WO2022011298A1 - Microalgae and fertilizer mixtures and methods of use thereof to enhance plant characteristics - Google Patents

Abstract

The present invention provides a mixture comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; wherein a combination of the first composition and the second composition exhibits synergy. Also provided is a method of treating a plant, a plant part, or the locus surrounding the plant to enhance plant growth and/or yield, the method comprising applying an effective amount of the disclosed mixture to the plant, plant part, or the locus surrounding the plant.

Description

MICROALGAE AND FERTILIZER MIXTURES AND METHODS OF USE THEREOF TO ENHANCE PLANT CHARACTERISTICS CROSS-REFERENCE TO RELATED APPLICATIONS The present application claims priority to U.S. Provisional Patent Application No. 63/050,557, filed July 10, 2020, and U.S. Provisional Patent Application No.63/057,007, filed July 27, 2020, the contents of each of which are hereby incorporated by reference in their entireties. TECHNICAL FIELD The present invention relates generally to mixtures and methods for stimulating and maintaining enhanced growth and yields in plants. More particularly, the present invention relates to mixtures comprising microalgae cells and fertilizers. BACKGROUND Fertilizers are one of the major factors that influence agriculture efficiency at the base of the food security chain. Fertilizers may be delivered to farmers in a granular, powder, or liquid form. Unfortunately, many agricultural practices focus entirely on synthetic fertilizers that accelerate plant growth but produce crops with limited nutrient value. Repeated and exclusive application of synthetic fertilizers compounds the problem of crop production with limited nutrient value as soil health declines. It has now been recognized that various characteristics including the quality, health, color, and yield of plants can be improved through the application of effective amounts of biomass obtained from the cell tissue of microalgae species including Chlorella. Application of microalgal biomass to soil increases soil aggregation and water retention thereby providing a more productive growth medium for plants. The resulting healthy soil provides the nourishment required to produce crops with high nutrient value. There is a growing need to develop mixtures and co-formulations of synthetic fertilizers with microalgae-based agricultural products to enhance plant growth while producing highly nutritious crops and building healthy soils. SUMMARY The present invention provides a mixture comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; wherein a combination of the first composition and the second composition exhibits synergy. In certain aspects, the culture of microalgae comprises Aurantiochytrium, Botryococcus, Chlorella, Chlamydomonas, Desmodesmus, Dunaliella, Scenedesmus, Pavolv, Phaeodactylum, Nannochloropsis, Spirulina, Galdieria, Haematococcus, Isochrysis, Porphyridium, Schizochytrium, Thraustochytrium, Tetraselmis, or combinations thereof. In one aspect, the culture of microalgae comprises Chlorella. The Chlorella can be whole cells, lysed cells, dried cells, cells that have been subjected to an extraction process, or a combination thereof. In other aspects, the fertilizer is an organic fertilizer, inorganic fertilizer, urea- containing fertilizer or combination thereof. In one aspect, the fertilizer is an organic fertilizer selected from the group consisting of humic acid, kelp, seaweed extract, fulvic acid, fish emulsion/fish meal, soy hydrolysate, protein hydrolysate, AMINO PRIME^TM (2.5-0-1 derived from sugarcane protein hydrolysate), corn steep liquor, compost, manure, biochar, sugar beet or sugarcane vinasse, sewage sludge, blood meal, bone meal, worm castings, peat moss, leaf compost, rice hull, coffee chaff, buckwheat hull, chicken manure, tree bark with or without composting, mushroom composts, black soldier fly frass, and combinations thereof. In another aspect, the fertilizer is an inorganic fertilizer selected from the group consisting of CAN17, CN9, CaTS^® (6% calcium and 10% sulfur calcium thiosulfate fertilizer solution), AN20, AMS, ATS, UN32, 10-34-0, 11-37-0, 6-26-5, 5-28-0-10 Mg, 44-0-0, 14-24- 0-10S, 17-1-0-20S, KTS^® blend (0-0-25+17S derived from potassium thiosulfate), K-ROW 23^® (0-0-23-8S derived from postassium sulfite), liquid ammonia, ammonium nitrate, calcium nitrate, potassium nitrate, calcium ammonium nitrate, phosphoric acid, ammonium phosphate, monoammonium phosphate, diammonium phosphate, ammonium polyphosphate, single super phosphate, triple super phosphate, ground rock phosphate, a phosphite salt, an ammonium iron salt, potassium chloride, muriate of potash, sulfate of potash, sulfate of potash magnesia, soluble potash, ammonium sulfate, ammonium thiosulfate, potassium sulfate, ammonium sulfate-nitrate, calcium cyanamide, potassium hydroxide, potassium acetate, ammonium carboxylate, cyclohexanediaminepentaacetic acid (CDTA), citric acid (CIT), diethylenetriaminepentaacetic acid (DTPA), ethylenediaminediaminedi-o- hydroxyphenylacetic acid (EDDHA), ethylenediamintetraacetic acid (EDTA), ethylene glycol bis(2-aminoethyl ether) tetraacetic acid (EGTA), hydroxyethylenediaminetriacetic acid (HEDTA), nitrilo-triacetic acid (NT A), oxalic acid (OX), pyrophosphoric acid (PPA), triphosphoric acid (TP A), vermiculite, perlite, Chilean nitrate, limestone, and combinations thereof.

In another aspect, the fertilizer is a urea-containing fertilizer selected from the group consisting of urea, urea formaldehyde, urea-acetaldehyde, ureaglyoxal condensate, urea sulfur, urea sulfuric acid (urea sulfate), urea ammonium sulfate, isobutylidene diurea, crotonylidene diurea, ethylene urea, methylene diurea, urea ammonium nitrate, and combinations thereof.

In some aspects, the fertilizer is a solid coated or uncoated granule; in a liquid or semi liquid form; formulated as a sprayable fertilizer; formulated for application via fertigation; and/or formulated as a slow release fertilizer.

In other aspects, the mixture further comprises a nitrification inhibitor, a urease inhibitor, a denitrification inhibitor, or a combination thereof.

In certain aspects, the present invention provides a method of treating a plant, a plant part, or the locus surrounding the plant to enhance plant growth and/or yield, the method comprising applying an effective amount of a mixture disclosed herein to the plant, plant part, or the locus surrounding the plant.

In other aspects, the present invention provides a method of plant enhancement comprising applying to a plant, seedling, plant propagation material, or the locus surrounding the plant material an effective amount of a mixture disclosed herein, wherein the plant characteristic is selected from the group consisting of seed germination rate, seed germination time, seedling emergence, seedling emergence time, seedling size, plant fresh weight, plant dry weight, utilization, fruit production, leaf production, leaf formation, leaf size, leaf area index, plant height, thatch height, plant health, plant resistance to salt stress, plant resistance to heat stress, plant resistance to heavy metal stress, plant resistance to drought, maturation time, yield, root length, root mass, color, insect damage, blossom end rot, softness, plant quality, fruit quality, flowering, sun bum, and any combination thereof.

In one aspect, the components in the mixture are applied simultaneously or subsequently.

In another aspect, the mixture is applied to soil in the immediate vicinity of the plant, seedling, or plant propagation material and/or as a foliar spray.

In yet other aspects, the present invention provides a kit-of-parts comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; in a spatially separated arrangement, wherein a combination of the first composition and the second composition exhibits synergy.

DETAILED DESCRIPTION

As used herein and in the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. For example, “a” or “an” means “at least one” or “one or more.”

Throughout this disclosure, various aspects of the claimed subject matter are presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the claimed subject matter. Accordingly, the description of a range should be considered to have specifically disclosed all the possible sub-ranges as well as individual numerical values within that range. For example, where a range of values is provided, it is understood that each intervening value, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the claimed subject matter. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the claimed subject matter, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the claimed subject matter. This applies regardless of the breadth of the range.

The term “microalgae” as used herein refers to microscopic single cell organisms such as microalgae, cyanobacteria, algae, diatoms, dinoflagellates, freshwater organisms, marine organisms, or other similar single cell organisms capable of growth in phototrophic, mixotrophic, or heterotrophic culture conditions.

The term “fertilizers” is to be understood as chemical compounds applied to promote plant and fruit growth. Fertilizers are typically applied either through the soil (for uptake by plant roots), through soil substituents (also for uptake by plant roots), or by foliar feeding (for uptake through leaves). The term also includes mixtures of one or more different types of fertilizers as mentioned below. The term “fertilizers” can be subdivided into several categories including: a) organic fertilizers (composed of decayed plant/animal matter), b) inorganic fertilizers (composed of chemicals and minerals) and c) urea-containing fertilizers.

As used herein, “phosphite” is defined as inclusive of the anion PO₃ ³ , a salt of P(OH)3 with the related acid being referred to herein as phosphorous acid being synonymously denoted as H3PO3. The term “nitrification inhibitors” is to be understood as any chemical substance which slows down or stops the nitrification process. Nitrification inhibitors retard the natural transformation of ammonium into nitrate, by inhibiting the activity of bacteria such as Nitrosomonas spp. and/or Archaea. The term “nitrification” is to be understood as the biological oxidation of ammonia (NH₃) or ammonium (NH₄ ⁺) with oxygen into nitrite (NO₂ ⁻) followed by the oxidation of these nitrites into nitrates (NO3⁻) by microorganisms. Besides nitrate (NO₃ ⁻) nitrous oxide is also produced though nitrification. Nitrification is an important step in the nitrogen cycle in soil. The term “denitrification” is to be understood as the microbiological conversion of nitrate (NO3⁻) and nitrite (NO2) to gaseous forms of nitrogen, generally N2 or N2O. This respiratory process reduces oxidized forms of nitrogen in response to the oxidation of an electron donor such as organic matter. The preferred nitrogen electron acceptors in order of most to least thermodynamically favorable include: nitrate (NO₃ ⁻), nitrite (NO₂ ⁻), nitric oxide (NO), and nitrous oxide (N2O). Within the general nitrogen cycle, denitrification completes the cycle by returning N₂ to the atmosphere. The process is performed primarily by heterotrophic bacteria (such as Paracoccus denitrificans and various pseudomonads), although autotrophic denitrifiers have also been identified (e.g. Thiobacillus denitrificans). Denitrifiers are represented in all main phylogenetic groups. When faced with a shortage of oxygen many bacterial species, are able switch from using oxygen to using nitrates to support respiration in a process known as denitrification, during which the water-soluble nitrates are converted to gaseous products, including nitrous oxide, that are emitted into the atmosphere. The term “plant propagation material” is to be understood to denote all the generative parts of the plant such as seeds and vegetative plant material such as cuttings and tubers (e. g. potatoes), which can be used for the multiplication of the plant. This includes seeds, roots, fruits, tubers, bulbs, rhizomes, shoots, sprouts and other parts of plants, including seedlings and young plants, which are to be transplanted after germination or after emergence from soil. The term “auxiliary” as used herein refers to an inert ingredient commonly used in agricultural compositions. Examples of auxiliaries include, but are not limited to, extenders, solvents, diluents, emulsifiers, dispersants, binders, fixing agents, wetting agents, dyes, pigments, antifoams, preservatives, secondary thickeners, and stickers. As used herein, the terms “10-34-0,” “APP,” and “Ammonium Polyphosphate 10-34- 0,” and “Ammonium Polyphosphate” are synonymous and refer to an ammonium polyphosphate solution containing about 10.0% total nitrogen as N and about 34.0% total P2O5. As used herein, the term “CAN17” refers to calcium ammonium nitrate 17-0-0 fertilizer.

As used herein, the term “CN9” refers to ammonium calcium nitrate 9-0-0 fertilizer containing about 11% Ca.

As used herein, the term “AN20” refers to ammonium nitrate 20-0-0 fertilizer.

As used herein, the term “AMS” refers to ammonium sulfate fertilizer.

As used herein, the term “ATS” refers to ammonium thiosulfate 12-0-0-26S fertilizer.

As used herein, the term “UN32” refers to urea ammonium nitrate solution fertilizer.

When a fertilizer is referenced herein by a set of three numbers (e.g., “10-34-0”), the numbers refer to the total percent of nitrogen (N), phosphorus (P), and potassium (K) present in the fertilizer. When the fertilizer has a fourth number (e.g., “5-28-0-10 Mg” and “14-24-0- 1 OS”) the fourth number will be followed by a symbol for an element and the number preceding that symbol will indicate the total percent of that element in the fertilizer. For example, in 5- 28-0-10 Mg there is a total of 5% N, a total of 28% P, a total of 0% K, and a total of 10% Mg.

Analysis of the DNA sequence of the strain of Chlorella sp. described herein was done in the NCBI 18s rDNA reference database at the Culture Collection of Algae at the University of Cologne (CCAC) and showed substantial similarity (i.e., greater than 95%) with multiple known strains of Chlorella and Micractinium. Those of skill in the art will recognize that Chlorella and Micractinium appear closely related in many taxonomic classification trees for microalgae, and strains and species may be re-classified from time to time within the Chlorella and Micractinium genera. As would be understood in the art, the reclassification of various taxa is not unusual, and occurs as developments in science are made. Any disclosure in the specification regarding the classification of exemplary species or strains should be viewed in light of such developments. While the exemplary microalgae strain is referred to in the instant specification as Chlorella, it is recognized that microalgae strains in related taxonomic classifications with similar characteristics to the exemplary microalgae strain would reasonably be expected to produce similar results. Accordingly, any mention of Chlorella herein should be understood to include Micractinium species genetically and morphologically similar to species classified within the genus Chlorella as of the filing date.

Taxonomic classification has been in flux for organisms in the genus Schizochytrium. Some organisms previously classified as Schizochytrium have been reclassified as Aurantiochytrium, Thraustochytrium, or Oblongichytrium. See Yokoyama et al. Taxonomic rearrangement of the genus Schizochytrium sensu lato based on morphology, chemotaxonomic characteristics, and 18S rRNA gene phylogeny (Thrausochytriaceae, Labyrinthulomycetes): emendation for Schizochytrium and erection of Aurantiochytrium and Oblongichytrium gen. nov. My coscience (2007) 48:199-211. Those of skill in the art will recognize that Schizochytrium, Aurantiochytrium, Thraustochytrium, and Oblongichytrium appear closely related in many taxonomic classification trees for microalgae, and strains and species may be re-classified from time to time. Thus, for references throughout the instant specification for Schizochytrium, it is recognized that microalgae strains in related taxonomic classifications with similar characteristics to Schizochytrium, such as Aurantiochytrium, would reasonably be expected to produce similar results.

By artificially controlling aspects of the microalgae culturing process such as the organic carbon feed (e.g., acetic acid, acetate), oxygen levels, pH, and light, the culturing process differs from the culturing process that microalgae experiences in nature. In addition to controlling various aspects of the culturing process, intervention by human operators or automated systems occurs during the non-axenic mixotrophic culturing of microalgae through contamination control methods to prevent the microalgae from being overrun and outcompeted by contaminating organisms (e.g., fungi, bacteria). Contamination control methods for microalgae cultures are known in the art and such suitable contamination control methods for non-axenic mixotrophic microalgae cultures are disclosed in W02014/074769A2 (Ganuza, et ah), hereby incorporated by reference. By intervening in the microalgae culturing process, the impact of the contaminating microorganisms can be mitigated by suppressing the proliferation of containing organism populations and the effect on the microalgal cells (e.g., lysing, infection, death, clumping). Thus, through artificial control of aspects of the culturing process and intervening in the culturing process with contamination control methods, the microalgae culture produced as a whole and used in the described inventive compositions differs from the culture that results from a microalgae culturing process that occurs in nature.

In some embodiments and Examples below, the microalgae composition may be referred to as PHYCOTERRA^®, PHYCOTERRA^® ORGANIC, OR PHYCOTERRA^® ST. The PHYCOTERRA^®, PHYCOTERRA^® ORGANIC, OR PHYCOTERRA^® ST Chlorella microalgae composition is a microalgae composition comprising Chlorella sp. The PHYCOTERRA^® and PHYCOTERRA^® ORGANIC products contain whole cell Chlorella biomass while the PHYCOTERRA^® ST contains lysed cell Chlorella biomass. The PHY COTERRA^® Chlorella microalgae composition treatments were prepared by growing the Chlorella in non-axenic acetic acid supplied mixotrophic conditions, increasing the concentration of Chlorella using a centrifuge, pasteurizing the concentrated Chlorella at between 65°C - 75°C for between 90 - 150 minutes, adding potassium sorbate and phosphoric acid to stabilize the pH of the Chlorella, and then adjusting the whole biomass treatment to the desired concentration. The PHYCOTERRA^® Chlorella microalgae composition may comprise approximately 10% w/w of Chlorella microalgae cells. Furthermore, the PHYCOTERRA^® Chlorella microalgae composition may comprise between approximately 0.3% potassium sorbate and between approximately 0.5%-1.5% phosphoric acid to stabilize the pH of the Chlorella to between 3.0-4.0 and 88.2%-89.2% water. It should be clearly understood, however, that other variations of the PHYCOTERRA^® Chlorella microalgae composition, including variations in the microalgae strains, variations in the stabilizers, and/or variations in the % composition of each component may be used and may achieve similar results.

