A possible mechanism for enzymic depilation of skins
- Authors: Brady, Dean
- Date: 1989
- Subjects: Chemistry, Technical , Tanning
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3877 , http://hdl.handle.net/10962/d1001611
- Description: Streptomyces fradiae is a bacterium which has been previously found to produce extracellular enzymes which are capable of wool degradation and skin depilation. Streptomyces fradiae 3739 and other strains of Streptomyces were found in this study to be able to degrade a keratin source (wool) to a considerable degree. However according to the evidence of SEM micrographs presented here the highly keratinised spindle cells of the paracortex are fairly resistant to protease attack, and it is the cementation material which binds these cells together which is initially degraded by the proteases. A large degree of correlation was found with the strains of Streptomyces studied, between the ability of the individual strains to degrade wool and the ability of their extracellular proteases to reduce the depilation load of sheepskins. With further analysis S. fradiae 3739 was found to produce at least one amylase and four or more proteases. The proteases as a group had maximal proteolytic activity in the 8.0-9.0 pH unit range, and were considerably thermostabilised by the inclusion of calcium ions into the reaction solution. The protease group was found to cause depilation of merino sheepskins. For comparative purposes a protease produced by a strain of Proteus vulgaris isolated from a staling hide with hair slip (natural depilation) was studied. The protease activity was maximal in the alkaline region between 8.0-9.0 pH units. Tbe protease appeared to be a single enzyme with a molecular mass of approximately 44 000 daltons. The protease was maximally active at 40°C, although it was only thermostable to 30°C. The enzyme was ineffectual as a depilant except when the skin was pre-treated with a strong alkali, preferably including sodium sulphite in the protease preparation. One of the most important differences between the extracellular proteases of S. fradiae and P. vulgaris was that the former were greater in variety and caused a greater decrease in the depilation load of sheepskins than the latter. Further research with mixtures of commercial proteases provided evidence that a synergistic depilatory effect occurs when proteases of complementary bond specificities are used in conjunction in enzymic depilatory preparations. Some form of strong alkali treatment of skins was found to be necessary to produce leather of the prerequisite quality when the skin was depilated by proteases, otherwise the skin was found to be depleted and stiff. Calcium hydroxide alone was found to be inadequate for this task, probably owing to the fact that it is less alkaline than the lime-sulphide mixture. The calcium hydroxide (lime) must therefore be used in conjunction with sodium hydroxide (which makes the solution as alkaline as that of the lime-sulphide solution) to produce leather comparable to that produced by the lime sulphide treatment. A combination of the information provided by the present research and that gleaned from the relevent literature allows for the construction of a model to represent the possible mechanism of enzymic depilation of skins, in which depilation is caused by the disruption of the basement membrane at the dermal-epidermal junction by the degradation of its constituent molecular components by general proteases, resulting in the removal of the epidermis and its associated wool or hair
- Full Text:
- Authors: Brady, Dean
- Date: 1989
- Subjects: Chemistry, Technical , Tanning
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3877 , http://hdl.handle.net/10962/d1001611
- Description: Streptomyces fradiae is a bacterium which has been previously found to produce extracellular enzymes which are capable of wool degradation and skin depilation. Streptomyces fradiae 3739 and other strains of Streptomyces were found in this study to be able to degrade a keratin source (wool) to a considerable degree. However according to the evidence of SEM micrographs presented here the highly keratinised spindle cells of the paracortex are fairly resistant to protease attack, and it is the cementation material which binds these cells together which is initially degraded by the proteases. A large degree of correlation was found with the strains of Streptomyces studied, between the ability of the individual strains to degrade wool and the ability of their extracellular proteases to reduce the depilation load of sheepskins. With further analysis S. fradiae 3739 was found to produce at least one amylase and four or more proteases. The proteases as a group had maximal proteolytic activity in the 8.0-9.0 pH unit range, and were considerably thermostabilised by the inclusion of calcium ions into the reaction solution. The protease group was found to cause depilation of merino sheepskins. For comparative purposes a protease produced by a strain of Proteus vulgaris isolated