Jeung-yil Park

A novel lime pretreatment for subsequent bioethanol production from rice straw--calcium capturing by carbonation (CaCCO) process.

In order to establish an efficient bioethanol production system for rice straw, we developed a novel lime-pretreatment process (CaCCO process) that did not require a solid-liquid-separation step. This process adopted a step in which after pretreatment lime was neutralized by carbonation, resulting in a final pH of about 6. CaCO(3) produced by the process was kept in the reaction vessel, and no significant inhibitory effects on enzymatic saccharification and fermentation were observed. In the CaCCO process, solubilized carbohydrates, such as xylan, starch, and sucrose were also kept in the vessel, enabling high recoveries of monomeric sugars. Simultaneous saccharification and fermentation (SSF) of pretreated rice straw, 10% (g-rice straw/g-water), using Saccharomyces cerevisiae and Pichia stipitis yielded 19.1 g L(-1) ethanol that was 74% of the theoretical yield from glucose and xylose. Thus, this process represents a novel pretreatment method to utilize not only cellulose but also xylan, starch, and sucrose from biomass.

Efficient Recovery of Glucose and Fructose via Enzymatic Saccharification of Rice Straw with Soft Carbohydrates

Soft carbohydrates, defined as readily-recoverable carbohydrates via mere extraction from the biomass or brief enzymatic saccharification, were found in significant amounts in rice straw as forms of free glucose, free fructose, sucrose, starch, and β-1,3-1,4-glucan. In this study, we investigated their amounts in rice straw (defined as culm and leaf sheath), and developed an easy method for glucose and fructose recovery from them with heat-pretreatment and subsequent 4-h enzymatic saccharification with an enzyme cocktail of cellulase and amyloglucosidase. The recovery of glucose and fructose exhibited good correlation with the amounts of soft carbohydrates. The maximum yields of glucose and fructose in the rice straw per dry weight at the heading stage and the mature stage were 43.5% in cv. Habataki and 34.1% in cv. Leafstar. Thus, rice straw with soft carbohydrates can be regarded as a novel feedstock for economically feasible production of readily-fermentable glucose and fructose for bioethanol.

Bioethanol production from rice straw by a sequential use of Saccharomyces cerevisiae and Pichia stipitis with heat inactivation of Saccharomyces cerevisiae cells prior to xylose fermentation.

In order to establish an efficient bioethanol production system from rice straw, a new strategy to ferment the mixture of glucose and xylose by a sequential application of Saccharomyces cerevisiae and Pichia stipitis was developed, in which heat inactivation of S. cerevisiae cells before addition of P. stipitis was employed. The results showed that heating at 50°C for 6h was sufficient to give high xylose fermentation efficiency. By application of the inactivation process, 85% of the theoretical yield was achieved in the fermentation of the synthetic medium. At the same time, the xylitol production was reduced by 42.4% of the control process. In the simultaneous saccharification and fermentation of the lime-pretreated and CO(2)-neutralized rice straw, the inactivation of S. cerevisiae cells enabled the full conversion of glucose and xylose within 80 h. Finally, 21.1g/l of ethanol was produced from 10% (w/w) of pretreated rice straw and the ethanol yield of rice straw reached 72.5% of the theoretical yield. This process is expected to be useful for the ethanol production from lignocellulosic materials in the regions where large-scale application of recombinant microorganisms was restricted.

RT-CaCCO process: an improved CaCCO process for rice straw by its incorporation with a step of lime pretreatment at room temperature.

We improved the CaCCO process for rice straw by its incorporation with a step of lime pretreatment at room temperature (RT). We firstly optimized the RT-lime pretreatment for the lignocellulosic part. When the ratio of lime/dry-biomass was 0.2 (w/w), the RT lime-pretreatment for 7-d resulted in an effect on the enzymatic saccharification of cellulose and xylan equivalent to that of the pretreatment at 120°C for 1h. Sucrose, starch and β-1,3-1,4-glucan, which could be often detected in rice straw, were mostly stable under the RT-lime pretreatment condition. Then, the pretreatment condition in the conventional CaCCO process was modified by the adaptation of the optimized RT lime-pretreatment, resulting in significantly better carbohydrate recoveries via enzymatic saccharification than those of the CaCCO process (120°C for 1 h). Thus, the improved CaCCO process (the RT-CaCCO process) could preserve/pretreat the feedstock at RT in a wet form with minimum loss of carbohydrates.

Culm in Rice Straw as a New Source for Sugar Recovery via Enzymatic Saccharification

Rice straw was manually dissected and two main fractions were recovered: a culm and a leaf sheath/blade fraction, in order to evaluate their potential as feedstocks for the recovery of fermentable sugars. In the case of cv. Koshihikari and Milkyqueen, most soft carbohydrates (SCs: glucose, fructose, sucrose, starch, and β-1,3-1,4-glucan) were present in the culms, reaching 47.9% and 89.2% of total SCs in the two main fractions. The results also indicated that β-glucans (cellulose and β-1,3-1,4-glucan) and xylan in the culms were more susceptible to direct enzymatic attack than those in the leaf sheath/blades. Thus the culm has high potential as a new feedstock for the extraction of fermentable sugars in a concentrated form, as compared to whole rice straw and the leaf sheath/blade. In this study, a novel method of separating a culm from the whole rice straw by means of wind power was also evaluated.

