Patent Description:
Popular smoking articles, such as cigarettes, have a substantially cylindrical rod shaped structure and include a charge, roll or column of smokable material such as shredded tobacco (e.g., in cut filler form) surrounded by a paper wrapper thereby forming a so-called "tobacco rod. " Normally, a cigarette has a cylindrical filter element aligned in an end-to-end relationship with the tobacco rod. Typically, a filter element comprises plasticized cellulose acetate tow circumscribed by a paper material known as "plug wrap. " Certain cigarettes incorporate a filter element having multiple segments, and one of those segments can comprise activated charcoal particles. Typically, the filter element is attached to one end of the tobacco rod using a circumscribing wrapping material known as "tipping paper. " It also has become desirable to perforate the tipping material and plug wrap, in order to provide dilution of drawn mainstream smoke with ambient air. A cigarette is employed by a smoker by lighting one end thereof and burning the tobacco rod. The smoker then receives mainstream smoke into his/her mouth by drawing on the opposite end (e.g., the filter end) of the cigarette.

The tobacco used for cigarette manufacture is typically used in blended form. For example, certain popular tobacco blends, commonly referred to as "American blends," comprise mixtures of flue-cured tobacco, burley tobacco, and Oriental tobacco, and in many cases, certain processed tobaccos, such as reconstituted tobacco and processed tobacco stems. The precise amount of each type of tobacco within a tobacco blend used for the manufacture of a particular cigarette brand varies from brand to brand. However, for many tobacco blends, flue-cured tobacco makes up a relatively large proportion of the blend, while Oriental tobacco makes up a relatively small proportion of the blend. See, for example, <NPL>), <NPL>) and <NPL>).

Through the years, various treatment methods and additives have been proposed for altering the overall character or nature of tobacco materials utilized in tobacco products. For example, additives or treatment processes have been utilized in order to alter the chemistry or sensory properties of the tobacco material, or in the case of smokable tobacco materials, to alter the chemistry or sensory properties of mainstream smoke generated by smoking articles including the tobacco material. The sensory attributes of cigarette smoke can be enhanced by incorporating flavoring materials into various components of a cigarette. Exemplary flavoring additives include menthol and products of Maillard reactions, such as pyrazines, aminosugars, and Amadori compounds. See also, <NPL>). In some cases, treatment processes involving the use of heat can impart to the processed tobacco a desired color or visual character, desired sensory properties, or a desired physical nature or texture. Various processes for preparing flavorful and aromatic compositions for use in tobacco compositions are set forth in <CIT>; <CIT>; <CIT>;<CIT>;<CIT>; <CIT>; <CIT>; <CIT>;<CIT>; <CIT>;<CIT>; <CIT>;<CIT>; <CIT><CIT>; and <CIT>; <CIT><CIT> Additionally, examples of representative components that can be employed as so-called natural tar diluents in tobacco products are set in <CIT>.

Tobacco also may be enjoyed in a so-called "smokeless" form. Particularly popular smokeless tobacco products are employed by inserting some form of processed tobacco or tobacco-containing formulation into the mouth of the user. Various types of smokeless tobacco products are set forth in <CIT>; <CIT>; <CIT>; <CIT><CIT>; <CIT>;<CIT>; <CIT><CIT>; <CIT><CIT>;<CIT>; <CIT>; <CIT>; and <CIT> See, for example, the types of smokeless tobacco formulations, ingredients, and processing methodologies set forth in <CIT><CIT>.

One type of smokeless tobacco product is referred to as "snuff. " Representative types of moist snuff products, commonly referred to as "snus," have been manufactured in Europe, particularly in Sweden, by or through companies such as <NPL>. See also, for example, <NPL>). In addition, certain quality standards associated with snus manufacture have been assembled as a so-called GothiaTek standard. Representative smokeless tobacco products also have been marketed under the tradenames Oliver Twist by House of Oliver Twist A/S; Copenhagen, Skoal, SkoalDry, Rooster, Red Seal, Husky, and Revel by U. Smokeless Tobacco Co. ; "taboka" by Philip Morris USA; Levi Garrett, Peachy, Taylor's Pride, Kodiak, Hawken Wintergreen, Grizzly, Dental, Kentucky King, and Mammoth Cave by Conwood Company, LLC; and Camel Orbs, Camel Sticks, and Camel Strips by R. Reynolds Tobacco Company.

The sensory attributes of smokeless tobacco can also be enhanced by incorporation of certain flavoring materials. See, for example, <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT><CIT><CIT>; <CIT>, <CIT><CIT><CIT><CIT><CIT>.

Because tobacco has long been cultivated throughout the world, though full utilization of tobacco biomass has yet to be attained, there is a long-felt need for a process for preparing from tobacco, or, more generally, from any one or more portions of any one or more members of genus Nicotiana, a material useful as a flavorant, inter alia, in the manufacture of smoking articles and/or smokeless tobacco products.

<CIT> discloses a method of producing biofuel from tobacco biomass including a solvent extraction of the tobacco biomass with methyl acetate or ethyl acetate, transesterification of the oil obtained from the biomass and separation of the biofuel from the transesterified product. Excellent yields of biofuel based on the weight of the biomass are obtained.

<CIT> provides a method of extracting ester from tobacco including the steps of: extracting water-soluble material by injecting a solvent into the tobacco and removing the extracted solution; drying the tobacco firstly; extracting hydrocarbon material by injecting nonpolar solvent and removing the extracted solution; drying tobacco secondly; injecting polar solvent into tobacco, extracting and removing tobacco; injecting acid into the extracted solution and forming crystal; removing the crystal; and condensing the extracted solution.

The present invention relates to a process for producing one or more sensory-positive ester flavorants from a quantity of biomass set out in claim <NUM>.

A process such as is described in various embodiments herein provides materials from Nicotiana species (e.g., tobacco-derived materials) comprising isolated components from plants of the Nicotiana species useful for incorporation into tobacco compositions utilized in a variety of tobacco products, such as smoking articles and smokeless tobacco products, or more generally into compositions that may comprise a flavorant. A process such as is described herein also provides processes for isolating components from Nicotiana species (e.g., tobacco materials), and processes for processing those components and tobacco materials incorporating those components. For example, tobacco-derived materials can be prepared by subjecting at least a portion of a tobacco plant (e.g., leaves, stalks, roots, or stems) to a separation process, which typically can include multiple sequential extraction steps, in order to isolate desired components of the tobacco material. For example, tobacco-derived materials can be prepared by subjecting at least a portion of a tobacco plant (e.g., leaves, stalks, roots, or stems) to a separation process, which typically can include multiple sequential extraction steps, in order to isolate desired components of the tobacco material.

