Process for the fractionation of seeds from oleaginous plants

Abstract

The invention relates to a process for the fractionation of seeds of oleaginous plants of to the Asteraceae family, comprising at least one mechanical pressing operation of the seeds and an extraction with polar organic solvent of the first residue obtained. Said process allows to separate oil, active substances and a solid residue which is particularly suitable to be used in animal feed. The present invention also relates to said solid residue and to its use for the production of animal feed, as well as to the extracted active substances and to their use as cosmetic and/or pharmaceutical ingredient.

Claims

1. A process for the fractionation of seeds of an oleaginous plant of the Asteraceae family into oil, solid residue and active substances comprising the steps of: (a) subjecting said seeds to at least one mechanical pressing operation, removing at least a portion of the oil and obtaining a first residue; (b) subjecting said first residue to extraction with a polar organic solvent and separating the remaining solid residue from the resulting liquid phase comprising oil and extracted active substances; (c) separating the active substances and oil from said liquid phase of step b), wherein the extraction of step b) is performed at temperatures above ambient temperature and below or equal to the boiling point of the solvent and wherein the weight ratio between the polar organic solvent and the first residue is from 1:1 to 4:1.

2. The process according to claim 1 wherein the step (c) comprises the addition of an apolar solvent and subsequent separation of an apolar fraction comprising oil from a polar fraction comprising the active substances.

3. The process according to claim 1 comprising, prior to step (a) at least a preliminary stage of treatment of the seeds selected from: (i) cleaning and sieving; (ii) decortication; (iii) drying; (iv) comminuting and/or grinding.

4. The process according to claim 1, comprising a single mechanical pressing operation before the step (b).

5. The process according to claim 1, comprising at least two operations of mechanical pressing before the step (b), at least one of which conducted at temperatures from 55-75 C.

6. The process according to claim 5 comprising at least a step of grinding and/or drying of the intermediate solid residue between the at least two operations of mechanical pressing.

7. The process according to claim 1, wherein said oleaginous plant of the Asteraceae family belongs to the Cardueae tribe.

8. The process according to claim 1, wherein said polar organic solvent is selected from, methanol, ethanol, isopropanol, isobutanol and higher alcohols, their esters with acetic and propionic acids, acetonitrile, acetone, methyl ethyl ketone, diethyl ketone, ethyl propyl ketone, THF, 2-methoxyethanol.

9. The process according to claim 8, wherein said polar organic solvent is ethanol or isopropanol.

10. The process according to claim 2, wherein said apolar solvent is selected from petroleum ether and/or hexane.

11. A solid residue resulting from the seeds of an oleaginous plant of the Asteraceae family and obtainable from step b) of the process according to claim 1, having a protein content higher than 20% by weight and an oil content of less than 10%, wherein said oleaginous plant is Cynara cardunculus.

12. A method for producing animal feed which comprises adding the solid residue according to claim 11 to the animal feed.

13. An animal feed comprising the solid residue according to claim 11, fiber, fat, minerals, carbohydrates, vitamins.

14. An active substance obtained from step c) of the process according to claim 1 in a form of a mixture rich in polyphenols characterized by the presence of Arctiin and Tracheloside in a weight ratio of 0.6 to 1.5, wherein said oleaginous plant is Cynara cardunculus.

15. The active substance according to claim 14 further comprising one or more substances selected from Arctigenin, Trachelogenin, Cynarine, Cynarinine, Chlorogenic acid, Nortrachelogenin guaiayacylglyceryl ether, Silibinin, Isosilibinin, Silicristin, Silidianin, N-(p-coumaroyl)serotonin glucoside, N-feruloylserotonin glucoside, and their isomers.

16. A composition comprising the active substance according to claim 14 as a cosmetic ingredient, or as biostimulant.

17. The process according to claim 2 comprising, prior to step (a) at least a preliminary stage of treatment of the seeds selected from: (v) cleaning and sieving; (vi) decortication; (vii) drying; (viii) comminuting and/or grinding.

18. The process according to claim 2, comprising a single mechanical pressing operation before the step (b).

19. The process according to claim 3, comprising a single mechanical pressing operation before the step (b).

20. The process according to claim 2, comprising at least two operations of mechanical pressing before the step (b), at least one of which conducted at temperatures from 55-75 C.

21. The active substance according to claim 14, wherein the weight ratio of Arctiin to Tracheloside is 0.7 to 1.3.

22. The active substance according to claim 14, wherein the weight ratio of Arctiin to Tracheloside is 0.8 to 1.2.

Description

EXAMPLES

Example 1

(1) (Stage a)

(2) 200 kg of seeds of Cynara cardunculus were cleaned, screened and subsequently dried to a water content of 6.6% by weight.

