Classifications

• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23D—EDIBLE OILS OR FATS, e.g. MARGARINES, SHORTENINGS OR COOKING OILS
  • A23D9/00—Other edible oils or fats, e.g. shortenings or cooking oils

Definitions

• This invention relates to environmentally preferred frying oils, such as high oleic oils.
• a Life Cycle Assessment (LCA) of high oleic oil compared to
• LCA Life Cycle Assessment
• Soybean oil is the most abundant vegetable oil in the world. Common soybean varieties produce an oil high in polyunsaturated fatty acids. This property makes the oil unstable, easily oxidized and subject to becoming rancid. When heated, soybean oil develops objectionable flavors and odors, making it unsuitable for many applications. Oils with high levels of polyunsaturated fatty acids are not often used in applications that require a high degree of oxidative stability, such as cooking for a long period of time at an elevated temperature.
• the present disclosure generally relates to a sustainable frying oil which reduces the environmental impact and carbon footprint in the food service industry.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil has an increased oleic content when compared to an ordinary frying oil.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil is useful as a blending source to make a blended environmentally preferred frying oil.
• the invention concerns an environmentally preferred frying oil obtained from a high oleic oilseed.
• the oilseed is one selected from the group consisting of: soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower.
• the invention concerns an environmentally preferred frying oil, wherein the oleic acid content of said oil comprises at least 60% of the fatty acid moieties in the oil.
• the invention concerns a method for frying with a reduced impact on the environment, comprising:
• a further embodiment of the invention concerns a method for frying with a reduced impact on the environment , comprising using an oil with an increased oleic acid content when compared to an ordinary oil and quantifying the reduction in environmental impacts when using a frying oil with an increased oleic acid content compared to an ordinary frying oil, wherein the reduction environmental impact is at least one selected from the group consisting of: reduced carbon footprint , reduced eutrophication potential, reduced air acidification potential, and reduced non- renewable energy consumption.
• High oleic oil obtained from seeds , soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower are also part of the invention.
• a further embodiment of the invention concerns the use of a high oleic oil, wherein the use of the oil for frying applications reduces land use pressure.
• An additional embodiment of the invention concerns use of a high oleic frying oil for reduction of environmental impact, such as a reduction of carbon footprint, reduction of eutrophication potential, reduction of air acidification potential, or reduction of non-renewable energy consumption.
• a further embodiment of the invention includes the use of a high oleic oil for frying , wherein the burden on the environment is reduced by at least 40% when compared to the use of a conventional oil.
• Fig .1 shows the climate change potential of high oleic oil compared to conventional oil during a 2 day fryer use.
• Fig.2 shows the non-renewable energy use of high oleic oil compared to
• Fig.3 shows the terrestrial acidification potential of high oleic compared to
• Fig.4 shows freshwater eutrophication potential of high oleic compared to
• Fig.5 shows a comparison of conventional, high oleic soy oil base case soy oil, and high oleic high price premium soy oil and their relative impact on climate change potential, non-renewable energy use, terrestrial acidification potential, and freshwater eutrophication potential.
• Fig.6 shows a sensitivity analysis of the allocation method of high oleic and conventional oil on climate change potential.
• Fig.7 shows a sensitivity analysis of the allocation method of high oleic and conventional oil on terrestrial acidification potential.
• Fig.8 shows a comparison of conventional oil base case, conventional oil base case using a 2-day wash cycle, and high oleic oil and their relative impact on climate change potential, terrestrial acidification potential, freshwater eutrophication potential , and non-renewable energy use.
• Figure 9 shows the climate change potential per 2 day fryer use in restaurant for high oleic canola oil compared to conventional canola oil.
• Figure 10 shows the non-renewable energy use per 2 day fryer use in a restaurant for high oleic canola oil compared to conventional canola oil.
• Figure 1 1 Terrestrial acidification potential per 2 day fryer use in a restaurant for high oleic canola oil compared to conventional canola oil.
• Figure 12 Freshwater eutrophication potential per 2 day fryer use in a restaurant for high oleic canola oil compared to conventional canola oil.
• FIG. 13 Flowchart: soybean oil life cycle system boundaries.
• “soybean” refers to the species Glycine max, Glycine soja, or any species that is sexually cross compatible with Glycine max.
• a “line” is a group of plants of similar parentage that display little or no genetic variation between individuals for a least one trait. Such lines may be created by one or more generations of self-pollination and selection, or vegetative propagation from a single parent including by tissue or cell culture techniques.
• An "agronomically elite line” or “elite line” refers to a line with desirable agronomic performance that may or may not be used commercially.
• a "variety”, “cultivar”, “elite variety”, or “elite cultivar” refers to an agronomically superior elite line that has been extensively tested and is or was being used for commercial soybean production. “Mutation” refers to a detectable and heritable genetic change (either spontaneous or induced) not caused by segregation or genetic recombination. “Mutant” refers to an individual, or lineage of individuals, possessing a mutation.
• LCA Life Cycle Assessment
• the term “environmentally preferred” or “sustainable” means reduction in the impact on the environment , such as, but not limited to a reduction in at least one of the following parameters: carbon footprint, eutrophication potential, air
• climate Change Potential refers to any significant change in the climate lasting for an extended period. climate change can be caused by natural factors, natural processes, and human activities. climate change potential (carbon footprint) is measured in terms of total greenhouse gas emissions, and takes into account the global warming potential of specific species known to contribute to climate change.
• Freshwater Eutrophication Potential refers to the overload of a waterbody with nutrients. This overload causes an increase in algal growth and a subsequent reduction in oxygen availability for aquatic life. Freshwater eutrophication is caused by phosphorous-containing emissions.
• Terrestrial acidification potential or "acid rain” - refers to emissions which alter optimum soil pH.
• the emissions that contribute to acidification are NOx, ammonia, and SO2.
• Non-renewable energy use or “fossile fuel depletion”- accounts for all of the coal, oil, natural gas, and uranium consumed in a particular supply chain.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil has an increased oleic content when compared to an ordinary frying oil.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil is useful as a blending source to make a blended environmentally preferred frying oil.
• the invention concerns an environmentally preferred frying oil obtained from a high oleic oilseed.
• the oilseed is one selected from the group consisting of: soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower.
• the invention concerns an environmentally preferred frying oil, wherein the oleic acid content of said oil comprises at least 60% of the fatty acid moieties in the oil.
• the invention concerns a method for frying with a reduced impact on the environment, comprising:
• a further embodiment of the invention concerns a method for frying with a reduced impact on the environment , comprising using an oil with an increased oleic acid content when compared to an ordinary oil and quantifying the reduction in environmental impacts when using a frying oil with an increased oleic acid content compared to an ordinary frying oil, wherein the reduction environmental impact is at least one selected from the group consisting of: reduced carbon footprint , reduced eutrophication potential, reduced air acidification potential, and reduced nonrenewable energy consumption.
• High oleic oil obtained from seeds , soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower are also part of the invention.
• a further embodiment of the invention concerns the use of a high oleic oil, wherein the use of the oil for frying applications reduces land use pressure.
• An additional embodiment of the invention concerns use of a high oleic frying oil for reduction of environmental impact, such as a reduction of carbon footprint, reduction of eutrophication potential, reduction of air acidification potential, or reduction of non-renewable energy consumption.
