Facile Solid-phase Research Articles

Improved liquid chromatography–tandem mass spectrometry method in clinical utility for the diagnosis of Cushing’s syndrome

Determination of urinary free cortisol is one of the first lines in screening for the diagnosis of Cushing's syndrome where its measurement is mostly done by immunoassay. Although easy to perform, immunoassays suffer from the problem of assay interferences and are unable to measure cortisone levels. To enhance such techniques for clinical diagnosis, an improved liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed for the simultaneous determination of urinary free cortisol and cortisone. The leftover urine samples from immunoassay were collected and subjected to facile solid-phase extraction cleanup. In the analysis of 130 urine samples from patients, 65 (50%) were found to have elevated urinary free cortisol (UFC) by immunoassay; but only 13 (10.8%) were found to have elevated UFC by this improved LC-MS/MS method. Nine out of the 13 patients, which showed elevated UFC by LC-MS/MS, were surgically confirmed to have Cushing's syndrome/disease. By setting a two times upper limit as a cut-off, the immunoassay gave a positive predictive value of 43.5%, whilst by using the improved method, a positive predictive value of 90% was obtained. Although several tests have been used extensively in first line screening for the diagnosis of Cushing's syndrome, none has ever shown with full capability of distinguishing all cases of Cushing's syndrome from normal and/or obese individuals. This method has shown superior analytical advantages over existing immunoassay type in terms of sensitivity, specificity and capability to diagnose Cushing's syndrome. Comparison between existing spectrometric methods, the reported developed method shown here, provides a simpler sample preparation procedure and meets with the high throughput demand of clinical laboratories.

“Four-Potential” Ferrocene Labeling of PNA Oligomers via Click Chemistry

The scope of the Cu(I)-catalyzed [2 + 3] azide/alkyne cycloaddition (CuAAC, click chemistry) as a key reaction for the conjugation of ferrocene derivatives to N-terminal functionalized PNA oligomers is explored herein (PNA: peptide nucleic acid). The facile solid-phase synthesis of N-terminal azide or alkyne-functionalized PNA oligomer precursors and their cycloaddition with azidoferrocene, ethynylferrocene, and N-(3-ethylpent-1-yn-3-yl)ferrocene-carboxamide (DEPA-ferrocene) on the solid phase are presented. While the click reaction with azidomethylferrocene worked equally well, the ferrocenylmethyl group is lost from the conjugate upon acid cleavage. However, the desired product was obtained via a post-SPPS conversion of the alkyne-PNA oligomer with azidomethylferrocene in solution. The synthesis of all ferrocene-PNA conjugates (trimer t(3)-PNA, 3, 4, 5, 6; 12mer PNA, 10 - t c t a c a a g a c t c, 11 - t c t a c c g t a c t c) succeeded with excellent yields and purities, as determined by mass spectrometry and HPLC. Electrochemical studies of the trimer Fc-PNA conjugates 3, 4, 5, and 6 with four different ferrocene moieties revealed quasi-reversible redox processes of the ferrocenyl redox couple Fc(0/+) and electrochemical half-wave potentials in a range of E(1/2) = -20 mV to +270 mV vs FcH(0/+) (Fc: ferrocenyl, C(10)H(9)Fe). The observed potential differences ΔE(1/2)(min) are always greater than 60 mV for any given pair of Fc-PNA conjugates, thus allowing a reliable differentiation with sensitive electrochemical methods like e.g. square wave voltammetry (SWV). This is the electrochemical equivalent of "four-color" detection and is hence denoted "four-potential" labeling. Preparation and electrochemical investigation of the set of four structurally different and electrochemically distinguishable ferrocenyl groups conjugated to PNA oligomers, as exemplified by the conjugates 3, 4, 5, and 6, demonstrates the scope of the azide/alkyne cycloaddition for the labeling of PNA with electrochemically active ferrocenyl groups. Furthermore, it provides a PNA-based system for the electrochemical detection of single-nucleotide polymorphism (SNP) in DNA/RNA.

