WO2015028942A1 - Biocidal compound, its production process and wood preservative composition - Google Patents

Abstract

A biocidal compound characterized in that it comprises the following molecule (Figure 2): wherein: R1 is selected from the group comprised of methyl and propylene, and R2 is selected from the group comprised of C12H25, C15H25 and C11F17H6 A process for making it, and a wood preservative composition comprising it.

Description

Title:

BIOCIDAL COMPOUND, ITS PRODUCTION PROCESS AND WOOD

PRESERVATIVE COMPOSITION

Field of the Invention

The present invention falls within the technical field of biocidal compounds, in particular those with low toxicity to the environment.

State of the Art

Toxic compounds are known and used as biocides which are slowly being replaced by synthetic compounds which are not as aggressive for the environment. In recent years the functional characteristics and biocides of the gemini compounds have been studied that are presented as ecological alternatives of preservation (Murguia, et al . 2008a) . Few patent documents have been identified which are related to the state of the art, such as US20040234492, PL383499, PL211725, US20050031653 and PL386714.

The harvested wood becomes a completely biodegradable material without defense against the action of a number of biological agents (Encinas et al . , 2004) . The fungus, by settling in timber, develops structures that rapidly canalize walls and cell lumens, producing a biochemical transformation with physical projection. This can cause great harm to economic status in the industry, since the percentages of lignin and cellulose of the wood affected are altered (Chaparro et al . 2009) .

Preservatives used in the industry of wood preservation are classified as creosote, oil-soluble compounds, and hydro-soluble compounds. According to the American standards AWPA (American Wood Protection Association) , creosote is obtained from the distillation of coal tar; it is an oily liquid, consisting of approximately 200 chemical compounds derived mostly from aromatic hydrocarbons, containing neutral, acidic and alkaline substances (Novoa Robles et al . 2006) . It is highly effective against fungi and insects; however, due to its high toxicity a gradual abandonment of its use has been observed since the late eighties. In addition, some components of creosote are not easily decomposed and are harmful to the environment (Fuentes-Salinas et al . , 2002) . The different formulations of oil-soluble organic preservatives consist of one or several synthetic, organic active principles, adjuvants and a solvent that allows the penetration into the wood. In general, these compounds have great toxic power and high penetrating power: pentachlorophenol, naphthenes and 2-

(thiocyanomethylthio) benzothiazole (TCMTB) (Fuentes- Salinas et al . 2002) . The hydro-soluble compounds consist of multiple inorganic salts, which show good fixability and high antifungal and insecticidal activity as chromium, copper and arsenic oxides (CCA) , the ammonium salts with copper and arsenic oxides (ACQ) , and copper, chromium, boron oxides (CCB) (Jimenez Borrallo et al . , 2007) .

In recent years, some of the main preservative substances used in the treatment of wood have been subject to restrictions on its application. Security and toxicity problems to which relative phenomena of resistance of these preservatives are added, maintain the current need for further development of new antifungal agents that provide significant advantages over existing ones; new substances that are less harmful to humans and the environment. In this context, some countries like USA, Germany and Sweden have chosen to replace the use of CCA for alternative products, such as CCB, ACQ. However, these treatments are more expensive and have limited data on its behavior and environmental impact (Borrallo Jimenez et al . 2007) .

There is currently a growing interest in the compounds of gemini type derived from quaternary bis- ammonium salts due to its remarkable antimicrobial activity, excellent surface active properties and some of them have favorable toxicological and biodegradability data (Perez et al . 1996) . Gemini compounds are comprised of two amphipathic molecules, connected at the level of the polar heads or close to them by a spacer group of variable nature: hydrophilic, hydrophobic, rigid or flexible. The main advantages of these in comparison to corresponding conventional surfactants are: increased activity of surface, lower critical micelle concentrations (CMC) , enhanced solubilization, interesting properties of viscosity and demonstrated antimicrobial activity. This leads to the fact that the gemini compounds may be used in lower quantities compared to conventional surfactants (Sekhon et al. 2004) . The structural nature of these molecules and their dissolution properties confer increased capacity of hydrophobic interactions within and between molecules and additionally are able to form three-dimensional clusters as micelles, vesicles or bilayers (Zhu et al . 1990) . The gemini type surfactants possess antimicrobial capabilities to both prokaryotes and eukaryotes (Massi et al . 2003) . Reported are di-and trimeric cationic gemini microbicidal activity against Gram (+) and Gram (-) bacteria, yeasts and filamentous fungi (Murguia et al . 2008b) . For all these reasons, our interest has been to design and synthesize new cationic gemini type antifungal compounds derived from bis-ammonium quaternary salts with potential use for preserving wood and non-toxic to the environment .

The present invention provides a biocidal compound type gemini; a process for obtaining the same and compositions for preserving wood comprising said compound.

Brief Description of the Drawings

Figure 1. Synthetic route of the gemini biocidal compounds of the present invention. Reagents and conditions: (i) 1, Acetic anhydride, 110 °C - 90 minutes; (ii) 2, N, iV-dialkyl amines, tetrabutylammonium bromide (TBABr) , ethanol, 30 °C - 8 hours.

Figure 2. Molecule corresponding to the biocidal compound of the present invention.

Figure 3. Characteristics of some of the tested yeast: a) macroscopic observation of C. parapsilosis colonies, b) microscopic observation of C. albicans.

Figure 4. Microscopic characteristics: a) Aspergillus niger and b) Alternaria sp .

Figure 5. Fuchs-Rosenthal chamber: a) Image of a Fuchs-Rosenthal chamber, b) counting grid: It shows 16 large squares, each of 1mm². Each square is divided into 16 small squares with a surface of 0.0625 mm².

Figure 6a. Results of chronic toxicity study in terms of survival. The number of living individuals in terms of days, for each concentration evaluated for the TCMTB-based compound was plotted.

Figure 6b. Results of chronic toxicity study in terms of survival. The number of living individuals in terms of days, for each concentration evaluated for the gemini BLV was plotted.

Figure 7a. Effect of compound based on TCMTB on the fertility of D. magna.

Figure 7b. Effect of compound gemini BLV on the fertility of D. magna.

Figure 8a. Growth of D magna for compound based on TCMTB.

Figure 8b. Growth of D. magna for compound gemini

BLV.

Figure 9. Tertiary fatty amines of various kinds used for the synthesis of gemini surfactants: a) N,N- dimethylfarnesylamine, b) N, iV-Perfluorooctyl dimethyl- propylamine, c) N, iV-dipropenylfarnesylamine, d) N, N- dipropenyl-perfluorooctyl-propylamine, and e) N,N- dipropenyldodecylamine .

Brief Description of the Invention

The biocidal compound, main object of the present invention comprises the following molecule (Figure 2) :

wherein :

Rl is selected from the group comprised of methyl and propylene,

R-i = CH₃ CH₂"CH—CH₂

and R2 is selected from the group comprised of C12H25, C15H25 and C11F17H6

Ci i F ₇H₆

Furthermore, the present invention comprises a salt derived therefrom.

