Subcritical water hydrolysis of brewer’s spent grains: Selective production of hemicellulosic sugars (C-5 sugars)

The Journal of Supercritical Fluids 145 (2019) 19–30

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Subcritical water hydrolysis of brewer’s spent grains: Selective production of hemicellulosic sugars (C-5 sugars)

T

P.C. Torres-Mayangaa, S.P.H. Azambujaa, M. Tyufekchievb, G.A. Tompsettb, M.T. Timkob, ⁎ R. Goldbecka, M.A. Rostagnoc, T. Forster-Carneiroa, a

School of Food Engineering, University of Campinas (UNICAMP), Rua Monteiro Lobato, n.80, 13083-862 Campinas, SP, Brazil Department of Chemical Engineering, Worcester Polytechnic Institute, 100 Institute Road, Goddard Hall 123, Worcester, MA, 01609, United States c School of Applied Sciences, University of Campinas (UNICAMP), Rua Pedro Zaccaria, n. 1300, 13484-350, Limeira, SP, Brazil b

GRAPHICAL ABSTRACT

ARTICLE INFO

ABSTRACT

Keywords: Supercritical technology Semi-continuous Flow-through hydrolysis reactor Fourier transform infrared spectroscopy Thermogravimetric analysis

The objective of this work was to produce C-5 sugars from the hydrolysis of the spent grains of the brewery. The total sugar recovery and formation of by-products were evaluated as a function of the reaction temperature (140, 160, 180 and 210 °C), flow rate (10 and 20 mL min–1) and the solvent/feed ratio (S/F: 64, 80 and 112). The total carbohydrate yields were dependent on the hydrolysis temperature. Arabinose was the main product, followed by xylose and the arabinose yield increased with increasing the S/F ratio and the flow rate. Yields of furfural were sensitive to flow rate. The hemicellulose content of the residual solids was reduced by approximately 90% after hydrolysis, with the parallel formation of carbonyl and carbonized species. The effects of flow rate on the efficiency of converting hemicellulose into simple sugars were complex, suggesting roles for the formation of autocatalytic acids.

1. Introduction The massive scale of agricultural waste generation makes imperative its environmentally responsible disposal or reuse. One of the main types of agricultural waste is the biomass plant matrix itself, for

⁎

example stalks, straws, leaves, and pods. Collectively, these residues are composed primarily of cellulose, hemicellulose, and lignin biopolymers [1–3]. While a portion of these residues can be left on the field after harvesting, these valuable biopolymers represent a potential source of renewable chemical and energy products that should be examined

Corresponding author. E-mail address: [email protected] (T. Forster-Carneiro).

https://doi.org/10.1016/j.supflu.2018.11.019 Received 26 September 2018; Received in revised form 19 November 2018; Accepted 20 November 2018 Available online 23 November 2018 0896-8446/ © 2018 Elsevier B.V. All rights reserved.

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(approximately 9 min at 10 mL min–1 compared to 4.5 min at 20 mL min–1) on sugar yields and selectivity. Finally, S/F was studied at 64, 80, and 112 g of solvent per gram of biomass to study the effects of dilution. These results guide future work in this area to help spur further development of flow-through subcritical water hydrolysis as a green biomass fractionation technology.

thoroughly to identify economically viable alternatives to burning in place and other disposal methods. An important by-product of the beer industry is brewer´s spent grain (barley), which represents approximately 85% of the raw material used in the process [4,5]. Brewer´s spent grains (BSG) is rich in protein and other organic biopolymers, with major components being cellulose (12–23%), hemicellulose (15–29%) and lignin (19–30%) [6–10]. In all plants, hemicellulose is a branched, heteropolymer consisting of C6 (hexose) and C5 (pentose) sugars, as well as abundant acetyl side chains [11,12], making hemicellulose a potential source of simple sugars and valuable byproducts such as furfural [12]. However, the ratio of different sugars contained within hemicellulose varies with the source, as xylans dominate the hemicellulose of woody biomass whereas grains is composed primarily of arabinoxylans, with greater arabinose content than xylan content [13,14]. The hemicellulose contained in grains can be used in a range of different bioprocess applications, such as in alcoholic fermentation [15], and in the production of lactic acid and xylitol by the action of microorganisms in the hemicellulosic hydrolysate [16–18]. The separation of the lignocellulosic complex into its components remains a technological barrier to economic utilization of biomass. Hemicellulose, cellulose, and lignin interact with one another physically and chemically to form a rigid complex [19]. Although methods have been proposed for fractionating biomass into subcomponents [20], none have yet proven economically viable at commercial scale. Among the options, subcritical water hydrolysis of lignocellulosic waste has potential as a green technology for the production of soluble chemical products, which can serve as feedstocks for biofuel and bioenergy sources, chemical products, and renewable materials [21–25]. In the subcritical state, water is a promising reaction solvent for hydrolytic depolymerization of biomass to produce soluble products [22] that does not require use of organic solvents or additional catalytic materials [26]. A natural consequence of the tunable properties of subcritical water is that the individual components of the lignocellulosic complex are converted to soluble products over distinct temperature ranges, suggesting selective optimization of hydrolysis conditions for each biopolymer. Hemicellulose, in particular, is solubilized at lower temperature (150–230 °C) than either cellulose or lignin [27,28]. A major challenge to any process that aims to fractionate lignocellulosic biomass into individual sugars is that the degradation reactivity of the sugar products is similar to, or even greater than, the hydrolysis reactivity of the parent polymers [29,30]. As a result, polymer hydrolysis and sugar degradation naturally occur in parallel at comparable rates [31]. At the extreme, accumulation of degradation products results in formation of char layers that coat the biomass particles, hindering hydrolysis of interior components [32,33]. Recent work using flow-through subcritical water reactors, in which subcritical water continuously flows over a packed bed consisting of biomass particles, has shown potential for minimizing sugar degradation and char deposition [33]. However, concerns about water and energy use in flow-through subcritical water reactors [34] necessitate careful investigation of the effects of flow rates and the solvent/feed (S/F) ratio used in the process. Moreover, organic acid byproducts, which accumulate to a much greater concentration in batch operation than in flowthrough mode, can act auto-catalytically to promote biopolymer depolymerization [27,35]. Flow-through operation thereby decreases the potential auto-catalytic benefit, making imperative the careful design of flow-through reactors to maximize potential benefits. In this work, production of hemicellulose (C-5 sugar) from brewer´s spent grains (BSG) was investigated in a laboratory-scale, flow-through subcritical water reactor. The aim of this work was to investigate the effects of hydrolysis temperature, flow rate, and S/F on sugar yields and selectivity. Experiments were performed at 140, 160, 180, and 210 °C hydrolysis temperatures, selected to span the range shown most effective for hemicellulose hydrolysis [30,36]. Flow rate was studied at 10 and 20 mL min–1 to investigate the effects of water residence time

2. Materials and methods 2.1. Raw material AMBEV CSC Brewery (Jaguariúna, São Paulo, Brazil) provided the brewer´s spent grains (BSG) Before use, the particle size was reduced using a knife mill (Marconi, model MA 340, Piracicaba, Brazil) equipped with a 1 mm sieve to produce particles in the 180 and 310 μm size range; this size range was selected to balance concerns about particle spillover (which is minimized with increasing particle size) and heat/mass transfer limitations (which are minimized with decreasing particle size) [37]. The milled brewer´s spent grains (BSG) were stored at –20 °C until use. 2.2. Chemicals Acetic acid, ethanol, sodium chlorite, and sodium hydroxide were obtained from Wako Pure Chem. Ind., Ltd., Osaka, Japan. Standards of L-(+)-arabinose, D-(+)-galactose, D-(+)-glucose, D-(+)-xylose, D(-)-fructose, D-(+)-cellobiose, sucrose, and maltose (all with > 99% purity), as well as 5-(hydroxymethyl)furfural (≥ 99%), furfural (≥ 98%), acetic acid (≥ 98%), and formic acid (≥ 97%) were obtained from Sigma Aldrich (St. Louis, MO, USA) and used as high pressure liquid chromatograph (HPLC) standards. Ultrapure water was used for standard preparation. 2.3. Characterization and chemical composition of the brewer´s spent grains Moisture, total solids, ash, total extractives, carbohydrates, lignin and protein analyses were performed using methodologies recommended by the National Renewable Energy Laboratory (NREL) [38–42]. Holocellulose content was determined gravimetrically after delignification with sodium hypochlorite (NaClO2) [43]. For the analysis, 2.5 g of brewer´s spent grains (BSG) was added to 150 mL of water at 75 °C, containing 0.2 mL of glacial acetic acid and 2.0 g of NaClO2. After 3 h, additional aliquots of glacial acetic acid (0.2 mL) and NaClO2 (1.0 g) were added every hour. The suspension was vacuum filtered with water and acetone. The delignified fraction was dried in an oven at 105 °C for 24 h, producing a white powder identified as holocellulose. The α-cellulose content of the holocellulose fraction was determined by adding 1 g of holocellulose to a 17.5% NaOH solution, which was stirred for 40 min at 20 °C and then vacuum filtered. The filtered residue was washed with 40 mL of a 10% glacial acetic acid solution and 1 L of boiling water in two different stages. The filtrate was oven dried at 105 °C for 24 h and weighed [44]. The hemicellulose content was determined by the difference, using these equations:

%

Cellulose =

WeightCellulose 1g

x

Weighthollocelulose Weightfree extractives sample

x 100% (1)

% Hemicellulose =

Weighthollocelulose Weightfree extractives sample

x 100%

(%

Cellulose ) (2)

All determinations were performed in triplicate. 20

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Fig. 1. Schematic diagram of the subcritical water technology unit: W- Water reservoir; B- HPLC Pump; V-1 - 3: Block valves; M1- Manometer; T- Thermocouples; FFilter; R1- reactor; HE- Heat exchanger; BPR- Back Pressure Regulator Valve; CV- Collecting vessel.

