Capillary gas chromatographic analysis of alditol acetates of neutral and amino sugars in bacterial cell walls

Fox, Alvin; Morgan, Stephen L.; Hudson, Joseph R.; Zhu, Zhong Tao; Lau, Pauline Y.

doi:10.1016/s0021-9673(01)88260-1

Cited by 52 publications

(13 citation statements)

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“…HF was removed by four or five cycles of freeze-drying or evaporation (Speed-Vac; Savant Instruments, Inc., Hicksville, N.Y.), followed by rehydration with distilled deionized water. The rate and efficacy of hydrolysis were monitored by treating samples of purified polysaccharide with 48% HF for 30 min and 1, 4,8,24, 48, 72, 96, and 144 h in a time course experiment; HF was removed as described above.Isolation of S. sanguis Hi CIP and HF-released oligosaccharide. Coaggregation-inhibiting material released from S. sanguis Hi cell walls by both the autoclaving and the mutanolysin methods yielded purified polysaccharide by anionexchange chromatography on a fast protein liquid chromatography column (MonoQ HR 5/5; Pharmacia, Inc., Piscataway, N.J.) mounted on a 1090L high-performance liquid chromatograph (HPLC) (Hewlett-Packard Co., Palo Alto, Calif.).…”

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confidence: 99%

Isolation of a coaggregation-inhibiting cell wall polysaccharide from Streptococcus sanguis H1

Cassels

London

1989

J Bacteriol

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Coaggregation between Streptococcus sanguis HI and Capnocytophaga ochracea ATCC 33596 cells is mediated by a carbohydrate receptor on the former and an adhesin on the latter. Two methods were used to release the carbohydrate receptor from the gram-positive streptococcus, autoclaving and mutanolysin treatment. The polysaccharide released from the streptococcal cell wall by either treatment was purified by ion-exchange chromatography; this polysaccharide inhibited coaggregation when preincubated with the gram-negative capnocytophaga partner. After hydrolysis of the polysaccharide by hydrofluoric acid (HF), the major oligosaccharide of the polysaccharide was purified by high-performance liquid chromatography. By analysis of the HF hydrolysis of the polysaccharide and the purified oligosaccharide, this major oligosaccharide appeared to be the repeating unit of the polysaccharide, with minor components resulting from internal hydrolysis of the major oligosaccharide. Gas chromatography results showed that the oligomer was a hexasaccharide, consisting of rhamnose, galactose, and glucose, in the ratio of 2:3:1, respectively. By weight, the purified hexasaccharide was a fourfold-more-potent inhibitor of coaggregation than the native polysaccharide. Resistance to hydrolysis by sulfuric acid alone and susceptibility to hydrolysis by HF suggested that oligosaccharide chains of the polysaccharide are linked by phosphodiester bonds. Studies with a coaggregationdefective mutant of S. sanguis Hi revealed that the cell walls of the mutant contained neither the polysaccharide nor the hexasaccharide repeating unit. The purification of both a polysaccharide and its constituent hexasaccharide repeating unit, which both inhibited coaggregation, and the absence of this polysaccharide or hexasaccharide on a coaggregation-defective mutant strongly suggest that the hexasaccharide derived from the polysaccharide functions as the receptor for the adhesin from C. ochracea ATCC 33596.Most bacteria isolated from the human oral cavity possess the ability to participate in intergeneric coaggregation. Coaggregation is characterized by a highly specific binding between stable surface components found on two different bacterial types. Intergeneric coaggregation is thought to play an important role in the formation of dental plaque deposits (11; P. E. Kolenbrander, Crit. Rev. Microbiol., in press). Streptococcus sanguis is one of the earliest colonizers of the clean tooth surface and is found in significant numbers in dental plaque (4,26 coaggregation-inhibiting polysaccharide (CIP) antigen from S. sanguis 34 has been isolated and characterized (18). This polysaccharide consists of rhamnose, glucose, galactose, and N-acetylgalactosamine in a hexasaccharide repeating unit (17). The polysaccharide inhibits coaggregation, contains saccharide components that by themselves are effective inhibitors of the interaction, and is a major cell surface antigen (18).To characterize the carbohydrate receptor from a different coaggregating pair, S. sanguis Hi and ...