In particular, it comprises a compound selected from the group comprised of: N,N-B±s (1,3- dodecyldimethylammonium) -2-O-acetyl-propane dichloride; N, iV-Bis [1, 3- (E,E) -3, 7, 1 l-trimethyl-2 , 6,10- dodecat rienyldimethylammonium] -2-O-acetyl-propane

dichloride; N, iV-Bis [ 1 , 3-

(4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldimethylammonium] -2-O-acethyl- propane dichloride; N,N-B±s (1,3- dodecyldipropenylammonium) 2-O-acetyl-propane dichloride; N, iV-Bis [1, 3- (E,E) -3, 7, 1 l-trimethyl-2 , 6,10- dodecat rienyldipropenylammonium] -2-O-acetyl-propane dichloride; N, iV-Bis [ 1 , 3-

(4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldipropenylammonium] -2-0-acethyl- propane dichloride.

Another object of the present invention is a wood preservative composition comprising an aqueous solution of said compound of the present invention in a concentration of between 0.1 and 15% W/V; preferably between 1 and 6% W/V.

Another object of the present invention is a process for obtaining said biocidal compound of the invention comprising the following steps:

1. Acetic anhydride is reacted, which preferably is added dropwise over 1 , 3-dichloro-2-propanol to give 2- O-acetyl-1, 3-dichloropropane, which preferably comprises heating the mixture at a temperature of 110 °C for a time between one and two hours, and more preferably further comprises removing the acetic acid formed in the reaction by distillation;

2. The 2-O-acetyl-l , 3-dichloropropane obtained in step 1 is reacted, which preferably is added in a mixture with ethanol and tetrabutylammonium bromide (TBABr) , with tertiary fatty amines to cause the aminoalkylation and give the biocidal compound of the invention, and more preferably it is carried out at a temperature of 30 °C for 18 hours with stirring.

Wherein, in said process, said fatty amines are selected from the group comprised of N, N- dimethyldodecylamine, N, iV-dimethylfarnesylamine, N, N- perfluorooctyl dimethyl-propylamine, N, N- dipropenyldodecylamine, N, iV-dipropenylfarnesylamine, N, N- dipropenyl-perfluorooctyl-propylamine, or mixtures thereof .

Wherein the process, preferably after step 2, ethanol is removed and the purification is performed by adsorption chromatography on silica gel.

Detailed description of the invention

The gemini compounds of the present invention are designed based on β-hydroxy amino/ammonium structures and alkylamino derived from dihalogenated glycerol, as the β- hydroxy amino/ammonium functionality causes disruption of membranes and inhibition of essential enzymes for the development of microorganisms, and functional groups derived from simple complex alcohols that may confer antifungal activity (Murguia et al. 2008b) . Additional, environmental impact studies were performed on the most active compounds using ecotoxicity testing and efficacy studies for the preservation of wood.

According to the literature, there are numerous and varied methods for the synthesis of gemini compounds. In general, the production of these compounds involves two or more reactions in which different kinds of reagents and techniques commonly used in chemical synthesis are involved .

For the synthesis of the new gemini molecules of the present invention, it was necessary to conduct a simple acetylation followed by an aminoalkylation shown in Figure 1.

The first step involves obtaining the 2-O-acetyl- 1 , 3-dichloropropane (2) from acetic anhydride and 1,3- dichloro-2-propanol (1) for a time between 50 and 120 minutes, preferably about 90 minutes.

Thus, a reaction intermediate which constitutes the spacer molecule, 2-O-acetyl-l , 3-dichloropropane (2) is obtained. Its synthesis was carried out in a simple way with a short reaction time and with good results, the pure product of interest was recovered by 98%. In addition, the reactants employed to give 1 , 3-dichloro-2- propanol and acetic anhydride are inexpensive and easily available compounds.

The next step consists of the aminoalkylation, at a temperature of between 20 and 40 °C for several hours, between 5 and 12 hours, of said intermediate with tertiary fatty amine (N, iV-dimethyl ) and (N, iV-dipropenyl ) of different nature to obtain the compounds of interest (3a, 3b and 3c) and (4a, 4b and 4c), respectively (Figure 1) . Said gemini compounds of the present invention are selected from the group comprised of N, iV-Bis (1, 3- dodecyldimethylammonium) -2-O-acetyl-propane dichloride (3a) , N,N-B±s [1,3- (£,£) -3, 7, ll-trimethyl-2, 6, 10- dodecatrienyldimethylammonium] -2-O-acetyl-propane dichloride (3b), N, iV-Bis [ 1 , 3-

(4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldimethylammonium] -2-O-acethyl- propane dichloride (3c), N, iV-Bis (1, 3- dodecyldipropenylammonium) -2-O-acetyl-propane dichloride

(4a) , N,N-B±s [1,3- (£,£) -3, 7, ll-trimethyl-2, 6, 10- dodecatrienyldipropenylammonium] -2-O-acetyl-propane dichloride (4b), N, iV-Bis [ 1 , 3-

(4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldipropenylammonium] -2-0-acethyl- propane dichloride (4c), or mixtures thereof. They were obtained in excellent yields, using a minimum amount of solvent (ethanol) and 2 equivalents of dialkylamine . The latter reaction was conducted under PTC conditions

(phase-transfer catalysis) , using 2 mol% of tetrabutylammonium bromide (TBABr) as phase-transfer catalyst .

Said tertiary fatty amines (Figure 9) (N, iV-dimethyl ) and (N, iV-dipropenyl ) are selected from the group comprised of N, iV-dimethylfarnesylamine (a), N, N- perfluorooctyl dimethyl-propylamine (b) , N, N- dipropenylfarnesylamine (c) , N, iV-dipropenyl- perfluorooctyl-propylamine (d) , N, N- dipropenyldodecylamine (e) , or mixtures thereof.

Notably, each of the intermediates as well as the final products obtained in the various synthesis steps were properly purified and characterized by spectroscopic techniques .

Said gemini compounds of the present invention are soluble in water and are formulated as biocidal compounds in a concentration of between 0.1 and 10% W/V in aqueous solution. This resulting aqueous compositions can be useful in many industrial applications such as the preservation of leather, wood, among others processes. The applications in the timber industry are of primary relevance to the present invention for preservation by impregnation .

Hereinbelow, the following examples describing in detail the manner of carrying out the present invention are presented.

Examples

Example 1

Examples of synthesis of the substances used as reagents for obtaining the biocide compound gemini of the present invention are shown below.

