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. 2023 Feb 28;13(3):106. doi: 10.1007/s13205-023-03526-x

Valorisation of wheat straw into paper with improved quality characteristics using ultrafiltered xylano-pectinolytic pulping approach

Sharad Agrawal 2, Divya Sharma 3, Raksha Nagpal 1, Amanjot Kaur 1, Nishikant Bhardwaj 4, Ritu Mahajan 1,✉
PMCID: PMC9975122  PMID: 36875962

Abstract

This study has been conducted to assess the pulpability of ultrafiltered pectinase and xylanase in pulping of wheat straw. Best biopulping conditions were achieved using 107 and 250 IU of pectinase and xylanase, respectively, per gram of wheat straw, 180 min of treatment period, one gram: 10 m1 material to liquor ratio, 8.5 pH and 55 °C temperature. Ultrafiltered enzymatic treatment improved the pulp yield (6.18%), brightness (17.83%), along with reduced rejections (61.01%) and kappa number (16.95%) as compared to chemically synthesized pulp. Biopulping of wheat straw saved 14% alkali dose, with nearly same optical properties, as obtained under 100% alkali dose. Bio-chemically pulped samples resulted an increase in breaking length, tear index, burst index, viscosity, double fold and Gurley porosity by 6.05%, 18.64%, 26.42%, 7.94%, 21.6% and 15.38%, respectively, in comparison to control pulp samples. Bleached-biopulped samples showed an improvement in breaking length, tear index, burst index, viscosity, double fold number, and Gurley porosity by 7.39%, 3.55%, 28.82%, 9.1%, 53.66%, and 30.95% respectively. Thus, biopulping of wheat straw with ultrafiltered enzymes lowers alkali consumption and also improves the paper quality. This is the first study reporting, eco-friendly biopulping, for producing better quality wheat straw pulp, using ultrafiltered enzymes.

Keywords: Wheat straw, Biopulping, Ultrafiltered xylano-pectinolytic enzymes, Soda-anthraquinone pulping, Alkali dose, Strength properties

Introduction

Increased requirement for paper production and insufficient wood supply has prompted researchers to focus on finding suitable nonwood material for pulp and paper processing. Various types of nonwoods, generated as agricultural by products, have been analysed by various researchers and wheat straw has been found to be one of the most predominant agro-based residue. Wheat cultivation occupies the largest agricultural land in the world. Currently, global wheat farm area exceeds 2.2 million ha, and an estimated production of wheat is 730 million t, comprising 1/3rd of the global food (Lu et al. 2017). From the perspective of production, world's largest producer of wheat is China, followed by India, Russia, America and nearly 50% of world’s wheat is being produced by these four countries (Liu et al. 2016; Zhao et al. 2019). Therefore, annually, above 650 million t of wheat straw is available (Li 2011). Due to shortage of forest land and ease of the availability of wheat straw, it could be a better raw source for the industry for manufacturing paper in future. Wheat straw is made up of cellulose (45.5%), xylan (25.5%), lignin (18%), ash (8.5%) and pectin (Petroudy et al. 2018; Volynets and Dahman. 2011).

In the conventional process of paper production, pulping and bleaching process involves huge quantity of strong pulping and bleaching chemicals. The use of chemicals in traditional paper manufacturing processes have created difficulties in disposal of hazardous waste water (Ayyachamy and Vatsala 2007; Verma and Satyanarayana 2013; Azevedo et al. 2020; Haq and Raj 2020). Due to lack of proper irrigation system, farmers use this toxic waste water in irrigation, which may further enter into the food chain and consequently, may affect soil, plant and animal health, as reported by World Health Organization (Ahmed et al. 2021). Hence, nowadays, more emphasis is being paid on the development of greener, cleaner techniques for the generation of pulp, keeping in mind their least hazardous impact on the environment (Varghese et al. 2020; Nagpal et al. 2022). Microbial degradation of lignocellulosic material has been significantly emphasized. Enzymatic treatment has been found to increase diffusion rate of NaOH in both softwoods as well as hardwoods (Woldesenbet et al. 2012).

