Amphiphilic β‐cyclodextrin‐based star‐like block copolymer unimolecular micelles for facile in situ preparation of gold nanoparticles

Multifunctional polymer unimolecular micelles, which are used as templates to fabricate stable gold nanoparticles (GNPs) in one‐step without external reductant, have been designed and prepared. Amphiphilic 21‐arm star‐like block copolymers β‐cyclodextrin‐{poly(lactide)‐poly(2‐(dimethylamino) ethyl methacrylate)‐poly[oligo(2‐ethyl‐2‐oxazoline)methacrylate]}₂₁ [β‐CD‐(PLA‐PDMAEMA‐PEtOxMA)₂₁] and the precursors are synthesized by the combination of ring‐opening polymerization (ROP) and activators regenerated by electron transfer atom transfer radical polymerization (ARGET ATRP). The tertiary amine groups of PDMAEMA block reduce the counterion to zerovalent gold in situ, and these gold atoms combine mutually to form final GNPs. GNPs with relatively small size and narrow size distribution can be obtained in longer DMAEMA block copolymer, larger molar ratio of DMAEMA to HAuCl₄ and smaller absolute concentrations of both polymer and HAuCl₄. These results showed that the unimolecular micelles can be used as templates for preparing and stabilizing GNPs in situ without any external reducing agents and organic solvents, suggesting that the nanocomposite systems are latent nanocarriers for further biomedical application. © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 186–196     

Selective Targeting of Dendritic Cell‐Specific Intercellular Adhesion Molecule‐3‐Grabbing Nonintegrin (DC‐SIGN) with Mannose‐Based Glycomimetics: Synthesis and Interaction Studies of Bis(benzylamide) Derivatives of a Pseudomannobioside

Dendritic cell‐specific intercellular adhesion molecule‐3‐grabbing nonintegrin (DC‐SIGN) and Langerin are C‐type lectins of dendritic cells (DCs) that share a specificity for mannose and are involved in pathogen recognition. HIV is known to use DC‐SIGN on DCs to facilitate transinfection of T‐cells. Langerin, on the contrary, contributes to virus elimination; therefore, the inhibition of this latter receptor is undesired. Glycomimetic molecules targeting DC‐SIGN have been reported as promising agents for the inhibition of viral infections and for the modulation of immune responses mediated by DC‐SIGN. We show here for the first time that glycomimetics based on a mannose anchor can be tuned to selectively inhibit DC‐SIGN over Langerin. Based on structural and binding studies of a mannobioside mimic previously described by us ( 2 ), a focused library of derivatives was designed. The optimized synthesis gave fast and efficient access to a group of bis(amides), decorated with an azide‐terminated tether allowing further conjugation. SPR inhibition tests showed improvements over the parent pseudomannobioside by a factor of 3–4. A dimeric, macrocyclic structure ( 11 ) was also serendipitously obtained, which afforded a 30‐fold gain over the starting compound ( 2 ). The same ligands were tested against Langerin and found to exhibit high selectivity towards DC‐SIGN. Structural studies using saturation transfer difference NMR spectroscopy (STD‐NMR) were performed to analyze the binding mode of one representative library member with DC‐SIGN. Despite the overlap of some signals, it was established that the new ligand interacts with the protein in the same fashion as the parent pseudodisaccharide. The two aromatic amide moieties showed relatively high saturation in the STD spectrum, which suggests that the improved potency of the bis(amides) over the parent dimethyl ester can be attributed to lipophilic interactions between the aromatic groups of the ligand and the binding site of DC‐SIGN.     

Compact,Polyvalent Mannose Quantum Dots as Sensitive,Ratiometric FRET Probes for Multivalent Protein–Ligand Interactions

A highly efficient cap‐exchange approach for preparing compact, dense polyvalent mannose‐capped quantum dots (QDs) has been developed. The resulting QDs have been successfully used to probe multivalent interactions of HIV/Ebola receptors DC‐SIGN and DC‐SIGNR (collectively termed as DC‐SIGN/R) using a sensitive, ratiometric Förster resonance energy transfer (FRET) assay. The QD probes specifically bind DC‐SIGN, but not its closely related receptor DC‐SIGNR, which is further confirmed by its specific blocking of DC‐SIGN engagement with the Ebola virus glycoprotein. Tuning the QD surface mannose valency reveals that DC‐SIGN binds more efficiently to densely packed mannosides. A FRET‐based thermodynamic study reveals that the binding is enthalpy‐driven. This work establishes QD FRET as a rapid, sensitive technique for probing structure and thermodynamics of multivalent protein–ligand interactions.     

