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Purpose To study the metagenomics of the microbes isolated from the canaliculus of patients with infective canaliculitis. Methods A prospective study was performed on five consecutive canalicular samples obtained for the metagenomic analysis from the patients with infective canaliculitis who underwent non-incisional canalicular curettage at a tertiary care Dacryology service. The canalicular concretions were collected intraoperatively soon after a canalicular curettage and immediately transported on ice to the laboratory. Following DNA extraction and library preparation, a whole shotgun metagenome sequencing was performed on the Illumina™ platform. The downstream processing and bioinformatics of the samples were performed using multiple software packaged in SqueezeMeta™ pipeline or MG-RAST™ pipeline. Results The taxonomic hit distribution across the samples showed that bacteria were the most common isolates (mean—80.5%), followed by viruses (mean—0.74%), and archaea (0.01%). The five major phyla identified across the samples of infective canaliculitis were, Fusobacteria, Bacteroidetes, Proteobacteria, Actinobacteria, and Firmicutes. The prevalent organisms include Fusobacterium nucelatum, Fusobacterium periodonticum, Parvimonas micra, Prevotella oris, Selonomonas noxia, Pseudopropionobacterium propoinicum, Campylobacter showae, and Streptococcus anginosus, amongst few others. Actinomycetes israelii was noted in all the samples, though it was not the most abundant. The microbial gene mapping and protein prediction demonstrated proteins with known functions to range from 69.91% to 87.09% across the samples. The functional subsystem profiling demonstrated genes associated with carbohydrate, amino acid, and co-enzyme transport and metabolism, cell wall or cell membrane biogenesis, energy production and conversion, transcription, translation, and cellular communications. Conclusion This is the first whole metagenome sequencing of infective canaliculitis. Infected canaliculi harbor diverse microbial communities, including bacteria, viruses, and archaea. Functional analysis has provided newer insights into the ecosystem dynamics and strategies of microbial communities.

HVPE is an excellent and often overlooked method for obtaining objective and meaningful information about cell-wall "building blocks" and their metabolic precursors. It provides not only a means of analysis of known compounds but also an insight into the charge and/or mass of any unfamiliar compounds that may be encountered. It can be used preparatively or analytically. It can achieve either "class separations" (e.g. delivering all hexose monophosphates into a single pool) or the resolution of different compounds within a given class (e.g. ADP-Glc from UDP-Glc; or GlcA from GalA). All information from HVPE about charge and mass can be obtained on minute traces of analytes, especially those that have been radiolabelled, e.g. by in-vivo feeding of a (3)H- or (14)C-labelled precursor. HVPE does not usually damage the substance under investigation (unless staining is used), so samples of interest can be eluted intact from the paper ready for further analysis. Although HVPE is a technique that has been available for several decades, recently it has tended to be sidelined, possibly because the apparatus is not widely available. Interested scientists are invited to contact the author about the possibility of accessing the Edinburgh apparatus.

Effect of paper mill effluents on Phaseolus aureus Roxb was studied. The effluent significantly inhibited germination of root and shoot length. The bio-chemical injury does not appear spontaneously but with the increase in effluent treatment there is reduction in observed biochemical parameters (chlorophyll, protein, amino acid, nuclic acids and carbohydrate) which are negatively correlated. The shoots of the seedlings were found to be resistant; whereas roots of the seedlings were susceptible to paper mill effluent treatment.

