Lysozyme was inkjet printed on two different polyester fabrics considering several challenges of printing enzymes on synthetic fabric surfaces. Wettability of both the fabrics were improved by alkaline pre-treatment resulting reduction in water contact angle to 60±2 from 95°±3 and to 80°±2 from 115°±2 for thinner and coarser fabric respectively. Activity of lysozyme in the prepared ink was 9240±34 units/ml and reduced to 5946±23 units/ml as of collected after jetting process (before printing on fabric). The formulated ink was effectively inkjet printed on alkali treated polyester fabric for antimicrobial applications. Retention of higher activity of the printed fabric requires further studies on enzyme-fibre binding mechanisms and understanding protein orientation on fabric surface after printing

This thesis explores the possibilities of printing enzymes using resource-efficient technologies to promote the binding of other proteins and biomaterials on synthetic textiles. This strategy can be used to develop advanced textiles for applications, for example, in antimicrobial, drug delivery and biosensing. Digital inkjet printing was combined with enzyme technology to ensure minimum use of water, chemicals and energy in textile manufacturing processes.  

Inks containing two enzymes, lysozyme and tyrosinase, were formulated by adjusting several rheological and ionic properties. The activity of these enzymes was optimised while being printed through two different industrial grade piezoelectric printheads. The theoretical printability of the prepared inks was calculated. The effect of printhead temperature and number of printing passes on the activity was evaluated. Polyester (polyethylene terephthalate) and polyamide-6,6 were pre-treated through several techniques to understand their effect on enzyme adhesion, binding and activity retention. Tyrosinase was used to bind lysozyme on plasma activated polyamide-6,6 surface. The effects of printing these two enzymes in various sequences, i.e. tyrosinase before lysozyme and vice-versa on binding stability and activity, were studied. Influence of the printing process on enzyme kinetics was evaluated. Ability to store and reuse printed fabrics was also studied.  

Lysozyme and tyrosinase containing inks showed activity retention of 85% and 60%, respectively. Activity of lysozyme containing ink was optimum at 10–15 mPa.s when glycerol was used as a viscosity modifier. However, the optimum viscosity for tyrosinase containing ink was at 6–9 mPa.s, and carboxymethyl cellulose was found to be the most favourable modifier. For both inks, a surfactant amount below the critical micelle concentration was considered to be the most effective for printing. Among the studied fabric pre-treatment methods (alkaline, cutinase and plasma), it was found that the activity and stability of the enzyme were dependent on the nature of the pretreatment processes, which can be beneficial for different application areas, e.g. drug release and bio-sensing. Upon printing both inks on a plasma treated polyamide-6,6, tyrosinase was able to catalyse lysozyme protein to bind it on fabric. A maximum of 68% lytic activity was retained by lysozyme when it was printed after tyrosinase. This fabric showed inhibition of bacterial growth and retained almost half of its initial activity when cold stored for a month. 

Enzymes can be immobilized on textiles to impart anti-microbial properties in a more environment-friendly manner compared to conventional biocide-based solutions. Such application requires ensuring precise, flexible and contamination-free immobilization methods that can be offered by digital printing compared to coating or screen-printing techniques. Drop-on-demand inkjet printing is a resource-efficient technology that can ensure these requirements. The use of polyester and polyamide-based fabrics is rising for applications ranging from apparel and home furnishing to hygiene and medical textiles. These fibers offer superior chemical, physical, and mechanical properties due to their inert nature but challenge the printing process due to hydrophobicity and lack of functional groups. Lysozyme and tyrosinase are two enzymes showing great potential for grafting on synthetic fabrics paving the way to use them for inkjet printing as well.

Challenges for inkjet printing of enzymes on synthetic fabric surfaces come in multiple forms i.e. ink recipe formation, printer mechanics and fabric surface characteristics. The ink must maintain a suitable viscosity and surface tension for effective drop ejection and a feasible ionic nature for enzyme activity. Then, the enzyme must be able to sustain the temperature and shear stress generated inside an inkjet printhead. Finally, influential fabric characteristics include surface structure, pore size distribution, evaporation rate and binding mechanism. By considering these parameters, lysozyme and tyrosinase were successfully printed on variously modified synthetic fabrics using a combination of sustainable technologies.