Compact,Polyvalent Mannose Quantum Dots as Sensitive,Ratiometric FRET Probes for Multivalent Protein–Ligand Interactions

A highly efficient cap‐exchange approach for preparing compact, dense polyvalent mannose‐capped quantum dots (QDs) has been developed. The resulting QDs have been successfully used to probe multivalent interactions of HIV/Ebola receptors DC‐SIGN and DC‐SIGNR (collectively termed as DC‐SIGN/R) using a sensitive, ratiometric Förster resonance energy transfer (FRET) assay. The QD probes specifically bind DC‐SIGN, but not its closely related receptor DC‐SIGNR, which is further confirmed by its specific blocking of DC‐SIGN engagement with the Ebola virus glycoprotein. Tuning the QD surface mannose valency reveals that DC‐SIGN binds more efficiently to densely packed mannosides. A FRET‐based thermodynamic study reveals that the binding is enthalpy‐driven. This work establishes QD FRET as a rapid, sensitive technique for probing structure and thermodynamics of multivalent protein–ligand interactions.     

Synthetic Glycopolypeptides as Potential Inhibitory Agents for Dendritic Cells and HIV‐1 Trafficking

Multivalent binding is a key for many critical biological processes and unique recognition and specificity in binding enables many of different glycans and proteins to work in a great harmony within the human body. In this study, the binding kinetics of synthetic glycopolypeptides to the dendritic cell lectin DC‐SIGN and their inhibition potential for DC‐SIGN interactions with the gp120 envelope glycoprotein of HIV‐1 (gp120) are investigated.

     

Synthesis and Predetermined Supramolecular Chirality of Carbohydrate‐Functionalized Perylene Bisimide Derivatives

Eight carbohydrate‐modified perylene bisimides ( PBI‐4 lac‐2 lac , PBI‐4 lac‐2 Man , PBI‐4 lac‐2 Gal , PBI‐4 lac‐2 Mal , PBI‐4 Man‐2 Man , PBI‐4 Man‐2 lac , PBI‐4 Man‐2 Gal and PBI‐4 Man‐2 Mal ) were synthesized, and the following predetermined supramolecular chirality rule was found: perylene bisimides modified with disaccharides (D ‐lactose and D ‐maltose) at the imide position generated right‐handed chirality, and those modified with monosaccharides (D ‐mannose and D ‐galactose) generated left‐handed chirality, when D ‐lactose or D ‐mannose was substituted in the bay positions of perylene bisimides with amide bonds as the linking spacers. These results may be because of the difference in the stacking angle of the perylene bisimide backbones induced by the steric effect and the additional hydrogen bonds between the disaccharide residues. This study provides an important design rule for predetermined chiral self‐assembly of perylene bisimides.     

Structural and Kinetic Dissection of the endo‐α‐1,2‐Mannanase Activity of Bacterial GH99 Glycoside Hydrolases from Bacteroides spp.

