Inoculum studies in production of penicillin g acylase by Bacillus megaterium ATCC 14945

This article reports studies concerning the production of penicillin G acylase (PGA) by Bacillus megaterium. This enzyme has industrial use in the hydrolysis of penicillin G to obtain 6-aminopenicillanic acid, an essential intermediate for the production of semisynthetic β-lactam antibiotics. Although most microorganisms produce the enzyme intracellularly, B. megaterium provides extracellular PGA. The enzyme production by microorganisms involves several steps, resulting in a many operational variables to be studied. The study of the inoculum is an important step to be accomplished, before addressing other issues such as culture optimization and downstream processing. In this study, using a standard inoculum as reference, several runs were performed aiming at the definition of operational conditions in the PGA production. Cell concentration and PGA activity in the production medium were measured after 24, 48, and 72 h of the beginning of the production phase. This study encompasses the duration of the inoculum germination phase and the concentration of cells used to startup the germination. Based on these results, PGA productivity during the production phase was maximized. The selected values for these variables were 1.5 × 10⁷ spores/mL of germination medium, germination during 24 h, and 72 h for the production phase.     

The Binary Oxide System TeO2?MoO3

Phase analysis of the mixed oxide system TeO₂? MoO₃ by means of x-ray diffraction and optical microscopy indicates the formation of a new phase, α-Te₂MoO₇, stable at room temperature. Below 500°C mixtures of crystalline products are obtained or complete devitrification can easily be induced in the system TeO₂? MoO₃. Above this temperature, tendency to glass formation is observed under the conditions employed, due to the liquidus temperature effect. Quenching of a melt of Te₂MoO₇ yields a dark yellow glass, α-Te₂MoO₇. X-ray and density measurements were used to explore the range of stoichiometry and exclude formation of solid solutions in the system; no apparent relationship exists between the crystal structures of the component oxides and the binary compound.     

Biosynthesis of aglajnes, polypropionate allomones of the opisthobranch mollusc Bulla striata

The biogenesis of aglajnes, polypropionate allomones of the cephalaspidean mollusc Bulla striata, has been investigated in vivo by feeding experiments. Incorporation of the committed precursor, [1-¹⁴C]-propionate, into aglajne-1 (1) and -3 (3) established the de novo origin of these compounds in B. striata. In the letter we also discuss briefly the ecological meaning of the origin of polypropionates in B. striata and in other cephalaspidean molluscs.     

Interactions between naproxen and maltoheptaose,the non-cyclic analog ofβ-cyclodextrin

The crystallinity of naproxen in solid combinations with amorphous maltoheptaose, the non-cyclic analog of -cyclodextrin, was assessed using differential scanning calorimetry supported by X-ray powder diffractometry. Cogrinding induced a decrease in drug crystalinity to an extent which depended on the grinding time, and was most pronounced for the combination of equimolecular composition. Thermal analysis showed that the mechanism behind the conversion of crystalline naproxen into the amorphous state by cogrinding with maltoheptaose differed from that with randomly substituted, amorphous -cyclodextrins. Interactions of naproxen with maltoheptaose in aqueous solution were studied by means of fluorescence spectroscopy, phase-solubility analysis, and computeraided molecular modelling. Maltoheptaose can wrap up naproxen, taking on a cyclic conformation and forming a pseudo inclusion complex (apparent binding constant K_{1: 1} = 1.0 × 10³ (–20%) L mol^–1 at 25 °C) which is about as stable as the true inclusion complex with -cyclodextrin in the lowest temperature range (0-100 K). A better complexing ability for naproxen in terms of binding constant values, however, was displayed by both native and derivatized -cyclodextrins, the hosts with covalently-bound cyclic structures.     

Quantitative analysis of 2-furfural and 5-methylfurfural in different Italian vinegars by headspace solid-phase microextraction coupled to gas chromatography-mass spectrometry using isotope dilution

A new method was developed for the determination of 2-furfural (2-F) and 5-methylfurfural (5-MF), two products of Maillard reaction in vinegar, with head-space solid-phase microextraction (HS-SPME) coupled to gas chromatography-mass spectrometry (GC-MS). A divinylbenzene (DVB)/carboxen (CAR)/polydimethylsiloxane (PDMS) fibre was used and SPME conditions were optimised, studying ionic strength effect, temperature effect and adsorption time. Both analytes were determined by calibration established on 2-furfural-d4 (2-F-d4). The method showed good linearity in the range studied (from 16 to 0.12 mg/l), with a regression coefficient r2 of 0.9999. Inter-batch precision and accuracy were found between 14.9 and 6.0% and between -11.7 and 0.2%, respectively. Detection limit was 15 microg/l. The method is simple and accurate and it has been applied to a series of balsamic and non-balsamic vinegars.     

