WO2013122890A1

WO2013122890A1 - System and method of bioavailability enhancement of plant phytonutrients involving shearing to alter stereochemical structure

Info

Publication number: WO2013122890A1
Application number: PCT/US2013/025631
Authority: WO
Inventors: Roger Duffield; Thomas Matula
Original assignee: Moor Holdings Limited
Priority date: 2012-02-13
Filing date: 2013-02-11
Publication date: 2013-08-22

Abstract

The disclosure relates to a method of altering the cis/trans ratio of phytonutrients from live plant materials, including green tea, tomatoes, and citrus fruits involving shearing. The process may be carried out at temperatures between 80°C and 95°C.

Description

SYSTEM AND METHOD OF BIOAVAILABILITY ENHANCEMENT OF PLANT PHYTONUTRIENTS INVOLVING SHEARING TO ALTER STEREOCHEMICAL STRUCTURE

CROSS-REFERENCE TO RELATED APPLICATION

This application claims priority to United States Application Serial No. 61 /598,261 filed February 13, 2012.

Aspects relate at least in part to a method of increasing bioavailability of phytonutrients from live plant materials. Other aspects relate at least in part to compositions of emulsions and resultant slurry produced.

Bioavailability of a substance relates to the amounts of the substance absorbed into the bloodstream and therefore to the availability of the substance for use by target tissues and cells in the body.

According to the Physician's Desk Reference, the absorption ratio of nutrients formulated into oral supplements as pills, tablets or soft gels varies between 1 -30%. Therefore, the real challenge is to overcome this problem of poor absorption and to efficiently transport and release the nutrients into the cells in order for the cells to do their healing work.

An objective of aspects of this process is to increase the bioavailability of phytonutrients from live plant materials.

Provided in one or more implementations at least in part is a method of processing live plant material containing isomeric phytonutrients, wherein the material is subjected to realignment to alter the stereochemical structure of the isomeric phytonutrients by changing frans-isomers into the cis form, and/or changing c/s-isomers into the trans form.

As provided in one or more implementations of this method, phytonutrients can be selected from a group consisting of polyenes having about 95% trans- structure and flavonoids having about 95% c/s-structure.

As further provided in one or more implementations, the polyenes can comprise at least one of carotenoids, lycopenes, catechins, bioflavonoids, limonoids.

Also as provided in one or more implementations of the process, a

composition produced contains at least in part isomeric phytonutrients from live plant material at a ratio of 20/80 to 80/20 cis/trans isomers; a ratio of 30/70 to 70/30 cis/trans isomers; or a ratio of 50/50 cis/trans isomers.

In aspects of one or more implementations of the process, shearing can be achieved by treating the live plant material in a High Shear Processor comprising a stator (42) having an inlet opening (43), a coaxial cylinder with teeth (47) defined by cuts (49), therein, and a rotor (44) which is made as a disk with blades (51 ) defined by cuts (52) in the cylinder and is brought in rotation with the help of a shaft (45), characterized in that installed additionally on the rotor (44), is an impeller comprised of straight or curved blades (50), and the stator (42), additionally has an outer concentric row of straightening blades (48) defined by the cuts (49) in the outer coaxial cylinder which encompasses the rotor (44) from the outside, the width of the radial cuts (49) between the straightening blades of the stator being at least two times smaller than their length for stabilizing the mechano-hydrodynamic effect produced on the product being processed; wherein the radial clearance between the teeth (47) of the stator (42) and the blades (51 ) of the rotor (44) does not exceed 0.2 mm.

In aspects of one or more implementations of the process, shearing can be carried out at a temperature of between 80^QC and 95^QC, and for a period of not less than 3 minutes, and the live plant material is comminuted to an average size 100- 120 μιτι and slurried in water prior to processed by high shear processing; the material can be slurried in water at a ratio of 10-50% live plant material and 90-50% water, by mass.

One or more implementations of the methods provided can be applied to live plant material including fruit, vegetable, legume, and/or cereal, wherein the plant material can include green tea of the plant Camellia senesis, tomatoes, or citrus fruits.

One or more implementations provided can include a method of changing the cis/trans ratio of catechins wherein the catechins are classified as c/^'s-type and trans- type from the configuration of the two hydrogen's at the 2 and 3 positions on the C- ring and are mainly in the c/^'s-type isomeric form and wherein, after processing, catechins in the process composition are raised to a ratio of about 20/80 to 80/20 cis/trans isomers; to a ratio of about 30/70 to 70/30 cis/trans isomers; or to a ratio of about 50/50 ratio cis/trans isomers. Also one or more implementations provided can include a method of changing the cis/trans ratio in live plant material including citrus fruit containing c/^'s-type and frans-type aglycones and glucosides wherein the aglycones and glucosides are classified into c/^'s-type and frans-type from the configuration of the two hydrogen's at the 2 and 3 positions on the C-ring and are mainly in the c/^'s-type isomeric form and wherein, after processing, aglycones and glucosides in the composition are raised to a ratio of 20/80 to 80/20 cis/trans isomers; to a ratio of 30/70 to 70/30 cis/trans isomers; or to a ratio of about 50/50 cis/trans isomers.

