Proton and metal‐ion activation of C–H exchange in five‐membered azoles

Factors influencing C–H isotopic exchange rates in five‐membered azoles, that is imidazoles and thiazoles, under catalysis by H⁺ and Mⁿ⁺, especially transition metals, Pt(II) and Co(III) are discussed. Hydrogen ion catalysis through N(3) protonation of azoles 1–3 is generally the most efficient, with rate enhancements in the range 10²–10⁹ over the neutral process being attained. Metal‐ion coordination also results in effective catalysis, though less so than catalysis by protons. Catalysis of C–H exchange by Mⁿ⁺ can be studied through addition of the metal salts to a buffered solution of the heterocycle in which labile complexes exist, or on synthesized complexes such as 4–13 which are substitution‐inert thus precluding complications from unknown dissociation equilibria. A delicate balance of factors influence the ease of C–H exchange, including: (1) the magnitude of the fractional charge located at N(3) of the heterocycle through Mⁿ⁺–N(3) σ bond polarization; (2) metal‐to‐ligand π back‐bonding; (3) the electronic structure of the metal ions. These considerations have obvious consequences for deuterium‐ and tritium‐labelling of a number of biomolecules, e.g. proteins, enzymes, nucleic acids, some vitamins, as well as drugs which incorporate five‐membered azoles in their structures. Copyright © 2002 John Wiley & Sons, Ltd.     

Evaluation of the efficiency of Pd/H2‐catalyzed benzylic H/D exchange of dehydroabietinal with D2O and synthesis of a tritium‐labeled analogue

Dehydroabietinal (DA) has been identified as an important signaling molecule in systemic acquired resistance in plants. Deuterium and tritium‐labeled DA were synthesized to confirm its role in signaling and to further elucidate the mechanism by which DA induces systemic acquired resistance. Pd/H₂‐catalyzed exchange of benzylic hydrogen atoms of DA with ²H‐H₂O or ³H‐H₂O was conducted with >97% label incorporation for ²H‐DA and a specific activity of 12.6 mCi/mmol for ³H‐DA synthesized from 90 mCi/mmol ³H‐H₂O. The extent of deuterium labeling at each benzylic position was determined via an inverse‐gated ¹³C NMR experiment. C7 and C15 were 87% and 81% labeled, respectively. Isotope‐induced chemical shift changes at C6 were used to approximate the amount of singly (66%) and doubly (17%) labeled ²H‐DA at C7. Results also indicated that two of the three benzylic protons in DA underwent facile exchange. Exchange at the remaining position was likely hampered by steric interactions of nearby methyl groups at the surface of the Pd catalyst.     

NMR analysis of t‐butyl‐catalyzed deuterium exchange at unactivated arene localities

Regioselective labelling of arene rings via electrophilic exchange is often dictated by the electronic environment caused by substituents present on the aromatic system. Previously, we observed the presence of a t‐butyl group, either covalently bond or added as an external reagent, could impart deuterium exchange to the unactivated, C1‐position of estrone. Here, we provide nuclear magnetic resonance analysis of this exchange in a solvent system composed of 50:50 trifluoroacetic acid and D₂O with either 2‐t‐butylestrone or estrone in the presence of t‐butyl alcohol has shed insights into the mechanism of this t‐butyl‐catalyzed exchange. Fast exchange of the t‐butyl group concurrent with the gradual reduction of the H1 proton signal in both systems suggest a mechanism involving ipso attack of the t‐butyl position by deuterium. The reversible addition/elimination of the t‐butyl group activates the H1 proton towards exchange by a mechanism of t‐butyl incorporation, H1 activation and exchange, followed by eventual t‐butyl elimination. Density functional calculations are consistent with the observation of fast t‐butyl exchange concurrent with slower H1 exchange. The σ‐complex resulting from ipso attack of deuterium at the t‐butyl carbon was 6.6 kcal/mol lower in energy than that of the σ‐complex resulting from deuterium attack at C1. A better understanding of the t‐butyl‐catalyzed exchange could help in the design of labelling recipes for other phenolic metabolites.     