In some embodiments and Examples below, the microalgae composition may be an OMRI certified microalgae composition referred to as TERRENE^®. The OMRI certified TERRENE^® Chlorella microalgae composition is a microalgae composition comprising Chlorella. The OMRI certified TERRENE^® Chlorella microalgae composition treatments were prepared by growing the Chlorella in non-axenic acetic acid supplied mixotrophic conditions, increasing the concentration of Chlorella using a centrifuge, pasteurizing the concentrated Chlorella at between 65°C - 75°C for between 90 - 150 minutes, adding citric acid to stabilize the pH of the Chlorella, and then adjusting the whole biomass treatment to the desired concentration. The OMRI certified TERRENE^® Chlorella microalgae composition may comprise approximately 10% w/w of Chlorella microalgae cells. Furthermore, the OMRI certified TERRENE^® Chlorella microalgae composition may comprise between approximately 0.5% - 2.0% citric acid to stabilize the pH of the Chlorella to between 3.0-4.0 and 88%-89.5% water. It should be clearly understood, however, that other variations of the OMRI certified TERRENE^® Chlorella microalgae composition, including variations in the microalgae strains, variations in the stabilizers, and/or variations in the % composition of each component may be used and may achieve similar results.

In some embodiments and Examples below, the microalgae composition may be an OMRI certified microalgae composition referred to as OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition or as TERRENE65. The OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition is a microalgae composition comprising Chlorella. The OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition treatments were prepared by growing the Chlorella in non- axenic acetic acid supplied mixotrophic conditions, increasing the concentration of Chlorella using a centrifuge, pasteurizing the concentrated Chlorella at 65°C for between 90 - 150 minutes, adding citric acid to stabilize the pH of the Chlorella, and then adjusting the whole biomass treatment to the desired concentration. The OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition may comprise approximately 10% w/w of Chlorella microalgae cells. Furthermore, the OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition may comprise between approximately 0.5% - 2.0% citric acid to stabilize the pH of the Chlorella to between 3.0-4.0and 88-89.5% water. It should be clearly understood, however, that other variations of the OMRI certified TERRENE^® Chlorella pasteurized at 65°C microalgae composition, including variations in the microalgae strains, variations in the stabilizers, variations in the pasteurization temperature, and/or variations in the % composition of each component may be used and may achieve similar results.

In some embodiments and Examples below, the microalgae composition may be an OMRI certified microalgae composition referred to as OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition or as TERRENE90. The OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition is a microalgae composition comprising Chlorella. The OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition treatments were prepared by growing the Chlorella in non- axenic acetic acid supplied mixotrophic conditions, increasing the concentration of Chlorella using a centrifuge, pasteurizing the concentrated Chlorella at 90°C for between 90 - 150 minutes, adding citric acid to stabilize the pH of the Chlorella, and then adjusting the whole biomass treatment to the desired concentration. The OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition may comprise approximately 10% w/w of Chlorella microalgae cells. Furthermore, the OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition may comprise between approximately 0.5% - 2.0% citric acid to stabilize the pH of the Chlorella to between 3.0-4.0 and 88-89.5% water. It should be clearly understood that other variations of the OMRI certified TERRENE^® Chlorella pasteurized at 90°C microalgae composition, including variations in the microalgae strains, variations in the stabilizers, variations in the pasteurization temperature, and/or variations in the % composition of each component may be used and may achieve similar results.

A composition comprising microalgae can be stabilized by heating and cooling in a pasteurization process. In certain aspects, the active ingredients of the microalgae based compositions maintain effectiveness in enhancing at least one characteristic of a plant after being subjected to the heating and cooling of a pasteurization process. In other embodiments, compositions with whole cells or processed cells (e.g., dried, lysed, extracted) of microalgae cells may not need to be stabilized by pasteurization. For example, microalgae cells that have been processed, such as by drying, lysing, and extraction, or extracts can include such low levels of bacteria that a composition can remain stable without being subjected to the heating and cooling of a pasteurization process.

In some embodiments, the composition is lysed. Lysing is a technique where the cell membrane of a cell is ruptured, which releases lysate, the fluid contents of lysed cells, from the cells. As an example, the lysing process can comprise anything suitable that ruptures a cell membrane. For example, a bead mill may be used for lysing, where feedstock biomass solids can be dispersed and wetted (e.g., placed into a liquid phase). In this example the bead mill can utilize ceramic, glass, or metal beats (e.g., of a suitable size for the desired result) disposed in a chamber, such as a rotating cylinder, to collide with and mechanically macerate the solid biomass in the mill, which can help rupture the cell walls (e.g., the hydrogen bonds that hold together a cell membrane). Accordingly, in this example, the whole biomass may be lysed with water at cooler temperatures, with the resulting lysate comprising lipids in the form of an oil, biomass cell contents and unbroken biomass solid (e.g., non-target portion of biomass), and water.

In another aspect, the biomass is lysed using a shear mill. A shear mill utilizes a rotating impeller or high-speed rotor to create flow and shear of its contents. This causes the solid particles, such as biomass solid, to rupture due to shear stress.

Microalgae

Non-limiting examples of microalgae that can be used in the compositions, mixtures, and methods of the invention are members of one of the following divisions: Chlorophyta, Cyanophyta (Cyanobacteria), and Heterokontophyta. In certain embodiments, the microalgae used in the compositions, mixtures, and methods of the invention are members of one of the following classes: Bacillariophyceae, Eustigmatophyceae, and Chrysophyceae. In certain embodiments, the microalgae used in the compositions and methods of the invention are members of one of the following genera: Nannochloropsis, Chlorella, Desmodesmus, Dunaliella, Scenedesmus, Spirulina, Chlamydomonas , Galdieria, Isochrysis, Porphyridium, Schizochytrium, Tetraselmis, Thraustochytrium, Botryococcus , md Haematococcus .

Non-limiting examples of microalgae species that can be used in the compositions, mixtures, and methods of the present invention include: Achnanthes orientalis, Agmenellum spp., Amphiprora hyaline, Amphora coffeiformis , Amphora coffeiformis var. linea, Amphora coffeiformis var. punctata, Amphora coffeiformis var. taylori, Amphora coffeiformis var. tenuis , Amphora delicatissima, Amphora delicatissima var. capitata, Amphora sp.,Anabaena, Ankistrodesmus , Ankistrodesmus falcatus, Aurantiochytrium, sp. Boekelovia hooglandii, Borodinella sp., Botryococcus braunii, Botryococcus sudeticus, Bracteococcus minor, Bracteococcus medionucleatus , Carteria, Chaetoceros gracilis, Chaetoceros muelleri, Chaetoceros muelleri var. subsalsum, Chaetoceros sp., Chlamydomonas sp., Chlamydomas perigranulata, Chlorella anitrata, Chlorella antarctica, Chlorella aureoviridis , Chlorella Candida, Chlorella capsulate, Chlorella desiccate, Chlorella ellipsoidea, Chlorella emersonii, Chlorella fusca, Chlorella fusca var. vacuolate, Chlorella glucotropha, Chlorella infusionum, Chlorella infusionum var. actophila, Chlorella infusionum var. auxenophila, Chlorella kessleri, Chlorella lobophora, Chlorella luteoviridis , Chlorella luteoviridis var. aureoviridis, Chlorella luteoviridis var. lutescens, Chlorella miniata, Chlorella minutissima, Chlorella mutabilis, Chlorella nocturna, Chlorella ovalis, Chlorella parva, Chlorella photophila, Chlorella pringsheimii, Chlorella protothecoides, Chlorella protothecoides var. acidicola, Chlorella regularis, Chlorella regularis var. minima, Chlorella regularis var. umbricata, Chlorella reisiglii, Chlorella saccharophila, Chlorella saccharophila var. ellipsoidea, Chlorella salina, Chlorella simplex, Chlorella s or okiniana, Chlorella sp., Chlorella sphaerica, Chlorella stigmatophora, Chlorella vanniellii, Chlorella vulgaris, Chlorella vulgaris fo. tertia, Chlorella vulgaris var. autotrophica, Chlorella vulgaris var. viridis, Chlorella vulgaris var. vulgaris, Chlorella vulgaris var. vulgaris fo. tertia, Chlorella vulgaris var. vulgaris fo. viridis, Chlorella xanthella, Chlorella zofmgiensis, Chlorella trebouxioides, Chlorella vulgaris, Chlorococcum infusionum, Chlorococcum sp., Chlorogonium, Chroomonas sp., Chrysosphaera sp., Cricosphaera sp., Crypthecodinium cohnii, Cryptomonas sp., Cyclotella cryptica, Cyclotella meneghiniana, Cyclotella sp., Desmodesmus sp., Dunaliella sp., Dunaliella bardawil, Dunaliella bioculata, Dunaliella granulate, Dunaliella maritime, Dunaliella minuta, Dunaliella parva, Dunaliella peircei, Dunaliella primolecta, Dunaliella salina, Dunaliella terricola, Dunaliella tertiolecta, Dunaliella viridis, Dunaliella tertiolecta, Eremosphaera viridis, Eremosphaera sp., Ellipsoidon sp., Euglena spp., Franceia sp., Fragilaria crotonensis, Fragilaria sp., Gleocapsa sp., Gloeothamnion sp., Haematococcus pluvialis, Hymenomonas sp., Isochrysis aff. galbana, Isochrysis galbana, Lepocinclis, Micractinium, Micractinium, Monoraphidium minutum, Monoraphidium sp., Nannochloris sp., Nannochloropsis salina, Nannochloropsis sp., Navicula acceptata, Navicula biskanterae, Navicula pseudotenelloides , Navicula pelliculosa, Navicula saprophila, Navicula sp., Nephrochloris sp., Nephroselmis sp., Nitschia communis, Nitzschia alexandrina, Nitzschia closterium, Nitzschia communis, Nitzschia dissipata, Nitzschia frustulum, Nitzschia hantzschiana, Nitzschia inconspicua, Nitzschia intermedia, Nitzschia microcephala, Nitzschia pusilla, Nitzschia pusilla elliptica, Nitzschia pusilla monoensis, Nitzschia quadrangular, Nitzschia sp., Ochromonas sp., Oocystis parva, Oocystis pusilla, Oocystis sp., Oscillatoria limnetica, Oscillatoria sp., Oscillatoria subbrevis, Parachlorella kessleri, Pascheria acidophila, Pavlova sp., Phaeodactylum tricomutum, Phagus, Phormidium, Porphyridium, Platymonas sp., Pleurochrysis camerae, Pleurochrysis dentate, Pleurochrysis sp., Prototheca wickerhamii, Prototheca stagnora, Prototheca portoricensis , Prototheca moriformis, Prototheca zopfii, Pseudochlorella aquatica, Pyramimonas sp., Pyrobotrys, Rhodococcus opacus, Sarcinoid chrysophyte, Scenedesmus armatus, Schizochytrium, Spirogyra, Spirulina platensis, Stichococcus sp., Synechococcus sp., Synechocystisf Tagetes erecta, Tagetes patula, Tetraedron, Tetraselmis sp., Tetraselmis suecica, Thalassiosira weissflogii, Thraustochytrium sp., and Viridiella fridericiana.

Fertilizers

According to the invention at least the following solid fertilizers or combinations thereof may be used:

Organic fertilizers include manure, e.g. liquid manure, semi-liquid manure, liquid dung-water, biogas manure, stable manure or straw manure, slurry, sewage sludge, worm castings, peat, seaweed, compost, sewage, and guano. Green manure crops are also regularly grown to add nutrients (especially nitrogen) to the soil. Manufactured organic fertilizers include compost, blood meal, bone meal and seaweed extracts. Further examples are enzyme digested proteins, fish meal, and feather meal. The decomposing crop residue from prior years is another source of fertility.

Inorganic fertilizers are usually manufactured through chemical processes (such as the Haber-Bosch process), also using naturally occurring deposits, while chemically altering them (e.g. concentrated triple superphosphate). Naturally occurring inorganic fertilizers include Chilean sodium nitrate, mine rock phosphate, limestone, and raw potash fertilizers.

Typical solid fertilizers are in a kristallin, prilled or granulated form. Typical nitrogen containing inorganic fertilizers are ammonium nitrate, calcium ammonium nitrate, ammonium sulfate, ammonium sulfate nitrate, calcium nitrate, diammonium phosphate, monoammonium phosphate, ammonium thiosulfate and calcium cyanamide.

The inorganic fertilizer may be an NPK fertilizer. “NPK fertilizers” are inorganic fertilizers formulated in appropriate concentrations and combinations comprising the three main nutrients nitrogen (N), phosphorus (P) and potassium (K) as well as typically S, Mg, Ca, and trace elements. “NK fertilizers” comprise the two main nutrients nitrogen (N) and potassium (K) as well as typically S, Mg, Ca, and trace elements. “NP fertilizers” comprise the two main nutrients nitrogen (N) and phosphorus (P) as well as typically S, Mg, Ca, and trace elements. NPK, NK and NP fertilizers can be produced chemically or by a mixture of its single components.

Urea-containing fertilizer may be urea, formaldehyde urea, urea sulfur, urea sulfuric acid (urea sulfate), urea based NPK-fertilizers, or urea ammonium sulfate. In some aspects, the fertilizer is urea sulfuric acid (urea sulfate) such as 15/49 (i.e., a 1:1 ratio of urea to sulfuric acid: 15-0-0-16S, 49% sulfuric acid), 10/55 or 28/27. Also envisaged is the use of urea as fertilizer. A urea-containing fertilizer may comprise at least one component selected from the group consisting of urea, urea ammonium nitrate (UAN), and suspensions of isobutylidene diurea (IBDU), crotonylidene diurea (CDU) and urea formaldehyde (UF), urea-acetaldehyde, and ureaglyoxal condensates. In case urea-containing fertilizers or urea are used or provided, it is particularly preferred that urease inhibitors as below may be added or additionally be present, or be used at the same time or in connection with the urea-containing fertilizers.

In further embodiments the fertilizer mixture may be provided as, or may comprise or contain a slow release fertilizer. The fertilizer may, for example, be released over any suitable period of time, e.g. over a period of 1 to 5 months, preferably up to 3 months. Typical examples of ingredients of slow release fertilizers are IBDU (isobutylidenediurea), e.g. containing about 31-32% nitrogen, of which 90% is water insoluble; or UF, i.e. an urea-formaldehyde product which contains about 38% nitrogen of which about 70% may be provided as water insoluble nitrogen; or CDU (crotonylidene diurea) containing about 32% nitrogen; or MU (methylene urea) containing about 38 to 40% nitrogen, of which 25-60% is typically cold water insoluble nitrogen; or MDU (methylene diurea) containing about 40% nitrogen, of which less than 25% is cold water insoluble nitrogen; or DMTU (dimethylene triurea) containing about 40% nitrogen, of which less than 25% is cold water insoluble nitrogen; or TMTU (tri methylene tetraurea), which may be provided as component of UF products; or TMPU (tri methylene pentaurea), which may also be provided as component of UF products. The fertilizer mixture may also be long-term nitrogen-bearing fertilizer containing a mixture of acetylene diurea and at least one other organic nitrogen-bearing fertilizer selected from methylene urea, isobutylidene diurea, crotonylidene diurea, substituted triazones, triuret or mixtures thereof.

In one embodiment, the fertilizer is a urea-containing fertilizer, and/or P-containing fertilizer, and/or a K fertilizer (potassium-containing fertilizer), and/or a N fertilizer (nitrogen- containing fertilizer), and/or a NK fertilizer (nitrogen-potassium fertilizer), and/or a NPK (nitrogen-phosphorous-potassium fertilizer), and/or a single or dual element fertilizer containing S, Ca, Mg, Fe, Mn, Cu, Zn, Mo, B, Ni, Cl, or a combination thereof.

In various embodiments, the fertilizer is selected from the group of nitrogen, phosphate, potash, sulfur, and combinations thereof. In certain embodiments, the fertilizer is nitrogen based. Examples of suitable nitrogen based fertilizers include anhydrous ammonia, urea, ammonium nitrate, urea ammonium nitrate, calcium ammonium nitrate, and combinations thereof. In specific embodiments, the fertilizer comprises, consists essentially of, or consists of, urea. In other embodiments, the fertilizer is phosphate based. Examples of suitable phosphate based fertilizers include phosphoric acid, mono-ammonium phosphate, ammonium polyphosphate, ammonium phosphate sulfate, and combinations thereof. In yet other embodiments, the fertilizer is potash based. Examples of suitable potash based fertilizers include potash, ammonium nitrate, and combinations thereof. In yet other embodiments, the fertilizer is sulfur based. Examples of suitable sulfur based fertilizers include ammonium sulfate, sulfuric acid, and combinations thereof. Various combinations of fertilizers can be utilized.

