FIELD OF INVENTION

This invention relates to oil extracts from isolated beta-Carotene crystals, beta-Carotene rich fruits, underground parts of plants, flowers and green leafy biomass, method of making the oil extracts and to standardize them for beta-Carotene content for health benefit. BACKGROUND OF INVENTION

The outbreak of Covid-19 was first identified in Wuhan, Hubei, China in December 2019, and was recognized as a pandemic by the World Health Organization (WHO) on 11 March 2020. WHO has acknowledged that the virus will likely spread to all countries on the globe. It is widely recognized that the life in general has been thrown out of gear in most countries. Covid-19 has no medicine nor vaccine. However, The Innate Immunity, also known as Non specific Immunity is a resource that is mediated through Natural Killer Cells that have capability to protect a person from not only Coronavirus from but any microbial pathogen that enters body as well as cancer cells. Vaccines are specific to the strain of microorganism against which they are prepared. Non-specific Immunity, as per its name, is not specific to any pathogen. Its property is that Natural Killer Cells, which mediate the non-specific immunity, detect any cell of the body as “non-self” when it is infected with any and every pathogen that enters in body, or it is an early stage cancer cell and is yet to multiply and establish a nodule of cancer. Natural Killer Cell attaches itself to that “non-self’, injects its lethal charge into the “non-self’ cell to not only kill it but also to destroy the nucleic acid of the cell itself as well as that of the pathogen contained in the “non-self” cell to make sure that the infection is eradicated completely. In a situation wherein vaccine is not available and there are no antibodies in the body that arise from prior infection of the pathogen, infection from a pathogenic microorganism leads to development of the disease if the Natural Killer Cells activity has declined. This declines in persons with age, diabetes and strenuous physical activity. Hence, keeping the Natural Killer Cells activity to normal is first step that should be taken as a matter of general healthcare measure and as a primary step / First Line step against any epidemic/pandemic.

However, the means for improving Natural Killer Cell Activity in those who have the same in compromised state was shown by Santos et al. 1996 ¹ confirmed that 10-12 y of beta-Carotene supplementation (50 mg on alternate days) on NK cell activity in 59 (38 middle-aged men, SI- 64 y; 21 elderly men, 65-86 y) had significantly greater NK cell activity than elderly men receiving placebo.

Hughes (1999) ² pointed out that “Many epidemiological studies have shown an association between diets rich in carotenoids and a reduced incidence of many forms of cancer, and it has been suggested that the antioxidant properties of these compounds are a causative factor. Attention has focused on the potential role of one specific carotenoid, beta-Carotene, in preventing cancer, and numerous publications have described in vitro experiments and animal studies which suggest that not only can this carotenoid protect against the development of cancer, but also several other chronic diseases. Since the immune system plays a major role in cancer prevention, it has been suggested that beta-Carotene may enhance immune cell function. Several human trials, using dietary beta-Carotene supplementation with a wide range of intakes, have been undertaken to address this hypothesis. The general conclusion of these studies is that this compound can enhance cell-mediated immune responses, particularly in the elderly.” However, he also pointed to “the major unresolved problems of beta-Carotene; i.e. what intake is required for optimal immune function and other healthy properties.”

However, crystalline beta-Carotene has very low bioavailability, consumption of doses of beta- Carotene to the tune of 50mg alternate day used by Santos et al ¹ . are too high as compared to epidemiological consumption of beta-Carotene, which in North America is 1.5 mg/day as mentioned by Hughes (1999) ² and also from the view of practical impossibility of availability of beta-Carotene to that extent for daily consumption from ordinarily available food sources; and further, whether that high a consumption of beta-Carotene would be safe in the long run is also not known.

Purified preparations of beta-Carotene are available in soft-gel form which usually carry 25000 IU of beta-Carotene, for which consumption of one capsule per day is recommended on the label. However, beta-Carotene has only 0.08% solubility in oil at room temperature. Consequently, purified Beta-Carotene capsules in oil suspension shall have a very low bioavailability. This is evident from very high doses of the beta-Carotene used in clinical trials on improvement of Natural Killer Cells in adults to a tune of 50mg per alternate day for 38 middle- aged men, 51-64 y; 21 elderly men, 65-86 y as disclosed by Santos et al. (1996) ¹ . Satia et al (2009) ³ noted that in two such clinical trials with such high doses, the beta-Carotene and Retinol Efficacy Trial (CARET) in the United States tested the efficacy of 30 mg of beta- Carotene plus 25,000 IU of retinyl palmitate daily in male and female heavy smokers and in men exposed to asbestos; and the ATBC trial tested 20 mg of beta-Carotene plus 50 I U of vitamin E daily in male heavy smokers; both trials found that beta-Carotene, alone or in combination with vitamin E or retinyl palmitate, increased the incidence of lung cancers by 36% (in CARET) (Omen, 1996) ⁴ and 16% in the ATBC trial reported by Albanes ⁵ who showed that when compared with placebo, smokers are adversely affected with increased risk of lung cancer. In later clinical trials with lower doses of beta-Carotene from natural sources, Samman et al (2003) ⁶ .This clinical trial has shown that bioavailability of beta-Carotene was better and resulted in decrease in risk of Cardiac Heart Disease as indicated by decreased homocysteine level when, instead of isolated crystalline beta-Carotene, the same was provided through a mixed fruit and vegetable dehydrated juice at a rate of a dose of 12mg per day (which is also accompanied by presence of vitamin C, vitamin E and folic acid) in a double blind placebo controlled trial of two 6 week durations with a wash-out period of 3 weeks. This trial had 13 smokers and 19 nonsmokers in the trial. The dose used was also much smaller than 50 mg/day used for crystalline beta-Carotene in earlier reported prospective trials and no adverse effect was reported on smokers. Thus, use of natural source of beta-Carotene rather than synthetic source and at a smaller dose than 30mg per day was seen to be safer and equally effective at about 2.5 times smaller dose.

