Dataset Information

Lime and Sulphide-Free Dehairing of Animal Skin Using Collagenase-Free Alkaline Protease from Vibrio metschnikovii NG155.

ABSTRACT: The objective of this work was to isolate a microorganism producing alkaline protease that can be used as an ecofriendly alternative to chemicals in dehairing process of leather manufacture. Alkaline protease producing bacterium Vibrio metschnikovii NG155 was isolated from soil samples of leather industry. The protease was highly effective in dehairing of goat skin, completely eliminating the use of lime and sulfide. Histological studies of the skin after dehairing showed that the enzyme did not damage the collagen layer and brought good fiber opening. Absence of collagenase activity was confirmed by reacting pure collagen with the enzyme and analyzing it on SDS PAGE, which showed no degradation of collagen. The enzyme was stable in a wide range of pH (7-11) and temperature (10-50 °C), which makes it suitable for industrial application.

SUBMITTER: George N

PROVIDER: S-EPMC4188488 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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Lime and Sulphide-Free Dehairing of Animal Skin Using Collagenase-Free Alkaline Protease from Vibrio metschnikovii NG155.

George Nancy N Sondhi Sonica S Soni Sanjeev Kumar SK Gupta Naveen N

Indian journal of microbiology 20131206 2

The objective of this work was to isolate a microorganism producing alkaline protease that can be used as an ecofriendly alternative to chemicals in dehairing process of leather manufacture. Alkaline protease producing bacterium Vibrio metschnikovii NG155 was isolated from soil samples of leather industry. The protease was highly effective in dehairing of goat skin, completely eliminating the use of lime and sulfide. Histological studies of the skin after dehairing showed that the enzyme did not ...[more]

PMID: 25320413

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Project description:In neuroscience, stereotactic brain surgery is a standard yet challenging technique for which laboratory and veterinary personnel must be sufficiently and properly trained. There is currently no animal-free training option for neurosurgeries; stereotactic techniques are learned and practiced on dead animals. Here we have used three-dimensional (3D) printing technologies to create rat and mouse skin-skull-brain models, specifically conceived for rodent stereotaxic surgery training. We used 3D models obtained from microCT pictures and printed them using materials that would provide the most accurate haptic feedback for each model-PC-ABS material for the rat and Durable resin for the mouse. We filled the skulls with Polyurethane expanding foam to mimic the brain. In order to simulate rodent skin, we added a rectangular 1mm thick clear silicone sheet on the skull. Ten qualified rodent neurosurgeons then performed a variety of stereotaxic surgeries on these rat and mouse 3D printed models. Participants evaluated models fidelity compared to cadaveric skulls and their appropriateness for educational use. The 3D printed rat and mouse skin-skull-brain models received an overwhelmingly positive response. They were perceived as very realistic, and considered an excellent alternative to cadaveric skulls for training purposes. They can be made rapidly and at low cost. Our real-size 3D printed replicas could enable cost- and time-efficient, animal-free neurosurgery training. They can be absolute replacements for stereotaxic surgery techniques practice including but not limited to craniotomies, screw placement, brain injections, implantations and cement applications. This project is a significant step forward in implementing the replacement, reduction, and refinement (3Rs) principles to animal experimentation. These 3D printed models could lead the way to the complete replacement of live animals for stereotaxic surgery training in laboratories and veterinary studies.

Project description:1. The neutral collagenase released into the culture medium by explants of human skin tissue was purified by ultrafiltration and column chromatography. The final enzyme preparation had a specific activity against thermally reconstituted collagen fibrils of 32mug of collagen degraded/min per mg of enzyme protein, representing a 266-fold increase over that of the culture medium. Electrophoresis in polyacrylamide disc gels showed it to migrate as a single protein band from which enzyme activity could be eluted. Chromatographic and polyacrylamide-gel-elution experiments provided no evidence for the existence of more than one active collagenase. 2. The molecular weight of the enzyme estimated from gel filtration and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis was approx. 60000. The purified collagenase, having a pH optimum of 7.5-8.5, did not hydrolyse the synthetic collagen peptide 4-phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-d-Arg-OH and had no non-specific proteinase activity when examined against non-collagenous proteins. 3. It attacked undenatured collagen in solution at 25 degrees C, producing the two characteristic products TC(A)((3/4)) and TC(B)((1/4)). Collagen types I, II and III were all cleaved in a similar manner by the enzyme at 25 degrees C, but under similar conditions basement-membrane collagen appeared not to be susceptible to collagenase attack. At 37 degrees C the enzyme attacked gelatin, producing initially three-quarter and one-quarter fragments of the alpha-chains, which were degraded further at a lower rate. As judged by the release of soluble hydroxyproline peptides and electron microscopy, the purified enzyme degraded insoluble collagen derived from human skin at 37 degrees C, but at a rate much lower than that for reconstituted collagen fibrils. 4. Inhibition of the skin collagenase was obtained with EDTA, 1,10-phenanthroline, cysteine, dithiothreitol and sodium aurothiomaleate. Cartilage proteoglycans did not inhibit the enzyme. The serum proteins alpha(2)-macroglobulin and beta(1)-anti-collagenase both inhibited the enzyme, but alpha(1)-anti-trypsin did not. 5. The physicochemical and enzymic properties of the skin enzyme are discussed in relation to those of other human collagenases.

Dataset Information

Lime and Sulphide-Free Dehairing of Animal Skin Using Collagenase-Free Alkaline Protease from Vibrio metschnikovii NG155.

Publications

Lime and Sulphide-Free Dehairing of Animal Skin Using Collagenase-Free Alkaline Protease from Vibrio metschnikovii NG155.

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