from a staling hide with hair slip (natural depilation) was studied. The protease activity was maximal in the alkaline region between 8.0-9.0 pH units. Tbe protease appeared to be a single enzyme with a molecular mass of approximately 44 000 daltons. The protease was maximally active at 40°C, although it was only thermostable to 30°C. The enzyme was ineffectual as a depilant except when the skin was pre-treated with a strong alkali, preferably including sodium sulphite in the protease preparation. One of the most important differences between the extracellular proteases of S. fradiae and P. vulgaris was that the former were greater in variety and caused a greater decrease in the depilation load of sheepskins than the latter. Further research with mixtures of commercial proteases provided evidence that a synergistic depilatory effect occurs when proteases of complementary bond specificities are used in conjunction in enzymic depilatory preparations. Some form of strong alkali treatment of skins was found to be necessary to produce leather of the prerequisite quality when the skin was depilated by proteases, otherwise the skin was found to be depleted and stiff. Calcium hydroxide alone was found to be inadequate for this task, probably owing to the fact that it is less alkaline than the lime-sulphide mixture. The calcium hydroxide (lime) must therefore be used in conjunction with sodium hydroxide (which makes the solution as alkaline as that of the lime-sulphide solution) to produce leather comparable to that produced by the lime sulphide treatment. A combination of the information provided by the present research and that gleaned from the relevent literature allows for the construction of a model to represent the possible mechanism of enzymic depilation of skins, in which depilation is caused by the disruption of the basement membrane at the dermal-epidermal junction by the degradation of its constituent molecular components by general proteases, resulting in the removal of the epidermis and its associated wool or hair
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Cleavage of the precursor coat protein of black beetle virus strain w17 in rabbit reticulocyte lysate
- Authors: Blackhurst, Diane Mary
- Date: 1988
- Subjects: Beetles , Insects -- Viruses
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3880 , http://hdl.handle.net/10962/d1001614
- Description: Black beetle virus (BBV) is a bipartite single-stranded RNA virus belonging to the family Nodaviridae. Its host range has been found to be limited to insects. RNA 1, the larger of the two RNA molecules, with a MW of 1,15 x 10⁶ and the smaller RNA 2 with a MW of 0,46 x 10⁶, are both packaged in the same virus particle. The two RNA molecules are translated separately, with RNA 1 coding for protein A of MW 105 x 10³ and RNA 2 coding for protein α of MW 47 x 10³. Protein α is the major capsid protein precursor, which during in vivo maturation is cleaved to form the coat protein β of MW 43 x 10³, and protein γ of MW 5 x 10³. Cell-free translation of BBV (strain W17) mRNA was carried out in rabbit reticulocyte lysates. Protein α was detectable between 0 and 30 minutes after RNA addition. A protein 'β', which was found to co-electrophorese on polyacrylamide gels with authentic β and which was immunoprecipitated by anti-BBV antiserum, was detectable after 30 minutes. Results of this work show that the formation of 'β' could be prevented by the addition of RNase to the lysate, indicating that intact RNA is necessary for α to β cleavage. Arresting protein synthesis by the addition of cycloheximide to the lysate mix did not inhibit the cleavage. The formation of β could also be prevented by cooling the lysate mix to 1°C. Cleavage of α to β still occurred when RNA 2, without the presence of RNA 1, was translated. Therefore the cleavage is not dependent on a translation product of RNA 1. Sedimentation of lysate on sucrose density gradients showed that α to β cleavage was not accompanied by assembly of BBV RNA and protein lnto a viral substructure as has been shown to occur with some viruses, for example certain picornaviruses. Serial dilution of lysate containing α showed that the level of β decreased with increasing dilution, indicating that the cleavage is not mediated by autocatalysis, but by some other unknown factor. Although much work has been carried out on black beetle virus, no work has been published to date concerning α to β cleavage as an indication of assembly in rabbit reticulocyte lysates. Results of these cell-free translation experiments thus indicate that BBV coat protein precursor α, in association with its messenger RNA 2, undergoes a maturation cleavage in the lysate to produce BBV coat protein β. In addition, this cleavage seems to occur without assembly into any intermediate viral structure
- Full Text:
- Authors: Blackhurst, Diane Mary
- Date: 1988
- Subjects: Beetles , Insects -- Viruses
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3880 , http://hdl.handle.net/10962/d1001614
