Ethyl chitosan synthesis and quantification of the effects acquired after grafting it on a cotton fabric, using ANOVA statistical analysis

Accepted Manuscript Title: Ethyl chitosan synthesis and quantification of the effects acquired after grafting it on a cotton fabric, using ANOVA statistical analysis Author: Vasilica Popescu Augustin Muresan Gabriel Popescu Mihaela Balan Marius Dobromir PII: DOI: Reference:

S0144-8617(15)01098-X http://dx.doi.org/doi:10.1016/j.carbpol.2015.11.009 CARP 10527

To appear in: Received date: Revised date: Accepted date:

11-5-2015 29-10-2015 4-11-2015

Please cite this article as: Popescu, V., Muresan, A., Popescu, G., Balan, M., and Dobromir, M.,Ethyl chitosan synthesis and quantification of the effects acquired after grafting it on a cotton fabric, using ANOVA statistical analysis, Carbohydrate Polymers (2015), http://dx.doi.org/10.1016/j.carbpol.2015.11.009 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

*Highlights (for review)

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Synthesis of triethyl chitosan using ethyl chloride as alkylation agent Alkylation and grafting were confirmed by FTIR, 1HNMR, XPS, DSC, TGA/DTG Quantified results: take-up degree, wrinkle-recovering angle, tensile strength ANOVA shows the maximal effects of chitosan, ethyl chloride and their interaction Durability of effects was highlighted with tinctorial and diffusimetric methods

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1  2  3 

Ethyl chitosan synthesis and quantification of the effects acquired after grafting it on a cotton

4 

fabric, using ANOVA statistical analysis

5 

Vasilica Popescua, Augustin Muresana*, Gabriel Popescub, Mihaela Balanc, Marius Dobromird

7 

a

8 

Management, 29 Blvd. Mangeron, TEX 1 Building, Iasi-700050, Romania;

cr

“Gheorghe Asachi” Technical University, Faculty of Textiles, Leather Engineering and Industrial

b

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“Gheorghe Asachi” Technical University, Faculty of Mechanical Engineering, 43 Blvd. Mangeron

Iasi-700050, Romania.

11 

c

12 

Romania

13 

d

14 

Blvd. Carol I, Iasi 700506, Romania.

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Correspondent author: [email protected]

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“Petru Poni” Institute of Macromolecular Chemistry, 41-A Alley Grigore Ghica Voda, Iasi-700487,

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Tel: +40 0721194298

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“Alexandru Ioan Cuza” University, Plasma Advanced Research Center (IPARC)-Faculty of Physics, 11

Abstract

18 

Three ethyl chitosans (ECSs) have been prepared using the ethyl chloride (AA) that was obtained in situ.

19 

Each ECS was applied on a 100% cotton fabric through a pad-dry-cure technology. Using the ANOVA as

20 

statistic method, the wrinkle-proofing effects have been determined varying the concentrations of AA

21 

(0.1- 2.1 mmols) and chitosan (CS) (0.1- 2.1 mmols). Alkylation and grafting mechanisms have been

22 

confirmed by the results of FTIR, 1HNMR, XPS, SEM, DSC and termogravimetric analyses. The

23 

performances of each ECS as wrinkle-proofing agent have been revealed through quantitative methods

24 

(taking-up degree, wrinkle-recovering angle, tensile strength and effect’s durability). The ECSs confer

25 

wrinkle-recovering angle and tensile strength higher than those of the witness sample. Durability of ECSs

26 

grafted on cotton have been demonstrated by a good capacity of dyeing with non-specific (acid/anionic

27 

and cationic) dyes under severe working conditions (100ºC, 60min.) and a good antimicrobial capacity.

28 

Keywords: ANOVA, ethyl chitosan, FTIR, wrinkle-proofing agent.

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Chemical compounds studied in this article

30 

Chitosan (PubChem CID: 71853); Acetic acid (PubChem CID: 176); Ethanol (PubChem CID: 702);

31 

Hydrochloric acid (PubChem CID: 313); Sodium hydroxide (PubChem CID: 14798); Zinc acetate

32 

(PubChem CID: 11192); C.I. Basic Blue 9 (PubChem CID: 104827).

33 

1. Introduction

34 

CS is an aminoglucopyran composed of N- acetyl glucosamine and glucosamine residues (Mourya &

35 

Inamdar, 2009). The most reactive functional groups are primary OH group (from C6) and NH2 bound to

36 

the C2. That is why in alkylation reactions these groups can react with alkylation agents producing O-

37 

alkyl chitosans and N-alkyl chitosans respectively, depending of the alkylation conditions.

38 

CS alkylation can be accomplished through three methods: 1). Reductive alkylation; 2). Michael addition;

39 

3). Direct alkylation (An, Dung, Thien, Dong, & Nhi, 2008).

40 

1). Reductive alkylation was initiated by Muzzarelli and Tanfani (Muzzarelli & Tanfani, 1985). They

41 

realized a Schiff base by means of formaldehyde, followed by its reduction with a reducer (sodium

42 

borohydride NaBH4). The obtaining of alkyl CS was possible using RI as alkylation agent, in the

43 

presence of NaOH and NaI. Iodine replacement by chlorine was realized through an ion exchange stage,

44 

because the utilization of methyl iodide is dangerous (even if it is very efficient), as it is readily volatile

45 

and produces cancer. This stage was accomplished by adding NaCl over trialkyl CS and depositing it for

46 

1- 2 days at room temperature. This method permitted to obtain N-alkylated products, without realizing

47 

also the O- alkylation. Thus N-alkylated products with identical (Xu, Xin, Li, Huang, & Zhou 2010;

48 

Verheul, Amidi, van der Wal, van Riet, Jiskoot, & Hennink, 2008) or different alkyl groups were

49 

synthesized (Bayat et.al., 2006; Zhang, Ding, Ping, & Yu, 2006).

50 

2). Michael addition is based on addition reaction of some α, β non- saturated carbonyl reagents (ethyl

51 

acrylate) to the NH2 group of CS (Mather, Viswanathan, Miller, & Long, 2006). The acrylic acid was

52 

used as reagent and N-carboxyethylated derivates were obtained (An, Dung, Thien, Dong, & Nhi, 2008).

53 

3). Direct alkylation was initiated by Dormand and coworkers, who realized the quaternization of amino

54 

group from CS by means of CH3I as alkylation agent (Domard, Rinaudo, & Terrasin, 1986).

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Direct alkylation has only one stage, namely direct/proper alkylation of the NH2 group in conditions of

56 

high temperature and alkaline medium. The obtained product is under the form of a quaternary salt of

57 

trialkyl CS iodide type. The conversion in chloride was performed through an ion-exchange stage in

58 

which the alkylated polymer was brought in contact with a solution of NaCl for a long period (1- 2 days)

59 

to accomplish the ion exchange (I- by Cl-). Even though the objective of direct alkylation consisted in

60 

obtaining only N-alkylated products, it was found that the O-alkylation phenomenon was still present.