When used in connection with a process such as is described in various embodiments herein, the term "biomass" denotes any one or more portions of a plant, and in particular denotes substantially the entirety of the superterranean portion of a plant, optionally including some or all of the subterranean portion of a plant. Accordingly, the term "biomass" may refer to flower or to leaf or to seed or to any other superterranean portion of a plant, or to any combination thereof, optionally including some or all of the subterranean portion of a plant. Accordingly, the term "biomass" and related terms such as "biomatter" and "plant source" may be properly understood to refer to any one or more portions of a harvested plant that may be processed to extract, separate, or isolate components of interest therefrom.

When used in connection with a process such as is described in various embodiments herein, the term "one or more plants of genus Nicotiana" denotes any one or more plants of the genus Nicotiana of family Solanaceae, including, for example, any one or more of the following: N. arentsii, N. excelsior, N. forgetiana, N. glutinosa, N. kawakamii, N. knightiana, N. langsdorffi, N. otophora, N. setchelli, N. sylvestris, N. tomentosa, N. tomentosiformis, N. undulata, and N. x sanderae, N. africana, N. amplexicaulis, N. benavidesii, N. bonariensis, N. debneyi, N. longiflora, N. maritina, N. megalosiphon, N. occidentalis, N. paniculata, N. plumbaginifolia, N. raimondii, N. rosulata, N. rustica, N. simulans, N. stocktonii, N. suaveolens, N. tabacum, N. umbratica, N. velutina, and N. wigandioides, N. acaulis, N. acuminata, N. attenuata, N. benthamiana, N. cavicola, N. clevelandii, N. cordifolia, N. corymbosa, N. fragrans, N. goodspeedii, N. linearis, N. miersii, N. nudicaulis, N. obtusifolia, N. occidentalis subsp. Hersperis, N. pauciflora, N. petunioides, N. quadrivalvis, N. repanda, N. rotundifolia, N. solanifolia, N. spegazzinii.

The use of Nicotiana-derived (e.g., tobacco-derived) materials produced by a process such as is described in various embodiments herein enables the preparation of tobacco compositions for smoking articles or smokeless tobacco compositions that are derived substantially or even entirely from Nicotiana materials. For example, a tobacco composition can incorporate tobacco or tobacco-derived material of some form, including isolated components from Nicotiana species, such that at least about <NUM> weight percent, more typically at least about <NUM> weight percent, or even at least about <NUM> weight percent (on a dry weight basis), of that tobacco composition consists of tobacco-derived material.

It has long been recognized that there is a need to make fuller use of material or substance from tobacco, and in particular from plants or portions of plants from Nicotiana species. Readily available starting materials or inputs from plants or portions of plants from Nicotiana species, such starting materials or inputs being useful in particular for inclusion as starting materials or inputs in a process whereby material or substance from tobacco can be more fully utilized, include inter alia tobacco biomass. Tobacco biomass can include for example the entirety of the substance of a tobacco plant that has been harvested whole. Tobacco biomass can include for example essentially all of the superterranean parts of a tobacco plant and optionally can include some or all of the subterranean parts of a tobacco plant. Tobacco biomass can include for example the solid portion of a tobacco plant that has been harvested whole, or the solid portion of essentially all of superterranean parts of a tobacco plant, and from which so-called "green juice" has been expelled for example through the action of a screw press. Tobacco biomass can include for example such a solid portion from which at least a portion of the water has been removed by drying.

Among ways in which fuller use can be made of material or substance from tobacco, and in particular from plants or portions of plants from Nicotiana species, are various physical and/or chemical transformations to which plants or portions of plants from Nicotiana species can be subjected. Such physical and/or chemical transformations may result in outputs or products having one or more desired or favorable properties. Such outputs or products may themselves be useful as starting material or inputs for further useful processes. Among physical transformations to which plants or portions of plants from Nicotiana species can be subjected are disruptions of the physical integrity of tobacco biomass, such as a disruption resulting from the action of a screw press against a quantity of tobacco biomass. Among physical transformations to which plants or portions of plants from Nicotiana species can be subjected are fractionations according to, for example, particle size, relative density, sedimentation velocity, or affinity for a fixed matrix.

In an aspect, a process such as is described in various embodiments herein provides a material for use in a smoking article or a smokeless tobacco composition comprising an additive derived from a flower of a Nicotiana species. A material can be a flower of a Nicotiana species or a portion thereof in particulate form or in the form of a flower derivative derived from a flower of a Nicotiana species. A flower derivative may be in the form of an extract from a flower of a Nicotiana species or in the form of a chemically transformed flower derivative, exemplary chemical transformations including acid/base reaction, hydrolysis, thermal treatment, enzymatic treatment, and combinations of such steps. A chemical transformation typically results in a change in chemical composition of a tobacco derivative, such as an increase in the amount of certain compounds that have desirable sensory characteristics (e.g., aromatic or flavorful compounds). In certain embodiments, a process such as is described in various embodiments herein provides techniques adapted for expressing lipids from biomass, such as from flower or from seed, such as high pressure squeezing or cold pressing. Alternatively, a component containing tobacco oil according to a process such as is described in various embodiments herein is formed by extracting components from biomass, such as from flower or from seed, using appropriate extraction techniques and solvents. Exemplary solvents include hydrocarbons such as heptane and hexane. Other separation processes can be used, such as chromatography, distillation, filtration, recrystallization, solvent-solvent partitioning, and combinations thereof. An oil-containing component formed using an extraction process can be either the solvent-soluble portion or the insoluble residue of biomass or seed material remaining after solvent extraction. An oil-containing component formed using a pressing process may be inter alia a lipid-containing portion of biomass, such as flower or seed, expressed from pressed biomass, such as flower or seed material.

In an aspect, a flower derivative is in the form of an extract of an enzymatically-treated flower of a Nicotiana species. Exemplary extraction solvents include hydrocarbons such as heptane and hexane.

In an aspect, a process such as is described herein provides a material for use in a smoking article or a smokeless tobacco composition comprising an additive derived from one or more flowers of a Nicotiana species such as described herein. For example a process such as is described in various embodiments herein provides a material wherein an additive is in the form of a casing formulation or a top dressing formulation applied to tobacco strip or wherein an additive is added to a reconstituted tobacco material. Smoking articles or smokeless tobacco compositions incorporating a flower additive derived from a process such as is described in various embodiments herein may comprise between about <NUM> ppm and about <NUM> weight percent of flower additive based on total dry weight of tobacco material in the smoking article or smokeless tobacco product.

