Influence of crosslinking agents on the pore structure of skin

Colloids and Surfaces B: Biointerfaces 57 (2007) 118–123

Influence of crosslinking agents on the pore structure of skin N. Nishad Fathima, Aruna Dhathathreyan ∗ , T. Ramasami ∗ Chemical Laboratory, Central Leather Research Institute, Adyar, Chennai 600020, India Received 10 January 2007; received in revised form 22 January 2007; accepted 23 January 2007 Available online 30 January 2007

Abstract Analysis of pore structure of skin is important to understand process of diffusion and adsorption involved during any application of the skin matrix. In this study, the effect of thermal shrinkage on the pore structure of chromium and vegetable treated skin has been analyzed as these tanning agents are known to bring about thermal stability to the matrix. The changes brought about in the pore structure have been studied using mercury intrusion porosimetry and scanning electron microscopy. Response of the chromium treated and vegetable tanning treated skin structure to heat has been found to be quite different from each other. About 41% decrease in porosity is observed for chromium treated skin as against 97% decrease for the skin treated with vegetable tannins. This is primarily attributed to the basic nature of these materials and the nature of interaction of them towards skin. © 2007 Elsevier B.V. All rights reserved. Keywords: Skin; Pore; Mercury intrusion porosity; SEM; Crosslinking agents

1. Introduction An understanding of the internal pore structure of skin as a matrix has far reaching implications in both biological and industrial applications of the material. Skin, is an architectural marvel. Many of the unique properties of leather like visco-elasticity and breathability seem to stem from the pore ˚ structure and connectivity with their sizes ranging from 7 A to 150 ␮m of the skin [1]. Pore structure renders skin many unique properties. Insight into the pore structure of the skin matrix is required to understand mass and heat transport properties as well as fracture mechanism of material under flexural stress [2,3]. Several studies have been carried out on the water vapour adsorption and transmission of leather [4–6]. Pore structures of skin and leather have been investigated previously [7,8]. Characterization of porous space inside matrices can be approached using different methods including gas adsorption, vapour sorption, thermoporometry, mercury porosimetry and image analysis. Typically, the distribution of pore sizes is char-

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acterized by mercury intrusion porosimetry (MIP) and nitrogen adsorption [9]. The basic assumption in all these techniques is that the geometry of the pores is regular, that the pores are interconnected and that the size distribution is not affected by the loss of water in the pores upon drying. MIP is one of the few analytical techniques that uses data over a wide dynamic range (0.003–10 ␮m) to investigate the porous structure of solid samples in a quantitative manner. This is widely used technique for the pore structure characteristics of many materials [10,11]. One of the characteristic features of skin is its dimensional change under heat. Skin undergoes length reduction at a characteristic temperature to a level of one third of its original dimension. Such shrinkage occurs at a characteristic temperature depending on the type and nature of skin [12]. Shrinkage temperature is considered an important parameter in leather making. Tanning, the process that converts raw hide/skin into leather, aims at bringing about thermal and enzymatic stability to the skin matrix [13,14]. The shrinkage temperature of skin is known to increase on tanning [15]. In our previous study on the shrinkage phenomenon of skin, the changes brought about in the porosity of the skin matrix as a result of thermal shrinkage has been reported [16]. The volume changes accompanying the removal of water from skin, expressed as partial fractions of

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Table 1 The intrusion data summary for native, chrome treated, vegetable treated and respective shrunken skin samples from the mercury porosimetry technique Property

Native

Native shrunk

Chrome

Chrome shrunk

Vegetable

Vegetable shrunk

Total intrusion volume (mL/g) Total surface area (m2 /g) Median pore diameter (volume) (␮m) Bulk density at 0.10 psia (g/mL) Total porosity (%)

0.2431 0.0986 14.60 1.3646 33.17

0.0446 1.7533 0.409 2.0925 9.33

0.4786 0.2430 10.65 0.7580 16.34

0.3280 0.6238 4.729 1.1514 9.5

0.5918 1.0602 5.152 0.9919 31.35

0.0145 0.0152 6.256 1.3486 0.85

volumes of solid, liquid and air has been studied using dilatometric technique [17]. A fundamental understanding of changes in shrinkage temperature on tanning in relation to pore size distribution is yet to be achieved. In the present work, an attempt

has been made to unravel relationships, if any, between porosity of various tanned matrices in relation to increases in their shrinkage temperature using mercury intrusion porosimetry and scanning electron microscopy.