2.4. Semi-continuous subcritical water hydrolysis

performed, during which hydrolysate samples were collected over time, corresponding to S/F ratios between 8 and 112. The total reducing sugars, specific sugar concentrations, degradation products, and organic acids present in the hydrolysates were quantified at S/F ratios of 64, 80 and 112. Temperature, residence time, and S/F all contribute to biopolymer hydrolysis and sugar degradation. In a reactor with flow-through, the relevant residence time is determined by the flow rate and water density, which in turn depends on hydrolysis temperature [27,45]. Table 1 shows the values of residence time (τ) and severity factor (R0), which are directly related to one another through reaction temperature and are useful variables for describing the sugar yields obtained from subcritical water hydrolysis. The residence time of the fluid was estimated with the following Eq. (3), following the work of Mayanga -Torres et al. [46]:

Fig. 1 shows the schematic diagram of the experimental system used for semi-continuous, sub-critical water hydrolysis experiments under flow-through conditions. The reactor itself is constructed of 316-stainless steel with tubular geometry, with an internal volume of 110 mL, a maximum working temperature of 400 °C, and a maximum working pressure of 40 MPa. The reactor is heated by a heating jacket, capable of delivering 1500 W. The reaction temperature was monitored by thermocouples (type K) located at the outlet of the reactor. A high-pressure liquid pump (double piston pump, Model 36 preparation pump, Apple Valley, MN, USA) was used for pressurization and liquid pumping. The pressure in the reactor was controlled by a micrometric valve located at the heat exchanger outlet. An in-line porous stainless steel filter with 2 μm pores was positioned at the outlet of the reactor to minimize particle spillover. The hot effluent exiting the hydrolysis reactor was cooled to less than 27 °C in a tube heat exchanger coupled to a recirculating bath (Marconi, model MA-184, Piracicaba, SP, Brazil). The heat exchanger working fluid was distilled water, and its temperature was controlled at 5 °C. Hydrolysate was sampled periodically at the outlet of the pressure valve. Subcritical water hydrolysis experiments were performed at a constant pressure of 15 MPa. This pressure was selected to ensure liquidphase operation at all times; previous work indicates that reaction pressure is a minor variable in subcritical water hydrolysis, provided that the pressure remains above the vaporization point. For each experiment, approximately 5 g of raw material were placed in the reactor. Hydrolysis experiments were performed over a range of temperatures (140–210 °C) and water flow rates (10 and 20 mL min–1), as presented in Table 1. All experiments presented here were performed in duplicate and ranges are presented as uncertainties. Kinetic rate tests were

=

140 °C, 160 °C, 180 °C, 210 °C, 140 °C, 160 °C, 180 °C, 210 °C, a b c d

10 mL 10 mL 10 mL 10 mL 20 mL 20 mL 20 mL 20 mL

min−1 min−1 min−1 min−1 min−1 min−1 min−1 min−1

τ

a

9.5 9.3 9.1 8.8 4.8 4.7 4.6 4.4

log(R0) 2.16 2.74 3.32 4.19 1.86 2.44 3.02 3.89

b

Densityc

S/Fd

934.0 915.9 896.1 862.8 934.0 915.9 896.1 862.8

64, 80 and 112

R (T , P ) v0 0

(3) 3

where VR is the reactor volume (m ), vo is the volumetric water flow rate (m3/s), ρ0 is the density of the liquid feed (1080 kg/m3), and ρR is the density of the fluid under reactor temperature and pressure, which was estimated using steam tables [47]. The severity factor, R0, depends on the temperature and residence time and was calculated using the following equation, following the work of Lachos-Perez et al. [33]:

R 0 = * exp

T 100 14.75

(4)

Where: T is the hydrolysis temperature in °C, and 14.74 is an empirical parameter related to the activation energy, assuming pseudo-first order kinetics. Severity factor is presented in base 10 logarithm units, i.e., logR0.

Table 1 Experimental conditions used for hemicellulosic sugars extraction. Experimental conditions

VR

2.5. Analytical methods 2.5.1. pH, reducing sugars, and total reducing sugars The pH of the hydrolysate was determined at ambient temperature using a digital pH meter (Digimed, model DM-22, Brazil). The reducing sugars (RS) and total reducing sugars (TRS) content of the hydrolysate were determined by the Somogyi-Nelson colorimetric method [48,49]. For TRS measurements, the hydrolysate was first subjected to acid hydrolysis to convert sugar oligomers into monomeric form. After the coloring reaction, the absorbance of the sample was measured at a wavelength of 540 nm using a spectrophotometer (Hach, model DR/ 4000U, Loveland, CO, USA). The TRS concentrations were calculated using a calibration curve based on standard glucose solutions (0.05 – 0.50 g L–1). TRS and RS yields are expressed as glucose equivalents

64, 80 and 112

Residence time value (min). Severity factor value. kg/m3. g solvent/g feed value. 21

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[46]. In some cases, hydrolysate samples were diluted with deionized water before photometric analysis to ensure that the measured absorbance fell within the calibration range.

Table 2 Proximate analysis of brewer´s spent grains in percentages by dry weight (average ± standard deviation).

2.5.2. Analysis of monosaccharides by high-performance liquid chromatography (HPLC) Identification and quantification of arabinose, galactose, glucose, xylose, fructose, and sucrose were performed by HPLC on a DIONEX DX - 500 system (Sunnyvale, CA, USA), equipped with an electrochemical detector (ED - 40). Separation was carried out on a Carbopac PA-1 column (4 x 250 mm) using a two-solvent gradient: (A) 150 mM NaOH solution and (B) 100 mM NaOH solution containing 500 mM sodium acetate with a flow of 1 mL min–1. Retention times and calibration factors were determined using analytical standards. The limits of the curve were [0.01 – 0.10 g/L] with an R2Arabinose = 0.9703, R2Galactose = 0.9414 R2Xylose = 0.9806, R2Glucose = 0.9612, R2Fructose = 0.9885 and with an R2Sucrose = 0.9754.

Component

%

Moisture Ash Protein Extractives in water Extractives in ethanol Soluble lignin Insoluble lignin Cellulose Hemicellulose

4.8 ± 0.1 3.9 ± 0.2 19.2 ± 0.2 5.7 ± 0.3 13.6 ± 0.2 6.1 ± 0.4 11.7 ± 0.1 17.90 ± 0.02 35.70 ± 0.02

3. Results and discussion Table 2 shows the initial characterization of brewer´s spent grains (BSG) used as raw material. The ash content of the BSG is comparable to previously reported values, which ranged from 1.2 to 6.4% [50–52]. The protein content in the BSG was 19.2%, which is similar to those reported by Linan-Montes et al. [53] and Poerschmann et al. [54] but 4.5 to 6.6% greater than those reported by Mussatto et al. [9], and Mello and Mali [52]. The protein content of BSG varies due to differences in process factors, including the use of additives and supplements, grinding, and crushing processes [55], and the discrepancy with prior analysis is consistent with this natural variability. Of the main biopolymers present in BSG, hemicellulose is abundant and cellulose and lignin are present in smaller quantities. The hemicellulose content of the sample was 35.7%, greater than previous reports, which vary between 22.46 and 28.97% [51–53,56]. The cellulose and lignin contents were 17.9% and 17.8%, respectively. Differences in the composition of the BSG may be caused by several factors, such as species, time of harvesting, differences in the malting process, and differences in the conditions used during production of beer [54,57]. The hemicellulose content of the sample used in this work suggests it as an especially attractive source of C5 sugars.

