In vivo assessment of the permeability of the blood--brain barrier and blood-retinal barrier to fluorescent indoline derivatives in zebrafish

ABSTRACT: BACKGROUND: Successful delivery of compounds to the brain and retina is a challenge in the development of therapeutic drugs and imaging agents. This challenge arises because internalization of compounds into the brain and retina is restricted by the blood--brain barrier (BBB) and blood-retinal barrier (BRB), respectively. Simple and reliable in vivo assays are necessary to identify compounds that can easily cross the BBB and BRB. METHODS: We developed six fluorescent indoline derivatives (IDs) and examined their ability to cross the BBB and BRB in zebrafish by in vivo fluorescence imaging. These fluorescent IDs were administered to live zebrafish by immersing the zebrafish larvae at 7--8 days post fertilization in medium containing the ID, or by intracardiac injection. We also examined the effect of multidrug resistance proteins (MRPs) on the permeability of the BBB and BRB to the ID using MK571, a selective inhibitor of MRPs. RESULTS: The permeability of these barriers to fluorescent IDs administered by simple immersion was comparable to when administered by intracardiac injection. Thus, this finding supports the validity of drug administration by simple immersion for the assessment of BBB and BRB permeability to fluorescent IDs. Using this zebrafish model, we demonstrated that the length of the methylene chain in these fluorescent IDs significantly affected their ability to cross the BBB and BRB via MRPs. CONCLUSIONS: We demonstrated that in vivo assessment of the permeability of the BBB and BRB to fluorescent IDs could be simply and reliably performed using zebrafish. The structure of fluorescent IDs can be flexibly modified and, thus, the permeability of the BBB and BRB to a large number of IDs can be assessed using this zebrafish-based assay. The large amount of data acquired might be useful for in silico analysis to elucidate the precise mechanisms underlying the interactions between chemical structure and the efflux transporters at the BBB and BRB. In turn, understanding these mechanisms may lead to the efficient design of compounds targeting the brain and retina.     

Preparation and performance of plasticizers for poly(vinyl chloride) from products of thermal decomposition of waste polystyrene

2,4-Diphenylbutyl-2,4-diphenylbutyrate (DPBDPB) and 2,4,6-triphenylhexyl-2,4,6-triphenylhexoate (TPHTPH), plasticizers for poly(vinyl chloride), were synthesized from the products of thermal decomposition of waste polystyrene. Their heat stabilities were studied by thermogravimetric analysis and differential thermal analysis, and compared with those of typical plasticizers for PVC such as dibutyl phthalate (DBP), dihexyl phthalate (DHP) and bis(2-ethylhexyl) phthalate (DOP). DPBDPB and TPHTPH showed much higher heat resistance than DOP. PVC was plasticized with a mixed system consisting of DOP as the primary plasticizer and DPBDPB as the secondary. It became clear that DPBDPB is an excellent heat-resistant plasticizer which does not affect the compatibility of PVC with DOP.     

DNA PHOTOREACTIVATING ENZYME FROM SILKWORM

An ammonium-sulfate-precipitable (33–70%) fraction in extracts from eggs of silkworm Bombyx mori contains photoreactivating enzyme that reactivates the transforming activity of UV inactivated Hemophilus influenzae DNA. The action spectrum for in vitro photoreactivation with the enzyme has a broad peak around 365–385 nm, with a shoulder extending to 460 nm. This relatively higher photoreactivation efficiency at wavelengths longer than 450 nm seems to be a unique feature of DNA photoreactivating enzyme of silkworm. Using gel filtration, a mol wt of 42,000 was estimated for the enzyme. Optimum and isoionic pH of the enzyme were 7.2 and 5.4, respectively. These properties of silkworm enzyme are within the range of variations in reported biochemical characteristics of photoreactivating enzymes from different species.     

Millimeter and Submillimeter Wave ESR System: Using 30 T Pulsed Magnetic Field

New systems for millimeter and submillimeter wave ESR have been developed in Kobe University. In the previous system the pulsed magnetic field was limited up to 17 T in the temperature range from 1.8 to 86 K. Using the new systems, we can measure in the field range up to 30 T in the temperature range from 1.8 K to 4.2 K and from 18 K to room temperature. The resolution of the magnetic field has been also improved in the new ESR system. The details of our new ESR systems are presented. In addition, the measurements of Yb₂Cu₂O₅ using these new systems are presented.     