In an aspect, a process such as is described herein provides a method for preparing an additive derived from a flower of a Nicotiana species for addition to a tobacco composition, the method comprising: i) receiving a harvested flower or a portion thereof; ii) processing the harvested flower or portion thereof by at least one of subdividing the harvested flower or portion thereof to form a particulate flower material or separating a flower derivative from the harvested flower by subjecting the harvested flower or a portion thereof to solvent extraction, chromatography, distillation, filtration, recrystallization, solvent-solvent partitioning, or a combination thereof; and iii) adding the particulate flower material or flower derivative produced in step ii) to a tobacco composition adapted for use in a smoking article or a smokeless tobacco composition.

In an aspect, a process such as is described herein provides a method for preparing an additive derived from a flower of a Nicotiana species for addition to a tobacco composition, the method comprising separating a flower derivative from a flower of the Nicotiana species, said separating step comprising one or more of the following steps: i) collecting vapor-phase components from the headspace surrounding a living flower; and ii) isolating components of a harvested flower by subjecting the harvested flower or a portion thereof to solvent extraction, chromatography, distillation, filtration, recrystallization, solvent-solvent partitioning, or a combination thereof.

A process such as is described in various embodiments herein now will be described more fully hereinafter. As used in this specification and the claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Reference to "dry weight percent" or "dry weight basis" refers to weight on the basis of dry ingredients (i.e., all ingredients except water). When used in this specification and the claims as an adverb rather than a preposition, "about" means "approximately" and comprises the stated value and every value within <NUM>% of that value; in other words, "about <NUM>%" includes <NUM>% and <NUM>% and every value in between.

The selection of the plant from a Nicotiana species can vary; and in particular, the types of tobacco or tobaccos may vary. Tobaccos that can be employed include flue-cured or Virginia (e.g., K326), burley, sun-cured (e.g., Indian Kurnool and Oriental tobaccos, including Katerini, Prelip, Komotini, Xanthi and Yambol tobaccos), Maryland, dark, dark-fired, dark air cured (e.g., Passanda, Cubano, Jatin and Bezuki tobaccos), light air cured (e.g., North Wisconsin and Galpao tobaccos), Indian air cured, Red Russian and Rustica tobaccos, as well as various other rare or specialty tobaccos. Descriptions of various types of tobaccos, growing practices and harvesting practices are set forth in <NPL>). Various representative types of plants from the Nicotiana species are set forth in <NPL>); <CIT>;<CIT>; <CIT>; <CIT><CIT> Of particular interest are N. arentsii, N. excelsior, N. forgetiana, N. glutinosa, N. kawakamii, N. knightiana, N. langsdorffi, N. otophora, N. setchelli, N. sylvestris, N. tomentosa, N. tomentosiformis, N. undulata, and N. x sanderae. Also of interest are N. africana, N. amplexicaulis, N. benavidesii, N. bonariensis, N. debneyi, N. longiflora, N. maritina, N. megalosiphon, N. occidentalis, N. paniculata, N. plumbaginifolia, N. raimondii, N. rosulata, N. rustica, N. simulans, N. stocktonii, N. suaveolens, N. tabacum, N. umbratica, N. velutina, and N. wigandioides. Other plants from the Nicotiana species include N. acaulis, N. acuminata, N. attenuata, N. benthamiana, N. cavicola, N. clevelandii, N. cordifolia, N. corymbosa, N. fragrans, N. goodspeedii, N. linearis, N. miersii, N. nudicaulis, N. obtusifolia, N. occidentalis subsp. Hersperis, N. pauciflora, N. petunioides, N. quadrivalvis, N. repanda, N. rotundifolia, N. solanifolia and N. spegazzinii.

Nicotiana species can be derived using genetic-modification or crossbreeding techniques (e.g., tobacco plants can be genetically engineered or crossbred to increase or decrease production of certain components or to otherwise change certain characteristics or attributes). See, for example, the types of genetic modifications of plants set forth in <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT><CIT>; <CIT>; and <CIT>.

For the preparation of smokeless and smokable tobacco products, it is typical for harvested plants of a Nicotiana species to be subjected to a curing process. Descriptions of various types of curing processes for various types of tobaccos are set forth in<NPL>). Exemplary techniques and conditions for curing flue-cured tobacco are set forth in <NPL>) and <CIT>. See, also, for example, <CIT> Representative techniques and conditions for air curing tobacco are set forth in <NPL>) and <NPL>). Certain types of tobaccos can be subjected to alternative types of curing processes, such as fire curing or sun curing. Preferably, harvested tobaccos that are cured are then aged.

At least a portion of the plant of a Nicotiana species (e.g., at least a portion of the tobacco portion) can be employed in an immature form. That is, the plant, or at least one portion of that plant, can be harvested before reaching a stage normally regarded as ripe or mature. As such, for example, tobacco can be harvested when the tobacco plant is at the point of a sprout, is commencing leaf formation, is commencing seeding, is commencing flowering, or the like.

At least a portion of the plant of a Nicotiana species (e.g., at least a portion of the tobacco portion) can be employed in a mature form. That is, the plant, or at least one portion of that plant, can be harvested when that plant (or plant portion) reaches a point that is traditionally viewed as being ripe, over-ripe or mature. As such, for example, through the use of tobacco harvesting techniques conventionally employed by farmers, Oriental tobacco plants can be harvested, burley tobacco plants can be harvested, or Virginia tobacco leaves can be harvested or primed by stalk position. After harvest, a plant of a Nicotiana species, or portion thereof, can be used in a green form (e.g., tobacco can be used without being subjected to any curing process). For example, tobacco in green form can be frozen, freeze-dried, subjected to irradiation, yellowed, dried, cooked (e.g., roasted, fried or boiled), or otherwise subjected to storage or treatment for later use. Such tobacco also can be subjected to aging conditions.

In accordance with a process such as is described in various embodiments herein, a tobacco product may incorporate tobacco that is combined with some form of biomass or one or more anatomical parts, such as a flower, obtained from, or derived from, a plant of at least one Nicotiana species. That is, a portion of a tobacco product according to a process such as is described in various embodiments herein can be composed of some form of biomass or one or more anatomical parts of a Nicotiana species, such as parts or pieces of biomass or one or more anatomical parts, or processed materials incorporating processed biomass or one or more anatomical parts or components thereof, such as a flower or one or more parts thereof. At least a portion of the tobacco product can be composed of components of biomass or one or more anatomical parts, such as a flower, such as ingredients removed from biomass or one or more anatomical parts, such as a flower (e.g., by extraction, distillation, or other types of processing techniques). At least a portion of the tobacco product can be composed of components derived from biomass or one or more anatomical parts, such as a flower, such as components collected after subjecting biomass or one or more anatomical parts to chemical reaction or after subjecting components collected from biomass or one or more anatomical parts, such as a flower, to chemical reaction (e.g., acid/base reaction conditions or enzymatic treatment).