• a further embodiment of the invention includes the use of a high oleic oil for frying, wherein the burden on the environment is reduced by at least 40% when compared to the use of a conventional oil.
• fatty acids refers to long-chain aliphatic acids (alkanoic acids) of varying chain length, from about C12 to C22 (although both longer and shorter chain-length acids are known).
• the predominant chain lengths are between C16 and C22-
• the structure of a fatty acid is represented by a simple notation system of "X:Y", where X is the total number of C atoms in the particular fatty acid and Y is the number of double bonds.
• fatty acids are classified as saturated or unsaturated.
• saturated fatty acids refers to those fatty acids that have no “double bonds” between their carbon backbone.
• unsaturated fatty acids have “double bonds” along their carbon backbones (which are most commonly in the cis- configu ration).
• “Monounsaturated fatty acids” have only one "double bond” along the carbon backbone (e.g., usually between the 9 tn and 10 tn carbon atom as for palmitoleic acid (16:1 ) and oleic acid (18:1 )), while “polyunsaturated fatty acids” (or “PUFAs”) have at least two double bonds along the carbon backbone (e.g., between the 9 th and 10 th , and 12 th and 13 th carbon atoms for linoleic acid (18:2); and between the 9 th and 10 th , 12 th and 13 th , and 15 th and 16 th for a-linolenic acid (18:3)).
• PUFAs polyunsaturated fatty acids
• total fatty acid content refers to the sum of the five major fatty acid components found in soybeans, namely C16:0, C18:0, C18:1 , C18:2, and C18:3.
• total polyunsaturated fatty acid content refers to the total C18:2 plus C18:3 content.
• the omega-reference system will be used to indicate the number of carbons, the number of double bonds and the position of the double bond closest to the omega carbon, counting from the omega carbon (which is the terminal carbon of the aliphatic chain and is numbered 1 for this purpose). This nomenclature is shown below in Table 1 , in the column titled "Shorthand Notation”. Table 1
• the term "desaturase” refers to a polypeptide that can desaturate, i.e., introduce a double bond, in one or more fatty acids to produce a mono- or polyunsaturated fatty acid or precursor which is of interest.
• omega-reference system throughout the specification in reference to specific fatty acids, it is more convenient to indicate the activity of a desaturase by counting from the carboxyl end of the substrate using the ⁇ -system.
• FAD fatty acid desaturase
• fatty acid desaturase are used interchangeably and refer to membrane bound microsomal oleoyl- and linoleoyl-phosphatidylcholine desaturases that convert oleic acid to linoleic acid and linoleic acid to linolenic acid, respectively, in reactions that reduce molecular oxygen to water and require the presence of NADH.
• high oleic oilseed refers to seeds that have an oleic acid content of at least 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, and 95% of the seed by weight.
• the high oleic oilseed can be one selected from the group consisting of: soybean, sunflower, palm, peanut, corn and canola. Preferred high oleic soybean oil starting materials are disclosed in World Patent Publication WO94/1 1516, the disclosure of which is hereby incorporated by reference.
• high oleic oil refers to an oil that has at least 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71 %, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, and 95% of its fatty acid moieties in the oleic acid.
• ordinary oil or “conventional oil” refers to an oil obtained from commodity oilseeds, wherein the oleic acid content of said oil comprises less than 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23% 22%, 21 %, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 1 1 %, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1 % of the fatty acid moieties in the oil.
• enzyme activity refers to the ability of an enzyme to convert a substrate to a product.
• polynucleotide polynucleotide sequence
• nucleic acid sequence nucleic acid fragment
• isolated nucleic acid fragment are used interchangeably herein. These terms encompass nucleotide sequences and the like.
• a polynucleotide may be a polymer of RNA or DNA that is single- or double- stranded, that optionally contains synthetic, non-natural or altered nucleotide bases.
• a polynucleotide in the form of a polymer of DNA may be comprised of one or more segments of cDNA, genomic DNA, synthetic DNA, or mixtures thereof.
• Nucleotides (usually found in their 5'-monophosphate form) are referred to by a single letter designation as follows: "A” for adenylate or deoxyadenylate (for RNA or DNA, respectively), "C” for cytidylate or deoxycytidylate, "G” for guanylate or
• deoxyguanylate "U” for uridylate, “T” for deoxythymidylate, “R” for purines (A or G), “Y” for pyrimidines (C or T), “K” for G or T, “H” for A or C or T, “I” for inosine, and “N” for any nucleotide.
• fragment that is functionally equivalent and “functionally equivalent subfragment” are used interchangeably herein. These terms refer to a portion or subsequence of an isolated nucleic acid fragment in which the ability to alter gene expression or produce a certain phenotype is retained whether or not the fragment or subfragment encodes an active enzyme.
• the fragment or subfragment can be used in the design of chimeric genes to produce the desired phenotype in a transformed plant.
• Chimeric genes can be designed for use in suppression by linking a nucleic acid fragment or subfragment thereof, whether or not it encodes an active enzyme, in the sense or antisense orientation relative to a plant promoter sequence.
• corresponding substantially are used interchangeably herein. They refer to nucleic acid fragments wherein changes in one or more nucleotide bases do not affect the ability of the nucleic acid fragment to mediate gene expression or produce a certain phenotype. These terms also refer to modifications of the nucleic acid fragments of the instant invention such as deletion or insertion of one or more nucleotides that do not substantially alter the functional properties of the resulting nucleic acid fragment relative to the initial, unmodified fragment. It is therefore understood, as those skilled in the art will appreciate, that the invention encompasses more than the specific exemplary sequences.
• Gene refers to a nucleic acid fragment that expresses a specific protein, including regulatory sequences preceding (5' non-coding sequences) and following (3' non-coding sequences) the coding sequence.
• “Native gene” refers to a gene as found in nature with its own regulatory sequences.
• “Chimeric gene” refers to any gene that is not a native gene, comprising regulatory and coding sequences that are not found together in nature. Accordingly, a chimeric gene may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source, but arranged in a manner different than that found in nature.
• a “foreign” gene refers to a gene not normally found in the host organism, but that is introduced into the host organism by gene transfer. Foreign genes can comprise native genes inserted into a non-native organism, or chimeric genes.
• a “transgene” is a gene that has been introduced into the genome by a transformation procedure.
• An “allele” is one of several alternative forms of a gene occupying a given locus on a chromosome. When all the alleles present at a given locus on a chromosome are the same that plant is homozygous at that locus. If the alleles present at a given locus on a chromosome differ that plant is heterozygous at that locus.
• a “codon-optimized gene” is a gene having its frequency of codon usage designed to mimic the frequency of preferred codon usage of the host cell.
• Coding sequence refers to a DNA sequence that codes for a specific amino acid sequence.
• Regulatory sequences refer to nucleotide sequences located upstream (5' non-coding sequences), within, or downstream (3' non-coding sequences) of a coding sequence, and which influence the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences may include, but are not limited to, promoters, translation leader sequences, introns, and polyadenylation recognition sequences.
• Promoter refers to a region of DNA capable of controlling the expression of a coding sequence or functional RNA.
• the promoter sequence consists of proximal and more distal upstream elements. These upstream elements are often referred to as enhancers.
• an “enhancer” is a DNA sequence that can stimulate promoter activity, and may be an innate element of the promoter or a heterologous element inserted to enhance the level or tissue-specificity of a promoter. Promoters may be derived in their entirety from a native gene, or be composed of different elements derived from different promoters found in nature, or even comprise synthetic DNA segments.
