Direct saccharification of lignocellulosic biomass by hydrolysis with formic acid solution

2.1 Samples

Japanese cedar trees were used as a raw lignocellulosic biomass material. A commercial microcrystalline cellulose (Nakalai Tesque Co., Kyoto, Japan) was also used to examine the hydrolysis behavior. The Japanese cedar tree was crushed into chips. All samples were dried in vacuo for 24 h at 70°C prior to use. The ultimate analyses of these samples are listed in Table 1.

2.2 Batch reaction system

An aqueous solution (21 ml), with a certain concentration of organic acid, was inserted into a stainless steel batch reactor (Swagelok, USA) which contained 0.5 g of the sample. The experiments were performed by using a stainless-steel batch reactor with an inner diameter of 11 mm and a length of 95 mm. The reactor was then heated by using an oil bath, at the fixed temperature for reaction time t=2 h. After the reaction, the product solution was filtered by suction filtration with filter paper (pore diameter 0.1 µm) and the solid residue was desiccated by using a vacuum oven for 24 h, to measure the weight by electronic balance. The organic acid aqueous solutions were acetic acid aqueous solution (Wako Pure Chemical Industries, Osaka, Japan) or formic acid aqueous solution (Wako Pure Chemical Industries).

2.3 Solvent flow reaction system

Figure 1 shows the schematic of the experimental set-up. The sample (0.3 g) was inserted onto the filter in the reactor. The experiments were performed by using a stainless-steel flow reactor, with an inner diameter of 4.6 mm and a length of 82 mm. The solvent was fed into the reactor with the LC pump. The pressure was held at 2 MPa with the back pressure regulator. Then, the flow rate was set to 0.2 ml/min. The residence time was 6.8 min. The reactor was then immersed into the oil bath with a certain temperature and the formic acid aqueous solution was fed for a reaction time t=2 h. Then, the reactor was taken out from the oil bath and cooled down with iced water. Distilled water was then fed to collect all of the remaining product solution in the tube.

Figure 1

Schematic apparatus of the solvent flow reaction system.

2.4 Analysis

Water-insoluble components were measured as follows. Soluble products were dissolved in formic acid aqueous solution and the solid solubles were obtained by evaporation of the solvent. We termed water-soluble parts “saccharides” and water-insoluble parts “water-insolubles”. This water-insoluble deposit was weighed and water-soluble components were quantified by the difference. The chemical structure, such as a functional group, was analyzed by an FTIR spectrometer (JIR-SPX60, JEOL. Ltd., Tokyo, Japan). All IR spectra were measured at 4 cm^-1 resolution, and were collected by acquisition of 100 scans. The product solution was analyzed to measure the glucose and xylose concentrations by using HPLC, and compared with the standard solution. Meanwhile, the solid residue was collected by suction filtration and desiccated by using a vacuum oven at 70°C for 24 h, to measure the weight with an electronic balance. The ultimate analysis of the residue was then carried out with an elemental analyzer. The experimental conditions were concentration C=25 and 50 vol%, T=150, 175 and 200°C and t=2 and 4 h. The solution from the hydrolysis of cedar was analyzed using gel permeation chromatography (GPC).

3.1 Effect of acidity of two organic acids on hydrolysis of cellulose

First, we examined the effect of the acidity of the organic acid on the hydrolysis behavior, using a batch reactor. Woody biomass contains cellulose, hemicellulose and lignin. Organic acids act as reagents for the hydrolysis of polysaccharides and also as solvents for the extraction of lignin. It is expected that the hydrolysis behavior of lignocellulosic biomass is complex, so we started the study on the hydrolysis of crystalline cellulose with an organic acid solution. For comparison, we also conducted hydrothermal solubilization or decomposition with hot compressed water. The properties of the liquid products change with time, so the fresh liquid products were analyzed immediately after collection [14]. As a result, only the formic acid aqueous solution decomposed cellulose effectively below 200°C. Distilled water and acetic acid aqueous solution did not show any sign of decomposition of cellulose below 200°C. Solid residue yields after 50 vol% formic acid hydrolysis were given as follows: 95.5% at 150°C, and 76.1% at 175°C. Formic acid has the lowest pKa=3.75 (4.75 for acetic acid) and thus it is the strongest organic acid to hydrolyze cellulose. Meanwhile, the boiling point of formic acid is almost the same (100.8°C) as that of water. Therefore, it is possible to recover and easily recycle formic acid after the reaction, by just evaporating or by distillation at around 100°C. Based on these two merits, formic acid aqueous solution was chosen as the main reagent for the hydrolysis of cellulose and lignocellulosic biomass.

3.2 Hydrolysis of cellulose with formic acid and comparison between reactor systems

Next, we investigated the effect of the reaction temperature and compared the reactor systems. The results are shown in Figure 2. With the increase in reaction temperature (up to 200°C), the solid residue yield decreased and the soluble components yield increased. This is mainly due to the increase in the hydrolysis rate of cellulose. However, the decomposition of glucose occurs simultaneously. The glucose yield increased below 175°C, then decreased above this temperature. It is known that glucose is converted into 5-hydroxymethyfurfural or levulinic acid at higher temperatures around 200°C [15]. Therefore, a maximum glucose yield can be found at 175°C. When comparing the results of the batch reactor and those of the flow reactor, the glucose and soluble yields of hydrolysis using the flow reactor were higher than those of hydrolysis using the batch reactor. This difference can be explained by the contact efficiency between the solvent and cellulose. A sufficient amount of flowing solvent could well wash out solubles and suppress the secondary decomposition of glucose. A secondary reaction does not seem to occur much in this residence time [16]. Therefore, it can be concluded that the flow reactor showed the behavior of the primary hydrolysis of solid biomass more clearly than the batch reactor did.

