Composition comprising polyphenolic aqueous extract derived from olive mill wastewater (alpechin) as a biostimulant and protective agent for plants

Abstract

The proposed invention relates to a composition having two phases (organic phase and aqueous phase), which is an organic product that enables crops to better adapt to environmental conditions and promotes increased tolerance to biotic and abiotic stress phases, especially those caused by environmental conditions or inherent conditions at critical moments of development, and also stimulates and strengthens crop growth and yield, thereby providing nutrients and strengthening plant immunity.

Description

[0001] COMPOSITION POLYPHENOLIC AND AQUEOUS EXTRACT DERIVED FROM ALPECHIN AS

[0002] BIOSTIMULANT AND PROTECTIVE AGENT FOR PLANTS

[0003] Field of Invention

[0004] The present invention relates to the field of agriculture, the cultivation of agricultural products, and the stimulation and protection of agricultural products against pathogens and microbiological agents that may affect crop fields and plant growth.

[0005] Background

[0006] First of all, it is important to consider that a biostimulant is any substance or microorganism that, when applied to plants, is capable of improving their efficiency in nutrient absorption and assimilation, tolerance to biotic or abiotic stress, or improving some of their agronomic characteristics, regardless of the substance's nutrient content. Furthermore, plant growth is influenced by a multitude of external factors. Individually or collectively, these factors can drive the evolution of plant development and adaptation to their environment. One of the major goals of agricultural research is to maximize the capacity of plants to enhance their self-defense and response potential in interaction with the factors that regulate the various events that characterize their biological cycle.

[0007] Regarding polyphenols, these are complex water-soluble substances essential for the physiological functions of plants. Thanks to their strong binding properties, they have a significant antioxidant effect by capturing free radicals. Furthermore, they can bind with certain lipids and enzymes, participating in signaling mechanisms; and they are capable of chelating some nutrients, increasing their availability. Plants produce them naturally, preparing them for biotic and abiotic stress conditions. The synthesis of secondary metabolites, such as polyphenols, has been shown to correlate positively with plant vigor. The exogenous application of phenolic antioxidants stimulates their endogenous concentration in plant cells, in turn activating enzymes that catalyze the synthesis of other antioxidant metabolites.

[0008] Based on what is known about polyphenols, we can affirm that we have found technologies that stimulate crop metabolism, generating more vigorous plants capable of more efficient resource use and tolerant to certain types of stress, whether biotic or abiotic, that may occur during the production cycle. However, there are no compositions containing polyphenolic compounds that ensure a biostimulant effect and, at the same time, protect against all types of biotic and abiotic factors present in different types of crops and plants.

[0009] For example, application CL 201403609, entitled “NATURAL FUNGICIDE FORMULA OF THE EMULSIFIED OR MICROENCAPSULATED TYPE BASED ON EXTRACTS OF GRAPE POMACE OR POMASA TO CONTROL INFESTATION BY BOTRYTIS CINEREA, IN PLANTS, PARTS OF PLANTS OR THEIR FRUITS, AND ITS PREPARATION PROCESS”, refers to a fungicide formulation of the emulsified or microencapsulated type comprising extracts of grape pomace or pomace and corresponds to a two-phase system: an oily phase consisting of concentrated extract of grape pomace polyphenols, and an aqueous phase consisting of one or more encapsulating agents selected from the group consisting of: gum arabic, ester gum, propyl cellulose, butylcellulose, capsul, purity gum 2000, purity gum ultra or purity gum bee, where said oily phase has a concentration that may vary between 1-5.000 ppm and said aqueous phase has a concentration that can vary between 1 and 20% w / w, and where the oily phase and the aqueous phase are in proportions in the range between 2 and 90% w / w, and also comprises one or more emulsifying and / or stabilizing agents, which are present in concentrations within the range of 0.1 and 20% w / w and 20% w / w of the aqueous phase, respectively, useful for controlling an infestation by Botrytis cinerea, in plants, parts of plants or their fruits, and can be applied by spraying as an aqueous suspension on the surface of the leaves and / or fruits of the plant to be protected, and its preparation process.In this case, wine industry waste is used, with a wide range of preparations, to try to cover different aspects and crops; however, the preparation is not specific to certain fruits or crops of commercial interest, and a long process of trial and error is required to find the proportions useful for different types of crops.