Glycoside hydrolase family 99 (GH99) was created to categorize sequence‐related glycosidases possessing endo‐α‐mannosidase activity: the cleavage of mannosidic linkages within eukaryotic N‐glycan precursors (Glc_1–3Man₉GlcNAc₂), releasing mono‐, di‐ and triglucosylated‐mannose (Glc_1–3‐1,3‐Man). GH99 family members have recently been implicated in the ability of Bacteroides spp., present within the gut microbiota, to metabolize fungal cell wall α‐mannans, releasing α‐1,3‐mannobiose by hydrolysing αMan‐1,3‐αMan→1,2‐αMan‐1,2‐αMan sequences within branches off the main α‐1,6‐mannan backbone. We report the development of a series of substrates and inhibitors, which we use to kinetically and structurally characterise this novel endo‐α‐1,2‐mannanase activity of bacterial GH99 enzymes from Bacteroides thetaiotaomicron and xylanisolvens. These data reveal an approximate 5 kJ mol^?1 preference for mannose‐configured substrates in the ?2 subsite (relative to glucose), which inspired the development of a new inhibitor, α‐mannopyranosyl‐1,3‐isofagomine (ManIFG), the most potent (bacterial) GH99 inhibitor reported to date. X‐ray structures of ManIFG or a substrate in complex with wild‐type or inactive mutants, respectively, of B. xylanisolvens GH99 reveal the structural basis for binding to D ‐mannose‐ rather than D ‐glucose‐configured substrates.     

Thermal and pH‐sensitive gold nanoparticles from H‐shaped block copolymers of (PNIPAM/PDMAEMA)‐b‐PEG‐b‐(PNIPAM/PDMAEMA)

Well‐defined H‐shaped pentablock copolymers composed of poly(N‐isopropylacrylamide) (PNIPAM), poly(N,N‐dimethylaminoethylacrylamide) (PDMAEMA), and poly(ethylene glycol) (PEG) with the chain architecture of (A/B)‐b‐C‐b‐(A/B) were synthesized by the combination of single‐electron‐transfer living radical polymerization, atom‐transfer radical polymerization, and click chemistry. Single‐electron‐transfer living radical polymerization of NIPAM using α,ω azide‐capped PEG macroinitiator resulted in PNIPAM‐b‐PEG‐b‐PNIPAM with azide groups at the block joints. Atom‐transfer radical polymerization of DMAEMA initiated by propargyl 2‐chloropropionate gave out α‐capped alkyne‐PDMAEMA. The H‐shaped copolymers were finally obtained by the click reaction between PNIPAM‐b‐PEG‐b‐PNIPAM and alkyne‐PDMAEMA. These copolymers were used to prepare stable colloidal gold nanoparticles (GNPs) in aqueous solution without any external reducing agent. The formation of GNPs was affected by the length of PDMAEMA block, the feed ratio of the copolymer to HAuCl₄, and the pH value. The surface plasmon absorbance of these obtained GNPs also exhibited pH and thermal dependence because of the existence of PNIAPM and PDAMEMA blocks. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2010     

Improved Access to Imidazole‐phosphonic Acids: Synthesis of D‐manno‐Tetrahydroimidazopyridine‐2‐phosphonates

The D ‐manno‐tetrahydroimidazopyridine‐2‐phosphonate 11 was prepared via a high‐yielding Pd(PPh₃)₄‐catalysed diphenylphosphonylation of the manno‐iodoimidazole 12 , followed by transesterification to the diethyl phosphonate 14 and dealkylation, providing 11 in eight steps from the thionolactam 1 and in an overall yield of 15%. Alternatively, a more highly convergent synthesis based on the HgCl₂/Et₃N‐promoted condensation of the thionolactam 1 with the α‐aminophosphonate 24 in THF led to 11 in four steps and in the same overall yield. In the presence of HgCl₂/Et₃N, the thionolactam 1 reacted at 80° with 2‐methoxyethanol to provide 66% of a 64 : 36 mixture of the gluco‐ and manno‐iminoethers 29 / 30 . Performing the reaction at 22° yielded preferentially the gluco‐isomer 29 (86%, 84 : 16).     

Isoquinuclidine Mimics of β‐D‐Glucopyranosides: Differences and Similarities in the Mechanism of Action of some β‐D‐Glucosidases and a β‐D‐Mannosidase