Inspired by dried blood spots (DBS), “dried spots of herbal medicines” (DSHM) concept was proposed here. In response to this superior sampling means, a new platform integrating dried spot, serially coupled reversed phase liquid chromatography and hydrophilic interaction liquid chromatography (RPLC-HILIC), and tandem mass spectrometry (MS/MS), was configured for directly, comprehensively chemical profiling of HM-derived liquid matrices. As an important original source of Cistanches Herba (Chinese name: Roucongrong) that is a well-known tonic HM, Cistanche sinensis (Csi) was employed to illustrate and validate the applicability. Dried spots (I.D. 3.0 mm) were prepared by loading 2 μL aliquots of Csi extract onto filter paper. Each dried spot was packed into an in-line filter holder (I.D. 3.0 mm × 4.0 mm) and then inserted behind the auto-sampler of a well-defined instrumentation named RPLC-HILIC–MS/MS. Hybrid ion trap-time of flight MS and hybrid triple quadrupole-linear ion trap MS were deployed in combination for the in-depth MS/MS data acquisition of diverse chemical families, such as phenylethanoid glycosides, lignans, iridoids, amino acids, and so forth. To assist chemical profiling, an in-house chemical library was built by collecting as much prior knowledge as possible. A total of 88 components were detected and tentatively annotated in Csi by matching their multi-stage MS spectra with those of authentic standards and literature data. Collectively, DSHM carried all merits of DBS, and the integrative dried spot-RPLC-HILIC-MS/MS platform was a promising analytical tool for direct chemical analysis and rapid quality evaluation of HMs, in particular those traditional Chinese medicine injections.

For investigations on the biological functions of oligosaccharides and peptidomimetics, new asparagine-based mono- and disaccharides containing glycopeptides were prepared in solution. The applicability of two common peptide coupling reagents, using an orthogonal Fmoc/t-Bu strategy along with acetyl protecting groups for the carbohydrate moiety, was studied. Thus, the prepared libraries of glycopeptides were designed as model systems of cell surfaces for future investigations by combined preparative mass spectroscopy and scanning tunneling microscopy (STM) using soft-landing electrospray beam deposition (ES-IBD), on metal surfaces.

Age can significantly affect human physiology and disease risk. Recent studies have shown that age may affect the composition and function of the gut microbiota, but the underlying mechanisms remain largely unknown. Non-human primates are an ideal model for uncovering how age shapes the gut microbiota, as their microbial composition is highly similar to that of humans and is not easily affected by confounding factors. Here, using the 16S rRNA and metagenomic sequencing methods, we characterized the microbial phenotypes of 16 female cynomolgus macaques from three age groups (young, adult and old). Our findings revealed significant differences in microbial composition among the three groups. With increased age, the relative abundances of Veillonellaceae, Coriobacteriaceae and Succinivibrionaceae were significantly increased, Ruminococcaceae and Rikenellaceae were significantly decreased at the family level. Functional enrichment showed that genes that differed among the three groups were mainly involved in arginine biosynthesis, purine metabolism and microbial polysaccharides metabolism. Moreover, CAZymes corresponding to polysaccharide degrading activities were also observed among the three groups. In conclusion, we characterized the composition and function of the gut microbiota at different ages, and our findings provide a new entry point for understanding the effects of age on the human body.

Highlight Changing leaf to air vapour pressure deficit (VPD) modifies primary metabolism of plants as demonstrated by excess starch synthesis and reduced amino acid, chlorophyll, and nitrogen concentrations in leaves under low VPD., Air humidity indicated as vapour pressure deficit (VPD) is directly related to transpiration and stomatal function of plants. We studied the effects of VPD and nitrogen (N) supply on leaf metabolites, plant growth, and mineral nutrition with young micropropagated silver birches (Betula pendula Roth.) in a growth chamber experiment. Plants that were grown under low VPD for 26 d had higher biomass, larger stem diameter, more leaves, fewer fallen leaves, and larger total leaf area than plants that were grown under high VPD. Initially, low VPD increased height growth rate and stomatal conductance; however, the effect was transient and the differences between low and high VPD plants became smaller with time. Metabolic adjustment to low VPD reflected N deficiency. The concentrations of N, iron, chlorophyll, amino acids, and soluble carbohydrates were lower and the levels of starch, quercetin glycosides, and raffinose were higher in the leaves that had developed under low VPD compared with high VPD. Additional N supply did not fully overcome the negative effect of low VPD on nutrient status but it diminished the effects of low VPD on leaf metabolism. Thus, with high N supply, the glutamine to glutamate ratio and starch production under low VPD became comparable with the levels under high VPD. The present study demonstrates that low VPD affects carbon and nutrient homeostasis and modifies N allocation of plants.

Urinary metabolic studies by means of high-voltage paper electrophoresis (HVPE) and some qualitative chemical tests were performed in 40 patients in a family with hereditary macular degeneration and in 40 nonaffected members of the same family. No biochemical abnormalities were observed in the examined urine samples. The results are discussed with regard to earlier published reports of aminoaciduria in hereditary macular degeneration.