61 

This phenomenon was the more pronounced, the higher was the degree of quaternization (DQ)

62 

(Jintapattanakit, Mao, Kissel, & Junyaprasert, 2008).

63 

The efficiency of the direct alkylation process depends on several factors: molar ratio: CS:CH3I:NaOH

64 

(Domard, Rinaudo, & Terrasin, 1986), reaction time (le Dung, Milas, Rinaudo, & Desbrieres,1994), the

65 

nature of the base which forms the alkaline medium (Hamman, & Kotze,2001),

66 

concentration (Curiti, Britto, & Compana-Filho, 2003), solvent type and temperature (Rúnarsson, et. al.,

67 

2007; Rúnarsson, Holappa, Jónsdóttir, Steinsson, & Másson, 2008), as well as the number of repeating

68 

stages (Sieval, et. al., 1998; Snyman, Hamman, Kotze, Rollings, &. Kotze, 2002).

69 

Regardless the utilized alkylation method, the trialkylated CS products have been used until now only for

70 

the synthesis of some drugs, as they are white powders readily soluble in water.

71 

In this paper we subjected CS to a direct alkylation with AA obtained in situ. The synthesized alkylation

72 

compounds (with various substitution degrees) were used as wrinkle-proofing agents for 100% cotton

73 

fabrics. The substitution degrees were determined from 1HNMR spectra.

74 

The technology used to treat the cotton fabric with products of alkylated CS was of pad-dry-cure type.

75 

The experimental protocol was realized such that to permit the application of ANOVA as statistical

76 

analysis method (with two independent variables).

77 

The mechanism of grafting the alkylated chitosan on cotton has been backed up by the results of FTIR,

78 

as well as its

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1

HNMR, XPS, SEM, DSC analyses. The effects produced by synthesized alkylated compounds are

79 

revealed by the wrinkle recovering angles (WRA), the values of tensile strength, durability of cotton

80 

chemical modification through grafting (tinctorial method). ANOVA (two-way analysis of variance) has 3   

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indicated with a 99% confidence the factors influencing chitosan alkylation and has quantified the

82 

obtained effects.

83 

2. Experiments

84 

2.1. Materials

85 

Highly viscous CS was obtained from Fluka AG, and the offered characteristics are presented in

86 

Supplementary data S1 (Fig. 1S). Reagents as hydrochloric acid 35%, ethanol 95%, glacial acetic acid

87 

100%, sodium hydroxide (p.a.) and sodium carbonate (p.a.) were obtained from Merck Company, zinc

88 

acetate 98% from Sigma Aldrich, and non-ionic surfactant (Romopal O) from Romtensid S.A. Timisoara,

89 

Romania. The dyestuffs C.I. Acid Red 88 (Acid Red AV) was obtained from Colorom S.A. Romania,

90 

while C.I. Basic Blue 9 (Methylene Blue) from Fluka A.G. The structures of these dyestuffs are presented

91 

in Table 1S from Supplimentary data.

92 

The 100% cotton fabric was obtained from IASITEX SA Romania and has the following characteristic:

93 

plain weave with a weight of 100 g/m2. The cotton fabric was prepared through an operation of scouring

94 

with 2% NaOH, 1% Na2CO3, 1% Romopal O, a liquor ratio of 1:50, at the temperature of 100oC for 2 h,

95 

followed by a hot and a cold rinsing and then drying at room temperature.

96 

2.2. Synthesis of ECSs necessary for spectroscopic analyses (FTIR, 1HNMR):

97 

Three ethyl chitosans (ECSs) have been prepared, namely: monoethyl chitosan (MECS), diethyl chitosan

98 

(DECS) and triethyl chitosan (TECS).

99 

AA was used as alkylation agent; this is a highly volatile gas with anesthetic properties. Even if it can be

100 

found in liquefied form, it is difficult to work with it because it volatilizes at 12oC. In order to avoid this

101 

inconvenient, we resorted to its in situ synthesis. Ethanol, HCl and zinc acetate (as catalyst) were put in

102 

contact (in equimolar ratio) in an Erlenmeyer flask perfectly sealed with a rubber stopper. The mixture

103 

was maintained at 40°C for 2h under continuous stirring. CS solution was also introduced in the flask by

104 

means of a syringe. The amount of CS used in these experiments ranged between 0.1- 2.1 mmol. The

105 

stirring continued for 60 minutes, at 40ºC. We have used various molar ratios CS: AA starting from 1:1

106 

to 1:11. NaOH was added also with a syringe for the precipitation of ECSs. The NaOH solution was used

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in concentration of 1M. The formation of a translucid gel penetrated by gas bubbles climbing to the top of

108 

the glass flask was noticed. As the quantity of NaOH increased, the gel transformed in a whitish

109 

precipitate, whose particles were the biggest in the case of trialkyl CS and the smallest for monoalkyl CS.

110 

After obtaining the precipitate, the stopper of the glass flask was removed and the content was transferred

111 

in a Berzelius beaker, where its pH was determined. It was found that pH varied between 4 and 5 (i.e. 4

112 

MECS, 4 for DECS and 4-5 for TECS). Using 1M NaOH solution, a pH > 8 was obtained and it has been

113 

noticed that the precipitate mass increased as NaOH was added. In this way ECSs were obtained, with

114 

various alkylations degrees, depending on the utilized CS: AA: NaOH molar ratios (Fig. 1). The afferent

115 

precipitates MECS, DECS and TECS (corresponding to molar ratios CS: AA: NaOH= 1:1:1;

116 

0.39:1.806:1.806 and 0.1:1.1:1.1 i.e. for 1 mmol CS it follows that CS: AA: NaOH= 1:1:1; 1:4.6:4.6 and

117 

1:11:11, respectively) were then stored in laboratory for 8 days, to evaporate the water. The obtained

118 

solid product was washed several times with ethanol solution until the pH of residual wash liquor equals

119 

7. It was then storred again to evaporate the ethanol, and the resulted solid form was subjected to

120 

spectroscopic analyses (FTIR, 1HNMR).

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2

3

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Fig. 1. Precipitates corresponding to MECS (1), DECS (2) and TECS (3) respectively.

122 

The process of chloride TECS production is presented in scheme (1) catalyst

123 

HCl

+

CH2 CH2

OH

(2)

OH O HO

O NH2 CS powder

124  125 

CH 2 CH 2

+ CH 3COOH

O

Cl

+

OH O HO

O +

(1)

H2O

O

-

H3N OOC

(3) + 3 ClC2 H5 + 2 NaOH - CH COOH 3 - 2 NaCl - 2 H2O

CH 3 Disolved CS

OH O

O HO +

O -

Cl C2H5 H5C2 C2H5 N

Chloride triethyl CS (TECS)

Scheme 1. Chemical reactions for synthesis of TECS. 5   

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These reactions have been proved by the results of spectroscopic analyses (FT-IR and 1HNMR).

127 

In order to have a basis for comparison, we have used the same methods to analyze the unmodified CS.