This study emphasizes on the applicability of partially purified xylano-pectinolytic enzymes as biopulping agent, before the traditional pulping, in order to bring down the consumption of toxic pulping chemical, while producing an upgraded quality of paper. Biocatalytic approach for preparing pulp from wheat straw using ultrafiltered xylano-pectinolytic enzymes has not been reported till date.

Materials and methods

Materials

Bacillus pumilus AJK was used for the production of xylanase and pectinase enzymes, using strain (MTCC accession no.10414) previously isolated by Kaur et al. (2010). All analytical reagent grade chemicals were used for this research work and purchased from Himedia (India). Agro-residues like citrus peel and wheat bran were procured locally. Assay chemicals (polymethylgalacturonic acid birchwood xylan, xylose, and galacturonic acid) were bought from Sigma chemicals Co. Ltd. (USA). The collection of straw samples was done from a village near Kurukshetra, India. After collection of straw, it was chopped into small pieces, washed, dried and used for further study.

Xylano-pectinolytic enzymes production, ultrafiltration step and enzymes activity estimation

Submerged fermentation approach was employed for the manufacturing of enzymes, using 50 ml media containing (g/l): MgSO4, 2.46; peptone, 5.0; citrus peel, 20; wheat bran, 20; pH 7.0. For inoculation, 2% of 21 h old bacterial culture was used and production of enzymes was done at 37 °C for 60 h with continuous shaking at 200 rpm. After 60 h, enzymes production was stopped by centrifuging at 8000g for 10 min (Kaur et al. 2011). The clear crude extract obtained, was further microfiltered and ultrafiltered, sequentially, under a transmembrane pressure of 5 psi (using a Quixstand model system of GE Healthcare Biosciences Ltd.), according to the methodology given by Agrawal et al. (2019).

The enzymes assay was performed by quantitative estimation of reducing sugars content using Miller reagent (Miller 1959) after enzyme substrate reaction under the same conditions, as given by Kaur et al. (2011). The batch of enzyme used contained 270 and 115 IU of xylanase and pectinase respectively, per mL of supernatant.

Absorption spectra of un-purified and partially purified enzymes

Absorption spectra of crude as well as ultrafiltered xylano-pectinolytic enzymes extract were taken between wavelength range of 200–800 nm, to analyse the removal of impurities in partially purified enzymes extract, which may affect the enzymes kinetics.

Evaluation of enzyme dose and treatment time for biopulping experiments

The treatment conditions for best biopulping were estimated, with ultrafiltered xylanase and pectinase enzymes dosage, ranging from 50 to 400 and 21.5 to 170 IU/g wheat straw, respectively and time from 60 to 360 min, keeping pH at 8.5, treatment temperature at 55 °C and material to liquid ratio of 1:10 g/ml. After treatment, pulp free filtrate samples were used for the assessment of biopulping process, by measuring the amount of hydrolysed reducing sugars using Miller's method, along with measurement of the absorbance of different non-cellulosic impurities like lignin, hydrophobic and phenolic compounds at various wavelengths from 200 to 465 nm (Miller 1959; Nissen et al. 1991; Patel et al.1993; Khandeparkar and Bhosle 2007). For the control samples, enzymes were not added. After each treatment, washing of samples was done with water for removal of enzymatic media components and then dried at 45 °C. Microscopic analysis was made in order to compare the efficacy of partly purified enzymes on the structure of wheat straw cell wall, with control wheat straw samples. All experiments were carried out in triplicates.

Chemical pulping

Chemical pulping experiments of control and biopulped wheat straw samples were carried, as mentioned by Varghese et al. (2020), using following parameters: wheat straw to liquor ratio (1:4 w/v), alkali dose (16%), anthraquinone (0.05%), maximum temperature/ramp time/ incubation time (160 °C/ 90 min/ 20 min), for comparison of both samples. After comparison, biopulped samples were treated with variable alkali doses (13.6–16%), so as to check the alkali dose reduction in biopulped samples. After cooking, pulp and black liquor analysis were done to optimize the chemical pulping process after biopulping. In analysis of black liquor, the pH and residual alkali content (TAPPI T625 2014) were measured. Pulp samples were thoroughly washed and screened before pulp analysis. The parameters like, pulp yield, viscosity (TAPPI T230 1976), pulp brightness and kappa number (TAPPI T236 2004) were also determined.

