Title of Invention

AN IMPROVED PROCESS FOR PRODUCTION OF TOCOPHEROL CONCENTRATES FROM DEODORIZED SOYA DISTILLATES

Abstract This invention relates to an improved process for production of tocopherol concentrates from deodorized soya distillates. The tocopherol concentrates produced according to the process of the invention are useful as pharmaceuticals, for cosmetic applications, and as food additive.
Full Text

Field of the Intention.
This invention relates to an improved process for production of tocopherol concentrates from deodorized Soya distillates. The tocopherol conccniraics produced according to the process of the invention are useful as pharmaceuticals such as for prevention and treatment of circulatory heart disease, providing protection against cognitive impairment in older people, cosmetic applications such as sunscreen preparation, skin conditioner, moisturizers, as food in food products that are high in fat content and animal farming as feed additive lo swine and poultry feeds due to their nutritional and antioxidant properties.
The natural mixed tocopherol (Vit.E) which are used in cosmetics, pharmaceuticals, food and animal farming industries is more expensive to manufacture because of their low content in raw materials and the complex chemical processes involved in their separation from the nontocophcrol components, yet has greater market value because of its higher efficiency as a nutrient and an antioxidant.
The natural form of Vitamin E is a mixture of four forms of tocopherols, viz. a-, P", y- and 6-tocopherol. Concentrates of these tocopherols in commercial vegetable oils vary widely. Soya bean oil contains up to 0.27% of mixed tocopherols. During refinement of the oil, a deodorized distillate or sludge is obtained (yield 1 % of the oil) in which the mixed tocopherol content ranges from 3-8%. The other components of the distillate are glycerides, free fatty acids, sterols, terpenes, hydrocarbons etc.
Soya sludge was considered as a waste product and used as an edible oil by poor people and its utility in processing for tocopherols is relatively recent in India. The invention can be considered as value addition to the waste product. The use

of sludge for Vit E production is now done by H.K. Agro hem, Hinkle and Sonia Biochem industries chiefly using molecular distillation process.
The synthetic Vitamin E, on the other hand, has a significantly lower value attached to it as it is used only as preservative. The a-tocopherol present in natural Vitamin E, functions as a nutrient, dietary supplement and as an antioxidant, while the other tocopherols present in it mainly function as antioxidants. Hence the higher value for natural form of Vitamin E.
The processes currently employed for the manufacture of natural tocopherol earn be classified into two which are:
(i) Process which converts a 3-8% mixed tocopherol containing raw material (deodorized vegetable oil distillate including the sludge obtained from soya bean oil refinement) to a 30% concentrate and (ii) Process which converts the product obtained the process (i) or directly converting the raw material employed in process (i) (deodorized vegetable oil distillate) into at least 50% tocopherol concentrate.
The main process step involved in the above said processes is the isolation of natural tocopherols by separating fiow the other components present in deodorized distillates. State of the art in the Field.
A process disclosed in US patent no. 2,704,764 dated. 22-3-1955 assigned to Refinery unincorporated involves esterification of the deodorized distillate followed by a series of steps to obtain tocopherol concentrate. The process consists of refluxing the deodorized soya sludge with methanol and cone. Sulphuric acid, cooling and filtering the precipitated sterols, distillation of methanol fiow the filtrate, washing the concentrate with water, followed by vacuum distillation of the methyl esters of fatty acids, addition of methanol lo

recover remaining sterols and distillation of methanol from the filtrate to get tocopherols.
Another US patent no 5,512,691 dt. 30-4-1996 assigned to Eastman Chemical CD. discloses a process for the preparation of tocopherol concentrates containing 20-80% tocopherols from vegetable oil distillates involving esterification followed by a series of distillation steps. The process comprises first of an esterification reaction where the more volatile alcohols are converted to the less volatile laity acid esters followed by a series of distillation steps where components boiling higher and lower than the tocopherols are separated from tocopherols and other like boiling substances.
The process of getting tocopherol concentrates involve esterification or saponification process to remove the fatty acids and the use of molecular distillation or short path distillation imit or wiped film reactor subsequently to obtain tocopherols of increased concentration. The two processes mentioned above are same as far as esterification and removal of fatty acids as esters is concemed. Subsequently the 2"^ process involves a series of (molecular) distillations to separate the components whereas the first process makes use of solvents to separate the components. The drawback, as per our observation, in the first process is formation of emulsions making it difficult to separate the organic and aqueous layers after esterification and washing to remove acid. Formation of emulsions results in the loss of tocopherols besides causing delay in processing . Further disadvantage is that the final methanol concentrate contains a lot of nO tocopherol material. The second process makes use of expensive equipment requiring high capital costs. There is no clear cut stage where sterols can be obtained as byproducts.