128 

Its presentation form was a film obtained as follows: 0.5 g CS powder was solved with a 6.2% acetic acid

129 

solution. A consistent, quite viscous solution was obtained. On a clock glass we put 5 ml of CS solution

130 

and deposited it for 2 days at room temperature. When water evaporated a film was formed, that was

131 

washed and then dried. Finally, this film was easily detached from the clock glass and subjected to

132 

spectroscopic analyses.

133 

2.3. Analysis method

134 

2.3.1. ANOVA Statistical method

135 

ANOVA represents nowadays a set of statistical method of major importance within the general frame of

136 

the procedures for experimental data study (Tovissi & Vodă, 1982). ANOVA is a statistical method that

137 

indicates the causes which explain the variation of a process and the factors with significant influence.

138 

One can quantify with ANOVA the main effects generated by the influencing factors, as well as the

139 

interactions between these factors. As the dispersions of statistical variables are not additive, for

140 

decomposition, in the process of dispersional analysis the quadratic variation is used, whence the name of

141 

“variation analysis” (Baron, 1979; Gluck, 1971; Rancu & Tovissi, 1963; Văduva, 1970). The principle of

142 

the method, experimental protocol and ANOVA components, together with the means of dispersion

143 

estimates are presented in Supplementary data /Tables 2S and 3S.

144 

Commonly, many scientists use the already realized software of ANOVA method to investigate certain

145 

topics; yet in this work the software for dimensional bifactorial analysis with systematic ANOVA effects

146 

has been elaborated in C++ language by one of the authors (Popescu G.). The logical diagram of this

147 

software is presented in Supplementary data/Fig. 2S. The software was tested on numerous themes of

148 

textile chemical finishing and the results were pertinent (Popescu, Butnaru, & Popescu, 2000; 2001a;

149 

2001b; 2002).

150 

The software of ANOVA method was conceived such that to permit the realization of a multiple

151 

comparison of factors’ levels by using the Scheffe method (Jaba, 1998; Mărgăritescu, 1981).

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2.3.2. Pad-dry-cure technology for wrinkle-proofing of cotton treated with ECS

153 

The mixture consisting in ECS precipitate and residual solution (obtained after NaOH adding, as in

154 

chapter 2.2) was stirred 15 minutes and then was applied on cotton samples. The application of each ECS

155 

on the cotton fabric was made according to a pad-dry-cure technology. The pad-dry-cure technological

156 

parameters (time, temperature) were the same for all the samples treated with ECSs (cotton samples of

157 

2 g each were taken on the direction of warp and weft respectively). After impregnation stage, the

158 

material was squeezed between padder’s cylinders to an 80% squeezing. The samples were dried at

159 

100oC for 3 minutes, followed by condensation/curing stage performed at 160oC for 3 min, on minitherm

160 

ERNST BENZ AG apparatus (Textilmachinen Rümlang – Zurich); due to high temperature and the

161 

presence of catalyst (remained from chemical reaction (1)), in this stage the water is eliminated between

162 

the primary OH groups of cellulose macromolecular chains and the OH groups from C6 of ECSs.

163 

Finally, the samples were washed with warm water at 40oC to remove the catalyst and the residual

164 

substances, cold washed and dried at room temperature.

165 

2.3.3. Analyses that confirm alkylation and grafting on cotton

166 

2.3.3.1. FT-IR analysis

167 

The FT-IR analyses of ECSs and of the samples treated with ECSs were performed on Multiple Internal

168 

Reflectance Accessory (SPECAC, USA) with ATR KRS- 5 crystal of thalium bromide- iodine, having 25

169 

reflections, and investigation angle of 45o. This accessory device was attached to the Spectrophotometer

170 

FTIR IR Affinity-1 Schimadzu (Japan); the spectra registration was realized with 250 scans in the 4000-

171 

600 cm-1 rang. After the registration, the absorption spectra have been electronically superposed using the

172 

LabCognition software.

173 

2.3.3.2. 1HNMR

174 

The 1H-NMR spectra were recorded on a Bruker Avance DRX 400 MHz Spectrometer equipped with a

175 

5mm BBFO direct detection probe and z-gradients. The samples were partially dissolved in D2O + HCl

176 

then filtered. Spectra were recorded at room temperature. The spectra were referenced relative to internal

177 

TSP.

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178 

Literature presents the relations used to compute the substitution degrees in the case of CS methylation

179 

using the information supplied by 1HNMR spectra (Cafaggi, et. al., 2007; Chen, Zhang, & Huang, 2007;

180 

Rúnarsson, et. al., 2007). Extrapolating the relations used by Rúnarsson (Rúnarsson, et. al., 2007) for the

181 

case of CS ethylation and using the integrals of the peaks from 1HNMR spectra (afferent to MECS,

182 

DECS and TECS), the substitution degree at N have been computed as follows:

183 

DM= (CH2+ CH3) / [H] • 6/5 • 100

184 

DE= 2 • (CH2 + CH3) / [H]• 6/10 • 100

[%]

(5)

185 

DQ= 3 • (CH2 + CH3) / [H] • 6/15) • 100 [%]

(6)

186 

where:

187 

DM is the degree of monoethylation;

188 

(CH2+CH3) = integrals of the peaks corresponding to monoethyl group attached at N (when H in CH2

189 

group appear between 2.83- 2.90 ppm and 1.10- 1.13 ppm respectively for H in methyl group);

190 

DE is the degree of diethylation;

191 

2 • (CH2 + CH3) = integrals of the peaks corresponding to diethyl groups attached at N (when H in CH2

192 

group appear at 3.0-3.05 ppm and 1.0-1.06 ppm respectively for H in methyl group);

193 

DQ is the degree of triethylation / quaternization;

194 

3 • (CH2+CH3)= integrals of the peaks corresponding to triethyl groups attached at N (when H in CH2

195 

group appear at 3.42-3.52 ppm and 1.32-1.34 ppm respectively for H in methyl group).

196 

[H] is the sum of the integrals of corresponding peaks H-2, H-3, H-4, H-5, H-6, H-6’;

197 

The alkylation phenomenon can also appear at the level of oxygen atoms, more precisely at O6 and O3

198 

(Rúnarsson, et. al., 2007).

199 

2.3.3.3. XPS Analysis

200 

The XPS analysis was carried out on the cotton samples grafted with ECSs. XPS analysis was possible

201 

using AXIS ULTRA DLD spectrometer of Kratos Analytical Ultra DLD with monochromatic aluminum

202 

source (power 150 W).

203 

2.3.3.4. SEM

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A QUANTA 200 3DDUAL BEAM electron microscope was used, which is a combination of two

205 

systems (SEM and FIB), by whose means, by sending an electron beam on the treated samples, three-

206 

dimensional images could be obtained, with a magnification of 100,000X. SEM analysis offers images

207 

which prove the modifications of the cotton fabric surface through ECSs grafting.

208 

2.3.3.5. Calorimetric and thermogravimetric analyses

209 

Calorimetric analysis (DSC): The DSC curves were recorded on a METTLER TOLEDO DSC1

210 

apparatus in inert atmosphere (nitrogen) at a flow rate of 150 mL/min, with a heating rate of 10ºC/min.