In conclusion, it seems that giving very large doses of synthetic/crystalline beta- capsules carrying 20 to 30 mg beta-Carotene is at least wasteful and at the worst, possibly an increased risk of cancer for smokers.

For this reason also, safer and efficacious compositions of beta-Carotene are required. In a cross-over clinical trial report of Chen et al ⁷ , which feeding of 23.42 mg of beta-Carotene per day through 200 g Purple Sweet Potato leaves (PSPL) elevated Natural killer Cell activity to about 3 time of baseline value within two weeks in Sixteen non-smoking healthy adults (seven men and nine women, aged 20-22 years), with normal weight (body mass index 20.9-21.6 kg/m ² ) Control group ate carrots providing same quantity of beta-Carotene. NK Cell activity increased significantly over the baseline and washout levels of in PSPL in the first week itself, whereas carrot diet required two weeks to improve the NK Cell Activity and that was at a level at which the PSPL resulted in 1 week. It was concluded that consumption of PSPL modulates various immune functions including increased proliferation responsiveness of PBMC, secretion of cytokines IL-2 and IL-4, and the lytic activity of NK cells. Increase in lytic activity of Natural Killer Cells was about 3 times over baseline within two weeks. This increase in such a short time as one week due to PSPL and that too in participants in young age group of 20-22 years is nothing short of being dramatic; since their Natural Killer Cell activity prior to the feeding trial is expected to be an optimum baseline for a healthy individual that requires no improvement; and it is not expected to increase substantially if the baseline level would be an optimum activity that would be a maximum achievable level for the Natural Killer Cell Activity in a healthy young person. However, by feeding PSPL 200g per day for one week resulted in increase of NK Cell Activity of about two and a half times and became three times at the end of two week feeding period PSPL is a very very good result that was available in 2005 itself. However, no further development occurred in the direction of developing methods of improvement of NK Cell Activity based on this disclosure. The reasons are obvious: everyday availability of 200 gram PSPL (or any other equivalent green leafy vegetable) round the year is not possible, eating 200 gram green leafy vegetable everyday is also not possible, and PSPL cannot provide a dosage form that can be manufactured and made available for daily consumption in a measured way. Hence, development of practical product and practical method to ensure everyday consumption of NK Cell Activity booster did not occur; despite its need in epidemics that came periodically thereafter such as Zica, Ebola, Swine Flue, Bird Flue, Nipah virus infection and encephalitis in children and more the ongoing COVID-19 which started in October, 2020 in China and thereafter spread very rapidly throughout the world prompting the World Health Organization to declare it as Pandemic on March 11, 2020; and world is suffering in absence of any NK Cell Booster developed thereafter. Surprisingly, although it is acknowledged that there is no vaccine, nor a medicine for the novel Coronavirus, the diseases is deadly, fast spreading, has hit global economy due to repeated lockdowns in various countries; the problem continues to exist and no one has any clue, except for waiting for a vaccine, which can never be 100% effective. SUMMARY OF INVENTION

This invention discloses a composition of oil with beta-Carotene dissolved in it in a quantity standardized for a quantity per dosage form that is effective for a health benefit available from beta-Carotene,. The composition of the oil according to this invention comprises, without limitation, beta- Carotene rich oil produced by a plant, extract of a plant part rich in beta-Carotene extracted in oil suitable for human consumption suitable for human consumption and the like. The beta- Carotene rich oil produced by a plant comprises, without limitation, Red Palm Oil/Palm Kernel oil and the like; the extract of a beta-Carotene rich plant part comprises, without limitation, beta- Carotene rich fruits, underground parts, green leafy biomass and flowers of plants. The source of composition of oil according to this invention also includes crystallized/solidified compositions of purified beta-Carotene. The beta-Carotene rich fruits comprise, without limitation, Tomato, (, Lycopersicon esculentum L.) Papaya ( Carica papaya L.) and Mango ( Mangifera indica L. ). The beta-Carotene rich underground parts of plants comprise, without limitations, carrot ( Daucus carota L.) and Sweet Potato ( Ipomoea batatas L) variety rich in beta-Carotene. The green leafy biomass comprise, without limitation Green leafy vegetable, Green leaves of cole crops, trees, shrubs. The beta-Carotene rich flowers comprise one or more, without limitations, Marigold (Tagetus erecta L.), Dandelion ( Taraxacum officinale L.), Saundersonia aurantiaca HOOK., Cameiiia crysantha HU., Ipomoea sp., Chrysanthemum (Chrysanthemum morifoliumindicum RAMAT.) and the like.

The health benefit available from beta-Carotene comprises, without limitation, improvement in Natural Killer Cell Activity, anti-allergy property, improving glow and health of skin, as a pro vitamin A for maintaining optimum level of Vitamin A within the body and the like. The health benefit from improvement in Natural Killer Cell Activity comprises, without limitation, First Line Innate Immunity from multiplication of pathogenic micro-organisms and pre-cancerous cells within the body.

Dosage form of the composition of the oil according to this invention comprises, without limitation, liquid oil, soft gelatine capsules, oil filled hard gelatine capsules, emulsions of various types, sachets of micro-capsules and gummies. In one embodiment, daily dose through the dosage forms comprises at least 418pg beta-Carotene with appropriate overages. The per day dosage of the composition of the oil according to this invention comprises, without limitation, 5ml of liquid oil, or 2 to 3 soft-gelatin capsules.

