Title of Invention

"A PROCESS FOR PREPARATION OF ODOURLESS SOYBEAN OIL"

Abstract This invention relates to a process for preparation odourless soybean oil which comprises fractionating soybean oil, characterized in that the fractionating soybean oil using at least 10% transition metal salt impregnated column and eluting the fractions using conventional non-polar solvents followed by polar solvent to get odourless soybean oil in first fraction and PUFA oil is second fraction.
Full Text The present invention relates to A PROCESS FOR PREPARATION OF ODOURLESS SOYBEAN OIL
Refined soybean oil is usually not preferred as a frying medium because it emits undesirable fishy odour during frying. It is generally accepted that linolenic acid and some associated unsaturates which are not removed during refining are the cause for this fishy odour. Effort have been made to remove or minimise them. This is the first time soybean oil free from odour particularly fishy odour has been prepared by using fractionation method.
Reference may be made to warner K, Mounts T.L., stability of soybean and canola oils with modified fatty acid composition, J. AM. Oil Chem. Soc. 70:983(1993), wherein pilot plant processed samples of soybean and canola (low erucic acid rape seed oil) with fatty acid composition modified by mutation breeding and hydrogenation were evaluated for frying stability. Linoleic acid content were 6.2% for standard soybean oil. The linolenic acid content were 10.1% for standard canola oil 1.7% for canola modified by breeding. All modified oils had significantly (p
Reference may be made to prevot A, perrin J L, Laclverie G., Auge ph and Coustille S I. A new variety of low-linolenic rape seed oil characteristics and room odour tests, J. Am. Oil Chem. Soc. 67:161(1990), wherein two Canadian rapeseed oil, "wester" and " Low linolenic" supplied by the canola Council were studied and compared with a French rapeseed. The linolenic acid content of the low-linolenic variety is about 3%. Seventy two percent of the triglycerides with at least one linolenic chain disappeared. The room odour test showed that the "low linolenic" had odour compared to other two varieties. A fruity odour dominated in the "low-linolenic" and the fishy painty odour were particularly reduced in this oil. The drawbacks here is that authors have not prepared any low-linolenic acid oil by themselves, but they received it form other sources which were also not devoid of fishy odour.
Reference may be made to Kim IH, and Yoon SH, Effect of extraction solvents on oxidative stability of crude soybean oil, J.Am.Oil Chem. Soc. 67:165 (1990), wherein oxidative stability of crude soybean oils obtained by different extraction solvents such as hexane, water and Folch's solvent (mixture of two volumes chloroform and one volume of methanol) were determined by gas chromatographic analysis of head space and peroxide value of oil samples. For the determination of oxidative stability of oil samples total volatile compounds formation, molecular oxygen disappearance in the headspace and peroxide value of samples were measured. Crude soybean oil obtained by Folch extraction was most stable-in oil oxidation, and oxidative stabilities of oils
obtained by hexane and aqueous extraction, which were significantly much less stable than Folch-extracted oil, were not significantly different during ten weeks storage. The drawbacks are the work is not directly concerned with the removal of fishy odour in soybean oil. However, indirectly autooxidation has a bearing on unsaturation vis-a-vis linolenic acid content of oil. If linolenic acid is more, extent of auto-oxidation is more and if linolenic acid is less, extent of auto-oxidation is less.
Reference may be made to Mounts TL, Warner K, and List GR. Performance evaluation of hexane extracted oils from genetically modified soybean, J.Am.Oil Chem. Soc. 71:157 (1994). Wherein soybeans produced by induced mutation breeding and hybridisation were cracked, flaked and hexane-extracted, and the recovered crude oils were processed to finished edible oils by laboratory simulation of commercial oil-processing procedures. Three lines yielded oils containing 1.7, 1.9 and 2.5% linolenic acid. These low linolenic oils were evaluated along with oil extracted from the cultivar Hardin, grown at the same time and location, and they were processed at the same time. The oil from Hardin contained 6.5% linolenic acid. Low linolenic acid oil showed improved flavour stability in accelerated storage tests. Room odour testing indicated that the low-linolenic oils showed significantly lower fishy odour after 1 hr at 190°C and lower pungent odour after 5 hour. The main drawback with this work is that method used to remove fishy odour is genetic modification. However, low linolenic acid oil showed low fishy odour as expected.