Synthesis of N, W-dimethyl and N, W-Dipropenylamines

Synthesis of N, W-dimethylfarnesylamine and N, W-dimethyl- perfluorooctyl-propylamine

Because of the synthesis of N,N- dimethylfarnesylamine (or N, iV-dimethyl- (E, E) -3, 7, 11- trimethyl-2 , 6, 10-dodecatrienylamine) and N, iV-dimethyl- perfluorooctyl propylamine (or N, iV-dimethyl-

(4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecylamine) (see Figure 9, amines and b) it was necessary to use a solution of dimethylamine in tetrahydrofuran (THF) , the preparation of said solution will be described first, then the dimethylated amine for each synthesis, unsaturated and perfluorinated .

In a two-necked ball containing 10 g of KOH and fitted with a condenser, an aqueous solution of Me₂ H HC1 (18 g; 0.22 mmol) was added dropwise through a glass column. The dimethylamine gas generated in the course of the reaction was passed through a drying column filled with KOH and was condensed and collected in a container with 20 g of THF in -80 °C (acetone/liquid air) . Finally, 20g more of cold THF were added, thereby obtaining a solution of 20% dimethylamine . Once the solution in question was obtained, the next step followed: the synthesis of unsaturated and fluorinated fatty amines.

Synthesis of N, IV-dimethylfarnesylamine (A) : alkyl halide ( (CH₃) ₂C=CH [CH₂CH₂C (CH₃) =CH] ₂CH₂Br ) , (983.25mg, 3.45 mmol) and a solution of dimethylamine in THF 20% (3.78 g, 84 mmol) were placed in a reaction flask. The reaction mixture was kept under magnetic stirring overnight at room temperature. Then, 70 mL of ethyl ether was added and the mixture was extracted with saturated sodium carbonate solution (3 x 15 mL) . Afterwards, the aqueous phase was extracted with ethyl ether (3 x 20 mL) . The organic fractions were combined, extracted with distilled water (2 x 15 mL) , brine (2 x 15 mL) and dried with anhydrous magnesium sulfate. Finally, the solvent was evaporated and the product was obtained as an orange oil with a yield of 90%. The N, iV-dimethylfarnesylamine was used for synthesis without further purification (Kaplanek R et al, 2007) .

Example 2

Synthesis of N, W-Perfluorooctyl dimethyl-propylamine (b) : perfluoroalkyl halide (CF₃ (CF₂) ₇ (CH₂) ₃I ) , (7.06 g, 12 mmol) and the solution of 20% dimethylamine in THF (2.61 g, 58 mmol) were placed in a reaction flask. The mixture was stirred for 24 hours at room temperature; the reaction product was treated with a solution of 1M NaOH and extracted with sulfuric ether (3 x 10 mL) . The ether phase was washed with distilled water and dried with anhydrous magnesium sulfate. Finally, the solvent was evaporated under reduced pressure and the product was obtained with a yield of 92%. The N, iV-dimethyl- perfluorooctyl-propylamine was used for synthesis without further purification (Szlavik Z et al, 2001) .

Example 3

Synthesis of N, W-dipropenylfarnesylamine (c) : a mixture of ethanol (12 mL) , tetrabutylammonium bromide (TBABr 2 mol%) (22.40 mg, 0.07 mmol) , alkyl halide

( (CH₃) ₂C=CH [CH₂CH₂C (CH₃) =CH] ₂CH₂Br), (983.25 mg, 3.45 mmol) and dipropenylamine (335.20 mg, 3.45 mmol) were placed in a reaction flask. The reaction mixture was maintained at a temperature of 30 °C for 8 hours with magnetic stirring. After the reaction (disappearance of visualized dipropenylamine by thin-layer chromatography (TLC) , ethanol was evaporated with the aid of a rotary evaporator and then vacuum was applied to ensure complete solvent removal. Finally, the product was obtained as a colorless oil with a yield of 95%. The N,N- dipropenylfarnesylamine was used for synthesis without further purification (Kaplanek R et al, 2007) .

Example 4

Synthesis of N, W-dipropenyl-perfluorooctyl-propylamine

(d) : a mixture of ethanol (48 mL) , tetrabutylammonium bromide (TBABr 2 mol%) (76.80 mg, 0.24 mmol), the halide of perfluoroalkyl (CF₃ (CF₂) ₇ (CH₂) ₃I ) , (7.06 g, 12 mmol) and dipropenylamine (1.16 g, 12 mmol) were placed in a reaction flask. The reaction mixture was maintained at a temperature of 30 °C for 8 hours with magnetic stirring. After the reaction (disappearance of visualized dipropenylamine by thin-layer chromatography (TLC) , ethanol was evaporated with the aid of a rotary evaporator and then vacuum was applied to ensure complete solvent removal. Finally, the product was obtained as a colorless oil with a yield of 98%. The N, iV-dipropenyl- perfluorooctyl-propylamine was used for synthesis without further purification (Kaplanek R et al, 2007) .

Example 5

Synthesis of N, W-dipropenyldodecylamine (e) : a mixture of ethanol (12 mL) , tetrabutylammonium bromide (TBABr 2 mol%) (22.40 mg, 0.07 mmol) , the alkyl halide (CH₃- (CH₂) iiBr) , (859.19 mg, 3.45 mmol) and dipropenylamine (335.20 mg, 3.45 mmol) were placed in a reaction flask. The reaction mixture was maintained at a temperature of 30 °C for 8 hours with magnetic stirring. After the reaction (disappearance of visualized dipropenylamine by thin-layer chromatography (TLC) , ethanol was evaporated with the aid of a rotary evaporator and then vacuum was applied to ensure complete solvent removal. Finally, the product was obtained as a colorless oil with a yield of 98%. The N, iV-dipropenyldodecylamine was used for synthesis without further purification (Kaplanek R et al, 2007) .

Example 6

Synthesis of the linker molecule 2-O-acetyl-l , 3- dichloropropane ( 2 )

In a 100 mL glass mini-reactor, equipped with a condenser, 1 , 3-dichloro-2-propanol (1) (9.5 mL, 0.1 mol) was placed. Using an ice bath (exothermic reaction) acetic anhydride (23.6 mL, 0.25 mol) was added dropwise. Then, the mixture was gradually heated to reflux and the reaction was maintained at 110 °C for half an hour with magnetic stirring. After this time, the mixture was cooled and the acetic acid formed during the reaction was removed with the aid of a Kurgelrohr micro-distiller (distilling at reduced pressure) . The yield obtained was 98% (percentage recovery of pure product) (Vogel AI, 1989) (see Figure 1) .

Example 7

Synthesis of the gemini type biocidal compound of the present invention using phase-transfer catalysis .

The experimental procedure described below allows obtaining the compound of the present invention (and is shown in Figure 1) .

