Classifications

• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
  • A23J1/00—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites

Definitions

• the present invention relates to a method and an apparatus for precipitating" protein from an aqueous protein-containing liquid.
• the term "precipitation” is intended to designate an operation by which protein which is present in. a liquid in dissolved form or in the form of such small particles that it cannot be separated by any filtration or normal industrial cen- trifugation technique, is brought into a form which can be sepa ⁇ rated by filtration and normal industrial centrifugation techniques .
• the term “precipitation” covers such phenomena as flocculation , coagulation , conglomeration , etc . , provided that these phenomena or combinations thereof will convert the protein contained in the liquid into a flocculate, coagulate, conglomerate, etc. which may easily be separated from the liquid by filtration or centrifugation .
• a special phenomenon covered by the term “precipitation” in the present context is the attraction of lipophilic proteins to oil droplets, which droplets with attracted protein may, if desired, be separated by centrifugation .
• the term "protein” is intended to designate not only proteins according to a strict chemical interpretation of that term, but also proteinaceous substances which, in addition to protein-characterizing moieties , may contain other chemical princi ⁇ ples .
• One major utilization of the present invention is in the field of separation of proteins and proteinaceous substances from aqueous liquids of vegetable origin, and one typical type of pro- tein precipitated by the process of the invention is the "chloro ⁇ plastic fraction" of the vegetable protein , that is , a fraction com ⁇ prising ruptured and unruptured chlorophyll granules and the single components associated therewith , including proteins , hydro ⁇ carbons , lipids , chlorophyll, carotinoids , etc .
• this chloroplastic fraction which has green colour, is termed "green protein” .
• the main aspect of the present invention concerns the precipitation of protein by heating.
• Precipitation by indirect heating incurs problems involving coagu ⁇ lation of the proteins on the heating surfaces with consequent reduced heat transfer and destruction of the protein burnt onto the heating surfaces .
• Precipitation by steam injection introduces large amounts of condensate which, again, requires large amounts of energy for the subsequent removal thereof, and in steam in ⁇ jection, it is generally difficult to control the temperature.
• Espe- cially in connection with the utilization of press juice from green plants as protein source which is a utilization of increasing im ⁇ portance, also for the production of proteins for human consump ⁇ tion, considering that compared to any other agricultural utili ⁇ zation, the highest protein yields per area are obtained by cul- tivation of green plants, e.g.
• the green chloroplast- containing fraction is a valuable product as animal feed, but is not immediately suitable for human consumption because of its bitter taste. If the green protein is not effectively separated from the juice, the quality and pro ⁇ perties of that fraction of the proteins contained in the juice which is suitable for human consumption, the so-called "white protein” , will be deteriorated. However, the green protein is especially difficult to handle technically as it shows an excessive tendency, even at very moderate temperature, to clot and deposit as a rubber-like substance on heating surfaces , thus virtually rende ⁇ ring the further use of the equipment in question impossible.
• an effective precipitation of proteins from an aqueous protein-containing solution is obtained under controllable conditions, avoiding deposit problems and per- mitting desired fractionation of the proteins contained in the pro ⁇ tein-containing liquid.
• the method of the invention comprises con ⁇ tacting the aqueous protein-containing liquid with a fluid which is immiscible with the said aqueous protein-containing liquid, main ⁇ taining the contact under conditions sufficient to cause precipi- tation of protein from the said protein- containing liquid, separating the aqueous phase from the said fluid, and, if desired, separating the precipitated proteins from the aqueous phase and/or the fluid phase.
• the aqueous protein-containing liquid subjected to the method of the invention is typically a green juice from vegetables as dis ⁇ cussed above, but it is also within the scope of the invention to precipitate protein from other aqueous protein- containing liquids where separation or fractionation of protein or a fraction thereof may present similar problems , including fermentation broths con ⁇ taining microorganisms (bacteria or fungi) , blood, whey, and waste water from starch production and from other processes processing cereals or vegetables .
• Aqueous protein-containing liquids for which the method of the invention is particularly suited are liquids where coagulation of protein occurs very readily even at a relatively low temperature such as , e.g. , 40 C .
