Autohydrolysis를 이용한 섬유성 바이오매스의 전처리

공창범, 최실호, 우창호, 윤현희

Pretreatment of Lignocellulosic Biomass by Autohydrolysis

Chang-Bum Kong, Shil Ho Choi, Chang Ho Wu, Hyon Hee Yoon

Department of Chemical Engineering, Kyungwon University, Seongnam 461-701, Korea

INTRODUCTION

Pretreatment is a necessary element in bioconversion of lignocellulosic materials to fuels and chemicals. The primary purpose of pretreatment is to make the lignocellulosic biomass accessible and reactive to allow high rates and yields on enzymatic hydrolysis. For the production of ethanol, product yield has a strong influence on process economics[1]. With the increasing availability of organisms capable of converting pentoses to ethanol[2], high recovery of hemicellulose sugars is a key feature of leading pretreatment methods.

When biomass is heated to a high temperature (above 150℃) with pressurized water, acids are formed by solubilization of acidic components and de-esterification of ester groups(like acetate groups) in hemicellulose. These acids catalyze the hydrolysis of the hemicellulose[3], and thus pretreatment of biomass with hot water is called autohydrolysis. The acids formed in autohydrolysis are weak acids (such as formic acid, acetic acid, and glucuronic acid etc.)[4] and are not strong enough to hydrolyze cellulose, therefore, autohydrolysis can selectively extract hemicellulose from the biomass[5]. Hemicellulose in biomass could be an important barrier to enzymatic hydrolysis of cellulose. The removal of hemicelluose from biomass, therefore, would be a prerequisite for complete hydrolysis of cellulose and recovery of hemicellulose.

The optimum conditions for autohydrolysis was investigated in terms of maximazing the solubilization and recovery of hemicellulose sugars. The effect of hemicellulose on enzymatic digestibility was also investigated.

MATERIALS AND METHODS

1. Materials

Hybrid poplar milled to the nominal size of 80 to 60 mesh was used as feedstock. Filte paper(Whatman No.1) and □-cellulose(Aldrich) was used as a reference substrate.  The cellulase enzyme, Spezyme-CP, was obtained from Environmental Biotechnology, Inc. The specific activity of the enzyme was determined by the supplier is as follows: filter pater activity = 64.5 FPU/mL,  □-glucosidase activity = 57.6 p-NPGU/mL. Birch wood xylan(Sigma) was used in hydrolysis experiment.

2. Experimental setup and operation

The process involves treating biomass with water at temperature above 150℃ and pressure around 325 psi. In the pretreatment operation, 5-7 g of biomass sample was packed into the reactor. For the autohydrolysis, two types of operation mode were employed, batch and percolation process. In batch operation, water was pumped into the reactor prepacked with biomass and stopped pumping during reaction. The liquid to solid weight ratio in the reactor was about 7-10. In the percolation process, a constant flow of water(1.0 mL/min) was applied during reaction.

3. Digestibility test and analytical methodes

Enzymatic hydrolysis of pretreated substrates was performed in 250mL glass bottle at 50℃ and pH 4.8(0.05 M sodium citrate buffer) on a shaker bath agitated at 150 rpm. The enzyme loading was 30 IFPU/g glucan, and the initial glucan concentration was 1%(v/v). Samples were taken periodically and analyzed for glucose content. The total glucose content after 72 hr of hydrolysis was taken to calculate the enzymatic digestibility.

The solid biomass samples were analyzed for sugar and Klason lignin content following the procedure described in NREL-CAT Standard Procedures.

RESULTS AND DISSCUSIONS

1. Effect of Hemicellulose on Cellulose Hydrolysis

In the previous study on ARP, it was found that the enzymatic digestibility of lignocellulosic biomass was greatly inhanced by removal of hemicellulose and lignin. Since the hemicellulose and lignin were removed simultaneously in the ARP process, the isolated effect of them could not be identified. The role of lignin in the enzymatic hydrolysis of lignocellulosic biomass has long been recognized. However, the role of hemicellulose is still uncertain. In this work, the effect of hemicellulose on the enzymatic digestibility was investigated. The enzymatic hydrolysis of cellulose was conducted at various levels of enzyme loading with and without hemicellulose addition. As shown in Fig. 1, the cellulose hydrolysis under the enzyme loadings of 10 and 20 IFPU/g cellulose was suppressed by the presence of hemicellulose. As the enzyme loading was increased to 40 and 60 IFPU/g cellulose, the effect of hemicellulose diminished.