Metabolic changes in Arabidopsis induced by periods of elevated heat and drought stress under ambient and elevated CO2, are dynamic and specific to different classes of molecules., As a consequence of global change processes, plants will increasingly be challenged by extreme climatic events, against a background of elevated atmospheric CO2. We analysed responses of Arabidopsis thaliana to periods of a combination of elevated heat and water deficit at ambient and elevated CO2 in order to gain mechanistic insights regarding changes in primary metabolism. Metabolic changes induced by extremes of climate are dynamic and specific to different classes of molecules. Concentrations of soluble sugars and amino acids increased transiently after short (4-d) exposure to heat and drought, and readjusted to control levels under prolonged (8-d) stress. In contrast, fatty acids showed persistent changes during the stress period. Elevated CO2 reduced the impact of stress on sugar and amino acid metabolism, but not on fatty acids. Integrating metabolite data with transcriptome results revealed that some of the metabolic changes were regulated at the transcriptional level. Multivariate analyses grouped metabolites on the basis of stress exposure time, indicating specificity in metabolic responses to short and prolonged stress. Taken together, the results indicate that dynamic metabolic reprograming plays an important role in plant acclimation to climatic extremes. The extent of such metabolic adjustments is less under high CO2, further pointing towards the role of high CO2 in stress mitigation.

New aromatic glycoconjugate building blocks based on the trifunctional 3-aminomethyl-5-aminobenzoic acid backbone and sugars linked to the backbone by a malonyl moiety were prepared via peptide coupling. The orthogonally protected glycoconjugates, bearing an acetyl-protected glycoside, were converted into their corresponding acids which are suitable building blocks for combinatorial glycopeptide synthesis.

High Resolution Magic Angle Spinning (HR-MAS) is an NMR technique that can be applied to semi-solid samples. Flavedo, albedo, pulp, seeds, and oil gland content of lemon and citron were studied through HR-MAS NMR spectroscopy, which was used directly on intact tissue specimens without any physicochemical manipulation. HR-MAS NMR proved to be a very suitable technique for detecting terpenes, sugars, organic acids, aminoacids and osmolites. It is valuable in observing changes in sugars, principal organic acids (mainly citric and malic) and ethanol contents of pulp specimens and this strongly point to its use to follow fruit ripening, or commercial assessment of fruit maturity. HR-MAS NMR was also used to derive the molar percentage of fatty acid components of lipids in seeds, which can change depending on the Citrus species and varieties. Finally, this technique was employed to elucidate the metabolic profile of mold flavedo.

Summary Four glycoconjugate building blocks for the construction of combinatorial PNA like glycopeptide libraries were prepared in 75–79% yield by condensing tert-butyl N-[2-(N-9-fluorenylmethoxycarbonylamino)ethyl]glycinate (AEG) 5 with 3-oxo-3-(2,3,4,6-tetra-O-acetyl-β-D-glucopyranosylamino)- (6a), 3-oxo-3-(β-D-galactopyranosylamino)- (6b), 3-oxo-3-(2-acetamido-2-deoxy-3,4,6-tetra-O-acetyl-β-D-glucopyranosylamino)- (6c) and 3-oxo-3-(2-acetamido-2-deoxy-3,4,6-tetra-O-acetyl-β-D-galactopyranosylamino)propanoic acid (6d), respectively. The resulting AEG glycoconjugates 1a–d were converted into the corresponding free acids 2a–d in 97–98% yield by treatment with aqueous formic acid. The Fmoc group of compound 1c was removed and the intermediate amine 9 was condensed with 2a to afford the corresponding glycosylated AEG dipeptide 4 in 58% yield. All glycoconjugate building blocks showed the presence of cis and trans rotamers. Compounds 1a, 1b and 4 were subjected to temperature dependent 1H NMR spectroscopy in order to determine the coalescence temperature which resulted in calculated rotation barriers of 17.9–18.3 kcal/mol for the rotamers.

