US 2010O227009 A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2010/0227009 A1 Spooner-Hart et al. (43) Pub. Date: Sep. 9, 2010
(54) METHODS AND COMPOSITIONS FOR Publication Classification CONTROLLING PESTS (51) Int. Cl. AOIN 65/00 (2009.01) (75) Inventors: Robert Neil Spooner-Hart, AOIN 35/06 (2006.01) Kurrajong (AU); Albert Habib AOIN3L/4 (2006.01) Basta, Glenwood (AU) AOIN 37/42 (2006.01) AOIN 33/10 (2006.01) Correspondence Address: AOIN 33/24 (2006.01) KNOBBE MARTENS OLSON & BEAR LLP AOIN3L/6 (2006.01) 2040 MAINSTREET, FOURTEENTH FLOOR AOIN 6.5/22 (2009.01) AOIN 65/10 (2009.01) IRVINE, CA 92.614 (US) AOIN 65/12 (2009.01) AOIN 65/04 (2009.01) (73) Assignees: BIOPROSPECT LIMITED, Perth AOIN 65/48 (2009.01) (AU); UNIVERSITY OF AOIP5/00 (2006.01) WESTERN SYDNEY, South AOIP 7/00 (2006.01) Penrith DC (AU) AOIP 9/00 (2006.01) (52) U.S. Cl...... 424/742: 514/678; 514/690: 514/689; (21) Appl. No.: 12/782,125 514/685; 514/718; 514/576; 514/646; 514/644; 514/706: 424/725; 424/747; 424/756 (22) Filed: May 18, 2010 (57) ABSTRACT Related U.S. Application Data The present invention is directed to pest-controlling compo sitions comprising as active ingredients one or more f3-di (63) Continuation of application No. 12/134,035, filed on ones, particularly B-diketones and B-triketones, and to the use Jun. 5, 2008, which is a continuation of application No. of these compositions inter alia for preventing, eradicating, 10/477.057, filed on Sep. 23, 2004, now abandoned, destroying, repelling or mitigating harmful, annoying or filed as application No. PCT/AU02/00569 on May 8, undesired pests including insects, arachnids, helminths, mol 2002. luscs, protozoa and viruses. The present invention is further directed to processes of preparing B-diones by de novo syn thesis or from natural Sources Such as Volatile oil-bearing (30) Foreign Application Priority Data plants from families including Alliaceae, Apiaceae, Aster aceae, Cannabinaceae, Lamiaceae, Pteridaceae, Myrtaceae, May 8, 2001 (AU) ...... PR 4842 Myoporaceae, Proteaceae, Rutaceae and Zingiberaceae.
O O C) O O O O O C) O or C) OCH tasman one angustione dehydroangsutione flawesome
O o C) O O O O O O O O O OHC CHO OCHs. O O O leptosperm one isoleptosperm one Jense none agglomerone
O O O O O O O O O O O O
O C) OCH3 xanthostem one papuanone grandiflorone lateriticone
HC) H O C) HCO OCH O O O Cha 3 3 H3CO OCH3
H H3C CH3 H3C CH3 OCH OCH3 OCH OCHs.
torquatone isolateriticone
O C) O HH OC OCCH3 H3CO OCH3 H3CO OCH3 O O H3C CH3 OCH3 CCH OCH3 O
baecked conglomerone Patent Application Publication Sep. 9, 2010 Sheet 1 of 4 US 2010/0227009 A1
O O O O O O O O O O O O
O OCH3 tasman one angustione dehydro angsutione flawesone
O O O O O O O O O O O O
OHC CHO OCH3 O O O leptosperm one isoleptospermone jense none agglomerone
O O O O O O O O O O O O
O CH O O 3 Xanthostem one papuanone grandiflorone lateriticone
HO HO O O HCO O CH O O H3CO OCH 3 3 H3CO OCH3 HC CH H3C CH 3 O CH 3 H3C CH3 O CH OCH3 3. OCH3 3
torquatone isolateriticone
O O O O HO OCH HCO OCH3 HCO OCH O O
H3C H3C CH3 OCHs OCH3 O CHs O
baeckeo conglomerone
FIGURE I Patent Application Publication Sep. 9, 2010 Sheet 2 of 4 US 2010/0227009 A1
Abundance
TC: 96O1 OO1 D 18OOOOO 17OOOOO 16OOOOO 15 OOOOO 14 OOOOO 13 OOOOO 12OOOOO 1 1 OOOOO 1 OOOOOO 9 OOOOO 8OOOOO 7OOOOO 6 OOOOO 5OOOOO 4OOOOO 3OOOOO 2OOOOO 6 5 1 OOOOO
I I I I I I I I I I I I It I, II, A I I I I I I I I I I I I I illI I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I I It 8 OO1 OOO12 OO14 OO16 O18O2OOO22 OC24 OC26 OC28 OOBOOCB2. OCB.4 OO Time --> FIGURE 2 Patent Application Publication Sep. 9, 2010 Sheet 3 of 4 US 2010/0227009 A1
H NMR Data (CDC13,500MHz)
"H (see below) Signal (ppm) | Integral Signal (ppm) | Integral 19.18 O.89 1845 O.48
3.98 109 3.85 O55
3.96 2.75 3.87 155
1.97 3.00 192 172
134 6.43 1.46 352
115 6.13 117 3.48
FIGURE 3 Patent Application Publication Sep. 9, 2010 Sheet 4 of 4 US 2010/0227009 A1
os-- os HO-le-SeO Os-S-OH -a-e- - e -2 - O CH O CH 3 3 A B C
FIGURE 4 US 2010/0227009 A1 Sep. 9, 2010
METHODS AND COMPOSITIONS FOR tones, such as those obtainable from Volatile oil-bearing CONTROLLING PESTS plants including plants from the families Alliaceae, Apiaceae, Asteraceae, Cannabinaceae, Lamiaceae, Pteridaceae, Myrta CROSS-REFERENCE TO RELATED ceae, Myoporaceae, Proteaceae, Rutaceae and Zingiber APPLICATIONS aceae, exhibit significant pesticidal, especially insecticidal, 0001. This application is a continuation of U.S. applica arachnicidal, helminthicidal and/or molluscicidal activity. tion Ser. No. 12/134,035, filed Jun. 5, 2008, which is a con This discovery has been reduced to practice in novel pest tinuation of U.S. application Ser. No. 10/477.057, filed Sep. controlling compositions and methods for their preparation 23, 2004. U.S. application Ser. No. 10/477,057 is the U.S. and use, as described hereinafter. National Phase under 35 U.S.C. S371 of International Appli cation No. PCT/AU02/00569, filed May 8, 2002 designating BRIEF DESCRIPTION OF THE DRAWINGS the U.S. and published in English as WO 02/089587, which 0005 FIG. 1 shows the structures relating to the major claims priority to Australian Patent Application PR 4842, constituents of the published Myrtaceae essential oils. filed May 8, 2001. This application incorporates herein by 0006 FIG. 2 is a representation of a GC-MS trace of E. reference U.S. application Ser. No. 12/134,035, U.S. appli cloeziana oil. cation Ser. No. 10/477.057, International Application No. 0007 FIG. 3 is a tabular and graphical representation PCT/AU02/00569 including the International Publication showing "H NMR data recorded on a fraction (F4) obtained No. WO 02/089587, and Australian Patent Application PR from silica gel chromatography of E. cloeziana oil and the 4842 in their entireties. structure of the major and minor isomers of the compound deduced from these data. FIELD OF THE INVENTION 0008 FIG. 4 is a diagrammatic representation showing 0002 This invention relates generally to methods and various tautomeric forms of an isolated B-triketone com compositions for controlling pests. More particularly, the pound in solution (CDC1). present invention relates to pest-controlling compositions comprising as active ingredients one or more f3-diones, par DETAILED DESCRIPTION OF THE INVENTION ticularly 3-diketones and B-triketones, and to the use of these 0009. One aspect of the present invention contemplates compositions inter alia for preventing, eradicating, destroy the use of a B-dione compound, particularly a 3-diketone or a ing, repelling or mitigating harmful, annoying or undesired B-triketone compound, in the preparation of a composition pests including insects, arachnids, helminths, molluscs, pro for controlling harmful, annoying or undesired pests, said tozoa and viruses. The present invention further relates to processes of preparing B-diones by de novo synthesis or from compound being represented by the general formula (I) natural sources such as Volatile oil-bearing plants from fami lies including Alliaceae, Apiaceae, Asteraceae, Cannabi (I) naceae, Lamiaceae, Pteridaceae, Myrtaceae, Myoporaceae, A B Proteaceae, Rutaceae and Zingiberaceae. Bibliographic details of various publications referred to in this specification are collected at the end of the description. Q BACKGROUND OF THE INVENTION 0003 Triketones have been used for many years as herbi wherein cides for the control of undesired vegetation. Herbicidal trike 0010) A is (C=O)R., (C=S)R, OR, SR (CRNRR), tones have been described, for example, in EP-A-33.8992, C(R)OR, NRRs (C=N Ra)R, N=O, N(EO), EP-A-336898, U.S. Pat. No. 4,869,748, EP-A-186118, EP NROR or SOR: A-186119, EP-A-186120, U.S. Pat. No. 4,202,840, U.S. Pat. 0011 R is selected from H. C-C alkyl, C-C aryla No. 4,695,673, U.S. Pat. No. 4,780,127, U.S. Pat. No. 4,921, lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla 526, U.S. Pat. No. 5,006,150, U.S. Pat. No. 5,545,607, U.S. lkyl, C-C haloalkyl, C-Co dihaloalkyl, C-C triha Pat. No. 5,925,795, U.S. Pat. No. 5,990,046, U.S. Pat. No. loalkyl, C-Co haloalkoxy, C-Co hydroxyalkyl, C-Co 6,218,579, EP-A-249.150, EP-A-137963, EP-A-394889, EP thioalkyl and C-C nitroalkyl, OR, SR (CRNRRs), A-506907 or EP-B-135191. Examples of herbicidal trike NRRs (C=N-R)R N=O, N(=O), NROR, or tones are interalia Sulcotrione (MIKADOR) whose chemical SOR7; designation is 2-(2-chloro-4-methanesulfonylbenzoyl)-1,3- 0012 R is selected from H. C-C alkyl, C-C aryla cyclohexandione, 2-(4-methylsulfonyloxy-2-nitrobenzoyl)- lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla 4,4,6,6-tetramethyl-1,3-cyclohexane dione: 3-(4-methylsul lkyl, C-C haloalkyl, C-Co dihaloalkyl, C-C triha fonyloxy-2-nitrobenzoyl)-bicyclo-3.2.1]octane-2,4-dione; loalkyl, (CRNRRs). NRRs (C=N R)R. N=O. 3-(4-methylsulfonyl-2-nitrobenzoyl)-bicyclo-3.2.1]octane N(-O), or NROR; 2,4-dione: 4-(4-chloro-2-nitrobenzoyl)-2,6,6-trimethyl-2H 0013 R is selected from H. C-C alkyl, C-C aryla 1.2-oxazine-3,5(4H, 6H)-dione: 3-(4-methylthio-2-nitroben lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla Zoyl)-bicyclo3.2.1]octane-2,4-dione; 4-(2-nitro-4- lkyl, C-C haloalkyl, C-Co dihaloalkyl, C-C triha trifluoromethoxybenzoyl)-2,6,6-trimethyl-2H-1,2-oxazine loalkyl, C-Chaloalkoxy, OR7, SR-7 (CRNRRs). NRRs. 3.5(4H, 6H)-dione. (C=N-R)R, N=O, N(-O), NROR, or SOR: 0014 Ra and Rs are independently selected from H. SUMMARY OF THE INVENTION C-Co alkyl, C-Co arylalkyl, C-C cycloalkyl, C-Coalk 0004. The instant invention is predicated in part on the enyl, C-Coheteroarylalkyl, C-Cohaloalkyl, C-Co diha discovery that 3-diones, particularly B-diketones and B-trike loalkyl, C-C trihaloalkyl, OR, or SR-7; US 2010/0227009 A1 Sep. 9, 2010
0015 R is selected from H. C-C alkyl, C-C aryla ally substituted with one or more groups, as defined above. lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla Further, bi- or tricyclic heteroaryl moieties may comprise at lkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C triha least one ring, which is either completely, or partially, Satu loalkyl, C-C haloalkoxy, OR, SR-7, (CRNRR), rated. Suitable heteroaryl moieties include, but are not limited NRoRo or NROR7; to oxazolyl, thiazaoyl, thienyl, furyl, 1-isobenzofuranyl. 0016 R, is selected from H. C-C alkyl, C-C aryla 2H-pyrrolyl. N-pyrrolyl, imidazolyl pyrazolyl, isothiazolyl, lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla isooxazolyl pyridyl, pyrazinyl, pyrimidinyl, pyradazinyl, lkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C triha indolizinyl, isolindolyl, indoyl, indolyl, purinyl, phthalazinyl. loalkyl; 0029. The articles “a” and “an are used herein to refer to 0017 R is selected from H. C-C alkyl, C-C aryla one or to more than one (i.e., to at least one) of the grammati lkyl, C-C cycloalkyl, C-C alkenyl, C-C heteroaryla cal object of the article. By way of example, “an element' lkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C triha means one element or more than one element. loalkyl, OR, SR or NRRo: 0030 A preferred carbocyclic ring formed by Q is an 0018 R, and Ro are independently selected from H. optionally substituted cyclohexanedione. C-Coalkyl, C-Co arylalkyl, C-C cycloalkyl, C-Coalk 0031. A preferred subgroup of compounds of formula (I) enyl, C-Coheteroarylalkyl, C-Cohaloalkyl, C-Co diha is represented by formula (II) loalkyl, C-Co trihaloalkyl, OR or SR; 0019 R is selected from H. C-C alkyl, C-C aryla lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla (II) lkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C triha loalkyl; 0020 R2 is selected from H. C-C alkyl, C-C aryla lkyl, C-C cycloalkyl, C-Coalkenyl, C-Co heteroaryla lkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C triha loalkyl; 0021 B is H. C-C alkyl, C-C alkenyl, aryl or het eroaryl; 0032 Such compounds may exist in a number of tauto 0022 X and Y are independently selected from oxygen, meric forms. For example, in the case wherein X and Y are Sulfur, —N-R, and each oxygen, and B is hydrogen, then the compounds of 0023. Q completes a 5-8-member saturated or unsaturated formula II may exist as one or more of the structural formulae carbocyclic or heterocyclic ring in which optionally one or shown below. more members comprise —C(=X)—, and wherein Q is optionally substituted with one or more substituents selected from C-Co alkyl, C-C cycloalkyl, C-Coalkenyl, C-Co O R O R HO R haloalkyl, C-Co dihaloalkyl, C-C trihaloalkyl, C-Co haloalkoxy, OR, SR (CRNRRs), NRRs (C=N R.) O O N - HO 7 O O O R, N=O, N(EO), NROR, SOR, C-C 1-arylalkyl, C-C 2-arylalkyl or (C=X)R. Q Q Q 0024 Heterocyclic systems can be optionally attached to a moiety other than those set forth above via a carbon atom or 0033. It is intended that all such tautomeric structures are a heteroatom of R to R. included within the scope of the present invention. 