Further one or more implementations provided can include a method of changing the cis/trans ratio in live plant material consisting of tomatoes containing lycopene, phytoene and phytofluene isomers which are mainly in the frans-type isomeric form, wherein, after processing, the lycopene, phytoene and phytofluene isomers in the composition are raised to a ratio of 20/80 to 80/20 cis/trans isomers, to a ratio of 30/70 to 70/30 cis/trans isomers, or to a ratio of 20 about 50/50 cis/trans isomers.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is a partly cut-away drawing of a processing mechanism

according to the present implementation process.

Figure 2 is a partial section of a High Shear Processor from the

Processing mechanism illustrated in Figure 1 .

Figure 3 shows a section taken along ll-ll in Figure 2.

Figure 4 is a flow diagram for a process according to the implementation process for treating plant material.

Figure 5 shows cis and trans orientation of 1 ,2-dichlorocyclohexane.

Figure 6 shows the effect of cis versus trans orientation on the position of the furan ring. Figure 7 shows the structure of limonin and nomilin. DETAILED DESCRIPTION

One or more implementations of one or more disclosed processes relate to one or more methods for improving bioavailability of released and stable

phytonutrients in microorganisms, cells and tissues that are produced from live plant materials. A goal of increased bioavailability can be to improve one or more pharmacological mechanisms of action of the phytonutrients. Implementations can cause one or more alterations of the stereochemical structure of the phytonutrients. Without being bound by a specific mechanism, at least one or more results could suggest that alteration of the stereochemical structure of phytonutrients could increase affinity with physiochemical molecular interactions of cell membranes by hydrogen bonding to the lipid bilayers.

Plant phytonutrients. Phenolic compounds occurring commonly in foods may be classified into simple phenols, hydroxybenzoic and hydrocinnamic acid

derivatives, flavonoids, stibenes, lignans and hydrolysable as well as condensed tannins. Phenolics in foods may occur in the free, esterified, etherified and insoluble- bound forms.

Abundant phenolic compounds in food are flavonoids. Flavonoids are present in at least some edible fruits, leafy vegetables, roots, tuber bulbs, herbs, spices, legumes, tea, coffee, cocoa, chocolate and red wine. They can be classified into seven groups: flavones, flavanones, flavonols, flavanonols, isoflavones, flavanols

(catechins) and anthocyanidins. In general, the leaves, flowers and fruits or plants themselves can contain flavonoid glucosides, woody tissues contain aglycones, and seeds may contain both glycosides.

Polyenes are poly-unsaturated organic compounds that contain one or more sequences of alternating double and single carbon-carbon bonds. These double carbon-carbon bonds interact in a process known as conjugation, which results in an overall lower energy state of the molecule.

Carotenoids and flavonoids (polyphenols) both belong to the natural polyenes. Members of the polyphenol group found in green tea are referred to as catechins. The carotenoid molecule is usually in the all frans-structure, whereas in polyphenols, the reverse occurs, where the catechin molecule is usually the all c/s-structure (95%). In both cases absorption is extremely low and the molecules can be oxidized by air quickly, affecting the biological properties, such as but not limited to antitumorigenic activities.

The polyene backbone consists of a pattern of conjugated double bonds, which allows the carotenoids to take up excess energy from other molecules. This characteristic may be responsible for the antioxidant activity seen in biological carotenoids. In addition to scavenging free radicals, other health benefits related to this observed antioxidative activity include protection from sunburn and inhibition of the development of certain types of cancers.

A cis isomer configuration means that adjacent hydrogen atoms are on the same side of the double bond. The rigidity of the double bond freezes its

conformation and, in the case of the cis isomer, causes the chain to bend and restricts the conformational freedom of the fatty acid. The more double bonds the chain has in the cis configuration, the less flexibility it has. When a chain has many cis bonds, it becomes quite curved in its most accessible conformations.

A trans isomer configuration, by contrast, means that the next two hydrogen atoms are bound to opposite sides of the double bond. As a result, they do not cause the chain to bend as much, and their shape is similar to straight saturated fatty acids.

