Appearance and color changings of the treated specimens
Reflectance curves were plotted to investigate the reflectance properties of the specimens. The involvement of the laccase enzyme in the discoloration process leads to a significant improvement in the reflectance of the specimens, as shown in Figure 4 (Reflectance curves for printed specimens). This phenomenon was enhanced for the specimens treated with both enzymes and sodium hydrosulfite. This result shows that the use of laccase enzymes and sodium hydrosulfite in combination with cellulase enzymes has a synergistic effect on the reflectance of the specimens. The washed specimens gave the same result (Figure 5). Comparing Figure 4 and Figure 5, it is clear that the printing method gives betterresults in terms of reflectance of the samples.
To gain a better understanding of the color changes of the treated specimens, color coordinates in CIELAB color space were obtained (Table 3). The results presented in Figure 6 and Table 3 show that the improvement in the lightness of the specimens ranged from 0.40 to 27.42. As shown in Figure 6, the lightness of the specimen treated with laccase enzyme was greater than that of the specimen treated with sodium hydrosulfite for both methods. The simultaneous use of laccase enzyme and sodium hydrosulfite resulted in a significant synergistic increase in the lightness of the specimens treated by both methods. The addition of cellulase enzyme enhanced the previously described synergistic effect, as shown in Table 3.
It can be concluded that the combination of laccase enzyme and sodium hydrosulfite is successful in discoloration, and the addition of cellulase enzyme increases the lightness (about 1 for the washed specimen and about 3 for the printed specimen). It should be noted that in previous experiments, the application of cellulase enzyme to denim without other chemicals resulted in a decrease in lightness . The addition of the cellulase enzyme to the chemicals used to discolor the jeans had no effect in the present study. It is believed that the mechanism of action of the chemicals is the probable cause of the synergistic effect of the combination of the two enzymes and sodium hydrosulfite. The indigo dye molecules were released by surface degradation of cellulose fibers by the cellulase enzyme. The laccase enzyme, on the other hand, destroyed the indigo dye molecules, and sodium hydrosulfite reduced the soluble dye molecules. It can be assumed that sodium hydrosulfite has a synergistic effect in the presence of the two enzymes (Figure 7).
As mentioned earlier, the printing process results in better discoloration performance, which could be due to the printing process. The chemicals act on the outer surface of the fabrics in this process, and it is likely that the printing process has a higher quality of enzyme reaction than the washing process.
In this study, the increase in lightness is achieved by preparing the specimens once. However, in previous studies, the increase in lightness was 21 after treating specimens four times . Therefore, this is one of the most remarkable achievements of the research.
Figure 8 shows optical images of the specimens. The accuracy of the above improvement in lightness is evident in this figure. In addition, the discoloration process changed the hues of the specimens (Table 3), as shown in the a*b* diagram (Figure 9).
As shown, the transition from redness to greenness was important, and the greenness of the treated specimens has increased. The blueness also improved in all treated specimens. In fact, the improvement in blueness appears to have been noticeable in the treated specimens (Figure 8). Finally, color differences (ΔE*) in CIELAB color-space were determined to investigate the color changes of the treated specimens compared to the untreated specimens (Figure 10).
As can be seen from Figure 10, the color differences were greater than three (ΔE*>3) in all specimens, which means that the color differences were evident in all specimens. The printed specimen with two enzymes and sodium hydrosulfite has the largest color difference. It should be noted that the aforementioned specimen showed the greatest improvement in terms of lightness. Moreover, the greatest influence on the color differences was associated with the improved lightness and then with the blueness of the treated specimens. As already mentioned, the printed specimens gave better results. It may be mentioned that in the printing process, a form of immobilization occurred, and therefore the action of the enzymes was much better than in the washing method. Since enzyme treatment is usually limited to the surface due to the size of the molecules, the printing method provided better conditions for enzymatic treatment of the specimens, resulting in highly efficient discoloration.