211 

Three cycles were applied: heating (25-250)ºC, cooling (250-25)ºC and re-heating (25-250)ºC. The

212 

samples (3.5mg) were placed in aluminum pan and sealed. We used aluminum pans with lids (Al

213 

crucibles 40 μL contact capacity, type ME-00027331, with pin). An empty pan was used as reference.

214 

Thermogravimetric analysis (TGA) was performed at a MATTLER TOLEDO TGA-SDTA851e

215 

derivatograph in nitrogen atmosphere with a flow rate of 20 mL/min and heating rate of 10ºC/min,

216 

between 25 and 800ºC. The samples were placed in crucibles (ME-00024123 aluminum oxide crucibles

217 

with 70 μL contact capacity).

218 

The operational parameters were maintained constant for all the samples to produce comparable data.

219 

Each DSC/TGA analysis was repeated twice to check the repeatability.

220 

2.4. Quantification of grafting effects

221 

2.4.1. Taking-up degree

222 

The taking up degree was determined using the relation (7):

223 

Υp

224 

Yp= taking-up degree;

225 

Wa= final weight of cotton sample (weight after wrinkle proofing);

226 

Wb= initial weight of cotton sample (weight before wrinkle proofing).

227 

2.4.2. Wrinkle recovering angle

228 

By using the Metrimpex FF-01 apparatus and keeping in line with German method standardized DIN

229 

53890, the wrinkle proofing angles (WRA) have been determined both on the warp and weft directions

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[%]

(7)

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(as arithmetic means of 5 determinations), as well as varying the conditions of the treated samples: dry

231 

and wet respectively.

232 

2.4.3. Mechanical properties

233 

According to ISO 9513 standard, the tensile strengths were determined on both warp and weft directions,

234 

as arithmetic means of 5 similar determinations. The H5K-T dynamometer equipped with QMAT

235 

TEXTILE software was used for the fabrics.

236 

2.4.4. Tinctorial method

237 

The tinctorial method was used to make evident the presence of amino groups (with different substitution

238 

degrees) existing in the ECSs grafted on the cotton fabric. Another objective of the tinctorial method was

239 

to prove the durability of the effects of grafting treatments with ECSs under severe conditions used for

240 

dyeing (T= 100oC, t= 60 minutes).

241 

Two classes of dyes (unspecific to cotton dying) were used to reach these objectives: anionic dyes (acid

242 

dyes) and cationic dyes. Regardless the class of used dyes, the dyeing was performed in two stages: 1)

243 

keeping the fabric (for 15 minutes) in a solution with a certain pH: neutral (pH=7) (to make evident only

244 

the substituted amino groups) or acid medium (pH=5) with a view to protonize the amino groups from

245 

ECSs; 2) proper dyeing: add the dye (3%) in the dyebath, increase the temperature at 100oC and keep it

246 

constant for 60 minutes; 3) cool down to 70oC and then wash warm and cold.

247 

All the dyeing operations were performed with Mathis Color machine. The values of color strength K/S

248 

offer indications on the durability of grafting treatments with ECSs; K/S depends on several factors: pH,

249 

concentration/alkylation degree, steric effects, dye nature and structure, etc.

250 

2.4.5. Testing of antimicrobial capacity: Diffusimetric method

251 

The antimicrobial capacity was tested on two series of samples: 1st series: cotton treated with ECSs; 2nd

252 

series: cotton treated with ECSs and then with a solution of 5g/l AgNO3 (20oC for 5 min.).

253 

The diffusimetric method consists in application of textile disks (15 mm) on the surface of a bacterial

254 

culture realized in agar (1 mL culture realized in standard nutrient medium I– 25 g/L added in 100 mL

255 

agar with standard nutrient I for microbiology (37 g/L), from Carl Roth GmbH&C0.KG- Germany). At

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about 37oC, above the first hardened agar layer, a second agar layer containing bacteria was applied. The

257 

bacterial cultures of 18 hours were obtained from bacterial inoculus, which was standardized according to

258 

McFarland scale, obtaining 107- 108 CFU/ml. The utilized culture medium was seeded with the

259 

corresponding inoculus, after which textile disks subjected to different treatments were applied.

260 

As the antibacterial action is conditioned by the diffusion capacity of the active substance, an analysis

261 

was carried out at the microscope for the inhibition zones and the adjacent surfaces (near the inhibition

262 

zone and underneath the fabric). Namely, in the case when the diffusion restricts the antibacterial activity,

263 

the zones under the textile material remain without cultures. In the case when cultures appear also under

264 

the textile material, the sample is completely inactive (EN ISO 20645:2004 (D)).

265 

3. RESULTS AND DISCUSSIONS

266 

3.1. Mechanism for cellulose grafting

267 

Since the condensation reaction occurs at 160oC, it is responsible for water elimination at the level of

268 

primary OH groups belonging to macromolecular chains of ECS and cellulose. Thus, appear ether

269 

bridges (-CH2-O-CH2-) and the architectonic structure of this product is presented in Scheme 2. O

-

Cl

OH HO

+

O

N

+

HO

O

Cellulose

270 

C2H5

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O

HO

O

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C2 H5

C2H5

O

OH

O HO

curing 160⁰C (CH3COO)2Zn HO (catalyst) O

O

O

O

O

+

N

C2H5 C2H5

C2H5 O

TECS

-

Cl

+ H 2O

(8)

OH

TECS grafted on cellulose

Scheme 2. Grafting reaction of TECS on cellulose.

271  272 

3.2. FT-IR spectra

273 

3.2.1. FT-IR spectra for synthetized ECS

274 

Figure 2a shows the FT-IR spectra of pure CS powder and of the synthesized ECSs (MECS, DECS and

275 

TECS) according to scheme 1. CS alkylation is proved by the appearance of a large absorption at 1339

276 

cm-1 (Fig. 2a), characteristic to methyl C-H band. As compared with CS, the IR spectra of ECSs products

277 

are characterized by the following aspects: 1)Peak at 3364 cm-1 decreases since the bands afferent to NH2

278 

and O-H, asymmetric and symmetric stretchings, decrease as the result of alkylation, both at the level of

279 

N atom, and at O; 2)Peaks from the range 2976-2860 cm-1 afferent to CH3 and CH2 increase; 11   

Page 12 of 30

3)Alkylation at the level of N is confirmed by the following aspects deduced when comparing with CS:

281 

increase of vibrations afferent to CH2 bending (at 1398 cm-1); significant increase at 1339 cm-1 due to

282 

CH2 bending and symmetric CH3 deformation (Badawy, 2010; Badawy, Rabea, & Taktak, 2014; Coates,

283 

2000); the diminution of vibrations of NH deformation, amide II, and N-H bending of NH2 from the

284 

ranges 1650-1584 cm-1 and 673-645 cm-1 (for NH, Wagging) (Supplementary data S3, Fig. 3Sa).

ip t

280 

M

an

us

cr

a) 

b) 

