Improvement of conventional leather making processes to reduce the environmental impact Eduard Hernàndez Balada Doctoral Thesis directed by Dr. José Costa López Dr. Jaume Cot Cosp Programa de Doctorat d’Enginyeria del Medi Ambient i del Producte Bienni 2006-2008 Barcelona, 5 de febrer 2009 FACULTAT DE QUÍMICA DEPARTAMENT D’ENGINYERIA QUÍMICA
The research included in this presentation was developed in its totality at the United States Department of Agriculture (USDA), Eastern Regional Research Center (ERRC) (Wyndmoor, PA).
Length of stay: From July 2005 thru October 2008
Research Unit: Fats, Oils and Animal Coproducts (FOAC)
Research Leaders: Dr. William N. Marmer and Dr. Daniel K.Y. Solaiman
CRIS (Current Research Information System) projects assigned:
New and efficient processes for making quality leather
Project number : 1935-41440-013-00D
Lead Scientists : Dr. William N. Marmer and Dr. Cheng-Kung Liu
Objectives : Develop new technology for preparing hides for tanning. Establish drying and finishing processes and develop in-line nondestructive tests for improving the quality and durability of leather. Additional funding was obtained to expand the scope of hide preparation research by investigating ways to impart efficiency to short-term hide preservation (brine-curing).
Sustainable technologies for processing of hides, leather, wool and associated byproducts
Project number : 1935-41440-014-00D
Lead Scientist : Dr. Eleanor M. Brown
Objectives : 1. Functional modification, leather and leather byproducts. Develop a foundation for the use of new chemical and biochemical technologies (a) in the production of high quality chrome-free leathers; (b) in expanding the range of high value biomaterial applications for solubilized proteins from leather byproducts. 2. Functional modification, wool: modify wool to impart functionality for improved performance and expanded uses of domestic wool.
Raw hides and skins are byproducts of the meat industry, and in turn are the raw material of the leather industry.
Hide, hair, bones and organs account for an approximate 23% of the animal’s weight.
The tanner employs the word hide to refer to the skin covering large animals, (e.g. cows, steers, horses) and the word skin is mostly used to refer to smaller animals (e.g. sheep, goats, pigs).
Schematic cross section of a bovine hide Composition of hide Collagenous fiber Fibril Microfibril Collagen molecule Polypeptide chain Introduction – Hides and skins
Hide preservation . Treatment given to raw hides or skins just removed from the carcass of the animal to minimize putrefaction. Beamhouse (Pretanning). Processes in the tannery that take place between the removal of the skins or hides from storage and their preparation for tanning. Tanning . Process by which the pelt is converted into a stable material which is resistant to microbial attack and has enhanced resistance to heat. Post tanning operations . Includes wringing, splitting, retanning, coloring, fatliquoring, setting out, drying. Finishing operations . The act of making completely tanned leather more attractive, serviceable and durable. Raw hides and skins Preserved hides and skins Wet blue leather Crust leather Finished leather Pelt Conversion of hides and skins into leather Introduction – Hides and skins
Conversion of hides and skins into leather Preserved Hides and Skins Pelt Wet Blue Leather Crust Leather Finished Leather Finishing Operations Preservation Pretanning Tanning Post tanning Operations Trimming Soaking Unhairing Liming Deliming Bating Scudding Pickling Wringing Splitting Retanning Dying Fatliquoring Setting Drying Conditioning Staking Toggling Buffing Spraying Plating Stages Unit Operations Outcome Introduction – Hides and skins
Economic figures on the American market of hides and skins Source: U.S. Leather Industry Statistics (2006 edition)
The United States is a hide exporting nation.
An 80% of the hides produced in the U.S. in 2008 were exported, half of which were shipped to China and Hong Kong ( Source: United States Hide Skin & Leather Association ).
