Biotransformation of Substituted Pyridines with Dioxygenase-containing Microorganisms.

A series of 2-, 3- and 4-substituted pyridines was metabolised using the mutant soil bacterium Pseudomonas putida UV4 which contains a toluene dioxygenase (TDO) enzyme. The regioselectivity of the biotransformation in each case was determined by the position of the substituent. 4-Alkylpyridines were hydroxylated exclusively on the ring to give the corresponding 4-substituted 3-hydroxypyridines, while 3-alkylpyridines were hydroxylated stereoselectively on C-1 of the alkyl group with no evidence of ring hydroxylation. 2-Alkylpyridines gave both ring and side-chain hydroxylation products. Choro- and bromo-substituted pyridines, and pyridine itself, while being poor substrates for P. putida UV4, were converted to some extent to the corresponding 3-hydroxypyridines. These unoptimised biotransformations are rare examples of the direct enzyme-catalysed oxidation of pyridine rings and provide a novel synthetic method for the preparation of substituted pyridinols. Evidence for the involvement of the same TDO enzyme in both ring and side-chain hydroxylation pathways was obtained using a recombinant strain of Escherichia coli (pKST11) containing a cloned gene for TDO. The observed stereoselectivity of the side-chain hydroxylation process in P. putida UV4 was complicated by the action of an alcohol dehydrogenase enzyme in the organism which slowly leads to epimerisation of the initial (R)-alcohol bioproducts by dehydrogenation to the corresponding ketones followed by stereoselective reduction to the (S)-alcohols.

Raman spectroelectrochemical studies and crystal structure of a binuclear copper(I) complex with a bridging diimine ligand

Raman spectroelectrochemical and X-ray crystallographic studies have been made for the binuclear copper(I) complex, [(Ph(3)P)(2)Cu(dpq)Cu(PPh(3))(2)][BF4](2), where dpq is the bridging ligand 2,3-di(2-pyridyl)quinoxaline. The X-ray data show that the pyridine rings are twisted out of plane with respect to the quinoxaline ring which is itself non-planar. The UV/VIS spectra of the metal-to-ligand charge-transfer excited state and those of the electrochemically reduced complex are similar. The resonance-Raman spectrum of the latter species exhibits little change in the frequency of the pyridinylquinoxaline inter-ring C-C bond stretching mode, compared to the ground electronic state. This suggests minimum change in the inter-ring C-C bond order in the electrochemically or charge-transfer generated radical anion. Semiempirical molecular-orbital calculations on both the neutral dpq and radical anion show two near-degenerate lowest unoccupied orbitals in the neutral species. One is strongly bonding across the inter-ring C-C bond while the other is almost nun-bonding. The Raman data suggest that it is this latter orbital which is populated in the transient and electrochemical experiments.

Guest sorption and desorption in the metal-organic framework [Co(INA)(2)] (INA = isonicotinate) - evidence of intermediate phases during desorption

The metal-organic framework [Co(INA)(2)].0.5EtOH (INA = isonicotinate, NC5H4-4-CO2-), 1 was synthesised under solvothermal conditions. Its X-ray crystal structure shows channels containing ethanol guests which are hydrogen-bonded to carboxylate oxygens of the framework. The pyridyl rings of the framework alternate between `open' and `closed' positions along the channels resulting in large variation in the channel cross-sectional area from ca. 1.4 by 2.3 at the narrowest point to 4.9 by 5.3 at the widest. Despite the very small windows, the ethanol guests (of van der Waals diameter ca. 4.2-6.1 Angstrom) may be reversibly desorbed and sorbed into the structure quantitatively, as shown by in situ variable-temperture IR spectroscopy and XRPD. The single-crystal structure of the desolvated form [Co(INA)(2)]2 shows that there is no change in the overall connectivity on desolvation, but the rotational positions of the pyridine rings are altered. This suggests that pyridyl rotation may occur to allow guests to pass in and out. When the synthesis was conducted in 1-propanol solvent [Co(INA)(2)].0.5Pr(n)OH.H2O 3, was obtained, and a single-crystal X-ray structure revealed the same overall connectivity as in 1 but with pyridine rings disordered over closed and open positions. There was no evidence of included guests from X-ray crystallography, suggesting that they are also highly disordered. Variable-temperature XRPD performed on bulk samples showed peaks which were unsymmetrical and exhibited shoulders, suggesting that for each pattern obtained the material actually consisted of several closely-related phases. The movements of the peaks during desolvation showed the presence of intermediate phases before the final desolvated product was formed. The peak positions of the intermediate phases matched more closely with the calculated pattern for 3 than with 1 or 2, suggesting that they may have disordered structures similar to 3. The results also suggest that the intermediate phase represents an initial increase in volume before a larger decrease in volume occurs to give the final desolvated material.