The demand for natural tocopherols appears to be growing and the feasibility of a modified process without the use of molecular distillation to produce 5()*X) and more of tocopherols can be of importance.
Preliminary market surveys indicated that marketing 30-40% tocopherol can be profitable if reliable raw material supply is assured. Higher tocopherol concentrates fetch even a better price. While surveying the literature we loud that both saponification and esterification processes can be used to remove the fatty acids and enrich the tocopherol content. While the saponification process provides 30% tocopherol concentrate further enrichment proved expensive requiring either short path distillation or extensive column chromatography. On the other hand the esterification route involved clear-cut stages for separation of fatty acids as esters, sterols, and tocopherols. It can also be less expensive provided the problems that are encountered are properly addressed.
Due to the increased demand for natural tocopherols in pharmaceuticals, cosmetics, food and animal farming industries and the availability of soya sludge fiow a number of oil refineries in horde (M.P., India) we started to investigate tlie intricacies involved in the development of a suitable process of obtaining tocopherol concentrates. Objective of the Invention
Therefore the main objective of the present invention is to provide an improved process for the production of tocopherol concentrates fiow deodorized soya distillates.
Another objective of the present invention is to provide an improved process for the production of 50% tocopherol concentrates besides providing valuable byproducts like sterols and fatty acids fiow deodorized soya distillates.

Another objective of the present invention is to provide an improved process (oi the production of 50% tocopherol concentrates which can be further enriched lo 70% or more tocopherol content by known methods, besides providing valuable byproducts like sterols and fatty acids from deodorized soya distillates which is simple and cost effective.
Yet another objective of the present invention is to provide an improved process for the production of 50% and more tocopherol concentrates besides providing valuable byproducts like sterols and fatty acids from deodorized soya distillates without the use of molecular distillation.
Still another objective of the present invention is to provide an improved process for the production of 50 % and more tocopherol concentrates.
Another objective of the present invention is to provide an improved process for the production of 50% and more tocopherol concentrates which can be used lor further enrichment of their a-tocopherol content by the known methods.
Another objective of the present invention is to provide an improved process lord the production of tocopherol concentrates in which formation of emulsion in lie working up of esterified product does not arise thereby avoiding employment of any means and /or steps for its prevention, thereby making the process not only economical but also environmentally safe.
Yet another objective of the present invention is to provide an improved process for the production of 50% and more tocopherol concentrates which can be scaled up for industrial application.
Still another objective of the present invention is to provide an improved process for the production of 50% and more tocopherol concentrates in which the

solvents employed are recovered and / or recycled thereby making the process further economical.
Another objective of the present invention is to provide an improved process lot the production of 50% tocopherol concentrates wherein the silica gel column can be used which can be used at least twice and can be regenerated, thereby making the process still more economical.
During our sustained research work in developing processes for natural tocopherols, we observed while working up of the esterified product, That the organic layer to be viscous and removing the acid from it by washing with water proved difficult and time consuming due to formation of emulsion. We tried adding non polar solvents like hexane or isopropyl ether to dilute the organic layer to overcome the problem. We also tried the molecular distillation process on a lab scale to enrich the tocopherols and found that the product gets distributed in a number of fractions. Besides the high cost of equipment, liberal use of liquid nitrogen for traps and other running costs made us try an alternative and less expensive process. Statement of Invention
Accordingly the present invention provides an improved process tour (he production of 50% and more tocopherol concentrates from deodorized soya distillates by novel method of isolation by adding non polar solvents like hexane or isopropyl ether to dilute the organic layer , and also the use of the molecular distillation process to enrich the tocopherols and found that the product gels distributed in a number of fractions to result in an improved process loi ihe production of 50% and more. The details of the process is described below.