0025 Preferred compounds represented by formula (I) are 0034. It should also be appreciated that some of the com 3-diketones and especially preferred are 3-triketones. pounds of formula (I) are capable of existing as different 0026. As used herein, the term “alkyl refers to linear or branched chains. The term “haloalkyl refers to an alkyl geometric isomers and diastereomers. The invention thus group Substituted by at least one halogen. Similarly the term includes both the individual isomers and mixtures of such “haloalkoxy' refers to an alkoxy group substituted by at least isomers. one halogen. As used herein the term “halogen refers to 0035 Another preferred subgroup of compounds of for fluorine, chlorine, bromine and iodine. mula (I) is represented by formula (III) 0027. As used herein the term “aryl refers to aromatic carbocyclic ring systems such as phenyl or naphthyl, anthra (III) cenyl, especially phenyl. Suitably, aryl is C-C with mono, A B di, tri, tetra and penta Substitution containing OR, F, Cl, Br, I, NO, CF, COR, NRRs, SOR, SR. X Y 0028. As used herein the terms "heterocycle”, “heterocy F C clic”, “heterocyclic systems’” and the like refer to a saturated, unsaturated, or aromatic carbocyclic group having a single E D ring, multiple fused rings (for example, bicyclic, tricyclic, or Z other similar bridged ring systems or Substituents), or mul tiple condensed rings, and having at least one heteroatom Such as nitrogen, oxygen, or Sulfur within at least one of the wherein rings. This term also includes "heteroaryl which refers to a 0036 X, Y and Z are each independently selected from heterocycle in which at least one ring is aromatic. Any het oxygen, sulfur, N—R or one of C=X, C=Y or C=Z is erocyclic or heteroaryl group can be unsubstituted or option CH: US 2010/0227009 A1 Sep. 9, 2010
0037 A is (C=O)R. (C=S)R, OR, SR (CRNRRs), C(R), OR, NRRs (C=N R)R, N=O, N(-O). -continued NROR or SOR: CF 0038 B is as defined above; S H 1. 0039 C, D, E and F are each independently selected from NN H, C-C alkyl, C-C cycloalkyl, C-C alkenyl, C-Co O O S S haloalkyl, C-Co dihaloalkyl, C-C trihaloalkyl, OR, SR, (CRNRRs), NRRs (C—N-R)R, N=O, N(-O), NROR, SOR, C-C 1-arylalkyl, C-C 2-arylalkyl or (C—X)R; and 0040. R. R. R. R. and Rs are as defined above. O O O 0041 Preferred B-diones represented by formula (III) are flavesone (1-isobutyroyl-3,3,5,5-tetramethylcyclohexan-2,4, CHSH 6-trione), isoleptospermone (1-isovaleroyl-3,3,5,5-tetram ethylcyclohexan-2,4,6-trione), leptospermone (1-Valeroyl-3, H3C NN 1. HO NN 1. O 3.5.5-tetramethylcyclohexan-2,4,6-trione), papuanone (1-pentoyl-3,3,5,5-tetramethylcyclohexan-2,4,6-trione), O O grandiflorone (1-(2-phenylethyl)-3,3,5,5-tetramethylcyclo hexan-2,4,6-trione) andjensenone (1-Valeroyl-3,5-dicarbon ylcyclohexan-2,4,6-trione), including analogues and deriva tives thereof. 0042. By way of example, flavesone analogues contem O O O plated by the present invention include, but are not restricted to, compounds having the following structural formulae, CHNO wherein the structural formula of flavesone is shown for com parative purposes:
O HO HN
O O O O O O O
0043. Non-limiting examples of isoleptospermone ana logues contemplated by the present invention include, but are O O O not restricted to, compounds having the following structural formulae, wherein the structural formula of isoleptosper mone is shown for comparative purposes: CHBr O
O O O HO HN
O O O O O O
O Flavesone
O O O CHBr CHCI CH2OH CHF O O O O
O O O O O O O O
O O O O US 2010/0227009 A1 Sep. 9, 2010
-continued Isoleptospermone CHCI CHOH O HO HN O O O O O O O O O O O O
O O O O O Leptospermone C HF CF CHBr
O S O S
O O O S S O O S S
O O O O O
n n H N 1. HO N 1. Ha r H3C N N r nN.1 No O O O O O O O O O O O O
O O O CHSH CHNO O O O
O O CHCI CH2OH
O O O O O O
O O O O
O O
O O
CHF CF
O O
O O O O 0044 Non-limiting examples of leptospermone analogues contemplated by the present invention include, but are not restricted to, compounds having the following structural for- O O mulae, wherein the structural formula of leptospermone is shown for comparative purposes: US 2010/0227009 A1 Sep. 9, 2010
-continued -continued CH2SH CHNO CHF CF
O O
O O O
OHC CHO OHC CO HO O O
S H YN r
S S O O
OHC CHO OHC CHO
O O
0045. Non-limiting examples of jensenone analogues contemplated by the present invention include, but are not H3C NN r H NNDC O restricted to, compounds having the following structural for mulae, wherein the structural formula of jensenone is shown O O O O for comparative purposes:
OHC CHO OHC CHO
O O CH2SH CHNO O HO O O O O O O O O O O
OHC CHO OHC CHO
O O OHC CHO OHC CHO Jensenone O O CHBr HO V HN O Nan
O O O O O O
OHC CHO OHC CHO OHC CHO
O O O CHCI CH2OH 0046. Another preferred subgroup of compounds of for mula (I) is represented by formula (IV) O O (IV) A B O O O O X Y
F OHC CHO OHC CHO E C O O US 2010/0227009 A1 Sep. 9, 2010
wherein 0047 X and Y are each independently selected from oxy -continued gen, sulfur - N R or one of C=X or C—Y is CH: CF3 0048 A is (C=O)R., (C=S)R, OR, SR (CRNRR), S H NN 1. C(R)OR, NRRs (C—N Ra)R, N=O, N(EO), NROR or SOR: O O S S 0049 B is as defined above; 0050 C, D, E and F are each independently selected from H, C-C alkyl, C-C arylalkyl, C-C cycloalkyl, C-Co alkenyl, C-Co heteroarylalkyl, C-Co haloalkyl, C-Co OCH OCH OCH dihaloalkyl, C-C trihaloalkyl, C-Co haloalkoxy, OR, CH2SH SR (CRNRR), NRRs (C=N-R)R, N=O, N(-O) NROR2, SOR, and HC NN 1. HO NN 1. O 0051 R. R. R. R. and Rs are as defined above. 0052 Preferred B-diones represented by formula (IV) are O O O O tasmanone (1-isobutroyl-4-methoxy-3.5.5-trimethylcyclo hex-3-en-2,6-dione), agglomerone(1-isobutroyl-4-methoxy 5,5-dimethylcyclohex-3-en-2,6-dione), lateriticone(1-vale royl-4-methoxy-3.5.5-trimethylcyclohex-3-en-2,6-dione), OCH OCH OCH isolateriticone(1-isovaleroyl-4-methoxy-3.5.5-trimethylcy CHNO HO clohex-3-en-2,6-dione and platyphyllol(6,6-dimethyl-2- acetyl-5-methoxycyclohex-4-ene-1,3-dione), including ana O logues and derivatives thereof. 0053 Non-limiting examples of tasmanone analogues O O O contemplated by the present invention include, but are not restricted to, compounds having the following structural for mulae, wherein the structural formula of tasmanone is shown for comparative purposes: OCH OCH
0054 Another preferred subgroup of compounds of for O HO mula (I) is represented by formula (V)
O O O O (V) A B X Y
H C OCH OCH OCH CHBr G D O F E
O O wherein 0055 X and Y are independently selected from oxygen, Sulfur or —N-R, and 0056 A is (C=O)R. (C=S)R, OR, SR (CRNRRs), OCH C(R)OR, NRRs (C=N Ra)R, N=O, N(EO), Tasmanone NROR or SOR: 0057 B is as defined above: CHCI CHOH CHF 0058 C, D, E, F, G and Hare each independently selected O O O from H. C-C alkyl, C-Co arylalkyl, C-C cycloalkyl, C-Co alkenyl, C-Co heteroarylalkyl, C-Co haloalkyl, O O O O O O C-Co dihaloalkyl, C-C trihaloalkyl, C-Cohaloalkoxy, OR, SR (CRNRRs), NRRs (C=N Ra)R, N=O, N(=O), NROR or SOR: and 0059 R. R. R. R. and Rs are as defined above. 0060 More specifically unsaturation, epoxides and OCH OCH OCH thioexpoxides may exist at positions designated by H (or G) connected to F (or E) or F (or E) connected to C (or D). A US 2010/0227009 A1 Sep. 9, 2010
four-membered ring forming a part of a bicyclic structure the following structural formulae, wherein the structural for may exist at positions designated by H (or G) connected to C mula of dehydroangustione is shown for comparative pur (or D). poses: 0061 Preferred B-diones represented by formula (V) are angustione (1-acetyl-3.5.5-trimethylcyclohex-2,6-dione), O HO HN dehydroangustione (1-acetyl-3,5,5-trimethylcyclohex-3-en 2,6-dione) and xanthostemone (1-isobutroyl-5,5-dimethyl cyclohex-3-en-2,6-dione), including their analogues and derivatives. 0062 By way of example, angustione analogues contem plated by the present invention include, but are not restricted to, compounds having the following structural formulae, Dehydroangustione wherein the structural formula of angustione is shown for O CHBr O CHCl2 comparative purposes:
HO HN OH OH r O CHOH O CHF
O CHBr
O CF S
Angustinone s HO 21 O HO 21 S O CHCl2 O CH2OH O CHF OH OH r HN1 N1 HO n1 HN1 HO O HO O
S
SH r O CH2SH O CHNO N 1 HON 1. O CH2SH N N r OH O OH
N o1 n O CHNO N HO1 n HO O
OH O OH
0.064 Non-limiting examples of XanthoStemone ana logues include, but are not restricted to, compounds having 0063) Non-limiting examples of dehydroangustione ana the following structural formulae, wherein the structural for logues include, but are not restricted to, compounds having mula of Xanthostemone is shown for comparative purposes: US 2010/0227009 A1 Sep. 9, 2010
potassium) and di-Valent metal salts (e.g., calcium, magne sium, iron or copper) and ammonium salts (e.g., isopropyl ammonium, trialkyl and tetraalkylammonium salts). O HO HN 0067. The compounds according to any one of formulae (I)-(V) can be prepared according to methods analogous to O O O O O O those known in the art for the preparation of B-diones. Exem plary methods are disclosed for example in EP-A-33.8992, EP-A-336898, U.S. Pat. No. 4,202,840, U.S. Pat. No. 4,869, 748, EP-A-186118, EP-A-186119, EP-A-186120, U.S. Pat. CHBr No. 4,695,673, U.S. Pat. No. 4,780,127, U.S. Pat. No. 4,921, 526, U.S. Pat. No. 5,006,150, U.S. Pat. No. 5,545,607, U.S. O Pat. No. 5,925,795, U.S. Pat. No. 5,990,046, U.S. Pat. No. 6,218,579, EP-A-249.150, EP-A-137963, EP-A-394889 EP O O A-5O6907 or EP-B-1351.91. 0068 More particularly, compounds according to formu lae (III)-(V) can be synthesised using the representative pro cedures outlined below. Xanthostemone 0069. For compounds according to formula (III), 1,3,5- trihydroxybenzene 1 is reacted with CHCN in the presence of Zinc chloride and hydrochloric acid, according to A. H. Blatt (1943, Org. Synth. Col. II, 522-523), affording 1-acetyl O 2,4,6-trihydroxybenzene 2 (phloroacetophenone) (R=Me) (Scheme 1). O O
OH OH O CF S H NN 1.
O O S S O O HO OH HO OH
0070 R groups other than methyl are depicted above. Reaction of 1-acetyl-2,4,6-trihydroxybenzene 2 affording 1-acetyl-3,3,5,5-tetramethylcyclohexan-2,4,6-trione 3 is a representative procedure for all compounds according to for mula (III) (R. A. Gray et al., U.S. Pat. No. 4,202,840) (Scheme 2). Anhydrous Mel (6 eq) is slowly added, at room temperature under an atmosphere of nitrogen, to a mechani cally stirred solution of 1-acetyl-2,4,6-trihydroxybenzene 2 (1 eq) and Sodium ethoxide (6 eq) in anhydrous methanol. The mixture is refluxed for 4 hours. On cooling the mixture is concentrated under vacuo, providing a residue, which is diluted with water and acidified with 2 M hydrochloric acid. Diethylether extracts are washed with saturated sodium sulfite solution, water and then dried (NaSO). Evaporation of the diethylether provides the desired product 3.