Reference may be made to Shatlory YE, Soliman MM and Saadia MA, Evaluation of compounds with fishy odour in soybean oil, J. Fd. Sci. Technol. 33:510 (1996), wherein degumming and refining of crude soybean oil were carried out individually in the laboratory to study the volatile compounds of both degummed soybean oil, before and after refining, along with the refined soybean oil from commercial unit. Hexanal, nonanal, decanal and hendecanal volatiles were found to be responsible for fishy odour. Studies on immersion of soybean in different ratios of ethyl alcohal, water for 4 weeks showed absence of principal compound with fishy, odour in the refined oil prepared from the seeds treated with 50 and 75% ethyl alcohol. These are the different set of aldehyde compounds suspected to contribute to the fishy odour of soybean oil. The draw back of the work is that linolenic acid has not been investigated, the work is not directly related with the work filed for patent but they do contribute to the same objective.
Reference may be made to Taneja V K, Sharma N K and Wagle R D, Partial hydrogenation and wintersation of Indian soybean oil. J. Oil Tech. Assn. (I) 18:79 (1986), wherein it has been reported to reduce the fishy odour, soybean oil was subjected to selective hydrogenation to IV around 110 varying conditions of temperature, pressure and type of catalyst. Liquid fraction with more unsaturation and solid fraction with less unsaturatioh were separated. This partially hydrogenated liquid oil remains clear at 20°C and can be used with winterisation as salad oil. The drawback of the work is that hydrogenation

technique is used here to reduce the amount of linolenic acid to produce a salad oil and not a frying oil.
Some of the salts like Zinc sulphate, cupric chloride and nickel catalyst hitherto not tried have been screened to assess their separation capacity. Among the three the Zinc sulphate found working satisfactorily. The Zinc sulphate columns were used for fractionation of soybean oil. Percent fraction collected were 82,15 and 2.5 for I, II and III respectively. It has been indicated that Zinc Sulphate column with 90% RH behaved in the same manner as silver nitrate. It has been indicated that III fraction which had been separated by Zinc sulphate was linolenic acid rich glycerides having, the highest iodine value (145) is the major fraction responsible for fishy odour as has also been indicated by room odour test. The first fraction which is major fraction (80-82%) didn't give any fishy odour.
The main object of the present invention is to provide a process for preparation of odourless soybean oil which obviates the drawbacks detailed above.
Another object of the present invention is to develop a process wherein column impregnated chromatography is used with salt of transition metals particularly zinc sulphate for fractionation of soybean oil.

90%) impregnated column (15%) and eluting with 200 ml of hexane (I fraction) and 200 ml of diethyl ether (II fraction), the percentage recovery was, first fraction 80% and second fraction 20%, it indicates that during fractionation, the separation has taken place as per degree of unsaturation where first fraction has mainly saturates and monoene like oleic acid and second fraction has mainly PUFA
including linolenic acid, which is the main component responsible for fishy odour, as has been confirmed by room odour test when first fraction didn't give fishy odour, where as second fraction gave fishy, painty odour when it was heated at 180 - 190°C in an open crucible in a closed room.
Accordingly, the present invention provides a process for preparation of odourless soybean oil which comprises fractionating soybean oil characterized in that the fractionating soyabean oil using at least 10% transition metal salt impregnated column and eluting the fractions using conventional non-polar solvents followed by polar solvent to get odourless soybean oil in first fraction and PUFA oil is second fraction.
In an embodiment of the present invention, one of the interesting properties of molecules having an ethylene linkage has been utilised as to make them form complex of relatively low stability with Zinc sulphate, cupric chloride and mercury salt. So far this type of separation of lipid material based on number, type and position of unsaturated centres they contain used to be carried out by complexing these unsaturated molecules with silver nitrate. Besides affecting separation according to degree of unsaturation to geometry of double bond, argentation chromatography has a third attribute, namely the