A 50-mL flask equipped with a condenser and magnetic stirrer was used to obtain the biocidal compound of the present invention. To a mixture of ethanol (2.5 mL) , 2-0- acetyl-1 , 3-dichloropropane (2) (0.923 g, 5.40 mmol) and tetrabutylammonium bromide (TBABr) (0.034 g, 0.108 mmol), the corresponding amine (N, iV-dimethyldodecylamine 2.305 g, N, iV-dimethylfarnesylamine 2.667 g, N, N-dimethyl- perfluorooctyl-propylamine 5.432 g, N, N- dipropenyldodecylamine 2.863 g, N, N- dipropenylfarnesylamine 3252 g, and N, iV-dipropenyl- perfluorooctyl-propylamine (6.017 g, 10.8 mmol) were added. The reaction mixture was maintained at a temperature of 30 °C for 18 hours with magnetic stirring. After the reaction (disappearance of (2) displayed by thin-layer chromatography (TLC) , ethanol was evaporated with the aid of a rotary evaporator and then vacuum was applied to ensure complete removal of solvent. Afterwards, the purification of the target compounds was performed using adsorption chromatography on silica gel, and the following mixed solvents were used as eluent : ethyl acetate/ethanol/distilled water/28 to 30% ammonium hydroxide (4:1:0.25:0.5) . Then, the solvent mixture was evaporated under reduced pressure. Finally, the dicationic gemini 3a—c and 4a—c were obtained as pure compounds isolated in 92, 91, 96% (3a-c) , and 92, 94 and 97% (4a-c) respectively.

Example 8

Characterization of the obtained products of the present invention

2-O-acetyl-l, 3-dichloropropane (2) :

Physical data of compound: colorless oil.

IR (KBr) : V (Crrf¹) = 3462.0, 2960.5, 1643.2, 1423.4, 1371.3, 1234.4, 1043.4, 607.5. ¹E NMR [200 MHz, (CDC1₃) ] : δ= 2.01 (s, 3H) , 3.27 (d, 2H J = 2.1), 3.65 (d, 2H J = 2.1), 4.59 (s, 1H) . ¹³C NMR [50 MHz, (CDC1₃) ] : δ= 17.60, 44.46, 47.44, 79.90, 169.10. MS: m/z (% rel. int.) = 171 (M⁺, 2), 129 (10), 116 (48), 104 (5), 87 (32), 74 (40), 56 (20), 43 (100) . (C₅H₈C1₂0₂ - MW: 171 021) .

N, W-Bis (1 , 3-dodecyldimethylammonium) -2-O-acetyl-propane dichloride (3a) :

Physical data of compound: white waxy product.

IR (KBr) : V (Crrf¹) = 760.1, 930.4, 1120.0, 1460.0, 1620.5, 2850.0, 2930.1, 3450.2. ¹E NMR [200 MHz, (D₂0) ] : δ= 0.84 to 0.91 (m, 10H) , 1.22-1.40 (m, 32H) , 1.94-2.01 (m, 4H) , 2.06 (s, 3H) , 3.12-3.16 (m, 6H) , 3.25-3.27 (m, 4H), 3.32 (s, 6H) , 3.36-3.45 (m, 4H) , 3.53-3.57 (m, 1H) . ¹³C NMR [50 MHz] : δ = 14.05, 20.70, 22.62, 23.79, 26.08, 27.88, 29.37, 29.45, 29.61, 29.62, 31.68, 41.64, 44.64, 44.64, 59.43, 63.50, 64.84, 66.93, 70.11, 70.25, 169.10. FAB-HRMS (M/2-2Cl)⁺: calculated for C₃3H₇oCl2N₂02 : 263.4605, found: 263.4675. N, W-Bis [ 1 , 3- (E, E) -3 , 7 , ll-trimethyl-2 ,6,10- dodecatrienyldimethylairimonium] -2-O-acetyl-propane dichloride (3b) :

Physical data of compound: white waxy product.

IR (KBr) : V (CrrT¹) = 1085.9, 2940.0, 3640.0. ¹H-NMR [200 MHz, (CDC1₃) ] : δ= 1.63 (s, 18H) , 1.69 (s, 6H) , 1.96- 2.02 (m, 16H), 2.05 (s, 3H) 3.14-3.54 (m, 12H) , 3.32 (s, 12H) , 3.87-4.06 (m, 3H) . ¹³C-NMR [50 MHz]: δ= 15.23, 17.56, 22.00, 23.22, 24.26, 25.56, 27.03, 34.22, 38.48, 39.95, 41.11, 53.58, 60.92, 62.77, 63.07, 66.95, 68.83, 71.20, 98.28, 124.17, 124.90, 129.89, 135.27, 144.63, 169.19 . FAB-HRMS (M/2-2Cl)⁺: calculated for C₃9H₇oCl_{2 2}0₂ : 299.4921, found: 299.4525.

N, W-Bis [1,3- (4, 4, 5, 5, 6, 6,7,7,8,8, 9, 9, 10, 10, 11, 11,11) - heptadecafluoroundecyldimethylammonium] -2-O-acetyl- propane Dichloride (3c) :

Physical data of compound: white waxy product.

IR (KBr) : V (Cm^-1) = 1085.9, 2940.0, 3640.0. ^XH-NMR [200 MHz, (CDCI₃) ] : δ= 2.05 (s, 3H) , 2.22-2.34 (m, 8H) , 3.23-3.54 (m, 16H), 3.34 (s, 3H) , 3.84-4.04 (m, 2H) . ¹³C- NMR [50 MHz, (CDCI₃ ) ] : δ= 14.00, 22.80, 25.95, 28.93, 29.23, 29.55, 29.60, 29.90, 32.20, 42.24, 62.74, 64.09, 70.11, 71.14, 72.71, 73.77, 98.28, 110.36, 115.24, 126.65, 130.50, 169.10. ¹⁹F-NMR [200 MHz, (CD₃OD/CFCI₃) ] : δ= -82.0 (M, 6F, 2CF₃), -115.1 (m, 4F, 2CF₂) , -123.1 (m, 12F, 6CF₂) , -123.3 (m, 4F, 2CF₂) , -124.0 (m, 4F, 2CF₂), - 124.9 (m, 4F, 2CF₂) , -127.5 (m, 4F, 2CF₂) . FAB-HRMS (M/2- 2C1)⁺: calculated for C₃iH₃₂F₃₄Cl_{2 2}0₂ : 555.2722, found: 555.2795. N, W-Bis (1 , 3-dodecyldipropenylainmonium) -2-O-acetyl-propane dichloride (4a) :

Physical data of compound: white waxy product.

IR (KBr) : V (CrrT¹) = 760.1, 930.4, 1120.0, 1460.0, 1620.5, 2850.0, 2930.1, 3450.2. ^XH NMR [200 MHz, (D₂0) ] : δ= 0.84-0.91 (m, 10H) , 1.22-1.40 (m, 12H) , 1.94-2.01 (m, 4H), 2.06 (s, 3H) , 3.12-3.16 (m, 6H) , 3.25-3.27 (m, 4H) , 3.36-3.45 (m, 2H) . ¹³C NMR [50 MHz] : δ = 14.05, 20.70, 22.62, 23.79, 26.08, 27.88, 29.37, 29.45, 29.61, 29.62, 31.68, 41.64, 44.64, 44.64, 59.43, 63.50, 64.84, 66.93, 70.11, 70.25, 169.10. FAB-HRMS (M/2-2Cl)⁺: calculated for C₄iH₇₈Cl_{2 2}0₂: 315.5356, found: 315.5375.

