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Enzyme unhairing - An eco-friendly biotechnological process
Abstract
Global environmental regulations are changing the leather industry. Conventional unhairing contributes most of the pollution in the industry, such as hydrogen sulfide and lime. Ever-increasing attention to the environmental impact of the leather industry has necessitated the development of enzymatic unhairing as a potent alternative to polluting chemicals. Enzymes are gaining increasing importance in the unhairing process, eliminating the need for sodium sulfide. This review discusses the enzymatic unhairing used in leather processing and introduces more information about enzymatic unhairing. This paper also summarises progress of enzymatic unhairing and looks ahead at its application prospect and development direction.
- ... Accordingly there are two most important reasons to develop hair-saving unhairing: i) the products of hair degradation complicate the cleaning of the unhairing solutions used, and ii) the saved hair can be used as a raw material for other applications [3]. A major part of hair-saving unhairing process investigations is devoted to enzymatic unhairing [4,5]. Despite the fact that the enzymatic process has a serious advantage from an environmental point of view, i.e. wastewaters after the process are free from sulphides and hair degradation products, pure enzymatic unhairing is not widely used in the industry. ...Since the environmental requirements for leather manufacture as well as for other industry branches are becoming stricter, investigations of unhairing have been directed towards conditions of the process which allow the saving of hair: enzymatic unhairing or unhairing with hair immunization conventionally achieved by using lime and sodium sulphide. The pro- blem is that lime forms a big quantity of liquid waste which contains lime sludge polluted by sulphides and protein degradation products. The research is devoted to the replacement of Ca(OH) 2 as hair immunization material with some other soluble non-hazardous material with a similar immunization effect. The preliminary tests showed that alkaline sodium alu- minate suits this purpose very well. Its immunisation efficiency increases with prolonged treatment duration and increased concentration of the treatment solution. Treatment with 20 g/l of sodium silicate solution for 3 h allows to reach a high immunization ability almost the same as that of calcium hydroxide under the same conditions. The immunisation effect lasts when the pH is approx. 13.
- ... Kinetics leading to enzymatic unhairing was carried out by other researchers that described linearity between release of total protein and square root of time at the initial stage that suggests that enzymatic unhairing by protease was a diffusion controlled process at the initial stage. The results showed that release of total protein further confirms that it is a hydrolysis of core protein by protease that leads to the degradation of proteoglycans and release of protein and saccharides (Song et al., 2011;Wang et al., 2009). It has been found from literature that several commercial enzyme formulations were available in combination with hydrosulfide and peroxides for better results. ...The leather industry deals with proteinous skin material for the conversion of leather and this generates huge amount of solid and liquid wastes giving rise to pollution that needs to be overcome by introducing sustainable cleaner technologies. This review describes various eco-friendly challenges and major achievements for abatement of pollution in leather processing. Different cleaner technological methods in preservation of raw hides/skins, unhairing, tanning and dyeing operations are discussed here. Cleaner-preservation techniques by using chemicals and biological agents have been developed to reduce pollution problems of salt up to a great extent in leather processing operations. Process intensified operations have helped to achieve better uptake and to manage pollution load. Enzymatic dehairing are implemented to reduce Bio-chemical-Oxygen-Demand (BOD) at the level of 40%, Chemical-Oxygen-Demand (COD) up to 50%. Improved biological methods for bio-degradation of dyes, azo-dyes and their mixtures and to reuse the liquors in the process to reduce the dye pollution load in the effluent streams are reviewed. Nano-Particle polymers and improved retanning materials have been synthesized for high exhaustion of dyeing and retanning properties. Mathematical models predicting kinetics and growth for the above processes are also reviewed. It could be concluded by utilization of these technologies, a possible reduction in pollution loads such as BOD & COD, upto levels of 50% & 40%, could be achieved in leather processing.
- ... The good linearity suggests that enzymatic unhairing by protease was a diffusion controlled process at the initial stage. The results showed that released total protein further confirms that it is a hydrolysis of core protein by protease that leads to the degradation of proteoglycans and release of protein and saccharides (Jians et al. 2010;Song et al. 2011;Wang et al. 2009). ...Dehairing is one of the important operations in leather processing where hair is removed by using suitable chemicals. Conventional dehairing is based on application of huge amount of chemicals such as lime and sodium sulfide that generate lot of pollution problem. Enzyme assisted dehairing is the eco-friendly option that reduces nearly 40% of BOD and 50% of COD in leather processing. For this; an alkaline protease producing strains namely Bacillus subtilis was isolated from fresh goat skins and the enzyme was extracted. The enzyme was applied on flesh side goat skins at different concentration (1–4% with 10% water) and dehairing was carried out. The complete dehairing was achieved at 2% enzyme for duration of 16 h. The pollution loads such as BOD, COD, TSS, and TDS were reduced at the level of 62.8, 79.0, 88.2, and 82.5% in comparison with the control sample. The enzymatic dehaired leather was analyzed by scanning electron microscopic (SEM) studies. The SEM analyses revealed similar fiber orientation and compactness compared with control sample. The color measurements of the enzyme dehaired leather showed better color properties than control sample.