Dichloridobis(picolinohydrazide)cadmium(II)

The title compound, [CdCl2(C6H7N3O)(2)], was obtained unintentionally as a product of an attempted reaction of CdCl2 center dot 2.5H(2)O and picolinic acid hydrazide, in order to obtain a cadmium(II) complex analogous to a 15-metallacrown-5 complex of the formula [MCu5L5]X-n, with M = a central metal ion, L = picolinic acid hydrazide and X = Cl- , but with cadmium the only metal present. The coordination geometry around the Cd-II atom can be considered as distorted octahedral, with two bidentate picolinic acid hydrazide ligands, each coordinating through their carbonyl O atom and amino N atom, and two chloride anions. In the crystal structure, intermolecular N-H center dot center dot center dot Cl and N-H center dot center dot center dot N hydrogen bonds link the molecules into a two-dimensional network parallel to the ( 100) plane. The pyridine rings of adjacent networks are involved in pi-pi stacking interactions, the minimum distance between the ring centroids being 3.693 (2) angstrom.

Dioxygenase-catalyzed cis-dihydroxylation of pyridine-ring systems

Toluene dioxygenase-catalyzed dihydroxylation, in the carbocyclic rings of quinoline, 2-chloroquinoline, 2-methoxyquinoline, and 3-bromoquinoline, was found to yield the corresponding enantiopure cis-5,6- and -7,8-dihydrodiol metabolites using whole cells of Pseudomonas putida UV4. cis-Dihydroxylation at the 3,4-bond of 2-chloroquinoline, 2-methoxyquinoline, and 2-quinolone was also found to yield the heterocyclic cis-dihydrodiol metabolite, (+)-cis-(3S,4S)-3,4-dihydroxy-3,4-dihydro-2-quinolone. Heterocyclic cis-dihydrodiol metabolites, resulting from dihydroxylation at the 5,6- and 3,4-bonds of 1-methyl 2-pyridone, were isolated from bacteria containing toluene, naphthalene, and biphenyl dioxygenases. The enantiomeric excess (ee) values (>98%) and the absolute configurations of the carbocyclic cis-dihydrodiol metabolites of quinoline substrates (benzylic R) and of the heterocyclic cis-diols from quinoline, 2-quinolone, and 2-pyridone substrates (allylic S) were found to be in accord with earlier models for dioxygenase-catalyzed cis-dihydroxylation of carbocyclic arenes. Evidence favouring the dioxygenase-catalyzed cis-dihydroxylation of pyridine-ring systems is presented.

A dynamic mechanical method for determining the silicone elastomer solubility of drugs and pharmaceutical excipients in silicone intravaginal drug delivery rings

The silicone elastomer solubilities of a range of drugs and pharmaceutical excipients employed in the development of silicone intravaginal drug delivery rings (polyethylene glycols, norethisterone acetate, estradiol, triclosan, oleyl alcohol, oxybutynin) have been determined using dynamic mechanical analysis. The method involves measuring the concentration-dependent decrease in the storage modulus associated with the melting of the incorporated drug/excipient, and extrapolation to zero change in storage modulus. The study also demonstrates the effect of drug/excipient concentrations on the mechanical stiffness of the silicone devices at 37°C.

