6816
Ind. Eng. Chem. Res. 2005, 44, 6816-6823
Comparison of Two Adsorbents for Sugar Recovery from Biomass Hydrolyzate Yi Xie,* Diana Phelps, Chong-Ho Lee, Miroslav Sedlak,† Nancy Ho,† and Nien-Hwa Linda Wang School of Chemical Engineering, Purdue University, West Lafayette, Indiana 47907
Two polymeric adsorbents, Dowex99 and poly(4-vinyl pyridine) (PVP), have been studied for the recovery of sugars from a corn-stover hydrolyzate. The major components of the hydrolyzate are five sugars, glucose, xylose, mannose, arabinose, and galactose, and four impurities, sulfuric acid, acetic acid, hydroxymethyl furfural (HMF), and furfural. In elution chromatography in a column packed with Dowex99, the five sugars are the “center-cut”, whereas sulfuric acid elutes earlier and the other three impurities elute later than the sugars. For a column packed with PVP, the sugars elute earlier than all the impurities. The intrinsic adsorption and mass-transfer parameters of the sugars and the major impurities were obtained from elution and frontal chromatography tests of single components. The experimental elution chromatograms of the hydrolyzate are in close agreement with the simulations based on a detailed rate model and the single-component intrinsic parameters. The results indicate that other unidentified impurities in the hydrolyzate do not affect the adsorption of the identified components. The hydrolyzate sugars recovered from the batch elution chromatography processes were fermented with genetically engineered yeast. The fermentation results show that the hydrolyzate sugars recovered from the PVP columns have the highest fermentability, compared with those for an overlimed hydrolyzate and the sugars recovered from the Dowex99 columns. 1. Introduction Cellulose and hemicellulose from plants and other biomass can be hydrolyzed to produce sugars (i.e., glucose and xylose; see Figure 1). Once these sugars are separated from other impurities, they can serve as a feedstock in fermentation to produce ethanol (as fuels), lactic acid, or other valuable chemicals.1-5 The need for producing fuels and chemicals from renewable biomass has become abundantly clear over the past decade. However, the cost of producing fermentable sugars from biomass hydrolyzate using existing technology is relatively high and has been a major obstacle.2 Sulfuric acid can hydrolyze the cellulose and hemicellulose in biomass to sugars,6 but this process can generate byproducts such as acetic acid and can lead to further degradation of the xylose to furfural and of the glucose to hydroxymethyl furfural (HMF). Also, lignin and other compounds in the biomass will degrade to various phenolic compounds. If the concentrations of these compounds exceed certain threshold levels, they will be toxic to the downstream fermentation and will severely limit the usefulness of the derived sugars.7 Standard posthydrolysis process involves neutralization of sulfuric acid, usually with lime (calcium hydroxide). A study by Wooley et al.8 showed that the limed hydrolyzate gave a low ethanol yield in a fermentation test (20% of theoretical yield compared to 77% of * To whom correspondence should be addressed. Current affiliation: Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285. Tel.: (317) 651-4315. Fax: (317) 276-1403. E-mail:
[email protected]. † Laboratory of Renewable Resources Engineering, Purdue University.
Figure 1. Schematic diagram of a simplified bioethanol production from biomass.
