lable at ScienceDirect

Polymer Degradation and Stability 111 (2015) 151e158

Contents lists avai

Polymer Degradation and Stability

journal homepage: www.elsevier .com/locate /polydegstab

The effect of electron beam on sheep wool

M�aria Porubsk�a a, *, Zuzana Hanzlíkov�a a, Jana Brani�sa a, Angela Kleinov�a b, Peter Hybler c,Marko Fül€op c, J�an Ondru�ska d, Klaudia Jomov�a a

a Constantine the Philosopher University in Nitra, Faculty of Natural Sciences, Department of Chemistry, Tr. A. Hlinku 1, 949 74 Nitra, Slovakiab Polymer Institute, Slovak Academy of Sciences, Dúbravsk�a cesta 9, 842 36 Bratislava, Slovakiac University Centre of Electron Accelerators in Tren�cín, Slovak Medical University, Limbov�a 12, 833 03 Bratislava, Slovakiad Constantine the Philosopher University in Nitra, Faculty of Natural Sciences, Department of Physics, Tr. A. Hlinku 1, 949 74 Nitra, Slovakia

a r t i c l e i n f o

Article history:Received 24 August 2014Accepted 14 November 2014Available online 22 November 2014

Keywords:Sheep woolElectron beamChemical changeSecondary structureMaterial testing

* Corresponding author. Tel.: þ421 37 6408 655; faE-mail address: mporubska@ukf.sk (M. Porubsk�a).

http://dx.doi.org/10.1016/j.polymdegradstab.2014.11.00141-3910/© 2014 Elsevier Ltd. All rights reserved.

a b s t r a c t

The effect of accelerated electron beam with absorbed doses of 0e400 kGy on wool fibres was investi-gated. The S-oxidized species, CH groups, secondary structure, temperature and enthalpy of crystalcleavage, strength and elongation of the fibres were monitored. All the properties showed fluctuationwith the absorbed dose and are related with change of the secondary structure. Increasing absorbed doseled to progressive predominance of b-sheet over a-helix conformation. The helical conformation fav-oured generation of S-sulphonate while the b-sheet suppressed it. Changes in the abundance of eCHegroups indicated a slight networking. High absorbed doses resulted in the polypeptide chain breakingand the formation of shorter fragments with helical conformation. The fibre strength was not changedsignificantly and the elongation, after an initial increase, monotonously decreased due to furtherdenaturation and chain breaking.

© 2014 Elsevier Ltd. All rights reserved.

1. Introduction

Nowadays traditional textile exploitation of sheep wool hasbeen considerably replaced with the use of using synthetic fibres.Wool is fully renewable material and deserves attention regardingsustainable development. Therefore present research should beaimed also in studying further and more detailed applications ofthis remarkable raw material.

The main component of wool fibre is hard keratin, having con-tent of the cysteine residual group. The principal structural units inthe native wool are successive turns of the alpha helix. The intrinsicstability of the alpha helix, and thus the fibre, results from intra-molecular hydrogen bonds. Both low-sulphur protein and high-sulphur protein exist in wool fibre, which play a key role in deter-mining super-molecular structure of wool fibre. The low sulphurcontent proteins have a a-helical crystalline structure, while theother proteins do not have a helical structure [1].

It is believed that in the low-sulphur protein of raw wool fibres,about 50% molecules exhibit alpha-helix structure in microfibrils,and the other 50% of molecules exhibit the form of irregular coils.

x: þ421 37 6408 020.

09

Mostmolecules exist inmatrix as irregular coils in the high-sulphurprotein, and disulphide bonds link low-sulphur protein moleculesto high-sulphur protein molecules. Therefore protofibrils in woolfibre are coupled with matrix via SeS bonds.

Wool has been an object of numerous studies due to its excep-tional properties. Many research studies have been focused onimproving shrink resistance and dyeability of wool fabric [2] in thetextile industry. Potential applications of wool have been examinedin composite materials [3,4], in medicine [5] and other fields. In-vestigations have also focused on the obtaining of keratin [6] aswell as keratin powdering [7]. A specific field dealing with super-ficial structure variations of wool due to physical-chemical modi-fication involves plasma application [8], corona charges [9],microwave [10] and UV radiation [11], etc. These mentioned radi-ation treatments modify only the wool surface.

Within the last decades development of radiation technologiesoffers further opportunities to study the wool property modifica-tions. The main feature and advantage of radiation initiation iscapability to generate active intermediate products in the solidphase. An electron beam can penetrate the whole volume of thefibres and affect all parts of the fibre substructures. Such treatmentdoes not require the use of any chemicals. It is environmentalfriendly dry process, which does not involve any solvent and re-agent of wet chemical processes.

Table 1Characteristic wave numbers for non-irradiated wool.

Form Structure Wave number (cm�1)

Cystine dioxide eSO2eSe 1128Cystine monoxide eSOeSe 1075Cysteic acid �SO�

3 1044S-sulphonate (Bunte salt) �S� SO�

3 1022Amide III as internal reference band eNeH mixed 1232

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158152

Although the impact of electron beam on synthetic polymershas been described in several papers, e.g. Ref. [12], according to ourbest knowledge, only one brief report on the electron beam effecton surface wool properties has been published [13]. The elucidationof such modification of whole wool fibre volume could show newalternatives for processing and exploitation of wool since it is a veryoften undesirable waste today. However, it is necessary to under-stand some fundamental variations induced by electron beam inwool.