• a fluid which is immiscible with the aqueous protein- containing liquid is intended to designate a fluid which, when contacted with the aqueous protein-containing liquid under the conditions prevailing during the contact, will not to any substantial extent become dissolved in the aqueous phase of the protein-containing liquid, but will, in contrast, exist as a separate phase different from the aqueous phase of the protein- containing aqueous liquid.
• the fluid is typically an edible oil or an edible fat which is fluid under the conditions prevailing during the contact, but also fluids (for example, water-immiscible inert non- toxic ingestible oils or fats such as paraffin oil) which may not normally be considered edible are suitable for the purpose of the present invention, provided that they are not of a character which would incur risk of contamination of the end products with con ⁇ stituents that are physiologically unacceptable.
• fluids for example, water-immiscible inert non- toxic ingestible oils or fats such as paraffin oil
• the fluid is suitably maintained at an elevated temperature which is effective to cause the desired degree of precipitation of proteins from the said protein-containing liquid, but typically, the temperature will not be so high as to result in a product having any "burnt” or "fried” character; on the contrary, the product precipitated by the method of the in- vention is preferably denatured to as low a degree as possible, thus rendering the product available for desired subsequent processing.
• the temperature of the fluid will normally be below the boiling point of the aqueous protein -containing liquid, and typical temperatures of the fluid are in the range of 60 - 95 C, most often 70 - 85 C.
• the conditions sufficient to cause precipitation of protein from the protein-containing liquid are primarily represented by the combi- nation of the temperature of the fluid, the ratio of the amount of fluid to the amount of aqueous liquid contacted therewith, and the time period during which the contact is maintained.
• the pH of the fluid the ratio of the amount of fluid to the amount of aqueous liquid contacted therewith, and the time period during which the contact is maintained.
• other factors will influence the precipitation, such as the pH of
• OMPI the aqueous protein-containing liquid, the degree of agitation during the contact, and the affinity between the fluid and the protein .
• the desirable pH of the aqueous pro ⁇ tein-containing liquid will depend on the nature of the liquid and may vary within relatively wide limits . When the liquid is green juice from plants such as lucerne or clover, a pH in the neutral range (5 - 8) will often be most suitable .
• the contact between the fluid and the aqueous liquid is performed by maintaining the fluid as a stationary layer in a reaction vessel and passing the protein- containing liquid through the layer dropwise or as a fine jet.
• the layer will tj ically be the top layer of a 2- ⁇ hase system where the lower layer is constituted by an aqueous phase where the protein-containing liquid with precipitated protein collects .
• the aqueous protein- containing liquid is suitably contacted with the fluid by being added dropwise or as a fine jet on top of the fluid phase.
• the droplets of the liquid When the fluid phase is kept under laminar conditions , that is , with slight or moderate agitation, the droplets of the liquid will pass substantially vertically through the layer, and if the walls of the reaction vessel are vertical at the levels where they contact the fluid layer, any contact between the protein-containing liquid and the walls of the vessel is minimized, thus minimizing deposits on the walls .
• the depth of the fluid layer is adapted to obtain satis- factory precipitation of the desired fraction during the passage of the droplets of the protein -con taining liquid through the layer. Normally, this depth will be between 10 and 50 cm . It is suitable to distribute the droplets of the aqueous liquid substantially uni ⁇ formly over the surface of the fluid layer. This may be obtained, e . g. , by means of suitable spraying devices or distributing devices with perforated bottoms .
• the fluid has a density which is higher than that of the aqueous liquid
• the fluid is suitably arranged on top of a distributing unit generating droplets of the aqueous liquid.
• the droplets of the aqueous liquid will then pass upwardly through the fluid layer, and the aqueous phase will collect on top of the fluid layer after having passed the layer.
• the aqueous protein-containing liquid constitutes a stationary phase through which the heated fluid is passed.
• the fluid has a lower density than the aqueous liquid, it is introduced from the bottom of the aqueous phase.
• the aqueous phase is separated from the fluid.
• the separation may be performed in a manner known per se for separating phases which are mutually immiscible, including allowing the phases to separate effectively by gravity and withdrawing one of the phases .
