Optimum conditions for polyamide fabric modification by ... - NOPR

Indian Journal of Fibre & Textile Research Vol. 39, March 2014, pp. 65-71

Optimum conditions for polyamide fabric modification by protease enzyme produced by Bacillus sp Samiha M Abo El-Ola1, Maysa E Moharam2, Maha M Eladwi3 & Magda A El-Bendarya,2 1

Proteinic and Synthetic Fibre Department, 2Microbial Chemistry Department, National Research Center, Dokki, Giza, Egypt 3 Women’s College, Ain Shams University, Helliopolis, Cairo, Egypt Received 19 October 2012; revised received and accepted 22 February 2013

The optimum conditions for surface modification of polyamide 6 (PA) by protease enzyme produced by Bacillus isolate 16P have been studied. These conditions are found to be 0.05 mg/mL enzyme concentration, 0.5 h treatment time, room temperature (30ºC) and pH 8 under shaking conditions. The effect of hydrolytic activity of enzymatic process on the weight loss of PA fabric after enzymatic treatment shows negligible difference. Printing both untreated and treated PA fabrics with transfer printing shows high leveling properties as regard to ∆E values which are considered as acceptable values of color differences. PA treated with different sources of protease enzyme under optimum conditions shows good physical properties. SEM of the surface of protease treated PA samples shows etches and voids compared to smooth surface of untreated PA fabric. Keywords: Bacillus sp, Physical properties, Polyamide modification, Protease enzyme

1 Introduction The most common synthetic polyamides (PA) are polyamide 6 (Nylon 6) and polyamide 6.6 (Nylon 6.6). The annual world production of polyamide is 4 million tonns1. Nylon filaments are used as yarns for textile, industrial and carpet applications and a growing demand has been reported especially for industrial and textile applications2. Nylon based textiles show the great disadvantage of being uncomfortable to wear and difficult to finish due to their hydrophobicity. Therefore, enhancement of the hydrophilicity of nylon is a key requirement for many applications and can be achieved by chemical, physical and enzymatic methods. Chemical modification requires harsh reaction due to which strength properties of polymer get affected. Physical methods include plasma treatment, UV irradiation, corona discharge and flame treatment. These methods are very difficult to optimize, require complicated machinery and are difficult to repeat. Enzyme treatments can be chosen as green alternatives for polymer surface modification as they offer many advantages over chemical and physical methods such as they are very specific, act under moderate reaction _________ a Corresponding author. E-mail: [email protected]

conditions which lead to less or negligible damage of the strength properties of synthetic polymers, cost effective, ecofriendly, the scale up is possible and gives reproducible results3,4. It is to be expected that, within 5-10 years, wet textile production processing will be shifted substantially towards sustainable processes, because of increasing governmental and environmental restrictions and the decreasing availability of fresh water. Biocatalysts have proven to be a flexible and reliable tool in wet textile processing and a promising technology for fulfilling expected future requirements5. Treatment of PA with hydrolases shows potential for targeted surface modification without changes of bulk properties6. Enzymes that can hydrolyze PA are proteases, cutinases and polyamidases7. These enzymes have been successfully used for surface targeted hydrolysis of PA leading to hydrophilization6,8,9. Surface hydrolysis of PA leads to increasing of polar groups (free amino and carboxylate end groups) on the surface. The enzymatic hydrolysis of surface moieties of PA has been demonstrated to be a powerful mild strategy for improving hydrophilicity and activating materials for further processing, which is the key requirement for many applications including painting,

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INDIAN J. FIBRE TEXT. RES., MARCH 2014

inking, antifogging, textile production, electronics and applications in the biomedical field7. The mechanism of action of protease, cutinase and amidase on PA is shown below4 :