OH O 0065 Derivatives of the above compounds include, but are not restricted to, ethoxylate derivatives, propoxylate deriva tives, hydrates, aldehyde derivatives, ester derivatives, ether derivatives, alcohol derivatives, phenol derivatives, amine derivatives, other biologically or chemically equivalent Sub HO OH stances, and any combination of two or more of the foregoing. 0066 Similarly effective as pesticides are salts of the above compounds, including mono-valent salts (e.g., Sodium, US 2010/0227009 A1 Sep. 9, 2010
0071. For mono, di, tri and tetra B, C, D, E, F substitution 0074 Compounds of formula (V) can be prepared accord patterns, reactions between one and seven mole equivalents ing to a first representative procedure, as follows: 4,4-dim of R-I and sodium ethoxide are used. Lawasson’s reagent is ethylcyclohexane-1,3-dione 7 (1 mole eq) (commercially used for conversion of oxygen into Sulfur groups and sodium available) is dissolved in anhydrous diethylether and hexam borohydride or sodium cyanoborohydride is used to reduce ethylphosphoramide (solvent ratio, 20:1 respectively) under ketone, thioketone and imino groups. When additional car an atmosphere of nitrogen. The mixture is cooled to 0°C. and bonyl groups are introduced into the cyclohexane ring system lithium diisopropylamide (2.1 mole eq) is added dropwise the procedure of Crow is utilised (M. L. Bolte et al., 1985, over 40 mins. The mixture is then stirred for a further 10 mins Agric. Biol. Chem., 49, 761). 0072 Compounds of formula (IV) can be prepared before the addition of methyl iodide (1 mole eq). The mixture according to a first representative procedure, as follows: is stirred for 12 h and then benzoyl cyanide (2 mole eq) is 3-methoxy-2,4,4-trimethylcyclohex-2-en-1,5-dione 4 (1 added and the mixture stirred for a further 24 h. The reaction mole eq), prepared according to Herzig (J. Herzig, and F. was quenched with water and the ether layer partitioned and Wenzel, Monatsh, 1903, 24, 101), is dissolved in anhydrous dried (NaSO). The solvent was removed and the residue diethylether and hexamethylphosphoramide (solvent ratio, Subjected to SiO2 column chromatography (hexane/ethyl 20:1 respectively) under an atmosphere of nitrogen. The mix acetate) affording 1-benzoyl-3,3,5-trimethylcyclohexan-2,6- ture is cooled to 0°C. and lithium hydride (1.1 mole eq) (60% dione 8 (Scheme 4). in mineral oil) is added in portions. After addition the mixture is stirred for a further 10 mins before the addition of benzoyl cyanide 5R CO CN, R is depicted above (1.1 mole eq). Scheme 4 The mixture is allowed to warm to room temperature over 12h at which time the reaction is quenched with water and parti O O tioned. The ether layer is dried (NaSO) and evaporated 1) LiN(iC3H7) He affording crude 1-benzoyl-3-methoxy-2,4,4-trimethylcyclo 2) Mel hex-2-en-1,5-dione 6 which is purified by SiO, column chro 3) 7 matography (hexane/ethyl acetate, gradient) (Scheme 3). O O O 7 8 Scheme 3 O 0075 Alternatively, the compounds of formula (V) can be O O prepared according to a second representative procedure, as follows: 4,4-dimethyl-1,3-cyclohexanedione 7 (1 mole eq) -- He (commercially available) and benzoyl cyanide are dissolved in anhydrous dichloromethane and cooled to 0°C. under an 1. or C’s atmosphere of nitrogen. To the cooled solution is added anhy O drous finely powdered zinc chloride (1.1 mole eq) followed 4 5 by slow addition of triethylamine (1.2 mole eq). The reaction mixture is stirred at room temperature for 5-6 hand then O O poured into 2 Mhydrochloric acid. The mixture is partitioned and the dichloromethane layer washed with 5% sodium car bonate. The aqueous carbonate phase is then acidified with 1. O hydrochloric acid and extracted with methylene chloride and dried (Na2SO). The solvent is removed and the residue sub O O jected to SiO, column chromatography (hexane/ethyl acetate) affording 1-benzoyl-3,3,5-trimethylcyclohexan-2,6- dione 8. (W. J. Michaely and G. W. Kraatz, EP-B-135191). 0073. Alternatively, compounds of formula (IV) can be 0076 Dehydroangustione and xanthostemone derivatives prepared according to a second representative procedure, as are simply derived from dehydrogenation of angustione follows: 3-methoxy-2,4,4-trimethylcyclohex-2-en-1,5-dione derivatives, for example, by treatment of 1-benzoyl-3,3,5- 4 (1 mole eq) (commercially available) and benzoyl cyanide trimethylcyclohexan-2,6-dione 8 with palladium on charcoal are dissolved in anhydrous dichloromethane and cooled to 0° in methanol, which thereby affords 1-benzoyl-3,5,5-trimeth C. under an atmosphere of nitrogen. To the cooled solution is ylcyclohex-3-en-2,6-dione 9 (Scheme 5). added anhydrous finely powdered zinc chloride (1.1 mole eq.) followed by slow addition of triethylamine (1.2 mole eq). The reaction mixture is stirred at room temperature for 5-6 hand then poured into 2 M hydrochloric acid. The mixture is par titioned and the dichloromethane layer is washed with 5% Sodium carbonate. The aqueous carbonate phase is then acidi fied with hydrochloric acid and extracted with methylene chloride and dried (NaSO). The solvent is removed and the residue subjected to SiO2 column chromatography (hexane/ ethyl acetate) affording 1-benzoyl-3-methoxy-2,4,4-trimeth ylcyclohex-2-en-1,5-dione 6 (W. J. Michaely and G. W. Kraatz, EP-B-135191). US 2010/0227009 A1 Sep. 9, 2010 10
be understood, in this regard, that the present invention is not -continued limited to the use or practice of any one particular method of extracting Volatile oils. I0081 Suitably, the compositions comprise naturally-oc curring compounds derived from a volatile oil-bearing organ ism. Thus, in a preferred embodiment, the pesticidal compo sition of the invention comprises one or more B-dione active compounds, particularly B-diketone- and/or B-triketone-ac tive compounds, that are derived from the volatile oil of a Volatile oil-bearing organism. In this embodiment, the com position may optionally contain a naturally-occurring carrier and/or other naturally-occurring additives. 0077 Metal salts (enolates) of the above compounds can I0082 Naturally-occurring additives contemplated by the be prepared by the reaction of triketone derivatives with the present invention include natural antioxidants, which can be corresponding metal hydroxides Suspended in methanol or used advantageously to reduce the effect of oxidation of the ethanol. Trialkylammonium salts can be prepared by the reac compounds of the invention. An example of a Suitable natu tion of triketone derivatives (e.g., 3) with trialkylamines in a rally-occurring antioxidant is C-tocopherol. Other additives, chlorinated solvent such as dichloromethane. Tetraalkylam Such as naturally-occurring stabilisers, are also contemplated, monium salts can be prepared by adding a halogenated tet which may desirably be added to improve the stability and raalkylammonium salt to a metal salt in dichloromethane, shelf life of the composition. Examples of suitable natural which precipitates the metalhalide removed by filtration. The stabilisers include gum arabic, guar gum, Sodium caseinate, pure material is obtained by evaporation of the filtrate. polyvinyl alcohol, locust bean gum, Xanthan gum, kelgum, 0078. The present inventors have discovered that the B-di and mixtures thereof. ones of the invention can be obtained from natural sources I0083. In an alternate embodiment, the naturally-occurring and, in particular, from Volatile oil-bearing organisms. compounds derived from a volatile oil may be modified or Accordingly, in another aspect, the present invention encom derivatised to improve, for instance, their shelf-life, stability, passes the use of a 3-dione compound, particularly a B-dike activity and/or bioavailability. tone or a f-triketone compound, obtainable from a volatile I0084. The compounds of the present invention are useful oil-bearing organism, including an analogue or derivative for controlling harmful, annoying or undesired pests. They thereof, in the preparation of a pesticidal composition for may be used singularly or in combination with other pest controlling harmful, annoying or undesired pests. controlling compounds of the invention. By “controlling' is 007.9 The present invention contemplates the use of any meant preventing, combating, eradicating, destroying, repel Volatile oil-bearing organism that produces B-diones, prefer ling, or mitigating pests or increasing the mortality or inhib ably the B-diones according to any one of formulae (I)-(V). iting the growth and/or development of pests. The term "pest' and especially B-diketones and/or B-triketones, for the prepa is used herein in its broadest sense and includes within its ration of the pesticidal compositions of the invention. Pre Scope insects, arachnids (e.g., acari, spiders), helminths (e.g., ferred volatile oil-bearing organisms are volatile oil-bearing nematodes), molluscs, protozoa (e.g., Plasmodium sp. Para plants including, but not restricted to, plants from the families mecium sp.), viruses (e.g., herpesviruses) and the like. Suit Alliaceae, Apiaceae, Asteraceae, Cannabinaceae, Lamiaceae, able applications for such control include, but are not limited Pteridaceae, Myrtaceae, Myoporaceae, Proteaceae, Rutaceae to, combating and/or eradicating infestations by pests in ani and Zingiberaceae. Preferably, the volatile oil-bearing plant mals (including humans) and/or plants (including trees) and/ is selected from genera of the Myrtaceae family including, or stored products, which includes the administration to the but not limited to, Angophora, Austromyrtus, Backhousia, animal or site of an effective quantity of a compound of the Baeckea, Callistemon, Corymbia, Darwinia, Eucalyptus, invention. Kunzea, Leptospermum, Melaleuca, Syzygium and Xanthos I0085. By “effective amount' is meant the administration teinOn. or application of that amount of active compound, either in a 0080 Thus, the compositions of the present invention may single dose or as part of a series, that is effective for control contain as active ingredients Substantially purified 3-diones ling a significant number of pests. Thus, for example, a “pes or crude B-dione-containing extracts obtained from a volatile ticidally-effective” amount is the amount of active compound oil-bearing organism, preferably a Volatile oil-bearing plant. that is effective for increasing the mortality or decreasing the Volatile oils, also known in the art as essential oils, typically growth of a significant number of pests. Alternatively, a "pest comprise a Volatile mixture of esters, aldehydes, alcohols, repelling effective amount is the amount of active compound ketones and terpenes, which can be prepared from botanical that is noxious to, and/or induces behavioural changes in, a materials or plant cell biomass from cell culture. Volatile oils significant number of pests. The effective amount will vary can be prepared by Subjecting botanical materials to a distil depending upon the taxonomic group of pest exposed to the lation process, for example. A number of different procedures active compound, the formulation of the composition, and can be used for distillation. For example, plant matter (e.g., other relevant factors. It is expected that the amount will fall foliage, stems, roots, seeds, bark etc) of a Volatile oil-bearing in a relatively broad range that can be determined through plant is placed in a suitable still and steam distillation is used routine trials. to break down the cells of the plant to release the oil. The I0086 Accordingly, the compounds of the present inven steam is then condensed and the oil phase is separated from tion can be used as pesticides, such as but not limited to the aqueous phase to obtain the volatile oil. It will be appre insecticides, arachnicides, anti-helminthics, molluscicides ciated that other methods of Volatile oil extraction (e.g., Sol antivirals, antiprotozoals and the like, or as pest repellents vent extraction) are known to those of skill in the art and it will including repellents of insects, arachnids, helminths, mol US 2010/0227009 A1 Sep. 9, 2010
luscs, protozoa and viruses. In especially preferred embodi carriers are water, alcohols such as methanol and ethanol, ments, the compounds of the present invention are used in the ketones such as acetone and methyl ethyl ketone, aromatic control of insects, arachnids, helminths or molluscs. In prac hydrocarbons such as benzene, toluene, Xylene, ethylbenzene tice, the compounds can be applied as formulations contain and methylnaphthalene, aliphatic hydrocarbons such as hex ing the various adjuvants and carriers known to or used in the ane, cyclohexane, kerosene and light oil, esters such as ethyl industry for facilitating bioavailability, stability and disper acetate and butyl acetate, nitriles such as acetonitrile and sion. The choice of formulation and mode of application for isobutyronitrile, ethers such as diisopropyl and dioxane, acid any given compound may affect its activity, and selection will amides such as N,N-dimethylformamide and N,N-dimethy be made accordingly. lacetamide, halogenated hydrocarbons such as dichlo 0087. In general, a pest-controlling compound of the romethane, trichloroethane and carbon tetrachloride, dim invention can be compounded with appropriate inert carriers ethyl sulfoxide, and fish oils, mineral oils, plant derived oils and additives in an appropriate ratio by means of dissolving, Such as canola oil, cotton-seed oil, soybean oil and sesame oil separating, Suspending, mixing, impregnating, adsorbing or as well as essential oils such as lavender oil, eucalyptus oil, precipitating operation to formulate into oil formulations, tea tree oil, citrus oil etc. Solid or liquid carriers can be used emulsifiable concentrates, wettable powders, flowables, alone or in combination. Examples of gas carriers, i.e., those granules, powders, dusts, Solutions, Suspensions, emulsions, of propellants, are butane gas, LPG (liquefied petroleum gas), controlled-release forms such as microcapsules, aerosols or dimethyl ether, fluorocarbons and carbon dioxide gas. fumigants. Typically, the compounds of the present invention 0093 Examples of surfactants are alkylsulfuric acid can be mixed with a solid carrier, liquid carrier orgas carrier, esters, alkylsulfonic acid salts, alkylarylsulfonic acid salts, optionally together with a Surfactant and other adjuvants use alkyl aryl ethers and polyoxyethylene adducts thereof, poly ful for such formulations. ethylene glycol ethers, polyhydric alcohol esters, Sugar alco 0088. The compounds of the invention can be used in an hol derivatives, sorbitane monolaurate, alkylallyl sorbitane amount from about 0.00005% to about 90% by weight as monolaurate, alkylbenzene Sulfonate, alkylnaphthalene Sul contained in these formulations as their active component. As fonate, lignin Sulfonate, and Sulfuric acid ester salts of higher used herein, the term “about” refers to a quantity, level, value alcohols. These surfactants may be used alone or in combi or amount that varies by as much as 30%, preferably by as nation. much as 20%, and more preferably by as much as 10% to a 0094. Examples of adjuvants for the formulations, such as reference quantity, level, value or amount. binders and dispersants, are casein, gelatin, polysaccharides I0089. Where the compounds are in the form of B-dione Such as starch, gum arabic, cellulose derivatives and alginic containing extracts, the formulations will usually comprise as acid, lignin derivatives, bentonite, Sugars and water-soluble their principal active ingredient from about 0.0001% to about synthetic high-molecular-weight Substances such as polyvi 90%, preferably from about 0.0001% to about 50%, more nyl alcohol, polyvinyl pyrrolidone and polyacrylic acids. preferably from about 0.0005% to about 10%, even more Examples of stabilisers are PAP (acid isopropyl phosphate), preferably from about 0.0005% to about 5%, even more pref BHT (2,6-di-tert-butyl-4-methylphenol), BHA (mixture of erably from about 0.0005% to about 1% and still even more 2-tert-butyl-4-methoxyphenol and 3-tert-butyl-4-methox preferably from about 0.001% to about 0.5% by weight of the yphenol), Synergists such as piperonyl butoxide, vegetable eXtract. oils, mineral oils, fish oils, Surfactants and fatty acids or esters 0090 Alternatively, where the compounds are in the form thereof. of substantially purified preparation of B-diones, the formu 0.095 Emulsifying agents that may be used are suitably lations will usually comprise as their principal active ingre one or more of those selected from non-ionic or anionic dient from about 0.00005% to about 90%, preferably from emulsifying agents. Examples of non-ionic emulsifying about 0.0001% to about 50%, more preferably from about agents include, but are not restricted to, polyoxyethylenealky 0.0005% to about 10%, even more preferably from about lphenylether, polyoxyethylenealkylether, polyethyleneglycol 0.001% to about 5% and still even more preferably from fatty ester, sorbitan fatty ester, polyoxyethylene sorbitan fatty about 0.001% to about 1% by weight of the substantially ester, polyoxyethylenesorbitol fatty ester, polyoxyethylenep purified f-dione. olyoxypropylenealkylether. Examples of anionic emulsify 0091. By “substantially purified' is meant a compound ing agents include alkyl Sulphates, polyoxyethylenealky which has been separated from components that naturally lether Sulphates, SulfoSuccinates, taurine derivatives, accompany it. Typically, a compound is Substantially pure sarcosine derivatives, phosphoric esters, alkylbenzene when at least 60%, more preferably at least 75%, more pref Sulfonates and the like. A mixture consisting of polyoxyeth erably at least 90%, and most preferably at least 99% of the ylenestyrylphenylether and calcium allylbenzenesulfonate is total material (by volume, by wet or dry weight, or by mole preferred. These emulsifying agents may be used in an percent or mole fraction) in a sample is the compound of amount of 5 to 20 weight parts per 100 weight parts of the interest. Purity can be measured by any appropriate method, compositions of the present invention. e.g., by chromatography or HPLC analysis. 