In an embodiment of the present invention, one of the interesting properties of molecules having an ethylene linkage has been utilised as to make them form complex of relatively low stability with Zinc sulphate, cupric chloride and mercury salt. So far this type of separation of lipid material based on number, type and position of unsaturated centres they contain used to be carried out by complexing these unsaturated molecules with silver nitrate. Besides affecting separation according to degree of unsaturation to geometry of double bond, argentation chromatography has a third attribute, namely the ability to separate suitable positional isomers of unsaturated fatty acids.
In another embodiment of the present invention the Zinc sulphate can replace the silver nitrate as an impregnating agent for argentation chromatography. Zinc sulphate which is an inexpensive (Rs. 290/Kg) chemical will be preferred in place of silver nitrate which is very expensive (Rs. 15,100/Kg) chemical.
In yet another embodiment of the present invention, the fraction rich in linolenic acid glyceride supposed to be responsible for fishy odour of soybean oil is separated out as a by-product can be used to fortify the PUFA poor oils.
Regarding the nature of silver-olefinic co-ordination bond, it has been demonstrated that silver-olefinic bond is similar in character to the co-ordination bond of the other metals like chromium, iron, nickel, lead, titanium and zinc.

Since silver nitrate is a very expensive chemical and it cannot be used on a large scale in industrial units, we had decided to search some other salt which may be having a similar effect like silver nitrate. Zinc sulphate, cupric chloride and nickel catalysts were found to be very powerful bonding agents with unsaturated fatty acids as is shown in Table - 1
TABLE -1 PERCENTAGE RECOVERY OF SOYBEAN OIL ON ARGENTATION COLUMN
(TABLE REMOVED)
The soybean oil fractionated over argentation column of these metal salts. Very little material (2-6%) was recovered with hexane as first fraction showing a very strong association with unsaturates. The II fraction which was between 98 to 94%, could be eluted with pure di-ethyl ether. Cupric chloride and nickel catalyst were leached out from the column during diethyl ether elution. Hence, they were discarded. But zinc sulphate column worked satisfactorily without getting leached out. Zinc sulphate also happens to be much cheaper compare to silver nitrate. Because the retention of the oil over zinc sulphate argentation column is very high, the polarity of zinc sulphate has to be reduced by adding moisture to the adsorbent at different level. Hence, mixture of silica gel with zinc sulphate (15%) was kept at different RH (43, 75 and 90%) to incorporate moisture before applying to column.

TABLE - 2
SEPARATION OF SOYABEAN OIL OVER ZINC SULPHATE COLUMN AT
DIFFERENT MOISTURE LEVEL
(TABLE REMOVED)
Table 2 shows the % moisture introduced to the zinc sulphate impregnated adsorbent at 43, 75 and 90% RH was 8.8, 10.9 and 37.6% respectively. The 90% RH column behaved very much similar to the silver nitrate column, yielding 78.6% of first fraction and 21.4% of second fraction.
To confirm the efficacy and separatebility of zinc sulphate column the fraction obtained were subjected for Iodine value estimation. In the present investigation where unsaturation is the criteria for fractionation of soybean oil, Iodine value plays an important role. The three fractions obtained from silver nitrate and zinc sulphate columns were subjected to iodine value estimation and results are given
in Table -3
TABLE - 3
IODINE VALUE OF SOYBEAN OIL AND ITS FRACTONS OBTAINED FROM SILVER NITRATE AND ZINC SULPHATE COLUMNS*