The D ‐gluco‐isoquinuclidines 3 and 4 were prepared and tested as inhibitors of the β‐glucosidases from Caldocellum saccharolyticum and from sweet almonds; the results are compared to the inhibition of snail β‐mannosidase by the D ‐manno‐isoquinuclidines 1 and 2 . Exploratory experiments in the racemic series showed that treatment of the ester epoxide 6 with benzyl alcoholates leads only to epimerisation, transesterification, and formation of the cyclopropane 9 . Ring opening of the reduced epoxide 13 by NaN₃ proceeded regioselectively to provide 14 . Treatment of the C(6)? O‐triflate 16 with AcOCs induced a rearrangement; the reaction with NaN₃ gave the C(5)‐azido derivative 14 . The acetoxy triflate 18 , however, reacted with AcOCs to provide the desired gluco‐isoquinuclidine 19 . Similarly, the enantiomerically pure acetoxy triflate 22 provided the D ‐gluco‐isoquinuclidine 24 , which was reduced and deprotected to provide 3 and 4 . The deoxy analogues 30 and 31 were obtained by reductive deiodination of the iodide 27 , derived from 22 . The D ‐gluco‐isoquinuclidines 3, 4, 30 , and 31 are much weaker inhibitors of β‐glucosidases than the D ‐manno‐analogues 1 and 2 of snail β‐mannosidase. The N‐benzyl derivative 3 is a weaker inhibitor than the N‐unsubstituted analogue in the gluco‐series, while it is a much stronger inhibitor in the manno‐series. A consideration of the pK_HA values of the isoquinuclidines 1 – 4 and the pH value of the enzyme assays suggests that the D ‐gluco‐isoquinuclidines are poor mimics of the shape of a reactive, enzyme‐bound gluco‐conformer, while the D ‐manno‐analogues are reasonably good mimics of a reactive, enzyme‐bound manno‐conformer. The inhibition results may also suggest that the glycosidase induced lengthening of the scissile bond and rehybridisation of the anomeric centre are more strongly correlated with the change of the ground‐state conformation during hydrolysis of β‐D ‐glucopyranosides than of β‐D ‐mannopyranosides.     

Synthesis and Evaluation as Glycosidase Inhibitors of Isoquinuclidines Mimicking a Distorted β‐Mannopyranoside

Racemic and enantiomerically pure manno‐configured isoquinuclidines were synthesized and tested as glycosidase inhibitors. The racemic key isoquinuclidine intermediate was prepared in high yield by a cycloaddition (tandem Michael addition/aldolisation) of the 3‐hydroxy‐1‐tosyl‐pyridone 10 to methyl acrylate, and transformed to the racemic N‐benzyl manno‐isoquinuclidine 2 and the N‐unsubstituted manno‐isoquinuclidine 3 (twelve steps; ca. 11% from 10 ). Catalysis by quinine of the analogous cycloaddition of 10 to (?)‐8‐phenylmenthyl acrylate provided a single diastereoisomer in high yield, which was transformed to the desired enantiomerically pure D ‐manno‐isoquinuclidines (+)‐ 2 and (+)‐ 3 (twelve steps; 23% from 10 ). The enantiomers (?)‐ 2 and (?)‐ 3 were prepared by using a quinidine‐promoted cycloaddition of 10 to the enantiomeric (+)‐8‐phenylmenthyl acrylate. The N‐benzyl D ‐manno‐isoquinuclidine (+)‐ 2 is a selective and slow inhibitor of snail β‐mannosidase. Its inhibition strength and type depends on the pH (at pH 4.5: K_i=1.0 μM , mixed type, α=1.9; at pH 5.5: K_i=0.63 μM , mixed type, α=17). The N‐unsubstituted D ‐manno‐isoquinuclidine (+)‐ 3 is a poor inhibitor. Its inhibition strength and type also depend on the pH (at pH 4.5: K_i=1.2?10³ μM , mixed type, α=1.1; at pH 5.5: K_i=0.25?10³ μM , mixed type, α=11). The enantiomeric N‐benzyl L ‐manno‐isoquinuclidine (?)‐ 2 is a good inhibitor of snail β‐mannosidase, albeit noncompetitive (at pH 4.5: K_i=69 μM ). The N‐unsubstituted isoquinuclidine (?)‐ 2 is a poor inhibitor (at pH 4.5: IC₅₀=7.3?10³ μM ). A comparison of the inhibition by the pure manno‐isoquinuclidines (+)‐ 2 and (+)‐ 3 , (+)‐ 2 /(?)‐ 2 1 : 1, and (+)‐ 3 /(?)‐ 3 1 : 1 with the published data for racemic 2 and 3 led to a rectification of the published data. The inhibition of snail β‐mannosidase by the isoquinuclidines 2 and 3 suggests that the hydrolysis of β‐D ‐mannopyranosides by snail β‐mannosidase proceeds via a distorted conformer, in agreement with the principle of stereoelectronic control.     