H N.M.R. STUDIES OF PROTECTED α-AMINOISOBUTYRIC ACID CONTAINING PEPTIDES

The benzylic methylene protons in a large number of benzyloxycarbonyl α-aminoisobutyric acid (Z-Aib) containing peptides, show chemical shift nonequivalence. The magnitude of the geminal nonequivalence is correlated with the involvement of the urethane carbonyl group, in an intramolecular hydrogen bond. Studies of the model compounds Z-Aib-Aib-Ala-NHMe, and Z-Aib-Aib-Aib-Pro-OMe clearly establish the presence of intramolecular hydrogen bonds, involving the urethane CO group. In both compounds marked anisochrony of the benzylic methylene protons is demonstrated. In Z-Aib-Aib-Pro-OMe, where a 4 → 1 hydrogen bonded β-turn is not possible, the benzylic -CH₂- protons appear as a singlet in CDCl₃ and have a very small chemical shift difference in (CD₃)₂SO. The observation of such nonequivalence is of value in establishing whether the amino terminal Aib-Pro β-turn is retained in large peptide fragments of alamethicin.     

Practical approaches to labelling terminal alkynes with deuterium

Base catalysed exchange with sodium hydroxide, calcium oxide or N,N,N,N-tetramethylguanidine in deuterium oxide is a viable procedure for the preparation of terminally deuterated alkynes for those alkynes stable to strong base. The use of silver perchlorate as a catalyst is an alternative practical option when labelling alkynes which are sensitive to base or contain functionalities which would lead to labelling elsewhere in the molecule. Labelling with this catalyst takes place smoothly at ambient temperature in a mixture of N,N-dimethylformamide and deuterium oxide.     

lonization and Solubilization of 4 Alkyl Benzole Acids and 4 Alkyl Anilines in Sodium Taurodeoxycholate Solutions

Purpose. The aqueous solubility and the extent of solubilization and ionization constant in sodium taurodeoxycholate (NaTDC) solutions of a series of benzoic acid and aniline derivatives were measured as a basis to characterize and thereby help predict the nature of the interaction of drugs with bile aggregates. Methods. The aqueous solubility and the solubilization of two series of compounds, 4-alkyl benzoic acids and 4-alkyl anilines, was measured as a function of NaTDC in 0 and 150 mM NaCl. The ionization constants were determined in water and in 50 mM NaTDC at sodium chloride concentrations of 0, 75 and 150 mM by spectrophotometric titration. The diffusion coefficients of NaTDC and the solutes were measured by pulsed-field gradient spin echo NMR spectroscopy. Results. The aqueous solubilities decreased with increasing alkyl chain length in both series, and the aniline derivatives had larger solubilities than the benzoic acid derivatives. The number of moles of solute solubilized per mole of bile salt ranged from 0.17 to 0.31 for the benzoic acid derivatives and from 1.3 to 3.0 for the aniline derivatives. The pKa values of the benzoic acid derivatives in the presence of NaTDC were higher relative to the controls, and the difference in the pKa (pKa,obs) increased with increasing chain length. With the aniline derivatives, the pKa values were also shifted to higher values in NaTDC relative to the control but only in the absence of salt. The presence of the solute caused a decrease in the diffusion coefficient of NaTDC, and the diffusion coefficients of the solutes decreased with increasing alkyl chain length. With the hexyl derivatives, the diffusion coefficient of the solute was smaller than the diffusion coefficient of the bile salt. The chemical shift of the protons attached to carbon 18 and 19 of the bile salt were decreased to a greater extent in the presence of the solutes than the protons attached to carbon 26. Conclusions. Both the solubilization and ionization behavior of solutes were affected by the presence of bile salt aggregates. The surface potential and effective polarity of NaTDC aggregates were found to be dependent on the alkyl chain length for these two homologous series of solutes. The solubilization ratio was largely independent of alkyl chain length, but the unitary partition coefficient was dependent on both the alkyl chain length as well as ionization state. The derivatives reduced the diffusivity of the micelles suggesting the formation of larger sized aggregates and the solutes (hexyl derivatives) appear to favor association with the larger sized aggregates. The phenyl ring of the solutes appears to be oriented parallel to the plane of the steroid frame with preferential positioning near the hydrophobic rings.     