In citrus fruits, bioflavonoids and limonoids found in the peels and seeds are expressed as aglycones or glucosides, with the difference being that the glucosides are attached to a sugar molecule. Usually the sugar is attached to the molecule at the seven position and is a disaccharide composed of glucose and rhamnose. It is known from published USDA scientific literature that aglycones are c/s-isomers and glucosides are trans-isomers. In the molecular structure there are double hydrogen bonds at both of or either/or of the C and D Rings and with different physical properties.

Aspects of one or more processes disclosed can relate to the conversion of the frans-isomers to c/s-forms or vice versa, of nutrients from live plant material to produce a mixture, preferably a 50/50 mixture of cis/trans isomers which leads to an increase in the bioavailability of these nutrients. This conversion is achieved through realigning of the C-Ring bond between carbon atoms; this enables the free rotation of these carbon atoms carrying substituent (an atom or group of atoms substituted in place of hydrogen on the parent chain of a hydrocarbon) around the axis represented by the C-Ring bond, and the alteration of the stereochemical structure of the compounds.

Related information about the role and importance of plant micronutrients in human health and illness is found in, for example, Hu, M., Molecular

Pharmaceuticals 4: 803-806 (2007); Fan Jiang et ai, Clinical Immunology 137: 347 - 356 (2010);

Stahl W., et ai, Arch. Biochem. Biophys. 294:173-177 (1992); Parker R. S., J. Nutr. 1 19:101 -104 (1989); and Parker R. S., FASEB J 10:542-551 (1996).

Apparatus and method of processing. One or more implementations regard disclosed alteration of the stereochemical structure of one or more compounds through hydrodynamics as as achieved by processing live plant materials using, for example, a processing mechanism comprising a High Shear Processor described in US 6,783,271 , with improvements as described herein.

This processing mechanism is indicated generally by the numeral 10 in Figure 1 . The mechanism 10 comprises a table frame 12 having legs 14 to which a support plate 16 is mounted. The support plate 16 supports a mixer component 18 and a processor component 20. The mixer component comprises a mixer motor 22 which drives two mixer blades 24 located within a reaction tank 26. The processor component 20 comprises a processor motor 28 which drives a High Shear Processor 30 located within the reaction tank 26. A funnel 32 is provided for introducing plant material into the reaction tank 26, and a valve outlet 34 is provided for removing processed plant material from the reaction tank 26.

With reference to Figures 2 and 3, the High Shear Processor comprises a stationary casing 40, a stator 42 having a central inlet opening 43 facing downwards and a rotor 44 set on a shaft 45 (which is driven by the mixer motor 28 shown in Figure 1 ), and secured on said shaft 45 with a nut 46.

The stator 42 is secured on the stationary casing 40 and has co-axial rows of teeth 47 and straightening blades 48. The teeth 47 and blades 48 of the stator 42 are defined by radial cuts 49 in cylinders; the width of the cuts between the

straightening blades 48 should be at least two times smaller than their length. The teeth 47 of the stator 42 have relieving along the inner surface at an angle of 15^Q to a tangent to the cylinder. The rotor 44 is a disc on which there are an impeller comprised of straight or curved blades 50 stored at an angel beta of not over 90^Q to the radius and a coaxial row of blades 51 . The blades are defined by radial cuts 52 in the cylinder. The cuts are made at angle delta not exceeding 60^Q. The impeller and teeth of the rotor may be made detachable.

In accordance with an improvement of the present implementation process, the radial clearance between the teeth 47 and stator 42 and the blades 51 of the rotor 44 does not exceed simultaneously 0.2 mm and 10% of the minimum width of the cuts 49 and 52; the clearances between the teeth 47 and stator 42 and the impeller blades 50 of the rotor 44 do not exceed 2/3 of the minimum width of the radial cuts 49 and 52. The teeth 47 of the stator and blades 51 of the rotor 44 are made such that as the rotor rotates, the radial flow of the medium should periodically be interrupted, thereby a variable sonic frequency pressure being set up in the medium. The number of teeth between the stator and rotor may vary between 60 and 90.

In use, and in accordance with one or more implementations, live comminuted plant material (with a size of 100-120 μιτι) is slurried in water and, at a temperature of not less than 80^QC and not more than 95^QC, is introduced to the reaction tank 26 through the funnel 32. In operation, slurry is fed through the inlet opening 43 of the stator 42 to the hollow interior of the rotor 44.

Solid inclusions are pressed by the centrifugal force and by the blades 50 to the stator and are intensively planed off by the teeth 47, whereby a preliminary comminution of the material is achieved. The relieving angle a makes the stator 42 operate like a file, reliably and quickly comminuting the material being treated, said material, being entrained by the liquid medium, passes through the cuts 49 and 52, being subjected in a stream to additional mechanical and hydrodynamic treatment.