Many proteases such as Protex Gentle L, Protex 40 L, Protex multiplus L, and Protex 50 FP were used to investigate changes in the nylon 6,6 polymer. Protease treatment of Nylon 6,6 showed significant decrease in thermal degradation temperature whereas reactive and acid dyes showed higher dye bath exhaustion on the protease-treated polymer10. Enzymatic surface modification of synthetic fibres can improve the present use of these fibres. A lot of experimental research work has been carried out on the surface modification of PA, PET and PAN but their large scale industrial application is yet to be undertaken and optimized4. The authors have previously screened 19 protease enzymes produced by isolated bacilli from Egypt for specific surface modification of PA fabric11. One of these enzymes showed good and promising results in modifiying PA surface. The purpose of this study is to establish the optimal conditions for PA surface modification by protease enzyme such as enzyme concentration, treatment temperature and treatment time by measuring the wettability of PA fabrics, staining by basic dyes and investigating the changes induced by the enzyme under study as well as comparing the efficiency of the enzyme with the standard protease. 2 Materials and Methods 2.1 Materials

In this study, protease enzyme produced by Bacillus isolate 16P (identified as Bacillus pumilus) was tested and standard protease enzyme was purchased from Sigma. The bacterial organism was isolated from rhizosphere of sugarcane roots in Egypt

and showed promising results in modification of PA fabrics11. Polyamide 6 (PA) knitted fabrics was used with number of wales 44, number of courses 52 and fabric weight 103 g/m2. It was purchased from Misr Rayon Co., Kafr El Dawar, Egypt. Sodium dihydrogen phosphate, disodium hydrogen phosphate, and non-ionic detergent Hostpal CVL-EL were produced by Clarient. Glacial acetic acid was purchased from El-Naser Pharmaceutical Chemical Company. Basic dyes C.I Basic Violet 14 (λmax 460) was supplied by European Colour Plc, Stockport, England. Methylene Blue (λmax 650) was purchased from Merck. Disperse dyes used in transfer printing paper were C.I. Disperse Red 60, C.I. Disperse Red 50 and C.I Disperse Blue 56. These dyes were supplied from Alwan Misr Company. The dye of this paper is a mixture of two Red Disperse dye. The thickener is fast print 54. All other chemicals were of analytical grade. 2.2 Methods

2.2.1 Production of Protease Enzyme

Protease enzyme of Bacillus isolate 16P was produced under solid state fermentation using 6% wheat germ meal with 10% moisture for 7 days. The enzyme was extracted by water and centrifuged at 6000 rpm. The filtrate was used as enzyme source. 2.2.2 Test Methods Enzymatic Treatments of Textile

Enzymatic treatments of textile were according to O'Neil et al12. The fabrics were washed with 10 g/l hostapal at 65°C for 1 h, followed by rinsing several times with tap water. Then the fabrics were washed in aqueous solution containing 2 g/L sodium carbonate for 1 h at 65°C, followed by rinsing several times with tap water. Finally, the fabrics were gently squeezed and air dried. One gram of the fabric was incubated in a glass vessel containing a solution of 0.05 M sodium phosphate buffer (pH 8) at liquor ratio 50:1, under continuous orbital shaking at 150 rpm. Surface modification of PA fabrics was studied under different conditions of enzyme concentration (0.025-0.2 mg/mL), treatment time (0.5-24 h) and treatment temperature (30-50°C). After enzymatic treatments, all samples were washed several times with tap water and then with 2 g/L sodium carbonate for 1 h at 65°C followed by washing with

EL-OLA et al: OPTIMUM CONDITIONS FOR POLYAMIDE FABRIC MODIFICATION BY PROTEASE

distilled water for 1 h at 65°C. Finally, the samples were washed with running tap water for 5 min and allowed to dry in open air. Staining the Fabrics

The staining was carried out below the glass temperature (Tg) of PA at 50°C for 90 min as described by O'Neil et al12. After enzymatic treatments, the fabric samples were stained together in the same sealed glass vessel (500 mL) on shaking water bath at proper pH (4.5 in case of Basic Violet 14 and 9.5 for Methylene Blue). The dyeing was performed with 0.5% shade (owf), liquor ratio 50:1 and at 200 rpm agitation. After dyeing process, samples were washed with aqueous solution containing 2 g/L non ionic detergent at 60°C for 1 h, then washed several times with tap water and dried in oven at 50°C for 6 h. Wettability Test

In order to obtain the degree of wettability (hydrophilicity) of PA fabric samples, a water droplet test was applied according to AATCC standard test method 27-1994. The wetting time was determined by placing a drop of distilled water on the stretched fabric sample (10 cm × 10 cm) from a burette held 1 cm from the fabric surface. The time of disappearance of the water-mirror on the surface was measured as the wetting time.