[18F]/19F exchange in fluorine containing compounds for potential use in 18F‐labelling strategies

Exchange of [¹⁸F]fluoride with ¹⁹F in various organofluorine compounds in concentrations ranging from 0.06 to 56 mM was explored. We aimed to explore whether exchange reactions can be a potential useful labelling strategy, when there are no requirement of high specific radioactivity. Parameters such as solvents, temperature, conventional vs microwave heating, and the degree of fluorine load in some aromatic and alkyl compounds were investigated with regard to radiochemical yield and specific radioactivity. A series of fluorobenzophenones (1–6), 1‐(4‐fluorophenyl)ethanone (7), various activated and deactivated fluoro benzenes (8–16), N‐(pentafluorophenyl)benzamide (17), (pentafluorophenyl)formamide (18), (tridecafluorohexyl)benzene (19) and tetradecafluorohexane (20) were subjected to [¹⁸F]/¹⁹F exchange. To test this strategy to label biologically active molecules containing fluorine atoms in an aryl group, two analogues of WAY‐100635 (21–22), Lapatinib (23), 2,5,6,7,8‐pentafluoro‐3‐methylnaphthoquinone (24) and 1‐(2,4‐difluorophenyl)‐3‐(4‐fluorophenyl)propan‐1‐one (25) were investigated. The multi‐fluorinated molecules containing an electron‐withdrawing group were successfully labelled at room temperature, whereas the monofluorinated, as well as those containing an electron‐donating group, required heating for the exchange reaction to take place. Copyright © 2009 John Wiley & Sons, Ltd.     

Convenient and efficient deuteration of functionalized aromatics with deuterium oxide: catalysis by cycloocta‐1,5‐dienyliridium(I) 1,3‐dionates

Aromatic compounds bearing an ortho‐directing substituent may be deuterated by exchange with deuterium oxide in the presence of a range of cycloocta‐1,5‐dienyliridium(I)1,3‐dionate catalysts. The exchange takes place in several dipolar aprotic solvents and is directly applicable to the deuteration of polar compounds. Isotope incorporation is efficient and regiospecific. The method is applicable to a wide range of ortho‐directing groups some of which are only weak directors for alternative ortho‐labelling approaches. In addition, the application of microwaves enables labelling within minutes even with sub‐stituents which are poor directors. Copyright © 2003 John Wiley & Sons, Ltd.     

INTERACTION OF CLONIDINE,ITS METHYLENE-BRIDGED ANALOG,St 1913, AND THE BENZYLIC HYDROXYL-SUBSTITUTED DERIVATIVE,St 1965, WITH α1- AND α2-ADRENORECEPTORS

• 1 The effects of benzylic hydroxyl substitution on the activity of a close structural analog of clonidine was assessed at α₁- and α₂-adrenoreceptors both in vitro and in vivo in order to uncover possible differences that this substitution may have on the effects of imidazolines and phenethylamines at adrenoreceptors.
• 2 In all test systems, the presence of the benzylic hydroxyl group was associated with a consistent and marked decrease in activity. These findings are in agreement with our previous studies with imidazolines having different pharmacological profiles and different physicochemical properties than the clonidine derivatives reported herein.
• 3 We conclude, therefore, that the deleterious effects of the benzylic hydroxyl group is ubiquitous among imidazolines and, more importantly, is in marked contrast to the 100–1000 fold enhancement in activity that the benzylic hydroxyl substituent (i.e. β-hydroxyl group) produces for the phenethylamines.
• 4 The results support the concept that imidazolines and phenethylamines may interact differently with α-adrenoreceptors.

     

Application of 1‐butyl‐3‐methylimidazolium hexafluorophosphate to Ir(I)‐catalyzed hydrogen isotope exchange labelling of substrates poorly soluble in dichloromethane

Substrate solubility remains a major limitation in Ir(I)‐catalyzed isotopic hydrogen exchange labelling. In the search for an alternative to the solvent dichloromethane, which is critical to the success of the reaction, we examined a series of ionic liquids for their suitability. Commercially available 1‐butyl‐3‐methylimidazolium hexafluorophosphate (abbreviated to [BMI][PF₆]) was found to support efficient deuterium and tritium exchange labelling of N‐(4‐methoxyphenyl)‐N‐methyl benzamide 1 under standard conditions. The solvent dissolves both polar hydroxyl and carboxylic acid substituted acetanilides, providing isotopomers in unprecedentedly high deuterium incorporation as compared to dichloromethane. We report the application of [BMI][PF₆] and its potential for extending the scope of Ir(I)‐catalyzed H/T exchange to more polar compounds. Copyright © 2003 John Wiley & Sons, Ltd.     

