Harnessing the potential of green leaves : agricultural biomass as a source of sustainable food protein

Demand for sustainable protein-rich food sources is currently increasing to meet the nutritional requirements of a growing population, while also considering climate challenges. Green leafy biomass (GLBM), in the form of side-streams and main crops, is a widely available protein source with potential food value. Extended use of GLBM, e.g. through a biorefinery process targeting leaf protein concentrates (LPCs), could add direct values, e.g. economic revenues from side-stream valorisation, or indirect values, e.g. reduced greenhouse gas emissions from higher resource utilisation. In this thesis, several types of GLBM were successfully subjected to an extraction protocol targeting water-soluble proteins, although the outcomes, e.g. yield, differed significantly between GLBM types. The major protein component in LPC was the enzyme RuBisCO. A pre-feasibility assessment revealed insufficient recovery rates on upscaling the process. To achieve economic viability, further process development is needed and additional compounds and products should be targeted. The use of LPCs in food applications is of interest due to their nutritional aspects, i.e. high protein content and good amino acid profile. Another area of interest is their potential as a functional ingredient, e.g. their foam stabilising ability, which was demonstrated for LPCs from several GLBM types in this thesis. Air-water interfacial properties, which can serve as an indicator of foam stabilising capacity, did not differ significantly between LPCs from the GLBM types evaluated. Further, no major differences in interfacial properties were observed between the LPCs and egg white. Green leafy biomass can be viewed as a valuable resource with great potential and extending the use of GLBM through LPC production could contribute to a more sustainable food production system

White proteins from green leaves in food applications

This introductory paper examines the current understanding of how green leaves could be utilized as a food source, and the importance of proteins in food structures. Green leaves have long been considered as a possible protein source for sustainable food and feed production. Proteins in green leaves can be divided into a white and green protein fraction. The white protein fraction is mainly RuBisCO, or ribulose-1,5-bisfosfat-caroboxylase/oxygenase, which has been called the most abundant protein in the world, while the green fraction consists of chlorophyll related proteins. A selection of reported leaf protein extraction methods are presented in this paper. Generally the first step of the process is the extraction of green juice, followed by a removal of the green protein fraction and a concentration and purification of the white protein fraction. The functional properties of proteins as gelling agents, emulsifiers and foam stabilizers are of great importance in many food systems. One example is the interfacial properties of a protein, which control the stabilization of foams and emulsions. In this paper the role of proteins in gels, emulsions and foams are reviewed, with emphasis on foams. Some of the methods and techniques used to observe and quantify these functional properties are mentioned, including imaging techniques and surface tensiometry analyses. Some methods used to assess the properties of the food structures are also presented

Protein Fractionation of Green Leaves as an Underutilized Food Source—Protein Yield and the Effect of Process Parameters

Green biomass has potential as a sustainable protein source for human consumption, due to its abundance and favorable properties of its main protein, RuBisCO. Here, protein fractionation outcomes of green leafy biomass from nine crops were evaluated using a standard protocol with three major steps: juicing, thermal precipitation, and acid precipitation. Successful protein fractionation, with a freeze-dried, resolubilized white protein isolate containing RuBisCO as the final fraction, was achieved for seven of the crops, although the amount and quality of the resulting fractions differed considerably between crops. Biomass structure was negatively correlated with successful fractionation of proteins from biomass to green juice. The proteins in carrot and cabbage leaves were strongly associated with particles in the green juice, resulting in unsuccessful fractionation. Differences in thermal stability were correlated with relatedness of the biomass types, e.g., Beta vulgaris varieties showed similar performance in thermal precipitation. The optimal pH values identified for acid precipitation of soluble leaf proteins were lower than the theoretical value for RuBisCO for all biomass types, but with clear differences between biomass types. These findings reveal the challenges in using one standard fractionation protocol for production of food proteins from all types of green biomass and indicate that a general fractionation procedure where parameters are easily adjusted based on biomass type should instead be developed

Protein Fractionation of Leafy Green Biomass at the Pilot Scale: Partitioning and Type of Nitrogen in the Fractions and Their Usefulness for Food and Feed

