TECHNICAL FIELD

The present invention relates to the field of organic chemistry and more specifically to a method of designing and preparing biologically active compounds, which undergo degradation easily in the environment. More specifically, the invention relates to strategic design of compounds that have a biological function of interest as well as to a functionality which enables subsequent photolytic decomposition under ambient conditions. Thus, the invention provides photolytically degradable compounds which may be used e. g. as parasiticides, pesticides, antimicrobial compounds, such as antibacterials, including antibiotics, and afterwards easily incapacitated by exposure to light.

BACKGROUND

An excessive use of antibiotics both in the human health care sector and agriculture leads to emergence of drug-resistant bacteria strains. Multi-drug resistant (MDR) bacteria are a widespread problem and several public-health organisations have described the situation as a crisis that would have catastrophic consequences. Alarming data from around the world have shown that over 100 pharmaceutical compounds have been detected in drinking water, wastewater, ground water and aquatic organisms. One example of the above pharmaceutical compounds is the antibiotic ciprofloxacin that has been detected in wastewater across the world. Conventional UV treatment of effluent water containing ciprofloxacin has been carried out but proved to have no effect. Another example of such compounds is the broad-spectrum antibiotic chloramphenicol.

To counteract further growth of MDR bacteria, it is necessary to develop antibiotics based on scaffolds with a structural motif that will enable their degradation under ambient conditions so that accumulation in the biosphere does not take place. Since most antibiotics act inside the body, conceivable new agents must be stable under aqueous and in particular physiological conditions.

Furthermore, the aquaculture industry is a large global business, which is estimated to continue growing. In Norway aquaculture products are the largest export commodity after oil and gas, and therefore it important to alleviate problems associated with production of the aquaculture products. One such problem is presence of salmon lice (Lepeophtheirus salmonis), which may induce epizootics in wild fish while feeding on its skin, mucus and blood. Since the salmon farms commonly use open-net pens, such lice infestations can easily move to adjacent farms and may also infect the local populations of fish, such as sea trout (Salmo trutta) and wild Atlantic salmon (Salmo salar).

In order to address the problems related to the salmon lice infestation, large quantities of pesticides, such as diflubenzuron and teflubenzuron (see below), are currently being applied. These compounds act as chitin synthase inhibitors preventing the salmon lice from forming the chitin-rich exoskeleton after moulting and ultimately leading to their death. However, there are many studies that show that the above compounds are stable with a half-life of 170 days. These compounds have been detected in water as far away as 1100 m from pens and have proven to exhibit negative impacts on king crab, shrimp, squat lobster, and European and Norway lobster. The detected levels of teflubenzuron were sufficient to induce mortality in salmon lice during the developmental stages when salmon lice lack exoskeleton.

Additionally, the maximum-residue limit for these pesticides for saithe and the above-mentioned crustaceans, respectively, exceeded that for Atlantic salmon. Therefore, the above discussion also directly concerns food safety. Thus, there is an urgent need to generate chitin synthase inhibitors that have a shorter half-life.

Diflubenzuron Teflubenzuron

Light-degradable compounds which decompose in the environment, such as phosphopyricin, have been described in e. g. CA2972079. Phosphopyricin is an antimicrobial compound with a photosensitive chemical architecture that reduces its accumulation in the environment.

Despite that there is still a need to protect the environment from harmful biologically active compounds by designing new strategies directed to creating efficient ways of reducing amount of or eliminating residues of pharmaceutical compounds and other harmful biologically active compounds in all aqueous compartments.

SUMMARY OF THE INVENTION

In the light of the above, it is an object of the present invention to provide a method for synthesising photolytically degradable compounds as well as photolytically degradable compounds and methods of utilising said compounds for managing water resources, reducing a population of salmon lice and diminishing direct or indirect damage salmon lice causes to fish.

Said invention alleviates at least part of the above-discussed problems and at least partially addresses one or more of the above-mentioned needs.

According to a first aspect of the invention a method is provided for synthesising a photolytically degradable compound, more precisely the method comprising: a) defining a biologically active compound which analogue is to be synthesised, wherein the biologically active compound comprises at least one aromatic group, the aromatic group being substituted, b) providing a reactant molecule comprising the at least one aromatic group, and c) synthesising a photolytically degradable compound by introducing in the reactant molecule at least one nitro substitution in said at least one aromatic group and introducing an oxazolidine group, wherein the photolytically degradable compound is defined by Formula I:

Formula I wherein at least one of A, B, C and D is selected from a group consisting of H, Cl, Br, B(OH) ₂ , CN, CH ₂ NR ¹ R ² and [CH ₂ NR ¹ R ² R ³ ] ⁺ , wherein each of R ¹ , R ² and R ³ is selected from a group consisting of H, branched or unbranched C-i-Ce alkyl, and

O(CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(C H ₃ ) _y , where o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x and y is 1 , while the other three of v, w, x and y are 0, and where X is selected from a group consisting of F and Cl, while each of the other three of A, B, C and D is

1 ) independently selected from a group consisting of H, F, Cl, I, Br, CN, CF ₃ , CI-C ₆ alkyl, alkoxy with a Ci-C ₆ alkyl, NH ₂ , NHR ¹ , NR ¹ R ² and [NR ¹ R ² R ³ ] ⁺ , where R ¹ , R ² and R ³ are as defined above, and

2) is attached to any carbon atom available for substitution in the phenyl group, n is 0-4,

Z is one selected from a group consisting of H, aryl, and NHC(O)R ⁷ , wherein R ⁷ is selected from a group consisting of R ⁵ , wherein R ⁵ is branched or unbranched Ci-C ₃ alkyl,

and derivatives thereof comprising one or more selected from a group consisting of F and Cl, and wherein