0096. Formulation thus obtained can be used solus or 0092. Examples of solid carriers useful in preparing the diluted, for example, with water or other diluent. The formu formulations are clays including kaolin clay, diatomite, lations can be used also as admixtures with other pesticides water-containing synthetic silicon oxide, bentonite, Such as insecticides, arachnids, anti-helminthics, mollusci Fubasami clay, and acid clay; talcs; ceramics; inorganic min cides, herbicides, plant growth regulators, Synergists, soil erals such as Celite, quartz, Sulfur, active carbon, calcium improvers, baits and the like, or can be used simultaneously carbonate and hydrated silica; and chemical fertilisers such as with Such agents without mixing. For example, the pest ammonium Sulfate, ammonium phosphate, ammonium controlling compounds of the invention can be combined nitrate, urea and ammonium chloride, these solid carriers with other naturally derived bioactive compounds or extracts being finely divided or granular. Examples of useful liquid Such as neem or its components, derris, pyrethrum; microbial US 2010/0227009 A1 Sep. 9, 2010 extracts such as avermectins or streptomycins; with synthetic 0104. The compositions and methods of the present inven insecticides, acaricides, molluscicides, anti-helminthics; tion may be applied to pests including, but not restricted to, anti-protozoals, antivirals or with microorganisms having insects, arachnids, helminths, molluscs, protozoa and viruses. insecticidal, acaricidal, molluscicidal, anti-helminthic anti For example, suitable insects that fall within the scope of the protozoal or antiviral activity e.g., bacteria Such as Bacillus present invention include those: thuringiensis, Bacillus popillae, entomogenous fungi such as 0105 (a) from the order of the lepidopterans (Lepi Metarhizium spp., Verticillium lecanii, nematodes such as doptera), for example, Adoxophyes Orana, Agrotis ypsilon, Steinernema spp and Heterorhabditis. Alternatively, or in Agrotis segetum, Alabama argillacea, Anticarsia gemmata addition, the pest-controlling compounds of the invention can lis, Argyresthia conjugella, Autographa gamma, Cacoecia be combined with synergists such as piperonylbutoxide, and murinana, Capua reticulana, Choristoneura filmiferana, with ultraviolet screening compounds of natural or synthetic Chilo partellus, Choristoneura Occidentalis, Cirphis uni origin. puncta, Cnaphalocrocis medinalis, Crocidolomia binotalis, 0097. When used as an agricultural pesticide, the com Cydia pomonella, Dendrolimus pini, Diaphania initidalis, pound of the invention is preferably applied usually in an Diatraea grandiosella, Earias insulana, Elasmopalpus amount of 0.01 to 500 g/100 m. When an emulsifiable con lignosellus, Eupoecilia ambiguella, Feltia subterranea, Gra centrate, wettable powder or flowables are used as diluted pholitha funebrana, Grapholitha molesta, Heliothis armig with water, the compound is applied usually at a concentra era, Heliothis virescens, Heliothis zea, Hellula undalis, tion of 0.1 to 1000 ppm, preferably 1 to 500 ppm. The granu Hibernia defoliaria, Hyphantria cunea, Hyponomeuta lar or dust can be applied without dilution. malinellus, Keiferia lycopersicella, Lambdina fiscellaria, Laphygma exigua, Leucoptera scitella, Lithocolletis blan 0098. The amount or concentration of application, cardella, Lobesia botrana, Loxostege Sticticalis, Lymantria although exemplified above, can be suitably increased or dispar; Lymantria monacha, Lyonetia clerkella, Mandu.ca reduced according to the type of preparation, time, place, sexta, Malacosoma neustria, Mamestra brassicae, Mocis method of application, kind of pest and extent of harm or repanda, Operophthera brumata, Orgvia pseudotsugata, annoyance Suffered. Ostrinia nubilalis, Pandemis heparana, Panolis flammea, 0099. The invention also contemplates the use of the above Pectinophora gossypiella, Phthorimaea operculella, Phyl described B-dione compounds in pest repellent, particularly locnistis citrella, Pieris brassicae, Plathypena scabra, insect repellent, compositions. Repellent compositions con Platynota Stultana, Plutella xylostella, Prays citri, Prays templated by the present invention include those that are Oleae, Prodenia Sunia, Prodenia Ornithogalli, Pseudoplusia noxious to, and/or induce behavioural changes in, a pest. The includens, Rhyacionia frustrana, Scrobipalpula absoluta, latter compositions suitably comprise an activity including, Sesamia inferens, Sparganothis pilleriana, Spodoptera fru but not restricted to, an antifeedant activity, an oviposition giperda, Spodoptera littoralis, Spodoptera litura, Syllepta deterrent activity and an insect growth regulatory activity. derogata, Synanthedon myopaeformis, Thaumatopoea pitvo Insect repellent compositions in various dosage forms can be campa, Tortrix viridana, Trichoplusia ni, Tryporyza incertu prepared by blending the above-described faione com las, Zeiraphera Canadensis, pounds as active ingredients with a base of cosmetics or 0106 (b) furthermore Galleria mellonella and Sitotroga pharmaceuticals, which are usually applied to human bodies cerealella, Ephestia cautella, Tineola bisselliella, or animals. They can be formulated in, for example, lotions, 0107 (c) from the order of the beetles (Coleoptera), for aerosols, milky lotions, creams or the like. These compounds example, Anthonomus grandis, Anthonomus pomorum, can be further incorporated with other insect repellents, anti Apion vorax, Atomaria linearis, Blastophagus piniperda, oxidants, UV-absorbers, humectants or other additives. Cassida nebulosa, Cerotoma trifiurcata, Ceuthorhynchus 0100. The above compounds or the above-prepared com assimilis, Ceuthorhynchus mapi, Chaetocnema tibialis, Con positions of the present invention can be applied directly to oderus vespertinus, Crioceris asparagi, Dendroctonus human bodies or animals. Besides, Substrates, such as sheets, refipennis, Diabrotica longicornis, Diabrotica 12-punctata, films, nets, timber or the like, which have preliminarily been Diabrotica virgifera, Epilachna varivestis, Epitrix hirtipen treated with the above compounds or compositions by means nis, Eutinobothrus brasiliensis, Hylobius abietis, Hypera of application, impregnation or blending, can also be used. brunneipennis, Hypera postica, Ips typographus, Lema bilin 0101 The quantity of the above compounds to be formu eata, Lema melanopus, Leptinotarsa decemlineata, Limonius lated in the noxious-insect repellents depends upon the dos Californicus, Lissorhoptrus Oryzophilus, Melanotus commu age form, usage or other conditions. Suitable dosages may be nis, Melligethes aeneus, Melolontha hippocastani, selected from about 0.1% to about 90% by weight. Melolontha melolontha, Oulema oryzae, Ortiorrhynchus sul 0102 Thus, in another aspect of the present invention catus, Otiorrhynchus ovatus, Phaedon cochleariae, Phyllop there is provided a method for controlling harmful, annoying ertha horticola, Phyllophaga sp., Phyllotreta chrysocephala, or undesired pests, said method comprising exposing said Phyllotreta memorum, Phyllotreta striolata, Popillia pests to a pest-controlling effective amount of a composition japonica, Psylliodes mapi, Scolytus intricatus, Sitona linea comprising a B-dione compound as broadly described above. titS, Preferred embodiments of this type include exposing said 0.108 (d) furthermore Bruchus rufimanus, Bruchus pests to a pesticidally effective amount or a pest-repelling pisorum, Bruchus lentis, Sitophilus granaria, Lasioderma effective amount of said composition. serricorne, Oryzaephilus surinamensis, Rhyzopertha 0103) The terms “comprise”, “comprises” and “compris dominica, Sitophilus Oryzae, Tribolium castaneum, Trogo ing” and the like refer, unless the context requires otherwise, derma granarium, Zabrotes subfasciatus, to the inclusion of a stated step or element or group of steps or 0109 (e) from the order of the dipterans (Diptera), for elements but not the exclusion of any other step or element or example, Anastrepha ludens, Ceratitis capitata, Contarinia group of steps or elements. Sorghicola, Dacus cucurbitae, Dacus oleae, Dasineura bras US 2010/0227009 A1 Sep. 9, 2010
sicae, Delia coarctata, Delia radicum, Hydrellia griseola, 0118 (n) from the order of the phthirapterans (Ph Hvlemyia platura, Liriomyza sativae, Liriomyza trifolii, thiraptera), for example, Mallophaga, Such as Damalina spp., Mayetiola destructor, Orseolia Oryzae, Oscinella frit, Pego and Anoplura Such as Linognathus and Haematopinus spp.; mya hyoscyami, Phorbia antiqua, Phorbia brassicae, Phor 0119 (o) from the order of the hemipterans (Hemiptera), bia coarctata, Rhagoletis cerasi, Rhagoletis pomonella, for example, Aphis, Bemisia, Phorodon, Aeneolamia, Empoasca, Parkinsiella, Pyrilla, Aonidiella, Coccus, 0110 (f) furthermore Aedes aegypti, Aedes vexans, Pseudococcus, Helopeltis, Lygus, Dysdercus, Oxycarenus, Anopheles maculipennis, Chrysomya bezziana, Chrysomya Nezara, Aleyrodes, Triatoma, Psylla, Myzus, Megoura, Phyl hominivorax, Chrysomya macellaria, Cordylobia anthro loxera, Adelges, Nilaparvata, Nephotettix or Cimwx spp.; pophaga, Culex pipiens, Fannia canicularis, Gasterophilus I0120 (p) from the order of the siphonapterans (Sipho intestinalis, Glossina morsitans, Haematobia irritans, Hap naptera), for example, Ctenocephalides or Pulex spp.; lodiplosis equestris, Hypoderma lineata, Lucilia caprina, I0121 (q) from the order of the thysanurans (Thysanura), Lucilia cuprina, Lucilia sericata, Musca domestica, Muscina for example, Lepisma spp.; stabulans, Oestrus ovis, Tabanus bovinus, Simulium damno 0.122 (r) from the order of the dermapterans (Der Stin, maptera), for example, Forficula spp.; and 0111 (g) from the order of the thrips (Thysanoptera), for I0123 (s) from the order of the psocopterans (Psocoptera), example, Frankliniella fisca, Frankliniella Occidentalis, for example, Peripsocus spp. Frankliniella tritici, Haplothrips tritici, Heliothrips haemor 0.124 Arachnids contemplated by the present invention rhoidalis, Scirtothrips citri, Thrips oryzae, Thrips palmi, include, but are not limited to, Spiders and Scorpions and Thrips tabaci, especially mites Such as phytophagous mites (Acari), such as 0112 (h) from the order of the hymenopterans (Hy Aculops lycopersicae, Aculops pelekassi, Aculus Schlechten menoptera), for example, Athalia rosae, Atta cephalotes, Atta dali, Brevipalpus phoenicis, Bryobia praetiosa, Eotetrany sexdens, Atta texana, Hoplocampa minuta, Hoplocampa chus carpini, Eutetranychus banksii, Eriophyes Sheldoni, testudinea, Iridomyrmes humilis, Iridomyrmex purpureus, Oligonychus pratensis, Panonychus ulmi, Panonychus citri, Monomorium pharaonis, Solenopsis geminata, Solenopsis Phyllocoptruta oleivora, Polyphagotarisonemus latus, Tar invicta, Solenopsis richteri, Technomyrmex albipes, sonemus pallidus, Tetranychus cinnabarinus, Tetranychus kanzawai, Tetranchus pacificus, Tetranychus urticae, ticks, 0113 (i) from the order of the heteropterans (Het Such as Amblyomma americanum, Amblyomma variegatum, eroptera), for example, Acrosternum hilare, Blissus leucop Argas persicus, Boophilus annulatus, Boophilus decoloratus, terus, Cyrtopeltis notatus, Dysdercus cingulatus, Dysdercus Boophilus microplus, Dermacentor Silvarum, Hvalomma intermedius, Eurygaster integriceps, Euschistus impictiven truncatum, Ixodes ricinus, Ixodes rubicundus, Ornithodorus tris, Leptoglossus phyllopus, Lygus hesperus, Lygus lin moubata, Otobius megnini, Rhipicephalus appendiculatus eolaris, Lygus pratensis, Nezara viridula, Piesma quadrata, and Rhipicephalus evertsi, and animal-parasitic mites, such Solubea insularis, Thyanta perditor, as Dermanyssus gallinae, Psoroptes Ovis and Sarcoptes sca 0114 () from the order of the homopterans (Homoptera), biei. for example, Acyrthosiphon Onobrychis, Acyrthosiphon 0.125 Helminths falling within the scope of the present pisum, Adelges laricis, Aonidiella aurantii, Aphidula mastur invention may be selected from cestodes such as fish tape tii, Aphis fabae, Aphis gossypii, Aphis pomi, Aulacorthum worm, pork tapeworm, beef tapeworm, and dwarf tapeworm; Solani, Bemisia tabaci, Brachycaudus Cardui, Brevicoryne trematodes Such as from the genera Metagonimus and Het brassicae, Dalbulus maidis, Dreyfusia nordmannianae, erophyes; and nematodes Such as but not limited to filariid, Dreyfusia piceae, Dysaphis radicola, Empoasca fabae, Erio ascarid, strongyle and trichoStrongyle nematodes of the gen Soma lanigerum, Laodelphax striatella, Macrosiphum ave era Acanthocheilonema, Aelurostrongylus, Ancylostoma, nae, Macrosiphum euphorbiae, Macrosiphon rosae, Angiostrongylus, Ascaris, Brugia, Bunostomum, Dictyocau Megoura viciae, Metopolophium dirhodium, Myzus persicae, lus, Dioctophyme, Dipetalonema, Dirofilaria, Dracunculus, Myzus cerasi, Nephotettix cincticeps, Nilaparvata lugens, Filaroides, Lagochilascaris, Loa, Mansonella, Muellerius, Perkinsiella saccharicida, Phorodon humuli, Psylla mali, Necator. Onchocerca, Parafilaria, Parascaris, Protostrongy Psylla piri, Psylla pyricola, Rhopalosiphum maidis, Schiza lus, Setaria, Stephanofilaria, Strongyloides, Strongylus, phis graminum, Sitobion avenae, Sogatella furcifera, Tox Thelazia, Toxascaris, Toxocara, Trichinella, Uncinaria and optera citricida, Trialeurodes abutilonea, Trialeurodes vapo Wuchereria. rariorum, Viteus vitifolii; 0.126 Suitable molluscs include those of the Gastropoda 0115 (k) from the order of the termites (Isoptera), for class examples of which include Snails, slugs, conchs, and example, Calotermes flavicollis, Coptotermes spp., Leucote whelks. rines flavipes, Macrotermes subhyalinus, Nasutitermes spp I0127 Protozoa may be selected for example from Plasmo Such as Nasutitermes walkeri, Odontotermes formosanus, dia, Toxoplasma, Leishmania, Trypanosoma, Giardia, Enta Reticulitermes lucifigus, Termes natalensis, moeba, Acanthamoeba, Nagleria, Hartmanella, Balan 0116 (1) from the order of the orthopterans (Orthoptera), tidium, Babesia, Cryptosporidium, ISOspora, for example, Gryllotalpa gryllotalpa, Locusta migratoria, Microsporidium, Trichomonas or Pneumocystis species. Melanoplus bivittatus, Melanoplus femur-rubrum, Melano I0128 Viruses may be selected from RNA viruses or DNA plus mexicanus, Melanoplus sanguinipes, Melanoplus spre viruses, which include but are not limited to Human Immu tus, Nomadacris Septemfasciata, Schistocerca americana, nodeficiency Virus (HIV), Poliovirus, Influenza virus, Rous Schistocerca peregrina, Stauronotus maroccanus, Schisto Sarcoma virus, Flaviviruses Such as Japanese encephalitis, Cerca gregaria, Influenza virus, Respiratory Syncytial Virus, Hepatitis virus, 0117 (m) furthermore Acheta domestica, Blatta Orienta Parvovirus, Rotavirus, Coronavirus, Adenovirus and Herpes lis, Blattella germanica, Periplaneta americana, viruses such as Papillomavirus and Epstein-Barr virus. US 2010/0227009 A1 Sep. 9, 2010