This invention also discloses a process of making composition of oil with beta-Carotene dissolved in it in a quantity standardized for a quantity per dosage form that is effective for a health benefit available from beta-Carotene. The process comprising steps of determining beta- Carotene content of beta-Carotene rich oil produced by a plant, or of an extract of a beta- Carotene rich plant part in oil suitable for human consumption, or of a solution of oil suspension of beta-Carotene crystals. If the content is greater than the target content of standard chosen for beta-Carotene, diluting the same by adding edible oil to adjust the content to the desired value is resorted to. If the content is lesser than the target content of the standard chosen for beta- Carotene, using the same extract to extract next batch of the plant parts and repeating these steps until the content of beta-Carotene, is raised to a level wherein standardized content can be adjusted; or partly removing the oil from the composition to get higher concentration of beta- Carotene. A part removal of the oil may be achieved by use of a detergent, soap or alkali to convert a part of the oil to water soluble component, and removing the water soluble component by adding water and removing the aqueous layer below the oil layer to leave behind more concentrated oil with respect to standardized content of beta-Carotene.

A beta-Carotene rich plant part in oil suitable for human consumption is made by steps comprising blanching a beta-Carotene rich plant part if required, comminuting the plant part, if required, removing as much moisture/water as possible by application of heat, contacting the comminuted plant part with an extractant comprising an edible oil or in combination a solvent for beta-Carotene and edible oil, allowing maximum possible dissolution of beta-Carotene in the extractant, removing the solvent for beta-Carotene, and recovering the oil as an extractthe solvent for beta-Carotene comprises a non-oil solvent permitted for solvent extraction of foods. The non-oil solvent comprises, without limitation, a group comprising hexane and supercritical Carbon Dioxide.

This invention also covers within its scope a composition comprising a composition of oil with beta-Carotene dissolved in it in a quantity standardized for a quantity per dosage form that is effective for a health benefit available from beta-Carotene as an ingredient, the composition of oil with beta-Carotene. Such compositions include, a composition for topical application to skin, a hair oil fortified with a composition of oil with beta-Carotene dissolved in it as an ingredient, and a cream or a lotion to be applied topically on the skin. The composition for topical application to skin being for anti-allergy, for skin nourishment or for cosmetic purpose. This invention also includes an extract of a plant part rich in beta-Carotene extracted in oil suitable for human consumption, the composition being made from a plant part other than green leafy biomass.

DETAILED DESCRIPTION OF THE INVENTION

Since PSPL increased their NK Cell Activity as much as three times in two weeks in healthy young adults, their decreased Natural Killer Cell Activity may reach equal to or near to what is considered as an optimum level, if not above the optimum/base level for healthy young adults. That shall also be a very good achievement because even in Innate immunity compromised persons (i.e. Natural Killer Cells Activity compromised persons), despite infection from a microbial pathogen, because the NK Cell Activity shall come very near to or equal to optimum level, the infected cells will be destroyed by the Natural Killer Cells before they release multiplied progeny of the infecting pathogen, and the disease will not flare up. Hence, this is considered as a very good performance of the dosage of beta-Carotene that was consumed during the clinical trial; and has been chosen at present as the standard at which the Innate Immunity Booster should be standardized. From green leafy material/vegetable, the bioavailability of beta-Carotene in terms of weight of Vitamin A converted in the body is 28: 1 (Tang 2010 ⁸ ). Since one molecule of Vitamin A is equivalent to half the molecule of beta-Carotene, the bioavailability in terms of weight of beta- Carotene molecules fed through green leafy vegetables to biologically available beta-Carotene is 56:1. Hence, daily feeding of bio-available beta-Carotene in the clinical trial of Chen et al. was 23.42/56 = 0.418 mg = 418 mg beta-Carotene per day. Allowing for overages of about 10%, the standard for Innate Immunity Booster works out to 460 pg beta-Carotene per day. If more overages are desired, the total required content will be higher proportionately. Providing this quantity of beta-Carotene per day through daily consumption of 200 gram green leafy vegetable is not a practical task. Enough availability of green leafy vegetable all over the world every day is not possible. Many people cannot eat this much green leafy vegetables every day. Many people do also not like green leafy vegetables. Further, as noted above, bioavailability of beta- Carotene provided as green leafy vegetable is very low. Hence, a practical alternative was needed. This problem continued to exist since the finding of Chen et al (2005) up to the date of this invention,

We noted that beta-Carotene in its natural plant derived raw materials are also known to be very low in bio-availability due to very low release in the digestive tract from the plant matrix in which beta-Carotene is embedded. Solubility in edible oils is also very low: 0.08% at room temperature, 0.2% at 60 °C and 0.8% at 100 °C. Hence, where beta-Carotene is the active ingredient of targeted health effect, its bioavailability in its carrier raw or processed food material is of paramount importance rather than its chemical content. For absorption in digestive tract, it is essential that the beta-Carotene gets dissolved in oil, the oil gets micronized during digestive process and the micronized oil droplet having beta-Carotene dissolved in it gets absorbed as a chylomicron from intestinal epithelium in blood stream. The link https://www.medicinenet.com/chylomicron/definition.htm ⁹ provides medial definition of chylomicron as: “A small fat globule composed of protein and lipid (fat). Chylomicrons are found in the blood and lymphatic fluid where they serve to transport fat from its port of entry in the intestine to the liver and to adipose (fat) tissue.” Thus, to ensure measured reasonable dose of beta-Carotene to get practically 100% absorbed, we decided to get edible oil solution of beta- Carotene. It may be noted here that although knowledge of Chylomicron exists for a long time, the problem of providing measured accurate easily manageable dosing of highly bioavailable beta-Carotene did exist since a long time, including Chen et al (2005), we are the first to devise oil solutions of beta-Carotene for this purpose.

Hence, in one aspect this invention discloses an oil comprising Beta-Carotene that is extracted. In a further embodiment, this invention discloses an oil extract made from a food source is standardized for quantity of beta-Carotene per dose effective for health benefit.

