Utilization of a leather industry waste

~ Pergamon

Waste Management,Vol. 16, No. 8, pp. 765-769, 1996 © 1997ElsevierScienceLtd All rights reserved.Printed in Great Britain 0956-053X/96 $15.00+ 0.00 PII: S0956-053X(97)00021k2

TECHNICAL NOTE

UTILIZATION OF A LEATHER INDUSTRY WASTE

L. S. S i m e o n o v a and P. G. Dalev* Department of Biology, Universityof Sofia, 8 Dragan Tzankovbvd, 1421 Sofia, Bulgaria ABSTRACT. In the production of leather the main waste that remains after splitting of limed hides before tanning is the lowest layer of the skin together with the underlying fatty tissue (subcutis), It is characterized by a very high water content (up to 870 g kg l) and a balanced content of protein (40-60 g kg-I of the dry mass), fat (10-20 g kg-1 of the dry mass) and carbohydrates. The object of this work was to elaborate a method to process this waste into useful products. The treatment proposed involves washing to remove the inorganic salts, separation of fat and extraction of collagen in hot water solution and additional extraction of protein from the insoluble residue after hydrolysis with alkaline proteinase. This results in the isolation of three fractions: fat--cattle tallow (4-12% of the total mass of the initial material), collagen hydrolysate--glue (5-10%) and protein concentrate for fodder (1-3% yield). Up to 95% of the protein in the initial material was extracted. Further purification of the collagen hydrolysate fraction into edible gelatin was achieved. The proposed method is applicable to every leather factory. © 1997 Elsevier Science Ltd

temperature for the removal of water and fats 3 or thermal treatment at high pressure.1 We are proposing a simple wasteless method for processing the material into fractions of high value at low costs, avoiding the use of reactive and toxic substances and extreme conditions and applying short procedures and simple equipment.

INTRODUCTION Animal skin has a complex morphological structure. It consists of an outer layer--epidermis, m i d d l e - dermis and a lower layer--subcutis. Dermis is the part of the skin from which leather is produced and is composed of thick connective tissue. It is bound to the body by loose fibrillar connective tissue containing bundles of collagen and elastin fibers. This latter tissue represents the underskin (subcutis). During the industrial processing of leather the subcutis is scraped off the skin as a waste product and substantial quantities of it are continuously being thrown away. However, it can be a source for the production of valuable products. A middle capacity leather factory which processes about 100 tonnes of hides daily, produces about 30 tonnes of this waste each day and this creates serious ecological problems. The purpose of this research is to suggest a method for its utilization. Different methods have been proposed for processing leather wastes: t physical (thermal), chemical (extraction in solutions of different reagents) and hydrolytic (treatment with enzymes). Efforts for the utilization of limed wastes are directed to extraction of the collagen after acid treatment with sulfuric acid and thermal hydrolysis at 125°C;2 pressing at high

M A T E R I A L S AND M E T H O D S

Raw Material Leather waste of cattle hides from the regular production of the leather factory 'Lion' in Gabrovo, Bulgaria was used. It is removed by splitting the limed hides before tanning and contains preservatives. Reagents Hydrochloric acid of technical grade and chemical reagents of analytical grade from Fluka were used. The enzyme, alkaline proteinase B72 from Bacillus subtilis of 50,000USg -1 activity, was produced in the enzyme plant in Botevgrad, Bulgaria. Analytical Procedures Standard methods for analysis of foodstuffs were used for assessment of protein, fat salt, sugar and water content 4 as well as of the parameters of the fat fraction. 5 Amino acid analysis was carried out on an automatic amino acid analyzer Microtechna

RECeiVED22 MAY 1996;ACCEPTED26 MARCH1997. *To whom correspondence may be addressed. Fax: 359-2-885-349 765

L. S. SIMEONOVA AND P. G. DALEV

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TABLE 1 Chemical Composition of the Initial Material

T339--Prague, after hydrolysis with 6N HC1 for 24 h at 115°C. Water

Electron Micrographs These were made by a PEM-100 electronic microscope---Ukraine. Ultra-thin cuts were made by ultramicrotome Reichert--Austria.

Proteins

Lipids

g kg -1 of the total mass 750-870

Ash

Polysaccharides

g kg -1 of the dry mass

350-600

100-350

50-70

100-140

Processing of the Leather Waste (Fig. 1)

RESULTS AND DISCUSSION

Characteristics of the Initial Material The chemical composition of the raw material varies over a wide range depending on the composition and pre-treatment of the hides and this influences the yield and composition of the products (Tables 1 and 2). As a result of the preservation of hides the material has a high content of mineral substances (sodium and calcium salts and mainly calcium hydroxide) which must be removed thoroughly. Apart from water, the main constituents of the waste are proteins (35-60% of the dry mass) and fats. Proteins show amino acid composition similar to that of collagens (Table 3). They can be separated and purified into gelatin and as the raw material contains much water, the easiest way is to extract the proteins in a water solution and remove the fats at high temperature.