Ac ce pt e

d

285 

286  287 

Fig. 2. FT-IR spectra for synthesized ECSs (a) and cotton samples treated with ECSs (b). 12   

Page 13 of 30

O-alkylation occurring at the level of primary/secondary OH group, confirmed by: 1)decrease of peaks

289 

from the range 1150-1200 cm-1 is due to overlapping asymmetric ether bridge (C-O-C) stretching and C-

290 

N stretching). Taking into account that -CH2-O-CH2- stretching determine an increase of vibrations, we

291 

infer that C-N stretching causes a significant decrease of the vibrations in this range, which confirms the

292 

destructuring of ECSs macromolecular chains (as a secondary effect of alkylation (Mourya, & Inamdar,

293 

2009)); 2)diminution of peaks at about 1042 cm-1 (for C-O stretching of C3-OH) and from 1013 cm-1 (for

294 

C-O stretching of C6-OH); 3)decrease of peaks from the range 620-617 cm-1 corresponding to OH

295 

deformation out of plane (Supplementary data S3, Fig. 3Sb).

cr

ip t

288 

3.2.2. FT-IR spectra for ECS grafted on cellulose

297 

Figure 2b shows the spectra of cotton treated with ECSs and of the untreated cotton. As at 3331 cm-1,

298 

bands afferent to NH2 and O-H asymmetric and symmetric stretchings are recorded, it follows that, by

299 

treating with ECSs, the number of free OH groups varies depending on the degree of implication in the

300 

formation of ether bonds realized between the macromolecular chains of cellulose and ECS. Other

301 

differences appeared between the spectra from Fig. 2b are: 1) within the range 2918-2851 cm-1 a

302 

vibration characteristic for C-C bond is recorded for the cotton samples treated with ECSs, the peaks are

303 

higher than for untreated cotton confirming the ECSs grafting on cotton; 2) at 1543 cm-1 one registers

304 

the absorption characteristic to NH2 group (N-H bending), which decreases in the direction MECS>

305 

DECS > TECS, because with the increase of the substitution degree, these NH2 groups were transformed

306 

more efficient into secondary or tertiary groups (i.e. groups which have no absorption within this range:

307 

-N-(CH2-CH3)2 and -N+-(CH2-CH3)3) respectively.

Ac ce pt e

d

M

an

us

296 

308 

Taking into account that part of the primary and secondary OH groups of ACSs are involved in O-

309 

alkylation, it follows that the number of free OH groups decrease; yet, this decrease is not visible due to

310 

the small degree of O-alkylation, such that some OH groups remain uninvolved in this alkylation process.

311 

The contribution of those OH groups uninvolved explains the slight increase of peaks from 1053 cm-1

312 

(assigned to secondary alcohol), 1024 cm-1 (assigned to the primary OH groups (C-O stretch). The slight

313 

increase of peak from 1159-1200 cm-1 is due the etheric bond, C-O-C (Supplementary data S3, Fig. 4S). 13   

Page 14 of 30

314 

3.3. 1HNMR for synthesized ECSs

315 

MECS

316 

1

317 

2.90 ( m, –CH2 from ethyl group, at N), 2.90-3.0 (m, H-2 GluN), 3.7–4.0 (m, H-2 (GluNAc), H- 3, H-4,

318 

H-5, H-6, H-6’), 4.5–5.0 (m, partly overlapped by water peak, H-1).

319 

DECS

320 

1

321 

3.05 ( m, –CH2 from ethyl group, at N),), 2.87-2.90 (m, H-2 GluN), 3.67–4.0 (m, H-2 (GluNAc), H- 3, H-

322 

4, H-5, H-6, H-6’), 4.5–5.0 (m, partly overlapped by water peak, H-1).

323 

TECS

324 

1

325 

3.52 ( m, –CH2 from ethyl group, at N),), 2.68-2.71 (m, H-2 GluN), 3.66–4.0 (m, H-2 (GluNAc), H- 3, H-

326 

4, H-5, H-6, H-6’), 4.5–5.0 (m, partly overlapped by water peak, H-1).

327 

These affirmations agree with those from literature which indicate the positions at which ethylene groups

328 

are bound especially to nitrogen atom (Avadi et al., 2004; Badawy, Rabea, & Taktak, 2014; Pretsch,

329 

Buhlmann, & Badertscher, 2009).

330 

Using the relations (4-6) it was found that the substitution degrees are 0.78 % for MECS, 2.40% for

331 

DECS and 20.73 % for TECS. In DECS and TECS were obtained ECSs mixtures with different

332 

substitution degrees. When alkylation / ethylation occurs at the oxygen atom, the substitution degrees

333 

vary between 6.97 to 19.29 % respectively (Table 4S in Supplementary data).

ip t

H NMR (400 MHz, D2O): δ ppm 1.10-1.13 ( (m, –CH3 from ethyl group at N , 2.00 (s, CH3C=O), 2.83-

us

cr

H NMR (400 MHz, D2O): δ ppm 1.00-1.06 ( (m, –CH3 from ethyl group at N , 2.00 (s, CH3C=O), 3.0-

Ac ce pt e

d

M

an

H NMR (400 MHz, D2O): δ ppm 1.32-1.34 ( (m, –CH3 from ethyl group at N , 2.00 (s, CH3C=O), 3.42-

334 

3.4. XPS results for cotton treated with TECS

335 

Table 1 presents the data which define the quantitative analysis of XPS spectra. Here are indicated the

336 

percentages of the atoms involved in each type of bond and the position at which they absorb. The

337 

presence of TECS on cotton is proved by the following:

338 

1) The increase of the percentage of atoms involved in C-C bond, from 50.8% in untreated cotton to 66.2% in cotton treated with TECS. This can be the result of bonding the radical -CH2- CH3 through

339 

14   

Page 15 of 30

340 

alkylation at the nitrogen atom from the position 2 (at the same time, a slight decrease of the number of

341 

C-C bonds from each glucopyrazine unit occurs as the result of destructuring; it seems that alkylation

342 

prevails against destructuring); 2) Non-existence of C-N bond in the untreated cotton and the appearance of C-N groups in the cotton

344 

treated with TECS. This is confirmed by the presence of the peak at 286.4 eV in which 26.1% N1s

345 

atoms are involved; 3) The increase of the percentage of atoms of O1s type involved in C-O-C bond (from 7.6% (atoms) in untreated cotton to 39.1% (at) in cotton treated with TECS);

347 

4) In the cotton treated with TECS, the N1s atom is implied in the following bonds: a)N-C from 398.72

us

348 

cr

346 

ip t

343 

eV: b) N-C=O, when its presence is remarked at 399.65 eV (Gervais, Douy, Gallot, & Erre,1986);

350 

c)+NH3-C group at 400.51 eV (from dissolving chitosan); d) +N-C, and the presence of this bond

351 

appears at 401.50 eV which confirms the TECS presence on the treated cotton; 5) Presence of clorine (Cl 2p) at 1305.69 eV, as prove of quaternization.

M

352 

an

349 

353 

Table 1. Data of XPS elemental analysis for TECS grafted on cotton fabric.