• promoters may direct the expression of a gene in different tissues or cell types, or at different stages of development, or in response to different environmental conditions. It is further recognized that since in most cases the exact boundaries of regulatory sequences have not been completely defined, DNA fragments of some variation may have identical promoter activity. Promoters that cause a gene to be expressed in most cell types at most times are commonly referred to as "constitutive promoters". New promoters of various types useful in plant cells are constantly being discovered; numerous examples may be found in the compilation by
• any seed-specific promoter can be used in accordance with the method of the invention.
• the origin of the promoter chosen to drive expression of the recombinant DNA fragment is not critical as long as it is capable of accomplishing the invention by transcribing enough RNA from the desired nucleic acid fragment(s) in the seed.
• promoters A plethora of promoters is described in WO 00/18963, published on April 6, 2000, the disclosure of which is hereby incorporated by reference.
• seed-specific promoters include, and are not limited to, the promoter for soybean Kunitz trypsin inhibitor (Kti3, Jofuku and Goldberg (1989) Plant Cell 11079-1093) ⁇ -conglycinin (Chen et al. (1989) Dev. Genet. 10: 1 12-122), the napin promoter, and the phaseolin promoter.
• promoters that may be useful in expressing the nucleic acid fragments of the invention include, but are not limited to, the SAM synthetase promoter (PCT Publication WO00/37662, published June 29, 2000), the CaMV 35S (Odell et al (1985) Nature 373:810-812), and the promoter described in PCT
• translation leader sequence refers to a polynucleotide sequence located between the promoter sequence of a gene and the coding sequence.
• the translation leader sequence is present in the fully processed mRNA upstream of the translation start sequence.
• the translation leader sequence may affect processing of the primary transcript to mRNA, mRNA stability or translation efficiency.
• the "3' non-coding sequences” or “transcription terminator/termination sequences” refer to DNA sequences located downstream of a coding sequence and include polyadenylation recognition sequences and other sequences encoding regulatory signals capable of affecting mRNA processing or gene expression.
• the polyadenylation signal is usually characterized by affecting the addition of
• the use of different 3'non-coding sequences is exemplified by Ingelbrecht et al. (1989) Plant Cell 1 :671 -680.
• an "intron” is an intervening sequence in a gene that does not encode a portion of the protein sequence. Thus, such sequences are transcribed into RNA but are then excised and are not translated. The term is also used for the excised RNA sequences.
• An "exon” is a portion of the sequence of a gene that is
• RNA derived from the gene transcribed and is found in the mature messenger RNA derived from the gene, but is not necessarily a part of the sequence that encodes the final gene product.
• RNA transcript refers to the product resulting from RNA polymerase- catalyzed transcription of a DNA sequence. When the RNA transcript is a perfect complementary copy of the DNA sequence, it is referred to as the primary transcript. An RNA transcript is referred to as the mature RNA when it is an RNA sequence derived from post-transcriptional processing of the primary transcript.
• Messenger RNA (mRNA) refers to the RNA that is without introns and that can be translated into protein by the cell.
• cDNA refers to a DNA that is complementary to and synthesized from a mRNA template using the enzyme reverse transcriptase. The cDNA can be single-stranded or converted into the double-stranded form using the Klenow fragment of DNA polymerase I.
• Sense RNA refers to RNA transcript that includes the mRNA and can be translated into protein within a cell or in vitro.
• Antisense RNA refers to an RNA transcript that is complementary to all or part of a target primary transcript or mRNA, and that blocks the expression of a target gene (U.S. Patent No. 5,107,065).
• the complementarity of an antisense RNA may be with any part of the specific gene transcript, i.e., at the 5' non-coding sequence, 3' non-coding sequence, introns, or the coding sequence.
• “Functional RNA” refers to antisense RNA, ribozyme RNA, or other RNA that may not be translated but yet has an effect on cellular processes.
• complementary and reverse complement are used interchangeably herein with respect to mRNA transcripts, and are meant to define the antisense RNA of the message.
• operably linked refers to the association of nucleic acid sequences on a single nucleic acid fragment so that the function of one is regulated by the other.
• a promoter is operably linked with a coding sequence when it is capable of regulating the expression of that coding sequence (i.e., that the coding sequence is under the transcriptional control of the promoter).
• Coding sequences can be operably linked to regulatory sequences in a sense or antisense orientation.
• the complementary RNA regions of the invention can be operably linked, either directly or indirectly, 5' to the target mRNA, or 3' to the target mRNA, or within the target mRNA, or a first complementary region is 5' and its complement is 3' to the target mRNA.
• endogenous RNA refers to any RNA which is encoded by any nucleic acid sequence present in the genome of the host prior to transformation with the recombinant construct of the present invention, whether naturally-occurring or non-naturally occurring, i.e., introduced by recombinant means, mutagenesis, etc.
• non-naturally occurring means artificial, not consistent with what is normally found in nature.
• PCR or “Polymerase Chain Reaction” is a technique for the synthesis of large quantities of specific DNA segments, consists of a series of repetitive cycles (Perkin Elmer Cetus Instruments, Norwalk, CT). Typically, the double stranded DNA is heat denatured, the two primers complementary to the 3' boundaries of the target segment are annealed at low temperature and then extended at an
• One set of these three consecutive steps is referred to as a cycle.
• recombinant refers to an artificial combination of two otherwise separated segments of sequence, e.g., by chemical synthesis or by the
• Plasmid refers to an extra chromosomal element often carrying genes that are not part of the central metabolism of the cell, and usually in the form of circular double-stranded DNA fragments.
• Such elements may be autonomously replicating sequences, genome integrating sequences, phage or nucleotide sequences, linear or circular, of a single- or double-stranded DNA or RNA, derived from any source, in which a number of nucleotide sequences have been joined or recombined into a unique construction which is capable of
• Transformation cassette refers to a specific vector containing a foreign gene and having elements in addition to the foreign gene that facilitates transformation of a particular host cell.
• Expression cassette refers to a specific vector containing a foreign gene and having elements in addition to the foreign gene that allow for enhanced expression of that gene in a foreign host.
• a recombinant construct comprises an artificial combination of nucleic acid fragments, e.g., regulatory and coding sequences that are not found together in nature.
• a chimeric construct may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source, but arranged in a manner different than that found in nature.
• Such construct may be used by itself or may be used in conjunction with a vector. If a vector is used then the choice of vector is dependent upon the method that will be used to transform host cells as is well known to those skilled in the art.
• a plasmid vector can be used.
• the skilled artisan is well aware of the genetic elements that must be present on the vector in order to successfully transform, select and propagate host cells comprising any of the isolated nucleic acid fragments of the invention.
• the skilled artisan will also recognize that different independent transformation events will result in different levels and patterns of expression (Jones et al., (1985) EMBO J. 4:241 1 -2418;
• Such screening may be accomplished by Southern analysis of DNA, Northern analysis of mRNA expression, immunoblotting analysis of protein expression, or phenotypic analysis, among others.
• expression refers to the production of a functional end-product e.g., a mRNA or a protein (precursor or mature).
• expression cassette refers to a discrete nucleic acid fragment into which a nucleic acid sequence or fragment can be moved.
• “Mature” protein refers to a post-translationally processed polypeptide; i.e., one from which any pre- or propeptides present in the primary translation product have been removed.
• Precursor protein refers to the primary product of translation of mRNA; i.e., with pre- and propeptides still present. Pre- and propeptides may be but are not limited to intracellular localization signals.
• Codon refers to the production of sense RNA transcripts capable of suppressing the expression of identical or substantially similar native genes (U.S. Patent No. 5,231 ,020, which issued to Jorgensen et al. on July 27, 1999). Co- suppression constructs in plants have been previously designed by focusing on overexpression of a nucleic acid sequence having homology to a native mRNA, in the sense orientation, which results in the reduction of all RNA having homology to the overexpressed sequence (see Vaucheret et al. (1998) Plant J. 76:651 -659; and Gura (2000) Nature 404:804-808).