0129. The present invention also extends to methods for percentage of the characteristics of the Susceptible taxon are producing resistance in plants to pests including, but not present in the progeny along with the gene or genes imparting limited to, insects, arachnids, helminths, molluscs, protozoa 3-dione activity. and viruses by crossing a plant expressing a B-dione com I0133. By the term “taxon herein is meant a unit of botani pound according to the invention with pest Susceptible lines. cal classification. It thus includes, genus, species, cultivars, Crossing a B-dione-producing plant into a pest Susceptible varieties, variants and other minor taxonomic groups which background would produce a resistant plant with a high level lack a consistent nomenclature. of pest resistance. Plants that could be made pest resistant I0134. In order that the invention may be readily under include, but are not limited to, dicotyledonous plants, espe stood and put into practical effect, particular preferred cially trees and more especially members of the Myrtaceae embodiments will now be described by way of the following family. For example E. cloeziana commonly known as Gym non-limiting examples. pie Messmate is one of the many Eucalyptus species grown EXAMPLES for hard wood production. However the oil present in this chemotype does not contain B-diones and hence an intra Example 1 species cross with the unique North Queensland tasmanone B-Triketone-Containing Oils Obtained from Austra chemotype would introduce this phenotypic trait. Such a pro lian Myrtaceae Species cess would be readily applicable to other Eucalyptus species of commercial interest. Interspecific crossing within the 0.135 Australia has an extensive number of volatile oils Myrtaceae family is well established to those skilled in the art from species of the Myrtaceae that are rich in a diversity of and inclusion of B-dione as an additional trait into formal structurally related constituents known as B-triketones. These breeding programs is acknowledged. oils often show not only a high yield of oil, but also a high degree of biosynthetic selectivity that produces B-triketones 0130 Suitable B-dione-producing plants may be selected in a high proportion of the oil composition. The major con from the families Alliaceae, Apiaceae, Asteraceae, Cannabi stituents of the published Myrtaceae essential oils (Hellyer, naceae, Lamiaceae, Pteridaceae, Myrtaceae, Myoporaceae, 1968; Boland and Brophy, 1990, 1993: Brophy, et al., 1995; Proteaceae, Rutaceae and Zingiberaceae. Preferably, the Bignallet al., 1997: Southwell and Brophy 2000) are listed in Volatile oil-bearing plant is selected from genera of the Table 1 and their structures are included in FIG. 1. Myrtaceae family including, but not limited to, Angophora, Austromyrtus, Backhousia, Baeckea, Callistemon, Corym TABLE 1 bia, Darwinia, Eucalyptus, Kunzea, Leptospermum, Mela leuca, Syzygium and Xanthostemon. Preferred B-dione-pro 3-Triketone Profiles of Australian Essential Oils ducing plants are Leptospermum morrisonii, Eucalyptus Species Yield B-Triketone (%) Distribn bensonii, Eucalyptus megacornuta, Eucalyptus pilularis, Backhousia anglisiifolia 2.5 Angustione (85) QLD Eucalyptus cornuta, Eucalyptus baxteri, Eucalyptus macro Backhousia anglisiifolia 2.5 Dehyroangustione (80) QLD rhyncha, Eucalyptus cloeziana, Melaleuca cajuputi, Euca Eucalyptus Cloeziana 3.0 Tasmanone (95) QLD lyptus jensenii, Backhousia angustifolia and Leptospermum Eucalyptus Suberea 1.4 Tasmanone (94) WA scoparium. A particularly preferred B-dione-producing plant Eucalyptus lateritica 0.9 Tasmanone (37), WA lateriticone (14) is Eucalyptus cloeziana. Eucalyptus Canfieldii Tasmanone (40) NSW Leptospermain scoparium 0.4 Leptospermone (19), NSW, NZ 0131. As used herein, the term “plant includes reference Flavesone (8), to whole plants, plant organs (e.g., leaves, stems, roots, etc.), Isoleptospermone (5) seeds and plant cells and progeny of same. Plant cell, as used Eucalyptus grandis 0.6 Leptospermone (20), NSW Flavesone (13), herein includes, without limitation, seeds Suspension cul Isoleptospermone (3) tures, embryos, meristematic regions, callus tissue, leaves, Eucalyptus agglomerata Agglomerone (40) NSW roots, shoots, gametophytes, sporophytes, pollen, and Eucalyptus mckieana Agglomerone (60) NSW microspores. The class of plants which can be used in the Eucalyptus bensonii 2.5 Agglomerone (72) Eucalyptus insularis 1 Agglomerone (19) WA methods of the invention is generally as broad as the class of Eucalyptusiensenii 0.3 Jensenone (70) NT higher plants amenable to transformation techniques, includ Eucalyptus paptiana 0.7 Papuanone (40) Nth Aus ing both monocotyledonous and dicotyledonous plants. Leptospermain morrisonii 1.8 Grandiflorone (58) NSW 0132) Thus, the present invention contemplates conven Melaletica Caiepitti Platyphyllol Nth Aus Xanthostenon Xanthostemone tional plant breeding methods to transfer the genetic material oppositifolius associated with B-dione production via crossing and back crossing. Such methods will comprise the steps of: (1) sexu ally crossing the B-dione-producing plant with a plant from a 0.136 The B-triketones obtained from selected Myrta pest Susceptible taxon; (2) recovering reproductive material ceous Volatile oils were shown to have significant insecticidal from the progeny of the cross; and (3) growing B-dione and/or acaricidal activity. producing/pest-resistant plants from the reproductive mate rial. Where desirable or necessary, the agronomic character Example 2 istics of the susceptible taxon can be substantially preserved by expanding this method to include the further steps of Insecticidal Activity repetitively: (1) backcrossing the pest-resistant progeny with 0.137 Initial insecticidal screening against two important pest-Susceptible plants from the Susceptible taxon; and (2) arthropod species, two-spotted mite (Tetranychus urticae) selecting for expression of a B-dione (or an associated marker and diamond back moth (Plutella xylostella) lavae high gene) among the progeny of the backcross, until the desired lighted three oils on the basis of efficacy, oil yield, oil profile US 2010/0227009 A1 Sep. 9, 2010
and ease of recollection (Table 2). Where feasible, the LDso 0143. The most promising oils were derived from B. and LDos values were determined. angustifolia, M. Cajuputi Subsp platyphylla and E. cloeziana and these were then subjected to additional chemical fraction TABLE 2 ation. The lower insecticidal activity observed in the recol Percentage Mortality of Three Efficacious Oils lected B. angustifolia (IV) was in part attributed to the lower levels of B-triketone. The level off-triketone was elevated by Two Spotted Mite Diamond Back Moth % Mortality % Mortality the removal of the more Volatile monoterpenes using vacuum Species (0.5%/1.0%) (0.5%/1.0%) distillation. Backhousia anglisiifolia 98.98 100,100 0.144 One of the most efficacious oils was from E. cloe Backhousia anglisiifolia 100,100 100,100 ziana, a tasmanone (84-96%) rich oil with additional terpenes Eucalyptus cloeziana 100,100 70,100 and B-triketones (FIG.2, Table 4), which displayed consistent activity at every stage of processing and formulation. This oil 0.138. Insecticidal tests using the oil from fresh plant rec was, therefore, fractionated using column chromatography ollections and steam distillations varied occasionally and, in on silica gel with a hexane-diethyl ether gradient and a final this regard, it is believed that improving storage conditions methanol elution. The profiles of the fractions are sum including temperature, light and exposure to air and inclusion marised in Table 5. of a dessicant can enhance the stability of the active fraction of such oils. TABLE 4 0139 E. cloeziana oil continued to show high potency against both insect tests. This oil exhibited an LDs between Chemical Profile of E. cloeziana Oil 0.04-0.20% (depending on formulation and treatment) against two-spotted mite. An LDs of 0.10% was observed Peak No Compound Composition (%) against 1 instar lavae of diamondback moth and this rose to 1 a-pinene O5-19 2 B-pinene 1.5-5.7 0.78% when tested against 3" instar lavae. In additional pre 3 Limonene O.1-0.6 liminary investigations with greenhouse thrips (Heliothrips 4 a-terpineol O.7-2.O haemorrhoidalis), a 0.1% concentration of E. cloeziana oil 5 Globulol O.O1-05 induced 100% mortality. 6 Agglomerone O.O1-0.6 7 Tasmanone 84-96 0140. Additional work was carried out on the E. cloeziana 8 Lateriticone O.2-0.7 oil to explore the contribution the various components make 9 Isolateriticone O3-12 to the overall efficacy of this oil. Fractions 1, 3, 4, and 5 outlined in Example 3 were screened against two-spotted mite. Fractions 1 and 5 showed no significant insecticidal effect. Fractions 3 and 4, comprising 98% and 99% tas TABLE 5 manone were active and showed little difference to the activ ity of the whole oil. Fractions Cut From E. cloeziana Oil 0141. This suggests that not only is tasmanone the prin Fraction No Solvent System Composition Amount ciple component in the oil, but it is also the principle bioactive F1 Hexane Hydrocarbons 73 mg constituent. It is also reasonable to assume that as the activity F2 Hex:Et2O (9:1) 80% Tasmanone 4 mg of E. cloeziana oil has been demonstrated against a number of F3 Hex:Et2O (1:1) 98% Tasmanone 3.66 g different arthropod species, namely a mite (T. urticae), a F4 Hex:Et2O (9:1) +99% Tasmanone 694 mg caterpillar (P xylostella) and a thrips (H. haemorrhoidalis), F5 MeOH Terpene alcohols 64 mg its insecticidal activity is broad in nature. (0145 H NMR data (FIG. 3) were recorded on F4 and Example 3 confirmed the structure of tasmanone. Moreover the com Chemistry pound exists in solution (CDC1) in tautomeric forms (FIG.4) in the ratio 2:0:1 (A:B:C). 0142 Chemical analysis (GC-MS) of the steam-distilled 014.6 Another efficacious oils was from M. cajuputisubsp oils from the collected plants in this work are summarised in platyphylla, a platyphyllol (64-71%) rich oil (Table 6), which Table 3. also displayed consistent activity at every stage of processing and formulation. TABLE 3 3-Triketone Profiles of Selected Oils TABLE 6 Plant Source Principle Component (%) Typical Chemical Profile of M. Caiupui subsp platyphylla Oil Backhousia anglisiifolia -I Dehydroangustione 85% Peak No Compound Composition (%) Backhousia anglisiifolia -II Dehydroangustione (80%) Backhousia anglisiifolia -III Angustinone (65%) 1 a-pinene Tr-O.8 Backhousia angustifolia -IV Angustinone (28%) 2 1,8-cineole Tr-O.7 Eucalyptus cloeziana Tasmanone (84-96%) 3 B-caryophyllene O6-32 Melaletica Cajuputti Subsp platyphylia Platyphyllol (64-71%) 4 Humulene O.6-1.3 5 Spathulenol 4.0-9.0 As determined by gas chromatography 6 Caryophyllene oxide Tr-36 US 2010/0227009 A1 Sep. 9, 2010
0151. There were no other clinical abnormalities in any TABLE 6-continued animal throughout the seven (7) day observation period. The stomach of one animal in the 250 mg/kg group (5F) had a Typical Chemical Profile of M. Caiuputti subsp platyphylla Oil single ulcer. There were no other gross abnormalities in the Peak No Compound Composition (%) major organs of any animal at autopsy. 6 Platyphyllol 64-71 Based on the results obtained from this study 50, 100 and 200 7 MW 234 - unknown 4.3 mg/kg can be selected for a main study as the maximum * As determined by gas chromatography non-toxic dose, intermediate dose and high dose, respec tively. Example 4 Full Study Phytotoxicity 0152 The acute oral toxicity of E. cloeziana oil was inves 0147 Initial investigations using leaves and leaf discs of tigated in thirty (30) Sprague Dawley Specific Pathogen Fee several plant species including French bean (Phaseolus vul rats (15 males and 15 females) at doses of 50, 100 and 200 garis) lemon (Citrus limon) Orange (Citrus sinensis) and mg/kg. These doses were chosen following a dose range Cabbage (Brassica oleracea) indicated that phytotoxicity did finding in the above study. The experimental procedure was not occur for most oils below 0.5%. More detailed investiga based on OECD guidelines for the testing of chemicals, No. tions using Soft intact leaves of young greenhouse-grown 420. French bean Subsequently showed E. cloeziana oil applied as a spray caused some phytotoxicity at concentrations of 0.5% 0153. The acute NOAEL of E. cloeziana oil was deter and above. mined to be 50 mg/kg, and the MTD was 200 mg/kg under the conditions of this study. Example 5 Acute Dermal Toxicity of E. cloeziana Oil 0154 The acute dermal toxicity of E. cloeziana oil was Bacterial Reverse Mutation Assay investigated in ten (10) Sprague Dawley Specific Pathogen Free rats (5 males and 5 females) at a dose of 2000 mg/kg. A 0148. This study investigated the potential of E. cloeziana preliminary study (SIGHTING) indicated no signs of toxicity oil to induce reverse mutations at the histidine locus in the at this dose. The experimental procedure was based on OECD genome of one strain of Salmonella typhimurium TA100 in guidelines for the testing of chemicals, No. 420. the presence and absence of a metabolic activation system (mammalian microsomal enzymes, S9 mix). The test sample 0.155. No clinical abnormalities, skin irritations or body was dissolved in dimethyl sulfoxide (DMSO). In this assay, weight losses were observed in any animal throughout the an E. cloeziana oil test sample did not induce an appropriate fourteen (14) day observation period. No deaths occurred. No fold increase (a 2-fold increase for TA100) in the mean rever abnormalities were seen in the major organs at necropsy. The tants per plate in the tester strain TA100 over the mean rever rat acute dermal LD50 of E. cloeziana oil was determined to tants perplate of the appropriate vehicle control. Accordingly, be greater than 2000 mg/kg under the conditions of this study. the test sample was considered to be non-mutagenic under the conditions of the assay. Skin Irritancy/Corrosion 0156 The potential of a test sample of E. cloeziana oil to Acute Oral provoke skin irritation/corrosion reactions was investigated Sighting using a primary dermal irritation/corrosion test in three (3) New Zealand White albino rabbits (OECD Guidelines for the 014.9 The acute oral toxicity of E. cloeziana oil was inves tigated in ten (10) Sprague Dawley Specific Pathogen Free Testing of Chemicals, No. 404). The results obtained from female rats (groups of 2) at doses of 500, 250, 125 and 50 this study indicated that E. cloeziana oil is a non-irritant mg/kg. The experimental procedure was based on OECD according to the National Occupational Health and Safety guidelines for the testing of chemicals, No. 420. Commission (NOHSC) Approved Criteria for Classifying 0150 Clinical signs of toxicity occurred between one (1) Hazardous Substances (NOHSC:1008 (1999). and twenty-four (24) hours after dosing. Both animals in the 500 mg/kg group exhibited subdued behaviour, partial eye Example 6 closure, slow breathing, reduced motor activity, ataxia fol lowed by death within 24 hours. The animals in the 250 mg/kg Isolation and Purification of B-Triketones from Aus group exhibited subdued behaviour, partial eye closure, slow tralian Myrtaceae Species breathing, social isolation, and reduced motor activity, and had returned to normal by 24 hours after dosing. The animals O157 Plant collections were commissioned at the Mt. in the 125 mg/kg group exhibited subdued behaviour, partial Annan Botanic Garden, Sydney, Australia, and the Darwin eye closure, slow breathing, piloerection and reduced motor Botanic Garden, Northern Territory, Australia to provide activity, and had returned to normal by 24 hours after dosing. potential sources of a range of B-triketone isolation. The The animals in the 50 mg/kg group did not show any signs of plants investigated for B-triketone exploration are listed in toxicity during the seven day experimental period. Table 6. US 2010/0227009 A1 Sep. 9, 2010 17
TABLE 6 Plants Sourced for B-Triketone Exploration B-triketone # Source Plant Plant Location Accession # anticipated 1 Leptospermain morrisonii Mt. Annan, NSW 873247 grandiflorone 2 Eucalyptus benisonii Mt. Annan, NSW 88.1045 agglomerone 3 Eucalyptus megacornia Mt. Annan, NSW 831078 jensenone 4 Eucalyptus pilularis Mt. Annan, NSW 873166 torquatone 5 Eucalyptus cornuta Mt. Annan, NSW 852618 jensenone 6 Eucalyptus baxteri Mt. Annan, NSW 860999 agglomerone taSmalOne 7 Eucalyptus macrorhyncha Mt. Annan, NSW 860896 conglomerone 8 Eucalyptus cloeziana Lappa, QLD PF 2513 taSmalOne 9. Melaletica Cajuputti SSp Bensbach, WP, PNG KW16-19 platyphyllol platyphyla 10 Eucalyptusiensenii NT Botanic Gardens, RK114 jensenone Darwin, NT LUS 11 Backhousia anglisiifolia Wilgavale, Texas, PF1712 dehydroangustione QLD 12 Backhousia anglisiifolia Didcott Creek, QLD PF 1708 angustione 13 Leptospermin Scoparium NA commercial oil NA flavesone, sample isoleptospermone, leptospermone 14 Eucalyptus conjuncta Mt Annan 854097 conglomerone "Oil contains other B-triketone constituents