In view of these remarks, the aim of our study is the irradiationof wool by different absorbed doses of electron beam and, in par-allel, observation of the wool property variations using FTIR spec-troscopy, microcalorimetry and tensile properties.

2. Materials and methods

2.1. Materials

In the work industrially scoured wool with fibre thickness of22e27 mmwas used in the form of wool combed sliver, supplied bythe company Pradiare�n vlny JK, Nov�e Mesto nad V�ahom, Slovakia.

2.2. Sample preparation

The wool was extracted with dichloromethane in Soxhlet using9 flow-off, then rinsed 2� in distilled water, first dried freely andfinally dried in a laboratory oven at 60 �C for 4 h. Such wool beingstill warm was put into a zip PE pockets with dimension of18 � 25 cm, closed and saved in a dessicator till irradiation.

2.3. Sample irradiation

For each irradiation dose, the samples of about 12 g in mass putin PE pockets were placed into carton boxes grouped according toselected absorbed doses. The exposure in air was conducted at theUniversity Centre of Electron Accelerators in Tren�cín in linearelectron accelerator UELR-5-1S (manufacturer FGUP “NIIEFA”,Petersburg, Russia) with 5 MeV of installed energy, an electronbeam (hereafter, EB) intensity of 200 mA, mean power of 1 kW andmean dose rate of 750 kGy/h. The doses applied were 0-16-25-40-63-100-156-250-400 kGy repeating 100 kGy cycles plus neededsupplementing dose, if necessary. Between individual irradiationcycles, the samples were allowed to cool down for 30 min tomaintain the temperature below 50 �C.

2.4. FTIR spectral analysis

Spectral analysis was carried out 9 days after the irradiation. Forthe measurements, the fine-cut wool samples were blended withKBr in a small grinder and pressed into discs in a pellet press. Themoulded discs were kept in the dessicator till spectral measure-ment. Infrared spectra were collected using AVATAR 330 ThermoNicolet FTIR Spectrometer (manufacturer Thermo Nicolet Corpo-ration, Madison, USA) in the range of 400e4000 cm�1 at 4 cm�1

resolution and 3 scan repetitions per analysis.

2.5. Thermal analysis

Differential scanning calorimetry (DSC) was performed on1e2 mm wool cut-offs using Mettler Toledo DSC822e device in airatmosphere. For analysis thewool sample, in quantities about 5mg,was moulded into an aluminium crucible 40 ml in volume. Theheating rate was 20 �C/minwithin the interval from 25 �C to 300 �Cwith 3 repetitions per analysis.

2.6. Tensile properties

Before testing, the samples were conditioned at 21 �C and 63%relative humidity for 48 h. The strength at break and elongationwere measured on twenty specimens following ISO 5079: 1999using the tearing machine Testometric (manufacturer TestometricCompany Ltd., Lancashire, United Kingdom) at 21 �C and 63% hu-midity, clamp speed movement 10 mm/min and working length10 mm. The thickness was scanned by the digital thickness meterSylvac (manufacturer Sylvac SA, Crissier, Switzerland).

3. Results and discussion

Absorption of electron particle energy in the material iscomplicated by X-ray productions, liberations of high energy sec-ondary electrons, photo- and Crompton processes. Therefore inradiation chemistry mechanism of chemical reactions is no explicitEB energy dependence. However in this work keeping all remainingprocess parameters to be constant, the only variable was absorbeddose. Therefore observed variations have been relative to theabsorbed dose.

3.1. FTIR analysis of sulphur-oxidized products

The disulphide bond SeS is themost reactive part of keratin and,after being initiated in air, gives to arise several sulphur-oxidizedspecies such as S-sulphonate (S-sulph), cysteic acid (CA), cystinemonoxide (CMO) and cystine dioxide (CDO). Characteristic IRabsorbance wave numbers used to estimate the sulphur-oxidativespecies are given in Table 1 and they are closed to data from pa-pers [4,14].

Collected FTIR spectra were inverted in the second-order de-rivative spectra, the related signals were read and divided by thesignal for reference Amide III. These absorbance ratios were relativeto the corresponding ratios of the non-irradiated sample. EBabsorbed doses resulted in some shift almost of all bands towardshigher frequencies and the ultimate shift up to 12 cm�1 wasobserved for 25-40-63-100 and 400 kGy doses. However, based onthe second-order derivative treatment, there were not any doubtson the assignment of related functional groups.

The results from FTIR analysis for the wool irradiated by gradedabsorbed doses represent a relative variation of individual func-tional groups when compared with the parent wool and are shownin Fig. 1. It can be seen that composition of the oxidized sulphurproducts was changing the each species considerably. Followingthe amount of the generated species (S-sulph, CA, CMO and CDO)the region of the absorbed doses can be divided roughly into twoparts, 0e220 kGy and 220e400 kGy.

In the range of 0e220 kGy, we observed a fluctuation of alloxidized products of which S-sulph and CMOwere shown to be thelargest. At the lowest absorbed dose of 16 kGy, the amounts of CAand CDO decreased below the initial level, indicating trans-formation into other products and temporary increase of CMO. Atthe next absorbed doses 25e40 kGy the formation of CMO togetherwith CA and CDO were reduced to a minimal level at 40 kGy while

Fig. 1. Dependence of the relative variations of oxidized sulphur species on absorbeddose in the irradiated wool.