Biomonitoring of 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and Its Carcinogenic Metabolites in Urine

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is a carcinogenic heterocyclic aromatic amine that is produced in cooked meats. The simultaneous analysis of PhIP and its metabolites in human urine is a challenge, because these biomarkers only occur in urine at parts per billion or lower concentrations and must be selectively purifed from thousands of other urinary constituents. We have developed a facile solid-phase extraction method, employing a mixed-mode reverse-phase cation exchange resin, to simultaneously isolate PhIP, its glucuronide conjugates, and the glucuronide conjugates of the genotoxic metabolite 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine from the urine of meat eaters. PhIP and its metabolites were quantified by liquid chromatography-electrospray ionization/tandem mass spectrometry (LC-ESI/MS/MS), using a triple stage quadrupole mass spectrometer in the selected reaction monitoring scan mode. The lower limit of quantification (LOQ) of PhIP is 5 parts per trillion (ppt), and the LOQ values for the glucuronide conjugates are 50 ppt, when 25 microL of urine is employed for assay. The extraction scheme is versatile and has been employed to isolate other ring-hydroxylated and glucuronidated metabolites of PhIP, for characterization by LC-ESI/MS/MS.

Solid-phase synthesis of aryl vinyl ethers based on polystyrene-supported β-phenylselenoethanol

A novel facile solid-phase organic synthesis of aryl vinyl ethers by reaction of polystyrene-supported β-phenylselenoethanol with phenols under Mitsunobu conditions and subsequent oxidation-elimination with 30% hydrogen peroxide has been developed. The advantages of this method include straightforward operation, lack of odour, good yield and high purity of the products.

A facile three-component solid phase synthesis of steroidal A-ring fused pyrimidines under microwave irradiation

The preparation of ring-A fused pyrimidines at the steroidal 2,3-position is herein described. The novel steroidal pyrimidines were prepared from the solid phase three-component reaction of 2-hydroxymethylene-3-keto steroids, arylaldehydes and ammonium acetate under microwave irradiation.

Oligo(N-aryl glycines): A New Twist on Structured Peptoids

We explore strategies to enhance conformational ordering of N-substituted glycine peptoid oligomers. Peptoids bearing bulky N-alkyl side chains have previously been studied as important examples of biomimetic "foldamer" compounds, as they exhibit a capacity to populate helical structures featuring repeating cis-amide bonds. Substantial cis/trans amide bond isomerization, however, gives rise to conformational heterogeneity. Here, we report the use of N-aryl side chains as a tool to enforce the presence of trans-amide bonds, thereby engendering structural stability. Aniline derivatives and bromoacetic acid are used in the facile solid-phase synthesis of a diverse family of sequence-specific N-aryl glycine oligomers. Quantum mechanics calculations yield a detailed energy profile of the folding landscape and substantiate the hypothesis that the presence of anilide groups establishes a strong energetic preference for trans-amide bonds. X-ray crystallographic analysis and solution NMR studies verify this preference. Molecular modeling indicates that the linear oligomers can adopt helical structures resembling a polyproline type II helix. High resolution structures of macrocyclic oligomers incorporating both N-alkyl and N-aryl glycine units confirm the ability to direct the presence of trans-amide bonds specifically at N-aryl positions. These results are an important step in developing strategies for the rational de novo design of new structural motifs in biomimetic oligopeptoid systems.

Peptoids that mimic the structure, function, and mechanism of helical antimicrobial peptides

Antimicrobial peptides (AMPs) and their mimics are emerging as promising antibiotic agents. We present a library of "ampetoids" (antimicrobial peptoid oligomers) with helical structures and biomimetic sequences, several members of which have low-micromolar antimicrobial activities, similar to cationic AMPs like pexiganan. Broad-spectrum activity against six clinically relevant BSL2 pathogens is also shown. This comprehensive structure-activity relationship study, including circular dichroism spectroscopy, minimum inhibitory concentration assays, hemolysis and mammalian cell toxicity studies, and specular x-ray reflectivity measurements shows that the in vitro activities of ampetoids are strikingly similar to those of AMPs themselves, suggesting a strong mechanistic analogy. The ampetoids' antibacterial activity, coupled with their low cytotoxicity against mammalian cells, make them a promising class of antimicrobials for biomedical applications. Peptoids are biostable, with a protease-resistant N-substituted glycine backbone, and their sequences are highly tunable, because an extensive diversity of side chains can be incorporated via facile solid-phase synthesis. Our findings add to the growing evidence that nonnatural foldamers will emerge as an important class of therapeutics.