• Antisense inhibition refers to the production of antisense RNA transcripts capable of suppressing the expression of the target protein.
• Plant viral sequences may be used to direct the suppression of proximal mRNA encoding sequences (PCT Publication WO 98/36083 published on August 20, 1998).
• "Hairpin" structures that incorporate all, or part, of an mRNA encoding sequence in a complementary orientation resulting in a potential “stem-loop" structure for the expressed RNA have been described (PCT Publication WO
• the stem is formed by polynucleotides corresponding to the gene of interest inserted in either sense or anti-sense orientation with respect to the promoter and the loop is formed by some polynucleotides of the gene of interest, which do not have a complement in the construct.
• This increases the frequency of cosuppression or silencing in the recovered transgenic plants.
• Yet another variation includes using synthetic repeats to promote formation of a stem in the stem-loop structure.
• Transgenic organisms prepared with such recombinant DNA fragment show reduced levels of the protein encoded by the polynucleotide from which the nucleotide fragment forming the loop is derived as described in PCT Publication WO 02/00904, published January 3, 2002.
• the use of constructs that result in dsRNA has also been described. In these constructs convergent promoters direct transcription of gene-specific sense and antisense RNAs inducing gene suppression (see for example Shiet al. (2000) RNA 6:1069-1076; Bastinet al. (2000) J. Cell Sci. 773:3321 -3328; Giordanoet al. (2002) Genetics 760:637-648; LaCount-and
• miRNAs are small regulatory RNAs that control gene expression. miRNAs bind to regions of target RNAs and inhibit their translation and, thus, interfere with production of the polypeptide encoded by the target RNA. miRNAs can be designed to be complementary to any region of the target sequence RNA including the 3' untranslated region, coding region, etc. miRNAs are processed from highly structured RNA precursors that are processed by the action of a ribonuclease III termed DICER. While the exact mechanism of action of miRNAs is unknown, it appears that they function to regulate expression of the target gene. See, e.g., U.S. Patent Publication No. 2004/0268441 Al which was published on December 30, 2004.
• expression refers to the production of a
• Antisense inhibition refers to the production of antisense RNA transcripts capable of suppressing the expression of the target protein.
• Co-suppression refers to the production of sense RNA transcripts capable of suppressing the expression of identical or substantially similar foreign or endogenous genes (U.S. Patent No. 5,231 ,020).
• “Overexpression” refers to the production of a functional end-product in transgenic organisms that exceeds levels of production when compared to expression of that functional end-product in a normal, wild type or non-transformed organism.
• “Stable transformation” refers to the transfer of a nucleic acid fragment into a genome of a host organism, including both nuclear and organellar genomes, resulting in genetically stable inheritance.
• “transient transformation” refers to the transfer of a nucleic acid fragment into the nucleus, or DNA-containing organelle, of a host organism resulting in gene expression without integration or stable inheritance.
• Host organisms containing the transformed nucleic acid fragments are referred to as "transgenic" organisms.
• Oilseed plant cells are the preferred plant cells.
• the transformed plant cell is then cultured and regenerated under suitable conditions permitting expression of the recombinant construct which is then recovered and purified.
• Recombinant constructs may be introduced into one plant cell or,
• a construct may be introduced into separate plant cells.
• Expression in a plant cell may be accomplished in a transient or stable fashion as is described above.
• Plant parts include differentiated and undifferentiated tissues, including but not limited to: roots, stems, shoots, leaves, pollen, seeds, tumor tissue, and various forms of cells and culture such as single cells, protoplasts, embryos, and callus tissue.
• the plant tissue may be in plant or in organ, tissue or cell culture.
• plant organ refers to plant tissue or group of tissues that constitute a morphologically and functionally distinct part of a plant.
• the term “genome” refers to the following: 1 . The entire complement of genetic material (genes and non- coding sequences) is present in each cell of an organism, or virus or organelle. 2. A complete set of chromosomes inherited as a (haploid) unit from one parent.
• the term “stably integrated” refers to the transfer of a nucleic acid fragment into the genome of a host organism or cell resulting in genetically stable inheritance.
• the particular method of regeneration will depend on the starting plant tissue and the particular plant species to be regenerated.
• the regeneration, development and cultivation of plants from single plant protoplast transformants or from various transformed explants is well known in the art (Weissbach and Weissbach, (1988) In.: Methods for Plant Molecular Biology, (Eds.), Academic: San Diego, CA).
• This regeneration and growth process typically includes the steps of selection of transformed cells, culturing those individualized cells through the usual stages of embryonic development through the rooted plantlet stage. Transgenic embryos and seeds are similarly regenerated. The resulting transgenic rooted shoots are thereafter planted in an appropriate plant growth medium such as soil.
• the regenerated plants are self-pollinated to provide homozygous transgenic plants. Otherwise, pollen obtained from the regenerated plants is crossed to seed-grown plants of agronomically important lines. Conversely, pollen from plants of these important lines is used to pollinate regenerated plants.
• a transgenic plant of the present invention containing a desired polypeptide is cultivated using methods well known to one skilled in the art.
• Oxidation and therefore the shelf life of animal feed ingredients is a common problem in the industry. Oxidation is an irreversible chemical reaction in which oxygen reacts with feed and feed components and can result in decreased animal health and performance. The negative effects of oxidation can be seen in loss of palatability, degradation of the oil component, development of unwanted breakdown products, changes in color, and loss of energy. Meat obtained from animals grown on oxidized feed has significantly lower oxidative status compared to animals fed a feed that has not undergone significant oxidation. Meat from animals fed diets containing high oleic corn products show extended shelf life and greater oxidative stability (PCT Publication WO/2006/002052, published January 5 th , 2006), particularly when combined with antioxidants such as tocols. Therefore it is highly desirable to prevent oxidation of feed and feed ingredients to protect both nutritional value and organoleptic quality.
• Synthetic antioxidants are used to preserve feed quality by preventing the oxidation of lipids, which can lead to improved animal performance.
• synthetic antioxidants can act as free radical scavengers and thereby reduce lipid oxidation. Synthetic antioxidants can prolong animal feed shelf-life and protect nutritional and organoleptic quality
• oxidation status of solid materials including soybean meal and other soybean protein products
• accelerating aging methods which predict a material's shelf-life.
• One test which can be used is to age a material either at room temperature or elevated temperatures and to measure the oxidative status of the material at specific time points.
• the OSI instrument is useful in this regard in that it reflects the length of time needed to start the oxidation process known as the induction time. A longer induction time means that the material has greater oxidative stability and thereby shelf-life.
• Other methods include the measurement of volatiles and color change.
• soybean protein products can be obtained in a variety of ways. Conditions typically used to prepare soy protein isolates have been described by (Cho, et al, (1981 ) U.S. Patent No. 4,278,597; Goodnight, et al. (1978) U.S. Patent No. 4,072,670). Soy protein concentrates are produced by three basic processes: acid leaching (at about pH 4.5), extraction with alcohol (about 55-80%), and denaturing the protein with moist heat prior to extraction with water. Conditions typically used to prepare soy protein concentrates have been described by Pass ((1975) U.S. Patent No. ,897,574) and Campbell et al. ((1985) in New Protein Foods, ed. by Altschul and Wilcke, Academic Press, Vol., Chapter 10, Seed Storage Proteins, pp 302-338).
• Soybean-containing products or “Soy products” can be defined as those products containing/incorporating a soy protein product.
• “soy protein products” can include, and are not limited to, those items listed in Table 2.
