A general strategy for selective detection of hypochlorous acid based on triazolopyridine formation

Triazolopyridines are an important kind of fused-ring compounds. A HOCl-promoted triazolopyridine formation strategy is reported here for the first time in which hypochlorous acid (HOCl) mildly and efficiently promotes the formation of 1,2,4-triazolo[4,3-a]pyridines NT1-NT6 from various 2-pyridylhydrazones N1-N6. N6, a rhodol-pyridylhydrazone hybrid, was developed into a fluorescent probe for the selective detection of HOCl, and successfully applied to probe endogenous HOCl in living cells and zebrafish in situ and in real time. The present intramolecular cyclization reaction is selective and atom-economical, thereby not only providing an important approach for the convenient synthesis of triazolopyridines, but also offering a general strategy for sensitive, selective and biocompatible detection of endogenous HOCl in complex biosystems.     

Development of a new fluorescence ratiometric switch for endogenous hypochlorite detection in monocytes of diabetic subjects by dye release method

Increased oxidative stress in metabolic complications like type 2 diabetes, dyslipidemia and cardiovascular disorders exerts potential health hazards in many facets. Enhanced production of reactive oxygen species (ROS) due to increased oxidative stress promotes the damage of many biologically important macromolecules. Hypochlorous acid (HOCl), a microbicidal agent is also known to be an important ROS sub-species. An enhanced generation of endogenous HOCl due to diseased condition therefore can be detrimental to health. In present work, a new quinoline-diaminomaleonitrile based probe (HQMN) has been designed for the selective detection of hypochlorite. The probe in hand shows a selective ratiometric emission change towards OCl^?. The probe behaves as a highly selective and sensitive tool for the detection of OCl^? over other analytes with a fast response time (within 100 s). Bioimaging study revealed that HQMN can detect endogenous OCl^? in human monocytes and an increase in endogenous HOCl concentration has been witnessed in diabetic condition compared to healthy control. Thus HQMN can be used as an excellent fluorescent probe for dynamic tracking of hypochorite in living biological cells especially to identify diabetic conditions.     

Hypochlorous acid turn-on boron dipyrromethene probe based on oxidation of methyl phenyl sulfide

A boron dipyrromethene (BODIPY)-based fluorometric probe, HCS, has been successfully developed for the highly sensitive and selective detection of hypochlorous acid (HOCl). The probe is based on the specific HOCl-promoted oxidation of methyl phenyl sulfide. The reaction is accompanied by a 160-fold increase in the fluorescent quantum yield (from 0.003 to 0.480). The fluorescent turn-on mechanism is accomplished by suppression of photoinduced electron transfer (PET) from the methyl phenyl sulfide group to BODIPY. The fluorescence intensity of the reaction between HOCl and HCS shows a good linearity in the HOCl concentration range 1–10 μM. The detection limit is 23.7 nM (S/N = 3). In addition, confocal fluorescence microscopy imaging using RAW264.7 macrophages demonstrates that the HCS probe could be an efficient fluorescent detector for HOCl in living cells.     

A ratiometric fluorescent probe based on boron dipyrromethene and rhodamine Förster resonance energy transfer platform for hypochlorous acid and its application in living cells

We have developed a ratiometric fluorescent probe BRT based on boron dipyrromethene (BODIPY) and rhodamine-thiohydrazide Förster resonance energy transfer (FRET) platform for sensing hypochlorous acid (HOCl) with high selectivity and sensitivity. The probe can detect HOCl in 15 s with the detection limit of 38 nM. Upon mixing with HOCl the fluorescence colour of probe BRT changed from green to orange. Moreover, probe BRT was applied to successfully monitor HOCl in living RAW 264.7 cells.     

Spectroscopic studies of a BINAM-based sensor: Highly selective fluorescent recognition of lysine in water solution through a nucleophilic substitution reaction

A BINAM-based compound (R)-1 is found to show significant fluorescence enhancement in the presence of Lys in aqueous solution (1%DMF). This probe achieves highly selective fluorescent recognition of Lys even in the presence of other natural amino acids. It can be used as a sensitive as well as selective fluorescent probe for Lys. The mechanism for the interaction of (R)-1 with Lys was studied by NMR and HRMS.     