Fractionation of green biomass often results in fractions with insufficient protein content or quality for food or feed. To understand ways forward, we evaluated the fate of nitrogen (N) and the food or feed suitability of six pilot-scale fractions. The N was present mainly as amino acids (AA) in all fractions (<87%), however, the protein was partly degraded or insoluble in the majority of samples. All protein types and AAs traveled similarly through the fractionation process, giving insignificant separation of RuBisCO versus other proteins, and essential versus nonessential AAs. Water-soluble N compounds were enriched in juice fractions (90-95%), while the protein fractions contained the highest insoluble protein content (13-17%). AA composition in pulp and green juice verified their suitability as feed for ruminants and pigs, respectively. Fractionation of green biomass for food and feed is indeed important, although for sustainable industrial applications, further evaluations are required regarding process feasibility, antinutritional components, and brown juice uses

Applicability of leaf protein concentrates from various sources in food: Solubility at food-relevant pH values and air-water interfacial properties

Leaf protein concentrates (LPCs) from green leafy biomass have potential as a sustainable protein source. Here, protein composition, solubility, foamability, and interfacial properties of LPCs from six different biomass types (beetroot, kale, lucerne, mangold, spinach, and sugarbeet) were evaluated. Although the LPCs showed some differences in protein composition and solubility, the foamability and interfacial properties were strikingly similar. RuBisCO was the major protein in all LPCs. All LPCs demonstrated lower solubility at pH 5.0 and 3.0 than at pH 7.0. Both the highest and the lowest solubility was found for sugarbeet: 67.6 ± 7.9% at pH 7.0 and 9.2 ± 4.0% at pH 5.0. The LPCs formed aggregates of increasing size at pH 3–4.5, matching the solubility results. The LPCs stabilized foams in a preliminary foaming test, although the main focus was on evaluating air-water interfacial properties. The ability of the proteins to diffuse to and adsorb at the interface and their ability to form viscoelastic films thereat, using an optical tensiometry method, was investigated. All six LPCs showed an interfacial behavior resembling that of reference egg white protein (0.1–1.0 mg/ml). Our findings indicate good opportunities for using LPCs in food applications and in particular as an alternative to egg white in foams

Protein fractionation of broccoli (Brassica oleracea, var. Italica) and kale (Brassica oleracea, var. Sabellica) residual leaves — A pre-feasibility assessment and evaluation of fraction phenol and fibre content

This pre-feasibility study evaluates the use of residual leafy green biomass from broccoli (Brassica oleracea, var. Italica) and kale (Brassica oleracea, var. Sabellica) as feedstock for protein fractionation and potential application of the fractions in food and feed products. The protein concentration, protein recovery potential and the content of phenols and dietary fibre in these biomass sources and fractions were investigated. Field produce and side-stream analysis showed that among broccoli and kale side-streams the potentially suitable leaves for protein fractionation constitute up to 16 and 1.9 t/ha (DM content), respectively. Fractionation demonstrated that between 34–42 and 25–34 kg total protein could be extracted per t DM of broccoli and kale residue leaves, respectively. The amount of protein was generally high in green protein fraction (GPF) and the white protein concentrate (WPC) of both crops, although significantly higher in broccoli compared to kale. The recovery of bound and free phenolic compounds was up to 18% in the GPF of both crops, while only 0.4% ended up in the WPC. The economic assessment showed that the feedstock and processing costs of producing GPF and WPC, as well as of the combined protein fraction (CPF) 1.9–6.0 and 1.3–3.9 times higher than expected revenues for broccoli and kale, respectively, indicating that the production of protein fractions is not economically feasible with the current production scheme. However, potentially higher revenues may be obtained if value-added products such as fractionated phenols and dietary fibre components are also included and investigated in future production schemes. The pathway investigated, that included a direct drying and milling of leaf biomass showed a low processing cost and thereby the most favourable economic alternative, with approx. 7–30% profit for kale, while for broccoli revenues covered only 44–47% of the costs due to the extra harvest cost of the broccoli leaves