[0010] On the other hand, application CL 200501730 describes a fungicidal composition comprising an anhydrous milk fat (amf) and optionally one or more of soybean oil, olive oil and coconut fat, and one or more of a carrier, an emulsifier and an antioxidant, its use for the production of an antifungal agent and a method for preventing a fungal infection in a plant; however, it does not directly refer to the use of agricultural industry residues, but rather to known nutritional elements, which could have a positive effect when applied to plants and / or protect against fungal infection.

[0011] In application WO2012138806A2, entitled “Olive mill waste fractions, their preparation and uses”, which describes compositions with different components derived from olive mill wastewater, including hydroxytyrosol and oleuropein, as part of the components of compositions useful for their antimicrobial, antiviral and antifungal effect of said invention. In application BRPI0614917A2, a process for obtaining a natural concentrate rich in natural hydroxytyrosol from olive residues and by-products is described, a process by which hydroxytyrosol can be obtained from oleuropein, with use mainly in the food, pharmaceutical and cosmetic industries.

[0012] Finally, EP2526785B1, entitled "Process for obtaining a phytoextract from vegetable water and olive oil pomace," describes the process for obtaining a phytoextract from vegetable water (VW) and olive oil pomace from olive milling. This process is based on a combination of physical-chemical and enzymatic pretreatment methods, tangential membrane filtration, and vacuum evaporation. The method allows for the eco-sustainable and efficient extraction of the active ingredients involved. The final extract is characterized by the presence of a high polyphenol count and is used in the food, cosmetics, and phytotherapeutic industries.

[0013] In general, there is prior art related to the technology related to the proposed invention, directly or indirectly, especially regarding the use of organic fungicides based on industrial waste (such as, for example, from the wine industry) containing phenolic compounds, to contain or prevent the action of pathogens in plants or crops. Although there is currently greater stress in crop production, both due to water stress and climate change, there is currently no product on the market that can ensure protection against a broad spectrum of biotic and abiotic stresses, while also contributing to improving the biological processes of plants, increasing crop yield and quality.

[0014] Summary of the Invention

[0015] The proposed invention comprises a two-phase composition (organic phase and aqueous phase) corresponding to an organic product that allows crops to better adapt to environmental conditions and contributes to increasing tolerance to biotic and abiotic stress phases, especially due to environmental or plant-based conditions at critical times of development, as well as stimulating and strengthening crop growth and yield by providing nutrients and strengthening plant immunity.

[0016] Description of the Invention

[0017] In general terms, the invention corresponds to a two-phase composition (organic and aqueous) of polyphenolic compounds with a biostimulant effect, which allows crops to better adapt to environmental conditions and collaborate to increase tolerance to stress phases due to environmental or specific conditions at critical moments of development.Although plants can produce polyphenols naturally and prepare them to face biotic and abiotic stress conditions, the combination present in the proposed invention is not naturally found in cultivation conditions, since the composition of the invention contains a balance of elements in organic phase and aqueous phase, in a ratio of concentrations that allows obtaining a surprising effect for the protection of the plant with respect to stress factors and in turn, stimulates foliar and root growth of crops, in different ranges depending on the species, but in a manner superior to the particular properties of its components separately.

[0018] In particular, the proposed development corresponds to a composition comprising a polyphenolic extract with specific elements and in specific and determined concentrations for different types of crops, which allows maintaining good yields under unfavorable conditions caused by water or heat stress or by the attack of pests and pathogens, where the proposed invention is useful or can be used and / or applied directly to plant species to favor any of the following conditions: Tolerance to abiotic stress (Tolerance to salinity and drought, tolerance to freezing, tolerance to high temperatures and flooding) Greater growth and nutritional quality (better root and branch growth, early flowering, higher yield, uniform fruit size, increased antioxidants) Post harvest.

[0019] In another embodiment of the invention, a mixture of the organic extract and a mixture of the aqueous phase (derived from obtaining the extract) in specific proportions, allows for improved growth compared to the use of only the organic extract or commercial fertilizers, observing an additive and synergistic effect, when the organic extract is combined with its aqueous phase, in specific concentrations, for certain types of crops and even for certain specific tissues in the different crops.