Synthesis and Inhibition of Human Leucocyte Elastase by Functionalized N-Aryl Azetidin-2-ones: Effect of Different Substituents on the Aromatic Ring

N-aryl-3,3-difluoroazetidin-2-ones featured by a latent electrophilic methylene quinoniminium function have been synthesized and evaluated as inhibitors of human leucocyte elastase. To promote hydrophobic interactions with the enzyme, to increase the rates of β-lactam ring opening and of benzylic group departure, or to induce hydrosolubility, these compounds incorporate on their aromatic ring either an alkyl moiety, a methoxy substituent or a carboxylic group. Some of these β-lactams proved to be good inactivators of human leucocyte elastase.     

Homogeneous deuteriodeiodination of iodinated tyrosine in angiotensin‐I using synthesized triethyl[2H]silane and Pd(0)

In our efforts to develop new reactions for the efficient labelling of peptides and proteins with tritium, we now report the use of silane hydrides together with homogenous Pd(0) catalysis for the protio‐ and deuteriodeiodination of an o‐iodo‐tyrosine containing peptide (angiotensin‐I) performed at room temperature. Copyright © 2010 John Wiley & Sons, Ltd.     

Syntheses of novel N‐([18F]fluoroalkyl)‐N‐nitroso‐4‐methyl‐benzenesulfonamides and decomposition studies of corresponding 19F‐ and bromo‐analogues: potential new compounds for the 18F‐labelling of radiopharmaceuticals

N‐([¹⁸F]fluoroalkyl)‐N‐nitroso‐4‐methyl‐benzensulfonamides [n‐alkyl = (?CH₂)[^18,19F]F, n=2–4)] were synthesized in radiochemical yields ranging from 75–90% to provide new secondary labelling precursors for the syntheses of ¹⁸F‐labelled compounds. Preliminary decomposition studies utilizing the corresponding non‐radioactive ¹⁹F‐compounds as well as the bromo containing analogous compounds were performed to evaluate their stability regarding temperature and basicity of the labelling medium. Furthermore, initial difficulties in labelling these compounds led to the development of a modified labelling procedure applying nearly solvent‐free conditions. Extensive decomposition experiments of the new fluoro‐ as well as the bromo‐compounds were conducted under various conditions in order to get experimental data about their stability and reactivity. As a result, different trends for the stability of the bromo as well as the fluoro compounds could be observed. Copyright © 2003 John Wiley & Sons, Ltd.     

Preparation via coligand exchange and characterization of [99mTc‐EDDA‐HYNIC‐D‐Phe1,Tyr3]Octreotide (99mTc–EDDA/HYNIC–TOC)

[^99mTc‐EDDA–HYNIC‐D‐Phe¹,Tyr³]octreotide (^99mTc‐EDDA/HYNIC–TOC) is a promising new agent with the potential to replace [¹¹¹In‐DTPA‐D‐Phe¹]‐octreotide in somatostatin receptor scintigraphy. This hydrazinonicotinic acid derivatized somatostatin complex contains ethylenediamine N,N′ diacetic acid (EDDA) as a coligand resulting in a high in vitro and in vivo stability. Since direct ^99mTc‐labelling of HYNIC–TOC with EDDA results in low labelling yields, in this study we describe the preparation of ^99mTc‐EDDA/HYNIC‐TOC via coligand exchange from Tricine for EDDA. Exchange of coligands is achieved at elevated temperature and under optimized conditions of pH, EDDA concentration and stannous ion. High labelling yields (mean 95.9%) were achieved at high specific activities (>37GBq/µmol). Characterization via HPLC, receptor binding and LC–MS of the resulting complex is described. The formulation developed enables rapid and simple labelling of ^99mTc‐EDDA/HYNIC–TOC in a manner suitable for a clinical setting. Copyright © 2003 John Wiley & Sons, Ltd.     