In vitro release of nonoxynol-9 from silicone matrix intravaginal rings

The controlled-release characteristics of matrix silicone intravaginal rings loaded with between 100 and 971 mg of nonoxynol-9 have been investigated with a view to developing a ring that may offer a new female-controlled method for the prevention of transmission of sexually transmitted diseases, particularly HIV. Intravaginal rings containing 253, 487 and 971 mg of nonoxynol-9 provided a daily release of 2 mg or more over the 8-day release period, the minimal amount of nonoxynol-9 considered to provide an effective vaginal concentration for the prevention of HIV. Furthermore, the maximum daily release of N9 was about 6 mg, an amount significantly smaller than that observed for other nonoxynol-9 products whose large daily doses may in fact increase the occurrence of HIV by causing epithelial damage to the vaginal tissue. The release mechanism of the liquid nonoxynol-9 from the intravaginal rings has also been investigated and compared to models describing the release of solid drugs from the rings. It has been demonstrated through release studies and surface microscopy that a drug depletion zone is not established in such liquid-loaded intravaginal ring systems, with implications for the release kinetics. (C) 2003 Elsevier B.V. All rights reserved.

Influence of silicone elastomer solubility and diffusivity on the in vitro release of drugs from intravaginal rings

The in vitro release characteristics of eight low-molecular-weight drugs (clindamycin, 17beta-estradiol, 17beta-estradiol-3-acetate, 17beta-estradiol diacetate, metronidazole, norethisterone, norethisterone acetate and oxybutynin) from silicone matrixtype intravaginal rings of various drug loadings have been evaluated under sink conditions. Through modelling of the release data using the Higuchi equation, and determination of the silicone solubility of the drugs, the apparent silicone elastomer diffusion coefficients of the drugs have been calculated. Furthermore, in an attempt to develop a quantitative model for predicting release rates of new drug substances from these vaginal ring devices, it has been observed that linear relationships exist between the log of the silicone solubility of the drug (mg ml(-1)) and the reciprocal of its melting point (K-1) (y = 3.558x - 9.620, R = 0.77), and also between the log of the diffusion coefficient (cm(2) s(-1)) and the molecular weight of the drug molecule (g mol(-1)) (y = - 0.0068x - 4.0738, R = 0.95). Given that the silicone solubility and silicone diffusion coefficient are the major parameters influencing the permeation of drugs through silicone elastomers, it is now possible to predict through use of the appropriate mathematical equations both matrix-type and reservoir-type intravaginal ring release rates simply from a knowledge of drug melting temperature and molecular weight. (C) 2003 Elsevier Science B.V. All rights reserved.

Characterisation of Silicone Elastomer Vaginal Rings Containing HIV Microbicide TMC120 by Raman Spectroscopy

Silicone elastomer vaginal rings are currently being pursued as a controlled-release strategy for delivering microbicidal substances for the prevention of heterosexual transmission of HIV. Although it is well established that the distribution of drugs in delivery systems influences the release characteristics, in practice the distribution is often difficult to quantify in-situ. Therefore, the aim of this work was to determine whether Raman spectroscopy might provide a rapid, non-contact means of measuring the concentrations of the lead candidate HIV microbicide TMC120 in a silicone elastomer reservoir-type vaginal ring. Vaginal rings loaded with TMC120 were manufactured and sectioned before either Raman mapping an entire ring cross-section (100 µm resolution) or running line scans at appropriate time intervals up to 30 h after manufacture. The results demonstrated that detectable amounts of TMC120, above the silicone elastomer saturation concentration, could be detected up to 1 mm into the sheath, presumably as a consequence of permeation and subsequent reprecipitation. The extent of permeation was found to be similar in rings manufactured at 25 and 80°C.