2.5.3. Analysis of 5-Hydroxymetilfurfural (5-HMF) and furfural (FF) by HPLC The 5-HMF and furfural contents of hydrolysate samples were measured on an HPLC model Accela Autosampler/Accela Pump from Thermo Scientific (Thermo Scientific, San Jose, CA, USA). The separation was performed using a column packed with core-shell particles (Kinetex C18, Phenomenex) (Phenomenex, Torrance, CA, USA). A solution consisting of acetonitrile (12.5%) and water with 1% acetic acid (87.5%) was the mobile phase, and the mobile phase flow rate was 0.3 mL min–1. The total run time was 6 min and detection was by absorbance at 274 nm. Retention times and calibration factors were determined using standard solutions of 5-HMF and furfural. The limits of the curve were [0.005 – 0.020 g/L] with an R2HMF = 0.9880 and with an R2FF = 0.9743. 2.5.4. Analysis of organic acids by HPLC Acetic and formic acid concentrations were measured using HPLC on a Varian chromatography system (Varian Inc., Palo alto, CA, USA) equipped with a refractive index detector (RI 2000, Chrom Tech Inc., Dorfen, Germany). Separation was achieved on an Aminex HPX-87H column (BioRad, Hercules, CA, USA), maintained at 35 °C. An aqueous solution of H2SO4 at a pH of 2.6 was used as mobile phase, with a flow of 0.6 mL min–1. Concentrations were determined by refractive index measurements that were made at a wavelength of 205 nm. Retention times and calibration factors were determined using standard solutions of acetic acid and formic acid. The limits of the curve were [0.1–10 g/L] with an R2acetic acid = 0.9989 and with an R2formic acid = 0.9985.

3.1. Reducing sugars and total reducing sugars in the hydrolysate BSG was treated under flow-through conditions in subcritical water over a range of hydrolysis temperatures and at flow rates of 10 and 20 mL min–1. The high percentage of hemicellulose (35.7%) present in the BSG under study suggests its use as a potential source of sugars. Accordingly, hydrolysis temperatures were selected to maximize hemicellulose hydrolysis, i.e., 140–210 °C. Hydrolysate was analyzed for RS and TRS yields, yields of specific sugars, and yields of specific decomposition products. In all cases, yields are presented per 100 g of the initial feed. Lastly, the residual solids were analyzed using TGA and FTIR to increase understanding of the hydrolysis process. Fig. 2 provides RS and TRS yield data, showing that yields increase with increasing hydrolysis temperature and, to a lesser extent, increasing S/F ratio. Interestingly, RS yields increase rapidly with increasing S/F up to a value of approximately 64, at which point further increases in S/F yield diminishing returns and potentially suggesting that the majority of accessible hemicellulose is extracted at this S/F condition. Moreover, increasing hydrolysis times and/or S/F beyond the point of diminishing returns has the negative impact of increasing water use and reducing sugar concentrations. The maximum RS yields were approximately 6.4 g RS / 100 g BSG, measured at 210 °C and independent of flow rate. Similarly, the maximum TRS yields were 39.8 ± 0.9 and 38.3 ± 0.4 g TRS / 100 g. Varying flow rate exerted a minor effect on RS and TRS yields, at least when data are plotted in terms of S/F ratio, and only at mild hydrolysis conditions of 140 and 160 °C. This suggests that thermal effects dominate at temperatures of 160 °C and greater. Instead, the benefit of increasing flow rate would be

2.5.5. Infrared spectroscopy, raman spectroscopy and thermogravimetric analysis Fourier transform infrared spectroscopy (FTIR) and thermogravimetric analysis (TGA) were used to investigate the chemical composition of treated bagasse samples. FTIR spectra were obtained using a Bruker Vertex 70 FTIR spectrometer equipped with a La-DTGS detector operated at room temperature. A diamond attenuated total reflectance (ATR) cell, “Golden-Gate” manufactured by Specac, was used for all measurements. The resolution was 4 cm−1, and 512 scans were acquired and then averaged over the 600–4000 cm−1 spectral range. MagicPlot software was used to peak fit the infrared bands. A Netzsch thermogravimetric analyzer (TG 209 F1 Libra) was used for analysis of biomass materials and hydrolysis products. An alumina crucible was used for holding samples. The nitrogen flow was constant at 20 mL min−1. The initial oven temperature was 35 °C and it was increased to 800 °C at a constant rate of 5 °C min−1. Vendor software converted raw data into differential thermograms (DTG). MagicPlot software was used to fit peaks to the DTG curves and the composition of the materials was estimated from the peak areas. 22

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Fig. 2. The RS and TRS yield at different S/F in the experimental kinetics at 10 mL min−1: a) RS and b) TRS. At 20 mL min−1 c) RS and d) TRS. The columns presented the mean value with their respective uncertainty obtained from the hydrolysis and duplication analysis.

to decrease processing times, since doubling the flow rates allows the same S/F to be reached in half the time. Comparison of Fig. 2a and c with b and d shows that TRS yields were between 5- and 15-times greater than RS yields, with the greatest difference occurring at the lowest hydrolysis temperature (140 °C). The difference between TRS and RS yields indicates that the majority of hydrolysis products were formed as oligomeric sugars. Because the oligomeric sugar products of hemicellulose hydrolysis are likely to be soluble in subcritical water, they are removed as primary products before they hydrolyze to form simple sugars and/or degrade to form other byproducts. The fact that the difference between TRS and RS yields decreases with increasing temperature points to the effect of thermal conditions on oligomer hydrolysis. Hydrolysis results in Fig. 2 indicate that reaction temperature is the most important factor governing the solubilization and hydrolysis of the carbohydrates contained in the BSG. Increasing the temperature of the subcritical water treatment has many potential effects on hydrolysis rates and mechanisms. Most importantly, the Arrhenius effect is associated with increasing treatment temperature; for a representative activation energy for glycosidic bond hydrolysis of 80 kJ mol–1 [58], the effect of increasing the hydrolysis temperature from 140 to 160 °C is a three-fold increase in hydrolysis rate. Accordingly, treatment temperature controls reaction rates, both for hydrolysis and the conversion of oligomers and dimers into simple sugars [59]. In addition to the Arrhenius effect, the physicochemical properties of subcritical water are sensitive to temperature [45], and the temperature variation of these properties can influence relative hydrolysis and solubilization rates. Specifically, the dielectric constant and fluid density decrease with increasing temperature over the range from 140 to 210 °C, favoring the solubilization of non-polar organic compounds [60]. On the other hand, the auto-ionization constants increases over the same range, generating concentrations of H+ and OH– ions by a factor of about 10 relative to room temperature [33,61] and promoting rates of both acid and base catalyzed reactions such as glycosidic bond hydrolysis [30]. As a result, subcritical water has potential to act as an acid/ base catalyst as well as reactant in biopolymer hydrolysis [22]. Residence time is an important factor affecting RS and TRS yields under batch conditions. Under flow-through conditions, residence time can refer either to the total process time (i.e., the residence time experienced by the solids) or the fluid residence time (i.e., the residence

time experienced by the solvent and any soluble products). The fluid residence time is directly related to the flow rate, previous studies have reported that significant increases in sugar yields could be obtained by increasing water flow rate [60,62]. However in this study of BSG, both RS and TRS yields were similar at 10 and 20 mL min–1, suggesting that biopolymer hydrolysis to form soluble oligomers is more rapid than subsequent hydrolysis of oligomers to form simple sugars. 3.2. Quantification of monosaccharides in the hydrolysate RS and TRS yields are a convenient way to quantify hydrolysis. However, they are imperfect measurements and do not provide information on the source of the sugar, whether it be cellulose or hemicellulose. The lower thermal stability of hemicellulose compared to cellulose suggests it as the more likely source of sugars [63,64]. Because cellulose consists exclusively of glucose and hemicellulose is a heteropolymer consisting primarily of C5 sugars with some C6 sugar content, quantification of individual sugars in the hydrolysate can be used to confirm their source. Results are summarized in Table 3. As expected, the C5 sugars arabinose and xylose constitute > 75% of the sugars quantified in the hydrolysate, with minor amounts of galactose, glucose, and fructose also present. This confirms preferential solubilization of hemicellulose (composed mainly of arabinoxylans) [14] during the hydrothermal treatment. The temperature dependence of individual sugar yields is instructive. Sugar production was observed at the lowest temperature of 140 °C, at which condition arabinose and xylose represented as much as 88% of the total quantified sugar yield. All sugar yields generally increased with increasing hydrolysis temperature; however, on a percentage basis, the yields of xylose, galactose, glucose, and fructose were most sensitive to temperature, consistent with greater activation energies for hydrolysis and solubilization of these sugars compared to arabinose. Arabinose yield was optimized at temperatures of 160 °C and 180 °C, with maximum yields of 3.13 ± 0.13 and 2.92 ± 0.02 g/ 100 g, observed at an S/F ratio of 112 and at flow rates of 10 at 20 mL min–1, respectively. Interestingly, arabinose yields decreased when treatment temperatures are increased from 180 to 210 °C, suggestive of degradation which flow-through operation was intended to decrease. Accordingly, the maximum arabinose yield is greater at 10 than at 23

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Table 3 The sugars profile composition of brewer´s spent grains hydrolysates obtained under different experimental conditions (g compound/100 g Feed). Temperature 140 °C