Labelling of ceftriaxone with 99mTc and its bio‐evaluation as an infection imaging agent

Differentiation of bacterial and sterile inflammation will have a significant impact on the current clinical practice. Ceftriaxone (CTRX) was labelled with ^99mTc and assessed for its ability to depict infection on scintigraphy. Stoichiometry was performed to optimize labelling parameters. Stability and bacterial binding was verified and biodistribution pattern was seen in normal, infected/inflamed animal models. ^99mTc‐CTRX prepared at pH 7 with stannous chloride of 50 µg, ligand of 30 mg, and boiling for 10 min gave labelling yield of 96.2 ± 0.2% with good stability. In vitro binding was higher for Escherichia coli than Staphylococcus aureus. Biodistribution in normal rats showed high uptake in hepatobiliary system, gut and urinary system. In animal models induced with infection or inflammation, lesion to normal ratios at 4 h were 2.36 ± 0.21, 12.66 ± 1.44 and 1.40 ± 0.01 with S. aureus infection, E. coli infection and turpentine oil inflammation, respectively. Infection specificity especially for E. coli was also confirmed on scintigraphic findings. Ceftriaxone can be labelled with ^99mTc with high labelling yield at pH compatible with that of blood. Our preparation has shown stability in vitro and in human serum, and binds preferentially with bacteria. ^99mTc‐CTRX scintigraphy can be used to delineate sites of active infection and to differentiate infection and inflammation.     

Physicochemical Characterization and Solubility Analysis of Thalidomide and Its N-Alkyl Analogs

Purpose. The present study was primarily aimed at exploring the feasibility of improving percutaneous delivery via chemical manipulation of the thalidomide molecule to form analogs with improved physicochemical properties. N-Alkyl analogs were synthesized with the belief that these would be suitably hydrophobic and far less crystalline than the reference compound. This article presents their physicochemical properties. Methods. Thalidomide and three of its N-alkyl analogs were synthesized. Identification and levels of purity (>96%) were assured through element analysis, fast atom-bombardment mass spectrometry, nuclear magnetic resonance spectroscopy, and high-performance liquid chromatography. N-Octanol/water partition coefficients were determined at pH 6.4. Solubilities in water and a series of n-alkanols were obtained. Best-fit solubility parameters were determined from the solubilities of the respective compounds in London solvents and were also calculated from respective hexane solubilities, melting points and heats of fusion. Results. Methylation of the thalidomide molecule at its acidic nitrogen led to an aqueous solubility about 6-fold higher than thalidomide but, because the alkyl chain length was further extended from methyl to pentyl, aqueous solubilities decreased essentially exponentially. The destabilization of the crystalline structure with increasing alkyl chain length led to an increased solubility in nonpolar media. The log partition coefficient increased linearly with increasing alkyl chain length and the solubility parameters declined systematically through this series. By adding a methyl group to the thalidomide structure, the melting point dropped by more than 100°C. Adding to the alkyl chain length led to further, more modest decreases. Heats of fusion decreased dramatically upon thalidomide's alkylation as well. Conclusion. Alkylation of the thalidomide molecule resulted in compounds with physicochemical properties that appear to be markedly better suited for percutaneous delivery.     

Comparison of the organization of t-tubules, sarcoplasmic reticulum and ryanodine receptors in rat and human ventricular myocardium

1. It is apparent from the literature that there are significant differences in excitation–contraction coupling between species, particularly in the density of calcium transporting proteins in the t‐system and sarcoplasmic reticulum (SR) Ca²⁺ release channels. Unfortunately, there is a lack of information as to how the principal structures that link electrical excitation to the activation of calcium‐induced calcium release (CICR) are different between human and animal models (particularly rat). 2. Comparison of wheat germ agglutinin and caveolin‐3 labelling revealed a non‐uniform distribution of surface membrane glycosylation in the rat, rabbit and human, and that the rat t‐system appeared more complex in geometry than the latter species. Analysis of the t‐system skeleton showed that the t‐system was highly branched in the rat compared with that of the human (0.8 ± 0.08 and 0.2 ± 0.07 branch points per μm², respectively; P < 0.001). 3. We also compared the distribution of contractile machinery, sodium–calcium exchange, SR and ryanodine receptors (RyR) in rat and human. F‐Actin and RyR labelling was used to estimate the area of contractile apparatus supplied by each RyR cluster. In the rat, each RyR cluster supplied an average cross‐sectional area of contractile machinery of 0.36 ± 0.03μm² compared with 0.49 ± 0.04 μm² in human (P = 0.048). Sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA2a) labelling showed that the SR formed a tight network of loops surrounding contractile fibrils that were denser than the t‐tubule network, but otherwise appeared similar in both species. 4. In general, the results show a higher density in structures involved in CICR in the rat compared with human.     