N, W-Bis [ 1 , 3- (E, E) -3 , 7 , ll-trimethyl-2 ,6,10- dodecatrienyldipropenylairimonium] -2-O-acetyl-propane dichloride (4b) :

Physical data of compound: white waxy product.

IR (KBr) : V (Crrf¹) = 1085.9, 2940.0, 3640.0. ^XH-NMR [200 MHz, (CDC1₃) ] : δ= 1.63 (s, 18H) , 1.69 (s, 6H) , 1.96- 2.02 (m, 16H), 2.05 (s, 3H) 3.14-3.54 (m, 4H) . ¹³C-NMR [50 MHz] : δ= 15.23, 17.56, 22.00, 23.22, 24.26, 25.56, 27.03, 34.22, 38.48, 39.95, 41.11, 53.58, 60.92, 62.77, 63.07, 66.95, 68.83, 71.20, 98.28, 124.17, 124.90, 129.89, 135.27, 144.63, 169.19 . FAB-HRMS (M/2-2Cl)⁺: calculated for C 47H78C I 2 2O2 : 351.5675, found: 351.5617.

N, W-Bis [1,3- (4, 4, 5, 5, 6, 6,7,7,8,8, 9, 9, 10, 10, 11, 11,11) - heptadecafluoroundecyldipropenylammonium] -2-O-acetyl- propane dichloride (4c) :

Physical data of compound: white waxy product.

IR (KBr) : V (Crrf¹) = 1085.9, 2940.0, 3640.0. ^XH-NMR [200 MHz, (CDCI₃) ] : δ= 2.05 (s, 3H) , 2.22-2.34 (m, 9H) , 3.23-3.54 (m, 16H), 3.34 (s, 6H) , 3.84-4.04 (m, 6H) . ^1JC- NMR [50 MHz, (CDC1₃)] : δ= 14.00, 22.80, 25.95, 28.93, 29.23, 29.55, 29.60, 29.90, 32.20, 42.24, 62.74, 64.09, 70.11, 71.14, 72.71, 73.77, 98.28, 110.36, 115.24, 126.65, 130.50, 169.10. ¹⁹F-NMR [200 MHz, (CD₃OD/CFCI₃) ] : δ= -82.0 (M, 6F, 2CF₃), -115.1 (m, 4F, 2CF₂) , -123.1 (m, 12F, 6CF₂) , -123.3 (m, 4F, 2CF₂) , -124.0 (m, 4F, 2CF₂), - 124.9 (m, 4F, 2CF₂) , -127.5 (m, 4F, 2CF₂) . FAB-HRMS (M/2- 2C1)⁺: calculated for C₃9H₄oF₃₄Cl_{2 2}0₂ : 607.3465, found: 607.3495.

The NMR spectra ^XH, ¹³C and ¹⁹F were performed on a Bruker spectrometer FT-300, using CDCI₃ or D₂0 (depending on the solubility of the compound) as a solvent. All spectra are reported in parts per million (δ) , relative to tetramethylsilane . IR spectra were performed on a Shimadzu spectrometer 8201 PC. In the chromatographic analysis the following equipments were used: gas chromatograph Shimadzu model GC-17AATF; gas chromatograph-mass spectrometer Shimadzu model GCMS-QP 5000.

The column chromatography was performed on silica gel (70 to 230 mesh ASTM) .

Reagents and solvents of analytical grade were used. The purifications of the compounds were performed according to the methods described in the literature (Perrin DD et al, 1980) .

The reagents 1 , 3-dichloro-2-propanol (Sigma- Aldrich) , 3-chloro-l-propene (Sigma-Aldrich) , dodecylamine (Sigma-Aldrich), and N, N- dimethyldodecylamine (Sigma-Aldrich) 98% were purchased commercially . Both bisulfate and high purity tetrabutylammonium bromide (Strem) were used as catalysts dried under vacuum at 100 °C (Perrin DD et al, 1980) .

For the synthesis of the cationic Gemini, tertiary amines were used. The tertiary amines derived from farnesol and perfluoroalkyl were prepared from the corresponding alkyl and perfluoroalkyl halides, respectively (Kim T et al, 1996; Szlavik Z et al, 2001) .

Dimethylamine hydrochloride (Aldrich) 99%, trans bromide, trans-farnesyl (Aldrich) 95% and heptadecafluoro-l-iodoctane (Aldrich) 98% were purchased commercially .

Example 9

Antifungal Activity

For the determination of the minimum inhibitory concentration of the cationic and nonionic compounds synthesized against different fungal strains the following were used:

- Sabouraud dextrose broth (CS) from Britania Laboratories SA, acquired commercially.

- Sabouraud Agar (AS), prepared from CS and Agar- Agar (Britania Laboratories SA) .

- Commercial dimethyl sulfoxide (DMSO) (Sintorgan S.A. Argentina) .

- McFarland turbidity Standard 0.5 (barium sulfate), prepared from barium chloride and sulfuric acid as directed by CLSI (Clinical Laboratory Standard Institute) (for preparation, see ANNEX) .

- Reference microorganisms from the ATCC collection (American Type Culture Collection) : Candida albicans ATCC 64548, Saccharomyces cerevisiae ATCC 9763, Candida parapsilosis ATCC 22019, Aspergillus niger ATCC 16404. - Regional isolation of Candida parapsilosis and Alternaria sp. (Courtesy of Dr. Arturo Simonetta, from the Faculty of Chemical Engineering and Dr. Cristina Lura, from the Faculty of Biochemistry and Biological Sciences - Universidad Nacional del Litoral) .

- Fluconazole (Todofarma S.A.), which was purchased commercially .

- Bio F-36 (Pellital S.A.) acquired commercially. Experimental Procedures

The antifungal activity was evaluated using measurements of MIC expressed in g/mL. To determine the MIC, the broth dilution technique was used. These procedures conformed to the reference methods approved by the CLSI or formerly known as NCCLS (National Committee for Clinical Laboratory Standards) (NCCLS M27-A2 and M38- A, 2002) .

The general procedure involved the preparation of a battery of tubes, each tube containing culture medium (broth) with a different concentration of test compound. Sabouraud Dextrose Broth (Britania Laboratories S.A.) was used as a culture medium, to which its pH was adjusted to 7.0 ± 0.1 with a NaOH solution 3 M. Then, the inoculation of the tubes with the corresponding microorganism proceeded .

Once the tubes were inoculated, they were incubated at 35 ± 0.2 °C for a period that varied between 48 and 72 hours depending on the microorganism used in the assay. Once the incubation was elapsed, we proceeded to observe the presence or absence of growth of the microorganism in each of the tubes.