Bleaching of enzymatically treated and chemically treated (100%) wheat straw soda-anthraquinone pulps and testing of handsheets

Using conventional bleaching series, the control (100% alkali dose) and enzymatically treated (with 14% less alkali dose) samples were bleached after pulping, using DoEPD1D2 sequence, where Do, D1 and D2 stages represent ClO2 treatment stages and EP represents alkali—peroxide treatments. The amount of ClO2 dose required was calculated using Hise equation (Hise 1996). After each bleaching treatment, percentage consumption of ClO2 was determined. The pulp samples were washed after each step, and were stored for analysing their optical properties and kappa number.

Handsheets of both control and ultrafiltered enzymes treated samples were made using TAPPI T205 sp-02 (2002), after pulping as well as bleaching stages. For the comparison of optical and physical properties, handsheets of 60 GSM was used. The optical properties were measured using TAPPI T217 wd-77, (2004), while various physical properties like breaking length (TAPPI T494 2001), burst index (TAPPI T403 2010), Gurley porosity (TAPPI T460 2002), double fold number (TAPPI T511 2002), Schopper–Riegler freeness (°SR) (SCAN-C 1999), viscosity (TAPPI T230 1976), and tear index (TAPPI T414 2004) were measured using various TAPPI methods.

Results and discussion

The current study was undertaken in order to utilize easily available, agrowaste wheat straw, as raw material for paper production and simultaneously to reduce pollution generated by paper industry, which is the strong need of these industries these days. There is intense pressure on paper industries to reduce the toxic load. So, to reduce pollution, ultrafiltered xylano-pectinolytic enzymes cocktail was used before conventional pulping process, so as to decrease the pulping chemical demand and simultaneously increase the quality of paper or no negative impact on the paper. Data on biopulping of wheat straw using ultrafiltered pectinase and xylanase enzymes, for manufacturing paper is not available in the literature till date, so this study will definitely reduce chemical pollution and valorise wheat straw into better quality paper. The results are presented in the following sections.

Analysis of absorption spectra of crude and ultrafiltered enzymes

Absorption spectra of crude and ultrafiltered enzymes preparations were analysed. The ultrafiltered xylano-pectinolytic enzymes preparation obtained after passing through 1000 Da cut off membrane showed the removal of many impurities, as can be seen from the absorption spectra of crude and ultrafiltered enzymes preparations (Fig. 1a, b). Absorption spectrum of crude enzymes preparation gave many peaks, but in case of ultrafiltered enzymes preparation, only some absorption peaks were detected, which indicates the removal of impurities from the enzyme preparation, which may interfere with the catalytic efficacy of these enzymes. This ultrafiltered enzyme preparation was further used for pulping of wheat straw.

Fig. 1.

Fig. 1

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Absorption spectra of (A) crude enzymes and (B) ultrafiltered enzymes preparations

Assessment of biopulping process

Biopulping is mainly affected by these parameters, enzyme dose and treatment time. These parameters were checked, so as to finalize the best treatment conditions. Nearly 250 and 107 IU of xylanolytic and pectinolytic catalysts respectively, displayed the highest release of reducing sugars at 55 °C temperature, 8.5 pH and ratio of material to liquor 1:10 g/ml (Fig. 2a). An optimized enzyme dose of 60 IU of pectinolytic enzyme per g of bagase, at temperature 55 °C, pH 9.0 and ratio of material to liquor 1:7 (Kg/L) has been reported (Liu et al. 2017a). Varghese et al. (2020) used material to liquor ratio (1:10 g/ml), temperature 55 °C, pH 8.5 and crude xylanolytic-pectinolytic enzymes dose (400–120 IU) per g of wheat straw for biopulping. The alkaline conditions suitable for biological delignification have also been reported in few other studies (Sporck et al. 2017; Cornetti et al. 2020). Maximum efficacy of ultrafiltered enzymes in biopulping of wheat straw was obtained after treatment time of 180 min (Fig. 2b). Liu et al. (2017a) have mentioned treatment time of 60 min for best enzymatic interaction during biopulping of bagasse. Ultrafiltered xylano-pectinolytic enzymes treatment under the best conditions, released maximum reducing sugars (8.454 ± 0.06 mg/g of wheat straw). The optimized enzymatic treatment also released maximum non-cellulosic impurities, with absorbance values of 0.8742 (for phenolic compounds at λ 237 nm), 1.1298 (for lignin compounds at λ 280 nm), 0.2495 (for lignin compounds at λ 400 nm) and 0.1534 nm (for hydrophobic compounds at λ 465 nm), after 10 × dilution (Fig. 2b). Varghese et al. (2020) also reported higher release of reducing sugar (7.410 ± 0.06 mg/g wheat straw) and non-cellulosic impurities under best biopulping conditions, after treatment of wheat straw with crude enzymes. Microscopic images (at 400X magnification) have also shown the hydrolytic enzymatic activity on the cell wall of biopulped wheat straw samples, as the cell walls of raw wheat straw were intact, but in case of biopulped wheat straw sample, cell walls were broken, thus confirming the efficacy of ultrafiltered enzymes in biopulping of wheat straw (Fig. 3).