355 

 

356 

 

Ac ce pt e

d

354 

Sample

Cotton untreated

357  358  359 

Cotton treated with TECS

360  361  362 

Atoms Name Position %At. Conc. C 1s 285.0 74.8

O 1s

533.1

25.2

C 1s

285.0

81.5

O 1s

533.0

16.9

N 1s

398.72

1.3

Cl 2p

1305.69

0.3

363  364 

Bonds Name

Position

C 1s C-C C 1s C-O C 1s C=O/O-C-O C 1s O-C=O O 1s O-C O 1s C-O-C C 1s C-C C 1s C-O/C-N C 1s C=O/N-C=O O 1s O-C O 1s C-O-C N-C N-C=O + NH3-C + N-C Cl

285.0 286.7 288.2 289.3 533.1 531.5 285.0 286.4 288.6 533.0 532.0 398.72 399.65 400.51 401.50 1305.69

%At. Conc. 50.8 37.9 9.7 1.6 92.4 7.6 66.2 26.1 7.7 60.9 39.1 28,92 60.41 8.90 1.77 100

365  366  15   

Page 16 of 30

367 

3.5. SEM analysis

368 

The ECS presence on the cotton yarns can be clearly seen in Figure 3. a) Witness

b) MECS M.F  2500 

c) DECS

cr

ip t

M.F  1200

d) TECS

M.F  2400 

M

an

us

M.F  2500

Fig. 3. SEM images for cotton treated with ECSs.

370 

The visibility of the presence of ECSs on cotton depends on the CS concentration, the molar ratios

371 

CS:AA: NaOH and the degrees of substitution at N and O. Fig. 3 contains an image for witness (Fig.3a)

372 

and only a single variant for MECS, DECS and TECS respectively, grafted on cotton, even though there

373 

are several possibilities to obtain them, as indicates the experimental protocol from Table 2S of

374 

Supplementary data. In Fig. 3 were used the smallest concentrations of CS (0.1 mmols) implied in the

375 

molar ratios CS: AA: NaOH to obtain MECS (Fig.3b) and DECS (Fig.3c), as follows: 0.1: 0.1: 0.1 for

376 

MECS and 0.1: 0.39: 0.39 for DECS. In turn, for TECS 0.39 mmols were used, therefore the highest CS

377 

concentration that can generate TECS during alkylation, and the afferent molar ratios CS: AA: NaOH

378 

were 0.39: 2.1: 2.1. Under this circumstances, taking into account that the solutions used for padding

379 

(based on impregnation and then squeezing) contain mixtures of ECSs with different degrees of

380 

substitution (as indicate the data from Table 4S/Supplementary data), it follows that the grafting

381 

phenomenon is the most visible in Fig. 3d where TECS prevails.

Ac ce pt e

d

369 

16   

Page 17 of 30

3.6. DSC

383 

In Figure 4 one can notice the existence of endothermic peaks within the range 25-150oC, which confirms

384 

the evaporation of the humidity sorbed and bound to cellulose or to ECSs. The size of endothermic peaks

385 

is equivalent to the dehydration heat and offers indications concerning the moisture amount from the

386 

tested samples. The dehydration heat was determined for each tested sample by measuring the area of the

387 

afferent endothermic peak, i.e. by carrying out the normalized integral. The biggest dehydration heat was

388 

recorded for the untreated cotton (-116.5 J/g). In the case of cotton treated with ECSs, the type of ECS

389 

influences the dehydration heats, namely: -115.86 J/g (TECS), -111.45 J/g (DECS), -101.39 J/g (MECS).

390 

One can notice that the quaternized alkyl chitosan (TECS) contains the biggest amount of humidity out of

391 

the ethyl chitosan series, a value which is very close to that of the witness. These affirmations agree with

392 

those from literature, when in the methyl chitosan series, the biggest dehydration heat was recorded in the

393 

case of trimethyl chitosan, TMC (namely at the quaternized alkyl chitosan (Enescu, 2008)).

Ac ce pt e

d

M

an

us

cr

ip t

382 

394  395 

Fig. 4. DSC curves for untreated cotton samples and samples treated with ECSs respectively.

396  397 

3.7. TGA and DTG

398 

Two stages appear at the pyrolysis of the samples tested in this paper: 1) dehydration (25-150oC); 2)

399 

sample degradation (300-380oC). This fact is confirmed by DTG curves from figures 5S from

17   

Page 18 of 30

Supplementary data. In the dehydrating stage, mass losses due to evaporation were recorded at all the

401 

grafted samples; these are rendered evident by the endothermic peaks in the range 25-150oC.

402 

The main characteristics of the sample decomposition stage (300- 380oC) are presented in Table 2. One

403 

can notice that the untreated cotton sample begins to decompose at 327.6oC and the process ends at

404 

380.9oC. This agrees with the values from literature (Ibrahim, El-Amoudy & Shady, 2011), which

405 

indicates the interval 300- 380oC for an untreated cotton fabric. Thermal stability is indicated by the

406 

values of Tonset. By comparing the values of Tonset, one can notice that the sample treated with TECS has

407 

the lowest stability, and after burning it, the biggest residue remains. The data are also confirmed by the

408 

DTG curves presented in Figure 5S from Supplementary data.

409 

Table 2. The main characteristics of degradation stage (collected from DTG and TGA curves).

a

us

T onseta (ºC) T peakb (ºC) T endsetc (ºC) W d (%) W total e (%) R f (%)

Cotton grafted with: MECS DECS TECS 305.80 306.90 294.09 357.43 356.20 339.90 377.40 373.60 367.07 78.20 77.10 74.24 81.67 80.73 78.21 18.33 19.27 21.79

an

411  412  413  414  415  416  417  418  419  420  421  422  423  424 

Untreated cotton 327.60 363.00 380.90 84.90 89.48 10.52

M

Parameters

d

410 

cr

ip t

400 

Tonset is the temperature at which the degradation stage begins; Tpeak is the temperature at which the degradation rate is maximum; c Tendset is the temperature at which degradation stage is finished; d W is the weight loss (expressed in percent) during degradation stage; e ∆Wtotal is the total weight loss during the entire thermal process (25-800°C); f R is the residue (the mass remaining after heating of the samples at 800ºC).

Ac ce pt e

b

425 

3.8. Quantified results of ECSs grafting on cotton

426 

3.8.1. Taking up degree

427 

The taking up degree has been determined for the cases where the molar ratios were: CS: AA: NaOH =

428 

(0.1-2.1): 1.1: 1.1 (Figure 5).

429 

As the CS concentration increases, the taking-up degree increases (under the condition to maintain

430 

constant the concentrations of AA, NaOH and catalyst (Zinc acetate)). Therefore the taking up degree

431 

increases as the degree of substitution through alkylation decreases.

