Characterization of glutaraldehyde-immobilized chymotrypsin and an in-situ immobilized enzyme reactor using capillary electrophoresis-based peptide mapping

La digestion enzymatique des protéines est une méthode de base pour les études protéomiques ainsi que pour le séquençage en mode « bottom-up ». Les enzymes sont ajoutées soit en solution (phase homogène), soit directement sur le gel polyacrylamide selon la méthode déjà utilisée pour l’isolation de la protéine. Les enzymes protéolytiques immobilisées, c’est-à-dire insolubles, offrent plusieurs avantages tels que la réutilisation de l’enzyme, un rapport élevé d’enzyme-sur-substrat, et une intégration facile avec les systèmes fluidiques. Dans cette étude, la chymotrypsine (CT) a été immobilisée par réticulation avec le glutaraldehyde (GA), ce qui crée des particules insolubles. L’efficacité d’immobilisation, déterminée par spectrophotométrie d’absorbance, était de 96% de la masse totale de la CT ajouté. Plusieurs différentes conditions d’immobilisation (i.e., réticulation) tels que la composition/pH du tampon et la masse de CT durant la réticulation ainsi que les différentes conditions d’entreposage tels que la température, durée et humidité pour les particules GA-CT ont été évaluées par comparaison des cartes peptidiques en électrophorèse capillaire (CE) des protéines standards digérées par les particules. Les particules de GA-CT ont été utilisés pour digérer la BSA comme exemple d’une protéine repliée large qui requit une dénaturation préalable à la digestion, et pour digérer la caséine marquée avec de l’isothiocyanate de fluorescéine (FITC) comme exemple d’un substrat dérivé afin de vérifier l’activité enzymatique du GA-CT dans la présence des groupements fluorescents liés au substrat. La cartographie peptidique des digestions par les particules GA-CT a été réalisée par CE avec la détection par absorbance ultraviolet (UV) ou fluorescence induite par laser. La caséine-FITC a été, en effet, digérée par GA-CT au même degré que par la CT libre (i.e., soluble). Un microréacteur enzymatique (IMER) a été fabriqué par immobilisation de la CT dans un capillaire de silice fondu du diamètre interne de 250 µm prétraité avec du 3-aminopropyltriéthoxysilane afin de fonctionnaliser la paroi interne avec les groupements amines. Le GA a été réagit avec les groupements amine puis la CT a été immobilisée par réticulation avec le GA. Les IMERs à base de GA-CT étaient préparé à l’aide d’un système CE automatisé puis utilisé pour digérer la BSA, la myoglobine, un peptide ayant 9 résidus et un dipeptide comme exemples des substrats ayant taille large, moyenne et petite, respectivement. La comparaison des cartes peptidiques des digestats obtenues par CE-UV ou CE-spectrométrie de masse nous permettent d’étudier les conditions d’immobilisation en fonction de la composition et le pH du tampon et le temps de réaction de la réticulation. Une étude par microscopie de fluorescence, un outil utilisé pour examiner l’étendue et les endroits d’immobilisation GA-CT dans l’IMER, ont montré que l’immobilisation a eu lieu majoritairement sur la paroi et que la réticulation ne s’est étendue pas si loin au centre du capillaire qu’anticipée.

Oxidation kinetics of nickel nano crystallites obtained by controlled thermolysis of diaquabis(ethylenediamine) nickel(II) nitrate

The metal complex, [Ni(en)2(H2O)2](NO3)2 (en = ethylenediamine), was decomposed in a static furnace at 200 C by autogenous decomposition to obtain phase pure metallic nickel nanocrystallites. The nickel metal thus obtained was studied by XRD, IR spectra, SEM and CHN analysis. The nickel crystallites are in the nanometer range as indicated by XRD studies. The IR spectral studies and CHN analyses show that the surface is covered with a nitrogen containing species. Thermogravimetric mass gain shows that the product purity is high (93%). The formed nickel is stable and resistant to oxidation up to 350 C probably due to the coverage of nitrogen containing species. Activation energy for the oxidation of the prepared nickel nanocrystallites was determined by non-isothermal methods and was found to depend on the conversion ratio. The oxidation kinetics of the nickel crystallites obeyed a Johnson–Mehl–Avrami mechanism probably due to the special morphology and crystallite strain present on the metal.

Production, purification, genetic characterization and application studies of tannase enzyme from marine fungus Aspergillus awamori

Marine fungi remain totally unexplored as a source of industrial enzyme and prospective applications. Further tannase production by a marine organism has so far not been established. The primary objective of this study included the evaluation of the potential of Aspergillus awamori isolated from sea water as part of an earlier study and available in the culture collection of the Microbial technology laboratory for tannase production through different fermentation methods, optimization of bioprocess variables by statistical methods, purification and characterization of the enzyme, genetic study, and assessment of its potential applications.