The CCP4 template-restraint library defines restraints for biopolymers, their modifications and ligands that are used in macromolecular structure refinement. JLigand is a graphical editor for generating descriptions of new ligands and covalent linkages., Biological macromolecules are polymers and therefore the restraints for macromolecular refinement can be subdivided into two sets: restraints that are applied to atoms that all belong to the same monomer and restraints that are associated with the covalent bonds between monomers. The CCP4 template-restraint library contains three types of data entries defining template restraints: descriptions of monomers and their modifications, both used for intramonomer restraints, and descriptions of links for intermonomer restraints. The library provides generic descriptions of modifications and links for protein, DNA and RNA chains, and for some post-translational modifications including glycosylation. Structure-specific template restraints can be defined in a user’s additional restraint library. Here, JLigand, a new CCP4 graphical interface to LibCheck and REFMAC that has been developed to manage the user’s library and generate new monomer entries is described, as well as new entries for links and associated modifications.

To investigate how N-fertilization affects the growth, carbon and nitrogen (N) physiology, and wood properties of poplars with contrasting growth characteristics, slow-growing (Populus popularis, Pp) and fast-growing (P. alba×P. glandulosa, Pg) poplar saplings were exposed to different N levels. Above-ground biomass, leaf area, photosynthetic rates (A), instantaneous photosynthetic nitrogen use efficiency (PNUE (i)), chlorophyll and foliar sugar concentrations were higher in Pg than in Pp. Foliar nitrate reductase (NR) activities and root glutamate synthase (GOGAT) activities were higher in Pg than in Pp as were the N amount and NUE of new shoots. Lignin contents and calorific values of Pg wood were less than that of Pp wood. N-fertilization reduced root biomass of Pg more than of Pp, but increased leaf biomass, leaf area, A, and PNUE(i) of Pg more than of Pp. Among 13 genes involved in the transport of ammonium or nitrate or in N assimilation, transcripts showed more pronounced changes to N-fertilization in Pg than in Pp. Increases in NR activities and N contents due to N-fertilization were larger in Pg than in Pp. In both species, N-fertilization resulted in lower calorific values as well as shorter and wider vessel elements/fibres. These results suggest that growth, carbon and N physiology, and wood properties are more sensitive to increasing N availability in fast-growing poplars than in slow-growing ones, which is probably due to prioritized resource allocation to the leaves and accelerated N physiological processes in fast-growing poplars under higher N levels. peerReviewed

Phloem is a central conduit for the distribution of photoassimilate, nutrients, and signals among plant organs. A revised technique was used to collect phloem sap from small woody plants in order to assess changes in composition induced by water deficit and flooding. Bled phloem sap delta(13)C and sugar concentrations were compared to delta(13)C of bulk material, soluble carbon extracts, and the neutral sugar fraction from leaves. Amino acid composition and inorganic ions of the phloem sap was also analysed. Quantitative, systematic changes were detected in phloem sap composition and delta(13)C in response to altered water availability. Phloem sap delta(13)C was more sensitive to changes of water availability than the delta(13)C of bulk leaf, the soluble carbon fraction, and the neutral soluble fraction of leaves. Changes in water availability also resulted in significant changes in phloem sugar (sucrose and raffinose), inorganic nutrient (potassium), and amino acid (phenylalanine) concentrations with important implications for the maintenance of phloem function and biomass partitioning. The differences in carbohydrate and amino acid composition as well as the delta(13)C in the phloem, along with a new model system for phloem research, offer an improved understanding of the phloem-mediated signal, nutrient, and photoassimilate transduction in relation to water availability.

Intestinal and fecal contents were collected from germfree rats fed a chemically defined diet. The contents, considered to consist mainly of the epithelial mucin of the gastrointestinal tract, were separately extracted with water followed by dialysis and freezedrying. The products obtained had a similar composition, corresponding to substances of glyco-protein nature. The carbohydrate part, constituting about 80 per cent of the material, contained galactose, mannose, fucose, arabinose, xylose, N-acetylglucosamine, N-acetyl-galactosamine and sialic acid. In the peptide moiety serine, threonine, aspartic acid and glutamic acid were the major amino acids. Gel chromatographic experiments indicated, chemical as well as physical heterogeneity of the extracts.