[Note: 1,000 hides] [Note: 1,000 hides] 17,845 1,355 19,200 32,535 2005 17,388 1,316 18,704 32,880 2004 18,177 1,153 19,330 35,647 2003 19,484 1,299 20,783 35,734 2002 21,750 1,721 23,471 35,530 2001 Net exports Imports Exports Total slaughter Year 41 11 107 India 76 160 141 Brazil 1 15 165 Vietnam 1,343 475 333 Japan 920 826 594 Italy 888 819 651 Thailand 1,647 1,348 1,287 Mexico 1,382 2,534 1,331 Hong Kong 2,751 1,941 1,718 Taiwan 7,602 4,860 4,089 Korea 5,417 5,434 8,191 China 2001 2003 2005 Destination Introduction – Hides and skins
Goal -> To temporarily prevent deterioration of raw hides and skins from the time they are removed from the animal until they are processed into a product that is no longer susceptible to putrefaction or rotting.
Physical damage . Occurs before the slaughter of the animal. Includes tears, scratches, cuts, hook marks, contamination with dirt, insect attack, etc.
Putrefaction . Caused by bacteria and the proteolytic enzymes produced by them. Bacteria are one-celled microorganisms that multiply very rapidly when they feel comfortable in the surrounding environment.
Factors that affect the growth of bacteria:
Introduction – Preservation of raw hides and skins
Sodium chloride (NaCl) is the most popular and inexpensive material used to preserve hides and skins.
It reduces the water content of the hide and lowers the water activity of the remaining moisture.
1. Salt pack curing (Green salting)
The more antique method of salt preservation.
Consists of sprinkling solid salt onto the flesh surface of the hide
Not common in the United States or Europe but extensively applied in India and other Asian countries.
2. Brine curing
Extensively used in American and European hide processing facilities.
Huge vat containing an almost saturated solution of sodium chloride (brine) where hides are suspended for a minimum of 18 h.
Concentration of brine in the raceway is monitored with a salometer, which scale ranges from 0 ºSAL (pure water) to 100 ºSAL (saturated brine).
Preservation of raw hides and skins with common salt Introduction – Preservation of raw hides and skins
Brine curing of raw hides and skins Composition of the hide before the cure Composition of the hide after the cure Introduction – Preservation of raw hides and skins H 2 O H 2 O H 2 O H 2 O Hair side Flesh side NaCl NaCl NaCl NaCl
Source : J.A.Chittenden EPA (1976) Sat. ~ 90% Sat. ~ 60% Sat. ~ 1% Brine curing of raw hides and skins
The degree of curing of a hide can be assessed by means of the salt saturation level.
A standard of 85% salt saturation was established by the U.S. Hide, Skin & Leather Association. That means that the moisture that remains in the hide after the cure has to be 85% saturated with salt.
Introduction – Preservation of raw hides and skins 63.5 53.4 10.4 0.2 48.5 14.0
Capacity to process a high volume of hides and skins (various thousands per day).
No high tech knowledge involved.
Usage of safe chemicals.
Water pollution. Release of about 50% of the total dissolved solids of the whole leather making process during the soaking.
The large amount of salt required.
Increasing commodity prices for sodium chloride (a 10-15% increase over the past few years).
Batch process. Hides might be removed from the vat in a different order that they were put in.
Vat overload. Leads to a slower diffusion rate of salt into the hide.
Red heat damage. Caused by bacteria that grow in a concentrated salt environment (halophiles).
Brine curing of raw hides and skins Introduction – Preservation of raw hides and skins Red heat damage on salted skins
Split the hide in three layers (grain, middle, flesh) Piece of ~ 100 g Cure with 95 °SAL brine (initial concentration) for various time intervals (0.5, 1, 2, 4, 8, 16, 24 h) and 500% float Squeeze out the excess moisture % moisture % ash % salt saturation Wash raw hide @ 6 rpm for 2 hours, 100% float (with 0.15% Boron TS and 0.10% Proxel GXL) Flesh Middle Grain Flesh and adipose tissue Junction of grain and corium and epidermis Majority of the corium Stratrigraphic study – Experimental ( ρ hide = 1 g/cm 3 )
Water is desorbed very rapidly through both flesh and grain surfaces.
The diffusion of salt takes place in its majority from the flesh side.
The flesh split is the first to reach an 85% saturation level, followed by the middle and grain splits.