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158 153

S-sulph reached a maximum three times higher than that of theparent wool. However, at the next absorbed dose of 63 kGy S-sulphdecreased to about 1.6 times of the initial value. At the same time,the content of the others monitored products increased, obviouslyat the expense of S-sulph. But with the exception of CMO, theiramounts did not exceed the initial level. The absorbed dose of100 kGy brought the next maximum for S-sulph and simultaneousreduction of all monitored constituents. From this absorbed dosethe amount of S-sulphwas decreasingmonotonously up to 400 kGywhere it showed only 60% of the original value. The CMO contentexhibited values below the starting figure from absorbed dose of100 kGy. After a transitive reduction around 156 kGy absorbed dosenearly to zero, CDO started to increase sharply and, at the highestabsorbed dose of 400 kGy CDO concentration reached almost 8times of the parent wool. From 100 kGy absorbed dose a linearincreasing of CA amount started and reached 2.5 times of thestarting level at 400 kGy dose. Thus approximately from 220 kGyabsorbed dose range, CDO and CA were dominant S-oxidized spe-cies in the wool. Such variable development of S-oxidized groups israther surprising in the case of S-sulph particularly. The main re-action pathway of radiation damaged disulphide bonds involvesone electron addition that yields the radical anion (SS��) accordingto the reaction:

(1)

The unpaired electron in SS�� resides in a three e electron s e

bond. Generally, radical anion SS�� is highly reactive and in theoxygenated environment may react with dioxygen leading to the S-sulph (Bunte salt) formation according to the reaction (2a):

(2)

A cleavage product of a disulphide bond according to the reac-tion (2b) is CA. Douthwaite et al. [15] have suggested CMO and CDOmay be intermediates of a gradual oxidation of cystine to S-sulph.

We presume that, besides other factors, differences in oxidationat particular structural levels linked with distinct oxygen diffusioncan contribute to the observed fluctuations of the S-oxidizedproducts. The upper layer rich in sulphur e cortex oxidizes mosteasily, mainly because of easy oxygen access at the surface layer. It

was found [11] that molecular oxygen is necessary for the forma-tion of oxidized sulphur species during UVC exposure of dry wool.Thus when discussing the properties of wool fibres, radiation pro-cess and presence of oxygen appear to be of great importance. Theinner fibre layers are poorer in sulphur and limited source of oxy-gen is inside the fibre. Then the more distant sub-structuralcomponent from the fibre surface (and so from the source of oxy-gen) it is the more difficult to start oxidation. It is also found [16]that plasma treatment of wool provoked any chemical changesonly at the surface layer rich in sulphur and using plasma surfacetreatment, the generation of S-suph was among S-oxidized speciesthe most dynamic and achieved the highest value when comparedwith the initial concentration. Similarly, stability studies of keratincysteine-S-sulphonate [17] revealed that S-sulph formation occurspreferentially on or near the surface of the fibre. The S-sulph as wellas CA represent surface polar cystine residues which affect thewettability of wool fabric [16]. Besides S-sulph formation, CA wasformed as a result of the cleavage of disulphide linkage [14].

In our case the amounts of CA, CDO and partially also CM, rangenear or under the level of the non-irradiated wool up to theabsorbed dose between 200 and 250 kGy (Fig.1). Kan and Yuen [14]noted a similar fact when under plasma treatment of wool fabricsunder different atmospheres he observed that the amount of CDOwas less in treated than the untreated wool fabric after 8 mintreatment time. This decreased amount of CDO was assigned to thespontaneous conversion of CDO to CA, too.

Examining photodegradation of wool it was concluded [11] thatthe effect of UVC radiation at 254 nm on the SeS bond of the aminoacid cysteine in wool keratin is the production of CA and thepartially oxidized derivates CMO and CDO residues, but no cysteineS-sulph residues. In contrast, UVA radiation at 360 nm produced CAand S-sulph residues, but no partially oxidized cysteine derivate.Our findings (Fig. 1) are comparable with those results when theformation mainly of S-sulph and partially CMO responded to thelower absorbed doses (up to 200 kGy) and the content of CDO andCA increased at the higher absorbed doses (above 200 kGy).However, it is necessary to admit that in the wool, CDO and CA canbe stabilized also by interaction with other species e.g. oxidizedproducts of hydrocarbon components in the wool which were notstudied in this work.

Also many other studies showed that depending on suppliedenergy, all intermediate products (S-sulph, CMO and CDO) arefinally converted into CA. Examining reactions of woolwith peroxy-compounds, alsowork [15] attained that CAwas themain oxidationproduct within the wool fibre. However, the presence of CMO andCDO suggests that CA is probably formed from these intermediatesas proposed in a previous model. From the mentioned finding re-sults that cystine oxides are not stable and may easily be convertedto CA [14], therefore it is hard to quantify accurately the formationof each product in relation to CA at different absorbed doses. Bothcystine residues were believed to be intermediate cystine oxidationproducts. Based on results of the above mentioned studies it can beexpected that the high amount of CDO in our irradiatedwoolwill beprogressively transformed into a corresponding content of CA.

Processes initiated by EB in wool take place with a ratecontrolled, inter alia, by concentration of the sulphur bridges, vol-ume of available oxygen (diffusion from environment or humiditypresent in the fibre) and concentration of generated active species.Thus conditions of the reaction course (1) and (2) are differentiatedfor each sub-structure and absorbed dose. For generation of S-oxidized products the inner structures will be adequately behindcompared with the surface layer. Kinetics of individual trans-formative steps can vary, too. In our case, regarding the samecharacter of the treated fibre, the only variable is absorbed dosewhich determines the concentration of generated active

Table 2Vibrations used on hydrocarbon group analysis [18].