[0020] In another embodiment of the invention, the composition comprising the organic extract and the aqueous phase, at specific concentrations, in addition to its biostimulant effect, has been observed to have a protective effect on the plant against pests and other factors that normally negatively affect the plant under field cultivation conditions, including biotic and abiotic factors, as well as, in particular, protection against pathogens, including fungal pathogens that normally affect crops. In particular, the invention allows the plant to be protected and, at the same time, facilitate and stimulate intrinsic protection mechanisms against pathogens in different plants.

[0021] In one embodiment of the invention, the composition comprises a proportion of at least 75 parts of phenolic extract and 25 parts of aqueous solution, per 100 parts of the total composition. In another embodiment of the invention, the composition comprises a proportion of at least 90 parts of phenolic extract and 10 parts of aqueous solution, per 100 parts of the total composition.

[0022] In another embodiment of the invention, the composition comprises a concentration of at least 1 cc / L and up to 2 cc / L of phenolic extract.

[0023] In another embodiment of the invention, the composition comprises a list of components and / or subcompositions, wherein said components and / or subcompositions can be selected from the list comprising: a part of phenolic extract, a mixture of phenolic extract and aqueous solution, a part of aqueous solution, a fertilizer, a biofertilizer, a biostimulant, as well as any of the possible combinations of these elements with each other. Wherein addition, the mixture of phenolic extract can be selected from the list comprising: a mixture of 50 parts of phenolic extract, for every 50 parts of aqueous solution; a mixture of 75 parts of phenolic extract and 25 parts of aqueous solution; and, a mixture of 90 parts of phenolic extract and 10 parts of aqueous solution, where the concentration of the composition is preferably expressed in cc / L.

[0024] The composition of the invention, whose mixture is 75 / 25 of phenolic extract over aqueous phase, respectively (EF / FA), is effective in stimulating plant growth, increasing average plant weight, increasing the leaf / root ratio of plants, as well as in protecting and preventing plant infections by pathogens. Furthermore, said 75 / 25 composition is effective in reducing the percentage of browning in cherry fruits.

[0025] The composition of the invention, whose mixture is 90 / 10 of phenolic extract on aqueous phase respectively (EF / FA), is effective for stimulating plant growth, increasing average plant weight, increasing weight in leaf / root ratio of plants, as well as in the protection and prevention of plant infections by pathogens.

[0026] EXAMPLES

[0027] Example 1 - Extraction methods of the polyphenolic and aqueous phases

[0028] Specifically, in our case, the biostimulant we developed consists of a mixture of a polyphenolic extract, primarily containing hydroxytyrosol and oleuropein, and an aqueous extract containing amino acids and NPK. Extracted from a highly polluting and difficult-to-treat agro-industrial waste product, Alpechin (a watery residue from the olive oil process).In our search for an effective and unique biostimulant, we discovered a unique feature in our extracts: they work effectively on their own. However, when mixed with different concentrations depending on the crop type, they produce a heightened effect that would not be expected based on the results obtained separately. It is this synergy that makes the difference, achieving a high-quality biostimulant from a process that not only benefits the environment by recovering a complex waste product, but is also highly innovative.

[0029] Obtaining extracts:

[0030] From alpechin, a residue from the agricultural industry, two extracts can be obtained, differentiated in terms of their components and physicochemical characteristics, as detailed below:

[0031] A) POLYPHENOLIC EXTRACT

[0032] First, a liquid-liquid solvent extraction method is used, which allows us to extract the polyphenols in their organic phase. This classic extraction consists of the following steps:

[0033] 1) Extraction at room temperature with ethyl acetate / alpechin 1 is 2

[0034] 2) Phase separation at room temperature by decantation method

[0035] 3) Concentration by vacuum rotary evaporation method at a suitable temperature to avoid decomposing the polyphenols present.

[0036] 4) Obtaining the polyphenolic biostimulant by adding the extract to water plus a dispersant. This solution contains 8.45% of the polyphenolic extract.

