WO2010061987A1 - Procédé et milieu de fabrication de bioéthanol à l'aide de plantes du genre typha l. - Google Patents

Procédé et milieu de fabrication de bioéthanol à l'aide de plantes du genre typha l. Download PDF

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Publication number
WO2010061987A1
WO2010061987A1 PCT/KR2008/006986 KR2008006986W WO2010061987A1 WO 2010061987 A1 WO2010061987 A1 WO 2010061987A1 KR 2008006986 W KR2008006986 W KR 2008006986W WO 2010061987 A1 WO2010061987 A1 WO 2010061987A1
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WIPO (PCT)
Prior art keywords
bioethanol
production
hydrolysis
genus typha
typha
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PCT/KR2008/006986
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English (en)
Inventor
Tae Kwon Kim
Sung Eun Lee
Byeoung Soo Park
Wan Jung Kim
Mi Kyung Son
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Nanotox Tech
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Priority to PCT/KR2008/006986 priority Critical patent/WO2010061987A1/fr
Publication of WO2010061987A1 publication Critical patent/WO2010061987A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
    • C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
    • C12P7/06—Ethanol, i.e. non-beverage
    • C12P7/08—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate
    • C12P7/10—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate substrate containing cellulosic material
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
    • Y02E50/00—Technologies for the production of fuel of non-fossil origin
    • Y02E50/10—Biofuels, e.g. bio-diesel

Definitions

  • the present invention relates to a method and medium for the production of bioethanol using a perennial herbaceous plant, Genus Typha L. More specifically, the method for the production of bioethanol comprises the steps of a) hydrolyzing Genus Typha L. and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source, and the medium for the production of bioethanol contains the hydrolyzed Genus Typha L. solution as a carbon source.
  • bioethanol is the most actively raised material.
  • Bioethanol alone or in mixture with gasoline, can be used as an automobile fuel, and thus it is a representative recycled energy, in addition to biodiesel.
  • Carbon dioxide which is generated from the combustion of bioethanol, is one of the exceptions to the greenhouse gas calculation provided by the Kyoto Protocols because of its greenhouse gas-reducing effects.
  • bioethanol can be supplied through existing facilities (gas/filling stations), which enables early commercialization.
  • wood biomass has an advantage in that resources are abundant and the amount of ethanol produced is high because the content of cellulose and hemicellulose is about 75% or more.
  • wood biomass has a much sturdier physical and chemical structure than crop biomass, a chemical or enzymatic approach for saccharif ⁇ cation of cellulose and hemicellulose is difficult.
  • wood biomass has a disadvantage because it contains about 15-25% of lignin, which consists of numerous hydrophobic aromatic compounds, and the pre-treatment process for removing such lignin is complicated and expensive. Forest destruction to ensure a biomass supply would also accelerate global warming and environmental destruction, and it would be hard to justify bioethanol as an environment- friendly fuel in light of environmental and ethical issues.
  • Herbaceous plants which can be used as biomass, include rice straw and bagasse.
  • Herbaceous plants are characterized by relatively high content of cellulose and hemicellulose, and low content of lignin, and thus their pre-treatment is easier than that of conventional wood biomass.
  • these plants' growth rate is fast, and thus their supply is easy. They also have the advantage of being free of the ethical issues that plague wood biomass.
  • Genus Typha L. is a perennial herbaceous plant which belongs to the Typhaceae family, and about 9 to 18 species thereof are distributed worldwide. It has been known that three types of Genus Typha L., i.e., Typha orientalis, Typha angustata and Typha latifolia, grow naturally in Korea (Choi, 2000; Chung, 1957; Im, 1998; Kim and Choi, 2001 ; Lee, 1996; Mori, 1922; Nakai, 1911, 1951). Genus Typha L. is characterized by rigorous development of an aerial part and a rhizome (a root) that results in a great amount of biomass. Genus Typha L. contains a large amount of carbohydrate in its leaf, stem and root, which can be converted into ethanol.
