Development of novel economical methodologies for zymographic analysis of purified xylano–pectinolytic enzymes using agrowaste-based substrates

In this study, zymographic analysis for xylanase and pectinase enzymes has been carried out using agrowaste residues, wheat bran and citrus peel as well as their extracts. Isozymic forms of xylanase as well as pectinase enzyme displayed comparable zymographic bands onto agar petriplates containing either commercial substrates (xylan and pectin), agrowaste-based substrates (wheat bran and citrus peel), or polysaccharides extracted from these agrowastes (crude xylan and pectin extracted from wheat bran and citrus peel, respectively), indicating the fact that agro residues and their extracts can be utilized as a substitute of cost-intensive commercial substrates, xylan and pectin for zymographic analysis. This is the first report revealing the zymographic analysis of xylano–pectinolytic enzymes using agro-based solid residues particles or polysaccharides extracted from agro-based residues.     

Preclinical Formulations: Insight,Strategies, and Practical Considerations

A lot of resources and efforts have been directed to synthesizing potentially useful new chemical entities (NCEs) by pharmaceutical scientists globally. Detailed physicochemical characterization of NCEs in an industrial setup begins almost simultaneously with preclinical testing. Most NCEs possess poor water solubility posing bioavailability issues during initial preclinical screening, sometimes resulting in dropping out of an NCE with promising therapeutic activity. Selection of right formulation approach for an NCE, based on its physicochemical properties, can aid in improving its solubility-related absorption and bioavailability issues. The review focuses on preclinical formulations stressing upon different preclinical formulation strategies and deciphers the understanding of formulation approaches that could be employed. It also provides detailed information related to a vast pool of excipients available today, which is of immense help in designing preclinical formulations. Few examples mentioned, throw light on key aspects of preclinical formulation development. The review will serve as an important guide for selecting the right strategy to improve bioavailability of NCEs for academic as well as industrial formulation scientists.     

Frequency and significance of fungal isolations from conjunctival sac and their role in ocular infections

Five hundred conjunctival swabs, from 150 males and 100 females with no history of ocular infections, were collected and cultured for the isolations of fungi. Eighty (16 %) of the total specimens yielded positive fungal isolations. The isolation rate was more from the males than the females subjects. Mycelial fungi were predominant than the yeast organisms. Aspergillus species were the commonest isolates with A. flavus taking the lead in the isolations being positive 16 of the total 24 Aspergillus species isolated. A variable rate of fungal isolations was observed in different months of the year. The percentage of the isolations increases after the local use of Efcorlin-N. Nineteen of the 20 eyes studied did not yield the same fungal species in the repeated samples.Presented at the Second All India Congress of Medical Microbiologists at Hydrabad from December 22–24, 1978.     

Fast flow rate processes for purification of alkaline xylanase isoforms from Bacillus pumilus AJK and their biochemical characterization for industrial application purposes

This study shows the presence of five isozymic forms of alkaline xylanase from Bacillus pumilus using fast flow rate microfiltration, ultrafiltration, Q-sepharose, and phenyl sepharose chromatographic techniques. Polyacrylamide gel electrophoresis, high-performance liquid chromatography, and zymographic studies also revealed the purity of five isoforms of alkaline xylanases. Isoforms—X-I, X-III, and X-V exhibited optimum activity at pH 8.5, whereas X-II, X-IV showed maximum activity at pH 9. All isoforms were optimally active at temperature 55°C. Isoforms were found to be stable at pH 7–11, showed 92–100% residual activity after 3 hr, treatment time for most industrial applications. The isoforms retained nearly 80–86% residual activity after incubating at 45°C for 3 hr. Molecular weights of xylanase I–V, were 13.1, 15.3, 18.4, 20.1, and 21.0 kDa, respectively. Mg²⁺ ions were found to be potent activator for all isozymic forms. The Km and Vmax values of X-I, X-II, X-III, X-IV, and X-V were 6.71, 6.66, 7.14, 5.88, 6.25 mg/ml and 2,000, 1,695, 1,666.66, 1,428.57, and 1,408.45 IU/mg protein, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed the monomeric nature of all isoforms. The low-molecular masses, significantly enhanced activity in the presence of industrially suitable—low cost activator, better stability of all isoforms at pH 7–11 and at higher temperature, also presence of multiple forms of alkaline xylanase, makes this enzyme suitable for textile–paper industries. This is also the first report mentioning the purification of five isozymic forms of alkaline xylanase using fast flow rate techniques.     

A versatile tool in controlling aggregation and Ag nanoparticles assisted in vitro folding of thermally denatured zDHFR

BackgroundProteins have tendency to form inactive aggregates at higher temperatures due to thermal instability. Maintenance of thermal stability is essential to gain the protein in sufficient quantity and biologically active form during their commercial production.MethodsBL21-DE3 Rosetta E. coli cells which contains plasmid pET43.1a vector was used for producing zDHFR protein commercially. The purification of N-terminal Histidine tagged zDHFR was performed by Immobilized Metal Ion chromatography (IMAC). Investigations were performed in existence and non existence of Silver nanoparticles (AgNPs). The inactivation kinetics of zDHFR in existence and non existence of AgNPs were monitored over a range of 40–80 °C as monitored by UV–Visible absorption spectroscopy.ResultsThe protein completely lost its activity at 55 °C. Kinetics of inactivated zDHFR follows first order model in presence and absence of AgNPs. Decrease in rate constant (k) values at respective temperatures depicts that AgNPs contribute in the thermostability of the protein. AgNPs also assists in regaining the activity of zDHFR protein.ConclusionsAgNPs helps in maintaining thermostability and reducing the aggregation propensity of zDHFR protein.General significanceResult explains that AgNPs are recommended as a valuable system in enhancing the industrial production of biologically active zDHFR protein which is an important component in folate cycle and essential for survival of cells and prevents the protein from being aggregated.