The effect of salicylic acid (SA) on the metabolic profile of Catharanthus roseus suspension cells throughout a time course (0, 6, 12, 24, 48 and 72 h after treatment) was investigated using NMR spectroscopy and multivariate data analysis. When compared to control cell lines, SA-treated cells showed a high level of sugars (glucose and sucrose) up to 48 h after treatment, followed by a dynamic change in amino acids, phenylpropanoids, and tryptamine. Additionally, one compound--2,5-dihydroxybenzoic-5-O-glucoside--was detected solely in SA-treated cells.

Autophagy is a pathway in which a cell degrades part of its cytoplasm in vacuoles or lysosomes. To identify the physiological functions of autophagy in plants, we disrupted ATG5, an autophagy-related gene, in Physcomitrella, and confirmed that atg5 mutants are deficient in the process of autophagy. On carbon or nitrogen starvation medium, atg5 colonies turned yellow earlier than the wild-type (WT) colonies, showing that Physcomitrella atg5 mutants, like yeast and Arabidopsis, are sensitive to nutrient starvation. In the dark, even under nutrient-sufficient conditions, colonies turned yellow and the net degradation of chlorophyll and Rubisco protein occurred together with the upregulation of several senescence-associated genes. Yellowing reactions were inhibited by the protein synthesis inhibitor cycloheximide, suggesting that protonemal colonies undergo dark-induced senescence like the green leaves of higher plants. Such senescence responses in the dark occurred earlier in atg5 colonies than WT colonies. The sugar content was almost the same between WT and atg5 colonies, indicating that the early-senescence phenotype of atg5 is not explained by sugar deficiency. However, the levels of 7 amino acids showed significantly different alteration between atg5 and WT in the dark: 6 amino acids, particularly arginine and alanine, were much more deficient in the atg5 mutants, irrespective of the early degradation of Rubisco protein. On nutrient-sufficient medium supplemented with casamino acids, the early-senescence phenotype was slightly moderated. We propose that the early-senescence phenotype in atg5 mutants is partly explained by amino acid imbalance because of the lack of cytoplasmic degradation by autophagy in Physcomitrella.

Summary A long-term in vitro culture system of intact grape berries was developed which can serve to study the response of berry composition to various trophic factors, shown by sugar regulation of anthocyanin accumulation., Grape berry development and ripening are under complex regulation by the nutrients, hormones, and environment cues sensed by the berry. However, the biochemical and molecular mechanisms underlying these types of regulation are poorly understood. A simplified but realistic model system that enables fruit growth conditions to be modulated easily will facilitate the deciphering of these mechanisms. Here, an in vitro culture system of intact detached grape berries was developed by coupling the production of greenhouse fruiting-cuttings and in vitro organ culture techniques. 13C and 15N labelling experiments showed that this system enables the intact detached berries actively to absorb and utilize carbon and nitrogen from the culture medium. It was further used to study the effects of sugars on anthocyanin accumulation. A sucrose concentration >2% could induce anthocyanin synthesis in the absence of additional exogenous abscisic acid. The higher the sucrose concentration, the earlier was the induction of anthocyanin accumulation. Glucose, fructose, and sucrose increased anthocyanin accumulation, with glucose and fructose being more effective than sucrose. This increase was not due to an increase in its precursor level, since the phenylalanine content was decreased by a high sugar supply. Instead, genome-wide transcriptome analysis suggests that the sugar-induced enhancement of anthocyanin accumulation results from altered expression of regulatory and structural genes (especially UDP-glucose:anthocyanidin 3-O-glucosyltransferase), together with massive reprogramming in signalling transduction pathways. This in vitro system may serve to study the response of berry composition to nutrient factors and hormones, and their interaction with environmental factors (e.g. light and temperature), which can all be finely tuned and controlled.

Crude antigen extracted from mycelium of Trichophyton mentagrophytes var. erinacei by means of phenol and ethylene glycol was studied for sugar and amino acid constituents using DEAE-Sephadex A-50 column chromatography for fractionation. The results revealed that the crude antigen contains 13 sugars and 15 amino acids. It is presumed that the antigenicity is based on glycopeptide fractions.