  • Genus Typha L. can fairly compete with conventional crop or wood biomass because its growth rate is much faster compared with wood, and it grows naturally in the swampy land of the temperate regions so that it is easy to cultivate and collect. It has been reported that rice straw and reeds can produce biomass in an amount of 8.5-11.2 ton/ha and 6.4-12.5 ton/ha by dry weight, respectively, whereas Genus Typha L. can produce biomass in an amount of 10.6-14.7 ton/ha and, in case a fertilizer is used, 14.5-22.7 ton/ha (Ja-Hyeong Gu, et al, ARPC Report, 2007, p. 95). Thus, Genus Typha L. merits attention as a future bioenergy crop. Thus, it is understood that developing a method for the production of bioethanol using Genus Typha L., a representative herbaceous plant, would have an important meaning in securing a second-generation bioethanol.
  • the present inventors have performed continuous study on new biomasses that can resolve problems caused by conventional methods for the production of bioethanol using crop or wood biomass, such as rapid increase of crop price, weaponization of crop, problems associated with crop ethics, impairment of forest resources and global warming.
  • problems caused by conventional methods for the production of bioethanol using crop or wood biomass such as rapid increase of crop price, weaponization of crop, problems associated with crop ethics, impairment of forest resources and global warming.
  • an ethanol-producing strain is cultured using as a carbon source a Genus Typha L. solution that has been obtained by hydrolyzing Genus Typha L. with an acid and/or a saccharification enzyme, it exhibits excellent effects, including a significant increase in the amount of bioethanol produced, decreased cost of production and easy security of resources necessary for production, compared with conventional methods that utilized crop or wood biomass, and completed the present invention.
  • the first object of the present invention is to provide a method for the production of bioethanol, comprising the steps of a) hydrolyzing Genus Typha L. and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the second object of the present invention is to provide a method for the production of bioethanol, comprising the steps of a) hydrolyzing Genus Typha L. with an acid or a saccharification enzyme and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the third object of the present invention is to provide a method for the production of bioethanol, comprising the steps of a) first hydrolyzing Genus Typha L. with an acid and second hydrolyzing it with a saccharification enzyme, and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the fourth object of the present invention is to provide a medium for the production of bioethanol, containing the hydrolyzed Genus Typha L. solution as a carbon source.
  • Fig. 1 is a graph that shows the hydrolysis rate when sulfuric acid or hydrochloric acid is used in the method for the production of bioethanol of the present invention.
  • Fig. 2 is a graph that shows the hydrolysis rate change depending on the concentration of sulfuric acid in the method for the production of bioethanol of the present invention.
  • Fig. 3 is a graph that shows the hydrolysis rate change depending on the amount of
  • Genus Typha L. added in the method for the production of bioethanol of the present invention.
  • Fig. 4 is a graph that shows the hydrolysis rate change depending on the amount of a saccharification enzyme, Viscozyme, added in the method for the production of bioethanol of the present invention.
  • Fig. 5 is a graph that shows the hydrolysis rate change depending on the reaction time of a saccharification enzyme in the method for the production of bioethanol of the present invention.
  • Fig. 6 shows the results of HPLC confirming the existence of bioethanol that has been distilled according to the method for the production of bioethanol of the present invention.
  • the first aspect of the present invention relates to a method for the production of bioethanol, comprising the steps of a) hydrolyzing Genus Typha L. and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the second aspect of the present invention relates to a method for the production of bioethanol, comprising the steps of a) hydrolyzing Genus Typha L. with an acid or a saccharification enzyme and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the third aspect of the present invention relates to a method for the production of bioethanol, comprising the steps of a) first hydrolyzing Genus Typha L. with an acid and second hydrolyzing it with a saccharification enzyme, and b) culturing an ethanol-producing strain using said hydrolyzed Genus Typha L. solution as a carbon source.