Detailed description of the invention:
Accordingly the present invention provides an improved process for the production of 50% and more tocopherol concentrates from deodorized soya distillates which comprises.
a. Esterifying by refluxing the deodorized soya distillate containing 3-H%
tocopherols obtained during the refining of soya oil with a lower alcohol.
the alcohol containing one or two carbon atoms, in the presence ol an
acid catalyst.
b. Adding water to the resulting reflux mass and distilling to recover the
unused lower alcohol.
c. Separating the organic and aqueous layers by conventional methods.
d. Adding non polar solvents such as hexane or isopropyl ether lo the
organic layer, washing with water to remove the acid and distilling the
hexane or isopropyl ether from the organic layer.
e. High vacuum distilling the esterified product obtained in step (d) to
recover the fatty acids as esters as a byproduct,
f. Extracting the residue with warm polar solvent and cooling to recover
sterols as byproducts.
g. Distilling the polar solvent 95% ethanol to get 45% mixed tocopherol
concentrate.
h. Passing the 45% mixed tocopherol through a short silica gel column and eluting with acetone-hexane mixture or isopropyl ether-hexane mixture.
i. Evaporating the acetone -hexane eluate or isopropyl ether-hexane eluate to get 50% tocopherol concentrate.
Further enrichment of 50% tocopherol concentrate can be done by the known methods to get 70% or more tocopherol concentrate.

In a preferred embodiment of the invention the lower alcohol employed lord esterification may be either methanol or ethanol and the acid catalyst may be cone. HCl or cone. H2SO4. The alcohol used may be 3 to 5 times the weight of soya distillate and the amount of cone, acid used may be 15 to 30% of the weight o( soya distillate. The reaction mixture may be refluxed at a temperature in the range of 60-80°C for a period in the range of 6 to 8 hours.
The amoimt of water added may be 35 to 50% of the alcohol added and \he distillate collected may be about the same volume as the alcohol used. The hexane or isopropyl ether (the solvent) added may be about the same volume of the organic layer. The distillation of esterified product may be carried out at a pressure in the range of 0.1 to 0.5 mm. For the extraction of the residue wilt methanol or ethanol is carried out 3 to 5 times, with methanol or ethanol 6 to 8 times that of the residue, at a temperature in the range of 35 to 45° C for a period in the range of 15-20 minutes each time. The solvent extract solution may be kept at a temperature in the range of 0 to -5° C for a period in the range of 15-20 minutes under stirring, the methanolic or ethanolic suspension is fibered under reduced pressure to collect sterols.
The polar solvent filtrate is evaporated milder reduced pressure to recover methanol or ethanol and a residue with mixed tocopherol content of 42 to 46%. resultant product is dissolved in acetone-hexane mixture or isopropyl ether hexane mixture (1:99) in about 75 to 100% weight of the residue and passed over a column of silica gel. The quantity of silica-gel employed is in the range of 2 to 2.5 times the weight of the residue. The column is eluted with acetone-hexane mixtures (1:99 and 2:98).or isopropyl ether-hexane mixtures(l:99 and 5:95). 1 he tocopherol containing fractions are pooled and evaporated under reduced pressure to get the tocopherol concentrates.
The resulting product is in the form of yellow to light red colored viscous liquid analyzing for 48-54% mixed tocopherols (HPLC) with the approximate

composition a-7-8% ,p,y - 25-28%, and 5- 15-18%. The recovery of tocopherols is about 80-90%, The preparation is non toxic and produces no reaction on skin on topical application.
The invention is disclosed in detail in the following examples which is provided by way of illustration only and therefore should not be construed to limit the scope o(~ the invention.
Example: 1 Process for the preparation of tochopherol concentrate (50%)
a) Esterification of deodorized soya distillate:
150 go deodorized soya distillate (tocopherol content 6%) is taken in a 1 liter 3 necked round bottomed flask equipped with a mechanical stirrer, a thermometer socket and a reflux condenser and kept in a water bath. To this 450 ml methanol is added and stirring started. To the resulting solution 30 g Cone. H2SO4 is added drop- over 10 minis under continuous stirring. Heating is started after the addition of sulphuric acid. The reaction mixture is refluxed for 6 hours (keeping the inside temperature at 65 , Thereafter 150 ml of distilled water is added slowly and distillation started. About 450 ml of distillate is collected, the reaction mass is cooled to room temperature and the organic layer and aqueous layers are separated in a separating final. To the separated organic layer 100 ml of hexane is added and washed with distilled water three times each time using 100 ml of water. The hexane layer is dried over , filtered and washed with 30 ml hexane. Yield of esterified product is 155g.
b) Recovery of fatty acid esters by high vacuum distillation:
In a 500 ml 3-necked round bottomed flask kept in oil bath and with a mechanical stirrer, thermometer socket and distillation set up, the esterified product (155 g) obtained from step (a) is charged and when the oil bath temperature reaches high vacuum (0.5 mm pressure) is