K is same as L, wherein L is selected from a group consisting of H, branched or unbranched C1 -C6 alkyl, alkene and (CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(CH ₃ ) _y , wherein o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x, and y is

1 , while each of the other three of v, w, x, and y is 0, and wherein X is at least one selected from a group consisting of F, Cl, alkyne and

K is H and L is as defined above, or

K is H and L is CH2OR ⁴ , wherein R ⁴ is selected from a group consisting of H and branched or unbranched C-i-Ce alkyl, Q is one selected from a group consisting

According to further aspects of the invention photolytically degradable compounds obtained according to the above method are provided. More specifically, chloramphenicol analogues depicted by Formula VI C, wherein n = 0 or 1 :

Chloramphenicol analogues and

Formula VI C a photolytically degradable compound as defined by Formula VII:

Formula VII

According to another aspect of the invention a method for managing water resources is provided, more specifically by utilising the photolytically degradable compounds obtained by the method according to the first aspect of the invention in treatment of water, the water containing at least one selected from a group consisting of microorganisms and pests.

According to another aspect of the invention a method for at least reducing a population of salmon lice (Lepeophtheirus salmonis) is provided, more specifically by utilising the photolytically degradable compounds obtained by the method according to the first aspect of the invention. Alternatively, the photolytically degradable compound obtained by the method according to the first aspect of the invention is provided for use in at least reducing or controlling the size of a population of salmon lice.

According to yet another aspect of the invention a method for at least diminishing direct or indirect damage salmon lice causes to fish is provided, more specifically by utilising the photolytically degradable compounds obtained by the method according to the first aspect of the invention. Alternatively, the photolytically degradable compound obtained by the method according to the first aspect of the invention is provided for use in at least diminishing direct or indirect damage salmon lice causes to fish.

These and other aspects of the invention are apparent from and elucidated with reference to the embodiments described hereinafter.

BRIEF DESCRIPTION OF THE DRAWINGS

For exemplifying purposes, the invention will be described in closer detail in the following with reference to embodiments thereof illustrated in the attached drawings, wherein:

Figure 1 : ¹ H-NMR spectra before and after photolysis of compound 2.

Figure 2: UV-Vis spectra of compounds 2, 2a, 2b and 2c, respectively.

Figure 3: ¹ H-NMR spectra before and after photolysis of compound 2a.

DETAILED DESCRIPTION OF INVENTION

In the following detailed description the technical terms and expressions are defined, and embodiments of the invention are described.

Generally, all terms and expressions used in the application text are to be interpreted according to the meaning commonly applied to them in the pertinent prior art unless explicitly defined otherwise herein.

As used in this specification and the appended claims, the singular forms ”a”, ”an” and ’’the” include plural referents unless the context clearly dictates otherwise. Thus, e. g. ”a photolytically degradable compound” includes one or more photolytically degradable compound, “a molecule” includes one or more molecules and the like.

The term “biologically active compound” is understood within the scope of the invention to refer to a compound which may be capable of at least one selected from a group consisting of inhibiting the growth of organisms, interfering with normal functioning of organisms, destroying organisms and the like.

The term “light” is understood within the scope of the invention to refer to what is commonly known as “daylight” or light used in an artificial and/or closed environment corresponding to “daylight”; the light may comprise light of a specific wavelength or a range of wavelengths.

The term “analogue” is understood within the scope of the invention to refer to what is commonly known a structural analogue or a chemical analogue; i. e. an analogue of compound A is a compound in which one or more individual atoms have been replaced with either an atom or a functional group, the atom and the functional group being different from that in compound A.

Illustrative embodiments of the invention are described below.

According to a first aspect of the invention a method is provided for synthesising a photolytically degradable compound, the method comprising: a) defining a biologically active compound which analogue is to be synthesised, wherein the biologically active compound comprises at least one aromatic group, the aromatic group being substituted, b) providing a reactant molecule comprising the at least one aromatic group, and c) synthesising a photolytically degradable compound by introducing in the reactant molecule at least one nitro substitution in said at least one aromatic group and introducing an oxazolidine group, wherein the photolytically degradable compound is defined by Formula I:

Formula I wherein at least one of A, B, C and D is selected from a group consisting of H, Cl, Br, B(OH) ₂ , CN, CH ₂ NR ¹ R ² and [CH ₂ NR ¹ R ² R ³ ] ⁺ , wherein each of R ¹ , R ² and R ³ is selected from a group consisting of H, branched or unbranched C-i-Ce alkyl, and

O(CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(C H ₃ ) _y , where o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x and y is 1 , while the other three of v, w, x and y are 0, and where X is selected from a group consisting of F and Cl, while each of the other three of A, B, C and D is

1 ) independently selected from a group consisting of H, F, Cl, I, Br, CN, CF ₃ , CI-C ₆ alkyl, alkoxy with a Ci-C ₆ alkyl, NH ₂ , NHR ¹ , NR ¹ R ² and [NR ¹ R ² R ³ ] ⁺ , where R ¹ , R ² and R ³ are as defined above, and

2) is attached to any carbon atom available for substitution in the phenyl group, n is 0-4,

Z is one selected from a group consisting of H, aryl, and NHC(O)R ⁷ , wherein R ⁷ is selected from a group consisting of R ⁵ , wherein R ⁵ is branched or unbranched Ci-C ₃ alkyl,

and derivatives thereof comprising one or more selected from a group consisting of F and Cl, and wherein

K is same as L, wherein L is selected from a group consisting of H, branched or unbranched C1 -C6 alkyl, alkene and (CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(CH ₃ ) _y , wherein o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x, and y is

1 , while each of the other three of v, w, x, and y is 0, and wherein X is at least one selected from a group consisting of F, Cl, alkyne and

K is H and L is as defined above, or

K is H and L is CH2OR ⁴ , wherein R ⁴ is selected from a group consisting of H and branched or unbranched C-i-Ce alkyl,

Q is one selected from a group consisting wherein m=0-4.