A highly selective turn-on fluorescent probe for hypochlorous acid based on hypochlorous acid-induced oxidative intramolecular cyclization of boron dipyrromethene-hydrazone

A BODIPY-based fluorescent probe, HBP, was developed for the detection of hypochlorous acid based on the specific hypochlorous acid-promoted oxidative intramolecular cyclization of heterocyclic hydrazone in response to the amount of HOCl. The reaction is accompanied by a 41-fold increase in the fluorescent quantum yield (from 0.004 to 0.164). The fluorescence intensity of the reaction between HOCl and HBP is linear in the HOCl concentration range of 1–8 μM with a detection limit of 2.4 nM (S/N = 3). Confocal fluorescence microscopy imaging using RAW264.7 cells showed that the new probe HBP could be used as an effective fluorescent probe for detecting HOCl in living cells.     

Sensitive and selective off-on rhodamine hydrazide fluorescent chemosensor for hypochlorous acid detection and bioimaging

A rhodamine 6G hydrazide fluorescent chemosensor was prepared for the rapid HOCl detection in aqueous media. The system makes good use of the irreversible HOCl-mediated selective oxidation reaction to generate fluorescent response proportional to the amount of HOCl in neutral buffer. This probe exhibits great photostability, high sensitivity, and good selectivity for HOCl over other reactive species and most of the common metal ions. Furthermore, the probe is cell membrane permeable, and its applicability has been successfully demonstrated for fluorescence imaging of both exogenous and endogenous HOCl within living cells. Cytotoxicity assays prove that this probe is almost nontoxic to the cultured cell lines under the experimental conditions.     

A highly selective chemodosimeter for the rapid detection of Hg2+ ions in aqueous media

A novel S,S′-diallyl carbohydrazonodithioate derivative 3 of rhodamine B hydrazone was developed as a chemodosimeter for selective detection of mercury ions based on Hg²⁺ promoted cyclization. The allyl groups of 3 play a key role in the binding and selection of Hg²⁺ ions. The probe responds selectively to Hg²⁺ over various other competitive cations with marked chromo- and fluorogenic changes. The formation of stable oxadiazole derivative 8 was a strong driving force for this high selectivity. Practically, this probe is more promising because of the remarkable high selectivity, faster response, low detection limit, and aqueous solubility of 3.     

Selective Detection and Visualization of Exogenous/endogenous Hypochlorous Acid in Living Cells using a BODIPY‐based Red‐emitting Fluorescent Probe

Herein, a red‐emitting fluorescent probe DM‐BDP‐OCl containing a para‐DMTC benzyl pyridinium moiety at the meso position of BODIPY as self‐immolative portion for the detection of HOCl was designed and synthesized. DM‐BDP‐OCl exhibited excellent specificity and a fast response for HOCl beyond other ROS/RNS. It was used for the accurately measurable detection of HOCl with a linear range from 0 μM to 50 μM, and the detection limit for HOCl reached 60 nM. Moreover, the probe could directly monitor fluctuations of exogenous and endogenous HOCl in living HeLa and RAW 264.7 cells. This work provided a powerful and convenient imaging tool for probing pathological and physiological actions of HOCl.     

Development of a BODIPY-based ratiometric fluorescent probe for hypochlorous acid and its application in living cells

A BODIPY-based ratiometric fluorescent probe for HOCl has been designed based on the transduction of thioether to sulfoxide function. This probe features a marked absorption and emission blue-shift upon the HOCl-promoted rapid transduction, enabling the highly selective and ratiometric detection. In addition, the probe works excellently within a wide pH range of 4–10, addressing the existing pH dependency issue. Living cells studies demonstrate that the probe is cell membrane permeable and can be employed successfully to image endogenous HOCl generation in macrophage cells.     

Imidazo[1,5-α]pyridine-derived fluorescent turn-on probe for cellular thiols imaging with a large Stokes shift

We presented the design, synthesis and preliminary evaluation of the 2,4-dinitrobenzenesulfonate (DNBS) of imidazo[1,5-α]pyridine derivative, NIPY-DNBS, as a turn-on fluorescent probe for the detection of thiols in aqueous solution. The reaction mechanism was confirmed by means of fluorescence, absorption and HRMS. The large Stokes shift (201 nm), high sensitivity (the detection limit for Cys was calculated to be as low as 0.17 μM) and fast response (10 min) of NIPY-DNBS make it a practical and reliable method for fluorescence imaging. Furthermore, application of NIPY-DNBS for the selective detection of intracellular thiols has been successfully demonstrated in living A549 cells.     

