Brown juice is a byproduct of fractionated green biomass during leaf protein isolation. It represents approximately 45%–50% of the total pressed fresh biomass. Disposal of brown juice is a serious issue in leaf protein production due to its high biological oxygen demand and carbohydrates content. The current study aimed to find a possible potential use of brown juice. Therefore, chemical and biochemical properties of brown juice—derived from alfalfa green biomass—were determined before and after fermentation by lactic acid bacteria. Additionally, the growth stimulation potential of fermented brown juice on plumed cockscomb (Celosia argantea var. plumose ‘Arrabona’) plants were tested. Celosia seedlings were sprayed at different rates of fermented brown juice (i.e., 0.5%, 1%, 2.5%, 5%, and 10%) and tap water was applied as control. The results revealed that lactic acid bacteria successfully enhanced the stabilization of brown juice via reducing sugars content and increasing organic acids content. After fermentation, contents of glucose monomers were 15 times lower; while concentrations of lactic and acetic acids increased by 7- and 10-fold, respectively. This caused a reduction in the pH of fermented brown juice by 13.9%. Treating Celosia plants at lower rates of fermented brown juice (up to 1.0%) significantly induced their growth dynamics and antioxidant capacity. Higher values of vegetative parameters were measured in treated plants compared to control. The brown juice treatments caused significant changes in histological parameters as well. The activity of catalase and peroxidase increased in plants that received fermented brown juice especially at low rates. Moreover, an increase in water-soluble protein and phenol was measured in different tissues of plants sprayed with fermented brown juice. Malondialdehyde content was lowered in treated plants compared to control. Fermented brown juice at high rates slightly reduced the amount of photosynthetic pigments; however, this reduction was not reported for low rates of fermented brown juice. These results surely illustrate the potential use of fermented alfalfa brown juice as a growth stimulator for crops particularly at rates below 2.5%.

The main objective of this study was to increase the economic value of broccoli green agro-waste using three wet fractionation methods in the shadow of green biorefinery and the circular economy. Product candidates were obtained directly by using a mechanical press, and indirectly by using microwave coagulation or via lactic acid fermentation of green juice. The leaf protein concentrates (LPC) fractions displayed significantly higher dry matter content and crude protein content (34–39 m/m% on average) than the green juice fraction (27.4 m/m% on average), without considerable changes in the amino acids composition ratio. UHPLC-ESI-ORBITRAP-MS/MS analysis showed that kaemferol and quercetin are the most abundant flavonols, forming complexes with glycosides and hydroxycinnamic acids in green juice. Lacto-ermentation induced a considerable increase in the quantity of quercetin (48.75 μg·g⁻¹ dry weight) and kaempferol aglycons (895.26 μg·g⁻¹ dry weight) of LPC. In contrast, chlorogenic acid isomers and sulforaphane disappeared from LPC after lactic acid fermentation, while microwave treatment did not cause significant differences. These results confirm that both microwave treatment and lacto-fermentation coagulate and concentrate most of the soluble proteins. Also, these two processes affect the amount of valuable phytochemicals differently, so it should be considered when setting the goals.

A pot experiment, under greenhouse conditions, was carried out aiming at investigating the agronomic biofortification of alfalfa (Medicago sativa L.) with Se and monitoring the Se uptake and accumulation dynamics within four consecutive harvests within the same growing season. Two ionic Se forms, i.e., sodium selenate (Se (VI)) and sodium selenite (Se (IV)), were applied once at a rate of 1, 10, and 50 mg kg⁻¹ (added on Se basis), while 10 and 50 mg L⁻¹ of a red elemental Se (red Se⁰) were used; all Se treatments were added as soil application. Application of Se (VI) at the rate of 50 mg kg⁻¹ was toxic to alfalfa plants. The effect of Se forms on Se accumulation in alfalfa tissues, regardless of the applied Se concentration, follows: Se (VI) > Se (IV) > red Se⁰. The leaf, in general, possessed higher total Se content than the stem in all the treatments. The accumulation of Se in stem and leaf tissues showed a gradual decline between the harvests, especially for plants treated with either Se (VI) or Se (IV); however, the chemically synthesized red Se⁰ showed different results. The treatment of 10 mg kg⁻¹ Se (VI) resulted in the highest total Se content in stem (202.5 and 98.0 µg g⁻¹) and leaf (643.4 and 284.5 µg g⁻¹) in the 1st and 2nd harvests, respectively. Similar tendency is reported for the Se (IV)-treated plants. Otherwise, the application of red Se⁰ resulted in a lower Se uptake; however, less fluctuation in total Se content between the four harvests was noticed compared to the ionic Se forms. The Se forms in stem and leaf of alfalfa extracted by water and subsequently by protease XIV enzyme were measured by strong anion exchange (SAX) HPLC-ICP-MS. The major Se forms in our samples were selenomethionine (SeMet) and Se (VI), while neither selenocysteine (SeCys) nor Se (IV) was detected. In water extract, however, Se (VI) was the major Se form, while SeMet was the predominant form in the enzyme extract. Yet, Se (VI) and SeMet contents declined within the harvests, except in stem of plants treated with 50 mg L⁻¹ red Se⁰. The highest stem or leaf SeMet yield %, in all harvests, corresponded to the treatment of 50 mg L⁻¹ red Se⁰. For instance, 63.6% (in stem) and 38.0% (in leaf) were calculated for SeMet yield % in the 4th harvest of plants treated with 50 mg L⁻¹ red Se⁰. Our results provide information about uptake and accumulation dynamics of different ionic Se forms in case of multiple-harvested alfalfa, which, besides being a good model plant, is an important target plant species in green biorefining.

