• Vol 10, No 3 (2019)
  • Chemical Engineering

The Effect of Mixed Biological Pretreatment and PEG 4000 on Reducing Sugar Production from Coffee Pulp Waste

Toto Iswanto, Nuniek Hendrianie, Maya Shovitri, Ali Altway, Tri Widjaja

Corresponding email: triw@chem-eng.its.ac.id


Cite this article as:
Iswanto, T., Hendrianie, N., Shovitri, M., Altway, A., Widjaja, T., 2019. The Effect of Mixed Biological Pretreatment and PEG 4000 on Reducing Sugar Production from Coffee Pulp Waste. International Journal of Technology. Volume 10(3), pp. 453-462
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Toto Iswanto Department of Chemical Engineering, Institut Teknologi Sepuluh Nopember, Sukolilo, Surabaya 60111 Indonesia
Nuniek Hendrianie Department of Chemical Engineering, Institut Teknologi Sepuluh Nopember, Sukolilo, Surabaya 60111 Indonesia
Maya Shovitri Department of Biology, Institut Teknologi Sepuluh Nopember, Sukolilo, Surabaya 60111, Indonesia
Ali Altway Department of Chemical Engineering, Institut Teknologi Sepuluh Nopember, Sukolilo, Surabaya 60111 Indonesia
Tri Widjaja Department of Chemical Engineering, Institut Teknologi Sepuluh Nopember, Sukolilo, Surabaya 60111 Indonesia
Email to Corresponding Author

Abstract
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Biological methods using bacteria and fungi are regarded as more economically viable and environmentally friendly alternatives for improving lignocellulosic degradation. Coffee pulp waste (CPW) as a lignocellulosic biomass is abundant and has potential as a reducing sugar feedstock. However, it contains lignin as a matrix polymer, which associated with pectin and cover the cellulosic microfibrils and make it difficult to be digested during the bioprocess. In this study, the performance of biological pretreatment in reducing lignin and pectin using a co-culture of Bacillus subtilis (BS), Aspergillus niger (AN), or Trichoderma reesei (TR) has been investigated. The pretreatment of the CPW was made using various microbial ratios in an aerobic stirred-bioreactor and incubated at 30o C, pH 5 for 7 days. Removal of lignin and pectin was analyzed during the pretreatment process. PEG 4000 as a surfactant was used and its effect on the yield of reducing sugar production from pretreated CPW using a A. niger and T. viride (TV) co-culture with a surfactant to substrate ratio of 1:1 (w/w) was investigated. A culture without surfactant was used as a control. The results reveal that the best lignin and pectin removal was 99.9%, when using a co-culture of AN and TR with a ratio of 1:1 (v/v) and of BS and TR with a ratio of 2:1 (v/v). The cellulose content of CPW in these co-cultures was 86.99% (w/w) and 81.61% (w/w), respectively, and the reducing sugar concentration obtained was 12.5 g/L and 9.74 g/L respectively. In further hydrolysis of pretreated CPW using a AN:TV (2:1) co-culture with the addition of surfactant, the yield of reducing sugar obtained was higher than that of the control, at 20.69%. Use of PEG 4000 as a surfactant had a positive effect on enhancing the yield of reducing sugar from coffee pulp waste.

Biological pretreatment; Coffee pulp waste; Hydrolysis; Reducing sugar; Surfactant

Introduction

The utilization of agro-industrial residues as lignocellulosic biomass to produce more valuable material has encouraged the improvement of advanced biotechnological innovations, principally in enzyme and fermentation technology, which have ultimately had an impact on the reduction of environmental pollution problems (Pandey et al., 2000). One of the agro-industrial residues that is available in abundance in Indonesia, but which has particular problems associated with its utilization due to the presence of anti-nutritional factors such as caffeine, tannin, and polyphenols, is coffee pulp waste (CPW). Coffee is an important plantation commodity in Indonesia (Haryuni et al., 2019); the country produces an average amount of 686×106 kg of coffee each year (Widjaja et al., 2017a). Without proper handling, the abundant untreated CPW will release xenobiotics as toxic substances into the soil, which kill the saprophytic microorganisms, causing disruption to important biotransformation stability in the environment (Ibrahim et al., 2014).