[0037] *Polyphenolic extract contains between: 0.5 - 10% of Total Polyphenols (TPP)

[0038] 1 - 1.6% Hydroxytyrosol of the % PFT.

[0039] X% Oleuropein, where X corresponds to a minimum of 0.5 and a maximum of 0.8% PFT.

[0040] B) AQUEOUS EXTRACT

[0041] In the previous process, after the phase separation step, we work with the separated aqueous phase:

[0042] 1) Concentration by evaporation, up to 10% of the original volume C) MIXTURE OF POLYPHENOLIC EXTRACT + AQUEOUS PHASE: i) 50% POLYPHENOLIC EXTRACT AND 50% AQUEOUS EXTRACT

[0043] The mixture of both is carried out for subsequent dilution to the optimal concentration, read per liter ii) 75% POLYPHENOLIC EXTRACT AND 25% AQUEOUS EXTRACT iii) 90% POLYPHENOLIC EXTRACT AND 10% AQUEOUS EXTRACT iv) 25% POLYPHENOLIC EXTRACT AND 75% AQUEOUS EXTRACT v) 10% POLYPHENOLIC EXTRACT AND 90% AQUEOUS EXTRACT

[0044] The extraction process can be observed in detail and sequentially in Figure 1.

[0045] Although the concentrations of hydroxytyrosol and oleuropein in the % PFT are variable, at least a ratio of 2:1 respectively has been identified, without prejudice to the fact that in different extracts, other ratios between hydroxytyrosol / oleuropein relevant to the proposed invention could be identified and / or used, in specific species and / or varieties.

[0046] Alternatively, there are also other reasons or proportional relationships between hydroxytyrosol and / or oleuropein with respect to one or more of the polyphenols identifiable in the PFT extract, relevant and present in the proposed invention.

[0047] Alternatively or additionally, formulations with different product ratios (FA / FO) have been identified and / or used depending on the desired effect (biostimulant, protection of specific pathogens) and the particular species to be protected.

[0048] Alternatively or additionally, the present invention further comprises a composition of aqueous phase and organic phase in different proportions depending on the different types of crops, wherein said formulation provides a preventive-residual protective effect to the crops, and allows extending the protection time of the crop to external factors, especially post-harvest, including: oxidation, dehydration and other factors related to the transportation of agricultural products from producers to consumers.

[0049] Example 2 - First embodiment of the invention in lettuce crops To evaluate the effectiveness of the extract on the growth and development of lettuce plants, 1.5-liter pots with compost substrate were used. Applications were made via irrigation (80 cc per plant) and / or sprinkling (a hand-held sprinkler).

[0050] The evaluation measured the fresh and dry weight of the plant's shoots and roots. The plants were kept in greenhouse conditions for 60 days. At the end of the trial, the plants were removed from the pots and the roots were washed to determine the fresh weight of the roots and leaves separately. They were then allowed to dry until the weight was stable, and then their dry weight was determined.

[0051] As we can see in Figure 2, the application of the polyphenolic extract biostimulant, by spraying the aerial part at a dose of 2cc / L, obtains the best results, since with this treatment an increase in fresh weight of 31% is obtained, the applications of the other treatments increase the weight of the lettuce plant in the same way, but to a lesser extent.

[0052] Figure 3 shows the evaluation of dry weight in lettuce plants after applying a biostimulant at doses of 2 and 4 cc / L, applied via irrigation and by sprinkling to the aerial part of the plant. In the case of total dry weight, the spray treatment at a dose of 2 cc / L produces a weight increase of 30.5%, the irrigation treatment at 4 cc / L, and irrigation plus sprinkling with doses of 2 cc / L, also produce an increase in the dry weight of the plants, the other treatments are not different from the control.

[0053] Example 3 - Composition of the Invention

[0054] Product: Biostimulant

[0055] Chemical group: Polyphenols

[0056] Composition: i) Aqueous Extract.

[0057] Amino acids

[0058] Water . 84.5% ii) Organic Extract:

[0059] Polyphenolic extract (Total polyphenol content 0.5-10%) . 8.45%

[0060] Dispersant additive. 7.1%

[0061] Where the proportion between the aqueous and organic phase will be determined by the objective (desired technical effect) of the invention and / or depending on the type of plant to be used, and where the dispersing additive can be selected from the list comprising: tween 20, tween 80 and any other suitable polysorbate or equivalent available.