Stratrigraphic distribution of water Stratrigraphic distribution of ash (salt) Stratrigraphic distribution of salt saturation Stratrigraphic study – Results
CoroNa Green dye is a sodium ion indicator that exhibits an increase in green fluorescence emission intensity upon binding Na + with little shift in wavelength. Fluorescence imaging MW=586 Da Brine concentration: 30% (w/v) NaCl 500% (v/v) float 5 μM CoroNa Green Stereomicroscope equipped for epifluorescence Samples irradiated with blue light (470/40 nm) Magnification: 2.5x 15min 30min 1h 2h 3h 4h 5h 28h 48h Flesh Hair CoroNa Green Na + indicator Fluorescence emission spectra of the CoroNa Green indicator Stratrigraphic study – Results
Back-scattered/Low Vacuum Scanning Electron Microscope (SEM-BSE)
Backscattered electrons consist of high-energy electrons originating in the electron beam, that are reflected or back-scattered out of the specimen interaction volume.
The brightness of the BSE image tends to increase with the atomic number .
High-quality atomic number contrast images are produced.
No conductive coating is required.
Wet samples can be imaged directly as received.
Low Vacuum Mode ON (0.98 Torr)
HV: 25.0 kV
3.0 Spot Size
Stratrigraphic study – Results
Brine curing of raw hides and skins - Variables
Concentration of brine
Length of curing
Temperature of curing
Degree of fleshing of the hide
Usage of additives and/or biocides
Mathematical model – Experimental Lab scale drum Piece of ~ 100 g Cure with brine of various initial concentrations or floats (drum, 6rpm) Wash raw hide @ 6 rpm for 2 hours, 100% float (with 0.15% Boron TS and 0.10% Proxel GXL)
300% (v/v) float at initial brine concentrations of 64, 80, 96 and 100 ºSAL.
Initial brine concentration of 96 ºSAL at float percentages of 300, 500, 750 and 1000 % (v/v)
Collection of residual brine at different times Filtration Determination of chloride concentration (classical Mohr titration)
Continuous reaction model to describe the diffusion of NaCl from the bath containing brine solution to the surface of the solid phase (hide).
Salt will further diffuse into the hide’s inner volume where it will form a non-stationary concentration field.
Diffusion takes place only into the flesh side.
Hide parameters such as thickness, surface and properties of both hair and flesh sides will remain constant throughout the whole process.
Non-stationary one dimensional concentration field (Fick´s second law) Mass balance (closed system) Boundary conditions Dimensionless parameters Dimensionless model Transport coefficient Mathematical model – Results
Effect of initial brine concentration and %float on the diffusion coefficient Note : assuming an initial hide thickness of 5 mm.
Usage of commercial degreasers and microbial biosurfactants Brine curing of raw hides and skins - Variables
Degreasing study – Introduction Grease distribution in raw hide and wet blue Removal of fat Increase of NaCl uptake Reduction of turn around times in the raceway Creation of additional curing capacity
NPE-free. Growing concern regarding the environmental persistence of NP-related compounds as well as their toxicity.
Excellent wetting and grease removal properties.
Commercial degreasers Nonylphenol ethoxylate (NPE) Alkylphenol ethoxylate (APE) Alkylphenol (AP) Nonylphenol (NP) Ethoxylation Primarily used as surfactants The European Union severally restricted marketing and use of NPEs effective January 2005 Alternative : Linear Alcohol Ethoxylates (LAEs) Ethoxylation Hydrophobic Hydrophilic 0 12 20 14 Greater attraction for fatty matter More hydrophilic in nature Hydrophilic-Lipophilic balance (HLB) Degreasing study – Introduction
Microbial biosurfactants produced by the yeast Candida bombicola using renewable (non-petroleum) feedstocks. Typically grown in a medium composed of two different carbon sources (usually sugar and oil) and a nitrogen source (frequently yeast extract).
Disaccharide sophorose, typically with acetylated 6’ and 6’’ hydroxy groups.
A fatty acid (FA), linked to the disaccharide through a glycosidic bond. The FA chain length varies between 16 to 18 carbons, which may be saturated or unsaturated.