Type of vibration/groups ṽ (cm�1)

Deformation vibration d(CH) 1340Valence vibration n(CH2) 2850Valence vibration n(CH3) 2960

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158154

intermediate products and the rate of their conversion. Both factorsincrease with the absorbed dose intensity. Transmission FTIRspectroscopy registers information on immediate sum of partialprocesses running in individual layers cortex e macrofibrils e mi-crofibrils e protofibrils e keratin throughout the whole fibre. Wesuppose that this is one of possible cause for observed S-oxidizedspecies fluctuation.

According to our best knowledge the study [13] has been theonly which applied EB irradiation to modify wool. However, theused electron accelerator was with only 300 keV compared to ouronewith 5 MeV of installed energy and, to reach absorbed doses upto 400 kGy, it required time over 10 h, while in our case it took onlya fraction of a hour. In addition, in Fatarella's work [13], the spectracollected by means FTIR-ATR techniques could monitor an objectonly to the depth around 0.5 microns. Thus within the usual woolfibre thickness of about 25 microns, ATR spectra cannot display thesituation within whole fibre volume. Therefore our findings cannotbe confronted with the study [13] results. In those experiments inair, from 200 kGy absorbed dose the amount of S-sulph increasedmildly and CA content reduced while the amounts of CMO and CDOdid not change within whole 400 kGy scale of the absorbed doses.On the other hand it is notable that under oxygen or nitrogenplasma treatment, the content of S-sulph indicated pulsing (S-curve) [13] similarly as in our case. Besides this the plasma treat-ment increased wettability of the wool significantly unlike anegligible EB effect. This result indicates a much smaller impact ofthe EB treatment used in that study compared with our experi-ment. Although the same absorbed doses were applied, in our casethe absorbed dose rate was incomparable higher and this is why ithas led to more pronounced variations of the fibres.

3.2. FTIR analysis of hydrocarbon groups

The EB energy affected the whole volume of the sample and hadimpact on hydrocarbons functional groups, too. It is known thatbesides oxidative scission in polyamides, similar to peptide chainsby eCOeNHe bond and then also keratin, EB initiates crossbondformation via hydrocarbon portion of the chain. It was found thatfor PA-6 the absorbed dose with detectable networked fraction, i.e.gel point, occurs about 200 kGy absorbed dose [12] and is precededby chain branching. As the formation of crossbonds through hy-drocarbon groups is linked with variation of eCH, eCH2 and eCH3groups, we analysed the change of the individual groups dependingon the absorbed doses (Fig. 2). The vibrations used on FTIR analysisare presented in Table 2.

Fig. 2. Dependence of the relative variations of selected hydrocarbon groups in theirradiated wool on absorbed dose.

It is generally accepted that under irradiation of hydrocarbon byEB the primary step is interaction of electrons with hydrocarbonand the result is excitation of CeH bond and next formation ofradicals as follows:

R1eCH3���!e�

R1eCH�

2 þ H�

(3)

R2eCH2eR3���!e�

R2eC�HeR3 þ H

�(4)

ðR4R5R6ÞCH���!e�

ðR4R5R6ÞC� þH

�(5)

The formed radicals can combine with each other:

2R1eCH�

2/R1eCH2eCH2eR1 (6)

and also give a three-dimensional network structure:

2R2eC�HeR3/R2eCHðR2eCHeR3ÞeR3 (7)

2ðR4R5R6ÞC�/ðR4R5R6ÞCeCðR6R5R4Þ (8)

Alkylradicals can terminate also accepting hydrogen:

R1eCH�

2 þ H�/R1 � CH3 (9)

etc.Reaction (3) implies decrease of eCH3 group content and reac-

tion (9) their increase,eCH2e groups reduce because of reaction (4)and grow with reaction (6) or by accepting H� by radical fromprocess (4). The decreasing of eCH group content occurs with re-action (5) while they increase with reaction (7) or by accepting H�

by radical from process (5).Besides the source of hydrocarbon groups available to create

crosslinks in keratin chain, also wool lipid components intimatelyassociated with the surface proteins, are present [19]. There is aconsiderable amount of hydrocarbon fraction in the lipids but,content of the lipids is low. Formation and oxidation of the gener-ated polymeric as well as lipid radicals by oxygen from aircontribute to the fluctuation of the hydrocarbon groups. Theircombination can prevail over scission producing crosslinks athigher absorbed doses and so higher stationary radical concentra-tion [20]. In our case in the absorbed dose region up to 150 kGy, theamounts of the followed hydrocarbon groups show the fluctuation(Fig. 2) however, without more detailed studies it is difficult toidentify their origin. At around 250 kGy absorbed dose the contentof CeH groups slightly grows, we believe that the formation ofcrosslinks occurs across the hydrocarbon groups (reaction (7)),most likely in the random region, but with a low network density.Just around the 200 kGy absorbed dose also the gel point for syn-thetic polyamides is found [12]. We assume that at higher absorbeddoses reaction (7) prevails whereas the content of CH groupsincreased over 1.45 times of the initial value. Also study [21]implied a similar finding and reports that a polymer-like sub-stance produced by decomposition of lipid was detected on thesurface of the plasma treated fibres which could come from thenetworking of the lipid components. We suppose that an

Fig. 3. Variation of secondary structure in the irradiated wool depending on absorbeddose.

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158 155

intermediate decrease of the CeH groups observed for the highestabsorbed dose of 400 kGy is connected with the breaking of thepolymeric chain into shorter segments. The observed small growthof eCH3 groups as ending groups of the formed fragments in termsof reaction (9) should correspond to this.

