HF Hydrolysis Research Articles

Acid hydrolysis conditions for quantification of meningococcal X polysaccharide in a pentavalent vaccine using HPAEC-PAD/ESI-MS

A selective and sensitive method was evaluated for quantitation of meningococcal X (Men X) polysaccharide in pentavalent meningococcal A, C, W, Y and X conjugate vaccine using different acid hydrolysis conditions like HCl, TFA, HF, HF-TFA, and HF–HCl. High-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) using CarboPac PA10 column was used to identify the hydrolyzed products based on retention time and its comparison with monosaccharide standards. Complete release of glucosamine (GlcN) from Men X in monovalent bulk and pentavalent vaccine samples was achieved using HF hydrolysis at 80 °C for 2 h. The Men X HF-hydrolyzed polysaccharide to glucosamine along with the reference standard was identified using collision-induced dissociation (CID) electrospray mass spectroscopy and the MS/MS fragments of m/z 162, m/z 144 and m/z 84. Meningococcal polysaccharide concentration was determined with a correlation coefficient r2 >0.99 using polysaccharide reference standard. The serogroups A, W, and Y were converted to their monosaccharides units and quantified using this method however, milder acid hydrolysis 0.1 M HCl 80 °C 2 h for release of sialic acid for Men C polysaccharide was found to be more suitable. These methods will provide necessary tools and prove to be beneficial to laboratories developing new saccharide-based vaccine combinations.

Constructing a stable interface by 2,4,6-trimethoxyboroxine as an electrolyte additive for Li-rich layered oxide cathode under high voltage

• 2,4,6-Trimethoxyboroxine (TMBX) is proposed as an effective electrolyte additive for Li 1.2 Mn 0.54 Co 0.13 Ni 0.13 O 2 /Li batteries. • TMBX suppresses the hydrolysis reaction of LiPF 6 and the generation of HF. • The TMBX-derived film can improve charge transport and surface stability of cathode and anode. • The batteries with TMBX additive exhibit enhanced electrochemical performances. • The mechanisms for the HF scavenging and interface formation are elucidated. Li-rich layered oxides (LLO) feature high delivery capacity are considered the most promising cathodes for the next-generation high-energy–density battery systems. Unfortunately, the LLO cathode suffers from poor cycling stability and fast voltage fading because of structural failure and unstable electrode/electrolyte interphase. Herein, we propose 2,4,6-trimethoxyboroxine (TMBX) as an effective electrolyte additive for enhancing the specific capacity and service life of LLO cathode. Impressively, the LLO/Li cell with 1 wt% TMBX exhibits a specific capacity of 226.3 mAh/g with a considerable cycling retention of 96.1% at 0.5 C after 200 cycles, while the capacity of the baseline cell is only 133.8 mAh/g and the cycling retention is less than 60%. Even at 1 C, the cell using TMBX additive still sustains a capacity retention of as high as 91.1% after 400 cycles. The cell with excellent performance is attributed to TMBX additive which can form a uniform, robust, and boron-rich interface film on both the cathode and anode surfaces simultaneously. Structural characterization and mechanism analysis reveal that TMBX additive can effectively eliminate HF hydrolysis from LiPF 6 , alleviate the loss of transition metal ions, maintain the integrity of crystal structure and improve the interfacial charge transfer.

Fourier‐transform surface‐enhanced Raman spectroscopy (FT‐SERS) applied to the identification of natural dyes in textile fibers: an extractionless approach to the analysis

In the present study, an application of a silver colloid substrate in order to obtain Fourier‐transform surface‐enhanced Raman (FT‐SER) spectra of natural historical dyes is presented. In detail, we collected a spectral database from solutions of pure dyes and then we carried out extractionless both hydrolysis and non‐hydrolysis FT‐SERS analyses on wool fibers previously dyed in our laboratory and on ancient textiles. The term ‘extractionless’ refers to a method of SERS analysis applied directly on the fiber, thus avoiding the extraction of dyes from textile samples. The combination of a low‐energy source of radiation, as in the FT‐Raman technique, with SER spectroscopy can bring the important advantage of reducing the fluorescence typical of ancient samples and organic dyes. In some historical textile samples, for which SER spectra by use of visible excitation could not be obtained, the FT‐SER spectrum of an iron‐gall dye was recorded without hydrolysis, while, with an HF hydrolysis pre‐treatment on ancient fibers, madder, lac dye and brazilwood were clearly recognized. Copyright © 2014 John Wiley & Sons, Ltd.