Isobornyl acrylate: an impurity in alkyl glucosides

Context: Alkyl glucosides and alkyl poly-glucosides are widely used as wetting agents, surfactants and emulsifiers in several industrial and cosmetic products. They are known as well-tolerated and are usually added to the primary surfactants in order to reduce the irritating potential of the main foaming agents.

Objective: Recently, some authors suggested that allergic contact dermatitis to alkyl glucosides might be more frequent than suspected. On the other hand, the chemical structures of glucosides do not show potentially allergenic chemical groups or strongly polarized structures. The aim of our study is to investigate alkyl glucosides carrying out a detailed chemical analysis on samples of raw materials to identify potentially allergenic impurities or by-products contained in commercial samples of alkyl glucosides.

Materials and methods: We chemically analyzed samples of cocoyl glucoside, decyl glucoside and lauryl glucoside by three different analytical methods, in order to identify any undesired or polluting substances.

Result: In each of the three samples, we detected the presence of isobornyl acrylate. Its approximate content in the tested samples is 500?ng/g of the product.

Discussion: Isobornyl acrylate is not used in the synthesis of alkyl glucosides, but as a plasticizer in many plastic materials. It can be easily released to materials flowing over these surfaces when they have high extraction power, as glucosides.

Conclusion: Isobornyl acrylate may play a role as hidden allergen, in the form of an impurity collected during the industrial process, explaining some cases of allergic reaction to alkyl glucosides.     

Time taken to produce dispensing labels in hospital pharmacies,and factors affecting it

The time taken in hospital pharmacies to produce labels for individual patients' medication was measured, and factors affecting the labelling process investigated. Labelling time was measured by direct observation using a stopclock at randomly chosen semi-stratified time periods. Four combinations of major London hospitals and computer systems were studied. The time to produce 2,167 labels was measured and 59 operators were observed. There were significant differences in average labelling time between the studied hospitals/systems (16.6 to 39.3 seconds per label). Operators' experience with their system and the occurrence of interruptions were found to affect labelling significantly (P<0.0001 in both cases). There was an overall trend for labelling time to decrease with increasing experience (P<0.0001), and interruptions added 11 to 12 seconds on average. Operator experience also affected the rate and duration of interruptions, which subsequently affected labelling time. Fewer interruptions occurred with more experienced staff (P=0.0015) and when interrupted, they took less time than inexperienced staff to complete the labelling process. A performance indicator of person-days per 100,000 labels varied from 62.3 to 147.6. Pharmacy managers should be aware that there are significant differences in performance using different labelling systems and that staff training and systems of work may have a marked effect on labelling time.     

Fast and efficient tritium labelling of the nonsteroidal anti‐inflammatory drugs naproxen,tolmetin, and zomepirac

Fast and efficient tritium labelling of the nonsteroidal anti‐inflammatory drugs naproxen, tolmetin and zomepirac is reported. Naproxen along with its (R)‐enantiomer were labelled by catalytic tritium–halogen exchange of the corresponding 5‐bromo derivatives providing [³H]naproxen with a specific activity of 25.4 Ci/mmol. Tolmetin and zomepirac were labelled by the hydrogen isotope exchange reaction using Crabtree's catalyst. This provided [³H]tolmetin and [³H]zomepirac with specific activities of 80.8 and 64.3 Ci/mmol, respectively. All compounds were obtained in high radiochemical purity (>98%). Copyright © 2005 John Wiley & Sons, Ltd.