Analysis by high-performance gel permeation chromatography of hyaluronic acid in animal skins and rabbit synovial fluid

The simultaneous analysis of the molecular weight and concentration of hyaluronic acid in biological samples using high-performance liquid chromatography with two gel permeation columns is described. The elution volumes of various molecular weights of hyaluronic acids were linearily related to the logarithms of their molecular weights up to 600,000. The concentration of hyaluronic acid could be determined in the range from 20 to 100 micrograms/ml, i.e., from 4 to 20 micrograms per 200 microliter injected. The method was applied to the analysis of several animal skin extracts and rabbit synovial fluid. Skin extracts from mouse, rat, guinea-pig and rabbit could be chromatographed without prior isolation and purification. Hyaluronic acids in skin were separated clearly from chondroitin sulphates and their concentrations were determined. The molecular weights were estimated simultaneously to be more than 10(6). Rabbit synovial fluids from intact joints and saline- and carrageenin-treated joints could be chromatographed directly. The chromatograms showed that the concentration of hyaluronic acid in carrageenin-treated synovial fluid is lower than that in saline-treated fluid and the molecular weight distribution is broader. This technique enabled the rapid analysis of hyaluronic acid present at low levels in biological samples.     

Formes linéaires de logarithmes de points algébriques sur les groupes algébriques

Dans ce papier, nous donnons des minorations de combinaisons linéaires à coefficients algébriques de logarithmes de points algébriques sur les groupes algébriques commutatifs: soitG un groupe algébrique commutatif connexe défini sur la clôture algébriqueQ de dans , et soitvT _G () un point de l'espace tangent dont l'image exp _G (v) par l'exponentielle deG appartient àG(Q). On cherche à minorer la distance du pointv aux hyperplansW deT _G (), rationnels surQ, qui ne passent pas parv. Cette minoration dépend, entre autres, d'un majorantB de la hauteur usuelle des coefficients d'une forme linéaire définissantW. LorsqueG est un groupe linéaire, il s'agit de minorer une combinaison linéaire de logarithmes de nombres algébriques, et la méthode de A. Baker permet d'obtenir une minoration

Automated high-performance liquid chromatographic method for the simultaneous determination of cefotiam and delta 3-cefotiam in human plasma using column switching.

An automated high-performance liquid chromatographic method using column switching was established for the simultaneous determination of cefotiam (I) and delta 3-cefotiam (II) in human plasma after oral administration of cefotiam hexetil dihydrochloride. The method allowed the determination of analytes in plasma by the direct injection of diluted specimen with phosphate buffer. The analytes were enriched onto the C18 short pretreatment column by 0.05 M phosphate buffer (pH 7.7), while proteins and endogenous hydrophilic substances in plasma were washed off to waste. The enriched analytes were then back-flushed onto the analytical C18 column, separated by a mixture of 0.05 M phosphate buffer (pH 7.7)-acetonitrile (88:12, v/v) and detected by the ultraviolet absorbance at 254 nm. Recoveries from spiked plasma were quantitative, and the coefficients of variation were below 4%. The lower detection limits in plasma were 10 ng/ml for both I and II. Concentrations of I and II in plasma determined by the present method were in good agreement with those obtained by the conventional deproteinization method.     

Absolute stereochemistries and total synthesis of (+)/(−)-macrosphelides, potent, orally bioavailable inhibitors of cell-cell adhesion

In the current studies, we used the single-crystal X-ray analysis and Kakisawa-Kashman modification of the Mosher NMR method to determine the complete relative and absolute stereochemistries of the (+)-macrosphelides A (+)-1 and B (+)-2. The stereostructure of (+)-2 was determined by chemical comparison with artificial (+)-2 from (+)-1. We also report the convergent total synthesis of (+)-1 and (+)-3, as well as their antipodes, utilizing an asymmetric dihydroxylation for introduction of chirality and Yamaguchi macrocyclization to form the 16-membered trilactone macrolides.     

Topological dependency of the aromatic sextets in polycyclic benzenoid hydrocarbons. Recursive relations of the sextet polynomial

Mathematical meaning of the Clar's “aromatic sextet” is clarified by analysing the topological dependency of the sextet polynomial. Generalised recursive method for obtaining the sextet polynomial of a polyhex graph is presented. It is shown that the concept of the “super sextet” is necessarily introduced, if one-to-one correspondence between the Kekulé and sextet patterns is assumed. Topological dependency of the maximum number of resonant sextets is clarified and discussed in relation to the aromaticity and stability of polycyclic benzenoid hydrocarbons.