Construction of a High‐Mannose‐Type Glycan Library by a Renewed Top‐Down Chemo‐Enzymatic Approach

A comprehensive method for the construction of a high‐mannose‐type glycan library by systematic chemo‐enzymatic trimming of a single Man9‐based precursor was developed. It consists of the chemical synthesis of a non‐natural tridecasaccharide precursor, the orthogonal demasking of the non‐reducing ends, and trimming by glycosidases, which enabled a comprehensive synthesis of high‐mannose‐type glycans in their mono‐ or non‐glucosylated forms. It employed glucose, isopropylidene, and N‐acetylglucosamine groups for blocking the A‐, B‐, and C‐arms, respectively. After systematic trimming of the precursor, thirty‐seven high‐mannose‐type glycans were obtained. The power of the methodology was demonstrated by the enzymatic activity of human recombinant N‐acetylglucosaminyltransferase‐I toward M7–M3 glycans, clarifying the substrate specificity in the context of high‐mannose‐type glycans.     

Effect of oligonucleotide conformation on its facilitation efficiency on negatively charged micelle‐to‐vesicle transition

In our previous article, we reported for the first time that the oligonucleotides composed of one nucleotide species, for example, oligo d(A)_n, oligo d(C)_n, and oligo d(T)_n, could facilitate negatively charged sodium dodecyl sulfate/dodecyl triethyl ammonium bromide mixed micelles to transform to vesicles. In this study, we will report the facilitation ability of self‐complementary hairpin‐structured oligonucleotides, oligo d(A_nCT_n) and oligo d(AT)_nACT(AT)_n (or oligo d(AT)_nC(AT)_n), on micelle‐to‐vesicle transition. It is found that the facilitation behavior of hairpin‐structured oligonucleotide is different from that of the oligonucleotide comprising one base species, and the facilitation efficiency of hairpin‐structured oligonucleotide is closely dependent on the sequence of bases A and T; oligo d(A_nCT_n) is more efficient than oligo d(AT)_nACT(AT)_n (or oligo d(AT)_nC(AT)_n). Moreover, oligo d(A_nCT_n) is more efficient than oligo d(A)_n, oligo d(C)_n, and oligo d(T)_n. Since so far, there is very limited report about the facilitation effect of oligonucleotide and DNA on vesicle formation as well as the role of their conformation in their interaction with surfactant, this study should be expected to provide some helpful information for the application of DNA/amphiphile system. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 48: 852–860, 2010     

Synthesis of C(2)‐Substituted manno‐Configured Tetrahydroimidazopyridines and Their Evaluation as Inhibitors of Snail β‐Mannosidase

It was shown that retaining β‐glucosidases and galactosidases of families 1–3 feature a strong interaction between C(2)OH of the substrate and the catalytic nucleophile. An analogous interaction can hardly take place for retaining β‐mannosidases. A structure? activity comparison between the inhibition of the β‐glucosidase from Caldocellum saccharolyticum (family 1) and β‐glucosidase from sweet almonds by the gluco‐imidazoles 1 – 6 , and the inhibition of snail β‐mannosidase by the corresponding manno‐imidazoles 8 – 13 does not show any significant difference, suggesting that also the mechanisms of action of these glycosidases do not differ significantly. For this comparison, we synthesized and tested the manno‐imidazoles 9 – 13, 28, 29, 32, 35, 40, 41, 43, 46, 47 , and 50 . Among these, the alkene 29 is the strongest known inhibitor of snail β‐mannosidase (K_i=6 nM , non‐competitive); the aniline 35 is the strongest competitive inhibitor (K_i=8 nM ).     