In certain aspects, the fertilizer is potash based. In one aspect, the feritilizer is soluble potash (e.g., LIQUID CHISEL^® (soluble potash)).

P fertilizers, K fertilizers, and N fertilizers are straight fertilizers, i.e. fertilizers that contain only one of the nutritive elements P, K, and N. It is to be understood, however, that these fertilizers may additionally comprise at least one additional nutritive element selected from S, Ca, Mg, Fe, Mn, Cu, Zn, Mo, B, Ni, and Cl.

NPK fertilizers, NP fertilizers, and PK fertilizers are multinutrient fertilizers, i.e., fertilizers that comprise combinations of the nutritive elements P, K, and N as indicated by the terms “NPK”, “NP”, and “PK”. It is to be understood, however, that these fertilizers may additionally comprise at least one additional nutritive element selected from S, Ca, Mg, Fe, Mn, Cu, Zn, Mo, B, Ni and Cl. The NPK fertilizers, NP fertilizers, and PK fertilizers may be provided as complex fertilizers or bulk-blend or blended fertilizers. The term complex fertilizer refers to a compound fertilizer formed by mixing ingredients that react chemically. In bulk-blend or blended fertilizers, two or more granular fertilizers of similar size are mixed to form a compound fertilizer.

Dual element fertilizers are preferably dual element fertilizers with Ca, Mg, Fe, Mn, Zn or Ni which may be applied as soluble salts of chloride, sulfate, nitrate or in chelated form (e.g., MnEDTA, Fe EDTA, FeEDDHA). Single or dual element fertilizers of Mo are available as salts of molybdate, B as boric acid or borates.

In some embodiments, the fertilizer can be any nitrogen containing fertilizer. For example, the fertilizer can be selected from anhydrous ammonia, urea, manure, ammonium nitrate, UAN (urea ammonium nitrate), mono ammonium phosphates, diammonium phosphates, organic fertilizers or mixtures thereof.

Such nitrogen containing fertilizers are often used in combination with other types of fertilizers, such as phosphorous containing fertilizers or potassium containing fertilizers. In one embodiment, the fertilizer is anhydrous ammonia or UAN. In another embodiment, the fertilizer is anhydrous ammonia.

“Liquid ammonia” is often also referred to as “anhydrous ammonia”, since it is not applied as an aqueous solution but rather as liquified ammonia that contains no or only small amounts of water that have condensed in the liquid ammonia from the air. Normally the amount of water in “anhydrous ammonia” upon its application to the soil is below 1% by weight based on the ammonia. Liquid ammonia is normally stored in a pressurized container.

In certain aspects, the mixtures disclosed herein further comprise a micronutrient product. For example, micronutrient products can include trace elements such as copper, iron, manganese, zinc, cobalt, molybdenum, and/or boron. These trace elements can be referred to as “micronutrients” because of the relatively small amounts required by plants for growth. In some embodiments, a micronutrient product is selected from Max In Ultra ZMB, First Choice Foliar Nutrient, Foli Gro, Versa Max Soybean, Versa Max Com, Ultra Che Com Mix EDTA, ManniPlex for Beans, ManniPlex for Com, Tracite LF Row Crop Mix, and KickStand Micro Mix. In one embodiment, the micronutrient product is Max In Ultra ZMB. In another embodiment, the micronutrient product is First Choice Foliar Nutrient. In yet another embodiment, the micronutrient product is Foli Gro. In one embodiment, the micronutrient product is Versa Max Soybean. In another embodiment, the micronutrient product is Versa Max Com. In yet another embodiment, the micronutrient product is Ultra Che Com Mix EDTA. In one embodiment, the micronutrient product is ManniPlex for Beans. In another embodiment, the micronutrient product is ManniPlex for Com. In yet another embodiment, the micronutrient product is Tracite LF Row Crop Mix. In one embodiment, the micronutrient product is KickStand Micro Mix. Fertilizer Formulations

Fertilizers may be provided in any suitable form, e.g. as solid coated or uncoated granules, in liquid or semi-liquid form, as sprayable fertilizer, or via fertigation etc.

Coated fertilizers may be provided with a wide range of materials. Coatings may, for example, be applied to granular or prilled nitrogen (N) fertilizer or to multi-nutrient fertilizers. Typically, urea is used as base material for most coated fertilizers. Alternatively, ammonium or NPK fertilizers are used as base material for coated fertilizers. The present invention, however, also envisages the use of other base materials for coated fertilizers, any one of the fertilizer materials defined herein. In certain embodiments, elemental sulfur may be used as fertilizer coating. The coating may be performed by spraying molten S over urea granules, followed by an application of sealant wax to close fissures in the coating. In a further embodiment, the S layer may be covered with a layer of organic polymers, preferably a thin layer of organic polymers.

Further envisaged coated fertilizers may be provided by reacting resin-based polymers on the surface of the fertilizer granule. A further example of providing coated fertilizers includes the use of low permeability polyethylene polymers in combination with high permeability coatings.

In specific embodiments the composition and/or thickness of the fertilizer coating may be adjusted to control, for example, the nutrient release rate for specific applications. The duration of nutrient release from specific fertilizers may vary, e.g. from several weeks to many months. The presence of nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors in a mixture with coated fertilizers may accordingly be adapted. It is, in particular, envisaged that the nutrient release involves or is accompanied by the release of a nitrification inhibitor according to the present invention.

Coated fertilizers may be provided as controlled release fertilizers (CRFs). In specific embodiments these controlled release fertilizers are fully coated urea or N — P — K fertilizers, which are homogeneous and typically show a pre-defmed longevity of release. In further embodiments, the CRFs may be provided as blended controlled release fertilizer products which may contain coated, uncoated and/or slow release components. In certain embodiments, these coated fertilizers may additionally comprise micronutrients. In specific embodiments these fertilizers may show a pre-defmed longevity, e.g. in case of N — P — K fertilizers.

Additionally, envisaged examples of CRFs include patterned release fertilizers. These fertilizers typically show a pre-defmed release patterns (e.g. hi/standard/lo) and a pre- defined longevity. In exemplary embodiments fully coated N — P — K, Mg and micronutrients may be delivered in a patterned release manner.

Any of the herein mentioned fertilizers or fertilizer forms may suitably be combined.

In the terms of the present invention “mixture” or “agrochemical mixture” means a combination of at least two active compounds, such as several fertilizers or a fertilizer and a microalgae-based composition. The terms “mixture” and “agrochemical mixture” are interchangeable. The agrochemical mixture may be co-formulated or formulated separately. If the agrochemical mixture is formulated separately, the fertilizer and the microalgae-based composition are applied in a temporal relationship, i.e. simultaneously or subsequently, whereas the subsequent application is carried out within a time interval which allows the combined action of the fertilizer and microalgae-based composition on the soil.

Also envisaged are double coating approaches or coated fertilizers based on a programmed release.

In certain aspects, the fertilizer is coated with a composition comprising a culture of microalgae. The microalgae can be Aurantiochytrium, Botyococcus, Chlorella, Chlamydomonas, Scenedesmus, Pavolva, Phaeodactylum, Nannochloropsis, Spirulina, Galdieria, Haematococcus, Isochrysis, Porphyridium, Schizochytrium, Thraustochytrium, Tetraselmis or a combination thereof.

In further embodiments the fertilizer mixture may be provided as, or may comprise or contain a slow release fertilizer. The fertilizer may, for example, be released over any suitable period of time, e.g. over a period of 1 to 5 months, preferably up to 3 months. Typical examples of ingredients of slow release fertilizers are IBDU (isobutylidenediurea), e.g. containing about 31-32% nitrogen, of which 90% is water insoluble; or UF, i.e. an urea-formaldehyde product which contains about 38% nitrogen of which about 70% may be provided as water insoluble nitrogen; or CDU (crotonylidene diurea) containing about 32% nitrogen; or MU (methylene urea) containing about 38 to 40% nitrogen, of which 25-60% is typically cold water insoluble nitrogen; or MDU (methylene diurea) containing about 40% nitrogen, of which less than 25% is cold water insoluble nitrogen; or MO (methylol urea) containing about 30% nitrogen, which may typically be used in solutions; or DMTU (diimethylene triurea) containing about 40% nitrogen, of which less than 25% is cold water insoluble nitrogen; or TMTU (tri methylene tetraurea), which may be provided as component of UF products; or TMPU (tri methylene pentaurea), which may also be provided as component of UF products; or UT (urea triazone solution) which typically contains about 28% nitrogen. The fertilizer mixture may also be long- term nitrogen-bearing fertilizer containing a mixture of acetylene diurea and at least one other organic nitrogen-bearing fertilizer selected from methylene urea, isobutylidene diurea, crotonylidene diurea, substituted triazones, triuret or mixtures thereof. Any of the herein mentioned fertilizers or fertilizer forms may suitably be combined. For instance, slow release fertilizers may be provided as coated fertilizers. They may also be combined with other fertilizers or fertilizer types. The same applies to the presence of a nitrification inhibitor according to the present invention, which may be adapted to the form and chemical nature of the fertilizer and accordingly be provided such that its release accompanies the release of the fertilizer, e.g. is released at the same time or with the same frequency. The present invention further envisages fertilizer or fertilizer forms as defined herein in combination with nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors. Such combinations may be provided as coated or uncoated forms and/or as slow or fast release forms. Preferred are combinations with slow release fertilizers including a coating. In further embodiments, also different release schemes are envisaged, e.g. a slower or a faster release. Mixtures In certain aspects, the culture of microalgae in the mixture is one or more of the following: (1) Aurantiochytrium; (2) Botryococcus; (3) Chlorella; (4) Chlamydomonas; (5) Desmodesmus; (6) Dunaliella; (7) Scenedesmus; (8) Pavolv; (9) Phaeodactylum; (10) Nannochloropsis; (11) Spirulina; (12) Galdieria; (13) Haematococcus; (14) Isochrysis; (15) Porphyridium; (16) Schizochytrium; (17) Thraustochytrium; or (18) Tetraselmis .

In other aspects, the fertilizer in the mixture is one or more of the following:

(a) CAN17; (b) CN9;

(c) CaTS^® (6% calcium and 10% sulfur calcium thiosulfate fertilizer solution);

(d) AN20;

(e) AMS;

(f) ATS; (g) UN32;

(h) 10-34-0;

(i) 11-37-0;

(j) 6-26-5;

(k) 5-28-0-10 Mg; (1) 44-0-0;

(m) 14-24-0-1 OS;

(n) 17-1-0-20S;

(o) KTS^® blend (0-0-25+17S derived from potassium thiosulfate);

(p) K-ROW 23^® (0-0-23-8S derived from postassium sulfite); (q) humic acid;

(r) kelp;

(s) seaweed extract;

(t) fulvic acid;

(u) fish emulsion/fish meal; (v) soy hydrolysate;

(w) protein hydrolysate;

(x) AMINO PRIME™ (2.5-0-1 derived from sugarcane protein hydrolysate);

(y) com steep liquor;

(z) urea; (aa) urea formaldehyde;

(bb) urea-acetaldehyde;

(cc) ureaglyoxal condensate;

(dd) urea sulfur;

(ee) urea sulfuric acid (urea sulfate); (ff) urea ammonium sulfate;

(gg) isobutylidene diurea;

(hh) crotonylidene diurea;

(ii) ethylene urea;

(jj) methylene diurea;

(kk) urea ammonium nitrate;

(II) liquid ammonia;

(mm) ammonium nitrate;

(nn) calcium nitrate;

(oo) potassium nitrate;

(pp) calcium ammonium nitrate;

(qq) phosphoric acid;

(rr) ammonium phosphate;

(ss) monoammonium phosphate;

(tt) diammonium phosphate;

(uu) ammonium polyphosphate;

(vv) single super phosphate;

(ww) triple super phosphate;

(xx) ground rock phosphate;

(yy) a phosphite salt;

(zz) an ammonium iron salt;

(aaa) potassium chloride;

(bbb) muriate of potash;

(ccc) sulfate of potash;

(ddd) sulfate of potash magnesia;

(eee) soluble potash;

(fff) ammonium sulfate;

(ggg) ammonium thiosulfate;

(hhh) potassium sulfate;

(iii) ammonium sulfate-nitrate;

(jjj) calcium cyanamide;

(kkk) potassium hydroxide;

(III) potassium acetate;