The carboxylic acid portion of the fatty acid can be lactonized to the disaccharide ring or remain as a free acid.
They are non-toxic and biodegradable.
They have antimicrobial properties.
They have a great potential for large scale commercialization.
They are currently used in the cosmetic industry, in the formulation of high value products, and as an active ingredient in detergent composition.
Sophorolipids (SLs) Lactonized form of C18:1 SL Free acid form of C18:1 SL Degreasing study – Introduction
Piece of ~ 100g Cure with 100 °SAL brine (initial concentration) for 16 h @ RT and 6 rpm, 500% (v/v) float Squeeze out the excess moisture % moisture % ash % salt saturation % fat content Addition of commercial degreasers OR sophorolipid
Commercial degreaser 1, 2 and 3 at 0.5% (w/w), with respect to combined weight of solution plus hide.
Commercial degreaser 1 at a 0.25, 0.5 and 1% (w/w) concentration level.
Sophorolipid at 0.5% (w/w), filtered or unfiltered.
Wash raw hide @ 6 rpm for 2 hours, 100% float (with 0.15% Boron TS and 0.10% Proxel GXL) Degreasing study – Experimental
Commercial Degreasers - Results Degreasing study – Results
Commercial degreasers and SL: 0.5% (w/w) Sophorolipids - Results 40.4 38.5 36.6 SL-unfiltered 27.2 32.3 36.9 SL-filtered 46.1 47.8 48.7 Degreaser 3 10.9 11.5 9.5 Degreaser 2 42.5 48.3 51.8 Degreaser 1 Moisture and ash free basis (MAFB) Moisture free basis (MFB) As it is Fat Removal (%)/Degreaser Degreasing study – Results
The addition of a 0.5% (w/w) of a commercial degreaser made of a blend of nonionic surfactants to the brine, significantly decreased the fat content of the hide and significantly enhanced the uptake of salt as well.
The composition of the degreaser was a critical parameter for the purpose of defatting the hide. It is likely that different values of HLB amongst the various commercial degreasers affected their activity onto the hides.
The sophorolipid tested showed remarkable degreasing properties and enhanced the uptake of salt by the hide if it was used above the solubility limit. These facts along with its low-foam properties and low cost (from $1 to 3/kg) make it an attractive choice of surfactant.
It would be interesting to remake the mathematical model in which the thickness of the hide would be a variable of the process instead of a parameter.
More research needs to be done in order to establish a minimum salt saturation level that ensures a proper preservation of the hides, currently set at 85%.
More research needs to be done to increase the solubility of sophorolipids. By accomplishing this, their field of applications would be greatly widened and hide dealers would be more receptive towards its usage in an industrial scale.
It is essential to find a way to remove the fat that builds up in the curing vats or raceways, which are operated continuously.
A non-destructive rapid test method for cure validation is needed.
Enzyme capable of forming inter- or intra-molecular crosslinks in many proteins.
Catalyzes an acyl transfer reaction between the γ-carboxamide group of peptite-bound glutamine residues as acyl donors and primary amines as acceptors.
Introduction – Fillers in the leather industry
Enhancement of heat stability, increase of viscosity and gel forming ability.
Improvement of solubility, foaming, emulsifying and surface functional properties.
Increase in solubility, decrease in surface hydrophobicity, improvement of emulsifying and foaming properties, reduction of bitterness.
Modification of heat stability, emulsifying properties and foaming capacity and stability of formed biopolymers.
Obtaining of films with better mechanical properties and more resistance to solubilization.
Whey protein + Soybean 11S globulin
Increase of gel forming ability, modification of breaking strength, strain and cohesiveness of the gels formed.