Fig. 1. Mercury intrusion volume to pore size curves for (a) native skin, (b) shrunk skin, (c) chrome treated, (d) chrome treated shrunk, (e) vegetable treated, and (f) vegetable treated shrunk.

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2. Materials and methods 2.1. Materials Wet salted cow hides were taken and processed to remove hair and flesh by conventional procedure [18]. Small pieces, of dimension 2 cm × 2 cm × 0.3 cm, were cut from the butt portion of the hides. Chromium and vegetable treatment to the hides were carried out as per standard procedures [18]. The untreated, chrome treated and vegetable tannin treated material was shrunk by heating in the presence of water. Pore size data in a dry sample do not truly represent the size distribution in a wet sample due to the matrix shrinkage associated

with the removal of bound water [8]. Hence, samples were prepared for the analysis by gradual dehydration with acetone and methanol solutions such that the pores were not altered [19]. 2.2. Mercury intrusion porosimetry measurement (MIP) The pore size distribution measurements of the samples before and after shrinkage were performed using mercury porosimetry (Quantachrome poremaster). The contact angle used was 140◦ . The samples were evacuated for 5 min at an evacuation pressure of about 50 mm Hg for low-pressure run. The mercury filling pressure was about 0.42 psia.

Fig. 2. Scanning electron micrographs (1000× magnification) showing the grain view of (a) native skin, (b) shrunk skin, (c) chrome treated, (d) chrome treated shrunk, (e) vegetable treated, and (f) vegetable treated shrunk.

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2.3. Scanning electron microscopy (SEM) Samples were cut into specimens of uniform thickness. A Polaron SC500 ion sputtering device was used for sputter˚ onto the samples. A Leica ing gold film of thickness 250 A Stereoscan 440 scanning electron microscope was used for the analysis. 3. Results and discussions An insight into the shrinkage phenomenon of collagen matrix is of interest owing to the varied applications of the matrix. Distribution of pores and effect of temperature on the pores and pore distribution is also of direct interest. In this study, effects

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of increases in temperature (20–120 ◦ C) on the pore connectivity of the collagen matrix has been investigated using mercury intrusion porosimetry and scanning electron microscopy and the Ritter and Drake method of obtaining pore volume distributions by the use of a high-pressure mercury porosimeter has been used [20]. When a liquid meniscus is at equilibrium in a cylindrical capillary tube, of radius r, and the pressure difference across the meniscus is P, then they are related by the equation developed by Washburn as [21]: P =−

2γ cos θ r

(1)

where θ is the angle of contact between the solid and the mercury and γ is the surface tension of mercury. The pore size distribu-

Fig. 3. Scanning electron micrographs (2500× magnification) showing the cross-section view of (a) native skin, (b) shrunk skin, (c) chrome treated, (d) chrome treated shrunk, (e) vegetable treated, and (f) vegetable treated shrunk.

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tion is determined from the volume intruded at each pressure increment. Total porosity is determined from the total volume intruded. The numerical value of the ratio of pore volume, Vp , to internal surface, SI (Vp :SI ), gives a useful general guide to the pore size of the solid; from simple geometrical considerations, if the pores were all cylindrical tubes of same radius r, then there is a fundamental assumption in this technique that the pores are circular, which is an approximation of the true shape. An MIP test demands complete water removal from the specimen. Changes in pore structure could occur on drying samples. However, a relative assessment of results obtained by applying the same technique seems appropriate. The total intrusion volume, total surface area, median pore diameter (volume), bulk density, percent porosity for native, chrome tanned, vegetable tanned and their respective shrunken samples are given in Table 1. In general, it is apparent from these values that the percentage porosity decreases on shrinkage. The % porosity reduction for native, chrome treated and vegetable tannin treated on shrinkage is 71.8, 41 and 97.2%, respectively. This is in accordance with the results obtained previously from scanning electron microscopy analysis, where the macropores were shown to close on shrinkage [16]. The difference in the % porosity reduction between native, chrome and vegetable treated is due to the crosslinking effect brought about by these agents. It can be seen that % reduction for chromium treated sample is less compared to vegetable treated owing to the reason that chromium forms coordinate covalent crosslinking and vegetable forms hydrogen bonds and also it coats the fibers. The median pore diameter (volume) of the sample register decreases on shrinkage. The total intrusion volume for unshrunk samples is more than the respective shrunk samples. This can be correlated with the reduction in pores on shrinkage. It can be noted that the difference in intrusion volume between vegetable and vegetable shrunk collagen matrix is more. This is possibly due to the fact that vegetable tanning leads to filling up of voids spaces [22]. For chrome tanned collagen matrix, there is less reduction in % porosity in comparison with native and vegetable treated. This is again due to the nature of interaction of chromium with skin matrix. It is known that chromium treatment does not lead to filling up of void spaces as in the case of vegetable treated [23]. Hence, when this skin is subject to heat, not much appreciable changes in porosity of skin matrix are observed. The intrusion–extrusion profiles of native and shrunken samples of skin are presented in Fig. 1. It is seen from Fig. 1, that the cumulative intrusion is larger for native compared to the shrunken samples. This not only shows that the pores present in native sample undergo changes on heating but also proves that pores are getting closed on increasing the temperature. It can also be seen that pores in the diameter range of 10–30 ␮m are affected more on shrinkage. Fig. 2 shows the grain surface of native, chromium treated and vegetable tannin treated and their respective shrunk samples at a magnification of 1000×. The grain surface of hide consists of hair pores, which are characteristic of the nature of the animal. It is seen from the micrographs that the mouths of the microsize pores are closed on shrinkage for all samples, i.e.

