Degradation of keratin containing wastes generated in the Hair -Saving Unhairing process by Paecilomyces lilacinus.

CAVELLO, IVANA; GORTARI, CECILIA; GALARZA, BETINA; CANTERA, CARLOS; HOURS, ROQUE ALBERTO; CAVALITTO, SEBASTIÁN FERNANDO

Curitiba, Brasil

Congreso; 4th International Congress on Bioprocesses in Food Industries; 2010

Federal University of Parana

 The reduction of the organic load in beamhouse effluents, expressed in terms of Chemical Oxygen Demand (between 40 and 60%), of the sulphide content (approx.50%) and the settleable suspended solid (almost 70%) is an essential feature of modern hair–saving unhairing, as far as effluents are concerned, but a “new” solid waste is generated which has to be properly disposed of: “hair waste”. It is estimated that a tannery processing 25 tons of salted cow hides recover about 2.5 tons of wet hair per day. An analysis of the “hair waste” after a hair-saving unhairing process using sodium sulphide / lime assisted by mercaptans / amines unhairing agents revealed that about 95% of the organic component is keratin protein. The remaining 5 % are non keratin proteins, these may be easily extracted with buffer solutions in an aqueous medium. Keratin is the insoluble structural protein of feathers, wool, hair and nails, it is known for its high stability and its resistance to proteolityc degradation. Degradation of keratinous material has been associated with dermatophytic fungi, yeast and bacteria. However, there are relatively few reports on the production and characterization of the keratinases produced by non-dermatophytic fungi. The aim of this study was to analyse the ability of Paecilomyces lilacinus (Thom) Samson LPSC # 876 to produce keratinolityc enzymes when it was grown on basal medium containing 10g/L of “hair waste” as a sole source of energy, carbon and nitrogen. The capability of the enzyme preparation to hydrolyse different keratinous and non-keratinous substrates and a comparison of the enzyme prepation with knwon proteinases were tested. The effect of enzyme inhibitors on the enzyme preparation was also studied. P. lilacinus were cultivated for up 5 days in whole hair medium and the resulting supernatant was used as enzyme preparation with a protein concentration of 167 mg/ml using Bradford´s method. Proteolytic activity was determined using azokeratin, hair waste and keratin azure as keratinous substrates and azocasein as non keratinous one. Among the keratinous susbtrates, the most hydrolysed one was azokeratin followed by hair waste. The activity of the enzyme preparation on azokeratin and azocasein and keratin/casein ratio were compared with commercially available proteases such Protease (Sigma), Trypsin (Sigma) and Proteinase K (Promega). All tested enzymes were compared on the basis of their specific activity. It was found that the enzyme preparation was superior in the hydrolysis of both substrates and it had the highest keratin/casein ratio. The effect of enzyme inhibitors was tested by adding them at the following working concentrations: 2 mM PMSF, 5 mM EDTA, 1 mM 1,10-Phenanthroline, 100 μg/ml Pepstatine, and 10 mM Iodoacetate. The enzyme preparation was inhibited only by PMSF suggesting that P. lilacinus produces serin-proteases. The enzymatic preparation obtained of P. lilacinus demonstrated to be broadly specific; it degrades keratinous proteins. These results suggest that keratinases produced by P.lilacinus have great potential application for biotechnological processes involving keratin hydrolysis, such as in the bioconversion of poultry waste and leather manufacturing. Key words: hair waste, tannery, keratinases, Serin proteases.