(477e) Fractionation of Cowpea Shells (Vigna unguiculata) Using Natural Deep Eutectic Solvent (Lactic acid-Proline)

For sustainability in biomass conversion, it is highly recommended to adopt pretreatment methods that do not threaten the environment. Natural deep eutectic solvents are made from substances found in plant cells Choi et al.(2011) and are environmentally friendly. They have been studied as a chemical route for biomass pretreatment because of their high solubilisation power and tunability (Abbott et al., 2011). These solvents are also easily produced by combining hydrogen bond donors and bond acceptors commonly found in plants in specific molar ratios.

This study applied a natural deep eutectic solvent that combined lactic acid and proline to fractionate cowpea shells. Cowpea (Vigna unguiculata) is one of the major sources of plant protein and generates a lot of waste from the deshelling process. According to FAO (2021), 8.9 million tonnes of cowpea were produced globally in 2020, of which Africa accounts for 97 %. Nigeria and the Republic of the Niger as the highest producers: 42 % and 31 %, respectively. A study by Jekayinfa et al. (2020) estimated that tonnes of cowpea shells are generated yearly in Nigeria and can serve as feedstock for biorefineries.

Factors such as temperature, time, and water dilution have been established as some of the factors that affect the use of NADES in biorefinery (New et al., 2019). The addition of water to solvents is done for several reasons, including reducing viscosity (Dai, Witkamp, Verpoorte, & Choi, 2015), to inhibit the solubility of cellulose. (Swatloski, Spear, Holbrey, & Rogers, 2002) and to reduce the volume of solvent used in a process (New et al., 2019). However, discrepancies in the research findings of different authors (Kumar, Parikh, & Pravakar, 2016; Li, Hou, Lin, Zhang, & Fu, 2018) on the effect of these factors necessitate this study.

Therefore, this work aims to apply NADES as a solvent for fractionating cowpea shells. The main objectives are to analyse the interactions between factors and to determine the optimal operating conditions using a central composite design of the response surface methodology. To the best of the authors knowledge, the effect of the interaction between temperature, time and water dilution on the fractionation of cowpea shells has not been explored. This study will provide more insight into applying NADES to pretreat cowpea shells.

Materials and Methods

The cowpea shells (Vigna unguiculata shells) were obtained from farms in Tunga in Chanchaga Local Government Area, Minna, Niger State, Nigeria and preserved in sample bags. Lactic acid (90 %) was purchased from Thermo Scientific (USA). Proline (99 %), Sulphuric acid (98 %), D- (+)-glucose and D- (+) -xylose were purchased from Sigma Aldrich Company Ltd, Germany.

The substrate was washed with deionised water and sundried for two days before milling with a Retsch PM 100 Planetary ball mill (Haan, Germany) and sieved with a 425 µm laboratory test before storage.

Lactic acid and proline were combined in a molar ratio of 4: 1 (Francisco, Van Den Bruinhorst, & Kroon, 2012) using the protocol described by Dai et al. (2013). The density, conductivity, and viscosity of the prepared NADES were analysed using the Attension Sigma force tensiometer (Sigma 700, Biolin Scientific USA), Jenco instruments (Model: 3020M) and RV-Viscometer (Model: NDJ-8S; W&J Instrument Co., Ltd. China), respectively.

The substrate was combined with the solvent at a 5: 100 (w/w), as determined by Kumar et al. (2016), in a capped Schott bottle and placed in a high-temperature bath circulator (Daihan Scientific, Korea). At the end of the experiment, the solid was recovered from the slurry based on the procedure reported by Procentese et al. (2015) and Li et al. (2018). The recovered solid residue was dried at 35 °C and analysed using the National Renewable Energy Laboratory (NREL) method (Sluiter et al., 2011) for its carbohydrate composition (glucan and xylan) and lignin composition. The upper and lower limits for the central composite design (CCD) were chosen based on the preliminary experimental results carried out. Temperature range (130 -150) ⁰C, time (4 -6) h and percentage water dilution of solvent (1.5 – 3.5) % (w/w). The results were analysed using Design Expert software 13. All experiments were carried out in duplicates.

Results

The viscosity, conductivity, and density of the lactic acid-Proline solvent (4:1) prepared were recorded as 638.75 ± 0.35 mPa.s, 195.5 ± 0.28 µS/cm, and 1.26 ± 0.04 g/ml, respectively. The chemical composition of the cowpea shell was determined to comprise glucan (21.32 ± 0.39 %), xylan (21.46 ± 0.29 %), acid-insoluble lignin (28.40 ± 0.01 %), acid-soluble lignin (8.63 ± 0.02 %), extractives (2.10 ± 0.14 %), moisture (5.96 ± 0.65 %), Ash (1.05 ± 0.14 %). The central composite design was applied with design Expert 13 to obtain the quadratic model equation relating independent variables to the percentage responses measured ( glucan, xylan and lignin). The statistical tests (ANOVA) were performed on the quadratic models to assess their accuracy based on the probability value (P) > 0.05, R-square (determination coefficient), adjusted R-square and predicted R-square. Percentage glucan yield in pretreated cowpea shell is significantly affected by the interactions between temperature and water dilution, time and water dilution. The interaction between temperature and water dilution significantly affects the xylan percentage composition of the pretreated sample. Lignin content percentage showed that interaction between temperature and time, temperature and water dilution, and time and water dilution were also statistically significant. These results imply that water dilution is key in lactic-proline solvent biomass pretreatment application. The process was optimised by applying the numerical method to obtain a solution satisfying the three quadratic equations by maximising glucan and xylan and minimising lignin. The optimal condition was 38.4 % glucan, 24.8 % xylan and 27.06 % total lignin at temperature: 146.4 ℃, time: 6 h and water dilution at 1.5 % w/w. The models were validated by experimenting at optimal conditions. From the experimental results obtained, % glucan: 38.48 ± 0.21, % xylan: 24.66 ± 1.46 and % total lignin: 27.27 ± 1.24 shows that the model and the experimental values are in good agreement.

Understanding the effect of variation of factors on the ability of Lactic-Proline, natural deep eutectic on the fractionation of cowpea shells plays a crucial role in its industrial application. This study shows that NADES can be applied for the pretreatment of cowpea shells.

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