- Description: Black beetle virus (BBV) is a bipartite single-stranded RNA virus belonging to the family Nodaviridae. Its host range has been found to be limited to insects. RNA 1, the larger of the two RNA molecules, with a MW of 1,15 x 10⁶ and the smaller RNA 2 with a MW of 0,46 x 10⁶, are both packaged in the same virus particle. The two RNA molecules are translated separately, with RNA 1 coding for protein A of MW 105 x 10³ and RNA 2 coding for protein α of MW 47 x 10³. Protein α is the major capsid protein precursor, which during in vivo maturation is cleaved to form the coat protein β of MW 43 x 10³, and protein γ of MW 5 x 10³. Cell-free translation of BBV (strain W17) mRNA was carried out in rabbit reticulocyte lysates. Protein α was detectable between 0 and 30 minutes after RNA addition. A protein 'β', which was found to co-electrophorese on polyacrylamide gels with authentic β and which was immunoprecipitated by anti-BBV antiserum, was detectable after 30 minutes. Results of this work show that the formation of 'β' could be prevented by the addition of RNase to the lysate, indicating that intact RNA is necessary for α to β cleavage. Arresting protein synthesis by the addition of cycloheximide to the lysate mix did not inhibit the cleavage. The formation of β could also be prevented by cooling the lysate mix to 1°C. Cleavage of α to β still occurred when RNA 2, without the presence of RNA 1, was translated. Therefore the cleavage is not dependent on a translation product of RNA 1. Sedimentation of lysate on sucrose density gradients showed that α to β cleavage was not accompanied by assembly of BBV RNA and protein lnto a viral substructure as has been shown to occur with some viruses, for example certain picornaviruses. Serial dilution of lysate containing α showed that the level of β decreased with increasing dilution, indicating that the cleavage is not mediated by autocatalysis, but by some other unknown factor. Although much work has been carried out on black beetle virus, no work has been published to date concerning α to β cleavage as an indication of assembly in rabbit reticulocyte lysates. Results of these cell-free translation experiments thus indicate that BBV coat protein precursor α, in association with its messenger RNA 2, undergoes a maturation cleavage in the lysate to produce BBV coat protein β. In addition, this cleavage seems to occur without assembly into any intermediate viral structure
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The study of hydroxyoximes and hydroxamic acids supported on macroporous resins and their use in the rapid seperation of metals
- Authors: Hemmes, Marlene
- Date: 1979
- Subjects: Chromatographic analysis , Separation (Technology) , Metal ions , Solvent extraction , Extraction (Chemistry) , Oximes , Metals -- Analysis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4453 , http://hdl.handle.net/10962/d1009512
- Description: Introduction: The macroporous Amberlite XAD resins were coated with LIX-64N and examined for the rate of uptake of copper . XAD-7 was by far the best support and gave a satisfactory rate of uptake up to loadings of 60% (w/w). The specific surface area of XAD-7 was measured by the adsorption of methylene blue from aqueous solution. The area of the wetted resin was five times less than that of the dry resin. LIX-65N was purified and the anti isomer characterised using spectroscopic techniques . The rate of uptake of copper was not improved by use of purified LIX-65N or by addition of LIX-63. XAD-7 coated with LIX-65N was used in columns. Elution curves for copper showed negligible tailing, and rapid separations of copper from iron (111), nickel, cobalt and magnesium by selective absorption were achieved. Copper was concentrated from very dilute solution at a flow rate of 50 ml min -1 ,and a 99% recovery was obtained. The method was applied to the rapid determination of copper in brass and bronze. A series of long-chain hydroxamic acids were synthesised and tested for suitability as stationary phase on XAD-7. Oleohydroxamic acid and naphthenohydroxamic acid were the most promising. The r ate of uptake of copper was reduced by the use of nonylphenol or amyl alcohol as a diluent. The capacities for copper of the hydroxamic acids were less when supported on XAD-7 than when used as liquid ionexchangers. The distribution coefficients of cobalt, nickel, zinc, lead, vanadium, uranium, iron (111) and copper were measured as a function of pH. XAD-7 coated with oleohydroxamic acid was used in columns for the rapid separation of iron (111) from copper and of copper from nickel, cobalt, lead and zinc. Copper was concentrated from very dilute solution at a flow rate of 45 ml min -1 and a 100,8% recovery was obtained. Copper was successfully separated from nickel by selective elution. The elution curves obtained show negligible tailing. The resin loaded with oleohydroxamic acid lost capacity due to chemical instability. Naphthenohydroxamic acid supported on XAD-7 was not suitable for use in columns, because it was physically unstable.