In accordance with one or more implementations, the slurry is treated in the High Shear Processor 30 described above which is operated at an intensity of 10 to 50 w/kg of product for not less than 3 minutes. Operation of the mechanism 10 under these conditions (in particular at a temperature of not less than 80^QC and not more than 95^QC which is the optimum temperature range for molecular changes to take place) physically changes the stereochemistry of phytonutrients through realignment of the C-Ring bond between carbon atoms. This enables the free rotation of these carbon atoms carrying substituent (an atom or group of atoms substituted in place of hydrogen on the parent chain of a hydrocarbon) around the axis represented by the C Ring bond, and the alteration of the stereochemical structure of the compounds, to provide a composition from live plant material containing phytonutrients which are substantially a 50/50 mixture of cis and trans isomers.

One or more implementations may be used on any live plant material such as: any fruit, vegetable, legume, cereal, and in particular live citrus fruits, or live tomatoes, and has particular application to live green tea. Fresh green tea leaves in general contain 36% polyphenols among which catechins prevail and have a wide range of pharmacological properties such as antimutagenicity, anticarcinogenicity antitumorigenicity, antioxidant, antihypercholesterolemia and antibacterial activities. As reported by Katsuko Kajia et al. (Biotechnol Biochem 65:2638-2643, 2001 ), tea catechins are classified into c/^'s-type and frans-type from the configuration of the 2 hydrogen's at the 2 and 3 positions on the C-ring. Green tea infusion consists of almost c/^'s-type catechins and a small amount of frans-type catechins.

Green tea contains, among others, the flavonoid epigallocatechin gallate (ECGC). ECGC has biological effects on several stages of cancer, including but not limited to reacting with chemical carcinogens, the suppressing the spread of tumors. ECGC is as much as 100 times more powerful an antioxidant as vitamin C, and 25 times more powerful than vitamin E. ECGC also may account for the antibacterial properties of green tea.

Treatment of fresh green tea leaves using one or more implementations can produce a composition containing cis/trans catechins in a 50/50 ratio.

Successful recovery of the polyphenol EGCG in a stable form from green tea leaves can be important to the commercial utilization of this compound in the wellness and health industry. Tea catechin is a major component of tea leaf, constituting approximately 20 to 30 % of dry leaf. Catechins are water soluble and colorless flavonoids derived from the shikimic and acetate- mal on ate biosynthetic pathway.

Catechins are grouped into at least four types based on structure, including: epigallocatechin gallate (EGCG); epigallocatechin (EGC); epicatechingallate (ECG) and epicatechin (EC), of which EGCG is the dominant one. EGCG consists of 80% of 95% cis isomers, and GCG, the stereoisomer of EGCG, consists of 80% of 5% trans isomers. EGCG can form multiple hydrogen bonds without a substantial structural change, whereas GCG forms substantially less hydrogen bonds, suggesting the phenolic OH₄ group (catechin OH phenolic groups with lipid oxygen atoms) of EGCG and GCG may affect the interactions of the catechins with the lipid bilayer.

Processing Tea Leaves. Catechins are immature tannins peculiar to the leaves of the tea bush and are easily oxidized when the leaves detached from the plant. The readily oxidizing properties require careful handling when live tea leaves are harvested for the processing and recovery of catechins. In black tea production, catechin is reported to influence the color of the product through the tea pigment which is composed of orange color derived from the compound Theaflavin (TF) and a brown color derived from Thearubigins (TR). During the fermentation process in black tea production, TF is converted to TR suggesting that TF is short lived and probably acts as an intermediate molecule. The instability of tea catechins therefore requires skilful management of the live tea leaf in order to produce a stable and active compound. Several factors have been identified that influence the rapid degradation of tea catechins in live tea leaf.

The picking and handling of green tea leaves used in the process of this implementation process is thus very important as discussed below.

Temperature. High temperatures promote the initial formation of catechins within the tea leaf theaflavin, but they also favor further conversion of this TF to thearubigins pigments. This conversion results in reduced catechins in the tea leaf. Polyphenol oxidase (PPO) is one of the main enzymes responsible for the oxidation of the flavonols (polyphenols) to theaflavins (TF) and thearubigins (TR). The polyphenol oxidase activity is particularly affected by temperature.

Owuor et al. (Owuor, P.O., and Obanda, A.M., The Annual Report, TRF Kenya, 93-96, 1992) of the Tea Research Foundation in Kenya carried out experiments with different withering temperatures on clonal tea. They withered the leaf at ambient temperatures of 8-10°C, 15°C, 21 -23 °C and at 30 °C, and found that the chemical withering temperatures had significant effects on TF and TR brightness. Chemical withering at the highest temperatures of 21 -23°C and at 30 °C reduced the TF levels and increased the TC levels.