- Recently, increasing interests have been directed towards incorporating nanotechnologies into leather manufacturing, which offer cost-effective improvements in leather performances, as well as enabling the industry to meet stricter legislation regarding environmental safety. In order to guide the leather technologists and chemists worldwide, this article reviews state of the art related to existing nanotechnologies in leather manufacturing processes, including tanning, finishing and effluent treatment, over the past decade. The emphasis is placed on the mechanism by which nanotechnologies improve conventional processes, impressive performances of the resultant leather and possible obstacles that cloud technical development in this field. In the end, potential health and environment risks associated with the incorporation of nanotechnologies, which are particularly important for daily necessities such as leather but have been basically overlooked by many researchers involved, are also addressed.
- Although the loss of scalp hair is distressing and many medical treatments focus on its restoration, the removal of body hair has been adopted since ancient times. Beauty standards, which r eflect the culture of each society, have been presenting the depilated body as absolutely desirable. Through the ages various methods of hair removal have been used depending on the requirements of the individuals. In recent years, Laser and Intense Pulse Light devices have been considered as the most promising solution for excess hair growth, without excluding the efficacy of other methods to induce satisfactory epilatory results. The enzyme-based hair removal method has received little recognition even though experimental and clinical data support its efficacy to provide long term or even permanent epilation. The present review presents these data and examines the likelihood of considering the aforementioned method as ideal.
- The enzymatic treatment of hides in tannery processes is a promising technology. However, the reaction kinetics of commercial enzymes available to the leather industry are not fully understood and their activities have been mainly determined with model proteins such as casein as substrate, which are not of direct relevance for cattle hides. Therefore, it is important to determine their activities on collagen and keratin, the main proteins of skin, in order to use these enzymes in leather processing. This work describes the study of five proteases, used commercially in tanneries, to assess their ability to act upon collagen and keratin and to determine their unhairing. Results showed that all commercial enzymes tested had more activity on collagen than on keratin. Unhairing was also tested and four out of the five enzymes tested showed some unhairing activity. Optima of the temperature and pH of the enzymes were very similar for all five enzymes, with maximal activities around 55ºC and pH 9 to 12, respectively.
- Usually, beamhouse processes are carried out by varying a medium from strongly alkaline (unhairing-liming) up to strongly acid (pickling). This study is designed to develop a preparation of hide for tanning via processes that avoid a sharp change of hide pH. Enzymes active in acid medium are employed for hide unhairing in a buffer system containing 2.5% acetic acid and 0.3% sodium acetate. An oxidative treatment with peracetic acid then allows the complete removal of residual hair and scud. During both processes, unhairing and oxidative treatment, the opening of the derma structure occurs: 13.3-14.6g of non-collagen proteins are removed and amount of dermatan sulphate decreases by 35%. The hide obtained can be chromed directly after the above processes and subsequent treatment with sodium chloride solution omitted conventional pickling. The pH of the hide varies in the range of 8.3–3.7 during the processes of preparation for tanning.
- We have developed a sensitive and highly efficient whole-cell methodology for quantitative analysis and screening of protease activity in vivo. The method is based on the ability of a genetically encoded protease to rescue a coexpressed short-lived fluorescent substrate reporter from cytoplasmic degradation and thereby confer increased whole-cell fluorescence in proportion to the protease's apparent activity in the Escherichia coli cytoplasm. We demonstrated that this system can reveal differences in the efficiency with which tobacco etch virus (TEV) protease processes different substrate peptides. In addition, when analyzing E. coli cells expressing TEV protease variants that differed in terms of their in vivo solubility, cells containing the most-soluble protease variant exhibited the highest fluorescence intensity. Furthermore, flow cytometry screening allowed for enrichment and subsequent identification of an optimal substrate peptide and protease variant from a large excess of cells expressing suboptimal variants (1:100,000). Two rounds of cell sorting resulted in a 69,000-fold enrichment and a 22,000-fold enrichment of the superior substrate peptide and protease variant, respectively. Our approach presents a new promising path forward for high-throughput substrate profiling of proteases, engineering of novel protease variants with desired properties (e.g., altered substrate specificity and improved solubility and activity), and identification of protease inhibitors.