applied. The distillate is collected over a temperature in the range of 165-180^ C when the distillation process will be over. The residue is cooled to room temperature.
Distillate is light yellow and weighs 92.9 g (methyl esters of fatty acids). Residue is dark brown and weighs 43.8 g. which on analysis indicates a content of 24% mixed tocopherols (HPLC).
c) Extraction of tocopherols and sterols from the residue obtained in step
(b) above:
The residue from the step (b) (43.8g) is taken in a 500 ml 3-necked round bottomed flask equipped with a mechanical stirrer and a reflux condenser. 300 ml of Methanol is added and the mixture heated in a water bath under stirring for 20 minutes with water bath temperature maintained at a temperature in the range of 40-45°C. Methanol-soluble part is separated by careful decantation and kept at a temperature in the range 0 to -5° C milder stirring when a solid (sterols) separate which is filtered and washed with cold methanol. The process of extraction is repeated 3 more times.
d) Preparation of 45% tocopherol product and sterols:
All the methanol filtrates containing tocopherols (obtained in step c) are mixed and concentrated under reduced pressure. Similarly all the sterol fractions (obtained in step c) are combined and dried. Weight of methanol soluble (containing tocopherols) 18.6 g, mixed tocopherols content 45% (HPLC), Weight of sterols (crude) 6.0 g. Weight of methanol insoluble residue 18.0 g.
e) Preparation of 50% tocopherol product:
18.6 g mixed tocopherol product obtained in step (d) is dissolved in acetone-hexane (1:99) mixture and passed through a silica-gel (36 g)

column of 3 cm diameter. Elution is done with the same solvent mixture followed by acetone-hexane (2:98), collecting fractions of 50 ml each. The first fraction contains hydrocarbons. The firaction 2-7 on removal of solvent under reduced pressure give 15.6 g of 50% mixed tocopherols concentrate (HPLC).
The composition of the mixed tocopherol concentrate is a-tocopherol 8%, P,g -tocopherols 27% and 5-tocopherol 15%. The recovery of tocopherols fiow the soya distillate is ca 87%. It appears as yellowish-red viscous oil.
Example: 2 Process for the preparation of tochopherol concentrate (50%)
a) Esterification using ethanol and sulphuric acid:
100 g of Deodorized soya distillate (tocopherol content 5.2%) is taken in a 1 liter 3 necked round bottomed flask equipped with a mechanical stirrer, a thermometer socket and a reflux condenser and kept in a water bath. To this 325 ml 95% ethanol is added and stirring started, lo the resulting solution 30 g Cone. H2SO4 is added drop-wise over 10 minutes under constant stirring and heating is started after the addition of sulphuric acid. The reaction mixture is refluxed for 7 hours (keeping tiie inside temperature at 80®C) thereafter 100 ml of distilled water is added slowly and distillation started. About 300 ml of distillate is collected, the reaction mass is cooled to room temperature and the organic and aqueous layers are separated in a separating final. To the separated organic layer 70 ml of isopropyl ether is added and washed with distilled water three times each time using 120 ml of water. The isopropyl ether is dried over Na2S04, filtered and washed with 20 ml isopropyl ether. The filtrate is concentrated under reduced pressure to remove isopropyl ether. Yield of esterified product is 105 g.