A flower is a characteristic reproductive structure (e.g., seed producing structure) of a plant of a Nicotiana species. For example, a tobacco flower is the flower characteristic of a tobacco plant. Flowers of various types of representative Nicotiana species are depicted in,<NPL>).

A Nicotiana species can be selected for the type of biomass or anatomical part that it produces. For example, plants can be selected on the basis that those plants produce relatively abundant biomass or seed, produce biomass or seed that incorporate relatively high levels of specific desired components, and the like.

A Nicotiana species of plant can be grown under agronomic conditions so as to promote development of biomass or one or more anatomical parts. Tobacco plants can be grown in greenhouses, growth chambers, or outdoors in fields, or grown hydroponically.

According to a process such as is described in various embodiments herein, biomass or one or more anatomical parts, such as a flower, are harvested from a Nicotiana species of plant. The manner by which biomass or one or more anatomical parts are harvested can vary. Typically, essentially all the biomass or anatomical parts, such as a flower, can be harvested, and employed as such.

A flower can be harvested from a Nicotiana species of plant. The manner by which a flower is harvested can vary. Harvest of flowers traditionally has been referred to as "picking " As such, a flower is removed from the rest of the plant by cutting or breaking the stem or pedicle that connects the flower from the rest of the plant. Alternatively, components of a flower can be derived by collecting vapor-phase components from the headspace in the vicinity of a living flower (i.e., a flower that has not been removed or picked from the plant), such as by capturing vapor-phase components from the headspace of a growth chamber containing a living flower.

Any one or more of various parts or portions of a flower can be employed. For example, virtually all of a flower (e.g., the whole flower) can be harvested, and employed as such. Alternatively, various parts or pieces of a flower can be harvested or separated for further use after harvest. For example, a petal, corolla, sepal, receptacle, anther, filament, stigma, stamen, style, pistil, pedicel, ovary, or any of various combinations thereof can be derived for further use or treatment.

Time of harvest during the life cycle of the plant can vary. For example, biomass or one or more anatomical parts, such as a flower, can be harvested when immature. Alternatively, biomass or one or more anatomical parts, such as a flower or a seed, can be harvested after the point that the plant has reached maturity.

With respect to a flower, time of harvest during the life cycle of the flower can vary. For example, a flower can be harvested when it is in the form of a bud, when it is closed prior to bloom, during bloom, or after bloom is complete. Timing of harvest can affect yield of certain desirable compounds derived from a flower, with harvesting late in a growing season toward the end of the plant life being less preferred.

A flower can be harvested at any of various times of day. For example, a flower can be harvested during morning hours or afternoon hours (i.e., during daylight hours), or at nighttime (i.e., when it is dark). A flower can be harvested when it is dry, or when it is wet (e.g., after being exposed to rain or irrigation).

Post-harvest processing of biomass or one or more anatomical parts, such as a flower or a seed, can vary. After harvest, the biomass or one or more anatomical parts, such as a flower or a seed, or portion thereof, can be used in the harvested form (e.g., the biomass or one or more anatomical parts, such as a flower or a seed, or portion thereof, can be used without being subjected to any curing and/or aging process steps). For example, biomass or one or more anatomical parts, such as a flower or a seed, can be used without being subjected to significant storage, handling or processing conditions. In certain situations, it is preferable that fresh biomass or one or more anatomical parts, such as a flower or a seed, be used virtually immediately after harvest. Alternatively, for example, biomass or one or more anatomical parts, such as a flower or a seed, for example, a flower in green form, can be refrigerated or frozen for later use, freeze dried, subjected to irradiation, yellowed, dried, cured (e.g., using air drying techniques or techniques that employ application of heat), heated or cooked (e.g., roasted, fried or boiled), or otherwise subjected to storage or treatment for later use.

Harvested biomass, such as a flower or a seed, can be physically processed. Biomass or one or more anatomical parts, or one or more parts thereof, can be further subdivided into parts or pieces (e.g., biomass can be comminuted, pulverized, milled or ground into pieces or parts that can be characterized as granules, particulates or fine powders, or, e.g., petals can be removed from remaining portion of a flower). Biomass or one or more anatomical parts, such as a flower or a seed, or one or more parts thereof, can be subjected to external forces or pressure (e.g., by being pressed or subjected to roll treatment). When carrying out such processing conditions, biomass or one or more anatomical parts, such as a flower or a seed, can have a moisture content that approximates its natural moisture content (e.g., its moisture content immediately upon harvest), a moisture content achieved by adding moisture to the biomass, such as a flower or a seed, or a moisture content that results from the drying of the biomass, such as a flower or a seed. For example, powdered, pulverized, ground or milled pieces of biomass or one or more anatomical parts, such as a flower or a seed, can have moisture contents of less than about <NUM> weight percent, often less than about <NUM> weight percent, and frequently less than about <NUM> weight percent. Parts or pieces of biomass or one or more anatomical parts, such as a flower or a seed, can be used as components of tobacco products without further processing, or alternatively the particulate biomass or anatomical part material can be processed further prior to incorporation into a tobacco product.

Harvested biomass or one or more anatomical parts, such as a flower or a seed, or components thereof, can be subjected to other types of processing conditions. For example, components of biomass or one or more anatomical parts, such as a flower or a seed, can be separated from one another, or otherwise fractionated into chemical classes or mixtures of individual compounds. As used herein, an "isolated biomass component," "isolated component of one or more anatomical parts," "biomass isolate," "isolate of one or more anatomical parts," or "isolate" when used as a noun is a compound or complex mixture of compounds separated from biomass or one or more anatomical parts, such as a flower or a seed, of a plant of a Nicotiana species. Accordingly, a "flower isolate" is a compound or complex mixture of compounds derived from a flower of a plant of a Nicotiana species. The isolated biomass component or isolated component of one or more anatomical parts, such as a flower or a seed, can be a single compound, a homologous mixture of similar compounds (e.g., isomers of a flavorful or aromatic compound), or a heterologous mixture of dissimilar compounds (e.g., a complex mixture of various compounds of different types, preferably having desirable sensory attributes).