Fig. 2.

Fig. 2

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Effect of (A) ultrafiltered enzymes quantity and (B) duration of enzymatic treatment on wheat straw biopulping

Fig. 3.

Fig. 3

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Microscopic pictures (A) raw wheat straw (B) enzymatically treated wheat straw at 400X magnification

Chemical pulping versus bio-chemical pulping

The control and enzymatically treated samples were chemically pulped under conventional cooking conditions, to analyse the effectiveness of enzymatic pulping process. Post-chemical pulping process of enzymatic pulp samples, resulted in the reduction of kappa number (16.95%), total solids (4.44%) and rejects (61.01%) (Table 1). Pectinase treated sugarcane bagasse displayed a significant reduction of 17.8% in the kappa number values, after 60 min of conventional pulping treatment process (Liu et al. 2017a). Using crude pectinase and xylanase enzymes in wheat straw pulping, also showed reduction of kappa number by 15.67% (Varghese et al. 2020). The removal of lignin, significantly improves the brightness in the enzymatic treated samples. The brightness of biopulped samples improved by 17.83% in comparison to chemically treated pulp samples, under similar cooking conditions (Table 1). Similarly, improvement of 16.04% in the brightness of pulp of wheat straw is reported, using crude enzymes (Varghese et al. 2020). Liu et al. (2017b) demonstrated an increase of 5.5% in the brightness of pectinase assisted biopulped bagasse samples. The higher screened pulp yield (6.18%) and residual alkali content (3.6 g/l as NaOH) was observed in biopulped samples after conventional pulping (Table 1). The reason for higher alkali content in black liquor, is due to less requirement of alkali dose in bio-catalytically treated straw samples as compared to control wheat straw samples. After pectinase treatment of bagasse for 100 min, residual alkali content in black liquor was higher (Liu et al. 2017a). These improvements in the pulp properties of biopulped samples (i.e. reduction in rejects, kappa number and improved pulp yield, brightness as compared to control wheat straw samples) indicate effective enzymatic assisted cooking. These results depict that, ultrafiltered enzymatic treatment increased the permeability of pulping chemicals onto the lignin polysaccharides present in straw cell wall and removed more lignin after chemical pulping.

Table 1.