• the precipitated proteins Depending upon the character of the precipitated proteins, they will collect in the aqueous phase or in the fluid phase, or in both. In the above-discussed precipitation of green proteins from vege ⁇ table juice, the predominant proportion of the green protein will follow the aqueous phase, although a minor proportion thereof will sometimes collect in the fluid phase.
• the separation of the precipi ⁇ tated protein from the aqueous phase and/or from the fluid phase is performed by filtration, decantation, centrifugation, etc. , in a manner known per se .
• the heating medium is a liquid, it is very easy to control the temperature to a level which is optimal for the protein fraction in question . Furthermore, any problems arising from "burning" of proteins onto heating surfaces are obviated as the heat transfer takes place at the interface between the heating fluid and the
• OM aqueous liquid OM aqueous liquid.
• the heat transfer is easy to control, in particular when the heating fluid is maintained as a stationary layer where the temperature and thickness of the layer can be optimally ad ⁇ justed.
• the green proteins tend to have a lipophilic cha ⁇ racter in the contact with the fluid.
• the precipitated green protein collects at the interface be ⁇ tween the . aqueous droplets and the heating fluid.
• a heated fluid e.g. a heated fat or oil
• a product consisting of oil droplets surrounded by attracted proteins will be obtained.
• This embodiment of the method of the invention will not necessarily incur coagulation of the proteins . Rather, the desired effect in certain cases is an effect where the lipophilic character of the proteins is utilized to "extract" the proteins from the aqueous phase onto or into the oil droplets .
• the "cream-like" product ob ⁇ tained in this manner may be used for human or animal consump ⁇ tion, depending upon the identity of the oil and the proteins .
• the cream phase may be used per se, or may be redispersed in water or other suitable liquid. Depending upon the identity of the oil and the proteins in the aqueous protein -containing liquid, it may or may not be necessary to perform any heating of the oil passed into the aqueous liquid .
• microorganisms may, e. g. , utilize nutrients in waste water or waste products . It has been a great problem to separate these microorganisms after completion of their cultivation, the reason for this being that the concentration of the microorganisms is so small and/or the microorganisms themselves are so small that normal filtering equipment cannot be used. Furthermore, the difference between the density of the microorganisms and the density of the fermentation liquid in which they are present is usually so small that it is extremely difficult to obtain effective separation by cen ⁇ trifugation.
• Such microorganism-containing fermentation liquid is an interesting aqueous protein-containing liquid subjected to the me ⁇ thod of the present invention .
• the filtrate after removal of the pre ⁇ cipitated green protein therefrom, is a liquid contain a high pro ⁇ portion of. protein which can be separated and used for human consumption, the so-called "white protein" , which, when uncon- taminated with green protein, is a substantially tasteless and odourless protein of optimum nutritional composition.
• This white protein can be separated from the filtrate by acidification, heating, etc. , the separation by acidification, e. g. , to pH 3, presently being preferred because it results in a white protein fraction which is easily soluble.
• the white protein precipitated in this manner may be separated by centrifugation or filtration.
• the centrifugate or filtrate is extremely suitable as a substrate for fermentation of microorganisms or as an addition to substrates for the fermentation of microorganisms .
• the brown juice is also useful as a basis for "liquid supplement” (li ⁇ quid feed for ruminants) .
• the method of the invention is particularly attractive for this fractionated precipitation of protein from juice from green plants .
• the method does not introduce any undesired substances into the protein fractions , considering that any absorbed fat or oil in fact increases the nu- tritional value of the end products .
• Secondlj* * due to the effective temperature control rendered possible through the method of the invention, the lipophilic green chloroplast fraction may be preci ⁇ pitated at such low temperatures that only a minimum of the white protein fraction is entrained with the green protein fraction. At the same time, the green protein fraction may be removed so efficiently that the white protein fraction will be immediately useful for human consumption.
• green juices may have a character which makes them less suitable for the immediate fractionation by the method of the invention.
• a green juice which is less suitable for immediate treatment by the method of the invention is typically characterized by a somewhat elevated viscosity, but a general indicative expe ⁇ riment for assessing the suitability for the green juice for treat ⁇ ment by the method of the invention is to subject the green juice to high speed centrifugation (centrifugation at about 25.000 rpm. ) .