Typical separation processes can include one or more process steps such as solvent extraction (e.g., using polar solvents, non-polar organic solvents, or supercritical fluids), chromatography, distillation, filtration, cold pressing or other pressure-based techniques, recrystallization, and/or solvent-solvent partitioning. Exemplary extraction and separation solvents or carriers include water, alcohols (e.g., methanol or ethanol), hydrocarbons (e.g., heptane and hexane), diethyl ether, methylene chloride and supercritical carbon dioxide. Exemplary techniques useful for extracting components from Nicotiana species are described in <CIT>; <CIT>;<CIT>; <CIT>;<CIT>; <CIT>;<CIT>; <CIT>; <CIT>;<CIT>; <CIT>;<CIT>;<CIT>; <CIT>;<CIT>; <CIT>; <CIT>; <CIT>;<CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>;<CIT>; <CIT>;<CIT>; <CIT>; <CIT>;<CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT><CIT>. See also, the types of separation techniques set forth in <NPL>) and <NPL>). In addition, the biomass or components thereof can be subjected to the types of treatments set forth in <NPL>); <NPL>); <NPL>); <NPL>); <NPL>); <NPL>); <NPL>); and <CIT> See also, for example, the types of processing techniques set forth in <NPL>); <NPL>); <NPL>); <NPL>); and <NPL>).

Any one or more components of a flower, or any one or more portions of a flower, can be isolated. As used herein, an "isolated component" or "flower isolate" is a compound or complex mixture of compounds separated from a flower of a plant of a Nicotiana species. An isolated component can be a single compound, a homologous mixture of similar compounds (e.g., isomers of a flavor compound), or a heterologous mixture of dissimilar compounds (e.g., a complex mixture of various compounds of different types, preferably having desirable sensory attributes). Likewise, any one or more components of a seed, or any one or more portions of a seed, can be isolated. As used herein, an "isolated component" or "seed isolate" is a compound or complex mixture of compounds separated from a seed of a plant of a Nicotiana species. An isolated component can be a single compound, a homologous mixture of similar compounds (e.g., isomers of a flavor compound), or a heterologous mixture of dissimilar compounds (e.g., a complex mixture of various compounds of different types, preferably having desirable sensory attributes). Accordingly, an "isolate" according to a process such as is described in various embodiments herein may be a flower isolate, a seed isolate, or, more generally, a biomass isolate.

Multiple sequential separation processes can be employed to purify and refine a flower isolate or a seed isolate in a desired manner. For example, a solvent extract of a flower or of a seed of a Nicotiana species can be subjected to additional separation steps to change the chemical composition of the extract, such as by increasing the relative amount of certain desirable compounds, such as certain flavorful or aromatic compounds. In one embodiment, a flower extract or a seed extract is processed using molecular distillation, which typically involves vacuum distillation at a pressure of less than about <NUM> Ton.

Examples of types of components that can be present in isolates include terpenes, sesqui-terpenes, diterpenes, esters (e.g., terpenoid esters and fatty acid esters), alcohols, aldehydes, ketones, carboxylic acids, lactones, anhydrides, phenols quinones, ethers, nitrites, amines, amides, imides, nitroalkanes, nitrophenols, nitroarenes, nitrogen-containing heterocyclics, lactams, oxazoles, aza-arenes, sulfur-containing compounds, alkaloids (e.g., nicotine), plastid pigments (e.g., chlorophylls or carotenoids), lipids (e.g., phytosterols), and derivatives thereof. Additional examples of representative components that can be employed are described as natural tar diluents in <CIT>.

Any one or more components of a flower or a seed can be subjected to conditions so as to cause those components (whether as part of the flower or of the seed or in the form of an isolated component) to undergo chemical transformation. For example, flower isolates that have been separated from the flower can be treated to cause chemical transformation or be admixed with other ingredients. The chemical transformations or modification of the flower isolate can result in changes of certain chemical and physical properties of those flower isolates (e.g., the sensory attributes of those isolates). For example, seed isolates that have been separated from the seed can be treated to cause chemical transformation or be admixed with other ingredients. The chemical transformations or modification of the seed isolate can result in changes of certain chemical and physical properties of those seed isolates (e.g., the sensory attributes of those isolates). Exemplary chemical modification processes can be carried out by acid/base reaction, hydrolysis, heating (e.g., a thermal treatment where the flower isolate is subjected to an elevated temperature such as a temperature of at least about <NUM> degrees Celsius, or at least about <NUM> degrees Celsius, or at least about <NUM> degrees Celsius), and enzymatic treatments (e.g., using glycosidase or glucocidase); and as such, components of the flower isolate can undergo esterification, transesterification, isomeric conversion, acetal formation, acetal decomposition, invert sugar reactions, and the like. Exemplary types of further ingredients that can be admixed with the isolates include flavorants, fillers, binders, pH adjusters, buffering agents, colorants, disintegration aids, antioxidants, humectants and preservatives.

Flowers and components of flower isolates are useful as additives for tobacco compositions, particularly tobacco compositions incorporated into smoking articles or smokeless tobacco products. Addition of one or more flower isolates to a tobacco composition can enhance a tobacco composition in a variety of ways, depending on the nature of the flower isolates and the type of tobacco composition. Exemplary flower isolates can serve to provide flavor and/or aroma to a tobacco product (e.g., composition that alters the sensory characteristics of tobacco compositions or smoke derived therefrom). Likewise, components of seed isolates are useful as additives for tobacco compositions, particularly tobacco compositions incorporated into smoking articles or smokeless tobacco products. Addition of one or more seed isolates to a tobacco composition can enhance a tobacco composition in a variety of ways, depending on the nature of the seed isolates and the type of tobacco composition. Exemplary seed isolates can serve to provide flavor and/or aroma to a tobacco product (e.g., composition that alters the sensory characteristics of tobacco compositions or smoke derived therefrom).

A variety of compounds having distinctive flavor and aroma characteristics can be isolated from flowers or seeds or, more generally, from biomass of plants of Nicotiana species. Certain of those compounds can be considered to be volatile under normal ambient conditions of temperature, humidity and air pressure. Preferred compounds exhibit positive sensory attributes at relatively low concentrations. For example, a suitable flower can provide compounds such as <NUM>-ketosiophorone, phytol, phenethyl alcohol, benzyl alcohol, linalool, various cembrenol isomers, various cembrenediols, isophorone, methylbenzoate, salicylaldehyde, benzylsalicylate, methoxy eugenol, thunbergol, various carboxylic acids, various oximes, benzaldehyde, benzylbenzoate, scaral, acetophenone, caryophyllene, cinnamaldehyde, cinnamyl alcohol, various cyclohexene-butanone isomers, solavetivone, farnesal, farnesol, and the like. Additional exemplary compounds include <NUM>,<NUM>-cineole, cis-<NUM>-hexen- <NUM>-ol, methylsalicylate, b-ionone, acetovanillone, b-damascone, b-damascenone, dihydroactinidiolide, vanillylacetone, sclareolide, sclareol, cis-abienol, cembrene isomers, cembratriene diol isomers (e.g.,. -cembratriendiol,. -cembratrienediol), megastigmatrienones, norsolanadione, solanone, caryophyllene oxide, ionol derivatives, and the like. Each of those types of compounds can be isolated in relatively pure form. See, for example, <NPL>) and <NPL>). In addition, compounds having distinctive flavor and aroma characteristics can be chemically bound, such as in the form of glycosidically bound compounds. Many different compounds of interest can be present in tobacco flowers in a glycoside form, such as benzaldehyde, benzyl alcohol, phenethyl alcohol, ethyl acetophenone, <NUM>-ketoisopherone, benzyl acetate, <NUM>,<NUM>-cineol, linalool, geraniol, eugenol, nerolidol, cembrenediols, terpineol, megastigmatrienones, and other compounds noted herein. See, for example, <NPL>); <NPL>); <NPL>); <NPL>); and <NPL>).