According to further aspects of the invention photolytically degradable compounds obtained according to the above method are provided. More specifically, chloramphenicol analogues depicted by Formula VI C, wherein n = 0 or 1 :

Chloramphenicol analogues

Formula VI C and a photolytically degradable compound as defined by Formula VII:

Formula VII

It is appreciated that a person skilled in the art realises that K and L may be in any spatial orientation and hence various stereoisomers may be envisaged according to the present invention.

The wording “defining a biologically active compound which analogue is to be synthesised” implies that a user of the present method decides which biologically active compound to use as a target compound in the method of the present invention.

The synthesising may include a selective synthesis.

The biologically active compound may be selected from a group consisting of antimicrobial compounds and pesticidal compounds.

It is appreciated that the person skilled in the art can determine in view of a relevant biologically active compound, i. e. in this case the biologically active compound which analogue is to be synthesised, the most advantageous parameters for the synthesis according to the first aspect of the invention, such as necessary amounts and concentrations of reagents and appropriate reaction conditions, including temperatures at which the steps of the synthesis process are to be conducted and duration of the steps thereof. For example, the present method may be performed at room temperature. For synthesising some photolytically degradable compounds, temperatures higher than the room temperature, temperatures above about 30 °C, above about 40 °C, above about 50 °C, above about 60 °C may be chosen.

The step c) of the method may include first synthesising a photolytically degradable compound by introducing at least one nitro substitution in said at least one aromatic group of the reactant molecule and then introducing an oxazolidine group. In this case introducing at least one nitro substitution in said at least one aromatic group of the reactant molecule may lead to formation of a first photolytically degradable compound, whereas the following introducing an oxazolidine group in the first photolytically degradable compound may lead to formation of a second photolytically degradable compound.

The at least one nitro substitution in said at least one aromatic group of the reactant molecule may be at least one selected from a group consisting of a metanitro substitution, a para-nitro substitution and an ortho-nitro substitution. Advantageously said at least one aromatic group of the reactant molecule may have both a para-nitro substitution and a meta-nitro substitution.

The photolytically degradable compound may be defined by Formula I A:

Formula I A wherein at least one of A, B, C and D is selected from a group consisting of B(OH)2, CN, CH ₂ NR ¹ R ² and [CH ₂ NR ¹ R ² R ³ ] ⁺ , wherein each of R ¹ , R ² and R ³ is selected from a group consisting of H, branched or unbranched C-i-Ce alkyl, and O(CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(C H ₃ ) _y , where o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x and y is 1 , while the other three of v, w, x and y are 0, and where X is selected from a group consisting of F and Cl, while each of the other three of A, B, C and D is

1 ) independently selected from a group consisting of H, F, Cl, I, Br, CN, CF ₃ , CI-C ₆ alkyl, alkoxy with a Ci-C ₆ alkyl, NH ₂ , NHR ¹ , NR ¹ R ² and [NR ¹ R ² R ³ ] ⁺ , where R ¹ , R ² and R ³ are as defined above, and

2) is attached to any carbon atom available for substitution in the phenyl group, n is 0-4,

Z is one selected from a group consisting of H, aryl and NHC(O)R ⁷ , wherein R ⁷ is selected from a group consisting of R ⁵ , a monohalogenated derivative thereof and perhalogenated derivative thereof, wherein R ⁵ is branched or unbranched Ci-C ₃ alkyl, and wherein

K is same as L, wherein L is selected from a group consisting of H, branched or unbranched C1 -C6 alkyl, alkene and (CH2)o(CX2)p(CH2) _q (CHX) _r (CH2)s(CX2)t(CH2)u(CX ₃ )v(CHX2)w(CH ₂ X)x(CH ₃ ) _y , wherein o = 0-5, p = 0-5, q = 0-5, r = 0-5, s = 0-4, t = 0-4, u = 0-4, v = 0-1 , w = 0-1 , x = 0-1 , y = 0-1 , provided that only one of v, w, x, and y is 1 , while each of the other three of v, w, x and y is 0, and wherein X is one selected from a group consisting of F, Cl, alkyne and

Formula IV K is H and L is as defined above, or

K is H and L is CH2OR ⁴ , wherein R ⁴ is selected from a group consisting of H and branched or unbranched C-i-Ce alkyl.

The photolytically degradable compound may comprise more than one aromatic group. The substituent Z in Formula I may be aryl as defined by Formula II:

Formula II wherein each of the substituents E, F, G and H may be as defined above for the substituents A, B, C and D. The nitro group may be in meta-, ortho- or paraposition to the carbon attaching Z to the carbon chain of the photolytically degradable compound.

Z may be beta lactam. The photolytically degradable compound, wherein Z is beta lactam, may be defined by either Formula III a or Formula III below, wherein n = 0 or 1.

Formula III a Formula III

The substituents A, B, C, D, L, K and H may be same.

Alternatively, it may be said that this is the photodegradable compound, wherein A, B, C, D, L, K and H are same The substituents K and L in Formula I may have various forms.

K may be same as L, wherein L may be branched or unbranched C-i-Ce alkyl, branched or unbranched C1-C3, and C1-C2 alkyl.

K may be H and L may be CH2CH2OR or CH2CH2CH2OR, wherein R one selected from a group consisting of H, and branched or unbranched C-i-Ce alkyl.