3.3. Secondary structure

Conception of EB effect on the wool secondary structure couldcontribute in explaining of the fluctuation of the resulting chemicalspecies. Therefore we used FTIR data to analyse the respondingconformations. The shape of bands with frequency near 1600 cm�1

assigned to Amide I and II vibrations was differentiated. The fre-quency is sensitive to protein conformation (a-helix, b-sheet,random). However it is suspected that the differences can beascribed to the differences in the water content of fibres due to anHeOeH bending mode at about 1640 cm�1. This is supposed topush up the intensity of the amine I band [21]. Therefore in thisstudy, we used Amide II band to estimate the particular confor-mations. The applied wavenumbers are showed in Table 3. TheAmide II absorption bands were curve-fitted into individualGaussian components regarding the second derivation of spectraand the adequate peaks listed in Table 3. After adding the relatedareas, a-helix, b-sheet and random conformations were estimatedas the percentage of the total area (Fig. 3).

The conformation of a-helix is characteristic for thewool fibre inits natural state, therefore it is found in the fibre which is notstretched along its axis. During stretching, the a-helix declines andb-keratin appears. Several authors pointed out that the secondarystructure of wool is variable depending on the wool origin and isconsiderably sensitive to chemical treatments [22,23]. In our casethe wool treatment, before being supplied, involved somestretching within carding. A direct consequence could be a partialtransformation of a-coil to b-strand. We suppose that this is thereasonwhy our starting sample showed secondary structure with alower portion of a-helix (less than expected 50%) giving ratio a-helix: b-sheet: random ¼ 35: 30: 35. The next measurements led tothe observation of fluctuation in the secondary structure, too(Fig. 3). The disulphide bridges, H-bonds, electrovalent bonds, etc.,in wool fibre, all represent a hindrance to the movement of themolecular segments, and thus to the secondary structure trans-formation. The presence of the interfibrillar linkages by the non-helical “tails” of the low-sulphur microfibrillar proteins may giverise to physical entanglement in the keratin systems [24]. In ourexperiment already the first absorbed dose of 16 kGy provided thewool with enough energy in order to release the physical entan-glements, and originally a forced partial change of the a-helical intothe b-sheet conformation under the wool-carding, to transformback into the a-helical. As the portion of the random conformationdid not change, the a-helix recovered going to the 65% level and theb-sheet practically disappeared. The a-helix was in predominancealso for 25 kGy and 40 kGy absorbed doses. For the 63 kGy absorbeddose a change happened where a substantial transformation of thea-helix into the b-sheet occurred, thus the wool denatured. Underthis absorbed dose the b-sheet portionwas at 50% level, the highestone of all the absorbed doses while the a-helix ratio was only 11%.Just for this 63 kGy absorbed dose we observed significantly lower

Table 3Wavenumbers of Amide II for estimation of secondary structure conformations [21].

Peak (cm�1) Species Groups Conformation

1500e1600 Amide II eCONHe1545 a-helix1540 Random1530 b-sheet

generation of S-sulph and a moderate increase of CMO, CDO and CAformation. It is obviously that energy of this absorbed dose wasused for the change of the secondary structure as well as generationof the S-oxidized species. The next absorbed dose of 100 kGy pro-voked the increasing of the a-helix to 32% and the decreasing of theb-sheet to 41%, what was followed by repeated growth of S-sulphand reduction of CMO, CDO and CA amounts. In the range from100 kGy absorbed dose with the prevailing b-sheet over the a-helixwe measured also decreasing S-sulph formation. Simultaneouslyfrom the dependence shown in Fig. 1, it is possible to deduce a clearincrease in production of CA and CDOwith no generation of S-sulphand CMO. The a-helix fraction attained the level of the originalcombing wool at the highest absorbed dose only. We interpret thisa-helix increasing by the beginning of the breaking chains whenthe shorter protein segments could more easily take the energeticpreferable a-helix conformation thanks to better mobility. Thesmall growth of the ending eCH3 groups and simultaneousreduction of CeH groups (Fig. 2) support this interpretation.

From the results obtained it is obvious that, the EB irradiation ofwool induces changes in distribution of secondary structure con-formations as well as composition of S-oxidized species. Unless theinitial chain length changed, a higher content of a-helix affectedgeneration of S-sulph favourably and vice versa while the b-sheetpredominance facilitated the formation of CDO and CA. The resultsindicate that the highest absorbed dose of 400 kGy alreadyinitialized the breaking of the peptide chain. The shifts of the bandwavenumbers for the followed functional groups in the FTIRspectra can be assigned to the variation in the secondary structure,too.

3.4. Thermal properties

Additional information concerning the effect of irradiation onthe material structure was obtained from DSC measurementsproviding useful conception of thermal processes running in thetreated wool. The process characterized by a DSC maximum in thetemperature range 210e250 �C for different keratins is variablyreferred to as crystal cleavage [7] or denaturation of the a-helicalfraction of keratin [25]. Also other authors [3,26] consider theendothermal peak to be the a-helix denaturation or helix uncoiling[27]. Unlike synthetic polymers, natural biological materials likehuman hair or wool are often difficult to characterise thermally, inparticular when a quantitative determination is required, due to theco-existence of various biological heterogeneous components.