theoretical yield from the fermentation of pure sugars). They showed that, instead of adding lime, an ion exclusion chromatography process9,10 could be used to remove acids, as well as to isolate the sugars from the biomass hydrolyzate. In this study, two polymeric adsorbents have been investigated: (1) Dowex99 and (2) a weak-base poly(4-vinyl pyridine) (PVP, Reilly Industries, Inc., Indianapolis, IN). The preferred adsorbent will be tested in a simulated moving bed (SMB) process in a separate study. Dowex99 has been used by Wooley et al. for the recovery of sugars from a hydrolyzate of yellow poplar sawdust.11 The Reilly PVP has excellent chemical and thermal stability. The maximum working temperature is up to 260 °C.12 A typical maximum working temperature for common anion-exchange resins is in the range of 60-70 °C.13 A high temperature is preferred in this
10.1021/ie049079x CCC: $30.25 © 2005 American Chemical Society Published on Web 07/07/2005
Ind. Eng. Chem. Res., Vol. 44, No. 17, 2005 6817
study to reduce the feed viscosity and inhibit microorganism growth during chromatographic separation. Moreover, several publications reported successful applications of PVP in the removal of phenols and carboxylic acids from industrial waste.14-16 Both phenols and carboxylic acids are likely to appear in the biomass hydrolyzate. For these reasons, PVP instead of other common anion-exchange resins was selected for study. The adsorption isotherms and mass-transfer parameters of the two polymeric adsorbents were estimated using single-component pulse tests and frontal tests. The parameters were then verified using a batch elution chromatography test of a corn-stover hydrolyzate provided by NREL (National Renewable Energy Laboratory). The sugars recovered in batch chromatography were then fermented using Saccharomyces cerevisiae 424A(LNH-ST) developed at LORRE (Laboratory of Renewable Resources Engineering).17-19 A standard mixture of pure sugars and an overlimed corn-stover hydrolyzate were fermented using the same procedure simultaneously. The fermentability of the overlimed hydrolyzate was the worst, and that of the sugars recovered using the PVP column was similar to that of a pure sugar mixture of similar composition. The sugars recovered using the Dowex99 column had an intermediate fermentability. 2. Theory 2.1. Isotherm Parameters. Isotherms are the most important parameters needed to design a chromatography or SMB process. The linear isotherm parameters can be obtained from a series of low-loading pulse tests. The retention time (tr) of the pulse is directly related to the linear isotherm parameter a (based on per solid volume) as shown in the following equation
u0b L ) uw ) tr b + (1 - b)p + (1 - b)(1 - p)a
(1)
where L is the column length, uw is the wave velocity, u0 is the interstitial velocity, b is the interparticle voidage, and p is the intraparticle voidage. Equation 1 shows that the retention time depends on only the interstitial velocity since the isotherm a is a constant. If a series of pulse tests at different flow rates are carried out, the isotherm parameter a (based on per solid volume) can be estimated from eq 1 with a linear regression technique. Chromatography separation is not always performed within the linear adsorption range. To increase the adsorbent productivity (or throughput per bed volume), a chromatography process is usually carried out in the nonlinear adsorption range (or high loading). The linear isotherms obtained from the low-loading pulse tests cannot accurately predict the adsorption at high loading. Other isotherm models can be used to correlate the nonlinear adsorption parameters. The Langmuir isotherm model (eq 2) is often used.20
qi )
aiCi N
1+
(i ) 1, 2, ..., N)
(2)
bjCj ∑ j)1
The shock wave velocity of a pure component with the
Langmuir isotherm can be described by the following equation21
L ) us ) tr,sh
u0b ∆q b + (1 - b)p + (1 - b)(1 - p) |C)Cp ∆C (3a)
where
∆q a | ) ∆C C)Cp 1 + bCp
(3b)
tr,sh is the shock wave retention time, and Cp is the plateau concentration. The Langmuir isotherm a and b parameters can be estimated from a series of frontal tests at different feed concentrations.20 2.2. Mass-Transfer Parameters. Mass-transfer parameters were needed in the simulation and design of both batch chromatography and SMB processes. The mass-transfer parameters included axial dispersion coefficient (Eb), film mass-transfer coefficient (kf), Brownian diffusivity (D∞), and intraparticle diffusivity (Dp). The axial dispersion coefficient was calculated from the Chung and Wen correlation22 unless noted otherwise. The film mass-transfer coefficient was estimated from the Wilson and Geankoplis correlation.23 The Brownian diffusivities of the carbohydrates and their derivatives were calculated from the Wilke and Chang correlation.24 The Brownian diffusivities of the electrolytes were found in the literature and adjusted for the operating temperature according to the Stokes-Einstein equation.25 The initial values of the intraparticle diffusivities were calculated from the Mackie and Meares correlation.26 They were fine-tuned by fitting the simulations from a rate model (described next) with the experimental data from single-component pulse and frontal tests. The aforementioned mass-transfer parameters can be related to HETP (height of a theoretical plate) in the following equation27