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Relative K/S (%) = [(K/S treated - K/Scontrol)/(K/S) control] × 100 Leveling Properties

CIELAB color system uses the L*, a*, b* and h, where L* stands for lightness and ranges from 0 for black to 100 for white. A color (hue) can also be specified either in terms of the coordinates a* or b* (for grey and dull colors). Leveling of printed fabrics was assessed by measuring the color differences, calculated from the CIELAB coordinates, at five separate points within each sample and the average color difference (∆E) between these points was determined according to Parvinzadeh et al.10 and Kothari and Kausik14. Printing of PA Fabrics

Enzymatic treated fabric samples (20 × 20 cm) were subjected to transfer printing using manual heat transfer press with a (40 × 40 cm) flat bed press at 190ºC for 30 s. SEM Study

To analyze the surface morphology of control and enzymatic treated fabric, the scanning electron microscopic pictures were obtained via scanning electron microscope model JEOL JXA-84OH Electron Prope Microanalyzer Japan operating at 19 kV. A thin coating (~10 nm) of gold was deposited onto the samples prior to examination.

Weight Loss

Weight loss, was evaluated according to the following equation:

Bursting Strength

where W1 and W2 are the weights before and after enzymatic treatment respectively.

Bursting strengths of protease treated PA and control fabric samples were measured according to ASTM D3786-87 by Mullen tester made in USA. Bursting strength results are the arithmetic means of three tests per sample. All bursting strength results are expressed as kg/M2.

Color Measurement

3 Results and Discussion

The K⁄S percentage increments after the enzymatic treatment and staining of the fabric samples were measured in order to detect differences in reactive groups formed on the surface according to Matama et al.13. The colourimetric data K⁄S of the dyed samples was collected using Mini Scan XE integrated with Hunter lab universal software at maximum absorption, as an average of three readings. The relative K⁄S of enzymatic treated samples was calculated as follow:

3.1 Effect of Protease Concentration

Weight loss (%) = [(W1-W2)/W1] × 100

The effect of protease enzyme concentrations on hydrophilicity, weight loss and the relative color strength of treated PA fabric is shown in Table 1. The drop absorption time in buffer treated PA fabric is found to be 57s. The optimum concentration of protease enzyme for enhancing the surface hydrophilicity is 0.05 mg/mL. The decrease in water drop absorption time is about 49% and 47% when the fabric is treated with 0.05 mg/mL protease


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protease treated PA fabric varies according to the basic dye used which may be attributed to the size of dye molecule and the pore size created on the fabric surface as a result of enzymatic hydrolysis. The enzymatic treatment increases the adhesion of cationic compounds probably by creating carboxyl groups on the surface of the treated substrates. It is reported that hydrolytic enzymes can act on the polymer surface resulting in an increased amount of free amino and carboxyl groups giving the PA desired properties17.

enzyme of Bacillus isolates 16p and the standard one respectively. The increment of enzyme concentration does not enhance the hydrophilicity. It is reported that there is a rapid initial decrease in the drop absorption time of PA 6 with increasing amidase activity which levels off to a plateau value once a certain amidase activity is reached15. In addition the excess enzyme could cause aggregation between enzyme molecules, leading to partially enzyme inactivation depending on the type of aggregation formed14. Weight loss per cent as a result of enzymatic treatment is negligible as shown in Table 1. It is found that Bacillus isolate 16P and standard protease enzymes have the same behavior and almost the same activity. In agreement with these results, it is reported that the effect of hydrolytic activity of enzymatic process on the weight loss after treatment does not show any significant differences and is independent on the source of enzymes16. A significant increase in dye binding is observed for the fabrics treated with 0.05 mg/mL of Bacillus isolate 16 P and standard protease enzymes as shown in Table 1. It is observed that the staining degree of

3.2 Effect of Treatment Time

Effect of treatment time is investigated at 0.05 mg/mL enzyme concentration and at room temperature (about 30°C). As shown in Table 2, the wettability time of protease treated PA fabric is decreased about 45% at 30 min treatment time in comparison with that of untreated fabric. This result is confirmed by increasing the relative colour strength of dyed PA after treatment for 30 min. Prolonging the reaction time more than 30 min does not improve the wettability and relative color strength.