18   

Page 19 of 30

ip t

432 

Fig.5. The values of taking-up degrees in terms of molar ratios CS: AA: NaOH.

cr

433 

us

434 

3.8.2. WRA

436 

ECSs grafted on cotton have determined the modification of the values of wrinkle recovering angles

437 

(WRA). In Fig. 6 one can notice that the WRA values depend on the molar ratios CS: AA: NaOH. The

438 

wet samples have smaller WRA than the dry samples. All the tested samples (both wet and dry) have

439 

recorded WRA values increasing as the concentration of CS used at alkylation increased. The increase of

440 

AA concentration results in the increase of substitution/alkylation degree and in slight variations between

441 

WRA corresponding to samples grafted with TECS, DECS and MECS (especially when the tested

442 

samples were dry).

Ac ce pt e

d

M

an

435 

443  444 

Fig. 6. Values of dry and wet WRA in terms of molar ratios CS: AA: NaOH. 19   

Page 20 of 30

By applying the bifactorial statistic ANOVA method with systematic effects, it was found that all the

446 

levels of the factors A (CS concentrations from 0.1 to 2.1mmols) and B (the 5 concentrations of AA from

447 

0.1 to 2.1 mmols) deny the null hypothesis; this means that the factors A and B have significant

448 

influences on WRA values. In ANOVA with systematic effects- the set of cells consists only of analyzed

449 

cells, no others, and the data grouped according to the two factors determine systematic deviations.

450 

In Tables 3-4 one can notice that the values computed for Fisher- Snedecor functions are higher than the

451 

tabulated values of Fisher- Snedecor functions for a significance level α= 0.01, then the null hypothesis is

452 

denied. By denying the null hypothesis (the null hypothesis supposes that the A and B factors and the AB

453 

interaction do not influence the studied parameter, having values equal to zero), one reaches to a

454 

negation of negation, therefore the factors A and B and the AB interaction do influence the values of wet

455 

and dry WRA.

456 

Table 3. ANOVA components together with the averages of dispersion estimates in the case of dry

457 

WRA.

an

Freedom degrees number

M

Sum of squared

Mean Square

Ac ce pt e

d

Variation source

us

cr

ip t

445 

458  459  460 

F Function calculated

F Function (in table)

Mean value of the estimates S2A, S2B, S2AB S2Amin = 1060.073583 S2Amax = 1060.215334 S2Bmin = 3018.145263 S2Bmax = 3018.287014 S2ABmin = 166.176481 S2ABmax = 166.318231 σmin2 = 0.077019 σmax2 = 0.218770 ……………..

Line (A is CS concentration)

SPL= 5299.0328

νL=4

S2L = 1324.7582

FL= 8883.3632

FL= 4.177

Column (B is AA concentration)

SPC= 15089.391 2

νC=4

S2C = 3772.3478

FC = 25296.039

FC = 4.177

Interaction (AB)

SPLC= 2656.1398

νLC=16

S2LC = 166.0087

FLC= 1113.1963

FLC= 2.813

Inside cells (errors )

SPrez= 3.7282

νrez=25

S2rez = 0.1491

-

-

Global

SPG = 23048.291

νG=49

………….

…………

……

Global relative error = 0.051550 %; **Same legend as in Supplementary data/Table 3S. *

20   

Page 21 of 30

461 

Table 4. ANOVA components together with the averages of dispersion estimates in the case of wet

462 

WRA.

S2C = 9730.6783

FC = 59551.274

SPLC= 495.1128

νLC=16

S2LC = 30.9446

FLC= 189.3791

Inside cells (errors )

SPrez= 4.0850

νrez=25

S2rez = 0.1634

Global

SPG = 44154.2562

νG=49

--------------

SPC= 38922.7132

Interaction (AB)

Global relative error =0.067021 %; **Same legend as in Supplementary data/Table 3S.

F Mean value of Function the estimates (in table) S2A, S2B, S2AB FL= S2Amin = 4.177 946.761629 S2Amax = 946.916945 FC = S2Bmin = 4.177 7784.835229 S2Bmax = 7784.990545 FLC= S2ABmin = 2.813 31.128346 S2ABmax = 31.283663 σmin2 = 0.08439 σmax2 = 0.23970 ----------------------

us

Column (B is (AA) concentration)

-

an

SPL= 4732.3452

M

Line (A is CS concentration)

Mean Square

Freedom degrees number νL=4

ip t

νC=4

Sum of squared

cr

S2L = 1183.0863

F Function calculated FL= 7240.4302

Variation source

--------------

*

467 

The multiple comparison carried out by applying the Scheffe test indicated with a 99% confidence that all

468 

the levels of A and B influences both wet and dry WRA.

469 

By analyzing the data in the Tables 5 and 6 result the following conclusions: 1) the effect exerted by CS

470 

concentration increases with increasing the CS concentration values; 2) the effect caused by AA

471 

concentration increases with increasing values of this agent.; 3) the effects of the A interaction with B

472 

have a positive influence (if they have values >0) or negative (if they have values<0).

Ac ce pt e

d

463  464  465  466 

473  Table 5. Maximal effects of the factors A, B and the interaction of A with B, for dry WRA.

Effects of factor: A B AB A B

B2 1

0.1

0.39

1.1

1.806

-16.1248 12.0152 -3.2972 -6.1948 12.0152

-16.1248 12.9372 -4.1192 -6.1948 12.9372

2.1

A

0.1 0.39

-16.1248 -32.6248 -14.5572 -6.19480 -32.6248

-16.1248 1.82520 10.9928 -6.1948 1.82520

-16.1248 16.9852 11.8328 -6.1948 16.9852

21   

Page 22 of 30

AB A B AB A B AB A B AB

1.806 2.1

-9.2372 2.39520 1.82520 2.37280 9.07720 1.82520 -1.5092 11.9852 1.82520 -1.7672

8.77280 2.39520 12.0152 1.28280 9.07720 12.0152 -3.4992 11.9852 12.0152 -2.4072

1.30080 2.39520 12.9372 10.7108 9.07720 12.9372 -2.5112 11.9852 12.9372 -4.5292

1

2.60280 2.39520 16.9850 -5.3372 9.07720 16.9852 -5.6692 11.9852 16.9852 -2.5772

ip t

A is CS concentration (mmols) B is AA (alkylation agent) concentration (mmols) 3 AB is the interaction of A with B (i.e. CS concentration with AA concentration).