Reduction in alkali dose after ultrafiltered enzymatic pulping

Conditions for cooking Chemical method Enzymes + different dose of alkali added
100% 100% 95% 90% 88% 86% 85%
Alkali dose (%)/anthraquinone (%) 16/0.05 16/0.05 15.2/0.05 14.4/0.05 14.08/0.05 13.76/0.05 13.6/0.05
Wheat straw to liquid ratio 1:4 1:4 1:4 1:4 1:4 1:4 1:4
Temperature (oC)/Heating time (min)/ Holding time at maximum temperature (min) 160/90/20 160/90/20 160/90/20 160/90/20 160/90/20 160/90/20 160/90/20
Pulp properties
 Unscreened pulp yield (%) 61.4 64.8 65.6 65.8 66.0 66.4 66.8
 Rejects (%) 0.59 0.23 0.32 0.44 0.54 0.63 0.71
 Screened pulp yield (%) 60.81 64.57 65.28 65.36 65.46 65.77 66.09
 Pulp brightness (% ISO) 39.8 46.9 43.8 41.2 40.6 39.9 39.4
 Kappa number 13.1 10.88 11.55 12.26 12.76 13.04 13.46
 Residual alkali (g/l as NaOH) 3.0 6.6 5.2 4.4 3.8 3.1 2.4
 Total solids (w/w) 13.5 12.9 12.2 12.0 11.8 11.5 11.2
 pH of black liquor 11.5 12.6 12.1 11.9 11.7 11.4 11.2
Physical properties
 °SR 23 22
 Double fold 37 45
 Gurley porosity (s) 39 45
 Breaking length (m) 3849 4082
Tear index (mNm2/g) 6.49 7.70
 Viscosity (cP) 18.9 20.4
 Burst index (kN/g) 2.46 3.11
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With the usage of ultrafiltered xylano-pectinolytic enzymes for wheat straw biopulping, consumption of pulping chemical (i.e. alkali dose) reduced up to 14%, which ultimately improved pulp yield and lowered total solids by 8.15% and 14.81%, respectively (Table 1). The ultrafiltered enzymes preparation with less impurities increased the pulping efficiency and decreased the consumption of alkali during pulping. Ultrafiltered enzymatic action on wheat straw improved the diffusion rate for pulping chemical, hence effective for delignification process, which further reduced the consumption of alkali and increased the yield of pulp. Pectinase assisted pulping of sugarcane bagasse reduced the alkali dose consumption up to 1% and improved the pulp yield by 15.8% (Liu et al. 2017a), while Varghese et al. (2020) have reported 12% mitigation of pulping chemical, after conventional pulping of crude enzymes treated wheat straw pulp samples. The enzymatic treatment improves the delignification from lignocellulosic material, hence, it lowers the consumption of alkali content during conventional pulping process.

Different physical properties of chemically treated pulp (with 100% pulping chemicals) were compared with ultrafiltered enzymes plus 86% chemically treated pulp samples (Table 1). Bio-chemically pulped samples with 14% less alkali dose, resulted an increase in breaking length, tear index, burst index and viscosity by 6.05%, 18.64%, 26.42%, and 7.94%, respectively in comparison to control pulp samples. The double fold and Gurley porosity also increased from 37 to 45 and 39 to 45, respectively. Bio-pulping has been introduced as a means of producing a stronger paper product (Akhtar et al. 1998, 2000). Paper strength properties like tear factor (7.0%), burst factor (16.5%), and breaking length (17.1%) were improved in pectinase assisted biopulping of sugarcane bagasse (Liu et al. 2017b). Yang et al. (2019) reported that, the enzymatic action increases the permeability of chemicals to the fiber surface, thereby improves the strength of the paper. Varghese et al. (2020) also displayed improvement in various strength parameters like tear index (18.22%), burst index (26.50%), and breaking length (5.56%) after biocatalytic treatment of wheat straw with crude enzymes. The reason for the improvement of strength properties after enzymatic treatment was selective removal of polysaccharides like xylan and pectin after hydrolytic reaction by ultrafiltered enzymes. This selective action of enzymes helps in increasing the porosity of straw, which improves the impregnation of pulping chemicals, and hence removes non-cellulosic impurities without affecting any strength properties.

Bleaching of enzyme treated and chemically treated wheat straw pulps

Biopulp (treated with 86% alkali dose) and control (treated with 100% alkali dose) samples were further bleached using DoEPD1D2 sequence. It was observed that, biopulped samples have higher bleaching efficiency over control pulp samples (Table 2). After the EP stage, an increase of 10.91% and 10.33% in brightness and whiteness, along with 15.27 and 11.76% decrease in yellowness and kappa number, respectively was observed in bleached- biopulped samples. Similarly, after stage D1, bleached- biopulped samples showed higher brightness (2.48%), whiteness (6.14%) and decreased yellowness (33.05%). After stage D2, biopulping resulted in decrease of yellowness (11.11%), which depicts that, biopulping with ultrafiltered xylanase-pectinase synergism have capability to reduce the consumption of alkali charge (14%) as well as ClO2 dose (6.61%), along with increase in the optical properties after bleaching. Crude enzymatic treatment of wheat straw also resulted in the decrease of pulping and bleaching chemicals, nearly by 12% and 4.42%, respectively (Varghese et al. 2020).

Table 2.