The form of an isolate can vary. Typically, an isolate is in a solid, liquid, or semi-solid or gel form. An isolate can be used in concrete, absolute, or neat form. Solid forms of an isolate include spray-dried and freeze-dried forms. Liquid forms of an isolate include isolates contained within aqueous or organic solvent carriers.

A flower, a processed flower or a flower isolate, or a seed, a processed seed or a seed isolate, can be employed in any of a variety of forms. A harvested flower or flower isolate or harvested seed or seed isolate can be employed as a component of processed tobaccos. In one regard, a flower, or any one or more components thereof, or a seed, or any one or more components thereof, can be employed within a casing formulation for application to tobacco strip (e.g., using the types of manners and methods set forth in <CIT>) or within a top dressing formulation. Alternatively, a flower, or any one or more components thereof, or a seed, or any one or more components thereof, can be employed as an ingredient of a reconstituted tobacco material (e.g., using the types of tobacco reconstitution processes generally set forth in <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; and <CIT>). A flower, or any one or more components thereof, or a seed, or any one or more components thereof, also can be incorporated into a cigarette filter (e.g., in the filter plug, plug wrap, or tipping paper) or incorporated into cigarette wrapping paper, preferably on the inside surface, during the cigarette manufacturing process.

A flower, processed flower or flower isolate, or a seed, processed seed or seed isolate, can be incorporated into smoking articles. Representative tobacco blends, non-tobacco components, and representative cigarettes manufactured therefrom, are set forth in <CIT>; <CIT>; <CIT>; <CIT>; and <CIT>; <CIT>; and <CIT>. Those tobacco materials also can be employed for the manufacture of those types of cigarettes that are described in <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; and <CIT>; <CIT>; and <CIT> and <CIT>. See, also, those types of commercially marketed cigarettes described <NPL>) and <NPL>).

A flower, processed flower or flower isolate, or a seed, processed seek or seed isolate, can be incorporated into smokeless tobacco products, such as loose moist snuff, loose dry snuff, chewing tobacco, pelletized tobacco pieces (e.g., having the shapes of pills, tablets, spheres, coins, beads, obloids or beans), extruded or formed tobacco strips, pieces, rods, cylinders or sticks, finely divided ground powders, finely divided or milled agglomerates of powdered pieces and components, flake-like pieces, molded processed tobacco pieces, pieces of tobacco-containing gum, rolls of tape-like films, readily water-dissolvable or water-dispersible films or strips (e.g., <CIT>), or capsule-like materials possessing an outer shell (e.g., a pliable or hard outer shell that can be clear, colorless, translucent or highly colored in nature) and an inner region possessing tobacco or tobacco flavor (e.g., a Newtonian fluid or a thixotropic fluid incorporating tobacco of some form). Various types of smokeless tobacco products are set forth in <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; <CIT>; and <CIT>; <CIT> and <CIT>; <CIT>; <CIT>; <CIT>; and <CIT> See also, the types of smokeless tobacco formulations, ingredients, and processing methodologies set forth in <CIT> and <CIT>; <CIT>; <CIT>; <CIT><CIT>, <CIT><CIT><CIT><CIT><CIT><CIT><CIT><CIT><CIT><CIT>.

An amount of a flower or a flower isolate, or of a seed or a seed isolate, added to a tobacco composition, or otherwise incorporated within a tobacco composition or tobacco product, can depend on the desired function of that flower or seed component, the chemical makeup of that component, and the type of tobacco composition to which the flower or seed component is added. The amount added to a tobacco composition can vary, but will typically not exceed about <NUM> weight percent based on the total dry weight of the tobacco composition to which the flower or flower isolate or seed or seed isolate is added. When the flower is employed within a smoking article, the amount of flower will typically be at least about <NUM> ppm, generally at least about <NUM> ppm, and often at least about <NUM> ppm, based on the total dry weight of the tobacco material within the smoking article; but will typically be less than about <NUM> percent, generally less than <NUM> percent, and often less than about <NUM> percent, based on the total dry weight of the tobacco material within the smoking article. When the flower is employed within a smokeless tobacco product, the amount of flower will typically be less at least about <NUM> ppm, generally at least about <NUM> ppm, and often at least about <NUM> ppm, based on the total dry weight of the tobacco material within the smokeless tobacco product; but will typically be less than about <NUM> percent, generally less than <NUM> percent, and often less than about <NUM> percent, based on the total dry weight of the tobacco material within the smokeless tobacco product.

Aspects of a process such as is described herein are further illustrated by the following examples, which are set forth to illustrate certain aspects of a process such as is described in various embodiments herein and are not to be construed as limiting thereof.

A flower absolute of Nicotiana alata contains a large quantity of octanoic acid (approximately <NUM>% isolated yield) along with other C<NUM> to C<NUM> acids in smaller percentages. These compounds are sensory neutral or sensory negative. Through esterification these compounds were transformed to sensory positive compounds.

a process was developed to synthesize esters of the aforementioned naturally occurring acids isolated from a N. alata flower absolute. The process was scaled-up to yield quantities of purified product.

Nicotiana flowers are, according to a process such as is descried in various embodiments herein, a source of compounds with positive sensory characteristics. Flash chromatography to separate the flower absolutes of N. sylvestris, N. suaveolens, and N. In the case of N. alata, the major isolated constituent was octanoic acid with trace quantities of other C<NUM>-C<NUM> acids. These compounds are sensory neutral or sensory negative (shorter chain acids have cheesy, sweaty socks aroma while C<NUM> and larger have no aroma). In contrast, the ethyl esters of these acids have very positive sensory characteristics: fruity pineapple, strawberry, apple, banana, coconut, wine, cognac, rum. Furthermore, these esters are very powerful with odor thresholds as low as <NUM> part per billion.

