EP4294188A1

EP4294188A1 - Use of cationic porphyrins as selective herbicide

Info

Publication number: EP4294188A1
Application number: EP22710652.3A
Authority: EP
Inventors: Mohamad ISSAOUI; Catherine Riou; Vincent SOL
Original assignee: Universite de Limoges
Current assignee: Universite de Limoges
Priority date: 2021-02-18
Filing date: 2022-02-17
Publication date: 2023-12-27
Also published as: US20240156098A1; WO2022175633A1; FR3119732B1; FR3119732A1

Abstract

The present invention relates to the use of cationic porphyrins, such as zinc-tetra (N-methylpyridyl) porphyrin tetrachloride and tetra (N-methylpyridyl) porphyrin tetrachloride or a mixture thereof, as a selective foliar herbicidal agent.

Description

Title: Use of cationic porphyrins as a selective weedkiller Technical area

The present invention relates to the use of cationic porphyrins such as zinc-tetra (N-methylpyridyl) porphyrin tetrachloride and tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture, as a selective foliar herbicidal agent.

Prior technique

[0002] Photosensitizers (PS) are molecules capable of inducing, thanks to a light stimulus and in the presence of oxygen, the formation of reactive oxygen species (ROS) which are more or less toxic for living cells, according to their physicochemical properties, the concentrations used as well as the nature of the cells and biological organisms. There are two mechanisms of production of EROs:

- The type I mechanism, which involves a transfer of electrons from the photosensitizer to biological substrates to form free radicals (superoxide anion (V, hydroxyl radical -OH and hydrogen peroxide H ₂ 0 ₂ );

- The type II mechanism which involves the transfer of energy to molecular oxygen, which passes from its ground state (triplet state: ³ 0 ₂ ) to the singlet state ¹ 0 ₂ .

[0003] These photosensitizers are mainly used in the medical field, in the context of photodynamic therapy in the treatment of cancer, but also for the fight against microorganisms such as bacteria and fungi, as disinfection therapy or for decontamination. water and food. In general, ROS are deleterious to all living cells.

[0004] These photosensitizers have therefore been considered for application as biocides in the agricultural field, and more particularly against crop pathogens such as bacteria and fungi, which cause reductions in agricultural yield. . The use of these photosensitizers as a biocide is also called antimicrobial photodynamic treatment.

[0005] However, and although necessary, studies evaluating the undesirable effects of photosensitizers on plants are rare. Moreover, the effects of molecules on plants are very difficult to transpose between different species. Typically, natural photosensitizers (coumarins and furocoumarins) have been evaluated as biocides on orange and strawberry trees. While orange leaves were unaffected by photosensitizers, strawberry leaves were damaged by coumarins. Cationic water-soluble porphyrins (zinc-tetra (N-methylpyridyl) porphyrin tetrachloride (Zn-TMPyP) and tetra (N-methylpyridyl) porphyrin tetrachloride (TMPyP)) have also been applied to tomato seedlings (Solanum lycopersicum ) grown in vitro under a photoperiod of 16 hours. Cationic porphyrins have been shown to significantly alter seedling growth in vitro. However, the authors suggest that these photosensitizers could be used as biocides against tomato pathogens (Guillaumot et al., Synergistic enhancement of tolerance mechanisms in response to photoactivation of cationic tetra (N-methylpyridyl) porphyrins in tomato plantlets , Journal of Photochemistry and Photobiology B: Biology, Volume 156, March 2016, Pages 69-78).

[0007] However, these same cationic porphyrins, also tested in vitro on Arabidopsis (Arabidopsis thaliana) seedlings, completely killed the plant at a very low dose (3.5 mM) (Issawi et al., Responses of an adventitious fast-growing plant to photodynamic stress: comparative study of anionic and cationic porphyrin effect on Arabidopsis thaliana, Physiologia Plantarum 162: 379-390. 2018).

[0008] Whereas the in vitro root effect of photosensitizers on plants is unpredictable and difficult to extrapolate to field or greenhouse conditions, and whereas cationic porphyrins such as Zn-TMPyP applied in vitro at best inhibit the growth of the plants tested , at worst destroy them, the inventors have, unexpectedly, demonstrated that the cationic porphyrins and preferentially TMPyP and Zn-TMPyP are selective weedkillers when they are applied by foliar spraying.

Summary

The present invention therefore relates to the use of cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride (Zn-TMPyP), tetra (N-methylpyridyl) porphyrin tetrachloride (TMPyP) or their mixture, in as a selective foliar herbicide.

[0010] The inventors have indeed advantageously demonstrated that Zn-TMPyP can be used as a selective foliar herbicide. While Zn-TMPyP significantly inhibits the development of plants (weeds and plants of agronomic interest) when it is applied to the roots of plants in vitro, the inventors have demonstrated that Zn-TMPyP preserves plants of agronomic interest and n does not affect their development when Zn-TMPyP is applied by foliar spray, typically in a greenhouse. On the other hand, this same process (foliar application of Zn-TMPyP in a greenhouse) leads to the death of weeds. Zn-TMPyP is therefore a selective foliar herbicide which helps to control weeds while preserving crops of agronomic interest. [0011] Furthermore, TMPyP and Zn-TMPyP are advantageously photodegradable, thus making it possible to limit their long-term effect. Their non-toxicity has also been demonstrated towards fishing worms (maggots).

