WO2011012561A1

WO2011012561A1 - Treatment of hop plants with bioregulators

Info

Publication number: WO2011012561A1
Application number: PCT/EP2010/060765
Authority: WO
Inventors: Adam R. Kavalier; Paul D. Matthews; Wilhelm Rademacher; Edward Kennelly
Original assignee: Basf Se
Priority date: 2009-07-31
Filing date: 2010-07-26
Publication date: 2011-02-03
Also published as: AR077351A1

Abstract

The present invention relates to a method for treating hop plants comprising the application of an aqueous formulation containing acylcyclohexanedione of the formula I as defined below, or a salt of said acylcyclohexanedione of the formula I, onto the hop plants at their growth stage BBCH from 61 to 79. The invention further relates to a use of said acylcyclohexanedione for increasing the yield of cone mass of the hop plants. It also relates to a use of said acylcyclohexanedione for increasing the concentration of prenylated polyketides in the cones of the hop plants.

Description

Treatment of hop plants with bioregulators

The present invention relates to a method for treating hop plants comprising the application of an aqueous formulation containing acylcyclohexanedione of the formula I as defined below, or a salt of said acylcyclohexanedione of the formula I, onto the hop plants at their growth stage BBCH from 61 to 79. The invention further relates to a use of said acylcyclohexanedione for increasing the yield of cone mass of the hop plants. It also relates to a use of said acylcyclohexanedione for increasing the concentration of prenylated polyketides in the cones of the hop plants. Combinations of preferred em- bodiments with other preferred embodiments are within the scope of the present invention.

Hops (Humulus lupulus) is a medicinal plant that is also used in the brewing of beer for flavor, stability, and process. Valued secondary metabolites produced in floral organs and especially in trichomatous glands of cones are the focus of breeding and metabolic engineering.

Bioregulators of the class of acyclcyclohexanediones and some effects on plants are known: EP 0 598 404 discloses a plant growth regulator composition comprising a cyclohexane derivative and a nitrogen-containing water-soluble inorganic substance or urea. EP 0 344 533 discloses a plant growth regulator composition comprising a 3,5- dioxo-4-propionylcyclohexane carboxylic acid. EP 0 123 001 discloses a plant growth regulating composition comprising a cyclohexane compound. WO 2000/78143 discloses a method for increasing and qualitatively modifying the content of flavonoids and phenolic constituents in various plants, such as hops, by treating the plants with an acyclcyclohexanedione, such as prohexadion-calcium.

It is an object of the present invention to improve the yield and/or the quality of hop cones. Especially the concentration of bitter acids in the hop cones should be in- creased.

The object was achieved by a method for treating hop plants comprising the application of an aqueous formulation containing acylcyclohexanedione of the formula I

where R is hydrogen or Ci-Cβ-alkyl, and R' is Ci-Cβ-alkyl or Cs-Cβ-cycloalkyl, or a salt of said acylcyclohexanedione of the formula I, onto the hop plants at their growth stage BBCH from 61 to 79. Hop plants of all varieties might be treated with the method according to the present invention, for example Admiral, Ahtanum, Amarillo, Apollo, Brewer's Gold, Bullion, Cascade, Centennial, Challenger, Chinook, Cluster, Columbus, Crystal, Eroica, First Gold, Fuggles, Galena, Glacier, Goldings, Greenburg, Hallertau / Hallertauer Mittelfruh, Herald, Hersbrucker, Horizon, Liberty, Lublin, Magnum, Millennium, Mount Hood, Nelson Sauvin, Newport, Northdown, Northern Brewer, Nugget, Pacific Gem, Palisade, Perle, Pioneer, Polnischer Lublin, Pride of Ringwood, Progress, Saaz, Santiam, Saphir, Satus, Select, Simcoe, Spalt, Sterling, Strisselspalt, Styrian Goldings, Summit, Tardif de Bourgogne, Target, Tettnang, Tomahawk, Tradition, Ultra, Vanguard, Warrior, Willamette, Zeus. Preferably, the varieties Willamette and Zeus are treated, more preferably Willamette.