Production of hydrogen by steam reforming of methanol over Cu/ZnO catalysts prepared via a practical soft reactive grinding route based on dry oxalate-precursor synthesis

Binary Cu/ZnO catalysts (with a Cu/Zn atomic ratio of 50/50) prepared via a novel dry synthetic approach based on solid-state oxalate-precursor synthesis were studied in regard to their performance in the steam reforming of methanol (SRM). The synthesis route involves facile solid-phase mechanochemical activation of a physical mixture of simple copper/zinc salts and oxalic acid, followed by calcination of the as-ground oxalate precursors at 350 °C. For comparison, their conventional analogues obtained by aqueous coprecipitation techniques were also examined. Structural characterization of the samples was performed by means of N 2 adsorption, X-ray diffraction (XRD), diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS), thermal gravimetric and differential thermal analysis (TG/DTA), scanning electron microscopy (SEM), temperature-programmed reduction (H 2-TPR), N 2O titration, and X-ray photoelectron spectroscopy (XPS). The results show that the grinding-derived Cu/ZnO catalysts exhibit superior SRM performance to their conventional counterparts obtained by wet-chemical methods. The enhanced performance of the grinding-derived catalysts can be attributed to a higher copper dispersion as well as the beneficial generation of highly strained Cu nanocrystals in the working catalyst. It is proposed that the present soft reactive grinding route based on dry oxalate-precursor synthesis can allow the generation of a new type of Cu/ZnO materials with favorable surface and structural properties, providing an attractive alternative for preparation of improved heterogeneous catalysts.

Characterization and field-emission properties of carbon nanotube arrays in nanoporous alumina template and on blank Si substrate

Ordered carbon nanotube (CNT) arrays were synthesized within anodized aluminum oxide template by thermal decomposition of hydrocarbon precursor with hydrogen ambient at growth temperature as low as 500°C. Excess hydrogen in precursor mixture enables a steady supply of mobile hydrocarbon reactant which promotes facile solid-phase diffusion. The activation energy for CNT growth was determined to be 0.55eV, a number smaller than 1.02eV for similar precursor in nitrogen ambient. Moreover, CNTs grown in anodized aluminum oxide nanopores in this low temperature process were found to exhibit unusually high field-emission current of 100mA∕cm2 at 8V∕μm.

A facile solid-phase synthesis of Baylis–Hillman products from polymeric selenium reagents

Treatment of lithio derivatives of polystyrene-supported methyl α-selenopropionate, 2-selenopropiophenone, 1-selenoethyl phenyl sulfone and diethyl 1-selenoethylphosphonate with aldehydes respectively, followed by cleavage from the polymer by oxidation-elimination with 30% hydrogen peroxide efficiently afforded Baylis–Hillman products in good yields and high purities.

Facile solid-phase synthesis of biotinylated alkyl thiols

Biotinylated alkyl thiols with the capacity to graft avidin proteins are in increasing demand for the development of self-assembled monolayers on gold. Here we propose 2-Chlorotrityl Chloride solid-phase resin as a new platform to produce these functionalized alkyl thiols. Biotinylated alkyl thiols of non-obvious solution synthesis were obtained rapidly using this method and without previous purification steps.

Facile preparation of new unsymmetrical curcumin derivatives by solid-phase synthesis strategy

Seventeen unsymmetrical curcumin derivatives were synthesized in good yield and purity by a facile solid phase synthesis strategy.

Facile solid-phase synthesis of 2,3-disubstituted 6 H-pyrano[2,3- f]benzimidazole-6-ones

We report herein the facile solid-phase synthesis of 2,3-disubstituted 6 H-pyrano[2,3- f]benzimidazole-6-ones using 7-fluoro-4-methyl-6-nitro-2-oxo-2 H-1-benzopyran-3-carboxylic acid as the scaffold. The fluorine of the resin-bound scaffold was first replaced by a primary amine. Reduction of the nitro group with tin(II) chloride afforded an o-dianilino intermediate that was treated with an aldehyde followed by the addition of 2,3-dichloro-5,6-dicyanoquinone (DDQ). 2,3-Disubstituted 6 H-pyrano[2,3- f]benzimidazole-6-ones were obtained in high purity and good yield after cleavage.