(mmm) ammonium carboxylate; (nnn) cyclohexanediaminepentaacetic acid (CDTA); (ooo) citric acid (CIT); (ppp) diethylenetriaminepentaacetic acid (DTPA); (qqq) ethylenediaminediaminedi-o-hydroxyphenylacetic acid (EDDHA); (rrr) ethylenediamintetraacetic acid (EDTA); (sss) ethylene glycol bis(2-aminoethyl ether) tetraacetic acid (EGTA); (ttt) hydroxyethylenediaminetriacetic acid (HEDTA); (uuu) nitrilo-triacetic acid (NTA); (vvv) oxalic acid (OX); (www) pyrophosphoric acid (PPA); (xxx) triphosphoric acid (TPA); (yyy) compost; (zzz) manure; (aaaa) biochar; (bbbb) sugar beet or sugarcane vinasse; (cccc) sewage sludge; (dddd) blood meal; (eeee) bone meal; (ffff) worm castings; (gggg) peat moss; (hhhh) leaf compost; (iiii) vermiculite; (jjjj) perlite; (kkkk) Chilean nitrate; (llll) limestone; (mmmm) rice hull; (nnnn) coffee chaff; (oooo) buckwheat hull; (pppp) chicken manure; (qqqq) tree bark with or without composting; (rrrr) mushroom composts; or (ssss) black soldier fly frass. In certain aspects, the mixture is (1)+(a); (1)+(b); (1)+(c); (1)+(d); (1)+(e); (1)+(f); (1)+(g); (1)+(h); (1)+(i); (1)+(j); (1)+(k); (1)+(l); (1)+(m); (1)+(n); (1)+(o); (1)+(p); (1)+(q); (1)+(r); (1)+(s); (1)+(t); (1)+(u); (1)+(v); (1)+(w); (1)+(x); (1)+(y); (1)+(z); (1)+(aa); (1)+(bb); (1)+(cc); (1)+(dd); (1)+(ee); (1)+(ff); (1)+(gg); (1)+(hh); (1)+(ii); (1)+(jj); (1)+(kk); (1)+(ll); (1)+(mm); (1)+(nn); (1)+(oo); (1)+(pp); (1)+(qq); (1)+(rr); (1)+(ss); (1)+(tt); (1)+(uu); (1)+(vv); (1)+(ww); (1)+(xx); (1)+(yy); (1)+(zz); (1)+(aaa); (1)+(bbb); (1)+(ccc); (1)+(ddd); (1)+(eee); (1)+(fff); (1)+(ggg); (1)+(hhh); (1)+(iii); (1)+(jjj); (1)+(kkk); (1)+(lll); (1)+(mmm); (1)+(nnn); (1)+(ooo); (1)+(ppp); (1)+(qqq); (1)+(rrr); (1)+(sss); (1)+(ttt); (1)+(uuu); (1)+(vvv); (1)+(www); (1)+(xxx); (1)+(yyy); (1)+(zzz); (1)+(aaaa); (1)+(bbbb); (1)+(cccc); (1)+(dddd); (1)+(eeee); (1)+(ffff); (1)+(gggg); (1)+(hhhh); (1)+(iiii); (1)+(jjjj); (1)+(kkkk); (1)+(llll); (1)+(mmmm); (1)+(nnnn); (1)+(oooo); (1)+(pppp); (1)+(qqqq); (1)+(rrrr); or (1)+(ssss). In other aspects, the mixture is (2)+(a); (2)+(b); (2)+(c); (2)+(d); (2)+(e); (2)+(f); (2)+(g); (2)+(h); (2)+(i); (2)+(j); (2)+(k); (2)+(l); (2)+(m); (2)+(n); (2)+(o); (2)+(p); (2)+(q); (2)+(r); (2)+(s); (2)+(t); (2)+(u); (2)+(v); (2)+(w); (2)+(x); (2)+(y); (2)+(z); (2)+(aa); (2)+(bb); (2)+(cc); (2)+(dd); (2)+(ee); (2)+(ff); (2)+(gg); (2)+(hh); (2)+(ii); (2)+(jj); (2)+(kk); (2)+(ll); (2)+(mm); (2)+(nn); (2)+(oo); (2)+(pp); (2)+(qq); (2)+(rr); (2)+(ss); (2)+(tt); (2)+(uu); (2)+(vv); (2)+(ww); (2)+(xx); (2)+(yy); (2)+(zz); (2)+(aaa); (2)+(bbb); (2)+(ccc); (2)+(ddd); (2)+(eee); (2)+(fff); (2)+(ggg); (2)+(hhh); (2)+(iii); (2)+(jjj); (2)+(kkk); (2)+(lll); (2)+(mmm); (2)+(nnn); (2)+(ooo); (2)+(ppp); (2)+(qqq); (2)+(rrr); (2)+(sss); (2)+(ttt); (2)+(uuu); (2)+(vvv); (2)+(www); (2)+(xxx); (2)+(yyy); (2)+(zzz); (2)+(aaaa); (2)+(bbbb); (2)+(cccc); (2)+(dddd); (2)+(eeee); (2)+(ffff); (2)+(gggg); (2)+(hhhh); (2)+(iiii); (2)+(jjjj); (2)+(kkkk); (2)+(llll); (2)+(mmmm); (2)+(nnnn); (2)+(oooo); (2)+(pppp); (2)+(qqqq); (2)+(rrrr); or (2)+(ssss). In other aspects, the mixture is (3)+(a); (3)+(b); (3)+(c); (3)+(d); (3)+(e); (3)+(f); (3)+(g); (3)+(h); (3)+(i); (3)+(j); (3)+(k); (3)+(l); (3)+(m); (3)+(n); (3)+(o); (3)+(p); (3)+(q); (3)+(r); (3)+(s); (3)+(t); (3)+(u); (3)+(v); (3)+(w); (3)+(x); (3)+(y); (3)+(z); (3)+(aa); (3)+(bb); (3)+(cc); (3)+(dd); (3)+(ee); (3)+(ff); (3)+(gg); (3)+(hh); (3)+(ii); (3)+(jj); (3)+(kk); (3)+(ll); (3)+(mm); (3)+(nn); (3)+(oo); (3)+(pp); (3)+(qq); (3)+(rr); (3)+(ss); (3)+(tt); (3)+(uu); (3)+(vv); (3)+(ww); (3)+(xx); (3)+(yy); (3)+(zz); (3)+(aaa); (3)+(bbb); (3)+(ccc); (3)+(ddd); (3)+(eee); (3)+(fff); (3)+(ggg); (3)+(hhh); (3)+(iii); (3)+(jjj); (3)+(kkk); (3)+(lll); (3)+(mmm); (3)+(nnn); (3)+(ooo); (3)+(ppp); (3)+(qqq); (3)+(rrr); (3)+(sss); (3)+(ttt); (3)+(uuu); (3)+(vvv); (3)+(www); (3)+(xxx); (3)+(yyy); (3)+(zzz); (3)+(aaaa); (3)+(bbbb); (3)+(cccc); (3)+(dddd); (3)+(eeee); (3)+(ffff); (3)+(gggg); (3)+(hhhh); (3)+(iiii); (3)+(jjjj); (3)+(kkkk); (3)+(llll); (3)+(mmmm); (3)+(nnnn); (3)+(oooo); (3)+(pppp); (3)+(qqqq); (3)+(rrrr); or (3)+(ssss). In other aspects, the mixture is (4)+(a); (4)+(b); (4)+(c); (4)+(d); (4)+(e); (4)+(f); (4)+(g); (4)+(h); (4)+(i); (4)+(j); (4)+(k); (4)+(l); (4)+(m); (4)+(n); (4)+(o); (4)+(p); (4)+(q); (4)+(r); (4)+(s); (4)+(t); (4)+(u); (4)+(v); (4)+(w); (4)+(x); (4)+(y); (4)+(z); (4)+(aa); (4)+(bb); (4)+(cc); (4)+(dd); (4)+(ee); (4)+(ff); (4)+(gg); (4)+(hh); (4)+(ii); (4)+(jj); (4)+(kk); (4)+(ll); (4)+(mm); (4)+(nn); (4)+(oo); (4)+(pp); (4)+(qq); (4)+(rr); (4)+(ss); (4)+(tt); (4)+(uu); (4)+(vv); (4)+(ww); (4)+(xx); (4)+(yy); (4)+(zz); (4)+(aaa); (4)+(bbb); (4)+(ccc); (4)+(ddd); (4)+(eee); (4)+(fff); (4)+(ggg); (4)+(hhh); (4)+(iii); (4)+(jjj); (4)+(kkk); (4)+(lll); (4)+(mmm); (4)+(nnn); (4)+(ooo); (4)+(ppp); (4)+(qqq); (4)+(rrr); (4)+(sss); (4)+(ttt); (4)+(uuu); (4)+(vvv); (4)+(www); (4)+(xxx); (4)+(yyy); (4)+(zzz); (4)+(aaaa); (4)+(bbbb); (4)+(cccc); (4)+(dddd); (4)+(eeee); (4)+(ffff); (4)+(gggg); (4)+(hhhh); (4)+(iiii); (4)+(jjjj); (4)+(kkkk); (4)+(llll); (4)+(mmmm); (4)+(nnnn); (4)+(oooo); (4)+(pppp); (4)+(qqqq); (4)+(rrrr); or (4)+(ssss). In other aspects, the mixture is (5)+(a); (5)+(b); (5)+(c); (5)+(d); (5)+(e); (5)+(f); (5)+(g); (5)+(h); (5)+(i); (5)+(j); (5)+(k); (5)+(l); (5)+(m); (5)+(n); (5)+(o); (5)+(p); (5)+(q); (5)+(r); (5)+(s); (5)+(t); (5)+(u); (5)+(v); (5)+(w); (5)+(x); (5)+(y); (5)+(z); (5)+(aa); (5)+(bb); (5)+(cc); (5)+(dd); (5)+(ee); (5)+(ff); (5)+(gg); (5)+(hh); (5)+(ii); (5)+(jj); (5)+(kk); (5)+(ll); (5)+(mm); (5)+(nn); (5)+(oo); (5)+(pp); (5)+(qq); (5)+(rr); (5)+(ss); (5)+(tt); (5)+(uu); (5)+(vv); (5)+(ww); (5)+(xx); (5)+(yy); (5)+(zz); (5)+(aaa); (5)+(bbb); (5)+(ccc); (5)+(ddd); (5)+(eee); (5)+(fff); (5)+(ggg); (5)+(hhh); (5)+(iii); (5)+(jjj); (5)+(kkk); (5)+(lll); (5)+(mmm); (5)+(nnn); (5)+(ooo); (5)+(ppp); (5)+(qqq); (5)+(rrr); (5)+(sss); (5)+(ttt); (5)+(uuu); (5)+(vvv); (5)+(www); (5)+(xxx); (5)+(yyy); (5)+(zzz); (5)+(aaaa); (5)+(bbbb); (5)+(cccc); (5)+(dddd); (5)+(eeee); (5)+(ffff); (5)+(gggg); (5)+(hhhh); (5)+(iiii); (5)+(jjjj); (5)+(kkkk); (5)+(llll); (5)+(mmmm); (5)+(nnnn); (5)+(oooo); (5)+(pppp); (5)+(qqqq); (5)+(rrrr); or (5)+(ssss). In other aspects, the mixture is (6)+(a); (6)+(b); (6)+(c); (6)+(d); (6)+(e); (6)+(f); (6)+(g); (6)+(h); (6)+(i); (6)+(j); (6)+(k); (6)+(l); (6)+(m); (6)+(n); (6)+(o); (6)+(p); (6)+(q); (6)+(r); (6)+(s); (6)+(t); (6)+(u); (6)+(v); (6)+(w); (6)+(x); (6)+(y); (6)+(z); (6)+(aa); (6)+(bb); (6)+(cc); (6)+(dd); (6)+(ee); (6)+(ff); (6)+(gg); (6)+(hh); (6)+(ii); (6)+(jj); (6)+(kk); (6)+(ll); (6)+(mm); (6)+(nn); (6)+(oo); (6)+(pp); (6)+(qq); (6)+(rr); (6)+(ss); (6)+(tt); (6)+(uu); (6)+(vv); (6)+(ww); (6)+(xx); (6)+(yy); (6)+(zz); (6)+(aaa); (6)+(bbb); (6)+(ccc); (6)+(ddd); (6)+(eee); (6)+(fff); (6)+(ggg); (6)+(hhh); (6)+(iii); (6)+(jjj); (6)+(kkk); (6)+(lll); (6)+(mmm); (6)+(nnn); (6)+(ooo); (6)+(ppp); (6)+(qqq); (6)+(rrr); (6)+(sss); (6)+(ttt); (6)+(uuu); (6)+(vvv); (6)+(www); (6)+(xxx); (6)+(yyy); (6)+(zzz); (6)+(aaaa); (6)+(bbbb); (6)+(cccc); (6)+(dddd); (6)+(eeee); (6)+(ffff); (6)+(gggg); (6)+(hhhh); (6)+(iiii); (6)+(jjjj); (6)+(kkkk); (6)+(llll); (6)+(mmmm); (6)+(nnnn); (6)+(oooo); (6)+(pppp); (6)+(qqqq); (6)+(rrrr); or (6)+(ssss). In other aspects, the mixture is (7)+(a); (7)+(b); (7)+(c); (7)+(d); (7)+(e); (7)+(f); (7)+(g); (7)+(h); (7)+(i); (7)+(j); (7)+(k); (7)+(l); (7)+(m); (7)+(n); (7)+(o); (7)+(p); (7)+(q); (7)+(r); (7)+(s); (7)+(t); (7)+(u); (7)+(v); (7)+(w); (7)+(x); (7)+(y); (7)+(z); (7)+(aa); (7)+(bb); (7)+(cc); (7)+(dd); (7)+(ee); (7)+(ff); (7)+(gg); (7)+(hh); (7)+(ii); (7)+(jj); (7)+(kk); (7)+(ll); (7)+(mm); (7)+(nn); (7)+(oo); (7)+(pp); (7)+(qq); (7)+(rr); (7)+(ss); (7)+(tt); (7)+(uu); (7)+(vv); (7)+(ww); (7)+(xx); (7)+(yy); (7)+(zz); (7)+(aaa); (7)+(bbb); (7)+(ccc); (7)+(ddd); (7)+(eee); (7)+(fff); (7)+(ggg); (7)+(hhh); (7)+(iii); (7)+(jjj); (7)+(kkk); (7)+(lll); (7)+(mmm); (7)+(nnn); (7)+(ooo); (7)+(ppp); (7)+(qqq); (7)+(rrr); (7)+(sss); (7)+(ttt); (7)+(uuu); (7)+(vvv); (7)+(www); (7)+(xxx); (7)+(yyy); (7)+(zzz); (7)+(aaaa); (7)+(bbbb); (7)+(cccc); (7)+(dddd); (7)+(eeee); (7)+(ffff); (7)+(gggg); (7)+(hhhh); (7)+(iiii); (7)+(jjjj); (7)+(kkkk); (7)+(llll); (7)+(mmmm); (7)+(nnnn); (7)+(oooo); (7)+(pppp); (7)+(qqqq); (7)+(rrrr); or (7)+(ssss). In other aspects, the mixture is (8)+(a); (8)+(b); (8)+(c); (8)+(d); (8)+(e); (8)+(f); (8)+(g); (8)+(h); (8)+(i); (8)+(j); (8)+(k); (8)+(l); (8)+(m); (8)+(n); (8)+(o); (8)+(p); (8)+(q); (8)+(r); (8)+(s); (8)+(t); (8)+(u); (8)+(v); (8)+(w); (8)+(x); (8)+(y); (8)+(z); (8)+(aa); (8)+(bb); (8)+(cc); (8)+(dd); (8)+(ee); (8)+(ff); (8)+(gg); (8)+(hh); (8)+(ii); (8)+(jj); (8)+(kk); (8)+(ll); (8)+(mm); (8)+(nn); (8)+(oo); (8)+(pp); (8)+(qq); (8)+(rr); (8)+(ss); (8)+(tt); (8)+(uu); (8)+(vv); (8)+(ww); (8)+(xx); (8)+(yy); (8)+(zz); (8)+(aaa); (8)+(bbb); (8)+(ccc); (8)+(ddd); (8)+(eee); (8)+(fff); (8)+(ggg); (8)+(hhh); (8)+(iii); (8)+(jjj); (8)+(kkk); (8)+(lll); (8)+(mmm); (8)+(nnn); (8)+(ooo); (8)+(ppp); (8)+(qqq); (8)+(rrr); (8)+(sss); (8)+(ttt); (8)+(uuu); (8)+(vvv); (8)+(www); (8)+(xxx); (8)+(yyy); (8)+(zzz); (8)+(aaaa); (8)+(bbbb); (8)+(cccc); (8)+(dddd); (8)+(eeee); (8)+(ffff); (8)+(gggg); (8)+(hhhh); (8)+(iiii); (8)+(jjjj); (8)+(kkkk); (8)+(llll); (8)+(mmmm); (8)+(nnnn); (8)+(oooo); (8)+(pppp); (8)+(qqqq); (8)+(rrrr); or (8)+(ssss). In other aspects, the mixture is (9)+(a); (9)+(b); (9)+(c); (9)+(d); (9)+(e); (9)+(f); (9)+(g); (9)+(h); (9)+(i); (9)+(j); (9)+(k); (9)+(l); (9)+(m); (9)+(n); (9)+(o); (9)+(p); (9)+(q); (9)+(r); (9)+(s); (9)+(t); (9)+(u); (9)+(v); (9)+(w); (9)+(x); (9)+(y); (9)+(z); (9)+(aa); (9)+(bb); (9)+(cc); (9)+(dd); (9)+(ee); (9)+(ff); (9)+(gg); (9)+(hh); (9)+(ii); (9)+(jj); (9)+(kk); (9)+(ll); (9)+(mm); (9)+(nn); (9)+(oo); (9)+(pp); (9)+(qq); (9)+(rr); (9)+(ss); (9)+(tt); (9)+(uu); (9)+(vv); (9)+(ww); (9)+(xx); (9)+(yy); (9)+(zz); (9)+(aaa); (9)+(bbb); (9)+(ccc); (9)+(ddd); (9)+(eee); (9)+(fff); (9)+(ggg); (9)+(hhh); (9)+(iii); (9)+(jjj); (9)+(kkk); (9)+(lll); (9)+(mmm); (9)+(nnn); (9)+(ooo); (9)+(ppp); (9)+(qqq); (9)+(rrr); (9)+(sss); (9)+(ttt); (9)+(uuu); (9)+(vvv); (9)+(www); (9)+(xxx); (9)+(yyy); (9)+(zzz); (9)+(aaaa); (9)+(bbbb); (9)+(cccc); (9)+(dddd); (9)+(eeee); (9)+(ffff); (9)+(gggg); (9)+(hhhh); (9)+(iiii); (9)+(jjjj); (9)+(kkkk); (9)+(llll); (9)+(mmmm); (9)+(nnnn); (9)+(oooo); (9)+(pppp); (9)+(qqqq); (9)+(rrrr); or (9)+(ssss). In other aspects, the mixture is (10)+(a); (10)+(b); (10)+(c); (10)+(d); (10)+(e); (10)+(f); (10)+(g); (10)+(h); (10)+(i); (10)+(j); (10)+(k); (10)+(l); (10)+(m); (10)+(n); (10)+(o); (10)+(p); (10)+(q); (10)+(r); (10)+(s); (10)+(t); (10)+(u); (10)+(v); (10)+(w); (10)+(x); (10)+(y); (10)+(z); (10)+(aa); (10)+(bb); (10)+(cc); (10)+(dd); (10)+(ee); (10)+(ff); (10)+(gg); (10)+(hh); (10)+(ii); (10)+(jj); (10)+(kk); (10)+(ll); (10)+(mm); (10)+(nn); (10)+(oo); (10)+(pp); (10)+(qq); (10)+(rr); (10)+(ss); (10)+(tt); (10)+(uu); (10)+(vv); (10)+(ww); (10)+(xx); (10)+(yy); (10)+(zz); (10)+(aaa); (10)+(bbb); (10)+(ccc); (10)+(ddd); (10)+(eee); (10)+(fff); (10)+(ggg); (10)+(hhh); (10)+(iii); (10)+(jjj); (10)+(kkk); (10)+(lll); (10)+(mmm); (10)+(nnn); (10)+(ooo); (10)+(ppp); (10)+(qqq); (10)+(rrr); (10)+(sss); (10)+(ttt); (10)+(uuu); (10)+(vvv); (10)+(www); (10)+(xxx); (10)+(yyy); (10)+(zzz); (10)+(aaaa); (10)+(bbbb); (10)+(cccc); (10)+(dddd); (10)+(eeee); (10)+(ffff); (10)+(gggg); (10)+(hhhh); (10)+(iiii); (10)+(jjjj); (10)+(kkkk); (10)+(llll); (10)+(mmmm); (10)+(nnnn); (10)+(oooo); (10)+(pppp); (10)+(qqqq); (10)+(rrrr); or (10)+(ssss). In other aspects, the mixture is (11)+(a); (11)+(b); (11)+(c); (11)+(d); (11)+(e); (11)+(f); (11)+(g); (11)+(h); (11)+(i); (11)+(j); (11)+(k); (11)+(l); (11)+(m); (11)+(n); (11)+(o); (11)+(p); (11)+(q); (11)+(r); (11)+(s); (11)+(t); (11)+(u); (11)+(v); (11)+(w); (11)+(x); (11)+(y); (11)+(z); (11)+(aa); (11)+(bb); (11)+(cc); (11)+(dd); (11)+(ee); (11)+(ff); (11)+(gg); (11)+(hh); (11)+(ii); (11)+(jj); (11)+(kk); (11)+(ll); (11)+(mm); (11)+(nn); (11)+(oo); (11)+(pp); (11)+(qq); (11)+(rr); (11)+(ss); (11)+(tt); (11)+(uu); (11)+(vv); (11)+(ww); (11)+(xx); (11)+(yy); (11)+(zz); (11)+(aaa); (11)+(bbb); (11)+(ccc); (11)+(ddd); (11)+(eee); (11)+(fff); (11)+(ggg); (11)+(hhh); (11)+(iii); (11)+(jjj); (11)+(kkk); (11)+(lll); (11)+(mmm); (11)+(nnn); (11)+(ooo); (11)+(ppp); (11)+(qqq); (11)+(rrr); (11)+(sss); (11)+(ttt); (11)+(uuu); (11)+(vvv); (11)+(www); (11)+(xxx); (11)+(yyy); (11)+(zzz); (11)+(aaaa); (11)+(bbbb); (11)+(cccc); (11)+(dddd); (11)+(eeee); (11)+(ffff); (11)+(gggg); (11)+(hhhh); (11)+(iiii); (11)+(jjjj); (11)+(kkkk); (11)+(llll); (11)+(mmmm); (11)+(nnnn); (11)+(oooo); (11)+(pppp); (11)+(qqqq); (11)+(rrrr); or (11)+(ssss). In other aspects, the mixture is (12)+(a); (12)+(b); (12)+(c); (12)+(d); (12)+(e); (12)+(f); (12)+(g); (12)+(h); (12)+(i); (12)+(j); (12)+(k); (12)+(l); (12)+(m); (12)+(n); (12)+(o); (12)+(p); (12)+(q); (12)+(r); (12)+(s); (12)+(t); (12)+(u); (12)+(v); (12)+(w); (12)+(x); (12)+(y); (12)+(z); (12)+(aa); (12)+(bb); (12)+(cc); (12)+(dd); (12)+(ee); (12)+(ff); (12)+(gg); (12)+(hh); (12)+(ii); (12)+(jj); (12)+(kk); (12)+(ll); (12)+(mm); (12)+(nn); (12)+(oo); (12)+(pp); (12)+(qq); (12)+(rr); (12)+(ss); (12)+(tt); (12)+(uu); (12)+(vv); (12)+(ww); (12)+(xx); (12)+(yy); (12)+(zz); (12)+(aaa); (12)+(bbb); (12)+(ccc); (12)+(ddd); (12)+(eee); (12)+(fff); (12)+(ggg); (12)+(hhh); (12)+(iii); (12)+(jjj); (12)+(kkk); (12)+(lll); (12)+(mmm); (12)+(nnn); (12)+(ooo); (12)+(ppp); (12)+(qqq); (12)+(rrr); (12)+(sss); (12)+(ttt); (12)+(uuu); (12)+(vvv); (12)+(www); (12)+(xxx); (12)+(yyy); (12)+(zzz); (12)+(aaaa); (12)+(bbbb); (12)+(cccc); (12)+(dddd); (12)+(eeee); (12)+(ffff); (12)+(gggg); (12)+(hhhh); (12)+(iiii); (12)+(jjjj); (12)+(kkkk); (12)+(llll); (12)+(mmmm); (12)+(nnnn); (12)+(oooo); (12)+(pppp); (12)+(qqqq); (12)+(rrrr); or (12)+(ssss). In other aspects, the mixture is (13)+(a); (13)+(b); (13)+(c); (13)+(d); (13)+(e); (13)+(f); (13)+(g); (13)+(h); (13)+(i); (13)+(j); (13)+(k); (13)+(l); (13)+(m); (13)+(n); (13)+(o); (13)+(p); (13)+(q); (13)+(r); (13)+(s); (13)+(t); (13)+(u); (13)+(v); (13)+(w); (13)+(x); (13)+(y); (13)+(z); (13)+(aa); (13)+(bb); (13)+(cc); (13)+(dd); (13)+(ee); (13)+(ff); (13)+(gg); (13)+(hh); (13)+(ii); (13)+(jj); (13)+(kk); (13)+(ll); (13)+(mm); (13)+(nn); (13)+(oo); (13)+(pp); (13)+(qq); (13)+(rr); (13)+(ss); (13)+(tt); (13)+(uu); (13)+(vv); (13)+(ww); (13)+(xx); (13)+(yy); (13)+(zz); (13)+(aaa); (13)+(bbb); (13)+(ccc); (13)+(ddd); (13)+(eee); (13)+(fff); (13)+(ggg); (13)+(hhh); (13)+(iii); (13)+(jjj); (13)+(kkk); (13)+(lll); (13)+(mmm); (13)+(nnn); (13)+(ooo); (13)+(ppp); (13)+(qqq); (13)+(rrr); (13)+(sss); (13)+(ttt); (13)+(uuu); (13)+(vvv); (13)+(www); (13)+(xxx); (13)+(yyy); (13)+(zzz); (13)+(aaaa); (13)+(bbbb); (13)+(cccc); (13)+(dddd); (13)+(eeee); (13)+(ffff); (13)+(gggg); (13)+(hhhh); (13)+(iiii); (13)+(jjjj); (13)+(kkkk); (13)+(llll); (13)+(mmmm); (13)+(nnnn); (13)+(oooo); (13)+(pppp); (13)+(qqqq); (13)+(rrrr); or (13)+(ssss). In other aspects, the mixture is (14)+(a); (14)+(b); (14)+(c); (14)+(d); (14)+(e); (14)+(f); (14)+(g); (14)+(h); (14)+(i); (14)+(j); (14)+(k); (14)+(l); (14)+(m); (14)+(n); (14)+(o); (14)+(p); (14)+(q); (14)+(r); (14)+(s); (14)+(t); (14)+(u); (14)+(v); (14)+(w); (14)+(x); (14)+(y); (14)+(z); (14)+(aa); (14)+(bb); (14)+(cc); (14)+(dd); (14)+(ee); (14)+(ff); (14)+(gg); (14)+(hh); (14)+(ii); (14)+(jj); (14)+(kk); (14)+(ll); (14)+(mm); (14)+(nn); (14)+(oo); (14)+(pp); (14)+(qq); (14)+(rr); (14)+(ss); (14)+(tt); (14)+(uu); (14)+(vv); (14)+(ww); (14)+(xx); (14)+(yy); (14)+(zz); (14)+(aaa); (14)+(bbb); (14)+(ccc); (14)+(ddd); (14)+(eee); (14)+(fff); (14)+(ggg); (14)+(hhh); (14)+(iii); (14)+(jjj); (14)+(kkk); (14)+(lll); (14)+(mmm); (14)+(nnn); (14)+(ooo); (14)+(ppp); (14)+(qqq); (14)+(rrr); (14)+(sss); (14)+(ttt); (14)+(uuu); (14)+(vvv); (14)+(www); (14)+(xxx); (14)+(yyy); (14)+(zzz); (14)+(aaaa); (14)+(bbbb); (14)+(cccc); (14)+(dddd); (14)+(eeee); (14)+(ffff); (14)+(gggg); (14)+(hhhh); (14)+(iiii); (14)+(jjjj); (14)+(kkkk); (14)+(llll); (14)+(mmmm); (14)+(nnnn); (14)+(oooo); (14)+(pppp); (14)+(qqqq); (14)+(rrrr); or (14)+(ssss). In other aspects, the mixture is (15)+(a); (15)+(b); (15)+(c); (15)+(d); (15)+(e); (15)+(f); (15)+(g); (15)+(h); (15)+(i); (15)+(j); (15)+(k); (15)+(l); (15)+(m); (15)+(n); (15)+(o); (15)+(p); (15)+(q); (15)+(r); (15)+(s); (15)+(t); (15)+(u); (15)+(v); (15)+(w); (15)+(x); (15)+(y); (15)+(z); (15)+(aa); (15)+(bb); (15)+(cc); (15)+(dd); (15)+(ee); (15)+(ff); (15)+(gg); (15)+(hh); (15)+(ii); (15)+(jj); (15)+(kk); (15)+(ll); (15)+(mm); (15)+(nn); (15)+(oo); (15)+(pp); (15)+(qq); (15)+(rr); (15)+(ss); (15)+(tt); (15)+(uu); (15)+(vv); (15)+(ww); (15)+(xx); (15)+(yy); (15)+(zz); (15)+(aaa); (15)+(bbb); (15)+(ccc); (15)+(ddd); (15)+(eee); (15)+(fff); (15)+(ggg); (15)+(hhh); (15)+(iii); (15)+(jjj); (15)+(kkk); (15)+(lll); (15)+(mmm); (15)+(nnn); (15)+(ooo); (15)+(ppp); (15)+(qqq); (15)+(rrr); (15)+(sss); (15)+(ttt); (15)+(uuu); (15)+(vvv); (15)+(www); (15)+(xxx); (15)+(yyy); (15)+(zzz); (15)+(aaaa); (15)+(bbbb); (15)+(cccc); (15)+(dddd); (15)+(eeee); (15)+(ffff); (15)+(gggg); (15)+(hhhh); (15)+(iiii); (15)+(jjjj); (15)+(kkkk); (15)+(llll); (15)+(mmmm); (15)+(nnnn); (15)+(oooo); (15)+(pppp); (15)+(qqqq); (15)+(rrrr); or (15)+(ssss). In other aspects, the mixture is (16)+(a); (16)+(b); (16)+(c); (16)+(d); (16)+(e); (16)+(f); (16)+(g); (16)+(h); (16)+(i); (16)+(j); (16)+(k); (16)+(l); (16)+(m); (16)+(n); (16)+(o); (16)+(p); (16)+(q); (16)+(r); (16)+(s); (16)+(t); (16)+(u); (16)+(v); (16)+(w); (16)+(x); (16)+(y); (16)+(z); (16)+(aa); (16)+(bb); (16)+(cc); (16)+(dd); (16)+(ee); (16)+(ff); (16)+(gg); (16)+(hh); (16)+(ii); (16)+(jj); (16)+(kk); (16)+(ll); (16)+(mm); (16)+(nn); (16)+(oo); (16)+(pp); (16)+(qq); (16)+(rr); (16)+(ss); (16)+(tt); (16)+(uu); (16)+(vv); (16)+(ww); (16)+(xx); (16)+(yy); (16)+(zz); (16)+(aaa); (16)+(bbb); (16)+(ccc); (16)+(ddd); (16)+(eee); (16)+(fff); (16)+(ggg); (16)+(hhh); (16)+(iii); (16)+(jjj); (16)+(kkk); (16)+(lll); (16)+(mmm); (16)+(nnn); (16)+(ooo); (16)+(ppp); (16)+(qqq); (16)+(rrr); (16)+(sss); (16)+(ttt); (16)+(uuu); (16)+(vvv); (16)+(www); (16)+(xxx); (16)+(yyy); (16)+(zzz); (16)+(aaaa); (16)+(bbbb); (16)+(cccc); (16)+(dddd); (16)+(eeee); (16)+(ffff); (16)+(gggg); (16)+(hhhh); (16)+(iiii); (16)+(jjjj); (16)+(kkkk); (16)+(llll); (16)+(mmmm); (16)+(nnnn); (16)+(oooo); (16)+(pppp); (16)+(qqqq); (16)+(rrrr); or (16)+(ssss). In other aspects, the mixture is (17)+(a); (17)+(b); (17)+(c); (17)+(d); (17)+(e); (17)+(f); (17)+(g); (17)+(h); (17)+(i); (17)+(j); (17)+(k); (17)+(l); (17)+(m); (17)+(n); (17)+(o); (17)+(p); (17)+(q); (17)+(r); (17)+(s); (17)+(t); (17)+(u); (17)+(v); (17)+(w); (17)+(x); (17)+(y); (17)+(z); (17)+(aa); (17)+(bb); (17)+(cc); (17)+(dd); (17)+(ee); (17)+(ff); (17)+(gg); (17)+(hh); (17)+(ii); (17)+(jj); (17)+(kk); (17)+(ll); (17)+(mm); (17)+(nn); (17)+(oo); (17)+(pp); (17)+(qq); (17)+(rr); (17)+(ss); (17)+(tt); (17)+(uu); (17)+(vv); (17)+(ww); (17)+(xx); (17)+(yy); (17)+(zz); (17)+(aaa); (17)+(bbb); (17)+(ccc); (17)+(ddd); (17)+(eee); (17)+(fff); (17)+(ggg); (17)+(hhh); (17)+(iii); (17)+(jjj); (17)+(kkk); (17)+(lll); (17)+(mmm); (17)+(nnn); (17)+(ooo); (17)+(ppp); (17)+(qqq); (17)+(rrr); (17)+(sss); (17)+(ttt); (17)+(uuu); (17)+(vvv); (17)+(www); (17)+(xxx); (17)+(yyy); (17)+(zzz); (17)+(aaaa); (17)+(bbbb); (17)+(cccc); (17)+(dddd); (17)+(eeee); (17)+(ffff); (17)+(gggg); (17)+(hhhh); (17)+(iiii); (17)+(jjjj); (17)+(kkkk); (17)+(llll); (17)+(mmmm); (17)+(nnnn); (17)+(oooo); (17)+(pppp); (17)+(qqqq); (17)+(rrrr); or (17)+(ssss). In other aspects, the mixture is (18)+(a); (18)+(b); (18)+(c); (18)+(d); (18)+(e); (18)+(f); (18)+(g); (18)+(h); (18)+(i); (18)+(j); (18)+(k); (18)+(l); (18)+(m); (18)+(n); (18)+(o); (18)+(p); (18)+(q); (18)+(r); (18)+(s); (18)+(t); (18)+(u); (18)+(v); (18)+(w); (18)+(x); (18)+(y); (18)+(z); (18)+(aa); (18)+(bb); (18)+(cc); (18)+(dd); (18)+(ee); (18)+(ff); (18)+(gg); (18)+(hh); (18)+(ii); (18)+(jj); (18)+(kk); (18)+(ll); (18)+(mm); (18)+(nn); (18)+(oo); (18)+(pp); (18)+(qq); (18)+(rr); (18)+(ss); (18)+(tt); (18)+(uu); (18)+(vv); (18)+(ww); (18)+(xx); (18)+(yy); (18)+(zz); (18)+(aaa); (18)+(bbb); (18)+(ccc); (18)+(ddd); (18)+(eee); (18)+(fff); (18)+(ggg); (18)+(hhh); (18)+(iii); (18)+(jjj); (18)+(kkk); (18)+(lll); (18)+(mmm); (18)+(nnn); (18)+(ooo); (18)+(ppp); (18)+(qqq); (18)+(rrr); (18)+(sss); (18)+(ttt); (18)+(uuu); (18)+(vvv); (18)+(www); (18)+(xxx); (18)+(yyy); (18)+(zzz); (18)+(aaaa); (18)+(bbbb); (18)+(cccc); (18)+(dddd); (18)+(eeee); (18)+(ffff); (18)+(gggg); (18)+(hhhh); (18)+(iiii); (18)+(jjjj); (18)+(kkkk); (18)+(llll); (18)+(mmmm); (18)+(nnnn); (18)+(oooo); (18)+(pppp); (18)+(qqqq); (18)+(rrrr); or (18)+(ssss). In certain aspects, the ratio by weight of cultured microalgae cells to fertilizer is from 1:500 to 500:1, 1:500 to 400:1, 1:500 to 300:1, 1:500 to 200:1, 1:500 to 100:1, 1:500 to 50:1, 1:500 to 10:1, 1:250 to 500:1, 1:250 to 400:1, 1:250 to 300:1, 1:250 to 250:1, 1:250 to 200:1, 1:250 to 100:1, 1:250 to 50:1, 1:100 to 500:1, 1:100 to 400:1, 1:100 to 300:1, 1:100 to 200:1, 1:100 to 100:1, 1:100 to 50:1, 1:10 to 500:1, 1:10 to 400:1, 1:10 to 400:1, 1:10 to 300:1, 1:10 to 200:1, 1:10 to 100:1, 1:10 to 50:1, 1:10 to 500:1, 1:10 to 400:1, 1:10 to 300:1, 1:10 to 200:1, 1:10 to 100:1, 1:10 to 10:1, 1:1 to 500:1, 1:1 to 400:1, 1:1 to 300:1, 1:1 to 200:1, 1:1 to 100:1, 1:1 to 50:1, or 1:1 to 10:1. In one aspect, the ratio by weight of cultured microalgae cells to fertilizer is from 1:500 to 500:1. In another aspect, the ratio by weight of cultured microalgae cells to fertilizer is from 1:100 to 100:1. In one aspect, the fertilizer is ESN^®, or Environmentally Smart Nitrogen, a slow-release nitrogen fertilizer containing polymer-coated urea with an analysis of 44-0-0. In another aspect, the fertilizer is CRYSTALGREEN^®, a slow-release granule nitrogen fertilizer with an analysis of 5-28-0-10 Mg. In another aspect, the fertilizer is SymTRX12S^TM a slow-release nitrogen fertilizer with an analysis of 14-24-0-10S. In another aspect, the fertilizer is SymTRX20S^TM, a slow-release nitrogen fertilizer with an analysis of 17-1-0-20S. In another aspect, the fertilizer is Organic MaTRX^TM, an organic complexed ammonium sulfate and ammonium iron with organic content from municipal biosolids, food waste digestate and/or animal residuals. Nitrification Inhibitors Nitrification of nitrogen containing compounds is a phenomenon that reduces the efficiency of fertilization of soil with nitrogen containing compounds. Through nitrification, N-containing compounds are decomposed by bacteria. Thereby, the nitrogen contained therein is oxidized and is no longer available for take-up by the crops. One common approach to reduce nitrification is to apply nitrification inhibitors to the soil. US 2003/14561 discloses the use of pyrazoles like 3,4-dimethylpyrazole (DMP) as nitrification inhibitors. WO 2017/069828 discloses formulations of nitrification inhibitors with solvent mixtures and corrosion inhibitors. WO 2015/81116 discloses formulations of nitrification/urease inhibitors in organic liquid solvating systems comprising a mixture of aprotic solvents.