The CIM value corresponded to the tube with the lowest concentration of antimicrobial agent at which no visible growth of the microorganism used in the test was observed.

For the broth dilution test, yeast and filamentous fungi were used.

The following fungi species were used, the preparation of the inoculum and the CIM value display for each case (yeasts and molds) are described.

The strategy (sequence in which the compounds were tested) used for determining the antifungal activity of the synthesized compounds will be explained later, in Section 4.4.

Yeast fungi :

During the study of the biological activity of the synthesized compounds, the tested yeasts were:

❖ Candida albicans ATCC 64548

^♦♦^♦ Saccharomyces cerevisiae ATCC 9763

❖ Candida parapsilosis ATCC 22019

^♦♦^♦ Candida parapsilosis (Regional isolation) In Figure 3 a-b we can observe the macroscopic and microscopic appearance of two of the above yeasts: C. parapsilosis and C. albicans, respectively.

Inoculum:

Regarding the preparation of the inoculum, in each case, it was made using groove tubes with young cultures between 24 - 48 hours (incubated at 35 °C) from the different species, from which a suspension was prepared in saline (0.85% NaCl) . The cell density of the suspension was adjusted with the aid of a 0.5 McFarland standard. Then, the resulting suspension was suitably diluted to obtain 5.0xl0² - 2.5xl0³ cells/mL (range of concentration which provides reproducible CIM values) .

Because the cell density of the inoculum could not be verified with a spectrophotometer, a microbiological surface count was performed in order to verify the correct preparation.

For this, Petri dishes with 10 mL of AS were prepared, and on the other hand, the inoculum dilutions (lCT¹, 1(T²) . 0.1 mL of each dilution was plated on the surface of a plate and distributed evenly with a Drigalski spatula previously flamed with 70% ethyl alcohol (at all times we worked in duplicate) .

After 48-72 hours of incubation (at 28-30 °C) the plates which had between 30 and 300 colonies were selected, the number of colonies among them was averaged and the initial inoculum concentration was calculated.

Display of the CIM value :

In the case of CIM in liquid media, the cutoff point for determination was made by evaluating if there was turbidity in the tubes. Turbidity measurements are a very quick and useful method to obtain estimates of the number of cells.

A cell suspension appears cloudy to the eye because the cells scatter the light passing through the suspension. The more cells are present be greater scattered light, hence greater the turbidity will be. It can be measured with devices such as a spectrophotometer (optical density, OD) which passes light through cell suspensions and detects the amount of emergent light, unscattered .

Within certain limits, turbidity measurements can be reasonably accurate and have the virtue of being quick and easy to make. In addition, these determinations can usually be done without destroying or significantly modify the sample.

Thus, after the adequate incubation time was elapsed, the CIM value determined in each particular case corresponded to the tube with the lowest concentration of antimicrobial agent at which no turbidity was observed.

Filamentous Fungi :

The filamentous fungal species used to determine the antimicrobial activity of the synthesized molecules were:

❖ Aspergillus niger ATCC 16404

♦♦^♦ Alternaria sp.

In Figure 4 a-b we can observe the microscopic appearance of the aforementioned mold species.

Inoculum:

Groove culture tubes with 7 days of incubation were used for preparation of the inoculum at 35 °C from the different species. From them, a conidia suspension in saline solution (NaCl 0.85%) was prepared. For A. niger ATCC 16404, the addition of one drop of Tween 20 may facilitate the preparation of the inoculum. The resulting mixture of conidia and hyphal fragments was transferred to a sterile tube and allowed to stand for 3 to 5 minutes for the heavier particles to settle. Then, the homogeneous suspension (without the heavy particles) was poured into another sterile tube and appropriately diluted .

The inoculum concentration was adjusted by counting in Fuchs-Rosenthal chamber (Figure 5 a-b) to achieve a final concentration of 0.4xl0⁴ - 5, 10⁴ CFU/mL (range of concentration which provides reproducible CIM values) .

Furthermore, the inoculum control was effected by a microbiological count on surface. Petri dishes were prepared with 10 mL of AS and on the other hand, the inoculum dilutions (10^_1, 10^~2) . 0.1 mL of each dilution was plated on the surface of a plate and distributed evenly with a Drigalski spatula previously flamed with 70% ethyl alcohol (here we also worked in duplicate) . After 48-72 hours of incubation (at 28-30 °C) , the plates which had between 30 and 300 colonies were selected, the number of colonies among them were averaged and the initial inoculum concentration was calculated.

Display of the CIM value :

To determine the CIM value, the tube with the lowest concentration of antifungal agent that inhibited visible growth of the organism in question was taken into account .

It is important to make clear that the broth dilution of the gemini compounds of the present invention was performed according to the following methodology:

For molecules of cationic nature, as they are water- soluble, no solubility problems arose when diluted with the previously prepared broth. The procedure used in the preparation of the experience can be seen in Table 1.

Table 1 Scheme used to prepare the dilutions of cationic soluble gemini compounds of the present invention in water. Susceptibility testing in broth.

Step Concentration Source Volume (+) Middle ■) Intermediate (=) Final

^g/mL) (mL) (mL) Concentration Concentration

^g/mL) (1/10) ^g/mL)

1 5120 Stock 1 7 640 64

2 640 Step 1 1 1 320 32

3 640 Step 1 1 3 160 16

4 160 Step 3 1 1 80 8

5 160 Step 3 0.5 1.5 40 4

6 160 Step 3 0.5 3.5 20 2

7 20 Step 6 1 1 10 1

8 20 Step 6 0.5 1.5 5 0.5

9 20 Step 6 0.5 3.5 2.5 0.25

10 2.5 Step 9 1 1 1.25 0,125

11 2.5 Step 9 0.5 1.5 0,625 0.0625 2.5 Step 9 0.5 3.5 0.3125 0.03125

It is important to note that a positive control or inoculum development (no antimicrobial agent) and a negative control or drug (no inoculum) was performed in all assays.

Additionally, to compare the obtained CIM values, two reference compounds were tested:

^♦♦^♦ Fluconazole, azole derivative of clinical use and high antifungal activity.

^♦♦^♦ Bio F-36, a preservative used in leather tanning, TCMTB-based and a carbamic derivative.

The azole compound was faced to each of the microorganisms (yeasts and molds) listed above. Instead, the TCMTB (corrosive compound and harmful for people and the environment) was used with some of the microorganisms tested, on the final stage of this microbiological screening .

In general, the range of concentrations used in the broth dilution tests was of 0.03125 to 64 mg/mL, as shown in Table 1.

However, in some cases it was necessary to modify the range from 0.25 to 512 g/mL due to the manifestation of a lower sensitivity by some microorganisms.

The obtained results are presented and analyzed below .

The obtained results (CIM values) for each compound and each of the studied strains are presented in Table 2.

Table 2. CIM values expressed in mcg/mL for the tested compounds (3a-c) and (4a-c) .

