Truong Thi Hoa

称号及び氏名博士（工学）

学位授与の日付

平成 30 年 3 月 31 日論

「 Utilization of Oil-Rich Biomass for the Production ofBiodiesel Fuel and Exploration of the Multi-BeneficialComponents 」

文

論文審査委員

名

竹中

規訓教授

小川昭弥教授長岡勉教授

論文要旨
The great attentions have been received on biomass energy due to its benefits on sustainabledevelopment. It gives opportunities in energy security, social and economic development, climate change
mitigation, and reduction of environmental and health impacts. Biomass has a large potential, which meetsthe goal of reducing greenhouse gases and could insure fuel supply in the future. The oils come from theoil-rich biomass divided in two groups: edible and non-edible oil. Focus should be shifted to exploit not
only non-edible resources that do not compete with edible vegetable oil in market, but also
multi-beneficial compounds in oil plants. It is imperative to have as much information on biochemical
applications and pharmaceutical uses as possible, so that the potential of plant could be utilized efficiently.
Candlenut tree (Aleurites moluccana) of which seeds contained 20  30 % oil is a resource of non-edibleoils, and its oil has been used for pharmaceutical, cosmetic, industrial, and dietary purpose. Jatropha
curcas is a multi-purpose and perennial plant belonging to a spurge Euphorbiaceae family. Its seed kernels
contain up to 60 % oil that is non-edible. Rice bran contains not only an edible oil (20 %), but also highlevels of various antioxidants, such as vitamin E, γ-oryzanol, and phenolic acids (especially ferulic acid(FA)). In Vietnam, an estimated 4.2 million tons of rice bran are generated each year. Much of rice bran
has been discarded, and thus represents a large untapped potential source of a valuable commodity.
Knowing their multi-beneficial compounds in oil-rich seeds and crops must be necessary for evaluating
the price of raw materials, the quality of BDF and improving its economic efficiency.