The measurement of the crystal cleavage enthalpy (DHc) and thecrystal cleavage temperatures Tc in our experiments are shown in

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158156

Fig. 4. The dependencies of both parameters on absorbed dosemeasured from the first DSC heating reflect immediate changes inthe original crystalline structure including oxidation effectsoccurring during irradiation.

The Tc indicates size of crystalline units. The decrease of Tc fromthe highest temperature of 237 �C for the parent wool to the lowesttemperature of 232 �C for the wool with absorbed dose of 400 kGyindicates a reduction of size of the ordered regions due to change inthe crystalline structures and oxidation processes during irradia-tion. We believe that the early decrease of the Tc for 16 and 25 kGyabsorbed doses is due to formation of new smaller a-helicesderivable from the disrupted entanglements as well as startingoxidation. The oxidation products increase the probability ofcreating H-bonds that can “reinforce and aggregate” transitionallyall forms of the secondary structure even if to different measure.Structure of such forms has not been specified yet. As the maximalcontent of the S-sulph corresponds to absorbed dose of 40 kGy(Fig. 1), a maximum for Tc and DHc is observed there. A repeateddecreasing in Tc over 40 kGy implies a progressive reduction of thecrystalline unit sizes connected with clearer changes in the sec-ondary structure. In the region around 250 kGy absorbed dose, asmall change is observed in the slope in the Tc curve. Possibly this isdue to a small portion of crosslinks which should occur there andcontribute to a lowering of size of the ordered regions.

The running of the DHc curve reflects mainly the crystallineportions and covers thermal processes taking place in all presentforms of the secondary structure. However, also some thermaldegradation of other structural components affecting the enthalpyshould be considered within DSC heating. Therefore it is impossibleto assigned right figures to some specific conformation due to thea-helix structure being a thermally less stable secondary structurethan the b-sheet conformation [26]. In the range of the earlyabsorbed doses up to 40 kGy, dominance of the a-helix (45%) withthe b-sheet contribution (27%) corresponds to the first maximumon the DHc curve. Around 63 kGy absorbed dose, the EB energy ispreferentially consumed in inversion of a-helix into b-sheetstructure (Fig. 3) and, as visible in Fig. 1, generation of the S-sulph isretarded while CMO creation is a little higher. Continuing to100 kGy, the movement in composition of the secondary structureas well as repeated S-sulph growth caused minimization of the DHc

(Fig. 4) because those various simultaneous processes brought upconsiderable irregularity in the material. Within the range of156e250 kGy, the b-sheet conformation is dominant (Fig. 3) andright here, theDHc dependency shows the nextmaximum (Fig. 4). Itcan be supposed that thermally more stabile b-sheet shows higherspecific (melting) enthalpy than a-helix does. The small portion of

Fig. 4. Variation of crystal cleavage temperatures Tc and enthalpy DHc in the irradiatedwool depending on absorbed dose.

crosslinked species is of negligible effect on DHc. The decrease ofthe DHc value for 400 kGy is linked with degradation includingoxidation due to the irradiation and the formation of the shorterchains with helical structure. The DHc figure for the highestabsorbed dose corresponds with DHc for the parent wool (Fig. 4)and also a-helix portion is the same (Fig. 3). However, as indicatedby the lowest Tc (Fig. 4), the helical structure belongs to the shorterpeptide chains formed under the breaking of the original chains. Inaccordance with that observation also the decrease of CH-groupsand simultaneous increase of the ending CH3 groups are observeddue to the chain breaking (Fig. 2).

3.5. Tensile strength

All SeS bonds in wool act as elastically effective crosslinks [24]and structural variations have to manifest themselves in the me-chanical properties of the fibres. It is generally known that varianceof individual measurements of tensile characteristics within onesort of fibre is rather large. That is why more measurements areneeded for one result value. In this study the intervals of the valuesðx±sÞ for results of tensile strength (Fig. 5) overlap within the entireabsorbed dose scale. So it cannot be said that there are statisticallysignificant differences. However, certain tendency is observable inthis dependence and we can conclude that within the appliedabsorbed doses, the EB effect did not reduce the strength. All dataare over level of the initial strength and only the strength for thefinal absorbed dose is next to figure for the untreated wool.

The tensile test is analogous with stretching slenderizationtechnology [28] where, under wool stretching, the secondarystructure transforms from a-helix to b-sheet and, the stretchedfibre becomes thinner. The broken fibre with the increasing portionof the b-sheet shows a higher strength and lower elongation atbreak [29]. The more of the b-sheet conformation, the higherstrength and lower elongation at break are. However, this consid-eration is applicable fully only to wool of the same chemicalcomposition. In this work each absorbed dose treated the wooldifferently, so that the responding observations (Figs. 5 and 6)represent a mixed effect of the fluctuation of both secondarystructure and chemical species.

Variations of the strength (Fig. 5) can be assigned to the mixedeffect of transition of the individual conformations from a-helix tob-sheet as well as intensive generation of polar species creatingconditions to increase number of H-bonds. Both have a veryimportant influence on the mechanical properties of the wool fibrein the whole absorbed dose range. At the maximal increasing by35% for 156 kGy absorbed dose, the b-sheet conformation is

Fig. 5. Variation of tensile strength at break of the irradiated wool depending onabsorbed dose.

Fig. 6. Variation of elongation at break of the irradiated wool depending on absorbeddose.