( )( ) ( )( )
u0b Rp u0b Rp2 HETP Eb + + ) 2 u0 1 - b 3kf 1 - b 15pDp
(4)
where Rp is the particle radius. On the right-hand side of eq 4, the three additive terms define the contributions from axial dispersion, film mass transfer, and intraparticle diffusion, respectively. Once Eb, kf, and Dp are known, the three terms can be calculated. The limiting factor for the column performance will be determined by the term that has the highest value. 2.3. Versatile Reaction and Separation (VERSE). A VERSE simulation package has been developed in the Bioseparation Laboratory of Purdue University since the 1980s. VERSE is based on a detailed rate model, which takes into account convection between particles, axial dispersion, film mass transfer, intraparticle diffusion (pore, surface, or parallel diffusion), intrinsic adsorption/desorption kinetics, competitive adsorption, and reactions occurring in the mobile phase or among solutes adsorbed on the stationary phase. The partial differential equations of the rate model and associated boundary and initial conditions are discretized by orthogonal collocation on finite elements. The resulting ODE’s and algebraic equations are solved using an equation solver. The model equations and the numerical solution procedure can be found in the literature.28,29
6818
Ind. Eng. Chem. Res., Vol. 44, No. 17, 2005
VERSE has been verified with many different systems and processes.30-34 If the adsorption isotherms and mass-transfer parameters are accurate and the assumptions of the rate model are valid, VERSE simulations give reliable predictions of transient column profiles, effluent histories, product purity, and yield. 3. Experimental Section 3.1. Materials and Equipment. Dowex99 adsorbents in calcium form were purchased from Supelco (St. Louis, MO). Poly(4-vinyl pyridine) (PVP) adsorbents, Reillex HP, were purchased from Reilly Industries, Inc. (Indianapolis, IN). Furfural, hydroxymethyl furfural (HMF), blue dextran, 0.507 N sulfuric acid aqueous solution, and pure sugars, such as glucose, xylose, galactose, etc., were purchased from Sigma-Aldrich (St. Louis, MO). Sodium chloride, sodium hydroxide (pellet), 37% hydrochloric acid, 29.8% ammonia hydroxide, and glacial acetic acid were purchased from Mallinckrodt Baker, Inc. (Paris, KY). Sodium hydroxide aqueous solution (50% w/w) was purchased from Fisher Scientific (Fair Lawn, NJ). Distilled deionized water (DDW) was obtained from a Milli-Q system by Millipore (Bedford, MA). Glass columns were purchased from Ace Glass, Inc. (Louisville, KY). Yeast extract and peptone were purchased from Difco (Becton Dickinson Microbiology Systems, Sparks, MD).The corn-stover hydrolyzate was provided gratis by NREL. A Perkin-Elmer HPLC pump (model IC200) was used to control the flow rates in the batch chromatography experiments. Temperature was controlled by either a Cole-Parmer circulation water bath (model 10106-50) or a column heating mantle from Glas-Col. The detectors used in the experiments include a Waters 431 conductivity detector, a Waters 990 PDA detector, a Waters 2414 RI detector, and a Hitachi RI detector. A New Brunswick Floor incubator (model G-25) was used to incubate fermentation flasks. The fermentation was performed in a 300 mL Nephelo culture flask with 3 baffles (Bellco Glass, Inc., Vineland, NJ). 3.2. Assays. Two different methods were used to measure the sugar concentrations. The first method used a Dionex ion chromatography system. The Dionex system consisted of an ED50 electrochemical detector, a GP50 gradient pump, and an LC30 chromatography oven. A Carbopac PA-1 analytical column (250 × 4 mm) and a guard column (50 × 4 mm) were used in the Dionex system for the analysis of sugars. The column temperature was maintained at 30 °C. The mobile phase flow rate was 1 mL/min. The sample injection volume was 25 µL and the effluent was monitored with an amperometry detector. Before injection, the column was equilibrated with 6 mM NaOH. After injection, the mobile phase remained at 6 mM NaOH for 30 min, followed by a step change to 200 mM NaOH for 15 min. Then the mobile phase composition was changed back to 6 mM NaOH to reequilibrate the column for 30 min before the next sample injection. The NaOH aqueous solution was degassed for 15 min before usage and was sparged with nitrogen during the assay period and overnight. The second method measured sugar concentrations using a Bio-Rad Aminex HPX-87H column (300 × 7.8 mm) and a guard column (30 × 4.6 mm) in a Waters HPLC system. The Waters HPLC system consisted of two 515 HPLC pumps, a 2414 RI detector, a 996 PDA detector, and an Eppendrof column heater. The column
