Phenacyl Esters Research Articles

Copper-Catalyzed Azide-Alkyne Cycloaddition-Oriented Multifunctional Bio-Orthogonal Linker BPPA: Design, Synthesis and Evaluation.

The multifunctional linker molecules are crucial for the bio-orthogonal reaction for proteomic target profiling. Herein, we wish to present a novel type of biotin-based tetra-functional bio-orthogonal linkers 3a-3h named BPPA which, possessing a unique photolabile phenacyl ester motif, were readily prepared in 85-90% yields by a simple and green one-step protocol from commercially available and inexpensive reagents of biotin acids and 4'-ethynyl/azido 2-bromoacetophenones. The typical click reaction of BPPA linkers 3a and 3e via copper-catalyzed azide-alkyne cycloaddition (CuAAC) took place easily, resulting in the corresponding BPPA-triazole adducts 4a and 4b in nearly quantitative yields. A further cleavability evaluation of 4a and 4b demonstrated that the expected C-O bond detachment could be accomplished efficiently and rapidly by UV irradiation or by ammonia hydrolysis, respectively, resulting in the residual (hydroxyl)acetylphenyl triazole fragment supposed to be attached to proteins during biological manipulations. The BPPA linkers, with dual clickable options of either the terminal azide or alkyne clickable group, exhibit high potentials for various CuAAC-oriented bio-orthogonal reactions.

Comprehensive insight on protein modification by V-type agent: A chemical proteomic approach employing bioorthogonal reaction.

Organophosphorus compounds (OPs) such as chemical agents and pesticides are posing critical threats to civilians due to their irreversible phosphonylation of diverse amino acids residues forming different protein adducts. However, traditional analytical approaches are quite limited in capturing the myriad of post-translational events that affect protein functions, especially in identifying the low-abundance OP adducts. Herein a systematic proteomic strategy based on a typical click-enrich-release-identify bioorthogonal operation was firstly developed by employing an alkynyl-tagged V-type agent probe (AVP) and a biotin-based azido-enrichment linker (BTP-N3 ). AVP targeting peptides from human serum albumin (HSA) or plasma were captured by BTP-N3 via CuAAC click reaction, enriched by streptavidin beads, released by selective alkaline hydrolysis of phenacyl ester bond, and subsequently sequenced by LC-MS/MS. This strategy has helped identifying 1115 unique OP adduction sites on 163 proteins in human plasma, and covers lots of OP adducts that cannot be achieved by traditional detection methods. The comprehensive coverage of novel OP substrates provided a general and sensitive approach to retrospective verification and/or dose assessment of toxic OPs.

The Glycine-Glucolipid of Alcanivorax borkumensis Is Resident to the Bacterial Cell Wall.

The marine bacterium Alcanivorax borkumensis produces a surface-active glycine-glucolipid during growth with long-chain alkanes. A high-performance liquid chromatography (HPLC) method was developed for absolute quantification. This method is based on the conversion of the glycine-glucolipid to phenacyl esters with subsequent measurement by HPLC with diode array detection (HPLC-DAD). Different molecular species were separated by HPLC and identified as glucosyl-tetra(3-hydroxy-acyl)-glycine with varying numbers of 3-hydroxy-decanoic acid or 3-hydroxy-octanoic acid groups via mass spectrometry. The growth rate of A. borkumensis cells with pyruvate as the sole carbon source was elevated compared to hexadecane as recorded by the increase in cell density as well as oxygen/carbon dioxide transfer rates. The amount of the glycine-glucolipid produced per cell during growth on hexadecane was higher compared with growth on pyruvate. The glycine-glucolipid from pyruvate-grown cells contained considerable amounts of 3-hydroxy-octanoic acid, in contrast to hexadecane-grown cells, which almost exclusively incorporated 3-hydroxy-decanoic acid into the glycine-glucolipid. The predominant proportion of the glycine-glucolipid was found in the cell pellet, while only minute amounts were present in the cell-free supernatant. The glycine-glucolipid isolated from the bacterial cell broth, cell pellet, or cell-free supernatant showed the same structure containing a glycine residue, in contrast to previous reports, which suggested that a glycine-free form of the glucolipid exists which is secreted into the supernatant. In conclusion, the glycine-glucolipid of A. borkumensis is resident to the cell wall and enables the bacterium to bind and solubilize alkanes at the lipid-water interface. IMPORTANCE Alcanivorax borkumensis is one of the most abundant marine bacteria found in areas of oil spills, where it degrades alkanes. The production of a glycine-glucolipid is considered an essential element for alkane degradation. We developed a quantitative method and determined the structure of the A. borkumensis glycine-glucolipid in different fractions of the cultures after growth in various media. Our results show that the amount of the glycine-glucolipid in the cells by far exceeds the amount measured in the supernatant, confirming the proposed cell wall localization. These results support the scenario that the surface hydrophobicity of A. borkumensis cells increases by producing the glycine-glucolipid, allowing the cells to attach to the alkane-water interface and form a biofilm. We found no evidence for a glycine-free form of the glucolipid.