Suitable acylcyclohexandione are those of the formula I

where R is hydrogen or Ci-Cβ-alkyl, and R' is Ci-Cβ-alkyl or Cs-Cβ-cycloalkyl, or a salt of said acylcyclohexanedione of the formula I. R is preferably hydrogen. R' is preferably d-Cβ-alkyl, more preferably ethyl. The compounds of the formula I may be present both in the trione form (triketo form) I. a and in the tautomeric keto-enol forms l.b and l.c:

The salts of the acylcyclohexanedione compounds I may be the salts either of the monoanions or of the dianions of these compounds. The monoanions may be present

In the dianion, the carboxylate and the enolate groups are accordingly present alongside one another. Preferred cations in the salts of the acylcyclohexanedione of the formula I are the ions of the alkali metals, preferably of lithium, sodium and potassium, of the alkaline earth metals, preferably of calcium and magnesium, and of the transition metals, preferably of manganese, copper, zinc and iron, and also ammonium (NH₄ ⁺) and substituted ammonium, in which one to four hydrogen atoms are replaced by Ci-C4-alkyl, hydroxy-Ci- C4-alkyl, Ci-C4-alkoxy-Ci-C4-alkyl, hydroxy-Ci-C4-alkoxy-Ci-C4-alkyl, phenyl or benzyl, preferably ammonium, methylammonium, isopropylammonium, dimethylammonium, diisopropylammonium, trimethylammonium, tetramethylammonium, tetraethylammo- nium, tetrabutylammonium, 2-hydroxyethylammonium, 2-(2-hydroxyeth-1-oxy)eth-1- ylammonium, di(2-hydroxyeth-1-yl)ammonium, benzyltrimethylammonium, benzyl- triethylammonium, and additionally phosphonium ions, sulfonium ions, preferably tri(Ci- C4-alkyl)sulfonium such as trimethylsulfonium, and sulfoxonium ions, preferably tri(Ci- C4-alkyl)sulfoxonium. Preferred cations are also chlormequat [(2-chloroethyl)tri- methylammonium], mepiquat (N,N-dimethylpiperidinium) and N,N-dimethyl- morpholinium. Particularly preferred cations are the alkali metal cations, the alkaline earth metal cations and the ammonium cation (Nl-U⁺). The compound is especially the calcium salt.

The acylycyclohexandione is preferably prohexadione-calcium. Prohexadione-calcium (3,5-dioxo-4-propionylcyclohexanecarboxylic acid) is a well known plant growth regulator, which is commercially available for example as Regalis® or Apogee® from BASF SE.

Typically, the acylcyclohexandione is applied in form of an aqueous formulation containing said acylcyclohexanedione. Preferably, the aqueous formulation comprises at least 80 wt%, more preferably at least 95 wt% water. The aqueous formulation is usually prepared immediately prior to use (so called "tank mix").

Various types of oils, wetters, adjuvants, herbicides, bactericides, other fungicides and/or pesticides may be added to the tank mix. These agents can be admixed with the compositions according to the invention in a weight ratio of 1 :100 to 100:1 , preferably 1 :10 to 10:1. Adjuvants which can be used are usually anionic surfactants (e.g. dioctyl sulfosuccinate sodium, such as Leophen RA^®,) or non-ionic surfactants, in particular non-ionic surfactants. Examples are polyether modified polysiloxanes such as Break Thru® S 240; fatty alcohol alkoxylates such as Plurafac® LF 120 (BASF) and Luten- sol® ON 30 (BASF); ethyleneoxide/propyleneoxide (EO/PO) block polymers, e. g. PIu- ronic® RPE 2035, Regulaid® and Genapol B, alcohol ethoxylates such as Lutensol XP 80^®; polyvinylalcohols, such as Plurafac® LF 240 (BASF). Especially preferred non- ionic surcactants are fatty alcohol alkoxylates, EO/PO block polymers and polyether modified polysiloxanes. Most preferred are EO/PO block polymers. The aqueous formulation comprises preferably 0.01 to 20 wt%, more preferably 0.1 to 5 wt% and especially 0.5 to 2 wt% of a non-ionic surfactant.