In some embodiments, the compositions and mixtures disclosed herein include a nitrification inhibitor as an additional component. The nitrification inhibitor can in principle be any compound capable of reducing the activity of bacteria in the nitrification process. Fertilizers which are suitable to combine with nitrification inhibitors include urea and/or ammonium-containing N-organic and inorganic fertilizers.

In some embodiments, the nitrification inhibitor is selected from pyrazoles like 3,4- dimethyl- 1-H-pyrazole (DMP), 2-Chloro-6-(trichloromethyl)pyridine (Nitrapyrin), dicyandiamide, ammoniumthiosulfate, or mixtures thereof. In other embodiments, the nitrification inhibitor in selected from nitrapyrin, DMP or mixtures thereof. In one embodiment, the nitrification inhibitor is DMP.

Examples of envisaged of alternative or additional nitrification inhibitors are linoleic acid, alpha-linolenic acid, methyl p-coumarate, methyl ferulate, methyl 3-(4-hydroxyphenyl) propionate (MHPP), Karanjin, brachialacton, p-benzoquinone sorgoleone, 2-chloro-6- (trichloromethyl)-pyridine (nitrapyrin or N-serve), dicyandiamide (DCD, 3,4-dimethyl pyrazole (DMP), 3,4-dimethyl pyrazole derivatives, 3,4-dimethyl pyrazole phosphate (DMPP, ENTEC), 4-amino- 1,2, 4-triazole hydrochloride (ATC), l-amido-2-thiourea (ASU), 2-amino- 4-chloro-6-methylpyrimidine (AM), 2-mercapto-benzothiazole (MBT), 5 -ethoxy-3 - trichloromethyl-l,2,4-thiodiazole (terrazole, etri diazole), 2-sulfanilamidothiazole (ST), ammoniumthiosulfate (ATU), 3-methylpyrazol (3-MP), 3,5-dimethylpyrazole (DMP), 1,2,4- triazol thiourea (TU), N-(lH-pyrazolyl-methyl)acetamides such as N-((3(5)-methyl-lH- pyrazole-l-yl)methyl)acetamide, and N-(lH-pyrazolyl-methyl)formamides such as N-((3(5)- methyl-lH-pyrazole-l-yl)methyl formamide, N-(4-chloro-3(5)-methyl-pyrazole-l-ylmethyl)- formamide, N-(3 (5),4-dimethyl-pyrazole-l-ylmethyl)-formamide, mixtures of 3,4- dimethylpyrazole phosphate succinic acid and 4,5-dimethylpyrazole phosphate succinic acid, neem, products based on ingredients of neem, cyan amide, melamine, zeolite powder, catechol, benzoquinone, sodium tetra borate, and zinc sulfate.

In certain aspects, the composition or mixture disclosed herein comprises at least one nitrification inhibitor selected from the group consisting of 2-(3,4-dimethyl-pyrazol-l-yl)- succinic acid, 3,4-dimethyl pyrazole (DMP), 3,4-dimethyl pyrazole derivatives, 3,4- dimethylpyrazolephosphate (DMPP), dicyandiamide (DCD), 1H- 1,2, 4-triazole, 3- methylpyrazole (3-MP), 2-chloro-6-(trichloromethyl)-pyridine, 5-ethoxy-3-trichloromethyl- 1,2,4-thiadiazol, 2-amino-4-chloro-6-methyl-pyrimidine, 2-mercapto-benzothiazole, 2- sulfanilamidothiazole, thiourea, sodium azide, potassium azide, 1-hydroxypyrazole, 2- methylpyrazole-1 -carboxamide, 4-amino- 1,2, 4-triazole, 3-mercapto-l, 2, 4-triazole, 2,4- diamino-6-trichloromethyl-5-triazine, carbon bisulfide, ammonium thiosulfate, sodium trithiocarbonate, 2,3-dihydro-2,2-dimethyl-7-benzofuranol methyl carbamate and N-(2,6- dimethylphenyl)-N-(methoxyacetyl)-alanine methyl ester.

Denitrification Inhibitors

In some embodiments, the compositions and mixtures disclosed herein include a denitrification inhibitor as an additional component. Fertilizers which are suitable to combine with denitrification inhibitors include all N-containing fertilizers. Examples of denitrification inhibitors are strobilurin components selected from the group consisting of pyraclostrobin, azoxystrobin, dimoxystrobin, enestroburin, fluoxastrobin, kresoxim-methyl, metominostrobin, orysastrobin, picoxystrobin, trifloxystrobin, pyrametostrobin, pyraoxystrobin, coumoxystrobin, coumethoxystrobin, fenaminostrobin (=diclofenoxystrobin), flufenoxystrobin, 2-(2-(6-(3-chloro-2-methyl-phenoxy)-5-fluoro-pyrimidin-4-yloxy)-phenyl)- 2-methoxyimino-N-methyl-acetamide, 3-methoxy-2-(2-(N-(4-methoxy-phenyl)- cyclopropane-carboximidoylsulfanyl-methyl)-phenyl)-acrybc acid methyl ester, methyl (2- chloro-5-[l-(3-methylbenzyloxyimino)ethyl]benzyl)carbamate and 2-(2-(3-(2,6- dichlorophenyl)-l-methyl-allybdeneaminooxymethyl)-phenyl)-2-methoxyimino-N methyl- acetamide.

Urease Inhibitors

In some embodiments, the compositions and mixtures disclosed herein include a urease inhibitor as an additional component. The urea-containing fertilizers disclosed herein may be used together with a urease inhibitor. Urease is an enzyme which hydrolyzes urea to ammonia and carbon dioxide. In agriculture, high urease activity during treatment with urea- containing fertilizers causes significant environmental and economic problems due to the release of ammonia which may be toxic to the plants and which deprives the plants of urea. Accordingly, it is desirable to inhibit the action of urease. Inhibitors of urease activity may comprise (i) substrate structural analogs of urea as e.g. hydroxyurea or hydroxamic acid or (ii) inhibitors which affect the mechanism of the urease reaction. The latter may be divided in the four groups of (i) phosphorodiamidates or phosphorotriamidiates as e.g., N-(n-butyl)thiophosphoric triamide, (ii) thiols as e.g., cysteamine, (iii) hydroxamic acids and its derivatives as e.g. acetohydroxamic acid, and (iv) ligands and chelators of the nickel ion in the active center of ureases as e.g., fluoride ions. Urease inhibitors are also discussed in Upadhyay (2012) Ind. J. Biotechnol. 1 1 : 381-388 and in "Improving Efficiency of Urea Fertilizers by Inhibition of Soil Urease Activity" by Kiss and Simihaian (2002), Springer Netherlands, ISBN 978-1-4020- 0493-3.

Additional examples of envisaged urease inhibitors include N-(n-butyl) thiophosphoric acid triamide (NBPT, Agrotain), N-(n-propyl) thiophosphoric acid triamide (NPPT), 2- nitrophenyl phosphoric triamide (2-NPT), further NXPTs known to the skilled person, phenylphosphorodiamidate (PPD/PPDA), hydroquinone, ammonium thiosulfate, and mixtures ofNBPT and NPPT (see e.g. U.S. Pat. No. 8,075,659). Such mixtures ofNBPT andNPPT may comprise NBPT in amounts of from 40 to 95% wt.-% and preferably of 60 to 80% wt.-% based on the total amount of active substances. Such mixtures are marketed as LIMUS, which is a composition comprising about 16.9 wt.-% NBPT and about 5.6 wt.-% NPPT and about 77.5 wt.-% of other ingredients including solvents and adjuvants. Furthermore, urease inhibitors can be neem and products based on ingredients of neem. Particularly preferably, the composition comprises NBPT and NPPT, wherein NBPT is present in amounts of from 1 to 99.99 wt. %, more preferably from 10 to 99.9 wt. %, most preferably from 20 to 99 wt. %, particularly preferably from 30 to 98 wt. %, more particularly preferably from 40 to 95 wt. %, most particularly preferably from 50 to 90 wt. %, especially from 60 to 85 wt. %, especially preferably from 72 to 80 wt. %, for example from 74 to 77 wt. %, in each case based on the total weight of the (thio)phosphoric acid triamides contained in the composition.

Kit of Parts

According to one embodiment, individual components of the composition according to the invention such as parts of a kit or parts of a binary mixture may be mixed by the user himself in a spray tank or any other kind of vessel used for applications (e.g., seed treater drums, seed pelleting machinery, knapsack sprayer) and further auxiliaries may be added, if appropriate. Consequently, one embodiment of the invention is a kit for preparing an agricultural composition, the kit comprising the individual components in a mixture comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; as defined herein, in a spatially separated arrangement, and at least one auxiliary.

Moreover, the kit of parts according to the present invention can additionally comprise at least one auxiliary selected from the group consisting of extenders, solvents, spontaneity promoters, carriers, emulsifiers, dispersants, frost protectants, thickeners and adjuvants. Thus, at least one auxiliary can be present either in the microalgae component of the kit of parts or in the fertilizer component of the kit of parts being spatially separated or in both of these components.

Methods of Application

The term “fertigation” as used herein refers to the application of fertilizers, optionally soil amendments, and optionally other water-soluble products together with water through an irrigation system to a plant or to the locus where a plant is growing or is intended to grow, or to a soil substituent as defined herein below. For example, liquid fertilizers or dissolved fertilizers may be provided via fertigation directly to a plant or a locus where a plant is growing or is intended to grow. Likewise, nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors may be provided via fertigation to plants or to a locus where a plant is growing or is intended to grow. Fertilizers with nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors may be provided together, e.g. dissolved in the same charge or load of material (typically water) to be irrigated. In further embodiments, fertilizers with nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors may be provided at different points in time. For example, the fertilizer may be ferti gated first, followed by the nitrification inhibitor, or preferably, the nitrification inhibitor may be fertigated first, followed by the fertilizer. The time intervals for these activities follow the herein outlined time intervals for the application of fertilizers with nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors. Also envisaged is a repeated fertigation of fertilizers with nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors according to the present invention, either together or intermittently, e.g. every 2 hours, 6 hours, 12 hours, 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days or more.

According to a preferred embodiment of the present invention the application of the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors and of said fertilizer as defined herein is carried out simultaneously or with a time lag. The term “time lag” as used herein means that the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors are applied before the fertilizer to the plant growing on soil or soil substituents and/or the locus where the plant is growing or is intended to grow; or the fertilizer is applied before the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors to the plant growing on soil or soil substituents and/or the locus where the plant is growing or is intended to grow. Such time lag may be any suitable period of time which still allows to provide a nitrification, denitrification, and/or urease inhibiting effect in the context of fertilizer usage. For example, the time lag may be a time period of 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months or more or any time period in between the mentioned time periods. Preferably, the time lag is an interval of 1 day, 2 days, 3 days, 1 week, 2 weeks or 3 weeks. The time lag preferably refers to situations in which the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors as defined herein are provided 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks,

11 weeks, 12 weeks, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months or more or any time period in between the mentioned time periods before the application of a fertilizer as defined herein.

In another specific embodiment of the invention the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors are applied between GS 00 to GS 33 BBCH of the plant, or between GS 00 and GS 65 BBCH of the plant, provided that the application of at least one fertilizer as defined herein is carried out with a time lag of at least 1 day, e.g. a time lag of 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days,

12 days, 13 days, 14 days, 3 weeks 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, or more or any time period in between the mentioned time periods. It is preferred that the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors, which are applied between GS 00 to GS 33 BBCH of the plant, are provided 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, or 12 weeks before the application of a fertilizer as defined herein.

In another specific embodiment of the invention, at least one fertilizer as defined herein is applied between GS 00 to GS 33 BBCH of the plant or between GS 00 and GS 65 BBCH of the plant, provided that the application of the nitrification inhibitors, denitrification inhibitors, and/or urease inhibitors as defined herein is carried out with a time lag of at least 1 day, e.g. a time lag of 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks or more or any time period in between the mentioned time periods.

According to certain aspects of the invention, a first composition comprising a culture of microalgae is applied to the soil in combination with a second composition comprising a fertilizer. “In combination” in context shall mean that the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil simultaneously or with a time span of no more than 14 days, preferably no more than 7 days or 3 days.

In some embodiments, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil simultaneously. “Simultaneously” in this context means that the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are mixed before being applied to the soil or that they are applied within a time span of less than 30 seconds, preferably less than 10 seconds, for example through separate application nozzles.

The application of the first composition comprising a culture of microalgae and the second composition comprising a fertilizer can for example be by spraying or by injection in the soil and optionally knifing.

In one embodiment, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil by a device that is physically connected to a container comprising the first composition comprising a culture of microalgae and to another container comprising the second composition comprising a fertilizer, preferably in liquid form.

In one embodiment, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil by a device that is physically connected to a container comprising the first composition comprising a culture of microalgae and to another container comprising the second composition comprising a fertilizer, preferably in liquid form, wherein the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are continuously mixed during the application in a mixing device, for example a mixing chamber, and wherein the so obtained mixture of the first composition and the second composition is then applied to the soil through a nozzle (“inline mixing”).

In one embodiment, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil by a device that is physically connected to a container comprising the first composition comprising a culture of microalgae and to another container comprising the second composition comprising a fertilizer, preferably in liquid form, wherein the first composition and the second composition are applied to the soil simultaneously through separate nozzles (“co-injection”).

In one embodiment, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied to the soil after preparing a tank mix, meaning that a mixture of the first composition comprising a culture of microalgae and the second composition comprising a fertilizer, preferably in liquid form, is prepared before its application. In this embodiment, the first composition comprising a culture of microalgae and the second composition comprising a fertilizer are applied by a device that does not continuously mix the two compositions but rather bears a tank comprising a readily prepared tank mix of the two compositions, preferably in liquid form, that is than applied to the soil, for example by injection into the soil.

In some embodiments, the first composition comprising a culture of microalgae can include 2.5-30% solids by weight of microalgae cells (i.e., 2.5-30 g of microalgae cells/100 mL of the composition). In some embodiments, the composition can include 2.5-5% solids by weight of microalgae cells (i.e., 2.5-5 g of microalgae cells/100 mL of the composition). In some embodiments, the composition can include 5-20% solids by weight of microalgae cells. In some embodiments, the composition can include 5-15% solids by weight of microalgae cells. In some embodiments, the composition can include 5-10% solids by weight of microalgae cells. In some embodiments, the composition can include 10-20% solids by weight of microalgae cells. In some embodiments, the composition can include 10-20% solids by weight of microalgae cells. In some embodiments, the composition can include 20-30% solids by weight of microalgae cells. In some embodiments, further dilution of the microalgae cells percent solids by weight can occur before application for low concentration applications of the composition.