In this study, the exploration of antioxidants in rice bran, and utilization of valuable compounds ofby-products (soapstock) from rice bran oil processing is discussed. In addition, the BDF production fromcandlenuts oil, the behavior of toxic phorbol esters (PEs) in Jatropha curcas oil and BDF, and how toanalyze these kinds of toxic compounds are also concerns. The utilization of oil-rich biomass is an
important driving force for the socio-economic development by means of efficient uses of these resourcesfrom the point of view of biodiesel production associating with the exploration of multi -beneficialcompounds. More specific discussions are outlined below.
In Chapter 1, general considerations on renewable energy, especially on biomass energy aresummarized to highlight how they are important for human life and climate changes in aspect ofsustainable developments. Then, the overview of biomass-based oil and biodiesel production is presented.
Markedly, the utilization of biomass to improve its economy by exploring multi-beneficial compounds
including the potential pharmaceutical compounds in biomass sources (rice bran) and toxic ones in
non-edible oil seed (candlenut and Jatropha curcas) are summarized. The final part of this chapter
presents the objective and organization of this thesis.
In Chapter 2, various methods for simultaneous extraction of tocols (tocopherols and tocotrienols)and γ-oryzanol, and FA from rice bran were examined using binary solvents of ethyl acetate and water,
The former tocols and γ-oryzanol were distributed into the upper organic phase, and the latter FA was intoaqueous phase. The procedures examined were as follows: alkaline treatment (Alk), ultrasound -assisted
extraction (UAE), and Alk combined with UAE (Alk+UAE). The experiments were conducted by va rying
the temperature and the extraction duration. The maximum yields of tocols and γ-oryzanol were obtained
at 25 ºC over a time span of 30 min. Upon raising the temperature to 80 ºC, the yield of FA increased
dramatically, while the recovery of γ-oryzanol slightly decreased. Employing the Alk+UAE procedure at
25 ºC, the concentrations of tocols, -oryzanol, and FA from various species of white rice brans in
Vietnam and Japan were in the ranges of 146 – 518, 1591 – 3629, and 352 – 970 µg / g, respectively.
In Chapter 3, the alkaline hydrolysis of γ-oryzanol was investigated using a homogeneous reaction
system in order to obtain the maximum yield of FA. The parameters examined were as follows: ratios ofpotassium hydroxide (KOH) to γ-oryzanol and ratios of ethanol (EtOH) to ethyl acetate (EtOAc).
Experiments were conducted by varying temperature, duration, and assisting with ultrasonication (26, 78,and 130 kHz, 50 W, a planar type). Under homogeneous conditions (concentration of γ-oryzanol 12 mg /mL, KOH / γ-oryzanol ratio (wt / wt) of 10 / 1, and EtOH / EtOAc ratio (v / v) of 5 / 1), a 45 % yield of FA
was obtained at 60 °C for 3 hours, and 73 % yield at 75 °C, respectively. With assistance of 78 and 130
kHz irradiation, the yield increased 1.6 fold at 60 ºC, while it was only 1.2 fold with assistance of lo wer
frequency of 26 kHz. Further with raising temperature to 75 °C (78 and 130 kHz), the yield reached to90%. From soapstock, the 74.3% yield of FA was obtained without assistance of ultrasonication. During
one month storage of reaction mixture, 20% of the trans-FA from -oryzanol was transformed into its
cis-form.
In Chapter 4, we investigated the effect of temperatures and ultrasonic irradiation (20 kHz, a horntype) with different powers (50 and 180 W) on the base-catalysed hydrolysis of γ-oryzanol in a
homogeneous reaction system (EtOH / EtOAc / H 2O). The experiments were conducted with various ratios(wt / wt) of KOH to γ-oryzanol amount (concentration of γ-oryzanol 6 mg / mL), temperature, andultrasonic irradiation powers. Without assistance of ultrasonic irradiation, 83.2 % yield of FA wasobtained under the conditions as follows: KOH / γ-oryzanol ratio (wt / wt) of 20 / 1, temperature of 75 ºC,duration of 3 hours. The yield of FA incredibly increased to 93.3 % in 2.5 h at 60 °C using ultrasonicirradiation of a horn-type 20 kHz with power of 180 W. These results indicated that using the highfrequency of horn-type 20 kHz, the high yield (more than 90 %) of FA was obtained at relatively low
temperature and in short time. The method can be used to produce FA from γ-oryzanol in soapstock fromrice bran oil processing efficiently.
In Chapter 5, biodiesel (BDF) production from candlenut oil (CNO) was investigated by two -step
co-solvent method in order to obtain the high quality of BDF. Firstly, esterific ation of FFAs (free fattyacids) (7 wt%) contained in CNO was conducted using a co-solvent of acetonitrile (30 wt%) and H 2SO4catalyst. The content of FFAs was reduced to 0.8 wt% in 1 hour at 65 °C. Subsequent transesterification ofthe produced crude oil was conducted using a co-solvent of acetone (20 wt%) and 1 wt% KOH. The BDF
content of 99.3 % was obtained at 40 °C in 45 minutes. After cleaning-up under vacuum condition at 5 kPa,
the water content in BDF was attained 0.023 %. The components of CNO BDF were characterized using a
Fourier-transform infrared (FTIR) spectrometry and gas chromatograph-flame ionization detector(GC-FID). The physicochemical properties of BDF satisfied the ASTM D6751-02 standard. The gaseous
exhausts from the diesel engine by combustion of the BDF blends (B0 − B100) with petro diesel wereexamined. The emissions of carbon monoxide (CO), and hydrocarbons (HC) were clearly lower, but thatof nitrogen oxides (NO x) was higher in comparison to those from petro diesel.
In Chapter 6, we investigated a series of PEs extracted from Jatropha curcas seed kernels cultivatedin Trang Bang, Vietnam with MeOH, and identified more than seven components contained in the PEs.
The isolation of main five components of a series of PEs was conducted using a semi -preparative reversed
phase HPLC analysis of ODS-3 column. The five peaks of components were successfully isolated, andpeaks of J2, J3, J5, and J7 were assigned to be Jatropha factors C 1, C2, C3, and C4/5, that has been alreadyreported by M. Hirota et al. in 1988 and W. Haas et al. in 2002, but J6 was a mixture of Jatropha factor C 6and its isomer based on the data of UV and LC-tandem mass spectrometry (LC-MS/MS), and J2 was
identified using 1H NMR analysis. By characterization using LC-MS/MS analysis, all components of aseries of PEs were elucidated to be the 12-deoxy-16-hydroxyphorbol esters composed of isomeric form of
dicarboxylic groups with same m / z value of 380.
In Chapter 7, the behaviors of toxic PEs in the Jatropha curcas oil were investigated during BDF
production in order to certificate the safety use of the produced biodiesel fuel (BDF). LC -MS/MS andphoto-diode array detector (PDA) analyses revealed that the PEs contained in Jatropha curcas oil had atigliane skeleton. The partition coefficients (KMeOH/oil) of several kinds of PEs between MeOH and the oil
ranged from 2.4 to 20. As a result, the PEs in the oil were largely partitioned into the MeOH phase. ThePEs in the oil were converted stoichiometrically into one molar phorbol and the corresponding three molarfatty acid methyl esters via a transesterification reaction in a KOH/MeOH solution. The phorbol produced
predominantly partitioned into the glycerin phase. A small amount of phorbol residue contained in theBDF could be removed by washing with water. These results suggest that it is safe to use BDF producedby the aforementioned transesterification reaction and purification process. However, phorbol
contamination of glycerin and wastewater from the production process should not be ignored.
In Chapter 8, conclusion of this thesis and summary of the previous chapters are described.

