Enantiorecognition of planar "metallocenic" chirality by a nitrile hydratase/amidase bienzymatic system.

For the first time, a bacterial strain expressing a nitrile hydratase/amidase activity was able to recognize a planar element of chirality: Rhodococcus rhodochrous PA-34 whole cells catalysed with a high level of enantioselectivity the biotransformation of a novel nitrile ferrocene derivative into its corresponding amide and/or acid. An important parameter in the enzymatic recognition is the choice of the inducer selected for the bacterial growth phase.

Synthesis, characterization and in vitro evaluation of dimethyl-beta-cyclodextrin-4-biphenylylacetic acid conjugate.

Biphenylylacetic acid (BPAA) was linked to the free hydroxyl group of 2,6-di-O-methyl-beta-Cyclodextrin (DM-beta-CyD) through an ester linkage to obtain the site specific release of the drug to the colon. The conjugate at 1:1 mole ratio was separated from the reaction mixture by semipreparative reverse-phase HPLC and characterized by 1H-NMR, 13C-NMR, IR spectroscopy, mass spectrometry and elemental analysis. Chemico-physical characteristics, such as water solubility and dissolution rate, were evaluated comparatively to the BPAA-DM-beta-CyD inclusion complex. Hydrolysis rates were investigated in media simulating gastro-intestinal fluids and at pH 7.4 in the presence of porcine liver esterase. A rapid release of the drug was observed at acid pH value. In all cases a first order kinetic was observed, characterized by t1/2 value of 1.19, 19 and 4 h for chemical hydrolysis at pH 1.1, at pH 7.4 and enzymatic hydrolysis, respectively. In vitro permeation studies through caco-2 cells confirmed the ability of DM-beta-CyD to increase the absorption of included BPAA. A slow permeation was observed for the drug conjugate to DM-beta-CyD due to the slow release of BPAA.

Biomembrane model interaction and percutaneous absorption of papaverine through rat skin: effects of cyclodextrins as penetration enhancers.

The effects of different concentrations of beta-cyclodextrin (beta-CyD), hydroxypropyl-beta-cyclodextrin (HP-beta-CyD) and 2,6-di-O-methyl-beta-cyclodextrin (DM-beta-CyD) on percutaneous absorption of papaverine hydrochloride (PAP) were investigated. Abdominal rat skin mounted in Franz cells was used for in vitro experiments. To evaluate CyD interaction with a bilayer structure model, dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and DPPC-Chol (8:2 mole ratio) vesicles were used. CyD vesicle interaction was evaluated by differential scanning calorimetry. Permeation through rat skin and calorimetric experiments demonstrated that at low concentrations DM-beta-CyD shows higher enhancer activity as a possible result of a perturbing action on the skin by a complexation of its lipid components, but at higher concentrations HP-beta-CyD is the most effective. By considering that HP-beta-CyD presents a very moderate destabilizing action on the skin, we conclude that a 10% aqueous solution of this macrocycle appears to be the most suitable transdermal absorption enhancer for PAP.

Synthesis and structural characterization of 6I,6II-diamino-6I,6II-dideoxy-cyclomaltoheptaose, a difunctionalized beta-cyclodextrin.

6I,6II-Diamino-6I,6II-dideoxy-cyclomaltoheptaose was prepared using the regioselective procedure described by Tabushi. The difunctionalized beta-cyclodextrin crystallizes as hexadecahydrate in the orthorhombic space group P2(1)2(1)2(1), with a = 11.395(3), b = 32.989(9), c = 17.560(5) A, V = 6601 A3, Z = 4. The structure was solved by molecular replacement techniques using the program PATSEE and was refined to a conventional final R = 0.058 for the 5031 observed reflections with I > or = 3 sigma(I). The beta-CD macrocycle presents only slight differences with respect to uncomplexed hydrated or methylated beta-CD. The macrocycle structure maintains an approximate seven-fold symmetry. The round shape of the cyclodextrin ring is stabilized by intramolecular O-H ... O H-bonds between the secondary hydroxyl groups of neighbouring glucose residues. Along the a axis, the beta-CD molecules are arranged in columns; the macrocycles form a herring-bone pattern, so that the cavity of each beta-CD molecule is closed at each end by neighbouring molecules. The macrocycles are directly linked to each other by H-bonds involving either primary and secondary hydroxyl or amino groups of symmetry-related molecules. The resulting layers are connected to each other by a dense intermolecular hydrogen-bond network, in which solvent molecules participate.