The photolytically degradable compound obtained by the above method may be as defined by Formula V:

Formula V

Alternatively, it may be said that this is the photolytically degradable compound, wherein Q is H and K is

The person skilled in the art will appreciate that as the advantages of the present invention are closely related to the chemical structure of the compounds, there may be any organic compound that fulfils the structural criteria presented herein. Non-limiting examples of such organic compounds are a pesticide, exemplified by diflubenzuron or teflubenzuron, and an antibiotic, exemplified by chloramphenicol.

The idea concerning photolytic decomposition has been extended to antibiotics as wastewater treatment plants do not have the necessary design to completely remove the antibiotics from the water. For example, ciprofloxacin has been detected in the effluent from a treatment plant, and another study investigated commonly used antibiotics in the U.S. and showed that even though the concentration of the antibiotics decreased during wastewater treatment they remain in the water. Another antibiotic that causes similar issues is chloramphenicol. The method according to the first aspect of the invention allows for synthesis of a compound with a functionality similar to that of chloramphenicol. The photolytically degradable compound may be chloramphenicol analogue as depicted by Formula VI A.

Formula VI A

The photolytically degradable compound may be a chloramphenicol analogue defined by Formula VI B below, wherein n = 0 or 1 . One or more Cl may be substituted with F.

Formula VI B

According to further aspects of the invention photolytically degradable compounds obtained according to the above method are provided. The photolytically degradable compound may be a chloramphenicol analogue as one selected from those depicted by Formula VI C:

Chloramphenicol analogues

Formula VI C

Additional substitution patterns on the aromatic group may also be envisaged within the scope of the present invention. The photolytically degradable compound may have one substituent selected from A, B, C, D, K, L and Z and have its ethanolamine-based compound-precursor defined by Formula VII a: In this case the photolytically degradable compound will be defined by Formula VII:

Formula VII

The photolytically degradable compound according to the invention may be one or more selected from a group consisting of compounds 2, 2a, 2b, 3, 4, 5, 6, 7, 8, 9 and 10 (Table 1 ).

Table 1 . Compounds 2, 2a, 2b and 3 At least one benzylic C-C bond of said photolytically degradable compound may be heterolytically cleaved by exposure to light at basic pH in aqueous environment. Said basic pH may be between about 7 and about 13, between about 7 and about 11 , between about 7 and 8.

In its broadest sense, the present invention relates to synthesis of photolytically degradable compounds resembling other biologically active compounds, for example antimicrobial compounds, such as antibiotics, and pesticides, which can undergo degradation by exposure to light.

The biologically active compound which analogue is to be synthesised may have at least one biological activity selected from a group consisting of an antimicrobial activity and a pesticidal activity. The biologically active compound which analogue is to be synthesised may be an antibiotic, e. g. ciprofloxacin, chloramphenicol or penicillin. The biologically active compound which analogue is to be synthesised may be a pesticide, e. g. diflubenzuron, teflubenzuron or lufenuron. The photolytically degradable compound synthesised according to the first aspect of the invention may have a biological activity substantially similar to that of its non- photolytically degradable analogue, i. e. the biological activity may be at least 80 %, at least 90 % or at least 99 % of that of the non-photolytically degradable analogue.

The chemical structure of the photolytically degradable compound may be defined by retrosynthetic analysis. As the person skilled in the art knows, the retrosynthetic analysis involves subjecting a target structure, in this case the biologically active compound which analogue is to be synthesised, to a deconstruction process, which corresponds to the reverse of a synthetic reaction. The retrosynthetic analysis includes providing a reactant compound and then devising a path to convert it to the target compound, e. g. identifying an immediate precursor that can be converted to the target compound using a known reaction.

According to another aspect of the invention a method for managing water resources is provided, more specifically by utilising the photolytically degradable compounds provided by the present invention in treatment of water; the water may contain microorganisms as well as pests. The water may be wastewater. The water may be drinking water. The pH of the water may be adjusted as appropriate for example to be between about 7 and about 13, between about 7 and about 11 , between about 7 and about 8. According to another aspect of the invention a method for at least reducing the size of a population of salmon lice (Lepeophtheirus salmonis) is provided, more specifically by utilising the photolytically degradable compounds provided by the present invention. Alternatively, the photolytically degradable compound obtained by the method according to the first aspect of the invention is provided for use in at least reducing or controlling the size of a population of salmon lice. A pesticide or a parasiticide obtained and synthesised according to the method of the invention may be used to treat salmon in aquaculture or open water to reduce the number of or eliminate salmon lice. Following the treatment, the water containing the pesticide may be subjected to light to cause photolytic degradation of any remaining pesticide. The pH of the water may be adjusted as appropriate, for example to be between about 7 and about 13, between about 7 and about 11 , between about 7 and about 8. The person skilled in the art will appreciate that pH conditions may be adjusted depending on the pesticide or parasiticide used.

According to yet another aspect of the invention a method for at least diminishing direct or indirect damage salmon lice causes to fish is provided, more specifically by utilising the photolytically degradable compounds provided by the present invention. Alternatively, the photolytically degradable compound obtained by the method according to the first aspect of the invention is provided for use in at least diminishing direct or indirect damage salmon lice causes to fish. The fish may be selected from a group consisting of salmon, trout and the like.

It is appreciated that the above-mentioned embodiments of the first aspect of the present invention may also apply to the further aspects of the present invention.

The invention has mainly been described above with reference to some embodiments. However, as it is readily appreciated by the person skilled in the art, other embodiments than the ones disclosed above are equally possible within the scope of the invention as defined by the appended claims.