Sample Treatment Considerations in the Analysis of Organic Colorants by Surface-Enhanced Raman Scattering

The introduction of surface-enhanced Raman spectroscopy (SERS) in the field of cultural heritage has significantly improved the analysis of the organic dyes and their complexes that have been used as textile dyes and pigments in paintings and other polychrome works of art since antiquity. Over the last five years, a number of different procedures have been developed by various research groups. In this Article, we evaluate the effect of pretreating samples by exposing them to hydrofluoric acid (HF) vapor prior to SERS analysis, a step designed to hydrolyze the dye-metal complexes and increase analyte adsorption on the nanosized metallic support, thus enhancing the SERS signal. Materials studied include pure colorants, commercial lake pigments, and fibers from dyed textiles, as well as actual aged samples, such as microscopic fragments of lakes on paper and ancient pigments and glazes from several works of art, covering a wide range of time, from the second century B.C. to the early 20th century. In each case, SERS spectra obtained with or without HF hydrolysis were critically evaluated. The pretreatment with HF vapor resulted in faster analysis and increased sensitivity in most cases, with the exception of dyed silk fibers, where silk protein hydrolyzates were found to interfere with SERS analysis. As a final point, a two-step procedure including SERS on untreated and treated samples is proposed as a standard approach: by analyzing a sample first without hydrolysis, and then, following removal of the colloid, upon HF treatment, the best and most reliable results for a great number of dyes and substrates are assured.

Phosphatidylglyceroylalkylamine, a novel phosphoglycolipid precursor in Deinococcus radiodurans

We report here the structure of a previously uncharacterized phospholipid in the radiation-resistant bacterium Deinococcus radiodurans. This phospholipid, designated lipid 4, was shown by chemical analysis, HF hydrolysis, and nuclear magnetic resonance spectroscopy to be phosphatidylglyceroylalkylamine. Lipid 4 thus contains the unusual lipid constituents glyceric acid and alkylamines, which have previously been identified in two complex phosphoglycolipids from this organism. By [32P]phosphate pulse-chase labeling techniques, lipid 4 was shown to be the precursor of the complex phosphoglycolipids alpha-galactosyl- and alpha-N-acetylglucosaminylphosphatidylglyceroylalkylamine. While phosphatidylglyceroylalkylamine is rapidly biosynthesized from Pi, its subsequent glycosylation occurs much more slowly. Therefore, we conclude that the final glycosylation step is the rate-limiting event in the biosynthesis of the complex phosphoglycolipids alpha-galactosyl- and alpha-N-acetylglucosaminyl-phosphatidylglyceroylalkylamine.

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

Coaggregation between Streptococcus sanguis H1 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 coaggregation-defective mutant of S. sanguis H1 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.

Lipophosphoglycan of Leishmania major that vaccinates against cutaneous leishmaniasis contains an alkylglycerophosphoinositol lipid anchor.

The major cell surface glycoconjugate of Leishmania major, a putative parasite receptor for macrophages, is a lipophosphoglycan containing 81.6% (wt/wt) carbohydrate, 17.0% (wt/wt) phosphate, and 1.4% (wt/wt) lipid. It has been purified to homogeneity by hydrophobic chromatography and consists of a polydisperse family of molecules with Mr 5000-40,000. It contains galactose, mannose, glucose, arabinose, glucosamine, and inositol in the molar ratio of 51:21:5:6:1:1. The lipophosphoglycan has a complex structure, consisting mainly of tri- and tetrasaccharide units linked by phosphodiester bonds, which are cleaved by HF hydrolysis. The phosphate groups are located on the 6-hydroxyl of both galactose and mannose residues. The lipophosphoglycan is anchored to the parasite surface by a 1-O-alkyl-sn-glycero-3-phosphoinositol moiety. This conclusion is supported by analysis of the products of nitrous acid deamination, HF hydrolysis, and Staphylococcus aureus phosphatidylinositol specific-phospholipase C treatment. The 24:0 and 26:0 alkyl chains accounted for 93% of the ether-linked fatty acids in the lipid anchor. The results are also consistent with a glycosidic linkage between the inositol and a non-N-acetylated glucosamine residue. The lipophosphoglycan membrane anchor shares limited structural homology with the glycosylphosphatidylinositol anchors of several eukaryotic proteins, indicating that this type of membrane anchor is not limited to proteins. Vaccination of mice with the purified L. major lipophosphoglycan in liposomes induced resistance against cutaneous leishmaniasis.

Occurrence of ribitol-containing lipoteichoic acid in Staphylococcus aureus H and its glycosylation.

A ribitol-containing lipoteichoic acid was obtained from the 20,000 x g supernatant fraction of Staphylococcus aureus H by extraction with Triton X-100 followed by fractionation on Sepharose 6B and DEAE-cellulose columns. The purified lipoteichoic acid was composed of phosphate, glycerol, glucose, glucosamine, ribitol, and fatty acids in a molar ratio of 1 : 0.9 : 0.06 : 0.03 : 0.09 : 0.07. Based on the structural analysis of fragments from alkali and HF hydrolysis, the lipoteichoic acid appears to consist of three moieties, namely a ribitol phosphate oligomer, poly(glycerol phosphate) which has about 30 glycerol phosphate units, and beta-glucosyl-beta-glucosyl(1 leads to 1)diacylglycerol. N-Acetylglucosamine was linked to the ribitol residues. The lipoteichoic acid serves as an acceptor of glycosyl moieties from UDP-glucose and UDP-N-acetylglucosamine in the enzyme reaction catalyzed by the membrane preparation. The rate of enzymatic glycosylation was increased by prior treatment of the lipoteichoic acid with N-acetyl-beta-D-glucosaminidase. The glycosylation seems to occur at the ribitol residues of the lipoteichoic acid.