- In this new article of the series "Hair saving unhairing process" the development of the theme of enzyme unhairing is initiated. It gathers three curious features: it has been extensively studied by many researchers in different countries during the last century, it is scarcely used in the industry and it still keeps the attraction, as a challenge, to keep on calling together the interest of research groups that find in this process the ideal unhairing process. Enzymatic unhairing will be the ideal unhairing process when the enzyme specificity is guided specially towards the components of the epidermis system, its action on the structured proteins-especially the collagen-is minimized or avoided, and when leather is developed with desirable properties. The aim of this first article on the enzyme unhairing is to show with no intention of being exhaustive-the research evolution of those work groups that showed relevant activity, adding in each of the sections comments and complementary suggestions from other work. Experimental studies and theoretical speculation at the Leather Research and Development Centre (CITEC) on the hair-saving unhairing process, particularly enzyme unhairing, tries to contribute to better knowledge on mechanisms (diffusive and chemical) involved in the unhairing, with the purpose of proposing the design of enzyme preparations to bring us closer to ideal unhairing. The introduction of the research evolution on the enzyme unhairing, the behaviour of enzyme preparations on different substrates, the mechanisms that can take place in the process and the desirable/undesirable properties of resultant leather come before the presentation of the experimental results that are being obtained at CITEC. Thus, we will show in a further article, the results of the characterisation of commercially available enzyme preparations through the evaluation of proteolytic activities against a representative substrate of animal hide components and the activation/inhibition effects that an unhairing sy stem has on enzymes.
- Since the time of Henry Procter, there have been important developments in early stage processing of hides and skins but there have been no fundamental changes in the process. The developments have mainly been the result of economic and environmental pressures to reduce labour requirements and processing times and to meet environmental regulations. The major unhairing developments have been the rapid hair-destroying drum processes and the subsequent rapid hair-saving systems. The promise of enzymes, amines and other systems for unhairing has not yet been fulfilled. Despite enormous research effort, most unhairing and dewooling processes still use lime and sulphides. It can be argued that unhairing is now the least environmentally acceptable tannery process. Although the adoption of one of the several rapid hair-saving processes now available significantly reduces effluent loads, a better, cleaner system for unhairing is still needed. Ideally, the hair should be recovered in a fibrous form in a sulphide-free process. Unless handled appropriately, sulphides create toxicity, odour and effluent problems. Exposure standards for gases and vapours in the occupational environment and industrial odour emission limits, incorporating plume modelling, are being enforced. Unhairing systems must be effective on all hides and skins irrespective of breed, age, sex, nutrition and season. Skin and hair structure and depilation processes and their environmental effects are reviewed.
- Alkaline protease isolated from a new strain Alcaligenes faecalis was evaluated for its unhairing potential in leather processing. The enzyme is relatively stable in the pH range of 8-11 and at temperature up to 30°C for 24 h. The enzyme can be stored at 0-4°C up to 6 months without much loss of activity. Unhairing action was found to be best at pH 9 and at the temperature of 30 - 37°C with 0.5% w/w enzyme concentration. Microscopic studies of the unhaired skins and the physical properties of the enzyme-treated finished leathers were well comparable with leather processed by conventional methods. These results indicate that the protease produced by A. faecalis is suitable for unhairing and can be exploited as an eco-friendly unhairing agent in leather processing.
- The conventional process of unhairing animal skins/hides by the lime-sulphide process generates enormous problems in tannery effluent treatment. It has been established that enzymatic unhairing is an environmentally friendly alternative to the conventional process. The enzyme, an alkaline protease (Clarizyme) is produced by Aspergillus flavus isolate using solid substrate fermentation (SSF). Since SSF has distinct economic advantages over conventional methods, it is hoped that this alkaline protease will be available at a commercially attractive price. Evolution of this enzyme as a depilating agent for goat skins indicates that its dehairing capacity is satisfactory. The physico-chemical properties of finished leather establishes that the enzymatically dehaired skins have better strength compared to the controls. This paper highlights the process development and scaling-up strategies adopted in the enzyme production and its application.
- Solid-state fermentation (SSF) for the production of an alkaline protease was carried out using the microorganism Rhizopus oryzae. The enzyme is stable from pH 3-11 and at temperature up to 80°C. During optimization of the different environmental parameters for the unhairing of skins, 46% hydration, 83% humidity, pH 8.0 and incubation period of 2 hours were found to produce the best results. Then the physical properties of the skins unhaired by enzymatic treatment and the chemical treatment were compared, the former was found to give better quality leather.
