negatively charged groups, metachromatic staining was found mixed with an orthochromatic hue and a birefringence effect weaker than in the case of staining on the vicinal OH groups only. The need of poststaining stabilization of the labile metachromatic staining reactions is emphasized.

A microbiological assay is described for determining gas produced by Clostridium perfringens (Veilon and Zuber) Holland from materials believed to be flatulent. A rationale for its use is given and analyzed. The fractionation and the results of the assay for several components of lima beans, Phaseolus lunatus L., and navy beans, P. vulgaris L., are given. In particular, a polysaccharide was isolated from dried P. vulgaris seeds. Only about one-seventh as much could be isolated from green P. lunatus seeds as from P. vulgaris seeds. The polysaccharide stimulates voluminous gas production by C. perfringens and meets all criteria of the rationale for flatulent sus pects. It has not been tested on humans; however, data from the microbiological assay correlate well with human results for green and dry lima beans and navy beans.