In some embodiments, the composition can include less than 1% by weight of microalgae biomass or extracts (i.e., less than 1 g of microalgae derived product/100 mL of the composition). In some embodiments, the composition can include less than 0.9% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.8% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.7% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.6% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.5% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.4% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.3% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.2% by weight of microalgae biomass or extracts. In some embodiments, the composition can include less than 0.1% by weight of microalgae biomass or extracts. In some embodiments, the composition can include at least 0.0001% by weight of microalgae biomass or extracts. In some embodiments, the composition can include at least 0.001% by weight of microalgae biomass or extracts. In some embodiments, the composition can include at least 0.01% by weight of microalgae biomass or extracts. In some embodiments, the composition can include at least 0.1% by weight of microalgae biomass or extracts. In some embodiments, the composition can include 0.0001-1% by weight of microalgae biomass or extracts. In some embodiments, the composition can include 0.0001-0.001% by weight of microalgae biomass or extracts. In some embodiments, the composition can include 0.001-.01% by weight of microalgae biomass or extracts. In some embodiments, the composition can include 0.01-0.1% by weight of microalgae biomass or extracts. In some embodiments, the composition can include 0.1-1% by weight of microalgae biomass or extracts.

In some embodiments, an application concentration of 0.1% of microalgae biomass or extract equates to 0.04 g of microalgae biomass or extract in 40 mL of a composition. While the desired application concentration to a plant can be 0.1% of microalgae biomass or extract, the composition can be packaged as a 10% concentration (0.4 mL in 40 mL of a composition). Thus, a desired application concentration of 0.1% would require 6,000 mL of the 10% microalgae biomass or extract in the 100 gallons of water applied to the assumption of 15,000 plants in an acre, which is equivalent to an application rate of about 1.585 gallons per acre. In some embodiments, a desired application concentration of 0.01% of microalgae biomass or extract using a 10% concentration composition equates to an application rate of about 0.159 gallons per acre. In some embodiments, a desired application concentration of 0.001% of microalgae biomass or extract using a 10% concentration composition equates to an application rate of about 0.016 gallons per acre. In some embodiments, a desired application concentration of 0.0001% of microalgae biomass or extract using a 10% concentration composition equates to an application rate of about 0.002 gallons per acre.

In another non-limiting embodiment, correlating the application of the microalgae biomass or extract on a per plant basis using the assumption of 15,000 plants per acre, the composition application rate of 1 gallon per acre is equal to about 0.25 mL per plant = 0.025 g per plant = 25 mg of microalgae biomass or extract per plant. The water requirement assumption of 100 gallons per acre is equal to about 35 mL of water per plant. Therefore, 0.025 g of microalgae biomass or extract in 35 mL of water is equal to about 0.071 g of microalgae biomass or extract per 100 mL of composition equates to about a 0.07% application concentration. In some embodiments, the microalgae biomass or extract based composition can be applied at a rate in a range as low as about 0.001-10 gallons per acre, or as high as up to 150 gallons per acre.

In some embodiments, the applications are performed using a 10% solids solution by weight microalgae composition. For greenhouse trials, the rates vary and essentially refer to how much volume of the 10% solids solution are added in a given volume of water (e.g. 2.5% v/v - 5.0% v/v).

Additionally, the present invention is directed to a method of treating a plant, a plant part, such as a seed, root, rhizome, corm, bulb, or tuber, and/or a locus on which or near which the plant or the plant parts grow, such as soil, to enhance plant growth and/or yield comprising the step of simultaneously or sequentially applying to a plant, a plant part and/or a plant loci: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer.

The compositions disclosed herein may be applied in any desired manner, such as in the form of a seed coating, soil drench, and/or directly in-furrow and/or as a foliar spray and applied either pre-emergence, post-emergence or both. In other words, the compositions can be applied to the seed, the plant or to the soil wherein the plant is growing or wherein it is desired to grow (plant's locus of growth).

In some embodiments, the microalgae based composition and/or the fertilizer composition may be applied to soil, seeds, and plants in an in-furrow application. An application of the microalgae based composition and/or the fertilizer composition in-furrow requires a low amount of water and targets the application to a small part of the field. The application in-furrow also concentrates the application of the compositions at a place where the seedling radicles and roots will pick up the material in the compositions or make use of captured nutrients, including phytohormones.

In some embodiments, the microalgae based composition and/or the fertilizer composition may be applied to soil, seeds, and plants as a side dress application. One of the principals of plant nutrient applications is to concentrate the nutrients in an area close to the root zone so that the plant roots will encounter the nutrients as the plant grows. Side-dress applications use a “knife” that is inserted into the soil and delivers the nutrients around 2 inches along the row and about 2 inches or more deep. Side-dress applications are made when the plants are young and prior to flowering to support yield. Side-dress applications can only be made prior to planting in drilled crops, i.e. wheat and other grains, and alfalfa, but in row crops such as peppers, com, tomatoes they can be made after the plants have emerged.

In some embodiments, the microalgae based composition and/or the fertilizer composition may be applied to soil, seeds, and plants through a drip system. Depending on the soil type, the relative concentrations of sand, silt and clay, and the root depth, the volume that is irrigated with a drip system may be about ½ of the total soil volume. The soil has an approximate weight of 4,000,000 lbs. per acre one foot deep. Because the roots grow where there is water, the plant nutrients in the microalgae based composition would be delivered to the root system where the nutrients will impact most or all of the roots. Experimental testing of different application rates to develop a rate curve would aid in determining the optimum rate application of the compositions in a drip system application.

In some embodiments, the microalgae based composition and/or the fertilizer composition may be applied to soil, seeds, and plants through a pivot irrigation application. The quantity and frequency of water delivered over an area by a pivot irrigation system is dependent on the soil type and crop. Applications may be 0.5 inch or more and the exact demand for water can be quantitatively measured using soil moisture gauges. For crops such as alfalfa that are drilled in (very narrow row spacing), the roots occupy the entire soil area. Penetration of the soil by the microalgae based composition may vary with a pivot irrigation application but would be effective as long as the application can target the root system of the plants. In some embodiments, the microalgae based composition and/or the fertilizer composition may be applied in a broadcast application to plants with a high concentration of plants and roots, such as row crops.

In certain aspects, the microalgae based composition and/or the fertilizer composition are applied at 0.1-150 gallons per acre, 0.1-50 gallons per acre, or 0.1-10 gallons per acre.

The present invention involves the use of a microalgae composition. Microalgae compositions, methods of preparing microalgae compositions, and methods of applying the microalgae compositions to plants are disclosed in WO 2017/218896 A1 (Shinde et al.) entitled “Microalgae-Based Composition, and Methods of its Preparation and Application to Plants,” which is incorporated herein in full by reference. In one or more embodiments, the microalgae composition may comprise approximately 10%-10.5% w/w of Chlorella microalgae cells. In one or more embodiments, the microalgae composition may also comprise one of more stabilizers, such as potassium sorbate, phosphoric acid, ascorbic acid, sodium benzoate, citric acid, or the like, or any combination thereof. For example, in one or more embodiments, the microalgae composition may comprise approximately .3% w/w of potassium sorbate or another similar compound to stabilize its pH and may further comprise approximately 0.5-1.5% w/w phosphoric acid or another similar compound to prevent the growth of contaminants. As a further example, in one or more embodiments where it is desired to use an OMRI (Organic Materials Review Institute) certified organic composition, the microalgae composition may comprise 1.0-2.0% w/w citric acid to stabilize its pH, and may not contain potassium sorbate or phosphoric acid. In one or more embodiments, the pH of the microalgae composition may be stabilized to between 3.0-4.0.

In some embodiments, the composition is a liquid and substantially includes water. In some embodiments, the composition can include 70-99% water. In some embodiments, the composition can include 85-95% water. In some embodiments, the composition can include 70-75% water. In some embodiments, the composition can include 75-80% water. In some embodiments, the composition can include 80-85% water. In some embodiments, the composition can include 85-90% water. In some embodiments, the composition can include 90- 95% water. In some embodiments, the composition can include 95-99% water. The liquid nature and high-water content of the composition facilitates administration of the composition in a variety of manners, such as but not limit to: flowing through an irrigation system, flowing through an above ground drip irrigation system, flowing through a buried drip irrigation system, flowing through a central pivot irrigation system, sprayers, sprinklers, and water cans.

In some embodiments, administration of the microalgae based composition and/or the fertilizer composition to soil, a seed or plant can be in an amount effective to produce an enhanced characteristic in plants compared to a substantially identical population of untreated seeds or plants. Such enhanced characteristics can include accelerated seed germination, accelerated seedling emergence, improved seedling emergence, improved leaf formation, accelerated leaf formation, improved plant maturation, accelerated plant maturation, increased plant yield, increased plant growth, increased plant quality, increased plant health, increased fruit yield, increased fruit sweetness, increased fruit growth, and increased fruit quality. Non limiting examples of such enhanced characteristics can include accelerated achievement of the hypocotyl stage, accelerated protrusion of a stem from the soil, accelerated achievement of the cotyledon stage, accelerated leaf formation, increased marketable plant weight, increased marketable plant yield, increased marketable fruit weight, increased production plant weight, increased production fruit weight, increased utilization (indicator of efficiency in the agricultural process based on ratio of marketable fruit to unmarketable fruit), increased chlorophyll content (indicator of plant health), increased plant weight (indicator of plant health), increased root weight (indicator of plant health), increased shoot weight (indicator of plant health), increased plant height, increased thatch height, increased resistance to salt stress, increased plant resistance to heat stress (temperature stress), increased plant resistance to heavy metal stress, increased plant resistance to drought, increased plant resistance to disease, improved color, reduced insect damage, reduced blossom end rot, and reduced sun bum. Such enhanced characteristics can occur individually in a plant, or in combinations of multiple enhanced characteristics.

In some embodiments, the microalgae based composition and/or the fertilizer composition can be administered before the seed is planted. In some embodiments, the microalgae based composition and/or the fertilizer composition can be administered at the time the seed is planted. In some embodiments, the microalgae based composition and/or the fertilizer composition can be applied by dip treatment of the roots. In some embodiments, the microalgae based composition and/or the fertilizer composition can be administered to plants that have emerged from the ground.

In another non-limiting embodiment, the administration of the microalgae based composition and/or the fertilizer composition can include contacting the soil in the immediate vicinity of the planted seed with an effective amount of the composition. In some embodiments, the microalgae based composition and/or the fertilizer composition can be supplied to the soil by injection into a low volume irrigation system, such as but not limited to a drip irrigation system supplying water beneath the soil through perforated conduits or at the soil level by fluid conduits hanging above the ground or protruding from the ground. In some embodiments, the microalgae based composition and/or the fertilizer composition can be supplied to the soil by a soil drench method wherein the composition is poured on the soil.

The microalgae based composition and/or the fertilizer composition can be diluted to a lower concentration for an effective amount in a soil application by mixing a volume of the composition in a volume of water. The percent solids of microalgae sourced components resulting in the diluted composition can be calculated by the multiplying the original concentration in the composition by the ratio of the volume of the composition to the volume of water. Alternatively, the grams of microalgae sourced components in the diluted composition can be calculated by the multiplying the original grams of microalgae sourced components per 100 mL by the ratio of the volume of the composition to the volume of water.

The rate of application of the microalgae based composition and/or the fertilizer composition at the desired concentration can be expressed as a volume per area. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 50-150 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 75-125 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 50-75 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 75-100 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 100-125 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 125-150 gallons/acre.

In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 10-50 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 10- 20 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 20-30 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 30- 40 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 40-50 gallons/acre.

In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 0.01-10 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 0.01-0.1 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 0.1-1.0 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 1-2 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 2-3 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 3-4 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 4-5 gallons/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 5-10 gallons/acre.

In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 2-20 liters/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 3.7- 15 liters/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 2-5 liters/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 5-10 liters/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 10-15 liters/acre. In some embodiments, the rate of application of the microalgae based composition and/or the fertilizer composition in a soil application can include a rate in the range of 15-20 liters/acre.

Plants

Many plants can benefit from the application of compositions that provide a bio stimulatory effect. Non-limiting examples of plant families that can benefit from such compositions include plants from the following: Solanaceae, Fabaceae (Leguminosae), Poaceae, Roasaceae, Vitaceae, Brassicaeae (Cruciferae), Caricaceae, Malvaceae, Sapindaceae, Anacardiaceae, Rutaceae, Moraceae, Convolvulaceae, Lamiaceae, Verbenaceae, Pedaliaceae, Asteraceae (Compositae), Apiaceae (Umbelliferae), Araliaceae, Oleaceae, Ericaceae, Actinidaceae, Cactaceae, Chenopodiaceae, Polygonaceae, Theaceae, Lecythidaceae, Rubiaceae, Papveraceae, Illiciaceae Grossulariaceae, Myrtaceae, Juglandaceae, Bertulaceae, Cucurbitaceae, Asparagaceae (Liliaceae), Alliaceae (Liliceae), Bromeliaceae, Zingieraceae, Muscaceae, Areaceae, Dioscoreaceae, Myristicaceae, Annonaceae, Euphorbiaceae, Lauraceae, Piperaceae, Proteaceae, and Cannabaceae.

The Solanaceae plant family includes a large number of agricultural crops, medicinal plants, spices, and ornamentals in its over 2,500 species. Taxonomically classified in the Plantae kingdom, Tracheobionta (subkingdom), Spermatophyta (superdivision), Magnoliophyta (division), Manoliopsida (class), Asteridae (subclass), and Solanales (order), the Solanaceae family includes, but is not limited to, potatoes, tomatoes, eggplants, various peppers, tobacco, and petunias. Plants in the Solanaceae can be found on all the continents, excluding Antarctica, and thus have a widespread importance in agriculture across the globe.

The Rosaceae plant family includes flowering plants, herbs, shrubs, and trees. Taxonomically classified in the Plantae kingdom, Tracheobionta (subkingdom), Spermatophyta (superdivision), Magnoliophyta (division), Magnoliopsida (class), Rosidae (subclass), and Rosales (order), the Rosaceae family includes, but is not limited to, almond, apple, apricot, blackberry, cherry, nectarine, peach, plum, raspberry, strawberry, and quince.

The Fabaceae plant family (also known as the Leguminosae) comprises the third largest plant family with over 18,000 species, including a number of important agricultural and food plants. Taxonomically classified in the Plantae kingdom, Tracheobionta (subkingdom), Spermatophyta (superdivision), Magnoliophyta (division), Manoliopsida (class), Rosidae (subclass), and Fabales (order), the Fabaceae family includes, but is not limited to, soybeans, beans, green beans, peas, chickpeas, alfalfa, peanuts, sweet peas, carob, and liquorice. Plants in the Fabaceae family can range in size and type, including but not limited to, trees, small annual herbs, shrubs, and vines, and typically develop legumes. Plants in the Fabaceae family can be found on all the continents, excluding Antarctica, and thus have a widespread importance in agriculture across the globe. Besides food, plants in the Fabaceae family can be used to produce natural gums, dyes, and ornamentals.

The Poaceae plant family supplies food, building materials, and feedstock for fuel processing. Taxonomically classified in the Plantae kingdom, Tracheobionta (subkingdom), Spermatophyta (superdivision), Magnoliophyta (division), Liliopsida (class), Commelinidae (subclass), and Cyperales (order), the Poaceae family includes, but is not limited to, flowering plants, grasses, and cereal crops such as barely, com, lemongrass, millet, oat, rye, rice, wheat, sugarcane, and sorghum. Types of turf grass found in Arizona include, but are not limited to, hybrid Bermuda grasses (e.g., 328 tifgm, 419 tifway, tif sport).

The Vitaceae plant family includes flowering plants and vines. Taxonomically classified in the Plantae kingdom, Tracheobionta (subkingdom), Spermatophyta (superdivision), Magnoliophyta (division), Magnoliopsida (class), Rosidae (subclass), and Rhammales (order), the Vitaceae family includes, but is not limited to, grapes.

Compositions, mixtures, and methods according to the invention can be used for growing a broad variety of crops, such as potatoes, sugar beets, wheat, barley, rye, oat, sorghum, rice, maize, cotton, rapeseed, oilseed rape, canola, soybeans, peas, field beans, sunflowers, sugar cane; cucumbers, tomatoes, onions, leeks, lettuce, squashes; com, wheat, soy, cereals, and row crops.