b) Recovery of fatty acid esters by high vacuum distillation:
In a 500ml 3-necked round bottomed flask kept in oil bath and equipped with a mechanical stirrer, thermometer socket and distillation set u]% the esterified product from step (a) is charged and when the oil hath temperature reaches 100°C. high vacuum (0.3mm pressure) is applied. The distillate is collected over a temperature in the range of 16()-175°C when the distillation process will be over. The residue is cooled to room temperature.
Distillate is light yellow and weighs 68.2 g (ethyl esters of fatty acids). Residue is dark brown and weighs 27.3 g. which on analyses indicates mixed tocopherols content of 23.5% (HPLC).
c) Extraction of tocopherols and sterols from the residue obtained in step
(b) above:
27.3 g of the residue obtained from the step (b) is taken in a 500 ml 3-necked round bottomed flask equipped with a mechanical stirrer and a reflux condenser. 160 ml of 95% ethanol is added and the mixture is heated in a water bath under stirring for 15 minutes with water bath temperature in the range of 42-45**C. Ethanol-soluble part is separated by careful decantation and kept at a temperature of 0°C under stirring when sterols separated which are filtered and washed with cold ethanol. The process of extraction is repeated 2 more times.
d) Preparation of 45% tocopherol product and sterols:
All the ethanol filtrates (obtained in step c) containing tocopherols are mixed and concentrated under reduced pressure. Similarly all the sterol flections (obtained in step c) are also combined and dried. Weight ethanol soluble containing tocopherols 11.9 g, mixed tocopherols content

43.5% (HPLC) Weight of sterols (crude) 4.1 g. Weight of ethanol insoluble residue 10.5 g.
e) Preparation of 50% tocopherol product:
11.9 g of the mixed tocopherol product obtained in step (d) is dissolved in 10 ml of isopropyl ether-hexane (1:99) mixture and passed through a silica-gel (30 g) column of 3 cm diameter. Elution is done with the same solvent mixture followed by isopropyl ether-hexane (5:95), colleting fractions of 40 ml each. The first fraction contains hydrocarbons. I he 2-5 on removal of solvent under reduced pressure give 9A g of 49% mixed tocopherols concentrate (HPLC).
The composition of the mixed tocopherol concentrate is a-tocopherol 7.5%, p,Y-tocopherols 26.5% and 5-tocopherol 15%. The recovery tocopherols from the soya distillate is ca 86%. It appears as yellows-red viscous oil.
Oral administration of the mixed tocopherol concentrate obtained in step (e) in examples (1) and (2) to mice in a dose of 2 g /kg showed no toxicity. Repeated topical application on animal skin showed no adverse reaction.
Advantages of the invention:
1. The product is safe and stable which can be used for pharmaceutical, food, cosmetic applications and in animal farming industries.
2. The process is simple and does not involve the use or maintenance of costly equipment.
3. The process can be scaled up for industrial use.
4. Solvents can be recovered and / or recycled in the operation thereby making the process economical.

5. The same silica gel column can be used at least twice and degeneracy, thereby making the process further economical.
6. Formation of emulsion in the working up of esterified product does not arise thereby avoiding employment of any means and /or steps for its prevention, thereby making the process not only economical but also environmentally safe.





We claim
1. An improved process for the production of 50% tocopherol concentrates
from deodorized soya distillates which comprises
a) Esterifying by refluxing the deodorized soya distillate containing 3-8% tocopherols obtained during the refining of soya oil with a lower alcohol, the alcohol containing one or two carbon atoms in the presence of an acid catalyst.
b) Adding water to the resulting reflux mass and distilling to recover the unused lower alcohol.
c) Separation of organic and aqueous layers by conventional methods.
d) Adding non polar solvents such as hexane or isopropyl ether to the organic layer; washing with water to remove the surplus acid and distilling the unused hexane or isopropyl ether from the organic layer
e) High vacuum distilling the esterified product obtained in step (d) to recover the fatty acid esters as byproduct.
f) Extracting the residue with warm polar solvent and cooling to recover sterols as by products.
g) Distilling the polar solvent such as 95% ethanol to get 45% mixed tocopherol concentrate.
h) Passing the distillate product through a short silica-gel molecular column and eluting with acetone-hexane mixture or isopropyl ether-hexane mixture.
i) Evaporating the acetone-hexane eluate or isopropyl ether-hexane eluate to get 50% tocopherol concentrate.
2. An improved process as claimed in 1 wherein the lower alcohol such as methanol or ethanol is used as the esterifying agent.
3. An improved process as claimed in claims 1 & 2 wherein the mineral acid such as concentrated sulphuric acid or concentrated hydrochloric acid is used as the acid catalyst.