Constrained cycloalkyl analogues of glutamic acid: stereocontrolled synthesis of (+)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740) and its 6-phosphonic acid analogue

A new stereocontrolled synthesis of (+)-2-aminobicyclo[3.1.0]hexane-2.6-dicarboxylic acid (LY354740) 1, a potent and selective 2mGluR agonist, has been accomplished in four steps with an overall yield of 27% starting from the enantiopure (+)-(R)-2-(p-tolylsulfinyl)cyclopent-2-enone 3. The key steps include asymmetric cyclopropanation of 3 with (dimethylsulfuranylidene)acetate (EDSA) and removal of the chiral p-tolylsulfinyl auxiliary from the cycloadduct ent-4c upon treatment with iso-propylmagnesium chloride. The stereoselective hydantoin formation from the bicyclic ketone 6 formed (Bucherer–Bergs reaction) and subsequent hydrolysis completed the synthesis of 1. The same reaction sequence has been applied in the first synthesis of enantiopure (+)-2-amino-6-phosphonobicyclo[3.0.1]hexane-2-carboxylic acid 2, a structural 6-phosphono analogue of 1. The starting bicyclic ketophosphonates 9–11 have been obtained by asymmetric cyclopropanation of (?)-(S)-3 with phosphoryl sulfonium ylides, producing only two endo-isomers. The major endo-isomer (+)-11a containing the 6-diisopropoxyphosphoryl group has been converted in three steps into (+)-endo-2 in 46% overall yield.     

Simultaneous stereoselective 4-amination with cyclic secondary amines and 2-O-deacetylation of peracetylated sialic acid derivatives

Treatment of methyl 5-acetamido-2,4,7,8,9-penta-O-acetyl-3,5-dideoxy-β-l-glycero-d-galacto-2-nonulopyranosidonate (1) with cyclic secondary amines in pyridine at room temperature for 24 h afforded unusual products (2a-g). Related experiments were carried out to explain the formation of 4-amination and 2-O-deacetylation of peracetylated sialic acid derivatives (2a-g). This reaction may provide a new strategy for the preparation of Zanamivir analogues as neuraminidase inhibitors for anti-H5N1 subtype of avian influenza virus (AIV).     

Synthesis of UDP-GalNAc analogues as probes for the study of polypeptide-α-GalNAc-transferases. Part 2

The synthesis of four UDP-GalNAc analogues (1-4) is described. The 3-, 4- and 6-deoxygenated analogues 1-3 were obtained by way of a divergent strategy starting from a 3,6-di-O-pivaloyl GlcNAc derivative as a common precursor. Analogue 4 bearing a N-trifluoroacetamido group was prepared from the trifluoromethylated oxazoline 24 as key intermediate. These compounds were designed to probe the substrate specificity of polypeptide-α-GalNAc transferases.     

The enzymatic glucuronidation of 3-O-protected morphine—a new route to 7,8-dihydromorphine-6-glucuronide

The selective enzymatic glucuronidation of the 6-position of morphine was probed by the glucuronidation of a series of 3-O-protected morphine derivatives. 3-O-Benzylmorphine 5 was converted to the corresponding 3-O-benzylmorphine-6-glucuronide 8 by human liver microsomes. The identity of 8 was confirmed by independent chemical synthesis. The glucuronide 8 was subsequently converted to 7,8-dihydromorphine-6-glucuronide.     

Stereoselective alkylation of C2-symmetric chiral N-phthaloylglycinamides in the preparation of enantiopure α-amino acids

The novel, chiral glycinamides (S,S)-3 and (S,S)-4 were prepared in good yields from C₂-symmetric chiral amines (S,S)-1 and (S,S)-2, respectively. Enolate formation and addition to methyl iodide and benzyl bromide proceeded in good yield and high diastereoselectivity, especially in the presence of LiCl or DMPU. Removal of the phthaloyl protecting group with hydrazine, followed by hydrolysis with 6N HCl, converted the benzylated product (S,S,S)-7 to enantiopure (S)-phenylalanine.     