This invention also embodies process/es meant for making oils and oil extracts from beta- Carotene standardized for quantity of beta-Carotene per dose effective for health benefit rich food sources. In a further embodiment of this invention the oil extracts of this invention are made from beta-Carotene rich plant parts. The beta-Carotene rich plant parts include, without limitations, fruits, underground parts, green leafy biomass and flowers. In this inventions, from these plant parts oil extracts can be prepared to extract beta-Carotene dissolved in edible oil, define what is an effective level of the same for health benefit and standardize the oil extract to the required concentration in volume suitable for daily consumption. The health benefit that can be targeted would be, without limitation, for modulating regulating improving Natural Killer Cell Activity, anti-allergy activity, cosmetic applications. The Natural Killer Cell Activity includes immunity from infectious diseases and keeping body free from cancer cells.

Savangikar & Savangikar (2007) ¹⁰ (IN 291544, having priority date of 28 ^th December, 2006) have disclosed oil extract of leaf protein concentrate made from green leafy vegetation. However, this is an elaborate process of fractionation, which is tedious and expensive due to hard mechanical texture of the leafy shoots as raw material which are difficult for pulping for extracting leaf protein concentrate. If solvent extraction is envisaged, solvent recovery from massive residues that remain behind make organic solvent extraction process expensive. Further solvent residues in the final product is difficult and that makes the final product unhealthy. Hence, despite this disclosure, the problem of availability of convenient, affordable and easy to scale up product containing beta-Carotene in a quantity effective for improving Natural Killer Cell Activity remained without a solution; and the instantly described invention has solved the long standing problem. Based on Chen et al.’s results of clinical trial, 418 pg beta-Carotene per day was selected for developing per day dosage of the intended NK (Natural Killer) Cell Activity Booster. For oil extract, a dose of 5 ml was chosen for dissolving 418 pg beta-Carotene with desired overages. For making softgel, the extract obtained as above was decided to be standardized to contain 418 pg beta-Carotene in oil volume that can be filled in two or three softgels / soft gelatin capsules.

In one aspect, this invention discloses an oil solution comprising beta-Carotene. In another aspect, this invention discloses an oil solution standardized for an amount of beta-Carotene effective in a daily dose for improvement in Natural Killer Cells Activity. In a further another aspect, this invention discloses an oil solution standardized for an amount of beta-Carotene effective in a daily dose for improvement in Natural Killer Cells Activity for reduction in the risk of infection from pathogenic microorganisms and the risk of cancer. In a further embodiment, a composition for topical application comprising the oil of this invention comprising effective level of Beta-Carotene would be useful for skin health for cosmetic purposes and also as First Line response to skin diseases arising from pathogenic microorganisms, including, without limitation, herpis zoster, eczema or rash and hives arising from allergies. This invention also includes the beta-Carotene rich fruit oil when it is standardized to contain an amount of beta-Carotene effective for improvement in Natural Killer Cells Activity. This invention further also includes the beta-Carotene rich fruit oil includes Crude Palm Oil, also known as Red Palm Oil when it is standardized to contain an amount of beta-Carotene effective for improvement in Natural Killer Cells Activity for reduction in the risk of infection from pathogenic microorganisms and the risk of cancer. The beta-Carotene rich fruit oil includes, without limitation. Crude Palm Oil, also known as Red Palm Oil also known as Palm Kernel Oil.

Oil extracts of beta-Carotene rich fruits, underground storage organs and flowers can be made in several different ways. Some of which are illustrated below, which are non-limiting examples; and these examples do not restrict the scope of the claims of this invention.

Crude Palm oil, also known as Red Palm Oil extracted from mesocarp of the fruit of palm tree (Elaeis guineensis) is rich in beta-Carotene to a tune of around more than 250pg/g oil. It is an aspect of this invention that Crude Palm oil /Red Palm oil, when standardized for a dosage per day that has beta-Carotene content in daily dose effective for improvement in Natural Killer Cell Activity is useful for reducing the risk of infection from pathogenic microorganisms. This also includes reduction in the risk of infection from the novel Coronavirus.

Above claim on Crude Palm Oil / Red Palm Oil for reduction in the risk of infection from pathogenic microorganisms; in general, and from the novel Coronavirus infection in particular has inventive step because this has not been contemplated by any person ordinarily skilled in the art of Clinical Nutrition or healthcare before it is proposed here. It is surprising that Crude Palm Oil or Red Palm oil has never been proposed/contemplated for use for reduction in the risk of infection from pathogenic microorganisms in general. Particularly, as on 28th April, 2020 also, despite incidence of the deadly COVID-19 to 30,42,444 cases, leading to 2,11 ,216 deaths in more than 180 countries, although the knowledge exists that Crude Palm oil or Red Palm Oil does have beta-Carotene more than 250pg/g of the oil, the people are suffering, world is spending on Personal Protective Equipment, masks for prevention of the infection and 3 million people have been hospitalized with nothing on hand as medical treatment except for ventilator support to the advanced stage patients, no health expert, including World Health Organization and health experts in any country including health experts in India has contemplated use of the Red Palm oil for reducing the risk of infection from Coronavirus.

Crude Palm Oil / Red Palm Oil may be used as an oil standardized for effective dosage volume or as encapsulated for reduction in the risk of infection from pathogenic microorganisms.

In essence, it is noted that a solution of beta-Carotene in oil shall be in bio-available form and the same provided from any source of beta-Carotene at an effective daily dose for improving activity of Natural Killer Cells would be an Innate Immunity Booster/Natural Killer Cells Activity Booster useful as first line response for any viral/bacterial communicable disease, including Novel Coronavirus.