(CIM) (Mg/mL)

Compound C. parapsilosis ^al C. albicans ^bl C. parapsilosis ^bl S. cerevisiae ^bI

ATCC 22019 ATCC 64548 Regional ATCC 9763 3 a 8 8 16 4

3 b 64 32 64 8

3 c 64 32 64 32

4 a 8 8 16 8

4 b 0.25 32 16 2

4 c 64 32 64 16

Fluconazole 16 <5 8 16

a Reading taken at 72 hours. Development was observed in the positive control after 48 hours.

b Reading taken at 48 hours. Development was observed in the positive control after 24 hours.

¹ Corresponds to the Candida genus.

² Corresponds to the Saccharomyces genus.

In general, from the values presented in Table 2, we can state that the tested compounds (3a-c) and (4a-c) were shown to be biologically active against the four tested yeasts.

For compounds with an apolar region of 12 carbons in length (3a and 4a) we can see that both showed similar activity together. In these two molecules, the length of the hydrophobic chains is identical and they manage to achieve the desired effect.

If we compare the performance of these compounds with that found for Fluconazole, we can claim pleasantly that the gemini compounds exhibit a very good antifungal activity with CIM values comparable to those of this effective azole derivative.

Due to their excellent performance as antifungal agents, the 3rd gemini is selected as a candidate for efficacy testing in INTI-Wood and toxicity. From now on, the gemini 3a will be referred to as compound BLV.

Example 11 In vitro toxicity assessment

Toxicity assessment of gemini 3e (BLV) was performed by acute and chronic toxicity tests using Chlorella vulgaris and Daphnia magna as test organisms. The results obtained with the gemini were compared with the corresponding to a commercial control based on TCMTB. Acute toxicity tests with C. vulgaris

Increasing dilutions of gemini BLV (0.32- 20.48 mg/L) and TCMTB (1.562 x 10^~4mg/L -0.02 mg /) prepared from a concentrated stock solution were used, each in distilled water, to which the culture medium of C. vulgaris inoculum was added at a concentration of 5 x 10⁴ cells/mL, incubated for 96 hours using a static type assay. Growing conditions: continuous lighting with white light - cold, 3000 lux intensity and a temperature of 23 ± 1 °C. Three replicates per concentration of the compound being evaluated were performed. The cell count per milliliter was performed with a Neubauer chamber

(Ronco et al . , 2004) calculating the growth rate (μ = 1ηΝ_η- InNo/tn) , the percentage of growth inhibition (1% =

[μ_α-μ±/μα] x 100) and the 96 hour EC₅₀ (by PROBALG2 program) for each of the compounds expressed in mg/L. Trials of acute and chronic toxicity with D. magna

The cultivation of D. magna was performed with reconstituted hard water with nutrients necessary for normal development at pH 7.0 ± 0.1; 10 infants were taken from there and different dilutions of the test compounds were added: gemini BLV (7.68 mg/L-0.48 mg/L) and of TCMTB

(l,125xl0^~3 mg/L 1, 755 x 10^~5 mg/L) . a static test was performed, incubated 48 hours at 23 ± 1 °C in the presence of white light, cold during 18 hours and 6 hours of darkness. A count of the living and dead organisms

(immobile) at 24 and 48 h was performed. Determinations were done in triplicate and a control was carried out without the addition of test compounds. Acute toxicity was assessed using measures of CL₅₀ 24 and 48 h, expressed in milligrams per liter (mg/L) using the Probit model (Diaz Baez and et al . 2004) .

For chronic toxicity assays (sublethal) cultures were performed in the same culture conditions than in the case of acute, but with different concentrations of the test compounds: gemini BLV: 0.085 mg/L; 0.17 mg/L and 0.34 mg/L; commercial TCMTB: 3,125 x 10^~6 mg/L; 6.25 x 10^{" 6} mg/L; 1.25 x 10^~5 mg/L; 2.5 x 10^~5 mg/L; 5 X 10^~5 mg/L; 1 x 10^~5 mg/L. In addition, a control without the addition of drugs and significantly longer time period was performed. One test was performed with culture medium renewal every other day (Gomez et al. 2001), setting the exposure levels based on the results obtained in acute assays. It was incubated during 21 days with periodic monitoring of survival, growth and fertility of the organisms. ANOVA (a = 0.05) was implemented with Dunnett ' s post-test using Graphpad InStat program.

Ecotoxicity results

Acute toxicity tests with C. vulgaris:

The tests conducted met the acceptability conditions of the test: the increase in the number of cells per milliliter was a factor of 16 at the end of the test and the variation between replicates was not more than 20%. EC₅₀ values calculated at the request of the growth rate (μ) were 2.172 x 10^~4 mg/L for TCMTB and 0.6568 mg/L for gemini BLV, yielding a difference of three orders of magnitude between the two. C. vulgaris turned out to be less sensitive to increases in the concentration of BLV, that at increasing concentrations of the commercial formulation (TCMTB) . It is noteworthy that the concentrations used for each compound differ widely because when testing the commercial compound similar to those used for the compound of the present invention concentrations, the organisms were not viable, a high mortality was recorded. LC₅₀ values of 24 hours showed a difference in four orders of magnitude between the compound and the synthesis BLV compound and the commercial (3.397 mg/L and 3.4 x 1(T⁴ mg/L respectively), whereas in the LC₅o for 48 hours, the difference was 0.21 mg/L and 1 x 10^~4 mg/L respectively. The LC₅o for 48 hours reported in the literature for the TCMTB is 0.06 mg/L

(Buckman Laboratories, Inc. 2007); however, our trials had to be performed well below this concentration, to avoid obtaining 100% mortality. This could be due to the toxicity of the excipients used in commercial formulation employed, among which carbamates stand out.

Trials of acute and chronic toxicity to D. magna:

In sublethal tests with D. magna, the commercial compound always turned out to be toxic at lower concentrations than the gemini BLV, detecting an increase in mortality with increasing exposure level at TCMTB

(Figure 6a) . As can be seen, for concentrations above 6.25 x 10^~6 mg/L of the commercial preparation, at 21 days the survival was less than 50%; however, Dunnett post test showed significant differences from control concentrations higher or equal to 2.5 x 10^~5 mg/L (p values <0.01 in all cases) . In turn, the statistical analysis of variance showed highly significant differences (ANOVA, p <0.0001) . Furthermore, for the gemini BLV, we can see that although a decrease in the survival of 21 days elapsed is evident, Dunnett post test indicated that significant differences were observed for concentrations greater than 0.34 mg/L (Figure 6b) .

In studies on the fertility of D. magna, the commercial compound and the gemini BLV showed significant differences from the control (ANOVA, p <0.0001) (Figures 7a and 7b) . In Dunnett ' s test, BLV showed significant differences from the control without drug (p <0.01) throughout the range of concentrations tested while the commercial preparation only at higher concentrations 3, 13xl0^~6 mg/L (p <0.05) in contrast to what was observed for survival where the limit for the commercial preparation gave 2.5 ^χ 10^~5 mg/L (Figure 7th) .

