27 research outputs found

    Development and In-Vitro Evaluation of Plantago ovata Based Rapid Disintegrating Tablets of Labetalol Hydrochloride

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    Objectives: To avoid swallowing problems of conventional tablets and improved patient compliance Plantago Ovata based Labetalol HCl Rapid disintegrating tablets have been prepared. Methods: Six different (F1 to F6) batches of Labetalol HCl Rapid disintegrating tablets were developed by ‘direct compression method’ using Plantago ovata as a natural super-disintegrating agent. The formulated RDT were tested for angle of repose’, densities like tapped and bulk density, Hausner’s ratio, Carr’s index like pre-compression parameters and for thickness, weight variation or weight uniformity, tablet hardness, % drug content or content uniformity, water absorption ratio’, time require for wetting of tablets’ means wetting time, in-vitro drug disintegration time and in-vitro drug dissolution studies under post-compression parameters of evaluation. Results: It was found that the all the results of these pre-compression and post-compression parameters comply with official standards. The drug release was determined using dissolution media of pH 6.8 phosphate buffer through in-vitro dissolution of drug. This study showed that a rapid drug release by prepared tablets. The optimized formulation F6 showed higher water absorption ratio`, lower wetting time, minimum in-vitro disintegration time’ and higher drug release amongst all the formulations. The F6 formulation was considered the best among all formulations. Conclusion: The prepared rapid disintegrating tablets shows rapid onset of action by quick drug release, minimize side effects and enhanced patient compliance. These prepared tablets containing selective alpha-1 and non-selective beta adrenergic antagonist’ drug candidate Labetalol HCl, will be very useful in the treatment of high blood pressure with enhanced bioavailability. Keywords: Rapid disintegrating tablets, Labetalol Hydrochloride, Bioavailability Enhancement, Natural Superdisintegrant, Plantago Ovata, High Blood Pressure, RDT, Patient Complianc

    Immobilization of procerain B, a cysteine endopeptidase, on amberlite MB-150 beads.

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    Proteases are involved in several crucial biological processes and reported to have important physiological functions. They also have multifarious applications in different industries. The immobilized form of the enzyme further improves its industrial applicability. Here, we report covalent immobilization of a novel cysteine endopeptidase (procerain B) on amberlite MB-150 beads through glutaraldehyde by Schiff base linkage. The immobilized product was examined extensively by Fourier Transform Infrared Spectroscopy (FTIR), Scanning electron microscopy (SEM) and Energy Dispersive X-ray (EDX) analysis. The characterization of the immobilized product showed broader pH and thermal optima compared to the soluble form of the enzyme. The immobilized form of procerain B also showed lower Km (180.27±6 µM) compared to the soluble enzyme using azocasein as substrate. Further, immobilized procerain B retains 38.6% activity till the 10(th) use, which strongly represents its industrial candidature

    Effects of pH on activity of immobilized procerain B.

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    <p>For the effect of pH on activity the substrate was also prepared in the buffers of respective pH. Stability was determined by overnight incubating the enzyme at room temperature at different pH conditions and next day activity was taken as described in method section.</p

    Biochemical characterization of immobilized procerain B.

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    <p>(<b>A</b>) Effect of temperature on activity of immobilized procerain B at pH 7.5, the substrate was also pre-incubated at respective temperatures and reactions were also carried at respective temperatures. (<b>B</b>) Effect of temperature on the stability of immobilized enzyme. For stability measurements, the enzyme was first incubated at required temperature for 15 min and then, the activity was measured at 37°C and pH 7.5. (<b>C</b>) Effect of substrate concentration on reaction velocity of immobilized procerain B. The Km value for azocaseine as substrate was calculated from the Lineweaver-Burk plot showed in subset of the graph.</p

    Physiochemical properties of Procerain B derived from amino acid sequencing data.

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    <p>Physiochemical properties of Procerain B derived from amino acid sequencing data.</p

    Confirmation of pET28<sub>(a)</sub>ProB and pET22<sub>(b)</sub>ProB clones with PCR and double digestion.