• Soy Protein Council aSee Soy Protein Products: Characteristics, Nutritional Aspects and Utilization (1987). Soy Protein Council.
• Processing refers to any physical and chemical methods used to obtain the products listed in Table 2 and includes, and is not limited to, heat conditioning, flaking and grinding, extrusion, solvent extraction, or aqueous soaking and extraction of whole or partial seeds. Furthermore, “processing” includes the methods used to concentrate and isolate soy protein from whole or partial seeds, as well as the various traditional Oriental methods in preparing fermented soy food products. Trading Standards and Specifications have been established for many of these products (see National Oilseed Processors Association Yearbook and Trading Rules 1991 -1992).
• Defatted flakes refer to flaked, dehulled cotyledons that have been defatted and treated with controlled heat to remove the remaining hexane. This term can also refer to a flour or grit that has been ground.
• White flakes refer to flaked, dehulled cotyledons that have been defatted and treated with controlled heat to remove the remaining hexane. This term can also refer to a flour that has been ground.
• Grits refer to defatted, dehulled cotyledons having a U.S. Standard screen size of between No. 10 and 80.
• Soy Protein Concentrates refer to those products produced from dehulled, defatted soybeans and typically contain 65 wt % to 90 wt % soy protein on a moisture free basis. Soy protein concentrates are typically manufactured by three basic processes: acid leaching (at about pH 4.5), extraction with alcohol (about 55-80%), and denaturing the protein with moist heat prior to extraction with water. Conditions typically used to prepare soy protein concentrates have been described by Pass (1975) U.S. Patent No. 3,897,574; Campbell et al., (1985) in New Protein Foods, ed. by Altschul and Wilcke, Academic Press, Vol. 5, Chapter 10, Seed Storage Proteins, pp 302-338). As used herein, the term “soy protein isolate” or “isolated soy protein” refers to a soy protein containing material that contains at least 90% soy protein by weight on a moisture free basis.
• Extrusion refers to processes whereby material (grits, flour or concentrate) is passed through a jacketed auger using high pressures and temperatures as a means of altering the texture of the material.
• Textturing and structuraluring refer to extrusion processes used to modify the physical characteristics of the material. The characteristics of these processes, including thermoplastic extrusion, have been described previously (Atkinson (1970) U.S. Patent No. 3,488,770, Horan (1985) In New Protein Foods, ed. by Altschul and Wilcke, Academic Press, Vol. 1A,
• Residual fatty acid analysis The commercial process used to de-fat soy flakes with hexane leaves a residue of fatty acids that can act as substrate for generation of off-flavor compounds.
• the residual fat content of hexane-defatted soy flakes can range from, 0.6-1 .0% (W:W) (ether extractable; AOCS Method 920.39 (Official Methods of Analysis of the AOAC International (1995), 16 th Edition, Method 920.39C, Locator #4.2.01 (modified)) to 2.5 - 3% (W:W) (acid hydrolysable; AOAC Method 922.06 (Official Methods of Analysis of the AOAC International (1995), 16 th Edition, Method 922.06, Locator 32.1 .13 (modified)).
• the principle reason for the discrepancy between these two methods of estimating residual fatty acids is the chemical nature of the fat classes associated with the protein matrix after hexane extraction.
• a small proportion of the residual fatty acid is in the form of neutral lipid (i.e., triglyceride) and the remainder is present as polar lipid (e.g., phospholipids, a.k.a., lecithin). Because of its polar nature the phospholipid is inaccessible to ether extraction and is only removed from the protein matrix if acid hydrolysis or some other stringent extraction protocol is performed.
• the ether extraction technique gives an estimation of the neutral lipid fraction whereas the acid hydrolysable method gives a better estimate of the total residual fatty acid content (i.e., neutral and polar fractions).
• Both of the AOAC methods described above rely on gravimetric determinations of the residual fatty acids and, although in combination they give an indication of the fat classes (neutral vs. polar), such estimates are crude and are subject to interference from other hydrophobic materials (e.g. saponins). Further, no information is obtained on the fatty acid composition and how it may have been affected by various experimental treatments or by the genetics of the starting material.
• AOAC methods for the determination of the fatty acid composition of residual fatty acids are available (Official Methods of Analysis of the AOAC
• Locator 41 .1 .28A Locator 41 .1 .28A.
• These are based on the conversion of residual fatty acids, extracted by acid hydrolysis, to fatty acid methyl esters prior to analysis by gas chromatography. Such techniques are rarely used to assess the residual fatty acid content of food materials in commercial settings although they are used for fatty acid evaluations in support of nutritional labeling. A report in which these methods have been used to determine the residual fatty acid composition of commercial soy protein isolates has recently been published (Solina et al. (2005) Volatile aroma components of soy protein isolate and acid-hydrolysed vegetable protein Food Chemistry 90: 861 -873)
• Example 24 A facile method for determining the fatty acid composition of the residual fats in soy protein products is described in Example 24.
• the advantage of this method over others is that it requires no extraction of the residual fats from the matrix prior to derivatization for GC analysis.
• the technique is suitable for all forms of fatty acids i.e., whether they are initially present as free fatty acids or as fatty acid esters e.g., tri-glycerides or phospholipids (Chistie (1989) Gas Chromatography and Lipids; The Oily Press. Ayr, Scotland).
• the technique will also remove fatty acids from the protein matrix even if the polar head group of the phospholipid is covalently bound to the protein.
• a soybean protein product of the invention can be in a liquid or in a dry powdered form.
• the foods to which the soybean protein product of the invention can be incorporated/added include almost all foods, beverages and feed (such as pet foods).
• food supplements, food bars, meats such as meat alternatives, ground meats, emulsified meats, marinated meats, and meats injected with a soybean protein product of the invention.
• beverages such as nutritional beverages, sports beverages, protein-fortified beverages, juices, milk, milk alternatives, and weight loss beverages.
• cheeses such as hard and soft cheeses, cream cheese, and cottage cheese. Included may also be frozen desserts such as ice cream, ice milk, low fat frozen desserts, and non-dairy frozen desserts. Finally, yogurts, soups, puddings, bakery products, salad dressings, spreads, and dips (such as mayonnaise and chip dips) may be included.
• a soy protein product can be added in an amount selected to deliver a desired amount to a food and/or beverage.
• soybean protein product and “soy protein product” are used interchangeably herein.
• Any high oleic soybean seed whether transgenic or non-transgenic, can be used as a source of soy protein product.
• Soybeans with decreased levels of saturated fatty acids have been described resulting from mutation breeding (Erickson et al. (1994) J. Hered. 79:465-468;
• Soybeans with decreased levels of polyunsaturated fatty acids have been described resulting from mutation breeding and selection. Reduced levels of linolenic acid have been achieved at relatively constant linoleic acid (U.S. Patent No. 5,710,369 and U.S. Patent No. 5,986,1 18). Decreased linoleic and linolenic acids combined have also been achieved using mutation breeding, genetic crosses and selection (Rahman, S. M. et al. (2001 ) Crop Sci. 41 :26-29).
• soybean seeds with oil profiles having linolenic acid contents of from 1 % to 3% of the total fatty acids and total levels of polyunsaturated fatty acids of about 30 to 35% as compared to greater than 6% linolenic acid and greater than 50% total polyunsaturated fatty acids in commodity soybeans.
• WO 94/1 1516 can be used to make a high oleic acid soybean variety.
• the resulting high oleic acid soybean variety was one in which the polyunsaturated fatty acids were reduced from 70% of the total fatty acids to less than 5%.
• FAD2-1 and FAD2-2 Two soybean fatty acid desaturases, designated FAD2-1 and FAD2-2, are ⁇ -12 desaturases that introduce a second double bond into oleic acid to form linoleic acid, a polyunsaturated fatty acid.
• FAD2-1 is expressed only in the developing seed (Heppard et al. (1996) Plant Physiol. 1 10:31 1 -319). The
• GmFad 2-1 is described in detail by Okuley, J. et al. (1994) Plant Cell 6:147-158 and in WO94/1 1516. It is available from the ATCC in the form of plasmid pSF2-169K (ATCC accession number 69092).
• FAD 2-2 is expressed in the seed, leaf, root and stem of the soy plant at a constant level and is the "housekeeping" 12-desaturase gene.