The first enantioselective synthesis of imino-deoxydigitoxose and protected imino-digitoxose by using l-threonine aldolase-catalyzed aldol condensation

The first enantioselective synthesis of protected imino-digitoxose (?)-16 was attained starting with a synthetic intermediate of polyoxin C prepared by the l-threonine aldolase-catalyzed aldol condensation of (2S,3S)-2,3-O-isopropyriden-4-penten-1-al 8 and glycine. The strategy took advantage of an intramolecular nucleophilic attack by a Cbz-protected amino group on the hemiacetal carbon, a side reaction in the synthesis of natural products, for the formation of the piperidine ring of the imino-sugar. Imino-deoxydigitoxose (+)-18 was also synthesized from (?)-16 by reduction and acid hydrolysis.     

First synthesis of nitro-substituted 2,2-diphenyl-2H-1-benzopyrans via the ipso-nitration reaction

The first synthesis of a series of nitro-substituted 2,2-diphenyl-2H-1-benzopyrans is reported. Our synthetic approach is based on a linear synthesis in two steps from appropriate brominated 2,2-diphenyl-2H-1-benzopyrans 12-17, which requires the preliminary preparation of bromophenols 7-11. These latter were easily obtained by the reaction of phenols 1-5 with a mild and selective brominating agent tetrabutylammonium tribromide (TBA·Br₃). The key intermediates 12-17 were efficiently elaborated through an univocal classic chromenization between the commercially available 1,1-diphenyl-2-yn-1-ol and the brominated phenols 6-11. The compounds 12-17 so obtained were converted into arylboronic acids 18-23 by a metalation/boronylation sequence, followed by acid hydrolysis. From advanced building blocks 18-23, the introduction of nitro group, which constitutes the ultimate step of our strategy, was achieved by an ipso-nitration reaction using the Crivello's reagent. This highly selective method provides only the ipso-nitrated products 24-29 in moderate to high yield.     

Reactions of β-substituted amines—III: Complete product study of the reactions of 3-chloro-1-ethylpiperidine and 2-chloromethyl-1-ethylpyrrolidine with hydroxide ion

A complete product study of the reaction of 3-chloro-1-ethylpiperidine (1) in aqueous sodium-hydroxide showed the formation of three bis-(β-aminoethers), 2,2′-bis-(N-ethyl-2-pyrrolidinomethyl) ether (6), 3-(N′-ethyl-2′-pyrrolidinomethoxy)-N-ethylpiperidine (7) and 3,3′-bis-N-ethyl-3-piperidinyl ether (8) in addition to the two previously reported products. The structures of these three ethers are demonstrated and their formation is shown to be consistent with the previously suggested mechanism¹ of the reaction. Reactions of 2-chloromethyl-1-ethylpyrrolidine (2) are shown to proceed by the same mechanistic pathways with the formation of the same products. Reaction of 1 or 2 with two other nucleophiles, methoxide and ethoxide are also examined.     

Orbital control of stereochemistry in acid-catalysed addition reactions of endo -tricyclo[3.2.1.02,4]0ct-6-ene

The reaction of endo-tricyclo[3.2.1.0^2,4]oct-6-ene 1 with methanol in the presence of catalytic amounts of toluene-p-sulphonic acid has been shown to give 2-exo- and endo-methoxybicyclo[3.2.1]oct-3-ene (2c) and (2d) and 2-endo-methoxybicyclo[3.2.1]oct-6-ene (13). The formation of 2-exo- methoxybicyclo[3.2.1]oct-3-ene (2c), the major product of reaction, has been probed by deuterium labelling experiments and a series of 6-exo-7-exo- dideuterobicyclo[3.2.1]oct-3-enes synthesised for ²H, ¹H and ¹³C NMR spectral analysis in order unambiguously to determine the stereochemistry of proton attack on endo-tricyclo[3.2.1 0^2,4]oct-6-ene (1). The formation of 2-exo-methoxybicyclo[3.2.1]oct-3-ene (2c) has been determined to involve corner protonation of the cyclopropyl moiety and skeletal rearrangement to an allylic cation with a small but measurable memory effect