• the green juice is indicated as less suitable for treatment by the method of the invention .
• Such green juice can be rendered suitable for the treatment by the method of the invention by treatment with a base such sodium hydroxide to pH 8 - 10, e.g. for about half an hour, and subsequent readjustment of the pH, that is , to about 6. After such pH cycling, the green juice will be found to give a clear supernatant in the high speed cen ⁇ trifuge treatment ans will be suitable for the method of the in- vention .
• the protein-containing aqueous liquid is passed to a liquid distributor 1, exemplified as distributor vessel having a bottom 2 provided with uniformly spaced perforations .
• a liquid distributor exemplified as distributor vessel having a bottom 2 provided with uniformly spaced perforations .
• the droplets or thin jets passing through the perfo ⁇ rations enter into a precipitation vessel 3 having an upper part
• the vessel 3 contains an oil phase 4 consisting of, e . g. , vegetable oil or molten lard, etc. , located in the upper part of the vessel.
• the oil phase is heated to. a suitable protein precipitation temperature by means of a heating and ther ⁇ mostating unit 24 immersed in the oil phase, e.g. , a thermostating unit with pumping or stirring means for circulating the oil and ope ⁇ rating at a rate which will not induce turbulent conditions in the oil phase.
• the oil phase 4 floats on a water phase 5 which, at the upstart of the apparatus, may be tap water, but which eventually will be constituted by the aqueous phase of the droplets having passed the oil phase.
• a water phase 5 which, at the upstart of the apparatus, may be tap water, but which eventually will be constituted by the aqueous phase of the droplets having passed the oil phase.
• the droplets of the juice Upon passage through the oil phase, the droplets of the juice will substantially entrain precipitated green proteins which will then fall down through the aqueous phase 5 as the droplets "break" substantially immediately upon contact with the aqueous phase.
• the aqueous phase with precipitated protein is withdrawn, either continuously or at suitable intervals, through a valve 6 and is passed into a sedimentation vessel 7 in which no stirring is performed. In the sedimentation vessel 7, the aqueous phase divides into an upper, substantially clear liquid 8 and a sediment-containing lower aqueous phase 9.
• the aqueous phase 9 with sedimented green protein is passed through a pump 10 to a separating unit 11 such as a filter, a centrifuge, or a decanter.
• the protein separated in the separator 11 is passed to a dehy- drator 12, e .g. drum drier, a spray drier, or a fluid bed drier, and is withdrawn as dehydrated green protein concentrate through a conduit 13.
• the liquid 8 is passed through a conduit 14 to a filter 15 where any content of precipitated protein is separated from the liquid. From the separator 11, the liquid separated from the green protein is passed through a conduit 17 and is combined with the liquid 8 subjected to the filtration .
• the protein filtered off from the liquid in the filter 15 is passed through a conduit 18 to the pump 10 and is combined with the main sediment of green protein.
• the filtrate from the filter 15 is withdrawn through a conduit 19 as a white protein solution which can be further pro ⁇ Ded in a manner known per se to obtain white, protein and brown juice.
• the apparatus suitably further comprises . separating means (not shown) , such as filters or centrifuges , for separating precipitated protein from the oil phase.
• Green juice from lucerne, prepared by desintegration and subse ⁇ quent pressing in an industrial leaf protein concentrate-producing plant was frozen immediately after preparation and was thawn prior to this experiment.
• the juice was sprayed dropwise into a vessel containing an aqueous phase of a depth of about 2_ cm and, on top of the aqueous phase, a phase of molten lard of a depth of 12.5 cm.
• the lard was heated in a thermostat bath outside the reaction vessel, and from this bath, it was continuously recycled into the reaction vessel, thereby maintaining the depth of the lard phase and controlling its temperature to 70 C .
• the temperature of the juice added was 5 C. After passage of all the juice through the fat layer, the temperature of the water phase was 38 - 39 C .
• the water phase with precipitated green protein was sucked off and filtered.