Flow rate 10 mL min

−1

160 °C 180 °C 210 °C 140 °C 160 °C 180 °C 210 °C

20 mL min−1

S/F

Arabinose

Galactose

Glucose

Xylose

Fructose

Sucrose

Total carbohydrates

64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112

1.2 ± 0.1 1.3 ± 0.1 1.6 ± 0.1 2.55 ± 0.04 2.8 ± 0.1 3.1 ± 0.1 2.4 ± 0.1 2.6 ± 0.1 3.1 ± 0.2 1.9 ± 0.2 2.0 ± 0.1 2.0 ± 0.1 1.48 ± 0.01 1.72 ± 0.01 1.3 ± 0.3 1.5 ± 0.1 1.7 ± 0.1 1.8 ± 0.1 2.5 ± 0.1 2.76 ± 0.05 2.92 ± 0.02 2.2 ± 0.1 2.3 ± 0.2 2.3 ± 0.3

n.d n.d n.d 0.13 0.14 0.15 0.14 0.15 0.15 0.16 0.18 0.19 n.d n.d n.d 0.07 0.07 0.07 0.12 0.13 0.14 0.15 0.17 0.16

n.d n.d n.d 0.18 ± 0.02 0.19 ± 0.03 0.21 ± 0.05 0.192 ± 0.02 0.21 ± 0.03 0.20 ± 0.00 0.3 ± 0.1 0.4 ± 0.1 0.5 ± 0.1 n.d n.d n.d 0.11 ± 0.02 0.12 ± 0.02 0.11 ± 0.03 0.2 ± 0.0 0.2 ± 0.0 0.3 ± 0.0 0.25 ± 0.03 0.29 ± 0.02 0.27 ± 0.03

0.3 ± 0.1 0.3 ± 0.1 0.3 ± 0.1 0.9 ± 0.2 0.9 ± 0.2 0.9 ± 0.2 1.1 ± 0.0 1.1 ± 0.1 0.8 ± 0.1 1.6 ± 0.0 1.6 ± 0.1 1.6 ± 0.2 0.4 ± 0.1 0.4 ± 0.1 0.3 ± 0.0 0.38 ± 0.00 0.41 ± 0.01 0.42 ± 0.02 0.8 ± 0.0 0.9 ± 0.0 0.9 ± 0.0 1.1 ± 0.2 1.1 ± 0.1 1.1 ± 0.2

0.07 0.08 n.d 0.14 0.14 0.15 0.16 0.18 0.18 0.33 0.38 0.42 n.d n.d n.d 0.10 0.11 n.d n.d n.d n.d 0.30 0.31 0.30

0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.2 0.2 n.d n.d n.d

1.7 1.8 2.0 4.0 4.2 4.7 4.1 4.3 4.5 4.4 4.7 4.8 2.0 2.2 1.7 2.2 2.5 2.5 3.7 4.2 4.4 4.0 4.2 4.2

± ± ± ± ± ± ± ± ±

0.01 0.01 0.02 0.01 0.01 0.02 0.01 0.01 0.01

± ± ± ± ± ± ± ± ±

0.00 0.00 0.00 0.01 0.00 0.01 0.01 0.01 0.03

± 0.01 ± 0.01 ± ± ± ± ± ± ± ± ±

0.01 0.01 0.01 0.01 0.01 0.02 0.01 0.02 0.02

± 0.00 ± 0.01

± 0.03 ± 0.05 ± 0.04

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0

± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

0.1 0.1 0.1 0.2 0.2 0.2 0.1 0.1 0.2 0.2 0.2 0.2 0.1 0.1 0.3 0.1 0.1 0.1 0.1 0.0 0.0 0.2 0.2 0.4

- n.d: not detected - below detection limits.

20 mL min–1, suggesting that increasing the flow rate and decreasing the fluid residence time can offset the degradation of arabinose observed at 210 °C. Interestingly, xylose yields did not show an optimum at intermediate temperatures but did show a progressive increase. The low yields of xylose may be due to the low proportion of xylose relative to arabinose in the feed. Moreover, differences in the temperature dependence of arabinose and xylose are consistent with the greater thermal stability of xylose compared to arabinose, as previously reported [65]. In terms of process optimization, Getachew and Chun recommend high temperatures and longer residence times, and hence high severity factors, for recovery of valuable compounds biomass [45]; however, for thermally sensitive products, such as arabinose, increasing severity factor must balance with minimizing thermal degradation. Flow-through conditions offer an approach that can de-couple fluid residence time from solids residence time, thereby providing a degree of freedom for maximizing yields that is independent of the batch-mode severity factor. Liu and Wyman [66] go a step further, suggesting mixing batch and flow-through conditions to gain advantages of each reactor configuration, a strategy which may be worth considering for BSG in future work. The effects of hydrolysis time and water consumption on sugar yields were considered. For example, doubling S/F from 64 to 112 increased arabinose yield by only 16% (2.6 compared to 3.1 g arabinose/ 100 g BSG). Similarly, Fig. 3 shows that increasing S/F decreases TRS concentrations at all conditions, with the most substantial effect observed at 210 °C. Since increasing S/F increases both water consumption and hydrolysis time, while providing minimal benefit to the arabinose yield and actually decreasing sugar concentrations, optimum conditions may be selected that do not simply maximize sugar yield but instead balance the benefits of minimizing solvent use, maximizing to the extent possible production rates, sugar concentrations, and sugar yields.

Fig. 3. The concentration of Total reducing sugars (TRS) equivalent in glucose as a function of the temperature and different S/F collected from hydrolysis kinetics at flow rates of a) 10 mL min−1 and b) 20 mL min−1.

oligomers. Flow-through conditions are designed to minimize degradation of simple sugars; hence, a complete study must include quantification of degradation products. Table 4 summarizes degradation product yields, including the organic acids (formic acid and acetic acid) and the furanic compounds, HMF and FF. Yields of degradation products were strongly influenced by hydrolysis temperature and well correlated with decreases in hydrolysate pH, measured at S/F (64, 80 and 112). The decreasing pH is consistent with the release of acetyl groups present in the hemicellulose structure [33,67]; their release may promote a catalytic and favorable medium for the hydrolysis reactions

3.3. Degradation products and organic acids Results presented on RS, TRS, and individual sugar yields suggest that sugar oligomers are the most abundant primary hydrolysis product; simple sugars form as minority products presumably directly from hemicellulose hydrolysis and also from hydrolysis of the sugar 24

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Table 4 The composition of organic compounds and pH of brewer´s spent grains hydrolysates obtained under different experimental conditions (g compound / 100 g feed). Flow rate

Temperature

−1

10 mL min

S/F

140 °C

64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112 64 80 112

160 °C 180 °C 210 °C 20 mL min−1

140 °C 160 °C 180 °C 210 °C

Organic acids

Degradation products

pH

Formic acid

Acetic acid

HMF

Furfural

n.d n.d n.d 0.4 ± 0.1 0.33 ± 0.00 0.37 ± 0.03 0.42 ± 0.00 0.45 ± 0.04 0.57 ± 0.04 0.8 ± 0.2 1.05 ± 0.01 1.5 ± 0.3 n.d n.d n.d 0.3 ± 0.2 0.24 ± 0.01 0.22 ± 0.02 0.34 ± 0.05 0.4 ± 0.1 0.5 ± 0.1 0.3 ± 0.1 0.7 ± 0.1 0.6 ± 0.2

n.d n.d n.d 0.7 ± 0.2 0.6 ± 0.1 0.6 ± 0.1 1.0 ± 0.1 1.09 ± 0.01 1.0 ± 0.1 1.7 ± 0.1 1.8 ± 0.1 1.78 ± 0.04 n.d n.d n.d 0.4 ± 0.1 0.43 ± 0.04 0.52 ± 0.01 0.7 ± 0.1 0.7 ± 0.2 0.7 ± 0.2 1.3 ± 0.1 1.3 ± 0.2 1.4 ± 0.1

n.d n.d n.d n.d n.d n.d 0.13 ± 0.00 0.16 ± 0.00 0.23 ± 0.00 0.39 ± 0.01 0.4 ± 0.1 0.46 ± 0.00 n.d n.d n.d n.d n.d n.d 0.13 ± 0.00 0.16 ± 0.00 0.22 ± 0.00 0.19 ± 0.08 0.24 ± 0.1 0.22 ± 0.01

0.2 ± 0.1 0.2 ± 0.1 n.d 0.7 ± 0.1 0.7 ± 0.1 0.8 ± 02 1.0 ± 0.1 1.1 ± 0.1 1.2 ± 0.3 1.37 ± 0.07 1.5 ± 0.1 1.7 ± 0.1 0.2 ± 0.1 0.2 ± 0.1 0.2 ± 0.0 0.51 ± 0.02 0.6 ± 0.1 0.62 ± 0.02 0.76 ± 0.02 0.80 ± 0.02 0.89 ± 0.02 1.1 ± 0.1 1.2 ± 0.1 1.2 ± 0.1