Fertilization management is a key issue in plant nutrition to produce plants with good quality and quantity. Deproteinized leaf juice or brown juice (BJ) is a by-product during the isolation of leaf protein from biomass crops such as alfalfa. The idea of using BJ as a biostimulant fits well in the aspect of circular economy since BJ is currently a problematic issue of the leaf protein production approach. Fractionation of one-kilogram fresh biomass results in approximately 500 cm³ BJ. Due to fast spoil of fresh BJ, if left on room temperature, it is found that fermentation of fresh BJ using lactic acid bacteria and reducing its pH increases its stability and storage on room temperature. In the present study, we examined the effect of fermented alfalfa BJ on vegetative, physiological, and anatomical properties of the versatile sweet basil (Ocimum basilicum L. ‘Bíborfelhő’) plants. Sweet basil seedlings were sprayed at different doses of fermented alfalfa BJ (i.e., 0.5%, 1.0%, and 2.5%) and tap water served as a control (0.0% BJ). The results revealed that foliar application of fermented alfalfa BJ significantly improved the biometrical features of sweet basil plants. Plants treated with fermented BJ showed significantly higher values of all the measured parameters compared to the control (0.0%), except for the number of leaves per plants where control plants (0.0%) had more leaves. However, the leaves of control plants (0.0%) were smaller than treated plants as data of leaf area showed. Fermented alfalfa BJ significantly increased the content of photosynthetic pigments (chl a and chl b), relative chlorophyll (SPAD value), lengths of stem and root, fresh masses of stem, root, and leaves, volumes of stem and root, and leaf area. Despite all rates of fermented BJ displayed higher values over control plants (0.0%), the rate of 0.5% was the best one supported by results. Application of fermented alfalfa BJ influenced the anatomical parameters as well. These findings demonstrate the possible use of fermented alfalfa BJ as a promising novel plant biostimulant.

ALFALFA is considered as “the Queen of the Forages” due to its high protein content and nutritional value. The production of alfalfa under different stressful environments is a great challenge, representing a serious threat to global food security. To improve the production of alfalfa under stressful conditions, there is a crucial need to understand the response of alfalfa plants to stresses, the mechanisms of tolerance and the management options. Alfalfa is a promising biorefinery crop from which leaf protein concentrates as well as bio-organic fertilizers can be produced. The potential of leaf-derived protein concentrate for human and animal nutrition is huge, yet unexploited. The bio-organic fertilizers derived from alfalfa plants represent a sustainable way to supply the plant with some nutrients. This review article highlights the sustainable use of alfalfa in biorefinery and bio-organic fertilizer production. Moreover, scattered knowledge about its use for production is collected supporting its suitability for improving soil fertility. Last, we discuss the suitability of alfalfa to produce stable yields in the light of changing climate and stressful conditions like salinity.

Organic and ecological farming programs require new and efficient biostimulants with beneficial properties for the sustainable and safe production of seedlings and ornamental plants. We examined the effect of non-fermented and lacto-fermented alfalfa brown juice (BJ) on seed germination and the vegetative, physiological, and anatomical properties of French marigold (Tagetes patula L. ’Csemő’) plants which were treated with 0.5–10% fermented and non-fermented BJ, with tap water applied as a control. Applying 0.5% fermented BJ significantly improved seed germination compared with non-fermented BJ, resulting in an increase of 9.6, 11.2, 10.9, and 41.7% in the final germination percent, germination rate index, germination index, and vigor index, respectively. In addition, it increased the root and shoot length by 7.9 and 16.1%, respectively, root and shoot dry mass by 20 and 47.6%, respectively, and the number of leaves by 28.8% compared to the control. Furthermore, an increase in contents of water-soluble phenol, chlorophyll a and b, and carotenoid was reported upon the application of 0.5% fermented BJ, while peroxidase activity decreased. Our results prove that alfalfa BJ can be enrolled as a biostimulant as part of the circular farming approach which supports the sustainable horticultural practice.