Column chromatographic purification free synthesis of long-chain monodisperse oligo(1,4-phenyleneethynylene)s: towards large-scale automatic synthesis of molecular wires

A facile, mild and rapid solid phase synthetic route free of column chromatographic purification to the synthesis of soluble monodisperse long-chain oligo(1,4-phenyleneethynylene)s is presented.

Amino acid bromides: their utilization for difficult couplings in solid-phase peptide synthesis

Use of N-protected-α-amino acid bromides for facile solid-phase synthesis of peptides (SPPS) containing extremely sterically hindered non-proteinogenic amino acids is presented. Amino acid bromides (Aaa-Br), generated in situ, were used for the synthesis of long chain homopeptides containing up to eight successive α-MeVal or Aib residues. SPPS of a heteropeptide containing a very bulky amino acid building block is also described. The choice of suitable N-protections is discussed.

A Facile Solid-Phase Synthesis of Substituted 2(5H)-Furanones with Sulfone Traceless Linker

A novel solid-phase synthetic method for substituted 2(5H)-furanones with traceless sulfone linker strategy has been developed. The products were obtained in good yields and excellent purities.

Synthesis and characterization of oligonucleotides containing conformationally constrained bicyclo[3.1.0]hexane pseudosugar analogs.

Oligodeoxyribonucleotides containing pseudorotationally locked sites derived from bicyclo[3.1.0]hexane pseudosugars have been synthesized using adenosine, thymidine and abasic versions of North- and South-methanocarba nucleosides. The reaction conditions for coupling and oxidation steps of oligonucleotide synthesis have been investigated and optimized to allow efficient and facile solid-phase synthesis using phosphoramidite chemistry. Our studies demonstrate that the use of iodine for P(III) to P(V) oxidation leads to strand cleavage at the sites where the pseudosugar is North. In contrast, the same cleavage reaction was not observed in the case of South pseudosugars. Iodine oxidation generates a 5'-phosphate oligonucleotide fragment on the resin and releases the North pseudosugar into the solution. This side reaction, which is responsible for the extremely low yields observed for the incorporation of the North pseudosugar analogs, has been studied in detail and can be easily overcome by replacing iodine with t-butylhydroperoxide as oxidant.

The facile solid-phase synthesis of cholesterol-based polyamine lipids

A facile solid-phase methodology for the production of cholesterol-based polyamines useful in mediating nucleic acid delivery for gene therapy is described. The methodology is compatible with a range of polyamines producing a library of lipids in excellent yields (>87%) and purity.

Solid Phase Synthesis of 2‐Substituted1, 3‐Oxazin‐6‐ones Using Resin‐bound Cyclic Malonic Acid Ester

AbstractA facile solid phase. synthesis of 2‐substituted1,3‐oxazin‐6‐ones using polymer‐supported Meldrum's acid has been reported. Reaction of the resin‐bound cyclic malonic acid ester with triethyl orthoformate and subsequent double substitution with amide, afforded the corresponding polymer‐supported acylaminomethylene cyclic malonic acid ester, which upon thermal treatment led to1, 3‐oxazin‐6‐ones in good yields and with high purity.

Preparation of Resin-Bound Bismethylene Cyclic Malonic Acid Ester and Facile Solid-Phase Synthesis of 2-Alkylthio-4(1H)-quinolone and 2-Alkyl-4(1H)-quinolone

The resin-bound bismethylthiomethylene cyclic malonic acid ester 3 was built up efficiently. Resin 3 can react with arylamines and subsequent thermal cyclizations give 2-alkylthio-4-(1H)-quinolones. Furthermore, conjugate addition of Grignard reagents to the resin 3 forms the resin 6 which was reacted with aryl amines and subsequent thermal-cyclization-cleavages afford the 2-alkyl-4-(1H)-quinolones.