0158 All plants, apart from 9 and 13, were steam distilled to yield various quantities of essential oil. All oils were analy TABLE 7-continued sed by Gas Chromatography Mass Spectrometry (GCMS) to determine the presence and abundance off-triketones. Based 3-Triketones Isolated for Insecticidal Screening on this information, particular oils were targeted for isolation Amount sent and purification off-triketones using wet chemistry and pre Sample Purity % by for screening parative HPLC techniques. B-triketones Plant source GCMS (mg) leptospermone Leptospermain 95.33 77.0 0159. The B-triketones listed in Table 7 were isolated in Scoparitim quantities adequate for insecticidal screening. A minimum of taSmanone Eucalyptus 99.92 110.1 50 mg of each compound was required. Isoleptospermone cioeziana platyphyllol Meiaieuca 99.62 255.6 and leptospermone were difficult to separate due to their cajuputti Subsp. structural similarity as were angustione and dehydroangus platyphylia tione. Consequently, mixtures of these compounds, where one isomer was significantly more abundant, were provided 0160 The chemical structures and identities of the B-trike for insecticidal screening as this will still allow for differen tones isolated were confirmed by GCMS and Nuclear Mag tiation in observed activity. netic Resonance (NMR) analysis. TABLE 7 Example 7 3-Triketones Isolated for Insecticidal Screening Efficacy of E. cloeziana Oil on Target Organisms Amount sent Sample Purity % by for screening Target Organisms B-triketones Plant source GCMS (mg) grandiflorone Leptospermain 100.00 99.5 0.161 Two spotted mite (TSM) Tetranychus urticae Koch morrisonii Acarina: Tetranychidae were collected from a mass culture Jensenone Eucalyptus 100.00 67.9 maintained at the University of Western Sydney's Hawkes dehydroangustione fensentiBackhousia 95.02 16S.O2 bury Campus in Richmond, NSW, Australia. They Were angustifolia reared on potted French beans (Phaseolus vulgaris L. Fa PF1712 bales: Fabaceae in a glasshouse maintained at 25+5° C. angustione Backhousia 66.30 (33.70% 107.1 65+5% RH and 14 h D:L photoperiod. Only young females angustifoliaPF1708 dehydroangustione) were selected for bioassay. agglomerone Eucalyptus 99.28 109.2 0162 Adult parthenogenetic female greenhouse thrips f bensonii (GHT), Heliothrips haemorrhoidalis Bouché (Thysanoptera: 8WCSOile tropi" 99.32 101.8 Thripidae) of similar age were obtained from a colony reared isoleptospermone Leptospermain 33.10 (66.90% 113.4 on Orange fruits and maintained in an insectary at UWS Scoparitim leptospermone) Hawkesbury under conditions of 25+3°C., 65% RH and 16h D:L photoperiod US 2010/0227009 A1 Sep. 9, 2010
0163 Young nymphs of citrus aphids Toxoptera citricida discs. Five mL aliquots (unless otherwise stated) were (Kirkaldy) (Hemiptera: Aphididae) were collected from applied to each petri dish with a Potter precision spray tower lemon seedlings grown under glasshouse conditions at UWS as described by Herron et al (1995). The average mass of Hawkesbury. solution applied to each dish was calculated to be 3.95 0164 Workers of the termite Nasutitermes walkeri Hill mg/cm. Mortality was recorded 24 h after treatment. Death (Isoptera: Termitidae) were collected from a laboratory cul was recognised by the absence of movement when the test ture at UWS Richmond which was initiated from soil, termi organisms were mechanically stimulated. Data were analy taria and wood on which termites were feeding were field sedusing SPSS for WindowsTMVersion 7. Probit analysis was collected at Richmond NSW, and maintained in a darkened carried out for dose-mortality data and heterogeneity of container under conditions of 25+2° C., 35-68% RH. Ter regressions was determined by the Pearson chi-squared char mites were fed on wood collected from near the original nest. acteristic. Moistened soil from the nest together with paper towel were 0.174. In addition, TSM was treated with E. cloeziana oil placed on top of the nest and made it possible to maintain this extract in combination with paraffin oil. In particular, E. cloe culture for several months in the laboratory at UWS Hawkes ziana oil extract at levels of 0.1, 0.2,0.3, 0.4 and 0.5 g were bury. weighed out and each was made up to a weight of 10.0 g with 01.65 Workers of the whitefooted house ant Techno a formulated paraffin oil (BioPest(R), which was then soni myrmex albipes (F. Smith) (Hymenoptera: Formicidae) were cated for 10 min. A 1.0% V/v of each blend was prepared by field collected at UWS Hawkesbury by baiting in an empty mixing 1.0 mL with distilled water in a 100.0 mL volumetric glass jar containing Sugar granules. flask. TSM were transferred to the petri dish following the 0166 Pupae of housefly, Musca domestica L (Diptera: same standard method. Five-mL aliquots were applied to Muscidae) and different stages of American Cockroach, each petri dish and mortality was recorded 24 h after treat Periplaneta americana L (Blattodea: Blattidae) were initially ment. All blends of E. cloeziana oil extract with paraffin oil supplied by C.E.R.I.T and maintained in laboratory culture at produced 100.0% mortality, compared with Biopest(R) alone UWS Hawkesbury. which caused only 35.5-6.9 mortality. The lowest concentra 0167 Tomato russet mites (TRM) Aculops lycopersici tion of E. cloeziana oil extract tested in this combination (i.e. (Massee) (Acarina: Eriophyidae) were collected from 0.01%) resulted in 100% mortality in TSM, which is signifi infested tomato plants near Riverstone, NSW. cantly lower than that reported earlier in this document to 0168 Mixed sex adults of the mosquito Culex quinquefa produce 100% mortality with E. cloeziana oil extract alone ciatus (Diptera: Culicidae) were supplied by C.E.R.I.T., held (>0.06%). at the in the Centre for Horticulture & Plant Science, Univer sity of Western Sydney, Richmond NSW and treated one day Greenhouse Thrips after arrival. (0175. The same experimental procedure used for TSM (0169 Workers of the honeybee Apis mellifera (Hy was repeated for GHT except that lemon leaf discs were used menoptera: Apidae) were collected from several field hives instead of French beans. The required number of adult thrips maintained at the apiary in the Centre for Horticulture & Plant for each treatment was transferred with a fine brush to the Sciences, University of Western Sydney, Richmond NSW underside of a lemon disc of the same diameter (6 cm) as the 0170 Adults of ash-white-flies Aleaurocanthus woglumi base of a petridish. The lemon disc was mounted on agar with (Homoptera: Aleyrodidae) were field collected from an orna its adaxial side uppermost. Immediately after treatment the mental pear (Prunus sp) in the Centre for Horticulture & Plant petridish was covered with perforated plastic wrap. Mortality Sciences, University of Western Sydney, Richmond NSW. was assessed 24 h after treatment. 0171 Drug store beetle Sitodrepa panicea (Coleoptera: Anobiidae) were reared on curry powder under laboratory Tomato Russet Mite conditions of 25-1° C. and 65.5% RH. 0176 The same experimental procedure for TSM was 0172 Mixed age groups of snails Helix apersa (Mollusca: repeated except that tomato leaf discs were used instead of Gastropoda) were collected from infested plants in the Centre French bean. for Horticulture & Plant Science, University of Western Syd ney, Richmond NSW. Brown Citrus Aphid Bioassays 0177 Lemon leaf discs 2.5 cm diam. were cut from tender young leaves and mounted on moistened absorbent cotton TSM wool in 90 mm petri dish with their adaxial surface upper 0173 From an E. cloeziana oil extract (containing 85% most. Each petridish contained four leaf discs. Uniform early tasmanone), 1.1765 g was dissolved in 5 mL ethyl alcohol and instar nymphs were then transferred with a fine brush to the distilled water containing 200 ppm Triton X-100TM was leaf discs (each containing 8-10 nymphs). A Potter tower was added to prepare a 1% stock solution. From this homogenised used to apply 5 mL aliquots to each petri dish. A control stock solution, further serial dilutions of 0.0125, 0.025, 0.05, (solvent and Surfactant only) was also included in the assess 0.10, and 0.12% were prepared by mixing the required ment. Mortality was assessed 24 h after treatment. amount of stock solution in distilled water and Triton X-100 solution. Each treatment was conducted on 60-80 TSM, Termites which were evenly distributed on four French bean leaf discs 0.178 Twenty uniform termite workers were transferred to (25 mm diam) contained in 90 mm diam. petridishes. The leaf 90 mm petri dishes lined with the same diameter moistened discs were placed with their underside uppermost on moist filter paper (Whatman No 2). A preliminary trial was carried absorbent cotton wool covered with muslin netting. Water out using a Potter tower to apply 5 mL aliquots of each was added to the dishes daily to prevent desiccation of the leaf concentration. Using this method, all termite workers died 4 US 2010/0227009 A1 Sep. 9, 2010
h after treatment in all concentrations, including the lowest acetone only (i.e., minus E. cloeziana oil extract). After com concentration of 0.015%. There was no mortality recorded in plete dryness of the filter paper, the required number of cock the blank control treatment, and all workers remained alive roaches was transferred inside the kilner jars, and were fed and active for >48 h after treatment. Identical results were dry dog food. Kilner jar necks were covered with muslin obtained from these investigations, whether the petri dishes netting supported by rubber bands. Mortality was assessed 24 were covered or uncovered after application of the E. cloezi h after releasing the cockroaches, and death was recognised ana oil extract. by the absence of movement when the test animals were 0179 Subsequent investigations further assessed efficacy mechanically stimulated. of E. cloeziana oil extract by releasing termite workers on fresh dried residues. This was carried out by uniformly dis Adult Mosquitoes tributing one mL of each concentration over the entire Surface 0184. A 0.3576 g of 85% ai of E. cloeziana oil extract was area of a 90 mm diam filter paper. When the paper was air dry, dissolve in pure acetone as a diluent to give approximately 20 termite workers were placed in each petri dish. One hun 0.304% concentration stock solution. From this solution, fur dred percent mortality was recorded in all four replicates even ther serial dilutions 0.152, 0.076, 0.0043 and 0.00215 were at the lowest concentration applied (0.015% w/v=150 ppm). prepared by adding the required amount in acetone. Aliquots This suggests that the plant extract is a highly toxic contact (2.5 mL) of each concentration of E. Cloeziana oil extract poison to termites. were dispensed into 500 mL kilner jars with total internal surface area as 286.53 cm. The kilnerjars were immediately Ants rotated to coat the inner surface uniformly with the E. cloe 0180. The same experimental procedure for termites was Ziana oil extract residue, until dry. Once completely dry, repeated for ants, with the required number of worker ants for 10-25 mixed sex adult mosquitoes were released into each each treatment transferred with a fine brush to the filter paper kilner jar by allowing the mosquitoes to fly from a darkened containing a fresh dried residue of the E. cloeziana oil extract. cage into the naturally lit jars. The jar mouth was Subse Immediately after release of the ants the petri dish was cov quently placed on 110 mm diam. filter paper onto which had ered with perforated plastic wrap, which enabled any excess been placed a yellow sponge soaked in 7.0% Sugar Solution. vapours to escape while retaining the ants. Mortality was Treated jars were kept under laboratory temperature and assessed 4 hafter treatment. In all 4 replicates 100% mortality humidity conditions, (viz. 24+1° C. and 65+5% RH respec was obtained at concentrations as low as 0.0075% w/v ai tively). Mortality was assessed 24 h after releasing the mos when applied at a rate of 1.0 mL to a 90 mm diam. filter paper. quitoes in the jars. Houseflies Honey Bees 0181. One percent E. cloeziana oil extract was prepared 0185. Ten worker honey bees, anaesthetised with carbon using pure acetone as a diluent. From this solution, further dioxide, were transferred to 90 mm diam. petri dishes lined serial dilutions were prepared by adding the required amount with moistened filter paper. Five mL aliquots were applied in acetone. Five mL aliquots of each concentration of E. using a Potter Spray Tower while the bees were still anaes cloeziana oil extract were dispensed into 500 mL kilner jars. thetised. Yellow sponges soaked in 7% sugar solution were The kilnerjars were immediately rotated until dryness to coat then placed inside petri dishes for bee Sustenance, and the the inner surface uniformly with the E. cloeziana oil residue. bees after treatment were placed in these dishes. The lids that After complete dryness, 30-50 pupae were transferred to a were perforated and covered with muslin netting were placed series of clean uncovered petri dishes (45 mm diam.), one of on the dishes, and mortality was assessed 24 hours after which was placed inside each jar. The jar mouth was then treatment. covered with nylon netting supported by a rubber band. All adulthouseflies started to emerge from pupae after 48 h and Ash White Flies most emerged within a 3 h period. Flies were fed 5% sugar 0186 For each treatment, a 2.5 cm diam. leaf disc was Solution soaked in absorbent cotton wool. Jars were main mounted on a moistened Whatman #2 filter paper lining the tained in an incubator at 29°C. bottom of petri dishes. Adult white flies were anaesthetised 0182 Mortality was assessed at the end of the third day using carbon dioxide and 30-50 adults were transferred onto (i.e., approx. 72 h) after application of the E. cloeziana oil leaf discs. Serial dilutions of 0.00546, 0.0220, 0.0894 & residues in the kilnerjar and the placement of pupae inside the 0.1788% concentration of E. cloeziana oil extract were pre jars. (This comprised 48 h for pupae to emerge and 24 h. pared using distilled water containing 200 ppm Triton X100. exposure to E. cloeziana oil residues which were now 48 h. Three replicates were treated for each concentration. Five old). Flies were observed to die within a few hours after mL aliquots were applied to each petri dish using a Potter emergence, whereas in the control they remained alive for Spray Tower. After treatment, petridishes were left to dry and >48 h after emergence. The total number of adult houseflies then covered with muslin netting. Mortality assessment was that emerged in each kilnerjar was counted and their mortal assessed 24 h after white flies had been transferred to petri ity was recorded. dishes. American Cockroaches Drug Store Beetle 0183 Tests were conducted on 10-20 three months old 0187. One mL of each concentration of E. cloeziana oil nymphs (meanindividual mass 0.2-0.3 g) and replicated three extract in pure acetone was uniformly dispensed on 90 mm times. One mL of 1.0% E. Cloeziana oil extract inacetone was diam. Whatman No 2 filter paper. The latter was left for 1 h to uniformly distributed on 90 mm diam. Whatman No 2 filter air dry before being used to line the lid of a 90 mm diam. petri paper. A control treatment was also carried out using 1 mL dish. Between 10-15 mixed adults were then transferred into US 2010/0227009 A1 Sep. 9, 2010 20 the petri dish, which was sealed with ParafilmTM. Mortality 0189 Allowing the snails to crawl on a E. cloeziana oil assessment was carried out 24 h after sealing the petri dish. extract contaminated Surface carried out the second method. Snails Two mL of 0.08% concentration of E. cloeziana oil extract in 0188 Two methods were used to assess the efficacy of E. pure acetone were dispensed in 500 mL kilner jars which cloeziana oil extract against Snails. In the first method differ were rotated to uniformly cover all inner surfaces until dry. ent age groups of adult snails were dipped directly into a Six different age groups of snails were put into each jar along solution of 0.5% concentration of E. Cloeziana oil extract in with plant material for food and covered with muslin netting distilled water containing 200 ppm Triton X 10. Snails were held by a rubber band. Three replicates of each treatment (0.0 dipped for 10 seconds and thereafter immediately filtered in a sieve. The treated snails were divided in three 500 mL kilner & 0.08% concentration) were carried out. Mortality was jars containing French bean leaves as a food source. The assessed 24 h after releasing the Snails inside the jars. control was carried out using 0%. E. cloeziana oil extract. 0190. The above bioassay results are summarised in Table Mortality was assessed 24 h after treatment. 8.