The aqueous formulation may also comprise auxiliaries which are customary in agro- chemical compositions. Examples for suitable auxiliaries are solvents, solid carriers, dispersants or emulsifiers (such as further solubilizers, protective colloids, surfactants and adhesion agents), organic and inorganic thickeners, bactericides, anti-freezing agents or anti-foaming agents. Suitable solvents are water, organic solvents such as mineral oil fractions of medium to high boiling point, such as kerosene or diesel oil, furthermore coal tar oils and oils of vegetable or animal origin, aliphatic, cyclic and aromatic hydrocarbons, e. g. toluene, xylene, paraffin, tetrahydronaphthalene, alkylated naphthalenes or their derivatives, alcohols such as methanol, ethanol, propanol, butanol and cyclohexanol, glycols, ke- tones such as cyclohexanone and gamma-butyrolactone, fatty acid dimethylamides, fatty acids and fatty acid esters and strongly polar solvents, e. g. amines such as N- methylpyrrolidone.

Suitable surfactants (adjuvants, wetters, tackifiers, dispersants or emulsifiers) are alkali metal, alkaline earth metal and ammonium salts of aromatic sulfonic acids, such as ligninsoulfonic acid (Borresperse^® types, Borregard, Norway) phenolsulfonic acid, naphthalenesulfonic acid (Morwet^® types, Akzo Nobel, U.S.A.), dibutylnaphthalene- sulfonic acid (Nekal^® types, BASF, Germany), and fatty acids, alkylsulfonates, alkyl- arylsulfonates, alkyl sulfates, laurylether sulfates, fatty alcohol sulfates, and sulfated hexa-, hepta- and octadecanolates, sulfated fatty alcohol glycol ethers, furthermore condensates of naphthalene or of naphthalenesulfonic acid with phenol and formaldehyde, polyoxy-ethylene octylphenyl ether, ethoxylated isooctylphenol, octylphenol, nonylphenol, alkylphenyl polyglycol ethers, tributylphenyl polyglycol ether, tristearyl- phenyl polyglycol ether, alkylaryl polyether alcohols, alcohol and fatty alcohol/ethylene oxide condensates, ethoxylated castor oil, polyoxyethylene alkyl ethers, ethoxylated polyoxypropylene, lauryl alcohol polyglycol ether acetal, sorbitol esters, lignin-sulfite waste liquors and proteins, denatured proteins, polysaccharides (e. g. methylcellulose), hydrophobically modified starches, polyvinyl alcohols (Mowiol^® types, Clariant, Switzerland), polycarboxylates (Sokolan^® types, BASF, Germany), polyalkoxylates, polyvinyl- amines (Lupasol^® types, BASF, Germany), polyvinylpyrrolidone and the copolymers therof.

Examples for thickeners (i. e. compounds that impart a modified flowability to compositions, i. e. high viscosity under static conditions and low viscosity during agitation) are polysaccharides and organic and anorganic clays such as Xanthan gum (Kelzan^®, CP Kelco, U.S.A.), Rhodopol^® 23 (Rhodia, France), Veegum^® (RT. Vanderbilt, U.S.A.) or Attaclay^® (Engelhard Corp., NJ, USA). Bactericides may be added for preservation and stabilization of the composition. Examples for suitable bactericides are those based on dichlorophene and benzylalcohol hemi formal (Proxel^® from ICI or Acticide^® RS from Thor Chemie and Kathon^® MK from Rohm & Haas) and isothiazolinone derivatives such as alkylisothiazolinones and benzisothiazolinones (Acticide^® MBS from Thor Chemie). Examples for suitable anti-freezing agents are ethylene glycol, propylene glycol, urea and glycerin. Examples for anti-foaming agents are silicone emulsions (such as e. g. Silikon^® SRE, Wacker, Germany or Rhodorsil^®, Rhodia, France), long chain alcohols, fatty acids, salts of fatty acids, fluoroorganic compounds and mixtures thereof.