Small-angle neutron scattering of ionic perfluoropolyether micellar solutions: role of counterions and temperature

This paper reports a small-angle neutron scattering (SANS) characterization of perfluoropolyether (PFPE) aqueous micellar solutions with lithium, sodium, cesium and diethanol ammonium salts obtained from a chlorine terminated carboxylic acid and with two perfluoroisopropoxy units in the tail (n(2)). The counterion and temperature effects on the micelle formation and micellar growth extend our previous work on ammonium and potassium salts n(2) micellar solutions. Lithium, sodium, cesium and diethanol ammonium salts are studied at 0.1 and 0.2 M surfactant concentration in the temperature interval 28-67 degrees C. SANS spectra have been analyzed by a two-shell model for the micellar form factor and a screened Coulombic plus steric repulsion potential for the structure factor in the frame of the mean spherical approximation of a multicomponent system reduced to a generalized one component macroions system (GOCM). At 28 degrees C, for all the salts, the micelles are ellipsoidal with an axial ratio that increases from 1.6 to 4.2 as the counterion volume increases. The micellar core short axis is 13 A and the shell thickness 4.0 A for the alkali micelles, and 14 and 5.1 A for the diethanol ammonium micelles. Therefore, the core short axis mainly depends on the surfactant tail length and the shell thickness on the carboxylate polar head. The bulky diethanol ammonium counterion solely influences the shell thickness. Micellar charge and average aggregation number depend on concentration, temperature and counterion. At 28 degrees C, the fractional ionization decreases vs the counterion volume (or molecular weight) increase at constant concentration for both C = 0.1 M and C = 0.2 M. The increase of the counterion volume leads also to more ellipsoidal shapes. At C = 0.2 M, at 67 degrees C, for sodium and cesium micelles the axial ratio changes significantly, leading to spherical micelles with a core radius of 15 A, lower average aggregation number, and larger fractional ionization.     

Thermal analysis of a drug-polymeric excipient solid system

Differential scanning calorimetry has been applied to the study of the solid system cross-linked polyvinylpyrrolidone/trimethoprim.Different results have been systematically obtained on co-ground and not co-ground mixtures, suggesting some type of interaction to take place between components as a consequence of co-grinding.A simple phenomenological model of the interaction is proposed, which quite satisfactorily agrees with thermal data.

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Zusammenfassung Das feste System von vernetztem Polyvinyl-pyrolidin/Trimethoprim wurde mittels DSC untersucht. Unterschiedliche Ergebnisse wurden systematisch für zusammen und nicht zusammen verriebene Mischungen gefunden, was auf eine Wechselwirkung zwischen Komponenten beim Verreiben hindeutet. Ein einfaches phänomenologisches Modell dieser Wechselwirkung wird vorgeschlagen, das mit thermischen Daten befriedigend übereinstimmt.

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This work has been partially supported by Ministero Pubblica Istruzione (Fondi 60%).     

Sensitivity improvement in 19F NMR-based screening experiments: theoretical considerations and experimental applications

NMR-based binding and functional screening performed with FAXS (fluorine chemical shift anisotropy and exchange for screening) and 3-FABS (three fluorine atoms for biochemical screening) represents a potential alternative approach to high-throughput screening for the identification of novel potential drug candidates. The major limitation of this method in its current status is its intrinsic low sensitivity that limits the number of tested compounds. One approach for overcoming this problem is the use of a cryogenically cooled (19)F probe that reduces the thermal noise in the receiver circuitry. Sensitivity improvement in the two screening techniques achieved with the novel cryogenic (19)F probe technology permits an increased throughput, detection of weaker binders and inhibitors (relevant in a fragment-based lead discovery program), detection of slow binders, and reduction in protein and substrate consumption. These aspects are analyzed with theoretical simulations and experimental quantitative performance evaluation. Application of 3-FABS combined with the cryogenic (19)F probe technology to rapid screening at very low enzyme concentrations and the current detection limits reached with this approach are also presented.