Synthesis and Pancreatic Lipase Inhibitory Activity of Phenacyl Esters of N-Aroyl Amino Acids

Background:Obesity has become a major life-threatening disorder as it is associated with many risk factors such as coronary heart disease, hypertension, type II diabetes and certain forms of cancer. Pancreatic lipase inhibition is one of the alternate strategies to treat obesity. In our previous study, phenacyl ester of N-sulphonyl beta alanine was obtained as a hit molecule when pharmacophore- based virtual screening was carried out using reported pancreatic lipase inhibitors.Objective:In search of novel pancreatic lipase inhibitors, three series of phenacyl esters of Nsubstituted amino acids were synthesized and evaluated for their pancreatic lipase inhibitory activity.Methods:N-aroylation of amino acids and subsequent esterification with different phenacyl bromides in the presence of N,N-Dimethylformamide (DMF) afforded the corresponding phenacyl esters in quantitative yields.Results:The synthesized compounds were found to have good pancreatic lipase inhibitory activity and showed IC50 in the range of 0.036 and 84 μg/ml.Conclusion:All the synthesized compounds showed better pancreatic lipase inhibitory activity and it indicates that the N-aroylated phenacyl esters could be considered as lead moiety for further studies to develop novel anti-obesity potentials.

Synthesis of 4-(1',3'-dioxo-2',3'-dihydro-1H-benzo[de]isoquinolin-2'-yl)-2-butenoic acid and its phenacyl esters

Synthesis of 4-(1',3'-dioxo-2',3'-dihydro-1H-benzo[de]isoquinolin-2'-yl)-2-butenoic acid and its phenacyl esters

Absolute Configurations of Naturally Occurring [5]- and [3]-Ladderanoic Acids: Isolation, Chiroptical Spectroscopy, and Crystallography.

We have isolated mixtures of [5]- and [3]-ladderanoic acids 1a and 2a from the biomass of an anammox bioreactor and have separated the acids and their phenacyl esters for the first time by HPLC. The absolute configurations of the naturally occurring acids and their phenacyl esters are assigned as R at the site of side-chain attachment by comparison of experimental specific rotations with corresponding values predicted using quantum chemical (QC) methods. The absolute configurations for 1a and 2a were independently verified by comparison of experimental Raman optical activity spectra with corresponding spectra predicted using QC methods. The configurational assignments of 1a and 2a and of the phenacyl ester of 1a were also confirmed by X-ray crystallography.

Synthesis, biological evaluation, molecular docking and in silico ADME studies of phenacyl esters of N-Phthaloyl amino acids as pancreatic lipase inhibitors

In continuation of our previous study, a series of phenacyl esters of N-phthaloyl amino acids were synthesized and screened for their in vitro pancreatic lipase inhibitory activity. N-phthloylation of amino acids and subsequent esterification with different phenacyl bromides in presence of N,N-Dimethylformamide (DMF) afforded the corresponding phenacyl esters in quantitative yields. The synthesized compounds were found to have potent in vitro pancreatic lipase inhibitory activity in 0.9–2.5 μM range. The most active compound 2‐(4‐bromophenyl)‐2‐oxoethyl 3‐(1,3‐dioxo‐2,3‐ dihydro‐1H‐isoindol‐2‐yl) propanoate (4g) displayed very good pancreatic lipase inhibitory activity with the IC50 value 0.9 μM. To understand the enzyme inhibition and to ensure the lead likeness, in silico docking and ADME studies were carried on title compounds. The in vitro pancreatic lipase inhibitory activities and in silico studies suggested that this N-phthaloyl phenacyl ester analogues could be a potential lead and may be taken to the next level of antiobesity drug development.