Ten kilograms of the material were cut into pieces (5-10cm), put in a rotating perforated cylinder and rinsed continuously with 10 litres of water, 12 litres of 0.4N HCI and then again with 10 litres of water. The washed material was heated in a reactor at 90°C with continuous stirring for 1 h. Under these conditions the material was easily liquefied and represented a dark viscose mass with some insoluble particles in it, which however did not hinder its agitation. Because of the high water content of the material, addition of more water for the first extraction was not necessary. After separation in a three stage centrifugal separator three fractions were obtained: light fraction--fat, gelatin solution and insoluble residue. Some of the initial protein was extracted in the water phase, but a considerable part remained in the pellet-insoluble residue. Water was added to it (5 litres each time) and the extraction

TABLE 2 Yield (% of the Total Mass of the Initial Material) and Chemical Composition (g kg -l) of the Fractions Fraction Collagen hydrolysate Enzyme hydrolysate Protein concentrate Fat fraction

Yield

Water

Proteins

Lipids

Ash

Sugars

5-9 1-2 1-3 4-12

30-50 30-60 40-80 20-60

910-930 600-630 350-450 --

20-30 20-30 30-80 930-950

30-50 30-50 50-90 2~,

10-30 150-190 120-250 --

TABLE 3 Amino Acid Composition of the Initial Material and of the Fractions Compared to that of Skin Collagen (grams Amino Acid per 100 g Protein, Cys and Trp are Omitted) Amino acid Asp Thr Ser Glu Pro Gly Ala Val Met lie Leu Tyr Phe His Lys Arg

Initial material

Collagen hydrolysate

Protein concentrate

Skin collagen8

8.67 3.83 5.05 10.76 11.67 17.36 8.82 4.83 1.19 3.19 5.17 2.58 3.83 2.43 4.71 5.99

7.34 2.37 5.52 10.49 13.15 22.88 9.46 3.08 1.31 2.13 3.68 1.42 2.50 2.13 3.92 8.62

7.05 6.29 6.49 10.76 12.38 19.01 8.63 4.26 1.39 2.67 4.74 2.67 3.08 4.80 3.69 8.09

6.66 2.11 4.16 11.04 15.48 26.14 9.75 2.32 1,03 1.50 3.42 0,89 2,24 0,74 3.81 8.71

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UTILIZATION OF A LEATHER INDUSTRY WASTE WASTE

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THREEFOLDEXTRACTIONOFCOLLAGENINHOTWATER

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SEPARATIONINTOFRACTIONS

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FAT FRACTION /'TALLOW/

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INSOLUBLE RESIDUE

COLLAGEN HYDROLYSATE .... I

I PURIRCATION

ENZYMETREATMENT

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CONCENTRATION

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FIGURE 1. Schemeof the process of fractionationof the leather waste.

was repeated twice for 30 min at the same temperature. The three extracts were gathered, concentrated by vacuum evaporation and freeze dried. The resulting collagen hydrolysate meets the requirements for leather glue. An almost complete solubilization of the proteins in the insoluble residue was reached by the addition of water and alkaline proteinase (2.5g per kilogram wet material) to it and agitation at 60-70°C for 1 h. After filtration the insoluble residue was dried at 100°C. The extraction of collagen can be followed on the successive electron micrographs of the raw material and of the insoluble residues after the extractions and the enzyme treatment (Fig. 2). In the first photomicrograph (before extraction) bundles of collagen fibers can be seen clearly in the connective tissue of the subcutis. On the next pictures gradual denaturation and destruction of the fibers is observed and after the final stage the fibers are almost completely destroyed and leave cavities throughout the remaining connective tissue. In Fig. 3 the rate of extraction at each stage of the process can be seen. The first extract removes 59% of the proteins of the raw material, 18.5% more are extracted in the second, 6.2% in the third and after the enzyme treatment only 5% of the raw material

proteins remain in the insoluble residue. This complicated procedure is worth consideration. It can be stopped after the second or after the third extraction when 67.5% or 73.7% of the initial proteins, respectively, are extracted. This will reduce the yield of the collagen fraction but it will raise the yield and the protein content of the residue that is a protein concentrate of good nutritional value and can be used as a fodder product. The process may be optimized by using the third extract as a solvent in the second extraction instead of water. The enzyme hydrolysate can be added to the water extracts to raise the yield of the collagen hydrolysate fraction. If however this fraction is to be purified to produce edible gelatin, it would be better to dry and use the enzyme hydrolysate separately, as the presence of low molecular weight elements in it would reduce the gelling ability of the final product. The residue after the enzyme treatment can be dried without filtration and be used as an additive to forages with higher digestibility.