Cloughley (Cloughley, J.B., Journal of Science, Food and Agriculture 31 :920- 923,1979) of the Tea Research Center of Central Africa completed work on the effects of temperature on the enzyme activity during the fermentation phase of black tea manufacture. Fermentation was carried out at 15°C, 25 °C and at 35 °C. It was found that the rate of loss of PPO activity was markedly temperature dependent being more rapid at high temperatures. At the fermentation temperature of 35 °C, Clone SF2-204 had lost over 70% of its original PPO activity at the termination of fermentation after 150 minutes.

Molla (Molla, M.M., Sri Lanka Journal of Tea Science 61 :15-19, 1992) showed that there was no appreciable loss in PPO activity from 10°C to 32°C, but at temperatures higher then 32 °C there was a considerable decline in activity.

This scientific evidence supports an understanding that PPO activity is dramatically reduced when leaf is held at high temperatures, with the same loss of activity encountered in the field. However, for the purposes of commercially recovering catechins at such high temperatures, the live leaf suffers extensive moisture loss resulting leaf withering and rapid degradation. On the other hand, live leaf that is held at low temperatures, ideally 10°C maintains its high levels of catechin activity through the reduced PPO activity.

Leaf Handling. The start of heat build-up in harvested tea leaves commences in the tea field through poor leaf handling and transportation. This process starts in the puckers' basket and continues until arrival at the factory. Pickers are normally paid by weight of harvested leaf and therefore tend to press the leaf down to the bottom of the basket in order that their loads weigh more. Unfortunately this process allows the buildup of respiratory heat and physical damage to the leaf. The leaf can reach temperatures up to 50 °C which under these conditions, incipient fermentation takes place resulting in considerably lower PPO activity and much higher TF levels.

These results are relevant to biochemical processes that take place under certain conditions in the field. These results indicate that there is a loss of PPO activity under temperature stress, loss of moisture and a reduction in total oxygen uptake (TOU). However, there must be a certain balance between all these reactions in order to contain PPO activity.

On the opposite end of the scale, if leaf is left open in a pile, the TOU will be very high due to easy access of oxygen to all the shoots. However on a hot dry day, the open leaf loses moisture through evaporation, which prevents the leaf from heating up to such high temperatures. The perfect situation for the recovery of stable and active catechins is for the live leaf to be stored in a specific environment that prevent significant heat generation, allows a high degree of oxygen intake, and contains moisture loss.

Bruising. During poor leaf plucking and handling, the live leaf is exposed to bruising and cell damage which is not immediately apparent. Once the cell sap is exposed to the atmosphere, a change in the pigmentation of the tissue takes place, characteristically transforming the chloroplasts (green) into chromoplasts (red). Hence the appearance of "red leaf" is physical damage and heat abused leaf.

Melican et al. (Melican, N.J.T., and Mallows, R., Teacraft Technical Services Company, U.K., 1992) studied the causes of red leaf and concluded that particular care should be taken to prevent any leaf exceeding temperatures of 35 °C during the storage and transport of the plucked leaf. Red leaf occurs naturally under certain conditions and the employment of stringent management controls is necessary to prevent incipient damage taking place.

With reference to Figure 4, when green tea leaves (two - four leaves and a bud) are picked at the farm the leaves are collected and packed into baskets with care, to ensure that the leaves are not bruised, and immediately transferred to trailer 60 on a truck 62. .

The truck 62 transports the live leaves to a processing plant according to the implementation process, where the leaves are conveyed by a conveyor 64 into a chopper 66 (in this case a leaf chopper). Temperature controlled Water 68 from a water tank 70 is supplied to the leaf chopper where the leaves are cut into sizes of 100 to 200 μιτι, and slurried in the water. The amount of water added depends on the plant material being processed. In the case of green tea leaves, the ratio is between 40-60% fresh green leaves which contain 80% liquid (pectin and

surfactants are insignificant), and between 40-60%, by mass, water.

The slurried tea leaves are pumped by a pump 80 into a holding tank 82. From the holding tank 82, the slurried tea leaves are pumped by a pump 84 through a heat exchanger 86 where the emulsion is heated to a temperature of no less than 85^QC and no more than 92^QC. The emulsion is then added to the processing mechanisms 10A, 10B and 10C (as described above and illustrated in Figures 1 -3) where it is processed, for a period of no less than 3 minutes.