The minor salivary glands contribute to the composition of whole saliva, but little information has been available about their chemical constituents. Pilocarpine-stimulated labial and palatine secretion from 4 human subjects was investigated by paper and disc electrophoresis, immunochemical analysis, and for content of carbohydrates, amino acids, lipids, hexuronic acids and sulphate. No significant differences were noted between the labial and palatine secretions by any of the methods employed. The minor gland secretions appeared to consist mainly of mucosubstances, possibly with blood group specificity. In addition, three water-soluble components with the characteristics of albumin, alpha-amylase and secretory IgA were seen. The minor gland secretions had an amino acid profile different from those of the major salivary glands and contained higher proportions of carbohydrate. Only one lipid component, with the characteristics of a polar lipid, was seen. Hexuronic acids were not detected in either secretion, whereas both contained sulphate. It would appear that the minor mucous glands contribute to the content of mucosubstances in whole saliva, whereas their content of water-soluble material is negligible in this respect.

Human C3, the third component of human complement, contained mannose and N-acetylglucosamine as sugar components. The sugar chains were liberated from the polypeptide chains by hydrazinolysis, and the free amino groups were N-acetylated. The reducing end residues of the sugar chains thus obtained were tagged with 2-aminopyridine, and the pyridylamino (PA-) derivatives of sugar chains were separated by high-performance liquid chromatography. The structures of purified PA-sugar chains were analyzed by a combination of stepwise exoglycosidase digestions, size determination by paper electrophoresis, methylation analysis, Smith degradation, and partial acetolysis. These results showed that C3 contained two high-mannose type sugar chains ranging from Man5GlcNAc2 to Man9GlcNAc2. Analyses of the sugar chains of alpha- and beta-chains of C3 indicated that the alpha-chain contained mainly Man8GlcNAc2 and Man9GlcNAc2, while the beta-chain contained mainly Man5GlcNAc2 and Man6GlcNAc2.

An abnormal protein with similar antigenic properties to Fc fragments of IgG, was found in the serum and urine of an eleventh case of heavy (Hgamma1) chain disease (Yok). This protein was purified with ammonium sulfate precipitation and by column chromatography of DEAE cellulose, CM cellulose and Sephadex G-200. The purity of the protein obtained was 98.5%. It was crystallized easily, forming thin hexagonal plates of various sizes. The chemical compositions and physical properties of the protein including viscosity, partial specific volume, diffusion constant, sedimentation constant, frictional ratio, extinction coefficient and iso-ionic point are reported.

A small molecular weight structural glycopeptide was solubilized after collagenase digestion of the connective tissue capsule surrounding the 5-day sponge-implant of the rat. The major amino acids are one residue each of aspartic and glutamic acids, proline, hydroxyproline and alanine and two residues of glycine, and the carbohydrates are one residue each of glucose, xylose and hexosamine and two residues of mannose. The sum of the amino acid and carbohydrate residues gives a molecular weight of 1635. Dansylation of the glycopeptide produces a single strongly fluorescent yellow-orange amino-terminal spot, not positively identified. The solubilization of the granuloma glycopeptide by collagenase and its composition are suggestive of its association with an immature form of collagen in early granulation tissue.

Sugar and aminoacids were investigated in sterile distilled water after Pinus pinea seeds had been soaked in it during 24 hours at 37 degrees C. Germination viability decreased with the ageing of the seeds and was accompanied by sugar and aminoacids increase in the exudate. The sugar content from seeds with loss of germinability was about 50 to 80 times greater than that from seeds with good germinability. Paper chromatography showed that there was an increase in mono, di, oligosaccharides and amino-acids in seeds without germinability, but these substances were only found as trace quantities in viable seeds. The methods already described by TKAYANAGI and MURAKAMI to determine germinability could be applied with some modifications to Pinus pinea seeds. The presence of sugar in the exudate could be detected by urine sugar test after 24 hours at 37 degrees C. It was necessary to concentrate the exudate till a final volume of 2 ml.