Figure 2

Product distributions through the hydrolysis of cellulose (2 h, 50 vol% formic acid).

3.3 Hydrolysis of wood chips with formic acid (aq)

As mentioned above, organic acid hydrolysis of lignocellulosic biomass brings about the depolymerization of polysaccharides and the elution of lignin. Furthermore, the hydrolysis of hemicellulose into soluble saccharides, such as xylose, starts at a lower temperature than that of cellulose. Figure 3 shows the distributions of the products through formic acid hydrolysis of the cedar chips, using the flow reactor. At a high temperature and with high formic acid concentration, the hydrolysis and solubilization of wood chips were promoted. With the 50 vol% formic acid solution, the solid residue yield decreased to 7% at 200°C. This proved that the treatment with formic acid was an effective method for direct hydrolysis and solubilization of all components of wood. However, the yields of monosaccharides, such as glucose and xylose, were not so high. This result suggests that the depolymerization into monomer is almost balanced with the secondary decomposition of monomer. Under these conditions, a very small amount of black solid was deposited on the filter that was set upstream of the regulator valve. From the elemental analysis and FTIR measurements, this deposit turned out to be lignin fraction. This shows that lignin was slightly eluted in the hot reactor and washed down, cooled, and deposited on the filter at room temperature. Figure 4 summarizes the soluble saccharide yield, which includes the sugar derived from hemicellulose and the yield of lignin eluted under each condition. The optimal condition of direct saccharification was studied by examining the change in saccharification and the elution behavior with temperature. Lignin was eluted as a soluble product at high temperature. The lignin content in the residue was high at a high temperature, although the residue yield was low. This indicates that in order to suppress lignin elution, mild conditions (a low temperature and low acid concentration) are desirable. Saccharification mainly proceeded at temperatures between 120°C and 175°C. By contrast, lignin was eluted at temperatures >150°C. At 150°C, high saccharides yield, with little elution of lignin, could be obtained. Based on this idea, approximately 47 wt% of soluble saccharides, keeping a minimum lignin elution, was successfully obtained at 150°C with the 25 vol% formic acid solution. As compared with the soluble yield at 150°C from cellulose in Figure 1, the saccharide yield from wood chips was obviously higher, excluding the soluble hemicellulose even under the same conditions. This is because cellulose of woody chips contains amorphous parts. Hydrolysis of these amorphous parts of cellulose start at a low temperature [17]. It is a key factor to relax the rigid structure of biomass for easy saccharification at a low temperature. This can be considered as one of the direct saccharification options that does not require a lignin separation process.

Figure 3

Product distributions through the hydrolysis of wood chips.

Figure 4

Change in the yield of (A) saccharide and (B) lignin eluted with the temperature and the formic acid concentration.

3.4 Effect of organic acids on hydrolysis of wood chips and the possibility of lignin separation

Hydrolyses of wood chips with formic acid or acetic acid were carried out and the effects of organic acid type were examined again for woody biomass. The results are shown in Figure 5. Hydrolysis with formic acid solubilized more wood chips than with acetic acid. This is because the acidity of formic acid is much higher than that of acetic acid. On the contrary, the water-insoluble components yield was higher for the acetic acid hydrolysis. The water-insoluble components correspond to the matter which was solubilized into 50 vol% acetic acid solution, but was deposited with pure water. From this result, it is estimated that acetic acid works as a good solvent for the elution of lignin. Therefore, formic acid is favorable for obtaining much water-soluble saccharides from woody biomass, as expected. GPC measurements of water-soluble components by formic acid hydrolysis showed that the molecular weight of the main water-soluble was approximately 1000, which corresponds to the hexaose (oligomer which consists of six monosaccharides). For the acetic acid hydrolysis, xylotriose was the main water-soluble component, because of hemicellulose saccharification.

Figure 5

Product distributions through the hydrolysis of wood chips with the two types of acids.

Lastly, we tried the severe conditions for the hydrolysis of wood chips. Figure 6 shows the products yields for a long period, 4 h. Wood chips were solubilized almost completely and a large amount of water-soluble product was obtained with a 50 vol% formic acid solution at 175°C. However, the solubilization of wood chips was not so high (60%) and the water-soluble oligosaccharide yield was small with 25 vol% formic acid solution at 150°C. This indicates that severe conditions enable a nearly complete hydrolysis in a high saccharide yield. However, a large amount of lignin was simultaneously eluted. Therefore, separation of lignin at the subsequent stage is required in this case.

Figure 6

Product distributions through the hydrolysis of wood chips under severe conditions (4 h).

The possibility of lignin separation was investigated. The hydrolysis residues from wood chips under severe conditions, water-soluble products and water-insoluble components, were examined by using FTIR and the elemental analyzer. From the results, the solid residue and the water-insoluble components contained lignin, but the water-soluble ones did not, which suggests that lignin fraction is water-insoluble and only exists in residue and water-insoluble extracts. Therefore, by diluting or evaporating formic acid aqueous solution, lignin can be recovered and separated from water-soluble saccharides as a deposit.