DETAILED DESCRIPTION OF THE DRAWINGS

Figures 1-3 all illustrate results obtained as described in the experimental section below:

Figure 1 shows the ¹ H-NMR spectra before and after photolysis of compound 2 (see Table 1 above). The y-axis shows relative intensity, and the X-axis shows [ppm]. From bottom to top, the spectra are obtained before and after photolysis, respectively. The key CH2-oxazolidine group signals are highlighted in grey. Full decomposition of compound 2 at pH 8 was observed after 2 h.

Figure 2 shows UV-Vis spectra of compounds 2, 2a, 2b and 2c, respectively. The y-axis shows absorbance A (mAU), and x-axis shows wavelength A (nm). Compounds 2, 2a, 2b and 2c had a max of 304 nm, 414 nm, 291 nm and 466 nm, respectively.

Figure 3 shows ¹ H-NMR spectra before and after photolysis of compound 2a (see Table 1 above). From bottom to top, the spectra are obtained before and after photolysis, respectively. The y-axis shows relative intensity, and the X-axis shows [ppm]. Full decomposition of compound 2a at pH 8 was observed after two hours.

Experimental section

The present examples are provided for illustrative purposes only and are not to be construed as limiting the scope of the present invention as defined by the appended claims.

In one embodiment a final product of the synthesis, the final product being a photolytically degradable compound, was obtained by first introducing at least one nitro substitution in at least one aromatic group of a reactant molecule to obtain a first photolytically degradable compound and then introducing an oxazolidine group in the first photolytically degradable compound to obtain the final product of the synthesis.

Results

I. Synthesis of model compounds:

The first photolytically degradable compound, i. e. an aminol (compound 1 ), was prepared using the procedure described below (Scheme 1 ). First, a Suzuki- Miyaura cross-coupling reaction with allyl boronic acid pinacol ester was used according to a method described by Kotha and co-workers (Kotha, S.; Behera, M.; Shah, V., A. Synlett 2005, 12, 1877-1880). Subsequent treatment with mCPBA gave the corresponding epoxide. A Lewis acid-promoted epoxide ring-opening reaction using 5 M lithium perchlorate-diethyl ether (LPDE) solution provided compound 1.

Scheme 1 : Synthesis of compound 1. Reagents and conditions: (i) 1 ) Pd(PPh ₃ ) ₄ , CsF, AllyIBpin, THF, reflux, 2) mCPBA, DCM, rt; (ii) 5 M LPDE, 40 °C; (iii) H2SO4, HNO ₃ , 0 °C

The final product of the synthesis (compound 2) was prepared from the corresponding ethanolamine-based compound (compound 1 ) by treating the latter with formaldehyde and formic acid in DCE at room temperature (Scheme 2).

Scheme 2. Formation of oxazolidine analogue 2 from ethanolamine 1

Using the same procedure as above, the appropriate ethanolamines could be converted into the corresponding oxazolidines in yields ranging from 88-91 % (Scheme 3).

CH ₂ O (37% aq.),

Scheme 3. Synthesis of model compounds containing the oxazolidine group By replacing formaldehyde with acetaldehyde in the reaction with ethanolamine it was possible to obtain the methyl substituted oxazolidine 2c in 90% yield (Scheme 4).

Scheme 4. Formation of methyl substituted oxazolidine 2c

II. Photolysis:

Subjecting compound 2 to photodecomposition in MeCN/water (7:3) at pH 8 resulted in full decomposition after two hours (Figure 1 ). The same treatment of the precursor compound (compound 1 ) resulted in full decomposition only after 24 h. Thus, compound 2 decomposed 12 times faster under light (laboratory conditions) than the corresponding ethanolamine-based compound 1.

Subjecting oxazolidine 2a to photodecomposition in MeCN/water (7:3) at pH 8 resulted in full decomposition after two hours (Figure 3).

UV-Vis analysis showed that the four compounds 2, 2a, 2b and 2c had a max of 304 nm, 414 nm, 291 nm and 466 nm, respectively (Figure 2).

III. Biological activity:

Compounds 2 and 12 were tested against both Gram-positive and Gramnegative bacteria, including E. faecalis, E. coli, P. aeruginosa, S. aureus, S. agalactiae and S. epidermidis. For S. agalactiae compound 2 showed a minimum inhibitory concentration of 25 pM, whereas its precursor compound (compound 1 ) showed a minimum inhibitory concentration of 6.3 pM. For S. agalactiae compound 12 showed a minimum inhibitory concentration of 75 pM, whereas its precursor compound (corresponding ethanolamine-based compound) showed a minimum inhibitory concentration of 12.5 pM.

The following tests can be conducted to evaluate the above-mentioned compounds for chitin synthetase inhibitor properties. This biological activity may result in death of pests containing a chitin exoskeleton. This includes pests on land and in water. Salmon lice (Lepeophtheirus salmonis) is used as a model organism for testing of chitin synthetase activity. All lice used for testing belongs to an established lice strain (LsGulen) and all information of breeding and maintenance of the lice is outlined by Hamre et al. (Parasitol. Int. 2009, 58, 451 -460).

General procedure for testing with the live-dead assay as follows. 5-30 nauplius I stage of the salmon lice are exposed for 1 h at 10 °C at concentrations 1 .0 ppt and 0.1 ppt according. The lice are subsequently transferred to incubators with a constant flow of seawater at 9 °C. After 7 days the number of surviving nauplii or copepodids are counted and their condition is subjectively evaluated.

Compounds that show activity in the live-dead assay and the corresponding photodecomposition mixtures are dissolved in DMSO to a concentration of 39-40 ppt. These samples are then diluted with seawater to a concentration of 0.1 ppt, 0.01 ppt, 0.001 ppt and 0.0001 ppt. Diflubenzurone samples prepared by the same method as described for the oxazolidines are used as positive control and DMSO (40 ppt) is used as negative control. All solutions are tested using the live-dead assay described above. An active compound would prevent formation of the exoskeleton when nauplii develop resulting in observation of dead nauplii and no copepodids.