• the filtrate was heated to 58°C which resulted in the precipitation of a voluminous white precipitate. After removal of this precipitate by filtration, the filtrate was heated to about 95 C, and an additional white protein coagulate was obtained.
• the white protein was precipitated by passing the filtrate from the removal of the green protein through the lard layer heated to 85 C .
• the properties of the juice were as ollows :
• the aqueous phase contain ⁇ ing precipitated protein had a temperature of 40 C .
• 770 ml of the aqueous phase containing protein sediment was withdrawn and was centrifugated at 2000 rpm . for 10 minutes in a laboratory centri- fuge.
• the sedimented green protein was separated as a green sludge (78.8 g) by this centrifugation .
• the green sludge contained 12.8% of dry matter including 5.75% of crude protein.
• the dry matter content in the green sludge corre- sponds to 28.6% of the total dry matter in the original green juice, and the crude protein content in the green sludge corresponds to 48% of the total crude protein content in the original green juice.
• the supernatant was subjected to acid precipitation bj ⁇ addition of HCl to pH 3 and centrifugation at 2000 rpm . for 10 minutes in a laboratory centrifuge and was thereafter freeze-dried.
• the yield of the white protein was 3.83 g having a dry matter content of 89.6%, corresponding to 10.5% of the total dry matter content in the ori- ginal green juice.
• the content of crude protein in the white pro ⁇ tein was 47.8%, corresponding to 20% of the crude protein content in the original green juice.
• a super ⁇ natant of 710 g of brown juice having a dry matter content of 2.8% was obtained.
• the dry matter content of the brown juice was 60.9% of the total dry matter content of the original green juice .
• the brown juice contained 0.4% of crude protein, corresponding to 32% of the total crude protein content in the original green juice .
• a "white protein" fraction (slightly green) having a dry matter content corresponding to 12.8% of the dry matter content of the original green juice and a crude protein content of 7.0%, corres ⁇ ponding to 20% of the crude protein content of the original green juice,
• a brown juice having a dry matter content corresponding to 56% of the dry matter content of the original green juice and a crude protein content of 0.47%, corresponding to 34% of the crude protein contained in the original green juice.
• Fresh green juice obtained from lucerne by mincing in a mincing machine and subsequent pressing in a juice press , was used as starting material.
• the properties of the juice were as follows : Dry matter content: 8.39% by weight
• the green sludge contained 9.4% of dry matter and 4.9% of crude protein.
• the dry matter content in the green sludge corresponds to 34% of the total dry matter in the original green juice
• the crude protein content in the green sludge corresponds to 44% of the total crude protein content in the original green juice .
• the supernatant was subjected to acid precipitation by addition of HCl to pH 3 and standing for 20 hours at 5 C .
• a pure white pro ⁇ tein precipiate was obtained.
• the white protein was separated by centrifugation at 2000 rpm for 10 minutes in a laboratory centri ⁇ fuge and ⁇ vas thereafter freeze-dried.
• the yield of white protein was 3.74 g having a dry matter content of 95%, corresponding to 15% of the total dry matter content in the original green juice.
• the content of crude protein in the white protein was 49.3%, corres ⁇ ponding to 20% of the crude protein content in the original green juice.
• a supernatant of 619 g of brown juice having a dry matter content of 1.97% was obtained.
• the dr 1 " matter content of the brown juice was 51% of the total dry matter content of the original green juice .
• the brown juice contained 0.56% of crude protein, corresponding to 36% of the total crude protein content in the original green juice.
• Example 2 In the same manner as described in Example 2, a series of further experiments were performed. In all the experiments , the green juice added had room temperature, and after passage of all of the green juice through the oil phase, the aqueous phase had a tempe ⁇ rature in the range of 40 - 44 C. Examples 4 - 8 were performed with green juice from the same batch and illustrate the influence of the oil phase temperature on the yield of white and green proteins. The results appear from Table I below where all percentages are by weight, and where the term "% tot. orig. " designates percent by weight of the total amount of the substance in question in the original fresh green juice:

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• 1981
  • 1981-01-02 EP EP19810900222 patent/EP0042856A1/de not_active Withdrawn
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