Economical process for the isolation of functional protein from plants

Abstract

The invention relates to a method for obtaining a protein from a plant material, wherein the method comprises the steps of i) mechanically disrupting the plant cells to obtain a plant juice in the presence of a reducing agent, ii) treating the plant juice to cause aggregation of chloroplast membranes, iii) removing the aggregated chloroplast membranes by precipitation and/or microfiltration, iv) subjecting the plant juice to ultrafiltration, and v) subjecting the soluble plant protein concentrate to hydrophobic column adsorption to remove residual chlorophyll, phenolic compounds and off-odors in a single column passage. The present invention also pertains to an apparatus and system for plant protein isolation based on this method. The isolated proteins can be economically obtained in batch scale and in large scale. Further, the invention is directed to a protein obtained by the method of the invention, a food product comprising thereof, and a use thereof.

What is claimed: 
     
         1 . A method for isolating a soluble plant protein from a plant material on an industrial scale, wherein the method comprises the steps of:
 i) mechanically disrupting the plant cells of said plant material to obtain a plant juice, wherein before, during, or after the step of disrupting the plant cells an extraction composition comprising a reducing agent is added to said plant material, said plant cells and/or said plant juice,   ii) treating the plant juice to cause aggregation of chloroplast membranes,   iii) separating said aggregated chloroplast membranes from the soluble plant protein in said treated plant juice by precipitation and/or microfiltration to provide a plant juice supernatant or plant juice permeate comprising the soluble plant protein,   iv) subjecting the plant juice supernatant or plant juice permeate to ultrafiltration, optionally in diafiltration mode, to provide a soluble plant protein concentrate, preferably comprising 25-50 wt % of protein, wherein said soluble plant protein concentrate is essentially free of salts and phenolic compounds, and   v) subjecting the soluble plant protein concentrate to hydrophobic column adsorption to thereby separate in a single column passage the residual chlorophyll, phenolic compounds and off-odors from the soluble plant proteins to provide a column permeate comprising isolated soluble plant protein that is essentially free of chlorophyll and off-odors,   
       and wherein said method further optionally comprises a steps of:
 vi) drying the said column permeate comprising soluble plant protein to provide a powder of functional plant protein, preferably said drying is performed by lyophilisation or spray drying, and/or 
 (vii) regenerating the hydrophobic adsorption column, wherein the regenerating is preferably accomplished by the use of ethanol as an eluent of column-adsorbed compounds. 
 
     
     
         2 . A method according to  claim 1 , wherein the extraction composition comprising a reducing agent, further comprises at least one selected from a divalent ion source, water and a buffering agent, and a combination thereof, preferably wherein said chelating agent, buffering agent and/or divalent ion source is food grade, more preferably wherein said divalent ion source is calcium chloride. 
     
     
         3 . A method according to any of  claim 1  or  2 , wherein the reducing agent is metabisulfite. 
     
     
         4 . A method according to any one of the proceeding claims, wherein said plant juice after addition of said extraction composition comprises a dry matter content of up to 10 wt % based on the total weight of the plant juice. 
     
     
         5 . A method according to any one of the proceeding claims, wherein said treating step involves heating the plant juice to a temperature of between about 40° C. to 60° C., preferably wherein after the heating said the heated juice is cooled by forced cooling, and/or wherein said treating step involves the use of an extraction comprising a divalent ion in an amount that causes flocculation of chloroplast membranes and allowing the plant juice to flocculate. 
     
     
         6 . A method according to any one of the proceeding claims, wherein said precipitation involves centrifugation of the heated and optionally cooled juice and/or the divalent ion-flocculated juice to provide a pellet of aggregated chloroplast membranes and a plant juice supernatant comprising the soluble plant protein, or wherein said microfiltration comprises filtering the heated and optionally cooled juice and/or the divalent ion-flocculated juice through a filter having a pore size in the range of 0.1-0.5 μm, preferably a pore size that retains membranes, chlorophyll, tannin, virus, bacteria and/or aggregates thereof from said juice while allowing passage of soluble proteins. 
     
     
         7 . A method according to any one of the proceeding claims, wherein hydrophobic column adsorption comprises the use of a column packed with a hydrophobic adsorptive resin, preferably a non-ionic crosslinked polystyrene resin, even more preferably a macroreticular styrene-divinylbenzene copolymer matrix, still more preferably Amberlite™ XAD 16 resin. 
     
     
         8 . A method according to any one of the proceeding claims, wherein the steps i), iii), iv), and v), and preferably also vi), are performed under low temperature, preferably a temperature in the range of about 0-15° C., preferably the temperature in the range of about 1-10° C., and preferably the temperature in the range about 2-5° C. 
     
     
         9 . A method according to any one of the proceeding claims, wherein the processing time to complete process steps i)-v), and preferably also vi), is no more than 1 day, preferably wherein the processing time from harvest of the plant material to completion of process step v), and preferably also vi), is no more than 1 day. 
     
     
         10 . A method according to any one of the proceeding claims, wherein the protein is ribulose 1,5-diphosphate carboxylase oxygenase (Rubisco). 
     
     
         11 . A method according to any one of  claims 2 - 10 , wherein the extraction composition comprises a divalent ion source and wherein step iii) comprises separating said aggregated chloroplast membranes from the soluble plant protein in said heated plant juice by precipitation to provide a plant juice supernatant comprising the soluble plant protein. 
     
     
         12 . A protein isolate obtained by a method according to any one of the proceeding claims, preferably wherein said protein isolate comprises low amounts of tannin. 
     
     
         13 . A food product comprising a protein isolate according to  claim 12 . 
     
     
         14 . Use of the protein isolate according to  claim 12  as a thickening agent, a foaming agent, an emulsifier, and/or a texturizing agent.