DiSC (direct saccharification of culms) process for bioethanol production from rice straw.

A simple process (the direct-saccharification-of-culms (DiSC) process) to produce ethanol from rice straw culms, accumulating significant amounts of soft carbohydrates (SCs: glucose, fructose, sucrose, starch and β-1,3-1,4-glucan) was developed. This study focused on fully mature culms of cv. Leafstar, containing 69.2% (w/w of dried culms) hexoses from SCs and cellulose. Commercially-available wind-separation equipment successfully prepared a culm-rich fraction with a SC recovery of 83.1% (w/w) from rice straw flakes (54.1% of total weight of rice straw). The fraction was suspended in water (20%, w/w) for starch liquefaction, and the suspension was subjected to a simultaneous saccharification and fermentation with yeast, yielding 5.6% (w/v) ethanol (86% of the theoretical yield from whole hexoses in the fraction) after 24h fermentation. Thus, the DiSC process produced highly-concentrated ethanol from rice straw in a one vat process without any harsh thermo-chemical pretreatments.

Controlled Preparation of Cellulases with Xylanolytic Enzymes from Trichoderma reesei (Hypocrea jecorina) by Continuous-Feed Cultivation Using Soluble Sugars

The objective of this study was to develop an efficient production system for cellulase preparation with a high level of xylanolytic enzymes using soluble carbon sources. When xylose and arabinose were simultaneously fed with glucose and cellobiose, a mutant of Trichoderma reesei, M3-1, showed sufficient levels of cellulolytic and xylanolytic activities, indicating that xylose and arabinose are good inducers for the production of xylanolytic enzymes. In a continuous feeding experiment using glucose/cellobiose and glucose/xylose/cellobiose, cellulase preparations with various levels of xylanolytic enzymes were obtained by altering the feeding solutions and the timing of their addition. The volumetric production rates for xylanolytic activities at the glucose/xylose/cellobiose-feeding phase were significantly higher than at the glucose/cellobiose-feeding phase, while those for cellulolytic activities were comparable under the two conditions. Thus the composition of the enzyme preparation produced by the mutant was readily controlled by varying the inducers and the pattern of their addition, facilitating the tailored production of enzymes in a diversity of bioconversion processes.

Characterization of Starch Granules in Rice Culms for Application of Rice Straw as a Feedstock for Saccharification

Rice plants are known to accumulate starch in leaf sheaths and culms, and in some cultivars significant amounts of starch are present at the mature stage. This can be considered as potential feedstock for the recovery of fermentable sugars. We isolated starches from the culms of cultivars Yumeaoba, Koshihikari, and Leafstar to investigate their structural and physical features. Yumeaoba culm starch contained 20.2% amylose, whereas Koshihikari and Leafstar contained 25.8% and 25.2%. Yumeaoba culm starch was found by chain-length distribution analysis to contain higher amounts of short chains, resulting in lower gelatinization temperature by 7 °C, as compared to Koshihikari and Leafstar. Consequently, the rate of enzymatic hydrolysis of Yumeaoba culm starches reached maximum at a lower temperature than Leafstar. Rice culm starch, with a lower gelatinization temperature, can provide an advantageous material for feedstock for bioethanol production in terms of energy conservation.

Readily-milled fraction of wet sugarcane bagasse as an advanced feedstock for monosaccharide production via the RT-CaCCO process

The RT-CaCCO process for enzymatic saccharification was applied to readily-milled fractions of wet sugarcane bagasse. Wet bagasse immediately after juice extraction was crushed with shark-mill blades to prepare two fractions referred to as readily-milled (RF) and hardly-milled fraction (HF). Monosaccharide recoveries from RFs via the RT-CaCCO process were 1.03-1.21 times higher than those from HFs. Moreover, when the wet weight ratio of RF/HF was adjusted to 2/8, the hexose recovery from RF was 90.9%, which was 1.3 times higher than that of the wet bagasse before fractionation. The results show that this process can be used for efficient monosaccharide recovery from RF of wet bagasse. In addition, the process can be adapted to more fibrous HF for multiple uses such as fuel for boilers and fibers for particleboards.

An Improved CARV Process for Bioethanol Production from a Mixture of Sugar Beet Mash and Potato Mash

A mixed mash of sugar beet roots and potato tubers with a sugar concentration of 23.7% w/v was used as a feedstock for bioethanol production. Enzymatic digestion successfully reduced the viscosity of the mixture, enabling subsequent heat pretreatment for liquefaction/sterilization. An energy-consuming thick juice preparation from sugar beet for concentration and sterilization was omitted in this new process.