4.6 ± 0.1 4.3 ± 0.2 4.45 ± 0.01 4.04 ± 0.01 4.04 ± 0.06 4.03 ± 0.04 3.83 ± 0.01 3.88 ± 0.00 3.92 ± 0.03 3.44 ± 0.00 3.7 ± 0.2 3.7 ± 0.2 4.2 ± 0.1 4.3 ± 0.2 4.3 ± 0.1 4.28 ± 0.01 4.3 ± 0.2 4.1 ± 0.1 3.9 ± 0.1 3.9 ± 0.1 3.98 ± 0.00 3.8 ± 0.0 3.9 ± 0.1 3.95 ± 0.01

- n.d: not detected - below detection limits.

of hemicellulose into simple sugars [68], which, in turn, can be degraded in other compounds such as furfural, hydroxymethylfurfural (HMF) and organic acids [30,69]. Degradation products also include organic acids, which contribute to further decreases in pH with increasing temperature. Although acidic conditions were not used, humins may also be formed by the condensation of glucose and C5 sugars with furfural and HMF, formation of which is catalyzed by organic acids formed by hydrolysis of simple sugars [70]. The degradation reaction mechanism likely occurs via the parallel channels shown in equations 5 and 6. Sugar degradation products, such as HMF, originate from dehydration of hexoses, specifically fructose but also glucose [30,71]. The dominant source of HMF depends both on the relative concentrations of glucose and fructose, an the relative dehydration rates of glucose and fructose, as fructose is much more readily dehydrated than glucose [27,72,73]. For BSG, glucose may be the product of either cellulose or hemicellulose hydrolysis. Regardless of the source, a fraction of the glucose produced is transformed into fructose, which may be consistent with the data in Table 3. Xylose and arabinose undergo dehydration to produce FF [74–76].

Xylan

Hydrolysis

Glucose

Xylose

isomerization

H2 O

Fructose

Furfural H2 O

HMF

hydrolysate pH. With the exception of hydrolysate obtained at 140 °C, the pH of hydrolysates obtained at 10 mL min–1 was always less than those obtained at 20 mL min–1, consistent with production of other byproducts. Since hemicellulose hydrolysis is pH sensitive, formation of organic acids is auto-catalytic [77,78]. On the other hand, the degradation reactions shown in Eqs. (5) and (6) are also promoted by acids, so the effect of acid must balance initial hydrolysis rates with degradation rates. When considered together, Tables 3 and 4 suggest that optimal flow-through treatment maximizes sugar yield and minimizes degradation by operating at high flow rates that allow operation at optimal pH, at high temperatures (on the order of 210 °C), and S/F equal to or less than 64. 3.4. Structural modification of brewer´s spent grains The next step to analyzing the flow-through process is characterization of the residual solids. Fig. 4 shows the mass of remaining solids collected after treatment after drying at 75 °C. The yield of solid residue decreased with increasing process temperature, as expected for a process involving hydrolysis and solubilization of hemicellulose during treatment [79] and in agreement with RS and TRS yields. The maximum biomass removal was observed at a treatment temperature of 210 °C, at which condition approximately 75% of the biomass had been removed from the reactor. Since TRS yields account for only 35–40% biomass deconstruction, the remaining mass loss must be due to formation of compounds not quantified in the TRS measurement and/or particle removal. The flow-through reactor was designed to prevent removal of primary biomass particles during hydrolysis. However, Mosteiro-Romero et al. [80] studied biomass hydrolysis in hydrothermal water, reporting formation of secondary particles during the hydrolysis process. The un-accounted for mass may be attributable to formation of secondary particles which are then carried out of the reactor. Interestingly, the mass of residual solids was greater at a flow rate of 20 mL min–1 compared to that found at 10 mL min–1, suggesting that particle removal, if/when it occurs, is not a hydrodynamic phenomenon and instead may be influenced by thermochemical factors.

(5) (6)

In all cases, FF yields were greater than HMF yields, generally by about a factor of 10 or more at 140 and 160 °C and by 3–5 at temperatures of 180 and 210 °C. The greater yield of FF is consistent with the greater yields of C5 sugars compared to C6 sugars. At 140 °C, HMF yields were less than detection limits (0.01 – 0.2 g L−1), presumably due both to the low fructose/glucose yields and because the temperature was not sufficient for dehydration of the primary sugars before they were removed from the reactor. Most interestingly, HMF and FF yields were sensitive to flow rates, with increasing flow rate decreasing both HMF and FF yields. HMF yields were especially sensitive to flow rate, with an increase from 10 to 20 mL min–1 decreasing HMF yields by a factor of 2 and FF by about 25%. As with sugar yields, HMF and FF yields did not vary greatly for S/F > 64. Lastly, Table 4 lists 25

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temperature, an observation which is consistent with progressive removal of glycosidic bonds associated with hemicellulose. As the hemicellulose features are removed from the IR spectra, the remaining cellulose features become increasingly distinct [84]. Fig. 6b provides a closer view of the aromatic and carbonyl region of the IR spectra, from 1400 to 1900 cm–1. Interestingly, the intensity of a prominent band at approximately 1650 cm–1 decreases after treatment at 140 °C. The band at 1650 cm–1 may be attributable to either lignin or bound water [53]; however, given that lignin is not hydrolyzed at these conditions, the attribution is more likely to be bound water, and the decrease in the bound water signal in the treated biomass may suggest that it is more hydrophobic than the feed. Inspection of the carbonyl bands, shown in Fig. 6b, provide additional insight. Specifically, the intensity of the band at approximately 1740 cm–1 decreases after treatment, and a new band at 1710 cm–1 appears. The band at 1740 cm–1 is associated with acetyl side chains, which are present in hemicellulose and easily hydrolyzed to produce acetic acid [85]. The band at 1710 cm–1, on the other hand, is likely attributable to carboxylic acid or keto- groups [86]. In their study of coffee powder hydrolysis, Ma et al. [32] observed formation of carboxylic acid groups at temperatures coincident with appearance of char. Likewise, Fig. 5 suggests onset of char formation when BSG is treated at 140 °C, the same temperature at which the carboxylic acid band appears in the IR spectrum of treated feed. Therefore, the current results obtained from flow-through hydrolysis of BSG corroborate earlier findings obtained from coffee powder hydrolysis that suggest formation of surface-bound carboxylic acids occurs at temperatures coincident with char formation [32].

Fig. 4. Solid residue remaining inside the reactor after subcritical water hydrolysis of brewer’s spent grains.

3.4.1. Thermogravimetric analysis Residual solids were studied using TGA. Fig. 5a, b show the raw thermograms, plotted as differential thermograms (DTG). DTG peaks of biomass are typically differentiated into 3 main components, representing cellulose, hemicellulose, and lignin. Of these, hemicellulose volatilizes over the temperature range from 250 to 300 °C, cellulose at approximately 320–340 °C, and lignin over a broad range from 200 to 450 °C [81]. DTG curves can be fit to estimate the solid composition; Fig. 5c, d show the results of the curve fitting exercise. Roughly consistent with bulk analysis, DTG indicates that approximately 30% of the feed is hemicellulose. The feed also contains a component that volatilizes at approximately 150–200 °C, which is termed “semi-volatile”; the semi-volatile component is likely some combination of free sugars, resins, and similar components. After treatment at 140 °C, the hemicellulose component decreases and the semi-volatile component disappears entirely. The cellulose and lignin contents increase accordingly, and a contribution from a component with volatility characteristics consistent with char [82] appears. With increasing treatment temperature, the hemicellulose component continues to decrease and the char component increases (Fig. 5c, d). Though always a minor component (< 10%), char content is less in samples treated at 20 mL min–1 compared to those treated at 10 mL min–1, suggesting that flow effectively decreases accumulation of char precursors. Lastly, the semi-volatile component, which is present in the feed but not in the sample treated at 140 °C, re-appears after treatment at 160 °C, and its contribution increases with increasing treatment temperature. Ma et al. [32] observed similar behavior in the semi-volatile components of coffee powder treated hydrothermally. Interestingly, unlike char, the semi-volatile content is greater in samples treated at a flow rate of 10 mL min–1 than in those treated at 20 mL min–1.