審査結果の要旨
本論文は、バイオディーゼル燃料(BDF)を合成するにあたり、油の豊富なバイオマス中の有用物質の利用および非食用油に含まれる油中の毒性物質の BDF 製造過程での挙動について詳しく調べた論文であり、次のような成果を得ている。
１）食用に供することができるがその多くが未利用で世界で最も多くの潜在的生産量を有する米
ぬか中の抗酸化剤の tocol (α､β、γトコフェロールとα、β、γトコトリエノールの総称)、γｰオリザノールとフェルラ酸の抽出方法を確立した。アルカリ条件下で超音波照射することで、米ぬか中に含まれる抗酸化剤の全量に対して、ほぼ８０％－９０％の抽出が可能となることを明らかにした。
２）上記の研究においてアルカリ条件下の抽出でγｰオリザノールから抗酸化剤として有用なフェルラ酸への加水分解を発見し、超音波照射下で 2 種の混合溶媒(エタノール/酢酸エチル)を用いることで 70％～９０％がフェルラ酸に加水分解できることを見出した。
３）Candle nuts 油を原料とした BDF の製造を検討した。Candle nuts 油には油分の分解した遊離脂肪酸が７－１０％も含まれているため、通常のアルカリ触媒法のトランスエステル化では純度のよい BDF が製造できなった。しかし、遊離脂肪酸を硫酸触媒によるエステル化でメチルエステルを生成し、分離後共溶媒法によるトランスエステル化を KOH 触媒で行なうことで高純度の BDF製造を確立した。
４）非食用油である Jatropha curcas は BDF 原料として期待されていたが、この実には毒性の強いフォルボールエステルが含まれていることが問題であった。本研究では、まずこのフォルボールエステルの分析方法を検討し、確立した。この確立した分析法を用いることで、フォルボールエステルがアルカリ触媒によるトランスエステル化中で毒性の低いフォルボールに変化すること、生成した BDF 中にはフォルボールエステルは存在しないことを明らかにした。
以上の諸成果は、経済的理由で未だに軽油の代替となっていない BDF を、その製造過程で高価な薬効成分を抽出・利用し、毒性物質を含まない非食用油－BDF を合成することで、安心して軽油と競合できる BDF とするためのブレークスルーに繋がる極めて重要な知見を得たものである。
石油の枯渇が懸念される中、人類の未来の燃料として使うことができ、さらに地球温暖化対策にも繋がるため、人類の持続可能な発展に対して貢献するところ大である。また、申請者が自立して研究活動を行うのに必要な能力と学識を有することを示したものである。