Description of the general procedure for synthesis of model compounds

An appropriate aldehyde (4 equiv.) and formic acid (1.1 equiv.) in 1 ,2- dichloroethane (DCE) were added to a solution of amino alcohol (1 .0 equiv.). The resulting reaction mixture was then stirred under an argon at room temperature until completion of the reaction as determined by TLC analysis. The crude mixture was diluted by addition of water (5 mL) and the aq. layer was extracted using CH2CI2 (3 x 5 mL). The combined organic layers were washed with water (1 x 5 mL) and brine (1 x 5 mL), dried (MgSC ), filtered and evaporated onto celite and purified by silica gel column chromatography using the eluents as indicated. The below procedure provides the target compounds (Scheme 5).

Scheme 5: Synthesis of oxazolidine resembling compound Description of spectral analysis

IR spectra were recorded on an Agilent Cary 630 FT-IR spectrophotometer equipped with an attenuated total reflectance (ATR) attachment. Samples were analysed neat on a ZnSe crystal, and the absorption frequencies are given in wave numbers (cm ^-1 ).

UV-Vis spectra were obtained on an Agilent 8453 single-beam UV-Vis spectrophotometer with a deuterium-discharge lamp for the UV range and a tungsten lamp for the visible wavelength range. Samples were analysed in an Agilent open-top UV quartz cell (10 mm, 3.0 mL) with ethanol as solvent. Wavelengths are reported in nm, and molar attenuation coefficients in M’ ¹ crrr ¹ .

NMR spectra were recorded on a Broker Ascend™ 400 spectrometer (400.13 MHz for ¹ H, 100.61 MHz for ¹³ C, 376.46 MHz for ¹⁹ F) or a Broker Ascend™ 850 spectrometer (850.13 MHz for ¹ H and 213.77 MHz for ¹³ C). Coupling constants (J) are given in Hz and the multiplicity is reported as singlet (s), doublet (d), triplet (t), sextet (sxt), multiplet (m) and broad singlet (bs). The chemical shift values are reported upfield to downfield in ppm, and calibration is done using the residual solvent signals for chloroform-d ( ¹ H 7.26 ppm; ¹³ C 77.16 ppm) or acetonitrile-d3 ( ¹ H 1 .94 ppm; ¹³ C 1 .32 ppm). ²⁷ Calibration for ¹⁹ F NMR is done using a,a,a- trifluorotoluene as internal standard in chloroform-d (-62.61 ppm) and acetonitrile-d3 (-63.10 ppm). High-resolution mass spectra were obtained on a JEOL AccuTOF™ T100GC mass spectrometer operated in ESI mode. Low-resolution mass spectra were recorded on an Advion expression compact mass spectrometer (CMS) operated in ESI mode equipped with a Plate Express® TLC plate reader for sample injection. A solution of ammonium acetate (3.0 mM) and formic acid (0.05%) in acetonitrile and water (95/5) was used as mobile phase for both positive and negative ESI modes.

Thin-layer chromatography (TLC) was carried out with silica gel (60 F254) on aluminium sheets with solvent systems consisting of various mixtures of petroleum ether, ethyl acetate and DCM. Staining was achieved with either exposure to UV light (254 and/or 365 nm) or a potassium permanganate stain. Flash chromatography was performed with a hand pump and 230-400 mesh silica gel.

Description of the detailed procedure for synthesis of some model compounds and results of spectral analysis thereof 3-(2 , 5-D ich loro~4-( 1 , 1 ,2,3,3,3-hexafluoropropoxy)phenyl)-5-(4-nitrobenzyl)oxazolid ine

(Compound 2):

Following the general procedure, the title compound was prepared from 1- ((2,5-dichloro-4-(1 ,1 , 2,3,3, 3-hexafluoropropoxy)phenyl)amino)-3-(4- nitrophenyl)propan-2-ol (30.0 mg, 0.059 mmol), formaldehyde (37%, 0.020 mL, 0.24 mmol), formic acid (2.4 pL, 0.065 mL) and DCE (2 mL). After a reaction time of 23 hours, workup was carried out according to the general procedure and purification by silica gel column chromatography (pet. ether/CFhCk, 7:3 v/v) and concentration of the relevant fractions [Rf = 0.20 (pet. ether/CFhCk, 7:3 v/v)] gave the target compound as a pale oil (13.0 mg, 43%). Spectral data are presented below.

IR (ATR): Vmax 2931 , 2879, 1599, 1519, 1350, 1198, 1113 cm- ¹ ;

¹ H NMR (400 MHz, CD ₃ CN): 5 8.14 (d, J = 8.8 Hz, 2H), 7.49 (d, J = 8.7 Hz, 2H), 7.40 (t, J = 0.9 Hz, 1 H), 7.04 (s, 1 H), 5.56 (dsxt, J = 42.7 Hz, 5.8 Hz, 1 H), 4.91 (dd, J = 9.0 Hz, 4.8 Hz, 2H), 4.41 -4.35 (m, 1 H), 3.59 (dd, J = 10.2 Hz, 6.2 Hz, 1 H), 3.26 (dd, J = 10.2 Hz, 7.5 Hz, 1 H), 3.11 -3.02 (m, 2H);

¹³ C NMR (100 MHz, CD ₃ CN): 5 147.8, 147.2, 146.5, 138.7, 131.4, 127.4, 126.9, 124.3, 124.2, 123.7, 120.3, 85.4 (dq, JCF = 35.1 Hz), 83.6, 77.9, 55.4, 39.7 (one carbon was obscured or overlapping);

¹⁹ F NMR (376 MHz, CD ₃ CN): 5 -75.7-75.8 (q, J = 9.5 Hz, 3F), -78.3-78.9 (m, 1 F), -79.6-80.3 (m, 1 F), -213.4-213.5 (m, 1 F);

UV (EtOH): Amax 304 nm.