In the biotechnological approach, CPW can be converted into reducing sugar as a substrate to produce biofuels such as ethanol and biogas. To obtain the optimum yield of reducing sugar, a pretreatment process is needed to remove the unnecessary matrix polymers such as lignin and pectin surrounding the cellulosic microfibrils. Pectin is a heterogeneous polysaccharide, whose molecular structure, weight and functional properties depend on its agro-waste source (Kusrini et al., 2018). Pectin has the function of cross-linking cellulose and hemicellulose fibers, providing rigidity to the cell wall (Abbott & Boraston, 2008). This is in line with Menon and Rao (2012), who state that hemicelluloses, amorphous polymers of different sugars, and other polymers such as pectin, attach the microfibrils of cellulose, which are stabilized by hydrogen bonds and covered by lignin, making the biomass difficult to be digested in the bioprocess.

Physicochemical pretreatment methods have been widely developed, such as steam explosion (Guo et al., 2011), or use of dilute acid (Dussán et al., 2014), alkali (Menezes et al., 2014) or oxidation, or varied combinations of these, but such processes usually involve high temperature, pressure and cost (Kumar et al., 2009). Since chemical or physical methods are not able to provide selective removal, are expensive, and are not environmentally friendly, microbial and enzymatic techniques for lignin and pectin removal could be alternative advantageous methods.

The co-culture strategy has been assessed as a more efficient way to treat lignocellulosic waste (Menon & Rao, 2012). The combination of cultures can consist of two, three, or a consortium of bacteria or fungi or a mixture of them, which can be applied in many biological production processes. In the case of lignin and pectin removal, previous studies have reported that A. niger and T. reesei are capable of degrading lignin (Adav & Sze, 2014; Asses et al., 2018). Moreover, Roussos et al. (1995) found that some strains of A. niger are capable of degrading caffeine. B. subtilis can degrade lignin and pectin (Gummadi & Kumar, 2005; Torimiro & Okonji, 2013; Cragg et al. 2015). Juliastuti et al. (2018) successfully achieved the combination of three strains, A. niger, Pseudomonas putida, and T. harzianum to treat coffee pulp in degrading lignin and anti-nutritional factor for a biogas feedstock.

Besides applying the pretreatment, the use of surfactants (i.e. PEG 4000) in the hydrolysis of lignocellulosic substrates has produced good effects. Surfactants, as surface-active additives, affect the interaction between enzymes such as cellulase and substrates (Li et al., 2012), leading to an increase in the total amount of reducing sugar after the hydrolysis process. PEG is presented in varying molecular weights (e.g. PEG 4000, 6000 and 8000); at the lowest molecular weight (PEG 4000) the amounts of reducing sugar produced were 7% and 13% higher than those from PEG 6000 and 8000 respectively (Iveti? et al., 2014). However, the effects of the addition of the same surfactants on the obtained hydrolysis product may be different, depending on the operating conditions, initial chemical content in the substrate, and treatment method during the hydrolysis process.

Therefore, this study conducts an investigation to determine the best co-culture ratio involving A. niger, T. reesei and B. subtilis to develop a co-culture for degrading lignin and pectin at a faster degradation rate and which is able to tolerate the presence of the anti-nutritional factors that effect their growth. The obtained CPW from the best pretreatment condition was further hydrolyzed using a co-culture of A. niger and T. viride with the addition of PEG 4000. The effects of the co-culture on the reducing sugar obtained from the hydrolysis process were also investigated and described.