[0062] EXAMPLE 4 - Tests of the invention applied to hydroponic crops

[0063] Methodology:

[0064] The following products were used to evaluate the effectiveness of applying a biostimulant based on olive oil industry waste:

[0065] 1. Polyphenolic Extract (EP)

[0066] 2. Aqueous phase of the extraction of the polyphenolic extract (FA)

[0067] 3. Mix of polyphenolic extract plus aqueous phase

[0068] These were applied in four different applications, separated by at least 15 days and finally harvested at least 5 days after the last application.

[0069] Treatments used

[0070] Table 1.- Summary of results obtained from successive applications of products based on the olive oil industry (FIGURE 4). From the data in Table 1, a synergistic effect can be observed in treatments 6 and 7 with respect to treatments with phenolic extract only (treatment 2 and 3) and treatment with aqueous phase (treatment 4) respectively.

[0071] Table 2. Percentage increase in the weight of lettuce leaves treated with products based on olive oil industry waste, compared to a control with water application.

[0072] According to the data obtained in Table 2 (FIGURE 5), we can observe that the treatment with a mixture of 0.75 cc of the polyphenolic extract + 0.25 cc of the aqueous phase per liter is the one that shows the best results in the growth of the vegetative part in hydroponically grown lettuce cv. Lozano, very similar to what was observed in the treatment 0.9 cc of the polyphenolic extract + 0.1 cc of the aqueous phase per liter. The other treatments did not differ from the control.

[0073] Similar to what was observed in Table 1, the 0.75 / 0.25 and 0.9 / 0.1 mixtures present a surprising effect, compared to the treatments with only the extract phase or the aqueous phase, respectively.

[0074] Table 3. Percentage increase in root weight of lettuce treated with products based on olive oil industry waste, compared to a control with water application.

[0075]

[0076] According to the data observed in Table 3 (FIGURE 6), the treatment with the mixture of 0.9 cc of polyphenolic extract + 0.1 cc of the aqueous phase shows the best results, reaching an average of 25.8 grams of root per plant. At an intermediate level of effectiveness, the treatment with the polyphenolic extract at a dose of 1 cc / L, the treatment with a mixture of 0.75 cc + 0.25 cc polyphenolic extract and the treatment with the aqueous phase at a dose of 1 cc / L. The other treatments do not differ from the control. The effect of the 0.9 / 0.1 mixture is surprising compared to the effect of the treatments with pure extract and pure aqueous phase, and any of their intermediate mixtures.

[0077] Table 4. Percentage increase in the total weight of lettuce plants treated with products based on olive oil industry waste, compared to a control with water application.

[0078] According to the data in Table 4 (FIGURE 7), treatments with a mixture of 0.75cc + 0.25cc polyphenolic extract per liter and the 0.9cc mixture of polyphenolic extract are those that show the best results in the total weight of lettuce plants of the Lozano variety, followed by the polyphenolic extract treatment at doses of Icc / L. The other treatments are not different from the control. Similar to what was observed in Tables 1 and 2, the 0.75 / 0.25 and 0.9 / 0.1 mixtures show a surprising effect, with respect to treatments with phenolic extract or aqueous extract respectively, as well as what was expected in a mixture of both.

[0079] EXAMPLE 5 - Application of the Invention in hydroponic cultivation of Racula

[0080] This trial was conducted in a hydroponic greenhouse in the town of Champa, Paine commune, in the Metropolitan Region. Three applications were made during the trial: the first in the seed trays and the remaining two during the crop's growth period. The plants were harvested when they reached the commercial requirement, and root and leaf weight were measured (FIGURE 8).