(3) The said seeds were fed to a single screw press (Mod. MIG PC25S screw diameter=110 mm; L/D=4.4) operating at 20 rpm at ambient temperature with a throughput of 75.8 kg/h yielding 38.8 kg of oil and approximately 158 kg of a first intermediate residue (containing the approximately 7.5% by weight of water). The said residue was dried in a 3 tray dryer (diathermic oil T=170 C.; residence time=55 minutes) at approximately 60 C. until the water content was 2.4%. The intermediate residue dried in this way was again fed to the same previously used single screw press operating at 10 rpm at a temperature of 60 C. with a throughput of 45 kg/h, yielding 5.35 kg of oil and 145.8 kg of a second residue comprising de-oiled seeds having an oil content of approximately 4% by weight.

(4) (Stage b)

(5) 1 kg of the de-oiled residue so obtained was then extracted with ethanol (residue/ethanol ratio=1/2 by weight) under reflux for one hour in a reactor provided with a mechanical stirrer. The suspension obtained was filtered and washed with ethanol (200-300 ml). The solid phase was dried overnight in a stove at a temperature of 80 C.

(6) A solid residue weighing 835 g having the following composition was obtained: oil: 0.3% by weight; protein content: 27% by weight.
(Stage c)

(7) The ethanol extract (liquid phase) obtained from stage (b) was dried in a rotary evaporator yielding an orange-coloured pasty solid which was then extracted with petroleum ether (300-400 ml) at ambient temperature in a reactor fitted with a mechanical stirrer so as to obtain a homogeneous dispersion which was then filtered.

(8) The apolar fraction separated out in this way was then dried, yielding 42 g of oil.

(9) The polar fraction was twice washed with petroleum ether (200 ml) and dried to constant weight with air, yielding 112 g of an orange-coloured solid.

(10) A 200 ppm (w/vol) solution of the said solid in acetonitrile was analysed by HPLC-MS, performed using a liquid chromatograph fitted with a Kinetex 1.7 XB-C18 100 1002.10 mm Phenomenex column, with a UV-PDA recorder interfaced with an ion trap spectrometer (LCQ Fleet Thermo Scientific; ESI ionisation method, positive/negative ions), with the following instrument conditions: Eluents: (A) 1% aqueous solution of HCOOH; (B) acetonitrile Gradient: 0 min (A/B=95/5), 5 min (A/B=90/10), 20 min (A/B=60/40), 30 min (A/B=10/90), 40 min (A/B=10/90), 45 min (A/B=95/5); Flow rate 0.4 ml/min; Detector: UV-PDA 280 nm.

(11) HPLC/UV analysis of the said solid revealed the presence of Arctiin (30% by weight with respect to the solid), the identity of which was confirmed from the mass spectrum (m/z 535 [M+H].sup.+). The total content of glucosides, all quantified as Arctiin, on the other hand corresponded to approximately 60% by weight with respect to the weight of the solid.

Example 2

(12) (Stage a)

(13) 200 kg of seeds of Cynara cardunculus were cleaned and screened.

(14) The said seeds with a water content of 7.5% by weight were fed to a single screw press (Mod. MIG PC25S screw diameter=110 mm; L/D=4.4) operating at 20 rpm at ambient temperature with a throughput of 60.9 kg/h, yielding 32.7 kg of oil and approximately 167 kg of a residue (containing the approximately 9% by weight of water). The said residue comprising de-oiled seeds had an oil content of approximately 10% by anhydrous weight.

(15) (Stage b)

(16) 1 kg of the obtained residue was then extracted with 1.6 kg of anhydrous ethanol under reflux for one hour in a reactor provided with a mechanical stirrer. The suspension obtained was filtered and washed with ethanol (200-300 ml). The solid phase was dried overnight in a stove at a temperature of 80 C.

(17) A solid residue weighing 790 g was obtained.

(18) (Stage c)

(19) The ethanol extract (liquid phase) obtained from stage (b) was dried in a rotary evaporator yielding 240 g of an orange-coloured pasty solid which was then extracted with hexane (300-400 ml) at ambient temperature in a reactor fitted with a mechanical stirrer so as to obtain a homogeneous dispersion which was then filtered.

(20) The polar solid fraction was twice washed with hexane (200 ml) and dried to constant weight with air, yielding 106 g of an orange-coloured solid.

(21) The hexane liquid fractions were then collected and dried, yielding 98 g of oil.

Examples 3-6

(22) (Stage a)

(23) 200 kg of seeds of Cynara cardunculus were cleaned and screened.