Initial studies dealt with screening reaction conditions to determine the optimal parameters for synthesis of ethyl esters (not according to the invention). Optimization was guided by conversion of octanoic acid to ethyl octanoate and reaction time.

As evident in Table <NUM>, favorable results were obtained in trial D with approximately <NUM> equivalents of concentrated sulfuric acid, <NUM> equivalents of absolute ethanol, and no molecular sieves for water scavenging.

A subsequent objective was to synthesize a mixture of ethyl esters in a quantity large enough for sensory evaluation. To accomplish this, the starting material acid mixture (<NUM>, <NUM> mmol) was added to a <NUM>-L round bottom flask equipped with a magnetic stir bar and dissolved in absolute ethanol (<NUM>, <NUM> mol). After dissolution, concentrated sulfuric acid (<NUM>, <NUM> mmol) was added to the reaction mixture. The flask was then fitted with a condenser and heated to reflux. After <NUM> hours an aliquot of the reaction mixture was analyzed by GC-MS and determined to be completely converted to the ethyl esters. The reaction mixture was cooled to ambient temperature and concentrated using a Rocket evaporator to remove a majority of the ethanol (down to <NUM> volume). This concentrate was then poured into a <NUM>-L separatory funnel and diluted with methyl-tert-butyl ether (<NUM>). This organic layer was then washed once with a saturated sodium bicarbonate solution (<NUM>) and four times with deionized water (<NUM> x <NUM>). After the final wash the aqueous solution was observed to be neutralized (pH <NUM>), indicating removal of the sulfuric acid catalyst. The organic layer was then dried over anhydrous sodium sulfate and concentrated using a Rocket evaporator.

Crude product (<NUM>) was purified using an Interchim PuriFlash <NUM> flash chromatography system. This method employed a silica gel column (<NUM>, <NUM> particle size) and a hexane/ethyl acetate elution gradient. Fractions that were enriched in ethyl esters (as determined by GC-MS analysis) were then combined and concentrated using the Rocket evaporator to yield a pale yellow oil (<NUM>, <NUM>% yield). A GC-MS chromatogram of purified ethyl ester material yielded the data shown in <FIG>.

A process such as is described in various embodiments herein was further employed to synthesize corresponding isopropyl and isoamyl esters in scaled-up quantity. Esterifications with other alcohols were performed to demonstrate scope of process and to produce other unique sensory positive materials. As seen in Table <NUM>, isopropyl and isoamyl esters of tobacco-derived material were produced by a process such as is described in various embodiments herein.

A GC-MS chromatogram of purified ethyl isopropyl ester material yielded the data shown in <FIG>. The GC-MS chromatogram of purified isoamyl ester material yielded the data shown in <FIG>.

Flash chromatography on a silica gel column was employed to prepare a mixture of acids such as was used in the examples above from an absolute of a Nicotiana species. According to such a process, hexane/ethyl acetate solvent gradient facilitated separation of cembratriendiols from target short- to medium-chain aliphatic acids. Such a process yielded successful preparation for N. suaveoloens and N. sylvestris. Flowers were extracted with hexanes at ambient temperature and concentrated to produce a flower concrete. Each concrete was dissolved in a minimal quantity of ethanol and precipitated to precipitate a corresponding wax. Each remaining solution was vacuum filtered to produce a flower absolute. On average, flower absolute constituted <NUM>% of wet flower mass.

Various catalyses were undertaken to effectuate trans-esterification of tobacco oil triglycerides with ethanol to form fatty acid ethyl esters. For example, trans-esterification of tobacco seed oil triglycerides with boron trifluoride in the presence and absence of NaOH was undertaken. To <NUM> of oil in a small vial was added <NUM> of <NUM> NaOH. The vial was purged with N<NUM>, capped, and heated for <NUM> minutes at <NUM>. The resulting mixture was then cooled and <NUM> of <NUM>% BF<NUM> in ethanol was added to the solution. The vial was again purged with N<NUM>, capped, and heated for an additional <NUM> minutes at <NUM>. Next, the sample was cooled, and most of the ethanol was removed under vacuum. The mixture of fatty acid ethyl ester products was extracted. There was substantial conversion of triglyceride to corresponding fatty acid ethyl ester.

In like manner, sodium ethoxide / boron trifluoride catalysis was undertaken. <NUM> of either <NUM> or <NUM> NaOEt in ethanol was used with <NUM> of tobacco seed oil. The solution was purged with N<NUM>, capped, and heated at <NUM> for <NUM> minutes. Samples were cooled and a volume (<NUM>, <NUM>, or <NUM>) of <NUM>% BF<NUM> in ethanol was added to the reaction vessel. In addition to studying the effect of NaOEt on the reaction, three other experiments were performed to determine if a higher concentration of base and/or a higher volume of BF<NUM> would provide a more efficient reaction. Varying the concentration of NaOEt did not have a major effect on conversion of either the C<NUM> triglyceride or the various triglycerides in the tobacco seed oil. An increase in the volume of BF<NUM> from <NUM> to <NUM>, however, increased reaction yield. This catalytic method was found to be highly sensitive to trace amounts of moisture. Acceptable results were obtained when only boron trifluoride and not sodium ethoxide was used as catalyst.

Base catalysts such as sodium carbonate, potassium carbonate, sodium hydroxide, and sodium ethoxide were tested for trans-esterification of tobacco seed oil. However, none of these catalysts showed reaction yields greater than <NUM>-<NUM>%. These results seemed to contradict reports in the literature, wherein <NUM>% conversion of triglycerides to ethyl esters was observed. See "<NPL>); "<NPL>). The cited literature emphasized that the reaction must be completed under anhydrous and anaerobic conditions. It is, therefore, possible that some of the poor recoveries were due to either wet tobacco seed oil or the presence of air in the reaction chamber. It was concluded that a trans-esterification reaction which exhibited no notable sensitivity to the presence of moisture could have a distinct advantage. The presence of moisture, however, would be very difficult to control on an industrial production scale.