[0081] Table 5. Composition treatment in Raculas grown in hydroponic culture (total weight)

[0082] Table 6. Composition treatment in Raculas grown in hydroponic culture (leaf weight)

[0083] Table 7. Composition treatment in Rúenlas grown in hydroponic culture (root weight)

[0084] EXAMPLE 6 - Hydroponic lettuce

[0085] According to the data obtained (FIGURE 9), we can observe that the treatment with a mixture of 0.75cc polyphenolic extract + 0.25cc of the aqueous phase per liter is the one that shows the best results in the growth of the vegetative part in hydroponically grown lettuce cv. Lozano, very similar to what was observed in the treatment 0.9cc of polyphenolic extract + 0.1cc of the aqueous phase per liter, where both treatments show a surprising effect compared to any of the mixtures and treatments with phenolic extract and aqueous phase only. The other treatments did not differ from the control.

[0086] EXAMPLE 7 - Determination of Polyphenol Content in Alpechín The quantification of the content of Polyphenols present in the liquid waste of a Commercial Agricultural Company was determined in the Food Laboratory (Antioxidant Analysis) of the Institute of Nutrition and Food Technology (INTA) (FIGURE 10 and 11).

[0087] The methodologies used for all polyphenol trials included in the period's activities are those described below:

[0088] Extraction / Dissolution Medium: Hexane and Methanol / Ultra-pure double-distilled water (60 / 40 V / V). Total Polyphenols: They were tested as described in the internal procedure of the Antioxidant Analysis Laboratory (MME-Pro-001), which is based on the method of Wu et al. (J. Agrie. Food Chem. 52: 4026-4037; 2004).

[0089] Hydroxytyrosol and Oleuropein content: It was determined based on the methodology described by Hai-Wei et al (J. of Chromatography B, 78: 187-191; 2003) with some modifications.

[0090] Where Hydroxytyrosol and Oleuropein are the main polyphenols present in the organic fraction of the olive industry waste, therefore in the process of obtaining olive oil, they remain mostly in the Alpechín.

[0091] EXAMPLE 8 - Pathogen protection in Arugula

[0092] The pathogen that affected the arugula is Diamondback moth, where the treatment applied refers to the use of only the control, treating the plants with water without any type of insecticide or insect protection product (FIGURE 14 A); while in another example, a mixed treatment between aqueous and organic composition was used, which effectively kept the treated plants free from insect damage (FIGURE 14B).

[0093] EXAMPLE 9 - Pathogen Protection in Tomato

[0094] In this case (FIGURE 16), it can be observed that both the phenolic extract and the aqueous solution, as well as all mixtures of both in different proportions, are effective in protecting tomato plants against powdery mildew infection. However, the 75F / 25A mixture only presents an "unexpected" effect in the high incidence zone compared to a mixture of the aqueous and phenolic extracts, respectively, since the percentage of infection incidence is still lower than both cases separately. Even so, the aqueous extract, the phenolic extract, and the 50 / 50 mixture of both have an equally positive effect on pathogen protection. EXAMPLE 10 - Harvest Parameter Evaluation in Cherry Trees

[0095] Harvest parameters, soluble solids (sugar in the fruit) were evaluated with a refractometer, fruit color was measured according to the parameters in photograph 1, and fruit size at harvest as equatorial diameter.

[0096] Fruit was stored in a refrigerator for one month at zero degrees Celsius, then the browning of the fruit pedicel was evaluated to determine the antioxidant power of the treatments used.

[0097] 30 fruits were evaluated per repetition according to the following table:

[0098] Table 8.- Browning scale in cherry pedicels.

[0099] According to the data presented in FIGURE 17A, when applying the treatments at: beginning of flowering, full flower, petal fall, jacket fall, color break, and pre-harvest, polyphenolic extract shows an increase in the percentage of sugar present in the fruit, compared to the other treatments. The other treatments are the same as the control to which only water was applied. Additionally, as observed in FIGURE 17B, the treatment with the highest percentage of soluble solids is the 75 / 25 Mix, the other treatments are not statistically different from the control.

[0100] Finally, in FIGURE 18 we can see that the treatments with the best results are the Mix 75 / 25, where the browning of the pedicels is reduced by 40%, followed by the Mix 50 and extract treatments, which reduce the browning by 32 and 28% respectively, with applications of the treatments from the beginning of flowering to pre-harvest.

[0101] EXAMPLE 11 - Weight gain effect on Basil leaves

[0102] The mixture of the proposed invention was analyzed in hydroponic cultivation of basil to evaluate its effect on the final weight of the plant, especially on the leaves of the basil plant, whose data are described in the following table:

[0103] Table 9. Detail of basil leaf weight in percentages.