Tyrosinase enzyme was digitally printed on plasma pretreated polyamide-6,6 fabric using several sustainable technologies. Ink containing carboxymethyl cellulose was found to be the most suitable viscosity modifier for this enzyme. Before and after being deposited on the fabric surface, the printed inks retained enzyme activity of 69% and 60%, respectively, compared to activity prior printing process. A good number of the printed enzyme was found to be strongly adsorbed on the fabric surface even after several rinsing cycles due to surface activation by plasma treatment. Rinsed out fabrics retained a maximum activity of 34% resulting from the well-adsorbed enzymes. The activity of tyrosinase on printed fabrics was more stable than ink solution for at least 60 days. Effects of pH, temperature and enzyme kinetics on ink solution and printed fabrics were assessed. Tyrosinase printed synthetic fabrics can be utilized for a range of applications from biosensing and wastewater treatment to cultural heritage works.

An ink containing tyrosinase catalyzes the tyrosine residues on lysozyme protein to bind it on a plasma-treated polyamide-6,6 (PA) fabric. Inkjet printing enables controlled and sequential deposition of two enzymes on PA which is necessary for proper binding. The effect of different printing sequences on crosslinking stability and enzymatic activity is presented. The lysozyme bound on the fabric shows satisfactory antimicrobial activity. The printed fabric retains about 68% of the ink activity when tyrosinase is printed before lysozyme. Further, this fabric retains about 24% of the initial activity up to four reuses. The fabric shows acceptable inhibition of bacterial growth and retains almost half of its initial activity when cold stored for a month. This work shows the potential of protein binding on textile surface using various means of sustainable technologies, namely enzyme, inkjet, and plasma. 

Enzymes immobilized on synthetic polyethylene terephthalate (PET) textile surface by resource‐efficient inkjet printing technology can promote developments for various novel applications. Synthetic fabrics often require adequate pretreatments to facilitate such printing process. This work discusses PET–woven fabric pretreatment routes to improve wettability by alkaline, enzymatic, and plasma processes for effective printing of lysozyme using an industrial piezoelectric printhead. Results indicate that all pretreated samples contain a similar amount of enzymes upon printing. Plasma treated fabrics show relatively more hydrophilic surface characteristics, better protein binding stability, and lower retained activity. Alkali and cutinase‐treated samples possess relatively higher activity due to the greater amount of enzyme desorption to substrate solution. Depending on respective enzyme‐binding stability, a combination of a well-pretreated surface and inkjet as preferential placement technology, the approach of this study can be used as a facile enzyme immobilization method for suitable applications, for example, controlled‐release and bio‐sensing.

Inkjet printing of enzymes can facilitate many novel applications where a small amount of materials need to be deposited in a precise and flexible manner. However, maintaining the satisfactory activity of inkjet printed enzyme is a challenging task due to the requirements of ink rheology and printhead parameters. Thus to find optimum inkjetting conditions we studied the effects of several ink formulation and jetting parameters on lysozyme activity using a piezoelectric printhead. Within linear activity range of protein concentrations ink containing 50 µg/mL lysozyme showed a satisfactory activity retention of 85%. An acceptable activity of jetted ink was found at pH 6.2 and ionic strength of 0.06 molar. Glycerol was found to be an effective viscosity modifier (10–15 mPa.s), humectant and protein structure stabilizer for the prepared ink. A non-ionic surfactant when used just below critical micelle concentration was found to be favourable for the jetted inks. An increase in activity retention was observed for inks jetted after 24 hours of room temperature incubation. However, no additional activity was seen for inkjetting above the room temperature. Findings of this study would be useful for formulating other protein-based inks and setting their inkjet printing parameters without highly compromising the functionality.

Immobilization of enzymes on textiles can impart a range of advanced applications e.g. anti-microbial, controlled release, drug delivery and bio-sensing (Wehrschütz-Sigl et al., 2010). Such applications enable minimal consumption, recovery, and reusability of these valuable bio-materials compared to their conventional textile applications in surface cleaning and finishing (Araujo et al., 2008). Methods used for immobilization can play important roles to ensure precise, flexible and contamination free application. Compared to many of the conventional methods of textile immobilization such as coating and screen-printing, digital inkjet technology offers many benefits for such advanced applications (Kan and Yuen, 2012). Among various inkjet technologies, drop-on-demand piezoelectric printing is a promising resource-efficient technology for enzyme immobilization. 