An improved process as claimed in claims 1 to 3 wherein the alcohol used is 3 to 5 times the amount of soya distillate used and the amount of the concentrated acid used is in the range 15 to 30% of the soya distillate used. An improved process as claimed in claims 1 to 4 wherein the reaction mixture is heated at reflux temperature over a range of 60-80° C for a period of 6 to 8 hours. An improved process as claimed in claims 1 to 5 wherein the amount of water added is 35 to 50% of the alcohol added.
An improved process as claimed in claims 1 to 6 wherein the hexane or isopropyl ether added is about the same volume as the organic layer. An improved process as claimed in claims 1 to 7 wherein the distillation of the esterified product is carried out over a pressure in the range of 0.1 to 0.5 mm. An improved process as claimed in claims 1 to 8 wherein the extraction of the residue with methanol or ethanol is carried out 3 to 5 times, with methanol or ethanol 6 to 8 times that of the residue, at a temperature in the range of of 35 to 45° C for 15 to 20 minutes each time.
An improved process as claimed in claims 1 to 9 wherein the methanol or ethanol solution is cooled to temperature in the range of at 0 to -5°C for a period of 15 to 20 minutes under stirring.
An improved process as claimed in claims 1 to 10 wherein ethanolic or ethanolic suspension is filtered under reduced pressure to collect sterols. An improved process as claimed in claims 1 to 11 wherein the methanol or ethanol filtrate is evaporated under reduced pressure to recover methanol or ethanol and a residue with mixed tocopherol content of 42 to 46%. An improved process as claimed in claims 1 to 12 wherein the product is dissolved in acetone-hexane mixture or isopropyl ether -hexane mixture (1:99) in about 75 to 100% weight of the residue and passed over a column of silica gel. An improved process as claimed in claims 1 to 13 wherein the quantity of silica-gel employed is in the range of 2 to 2.5 times the weight of the residue.

An improved process as claimed in claims 1 to 14 wherein the column is eluted
with acetone-hexane mixtures (1:99 and 2:98).or isopropyl ether-hexane mixtures
(1:99 and 5:95).
An improved process as claimed in claims 1 to 15 wherein the tocopherol
containing fractions are pooled and evaporated under reduced pressure to get the
tocopherol concentrates with mixed tocopherol content of 48-54%.
An improved process as claimed in 16 wherein the tocopherol concentrate
obtained is enriched further elution methods to obtain 70% or more of
tocopherols.
An improved process as claimed in claims 16 and 17 wherein the tocopherol
concentrates obtained are enriched by elution methods for of-tocopherol content.
An improved process for the preparation of tocopherol concentrate (50%) from
deodorized soya distillate substantially herein described with reference to the
examples 1 and 2.


Documents:

300-mas-2001-abstract.pdf

300-mas-2001-claims filed.pdf

300-mas-2001-claims granted.pdf

300-mas-2001-correspondnece-others.pdf

300-mas-2001-correspondnece-po.pdf

300-mas-2001-description(complete)filed.pdf

300-mas-2001-description(complete)granted.pdf

300-mas-2001-form 1.pdf

300-mas-2001-form 19.pdf


Patent Number 210216
Indian Patent Application Number 300/MAS/2001
PG Journal Number 50/2007
Publication Date 14-Dec-2007
Grant Date 25-Sep-2007
Date of Filing 09-Apr-2001
Name of Patentee M/S. NATCO PHARMA LIMITED
Applicant Address NATCO HOUSE, ROAD NO.2, BANJARA HILLS, HYDERABAD 500 033
Inventors:
# Inventor's Name Inventor's Address
1 VENKATA RAO ERRAGUNTLA NATCO HOUSE, ROAD NO.2, BANJARA HILLS, HYDERABAD 500 033
2 SUDHEER PALADUGU NATCO PHARMA LIMITED, NATCO HOUSE, ROAD NO.2, BANJARA HILLS, HYDERABAD-33,
PCT International Classification Number C07 D 311/72
PCT International Application Number N/A
PCT International Filing date
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 NA