In one aspect of this invention, the composition of oil containing Beta-Carotene would enable maintaining safely Vitamin A level at optimum level throughout the pregnancy period, which is vitally necessary to ensure that all systems of the fetus develop to its full genetic potential. Managing to maintain optimum level of Vitamin A throughout pregnancy period is not possible with synthetic Vitamin A since it become readily toxic if it raises Vitamin A level within the body even a little above optimum level within the body, and to avoid such toxicity, if the synthetic Vitamin A is provided at defensive level, the level of Vitamin A in body readily drops down.

Shortfall in extent of growth of systems of the fetus occurring on account of lower than optimum level of Vitamin A in the body of pregnant woman can never be made up by Vitamin A supplementation after birth of the child in entire life. In one embodiment of this invention, from soft gelatin capsules available in market with 15mg natural beta-Carotene, recommended for daily consumption, substantially dilute but efficacious and safer oil solutions containing 460 pg beta-Carotene per 5 ml are made as Innate Immunity Booster/ NK Cell Activity Booster. This constitutes substantial technical advance against the prior art use of beta-Carotene.

In making oil solutions of beta-Carotene in oil, in illustrative examples, Rice Bran oil is used. However, any other edible oil can be used such as Groundnut oil, Safflower oil, Soybean oil, Coconut oil, Mustard oil, Fish oil, Algal oil, Castor oil, Sesame oil, Flaxseed oil, Palm oil, Olive Oil and any other edible oil. Borel et al (1998 ¹¹ ) have shown that Chylomicron beta-Carotene and retinyl palmitate responses are dramatically diminished when men ingest beta-Carotene with medium-chain rather than long-chain triglycerides. Hence edible oils rich in long chain fatty acids would be preferable; which include, without limitation, Soybean oil and Olive oil.,

If, in future, it seems that a different standard seems necessary for amount of beta-Carotene required for an Innate Immunity Booster, those standards can also be used instead of 418 pg beta-Carotene per day plus desired overages. In a further embodiment, this invention comprises oil extract of leaf nutrient concentrate, wherein the quantity of beta-Carotene dissolved in it is standardized per dosage form that is effective for a health benefit available from beta-Carotene,

In another embodiment, the oil extract according to above described invention has anti-allergy properties. Examples 7 and 8 illustrate this use. Thus, the oil extract of tomato of this invention would also have potential use for treating allergies; which is a novel property of the oil extract of tomato. This property is especially useful in the context of vaccination drive for COVID-19, since all vaccines have risk of allergies for those who are prone for allergies. This is particularly an aggravated risk for vaccines based on messenger RNA technology. This oil extract of tomato would potentially be an answer for this problem.

Extracting beta-Carotene from plant parts is conventionally done using organic solvents such as hexane, acetone and methanol and their mixtures. The isolated solvent extract with all solvent soluble components or crystallized beta-Carotene can thereafter be dissolved in oil after solvent removal to make oil solution of beta-Carotene. This method has also been illustrated and is one aspect of this invention. However, that is considered as a method of low in priority, since the residual raw material left after solvent extraction as well as the beta-Carotene-containing fractions retain solvent residues which are undesirable feature and solvent recovery of entire process becomes costly. Hence, although that route was available for making oil solution of beta-Carotene of this invention, the same has been illustrated on isolated beta-Carotene crystals available in market as encapsulated suspension of beta-Carotene crystals in oil, methods have been evolved for extraction of beta-Carotene in edible oil directly, so that an organic -solvent-less process be available for making the “Oil solution of beta-Carotene” product of this invention.

Making oil extracts from parts of plants was a challenging task, since the requirement was that for making an oil solution of beta-Carotene, if it is to be provided as a liquid dose, it should be possible to limit the same to a conveniently small volume dose per day, such as a 5ml once a day dose. However, the volume of the raw material that is required to be processed to get beta- Carotene in quantity effective for improving NK Cell Activity is far more in volume than 5 ml oil. Hence, getting an enough concentrated extract in oil from a voluminous raw material was a difficult task because the oil would get distributed over the entire surface of that mass and it is difficult to separate the same from the massive mixture of the raw material and oil added to it for extraction. Additional problem is most of the oil gets emulsified when mixed with raw material in a mixer, and recovering such oil extract from the mass of macerated raw material is a difficult task.

Weight of content of a 15mg beta-Carotene capsule in one such capsule sourced form market, excluding weight of the capsule shell is 160mg. No information is given on how much is Soybean oil in the capsule. Presuming that the same is 100mg, in view of solubility of 0.08mg beta-Carotene in 100 mg oil, the capsule contains 80 pg Beta-Carotene dissolved in oil (bio- available); and rest 14.92 mg beta-Carotene in a form that is not bioavailable. After consuming with food, further dissolution of the un-dissolved beta-Carotene derived from the soft-gelatin capsule will depend upon how much oil is contained in the food consumed concurrently with the capsule and only that much beta-Carotene will get dissolved in the available oils in the food. At 0.08% solubility in oil at room temperature for each 5ml oil, provided no other oil soluble material is dissolved in it, 4000 pg beta-Carotene will theoretically get absorbed from the 15000 pg beta-Carotene; however, in practice far less beta-Carotene will get dissolved due to competitive inhibition of dissolution if the oil has already dissolved other fat soluble components from the food. Thus there is an uncertainty of how much beta-Carotene will be bio-available from such a capsule for one individual, which shall differ from another individual due to differences in case-to-case for above variables. Thus, it is certain that a major portion of the beta-Carotene contained in a 15mg capsule will invariably be wasted. Further, even if, in some cases, it gets dissolved more than envisaged above, that may be injurious to smokers and it is not known how much is safe for a chronic overdose for non-smokers too. On the contrary, if target of daily dose for improving activity of Natural Killer Cells is 460 pg beta-Carotene per dose, a 15mg beta-Carotene capsule shall give 32.6 doses if its content dissolved in oil, which would give same effect as the consumption of one soft-gel containing 15mg beta-Carotene. Thus, instead of one soft-gel containing 15mg beta-Carotene oil suspension, 32.6 doses of 460 pg beta-Carotene each in oil form or in soft-gel form shall be a rational product for improving activity of compromised Natural Killer Cells.