The enzymes should retain high activity after the immobilization process. Various factors involved during inkjet printing (Saunders and Derby, 2014) and fabric characteristics (Mohamed et al., 2008) can influence this enzymatic activity. Factors concerning the inkjet procedure include rheology and ionic nature of ink along with the shear force and waveform generated inside a piezoelectric printhead (Magdassi, 2010). Factors dependent upon fabric characteristics include surface structure, pore size distribution, and binding mechanism (Nierstrasz and Warmoeskerken, 2003). In this work, we have studied the effects of inkjet procedures on enzymatic activity. Lysozyme being a stable and well-studied enzyme was chosen for our experiments. A Xennia Carnelian printer with a Dimatix QS10 industrial printhead was used for inkjetting. Lytic activity of lysozyme was studied by a UV-Vis spectrophotometer against decrease of Micrococcus lysodeikticus cell concentration at 450 nm. Results showed ca. 10-15% activity reduction of the jetted lysozyme ink. As all the ink and printer parameters were optimized, the probable reason for such reduction could be the effect of shear forces inside the printhead on three-dimensional conformation of lysozyme. In conclusion, our formulated lysozyme ink showed potential for printing textiles with probable activity reduction that require further investigation. 

Children suffering from autism spectrum disorder are often reported to encounter sleeping disorder several causes such as hypersensitivity as a result of irregular brain and muscle functions. Disturbance in sleep affects not only their health but also daytime activities including the risk of other cognitive and behavioral impairments. Such hindrance in sleep has been demonstrated to treat therapeutically by measures like the application of deep pressure touch and full body vibration which can be beneficially integrated into the sleeping environment such as on the textile-based platform around the bed. With such a vision, this pilot design project aimed to develop a smart textile based sleeping bag incorporated with sensors to detect awakening stage of the child and thereby actuating stimuli for assuaging the child to fall asleep. To serve the purpose, a micro-controllable body movement detection sensor, based on conductive yarns connected to a vibrating motor was prosperously embedded at the interior of the sleeping bag along with weighted slots to exert deep touch and soothing sensation in the form of wearable technology.

The catalytic activity of the enzymes can be introduced to textile surfaces for bio-sensing applications by immobilizing them through a resource-efficient deposition method such as inkjet printing [1]. Contrary to conventional dispensing methods, drop-on-demand inkjet printing can provide with high precision deposition of these enzymes along with flexibility for small-scale production [2]. To the best of our knowledge, studies on the inkjetting of enzymes are limited and often uses a modified/adapted commercial paper printer for jetting [3]. Additionally, the effect of ink formulation and printing condition variables on the activity of enzyme are not well explored. Many of such variables suggested for jetting of proteins [4] includes e.g. ink rheology, operating temperature, drop size retention, and the shear force acting on the ink. In our research effect of these variables are studied using a digital inkjet printer (Xennia Carnelian) with a Sapphire QS10 piezo-electric print head (Fujifilm Dimatix, USA). Lysozyme is used as a model enzyme for printing due to its well-known structure and catalytic mechanism. Effect of temperature and shear force development within the print head on lysozyme activity is investigated. Additionally, pre-treatment of the fabric to improve ink adhesion through various surface activation processes are studied. Finally, remaining activity of the printed enzymes over washing is evaluated to ensure the fastness property.

Acknowledgment

This research project is funded by University of Borås, Sweden.

References

[1]     Li J, Rossignol F, Macdonald J. Inkjet printing for biosensor fabrication: combining chemistry and technology for advanced manufacturing. Lab on a Chip 2015;15(12):2538-2558.

[2]     Nierstrasz V, Yu J, Seipel S. Towards more flexible, sustainable and energy-efficient textile functionalization processes: Digital inkjet in functional and smart textile production. In: 9th Aachen-Dresden International Textile Conference 2015; 2015.

[3]     Yamazoe H. Fabrication of protein micropatterns using a functional substrate with convertible protein-adsorption surface properties. J Biomed Mater Res A 2012;100(2):362-9.

[4]     Delaney JT, Smith PJ, Schubert US. Inkjet printing of proteins. Soft Matter 2009;5(24):4866-4877.

Development of biocide-based antimicrobial textiles is proving to be a concern for the economy, and more evidently, for the environment and health. On the contrary, probiotic (beneficial bacteria) can replace these traditional biocides in order to overcome the toxicity and resistance problems. This paper elaborates an adapted sol-gel coating process to embed such beneficial spores on the polyester woven surface, and their viability is studied along with the characterization of the physical properties of the coated fabric. The results illustrate the successful incorporation of the beneficial spores with an adequate number of living organisms (even after repeated washing cycles), sufficient tensile strength, and good abrasion resistance properties with the opportunity to improve surface wettability maintaining sufficient adhesion between the fibre and the coated layer.