Use of Phenacyl Thiosalicylates for the Preparation of 3-Hydroxybenzo[b]thiophene Derivatives

In this work, we attempted to synthesize thioflavonols using rearrangement of phenacyl thiosalicylates prepared by two different approaches and subjected to cyclization under acidic conditions. Contrary to our expectations, the isolated products were identified as (3-hydroxybenzo[b]thiophen-2-yl)(phenyl)methanones. The detailed reaction mechanism was elucidated by characterization of all reaction intermediates with HPLC and NMR spectroscopy. The applicability of the reaction using different phenacyl esters was tested.

High performance liquid chromatography-charged aerosol detection applying an inverse gradient for quantification of rhamnolipid biosurfactants

A method using high performance liquid chromatography coupled to charged-aerosol detection (HPLC-CAD) was developed for the quantification of rhamnolipid biosurfactants. Qualitative sample composition was determined by liquid chromatography coupled to tandem mass spectrometry (LC⿿MS/MS). The relative quantification of different derivatives of rhamnolipids including di-rhamnolipids, mono-rhamnolipids, and their precursors 3-(3-hydroxyalkanoyloxy)alkanoic acids (HAAs) differed for two compared LC⿿MS instruments and revealed instrument dependent responses. Our here reported HPLC-CAD method provides uniform response.An inverse gradient was applied for the absolute quantification of rhamnolipid congeners to account for the detector⿿s dependency on the solvent composition. The CAD produces a uniform response not only for the analytes but also for structurally different (nonvolatile) compounds. It was demonstrated that n-dodecyl-β-d-maltoside or deoxycholic acid can be used as alternative standards. The method of HPLC-ultra violet (UV) detection after a derivatization of rhamnolipids and HAAs to their corresponding phenacyl esters confirmed the obtained results but required additional, laborious sample preparation steps. Sensitivity determined as limit of detection and limit of quantification for four mono-rhamnolipids was in the range of 0.3⿿1.0 and 1.2⿿2.0μg/mL, respectively, for HPLC-CAD and 0.4 and 1.5μg/mL, respectively, for HPLC-UV. Linearity for HPLC-CAD was at least 0.996 (R2) in the calibrated range of about 1⿿200μg/mL. Hence, the here presented HPLC-CAD method allows absolute quantification of rhamnolipids and derivatives.

Macrocycle-functionalized polystyrene beads as specific absorbers

Crosslinked polystyrene beads were first functionalized with phenacyl ester linkers, and a series of four macrocycles with different ring sizes were synthesized from these linkers. The macrocycles were built from amide-linked monomers coupled by conventional peptide-synthesis methods. Annulation was achieved by copper(I)-catalyzed intramolecular azide–alkyne cycloaddition to give triazole linked macrocycles. The macrocycles were cleaved from the polymer beads with hydrazinolysis or saponification. The structures of macrocycles were confirmed by high-resolution nuclear magnetic resonance (NMR) and liquid chromatography–mass spectrometry (LC–MS) analysis. The ability of the polymers to selectively bind compounds from a mixture of aromatic derivatives in ethanol was tested. The prepared polystyrene supported macrocycles were found to selectively bind bromophenol blue and bromocresol green non-covalently with an association constant of 160–490M−1.

Photorelease of incarcerated guests in aqueous solution with phenacyl esters as the trigger.

We report the clean, efficient photorelease of a series of carboxylic acids embedded in octa acid (OA) host and protected by a p-hydroxyphenacyl cage. A key role is played by the cage by providing hydrophobicity for entry into the OA enclosure and yet readily removable as a photoactivated protecting group for release from the host. The rapid photo-Favorskii rearrangement of the departing chromophore does not react with the host OA but diminishes hydrophobicity of the OA contents, leading to their facile release into the solvent.

A simple and accurate HPLC method for fecal bile acid profile in healthy and cirrhotic subjects: validation by GC-MS and LC-MS

We have developed a simple and accurate HPLC method for measurement of fecal bile acids using phenacyl derivatives of unconjugated bile acids, and applied it to the measurement of fecal bile acids in cirrhotic patients. The HPLC method has the following steps: 1) lyophilization of the stool sample; 2) reconstitution in buffer and enzymatic deconjugation using cholylglycine hydrolase/sulfatase; 3) incubation with 0.1 N NaOH in 50% isopropanol at 60°C to hydrolyze esterified bile acids; 4) extraction of bile acids from particulate material using 0.1 N NaOH; 5) isolation of deconjugated bile acids by solid phase extraction; 6) formation of phenacyl esters by derivatization using phenacyl bromide; and 7) HPLC separation measuring eluted peaks at 254 nm. The method was validated by showing that results obtained by HPLC agreed with those obtained by LC-MS/MS and GC-MS. We then applied the method to measuring total fecal bile acid (concentration) and bile acid profile in samples from 38 patients with cirrhosis (17 early, 21 advanced) and 10 healthy subjects. Bile acid concentrations were significantly lower in patients with advanced cirrhosis, suggesting impaired bile acid synthesis.