Kinetics of Bacterial Colony Growth by Laser Induced Fluorescence

The growth kinetics of an aerial bacterial colony on solid agar media was studied using laser induced fluorescence technique. Fluorescence quenching of Rhodamin B by the bacterial colony was utilized for the study. The lag phase, log phase, and stationary phase of growth curve of bacterial colony was identified by measuring peak fluorescence intensity of dye doped bacterial colony.

Effect of Dietary Proteins on Growth, Food Conversion and Digestive Enzyme Activity of Juvenile Macrobrachium Rosenbergii

School of Industrial Fisheries, Cochin University of Science and Technology

On the crystallization kinetics and micro-structural transformations of Fe40Ni38B18Mo4 alloys

Activation energy for crystallization (Ec) is a pertinent parameter that decides the application potential of many metallic glasses and is proportional to the crystallization temperature. Higher crystallization temperatures are desirable for soft magnetic applications, while lower values for data storage purposes. In this investigation, from the heating rate dependence of peak crystallization temperature Tp, the Ec values have been evaluated by three different methods for metglas 2826 MB (Fe40Ni38B18Mo4) accurately. The Ec values are correlated with the morphological changes, and the structural evolution associated with annealing temperatures is discussed.

Characterization of laccase from Streptomyces psammoticus: an enzyme for eco-friendly treatment of environmental pollutants

The present study has identified an actinomycete culture (S. psammoticus) which was capable of producing all the three major ligninolytic enzymes. The study revealed that least explored mangrove regions are potential sources for the isolation of actinomycetes with novel characteristics. The laccase production by the strain in SmF and SSF was found to be much higher than the reported values. The growth of the organism was favoured by alkaline pH and salinity of the medium. The enzyme also exhibited novel characteristics such as activity and stability at alkaline pH and salt tolerance. These two characters are quite significant from the industrial point of view making the enzyme an ideal candidate for industrial applications. Many of the application studies to date are focused on enzymes from fungal sources. However, the fungal laccases, which are mostly acidic in nature, could not be used universally for all application purposes especially, for the treatment of effluents from different industries, largely due to the alkaline nature of the effluents. Under such situations the enzymes from organisms like S. psammoticus with wide pH range could play a better role than the fungal counterparts. In the present study, the ability of the isolated strain and laccase in the degradation of dyes and phenolic compounds was successfully proved. The reusability of the immobilized enzyme system made the entire treatment process inexpensive. Thus it can be concluded from the present study that the laccase from this organism could be hopefully employed for the eco-friendly treatment of dye or phenol containing industrial effluents from various sources.

Comment on "Kinetics of spinodal decomposition in the ising model with vacancy diffusion"

In a recent paper [Phys. Rev. B 50, 3477 (1994)], P. Fratzl and O. Penrose present the results of the Monte Carlo simulation of the spinodal decomposition problem (phase separation) using the vacancy dynamics mechanism. They observe that the t1/3 growth regime is reached faster than when using the standard Kawasaki dynamics. In this Comment we provide a simple explanation for the phenomenon based on the role of interface diffusion, which they claim is irrelevant for the observed behavior.

Nitrifying Bioreactors Integrated into Shrimp and Prawn Hatchery Systems: Molecular Characterization of the Nitrifying Consortia,Reactor Kinetics, Modeling and Validation

National Centre for Aquatic Animal Health, Cochin University of Science and Technology

α-Galactosidase from Streptomyces griseoloalbus: an enzyme with versatile applications

The present study is focused on the production, purification and characterization of multiple thermostable α-galactosidases from a novel actinomycete strain Streptomyces griseoloalbus. The Chapter I of the thesis covers the wide literature regarding α-galactosidases from various sources and their potential applications. The Chapter 11 deals with the isolation of α-galactosidase- producing actinomycetes and selection of the best strain. The Chapters III and IV describe the optimization of α-galactosidase production under submerged fermentation and solid-state fermentation respectively. The Chapter V describes the purification and characterization of multiple α-galactosidases and also the obvious existence of a novel galactose-tolerant enzyme. The Chapter VI illustrates the potential applications of α-galactosidases from S. griseoloalbus followed by the Chapter VII summarizing and concluding the results of the present investigation.

Kinetics of martensitic transitions in Cu-Al-Mn under thermal cycling: Analysis at multiple length scales

In this paper we study the evolution of the kinetic features of the martensitic transition in a Cu-Al-Mn single crystal under thermal cycling. The use of several experimental techniques including optical microscopy, calorimetry, and acoustic emission, has enabled us to perform an analysis at multiple scales. In particular, we have focused on the analysis of avalanche events (associated with the nucleation and growth of martensitic domains), which occur during the transition. There are significant differences between the kinetics at large and small length scales. On the one hand, at small length scales, small avalanche events tend to sum to give new larger events in subsequent loops. On the other hand, at large length scales the large domains tend to split into smaller ones on thermal cycling. We suggest that such different behavior is the necessary ingredient that leads the system to the final critical state corresponding to a power-law distribution of avalanches.