5-Benzyl-3-(4-nitrophenyl)oxazolidine (Compound 2a): Following the general procedure, the title compound was prepared from 1 -(4- nitrophenylamino)-3-phenylpropan-2-ol (100.0 mg, 0.37 mmol), formaldehyde (37%, 0.12 mL, 1 .47 mmol), formic acid (1 .5 pL, 0.41 mmol) and DCE (5 mL). After a reaction time of 20 hours, workup was carried out according to the general procedure to give the crude as a bright yellow solid (92.7 mg, 88%), which was essentially pure as shown by NMR, mp 125-127 °C. Spectral data are presented below.

IR (ATR): Vmax 3429, 3078, 2919, 2850, 1596, 908 cm’ ¹ ;

¹ H NMR (400 MHz, CDCI ₃ ): 6 8.09 (d, J = 9.2 Hz, 2H), 7.33-7.29 (m, 2H), 7.25-7.22 (m, 3H), 6.34 (d, J = 9.2 Hz, 2H), 5.06 (d, J = 3.0 Hz, 1 H), 4.88 (d, J = 2.9 Hz, 1 H), 4.52-4.45 (m, 1 H), 3.50 (dd, J = 8.9 Hz, 6.0 Hz, 1 H), 3.21-3.17 (m, 1 H), 3.12 (dd, J = 13.8 Hz, 6.7 Hz, 1 H), 2.94 (dd, J = 13.8 Hz, 6.5 Hz, 1 H);

¹³ C NMR (100 MHz, CDCI3): 6 149.2, 138.1 , 136.8, 129.2, 128.8, 127.1 , 126.5, 110.7, 80.2, 79.6, 50.5, 39.4;

UV (EtOH): Amax414 nm.

5-(4-Nitrobenzyl)-3-phenyloxazolidine (Compound 2b):

Following the general procedure, the title compound was prepared from 1 -(4- nitrophenyl)-3-(phenylamino)propan-2-ol (110.0 mg, 0.40 mmol), formaldehyde (37%, 0.13 mL, 1 .61 mmol), formic acid (0.02 mL, 0.44 mmol) and DCE (5 mL). After a reaction time of 2 days, workup was carried out according to the general procedure to give the crude as an orange solid (103.9 mg, 91 %), which was essentially pure as shown by NMR, mp 133-135 °C. Spectral data are presented below.

IR (ATR): Vmax 2912, 2832, 2751 , 1596, 1508, 1343, 1176, 855 cm’ ¹ ;

¹ H NMR (400 MHz, CDCI3): 6 8.19 (d, J = 8.8 Hz, 2H), 7.46 (d, J = 8.7 Hz, 2H), 7.25 (dd, J = 8.6 Hz, 7.4 Hz, 2H), 6.79 (tt, J = 8.2 Hz, 0.9 Hz, 1 H), 6.50 (dd, J = 8.7 Hz, 1 .0 Hz, 2H), 5.00 (d, J = 2.1 Hz, 1 H), 4.87 (d, J = 2.2 Hz, 1 H), 4.55-4.48 (m, 1 H), 3.52 (dd, J = 8.4 Hz, 6.1 Hz, 1 H), 3.18-3.13 (m, 2H), 3.08 (dd, J = 14.1 Hz, 5.3 Hz, 1 H); 1 ¹³ C NMR (100 MHz, CDCI3): 6 147.1 , 145.5, 145.5, 130.2, 129.5, 123.9, 117.9, 112.6, 81.3, 78.4, 51.1 , 39.7;

UV (EtOH): Amax 291 nm.

5-Benzyl-2-methyl-3-(4-nitrophenyl)oxazolidine (Compound 3):

Following the general procedure, the title compound was prepared from 1 -(4- nitrophenylamino)-3-phenylpropan-2-ol (100.0 mg, 0.37 mmol), acetaldehyde (0.083 mL, 1 .48 mmol), formic acid (1 .5 pL, 0.41 mmol) and DCE (5 mL). After a reaction time of 20 hours, workup was carried out according to the general procedure to give the crude as a bright yellow solid (99.7 mg, 90%), which was essentially pure as shown by NMR, mp 116-118 °C. Spectral data are presented below.

IR (ATR): Vmax3068, 3026, 2979, 2921 , 2883, 1595, 1481 , 1309, 746 cm’ ¹ ;

¹ H NMR (400 MHz, CDCI3): 6 8.14-8.09 (m, 4H), 7.35-7.32 (m, 4H), 7.29-7.24 (m, 6H), 6.45 (d, J = 9.2 Hz, 2H _mi nor), 6.43 (d, J = 9.4 Hz, 2H _ma jor), 5.48 (q, J = 5.4 Hz, 1 H major), 5.35 (q, J = 5.1 Hz, 1 Hminor), 4.72-4.65 (m, 1 H major), 4.35-4.28 (m, 1 Hminor), 3.60-3.53 (m, 2H), 3.39-3.30 (m, 1 H), 3.21 -3.09 (m, 3H), 2.95 (dd, J = 13.8 Hz, 6.7 Hz, 1 H), 2.87 (dd, J = 13.8 Hz, 7.0 Hz, 1 H), 1 .49 (d, J = 5.1 Hz, 3H _mi nor), 1 .45 (d, J =

5.5 Hz, 3Hmajor);

¹³ C NMR (100 MHz, CDCI3): 6 149.6, 149.3, 137.9, 137.9, 136.9, 136.9, 129.3, 129.2, 128.8, 128.8, 127.0, 126.4, 126.2, 111.6, 110.9, 87.2, 87.0, 77.8, 76.6, 52.2, 51.1 , 39.8, 39.6, 20.3, 19.5;

UV (EtOH): Amax 466 nm.