3.5. Integrating hydrolysate and residual solids analysis Previous work by Ma et al. [32], which is supported by chemical interpretation of TGA data obtained from analysis of carbonaceous solids [87], linked the semi-volatile component with surface bound acids; therefore, the effect of increasing flow rate is expected to decrease the contribution of surface bound acids in the treated samples. Fig. 7 is a direct comparison of the effects of flow rate on the carbonyl regions of the IR spectra obtained from samples treated at 140 (Fig. 7a) and 210 °C (Fig. 7b). In the samples treated at 140 °C, the intensity of the carboxylic acid band is not strongly dependent on flow rate. However, in the samples treated at 210 °C, the carboxylic band is far more intense in the sample obtained from hydrolysis at 10 mL min–1 than the one obtained at 20 mL min–1. Similarly, spectra obtained at intermediate temperatures are intermediate to the two extremes shown in Fig. 7, with the intensity of the carbonyl band in samples treated at 10 mL min–1 always comparable to or greater than that obtained at 20 mL min–1. Combining TGA data that indicate that char content is less in samples treated at 20 mL min–1 than in samples treated at 10 mL min–1 is consistent with removal of char precursors from the reaction mixture, preventing their reaction with reactive groups on the biomass particle surface (including carboxylic acids) to form char. Therefore, increasing flow rate decreases char yields, but increases the content of surface bound carboxylic acids present on the particle surface. Finally, comparing sugar yields obtained at different flow rates depends on what is held constant – S/F ratio or solids residence time. Integration of solids characterization data with hydrolysate data allows comparison of sugar yields as a function of hemicellulose removal, which can be obtained directly from Fig. 5. Fig. 8 shows the results of using hemicellulose removal as the independent variable for comparing sugar yields. First, Fig. 8a shows the fractional hemicellulose content remaining after treatment as a function of hydrolysis temperature. Interestingly, at temperatures less than 180 °C, hemicellulose removal is favored by operation at the lower flow rate (10 mL min–1), whereas at temperatures equal to and greater than 180 °C, hemicellulose removal is favored by the higher flow rate (20 mL min–1). This finding is consistent with the importance of acid auto-catalysis at low temperatures – where

3.4.2. Infrared spectroscopy Fig. 6 contains IR spectra obtained from analysis of the feed and treated biomass at a flow rate of 10 mL min–1. Similar spectral features were observed in samples treated at 20 mL min–1. Fig. 6a contains IR data for the glycosidic bond region, 800–1200 cm–1, showing that the main cellulose bands present at 1035, 1060, 1110, and 1165 cm–1 are visible in all of the samples [83]. The persistence of cellulose bands after treatment is consistent with the hydrothermal stability of cellulose. The main difference in the glycosidic region of the IR spectra is that the cellulose bands become increasingly distinct with treatment 26

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Fig. 5. Thermogravimetric analysis of residual solids: (a and b) raw differential thermograms (DTG) obtained from analysis of solids treated at different temperatures and a flow rate of 10 mL min–1 and 20 mL min–1; (c and d) compositions inferred from deconvolution of the data shown in (a) and (b), as obtained at 10 mL min–1 and 20 mL min–1, respectively.

Fig. 6. IR spectra obtained from analysis of residual solids collected after hydrolysis treatment at a flow rate of 10 mL min–1 and at various temperatures: (a) the glycosidic bond region and (b) the carbonyl region.

27

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10 mL min 20 mL min

8

–1

40

(b)

(c)

–1

25

–1

20 15 10

6

–1

30

(a)

4

2

10 mL min 20 mL min

5 0

TRS Yield (g 100 g )

35

RS Yield (g 100 g )

Fractional Hemicellulose Content (%)

Fig. 7. Carbonyl portion of the IR spectra obtained from analysis of residual solids collected after hydrolysis treatment at (a) 140 °C and (b) 210 °C at flow rates of 10 mL min−1 (green) and 20 mL min−1 (red).

–1

20

10

10 mL min

–1

20 mL min

–1 –1

0

0 140 160 180 200 Treatment Temperature (ºC)

30

0

10 20 30 Hemicellulose Content (%)

0

10 20 30 Hemicellulose Content (%)

Fig. 8. Analysis of the inter-relationships between hydrolysis temperature, hemicellulose content, and RS and TRS yields: (a) relationship between hemicellulose content and hydrolysis temperature; (b) relationship between RS yield and hemicellulose content, (c) relationship between TRS yield and hemicellulose content.

the hydrolysis rate is slow – and the importance of char removal at high temperatures, where degradation and char formation is most problematic. Plotting hemicellulose removal as a function of log R0 (not shown) results in similar conclusions to those drawn when data are plotted as a function of temperature. Fig. 8b and c show RS and TRS yields as functions of residual hemicellulose content, respectively. For RS yield, shown in Fig. 8b, high flow is preferred over low flow when the residual hemicellulose content is greater than about 15%. However, for more aggressive conditions which result in less than 15% residual hemicellulose, low flow results in greater RS yields. Again, this may be suggestive of the auto-catalytic role of both soluble and surface acids. TRS yields, in comparison, are nearly independent of flow rate, when the data are plotted as a function of hemicellulose content (Fig. 8c). Therefore, the effect of flow rate on TRS yields is expressed entirely by hemicellulose removal, which is consistent with rapid removal of TRS products once they are solubilized.

operating conditions were 210 °C, 20 mL min–1 water flow rate, and S/F of 64. The maximum RS yield of reducing sugars was 5.84 g per 100 g of feed and the maximum TRS yield was 35.11 g per 100 g of feed. Under all conditions, arabinose was the most abundant identified sugar product, with a maximum yield of approximately 3.1 g per 100 g of feed. Yields were strongly dependent on hydrolysis temperature and also S/F ratio, at least for S/F < 64. Instead, the main effect of increasing flow rate is to decrease the time required to reach maximum yields. In contrast, yields of degradation products, including soluble furfuralbased compounds and insoluble chars, were sensitive to both hydrolysis temperature and flow rate. Analysis of the residual solids indicated that flow rate plays a complex role in hemicellulose removal, suggesting potential phenomena such as auto-catalysis, char formation, and break down of primary products to form simple sugars and degradation products. The findings provide new insight in the design and operation of flow-through hydrolysis reactors, and emphasize the potential of brewer´s spent grains as a renewable source of C5 sugars.

4. Conclusions

Acknowledgments

Flow-through subcritical water technology has promise for production of hemicellulosic sugars (C5 sugar) from brewer´s spent grains (BSG). In terms of sugar yields that balance solvent use, the optimal

The authors acknowledge financial support from the São Paulo Research Foundation – FAPESP (2011/19817-1, 2018/05999-0, 2016/ 04602-3 and 2013/04304-4). This study was financed in part by the 28

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Coodenação de Aperfeiçoamento de Pessoal de Nivel Superior – BrazilFinance code 001. MAR are thankful to CNPq for the productivity grant (303568/2016-0). The WPI contribution was supported by the U.S. NSF (CBET 1554283). Constantina Drakontis, Sotirious Filippou, Stephanie Gulezian, and Carolyn Morales Collado contributed to the solids analysis as part of their senior projects at WPI.

[30] [31]

References

[32]