Cell cycle time and life-span of cells in the mouse eye. Measurements during the postfetal period using repeated 3H-thymidine injections.

In this investigation the cell cycle time and the life-span of cells in different tissues of the mouse eye were determined during postfetal development not with single but, instead, by means of repeated 3H-thymidine injections. The potential of this method applied for the first time in the mammalian eye, is thoroughly discussed. Essentially, four groups of 19-21 mice each, aged 1, 10, 20, and 60 days at the start of the experiment, received intraperitoneal injections of 3H-thymidine at a dose of 1 microCi/g body weight every 4 h for a maximum of 14 days, i.e. a total of up to 85 injections. Further animals were sacrificed after the 13th, 19th, 25th, 31st, 37th, 49th, 61st, 73rd, and 85th injections, i.e. 2, 3, 4, 5, 6, 8, 10, 12, and 14 days after the start of the experiment. When all the injections had been given, animals from each group were sacrificed on the 1st, 4th, 8th, 16th, and sometimes the 32rd, 48th, and 64th days, respectively, after the last injection. With this experimental paradigm it was possible: (1) to determine the cell cycle time and the life-span of the cells during postfetal development up to maturity without gaps; (2) to establish the end of the development by means of cell proliferation in various tissues of the mouse eye; herewith it was possible to determine the times at which the development by cell proliferation is replaced by development by cell differentiation, and (3) to clearly prove in which mature ocular tissues cell turnover still exists and in which it does not; this appears to be especially important, since in recent years the importance of cell proliferation process following injury and stimulation was also recognized in the eye, for example, in massive periretinal proliferation in connection with retinal detachment and retinal surgery, as well as in endothelial injuries following intraocular lens implantation; only when normal conditions are known can pathological proliferative processes be recognized as such and be distinguished from normal ones. The results are cumulatively represented in tabular form, from which details are to be extracted. As expected, cell cycle times are very short and seemingly homogeneous in tissues which develop within themselves, but become longer and inhomogeneous, except in the inner and outer granular layer of the retina, in which the cell proliferation comes to a particularly abrupt end. The shortest cell cycle time occurred in the cells of the vascular walls of the retina at the time of birth and was 24 h.(ABSTRACT TRUNCATED AT 400 WORDS)

[Mitotic activity of the pigment epithelium during embryonic and postembryonic development]. / Die mitotische Aktivität des Pigmentepithels während der embryonalen und postembryonalen Entwicklung. Untersuchungen mit Colchizin und 3H-Thymidin an Kaninchen, Ratten und Mäusen.

The purpose of the present study was to examine the mitotic activity of the normal pigment epithelium of the retina (RPE), the ciliary body and the iris of different animals during gestation and after birth by blocking the metaphase with colchicine and by marking the pigment epithelial nuclei with tritium-labeled thymidine. The colchicine examinations were made on 54 albino rabbits and 56 albino rats, the 3H-thymidine studies with 78 albino mice. In the rabbit the peak of mitotic activity (respectively the end) is found in the RPE at the beginning of the 2nd third of gestation (respectively at the 9th postnatal day), but in the pigment epithelium of the ciliary body and in the iris during the last third of gestation (respectively in the 2nd month of life and the 3rd postnatal week). In the rat the highest mitotic activity is reached in the RPE at the beginning of the 2nd half of gestation (respectively at the 13th postnatal day), in the ciliary body at the 3rd day of life (respectively the 24th postnatal day) and in the iris at the end of the gestational period (respectively the 17th postnatal day). In the mouse the highest rates of mitotic activity are found in the RPE from the 16th gestational day to the 8th day of life (respectively the 20th day of life), in the ciliary body from the 1st to the 4th day of life (respectively from the 12th to the 20th postnatal day) and in the iris on the 9th day of life (respectively the 12th to the 20th postnatal day). The present observations have also demonstrated that with maturation of all areas of the pigment epithelium, the mitotic activity stops. The cells of the pigment epithelium do not have an epithelial cell turnover but they are reversible postmitotic cells. Despite the enormous proliferative properties the pigment epithelium shows no regeneration by mitosis after severe damage.