Molecular cloning and characterization of thermostable beta-lactam acylase with broad substrate specificity from Bacillus badius

The gene (pac) encoding beta-lactam acylase from Bacillus badius was cloned and expressed in Escherichia coli. The pac gene was identified by polymerase chain reaction (PCR) using degenerated primers, on the basis of conserved amino acid residues. By using single specific primer PCR (SSP-PCR) and direct genome sequencing, a complete pac gene with its promoter region was obtained. The ORF consisted of 2415 bp and the deduced amino acid sequence indicated that the enzyme is synthesized as a preproenzyme with a signal sequence, an alpha-subunit, a spacer peptide and a beta-subunit. The pac gene was expressed with its own promoter in different E. coli host strains and a maximum recombinant PAC (1820 U l(-1)) was obtained in E. coli DH5alpha. The recombinant PAC was purified by Ni-NTA chromatography and the purified PAC had two subunits with apparent molecular masses of 25 and 62 kDa. This enzyme exhibited a high thermostability with a maximum activity at 50 degrees C. This enzyme showed stability over a wide pH range (pH 6.0-8.5) with a maximum activity at pH 7.0 and activity on a wide beta-lactam substrate range. The K(m) values obtained for the hydrolysis of penicillin G and a chromogenic substrate, 6-nitro-3-phenylacetylamidobenzoic acid, from B. badius PAC were 39 and 41 microM, respectively. The PAC activity was competitively inhibited by PAA (K(i), 108 microM) and noncompetitively by 6-APA (K(i), 17 mM). The constitutive production of B. badius PAC in E. coli and its easier purification together with the advantageous properties, such as thermostability, pH stability and broad substrate specificity, make this as a novel enzyme suitable for beta-lactam industry.     

Lipase immobilized on nanostructured cerium oxide thin film coated on transparent conducting oxide electrode for butyrin sensing

Nanostructured cerium oxide (CeO₂) thin films were deposited on transparent conducting oxide (TCO) substrate using spray pyrolysis technique with cerium nitrate salt, Ce(NO₃)₃·6H₂O as precursor. Fluorine doped cadmium oxide (CdO:F) thin film prepared using spray pyrolysis technique acts as the TCO film and hence the bare electrode. The structural, morphological and elemental characterizations of the films were carried out using X-ray diffraction (XRD), field emission scanning electron microscopy (FE-SEM) and energy dispersive X-ray analysis (EDX) respectively. The diffraction peak positions in XRD confirmed the formation of highly crystalline ceria with cubic structure and FE-SEM images showed uniform adherent films with granular morphology. The band gaps of CeO₂ and TCO were found to be 3.2 eV and 2.6 eV respectively. Lipase enzyme was physisorbed on the surface of CeO₂/TCO film to form the lipase/nano-CeO₂/TCO bioelectrode. Sensing studies were carried out using cyclic voltammetry and amperometry, with lipase/nano-CeO₂/TCO as working electrode and tributyrin as substrate. The mediator-free biosensor with nanointerface exhibited excellent linearity (0.33–1.98 mM) with a lowest detection limit of 2 μM with sharp response time of 5 s and a shelf life of about 6 weeks.     

Influence of heat pump drying on tomato flavor

ABSTRACT

Tomato flavor from a heat pump dehumidifier dryer was characterized and compared to fresh and freeze-dried tomato samples for the retention of fresh flavor compounds. The monitored quality parameters were nonvolatile, volatile, and odor intensity. There were no differences in nonvolatile profiles; however, significant changes in key odor-impact volatiles were observed. Commercial spray-dried tomato was also evaluated to examine thermally induced volatile changes. The volatile and sensory profiles of heat pump dried tomato were comparable to freeze-dried tomato, with good retention of fresh aroma. In contrast, loss of compounds contributing to fresh green aroma ((E)-2-hexenal, 1-penten-3-one, 1-hexanol) and presence of heat-induced compounds (dimethyl sulfide, furfural, pyrrole derivatives) were detected in the spray-dried tomato.     

A simple route towards the reduction of surface conductivity in gas sensor devices

Surface conductivity caused by adsorbed water ions and molecules affects the performance of capacitive field effect transistor (FET) based gas sensors. Surface conduction leads to a slow decay of any voltage difference built-up between the sensitive surface and the gate electrode of the FET and thus to a drift in the sensor signal. In humid environments this problem becomes even more significant, as more water is becoming adsorbed. This leads to a cross-sensitivity of the sensor signal towards moisture. In this work we report on the application of a simple room temperature violet photochemical process to yield chemically anchored thin fluoropolymer films on the surface of such FET devices. The modified areas of the sensor surface were strongly water repellent. This led to a decrease in surface conductivity by more than three orders of magnitude at high relative humidities. Making the surface of the sensor chips hydrophobic resulted in elimination of the surface discharge caused by the leakage current even at high relative humidities.     

Modification of micronozzle surfaces using fluorinated polymeric nanofilms for enhanced dispensing of polar and nonpolar fluids

In this work, we report on the surface modification of a micronozzle surface to enhance fluid dispensing in the nanoliter range. Unmodified dispensing chips usually suffer from lateral wetting of the nozzle surfaces by low surface tension liquids resulting in poor control of the volume of the dispensed fluid. Covalent attachment of a fluorinated acrylate polymer to the outer surface of the micronozzle using a novel UV irradiation process helps to overcome capillary and adhesive forces and results in an enhancement of the control on fluid dispensing in the nanoliter range. The modified nozzles have been tested with a variety of fluids having a wide range of surface tensions. The surface modification allows precise control of the dispensing of nanoliter droplets with a high degree of reproducibility.