Characteristics of the Products The fat fraction. Amounts to 4-12% of the total mass of the initial material (Table 2). It has a light

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L . S . S I M E O N O V A A N D P. G. D A L E V

(a)

(c)

(b)

(d)

FIGURE 2. Electron micrographs of the material at different stages of the extraction of collagen. A--raw material, 20,000 xmagnification; B--after the first extraction, 20,000xmagnification, transversally cut collagen fibers are seen; C--after the second extraction, 15,000 x magnification, some longitudinally cut fibers are present; D--after enzyme treatment, 10,000 x magnification.

UTILIZATION OF A LEATHER INDUSTRY WASTE Protein extracted (% of the total 9rotein in the initial material) 6O

\\

5o

\

30

enzyme

20'

~,

10' 0

0

~ i

1

I - -

P

2 3 Number of extractions

I

4

-q 5

FIGURE 3. Extraction of collagen at different stages of the process.

yellow colour and is solid at room temperature (20°C). It is cattle tallow of high quality. Its melting point is 40-45°C and its acid number is 10-20. This fraction can be applied as a fodder additive or as technical fat.

The insoluble residue (protein concentrate). Depending on the number of extractions, can reach a mass of 1-6% of the raw material. It has a comparatively high protein content (Table 2) which determines its nutritional value and the presence of fats and carbohydrates in it raise its energetic value. The amino acid composftion of its proteins shows that they are mainly collagen hydrolysis products (Table 3). The high content of threonine and methionine is notable. In experiments with mice (made in the Institute for Fodder Industry, Bulgarian Agricultural Academy) this product showed 76% digestibility and absence of toxic effects. So the insoluble residue can be recommended as a protein concentrate for fodder. The collagen hydrolysate fraction. Is a yellow powder containing 91-93% protein with a typical amino acid composition mainly of short amino acid residues. Their content rises and approaches that of pure collagen from skin, which indicates the removal of the proteins of non-collagen origin in the insoluble residue. The pH of a 1% solution of this fraction is 6.5-7. It meets the requirements for animal glue (relative viscosity of 7% solution at 40°C not less than 4.5; gluing ability over 110kgcm -2 and resistible to microbial growth for at least 72 h). 6 However it is possible to purify this product to obtain edible gelatin.

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The impurities of unpleasant smell and taste as well as the low molecular weight products of collagen degradation were removed by the addition of active coal and flocculation of calcium phosphate. The combined water extract was heated at 60°C and lgl -l active coal was suspended in it; 100g1-1 calcium hydroxide was added till pH reached 9, then it was adjusted to 4~.2 with 2M orthophosphoric acid and again raised to 5.7-6.2 with calcium hydroxide. The suspension was heated to 90°C for 10min and afterwards filtered. The calcium phosphate formed flocculated and settled easily with the active coal and the impurities thus facilitating their filtration. The purified extract was freeze-dried. The resulting product is a yellowish powder with no smell and taste-edible gelatin. 7 It contains only 1-2% ash, a hot 1% solution is free from any undesirable smell and taste and when cooled, forms a stable gel in the whole volume of the system. Microbiological tests showed absence of coli forms in 0.1 g; of salmonelli in 30g and of Staphylococci in 1 g of the product.

CONCLUSION If the proposed method is developed into a technology, it could solve both economic and environmental problems. From 30 tonnes of the waste at least 1-2 tonnes of edible gelatin as well as the same amount of tallow and protein concentrate as by-products, can be produced. Thus the environment of the region can get rid of a very problematic waste and this can help in the main leather production process by obtaining products that can be of commercial interest. REFERENCES 1. Mdinaradze, T. Processing of Wastes of Animal Origin. Agropromizdat, Moscow (1987). 2. Gaidouk, V., Kuzmich, M. and Bejidashvili, T. A Method for Processing Leather Production Wastes into Fodder. Patent SU 1132893 (1985). 3. Tkac, J., Pecha, F., Manas, M. and Komarek, A. Processing of lime leather wastes, Das Leder 6(21): (1987). 4. AOAC. Official Methods of Analysis (14th Edition.). Association of Official Analytical Chemists, Washington (1984). 5. Bulgarian states standard 1328-72 (1973). 6. Bulgarian states standard 543-72 (1973). 7. Bulgarian states standard 1560-74 (1975). 8. Fitton-Jackson S. The Cell, eds J. Wrachett and A. E. Mirsky. Academic Press, New York, V1 (1964).

Open for discussion until 31 December 1997