CONTENT, YIELD, AND HEALTH OF CROPS

RELATED APPLICATION

This application claims the benefit under 35 U.S.C. § 119(e) of U.S. provisional application number 61/979,053, filed April 14, 2014, which is incorporated by reference herein in its entirety.

SUMMARY OF THE INVENTION

Healthier and higher- yielding crops are constantly sought, as the global economy depends on agricultural products. Such dependence extends beyond food and feed to areas such as bioenergy, chemicals, fibers, among others. Because costs of production usually scale with area, but products sell per unit of mass, economic viability in the production of crops or crop-derived products depends on the productivity per unit area.

The present invention comprises the application of extracts or parts of a plant of the genus Stevia on plants of either the genus Sorghum or the genus Saccharum. The methods increase the sugar content and the sugar production of the treated plants, and increase the grain yield and quality, overall yield, productivity, root development, health, and general sanitary conditions of the treated plants. The present invention further comprises the utilization of sugars in subsequent processes.

The present invention deals in particular with the production and agriculture of sorghum and sugarcane, two staple crops of wide use in the production of sugar, sweeteners (e.g. sorghum syrup, molasses, etc.), grain, and fiber, in addition to green matter that can be used as feed or fuel. Derived products from the sugar or fiber are also of interest. For example, the sugars in sweet sorghum and sugarcane can be released and fermented into ethanol, butanol, propanediol, lactic acid, butanediol, succinic acid, farnesene, among other compounds. The leftover biomass (bagasse) or the green matter itself, depending on the process and type of crop, can be used as a feedstock for power production, as fodder, and as feedstock for a variety of biochemical or thermochemical processes. The present invention deals with the use of extracts of plants to enhance the sugar content, yield, productivity, health, and sanitary conditions of sorghum and sugarcane crops. In particular, the invention describes the use of plants of the genus Stevia or artificial mixtures containing some or all of the compounds present in that plant to serve the ends described by this invention.

The extract of the Stevia plant has been reported as a crop additive in JP 2002205907 by Sato Naohiko, which describes the explicit aim of preventing and treating the disease caused by the microorganism Helicobasidium mompa. The use of Stevia extracts for increasing or improving the sugar content, sugar production, grain yield and quality, overall yield, productivity, root development, health, and general sanitary conditions of sorghum or sugarcane (genus Sorghum and Saccharum, respectively) have not been reported to our knowledge.

Genetic methods to increase sugar content have also been reported, e.g., by publication US 2011/0179525. Genetic methods, including breeding for particular traits, are significantly different from the present invention and suffer from disadvantages as outlined in the next section.

According to one aspect of the invention, methods for producing a plant with increased sugar content or that produces an increased amount of sugar per unit area as compared to a non-treated plant comprising treating the plant with one or more extracts of a plant of the genus Stevia. In some embodiments, the treated plant belongs to the genus Sorghum (colloquial sorghum) or the genus Saccharum (colloquial sugarcane). In some embodiments, the extract is applied directly to the sorghum or sugarcane plant structures (leaves, shoots, roots, etc.). In some embodiments, the extract is applied to the soil where the sorghum or sugarcane plants to be treated will grow.

In some embodiments, the extract used is naturally-derived. In other embodiments, the extract is artificially synthesized. In some embodiments, the extract is a mixture of any, part, or all of the components present in the naturally-derived extract from the plant of the genus Stevia. In some embodiments, the extract or mixture contains any, part, or all of the following compounds: Steviol Glucoside, Dulcoside, Dulcoside A, Stevioside, Rebaudioside, Rebaudioside A, Rebaudioside B, Rebaudioside C, Rebaudioside D, Rebaudioside E, Rebaudioside F, Rubusoside, Steviolbioside, or Steviol.

In some embodiments, the extract is derived from the leaves of a plant of the genus Stevia, or from the shoots of a plant of the genus Stevia or from the roots of a plant of the genus Stevia. In some embodiments, the extract is an aqueous extract of the plant of the genus Stevia. In other embodiments, the extract is a partially aqueous or non-aqueous extract of the plant of the genus Stevia, optionally an ethanol, methanol, butanol, propanol, isopropanol, or mixed solvent extract. In other embodiments, the extract is a heat extract of the plant of the genus Stevia.