temperature was 55 °C. The sample injection volume was 20 µL and the mobile phase was a 0.01 N sulfuric acid aqueous solution. The mobile phase flow rate scheme was as follows: 0 f 25 min 25 f 30 min 30 f 70 min 70 f 75 min
0.3 mL/min 0.3 to 0.7 mL/min (linearly increased) 0.7 mL/min 0.7 to 0.3 mL/min (linearly decreased)
The Aminex column was also used for the assays of acetic acid, HMF, and furfural. The sugars and acetic acid were analyzed by the RI detector. HMF and furfural were analyzed by the PDA detector at 205 nm. A sulfuric acid peak was also observed in the Aminex column chromatogram. However, if the sulfuric acid sample concentration was lower than 0.01 N (mobile phase concentration), a negative peak appeared. The aforementioned Dionex system can be used to measure the sulfuric acid concentration. An Ionpac analytical column (150 × 3 mm) and a guard column (30 × 3 mm) were used. The column was maintained at 30 °C with a mobile phase of 20 mM NaOH at 0.6 mL/min flow rate. The sample injection volume was 25 µL. The effluent was monitored with a conductivity detector. Xylose, glucose, acetic acid, and their fermentation products such as xylitol, glycerol, and ethanol were analyzed as reported previously16 by HPLC using HPX 87H (300 × 8 mm, Bio-Rad Laboratories, CA), equipped with an autoinjector (Hitashi, model AS-4000), an isocratic liquid pump (Hitashi, model L-6000), an RI detector (Hitashi, model L-3350), and a computing integrator (Hitashi, model D-2500). Samples (1 mL) were centrifuged, and the supernatant was collected. Analysis was performed on 10 µL of the 10-fold diluted sample. 3.3. Resin Pretreatment and Packing. Dowex99 was received in the calcium form. The resin was converted to the hydrogen form prior to use. The resin was first stored in a long column. A 5% (w/v) hydrochloric acid aqueous solution was used to wash the resin. A peristaltic pump (Pharmacia) was used to pump HCl at 10 mL/min in up-flow. After five bed volumes of acid wash, the column was then washed with five bed volumes of deionized distilled water (DDW) to remove the excess acid. The PVP resin was washed with five bed volumes of 1 N HCl, 1N NaOH, and 100% ethanol. The resin was washed with DDW before each solvent change and finally equilibrated in DDW. A slurry packing technique was applied to pack the columns. The resin slurry was poured into an empty column quickly. If the slurry was slowly added into the column, the resin could be unevenly distributed; large particles would settle at the lower part of the column, and fine particles would settle at the upper part. After the excess mobile phase was removed, the packing was packed tightly by pumping in the mobile phase at the maximum flow rate that would be used in the experiments. An hour at the high flow rate was usually sufficient to achieve stable packing. If a gap appeared between the resin top and the column filter after compression, more resin slurry would be added, followed by recompression. 3.4. Void Fraction Measurement. Blue dextran pulses were injected to estimate the interparticle void fraction (b) of the columns packed with Dowex99. The concentration of the blue dextran aqueous solution was
Ind. Eng. Chem. Res., Vol. 44, No. 17, 2005 6819
2 g/L, and the pulse injection volume was 2 mL for a large column (60 × 2.65 cm) and 20 µL for a small column (12.5 × 1.5 cm). An FPLC pump (Pharmacia) was used to deliver the mobile phase (DDW) at 10 mL/ min for the large column and at 1 mL/min for the small column. To reduce backpressure caused by the detector, a flow-splitter was connected at the outlet of the column. A side-stream of 1 mL/min was delivered to a Waters 990 PDA detector. Effluent histories were monitored at 620 nm. Interparticle void fraction was calculated from the retention time of the blue dextran peak. The particle porosity (p) was reported by Wooley et al.11 Sodium chloride is a small solute and does not adsorb on PVP. It was used to measure the total column void fraction (t) of the PVP columns. The concentration of NaCl was 0.1 N and the injection volume was 1.1 mL. The flow rate was 