[1] J.S. White, B.K. Yohannan, G.M. Walker, Bioconversion of brewer’s spent grains to bioethanol, FEMS Yeast Res. 8 (2008) 1175–1184. [2] H. Chen, Chemical composition and structure of natural lignocellulose, Biotechnology of Lignocellulose, Springer, 2014, pp. 25–71. [3] V. Menon, M. Rao, Trends in bioconversion of lignocellulose: biofuels, platform chemicals & biorefinery concept, Prog. Energy Combust. Sci. 38 (2012) 522–550. [4] S.I. Mussatto, Brewer’s spent grain: a valuable feedstock for industrial applications, J. Sci. Food Agric. 94 (2014) 1264–1275. [5] G.A. Jacometti, L.R. Mello, P.H. Nascimento, A.C. Sueiro, F. Yamashita, S. Mali, The physicochemical properties of fibrous residues from the agro industry, LWT - Food Sci. Technol. 62 (2015) 138–143. [6] L.R.P.F. Mello, R.M. Vergílio, S. Mali, Caracterização Química e funcional do Resíduo Fibroso Da Indústria Cervejeira, BBR, Biochem. Biotechnol. Rep. 2 (2013) 191–194. [7] E.J. Pires, Hc.A. Ruiz, J.A. Teixeira, An.A. Vicente, A new approach on brewer’s spent grains treatment and potential use as lignocellulosic yeast cells carriers, J. Agric. Food Chem. 60 (2012) 5994–5999. [8] N.G. Meneses, S. Martins, J.A. Teixeira, S.I. Mussatto, Influence of extraction solvents on the recovery of antioxidant phenolic compounds from brewer’s spent grains, Sep. Purif. Technol. 108 (2013) 152–158. [9] S. Mussatto, G. Dragone, I. Roberto, Brewers’ spent grain: generation, characteristics and potential applications, J. Cereal Sci. 43 (2006) 1–14. [10] S.I. Mussatto, G.J. Rocha, I.C. Roberto, Hydrogen peroxide bleaching of cellulose pulps obtained from brewer’s spent grain, Cellulose 15 (2008) 641–649. [11] A. Ebringerová, Structural diversity and application potential of hemicelluloses, Macromolecular Symposia, Wiley Online Library, 2005, pp. 1–12. [12] Ž. Knez, E. Markočič, M.K. Hrnčič, M. Ravber, M. Škerget, High pressure water reforming of biomass for energy and chemicals: a short review, J. Supercrit. Fluids 96 (2015) 46–52. [13] Y. Yu, X. Lou, H. Wu, Some recent advances in hydrolysis of biomass in hot-compressed water and its comparisons with other hydrolysis methods, Energy Fuels 22 (2007) 46–60. [14] X. Zhao, L. Zhang, D. Liu, Biomass recalcitrance. Part I: the chemical compositions and physical structures affecting the enzymatic hydrolysis of lignocellulose, Biofuel Bioprod. Biorefin. 6 (2012) 465–482. [15] N. Kopsahelis, M. Kanellaki, A. Bekatorou, Low temperature brewing using cells immobilized on brewer’s spent grains, Food Chem. 104 (2007) 480–488. [16] N. Kopsahelis, L. Bosnea, A. Bekatorou, C. Tzia, M. Kanellaki, Alcohol production from sterilized and non-sterilized molasses by Saccharomyces cerevisiae immobilized on brewer’s spent grains in two types of continuous bioreactor systems, Biomass Bioenergy 45 (2012) 87–94. [17] S.I. Mussatto, M. Fernandes, G. Dragone, I.M. Mancilha, I.C. Roberto, Brewer’s spent grain as raw material for lactic acid production by Lactobacillus delbrueckii, Biotechnol. Lett. 29 (2007) 1973–1976. [18] S.I. Mussatto, I.C. Roberto, Acid hydrolysis and fermentation of brewer’s spent grain to produce xylitol, J. Sci. Food Agric. 85 (2005) 2453–2460. [19] D.A. Cantero, M.D. Bermejo, M.J. Cocero, Reaction engineering for process intensification of supercritical water biomass refining, J. Supercrit. Fluids 96 (2015) 21–35. [20] N. Mosier, C. Wyman, B. Dale, R. Elander, Y. Lee, M. Holtzapple, M. Ladisch, Features of promising technologies for pretreatment of lignocellulosic biomass, Bioresour. Technol. 96 (2005) 673–686. [21] A. Demirbas, Hydrogen production from biomass via supercritical water gasification, Energy Sourc. A Recovery Util. Environ. Effects 32 (2010) 1342–1354. [22] S.S. Toor, L. Rosendahl, A. Rudolf, Hydrothermal liquefaction of biomass: a review of subcritical water technologies, Energy 36 (2011) 2328–2342. [23] M. Mohan, T. Banerjee, V.V. Goud, Hydrolysis of bamboo biomass by subcritical water treatment, Bioresour. Technol. 191 (2015) 244–252. [24] M.A. Rostagno, J.M. Prado, A. Mudhoo, D.T. Santos, T. Forster–Carneiro, M.A.A. Meireles, Subcritical and supercritical technology for the production of second generation bioethanol, Crit. Rev. Biotechnol. 35 (2015) 302–312. [25] R.A. Sheldon, Green and sustainable manufacture of chemicals from biomass: state of the art, Green Chem. 16 (2014) 950–963. [26] H. Xu, W. Wang, X. Liu, F. Yuan, Y. Gao, Antioxidative phenolics obtained from spent coffee grounds (Coffea arabica L.) by subcritical water extraction, Ind. Crops Prod. 76 (2015) 946–954. [27] J.M. Prado, D. Lachos-Perez, T. Forster-Carneiro, M.A. Rostagno, Sub-and supercritical water hydrolysis of agricultural and food industry residues for the production of fermentable sugars: a review, Food Bioprod. Process. 98 (2016) 95–123. [28] H.A. Ruiz, R.M. Rodriguez-Jasso, B.D. Fernandes, A.A. Vicente, J.A. Teixeira, Hydrothermal processing, as an alternative for upgrading agriculture residues and marine biomass according to the biorefinery concept: a review, Renew. Sust. Energy Rev. 21 (2013) 35–51. [29] R. Lin, J. Cheng, L. Ding, W. Song, F. Qi, J. Zhou, K. Cen, Subcritical water

[33] [34] [35] [36] [37] [38] [39] [40]

[41] [42] [43] [44] [45] [46]

[47] [48] [49] [50] [51] [52] [53] [54]

[55] [56] [57] [58] [59] [60]

29

hydrolysis of rice straw for reducing sugar production with focus on degradation byproducts and kinetic analysis, Bioresour. Technol. 186 (2015) 8–14. F.M. Yedro, H. Grénman, J.V. Rissanen, T. Salmi, J. García-Serna, M.J. Cocero, Chemical composition and extraction kinetics of Holm oak (Quercus ilex) hemicelluloses using subcritical water, J. Supercrit. Fluids 129 (2017) 56–62. J.M. Prado, T. Forster-Carneiro, M.A. Rostagno, L.A. Follegatti-Romero, F. Maugeri Filho, M.A.A. Meireles, Obtaining sugars from coconut husk, defatted grape seed, and pressed palm fiber by hydrolysis with subcritical water, J. Supercrit. Fluids 89 (2014) 89–98. Z. Ma, P. Guerra, M. Tyufekchiev, A. Zaker, G.A. Tompsett, P.T. Mayanga, T. Forster-Carneiro, P. Wang, M.T. Timko, Formation of an external char layer during subcritical water hydrolysis of biomass, Sustain. Energy Fuels (2017). D. Lachos-Perez, G. Tompsett, P. Guerra, M. Timko, M. Rostagno, J. Martínez, T. Forster-Carneiro, Sugars and char formation on subcritical water hydrolysis of sugarcane straw, Bioresour. Technol. 243 (2017) 1069–1077. V. Archambault‐Léger, Z. Losordo, L.R. Lynd, Energy, sugar dilution, and economic analysis of hot water flow‐through pre‐treatment for producing biofuel from sugarcane residues, Biofuel Bioprod. Biorefin. 9 (2015) 95–108. O. Pourali, F.S. Asghari, H. Yoshida, Sub-critical water treatment of rice bran to produce valuable materials, Food Chem. 115 (2009) 1–7. D. Ciftci, M.D. Saldaña, Hydrolysis of sweet blue lupin hull using subcritical water technology, Bioresour. Technol. 194 (2015) 75–82. W. Abdelmoez, S.M. Nage, A. Bastawess, A. Ihab, H. Yoshida, Subcritical water technology for wheat straw hydrolysis to produce value added products, J. Clean. Prod. 70 (2014) 68–77. A. Sluiter, B. Hames, C.S.R. Ruiz, J. Sluiter, D. Templeton, Determination of Ash in Biomass, National Renewable Energy Laboratory, 2008, p. 5. R. Ruiz, C. Scarlata, J. Sluiter, D. Templeton, Determination of Extractives in Biomass, NREL Laboratory Analytical Procedure, National Renewable Energy Laboratory Golden, CO, 2005. A. Sluiter, B. Hames, D. Hyman, C. Payne, R. Ruiz, C. Scarlata, J. Sluiter, D. Templeton, J. Wolfe, Determination of Total Solids in Biomass and Total Dissolved Solids in Liquid Process Samples, National Renewable Energy Laboratory, 2008, pp. 1–6. A. Sluiter, B. Hames, R. Ruiz, C. Scarlata, J. Sluiter, D. Templeton, D. Crocker, Determination of Structural Carbohydrates and Lignin in Biomass, National Renewable Energy Laboratory, 2008. B. Hames, C. Scarlata, A. Sluiter, Determination of Protein Content in Biomass, National Renewable Energy Laboratory, 2008, pp. 1–5. L.E. Wise, Chlorite holocellulose, its fractionation and bearing on summative wood analysis and on studies on the hemicelluloses, Paper Trade 122 (1946) 35–43. Y. Teramoto, S.-H. Lee, T. Endo, Cost reduction and feedstock diversity for sulfuric acid-free ethanol cooking of lignocellulosic biomass as a pretreatment to enzymatic saccharification, Bioresour. Technol. 100 (2009) 4783–4789. A.T. Getachew, B.S. Chun, Influence of pretreatment and modifiers on subcritical water liquefaction of spent coffee grounds: a green waste valorization approach, J. Clean. Prod. 142 (2017) 3719–3727. P. Mayanga-Torres, D. Lachos-Perez, C. Rezende, J. Prado, Z. Ma, G. Tompsett, M. Timko, T. Forster-Carneiro, Valorization of coffee industry residues by subcritical water hydrolysis: recovery of sugars and phenolic compounds, J. Supercrit. Fluids 120 (2017) 75–85. S.I. Sandler, Chemical, Biochemical, and Engineering Thermodynamics, John Wiley & Sons, Hoboken, NJ, 2006. N. Nelson, A photometric adaptation of the Somogyi method for the determination of glucose, J. Biol. Chem. 153 (1944) 375–380. G.L. Miller, Use of dinitrosalicylic acid reagent for determination of reducing sugar, Anal. Chem. 31 (1959) 426–428. F. Carvalheiro, M. Esteves, J. Parajó, H. Pereira, F. Gırio, Production of oligosaccharides by autohydrolysis of brewery’s spent grain, Bioresour. Technol. 91 (2004) 93–100. N.C. Steinmacher, F.A. Honna, A.V. Gasparetto, D. Anibal, M.V. Grossmann, Bioconversion of brewer’s spent grains by reactive extrusion and their application in bread-making, LWT - Food Sci. Technol. 46 (2012) 542–547. L.R. Mello, S. Mali, Use of malt bagasse to produce biodegradable baked foams made from cassava starch, Ind. Crops Prod. 55 (2014) 187–193. A. Linan-Montes, S. De La Parra-Arciniega, M. Garza-González, R. García-Reyes, E. Soto-Regalado, F. Cerino-Córdova, Characterization and thermal analysis of agave bagasse and malt spent grain, J. Therm. Anal. Calorim. 115 (2014) 751–758. J. Poerschmann, B. Weiner, H. Wedwitschka, I. Baskyr, R. Koehler, F.-D. Kopinke, Characterization of biocoals and dissolved organic matter phases obtained upon hydrothermal carbonization of brewer’s spent grain, Bioresour. Technol. 164 (2014) 162–169. S.I. Mussatto, I.C. Roberto, Chemical characterization and liberation of pentose sugars from brewer’s spent grain, J. Chem. Technol. Biotechnol. 81 (2006) 268–274. L.G. Cordeiro, Â.A. El-Aouar, C.V.B. de Araújo, Energetic characterization of malt bagasse by calorimetry and thermal analysis, J. Therm. Anal. Calorim. 112 (2013) 713–717. M. Santos, J. Jiménez, B. Bartolomé, C. Gómez-Cordovés, M. Del Nozal, Variability of brewer’s spent grain within a brewery, Food Chem. 80 (2003) 17–21. M. Roman, W.T. Winter, Effect of sulfate groups from sulfuric acid hydrolysis on the thermal degradation behavior of bacterial cellulose, Biomacromolecules 5 (2004) 1671–1677. W. Yang, H. Wang, J. Zhou, S. Wu, Hydrolysis kinetics and structure changes of wood meal in subcritical water, ACS Sustain. Chem. Eng. 5 (2017) 3544–3552. A.G. Carr, R. Mammucari, N. Foster, A review of subcritical water as a solvent and