- Full Text:
- Authors: Hemmes, Marlene
- Date: 1979
- Subjects: Chromatographic analysis , Separation (Technology) , Metal ions , Solvent extraction , Extraction (Chemistry) , Oximes , Metals -- Analysis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4453 , http://hdl.handle.net/10962/d1009512
- Description: Introduction: The macroporous Amberlite XAD resins were coated with LIX-64N and examined for the rate of uptake of copper . XAD-7 was by far the best support and gave a satisfactory rate of uptake up to loadings of 60% (w/w). The specific surface area of XAD-7 was measured by the adsorption of methylene blue from aqueous solution. The area of the wetted resin was five times less than that of the dry resin. LIX-65N was purified and the anti isomer characterised using spectroscopic techniques . The rate of uptake of copper was not improved by use of purified LIX-65N or by addition of LIX-63. XAD-7 coated with LIX-65N was used in columns. Elution curves for copper showed negligible tailing, and rapid separations of copper from iron (111), nickel, cobalt and magnesium by selective absorption were achieved. Copper was concentrated from very dilute solution at a flow rate of 50 ml min -1 ,and a 99% recovery was obtained. The method was applied to the rapid determination of copper in brass and bronze. A series of long-chain hydroxamic acids were synthesised and tested for suitability as stationary phase on XAD-7. Oleohydroxamic acid and naphthenohydroxamic acid were the most promising. The r ate of uptake of copper was reduced by the use of nonylphenol or amyl alcohol as a diluent. The capacities for copper of the hydroxamic acids were less when supported on XAD-7 than when used as liquid ionexchangers. The distribution coefficients of cobalt, nickel, zinc, lead, vanadium, uranium, iron (111) and copper were measured as a function of pH. XAD-7 coated with oleohydroxamic acid was used in columns for the rapid separation of iron (111) from copper and of copper from nickel, cobalt, lead and zinc. Copper was concentrated from very dilute solution at a flow rate of 45 ml min -1 and a 100,8% recovery was obtained. Copper was successfully separated from nickel by selective elution. The elution curves obtained show negligible tailing. The resin loaded with oleohydroxamic acid lost capacity due to chemical instability. Naphthenohydroxamic acid supported on XAD-7 was not suitable for use in columns, because it was physically unstable.
- Full Text:
The development of a morphometric model for the estimation of mean annual sediment yield in ungauged catchments of South African river systems
- Authors: Roberts, Peter J T
- Date: 1975
- Subjects: Sedimentation and deposition -- South Africa -- Mathematical models
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4876 , http://hdl.handle.net/10962/d1013062
- Description: Hydrologists are regularly faced with the unenviable task of having to predict the magnitude and frequency of phenomena such as floods and droughts; and rates of erosion. If long records are available for analysis the hydrologist is able to base his predictions on the premise that the pattern of variation that has been observed in the past will persist in the future. The confidence that can be placed in any estimate consequently depends to a large extent on the length of time over which the phenomena have been measured at the problem site. Unfortunately the availability of adequate records tends to be the exception rather than the rule and in areas where there is inadequate data, it is necessary to resort to the hazardous procedure of transferring information from the gauged to the ungauged catchments. The transfer of information is accomplished by using empirical methods based on regionalised parameters, but the uncertainties involved together with the economic implications that could arise from a poor estimate, prompt the hydrologist to use as many methods as possible. The need for empirical methods of estimating mean annual sediment yield in ungauged catchments was first appreciated by the author when he was involved in the estimation of design floods and sediment accumulation at sites for proposed reservoirs. Empirical methods of estimating sediment yield are frequently used in an engineering context, but little attention has been given to the catchment surface from which the sediment supply is derived. It is perhaps in this often neglected field of research that the physical geographer can make a contribution. The principal aim of the thesis, more fully discussed in Chapter I, was the development of a morphometric model which could be used to estimate mean annual sediment yield in ungauged catchments in South Africa. The data used in the development of the model were drawn from the catchments, described in Appendix A, that cover a wide range of climate and topography. A description of the approaches adopted by other researchers for the development of empirical models of estimating sediment yield which forms the