[0104] As a result, FIGURE 19 shows that the best treatments observed are: the mixture of the extract with the aqueous in percentages of 0.75cc extract + 0.25cc of aqueous in 1 liter and the application of only the polyphenolic extract; where the mixture of the invention in the aforementioned proportion, presents a superior effect to the pure phenolic extract, which is not expected for a mixture considering the effect of the pure aqueous extract, as would be the result of the 0.5 / 0.5 mixture that is also observed in the graph.

[0105] Brief Description of Figures

[0106] FIGURE 1. Method of obtaining the extract

[0107] FIGURE 2. Total fresh weight of lettuce treated with the Biostimulant

[0108] FIGURE 3. Total dry weight of lettuce treated with the biostimulant

[0109] FIGURE 4. Evaluation in lush lettuce, where an unexpected effect (synergy) is observed between the extract and the fertilizer, especially in the weight of leaves and in the total weight of the plant.

[0110] FIGURE 5. Graph showing the percentage increase in the weight of lettuce leaves treated with products based on olive oil industry waste, compared to a control with water application.

[0111] FIGURE 6. Graph showing the percentage increase in root weight of lettuce treated with products based on olive oil industry waste, compared to a control with water application.

[0112] FIGURE 7. Graph showing the percentage increase in total weight of lettuce plants treated with olive oil industry waste products compared to a control treated with water. FIGURE 8. Evaluation of lettuce plants, showing the unexpected effect and synergy between the extract and the fertilizer on the lettuce root.

[0113] FIGURE 9. Leaf weight of Lozano lettuce, treated with phenolic, aqueous phase, and mixtures of these in different percentages. Means with a common letter are not significantly different (p > 0.05).

[0114] FIGURE 10. Molecules of some relevant components present in alpechin: A) Hydroxytyrosol molecule, and B) Oleuropein molecule.

[0115] FIGURE 11. Main polyphenolic compounds in alpechin.

[0116] FIGURE 12. Analysis of amino acids present in sample 1.

[0117] FIGURE 13. Analysis of Amino Acids present in sample 2

[0118] FIGURE 14. Ruenla plant: A) control treatment with application of water only, with insect damage to the leaves (ie: Diamondback moth), and B) with application of polyphenolic extract and FE / FA mixtures according to the proposed invention, with evident reduction of insect damage, and greater growth of the leaf part.

[0119] FIGURE 15. Examples of substances that can be identified from a sample of alpechin.

[0120] FIGURE 16. Graph depicting powdery mildew infection on tomato plant leaves.

[0121] FIGURE 17. A) Percentage of soluble solids in cherry fruit with applications from blossom onset to preharvest; B) Percentage of soluble solids in cherry fruit with applications from color break to preharvest.

[0122] FIGURE 18. Browning of pedicels in cherry fruits at harvest with applications from the beginning of flowering to pre-harvest.

[0123] FIGURE 19. Hydroponic basil leaf development growth.

Claims

CLAIMS 1. A two-phase composition extracted from vegetable water for the protection of crops against biotic and abiotic stress that also stimulates plant growth, CHARACTERIZED in that it comprises a specific proportion between the aqueous phase and the organic phase, wherein the organic phase in turn comprises specific polyphenolic components that allow the protection and stimulation of plant growth in different crops; wherein in addition, the aqueous and organic phases are obtained from a suitable extraction method, and wherein the composition of the phases comprises at least: a polyphenolic extract, an aqueous extract, at least one dispersant additive; wherein the polyphenolic extract contains at least: i. between 0.5 - 10% of Total Polyphenols (TPP), ii. between 1 - 1.6% of Hydroxytyrosol of the % TFP, and iii. X% of Oleuropein, where X corresponds to a minimum of 0.5 and a maximum of 0.8% of the TFP percentage.and where the aqueous extract corresponds to a concentration by evaporation, up to 10% of the original volume, which comprises amino acids and water.

2. The composition of claim 1, CHARACTERIZED in that the proportion of the composition is 75 parts of phenolic extract per 25 parts of aqueous solution.