Engineering Hydrogen Gas Production from Formate in a Hyperthermophile by Heterologous Production of an 18-Subunit Membrane-bound Complex

Biohydrogen gas has enormous potential as a source of reductant for the microbial production of biofuels, but its low solubility and poor gas mass transfer rates are limiting factors. These limitations could be circumvented by engineering biofuel production in microorganisms that are also capable of generating H2 from highly soluble chemicals such as formate, which can function as an electron donor. Herein, the model hyperthermophile, Pyrococcus furiosus, which grows optimally near 100 °C by fermenting sugars to produce H2, has been engineered to also efficiently convert formate to H2. Using a bacterial artificial chromosome vector, the 16.9-kb 18-gene cluster encoding the membrane-bound, respiratory formate hydrogen lyase complex of Thermococcus onnurineus was inserted into the P. furiosus chromosome and expressed as a functional unit. This enabled P. furiosus to utilize formate as well as sugars as an H2 source and to do so at both 80° and 95 °C, near the optimum growth temperature of the donor (T. onnurineus) and engineered host (P. furiosus), respectively. This accomplishment also demonstrates the versatility of P. furiosus for metabolic engineering applications.

An efficient method for the synthesis of phenacyl ester‐protected dipeptides using neutral alumina‐supported sodium carbonate ‘Na2CO3/n‐Al2O3’

In the synthesis of dipeptides (Boc-AA(1)-AA(2)-OPac: AA(1) and AA(2) represent amino acids) protected by phenacyl (Pac) ester, amines and solid bases as the base for the conversion of the trifluoroacetic acid (TFA) salt of the amino component (TFA·H-AA(2)-OPac) into the corresponding free amino component (H-AA(2)-OPac) were examined. The synthesis of a dipeptide (Boc-Ala-Gly-OPac) using amines for the conversion afforded an unsatisfactory yield with by-products. On the other hand, the use of neutral alumina-supported Na(2) CO(3) (Na(2)CO(3) /n-Al(2)O(3)) as a solid base for the conversion provided the dipeptide in a quantitative yield without by-products. The application of Na(2)CO(3) /n-Al2 O3 to the synthesis of some dipeptides protected by Pac ester gave the desired peptides in excellent yields.

Phenacyl ester derivatives bearing heterocycles as models for photocleavable linkers: synthesis and photolysis studies

The synthesis of several phenacyl ester derivatives linked to oxygen and nitrogen heterocycles, benzoquinolone and (benzo)coumarins, was carried out in an effort to obtain systems that could be applied as photocleavable (bi)functional linkers for solid phase peptide synthesis. The heterocycles were attached to a spacer, with different lengths, followed by coupling to 2-bromo-1-phenylpropan-1-one, acting as a model for the solid support. Photolysis studies of the resulting phenacyl ester derivatives were carried out by irradiation in a photochemical reactor at different wavelengths (300, 350 and 419nm), in methanol/HEPES buffer solution (80:20).

Characterization of Jamaican Agro-Industrial Wastes. Part II, Fatty Acid Profiling Using HPLC: Precolumn Derivatization with Phenacyl Bromide

This paper describes the determination of fatty acid composition of coffee, citrus and rum distillery wastes using reversed-phase high-performance liquid chromatography (RP-HPLC). Lipid extracts of the waste samples are derivatized with phenacyl bromide and their phenacyl esters are separated on a C8 reversed-phase column by using continuous gradient elution with water and acetonitrile. The presence of saturated and unsaturated fatty acids in quantifiable amounts in the examined wastes, as well as the high percentage recoveries, are clear indications that these wastes have potential value as inexpensive sources of lipids. The HPLC procedures described here could be adopted for further analysis of materials of this nature.