Table 1—Effect of protease enzyme concentration on wettability and weight loss of PA fabric and on the relative colour strength K/S of the dyed fabric [Treatment time 30 min., room temperature, material: liquor ratio 1:50] Enzyme conc. mg/mL

0.025 0.05 0.1 0.15 0.2

Enzyme source

Control

Bacillus isolate16P Wettability

time, s

Weight loss, %

44 29 32 37 37

1.9 1.8 1.76 1.76 1.8

Standard

Relative K/S % Methylene C.I. Basic Blue Violet 14 12.34 16.1 5.9 3.9 2.81

11.4 14.6 5.5 3.05 1.13

Wettability

time, s

Weight loss, %

45 30 33 37 39

1.5 1.9 2.1 2.1 2.05

Relative K/S % Methylene C.I. Basic Blue Violet 14 11.95 15.8 8.46 6.1 5.9

9.5 13.6 8.7 6.6 4.5

Wettability

Weight

time, s

loss %

57

1.4

Table 2—Effect of enzymatic treatment time on wettability and weight loss of PA fabric and on the relative colour strength K/S of dyed fabrics Treatment

Enzyme source

time, h

Bacillus isolate 16 P Wettability

Weight

time, s

loss, %

Relative K/S % Methylene

Blue 0.5 1 2 3 24

29 30 33 38 40

1.76 1.4 1.6 1.48 1.5

16.1 10.1 6.8 8.78 2.81

Control Standard

C.I. Basic Violet 14

14.6 6.9 4.6 3.05 1.13

Wettability

Weight

time, s

loss, %

30 33 32 33 38

1.85 1.57 1.5 1.4 1.57

Relative K/S % Methylene

Blue


15.6 10.3 7.7 7.1 5.9

13.6 7 4.8 3.6 3.3

Wettability

Weight

time, s

loss %

54 55 56 56 57

1.4 1.14 0.98 0.9 1.2


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Table 3—Effect of enzymatic treatment temperature on wettability and weight loss of PA fabric on the relative colour strength K/S of dyed fabrics Treatment temperature o C

30 35 40 50

Enzyme source

Control

Bacillus isolate16P

Standard

Wettability

Weight

time, s

loss %

Methylene

Blue


1.76 1.56 1.35 1.46

16.1 13.57 8.73 6.3

14.6 9.89 5.1 3.2

30 32 41 45

Relative K/S %

Wettability

Weight

time, s

loss %

Methylene

Blue


1.85 1.53 1.71 1.51

15.6 12.06 8.4 5.98

13.6 10.2 6.3 3.3

The enzymatic surface modification of polymers requires prolonged incubation due to which the desired enzyme gets denatured and inactivated due to their short half-life4. Therefore, enzymes with short incubation period and better stability are required for surface catalysis. It is concluded that the maximum increase in surface hydrolysis of PA after treatment with polyamidase is reached after 3 min of incubation with no further changes over a period of 60 min of incubation6. 3.3 Effect of Enzymatic Treatment Temperature

It is known that the temperature is very important to get the maximum enzyme activity4. The hydrophilicity and the staining with basic dyes are the maximum at 30ºC (Table 3) since protease actively hydrolyzes the amide linkages in polyamide fabrics at this critical conditions and the number of carboxylic groups increases, leading to increasing the hydrophilicity as well as the relative color strength. Increasing the treatment temperature more than 30°C decreases the hydrophilicity and the relative colour strength of enzymatic treated fabrics. 3.4 Calorimetric Measurement

The L* a* b* c*, H and ∆E values of printed protease treated PA fabrics are given in Table 4. The color values were evaluated in CIELAB color space, the three axes, namely L*, a* and b*. The results reveal that the lightness L* values decrease for samples treated with protease enzyme of B. sphaericus 16P and the standard one and printed with Disperse Red dye. Decrease in L* values could be explained on the basis that more disperse dye molecules penetrate into the enzymatic treated samples as more voids are formed on the fibres surface and so it is more open for dye molecules penetration. In the same time it can be seen that there is no considerable change in the color