In some embodiments, the extract is clarified before application. In some

embodiments, the extract is diluted before application, such as diluted to 0.5-50 wt or vol .

In another aspect of the invention, methods for producing a plant with increased sugar content or that produces an increased amount of sugar per unit area as compared to a non- treated plant are provided. The methods include incorporating parts of a plant of the genus Stevia in the soil where the treated plant will grow or placing parts of a plant of the genus Stevia on the soil where the treated plant will grow.

In some embodiments, the foregoing methods increase the productivity of the treated plant. In some embodiments, the foregoing methods increase the grain yield of the treated plant. In some embodiments, the foregoing methods increase the grain quality, nutritional value, or aspect of the treated plant. In some embodiments, the foregoing methods increase the overall yield of the treated plant. In some embodiments, the foregoing methods increase productivity of the treated plant. In some embodiments, the foregoing methods develop, fortify, and/or improve the root system of the treated plant. In some embodiments, the foregoing methods improve the general health of the treated plant. In some embodiments, the foregoing methods improve the sanitary conditions of the treated plant. In some

embodiments, the foregoing methods alter the growth cycle of the treated plant.

In another aspect of the invention, methods for obtaining the sugars from a sorghum or sugarcane plant are provided. The methods include treating the plant according to according to any of the foregoing methods, and obtaining sugar from the treated plant. In some embodiments, the sugars are obtained via milling. In some embodiments, the sugars are obtained via diffusion. In some embodiments, the sugars are obtained via pressing. In some embodiments, the sugars are obtained via extrusion.

In some embodiments of these methods, the obtained sugars are further processed. In some embodiments, the sugars are processed via fermentation. In some embodiments, the sugars are used to produce ethanol. In some embodiments, the sugars are used to produce butanol. In some embodiments, the sugars are used to produce propanediol. In some embodiments, the sugars are used to produce lactic acid. In some embodiments, the sugars are used to produce succinic acid. In some embodiments, the sugars are used to produce propanediol. In some embodiments, the sugars are used to produce butanediol. In some embodiments, the sugars are used to produce an isoprenoid compound. In some

embodiments, the sugars are used to produce crystalline sugar (table sugar). In some embodiments, the sugars are treated with a catalyst.

According to another aspect of the invention, methods for further processing the sorghum or sugarcane plants are provided. The methods include treating the plant according to any of the foregoing methods, and producing energy, chemicals, fuels, fibers, feed, or agricultural products from the treated plant.

DESCRIPTION OF THE INVENTION

Purposes and advantages of the invention

Application of Stevia-derived compounds on sorghum or sugarcane has several purposes as described herein, including improving the sugar content, sugar production, grain yield and quality, overall yield, productivity, root development, health, and general sanitary conditions. All of these have measurable and important advantages for the production of sorghum and sugarcane. In particular, the production of increased amounts of sugar and/or biomass using the same land area and comparable inputs reduces the cost of production and improves the crop economic value. The economic benefit trickles down to the products obtained from the plants, including sugars, grain, fiber, and the variety of products that can be produced from them (e.g. biofuels, chemicals, etc.). Furthermore, the methods here described have several health-improving effects on the crops. Additional advantages thus include the possibility to forgo or reduce the use of other additives that have overlapping effects, such as herbicides, fungicides, bactericides, nematicides, crop-enhancement agents (e.g.

thiamethoxam), among others.

Genetic methods have also been used to increase sugar content and yield of crops. These methods, however, are generally hard to implement and genetic manipulations have several consequences in plant physiology, some of which can be detrimental to commercial or large-scale applications. If the genetic methods involve a gene transfer, the crop becomes a "genetically-modified organism", which can limit the crop's usefulness and marketability and increases costs through compliance and registration burdens. The fact that the extracts here described are naturally-derived ensures that the production of the crops can remain under the denomination "organic".

Description and Operation

Many alternatives exist for the implementation of the method. In one embodiment, extracts of a plant of the genus Stevia are prepared by water or solvent extraction of the shoots, leaves, or roots of the sorghum or sugarcane plant. Alternatively, in other

embodiments a mixture of any, part, or all of the components present in the extract of the Stevia plant can be artificially prepared, such as by chemical synthesis, and used. This may include separation of desired compounds by chromatography, adsorption, or similar processes into fractions that can then be used to produce the desired mixture. The extract or mixture can be diluted if desired.