HPMA‐Based Nanocarriers for Effective Immune System Stimulation

The selective activation of the immune system using nanoparticles as a drug delivery system is a promising field in cancer therapy. Block copolymers from HPMA and laurylmethacrylate‐co‐hymecromone‐methacrylate allow the preparation of multifunctionalized core‐crosslinked micelles of variable size. To activate dendritic cells (DCs) as antigen presenting cells, the carbohydrates mannose and trimannose are introduced into the hydrophilic corona as DC targeting units. To activate DCs, a lipophilic adjuvant (L18‐MDP) is incorporated into the core of the micelles. To elicit an immune response, a model antigen peptide (SIINFEKL) is attached to the polymeric nanoparticle—in addition—via a click reaction with the terminal azide. Thereafter, the differently functionalized micelles are chemically and biologically characterized. While the core‐crosslinked micelles without carbohydrate units are hardly bound by DCs, mannose and trimannose functionalization lead to a strong binding. Flow cytometric analysis and blocking studies employing mannan suggest the requirement of the mannose receptor and DC‐SIGN for effective micelle binding. It could be suppressed by blocking with mannan. Adjuvant‐loaded micelles functionalized with mannose and trimannose activate DCs, and DCs preincubated with antigen‐conjugated micelles induce proliferation of antigen‐specific CD8+ T cells.     

Synthesis of amphiphilic block copolymer by metal‐free ring‐opening oligomerization of glycidyl phenyl ether initiated with tetra‐n‐butylammonium fluoride in the presence of poly(ethylene glycol) monomethyl ether

Metal‐free ring‐opening oligomerization of glycidyl phenyl ether (GPE) initiated with tetra‐n‐butylammonium fluoride (n‐Bu₄NF) (5.0 mol %) was performed in the presence of poly(ethylene glycol) monomethyl ether (PEGM) (5.0, 10, 20 mol %) as a chain transfer agent, by which the resulting polymers having narrow molecular weight distribution (M_w/M_n < 1.2) were obtained in 80–84% yield. Solubility of the obtained polymers in water increased with the increase of amount of PEGM, owing to an increase of number of PEGM‐block‐oligo(GPE) molecules compared to that of oligo(GPE) molecules having FCH₂– group at the initiating end as well as a decrease in degree of oligomerization of oligo(GPE). The PEGM‐block‐oligo(GPE) was isolated by filtration of the polymer aqueous solution, whose number‐average molecular weights determined by NMR spectroscopic analysis were almost consistent to the theoretical values. The PEGM‐block‐oligo(GPE) formed micelles in aqueous media, whose average particle diameter was 58 and 140 nm for the copolymers having a composition of PEGM:GPE = 62:38 and 53:47, respectively. © 2013 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2013, 51, 4451–4458     

Synthesis and properties of oligo(biradicals) composed of 1,3‐diphosphacyclobutane‐2,4‐diyl units and benzyl‐type linkers

We have succeeded in catenating two sterically encumbered 1,3‐di‐t‐butyl‐2,4‐bis(2,4,6‐tri‐t‐butylphenyl)‐1,3‐diphosphacyclobutane‐2,4‐diyl units with a spacer 1,2‐(CH₂)₂C₆H₄ to obtain bis(biradicals) as considerably stable compounds. We have discussed physicochemical properties of the dimer, together with DFT calculations of model compounds. Spectroscopic data, redox properties, and X‐ray structures of the oligo(biradicals) derivatives including other spacers like 1,3‐(CH₂)₂C₆H₄, 1,4‐(CH₂)₂C₆H₄, and 1,3,5‐(CH₂)₃C₆H₃, reveal that the P‐heterocyclic biradical moieties interact through nonconjugative pathways. These properties of oligo(biradicals) will facilitate to design novel molecular systems for electronics. © 2010 Wiley Periodicals, Inc. Heteroatom Chem 21:404–411, 2010; View this article online at wileyonlinelibrary.com . DOI 10.1002/hc.20625     

Divergent Synthesis of 3‐Deoxy‐d‐manno‐oct‐2‐ulosonic Acid (Kdo) Glycosides Containing α‐(2→4)‐Linked Kdo‐Kdo Unit