  • the fourth aspect of the present invention relates to a medium for the production of bioethanol, containing the hydrolyzed Genus Typha L. solution as a carbon source.
  • Genus Typha L. there is no limit on the kind of Genus Typha L. to be used.
  • Typha latifolia can be used. Any one part selected from leaf, stem and root, or all of them can be used.
  • leaf, stem and root of Genus Typha L. contains a considerable amount of carbohydrate, which can be converted into ethanol.
  • the composition of leaf, stem and root is shown in Table 1 below.
  • the first step of the method for the production of bioethanol according to the present invention is hydrolyzing Genus Typha L.
  • Genus Typha L. is saccharified into a reducing sugar (a fermenting sugar), which is easy for an ethanol-producing strain to use.
  • Genus Typha L. contains a relatively small amount of lignin compared with wood biomass, the hydrolysis process itself according to the present invention is sufficient for lignin treatment, and there is no need to additionally use various chemicals that were conventionally required for lignin treatment.
  • the hydrolysis of Genus Typha L. can be carried out by using either an acid or a saccharification enzyme, or both.
  • acid there is no limit on the kind of acid that can be used for the hydrolysis of the present invention.
  • sulfuric acid or hydrochloric acid can be used.
  • the method using an acid of weak concentration at high temperature is used more often because it is easy to neutralize and is safe, due to less addition of the acid.
  • it is more preferable to adopt the method using an acid of weak concentration at high temperature for the hydrolysis of the present invention.
  • hydrochloric acid which is weaker than sulfuric acid, is dangerous to use at high temperature due to its high volatility, and thus it is difficult to add 10% or more; thereby, its hydrolysis rate was observed to be low. Therefore, it was concluded that sulfuric acid is more preferable for hydrolysis at high temperature.
  • the present invention has established a preferable condition for hydrolyzing Genus Typha L. with an acid. Specifically, in order to determine the concentration of an acid for the hydrolysis of the present invention, the concentration of sulfuric acid was changed from 1% to 6%, and the concentration of a reducing sugar within the solution was measured to determine the hydrolysis rate. As the concentration of sulfuric acid increased, the hydrolysis rate also increased. When the concentration of sulfuric acid was 6%, the hydrolysis rate was about 20% (see Fig. 2).
  • the concentration of sulfuric acid used for hydrolyzing Genus Typha L. with an acid according to the present invention is preferably 5% to 6%, and more preferably 6%.
  • the concentration of hydrochloric acid which is another acid that can be used for the hydrolysis of the present invention, is preferably 10% to 20%.
  • the ratio of Genus Typha L. and the acid is preferably 1 : 60-70 (w/v).
  • this ratio of Genus Typha L. and the acid is applicable only when 6% sulfuric acid is used at high temperature of 120 ° C
  • the ratio of Genus Typha L. and an acid can vary when the hydrolysis is carried out at low temperature, sulfuric acid of different concentration is used, or an acid other than sulfuric acid is used.
  • the hydrolysis of Genus Typha L. of the present invention can also be carried by a saccharification enzyme.
  • saccharification refers to a process of hydrolyzing cellulose by enzymes to produce a sugar and is a different concept from glycosylation, which adds a sugar to a protein.
  • saccharification enzyme that can be used for the hydrolysis of the present invention.
  • one or more selected from the group consisting of carbohydrase, cellulase, amylase and glucosidase can be used.
  • carbohydrase cellulase
  • amylase glucosidase
  • glucosidase can be used.
  • all of the above-mentioned four kinds of saccharification enzymes exhibited a hydrolysis rate up to about 30%.
  • a carbohydrase-containing saccharification enzyme showed the highest hydrolysis rate.
  • the kind of a saccharification enzyme is not limited thereto.
  • Celluclast 1.5L is preferable as a cellulase, Viscozyme 2L as a carbohydrase, and Fungamyl 800L as an amylase and a glucosidase.
  • the present invention has established a preferable condition for hydrolyzing Genus Typha L. with a saccharification enzyme.