• the Fad 2-2 gene product is responsible for the synthesis of polyunsaturated fatty acids for cell membranes.
• FAD2-1 is the major enzyme of this type in soybean seeds, reduction in the expression of FAD2-1 results in increased accumulation of oleic acid (18:1 ) and a corresponding decrease in polyunsaturated fatty acid content.
• FAD2-2 Reduction of expression of FAD2-2 in combination with FAD2-1 leads to a greater accumulation of oleic acid and corresponding decrease in polyunsaturated fatty acid content.
• FAD3 is a ⁇ -15 desaturase that introduces a third double bond into linoleic acid (18:2) to form linolenic acid (18:3).
• Reduction of expression of FAD3 in combination with reduction of FAD2-1 and FAD2-2 leads to a greater accumulation of oleic acid and corresponding decrease in polyunsaturated fatty acid content, especially linolenic acid.
• Nucleic acid fragments encoding FAD2-1 , FAD2-2, and FAD3 have been described in WO 94/1 1516 and WO 93/1 1245.
• Chimeric recombinant constructs comprising all or a part of these nucleic acid fragments or the reverse complements thereof operably linked to at least one suitable regulatory sequence can be constructed wherein expression of the chimeric gene results in an altered fatty acid phenotype.
• a chimeric recombinant construct can be introduced into soybean plants via transformation techniques well known to those skilled in the art.
• the recombinant DNA are assayed to select plants with altered fatty acid profiles.
• the recombinant construct may contain all or part of 1 ) the FAD2-1 gene or 2) the
• Recombinant constructs comprising all or part of 1 ) the FAD2-1 gene with or without 2) all or part of the Fad2-2 gene with or without all or part of the FAD3 gene can be used in making a transgenic soybean plant having a high oleic phenotype.
• An altered fatty acid profile specifically an increase in the proportion of oleic acid and a decrease in the proportion of the polyunsaturated fatty acids, indicates that one or more of the soybean seed FAD genes (FAD2-1 , Fad2-2, FAD3) have been suppressed.
• Assays may be conducted on soybean somatic embryo cultures and seeds to determine suppression of FAD2-1 , Fad2-2, or FAD3.
• constructs comprising sequences other than those specifically exemplified which have similar functions, may be used. These constructs may include any seed- specific promoter. These constructs may or may not also include any nucleotides that promote stem-loop formation. These constructs may contain a polynucleotide having a nucleotide sequence identical to any portion of the gene or genes mentioned above inserted in sense or anti-sense orientation with respect to the promoter. Finally, these constructs may or may not contain any transcription termination signal.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil has an increased oleic content when compared to an ordinary frying oil.
• the environmentally preferred frying oil is also useful as a blending source to make a blended environmentally preferred frying oil.
• the environmentally preferred frying oils of the invention are obtained from high olei oilseeds, such as, but not limited to, soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower.
• the preferred frying oil of the invention comprises an oleic acid content of at least 60% of the fatty acid moieties in the oil.
• Another embodiment of the invention concerns a method for frying with a reduced impact on the environment , comprising: using an oil with an increased oleic acid content when compared to an ordinary oil and quantifying the reduction in environmental impacts when using a frying oil with an increased oleic acid content compared to an ordinary frying oil.
• the reduction of environmental impact can be at least one selected from the group consisting of: reduced carbon footprint , reduced eutrophication potential, reduced air acidification potential, and reduced nonrenewable energy consumption.
• Another embodiment of the invention concerns the use of the high oleic oil for frying applications, wherein the reduction of the impact on the environment is at least one selected from the group consisting of: reduced carbon footprint , reduced eutrophication potential, reduced air acidification potential, and reduced non- renewable energy consumption, when compared to the use of an ordinary oil for the same application.
• the environmentally preferred or high oleic oils of the invention can be used as a blending source with an ordinary oil.
• An additional embodiment of the invention concerns the use of an high oleic oil in a frying process, wherein the burden on the environment is reduced by at least 40% when compared to the use of a conventional oil .
• the goal of this LCA was to compare high oleic oil with conventional oil used in a large-scale restaurant fryer application in the United States
• the environmental impacts studied in this LCA were greenhouse gas emissions, non-renewable energy use, eutrophication potential, and acidification potential. These were selected as critical environmental issues associated with agriculture related processes.
• Losses to food were assumed to be 8 lbs per day. Top-off is required when oil is not replaced at the end of a day. For high oleic oil, 208 lbs of oil (8 lbs top-off for the fryers) are used for two days of frying in a fast food restaurant. For conventional oil, 400 lbs of oil are used for two days of frying in a fast food restaurant. Top off is not required for conventional oil since the oil is replaced each day. Washing of the fryer was included and assumed to be required every time the oil is changed.
• Soybean oil refining, bleaching, and deodorizing • Soybean oil use in a large-scale restaurant fryer
• Yellow grease (the waste oil from the restaurant) is of no economic value or cost for the restaurant. Therefore, it is assumed that the yellow grease has no burdens and gets no credits, per the attributional approach from the perspective of the restaurant
• Packaging is not included due to scope and time limitations. Including packaging would likely only improve the environmental profile of the high oleic oil further, since about half as much would need to be produced and therefore packaged, for the restaurant.
• Research Service Oil Crops yearbook for the U.S. provides price and supply history for the U.S. soybean oil and soybean meal on a monthly basis from 2004 through the summer of 2009 at the time when data was collected for this study.
• the mundi index provides a monthly price history for meal and oil for a ten year timespan through the end of 201 1 based on the Chicago Soybean Oil and Soybean Meal futures without associated supply volumes. Pricing agreed across both methods for the years where data was common. Using data that includes the most current pricing was deemed important for this analysis. As such we chose to use the data available through index Mundi.
• the entire supply chain from a restaurant's perspective for both high oleic oil and conventional oil was modeled.
• the supply chain included the farming of the crop, the pressing and extracting of the crop into crude oil and meal, refining the crude oil into food grade oil by neutralizing, bleaching and deodorizing, washing of the fryer, and end of life of the oil from the perspective of the restaurant.
• the model also included transportation for all inputs and the beans and oil.
• Conventional and high oleic soybean oils are assumed to be representative for other types of conventional and high oleic oils. It is assumed that there is no difference between processing a high oleic soybean and a conventional soybean. The soybean and oil are produced in mid-west USA, Illinois, and then transported via truck to a restaurant in New York City.
• Both base cases assume that environmental burdens are partitioned to co-products based on the co-product's economic value.
• Soy meal is produced during the pressing and extracting of the soybean into crude oil.
• a distillate material and soapstock are also sold as co-products, and are produced at the refining, bleaching and deodorizing of the refined oil.
• the base cases model two days at the restaurant, with washing procedures based on Stratus Foods Fryer Tips. Each time the fryer is emptied, it is cleaned and during cleaning the fryer is filled two times with water that is heated to 250 F and has some vinegar added to it. After the oil is used at the restaurant it is referred to as yellow grease, which has no economic value or cost for the restaurant. The yellow grease therefore receives no environmental benefit or burden in the LCA.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil has an increased oleic content when compared to an ordinary frying oil.
• the invention concerns an environmentally preferred frying oil, wherein said environmentally preferred frying oil is useful as a blending source to make a blended environmentally preferred frying oil.
• the invention concerns an environmentally preferred frying oil obtained from a high oleic oilseed.
• the oilseed is one selected from the group consisting of: soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower.