3-(2-bromo-4-(trifluoromethoxy)phenyl)-5-(4-nitrobenzyl)o xazolidine

Formaldehyde (0.137 mL, 4.0 eq., 1 .84 mmol) and formic acid (0.019 mL, 1.1 eq., 0.51 mmol) in 1 ,2-dichloroethane (9.2 mL, 0.05 M) were added to a solution of 1- ((2-bromo-4-(trifluoromethoxy)phenyl)amino)-3-(4-nitrophenyl )propan-2-ol (0.2 g, 1.0 eq., 0.46 mmol). The resulting mixture was stirred at room temperature for 2 days. The crude product was diluted with water (20 mL), and the aqueous layer was extracted using CH2CI2 (40 mL x 3). The combined organic layers were washed with water (20 mL), brine, dried over MgSC and concentrated under reduced pressure. The crude was purified by silica-gel column chromatography (pet. ether/EtOAc + 3% EtsN, PE 100-60:40-50:50-45:55-40:60-50:50, Rf. 0.54) to provide the title compound (0.45 g, 53%) as a sticky yellow oil.

IR: Vmax (film) 2924 (C-H, aromatic), 1516, 1343 (C-H, aromatic), 1248, 1213 (C-O-C), 1030 (C-Br);

¹ H N MR (400 MHz, CDCI3): 5 2.95-3.09 (2H, m), 3.21 -3.29 (1 H, m), 3.56- 3.63 (1 H, m), 4.27-4.36 (1 H, m), 4.87 (1 H, d, J = 5.8 Hz), 4.92 (1 H, d, J = 5.8 Hz), 7.02-7.08 (1 H, m), 7.08-7.12 (1 H, m), 7.39-7.47 (3H, m), 8.15 (1 H, d, J = 8.9 Hz);

¹³ C NMR (100 MHz, CDCI3): 5 40.1 , 56.9, 76.1 , 84.0, 117.6, 119.2, 120.4, 120.9, 121.8, 126.9, 130.2, 144.5, 144.5, 145.7, 147.0; ¹⁹ F{ ¹ H} NMR (470 MHz, CDCI3) 5 -58.6.

3-(6-bromo-2,3,4-trifluorophenyl)-5-(4-nitrobenzyl)oxazol idine

Formaldehyde (0.139 mL, 4.0 eq., 1 .88 mmol) and formic acid (0.020 mL, 1.1 eq., 0.52 mmol) in 1 ,2-dichloroethane (9.4 mL, 0.05 M) were added to a solution of 1 - ((6-bromo-2,3,4-trifluorophenyl)amino)-3-(4-nitrophenyl)prop an-2-ol (0.19 g, 1.0 eq., 0.47 mmol). The resulting mixture was stirred at room temperature for 2 days. The crude product was diluted with water (20 mL), and the aqueous layer was extracted using CH2CI2 (40 mL x 3). The combined organic layers were washed with water (20 mL), brine, dried over MgSC and concentrated under reduced pressure. The crude was purified by silica-gel column chromatography (pet. ether/EtOAc + 3% EtsN, PE 100-60:40-50:50-45:55-40:60-50:50, Rf. 0.70) to provide the title compound (81.6 mg, 42%) as a colourless oil.

IR: Vmax (film) 1927 (C-H, aromatic ), 1501 (N-O, nitro), 1201-1179 (C-O-C), 1343 (C-F), 654 (C-Br);

¹ H NMR (400 MHz, CDCI3): 6 2.98-3.16 (2H, m), 3.18-3.22 (1 H, m), 3.57-3.64 (1 H, m), 4.38-4.47 (1 H, m), 4.71 -4.74 (1 H, m), 4.84-4.86 (1 H, m), 7.22-7.29 (1 H, m), 7.45 (1 H, d, J = 9.1 Hz), 8.16 (1 H, d, J = 9.1 Hz);

¹³ C NMR (100 MHz, CDCI3): 6 39.9, 55.8, 84.1 , 93.2, 115.9, 116.0, 123.8,

130.3, 132.3, 138.6, 141.3, 145.9, 147.2; ¹⁹ F{ ¹ H} NMR (470 MHz,CDCI ₃ ) 6 -149.7, -

161.3.

3-(3-chloro-4-nitrophenyl)-5-(4-chlorobenzyl)oxazolidine

Formaldehyde (4 eq) and formic acid (1.1 eq.) in 1 ,2-dichloroethane (DCE) were added to a solution of amino alcohol (1 eq.). The resulting mixture was stirred under an N2 atmosphere at room temperature for 2 days. The crude product was diluted with water (5 mL), and the aqueous layer was extracted using DCM (3 x 5 mL). The combined organic layers were washed with brine (5 mL) and water (5 mL), dried over MgSC , and concentrated under reduce pressure. The crude product was pure based on ¹ H NMR analysis. The target compound was obtained as a sticky yellow oil (37.1 mg, 50%). IR: Vmax = 3092 (C-H, alkene), 2921 (C-H, alkane), 1900 (C-H, aromatic), 1594 (N-O, nitro), 1303-1264 (C-O-C), 802 (C-CI);

¹ H NMR (400 MHz, CDCI ₃ ): 6 8.04 (m, 1 H), 7.98 (m, 2H), 7.42 (m, 2H), 7.32 (m, 2H), 5.55-5.24 (m, 2H), 4.48 (m, 1 H), 3.97-85 (m, 2H), 2.76 (m, 2H).