Surface Morphology Evaluation
The SEM images of the untreated and treated specimens are shown in Figure 11. Since there was no chemical degradation by laccase enzyme and sodium hydrosulfite on the surface of cellulose fibers during the process, the images of SEM were madeonly from the specimens that had been decolorized with the cellulase enzyme. As can be seen, the application of the cellulase enzyme to the cellulose fibers of the denim is obviously results in degradation. It can be concluded that the anchor fibers were formed due to the surface destruction of the fibers by the cellulase enzyme . The surface destruction of the cellulose fibers resulted in a kind of fibrillation on the fiber surface, which can be clearly seen in the images of the specimens treated by both printing and washing methods. It appears that the degradation is more severe in the specimens treated with cellulase enzyme in the printing method than in the washing method, which can be seenwhen comparing Figure 11-b and Figure 11-d with Figure 11-c and Figure 11-e. It should be noted that some of the light spots on the longitudinal surface of the fibers in the printed specimens (Figure 11-b and Figure 11-d) may be the thickening agent remaining after the final treatment. Enzymes are large molecules whose treatment is mainly limited to the surface, and in the printing method, due to the placement of the chemicals in the presence of the printing paste, better conditions are provided to achieve higher efficiency in treating the surface of the specimens. It can be said that a kind of immobilization takes place in the printing process. Previous researches have shown that immobilization of enzymes, including their mutual presence, increases their efficiency [27, 55, 56]. When comparing the degree of degradation obtained by the two methods, it should be noted that according to the studies of Montazer and Sadighi, the presence of sodium hydrosulfite reduces oxidation by cellulase enzyme due to its reducing effect . Therefore, as expected, the degradation of the specimens treated with a combination of the two enzymes with sodium hydrosulfite was lower than the specimens treated with a combination of laccase and cellulase enzymes. This is evident when comparing Figure 11-b and Figure 11-c with Figure 11-d and Figure 11-e.
Moisture Content Evaluation
In order to investigate the effects of material type and the method of discoloration on the moisture content of the denim fabrics, their moisture content was determined and compared with that of the untreated specimen. As Table 4 shows, due to the lack of chemical interaction of cellulosic fibers with sodium hydrosulfite and laccase enzyme, there are no significant changes in the moisture content of the specimens treated either individually or in combination with these materials. There was a significan increase in moisture content of the samples treated with the combination of the two enzymes and the two enzymes with sodium hydrosulfite. This is due to the degradation of cellulose fibers by the cellulase enzyme. It can be said that hydroxyl groups were formed at both ends of the chain due to the destruction of the polymer chain of the fibers. Hydroxyl groups are hydrophilic functional groups and can help in moisture absorption, increasing the moisture content. The increased moisture absorption due to cellulase treatment of cellulose fabric has been observed in previous studies [41, 42]. Compared to a study by Ebrahimi et al., the present research revealed that the application of advanced oxidation process (AOP) did not result in any significant change in the water absorption capacity and water retention properties of the denim fabric . The specimens treated with the combination of cellulase-laccase enzymes and also the combination of the two enzymes with sodium hydrosulfite by washing method shows a slight increase in moisture content as compared to the printing method as compared to the untreated specimen. This may be due to the nature of the method and it can be said that when the samples are treated by the printing method, the enzymes reach the surface of the fabrics more easily and the surface of the fibers is degraded more by the cellulase enzyme. The specimens treated by both printing and washing methods with the combination of laccase-cellulase enzymes and also with the combination of the two enzymes with sodium hydrosulfite in both printing and washing methods, it is also observed that the specimens treated with the combination of enzymes without sodium hydrosulfite absorb more moisture than the other similar specimens. Due to its reducing effect, sodium hydrosulfite can prevent the oxidation of cellulose, which was also mentioned in the study conducted by Montazer and Siddiqui . Therefore, it can be concluded that the combination of sodium hydrosulfite and cellulase enzyme reduces the destructive effect of the enzyme as shown in Figure 13-d and Figure 13-e.
Crease Recovery Angle Evaluation
To investigate the effects of the materials and treatments on the wrinkling of the treated fabrics, the crease recovery angle of the specimens was analyzed and the results can be seen in Table 4. According to the results, sodium hydrosulfite increased the wrinkle resistance of the fabric. This problem was observed more in the printed specimens than in the washed specimens. No significant change in this parameter was observed in the Laccase-treated specimens, which is due to the fact that there is no interaction with cellulose fibers. This problem was also found in the research carried out by Maryan and Montazer . Specimens treated with cellulase enzymes were found to have a significant decrease in the crease recovery angle, which is due to the surface degradation of the cellulosic fibers, which in return reduces the resistance of the fabric to external forces to compress the fabric. In other methods, such as using oxidizing agents by increasing the treatment variables such as concentration, time, and other parameters, the wrinkling and bending stiffness of the fabrics continuously decreased, indicating further destruction of the fiber structure [21, 58]. The wrinkle recovery of the printed specimens (reduced by 14 and 20 degrees for the specimens printed using the two enzymes and sodium hydrosulfite and the speci,ents printed with laccase enzyme and sodium hydrosulfite, respectively) was greater than that of the washed specimens (reduced by 7 and 9 degrees in the crease recovery angle for the washed specimens using the two enzymes and sodium hydrosulfite and washed with laccase enzyme and sodium hydrosulfite, respectively), which can be attributed to the greater degradation of the fibers due to the nature of the method.