However, oil extract without encapsulation may be the most rational/preferred way of consumption than its soft-gel because it is possible to ensure that the oil extract is well mixed with some component of food, which can be eaten with rest of the food of a major meal to ensure that all of the oil extract gets to the small particulate level which gets converted and absorbed in blood stream from the digestive tract in the form of Chylomicron which ensures complete bioavailability of the beta-beta-Carotene content of the dose. A chylomicron is “a lipoprotein rich in triglyceride and common in the blood during fat digestion and assimilation” as per Merriam-Webster dictionary on the link https://www.merriam- webster.com/dictionary/chylomicron ¹² . In case of a capsule we will have to assume that it will get released and get mixed equally well in the digesting food and all the contents would get converted into Chylomicron. Of course, yet capsules would be next best when palatability of the oil extract become limiting for some people or carrying a bottle of oil extract is not feasible for one or the other reasons.

EXAMPLES

EXAMPLE 1

DETERMINATION OF CONCENTRATION OF BETA-CAROTENE BY SPECTROPHOTOMETER

This method was used as provided by Daugan et al. (2010) ¹³ ; details of which are provided below: beta-Carotene content in the samples were analysed by Ultraviolet-Visible (UV-vis) spectrophotometer at 446 nm using MPOB test method. The sample was homogenized and weighed to the nearest ±0.0001 g into a 25 ml. volumetric flask. The sample was dissolved with n-hexane and diluted to the mark. The solution was transferred into a 1 cm quartz cuvette and the absorbance was measured at 446 nm against n-hexane. The beta-Carotene content of different vegetable oils is defined and calculated as beta-Carotene in parts per million (ppm).

The calculation was as follows:

Carotenoids content = [V x 383 x (As-Ab)]/ (100*W) Where:

V = The volume used for analysis 383 = The extinction coefficient for carotenoids AS = The absorbance of the sample Ab = The cuvette error W = The weight of the sample in g

Spectrophotometric analyses done at 446 nm were outsourced and colorimetric analyses at 460nm were done in-house.

EXAMPLE 2

EXPLORATORY EXPERIMENT FOR MAKING OIL EXTRACT FROM FRESH TOMATO, PAPAYA AND CARROTS Tomato was pulped in kitchen blender, juice was separated from pomace by straining through cloth. The pomace was dried. The juice was heated to boiling. It resulted in red precipitate, which was separated by filtration through cloth. It was found that oil extract, using ground-nut oil, could be made from wet as well as dry portions of pomace and the precipitate by mixing well and separating the oil by filtration, straining out through cloth and also by centrifugation. . The oil extracts showed characteristic orange/reddish yellow color which was florescent when seen against light which showed presence of beta-Carotene in them. This oil extract is useful as against whole juice because the oil extract is a concentrated one, has better stability against spoilage, is small in volume than the juice or original raw material and can be made into dosage forms, such as soft-gels which can be stored and be made industrially applicable.

Oil extracts were made by using same method from the pomace and precipitate obtained from Papaya fruit and carrots.

EXAMPLE 3

MAKING OIL EXTRACT FROM VARIOUS FRACTIONS OF TOMATO PASTE Oil extracts were also made from Tomato paste by adding 10ml oil to about 9 gram paste, mixing well, with or without additional water, and heating the mix on water bath in boiling water bath. Caotenoids dissolved in the oil and were separated from the wet residue by centrifugation or decantation or filtering through a porous membrane such as a cloth.

EXAMPLE 4 CONCENTRATING OIL EXTRACTS WITH RESPECT TO BETA-CAROTENE

Just as concentrated oil solutions can be diluted if required to standardize their content for a 5 ml dosage, there would also be instances when concentrating an extract may be required for its use, such as for making soft-gel capsules limited to a dosage of 2 or 3 capsules per day. This has been achieved as follows.

6.5 g of oil containing 137.6 microgram beta-Carotene per gram oil was treated with soap for removing a part of the oil, the mix was washed with water, the emulsion was treated with 1.5 g. salt and the mix was washed with water. After removal of the aqueous wash, the remaining oil was 3.71 g. Thus concentration was 57%, This means that the concentrated oil extract now contains 318.6 microgram of beta-Carotene. This can be appropriately used to make two softgels of 0.5 gram each and appropriately diluted to provide 418 microgram beta-Carotene plus 10% to 30% overages, as required, for making two one gram softgels or three softgels of about 670 milligram oil in each.

Crude Palm oil, also known as Red Palm Oil extracted from mesocarp of the fruit of palm tree (Elaeis guineensis) is rich in beta-Carotene to a tune of around more than 250pg/g oil. To adjust the beta-Carotene content accurately to what is required for making softgels from the Red Palm Oil, if there is a requirement of concentrating the content to some extent, the above technique of concentration the Red Palm Oil or any other beta-Carotene containing oil is now available.

EXAMPLE 5

PILOT SCALE PRODUCTION OF OIL EXTRACT FROM TOMATO PASTE

On pilot scale, tomato paste 50 kg, sourced from market, was mixed with 25 kg rice bran oil in an industrial mixer for half an hour and left overnight. Next day, the mass was filtered and squeezed through cloth bag. The expressed oil was filtered through common salt to remove traces of water. Recovery of oil as extract was 48% The filtrate was optically clear red colored oil. beta-Carotene and lycopene content was determined by spectrophotometric method. The weight of the extract taken was accurately about 1 g, it was diluted with hexane to 100ml and absorbance reading taken on spectrophotometer was 0.871. The content of beta-Carotene was calculated to be 33.4 mg per 100g of the oil extract. This corresponds to 1537 pg / 5 ml of the extract. This was diluted 3.34 times by rice bran oil to standardize the content to 460 pg beta-Carotene/ 5 ml oil extract.