• the invention concerns an environmentally preferred frying oil, wherein the oleic acid content of said oil comprises at least 60% of the fatty acid moieties in the oil.
• the invention concerns a method to determine that an oil is environmentally preferred for a frying process, comprising:
• a further embodiment of the invention concerns a method to determine that an oil is environmentally preferred for a frying process, comprising using an oil with an increased oleic acid content when compared to an ordinary oil and quantifying the reduction in environmental impacts when using a frying oil with an increased oleic acid content compared to an ordinary frying oil, wherein the reduction environmental impact is at least one selected from the group consisting of: reduced carbon footprint , reduced eutrophication potential, reduced air acidification potential, and reduced non-renewable energy consumption.
• High oleic oil obtained from seeds , soybean, palm, peanut, canola, sunflower, corn, flax, cotton, and safflower are also part of the invention.
• a further embodiment of the invention concerns the use of a high oleic oil, wherein the use of the oil for frying applications reduces landuse pressure.
• An additional embodiment of the invention concerns use of a high oleic frying oil for reduction of environmental impact, such as a reduction of carbon footprint, reduction of eutrophication potential, reduction of air acidification potential, or reduction of non-renewable energy consumption.
• Soybean embryogenic suspension cultures are transformed by the method of particle gun bombardment using procedures known in the art (Klein et al. (1987) Nature (London) 327:70-73; U. S. Patent No. 4,945,050; Hazel et al. (1998) Plant Cell. Rep. 17:765-772; Samoylov et al. (1998) In Vitro Cell Dev. Biol.-Plant 34:8-13).
• particle gun bombardment procedures it is possible to use purified 1 ) entire plasmid DNA or, 2) DNA fragments containing only the recombinant DNA
• Stock tissue for transformation experiments are obtained by initiation from soybean immature seeds. Secondary embryos are excised from explants after 6 to 8 weeks on culture initiation medium.
• the initiation medium is an agar-solidified modified MS (Murashige and Skoog (1962) Physiol. Plant. 75:473-497) medium supplemented with vitamins, 2,4-D and glucose. Secondary embryos are placed in flasks in liquid culture maintenance medium and maintained for 7-9 days on a gyratory shaker at 26 +/- 2°C under -80 Em-2s-1 light intensity.
• the culture maintenance medium is a modified MS medium supplemented with vitamins, 2,4-D, sucrose and asparagine.
• clumps of tissue Prior to bombardment, clumps of tissue are removed from the flasks and moved to an empty 60X15 mm petri dish for bombardment. Tissue is dried by blotting on Whatman #2 filter paper. Approximately 100-200mg of tissue corresponding to 10-20 clumps (1 -5 mm in size each) are used per plate of bombarded tissue.
• tissue from each bombarded plate is divided and placed into two flasks of liquid culture maintenance medium per plate of bombarded tissue. Seven days post bombardment, the liquid medium in each flask is replaced with fresh culture maintenance medium supplemented with 100ng/ml selective agent (selection medium).
• selective agent used can be a sulfonylurea (SU) compound with the chemical name,
• benzenesulfonamide (common names: DPX-W4189 and chlorsulfuron).
• Chlorsulfuron is the active ingredient in the DuPont sulfonylurea herbicide
• GLEAN® The selection medium containing SU is replaced every week for 6-8 weeks. After the 6-8 week selection period, islands of green, transformed tissue are observed growing from untransformed, necrotic embryogenic clusters. These putative transgenic events are isolated and kept in media with SU at 100 ng/ml for another 2-6 weeks with media changes every 1 -2 weeks to generate new, clonally propagated, transformed embryogenic suspension cultures. Embryos spend a total of around 8-12 weeks in contact with SU. Suspension cultures are subcultured and maintained as clusters of immature embryos and also regenerated into whole plants by maturation and germination of individual somatic embryos.
• the relative amounts of the fatty acids, palmitic, stearic, oleic, linoleic and linolenic can be determined as follows.
• Fatty acid methyl esters are prepared from single, mature, somatic soybean embryos or soybean seed chips by transesterification. One embryo, or a chip from a seed, is placed in a vial containing 50 ⁇ _ of trimethylsulfonium hydroxide and incubated for 30 minutes at room temperature while shaking.
• Triplicate samples (approximately 100mg) were weighed, to a precision of 0.1 mg, into 13 x 100 mm screw capped (PTFE liners) tubes.
• C17:0 triacylglycerol internal standard (10 ⁇ , 5% W:V stock in toluene)
• 1 ml of fresh methanolysis solution (5% sulfuric acid in anhydrous methanol) was added to each tube.
• the tubes were capped, vortex mixed and heated at 80°C for 30 min, with vortex mixing every 10 minutes.
• the samples were cooled to room temperature and 1 ml of saline solution (25% sodium chloride in water), followed by 1 ml heptane, was added to each tube. After vortex mixing, the phases were separated by
• the residual fatty acids associated with the hexane-defatted white flake flour and soy protein isolate is principally in the form of phospholipid, and therefore derived from membrane lipids, while the hexane-extracted soy oil is principally composed of storage triglycerides.
• Prior to this work it was not known how closely the residual fatty acid profile would be related to the fatty acid profile of hexane- extracted soy oil. From the data shown in Table 9 it can be seen that the level of palmitic acid increases in the residual fatty acids present in soy white flake flour and soy protein isolate compared to hexane-extracted soy oil in the three genetically different soybean varieties tested.
• the level of oleic acid decreases in the residual fatty acids compared to hexane-extracted soy oil significantly in the commodity and low linolenic acid soybeans, but only marginally in the high oleic soybeans.
• the polyunsaturated fatty acids, linoleic and linolenic are at similar levels in the residual fatty acids and hexane-extracted soy oil from the three genetically different soybean varieties.
• soy white flake flour and soy protein isolate from low linolenic acid soybeans is lower in oxidatively unstable linolenic acid than that of commodity soy protein products, indicating that soy protein products produced from low linolenic acid soybeans are less likely to generate off-flavor compounds.
• the residual fatty acid content in soy white flake flour and soy protein isolate from high oleic acid soybeans is lower in both of the polyunsaturated fatty acids, linoleic and linolenic, than that of commodity soy protein products, indicating that soy protein products produced from high oleic acid soybeans are less likely to generate off-flavor compounds.
• fatty acid % relates the individual fatty acid to the sum of the five major fatty acids indicated. Other fatty acid types that are sometimes present and represent less than 3% of the total fatty acids are not considered for purposes of comparison. Value ranges for the five major fatty acids in commodity soy oil are taken from "The Lipid Handbook" 2 na ed., (1994) Gunstone, F.D., Harwood, J.L., Padley, F.B., Chapman & Hall.
• 3SPI Soy protein isolate produced from white flake flour
• Fryers are set to 176C (-350F).
• Fryers charged with oil to cold fill line and sample ( ⁇ 250ml) is collected for analysis. Samples taken into brown glass bottle and are nitrogen capped and stored at 4C until analysis. In most cases analysis is performed on the day of collection. Bring fryers up to temperature and measure polar compounds with quick test (Testo and FOM; instruments are inserted in oil to specified depth and moved slowly in a figure of 8 pattern prior to taking readings of temperature and total polar
• Oxidative Stability Index (OSI); AOCS Cd 12b-92
• %TPC refers to the percentage of Total Polar Compound s. This measuren used to estimate the degradation of the oil during frying. Once the %TPC reaches 25% the oil is considered spent.
• PlenishTM is a high oleic soybean oil.
• the anti-foaming agent silicon and TBQH are usually added to oil that are used for frying on a commercial scale. Data with and without addition of these additives are shown.
• the term “Farming” includes the production, transportation, application of the fertilizers, soil emissions, and the energy used for the tractors and driers at the farm.
• the term “Crude Oil” includes the transportation of the soybean from the farm to the crude oil production facility, and all energy and materials associated with the pressing and extracting of the soybean into the crude oil and meal.
• the term “Refined Oil” pertains to the neutralization, bleaching and deodorizing process steps to make food grade soybean oil.
• the “Transport” step of the supply chain includes the one-way transportation of the refined oil from the plant in Illinois to a New York City restaurant.
• the term “Washing” includes the cleaning of the fryer after each time the fryer is emptied.