In an embodiment of the disclosed process, each processing mechanism can hold and process up to 50 liters of slurry. Each processing mechanism 10A, 10B and 10C can produce up to 300kg of processed material per hour; the system can therefore handle up to 900kg of processed material per hour. The slurry is passed on to a holding tank 88 which has a cooling jacket for cooling the processed material to ambient temperature.

The slurry may then be passed on for further processing for example for the extraction of phytonutrients such as catechins. The stereochemistry of the

phytonutrients such as catechins extracted from the treated material can be from to 20/80 to 80/20, preferably 30/70 to 70/30, most preferably about 50/50 ratio cis/trans isomers.

Medicinal use of plant polyphenols. Several studies support the medicinal effects of plant polyphenols, and further support the need in the art for plant polyphenols with improved bioavailability as disclosed herein. Several non-limiting representative studies are discussed below.

A study in 201 1 by the Centre for Infection, Division of Cellular and Molecular Medicine, St. George's, University of London, and the Centre for Parasitic Zoonoses, Institute for Medical Research, University of Belgrade, suggests that catechins with the galloyi moiety could be a novel and effective drug class that naturally inhibits the mosquito parasite and mammalian hexose transporter. It was found that catechins containing a gallate group (epicatechin-gallate and epigallocatechin-gallate) inhibit hexose uptake processes in infected erythrocytes, the primary mechanism of their anti-malarial activity which may involve interaction with alternative higher affinity target/s. (Ksenija Slavic, et al. Malaria Journal 10:165, 201 1 )

In the case where the live plant material is citrus fruits, the whole live fruit is processed at a ratio of 20-40 % fruit (containing 40-60% percent liquid and high in pectin and surfactants), and between 60-80%, by mass, water. The slurry is processed through the system for a period of no less than 3 minutes. The resulting emulsion may then be pasteurized and discharged into suitable packaging for domestic marketing bottles or containers and thereafter rapidly cooled to ambient temperature.

The stable emulsion produced in this process is high in pectin and surfactants and contains over one hundred and seventy released, active and stable

phytonutrients that includes more than sixty bioflavonoids (polyphenols that act as epinephrine antioxidants) with cellular metabolic pathways, and with the action to address many health issues that include respiratory, sinus, flu, the common cold; blood, vascular and capillary integrity by counteracting inflammation and stress conditions. In addition, the emulsion contains as many as fifty three identified limonoids that are not antioxidants but serve as chemopreventative agents, that inhibit the activity of carcinogens through biosynthetic pathways which are enzyme regulated and occur in specific tissues. Carcinogens are any substance, radionuclide or radiation which is an agent directly involved in the promotion of cancer or in the facilitation of its propagation.

In a 2003 study by the Department of Pharmacology, University La Sapienza, Rome, two citrus limonoids, namely limonin and nomilin were screened for their anti- HIV-1 properties (Battinelli, L. et ai, Plasta Med. 69:910-913). The results obtained established a similar anti-HIV activity of both limonoids indicating that the activity was not drastically influenced by the structural difference between the two

compounds. Conversely, the moiety of the molecules formed by the rings C and D and the associated furan ring appears to play an important role in the antiviral activity here observed because of the similarity to the cyclopentanone PM-92131 that showed a strong anti-HIV-1 activity. This moiety is also considered critical for the antifeedant and antifungal activity of limonin and nomilin (Figure 7).

Where the live plant material is tomato, lycopene is the major carotenoid responsible for the red color of tomato fruit, and tomato products are the major source of lycopene in Western diets. Nutritional studies on humans and animal models show that consumption of lycopene-containing foods increases the plasma levels of lycopene, which has been correlated with increased protection against prostate and other cancers, cardiovascular disease, and common eye diseases such as cataracts and age-related macular degeneration.

Lycopene is the predominant carotenoid in tomatoes and tomato-based foods and is also a predominant carotenoid in human serum and tissues. Intake of lycopene-rich foods is associated with decreased risk for several chronic diseases. Through research it has been observation that serum and tissue lycopene is more than 50% c/^'s-lycopene, whereas tomatoes and tomato-based foods contain mainly all-trans-lycopene, has led to the understanding that c/^'s-isomers of lycopene are more bioavailable

(Khachik, F. et ai, J. Cell. Biochem. Suppl. 22:236-246, 1995; Khachik, F. et ai, Analytical Chem. 69:1873-1881 , 1997; Parker, R.S., J. Nutr. 1 19:101 -104, 1989; and Stahl, W. et ai Arch. Biochem. Biophys. 294:173-177, 1992). Concentrations in liver were studied by Kaplan et al. (1990), Schmitz, H. et al., J. Nutr. 121 :1613-1621 , 1991 ); in testes by Kaplan et al. 1990, and Stahl et al.