474  475  476  477 

cr

2

B2

1.1 1.806

0.39

-13.565 -33.585 -1.0676 -6.5957 -33.585 5.4124 0.3243 -33.585 2.4924 6.9943 -33.585 -0.7276 14.0343 -33.585 -5.2176

-13.565 -25.475 -0.7276 -6.5957 25.475 -3.6976 0.3243 25.475 1.3824 6.9943 25.475 2.2624 14.0343 25.475 1.6724

1.1

1

2.1

-13.565 5.6143 2.5324 -6.5957 5.6143 2.8624 0.3243 5.6143 0.7424 6.9943 5.6143 -0.8276 14.0343 5.6143 -4.4176

an

0.39

0.1

M

A1 0.1

Ac ce pt e

Effects of factor: A B AB A B AB A B AB A B AB A B AB

us

Table 6. Maximal effects of the factors A, B and the interaction of A with B, for wet WRA.

d

478 

1.1

-2.58720 2.395200 -32.6248 -8.17720 9.077200 -32.6248 14.04080 11.98520 -32.6248 12.13280

1.806

-13.565 10.1243 4.2224 -6.5957 10.1243 0.3524 0.3243 10.1243 -3.7676 6.9943 10.1243 -2.5376 14.0343 10.1243 2.6224

2.1 -13.565 44.5143 -4.0676 -6.5957 44.5143 -4.0376 0.3243 44.5143 0.0424 6.9943 44.5143 2.7224 14.0343 44.5143 6.2324

A is CS concentration (mmols) B is AA (ethyl chloride) concentration (mmols) 3 AB is the interaction of A with B (i.e. CS concentration with AA concentration)

479  480  481  482  483 

2

484 

3.8.3. Tensile strength on weft and warp

485 

Figure 7 shows the tensile strengths of the tested samples; one can notice that there are few situations

486 

when tensile strength are smaller than that of the witness, both on warp and weft; most of the samples

487 

have tensile strengths bigger than the witness because CS converted in ECS acts as a binder for the fibers

488 

in the cotton yarn determining the increase of the breaking force.

22   

Page 23 of 30

ip t cr us

489 

Fig. 7. Tensile strength values recorded on both directions (warp and weft) of the tested samples (coded

491 

according to molar ratios CS: AA: NaOH).

an

490 

492 

3.8.4. Tinctorial capacity of cotton functionalized with ECSs

494 

ECSs grafting on cotton resulted in the modification of cotton tinctorial capacity. On the other hand,

495 

dyeing cotton under severe conditions (100oC, 60 min.) proves once more a good durability of the effects

496 

of ECSs alkylation and grafting respectively on cotton (Fig. 8).

497 

The explanation of dyeing behavior is based on the nature of interactions appeared between amino

498 

groups (with different substitution degrees) and ionic groups from dyes; the pH of dyebath can generate

499 

the positivation of all amino groups (substituted through alkylation or not), which leads to the attraction

500 

of acid dye (because in the C.I. Acid Red 88 dye there are anionic groups type SO-3 ) and respectively to

501 

the rejection of cationic dye (since in the CI Basic Blue dye there are groups with positive charges).

502 

Significant influences are also exerted by the ECS concentration (approximated by the concentration of

503 

CS used to obtain ECS) and the steric effects manifested by the ethylene groups appeared after

504 

alkylation. Explanations justifying in details the color strength (K/S) values (at two different pH values

505 

(pH= 5 and pH=7 respectively) from figure 8 are presented in Supplementary data S6.

Ac ce pt e

d

M

493 

506  23   

Page 24 of 30

CS concentration varies

CS concentration is constant = 1.1 mmols b

c)

us

cr

ip t

a)

Ac ce pt e

d

M

an

d

507 

Fig. 8. Variation of color strength (K/S) after dyeing with non-specific dyes (C.I. Acid Red 88 (a, b) and

508 

C.I. Basic Blue 9 (c, d) in acid medium (pH=5) and neutral medium (pH=7), when CS concentration

509 

(used in the ECS synthesis) varies from 0.1 to 2.1 mmols.

510  511 

3.8.5. Antimicrobial capacity

512 

Microorganisms used for testing consisted in a Gram positive coccus (Micrococcus Luteus ATCC 934)

513 

and a Gram negative bacillus (Escherichia coli DSMZ 498) from the collection of Biology Laboratory of

514 

DTNW Krefeld, Germany.

24   

Page 25 of 30

One can notice in Table 7 and Fig. 6S (from Supplementary data) that the witness sample (M), which

516 

consists in untreated fabric, does not possess any antimicrobial effect against the both microorganisms

517 

used for test. At the same time, the samples 1 and 2 have no visible inhibition zone either for

518 

Micrococcus Luteus or for Escherichia coli. In the case of the samples 3 and 4 a diffuse inhibition zone

519 

appears (less sharp) for Micrococcus Luteus. The diffuse aspect of the inhibition zone can be interpreted

520 

as a retardation of coccus development, without producing a complete elimination, as in the case of

521 

samples 5, 6 and 7.

522 

In the case of activity against Escherichia coli, the samples 3 and 4 are inactive, showing no inhibition

523 

zone; in exchange, the samples 5, 6 and 7 are strongly antibacterial, the bacillus being completely

524 

destroyed in the inhibition zone.

525 

Table 7. Testing textile antimicrobial action against a Gram positive coccus (Micrococcus Lutes) and a

526 

Gram negative bacillus (Escherichia Coli).

529 

an

Sample number

Sample Type

0 1 2 3 4 5 6 7

Witness Cotton treated with CS Cotton treated with MECS Cotton treated with DECS Cotton treated with TECS Cotton treated with MECS and AgNO3 Cotton treated with DECS and AgNO3 Cotton treated with TECS and AgNO3

Ac ce pt e

530 

us

cr

528 

M

 

d

527 

ip t

515 

531  532  533 

Diameter of inhibition zone (mm) Micrococcus Escherichia Luteus Coli 0 0 0 0 0 0 20 0 16 0 18 16 19 17 19 17

534 

4. Conclusions

535 

The synthesis of ECSs is easy to perform, and their application on a cotton fabric is made through the

536 

classic pad-dry-cure procedure. An advantage of the proposed method consists in avoiding fabric

537 

yellowing, because a soft catalyst is used in the condensation stage (at 160oC it develops acetic acid

538 

which has no destructive effects on the fabric, as it happens in other procedures which used as catalyst

539 

an acid or a metallic salt which can generate an strong acid).

25   

Page 26 of 30

540 

Through ECSs grafting (with various degrees of substitution to N and O) on cotton, the approach of

541 

neighboring macromolecular chains that could generate an external network was detained, thus avoiding

542 

the formation of a rigid architectonic structure, which would have resulted in significant loss of tensile

543 

strength. Through grafting, the macromolecular chains flexibility was maintained and the brittle form was

544 

avoided, thus influencing positively the values of both WRA and tensile strength. Using the ANOVA method one could determine the effects exerted by CS and AA concentrations on the

546 

wrinkle-proofing degrees expressed by the WRA values and the tensile strengths (on the two directions:

547 

warp and weft). Each ECS can be considered as a good wrinkle-proofing agent, since it makes possible

548 

to obtain higher values for both WRA and tensile strength, as compared to the witness. Moreover, the

549 

cotton treated with TECS (i.e. CS with the highest substitution degree) manifests a wettability similar to

550 

that of the witness (fact confirmed by DSC, TGA/DTG) analyses) and a good antimicrobial effect.

an

us

cr

ip t

545 

551 

Appendix A. Supplementary data

553 

1) Materials; 2) ANOVA method; 3) FTIR; 4) Substitution degrees; 5) Termogravimetric results;

554 

6) Modification of tinctorial capacity; 7) Antimicrobial capacity.

555 

Supplementary data associated with this article can be found in the online version.

d

Ac ce pt e

556  557 

M

552 

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