Finally, to analyze the effect of the compounds on the growth of D. magna in the different checks made throughout the 21 days of incubation, the average number of seedlings was counted by organism. A statistical analysis was performed on these values to determine whether the concentrations used were able to cause significant differences in the growth of the organisms. Both the commercial compound and BLV threw significant differences from the control (p <0.0001 and ANOVA p = 0.2083, respectively) . In turn, in the Dunnett test, only the commercial preparation showed significant differences in concentrations greater than 2,5xl0^~5 mg/L (P <0.0001) (Figures 8a and 8b) .

We can say that of the three parameters, the most influenced one by the presence of these compounds turns out to be fertility. That is, in the presence of concentrations higher than 3,13xl0^~6 mg/L for the commercial TCMTB-based compound, or 0,085mg/L of the gemini BLV, the fertility of D. magna is significantly decreased, while in cases of growth and survival, higher drug concentrations were required for attaining a significant negative impact. The comparison of the commercial compound and the one synthesized in laboratory showed a clear difference between the effect of both, both in D. magna, as well as in C. vulgaris, being BLV much less toxic.

Example 12

Evaluating the effectiveness of gemini BLV as wood preservatives

The gemini BLV compound was obtained in 92% purity and in vitro, it inhibited the development of yeast strains as C. parapsilosis, C. albicans, A. alternata with significant CIM values two orders lower compared to other business assets, and considerably less toxic turned against D. magna in relation to TCMTB. As a wood preservative, it allowed a retarded growth and degradation (brown rot) in extreme conditions of temperature and humidity and with significantly similar results to commercial asset as the CCA.

Assays were performed in INTI woods under standards: ASTM D-2017.1981, 86- Standard Method of accelerated laboratory tests of Natural decay resistance of woods; IRAM 1962 9518. Toxicity, Permanence and Efficiency of Wood Preservers. Institute of Materials Rationalization. 12 pp .; IRAM 9532. 1963 Determination of moisture. Institute of Materials Rationalization. 14 pp .: IRAM 9544. 1985 Method for determination of the bulk density. Argentine Institute of Materials Rationalization. 10 pp.

The impregnation method employed was Bethell or filled cell, which was conducted in a pilot plant impregnation. We worked with wood ponderosa pine (Pinus ponderosa) from the province of Neuquen in the form of strips of square section of 20 mm side and 100 mm in length. To carry out the tests, a total of 30 pine slats were used, of which 20 were impregnated (10 per product type) and the rest was considered a witness.

Through these trials, the following conclusion was reached :

Regarding the toxicity of the BLV preservative 5% W/V, as measured by the resistance of the impregnated wood to fungal degradation - acquired durability - in the same wood and to the same test conditions - cell impregnation method It fills - the same kind of resistance that the CCA was determined.

Acknowledgements

The authors wish to thank the National Agency for Scientific and Technological Promotion (ANPCyT) , the National Council for Scientific and Technical Research (CONICET) and the Universidad Nacional del Litoral (UNL) Argentina, for their support and contribution to the development of this work.

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Claims

CLAIMS Having thus specially described and determined the nature of the invention and how the same may be carried into practice, it is stated and claimed as exclusive property right :

1. A biocidal compound characterized in that it comprises the following molecule (Figure 2) :

wherein :

Rl is selected from the group comprised of methyl and propylene,

R-i = CH₃ CH2"CH— CH2

and R2 is selected from the group comprised of C12H25, C15H25 and C11F17H6

Ci i F ₇H₆

2. A compound as claimed in claim 1 which comprises a salt derivative thereof.

3. A compound as claimed in claim 1, characterized in that it comprises N, iV-Bis (1, 3- dodecyldimethylammonium) -2-O-acetyl-propane dichloride.

4. A compound as claimed in claim 1, characterized in that it comprises N, iV-Bis [ 1 , 3- (E, E) -3, 7 , 11-trimethyl- 2, 6, 10-dodecatrienyldimethylammonium] -2-O-acetyl-propane dichloride .

5. A compound as claimed in claim 1, characterized in that it comprises N, iV-Bis [ 1 , 3- (4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldimethylammonium] -2-O-acethyl- propane dichloride.

6. A compound as claimed in claim 1, characterized in that it comprises N,N-B±s (1,3- dodecyldipropenylammonium) 2-O-acetyl-propane dichloride.

7. A compound as claimed in claim 1, characterized in that it comprises dichloride N, iV-Bis [ 1 , 3- (E, E) -3, 7 , 11- trimethyl-2, 6, 10-dodecatrienyldipropenylammonium] -2-O- acetyl-propane .

8. A compound as claimed in claim 1, characterized in that it comprises N, iV-Bis [ 1 , 3- (4,4,5,5,6,6,7,7,8,8,9,9,10,10,11,11,11)- heptadecafluoroundecyldipropenylammonium] -2-0-acethyl- propane dichloride.

9. A wood preservative composition, characterized in that it comprises an aqueous solution of the compound of claim 1 in a concentration of between 0.1 and 15% W/V.

10. A wood preservative composition according to the preceding claim characterized in that it comprises an aqueous solution of the compound of claim 1 in a concentration of between 1 and 6% W/V.

11. A process of preparing the compound of claim 1, characterized in that it comprises the following steps:

11.1. reacting acetic anhydride and 1,3- dichloro-2-propanol to give 2-O-acetyl-l , 3- dichloropropane,

11.2. reacting the 2-O-acetyl-l , 3- dichloropropane obtained in step 11.1 with tertiary fatty amines to cause aminoalkylation and give the compound of claim 1.

12. The process of claim 11, wherein said acetic anhydride is added dropwise on the 1 , 3-dichloro-2- propanol .

13. The process of claim 11, wherein step 11.1 further comprises heating the mixture at a temperature of 110 °C for a time between one and two hours.

14. The process of claim 11, wherein step 11.1 further comprises removing the acetic acid formed in the reaction by distillation.

15. The process of claim 11, wherein said fatty amines are selected from the group comprised of N, N- dimethyldodecylamine, N, iV-dimethylfarnesylamine, N, N- perfluorooctyl dimethyl-propylamine, N, N- dipropenyldodecylamine, N, iV-dipropenylfarnesylamine, N, N- dipropenyl-perfluorooctyl-propylamine, or mixtures thereof .

16. The process of claim 11, wherein in step 11.2 the said 2-O-acetyl-l , 3-dichloropropane is added in a mixture with ethanol and tetrabutylammonium bromide (TBABr)

17. The process of claim 11, wherein step 11.2 is performed at a temperature of 30 °C for 18 hours with stirring .

18. The process of claim 11, wherein after step 11.2 ethanol is eliminated and the purification is performed by adsorption chromatography on silica gel.