The Journal of Supercritical Fluids 145 (2019) 19–30

P.C. Torres-Mayanga et al.

[61] [62] [63] [64] [65] [66] [67] [68]

[69] [70] [71] [72] [73]

its utilisation for the processing of hydrophobic organic compounds, Chem. Eng. J. 172 (2011) 1–17. I. Sereewatthanawut, S. Prapintip, K. Watchiraruji, M. Goto, M. Sasaki, A. Shotipruk, Extraction of protein and amino acids from deoiled rice bran by subcritical water hydrolysis, Bioresour. Technol. 99 (2008) 555–561. T. Anekpankul, M. Goto, M. Sasaki, P. Pavasant, A. Shotipruk, Extraction of anticancer damnacanthal from roots of Morinda citrifolia by subcritical water, Sep. Purif. Technol. 55 (2007) 343–349. S. Evstafev, E. Chechikova, Transformation of wheat straw polysaccharides under dynamic conditions of subcritical autohydrolysis, Russ. J. Bioorganic Chem. 42 (2016) 700–706. F.M. Yedro, J. García-Serna, D.A. Cantero, F. Sobrón, M.J. Cocero, Hydrothermal fractionation of grape seeds in subcritical water to produce oil extract, sugars and lignin, Catal. Today 257 (2015) 160–168. Y. Gao, H. Wang, J. Guo, P. Peng, M. Zhai, D. She, Hydrothermal degradation of hemicelluloses from triploid poplar in hot compressed water at 180–340° C, Polym. Degrad. Stab. 126 (2016) 179–187. C. Liu, C.E. Wyman, Partial flow of compressed-hot water through corn stover to enhance hemicellulose sugar recovery and enzymatic digestibility of cellulose, Bioresour. Technol. 96 (2005) 1978–1985. J.V. Rissanen, H. Grénman, C. Xu, S. Willför, D.Y. Murzin, T. Salmi, Obtaining spruce hemicelluloses of desired molar mass by using pressurized hot water extraction, ChemSusChem 7 (2014) 2947–2953. F. Carvalheiro, L. Duarte, F. Gírio, P. Moniz, Hydrothermal/Liquid Hot Water pretreatment (autohydrolysis): a multipurpose process for biomass upgrading, Biomass Fractionation Technologies for a Lignocellulosic Feedstock Based Biorefinery, Elsevier, 2016, pp. 315–347. Y. Kim, T. Kreke, N.S. Mosier, M.R. Ladisch, Severity factor coefficients for subcritical liquid hot water pretreatment of hardwood chips, Biotechnol. Bioeng. 111 (2014) 254–263. B. Girisuta, B. Danon, R. Manurung, L. Janssen, H. Heeres, Experimental and kinetic modelling studies on the acid-catalysed hydrolysis of the water hyacinth plant to levulinic acid, Bioresour. Technol. 99 (2008) 8367–8375. D.A. Cantero, A. Álvarez, M.D. Bermejo, M.J. Cocero, Transformation of glucose into added value compounds in a hydrothermal reaction media, J. Supercrit. Fluids 98 (2015) 204–210. B.M. Kabyemela, T. Adschiri, R.M. Malaluan, K. Arai, Kinetics of glucose epimerization and decomposition in subcritical and supercritical water, Ind. Eng. Chem. Res. 36 (1997) 1552–1558. F. Salak Asghari, H. Yoshida, Acid-catalyzed production of 5-hydroxymethyl

[74] [75] [76] [77] [78] [79] [80] [81] [82] [83] [84] [85] [86] [87]

30

furfural from D-fructose in subcritical water, Ind. Eng. Chem. Res. 45 (2006) 2163–2173. M.J. Antal, T. Leesomboon, W.S. Mok, G.N. Richards, Mechanism of formation of 2furaldehyde from D-xylose, Carbohydr. Res. 217 (1991) 71–85. L. Hu, G. Zhao, W. Hao, X. Tang, Y. Sun, L. Lin, S. Liu, Catalytic conversion of biomass-derived carbohydrates into fuels and chemicals via furanic aldehydes, RSC Adv. 2 (2012) 11184–11206. H. Rasmussen, H.R. Sørensen, A.S. Meyer, Formation of degradation compounds from lignocellulosic biomass in the biorefinery: sugar reaction mechanisms, Carbohydr. Res. 385 (2014) 45–57. C. Pronyk, G. Mazza, Kinetic modeling of hemicellulose hydrolysis from triticale straw in a pressurized low polarity water flow-through reactor, Ind. Eng. Chem. Res. 49 (2010) 6367–6375. H.A. Ruiz, M.A. Cerqueira, H.D. Silva, R.M. Rodríguez-Jasso, A.A. Vicente, J.A. Teixeira, Biorefinery valorization of autohydrolysis wheat straw hemicellulose to be applied in a polymer-blend film, Carbohydr. Polym. 92 (2013) 2154–2162. K. Watchararuji, M. Goto, M. Sasaki, A. Shotipruk, Value-added subcritical water hydrolysate from rice bran and soybean meal, Bioresour. Technol. 99 (2008) 6207–6213. M. Mosteiro-Romero, F. Vogel, A. Wokaun, Liquefaction of wood in hot compressed water: part 1—experimental results, Chem. Eng. Sci. 109 (2014) 111–122. M. Carrier, A. Loppinet-Serani, D. Denux, J.-M. Lasnier, F. Ham-Pichavant, F. Cansell, C. Aymonier, Thermogravimetric analysis as a new method to determine the lignocellulosic composition of biomass, Biomass Bioenergy 35 (2011) 298–307. M.A. Islam, M. Asif, B. Hameed, Pyrolysis kinetics of raw and hydrothermally carbonized Karanj (Pongamia pinnata) fruit hulls via thermogravimetric analysis, Bioresour. Technol. 179 (2015) 227–233. F. Xu, J. Yu, T. Tesso, F. Dowell, D. Wang, Qualitative and quantitative analysis of lignocellulosic biomass using infrared techniques: a mini-review, Appl. Energy 104 (2013) 801–809. D. Lachos-Perez, F. Martinez-Jimenez, C. Rezende, G. Tompsett, M. Timko, T. Forster-Carneiro, Subcritical water hydrolysis of sugarcane bagasse: an approach on solid residues characterization, J. Supercrit. Fluids 108 (2016) 69–78. R.C. Pettersen, The chemical composition of wood, Chem. Solid Wood 207 (1983) 57–126. H. Schulz, M. Baranska, Identification and quantification of valuable plant substances by IR and Raman spectroscopy, Vib. Spectrosc. 43 (2007) 13–25. J. Figueiredo, M. Pereira, M. Freitas, J. Orfao, Modification of the surface chemistry of activated carbons, Carbon 37 (1999) 1379–1389.