Same method was reproduced on 100 kg tomato paste with 50 kg oil in second pilot scale experiment. The oil recovery as extract was 42%. Oil Extract contained 1336pg beta- Carotene/5ml.

In further pilot scale experiment, 50 kg tomato paste with 50kg oil, recovery of oil was 86% and oil extract contained 669 pg beta-Carotene/5ml. .

EXAMPLE 6

MAKING STANDARDIZED DOSES OF OIL SOLUTION OF BETA-CAROTENE FROM SOFT

GELATIN CAPSULES OF OIL SUSPENSION OF BETA-CAROTENE CRYSTALS Since at the time of this work there was a phase of withdrawal of lockdown and risk of infection had increased several-fold, one soft-gel of purified (100%) beta beta-Carotene having a label claim of 25,000 IU in one soft-gel that had been sourced in the past from market about 5 years ago was taken for this work. Weight of one capsule was 380 mg with shell with following ingredients: Soybean oil, gelatin, glycerin, Yellow Beeswax, Soy Lecithin. The weight of empty shell after taking out its content and drying of the shell was 120mg. The contents were dissolved in 20g Rice Bran oil. Of this 20g oil solution, 0.25 g was diluted to 25 ml with hexane and read at 460 nm in colorimeter, which gave absorbance reading of 0.43. This calculates to 758mg beta-Carotene /5ml of the oil solution. This 20 g solution upon dilution to 33ml shall provide an oil solution containing 460pg beta-Carotene/5ml. Thus, the 33 ml shall provide 33/5 = 6.6 doses.

It may be seen here that the soft-gel capsules were 5 year old. By taking recently made soft-gel containing 15mg shall provide 32 portions of oil solution containing 460pg beta-Carotene/5ml.

By appropriate variation/adaptation in above illustrated methods it is also possible to make oil soluble beta beta-Carotene wherein 1 ml of the oil extract shall have at least 460 pg beta- Carotene; so that two soft-gels would give daily dose of the Innate Immunity Booster/NK Cell Activity Booster made from the method of this invention. Soft-gels of oil solution would be convenient for those for whom oil extract is not palatable.

EXAMPLE 7

ANTI-ALLERGY EFFECT OF THE OIL EXTRACT COMPRISING 418pg BETA-CAROTENE/5 ML

In one self reported case, personally communicated by a person to the inventors, oil extract of tomato containing 418pg beta-Carotene/5 ml has stopped the person getting allergy from dust. The 69 year old person had dust allergy since 1998. He started using this oil extract product, named as Lelmmune-T, since last week of August 2020 mainly as a Natural Killer Cell Activity booster in view of the threat of COVID-19 pandemic. He also had dust allergy, due to which he used to start getting skin itching near the part of body which comes in contact with dust. Many times, this itching used to lead to swelling on his feet. Inhalation of air filled with dust used to lead to continuous sneezing. All above used to last for up to 30 minutes. Upon testing in an allergy clinic, he was tested for 18 different types of allergies including dust including paper dust, mosquito bites, groundnut and groundnut oil, some proteins, shell fish and some flowers. He took homeopathic medicines during the year 2000-2002. He was advised to avoid the articles that he was allergic to. He complied with that for years; but the allergies persisted to appear on and off. His major problem was dust and dust from old papers; which persisted until last week of August, 2020 when he started consuming this oil extract named as Lelmmune-T.

On 25 ^th November 2020, he shifted his factory to another location where he was forced to come in contact for long duration of time daily with dust and cement dust. He was surprised after a few days that there was no skin itching, which totally stopped until he reported this observation on 12 ^th January, 2021.

He consumed 5 ml of the oil extract of tomato once per day by applying the oil on bread slice daily for morning breakfast and have it with tea and milk.

These oil extracts are being studied for other phyto-ingredients also, other than beta beta- Carotene that bring health benefit including, without limitation, lycopene and vitamin E. EXAMPLE 8

ANTIALLERGY EFFECT OF THE OIL EXTRACT COMPRISING 418pg BETA-CAROTENE/5 ML

In a self reported case, a diabetic person aged 68 years undergoing a dental treatment was prescribed Metronidazole 200mg three times a day. He started getting random allergic patched of eruptions of blisters on his skin requiring scratching. He mixed applied moisturizer to such eruptions, which did not help. He mixed few drops of the oil containing 418pg beta-Carotene/5 ml with the moisturizer and applied it to these patches. The scratching stopped with immediate effect and the patched disappeared on next day. EXAMPLE 9

MAKING EDIBLE OIL EXTRACT COMPRISING BETA-CAROTENE USING BETA-CAROTENE DISSOLVING SOLVENT AND OIL AS EXTRACTANTS

Hexane 20ml was added to 25g Tomato paste (dry weight 12.21 g) and mixed well. Six gram Rice Bran oil was added to the mix and mixed further. The hexane with dissolved beta-Carotene and oil was poured out; hexane was allowed to evaporate. This yielded 6.75 g oil extract comprising beta-Carotene. The oil extract was determined to comprise 688pg/5ml beta- Carotene. This can be diluted to 418pg/5ml beta-Carotene plus desired overages, which would be 10% or 30% needing on the standard for overages chosen. In similar fashion, other solvents in which beta-Carotene is soluble, including supercritical extraction by supercritical fluids in which beta-Carotene is soluble can be done. Such supercritical fluids include, without limitation, Supercritical C0 ₂ .