This result can be clearly seen in the SEM images in Figure 11. More fibrillation occurred in the specimens treated by the printing method. Also, the higher wrinkle resistance of the specimens treated with two enzymes and sodium hydrosulfite compared to the specimens treated with two enzymes confirms the fact that sodium hydrosulfite reduces the destructive effect of the cellulase enzyme due to its reducing function. In a study by Montazer and Marian in which cellulase and laccase enzymes were used to decolorize indigo-dyed denim, the numerical values of the crease recovery angle increased for cellulase treated, two enzymes treated, laccase treated, and untreated specimens, respectively. The reason for the increase in wrinkle resistance of the specimens treated with the two enzymes compared to the specimens treated with cellulase is that the enzyme has a large molecule and its function is limited to the surface area, so with the addition of laccase, the available surface area of the cellulase enzyme was reduced and the degradation carried out by the enzyme decreased accordingly .
Air Permeability Evaluation
As shown in Table 4, in the specimens treated with the two enzymes as well as the two enzymes with sodium hydrosulfite, the air permeability was increased by 16 and 14 percent in the printed specimens and 19 to 16 percent in the washed specimens. It can be said that the pills can be removed by the action of cellulase enzyme on cellulose fabrics, therefore the air permeability is higher in the specimens treated by the washing method. It also seems that the reason for the higher air permeability of the washed specimens is due to the mechanical treatment during the process of discoloration as well as the removal of more fibers from the fabric surface , which may contribute to the improvement and completion of the cellulase enzyme function. In a study conducted by Nasrin Hagh et. al, after treating the denim fabric with calcium hypochlorite, it was observed that the air permeability decreased with time. The probable reason for this is the excessive damage, which led to the formation of too many anchor fibers on the surface of the fabric. Also, when a combination of ultraviolet irradiation and oxidizing agents was used with increasing treatment time, the air permeability remained constant, which may be due to relative destruction . No decrease in air permeability in the present study indicates controlled degradation (Figure 11). In concolusion, the lower air permeability in the specimens treated with the two enzymes without sodium hydrosulfite may be due to the greater degradation atfabricsurface (Figure 11-b and Figure 11-d). Therefore, the air permeability and moisture content improve the wearing comfort of the garment made from the fabric, both of which improved after the discoloration process in this study.
Abrasion Fastness Evaluation
Table 5 shows the results of the abrasion fastness test results of the specimens. Firstly, it can be seen that for the printed specimens treated with sodium hydrosulphite and / or the laccase enzyme, compared to the same washed specimens, the washed specimens have a higher abrasion fastness. It can be said that the washing method produced anchor fibers and fuzzy fibers and pills  due to surface degradation by the cellulase enzyme . They were removed from the fabric surface by the mechanical treatment, while this was not the case in the printing method. The fact that the mechanical treatments increase the areas on the fabric surface available for destructive treatment by the cellulase enzyme was mentioned in the studies of Kahn et al. . Moreover, the specimens treated with two enzymes and sodium hydrosulfite showed lower abrasion fastness with both methods. The reason may be the destructive effect of the cellulase enzyme on the celluose fibers, but this destructive effect is higher in the printing method and resulted in lower fastness compared to the washing method (Figure 11). It can be said that due to the limited effect of the enzyme on the fabric surface in the printing method, this opportunity was given to the cellulase enzyme as much as possible. Also, the abration fastness is higher for samples treated with two enzymes and sodium hydrosulfite compared to the specimens treated with the two enzymes. It can be said that the reason for this mechanism is the decrease of the destructive effect of the cellulase enzyme with sodium hydrosulfite by preventing the oxidation of cellulose by the enzyme .