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    <p>(<b>A</b>) Lane 1 & 2 represents PCR amplified 639 bp fragment using pET28<sub>(a)</sub>ProB construct as template. M, low range DNA ruler (Bangalore Genei). Lane 3 & 4 represents PCR amplified 639 bp fragment using pET22<sub>(b)</sub>ProB construct as template. (<b>B</b>)<b>.</b> Lane 1 represents pET28<sub>(a)</sub>ProB construct digested with Nhe1. The DNA fragment of 5974 bp represents the combined size of pET-28a(+) and Procerain B cDNA. Lane M represents 1 kb DNA ladder (NEB). Lane 2 represents release of 639 bp fragment (Procerain B cDNA) on double digestion (Nhe1/BamH1) of pET28<sub>(a)</sub>ProB construct. (<b>C</b>)<b>.</b> Lane 1 represents release of 639 bp fragment (Procerain B cDNA) on double digestion (BamH1/Xho1) of pET22<sub>(b)</sub>ProB construct. Lane M represents 1 kb DNA ladder (NEB). Lane 2 represents pET22<sub>(b)</sub>ProB construct digested with BamH1. The DNA fragment of 6092 bp represents the combined size of pET-22b(+) and Procerain B cDNA. (<b>D</b>) SDS-PAGE showing the over expression of recombinant procerain B in BL21 transformed with pET28<sub>(a)</sub>ProB. Lane M represents medium range protein molecular weight marker (Bangalore Genei). Lane 1 and 2 represent supernatant and pellet of induced culture (1 mM IPTG) at 37°C. Lane 3 and 4 represent supernatant and pellet of un-induced culture at 37°C. Lane 5 & 6 represents supernatant and pellet of induced (1 mM IPTG) at 25°C. Lane 7 and 8 represent supernatant and pellet of uninduced culture at 25°C. Lane 9 represents pellet of BL21 transformed with pET28a(+) at 25°C. (<b>E</b>)<b>.</b> Expression profile of recombinant procerain B in BL21 transformed with pET22<sub>(b)</sub>ProB. Lane 1 and 2 represent pellet and supernatant of induced culture (1 mM IPTG) at 25°C. <b>M</b>. Medium range protein molecular weight marker (Bangalore Genei). Lane 3 and 4 represent supernatant and pellet of un-induced culture at 20°C. (<b>F</b>)<b>.</b> Comparison of native and recombinant procerain B (with His-tag) on SDS-PAGE. Lane 1 represents purified native procerain B. Lane M represents medium range protein molecular weight marker (Bangalore Genei). Lane 2 represents purified recombinant procerain B (with His-tag). Slight difference in molecular weight of native and recombinant procerain B is due to presence of His-tag in recombinant procerain B. Arrow in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0059806#pone-0059806-g003" target="_blank">Figure 3</a> (D, E, F) indicates recombinant procerain B.</p

    Effect of pH on immobilization of procerain B on glutaraldehyde activated Amberlite MB-150 beads in the range of pH 4–10.

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    <p>The optimum pH for immobilization of procerain B was nearly 8.0 with 52.65% immobilization.</p

    Reusability of procerain B immobilized on amberlite MB-150 beads.

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    <p>The reusability of was tested by repeated use of same amberlite beads. After every use the beads were washed with Tris-HCl buffer pH 8 and reused for next batch of reaction.</p

    Primers used in cloning of procerain B.

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    <p>Primers used in cloning of procerain B.</p

    Comparison of FTIR spectra of normal and glutaraldehyde activated Amberlite beads.

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    <p>For FTIR analysis the beads were crushed with KBr and compressed to form a thin pellet. The pellet was used for FTIR analysis. (<b>A</b>) FTIR spectra of normal Amberlite beads. (<b>B</b>) FTIR spectra of glutaraldehyde activated Amberlite beads. Both spectra were compared for confirmation of glutaraldehyde activation of beads. The peaks at 2925, 1453 and 1121 are due to amberlite. The increase in 1637 peak intensity is due to activation of bead with glutaraldehyde.</p
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