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confidence: 99%

Isolation of a coaggregation-inhibiting cell wall polysaccharide from Streptococcus sanguis H1

Cassels

London

1989

J Bacteriol

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“…Fox et al, 1983), and the derivative of MurA exhibited poor stability compared with the corresponding basic amino sugar derivatives (data not shown). The addition of N-methylimidazole as a catalyst helped to accelerate the acetylation of basic amino sugars, but it failed to produce an appreciable signal peak for the MurA derivative (Whiton et al, 1985).…”

Section: Derivatization Of Amino Sugarsmentioning

confidence: 96%

“…A substantial effort has been made during the past few decades to optimize the pretreatment procedure for GC-based quantification of amino sugars, which requires three major steps: acid hydrolysis, purification, and derivatization, with hydrolysis being the key step for releasing amino sugars from biopolymers. Besides the most frequently used hydrolysis protocol, namely 6 M hot hydrochloric acid (HCl; Zhang and Amelung, 1996), less destructive procedures involving either hot trifluoroacetic acid (TFA; Neeser and Schweizer, 1984) or sulfuric acid (H 2 SO 4 ; Fox et al, 1983) have been proposed for the simultaneous extraction of neutral and amino sugars. Different purification protocols, such as neutralization with a base solution (Zhang and Amelung, 1996), precipitation of excess acid (Cowie and Hedges, 1984;Neeser and Schweizer, 1984), and deionization of hydrolysate (Cowie and Hedges, 1984), to name a few, have been used to reduce the content of acid and salts that are known to interfere with amino sugar derivatization.…”

Section: R Zhu Et Al: Compound-specific Stable Carbon Isotopic Analmentioning

confidence: 99%

“…Different purification protocols, such as neutralization with a base solution (Zhang and Amelung, 1996), precipitation of excess acid (Cowie and Hedges, 1984;Neeser and Schweizer, 1984), and deionization of hydrolysate (Cowie and Hedges, 1984), to name a few, have been used to reduce the content of acid and salts that are known to interfere with amino sugar derivatization. Conversion of amino sugars for GC analysis has been achieved via derivatization into alditol acetates (AA; Fox et al, 1983), aldononitrile acetates (ANA; Guerrant and Moss, 1984), or O-methyloxime acetates (Neeser and Schweizer, 1984). However, to perform isotopic analysis of amino sugars at trace levels, a systematic evaluation of these various methods with regard to the product recovery and pretreatment reproducibility is necessary.…”

Section: R Zhu Et Al: Compound-specific Stable Carbon Isotopic Analmentioning

confidence: 99%

“…Amino sugar standards were transformed to alditol acetate (AA) or aldononitrile acetate (ANA) derivatives in triplicate following the methods of Fox et al (1983) or Guerrant and Moss (1984), respectively. After being converted to ANA derivatives, amino sugars extracted from the environmental samples were further purified with a self-packed silica gel column (0.5 g; Kieselgel, 0.06-0.2 mm; Carl Roth GmbH, Karlsruhe, Germany; cf.…”

Section: Derivatization and Purificationmentioning

confidence: 99%

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Optimizing sample pretreatment for compound-specific stable carbon isotopic analysis of amino sugars in marine sediment

et al. 2014

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Abstract. Amino sugars are quantitatively significant constituents of soil and marine sediment, but their sources and turnover in environmental samples remain poorly understood. The stable carbon isotopic composition of amino sugars can provide information on the lifestyles of their source organisms and can be monitored during incubations with labeled substrates to estimate the turnover rates of microbial populations. However, until now, such investigation has been carried out only with soil samples, partly because of the much lower abundance of amino sugars in marine environments. We therefore optimized a procedure for compoundspecific isotopic analysis of amino sugars in marine sediment, employing gas chromatography-isotope ratio mass spectrometry. The whole procedure consisted of hydrolysis, neutralization, enrichment, and derivatization of amino sugars. Except for the derivatization step, the protocol introduced negligible isotopic fractionation, and the minimum requirement of amino sugar for isotopic analysis was 20 ng, i.e., equivalent to ∼ 8 ng of amino sugar carbon. Compoundspecific stable carbon isotopic analysis of amino sugars obtained from marine sediment extracts indicated that glucosamine and galactosamine were mainly derived from organic detritus, whereas muramic acid showed isotopic imprints from indigenous bacterial activities. The δ 13 C analysis of amino sugars provides a valuable addition to the biomarker-based characterization of microbial metabolism in the deep marine biosphere, which so far has been lipid oriented and biased towards the detection of archaeal signals.

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Experimental Uveitis

Fox¹

1984

Arch Ophthalmol

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Capillary gas chromatographic analysis of alditol acetates of neutral and amino sugars in bacterial cell walls

Cited by 52 publications

References 28 publications

Isolation of a coaggregation-inhibiting cell wall polysaccharide from Streptococcus sanguis H1

Isolation of a coaggregation-inhibiting cell wall polysaccharide from Streptococcus sanguis H1

Optimizing sample pretreatment for compound-specific stable carbon isotopic analysis of amino sugars in marine sediment

Experimental Uveitis

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