EXAMPLE 10

ANTI-MET ASTSIS EFFECT In a self reported case, a 56 year old woman to whom breast cancer was detected on 4 ^th October 2018 was treated with radiation, chemotherapy and surgery. To reduce the risk of metastasis, from August 2020 she started consuming daily 5 ml of the oil extract containing 418pg beta-Carotene/5 ml. Her annual examination was conducted on 5 ^th Match.2021. All PET scan and other reports were. Normal. Her current medication is Arimidex 1mg(Hormone therapy)and daily 5 ml of the oil extract containing 418pg beta-Carotene/5 ml. EXAMPLES 11

OIL EXTRACT OF TOMATO FOR COSMETICS

Application of few drops of the oil extract containing 418pg/5ml beta-Carotene plus 10% overage mixed with moisturizer to face everyday led to glow on the skin due to its beta- Carotene content. It is not necessary that the beta-Carotene content should be 418pg/5ml in the oil extract for giving the cosmetic effect.

EXAMPLE 12

OIL EXTRACT OF LEAF NUTRIENT CONCENTRATE STANDARDIZED FOR QUANTITY OF BETA-CAROTENE DISSOLVED IN IT EFFECTIVE FOR A HEALTH BENEFIT AVAILABLE FROM BETA-CAROTENE

5 kg of Methi (Fenugreek; Trigonell foenum-graecum L) green leafy vegetable biomass was pulped in a pulverizer, juice was pressed out from the pulp by squeezing through cotton cloth. The juice was added slowly to boiling water in a container to keep the water ina pan in boiling state. The juice coagulated to provide a green precipitate, the Leaf Nutrient Concentrate, which was removed by filtration through a cotton cloth and dried. The yield was about 60 gram after air drying.

To 39 g Leaf Nutrient Concetrate, 39 g of Rice Bran Oil was added and kept in freexe for 5 days. On 6 ^th day beta-Carotene content was determined on colorimeter at 460 nm wavelength; which was 849pg/5ml of beta-Carotene in the oil extract. This was diluted 1.8 times to get oil extract standardized for 460 pg/5ml beta-Carotene in the oil extract. REFERENCES

1. Santos Michelle S, Simm Nikbin Meydani, Lynette Leka, Dayong Wu, Nader Fotouhi, Mohsen Meydani, Charles H Hennekens, J Michael Gaziano, Natural killer cell activity in elderly men is enhanced by b -carotene supplementation, Am J Clin Nutr I996;64:7727

2. Hughes David A. Effects of carotenoids on human immune function, Proceedings of the Nutrition Society (1999), 58, 713-718

3. Satia, Jessie A, Alvson Littman, Christopher G. Slatore, Joseph A. Galanko, and Emily White. Long-term Use of b-Carotene, Retinol, Lycopene, and Lutein Supplements and Lung Cancer Risk: Results From the Vitamins And Lifestyle (VITAL) Study, Am J Epidemil. 2009 June 1 , 169(11): 1409

4. Omenn GS, Goodman GE, Thornquist MD, et al. Risk factors for lung cancer and for intervention effects in CARET, the Beta-Carotene and Retinol Efficacy Trial. J Natl Cancer Inst. 1996;88(21): 1550-1559.

5. Albanes D, Heinonen OP, Huttunen JK, et al. Effects of alpha-tocopherol and beta-carotene supplements on cancer incidence in the Alpha-Tocopherol Beta-Carotene Cancer Prevention Study. Am J Clin Nutr. 1995;62(6 suppl):1427S-1430S.

6. Samman Samir, Gayathri Sivarajah, June C. Man, Ziaul I. Ahmad, Peter Petocz and Ian D. Caterson], A Mixed Fruit and Vegetable Concentrate Increases Plasma Antioxidant Vitamins and Folate and Lowers Plasma Homocysteine in Men; Journal of Nutrition, 2003, 2188-2193, American Society for Nutritional Sciences

7. Chen Chiao-Ming, Sing-Chung Li, Ya-Ling Lin, Ching-Yun Hsu, Ming-Jer Shieh, Jen-Fang Liu. Consumption of purple sweet potato leaves, modulates human immune response: T- lymphocyte functions, lytic activity of natural killer cell and antibody production. World J Gastroenterol 2005; 11(37):5777-5781

8. Tang G. Bioconversion of dietary pro Vitamin A carotenoids to vitamin A in humans. Am J Clin Nutr. 2010 May;91(5):1468S-1473S. doi: 10.3945/ajcn.2010.28674G. Epub 2010 Mar 3.

9. https://www.medicinenet.com/chylomicron/definition.htm

10. SAVANGIKAR, Chitra, Vasant and SAVANGIKAR, Vasant, Anantrao, INTEGRATED PRODUCTION OF PHYTOCHEMICAL RICH PLANT PRODUCTSOR ISOLATES FROM GREEN VEGETATION IN IN 291544, 2007..

11. Borel, P, V Tyssandier, N Mekki, P G roller, Y Rochette, M C Alexandre-Gouabau, D Lairon, V AzaTs-Braesco Chylomicron beta-carotene and retinyl palmitate responses are dramatically diminished when men ingest beta-carotene with medium-chain rather than long- chain triglycerides. J Nutr 1998 Aug;128(8):1361-7. doi: 10.1093/jn/128.8.1361..

12. https://www.merriam-webster.com/dictionarv/chylomicron.

13. Dauqan, Eqbal, Halimah Abdullah Sani, Aminah Abdullah, Halimah Muhamad and Ab Gapor Md. Vitamin E and Beta Carotene Composition in Four Different Vegetable Oils. American Journal of Applied Sciences 8 (5): 407-412, 2011 ISSN 1546-9239 © 2010 Science Publications