Glycopeptides (Gp I and Gp II) were prepared from the proline-rich glycoprotein of human parotid saliva by extensive papain-pronase digestion, Sephadex G-25 gel filtration, and DE-52 cellulose chromatography. Their carbohydrate units contained N-acetylgluco-samine, mannose, fucose, galactose and sialic acid. Based upon minimal mol wt calculations, the salivary glycoprotein contained four heteropolysaccharide units. One glycopeptide (Gp I) was homogeneous by N-terminal analysis and descending paper chromatography. Using dansyl-Edman degradation and partial acid hydrolysis coupled with dansylation, the peptide sequence of Gp I was found to be GlyGlyProAsn (CHO)Gln. Alkaline borohydride treatment of Gp I at 80 °C verified a glycosylamine bond between asparagine and N-acetylglucosamine.

Lipopolysaccharides (LPSs) isolated from phase I and phase II Coxiella burnetii (LPS I and LPS II, respectively) were analyzed for chemical compositions, molecular heterogeneity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and immunological properties. The yields of crude phenol-water extracts from phase I cells were roughly three to six times higher than those from phase II cells. Purification of LPSs by ultracentrifugation gave similar yields for both LPS I and LPS II. Purified LPS I and LPS II contained roughly 0.8 and 0.6% protein, respectively. The fatty acid constituents of the LPSs were different in composition and content, with branched-chain fatty acids representing about 15% of the total. beta-Hydroxymyristic acid was not detected in either LPS I or LPS II. A thiobarbituric acid-periodate-positive compound was evident in the LPSs; however, this component was not identified as 3-deoxy-D-mannooctulosonic acid by gas and paper chromatographies. LPS II contained D-mannose, D-glucose, D-glyceromannoheptose, glucosamine, ethanolamine, 3-deoxy-D-mannooctulosonic acid-like material, phosphate, and fatty acids. LPS I contained the unique disaccharide galactosaminuronyl glucosamine and nine unidentified components in addition to the components of LPS II. The hydrophobic, putative lipid A fraction of LPS I and LPS II contained the above constituents, but the hydrophilic fraction was devoid of ethanolamine. The LPS I disaccharide galactosaminuronyl glucosamine was found in both fractions of the acetic acid hydrolysates. Analysis of LPSs by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by silver staining indicated that LPS II was composed of only one band, whereas LPS I consisted of six or more bands with irregular spacing. Ouchterlony immunodiffusion tests demonstrated that LPS I reacted with phase I but not with phase II whole-cell hyperimmune antibody, and LPS II reacted neither with phase I nor phase II hyperimmune antibody. From these results, it was concluded that the chemical structures of LPSs from C. burnetii were different from those of the LPSs of gram-negative bacteria; however, the LPS structural variation in C. burnetii may be similar to the smooth-to-rough mutational variation of saccharide chain length in gram-negative bacteria.

Chondroitin sulfite peptidoglycans were prepared by the pronase digestion of cartilages from human knee joints, porcine tracheae, and shark crania. The beta-elimination and sulfite addition reaction of these peptidoglycans yielded cysteic acid-containing peptides, and the amino acid sequences of some of them were determined. The finding that human and porcine peptidoglycans have the same sequence in the linkage region as that of the bovine peptidoglycan reported previously suggested that there is a common structural feature in the core proteins of these mammalian cartilage proteoglycans. The results also support the view that -Ser-Gly- is the minimum requisite for the formation of the xylosylserine linkage of proteochondroitin sulfates.

Fractional molar ratios of serine, threonine and aspartic acid to neutral sugars in the purified bovine vitreous humor hyaluronate, and a 4–5-fold increase in the percentage of these amino acids and the absence of sugar alditols in hyaluronate reduced with NaBH 4 PdCl 2 after alkali treatment indicated the absence of a carbohydrate—protein linkage. Gel filtration behavior, a decrease in intrinsic viscosity of reduced hyaluronate to about one-half and a significant decrease in its specific rotation suggested that the two antiparallel chains of the hyaluronate double helix may come apart upo reduction. The vitreous humor hyaluronate contained 109.2 ppm of “bound” silicon. It is suggested that the bound silicon may bridge the two antiparallel chains through the neutral sugars and/or through the hydroxyl group of the uronic acid moiety.