for both native and crosslinked hides. However, the amount of pores closed varies with the type of crosslinking. The same is observed in the cross-section view of the unshrunk and shrunk samples (2500×) as shown in Fig. 3. It is seen that the fibers coalesce completely on shrinkage. The volume changes accompanying removal of water from skin matrix can be expressed as volume fractions in terms of solid, liquid and air as shown in Eq. (2). It is expected that heat changes may influence the volume fractions of solids, liquid and air. Such changes in network may also be associated with alterations in protein–water interactions and hydration phenomenon. These results indicate that thermal shrinkage induces alterations in nano, meso- and microporic networks. Also, the difference in the way crosslinking affects shrinkage can be seen clearly from Fig. 3. Vegetable treated matrix on shrinkage (Fig. 3f) undergoes more changes than native and chromium treated, which was also observed from the porosity results: VTotal = Vsolid + Vliquid + Vair

(2)

4. Conclusion Drastic and irreversible changes in the pore structure of native and treated skins are observed on adsorption of heat. The response of chrome treated and vegetable tannin treated skin to heat varies due to the very nature of interaction of these two species towards skin. The results obtained from MIP and scanning electron microscopic studies reveal that distinct changes in pores structure like reduction in pore size diameter and coalescence of fiber occurs on shrinkage. These results should help in understanding the basic nature of interaction of tanning materials used in processing of leather. Work is presently underway to understand the effect of bound water in collagen on shrinkage and stability of skins. References [1] T. Ramasami, Approach towards a unified theory for tanning: Wilson’s dream, J. Am. Leather Chem. Assoc. 96 (2001) 290–304. [2] R.R. Stromberg, M. Swerdlow, Pores in collagen and leather, J. Am. Leather Chem. Assoc. 47 (1954) 336–351. [3] T.J. Carter, J.R. Kanagy, A flex tension test for leather, J. Am. Leather Chem. Assoc. 49 (1954) 23–42. [4] J.R. Kanagy, Influence of temperature on the adsorption of water vapor by collagen and leather, J. Am. Leather Chem. Assoc. 45 (1950) 12– 41. [5] J.R. Kanagy, Absorption of water vapor by untanned hide and various leathers at 100 degrees F, J. Am. Leather Chem. Assoc. 42 (1947) 98–117. [6] J.R. Kanagy, R.A. Vickers III, Factors affecting the water vapor permeability of leather, J. Am. Leather Chem. Assoc. 45 (1950) 211–247. [7] J.R. Kanagy, Macro pores in leather as determined with a mercury porosimeter, J. Am. Leather Chem. Assoc. 58 (1963) 524–550. [8] J.R. Kanagy, Sorption of water by collagen, in: H.R. Elden (Ed.), Biophysical Properties of the Skin, Wiley, New York, 1971, pp. 373–391. [9] L.G. Joyner, E.P. Barrett, R. Skold, The determination of pore volume and area distributions in porous substances. II. Comparison between nitrogen isotherm and mercury porosimeter methods, J. Am. Chem. Soc. 73 (1951) 3155–3158. [10] C. Gall´e, J. Sercombe, Permeability and pore structure evolution of silicocalcareous and hematite high-strength concretes submitted to high temperatures, Mater. Struct. 34 (2001) 619–628.

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