Tool In Pharmaceutical Industry Research Articles

A promising microbial α-amylase production, and purification from Bacillus cereus and its assessment as antibiofilm agent against Pseudomonas aeruginosa pathogen

Background and aimThe purpose of the current study is to isolate a heavily amylase-producing bacteria of the genus Bacillus from soil samples, optimize the production of the enzyme, purify it, and evaluate its activity against biofilm-producing bacteria. A total of 12 soil samples were collected and screened for promising Bacillus species with good amylolytic activity. Isolation was done by serial dilution and plating technique and amylolytic activity was determined by starch agar plate method. Among the 12 Bacillus isolates recovered from soil samples, 7 showed positive α-amylase production. The best isolate that recorded the greatest amylolytic activity was selected for further studies. This isolate was identified by 16S rRNA sequencing as Bacillus cereus and registered under gene bank accession number OP811897. Furthermore, the α-amylase enzyme was produced by a submerged fermentation technique using best production media and partially purified by ammonium sulfate and chilled ethanol and molecular weight had been determined by SDS-PAGE gel electrophoresis. The production of α-amylase was optimized experimentally by one-factor at a time protocol and statistically by Plackett–Burman design as well as RSM CCD design. Data obtained from OFAT and CCD revealed that α-amylase activities were 1.5- and twofold respectively higher as compared to un-optimized conditions. The most significant factors had been identified and optimized by CCD design.ResultsAmong the eleven independent variables tested by PBD, glucose, peptone, (NH4)2SO4, and Mg SO4 were the most significant parameters for α-amylase production with an actual yield of 250U/ml. The best physical parameters affecting the enzyme production were incubation time at 35 °C, and pH 5.5 for 48 h. The partially purified enzyme with 60% ammonium sulphate saturation with 1.38- fold purification showed good stability characteristics at a storage temperature of 4 °C and pH up to 8.5 for 21 days. Antibiofilm activity of purified α-amylase was determined against Pseudomonas aeruginosa (ATCC 35659) by spectrophotometric analysis and CLSM microscopic analysis. Results demonstrated biofilm inhibition by 84% of the formed Pseudomonas biofilm using a microtiter plate assay and thickness inhibition activity by 83% with live/Dead cells percentage of 17%/83% using CLSM protocol.ConclusionsA highly stable purified α-amylase from B. cereus showed promising antibiofilm activity against one of the clinically important biofilm-forming MDR organisms that could be used as a cost-effective tool in pharmaceutical industries.

Plasmonic Au nanoparticles embedded in glass: Study of TOF-SIMS, XPS and its enhanced antimicrobial activities

The Au nanoparticles (NPs) were formed near the surfaces of pre-synthesized sodium zinc –borate glass by an ion exchange process obtained by thermal heat treatment in an open air environment at various temperatures. The pre-heated Au doped glass samples were extensively tested by the different techniques such as optical absorption spectroscopy, Scanning Electron Microscope (SEM), X-ray photoelectron spectroscopy (XPS), Time of Flight Secondary Ion Mass Spectroscopy (TOF-SIMS), and these Au NPs were used for the antimicrobial applications. SEM confirmed the spherical shaped Au NPs with increasing thermal treatment up to 550 °C. The optical absorption findings showed that the as –synthesized Au NPs showed Localized Surface Plasmon Resonance behaviour, giving clear evidence of an Au NPs band formed in the glass matrix. The formation mechanism of the Au doped glass samples was studied theoretically from a thermodynamic point of view during heat treatment. XPS and TOF-SIMS were used to study the chemical state and the thermal stability of the pre-heated Au NPs doped glasses in an ultra-high vacuum. The effect of concentration changes in the line-shape and in binding energy as a function of thermal heat treatment suggests that the Au NPs formed near the glass surfaces and changes in the chemical composition as well as chemical structures of the Au doped glass samples occurred. Antimicrobial activity such as antibacterial as well as antifungal activity of pre-heated Au doped glass samples was tested against different strains by the disk diffusion method. The Au doped glass samples exhibited enhanced antibacterial as well as antifungal activities by the influence of thermal treatment at different temperatures. Thus, the Au doped glass sample could be efficiently explored as a medical tool in pharmaceutical industries, biotechnology industries and chemical laboratories based upon its antibacterial findings. • Au NPs were embedded in the glass matrix by ion-exchange technique. • Au NPs were uniformly distributed in the glass matrix as confirmed by TEM analysis. • The antimicrobial properties were enhanced by increasing the annealing temperatures. • Thermodynamic approach was performed for the growth of the Au NPs. • The Au NPs exhibited invitro antifungal activities.

Application of chemometrics using direct spectroscopic methods as a QC tool in pharmaceutical industry and their validation.