A convenient and divergent approach was developed to prepare diverse bacterial 3‐deoxy‐d ‐manno‐oct‐2‐ulosonic acid (Kdo) oligosaccharides containing a Kdo‐α‐(2→4)‐Kdo fragment. The orthogonal protected α‐(2→4) linked Kdo‐Kdo disaccharide 3 , serving as a common precursor, was divergently transformed into the corresponding 8‐, 8′‐, and 4′‐hydroxy disaccharides 5 , 7 , and 14 , respectively. Then, these alcohols were glycosylated, respectively, with the 5,7‐O‐di‐tert‐butylsilylene (DTBS) protected Kdo thioglycoside donors 1 or 2 in an α‐stereoselective and high‐yielding manner to afford a range of Kdo oligosaccharides. Finally, removal of all protecting groups of the newly formed glycosides resulted in the desired free Kdo oligomer.     

Travelling‐wave ion mobility mass spectrometry and negative ion fragmentation of hybrid and complex N‐glycans

Nitrogen collisional cross sections (CCSs) of hybrid and complex glycans released from the glycoproteins IgG, gp120 (from human immunodeficiency virus), ovalbumin, α1‐acid glycoprotein and thyroglobulin were measured with a travelling‐wave ion mobility mass spectrometer using dextran as the calibrant. The utility of this instrument for isomer separation was also investigated. Some isomers, such as Man₃GlcNAc₃ from chicken ovalbumin and Man₃GlcNAc₃Fuc₁ from thyroglobulin could be partially resolved and identified by their negative ion fragmentation spectra obtained by collision‐induced decomposition (CID). Several other larger glycans, however, although existing as isomers, produced only asymmetric rather than separated arrival time distributions (ATDs). Nevertheless, in these cases, isomers could often be detected by plotting extracted fragment ATDs of diagnostic fragment ions from the negative ion CID spectra obtained in the transfer cell of the Waters Synapt mass spectrometer. Coincidence in the drift times of all fragment ions with an asymmetric ATD profile in this work, and in the related earlier paper on high‐mannose glycans, usually suggested that separations were because of conformers or anomers, whereas symmetrical ATDs of fragments showing differences in drift times indicated isomer separation. Although some significant differences in CCSs were found for the smaller isomeric glycans, the differences found for the larger compounds were usually too small to be analytically useful. Possible correlations between CCSs and structural types were also investigated, and it was found that complex glycans tended to have slightly smaller CCSs than high‐mannose glycans of comparable molecular weight. In addition, biantennary glycans containing a core fucose and/or a bisecting GlcNAc residue fell on different mobility‐m/z trend lines to those glycans not so substituted with both of these substituents contributing to larger CCSs. Copyright © 2016 John Wiley & Sons, Ltd.     

Use of cyclodextrin‐modified gold nanoparticles for enantioseparations of drugs and amino acids based on pseudostationary phase‐capillary electrochromatography

The application of chemical‐modified gold nanoparticles (GNPs) as chiral selector for the enantioseparation based on pseudostationary phase‐CEC (PSP‐CEC) is presented. GNPs modified by thiolated β‐CD were characterized by NMR and FT‐IR. The nanoparticle size was determined to be of 9.5 nm (+2.5 nm) by Transmission Electron Microscopy (TEM) and UV spectra. Four pairs of dinitrophenyl‐labeled amino acid enantiomers (DL‐Val, Leu, Glu and Asp) and three pairs of drug enantiomers (RS‐chlorpheniramine, zopiclone and carvedilol) were analyzed by using modified GNPs as the chiral selector in PSP‐CEC. Good theoretical plate number (up to 2.4×10⁵ per meter) and separation resolution (up to 4.7) were obtained even with low concentration of modified GNPs (0.8–1.4 mg/mL). The corresponding concentration of β‐CD in the buffer was only 0.30?0.53 mM, which was much lower than the optimum concentration of 15 mM if pure β‐CD was used as chiral selector. Our results showed that thiolated β‐CD modified GNPs have more sufficient interaction with the analytes, resulting in significant enhancement of enantioseparation. The study shed light on potential usage of chemical modified GNPs as chiral selector for enantioseparation based on PSP‐CEC.