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158 157

dominant having higher strength than a-helix and reversely, a-helix shows higher elongation. In study [30] similar facts wereobserved when UV irradiation, and functionalization by graftingantibacterial agent, increased tensile strength and elongation ofwool fibre by 16% and 7% respectively. In another study [31], woolfabric treated with a roll-to-roll atmospheric plasma showedincreasing tensile strength by 13% and elongation at break by 19%.After that treatment by low temperature plasma it was noted thattensile energy increased steadily with prolonged treatment time.However, the increment was not so great when compared with theuntreated fabric [32]. The plasma treatment in the last mentionedstudies was not linkedwith energy supply intowhole volume of thefibre as it was under EB treatment and therefore the increase ofstrength was less profound than in our case. At the higher absorbeddoses, also the crosslinking came positively over to small measureversus negative effect of starting breaking of the carrying skeleton.Consequently the resulting demonstration is considered to be sumof the presented contributions.

3.6. Elongation

In general elongation is more sensitive to structural variationsthan strength. A change of a material elongation is observablealready under a mild structural variation while some strengthchange of the material may not yet be detected. The EB disruptsprogressively both intramolecular H-bonds and intermoleculardisulphide bonds in the treated wool. Thus under action of tensileforce, the a-helices begin to unfold more easily and the fibresextend more willingly. So in the range of the prevailed a-helicesup to 40 kGy absorbed doses (Fig. 3), the corresponding elongationgrows (Fig. 6). Above this absorbed dose, less ductile the b-sheetconformation predominates and the elongation decreases. Thepresent a-coils should fully extend to b-strands at 70 %e80 %elongation [1]. At the same time it is of worth to note that not onlyconformation transformation from a-helix to b-sheet exists, butalso structural slippage of wool fibre during stretching [28]. In ourcase, the elongation at level 70e80 % is up to the absorbed dose of250 kGy (Fig. 6), i.e. all elongation figures belong mostly to b-sheetconformation ultimately. Likewise for the strength, some althoughsmall contribution of the crosslinking has to be considered about250 kGy because the formatted crosslinks act like anchoringpoints reducing the fibre elongation. At the highest absorbed doseof 400 kGy, the beginning of breaking of the carrying peptideskeleton diminished the elongation as documented by the devel-opment of CH and CH3 groups (Fig. 2) as well as the lowest Tc value(Fig. 4).

4. Conclusions

Wool irradiated by an accelerated electron beam in air from5 MeV source with absorbed doses within 0e400 kGy and the doserate of 750 kGy/h showed some fluctuations in amount of the S-oxidized species as S-sulphonate, cystine monoxide, cysteic acid,cystine dioxide, temperature and enthalpy of the crystal cleavageand partially, in tensile properties. The fluctuations were associatedwith variations of the secondary structure relative to the absorbeddose. Increasing absorbed dose changed the a-helix from majorityinto minority non-monotonously. The a-helix conformation wasfavourable for the S-sulphonate formation while the b-sheetrestrained it. Over 220 kGy absorbed dose, only formation of thecystine dioxide and cysteic acid was observed. The development ofthe hydrocarbon groups indicated a mild formation of crosslinksaround 250 kGy and the beginning of breaking of the peptide chainwas observable for higher absorbed dose. The DSC measurementspointed out to formation of smaller fragments with the a-helixconformation and for the highest absorbed dose 400 kGy, portion ofthis conformation was consistent with the portion in the parentwool. Within thewhole range of doses, the tensile strength at breakdid not change statistically significantly while the original elonga-tion firstly increased and then, a monotonous reduction wasobserved.

References

[1] Yao J, Liu Y, Yang S, Liu J. Characterization of secondary structure trans-formation of stretched and slenderized wool fibers with FTIR spectra. J EngFibers Fabr 2008;3:1e10. http://www.jeffjournal.org. 1e10, [accessed10.02.14].

[2] Rippon JA, White MA. Curing of polyurethane prepolymers II-Thermal stabilityon wool. J Soc Dye Colour 1972;88:443e7. http://dx.doi.org/10.1111/j.1478-4408.1972.tb03055.x.

[3] Aluigi A, Vineis C, Varesano A, Mazzuchetti G, Ferrero F, Tonin C. Structure andproperties of keratin/PEO blend nanofibres. Eur Polym J 2008;44:2465e75.http://dx.doi.org/10.1016/j.eurpolymj.2008.06.004.

[4] Zoccola M, Montarsolo A, Aluigi A, Varesano A, Vineis C, Tonin C. Electro-spinning of polyamide 6/modified-keratin blends. E Polymers 2007;7:1204e22. http://dx.doi.org/10.1515/epoly.2007.7.1.1204.

[5] Rouse JG, Van Dyke MEA. Review of keratin-based biomaterials for biomedicalapplications. Materials 2010;3:999e1014. http://dx.doi.org/10.3390/ma3020999.

[6] Li R, Wang D. Preparation of regenerated wool keratin films from wool ker-atineionic liquid solutions. J Appl Polym Sci 2013;127:2648e53. http://dx.doi.org/10.1002/app.37527.

[7] Xu W, Wang X, Cui W, Peng X, Li W, Liu X. Characterization of superfine downpowder. J Appl Polym Sci 2009;111:2204e9. http://dx.doi.org/10.1002/app.29205.

[8] Kan CW, Chan K, Yuen CWM, Miao MH. Surface properties of low-temperatureplasma treated wool fabrics. J Mater Process Tech 1998;83:180e4. http://dx.doi.org/10.1016/S0924-0136(98)00060-0.

[9] Xu W, Shen X, Wang X, Ke G. Effective methods for further improving thewool properties treated by corona discharge. SENI' GAKKAISHI 2006;62:111e4.