1992); and the prostate by Clinton, S. K. et al. (FASEB J. 8:A423, 1994 and Can. Epidemiol. Biomarkers Prev. 5:823-833, 1996).

Cis-lycopene has been shown to predominate in both benign and malignant prostate tissues, demonstrating a beneficial effect of high c/s-isomer concentrations, and also the involvement of tissue isomerases in vivo isomerization from all trans to cis form (Clinton et al., Can. Epidemiol. Biomarkers Prev. 5:823-833, 1996).

The predominant form of lycopene found in tomatoes and other lycopene- containing fruits and vegetables is the a\\-trans stereoisomer, which is the straight chain form. Cooking increases the proportion of cis isomers of which there are several possible. C/^'s-lycopene isomers bend at the location of the cis double bond and have been found to be more bioavailable in human and animal studies. This means that cis isomers are more easily absorbed by intestinal cells and transported into the plasma. Hence, more lycopene in the diet is available for its utilization in protecting against degenerative diseases and the harmful effects caused by free radicals from sunlight and the metabolism of potential sources from ingested compounds.

Treated tomato emulsions and powders produced by shearing of tomato tissue, primarily tomato pericarp, were found to contain unusually high content of c/^'s- lycopene isomers. In addition, their resulting product also contains high amounts of released phytoene and phytofluene, compounds that are precursors in the

biosynthetic pathway to lycopene formation. Phytoene and phytofluene has been shown to have anti-proliferative effects in several human prostate cancer cell lines.

Lycopene has been identified as a protective agent against degenerative diseases caused by the action of free radicals produced in the body by metabolic reactions and exposure to ultraviolet light from the sun. Studies on human subjects in addition to other reports in the literature, show that cis isomers of lycopene are more bioavailable and therefore considered to be more bio-effective in protecting against damage caused in the body by free radical reactions.

Evidence in the literature also indicates that other carotenoids in tomato may work synergistically with lycopene in this protective action. Purified lycopene was shown to be ineffective against chemically induced prostate cancer in rats; whereas, a tomato powder and an oleoresin were considerably more effective in reducing tumorigenicity. Other carotenoids, notably phytoene and phytofluene, were shown to have anti-carcinogenic effectors.

In an independent study by the United States Department of Agriculture, Western Regional Research (Ishida, B., et al., Food Chemistry 132:1 156-1 160, 2012), fresh whole tomatoes were hydrodynamically processed using a process described herein. Changes in the ratio of natural 95% trans isomers and 5% cis isomers, to a ratio of 57% trans isomers and 43% cis isomers of total lycopene, were accomplished, thus confirming the commercial and industrial utility of the disclosed methods.

Having described the disclosure, the following general information further defines the data and compositions disclosed herein.

Unless otherwise indicated, all numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term "about." Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present disclosure. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.

Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the disclosure are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

The terms "a," "an," "the" and similar referents used in the context of describing the disclosure (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein is intended merely to better illuminate the disclosure and does not pose a limitation on the scope of the disclosure otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the disclosure.

Groupings of alternative elements or embodiments of the disclosure disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in any combination with other members of the group or other elements found herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and/or patentability. When any such inclusion or deletion occurs, the specification is deemed to contain the group as modified thus fulfilling the written description of all Markush groups used in the appended claims.

Certain embodiments of this disclosure are described herein, including the best mode known for carrying out the disclosure. Of course, variations on these described embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. Skilled artisans may employ such variations as appropriate, and the disclosure is intended to be practiced otherwise than specifically described herein. Accordingly, this disclosure includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the disclosure unless otherwise indicated herein or otherwise clearly contradicted by context.

Specific embodiments disclosed herein may be further limited in the claims using consisting of or and consisting essentially of language. When used in the claims, whether as filed or added per amendment, the transition term "consisting of" excludes any element, step, or ingredient not specified in the claims. The transition term "consisting essentially of" limits the scope of a claim to the specified materials or steps and those that do not materially affect the basic and novel characteristic(s). Embodiments of the disclosure so claimed are inherently or expressly described and enabled herein.

In closing, it is to be understood that the embodiments of the disclosure disclosed herein are illustrative of the principles of the present disclosure. Other modifications that may be employed are within the scope of the disclosure. Thus, by way of example, but not of limitation, alternative configurations of the present disclosure may be utilized in accordance with the teachings herein. Accordingly, the present disclosure is not limited to that precisely as shown and described.

Claims

1 . A method of processing live plant material containing isomeric phytonutrients, wherein the material is subjected to shearing to alter the

stereochemical structure of the isomeric phytonutrients by changing frans-isomers into the cis form, and/or changing c/s-isomers into the trans form.