The treatment of crop with the aqueous formulation may be done by applying said formulation by ground or aerial application, preferably by ground application. Suitable application devices are a predosage device, a knapsack sprayer, a spray tank or a spray plane.

The aqueous formulation is often applied onto the hop plants. This means, that the application is directed to the plants, but not to the soil where the plants grow. Preferably, the aqueous formulation is applied by spraying in the direction of the leaves of the hop plants.

The acylcyclohexanedione is typically applied in an amount from 300 to 1600 g/ha, preferably from 400 to 1500 g/ha, more preferably from 500 to 1400 g/ha.

The concentration of the acylcyclohexandione in the aqueous formulation is often from 10 to 200 ppm, preferably from 30 to 170 ppm, more preferably from 40 to 120 ppm, and most preferred from 40 to 70 ppm. Typically, 0,5 to 5 I, preferably 1 ,0 to 3,5 and more preferably 1 ,5 to 2,5 I of the aqueous formulation is applied per hop plant.

The term "growth stage" refers to the growth stages as defined by the BBCH Codes in "Growth stages of mono-and dicotyledonous plants", 2nd edition 2001 , edited by Uwe Meier from the Federal Biological Research Centre for Agriculture and Forestry. The BBCH codes are a well established system for a uniform coding of phonologically similar growth stages of all mono- and dicotyledonous plant species. In hop the BBCH codes are defined as follows:

Principal growth stage 5 (Inflorescence emergence): 51 Inflorescence buds visible; 55 Inflorescence buds enlarged.

Principal growth stage 6 (Flowering): 61 Beginning of flowering: about 10% of flowers open; 62 About 20% of flowers open; 63 About 30% of flowers open; 64 About 40% of flowers open; 65 Full flowering: about 50% of flowers open; 66 About 60% of flowers open; 67 About 70% of flowers open; 68 About 80% of flowers open; 69 End of flowering.

Principal growth stage 7 (Development of cones): 71 Beginning of cone development: 10% of inflorescences are cones; 75 Cone development half way: all cones visible, cones soft, stigmas still present; 79 Cone development complete: nearly all cones have reached full size. Typically, the acylcyclohexandione is applied at its growth stage BBCH from 61 to 79, preferably from 69 to 73, more preferably 69 to 71.

The acylcyclohexandione may be applied once or several times during the above men- tioned growth stages. Preferably, it is applied only once or twice, more preferably once.

In a preferred embodiment of the method according to the invention, the yield of the cone mass of the hop plants is increased. Typically, the yield is increased at least 3 %, preferably at least 7 %, more preferably at least 10 % and most preferably at least 13 %. The increase is usually compared to the yield without treating the hop plants according to the invention. The yield of the cone mass is measured in kg per hectare. To determine the yield, the cones from each bine are separated from stems and leaves using a single-bine picking machine, for example from Wolf, Germany. In another preferred embodiment of the method according to the invention, the concentration of prenylated polyketides in the cones of the hop is increased. Typically, the yield is increased at least 10 %, preferably at least 20 % and more preferably at least 30 %. The yield increase relates to the sum of the prenylated polyketides. The increase is usually compared to the yield without treating the hop plants according to the inven- tion. The concentration is measured in mg component per g kiln dried cone weight, wherein the cones have a moisture of 8-13 wt% and wherein the components are extracted with methanol. A suitable method for determining the concentration is presented in Example 2. Preferably, the prenylated polyketides are prenylflavonoids such as demythylxanthohumol (DMX) and xanthohumol; α-acids, such as cohumulone (Co- Hum), humulone (Hum) and adhumulone (AdHum); and β-acids, such as colupulone (CoLup) and lupulone (Lup). More preferably, the prenylated polyketides are α-acids and β-acids.