Subsequently, the extract or mixture is applied to the soil or over the plant to be treated, in one or several occasions during the crop's cycle. In preferred embodiments, the extract is applied between 1 and 6 times over the growth of the plant, and is applied directly on the plant leaves. It can also be applied on the soil where the sorghum or sugarcane crops will grow or during growth. In an alternate embodiment, the Stevia plant or plant parts can be mixed onto the soil or placed on the soil where the sorghum or sugarcane will grow or during growth. The resulting sorghum or sugarcane crop has improved qualities of sugar content, sugar production, grain yield and quality, overall yield, productivity, root development, health, and general sanitary conditions.

EXAMPLES Example 1

Materials and Methods

Solvent solution: the solvent solution is composed of 25% ethanol in water. Shoots of the Stevia rebaudiana plant were extracted with the solvent solution by contacting a 1.1 wt-% mixture of shoots and solvent for 24 hours. This solution was stored and later used in the sorghum gardens. Five sweet sorghum varieties were grown in three different plots of land (BI, BII and Bill) following protocols available to those ordinarily skilled in the art. The varieties used were as follows: Theis, Topper, M81, Dulce del Plata, and S. King. The extract was applied over the plants during the vegetative growth phase at a level of around 10 liters per hectare.

Results

The results of the assays are shown in Tables 1 and 2. The Brix content was used as a measure of the sugar content and was measured by a refractometer using a protocol available to persons with ordinary skill in the art. Measurements were taken after the plants reached maturity, which is 90-120 days after sowing depending on the variety. The comparison of the treated and non-treated controls shows an increase in sugar content and productivity per hectare.

Example 2: Stevia Extract Preparation and Use Thereof

Another process for preparing a Stevia extract was reported in "^,Εη que consiste el metodo de cultivo usando Stevia?" By Yasusada Que and Shinji Ueno, from the Office of Asistencia Oficial para el Desarrollo de Japon (Assistance for Japanese Development Office), Asuncion, Paraguay, March 2003. The process was performed as follows:

Experimental Protocol

1- The stems and leaves of the Stevia plant were cut at the time of flower induction at about 5 cm above the soil level. The point of cutting must be such that the plant can regrow and such that it affords the required amount of biomass for the procedure.

2- The leaves and stems were put to dry after cutting. Drying took place in rooms with low humidity and partial shading, such that the loss of water in the plant was relatively constant.

3- After having achieved a minimum of 70 % dry matter, the dry biomass was passed through a mill to achieve minced- sized particles smaller than 0.5 mm.

4- After grinding, 1 kg of dry matter was boiled in 12-13L of water for half an hour.

5- The liquid slurry was passed through a sieve, so that only the liquid passed. The

remaining liquid was left to evaporate to 10L of solution; its composition is detailed below (Compositional study performed by the research laboratories of JBB Stevia, Japan):

The solid that remains after filtration can be used in composting systems or directly on the base of plants to promote the growth of beneficial microorganisms.

The liquid extract can be used for two treatments of ~ 5 liters per hectare, 10 to 15 days apart from each other, in a volume of 100 liter water/ha on the foliage.

The application is done with the use of an adjuvant, which is typically an anionic surfactant. In a preferred embodiment, the surfactant' s active ingredients are Nonyl- phenoxy-polyethoxy-ethanol (300 g/L) and sodium dodecylbenzene sulfonate (150 g/L). This surfactant mixture is used at a rate of 1 mL per liter of solution. If long-term storage of the solution is required (up to two years), 20 mL of a 72% ethanol solution is added for preservation.

In addition to promoting sugar accumulation, the extract can promote the growth of beneficial microorganisms like fungi, whose action will be catalyzed in the presence of the extract.

The extract has also been used in combination with a solution of foliar fertilizer, which is a common raw material in agriculture. It can also be used in combination with garlic extract to promote the general health of the crop.

The extract can be applied together with or subsequently to foliar fertilizers in the same way in which foliar fertilizers are applied.