Conclusion

The microbial method in the form of a fungal and bacterial mixture has produced satisfying results in the degradation of lignin and pectin in coffee pulp waste, with more than 99% removal of those compounds. Due to the high cellulose concentration (more than 80%) in pretreated coffee pulp waste, two co-cultures, AN:TR (1:1) and BS:TR (2:1), were chosen for the following hydrolysis process. Subsequently, the hydrolysis of pretreated CPW using A. niger and T. viride with the addition of surfactant resulted a higher yield of reducing sugar than that without surfactant. The highest yield of reducing sugar resulted from the pretreated CPW by AN:TR (1:1) and hydrolyzed by the AN:TV (2:1) co-culture with the addition of PEG 4000, at 20.69%. In this co-culture, PEG 4000 as surfactant made a difference of 3.4% in reducing sugar, compared to without surfactant. Hopefully, the co-culture ratio obtained can indicate a direction for further study, focusing on the kinetics of its cell growth, which is important for developing an economical bioprocess of reducing sugar production.

 

Acknowledgement

The authors thank Atikah Badriya Husein and Dwi Ayu Primaningrum who have provided coffee pulp waste and selfless help. We also thank all members of Biochemical Technology Laboratory and Wastewater Treatment Laboratory, Institut Teknologi Sepuluh Nopember for their endless support.

References

Abbott, D.W., Boraston, A.B., 2008. Structural Biology of Pectin Degradation by Enterobacteriaceae. Microbiology and Molecular Biology Reviews, Volume 72(2), pp. 301–316

Abd-Elsalam, H.E., El-Hanafy, A.A., 2009. Lignin Biodegradation with Ligninolytic Bacterial Strain and Comparison of Bacillus subtilis and Bacillus sp. Isolated from Egyptian Soil. Journal of Agriculture & Environment Science, Volume 5(1), pp. 39–44

Adav, S.S., Sze, S.K., 2014. Trichoderma Secretome: An Overview. In: Biotechnology and Biology of Trichoderma, Gupta, V.K. Schmoll, M., Herrera-Estrella, A., Upadhyay, R.S., Druzhinina, I., Thouhy, M.G. Elsevier, pp. 103–114

Asses, N., Ayed, L., Hkiri, N., Hamdi, M., 2018. Congo Red Decolorization and Detoxification by Aspergillus niger: Removal Mechanisms and Dye Degradation Pathway. BioMed Research International, Volume 2018, pp. 1–9

Bandounas, L., Wierckx, N.J.P., de Winde, J.H., Ruijssenaars, H.J., 2011. Isolation and Characterization of Novel Bacterial Strains Exhibiting Ligninolytic Potential. BMC Biotechnology, Volume 11(94), pp. 1–11

Chang, Y.C., Choi, D.B., Takamizawa, K., Kikuchi, S., 2014. Isolation of Bacillus sp. Strains Capable of Decomposing Alkali Lignin and their Application in Combination with Lactic Acid Bacteria for Enhancing Cellulase Performance. Bioresource Technology, Volume 152, pp. 429–436

Cragg, S.M., Beckham, G.T., Bruce, N.C., Bugg, T.D.H., Distel, D.L., Dupree, P., Etxabe, A.G., 2015. Lignocellulose Degradation Mechanisms Across the Tree of Life. Current Opinion in Chemical Biology, Volume 29, pp. 108–119

Dussán, K.J., Silva, D.D.V., Moraes, E.J.C., Aruda P.V., Felipe, M.G.A., 2014. Dilute-acid Hydrolysis of Cellulose to Glucose from Sugarcane Bagasse. Chemical Engineering Transactions, Volume 38, pp. 433–438

Ferreira, S., Duarte, A.P., Ribeiro, M.H.L., Queiroz, J.A., Domingues, F.C., 2009. Response Surface Optimization of Enzymatic Hydrolysis of Cistus ladanifer and Cytisus striatus for Bioethanol Production. Biochemical Engineering Journal, Volume 45(3), pp. 192–200