(TABLE REMOVED)
* All the value are the average of three replications.
After examining the values it can be noticed that iodine value of fractions obtained by silver nitrate column and zinc sulphate columns are very close to each other. It confirms that zinc sulphate behaves very much similar to silver nitrate and that it can replace the silver nitrate column at a commercial level. As expected the Iodine value of first fraction was minimum and third fraction was maximum, indicating that most of linolenic acid had come to the third and the last fraction, as has also been confirmed by fatty acid composition Table-4.
TABLE - 4
FATTY ACID COMPOSITION OF SOYBEAN OIL AND ITS FRACTIONS OBTAINED FROM SILVER NITRATE COLUMN (WEIGHT %)
Refined soybean oil was procured from the local market, chemicals like silver nitrate, zinc sulphate, cupric chloride were AR grade, from BDH Chemical Co. where as Nickle catalyst was procured from Harshaw Catalyst NV, DE MERRN the Netherlands. The solvents like petroleum ether, diethyl ether were
t
from Qualigens Excellar R, grade. Ethyl alcohol was distilled over sodium hydroxide before use. Relative humidity chambers were prepared making use of

saturated aqueous solution of Potassium Carbonate, Sodium chloride and Barium Chloride which gave 43, 75, 90% RH respectively at 27°c.
As it is quite evident by the foregoing discussion that linolenic acid is the main component responsible for the fishy odour of soybean oil, major emphasis has been given to reduce the linolenic acid in the oil. As has been mentioned in the prior art portion mostly two methods namely genetic manipulation and hydrogenation have been tried to reduce the amount of linolenic acid. The previous workers were able to achieve this up to certain extent using the above two techniques. But here a new approach namely fractionation has bee employed by us. Separation and removal of glyceride containing linolenic acid has been achieved by fractination using a new agent zinc sulphate not used so far for this purpose. The first fraction obtained from zinc sulphate column was found deviod of fishy odour as has been confirmed by room odour test. The second fraction was rich in linolenic acid gave fishy, painty odour during heating and room odour test, which is undesirable. This can be used for fortification of PUFA poor oil.
The following examples are given by way of illustration of the present invention and therefore should not be construed to limit the scope of the present invention.

EXAMPLE -1
6.4 and 5.6 grams of soybean oil was loaded on silver nitrate and zinc
sulphate (15%) impregnated silica gel columns separately. The percentage
fraction collected from silver nitrate column and zinc sulphate columns were
82,15,2.5 and 80,12,8 respectively. Table 5 indicates that zinc sulphate column
with RH 90% behaved in the same manner as silver nitrate column and can be
used in place of silver nitrate column.
TABLE - 5
PERCENT SOYBEAN OIL FRACTION OBTAINED FROM SILVER NITRATE
AND ZINC SULPHATE COLUMNS*
(TABLE REMOVED)
* All the value are the average of three replication
EXAMPLE - 2
Preparation of silver nitrate impregnated column and fractionation of soybean oil : 100 gm of silica gel 60-120 mesh for column chromatography art.no. 194013, Sisco Research Labs. Pvt. Ltd., Bombay was mixed thoroughly with aqueous solution containing 15 gms of silver nitrate. The mixture was heated at 100°C for 1 hour and subsequently activated at 120°C for 3 hrs. After
activation the silver nitrate impregnated silica gel was allowed to cool in a desiccator. 50 gms of this coated silica gel was packed in a glass column (length 60 cms, dia 2 cms) with the help of 50 ml of dry hexane. The column was allowed to stabilised overnight in dark by covering with black paper. Fractionation of soybean oil was carried out by taking 10 gm of soybean oil mixed with 10 ml of dry hexane and loaded on silver nitrate column. Different components were eluted subsequently with hexane (first fraction), hexane + diethyl ether (1:1) - second fraction and only diethyl ether (third fraction). These fractions were recovered by removing solvents in a flash evaporator.
EXAMPLE - 3
Preparation of zinc sulphate impregnated column at different RH: Relative humidity chamber for 43, 75 and 90% RH was prepared using saturated aqueous solution of potassium carbonate, sodium chloride and barium chloride respectively kept at 27°C at an air tight condition. Silica gel (100 gms each) 60-120 mesh for column chromatography was mixed with zinc sulphate (15 g) 60-120 mesh thoroughly and kept at RH 43, 75 and 90% separately for 72 hours at 27°C. This silica gel impregnated with zinc sulphate (50 gms each) was packed in glass columns with the help of dry hexane. Soybean oil (10 g) along with dry hexane (10 ml) was loaded on columns an fractions eluted as has been done in the case of silver nitrate column.
EXAMPLE -4
Soybean oil 20 gms, was applied over the zinc sulphate column and eluted with 200 ml of hexane (first fraction) and 200 ml of diethyl ether (Second

