Production of biodiesel by yeast from lignocellulose and glycerol

A technology of microorganisms and free fatty acids, which is applied in the direction of enzymes, biofuels, biochemical equipment and methods, and can solve problems such as development, stability and high yield

Inactive Publication Date: 2013-05-01
DANMARKS TEKNISKE UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Although attempts have been made to produce biodiesel using microbial organisms, a major obstacle to success has been the development of stable, high-yielding microorganisms and methods of producing them

Method used

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  • Production of biodiesel by yeast from lignocellulose and glycerol
  • Production of biodiesel by yeast from lignocellulose and glycerol
  • Production of biodiesel by yeast from lignocellulose and glycerol

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0115] Example 1 Optimum Growth Conditions for Yeast Production of Fatty Acids Regarding Temperature, pH, Medium Composition and Dextrose Concentration

[0116] Experiments with growth conditions that maximize fatty acid production by yeast were performed with the aim of determining conditions that modulate biomass growth and enhance fatty acid synthesis and accumulation and / or secretion.

[0117] Two strains of S. cerevisiae were tested in parallel:

[0118] 1) National center for industrial microorganisms (NCIM) strain 3090,

[0119] (http: / / www.ncl-india.org / ncim / cataloguedetails.jsp?mid=29&category=yeast&ncimno=3090)

[0120] 2) Common baker's yeast strain

[0121] The strains were grown on Wickerham synthetic medium (Wickerham L J. 1951. Taxonomy of yeast. US Department of Agriculture, Technical Bulletin No. 1029.) the composition of which is given in Table 1, but with the carbon source changed as indicated.

[0122] Table 1

[0123]

[0124] Fermentation condition...

Embodiment 2

[0141] Example 2. Deletion of the FAA2 gene enhances fatty acid production in Saccharomyces cerevisiae

[0142] 2.0.0 method

[0143] Saccharomyces cerevisiae strain CEN-PK2 (MATa / MATa; ura3-52 / ura3-52; trpl-289 / trp1-289; leu2-3,112 / leu2-3,112; his3D1 / his3D1; The FAA2 gene in MAL2-8C / MAL2-8C; SUC2 / SUC2) was deleted and replaced with a 1.1 kbp URA3 marker gene (derived from Kluyveromyces marxii) to enable uracil synthesis. URA3 encodes orotidine 5-phosphate decarboxylase (ODCase), an enzyme involved in pyrimidine nucleotide synthesis.

[0144] Primer set 1 was designed to delete the FAA2 gene along the 150 bp upstream flank of the FAA2 gene and the 60 bp downstream flank of the FAA2 gene in the parental CEN-PK2 strain (hereinafter referred to as the WT strain). Primer set 3 (binding near the center of the FAA2 gene sequence) was used to amplify a ~1300 bp fragment of the FAA2 gene in order to detect the FAA2 gene in genomic DNA derived from the WT strain. Primers were design...

Embodiment 3

[0198] Example 3. Use of Candida tropicalis for the production of fatty acids suitable for diesel production

[0199] Candida tropicalis (DTU strain library) was cultured on the improved Wickerham synthetic medium with a specific carbon source, and the fatty acids in the growth medium and cells were extracted according to 2.0.0.

[0200] 3.0.0 When Candida tropicalis was grown on synthetic medium supplemented with glucose and the secreted fatty acids were extracted and analyzed by GC-MS, a total of 21 peaks were detected ( Figure 13 ). The most prominent peak corresponding to 35% of the total extracted fatty acids had a residence time of 28.84 minutes and was identified as hexadecanoic acid. The yield of saponifiable fatty acids after organic extraction was 7.5ml / 50 medium. The production of hexadecanoic acid was 50 ml / liter medium.

[0201] 3.0.1 When Candida tropicalis was grown on a synthetic medium (Table 5) supplemented with 5% glycerol and the secreted fatty acids we...

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Abstract

The invention is directed to a genetically modified microorganism for the extracellular production of free fatty acids and esters thereof, wherein said microorganism is characterised by a modified lipid biosynthesis metabolic pathway: for example reduced fatty acyl-coA synthetase activity that enables the microorganism to overproduce and secrete of esters of fatty acids (Biodiesel) into the surrounding medium, using one or more of: glucose, starch, lignocellulose and a glycerol-based substrate, as a carbon source. The invention further provides a method for the extracellular production of free fatty acids and esters thereof, comprising the use of said genetically modified organism, and a growth medium adapted for said method.

Description

technical field [0001] The present invention involves genetically engineering the lipid biosynthesis metabolic pathway in yeast to overproduce and secrete fatty acid esters using one or more of glucose, starch, lignocellulose, and glycerol-based substrates as carbon sources (biological Diesel) into the surrounding medium. Background technique [0002] Biodiesel refers to a fuel comprising mono-alkyl esters of long-chain fatty acids derived from vegetable oils or animal fats, known as B100, and meets the requirements of ASTM D6571. Currently, the most common method of producing biodiesel is the transesterification of edible and inedible vegetable oils or sometimes animal fats. The transesterification reaction converts triglycerides into fatty acid alkyl esters in the presence of an alcohol such as methanol or ethanol and a catalyst such as a base or acid, the by-product of which is glycerol. Biodiesel has a cetane number and energy content comparable to petroleum diesel and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/00C12P7/64C12N1/16C12N1/18C12N9/88
CPCY02E50/13C12N9/93C12P7/6409C12N9/88C12N1/18C12P7/649C12N1/16Y02E50/10
Inventor 安巴里什·戈文德·帕德纳维斯彼得·鲁达尔詹森
Owner DANMARKS TEKNISKE UNIV
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