Goyal, M., Kalra, K.L., Sareen, V.K., Soni, G., 2008. Xylanase Production with Xylan Rich Lignocellulosic Wastes by a Local Soil Isolate of Trichoderma viride. Brazilian Journal of Microbiology, Volume 39(3), pp. 535–541

Gummadi, S.N., Kumar, D.S., 2005. Microbial Pectic Transeliminases. Biotechnology Letters, Volume 27(7), pp. 451–458

Guo, P., Mochidzuki, K., Cheng, W., Zhou, M., Gao, H., Zheng, D., Wang, X., 2011. Effects of Different Pretreatment Strategies on Corn Stalk Acidogenic Fermentation using a Microbial Consortium. Bioresource Technology, Volume 102(16), pp. 7526–7531

Haryuni, Dewi, T.S.K., Suprapti, E., Rahman, S.F., Gozan, M., 2019. The Effect of Beauveria bassiana on The Effectiveness of Nicotiana tabacum Extract as Biopesticide Against Hypothenemus hampei to Robusta Coffee. International Journal of Technology, Volume 10(1), pp. 159–166

Ibrahim, S., Shukor, M.Y., Syed, M.A., Rahman, N.A.A., Khalil, K.A., Khalid, A., Ahmad, S.A., 2014. Bacterial Degradation of Caffeine: A Review. Asian Journal of Plant Biology, Volume 2(1), pp. 18–27

Iveti?, D.T., Š?iban, M.B., Antov, M.G., 2014. Enzymatic Hydrolysis of Pretreated Sugar Beet Shreds: Statistical Modeling of the Experimental Results. Biomass and Bioenergy, Volume 47, pp. 387–394

Juliastuti, S.R., Widjaja, T., Altway, A., Iswanto, T., 2017. Biogas Production from Pretreated Coffee-pulp Waste by Mixture of Cow Dung and Rumen Fluid in Co-digestion. In: AIP Conference Proceedings. Volume 1840(1)

Juliastuti, S.R., Widjaja, T., Altway, A., Sari, V.A., Arista, D., Iswanto, T., 2018. The Effects of Microorganism on Coffee Pulp Pretreatment as a Source of Biogas Production. In: MATEC Web of Conferences, Volume 156, pp. 1–7

Kumar, P., Barrett, D.M., Delwiche, M.J., Stroeve, P., 2009. Methods for Pretreatment of Lignocellulosic Biomass for Efficient Hydrolysis and Biofuel Production. Industrial and Engineering Chemistry Research, Volume 48(8), pp. 3713–3729

Kusrini, E., Wicaksono, W., Gunawan, C., Daud, N.Z.A., Usman, A., 2018. Kinetics, Mechanism, and Thermodynamics of Lanthanum Adsorption on Pectin Extracted from Durian Rind. Journal of Environmental Chemical Engineering, Volume 6(5), pp. 6580–6588

Li, J., Li, S., Fan, C., Yan, Z., 2012. The Mechanism of Poly(ethylene glycol) 4000 Effect on Enzymatic Hydrolysis of Lignocellulose. Colloids Surfaces B: Biointerfaces, Volume 89, pp. 203–210

Menezes, E.G.T., Carmo, J.R.C., Alves, J.G.L.F., Menezes, A.G.T., Guimarães, I.C., Queiroz, F., Pimenta, C.J., 2014. Optimization of Alkaline Pretreatment of Coffee Pulp for Production of Bioethanol. Biotechnology Progress, Volume 30(2), pp. 451–462

Menon, V., Rao, M., 2012. Trends in Bioconversion of Lignocellulose: Biofuels, Platform Chemicals & Biorefinery Concept. Progress in Energy and Combustion Science, Volume 38(4), pp. 522–550