  • Viscozyme 2L a carbohydrase-containing saccharification enzyme
  • the initial amount was 0.1 niL, which was increased by 0.1 mL.
  • the concentration of a reducing sugar was measured to determine the hydrolysis rate of Genus Typha L. It was observed that as the amount of added saccharification enzyme increased, the hydrolysis rate of Genus Typha L. increased.
  • the hydrolysis rate was about 35%.
  • the amount of a saccharification enzyme to be used for hydrolysis of a saccharification enzyme according to the present invention is preferably 0.4 to 0.6 mL, and more preferably 0.5 mL per 1 g of Genus Typha L.
  • the present inventors studied how the reaction time required for hydrolysis influences a hydrolysis rate in the case of hydrolyzing Genus Typha L. with a saccharification enzyme.
  • a saccharification enzyme which is a carbohydrase-containing saccharification enzyme, was added to a sample of crushed Genus Typha L. Every hour, the concentration of a reducing sugar was measured to determine the hydrolysis rate.
  • the hydrolysis rate increased and reached about 33% about 18 hours after the reaction was initiated. After 18 hours, the hydrolysis rate increased somewhat slowly (see Fig. 5).
  • the first hydrolysis with a weak acid or a saccharification enzyme is sufficient for hydrolyzing Genus Typha L.
  • the hydrolysis of Genus Typha L. of the present invention can be carried out by two-step processes that consist of the first hydrolysis by a weak acid followed by the second hydrolysis by a saccharification enzyme.
  • the second step is culturing an ethanol-producing strain using the above hydrolyzed Genus Typha L. solution as a carbon source.
  • ethanol-producing strain there is no limit on the kind of ethanol-producing strain that can be used for the method for the production of bioethanol according to the present invention.
  • one or more strains selected from the group consisting of Saccharomyces cerevisiae, Pichia stipitis and Zymomonas mobilis can be used.
  • the ethanol-producing strain is Saccharomyces cerevisiae.
  • the medium for the production of bioethanol of the present invention is one that contains the hydrolyzed Genus Typha L. solution which has been obtained according to the above-described methods.
  • the medium is preferably a complex medium consisting of natural ingredients, such as yeast extract, bean peptone, tripton and the like. Specific examples thereof include YP medium, YM medium and LB medium, which are conventionally used in the art.
  • the medium of the present invention is not limited to only a complex medium, and a semi-defined medium can also be used.
  • the medium for the production of bioethanol of the present invention essentially contains the hydrolyzed Genus Typha L. solution as a carbon source and may additionally contain conventionally used carbon sources.
  • the method and medium for the production of bioethanol according to the present invention exhibits excellent effects, including a significant increase in the amount of bioethanol produced, decreased cost of production and easy security of resources necessary for production, compared with conventional methods that utilized crop or wood biomass.
  • each of the two hydrolyzed Genus Typha L. solutions which had been obtained by hydrolysis with an acid or by hydrolysis with a saccharification enzyme, was concentrated and added to a YP medium (yeast extract 1.5%, peptone 1.5%), which was then used as the medium for the production of bioethanol of the present invention.
  • a YP medium yeast extract 1.5%, peptone 1.5%)
  • After pre-culturing Saccharomyces cerevisiae as an ethanol-producing strain in a YM medium it was inoculated into the above medium for the production of bioethanol and shaking-cultured for 80 hours.
  • the concentration of bioethanol was measured using HPLC.
  • the yield (mL) of converted ethanol per 1 g of a reducing sugar added was converted into percentage (%) to give an ethanol conversion rate. It was understood that the ethanol conversion rate for the acid-hydro lyzed Genus Typha L. solution was 30.3% and that for the saccharification enzyme-hydrolyzed Genus Typha L.