[10] Xue Z, Jin-xin H. Improvement in dyeability of wool fabric by microwavetreatment. Indian J Fibre Text Res 2011;36:58e62.

[11] Church JS, Millington KR. Photodegradation of wool keratin: part I. Vibrationalspectroscopic studies. Biospectroscopy 1996;2:249e58. doi:10.1002/(SICI)1520-6343(1996)2:4<249::AID-BSPY6>3.0.CO;2e1.

[12] Porubsk�a M, Janigov�a I, Jomov�a K, Chod�ak I. The effect of electron beamirradiation on properties of virgin and glass fiber-reinforced polyamide 6.Radiat Phys Chem 2014;102:159e66. http://dx.doi.org/10.1016/j.radphyschem.2014.04.037.

[13] Fatarella E, Ciabati I, Cortez J. Plasma and electron-beam processes as pre-treatments for enzymatic process. Enzyme Microb Tech 2010;46:100e6.http://dx.doi.org/10.1016/j.enzmictec.2009.10.004.

[14] Kan CW, Yuen CVM. Surface characterisation of low temperature plasma-treated wool fibre. J Mater Process Tech 2006;178:52e60. http://dx.doi.org/10.1016/j.jmatprotec.2005.11.018.

[15] Douthwaite FJ, Lewis DM, Schumacher-Hamedat U. Reaction of cystine resi-dues in wool with peroxy compounds. Text Res J 1993;63:177e83. http://dx.doi.org/10.1177/004051759306300308.

[16] Kan CW, Chan K, Yuen CWM. Application of low temperature plasma onwool - part III: surface chemical and structural composition. Nucl 2000;37:145e59.

M. Porubsk�a et al. / Polymer Degradation and Stability 111 (2015) 151e158158

[17] Erra P, G�omez N, Dolcet LM, Juli�a MR, Lewis DM, Willoughby JH. FTIR analysisto study chemical changes in wool following a sulfitolysis treatment. Text ResJ 1997;67:397e401. http://dx.doi.org/10.1177/004051759706700602.

[18] Milata V, Seg�la P, Brezov�a V, Gatial A, Kov�a�cik V, Miglierini M, et al. Aplikovan�amolekulov�a spektroskopia (Applied molecular spectroscopy). Bratislava: STUPublisher; 2008.

[19] Huson M, Evans D, Church J, Hutchinson S, Maxwell J, Corino G. New insightinto the nature of the wool fibre surface. J Struct Biol 2008;163:127e36.http://dx.doi.org/10.1016/j.jsb.2008.04.011.

[20] Laz�ar M, Rado R, Rychlý J. Crosslinking of polyolefins. Adv Polym Sci 1990;95:149e97.

[21] Mori M, Inagaki N. Relationship between anti-felting properties and physi-cochemical properties of wool treated with low-temperature plasma. Res JText Appar (RJTA) 2006;10:33e45.

[22] Cardamone JM. Investigating the microstructure of keratin extracted fromwool: peptide sequence (MALDI-TOF/TOF) and protein conformation (FTIR).J Mol Struct 2010;969:97e105. http://dx.doi.org/10.1016/j.molstruc.2010.01.048.

[23] Wojciechowska E, Włochowicz A, Wesełucha-Birczy�nska A. Application offourier-transform infrared and Raman spectroscopy to study degradation ofthe wool fiber keratin. J Mol Struct 1999;511e512:307e18. http://dx.doi.org/10.1016/S0022-2860(99)00173-8.

[24] Arai K, Sasaki N, Naito S, Takahashi T. Crosslinking structure of keratin. I.Determination of the number of crosslinks in hair and wool keratins from

mechanical properties of the swollen fiber. J Appl Polym Sci 1989;38:1159e72. http://dx.doi.org/10.1002/app.1989.070380613.

[25] Milczarek P, Zielinski M, Garcia ML. The mechanism and stability of thermaltransition in hair keratin. Colloid Polym Sci 1992;270:1106e15.

[26] Tonin C, Aluigi A, Vineis C, Varesano A, Montarsolo A, Ferrero F. Thermal andstructural characterization of poly(ethylene-oxide)/keratin blend films.J Therm Anal Calorim 2007;89:601e8.

[27] Rama Rao D, Gupta VB. Thermal characteristics of wool fibers. J Macromol SciPhys 1992;31:149e62. http://dx.doi.org/10.1080/00222349208215509.

[28] Tang F-J, Liu H-L, Yu W-D. Secondary structure analysis of stretched woolusing FT-IR and tensile stress-strain curve. Basic Sci J Text Univers 2007;20:412e7.

[29] Kwak SA, Lee JY, Lee DH, Jeon BS. Mechanical properties of wool fiber in thestretch breaking process. Fibers Polym 2007;8:130e3.

[30] Niu M, Liu X, Dai Hou W, Wei L, Xu B. Molecular structure and properties ofwool fiber surface-grafted with nano-antibacterial materials. SpectrochimActa A 2012;86:289e93. http://dx.doi.org/10.1016/j.saa.2011.10.038.

[31] Ceria A, Rombaldoni F, Rovero G, Mazzuchetti G, Sicardi S. The effect of aninnovative atmospheric plasma jet treatment on physical and mechanicalproperties of wool fabrics. J Mater Process Tech 2010;210:720e6. http://dx.doi.org/10.1016/j.jmatprotec.2009.12.006.

[32] Kan CW. KES-F analysis of low temperature plasma treated wool fabric. FibresText East Eur 2008;16:99e102.