The present invention is further illustrated by the following examples that should not be construed as limiting. The contents of all references, patents, and published patent applications cited throughout this application, as well as the Figures, are incorporated herein by reference in their entirety for all purposes.

EXAMPLES

Example 1. PHYCOTERRA^® ORGANIC (Whole Cell Chlorella Microalgae) in Combination with ENTERRA™ Fertilizer (Dried Trass' or Manure of Black Soldier Fly Larvae) Produces a Synergistic Effect

The advanced plant growth enhancement activity of the combinations of microalgae and fertilizer is evident from the example below. While each individual agent exhibits efficacy in enhancing plant growth, the combination has an activity which exceeds a simple addition of activities.

PHYCOTERRA^® ORGANIC (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce, tomato, and cauliflower plants at a rate of 2.5% (v/v) or 5% (v/v). ENTERRA™ fertilizer (dried Trass' or manure of black soldier fly larvae) was mixed with growing substrate prior to seeding at 7.42 g/L. Untreated control plants were compared to plants treated with PHYCOTERRA^® ORGANIC (whole cell Chlorella microalgae) alone, ENTERRA™ fertilizer (dried Trass' or manure of black soldier fly larvae) alone, and a combination of the two treatments. The average shoot weights and/or root weights were determined for the treated and untreated plants. Measurements were made 21 days after planting for lettuce and tomato plants and 16 days after planting for cauliflower plants.

A synergistic effect is present when the plant growth enhancement activity of the algal composition combination exceeds the total of the activities of the algal compositions when applied individually. The expected activity for a given combination of two plant growth enhancement agents can be calculated as follows (cf. Colby, S. R., “Calculating Synergistic and Antagonistic Responses of Herbicide Combinations,” Weeds, 1967, 15, 20-22):

If

• X is the plant growth enhancement activity when agent A is applied at an application rate of m gallons/acre (or liters/hectare),

• Y is the plant growth enhancement activity when agent B is applied at an application rate of n gallons/acre (or liters/hectare), • E is the plant growth enhancement activity when the active compounds A and B are applied at application rates of m and n gallons/acre (or liters/hectare), respectively,

• Then

The degree of plant growth enhancement activity compared to untreated control, expressed in %, is denoted. 0% means plant growth which corresponds to that of the untreated control while an activity of 100% means that the plant growth is twice that observed with the untreated control.

If the actual plant growth enhancement activity exceeds the calculated value, then the activity of the combination is super additive, i.e., a synergistic effect exists. In this case, the efficacy which was actually observed must be greater than the value for the expected efficacy (E) calculated from the above-mentioned formula. The results shown in Tables 1-4 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® ORGANIC (whole cell Chlorella microalgae) and ENTERRA™ fertilizer (dried Trass' or manure of black soldier fly larvae).

Table 1. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Table 2. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Table 3. Tomato shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

Table 4. Cauliflower shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 2. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with Organic Humic Acid Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine letuce plants at a rate of 1% (v/v), and organic humic acid was also applied at a rate of 1% (v/v). These greenhouse rates simulate application at 1 gallon/acre in the field. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, organic humic acid alone, and a combination of the two treatments. The average shoot weights were determined for the treated and untreated plants. Measurements were made 6 weeks after planting.

The results shown in Table 5 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and organic humic acid.

Table 5. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 3. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with LIQUID CHISEL^® (Soluble Potash) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 1 gallon/acre, and LIQUID CHISEL^® (soluble potash) was applied at a rate equivalent to 1 quart/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, LIQUID CHISEL^® (soluble potash) alone, and a combination of the two treatments. The average shoot weights were determined for the treated and untreated plants several weeks after planting.

The results shown in Table 6 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and LIQUID CHISEL^® (soluble potash).

Table 6. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 4. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with AN20 (Ammonium Nitrate 20-0-0) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre, and AN20 (ammonium nitrate 20-0-0) was applied at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, AN20 (ammonium nitrate 20-0-0) alone, and a combination of the two treatments. The average shoot weights and root weights were determined for the treated and untreated plants several weeks after planting.

The results shown in Tables 7 and 8 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and AN20 (ammonium nitrate 20-0-0).

Table 7. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Table 8. Romaine letuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 5. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with APP (Ammonium Polyphosphate 10-34-0) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre, and APP (ammonium polyphosphate 10-34-0) was applied at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, APP (ammonium polyphosphate 10-34-0) alone, and a combination of the two treatments. The average shoot weights and root weights were determined for the treated and untreated plants several weeks after planting. The results shown in Tables 9 and 10 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and APP (ammonium polyphosphate 10-34-0).

Table 9. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Table 10. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 6. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with CAN17 (Calcium Ammonium Nitrate 17-0-0) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre, and CAN 17 (calcium ammonium nitrate 17-0-0) was applied at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, CAN17 (calcium ammonium nitrate 17-0-0) alone, and a combination of the two treatments. The average shoot weights and root weights were determined for the treated and untreated plants several weeks after planting.

The results shown in Tables 11 and 12 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and CAN17 (calcium ammonium nitrate 17-0-0).

Table 11. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Table 12. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

Example 7. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with Potassium Nitrate (KNO3) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre, and potassium nitrate (KNO3) was applied at a rate equivalent to 0.5 gallon/acre or 1 gallon/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, potassium nitrate (KNO3) alone, and a combination of the two treatments. The average shoot weights and root weights were determined for the treated and untreated plants several weeks after planting. The results shown in Tables 13 and 14 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and potassium nitrate (KNO3). Table 13. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

Table 14. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

Example 8. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with UN32 (Urea Ammonium Nitrate Solution) Produces a Synergistic Effect PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to

Romaine lettuce plants in a greenhouse at a rate equivalent to 0.5 gallon/acre, and UN32 (urea ammonium nitrate solution) was applied at a rate equivalent to 0.5 gallon/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, UN32 (urea ammonium nitrate solution) alone, and a combination of the two treatments. The average shoot weights and root weights were determined for the treated and untreated plants several weeks after planting.

The results shown in Tables 15 and 16 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and UN32 (urea ammonium nitrate solution).

Table 15. Romaine lettuce shoot biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

Table 16. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula Example 9. PHYCOTERRA^® (Whole Cell Chlorella Microalgae) in Combination with NUTRASYST^® (Fulvic Acid) Produces a Synergistic Effect

PHYCOTERRA^® (whole cell Chlorella microalgae) was applied as a soil drench to Romaine lettuce plants in a greenhouse at a rate equivalent to 1 gallon/acre, and NUTRASYST^® (fulvic acid) was applied at a rate equivalent to 12.8 ounces/acre. Untreated control plants were compared to plants treated with PHYCOTERRA^® (whole cell Chlorella microalgae) alone, NUTRASYST^® (fulvic acid) alone, and a combination of the two treatments. The average root weights were determined for the treated and untreated plants several weeks after planting.

The results shown in Table 17 clearly indicate a synergistic effect resulting from the combination treatment of plants with PHYCOTERRA^® (whole cell Chlorella microalgae) and NUTRASYST^® (fulvic acid).

Table 17. Romaine lettuce root biomasses.

*Found = activity observed

**Calc. = activity calculated using Colby’s formula

All headings are for the convenience of the reader and should not be used to limit the meaning of the text that follows the heading, unless so specified.

Unless defined otherwise, all technical and scientific terms herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials, similar or equivalent to those described herein, can be used in the practice or testing of the present invention, the preferred methods and materials are described herein. All publications, patents, and patent publications cited are incorporated by reference herein in their entirety for all purposes.

The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention.

While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features hereinbefore set forth and as follows in the scope of the appended claims.

Claims

CLAIMS What is claimed is:

1. A mixture comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; wherein a combination of the first composition and the second composition exhibits synergy.

2. The mixture of Claim 1, wherein the culture of microalgae comprises Aurantiochytrium, Botryococcus, Chlorella, Chlamydomonas, Desmodesmus, Dunaliella, Scenedesmus, Pavolv, Phaeodactylum, Nannochloropsis, Spirulina, Galdieria, Haematococcus, Isochrysis, Porphyridium, Schizochytrium, Thraustochytrium, Tetraselmis, or combinations thereof.

3. The mixture of Claim 2, wherein the culture of microalgae comprises Chlorella.

4. The mixture of Claim 3, wherein the Chlorella are whole cells, lysed cells, dried cells, cells that have been subjected to an extraction process, or a combination thereof.

5. The mixture of any one of Claims 1 to 4, wherein the fertilizer is an organic fertilizer, inorganic fertilizer, urea-containing fertilizer or combination thereof.

6. The mixture of Claim 5, wherein the fertilizer is an organic fertilizer selected from the group consisting of humic acid, kelp, seaweed extract, fulvic acid, fish emulsion/fish meal, soy hydrolysate, protein hydrolysate, AMINO PRIME™ (2.5-0-1 derived from sugarcane protein hydrolysate), com steep liquor, compost, manure, biochar, sugar beet or sugarcane vinasse, sewage sludge, blood meal, bone meal, worm castings, peat moss, leaf compost, rice hull, coffee chaff, buckwheat hull, chicken manure, tree bark with or without composting, mushroom composts, black soldier fly frass, and combinations thereof.

7. The mixture of Claim 5, wherein the fertilizer is an inorganic fertilizer selected from the group consisting of CAN17, CN9, CaTS^® (6% calcium and 10% sulfur calcium thiosulfate fertilizer solution), AN20, AMS, ATS, UN32, 10-34-0, 11-37-0, 6-26-5, 5-28-0-10 Mg, 44-0-0, 14-24-0-10S, 17-1-0-20S, KTS^® blend (0-0-25+17S derived from potassium thiosulfate), K-ROW 23^® (0-0-23-8S derived from postassium sulfite), liquid ammonia, ammonium nitrate, calcium nitrate, potassium nitrate, calcium ammonium nitrate, phosphoric acid, ammonium phosphate, monoammonium phosphate, diammonium phosphate, ammonium polyphosphate, single super phosphate, triple super phosphate, ground rock phosphate, a phosphite salt, an ammonium iron salt, potassium chloride, muriate of potash, sulfate of potash, sulfate of potash magnesia, soluble potash, ammonium sulfate, ammonium thiosulfate, potassium sulfate, ammonium sulfate-nitrate, calcium cyanamide, potassium hydroxide, potassium acetate, ammonium carboxylate, cyclohexanediaminepentaacetic acid (CDTA), citric acid (CIT), diethylenetriaminepentaacetic acid (DTP A), ethylenediaminediaminedi-o- hydroxyphenylacetic acid (EDDHA), ethylenediamintetraacetic acid (EDTA), ethylene glycol bis(2-aminoethyl ether) tetraacetic acid (EGTA), hydroxyethylenediaminetriacetic acid (HEDTA), nitrilo-triacetic acid (NT A), oxalic acid (OX), pyrophosphoric acid (PPA), triphosphoric acid (TP A), vermiculite, perlite, Chilean nitrate, limestone, and combinations thereof.

8. The mixture of Claim 5, wherein the fertilizer is a urea-containing fertilizer selected from the group consisting of urea, urea formaldehyde, urea-acetaldehyde, ureaglyoxal condensate, urea sulfur, urea sulfuric acid (urea sulfate), urea ammonium sulfate, isobutylidene diurea, crotonylidene diurea, ethylene urea, methylene diurea, urea ammonium nitrate, and combinations thereof.

9. The mixture of any one of Claims 1 to 8, wherein the fertilizer is: a solid coated or uncoated granule; in a liquid or semi-liquid form; formulated as a sprayable fertilizer; formulated for application via fertigation; and/or formulated as a slow release fertilizer.

10. The mixture of any one of Claims 1 to 9, further comprising a nitrification inhibitor, a urease inhibitor, a denitrification inhibitor, or a combination thereof.

11. A method of treating a plant, a plant part, or the locus surrounding the plant to enhance plant growth and/or yield, the method comprising applying an effective amount of the mixture of any one of Claims 1 to 10 to the plant, plant part, or the locus surrounding the plant.

12. A method of plant enhancement comprising applying to a plant, seedling, plant propagation material, or the locus surrounding the plant material an effective amount of the mixture of any one of Claims 1 to 10, wherein the plant characteristic is selected from the group consisting of seed germination rate, seed germination time, seedling emergence, seedling emergence time, seedling size, plant fresh weight, plant dry weight, utilization, fruit production, leaf production, leaf formation, leaf size, leaf area index, plant height, thatch height, plant health, plant resistance to salt stress, plant resistance to heat stress, plant resistance to heavy metal stress, plant resistance to drought, maturation time, yield, root length, root mass, color, insect damage, blossom end rot, softness, plant quality, fruit quality, flowering, sun bum, and any combination thereof.

13. The method of Claim 11 or 12, wherein the components in the mixture are applied simultaneously or subsequently.

14. The method of any one of Claims 11 to 13, wherein the mixture is applied to soil in the immediate vicinity of the plant, seedling, or plant propagation material and/or as a foliar spray.

15. A kit-of-parts comprising: a) a first composition comprising a culture of microalgae; and b) a second composition comprising a fertilizer; in a spatially separated arrangement, wherein a combination of the first composition and the second composition exhibits synergy.

16. The kit-of-parts of Claim 15, wherein the culture of microalgae comprises

Aurantiochytrium, Botryococcus, Chlorella, Chlamydomonas, Desmodesmus, Dunaliella, Scenedesmus, Pavolv, Phaeodactylum, Nannochloropsis, Spirulina, Galdieria, Haematococcus, Isochrysis, Porphyridium, Schizochytrium, Thraustochytrium, Tetraselmis, or combinations thereof.

17. The kit-of-parts of Claim 16, wherein the culture of microalgae comprises Chlorella as whole cells, lysed cells, dried cells, cells that have been subjected to an extraction process, or a combination thereof.

18. The kit-of-parts of any one of Claims 15 to 17, wherein the fertilizer is an organic fertilizer selected from the group consisting of humic acid, kelp, seaweed extract, fulvic acid, fish emulsion/fish meal, soy hydrolysate, protein hydrolysate, AMINO PRIME™ (2.5-0- 1 derived from sugarcane protein hydrolysate), com steep liquor, compost, manure, biochar, sugar beet or sugarcane vinasse, sewage sludge, blood meal, bone meal, worm castings, peat moss, leaf compost, rice hull, coffee chaff, buckwheat hull, chicken manure, tree bark with or without composting, mushroom composts, black soldier fly frass, and combinations thereof.

19. The kit-of-parts of any one of Claims 15 to 17, wherein the fertilizer is an inorganic fertilizer selected from the group consisting of CAN17, CN9, CaTS^® (6% calcium and 10% sulfur calcium thiosulfate fertilizer solution), AN20, AMS, ATS, UN32, 10-34-0, 11- 37-0, 6-26-5, 5-28-0-10 Mg, 44-0-0, 14-24-0-10S, 17-1-0-20S, KTS^® blend (0-0-25+17S derived from potassium thiosulfate), K-ROW 23^® (0-0-23-8S derived from postassium sulfite), liquid ammonia, ammonium nitrate, calcium nitrate, potassium nitrate, calcium ammonium nitrate, phosphoric acid, ammonium phosphate, monoammonium phosphate, diammonium phosphate, ammonium polyphosphate, single super phosphate, triple super phosphate, ground rock phosphate, a phosphite salt, an ammonium iron salt, potassium chloride, muriate of potash, sulfate of potash, sulfate of potash magnesia, soluble potash, ammonium sulfate, ammonium thiosulfate, potassium sulfate, ammonium sulfate-nitrate, calcium cyanamide, potassium hydroxide, potassium acetate, ammonium carboxylate, cyclohexanediaminepentaacetic acid (CDTA), citric acid (CIT), diethylenetriaminepentaacetic acid (DTP A), ethylenediaminediaminedi-o-hydroxyphenylacetic acid (EDDHA), ethylenediamintetraacetic acid (EDTA), ethylene glycol bis(2-aminoethyl ether) tetraacetic acid (EGTA), hydroxyethylenediaminetriacetic acid (HEDTA), nitrilo-triacetic acid (NT A), oxalic acid (OX), pyrophosphoric acid (PPA), triphosphoric acid (TP A), vermiculite, perlite, Chilean nitrate, limestone, and combinations thereof.

20. The kit-of-parts of any one of Claims 15 to 17, wherein the fertilizer is a urea- containing fertilizer selected from the group consisting of urea, urea formaldehyde, urea- acetaldehyde, ureaglyoxal condensate, urea sulfur, urea sulfuric acid (urea sulfate), urea ammonium sulfate, isobutylidene diurea, crotonylidene diurea, ethylene urea, methylene diurea, urea ammonium nitrate, and combinations thereof.