  • Genus Typha L. was crushed without sorting leaf, stem or root thereof, and soaked in distilled water at 80 ° C for 1 hour to give a sample. To each 1 g of the sample was added 100 mL of 10% (w/w) hydrochloric acid or 100 mL of 6% (w/w) sulfuric acid.
  • the hydrolysis rate was about 8% when hydrochloric acid was added, whereas it was 20% when sulfuric acid was added. It was understood that because sulfuric acid is stronger than hydrochloric acid, it could give a relatively high hydrolysis rate even if a small amount were added. On the other hand, hydrochloric acid, which is weaker than sulfuric acid, is dangerous to use at high temperature due to its high volatility, and thus it is difficult to add 10% or more; thereby, its hydrolysis rate was observed to be low. Therefore, it was concluded that sulfuric acid is more preferable for hydrolysis at high temperature.
  • Example 2 Determination of a concentration of sulfuric acid for the hydrolysis of Genus Typha L.
  • Genus Typha L. was crushed without sorting leaf, stem or root thereof, and soaked in distilled water at 80 ° C for 1 hour to give a sample. To each 1 g of the sample was added 100 mL of sulfuric acid whose concentration varied from 1 to 6% (w/w). After treatment at 120 ° C for 30 min, the solution was filtered, and the concentration of a reducing sugar within the solution was measured to determine the hydrolysis rate. The results are shown in Fig. 2. As shown in Fig. 2, as the concentration of sulfuric acid increased, the hydrolysis rate also increased. When the concentration of sulfuric acid was 6%, the hydrolysis rate was maintained at around 20%. Thus, it was concluded that a proper concentration of sulfuric acid in the hydrolysis step is 5% to 6%.
  • Example 3 Determination of the ratio of Genus Typha L. and an acid for the hydrolysis of Genus Typha L.
  • Genus Typha L. was crushed without sorting leaf, stem or root thereof, and soaked in distilled water at 80 ° C for 1 hour to give a sample. To each 1 g, 1.5 g, 2 g, 2.5 g, 3 g,
  • Example 4 Determination of a saccharification enzyme for the hydrolysis of Genus Typha L. Further studies were carried out regarding how the addition of a saccharification enzyme, which is able to saccharify a fiber, influences the hydrolysis rate.
  • Genus Typha L. was crushed without sorting leaf, stem or root thereof, and soaked in distilled water at 80 ° C for 1 hour to give a sample. To each 1 g of the sample was added 0.5 mL of Celluclast 1.5 L, which is a cellulase, Viscozyme 2L, which contains various carbohydrases, or Fungamyl 800L, which contains an amylase and an amyloglucosidase.
  • Example 5 Determination of an amount of a saccharification enzyme added for the hydrolysis of Genus Typha L.
  • the amount of a saccharification enzyme to be used for the hydrolysis of Genus Typha L. is about 0.5 mL per 1 g of Genus Typha L.
  • Example 6 Determination of a reaction time of a saccharification enzyme for the hydrolysis of Genus Typha L. Further studies were carried out regarding how the reaction time required for hydrolysis influences the hydrolysis rate in the case of hydrolyzing Genus Typha L. with a saccharification enzyme. Under the same condition as Example 5, 0.5 mL of Viscozyme 2L was added and allowed for reaction. Every hour, the concentration of a reducing sugar was measured to determine the hydrolysis rate. The results are shown in Fig. 5. As shown in Fig. 5, as time passed, the hydrolysis rate increased and reached about 33% about 18 hours after the reaction was initiated. After 18 hours, the hydrolysis rate increased somewhat slowly.
  • the preferable reaction time of the saccharification enzyme is 18 to 24 hours.
  • Example 7 Production of bioetlianol using the hydrolyzed Genus Typha L. solution as a carbon source
  • Each of the two hydrolyzed Genus Typha L. solutions (treatment solutions), which had been obtained by hydrolysis with an acid or with a saccharification enzyme, was concentrated and added to a YP medium (yeast extract 1.5%, peptone 1.5%) so that the final concentration of a reducing sugar was 2% (w/w).