2. Effect of Temperature in Autohydrolysis

The effect of reaction temperature in autohydrolysis of hybrid poplar was investigated over 170-200℃. Autohydrolysis experiment was carried out under batch and percolation mode. Reaction time was kept for 1 hr for all runs. Table 1 summarizes the composition data for the remaining solids after autohudrolysis. As shown in Table 1, hemicellulose(xylan) was effectively solubilized(or removed) by the autohydrolysis pretreatment. A higher hemicellulose solubilization performance was observed in batch process than percolation process, probably due to the more acidic environment in the batch

Table 1. Effect of reaction temperature on composition of solid residues in

autohydrolysis operated under percolation and batch Modes

Temp., ℃	Percolation Mode					Batch Mode
	% Solid Remaining	% Lignin	% Glucan	% Xylan		% Solid Remaining	% Lignin	% Glucan	% Xylan
Untreated 170 180 190 200	100 73.70 66.10 59.60 61.50	26.73 16.80 18.30 15.70 18.50	43.52 42.50 42.30 38.55 39.36	14.43 6.60 3.40 2.20 1.80		100 79.40 71.20 70.40 67.40	26.73 26.40 22.90 25.40 26.70	43.52 42.90 40.20 38.60 35.50	14.43 6.10 3.10 1.50 trace

process. The results collectively indicate that the reaction temperature is an important operating variable in autohydrolysis. As reaction temperature increased, xylan remaining in the treated solid decreased, that is, the amount of hemicellulose solubilized increased. At 200℃, the extents of hemicellulose solubilized in the percolation and batch process were 88% and about 100%, respectively. During the autohydrolysis, some of lignin was also removed from the biomass. As much as 41% of lignin was removed by the percolation process. However, no significant amount of lignin was removed by the batch process. Due to the nature of the percolation process, the solubilized lignin was removed immediately from the reactor and re-precipitating on the substrate was prevented.

Fig. 2 shows the enzymatic digestibility of biomass pretreated by autohydrolysis. The biomass samples pretreated by autohydrolysis in either batch or percolation process exhibit enhanced enzymatic digestibility of cellulose. The cellulose digestibility increased with the reaction temperature of autohydrolysis. The digestibility at 72 hr of enzymatic hydrolysis increased from 41.5 to 92.5% as autohydrolysis temperature in percolation process increased from 170 to 200℃. Similar result was observed for samples treated by batch autohydrolysis. An important observation is that when the pretreatment temperature was above 190℃ in the batch process, almost no lignin was removed from the biomass, but the treated substrates exhibited very high digestibility close to that of the filter paper. These results indicate that thermal deformation of lignin, and not necessarily removal, may be a significant factor affecting the digestibility of biomass. The overall results indicate that the pretreatment temperature is a very important factor in autohydrolysis.

REFERENCES

[1] Lynd, L. R., Appl. Biochem. Biotechnol., 24/25, 695-719 (1990).

[2] Bealle, D.S., Ohta, K. and Ingram, I. O., Biotechnol. Bioengng., 38, 296-303      (1991).

[3] Lora, J. H. and Wayman, M., Tappi J., 61, 47-50 (1978)

[4] McGinnis, G. D., Wilson, W. W. and Mullen C. E., Ind. Eng. Chem. Prod.      Res. Dev., 22, 352-7 (1983).

[5] Mok, W.S.L. and Antal, M.J., Ind. Eng. Chem. Res., 31, 1157-61 (1992).

Figure 1. Enzymatic digestibility of hybrid poplar treated by autohydrolysis under batch(A) and percolation(B) operarion mode at various  temperatures.

Figure 2. Effect of hemicellulose on enzymatic digestibility of cellulose at different enzyme loading. weight ratio of hemicellulose to cellulose = 0.4 : 1.