The invention also relates to a use of an acylcyclohexanedione of the formula I for in- creasing the yield of cone mass of the hop plants. Preferably, the acylcyclohexanedione is applied onto the hop plants at their growth stage BBCH from 61 to 79, preferably from 69 to 73, more preferably 69 to 71.

The invention further relates to a use of an acylcyclohexanedione of the formula I for increasing the concentration of prenylated polyketides in the cones of the hop plants. Preferably, the acylcyclohexanedione is applied onto hop plants at their growth stage BBCH from 61 to 79, preferably from 69 to 73, more preferably 69 to 71.

Advantage of the present invention is that the quality of the hop cones is increased. Especially the concentration of bitter acids is increased. Another advantage is that the yield of the hop cones is increased. Yet another advantage is that this can be achieved with a single treatment of the plants. The invention is further illustrated but not limited by the following examples.

Examples

Regulaid®: a non-ionionic surfactant comprising a 91 wt% of a mixture of 2- butoxyethanol, poloxalene (a non ionic surfactant block copolymer of ethylene glycol and propylene glycol) and monopropylene glycol, commercially available from KaIo, Inc., USA.

Regalis®: Water-dispersible granules comprising 10 wt% prohexadione-calcium, commercially available from BASF SE, Germany.

Example 1 : Yield increase

Hop cultivars "Willamette" were grown in 2007 under standard agronomic conditions at Golden Gate Ranches, S. S. Steiner, Inc, Yakima, WA, USA. The plants were split into two experimental groups and a control group, wherein each group consisted of 37 plants. Control groups were exposed to mock treatments with the surfactant in de- ionized water. Plants were drenched "until run-off" at dusk to favor foliar absorption of prohexadione-calcium. The tank mix was prepared by mixing 1 kg Regalis® with 2000 I de-ionized water and 20 I Regulaid®. The application rate of the tank mix was 2.0 I per plant, which corresponded to 648 g prohexadione-calcium per hectare. The plants were treated once at growth stage BBCH 69 (on July 29, 2007) or, alternatively, once at growth stage BBCH 71 (on August 7, 2007).

To conduct yield analysis each plant was separately cut at a stem height of 0.5m and removed from the overhead trellis. Cones from each bine were separated from stems and leaves using a single-bine picking machine (Wolf, Germany) and total cone mass was weighed using digital scales coupled to a computer database. Samples consisted of single bine measurements of the plants.

A yield increase of 14,6 % of cone mass was found for the treatment on July 29, 2007 compared to the control group. This increase was significant as calculated by one-way ANOVA (Analysis Of Variance), where p=0.025. The standard error was +/- 3.05 %. An increase of 7,1 % was found for the treatment on August 7, 2007 with a standard error of +/-3.2 %. Example : Increase in bitter acids

Hop cultivars "Zeus" were grown under standard agronomic conditions at Golden Gate Ranches, S. S. Steiner, Inc, Yakima, WA, USA. The plants were split into a experimental group and a control group. Control groups were exposed to mock treatments with the surfactant in de-ionized water (Table 1 "Control"). Plants were drenched "until runoff" at dusk to favor foliar absorption of prohexadione-calcium. The tank mix was prepared by mixing 1 kg Regalis® with 2000 I de-ionized water and 20 I Regulaid®. The application rate of the tank mix was 2 I per plant, which corresponds to 648 g prohexadione-calcium per hectare. The plants were treated once at growth stage BBCH 69 (Table 1 "Treated").