30 31 32 33

Relative K/S %

Wettability

Weight

time, s

loss %

57 55 56 58

1.4 1.1 1.14 1.03

Table 4—Color co-ordinate of printed protease treated PA fabrics with disperse dye Sample treated with

Color co-ordinate L*

Untreated (blank) Control Bacillus isolate 16P protease Standard protease

a*

b*

C*

H

∆E

36.2

48.79

9.1

46.62

10.5

0.63

36.17 34.6

48.78 47.03

9.07 10.71

46.6 47.9

10.53 10.71

0.87 1.1

33.89

46.61

11.61

47.74

13.46

1.0

co-ordinates of control samples. Increase in C* values contributes to increase in brightness of the samples which is an important factor in the textile products. This could be considered as an advantage of protease treatment to improve the quality of PA fabric. The data show high leveling properties as regard to ∆E values which are considered as acceptable values of color differences. 3.5 Scanning Electron Microscopy

The fibre surface is analyzed in order to follow the surface changes and physical properties of the fibres. As shown in Fig. 1, the control sample (Fig. 1a) has a relatively smooth microscopic appearance. However, the surface micrographs of protease treated PA (Fig. 1b) show specific characteristics. The surface of PA samples treated with protease enzyme under optimum conditions shows etched surface. The surface of PA fibre is partially peeled off and small fragments remain on the surface. 3.6 Physical Properties

The enzyme treatment causes perceptible changes on the fibre surface. These changes can be positive (hydrophilic properties) and partly negative


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Fig. 1—SEM images of PA fabric after treatment with buffer solution (control) (a), protease enzyme of Bacillus isolate 16P (b), and standard enzyme (c) Table 5—Effect of protease enzyme treatment on some properties of PA fabrics Sample treated with

Untreated (blank) Control buffer Protease of Bacillus isolate 16P Standard protease

Bursting strength kg/cm2

Air permeability cm3/cm2

Thickness mm

9.33 8.8 8.8

282 280 302

0.35 0.35 0.34

8.8

291

0.35

(mechanical properties). In this study, no significant differences in bursting strength are detected among the enzymatic treated fabrics as long as the thickness of the samples is not changed as shown in Table 5. SEM micrographs (Fig. 1) show significant etching on the fabric surface which could lead to a reduction in the fabric bursting strength. However, the removal of a thin surface layer could reduce the thickness of the fabric only to some extent which would not make detectable reduction of the fibre strength. In agreement with these results, it is reported that the treatment of PA with hydrolases shows potential for targeted surface modification without changes in the bulk properties6,16.

and 30°C treatment temperatures. Under the optimum conditions, protease enzymes of different sources significantly improve the hydrophilicity and cationic dye ability of protease treated PA fabrics. Protease treated fabrics under optimum conditions show good physical and mechanical properties such as negligible weight loss, and good strength. The surface of protease modified PA fabric has unique characteristics as shown by SEM. Finally, there is no difference in the efficiency of protease enzymes produced by isolated bacteria under economic condition as compared to the standard one. References 1

2

3

4

4 Conclusion In view of obtained results, the use of protease enzyme for PA fabric modification can be considered as an effective ecological and energy saving process for changing fibre surface structure leading to the elimination of undesired properties of PA fibre without affecting its strength. The optimum conditions for modification of PA fabric by protease enzymes (produced and standard) are found to be 0.05 mg/mL enzyme concentration, 30 min reaction time,

5

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13 Matama T, Vaz F, Guebitz G M & Cavaco-Paulo A, Biotechnol J, 7 (2006) 842. 14 Kothari V K & Kausik B, J Eng Fibres Fabrics, 5 (2010) 22. 15 Almansa E, Heumann S, Eberl K F, Cavaco-Paulo A & Guebitz G M, Biocatalysis Biotransformation, 26 (2008) 371. 16 Behera B K, Mishra R & Naku S, Indian J Fibre Text Res, 33 (2008) 132. 17 Fischer-Colbrie G, Heumann S & Guebitz G, Enzymes for polymer surface modifications, in Modified Fibres with Medical and Specialty Applications, edited by JV Edwards (Springer, Netherlands), 2006, 181.