Optical and physical properties of pulp after biopulping followed by bleaching treatment

Bleaching Steps Pulping chemicals
(100%)		 Bio-pulping + 86% of pulping chemicals
Kappa number 13.07 13.04
D-0 ClO2 addition (%) 1.33 1.33
ClO2 used (%) 1.25 1.23
Brightness (% ISO) 59.46 60.21
EP Alkali added (%) 1.8 1.8
H2O2 addition (%) 0.5 0.5
Kappa number 3.4 3.0
Brightness (% ISO) 73.36 81.36
Whiteness (CIE) 55.76 61.52
Yellowness (ASTM) 14.54 12.32
D-1 ClO2 addition (%) 0.8 0.8
ClO2 used (%) 0.75 0.71
Brightness (% ISO) 82.56 84.61
Whiteness (CIE) 71.87 76.28
Yellowness (ASTM) 7.11 4.76
D-2 ClO2 addition (%) 0.3 0.3
ClO2 used (%) 0.27 0.18
Brightness (% ISO) 84.87 85.64
Whiteness (CIE) 79.76 80.47
Yellowness (ASTM) 4.32 3.84
Physical properties Pulping chemicals
(100%)		 Bio-pulping + 86% of chemicals
°SR 23 23
GSM 60.2 60.2
Bulk 1.63 1.63
Burst index (kN/g) 2.81 3.62
Breaking length (m) 4558 4895
Tear index (mNm2/g) 6.2 6.42
Double fold 41 63
Gurley porosity (s) 42 55
Viscosity (cP) 9.9 10.8
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After bleaching, strength properties of control and biopulped handsheets were compared. It has been resulted that, bleached- biopulped samples showed an improvement in tear index, breaking length, viscosity, Gurley porosity, burst index and double fold number by 3.55%, 7.39%, 9.1%, 30.95%, 28.82% and 53.66%, respectively (Table 2). Strength properties like double fold number (51.28%), burst index (24.31%), breaking length (6.63%), viscosity (6.12%), tear index (2.83%), and Gurley porosity (27.50%) also improved after crude enzymatic pulping of wheat straw (Varghese et al. 2020).

This study signifies the applicability of ultrafiltered xylano-pectinolytic enzymes in pulping of wheat straw. Wheat straw, an agro-based residue and abundantly available could be raw material of future for the paper industries. Adding biocatalysts to the process, improved the delignification efficiency and mitigated the usage of alkali dose, also improved the pulping—bleaching properties thus, reduced the demand of environment damaging chemicals, along with enhanced quality of the paper. Production of both enzymes by a bacterial isolate also reduces the fermentation associated cost and ultimately the overall application cost.

Conclusion

The addition of ultrafiltered xylano-pectinolytic enzymatic step before conventional pulping enhanced the effectiveness of chemical pulping in case of wheat straw, along with decline in the usage of both bleaching and pulping chemicals, hence ultimately produced paper with better optical, physical and strength properties. Production of both enzymes from a bacterial isolate, using agro-wastes based medium, would reduce the production cost and would eventually make the process economically viable. Thus, keeping in the mind the various benefits of these enzymes in paper making, this study would definitely help in manufacturing paper of better quality, along with reduced environmental pollution, which is the need of every paper industry and also valorising abundantly available, agro-residue wheat straw, for making paper.

Acknowledgements

The corresponding author is thankful to Avantha Center for Industrial Research and Development, Yamuna Nagar, for providing us the facilities for testing various properties of wheat straw pulp samples.

Author contributions

RM (Corresponding Author) given the idea of concept, planning and designing of various experiments done for this manuscript. SA (First Author)—Done all experimental work mentioned in this manuscript at research laboratory level. DS—Confirmed the results of first author. RN—Done the experimental work mentioned in this manuscript at Paper Industry level. AK—Done the microscopic analysis. NKB—supervised the research work done at Paper Industry level. All authors read and approved the final manuscript.

Funding

The financial support for this work was provided by Department of Biotechnology, Ministry of Science & Technology, Government of India (Grant number: BT/ PR 20438/BCE/8/1220/2016 for 3.5 years).

Data availability

All data generated or analyzed during this study are included in this published article.

Declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Consent to publication

Not Applicable.

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