Derivatization and liquid chromatography–UV–tandem mass spectrometric analysis of perfluorinated carboxylic acids

The presence of perfluorocarboxylates (PFCAs) in the environment is of increasing concern due to their possible toxicity to humans and bioaccumulation in organisms. PFCAs are frequently found in river water, sediment and organisms and sometimes even in groundwater. In order to quantitatively determine these PFCAs, a fast derivatization coupled with a liquid chromatography–ultraviolet detector–electrospray ionization-tandem mass spectrometry (LC–UV–ESI-MS/MS) method was developed. The PFCAs were quantitatively converted to their corresponding phenacyl esters using p-bromophenacyl bromide as the derivatization reagent. Under optimized reaction conditions, the conversion yield of the PFCAs ranged from 86 to 92% with low %RSD. The typical derivatization product (p-bromophenacyl bromide perfluorooctanoate) was characterized by 1H NMR, 13C NMR, FT-IR and mass spectrometry. UPLC with a BEH C18 column and CAN/H2O (8/2, v/v) as a mobile phase were used to separate the derivatives. The analytes were completely eluted within 6min and multidimensional detection using UV at 260nm and ESI–MRM in the negative ion mode were carried out. Bromide isotopic characteristic fragment ions appeared in the first Q1 scans, and four daughter ions of the MRMs at m/z [M−H−222]−, [M−H−250]–, [M−H−278]− and [M−H−316]− were used for quantification and confirmation. The mass spectral information ensured accurate identification of the analytes even when the sample matrices were complex. The method successfully eliminated the PFCAs background problems originating from polymeric parts in liquid chromatographic systems. The LODs of the method were lower than 5ngmL−1, and the relative standard deviation (RSD%) values ranged from 5.2 to 9.8%. The method was successfully applied for the quantification of PFCAs in river water contaminated by industrial wastewater, and this indicated that the method was useful in the determination of PFCAs in environmental samples.

An Efficient Preparation of N-Methyl-α-amino Acids from N-Nosyl-α-amino Acid Phenacyl Esters

In this paper we describe a simple and efficient solution-phase synthesis of N-methyl-N-nosyl-alpha-amino acids and N-Fmoc-N-methyl-alpha-amino acids. This represents a very important application in peptide synthesis to obtain N-methylated peptides in both solution and solid phase. The developed methodology involves the use of N-nosyl-alpha-amino acids with the carboxyl function protected as a phenacyl ester and the methylating reagent diazomethane. An important aspect of this synthetic strategy is the possibility to selectively deprotect the carboxyl function or alternatively both amino and carboxyl moieties by using the same reagent with a different molar excess and under mild conditions. Furthermore, the adopted procedure keeps unchanged the acid-sensitive side chain protecting groups used in Fmoc-based synthetic strategies.

A Photolabile Linker for the Mild and Selective Cleavage of Enriched Biomolecules from Solid Support

Selective release of enriched biomolecules from solid support is a desirable goal in proteomic and metabolomic studies. Here we demonstrate that photocleavage of a light-sensitive phenacyl ester bond is a suitable alternative cleavage strategy for the selective release of enriched biomolecules form avidin beads circumventing the disadvantages of conventional heat denaturation procedures.

Comprehensive supercritical fluid chromatography × reversed phase liquid chromatography for the analysis of the fatty acids in fish oil

The separation of the phenacyl esters of the fatty acids originating from a fish oil extract by means of a comprehensive analysis using silver-ion (SI) supercritical fluid chromatography (SFC) and reversed phase liquid chromatography (RP-LC) in the first and second dimensions, respectively, is described. The combination ensured a high orthogonality and peak capacity, particularly when compared to the comprehensive RP-LC × 2RP-LC separation achieved by using a configuration with two columns in parallel in the second dimension. The construction of the SI-SFC × RP-LC interface consists of two two-position/ten-port switching valves, of which one is equipped with two loops packed with octadecyl silica (ODS) particles. Compared to the SFC × RP-LC configuration described in an earlier publication, the peak capacity in the second dimension was increased. Water was not only added as make-up fluid to the SFC effluent to ensure analyte focusing, but also as rinsing medium of the loops prior to the transfer of the fractions to the second dimension. In the SFC dimension, high efficiency and loadability were obtained by coupling two wide-bore columns (4.6 mm ID) in series. Evaporative light scattering (ELSD) and ultraviolet (UV) detection with standard and high-pressure flow cells were evaluated in terms of data acquisition speed and suppression of signal interferences originating from the supercritical carbon dioxide (CO 2) expansion.