TABLE 8 Summary of E. cloeziana Oil Extract Efficacy Against Various Target Pests
Method of Target Pest LDso (95% CL) LDos (95% CL) application Remarks TSM O.O7 O.14 Potter spray Knockdown effect (0.06-0.08) (0.12–0.16) tower; Aliquot observed 2 h after applied 2.5 mL treatinent TSM <0.01 E. Cloeziana oil 100% mortality at extract + paraffin O.01% oil TRM O.O2 O.04 Potter spray OWe GHT O.10 O.12S Potter spray OW Aphid O.08 O.30 Potter spray OWe American 157.27 g/cm E. cloeziana oil cockroach residues on filter nymph paper Ant <0.0075 E. Cloeziana oil 100% mortality at residues on filter 0.0075% when 1 mL paper applied as residue on filter paper Termite <0.0075 E. Cloeziana oil 100% mortality at residues on filter 0.0075% when 1 mL paper applied as residue on filter paper DBM 1st O.09 O.20 5 mL Potter spray instar larva OWe Housefly 69.8 g/cm’ 130.87 g/cm E. cloeziana oil adult residues on kilner iar wall Adult O.OO387 O.OO694 Residues on kilner Apparently very mosquitoes (0.00328-0.00451) (0.00596-0.01051) jar wall rapid knock down. Ash white O.O1773 O.O.O3728 Potter spray Assessed 4h after fly (0.01179-0.03446) (0.02657-0.08337) tower, 2.5 mL treatinent aliquot Drug store O.O3S48 0.37275 Self Mortality assessed beetle contaminating by 24 h after releasing walking on filter adults. paper Honeybees O.12 O.40 Potter spray Knockdown effect tower observed 2 h after treatinent Aliquot applied 5 mL. Snails --O.O84 E. Cloeziana oil Three mL acetone residues on kilner solution for each of jar wall 3 replicates. Snails crawled on the wall on dried residues. Snails 100% mortality Dipping method Three replicates in 0.5% US 2010/0227009 A1 Sep. 9, 2010
Potted Plant Investigations aluminium foil. It was then placed under the lid of the petri 0191 The efficacy of E. cloeziana oil extract against TSM dishes containing the moist filter paper on their base. Twenty was further assessed under greenhouse conditions. French uniform worker termites were transferred to the moist base of bean plants were grown in 15 cm diam. plastic pots in an each test petri dish, which was subsequently covered with its insecticide-free glasshouse maintained at 27°C., RH 65% lid containing the treated filterpaper, and the dishes were then and natural light. Plants were used when they reached the two sealed with ParafilmTM. A similar series of control treatments true-leaf stage (i.e., before trifoliate leaves appeared). One was also prepared, using acetone only, for comparison. Five plant was maintained in each pot (i.e., two leaves/pot). replicates were used in each treatment. Twenty gravid TSM females were then transferred with a fine 0194 The termites did not move to the dry top surface and brush to the upper surface of each leaf. Mites were left to remained on the water-moistened filter paper lining the base settle for 4h before treatment, during which time they usually of the petri dishes throughout the experimental period. Mor settled on the lower leaf surfaces. E. cloeziana oil extractata tality was assessed 5 h after termite release. concentration of 0.07% as well as a blank control were pre pared using the same procedures described above for the 0.195. The results revealed that E. cloeziana oil extract has laboratory bioassays. A 400 mL hand sprayer was used to highly significant fumigant effects on termites. One hundred apply the pesticide evenly to all aerial Surfaces of the plants, percent mortality was recorded in all replicates of the E. to run-off. Each treatment was replicated four times. The cloeziana oil treatment whereas no mortality occurred in any mortality was assessed 24 h after treatment. The results were of the control (acetone only) replicates. recorded as mean percent mortality with Standard deviation. These results revealed that the mean percent mortality in E. Example 9 cloeziana oil extract treatment and control were 92.19+6.39 and 0.25+0.46, respectively. Efficacy of Purified B-Triketones Against TSM 0.192 In summary, E. cloeziana oil extract was efficacious against all pests tested namely TSM, TRM, GHT, aphids, 0196. The efficacy of purified f-triketones against TSM termites, houseflies, American cockroaches, whitefooted ants was investigated using the TSM bioassay described in adult mosquitoes, ash whitefly, drug store beetle and Snails. It Example 8. All B-triketones tested demonstrated a high level was also toxic to honey bees. of activity against TSM (see Table 9).
TABLE 9 Efficacy of B-Triketones Against TSM Sample wt No. (mg) Chemical Name LDso and 95% CL LDos and 95% CL 109.2 Agglomerone 0.15 (0.11-021) 0.33 (0.22-1.06) 99.5 Grandiflorone O.04 O.13 165.0 Dehydroangustione 0.36 (0.33-0.41) 0.69 (0.61-0.81) 107.1 Angustione 0.22 (0.21-0.24) 0.35 (0.31-0.40) 67.9 Jensenone No direct mortality occurred in any concentration tested (0.05-0.4%) within 24 h. However, all treated TSM were unable to move normally and continued to convulse until they commenced to die 72 h after treatment. There was no recovery. 110.1 Tasmanone 0.055 O.1SO 101.8 Flavesone 0.020 (0.006-0.043) 0.0876 (0.076-0.114) 77.0 Leptospermone O.O37 O.169 113.4 Isoleptospermone 0.043 (0.027-0.057) 0.071 (0.058–0.109) 10 141.1 Platyphyllol O.O70 O.23
Example 8 Example 10 E. cloeziana Oil Extract as a Fumigant Efficacy of Purified B-Triketones Against GHT 0197) The efficacy of purified f-triketones against GHT 0193 An investigation was undertaken to determine the was investigated using the GHT bioassay described in fumigation action of E. cloeziana oil extract against arthro Example 8. All B-triketones tested demonstrated a high level pods. The test organisms used were termites, Nasutitermes of activity against GHT (see Table 9). 0198 All B-triketones except jensenone caused 100% walkeri. A Whatman No 2 filter paper (90 mm diam.) was mortality on GHT at a concentration of 0.3% when 5 mL immersed in distilled water five seconds and left to drain aliquots were applied with a Potter spray tower. The latter excess water before placing it on the bottom of a 90 mm diam. B-triketone did not cause direct mortality within 24 h, but petri dish, to provide moisture for termite workers during the caused behavioural effects at all concentrations tested. Con experimental period. A 1.0% E. cloeziana oil solution was Vulsion and lack of movement were consistently observed prepared in pure acetone. One mL was uniformly spread on a and 60.0% mortality was recorded 72 h after application in second filter paper, which was allowed to air dry on a sheet of the 0.4% treatment. US 2010/0227009 A1 Sep. 9, 2010 22
Examples 11 Example 14 0199. 0203
Ready-to-use Initicide Spray - I Oil-based natural spray concentrate Ingredient Parts
E. Cloeziana extract O.1 Ingredient Parts Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O polyoxyethylenesorbitan, organosilicate E. Cloeziana extract 1O.O Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 5 Tannic acid 1.O Petroleum oil 89.0 Carrier e.g. water 92.9 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O polyoxyethylenesorbitan, organosilicate
Examples 12 Example 15 0200 0204
Concentrated natural emulsifiable concentrate spray (4.4%) - I Ingredient Parts Concentrated emulsifiable concentrate spray (10% E. Cloeziana extract 4.4 Ingredient Parts Pyrethrins 7.4 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 14.7 E. Cloeziana extract 1O.O polyoxyethylenesorbitan, organosilicate Permethrin 1O.O Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 73.5 Piperonyl butoxide 28.0 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 31.0 polyoxyethylenesorbitan, organosilicate Solvent e.g. ethyl alcohol, isopropyl alcohol etc 21 Example 12 0201 Example 16 0205 Concentrated emulsifiable concentrate spray (44% Ingredient Parts Molluscidal dust E. Cloeziana extract 22.0 Platyphyllol (natural or synthetic) 22.0 Ingredient Parts Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 34.0 polyoxyethylenesorbitan, organosilicate E. Cloeziana extract 2 Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 22 Anti-caking agent (e.g. silica gel) 2 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 3 polyoxyethylenesorbitan, organosilicate Inert carrier (talc, kaolin, diatomaceous earth) 93 Example 13 0202 Example 17 0206 Natural ready to use insecticide Spray Ingredient Parts Aerosol insecticidal and acaricidal Spray E. Cloeziana extract O.3 Lavender oil 1.O Ingredient Parts Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O polyoxyethylenesorbitan, organosilicate E. Cloeziana extract 1.O Solvent e.g. ethyl alcohol, isopropyl alcohol etc 40 Piperonyl butoxide O.9 Carrier: Water 57.7 Propellent hydrocarbon 98.1 US 2010/0227009 A1 Sep. 9, 2010
Example 18 Examples 22 0211 0207
Concentrated natural emulsifiable concentrate Spray (4.4%) - III Repellent Ingredient Parts
Ingredient Parts 99% Tasmanone 4.0 Pyrethrins 7.4 E. Cloeziana extract 19.5 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 14.7 Citronella oil 29.1 polyoxyethylenesorbitan, organosilicate Phthalic acid dibutyl ester 29.1 Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 73.9 N-octyl bicycloheptenedicarboxamide 22.3 Examples 23 Examples 19 0212 0208 Ready-to-use miticide spray - IV Ingredient Parts Ready-to-use miticide spray - II 99% Platyphyllol O.09 Ingredient Parts Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O polyoxyethylenesorbitan, organosilicate Melaletica Caieptiti extract O.2 Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 5 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O Tannic acid 1.O polyoxyethylenesorbitan, organosilicate Carrier e.g. water 92.91 Solvent: e.g. ethyl alcohol, isopropyl alcohol etc S.O Tannic acid 1.O Carrier e.g. water 92.8 Examples 24 0213 Examples 20
0209 Concentrated natural emulsifiable concentrate spray (4.4%) - IV Ingredient Parts 99% Platyphyllol 4.0 Concentrated natural emulsifiable concentrate spray (4.4%) - II Pyrethrins 7.4 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 14.7 Ingredient Parts polyoxyethylenesorbitan, organosilicate Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 73.9 Melaletica Caieptiti extract 8.8 Pyrethrins 7.4 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 14.7 polyoxyethylenesorbitan, organosilicate Example 25 Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 69.1 Intra-Specific Crosses for Imparting Pest Resistance to a Pest Susceptible Plant 0214 Controlled and wild pollination within Eucalyptus Examples 21 and other important commercial Myrtaceae are thoroughly addressed in the CRC for Sustainable Production Forestry 0210 Symposium on Hybrid Breeding and Genetics Controlled Pollination of Eucalypts on 12 Apr. 2000 and published as the proceedings of that symposium. Genetic Pollution from Farm Forestry (Potts et al., 2001) deals more specifically with intra Ready-to-use miticide Spray - III and inter species crosses occurring within the Myrtaceae. Ingredient Parts 0215 Using the protocols described in the above publica tions, an intra-specific Eucalyptus cross breeding technique 99% Tasmanone O.09 Emulsifier: e.g. t-octylphenoxypolyethoxyethanol, 1.O will employ the following protocol. This begins with the polyoxyethylenesorbitan, organosilicate selection of Supreme individuals as parent stock. Within a Solvent: e.g. ethyl alcohol, isopropyl alcohol etc 5 selected parent stock having Superior pest resistant character Tannic acid 1.O istics (e.g., E. Cloeziana) male and female trees are identified Carrier e.g. water 92.91 for cross-pollination experiments. Pollen is harvested from the male trees and either stored or directly transferred to the female trees if flowering is synchronous. Emasculation is US 2010/0227009 A1 Sep. 9, 2010 24 undertaken to preclude extraneous pollination occurring and maidenii E. grandisXE. globulus E. grandisXE. gunnii E. flowers are often bagged as a further precaution. Seed set and grandisXE. pulverulenta E. grandisXE. leucoxylon, E. gran Subsequent embryo development proceeds over the ensuing disxE. resinifera. In another example the Corymbia henryi/ 12-24 months and F1 seeds are collected at full maturation. variegata/maculata/citriiodora complex displays 14 natural 0216 Seeds are then germinated to produce seedlings that hybrids including C. citriodorax.C. catenaria, C. citriodorax are subjected to detailed analysis to assess the transfer of traits C. variegata, C. citriiodorax.C. maculata, C. maculataxC. from parent to progeny. If the F1 progeny show a desirable gummifiea, C. maculataxC. intermedia, C. maculataxC. cit mix of phenotypic traits these progeny can be used to vegeta riodora, C. maculataxC. variegata, C. variegataxC. blox tively propagate the new variety. If the F1 progeny show some Someri, C. variegataxC. Watsoniana, C. variegataxC. citri improvement in the selected phenotypic traits, but further Odora, C. variegataxC. maculata, C. HenryixC. torelliana, improvement is required selected F1 progeny can be back C. henryixC. variegata and onemanipulated hybrid C. torel crossed either within the F1 progeny or with one of the parent lianaxc. In one further example Eucalyptus cloeziana dis trees to produce an F2 progeny using the methodology out plays only 1 natural hybrid E. cloezianaxE. acnenoides. lined above. This iterative process can be continued ad infini 0218. The disclosure of every patent, patent application, tum until the desired characteristics are achieved. and publication cited herein is hereby incorporated herein by Example 26 reference in its entirety. 