3. The composition of claim 2, CHARACTERIZED in that said composition has a surprising effect on the growth of the leaf(s), as well as the proportion of leaf(s) vs root(s), increasing the average weight of hydroponic products, with respect to the effect of the phenolic extract, aqueous extract, and the expected effect of any of the mixtures with each other.

4. The composition of claim 1, CHARACTERIZED in that the proportion of the composition is 90 parts of phenolic extract per 10 parts of aqueous solution, has a surprising effect on plant growth and average weight increase of hydroponic products.

5. The composition of claim 4, CHARACTERIZED in that said composition favors leaf growth, as well as the leaf vs. root ratio, increasing the average weight of hydroponic products.

6. A two-phase composition (organic and aqueous) for the protection of crops against pathogens, including fungi, CHARACTERIZED in that it comprises a specific proportion between the aqueous phase and the organic phase, which in turn comprises specific polyphenols of the invention extracted from vegetable water, wherein the specific composition comprises 75 parts of phenolic extract and 25 parts of aqueous phase, and wherein said proportion has a preventive effect on plant infection against different pathogens.

7. The composition of claim 6, CHARACTERIZED in that said composition is effective against powdery mildew infection in leaves of tomato plants.

8. The composition of claim 7, CHARACTERIZED in that said composition is effective in the upper, middle and lower areas of the tomato plant.

9. The composition of claim 8, CHARACTERIZED in that said composition is preferably effective in the lower area of ​​the tomato plant.

10. The composition of claim 6, CHARACTERIZED in that said composition is also effective against infection by Diamondback moth, in ruenla.

11. A two-phase composition for improving plant growth, CHARACTERIZED in that said composition comprises at least two phases: i) phenolic phase, and ii) aqueous phase, wherein the proportion of both phases can be selected from the list comprising: composition 75% phenolic phase and 25% aqueous phase, phenolic composition 90% phenolic phase and 10% aqueous phase, and any of the intermediate mixtures; and wherein the improvement of plant growth of the invention includes: phenotypic improvements of plants and crops, yield, growth, nutritional quality and other visible and / or verifiable factors in plants in agricultural crops.

12. A two-phase composition for plant protection with a preventive-residual effect, promoting the absorption of nutrients by the plant and extending the protection time in crops, CHARACTERIZED in that said composition comprises at least two phases: i) phenolic phase, and ii) aqueous phase, wherein the proportion of both phases can be selected from the list comprising: composition 75% phenolic phase and 25% aqueous phase, phenolic composition 90% phenolic phase and 10% aqueous phase, and any of the intermediate mixtures; and wherein the protection time in crops where said composition of the invention has been applied is extended to include post-harvest.

13. A method for the separation of the organic phase and the aqueous phase from alpechin for the extraction of a phenolic extract and an aqueous extract, CHARACTERIZED in that the method comprises the following stages: a. Pretreatment, which comprises filtering the alpechin through a mesh of at least 2 mm, b. Solvent extraction, which comprises mixing the alpechin filtrate resulting from the previous stage with a suitable solvent in a 2: 1 ratio; subsequently, the mixture is stirred at a speed of at least 350 rpm, at room temperature and for a period of at least 30 minutes, c. Effective phase separation, wherein the organic phase corresponds to the resulting volume of the upper phase, and where the aqueous phase corresponds to the resulting volume of the lower phase, and both volumes are separated independently, d. In the case of the organic phase volume, a rotary evaporation of the organic phase is carried out to obtain the polyphenolic extract, wherein said extract is diluted in water with a dispersing additive, resulting in the organic component of the mixture of the invention, e. In the case of the aqueous phase volume, an evaporation of said phase is carried out until an aqueous extract is obtained, wherein said aqueous extract is decreased by evaporation to a volume of up to 10% of the initial volume, resulting in the aqueous component of the invention, and f. Finally, a mixture of the resulting components in d) and e) is made, in a specific proportion, depending on the type of effect and types of crops to be applied, which can be selected from the list comprising at least: 50% / 50%, 75% / 25%, and 90% / 10%, of phenolic component / aqueous component respectively.

14. The proposed invention, CHARACTERIZED as described herein.

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