• climate change refers to any significant change in the climate lasting for an extended period. climate change can be caused by natural factors, natural processes, and human activities. climate change potential is measured in terms of total greenhouse gas emissions, and takes into account the global warming potential of specific species known to contribute to climate change.
• the climate change potential for both conventional oil and high oleic oil base cases can be seen below in Figure 2.
• the climate change potential for the conventional oil is 220 kg CO2 eq per 2 day fryer use, and for high oleic oil the climate change potential is 120 kg CO2 eq per 2 day fryer use.
• the base case for high oleic oil is 45% lower than the base case for conventional oil in terms of climate change potential.
• "Farming” contributed roughly 50% and was the largest contributor to both oils.
• the "Crude Oil” piece of the supply chain is the second largest contributor at roughly 23% of the climate change potential.
• “Washing” of the fryer is also a significant contributor, which contributed 16% to conventional oil and 14% to high oleic oil.
• "Refined Oil” and “Transport” are both under 10% of the total climate change potential for a restaurant.
• Non-renewable energy use accounts for all of the coal, oil, natural gas, and uranium consumed in the supply chain.
• Conventional oil and high oleic oil base case non-renewable energy use from a restaurant's perspective for two day fryer use can be seen in Figure 3.
• the high oleic oil base case was 45% lower in non-renewable energy use than the conventional oil base case, with high oleic oil using 32 kg oil eq per 2 day fryer use and conventional oil using 59 kg oil eq per 2 day fryer use.
• "Farming" was the largest contributor for non-renewable energy, which contributed 35% for high oleic oil and 33% for conventional oil.
• the "Crude Oil” step contributed 31 % of the total non-renewable energy use for high oleic oil and 33% for conventional oil.
• the "Washing” step contributed 21 % of the total non-renewable energy use for high oleic oil and 23% for conventional oil.
• the "Refined Oil” and “Transport” steps contributed less than 10% of the total non-renewable energy use.
• Terrestrial acidification potential is caused by emissions which alter optimum soil pH.
• the emissions that contribute to acidification are NOx, ammonia, and SO2.
• Terrestrial acidification potential for both base cases can be seen in Figure 4.
• the terrestrial acidification potential for the conventional oil is 1 .6 kg SO 2 eq per 2 day fryer use, and for high oleic oil the terrestrial acidification potential is 0.91 kg SO2 eq per 2 day fryer use.
• the base case for high oleic oil is 44% lower than the base case for conventional oil.
• “Farming” was the largest contributor at 65% for conventional oil and 67% for high oleic oil.
• the “Crude Oil” step in the supply chain is the second largest contributor at roughly 14% of the terrestrial acidification potential.
• “Washing" of the fryer is also a significant contributor, which contributed 13% to conventional oil and 1 1 % to high oleic oil.
• "Refined Oil” and “Transport” are both under 5% of the total terrestrial acidification potential for a restaurant.
• Freshwater eutrophication occurs when a fresh waterbody is overloaded with nutrients. This overload causes an increase in algal growth and a subsequent reduction in oxygen availability for aquatic life. Freshwater eutrophication is caused by phosphorous-containing emissions.
• Freshwater eutrophication potential for conventional oil and high oleic oil use in a restaurant can be seen below in Figure 5.
• the high oleic oil base case was 43% lower in freshwater eutrophication potential than the conventional oil base case, with high oleic oil resulting in 0.1 1 kg P eq per 2 day fryer use and conventional oil resulting in 0.19 kg P eq per 2 day fryer use.
• "Farming" was by far the largest contributor for freshwater eutrophication potential, which contributed 82% for high oleic oil and 83% for conventional oil.
• the "Washing" phase contributed 8% of the total freshwater eutrophication potential for high oleic oil and 9% for conventional oil.
• the "Crude Oil” step contributed 7% of the total freshwater eutrophication potential for both high oleic oil and conventional oil.
• the "Refined Oil” and “Transport” pieces of the supply chain contributed less than 2% of the total freshwater eutrophication potential.
• the conventional oil economic allocation factor with respect to meal varied from 31 % to 47% over the past ten years with an average of 39%.
• the high oleic allocation factor with respect to meal varied from 33% to 49% with an average of 41 %.
• Table 8 Economic and mass allocation factors for the co-products at the oil refining process step for both conventional and high oleic oil
• the resulting economic allocation factor for high oleic soybean oil at a price adder of 15 cents per pound is 48% with respect to soybean meal.
• the resulting economic allocation factor for refined soybean oil with respect to soapstock and distillate is 97.4%.
• the price adder could be less than the assumed 6 cents per pound. However, this would only further favor high oleic oil since the allocation factors for the oil would be lower than they are for the base case.
• Mass allocation is used by the United Soybean board in its LCA on conventional soybean oil in the United States. This method distributes the environmental burdens based on the relative mass of products in each process. Tables 1 and 2 show that the soybean oil mass allocation factor with respect to soybean meal is 19% for both conventional and high oleic oil. This alternative allocation factor reduces the overall burdens for the cases studied based on the functional unit, but does not significantly change the relative performance of high oleic oil with respect to conventional soybean oil.
• Figure 7 shows climate change potential for both cases using all three allocation methods. Mass allocation showed the lowest overall burdens due to the significant mass of co-product meal produced from the soybean. System expansion showed the highest overall burdens due to the assumptions of marginal products in the market. Figure 8 highlights similar results for terrestrial acidification. For this metric, farming impacts were more important which resulted in larger relative differences among allocation methods. However for each allocation method and all impact categories studied, high oleic oil presented 43%-48% lower impact potential than conventional oil. This is mainly attributable to the reduced oil use rate for the high oleic oil case.
• the importance of washing varies by impact category.
• the reduction in washing results in a 5%-1 1 % reduction in overall burdens for the Conventional oil 2-day wash Cycle Case with respect to the base case for conventional oil.
• the high oleic oil case provides a 43%-45% reduction in burdens for the impact categories studied as compared to the conventional base case.
• These high oleic oil benefits are marginally reduced to 38%-40% compared to the conventional oil 2-day wash cycle case.
• washing intensity may also be reduced for high oleic oil compared to conventional oil due to the reduction in polymerization during use, this study assumed washing intensity was similar for both oil types. Only the frequency of washing was varied.
• Processing associated with farming of soybeans is consistently the highest contributor to burdens for both types of oil across all impact categories.
• farming represents 65% and 80% of the total burdens. While the refining process is relatively insignificant, the crude oil production process does contribute significantly to the overall burdens for the impacts measured. While not the controlling factor, washing impacts as defined in the base case are both significant and differentiating with respect to high oleic and conventional oil.
• Example 18 where conventional canola oil is compared to high oleic canola oil.
• the LCA report prepared for the Canola Council for North America on conventional canola oil was used as the primary data source for canola farming
• the terrestrial acidification potential results for 2 days of fryer use for conventional canola oil compared to high oleic canola oil are shown in Fig .1 1 .
• the base case for high oleic canola oil was about 44% lower than the base case for conventional canola oil.

Landscapes

• Chemical & Material Sciences (AREA)
• Oil, Petroleum & Natural Gas (AREA)
• Life Sciences & Earth Sciences (AREA)
• Engineering & Computer Science (AREA)
• Food Science & Technology (AREA)
• Polymers & Plastics (AREA)
• Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
• Edible Oils And Fats (AREA)

Priority Applications (2)

Applications Claiming Priority (2)

Publications (1)

Family

ID=46724672

Family Applications (1)

Country Status (3)

Cited By (2)

Citations (35)

• 2012
  • 2012-08-15 WO PCT/US2012/051005 patent/WO2014007832A1/fr not_active Ceased
  • 2012-08-15 CA CA2878383A patent/CA2878383A1/fr not_active Abandoned
  • 2012-09-21 AR ARP120103493 patent/AR087989A1/es unknown

Patent Citations (40)

Non-Patent Citations (62)

Also Published As

Similar Documents

Legal Events