Increase in breaking strength of gels
Gelatin, caseinate, soy protein isolate, egg yolk Effect of transglutaminase Substrate Acyl donor Acyl acceptor ε-(γ-glutamyl)lysine crosslink
Biopolymer study – Experimental And the next day… Characterization Incubation @ 45 ºC for 5 h Addition of mTGase solutions in selected samples 10 min @ 90 ºC Cool to RT 17 h @ 10 ºC
Whey protein isolate (from 1 to 10% w/w) Type B Gelatin (from 1 to 10% w/w) 10 mg Dithiothreitol (DTT)/g protein Swell Heated @38 ºC 1 h Cool to RT Adjustment of pH to 7.5 Store @ 4 ºC
Biopolymer study – Gel strength results G: Gelatin (from 0 to 3% w/w) W: Whey protein isolate (10% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Enzyme. Reacted with 2 U mTGase/g protein
Increasing gel strength with increasing gelatin concentration.
Significant effect of the reductant DTT at gelatin concentrations > 6% (w/w).
Significantly lower gel strength values in diluted (1 to 5% w/w) gelatin solutions reacted with the enzyme mTGase, with respect to samples of gelatin alone.
G: Gelatin (from 1 to 10% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Enzyme. Reacted with 10 U mTGase/g protein
Very weak gels for gelatin alone or blended with 10% (w/w) WPI, with or without mTGase in a non-reducing environment.
Dramatic increase in gel strength for mTGase-treated WPI-gelatin blends in a reducing environment .
Biopolymer study – Results
Biopolymer study – Viscosity results G: Gelatin (from 1 to 10% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Reacted with 10 U mTGase/g protein Viscosity measured at 60 ºC W: Whey (from 1 to 10% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Reacted with 10 U mTGase/g protein Viscosity measured at 25 ºC G: Gelatin (from 0 to 3% w/w) W: Whey protein isolate (10% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Reacted with 2 U mTGase/g protein Viscosity measured at 25 ºC
No effect of neither mTGase nor DTT alone on gelatin viscosity.
Dramatic increase in viscosity of gelatin solutions of concentrations ≥ 7% (w/w), reacted with mTGase in the presence of DTT.
Significant effect of mTGase on the viscosity of a 10% (w/w) WPI solution under reducing conditions.
Significantly higher viscosity values of WPI-gelatin blends than those of gelatin alone, only in a reducing environment.
Biopolymer study – Results
Biopolymer study – Rheology results W: WPI (10% w/w) G: Gelatin (3% w/w) R: Reducing conditions (10 mg DTT/g protein) E: Reacted with 10 U mTGase/g protein (for WE and WRE); or with 2 U mTGase/g protein (for WGE and WGRE) Time sweep measurements to study the evolution of the storage modulus (G’) as a function of time.
Different behavior between a sample of WPI treated with mTGase in a reducing or non-reducing environment.
Exponential increase of G’ in a WPI-gelatin blend incubated with mTGase in a reducing environment.
The increase of G’ is associated to the formation of a permanent and reticulated network that becomes more stable with time.
Biopolymer study – Results 2.5 mm
Biopolymer study – SDS-PAGE results Inter-protein crosslinking was evaluated by polyacrylamide gel electrophoresis in sodium dodecyl sulfate (SDS-PAGE) Lane 1 : molecular weight markers Lane 2 : gelatin 10% (w/w) Lane 3 : gelatin 10% (w/w) after treatment with mTGase (10 U/g) under reducing conditions Lane 4 : WPI 10% (w/w) Lane 5 : WPI 10% (w/w) after treatment with mTGase (5 U/g) Lane 6 : WPI 10% (w/w) after treatment with mTGase (5 U/g) under reducing conditions Lane 7 : WPI 10% (w/w) with gelatin 1.5% (w/w) Lane 8 : WPI 10% (w/w) with gelatin 1.5% after treatment with mTGase (2 U/g) Lane 9 : WPI 10% (w/w) with gelatin 1.5% after treatment with mTGase (2 U/g) under reducing conditions Biopolymer study – Results 1 2 3 4 5 6 7 8 9 0 Da 200 kDa
The addition of minor amounts of relatively low quality gelatin to whey protein improves the strength and stability of gels formed by the action of mTGase in a reducing environment.
When a small amount of gelatin was added to WPI, before mTGase treatment under reducing conditions, a dramatic rise in viscosity, higher gel strengths, and the appearance of high molecular bands due to inter-protein crosslinking in SDS-PAGE gel patterns than for either gelatin or WPI treated separately were observed.