Secondary metabolites were quantified by UHPLC. Kiln dried hop cones (9-12% mois- ture) were ground in a high-throughput system using 50 ml centrifuge tubes, a paint shaker, stainless steel balls, and liquid nitrogen. Ground tissue (0.25 g) was weighed and placed in 20 ml tubes in a 96-array rack. Methanol (10 ml) was added to each tube using robot-assisted pipetting on a Parkard Multiprobe NEx (Perkin Elmer, USA). Tissue was extracted by stir plate agitation for 20 minutes. The resulting extract was then transferred to a 96-well plate for analysis by Waters ACQUITY UPLC™ coupled to an ACQUITY PDA detector with a 1.7 micron BEH particle, 100 mm C18 column held at a constant temperature of 60⁰C. Efficient separation was achieved using a solvent system containing A:4% aqueous phosphoric acid (H3PO4) and B:4% H3PO4 in acetoni- trile at the following gradient: 0-0.7 min B:0-30%, 0.7-1.1 min B:30%, 1.1-1.9 min B:30- 60%, 1.9-2.0 min B:60-100%, 2.0-2.5 min B:100%. Empower Il software was used to quantify seven terpenophenolics, including: (1 ) prenylflavonoids, demythylxanthohumol (DMX) and xanthohumol (XN); (2) α-acids, cohumulone (CoHum), humulone (Hum) and adhumulone (AdHum); and (3) β-acids, colupulone (CoLup) and lupulone (Lup). Prenylflavonoids were quantified at a wavelength of 370 nm while α- and β-acids were quantified at a wavelength of 325 nm. These HTS methods of analysis allowed for sample sizes of 11 ,962 extracts from 5,981 samples equally divided between controls and treatments.

As outlined, seven terpenophenolic components were quantified; two prenylflavonoids, XN and DMX, and five prenylated acylphloroglucinols (α- and β-acids) Hum, CoHum, AdHum, Lup, and CoLup. Accumulation of all seven terpenophenolics increased, and highly statistically significant differences are shown (Table 1). Prenylflavonoid content increased by 39.1 to 48.6% while α- and β- acids increased by 30.5 to 46.7% when compared to controls (p < 0.001 , one-way ANOVA). Table 1 : Changes in metabolite concentration in hop cones (data as mg metabolite per g dried cones)

Claims

I . A method for treating hop plants comprising the application of an aqueous formulation containing acylcyclohexanedione of the formula I

where R is hydrogen or d-Cε-alkyl, and R' is d-Cε-alkyl or Cs-Cβ-cycloalkyl, or a salt of said acylcyclohexanedione, onto the hop plants at their growth stage BBCH from 61 to 79.

2. The method according to claim 1 , wherein the acylcyclohexanedione is applied in an amount from 300 to 1600 g/ha.

3. The method according to claims 1 or 2, wherein the acylcyclohexanedione is applied in an amount from 500 to 1400 g/ha.

4. The method according to claims 1 to 3, wherein the aqueous formulation comprises 10 to 200 ppm of the acylcyclohexanedione.

5. The method according to claims 1 to 4, wherein the aqueous formulation com- prises 0.01 to 20 wt% of a non-ionic surfactant.

6. The method according to claims 1 to 5, wherein the yield of cone mass of the hop plants is increased.

7. The method according to claims 1 to 6, wherein the concentration of prenylated polyketides in the cones of the hop is increased.

8. The method according to claims 1 to 7, wherein the acylcyclohexanedione is pro- hexadione-calcium.

9. The method according to claims 1 to 8, wherein the acylcyclohexanedione is applied at the growth stage BBCH from 69 to 73.

10. A use of an acylcyclohexanedione of the formula I according to claim 1 for in- creasing the yield of cone mass of the hop plants.

I 1. The use according to claim 10, wherein the acylcyclohexanedione is applied onto the hop plants at their growth stage BBCH 61 to 79.

12. A use of an acylcyclohexanedione of the formula I according to claim 1 for increasing the concentration of prenylated polyketides in the cones of the hop plants.

13. The use according to claim 12, wherein the acylcyclohexanedione is applied onto hop plants at their growth stage BBCH 61 to 79.