Studies on the relationship between glycerophosphoglycolipids and lipoteichoic acids: IV. Trigalactosylglycerophospho-acylkojibiosyldiacyl-glycerol and related compounds from streptococcus lactis kiel 42172

Streptococcus lactis Kiel 42172 contains at least six unusually polar glycerophosphoglycolipids. The predominant one was composed of d-ga1actose, d-glucose, glycerol, acyi groups and phosphorus in a molar ratio of approx. 3: 2: 2: 3: 1. By analysis of the breakdown products of HF hydrolysis and Smith-degradation the structure was established to be [Gal p( α1 → 6)Gal p( α1 → 3)- sn-glycero(2 ← 1 αGal p)-1-phospho] → 6Glc p( α1 → 2), acyl → Glc p( α1 → 3)acyl 2Gro. By HF hydrolysis the other compounds were shown to be in the main also derivatives of Gro P→-6Glc( α1→ 2), acyl → 6Glc( α1 → 3)acyl 2Gro but they released as water-soluble glycosides Gal( α1 → 2)Gro, Gal( α1 → 3)Gro, Gal( α1 → 3)Gro(2 ← 1 αGal), Gal( α1 → 6)Gal( α1 → 3)Gro and Gal( α1 → 6)Gal( α1 → 6)Gal( α1 → 3)Gro(2 ← 1 αGal), respectively. In the lipid extract Glc( α1 → 2), acyl → 6Glc(α1 → 3)acyl 2Gro and Gro P → 6Glc( α1 →2), acyl → 6Glc(α1 → 3)acyl 2Gro were a1so observed. This set of compounds is proposed to constitute a biosynthetic series reflecting the individua1 steps in the synthesis of the lipoteichoic acid of Streptococcus lactis Kiel 42172 which is made up by the same lipid anchor and a non-classical poly(ga1abiosyl, ga1actosyl glycerophosphate)-chain (Koch H.U. and Fischer W. (1978) Biochemistry 17, 5275–5281).

Introduction of O-glycosidically linked mannose into proteins via mannosyl phosphoryl dolichol by microsomes from Fusarium solani f. pisi

Cutinase, a glycoprotein containing O-glycosidically linked carbohydrates, is induced in glucose-grown Fusarium solani f. pisi by cutin hydrolysate. Microsomal preparations from the induced cells catalyzed mannosyl transfer from GDP-mannose to glycolipid and glycoprotein fractions but not into oligosaccharide lipids. Maximal rates of mannosyl transfer into glycolipids and glycoproteins were obtained with 5 m m Mg 2+ and 10 m m Mn 2+, respectively. Mannosyl transfer into glycolipids and glycoproteins showed pH optima of 8.0 and 7.0, respectively, and both transfers showed an apparent K m of about 2 μ m for GDP-mannose. The mannosyl lipid was identified as β- d-mannosyl phosphoryl dolichol by thinlayer and ion-exchange chromatography, as well as by analyses of the products derived from it by acid and base treatments. The fungal microsomal preparation also catalyzed mannosyl transfer from GDP-mannose to exogenous dolichol phosphate. This transfer was stimulated maximally by 0.09% Triton X-100 and showed a pH optimum at pH 8.0. The apparent K m values for dolichol phosphate and GDP-mannose were 120 and 2.3 μ m, respectively. The product derived from exogenous dolichol phosphate was identified as β- d-mannosyl phosphoryl dolichol as indicated above. The endogenous mannosyl acceptor lipid from this fungus was isolated by DEAE-cellulose chromatography. Analysis of the p-nitrobenzoyl derivatives of the base hydrolysis products of this acceptor lipid by highperformance liquid chromatography showed that the major components of this dolichol were C 95 and C 100. The microsomal preparation also catalyzed the transfer of mannose from exogenous mannosyl phosphoryl dolichol to glycoproteins with a pH optimum of 7.5 and an apparent K m of 1.7 μ m. Analyses of the β-elimination products of the glycoproteins generated from both GDP-mannose and dolichol phosphoryl mannose showed that single mannosyl residues were transferred to hydroxyl groups of the endogenous proteins. Exogenous cutinase was not glycosylated even after denaturation, sulfitolysis, or removal of carbohydrates by HF hydrolysis. Sodium dodecyl sulfate electrophoresis indicated that cutinase and its possible precursors were among the in vitro glycosylation products. Bacitracin and amphomycin but not tunicamycin inhibited the mannosyl transfer reactions.