  • the medium was then used for the production of bioethanol.
  • Saccharomyces cerevisiae as an ethanol-producing strain in a YM medium (yeast extract 0.3%, malt extract 0.3%, peptone 0.5%, dextrose 1%) for 18 hours, it was inoculated into 200 mL of the above medium for the production of bioethanol by 5% (v/w) of the volume of the cultured solution.
  • Genus Typha L. is characterized by rigorous development of an aerial part and a rhizome, which results in a great amount of biomass, and is characterized by a relatively low content of lignin compared with wood biomass. Due to the use of Genus Typha L., the method and medium for the production of bioethanol according to the present invention exhibits a significant increase of bioethanol produced and a decreased cost of production because the survival of ethanol-producing yeasts and microorganisms becomes easy due to the decreased use of various chemicals which were required for lignin treatment. In addition, Genus Typha L. provides easy security of resources because its growth rate is much faster compared with wood biomass and it grows naturally in swampy land in the temperate regions so that it is easy to cultivate and collect. Thus, the method and medium for the production of bioethanol according to the present invention would enable large-scale production of bioethanol in a more economical way, compared with conventional methods which utilized crop or wood biomass.

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Abstract

La présente invention concerne un procédé et un milieu de fabrication de bioéthanol à l'aide d'une plante herbacée pérenne du genre Typha L. Plus particulièrement, le procédé de fabrication du bioéthanol comporte les étapes a) d’hydrolyse de plantes du genre Typha L., et b) de culture d'une souche produisant de l'éthanol à l'aide de ladite solution de plantes du genre Typha L. hydrolysées en tant que source de carbone. En raison de l'utilisation de plantes du genre Typha L., qui sont caractérisées par un développement vigoureux d'une partie aérienne et d'un rhizome, par une vitesse de croissance rapide et une faible teneur en lignine, le procédé et le milieu de fabrication de bioéthanol selon la présente invention présentent d'excellent effets, comprenant une augmentation significative de la quantité de bioéthanol produite, un coût de fabrication réduit et une obtention facile des ressources nécessaires pour la fabrication, par comparaison avec les procédés classiques qui utilisaient une biomasse des cultures ou ligneuse.
PCT/KR2008/006986 2008-11-27 2008-11-27 Procédé et milieu de fabrication de bioéthanol à l'aide de plantes du genre typha l. WO2010061987A1 (fr)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2000053791A1 (fr) * 1999-03-11 2000-09-14 Dan Verser Processus de production d'ethanol
US20070134781A1 (en) * 2005-12-12 2007-06-14 Agblevor Foster A Method for producing bioethanol from a lignocellulosicbiomass and recycled paper sludge
WO2008064289A2 (fr) * 2006-11-21 2008-05-29 The Samuel Roberts Noble Foundation, Inc. Procédés et compositions de production de biocarburant
WO2008105618A1 (fr) * 2007-02-26 2008-09-04 Korea Institute Of Industrial Technology Méthode de production de biocarburant, à l'aide d'algues marines,

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2000053791A1 (fr) * 1999-03-11 2000-09-14 Dan Verser Processus de production d'ethanol
US20070134781A1 (en) * 2005-12-12 2007-06-14 Agblevor Foster A Method for producing bioethanol from a lignocellulosicbiomass and recycled paper sludge
WO2008064289A2 (fr) * 2006-11-21 2008-05-29 The Samuel Roberts Noble Foundation, Inc. Procédés et compositions de production de biocarburant
WO2008105618A1 (fr) * 2007-02-26 2008-09-04 Korea Institute Of Industrial Technology Méthode de production de biocarburant, à l'aide d'algues marines,

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
DAVID HULL ET AL.: "The feasibility of ethanol production from the cattail Typha latifolia.", FINAL REPORT, CALIFORNIA DEPARTMENT OF FOOD AND AGRICULTURE., June 1984 (1984-06-01) *

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