The reducing environment partially unfolds the whey proteins, increasing access to glutamine and
lysine side chains, favoring the gelatin chains to crosslink the whey proteins to form a network.
The improvement in physical properties over either protein component, given the same treatment, suggests the possibility of greater utilization and new products from these byproducts.
Biopolymer study – Conclusions Biopolymer study – Results
Extraction of aliquots (Protein determination assay) Filler study – Experimental
Grain tightness (break)
All weights calculated on the basis of the weight of wet blue Whey protein isolate Type B Gelatin Dithiothreitol (DTT) Swell in 200% float Heated @38 ºC 1 h Cool to RT Adjustment of pH to 7.5 Store @ 4 ºC Addition of WPI + gelatin (200% float) Retan-Color-Fatliquor Drain 1 h @ RT 5 h @ 45 °C Neutralization with 4% (w/w) NaHCO 3 Addition of mTGase solution (200% float) Wet Blue Drain Wash (x2) Dry Analyses Shoe upper Upholstery
Shoe upper wet blue – Protein uptake results Proteinaceous blend: 5% (w/w) WPI + 0.5% (w/w) gelatin, with respect to weight of wet blue Filler study – Results
Upholstery wet blue – Protein uptake results mTGase offer: 2.5% (w/w) Proteinaceous blend: 2.5% (w/w) WPI + 0.25% (w/w) gelatin, with respect to weight of wet blue Filler study – Results
Protein uptake results – Uptake rate coefficients One may consider that the absorption of protein by the wet blue follows a first order reaction kinetics, where k is the reaction rate coefficient. Filler study – Results Uptake rate coefficient k for various treatments x 0.68 x 0.78 x 0.53 x 3.77 Upholstery wet blue Shoe upper wet blue
Shoe upper wet blue – Mechanical properties Three dimensional regression plots of the mechanical properties of the belly area with respect to % WPI and % mTGase. Filler study – Results
Upholstery wet blue – Mechanical properties Filler study – Results
Shoe upper wet blue – Subjective properties All test samples treated with 5% (w/w) WPI + 0.5% (w/w) gelatin, with respect to weight of wet blue Improvement No effect Worsening Filler study – Results
Shoe upper wet blue – Subjective properties (Grain break) Control sample – Belly area Pretreated with 2.5% mTGase followed by treatment with 5% WPI + 0.5% gelatin (all weights with respect to weight of wet blue). Test sample – Belly area Filler study – Results
Shoe upper wet blue – Subjective properties (Color) Test : Pretreated with 2.5% mTGase followed by treatment with 5% WPI + 0.5% gelatin (all weights with respect to weight of wet blue). Filler study – Results
Shoe upper wet blue – SEM images Magnification: x1000 Test : Pretreated with 2.5% mTGase followed by treatment with 5% WPI + 0.5% gelatin (all weights with respect to weight of wet blue). Filler study – Results Belly Butt Neck Control Test 50.0 μ m
Upholstery wet blue – Subjective properties All test samples treated with 2.5% (w/w) WPI + 0.25% (w/w) gelatin, with respect to weight of wet blue Filler study – Results 2.5% mTGase 2.5% mTGase 0% mTGase 0% mTGase 0% mTGase 2.5% mTGase 2.5% mTGase 2.5% mTGase
It would be interesting to look into the possibility of using keratin, the main protein in the cattle hair and wool, as a cheap and readily available source of protein.
The reaction between the proteinaceous substrate and a carbodiimide (e.g. 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride, EDC) prior to the enzymatic treatment with mTGase would be an interesting approach to increase the reactivity of the proteins towards the enzyme.
Scaling up of the filling process. By doing so, a lesser concentrations of reactants and volume of float would be needed due to a stronger mechanical action.
It would be interesting to make the filler from the tannery's solid waste (e.g. chrome shavings). By doing this, they would not only adding value to a byproduct that it is usually landfilled, but they would also be increasing the value of low quality hides that will be treated with the product.
Future work – Renewable biopolymers as filling agents for leather