0219. The citation of any reference herein should not be Inter-Specific Crosses for Imparting Pest Resistance construed as an admission that Such reference is available as to a Pest Susceptible Plant “Prior Art” to the instant application 0220. Throughout the specification the aim has been to 0217 Inter-species hybridisation in the wild, which is a describe the preferred embodiments of the invention without common phenomenon within the Subgenera of the Myrta limiting the invention to any one embodiment or specific ceae, have been recorded at almost 40% in the Eucalypts, collection of features. Those of skill in the art will therefore 33% in Angophora 19% in Corymbia and 19% in Sym appreciate that, in light of the instant disclosure, various phomyrtus. Hybridisation between the major Subgenera may modifications and changes can be made in the particular also occur (Potts et al., 2001). For example, Eucalyptus cam embodiments exemplified without departing from the scope aldulensis displays 14 natural hybrid crosses including E. of the present invention. All Such modifications and changes camaldulensisXE. robusta, E. CamaldulensisXE. alba, E. are intended to be included within the scope of the appended camaldulensisXE. cladocalyx, E. camaldulensisXE. bigalat claims. All figures, tables, and appendices, as well as publi erita, E. CamaldulensisXE. tereticornis, E. CamaldulensisXE. cations, patents, and patent applications, cited herein are blakelyi, E. CamaldulensisXE. dwyeri, E. CamaldulensisXE. hereby incorporated by reference in their entirety for all pur rudis, E. camaldulensisXE. ovata, E. CamaldulensisXE. bridgesiana, E. CamaldulensisXE. viminalis, E. Camaldulen poses. sisXE. largiflorens, E. CamaldulensisXE. meliodora, E. cam aldulensisXE. leucoxylon, 15 manipulated hybrids including I. BIBLIOGRAPHY E. CamaldulensisXE. diversicolor, E. camaldulensisXE. gran dis, E. CamaldulensisXE. botryoides, E. CamaldulensisXE. 0221 Bignall, C. M., Dunlop, P. J., Brophy, J. J. and cladocalyx, E. camaldulensisXE. tereticornis, E. camaldu Fookes, C. J. R. (1997). Volatile Leaf Oils of some South lensisXE. blakelyi, E. CamaldulensisXE. urophylla, E. cam western and Southern Australian Species of the Genus aldulensisXE. macarthurii, E. CamaldulensisXE. exerta, E. Eucalyptus (Series I). Part XIV. Subgenus Monocalyptus. camaldulensisXE. maidenii, E. CamaldulensisXE. viminalis, Flay Frag. Journal, 12, 177-183. E. CamaldulensisXE. globulus, E. camaldulensisXE. gunnii, 0222 Boland, D. and J. Brophy (1993). Essential Oils of E. camaldulensisXE. laevOpinea, E. CamaldulensisXE. fasti the Eucalyptus and Related Genera Search for Chemical gata. In another example Eucalyptus globulus displays 15 Trends. Bioactive Volatile Compounds from Plants. R. Ter natural hybrids including E. globulusXE. barberi, E. globu anishi, R. G. Buttery and H. Sugisawa. Washington D.C., lusXE. brookeriana, E. globulusXE. ovata, E. globulusXE. American Chemical Society. 525: 72-87. kitsoniana, E. globulusXE.goniocalyx, E.globulusXE.. norto 0223 Brophy, J.J. and D. J. Boland (1990). “Leaf Essen nii, E. globulusXE. Cypellocarpa, E. globulusXE. pseudoglo tial Oil of Two Chemotypes of Eucalyptus cloeziana F. bulus, E. globulusXE. bicostata, E.globulusXE...johnstonii, E. Muell” Journal of Essential Oil Research 2(March/April): globulusXE. viminalis, E. globulusXE. Cordata, E. globulusX 87-90. E. rubida, E. globulusXE. urnigera, E. globulusXE. perrini ana and 13 Successful manipulated hybrids including E. 0224 Brophy, J. J., R. J. Goldsack, et al. (1995). “Leaf globulusXE. urophylla, E. globulusXE. grandis, E. globulusX Oils of the Genus Backhousia (Myrtaceae).” Journal of E. robusta, E. globulusXE. pellita, E. globulusXE. longifolia, Essential Oil Research 7(May/June): 237-254. E. globulusXE. loxophloeba, E. globulusXE. Camaldulensis, 0225 CRC for Sustainable Production Forestry. Sympo E. globulusXE. dunnii, E. globulusXE. Initens, E. globulusXE. sium on Hybrid Breeding and Genetics-Controlled Polli maidenii, E.globulusXE. bicostata, E.globulusXE. viminalis, nation of Eucalypts, Noosa Australia, 12 Apr. 2000 E. globulusXE. gunnii. In another example Eucalyptus gran 0226 Hellyer, R. (1968). “The Occurrence of B-Trike dis displays 4 natural hybrids including E. grandisXE. uro tones in the Steam-Volatile Oils of some Myrtaceous Aus phylla, E. grandisXE. robusta E. grandisXE. pellita E. gran tralian Plants.” Aust. J. Chem. 21 (11): 2825-2828. disxE. terreticornis and 14 manipulated hybrids including E. 0227. Herron GA, Beatie GAC, Parkes R A & Barchia I. grandisXE. urophylla E. grandisXE. botryoides E. grandisx 1995. Potter spray tower bioassay of selected citrus pests to E. pellita E. grandisXE. alba E. grandisXE. terreticornis E. petroleum spray oil. Journal of Australian Entomological grandisXE. Camaldulensis E. grandisXE. dunnii E. initensXE. Society 34: 225-263.
US 2010/0227009 A1 Sep. 9, 2010 26
RandRs are independently selected from H. C-C alkyl,
arylC-Coalkyl, C-C cycloalkyl, C-Coalkenyl, het -continued eroarylC-Co alkyl, C-Co haloalkyl, C-Co diha loalkyl, C-C trihaloalkyl, OR, or SR-7. 3. The method of claim 2, wherein the compound is selected from flavesone (1-isobutyroyl-3,3,5,5-tetramethyl cyclohexan-2,4,6-trione), isoleptospermone (1-isovaleroyl 3.3.5.5-tetramethylcyclohexan-2,4,6-trione), leptospermone (1-valeroyl-3,3,5,5-tetramethylcyclohexan-2,4,6-trione), papuanone (1-pentoyl-3,3,5,5-tetramethylcyclohexan-2,4,6- trione), grandiflorone (1-(2-phenylethyl)-3,3,5,5-tetrameth ylcyclohexan-2,4,6-trione) andjensenone (1-Valeroyl-3,5-di carbonylcyclohexan-2,4,6-trione). 5. The method of claim 2, wherein the compound has a structural formula selected from: 4. The method of claim 2, wherein the compound has a structural formula selected from: CHBr CHBr CHCI O O O O O O O O O O O O O O O
O O O O O
CHCI C HOH CH2OH CHF CF3 O O O O
O O O O O O O O O O
O O O O O
HF F. S H NN 1. H3C NN 1.
S S O S S
O O O
1. CHSH CHNO HO NN O O s t O O O O
O O O US 2010/0227009 A1 Sep. 9, 2010 27
-continued -continued CH2SH CHNO CHOH CHF
O O O O
O O O O O O O O
O O O O C F. CH2SH
O
O O O O
O 6.. ThThe method of claim 2, wherein the compound has a CHNO structural formula selected from: HON O N
CHBr O O O
O O
O O O O O
7. The method of claim 2, wherein the compound has a structural formula selected from: O O
r r O CHBr S H NN H3C NN O O O S S O O O O O O
OHC CHO OHC CHO O O O O O
DC CHCI CHCI CH2OH H O O O YN O
O O O O O O O O
OHC CHO OHC CHO
O US 2010/0227009 A1 Sep. 9, 2010 28
8. The method of claim 1, wherein the compound is repre -continued sented by the general formula (IV) CHF CF
(IV) O O A B X Y O O O O F
E C OHC CHO OHC CHO D O
wherein X and Y are each independently selected from oxygen, sulfur —N R or one of C=X or C=Y is CH: S H NN r A is (C=O)R. (C=S)R, OR, SR, NRRs (C=N- R)R, N=O, N(-O), NROR, or SOR: S S O O B is H. C-C alkyl, C-C alkenyl, aryl or heteroaryl; C, D, E and F are each independently selected from H. OHC CHO OHC CHO C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-Co alkenyl, heteroarylC-Co alkyl, C-Co O haloalkyl, C-Co dihaloalkyl, C-Co trihaloalkyl, C-Co haloalkoxy, OR, SR CRNRRs, NRRs. (C=N-R)R, N=O, N(=O), NROR, SOR: R is selected from H. C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-Co alkenyl, heteroary1C-Co H3C NN H NNDC O alkyl, C-Cohaloalkyl, C-Co dihaloalkyl, C-C tri O O O O haloalkyl, C-C haloalkoxy, C-C hydroxyalkyl, C-Co thioalkyl and C-Co nitroalkyl, OR2, SR, CRNRRs, NRRs (C—N—R)R N=O, N(EO), OHC CHO OHC CHO NROR, or SOR: R is selected from H. C-Co alkyl, arylC-Co alkyl, O O C-C cycloalkyl, C-C alkenyl, heteroarylC-Co alkyl, C-Cohaloalkyl, C-Co dihaloalkyl, C-C tri CHSH CHNO haloalkyl, CRNRRs, NRRs (C—N—R)RN=O, N(=O), or NROR; O O R is selected from H. C-Co alkyl, arylC-Co alkyl, C-C cycloalkyl, C-C alkenyl, heteroarylC-Co alkyl, C-Cohaloalkyl, C-Co dihaloalkyl, C-C tri O O O O haloalkyl, C-Co haloalkoxy, OR7, SR-7, CRNRRs, NRRs (C=N-R)R N=O, N(-O), NROR, or OHC CHO OHC CHO SOR; and RandRs are independently selected from H. C-C alkyl, O O arylC-Co alkyl, C-C cycloalkyl, C-Coalkenyl, het eroarylC-Co alkyl, C-Co haloalkyl, C-Co diha loalkyl, C-C trihaloalkyl, OR, or SR-7. HO 9. The method of claim 8, wherein the compound is V selected from the group consisting of agglomerone N N (1-isobutroyl-4-methoxy-5,5-dimethylcyclohex-3-en-2,6- O O dione), lateriticone (1-valeroyl-4-methoxy-3.5.5-trimethyl cyclohex-3-en-2,6-dione), isolateriticone (1-isovaleroyl-4- methoxy-3.5.5-trimethylcyclohex-3-en-2,6-dione) and OHC CHO platyphyllol (6,6-dimethyl-2-acetyl-5-methoxycyclohex-4- ene-1,3-dione). 10. The method of claim 8, wherein the compound is a tasmanone derivative having a structural formula selected from the group consisting of US 2010/0227009 A1 Sep. 9, 2010 29
11. The method of claim 1, wherein the compound is rep resented by the general formula (V) CHBr CHCI CH2OH O O O (V) A B O O O O O O X Y
H C
G D OCH OCH OCH F E
wherein X and Y are independently selected from oxygen, sulfur or CHF CF —N-R, and O O S A is (C=O)R. (C=S)R, OR, SR, NRRs (C=N- R)R, N=O, N(-O), NROR, or SOR: B is H. C-Co alkyl, C-Coalkenyl, aryl or heteroaryl; O O O O S S C, D, E, F, G and Hare each independently selected from H, C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-Co alkenyl, heteroarylC-Co alkyl, C-Co haloalkyl, C-Co dihaloalkyl, C-C trihaloalkyl, C-Co haloalkoxy, OR2, SR, CRNRRs, NRRs, OCH OCH OCH (C=N-R)R, N=O, N(-O), NROR or SOR: R is selected from H. C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-C alkenyl, heteroarylC-Co alkyl, C-C haloalkyl, C-Co dihaloalkyl, C-C tri haloalkyl, C-Co haloalkoxy, C-Co hydroxyalkyl, H- H3C HO- C-Co thioalkyl and C-Co nitroalkyl, OR2, SR, N N N CRNRRs, NRRs (C—N—R)R N=O, N(EO), O O O O O O NROR, or SOR: R is selected from H. C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-C alkenyl, heteroarylC-Co alkyl, C-Cohaloalkyl, C-Co dihaloalkyl, C-C tri haloalkyl, CRNRRs, NRRs (C—N—R)RN=O, OCH OCH OCH N(=O), or NROR; R is selected from H. C-C alkyl, arylC-C alkyl, C-C cycloalkyl, C-C alkenyl, heteroarylC-Co alkyl, C-C haloalkyl, C-Co dihaloalkyl, C-C tri CH2SH CHNO haloalkyl, C-Co haloalkoxy, OR7, SR-7, CRNRRs, NRRs (C=N-R)R N=O, N(-O), NROR, or O O SOR; and RandRs are independently selected from H. C-Coalkyl, O O O O arylC-Co alkyl, C-C cycloalkyl, C-Coalkenyl, het and eroarylC-Co alkyl, C-Co haloalkyl, C-Co diha loalkyl, C-C trihaloalkyl, OR, or SR-7. 12. The method of claim 11, wherein the compound is selected from angustione (1-acetyl-3,5,5-trimethylcyclohex OCH OCH 2,6-dione), dehydroangustione (1-acetyl-3,5,5-trimethylcy clohex-3-en-2,6-dione) and xanthostemone (1-isobutroyl-5, 5-dimethylcyclohex-3-en-2,6-dione). 13. The method of claim 11, wherein the compound has a HO structural formula selected from:
O O CHBr O CHCl2 O O. O OH O O O OH
OCH US 2010/0227009 A1 Sep. 9, 2010 30
-continued -continued O CH2OH O CHF O CF O CHNO HO1 Nn
O OH O OH O OH HO O HO O
S HN 1. N 1. 15. The method of claim 11, wherein the compound has a S SH O OH O OH structural formula selected from:
CHBr CHCI HOn 1 O CHSH O CHNO2 O O O N r O OH OH O O O O O O
CHF CF HO1 NN CHOH 2 O O O O OH
O O O O O O
14. The method of claim 11, wherein the compound has a structural formula selected from: S H na 3C N 1. H NN 1.
O O O O CHBr O CHCl2 S S O O
HO O HO O HO O 1. CHSH CHNO HO NN O O O CH2OH O CHF O CF O O O O O O HO O HO O HO O
HO S HN1 N1 V SH S t O
HON 1. O CHSH 16. The method of claim 1, wherein the compound is HO O HO O obtainable from a volatile oil-bearing organism. 17. The method of claim 16, wherein the volatile oil-bear ing organism is selected from Volatile oil-bearing plants. 18. The method of claim 16, wherein the volatile oil-bear ing organism is selected from plants from the families Alli US 2010/0227009 A1 Sep. 9, 2010
aceae, Apiaceae, Asteraceae, Cannabinaceae, Lamiaceae, 21. The method of claim 1, wherein the pest that is con Pteridaceae, Myrtaceae, Myoporaceae, Proteaceae, Rutaceae trolled is selected from insects, arachnids and molluscs. and Zingiberaceae. 22. The method of claim 1, wherein the helminth is a 19. The method of claim 16, wherein the volatile oil-bear nematode. ing organism is selected from genera of the Myrtaceae family. 23. The method of claim 1, wherein the compound is used 20. The method of claim 16, wherein the volatile oil-bear in the form of a pest-controlling composition which com ing organism belongs to a genus selected from Angophora, prises from about 0.00005% to about 90% by weight of said Austromyrtus, Backhousia, Baeckea, Callistenon, Coryn compound. bia, Darwinia, Eucalyptus, Kunzea, Leptospermum, Mela leuca, Syzygium and Xanthostemon.