fraction). The percentage recovery was : first fraction 79.80% and second action 20.21%. It can be inferred that during fractionation, the separation has taken place as per the degree of unsaturation. The first fraction has mainly saturates and monoene, like oleic acid and second fraction has mainly PUFA as has been seen by fatty acid composition. The second fraction is also rich in linolenic acid. As linolenic acid is main component responsible for fishy odour, the first fraction was found to be devoid of fishy odour where as second fraction was very rich in fishy odour as has been confirmed by room odour test. Room odour test of first and second fraction obtained from Zinc sulphate column was conducted by heating the oil in an open crucible at 180-190°C in a closed window room. The odour was tested in the room after heatings.
EXAMPLE - 5
Soybean oil 30 gms, was applied on a Zinc sulphate column, prepared as described earlier, and eluted with 200 ml of petroleum ether (b.p. 40-60°C) to obtain the first fraction and 200 ml of diethyl ether to get second fraction. The percentage recovery was: first fraction 80% and second fraction 20%. As usual the first fraction comprised mainly of saturates and monoenes and second fraction mainly of poly-unsaturated fatty acids. The main advantages of the present invention are: 1. The process developed removes fishy odour from the soybean oil, which is a
serious drawback, by fractionation method where no chemical reaction or
genetic manipulation is taking place.






We claim:
1. A process for preparation of odourless soybean oil which comprises fractionating soybean oil characterized in that the fractionating soyabean oil using at least 10% transition metal salt impregnated column and eluting the fractions using conventional non-polar solvents followed by polar solvent to get odourless soybean oil in first fraction and PUFA oil is second fraction.
2. A process is claimed in claims 1 and 2 wherein the transition metal salts such as sulphate with RH (Relative humidity) 90% is used.
3. A process is claimed in claim 1 to 3 wherein atleast 10% salt is impregnated in the chromatographic column.
4. A process is claimed in claims 1 to 4 where in non-polar solvents such as hexane or petroleum ether bp 40-60°C is used.
5. A process is claimed in claims 1 to 5 where in polar solvent such as diethyl ether is used.
6. A process for preparation of odourless soybean oil substantially as here in described with reference to the examples accompanying this specification.

Documents:

1195-del-1999-abstract.pdf

1195-del-1999-claims-cancelled.pdf

1195-del-1999-claims.pdf

1195-del-1999-complete specification(granted).pdf

1195-del-1999-correspondence-others.pdf

1195-del-1999-correspondence-po.pdf

1195-del-1999-description (complete).pdf

1195-del-1999-form-1.pdf

1195-del-1999-form-2.pdf

1195-del-1999-form-4.pdf

1195-del-1999-form-9.pdf


Patent Number 192842
Indian Patent Application Number 1195/DEL/1999
PG Journal Number 21/2004
Publication Date 22-May-2004
Grant Date 14-Oct-2005
Date of Filing 08-Sep-1999
Name of Patentee COUNCIL OF SCEINTIFIC AND INDUSTRIAL RESEARCH
Applicant Address RAFI MARG, NEW DELHI 110001, INDIA
Inventors:
# Inventor's Name Inventor's Address
1 JAMBUR VENKATESHIAH PRABHAKAR NO. 151, 7th MAIN ROAD, 3rd STAGE, GOKULAM, MYSORE-570 002 KARNATAKA, INDIA
2 NASIRULLAH CENTRAL FOOD TECHNOLOGICAL RESEARCH INSTITUTE, MYSORE-570 013, KARNATAKA, INDIA
PCT International Classification Number C11B 9/02
PCT International Application Number N/A
PCT International Filing date
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 NA