This present review described the application of chemometrics using direct spectroscopic methods at the quality control (QC) laboratory of Pharmaceutical Industries. Using chemometrics methods, all QC assessments during the fabrication processes of the drug preparations can be well performed. Chemometrics methods have some advantages compared to the conventional methods, i.e., non-destructive, can be performed directly to intake samples without any extractions, unnecessary performing stability studies, and cost-effective. To achieve reliable results of analyses, all methods must be validated first prior to routine applications. According to the current Pharmacopeia, the validation parameters are specificity/selectivity, accuracy, repeatability, intermediate precision, range, detection limit, quantification limit and robustness. These validation data must meet the acceptance criteria, that have been described by the analytical target profile (ATP) of the drug preparations.

Development of aqueous-based multi-herbal combination using principal component analysis and its functional significance in HepG2 cells

BackgroundThe present study was carried out to prepare multi-herbal combination via comparing antioxidant activity and polyphenolic composition of five medicinal plant extracts of Punica granatum L., Putranjiva roxburghii Wall., Swertia chirata Buch.-Ham., Tinospora cordifolia (Willd.) Miers and Trigonella corniculata L.MethodsThe herbs were individually evaluated using in vitro antioxidant assays and analyzed by HPLC-PDA. The resultant data was examined using principal component analysis (PCA). Further, herbal combination was prepared on the basis of PCA.ResultsThe PCA divided the plants into three groups. The leading or primary group contained P. granatum and P. roxburghii with the highest antioxidant activity strongly correlated with high amount of kaempferol. S. chirata was acknowledged as nourisher herb in one and T. cordifolia and T. corniculata were identified as stimulator herbs in other group. The herbal combination exhibited high antioxidant activity as compared to the individual plants. The combination revealed good antiproliferative efficacy against hepatocellular carcinoma (HepG2) cells with IC50 of 75.864 μg/ml.ConclusionsThe activity observed in vitro with HepG2 cells suggests that the herbal combination can provide therapeutic activity in vivo in future. The study may provide information regarding precise preparation of multi-herbal formulations using PCA as a tool in pharmaceutical industries.

Recrystallization of phenacetin and sulfathiazole using the sonocrystallization process

In this study, a cooling sonocrystallization process is used to recrystallize two active pharmaceutical ingredients (APIs), phenacetin and sulfathiazole, to modify the solid-state property and enhance the dissolution behavior. Recrystallization results using different organic solvents are presented and discussed. For both APIs, the crystal habit and mean particle size are modified significantly after the cooling sonocrystallization process. Recrystallized API particles show more regular habit and its mean particle size can be reduced to about 10 µm. In addition, for sulfathiazole, polymorph modification is also observed when different organic solvent is used. Furthermore, by dissolution studies, recrystallized APIs show enhanced dissolution behavior compared with the original samples. These results demonstrate that the sonocrystallization process is an efficient tool in pharmaceutical industry for solid state property control and dissolution rate enhancement.

Degradation kinetics of metronidazole and its mutual prodrug with ciprofloxacin: a calorimetric analysis

Calorimetric technique has aroused considerable interest as a versatile tool in pharmaceutical industry and academia to provide useful information about thermodynamic and kinetic aspects of drug molecules. The present paper utilizes this technique to monitor the hydrolytic degradation of metronidazole and its prodrug with ciprofloxacin, i.e. 2-(2-methyl-5-nitroimidazol-1-yl)ethyl-1-cyclopropyl-6-fluoro-1,4-dihydro-4-oxo-7-(1-piperazinyl)-quinoline-3-carboxylate. The synthesis of the present mutual prodrug was envisaged to combine the antiprotozoal and anaerobic antibacterial effects of metronidazole with antibacterial effects of ciprofloxacin. Heat flux microcalorimeter was used to determine the rate of heat evolved during the degradation of the drug and prodrug as a function of concentration, pH and temperature. In terms of enthalpy of hydrolysis the response is exothermic both for drug and prodrug. However, the absolute value of the enthalpy of reaction (ΔrH0) is low for the prodrug. The degradation followed pseudo first order kinetics, showed marked stability at pH 3-7 followed by accelerated hydrolysis at higher pH, characteristic of general acid-base catalysis. The catalytic rate constant for hydrogen ion (kH) and hydroxyl ion (kOH) were found to be 0.413 and 526.1 M-1h-1, respectively, at 318.15 K. The hydrolysis of the prodrug was found to be approximately 50-60 times faster than that of the drug. This may be attributed to the fact that hydrolysis of ester group in prodrug is assisted by keto group on the ciprofloxacin. However, there is no effect of protonation of nitrogen in piperazine ring in ciprofloxacin on the hydrolysis due to the distance from the ester moiety. Keywords: Calorimetry, stability studies, degradation kinetics, ciprofloxacin, metronidazole. International Journal of Biological and Chemical Sciences Vol. 1 (3) 2007: pp.197-210