2. The method as claimed in claim 1 , wherein said phytonutrients are selected from the group consisting of polyenes having about 95% frans-structure and flavonoids having about 95% c/s-structure.

3. The method as claimed in claim 2, wherein said polyenes comprise carotenoids.

4. The method as claimed in claim 3, wherein said polyenes comprise lycopenes.

5. The method as claimed in claim 2, wherein said flavonoids comprise catechins.

6. The method as claimed in claim 2, wherein said flavonoids comprise bioflavonoids.

7. The method as claimed in claim 2, wherein said flavonoids comprise limonoids.

8. The method of claim 1 , wherein a composition produced by the method contains isomeric phytonutrients from live plant material at a ratio of 20/80 to 80/20 cis/trans isomers.

9. The method as claimed in claim 8, wherein a composition produced by the method contains isomeric phytonutrients from live plant material at a ratio of 30/70 to 70/30 cis/trans isomers.

10. The method as claimed in claim 1 , wherein shearing is achieved by treating the live plant material in a High Shear Processor comprising a stator having an inlet opening, a coaxial cylinder with teeth defined by cuts therein, and a rotor which is made as a disk with blades defined by cuts in the cylinder and is brought in rotation with the help of a shaft, characterized in that installed additionally on the rotor, is an impeller comprised of straight or curved blades, and the stator, additionally has an outer concentric row of straightening blades defined by the cuts in the outer coaxial cylinder which encompasses the rotor from the outside, the width of the radial cuts between the straightening blades of the stator being at least two times smaller than their length for stabilizing the mechano-hydrodynamic effect produced on the product being processed; wherein:

the radial clearance between the teeth of the stator and the blades of the rotor does not exceed 0.2 mm.

1 1 . The method of claim 10, wherein said shearing is carried out at a temperature of between 80^QC and 95^QC.

12. The method of claim 10, wherein said shearing is carried out for a period of not less than 3 minutes.

13. The method of claim 10, wherein the live plant material is comminuted to an average size 100-120 μιτι, slurried in water, and the resulting slurry is processed by high shear processing.

14. The method of claim 10, wherein the live plant material is comminuted to an average size 100-120 μιτι and slurried in water at a ratio of 10-50% live plant material and 90-50% water, by mass, and said slurry is subjected to shearing.

15. The method of claim 1 , where said live plant material comprises at least one material selected from the group consisting of fruit, vegetable, legume, and cereal.

16. The method of claim 15, wherein said live plant material comprises green tea leaves containing c/s-type and trans-type catechins.

17. The method of claim 16, wherein said catechins are classified as c/s- type and trans-type from the configuration of the two hydrogen's at the 2 and 3 positions on the C-ring and are mainly in the c/s-type isomeric form and wherein, after processing, catechins in the process composition are raised to a ratio of about 20/80 to 80/20 cis/trans isomers.

18. The method of claim 18, wherein, after processing, catechins in the process composition are raised to a ratio of about 30/70 to 70/30 cis/trans isomers.

19. The method of claim 17, wherein, after processing, catechins in the process composition are raised to a ratio of about 50/50 ratio cis/trans isomers.

20. The method of claim 1 , wherein the live plant material is citrus fruit containing c/s-type and trans-type aglycones and glucosides.

21 . The method of claim 20, wherein said aglycones and glucosides are classified into c/s-type and trans-type from the configuration of the two hydrogen's at the 2 and 3 positions on the C-ring and are mainly in the c/s-type isomeric form and wherein, after processing, aglycones and glucosides in the composition are raised to a ratio of 20/80 to 80/20 cis/trans isomers.

22. The method of claim 21 , wherein, after processing, aglycones and glucosides in the composition are raised to a ratio of 30/70 to 70/30 cis/trans isomers.

23. The method as claimed in claim 22 wherein, after processing, aglycones and glucosides in the composition are raised to a ratio of about 50/50 cis/trans isomers.

24. The method of claim 1 , wherein the live plant material consists of tomatoes containing lycopene, phytoene and phytofluene isomers which are mainly in the frans-type isomeric form, wherein, after processing, said lycopene, phytoene and phytofluene isomers in the composition are raised to a ratio of 20/80 to 80/20 cis/trans isomers.

25. The method of claim 24 wherein, after processing, said lycopene, phytoene and phytofluene isomers in the composition are raised to a ratio of 30/70 to 70/30 cis/trans isomers.

26. The method as claimed in claim 25 wherein, after processing, said lycopene, phytoene and phytofluene isomers in the composition are raised to a ratio of about 50/50 cis/trans isomers.