Mood, S.H., Golfeshan, A.H., Tabatabaei, M., Jouzani, G.S., Najafi, G.H., Gholami, Ardjmand, M., 2013. Lignocellulosic Biomass to Bioethanol, A Comprehensive Review with a Focus on Pretreatment. Renewable and Sustainable Energy Reviews, Volume 27, pp. 77–93

Nathan, V.K., Rani, M.E., Rathinasamy, G., Dhiraviam, K.N., Jayavel, S., 2014. Process Optimization and Production Kinetics for Cellulase Production by Trichoderma viride VKF3. SpringerPlus, Volume 3(92), pp. 1–12

Ochiai, A., Itoh, T., Kawamata, A., Hashimoto, W., Murata, K., 2007. Plant Cell Wall Degradation by Saprophytic Bacillus subtilis Strains: Gene Clusters Responsible for Rhamnogalacturonan Depolymerization. Applied and Environmental Microbiology, Volume 73(12), pp. 3803–3813

Pandey, A., Soccol, C.R., Nigam, P., Brand, D., Mohan, R., Roussos, S., 2000. Biotechnological Potential of Co?ee Pulp and Co?ee Husk for Bioprocesses. Biochemical Engineering Journal, Volume 6(2), pp. 153–162

Park, S., Baker, J.O., Himmel, M.E., Parilla, P.A., Johnson, D.K., 2010. Cellulose Crystallinity Index: Measurement Techniques and their Impact on Interpreting Cellulase Performance. Biotechnology for Biofuels, Volume 3(10), pp. 1–10

Ranganna, S., 1979. Manual of Analysis of Fruit and Vegetable Products. Tata McGraw-Hill Publ Co Ltd, New Delhi

Roda, A., De Faveri, D.M., Dordoni, R., Lambri, M., 2014. Vinegar Production from Pineapple Wastes – Preliminary Saccharification Trials. Chemical Engineering Transactions, Volume 37, pp. 607–612

Roussos, S., Aquiáhuatl, M.D.L.A, Trejo-Hernández, M.D.R, Perraud, I.G., Favela, E., Ramakrishna, M., Raimbault, M., Viniegra-Gonzalez, G., 1995. Biotechnological Management of Coffee Pulp - Isolation, Screening, Characterization, Selection of Caffeine-degrading Fungi and Natural Microflora Present in Coffee Pulp and Husk. Applied Microbiology and Biotechnology, Volume 42(5), pp. 756–762

Seiboth, B., Ivanova, C., Seiboth, V.S., 2011. Trichoderma reesei: A Fungal Enzyme Producer for Cellulosic Biofuels. In: Biofuel Production-Recent Developments and Prospects, Vienna University of Technology, Austria, pp. 309–340

Sohail, M., Siddiqi, R., Ahmad, A., Khan, S.A., 2009. Cellulase Production from Aspergillus niger MS82: Effect of Temperature and pH. New Biotechnology, Volume 25(6), pp. 437–441

Sun, Y., Cheng, J., 2002. Hydrolysis of Lignocellulosic Materials for Ethanol Production: A Review. Bioresource Technology, Volume 83(1), pp. 1–11

Torimiro, N., Okonji, E.R., 2013. A Comparative Study of Pectinolytic Enzyme Production by Bacillus Species.  African Journal of Biotechnology, Volume 12(46), pp. 6498–6503

Widjaja, T., Iswanto, T., Altway, A., Shovitri, M., Juliastuti, S.R., 2017a. Methane Production from Coffee Pulp by Microorganism of Rumen Fluid and Cow Dung in Co-digestion.  Chemical Engineering Transactions, Volume 56, pp. 1465–1470

Widjaja, T., Iswanto, T., Agustiani, E., Altway, A., Silaban, B.M.J., Yuwono, L.F., 2017b. Optimization of Palmyra Palmsap Fermentation using Co-culture of Saccharomyces cerevisiae and Pichia stipitis. ARPN Journal of Engineering and Applied Sciences, Volume 12(23), pp. 6817–6824