With respect to acid catalysis, various concentrations of H<NUM>SO<NUM> in ethanol at different temperatures (<NUM>° and <NUM>) and different reaction times (<NUM>, <NUM>, <NUM>, and <NUM> hours) were tested. In order to achieve optimized reaction conditions, approximately <NUM> of oil was trans-esterified with <NUM> of ethanol containing <NUM>, <NUM>, or <NUM>% H<NUM>SO<NUM>. The triglyceride internal standard (<NUM> of glyceryl C<NUM>) was initially added to each reaction mixture. After each trans-esterification, GC/FID was used to estimate the percent conversion of the internal standard to the C<NUM> fatty acid ethyl ester. Subsequently, trans-esterification efficiency was determined via both gravimetry and GC/FID analysis. An object was to achieve high purity of fatty acid ethyl ester product. After each reaction, residual ethanol was removed under vacuum and the resulting mixture was washed with <NUM> of saturated NaCl solution. The vacuum-dried mixture of fatty acid ethyl esters was extracted with <NUM> x <NUM> of hexane. Next, the hexane containing fatty acid ethyl esters was dried over sodium sulfate, and the hexane was evaporated completely. The combined weight of fatty acid ethyl ester was obtained, then combined fatty acid ethyl esters were dissolved in <NUM> of dichloromethane and individually analyzed via GC/FID. For example, sn <NUM>% conversion for glyceryl C<NUM> to the corresponding fatty acid ethyl ester was obtained using <NUM>% H<NUM>SO<NUM> in ethanol at <NUM> for <NUM> hours.

In order to document trans-esterification of internal standard, three samples were trans-esterified as follows: <NUM>% H<NUM>SO<NUM> in ethanol at <NUM> for <NUM> hours. Recovery was as much as about <NUM> percent. <FIG> shows GC/FID of: (A) tobacco seed oil spiked with the glyceryl C<NUM> internal standard (<NUM>) after trans-esterification of the mixture; (B) reaction product of tobacco seed oil trans-esterified then spiked with C<NUM> fatty acid ethyl ester (<NUM>) which would be the same quantity as expected after trans-esterification of the internal standard. The C<NUM> fatty acid ethyl ester peak area for both chromatograms showed a similar area count. This experiment showed that the internal standard triglyceride conversion to C<NUM> fatty acid ethyl ester under these conditions was complete and no analyte was being lost during product work-up.

Three grams of tobacco seed oil were trans-esterified employing the above conditions employing H<NUM>SO<NUM> catalyst. Reactions were carried out in triplicate. A similar process was applied to <NUM> grams of the internal standard. For each reaction, <NUM> of <NUM>% H<NUM>SO<NUM> in ethanol was added. Each mixture was refluxed at <NUM> for <NUM> hours. After reaction was complete, most of the ethanol was removed via vacuum distillation followed by addition of <NUM>-<NUM> of saturated NaCl solution. Each sample was then extracted with 3x20 mL of hexane. The combined hexane solutions from each sample were next dried by passing them though sodium sulfate followed by evaporation of the hexane using vacuum distillation. The actual total weights of FAEE from both tobacco seed oil and the internal standard were obtained via gravimetry. GC/FID was also used to obtain the exact weight of each FAEE. Table <NUM> shows (a) the starting weight of oil or tri-undecanoin internal standard used, (b) the expected weight of FAEE obtained, (c) the combined weights of FAEE's via gravimetry and individual weights of FAEE via GC/FID.

Gravimetric analysis accordingly showed a recovery of <NUM>-<NUM>% of fatty acid ethyl esters. At the same time, GC/FID analysis of the same fatty acid ethyl esters showed only a recovery of <NUM>-<NUM>%. A high temperature GC/FID analysis by an independent laboratory (Medallion Labs, Minneapolis, MN) showed mostly the presence of fatty acid ethyl esters and less than <NUM>-<NUM>% of triglyceride. As shown in <FIG>, the GC/FID analysis of trans-esterified internal standard showed only the presence of C<NUM> fatty acid ethyl esters.

As shown in <FIG>, <NUM>C NMR of the fatty acid ethyl esters revealed: <NUM>) one carbonyl signal at ~<NUM> ppm, consistent with the presence of one structure, <NUM>) three signals around <NUM> ppm consistent with the alkene carbons of the long chain fatty acids, <NUM>) one signal at ~<NUM> ppm consistent with one type of C-O linkage, that is the α-carbon of the ethyl group, <NUM>) a group of signals between <NUM> and <NUM> ppm consistent with alkyl carbons of the long chain fatty acid groups.

As shown in <FIG>, proton NMR of the fatty acid ethyl esters revealed: <NUM>) a signal at <NUM> ppm consistent with a proton attached to an unsaturated carbon, <NUM>) a signal at <NUM> ppm consistent with protons attached to aliphatic carbons, and <NUM>) signals around <NUM> ppm, consistent with protons attached to the glycerin backbone.

As shown in <FIG>, <NUM>C NMR of tobacco seed oil revealed: <NUM>) three carbonyl signals at ~<NUM>-<NUM> ppm consistent with the presence of three carbonyl groups, although only two signals would have been predicted, <NUM>) three signals at ~<NUM> ppm consistent with the alkene carbons present in the alkyl side chains, <NUM>) multiple signals of varying intensity between <NUM>-<NUM> ppm consistent with carbons attached to oxygen, although only two signals would have been predicted, and <NUM>) multiple signals between <NUM>-<NUM> ppm consistent with alkyl carbons of the long chain fatty acid groups. An interpretation of these signals could be assigned to the presence of relatively small amounts of mono and diglycerides in the tobacco seed oil.

As shown in <FIG>, proton NMR of the tobacco seed oil revealed: <NUM>) a signal at <NUM> ppm consistent with a proton attached to an unsaturated carbon, <NUM>) signal at <NUM> ppm consistent with protons attached to aliphatic carbons, and <NUM>) signals around <NUM> ppm consistent with protons attached to the glycerin backbone.

Accordingly, the signals present in the trans-esterified reaction product are consistent with those of an ethyl ester of long chain unsaturated fatty acids. No other signals were present that would have suggested the presence of another structure.

Claim 1:
A process for producing one or more sensory-positive ester flavorants from a quantity of biomass of a plant of a Nicotiana species, the process comprising:
(a) contacting the quantity of biomass with a quantity of hexanes to form a first mixture;
(b) collecting a liquid phase of the first mixture to from a collected liquid;
(c) concentrating the collected liquid to form a concrete;
(d) dissolving the concrete in a minimal quantity of an alcohol solvent to form a reconstituted liquid;
(e) precipitating the reconstituted liquid to form a precipitated wax and an unprecipitated mixture;
(f) collecting the unprecipitated mixture;
(g) contacting the unprecipitated mixture with a quantity of an acidified alcohol mixture to form a reaction mixture for a period of time sufficient for the formation of ester linkages between one or more components of the unprecipitated mixture and one or more components of the acidified alcohol mixture, said acidified alcohol mixture comprising isopropanol and/or isoamyl alcohol;
thereby producing one or more sensory-positive ester flavorants.