background to the model has been included as Appendix B. The model was first developed in an elementary form as the focus of a research project which was documented in the form of three reports of research in progress (Roberts, 1973 a, b and c). Analysis of the pattern of variation of suspended sediment yield provided a better understanding of factors affecting sediment yield and supported the selection of the prediction variable (Horton's P ratio) which was used in the model. The concepts of network topology were utilised to gain insight into the environmental factors controlling both the P ratio and sediment yield. Reasons for the high correlation between the P ratio and sediment yield are suggested but it is felt that further research should be focused on this aspect. In order not to break the continuity and development of the steps taken in the derivation of the model details of the calculations are collected in Appendices C, D and E. While many of the figures and tables presented in the thesis appeared in technical notes prepared entirely by the author for the Department of Water Affairs, the views expressed in the thesis do not in any way, either explicitly or by implication, represent any official view or policy of the Department of Water Affairs.
- Full Text:
- Authors: Roberts, Peter J T
- Date: 1975
- Subjects: Sedimentation and deposition -- South Africa -- Mathematical models
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4876 , http://hdl.handle.net/10962/d1013062
- Description: Hydrologists are regularly faced with the unenviable task of having to predict the magnitude and frequency of phenomena such as floods and droughts; and rates of erosion. If long records are available for analysis the hydrologist is able to base his predictions on the premise that the pattern of variation that has been observed in the past will persist in the future. The confidence that can be placed in any estimate consequently depends to a large extent on the length of time over which the phenomena have been measured at the problem site. Unfortunately the availability of adequate records tends to be the exception rather than the rule and in areas where there is inadequate data, it is necessary to resort to the hazardous procedure of transferring information from the gauged to the ungauged catchments. The transfer of information is accomplished by using empirical methods based on regionalised parameters, but the uncertainties involved together with the economic implications that could arise from a poor estimate, prompt the hydrologist to use as many methods as possible. The need for empirical methods of estimating mean annual sediment yield in ungauged catchments was first appreciated by the author when he was involved in the estimation of design floods and sediment accumulation at sites for proposed reservoirs. Empirical methods of estimating sediment yield are frequently used in an engineering context, but little attention has been given to the catchment surface from which the sediment supply is derived. It is perhaps in this often neglected field of research that the physical geographer can make a contribution. The principal aim of the thesis, more fully discussed in Chapter I, was the development of a morphometric model which could be used to estimate mean annual sediment yield in ungauged catchments in South Africa. The data used in the development of the model were drawn from the catchments, described in Appendix A, that cover a wide range of climate and topography. A description of the approaches adopted by other researchers for the development of empirical models of estimating sediment yield which forms the background to the model has been included as Appendix B. The model was first developed in an elementary form as the focus of a research project which was documented in the form of three reports of research in progress (Roberts, 1973 a, b and c). Analysis of the pattern of variation of suspended sediment yield provided a better understanding of factors affecting sediment yield and supported the selection of the prediction variable (Horton's P ratio) which was used in the model. The concepts of network topology were utilised to gain insight into the environmental factors controlling both the P ratio and sediment yield. Reasons for the high correlation between the P ratio and sediment yield are suggested but it is felt that further research should be focused on this aspect. In order not to break the continuity and development of the steps taken in the derivation of the model details of the calculations are collected in Appendices C, D and E. While many of the figures and tables presented in the thesis appeared in technical notes prepared entirely by the author for the Department of Water Affairs, the views expressed in the thesis do not in any way, either explicitly or by implication, represent any official view or policy of the Department of Water Affairs.
- Full Text:
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