(24) The said seeds with a water content of 7.5% by weight were fed to a single screw press (Mod. MIG PC25S screw diameter=110 mm; L/D=4.4) operating at 34 rpm at ambient temperature with a throughput of 58.5 kg/h, yielding 41.3 kg of oil and approximately 157 kg of a residue (containing the approximately 8.5% by weight of water). The said residue comprising de-oiled seeds had an oil content of approximately 6% by anhydrous weight.

(25) (Stage b)

(26) Four polar organic solvents (anhydrous ethanol, hydrous ethanol (i.e. 95% ethanol and 5% water), isopropanol, acetone) were each tested for the extraction of 1 kg of the de-oiled residue obtained in stage a) as describe above.

(27) Each extraction was performed under reflux for one hour in a reactor provided with a mechanical stirrer; the residue/solvent ratio was 1/2 by volume.

(28) The suspensions obtained were filtered and washed with the same solvent used for the extraction.

(29) The solid phases were then dried overnight as in Examples 1-2; the weights of the obtained solid residues are reported in Table 1.

(30) (Stage c)

(31) The liquid phase of Examples 3-6 were dried in a rotary evaporator; the pasty solid was then washed three times with hexane and subsequently dried. The total weight of solid active substances and of the oil recovered after evaporation of the hexane fractions are reported in Table 1.

(32) The active substances obtained in Examples 3-6 were analysed by HPLC-UV as described in Example 1. The analysis of the samples revealed the presence of Arctiin and Tracheloside; the identity of the latter was confirmed from the mass spectrum (m/z 551 [M+H].sup.+). The amounts of Arctiin and Tracheloside (both quantified as Arctiin) within the extracted active substances are reported in Table 1: the Tracheloside/Arctiin weight ratio is of approximately of 1:1 (about 0.9) in all Examples 3-6.

(33) The protein content of the extracted active substances was calculated by determining the nitrogen content (by means of Kjeldhal method) and multiplying the obtained value by the coefficient 6.25.

(34) TABLE-US-00001 Extracted Active substances Solid Pro- Ex- residue Oil Tot. tein Arctiin Tracheloside ample Solvent (g) (g) (g) (g) (g) (g) 3 anhydrous 837 56 107 2.2 29.5 28.7 ethanol 4 ethanol 812 47 140 2.5 32.4 31.6 95%* 5 iso- 849 61 77 1.1 32.1 30.3 propanol 6 acetone 871 56 58 1.1 22.5 20.7 *water content of 5% by weight relative to the total weight of solvent.

(35) As can be seen from Table 1, isopropanol allowed to extract Arctiin with a purity of about 42% by weight (32.1 g over 77 g tot. active substances), i.e. a purity higher than that attained with other solvents.

Example 7

(36) (Stage a)

(37) 200 kg of seeds of Carthamus tinctorius were cleaned and screened.

(38) The said seeds having a water content of 7% by weight were fed to the same screw press of Examples 1-6, operating at 20 rpm at ambient temperature with a throughput of 49.7 kg/h, yielding 64.1 kg of oil and approximately 135.2 kg of a residue (containing the approximately 9.7% by weight of water). The said residue comprising de-oiled seeds had an oil content of approximately 11.5% by anhydrous weight.

(39) (Stage b)

(40) 1 kg of the de-oiled residue so obtained was then extracted with anhydrous ethanol (residue/ethanol ratio=1/1.6 by weight) under reflux for one hour in a reactor provided with a mechanical stirrer. The suspension obtained was filtered and washed with ethanol (200-300 ml). The solid phase was dried overnight in a stove at a temperature of 100 C.

(41) A solid residue weighing 864 g having a protein content of 21.5% by weight was obtained.

(42) (Stage c)

(43) The ethanol extract (liquid phase) obtained from stage (b) was dried in a rotary evaporator yielding a pasty solid which was then extracted with hexane at ambient temperature in a reactor fitted with a mechanical stirrer. The obtained emulsion was centrifuged at 3000 rpm for 10 minutes.

(44) The apolar fraction separated out in this way was then dried, while the polar solid fraction was washed with hexane and dried to constant weight with air, yielding 24.9 g of a yellow solid. The hexane fraction was also dried in the same manner, yielding 101 g of oil.

(45) The HPLC-MS analysis of the solid revealed the presence of Arctiin, Tracheloside and of the serotonin derivatives N-(p-coumaroyl)serotonin glucoside (m/z 485 [M+H].sup.+) and N-feruloylserotonin glucoside (m/z 515 [M+H].sup.+). According to the HPLC-UV analysis, the solid contained about 0.75% by weight of Arctiin and 16% by weight of Tracheloside with respect to the weight of the solid.