1.15 mL/min. The effluent was monitored with a Waters 431 conductivity detector. Once t was measured from the NaCl pulse, b was calculated from t and p (t ) b + (1 - b)p). The porosity (p) of PVP was reported by Wu et al.35 3.5. Single-Component Pulse Tests. Wooley et al. indicated that linear isotherms could be used to describe the adsorption of sugars and acid by the Dowex99 resin. Pulse tests are sufficient to obtain the isotherms. All 10 identified compounds in the biomass hydrolyzate were tested separately. The concentration of each component was kept the same as that of the corn-stover hydrolyzate. The small column (12.5 × 1.5 cm) packed with the Dowex99 resin was used. The mobile phase flow rate was 1 mL/min, and the injection volume was 20 µL. The column was immersed in a circulation bath kept at 65 °C. The eluent (DDW) was also maintained at 65 °C. A Waters refractive index (RI) detector was used to monitor the sugars, sulfuric acid, and acetic acid. Furfural and HMF were monitored with the PDA detector at 280 nm. 3.6. Single-Component Multiple Frontal Tests. Multiple frontal tests were used to measure the isotherms of individual components adsorbed onto the PVP resin. A Perkin-Elmer HPLC pump was used to deliver the solutions. One inlet stream was DDW and the other was the measured component at its maximum concentration. The two streams were mixed in the pump and loaded into the small column, which was equilibrated with DDW prior to the experiments. The output ratio of the two streams was controlled so that solutions of different concentrations were generated. The multiple frontal experiments started from the lowest concentration. Once the column was saturated with one concentration, the pump output ratio was changed to generate a higher concentration. Thus, the column was saturated at increasingly higher concentrations, resulting in a breakthrough curve between two consecutive concentration plateaus. The mobile phase, column, and sample solutions were kept at 65 °C. The effluent was monitored with a Waters 996 PDA and a Waters 2414 RI detector. The mass center of each breakthrough curve was used to calculate the amount of the adsorbed solutes as follows
qi ) qi-1 +
(Ci - Ci-1)F(tr,i - td - t0) BV
(5)
where qi is the adsorbed amount at the ith concentration (Ci); F is the volumetric flow rate; tr,i is the mass center of the ith breakthrough; td is the time for the mobile
phase to pass through the extra-column dead volume; t0 is the total void time (or time for the mobile phase to fill the interparticle void and the intraparticle void); and BV is the bed volume. 3.7. Batch Chromatography Experiments with Corn-Stover Hydrolyzate. The batch chromatography experiments with the corn-stover hydrolyzate had two purposes. One was to validate the intrinsic engineering parameters, and the other was to recover sugars from the hydrolyzate for fermentation tests. Two water-jacketed Ace glass columns (60 × 2.65 cm) were packed separately with Dowex99 and PVP, respectively. A Cole-Parmer circulation bath was used to maintain the column temperature at 65 °C. The feed and the eluent (DDW) reservoirs were kept at 65 °C with a VWR utility bath. A Perkin-Elmer pump was used for loading and elution. A Pharmacia LKB P-1 peristaltic pump was used for column regeneration. A flow splitter was connected to the column outlet, and the side-stream was connected to a Waters 990 photodiode array detector, which was used to monitor the effluent at 280 nm. Prior to the chromatography tests, the hydrolyzate was filtered with 0.2 µm filter paper and heated to 65 °C. The column was equilibrated with DDW before loading. The feed was delivered into the column by the Perkin-Elmer pump. The loading volume was 52.8 mL for the PVP column and 20 mL for the Dowex99 column. The flow rate was 8.8 mL/min for the PVP column and 8.5 mL/min for the Dowex99 column. After loading, the PVP column was eluted with DDW at the same loading flow rate for 300 min, while the Dowex99 column was eluted with DDW for 100 min. Samples were collected every minute for HPLC assay during the loading and elution steps. The batch chromatography experiment with the Dowex99 column was stopped after the DDW elution. The PVP column, however, needed caustic regeneration after the elution step. The PVP column was regenerated with 0.5 N NaOH at 8.5 mL/min for 80 min. Samples were collected for every 2 min and analyzed for sulfuric acid during the caustic regeneration step. After the regeneration, the PVP column was washed with more than four bed volumes of DDW until the effluent reached pH 7.0. 3.8. Concentration of Sugar Samples Prior to Fermentation. If the total sugar content in the collected sugar samples was