- The influence of enzyme action - during hair-saving unhairing - on the grain layer and grain surface of two kinds of leather (upholstery and a firm upper leather) is described in this paper. Two experiments were performed involving two variations of the unhairing process: hair-saving enzymeassisted unhairing and hair-saving "chemical" unhairing. The first experiment involved upholstery leather and the second involved firm upper leather. The unhairing processes were applied at CITEC's experimental plant, by using five cattle hides for each unhairing. The pelt was processed in the tannery's production line, to the semi-finished leather stage. At this stage, a subjective inspection was performed and an official sampling area was taken from every leather to evaluate their physical-mechanical properties. Then, both types of leather were finished at the tannery. The results obtained led to the following conclusions: • the physical-mechanical properties of the leather were not impaired when compared with the figures corresponding to normal production leather. • for both types of leather (upholstery and firm upper leather), the "buffing effect" on the grain was more noticeable in the "enzyme-unhaired" leathers. • In tanneries producing corrected grain leathers, an enzyme-assisted unhairing process could be used, especially for finished leather. Concerning articles made without a film-covering finish, our observations of an uneven dye absorption in enzyme-unhaired leather should be noted.
- For several centuries enzymes in the form of bate used in deliming have been part of the tanning process, and for the past seventy years research has been conducted on enzymes used for unhairing. These enzymes belong to a general classification known as proteolytic enzymes which catalyze the hydrolysis of a variety of different proteins. More recently, carbohydrases, which catalyze the breakdown of polysaccharides, and lipases, which hydrolyze fats into fatty acids and glycerol, have been proposed for the processing of hides and skins. These uses of enzymes have been the subject of a large amount of published literature from around the world. This review summarizes this research covering the period from 1965 to 1986 and is intended to provide some insight on the potential uses for enzymes in the tannery.
- Hydrogen sulfide (H2S) may produce deleterious effects on the developing central nervous system. The dendritic fields of developing cerebellar Purkinje cells were analyzed to determine the effects of chronic exposure to low concentrations of H2S during perinatal development. Treatment with two concentrations (20 and 50 ppm) of H2S produced severe alterations in the architecture and growth characteristics of the Purkinjec cell dendritic fields. The architectural modifications included longer branches, an increase in the vertex path length and variations in the number of branches in particular areas of the dendritic field. The treated cells also exhibited a nonsymmetrical growth pattern at a time when random terminal branching is normally occurring. These findings suggest that developing neurons exposed to low concentrations of H2S are at risk of severe deficits.
- Structural changes in the wool follicle during depilation with acetic acid, crystalline trypsin, and a commercial enzyme preparation, Pancrozyme C1A, are described. A number of the morphological changes produced by acetic acid and Pancrozyme C1A are similar. Both cause early separation of the epidermal and outer root sheath (ORS) structures from the underlying tissues, and the resistance to fibre withdrawal at this stage is due largely to the physical obstruction of the ORS material. In both cases the separated ORS assumes a tightly packed cylindrical form around the fibre and is gradually digested away until there is no longer any resistance to fibre removal. In both cases separation of the fibre ultimately occurs in the lower part of the prekeratinous zone just above the bulb.
- 1. Hydrogen sulphide (H2S) is a broad spectrum toxicant that occurs widely in nature and is also released by a variety of industrial activities and processes. 2. The central nervous system (CNS) appears to be the major target organ. 3. There is great potential for insult or injury to the developing or immature CNS. 4. The risks of chronic or repeated exposures to low concentrations have not been well defined. 5. Exposure to low concentrations of H2S to time-pregnant rats from day 5 postcoitus until day 21 postnatal results in architectural modification of cerebellar Purkinje cells, alteration of putative amino acid neurotransmitters and changes in monoamine levels in the developing rat brain up to day 21 postnatal. 6. H2S-induced alterations in monoamine tissue levels observed in the developing rat brain return to control values if exposure is discontinued during development, that is, at day 21 postnatal.
- Global environmental regulations are changing the leather-processing industry. Pre-tanning and tanning processes contribute 80-90% of the total pollution in the industry and generate noxious gases, such as hydrogen sulfide, as well as solid wastes, such as lime and chrome sludge. The use of enzyme-based products is currently being explored for many areas of leather making. Furthermore, enzymes are gaining increasing importance in the de-hairing process, eliminating the need for sodium sulfide. This review discusses emerging novel biotechnological methods used in leather processing. One significant achievement is the development of a bioprocess-based de-hairing and fiber-opening methodology to reduce toxic waste.