[0012] Still advantageously, TMPyP and Zn-TMPyP are effective at low concentrations. Typically, glyphosate, which remains the most widely used herbicide in the world, is used at extremely high concentrations (about 360g/L). The herbicidal compounds according to the invention are for their part used at concentrations of the order of a micromolar (mM).

[0013] These molecules therefore constitute an alternative to the herbicides present on the market, which are more respectful of the environment and have a reduced impact on human health and animal and plant biodiversity.

The present invention also relates to a selective weed control method consisting of the application of cationic porphyrins chosen from TMPyP, Zn-TMPyP or their mixture, said porphyrins being applied by leaf spraying.

Brief description of the drawings

[0015] Other characteristics, details and advantages will appear on reading the detailed description below, and on analyzing the appended drawings, in which:

Fig. 1

[0016] [Fig. 1] shows the in vitro biocidal effect of 3.5mM Zn-TMPyP on plants of agronomic interest (barley, tomato, potato, tobacco and grapevine).

Fig. 2

[0017] [Fig. 2] shows the in vitro biocidal effect of 3.5mM Zn-TMPyP on weeds (nettle, arabidopsis and poppy).

Fig. 3

[0018] [Fig. 3] shows the selective foliar herbicidal effect of Zn-TMPyP in greenhouse on two crop systems associated with their weeds (barley/poppy and tomato/millet) after two months of treatment with either 50mM Zn-TMPyP or water ( control culture). It should be noted that the poppy is completely destroyed.

Fig. 4

[0019] [Fig. 4] is a comparison demonstrating the in vitro biocidal effect of 3.5mM of Zn-TMPyP on 22-day-old tomato seedlings, and the absence of effect of 50mM of Zn-TMPyP on 22-day-old greenhouse tomato seedlings. days compared to their respective control media. Fig. 5

[0020] [Fig. 5] is a comparison demonstrating the in vitro biocidal effect of 3.5mM of Zn-TMPyP on 17-day-old barley seedlings, and the absence of effect of 50mM of Zn-TMPyP on barley seedlings in greenhouses 22 days compared to their respective control media. Fig. 6

[0021] [Fig. 6] is a comparison demonstrating the in vitro biocidal effect of 3.5mM of Zn-TMPyP on 22-day-old poppy seedlings, and the herbicidal effect of 50mM of Zn-TMPyP on poppy seedlings in greenhouses compared to their environments respective witnesses.

Fig. 7 [0022] [Fig. 7] shows the harmlessness of 50mM Zn-TMPyP on the viability of maggots in the presence and in the absence of light 24 hours after the spray treatment with 50mM Zn-TMPyP.

Fig. 8

[0023] [Fig. 8] corresponds to monitoring the photostability/photodegradation of Zn-TMPyP at 50 mM with or without surfactant in water under photoperiod (16 h light/8 h dark) or in the dark.

Fig. 9

[0024] [Fig. 9] corresponds to the dosage of total thiols as well as enzymatic activities (catalase and guaiacol peroxidase) on tomato seedlings cultivated for 1 month in vitro on the medium rich in Zn-TMPyP 3.5 mM.

Fig. 10

[0025] [Fig. 10] corresponds to the dosage of total thiols as well as enzymatic activities (catalase and guaiacol peroxidase) on tomato seedlings grown for two months in a greenhouse, after foliar application of 50 mM of Zn-TMPyP. Fig. 11

[0026] [Fig. 11] is a schematic comparison of the results presented in FIGS. 8 and 9 demonstrating, in particular, that the in vitro results cannot be extrapolated to the greenhouse results.

Fig. 12 [0027] [Fig. 12] shows the herbicidal effect of Zn-TMPyP against weeds at different concentrations (C1, C2, C3) compared to the control (T). Fig. 13

[0028] [Fig. 13] presents the safety of Zn-TMPyP against the plants of interest at different concentrations (C1, C2, C3) compared to the control (T).

Detailed Description The present invention therefore relates to the use of cationic porphyrins chosen from among zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture as a selective foliar herbicide.

[0030] Cationic porphyrins

The porphyrins used according to the invention are tetra (N-methylpyridyl) porphyrin tetrachloride (TMPyP or T4MPyP) of formula (I):

[0032] [chem 1]

(formula I) and zinc-tetra (N-methylpyridyl) porphyrin tetrachloride named Zn-TMPyP or Zn-T4MPyP of formula II, where X is a Zinc atom:

[chem 2]

(Form II).

Zn-TMPyP is a cationic porphyrin metallated at the level of the tetrapyrrole ring.

Zn-TMPyP and TMPyP are marketed by the company Frontier Scientific (Carnforth, UK).

[0035] According to a preferred embodiment, Zn-TMPyP is used as a selective foliar herbicide. [0036] Culture space

By culture or culture space is meant any space for cultivating plants and more particularly plants of agronomic interest. Mention will be made, by way of illustration, of a greenhouse, a field, a meadow, a courtyard, an alley, a garden, a vegetable patch, etc.

According to one embodiment, the growing space is chosen from among a greenhouse, a field, a meadow, a courtyard, an alley, a garden, a vegetable patch.

[0039] Selective foliar herbicide

A herbicide is a pesticide for agricultural use whose activity on the metabolism of plants leads to their death.

The cationic porphyrins according to the invention are selective herbicides when they are applied by foliar spraying.

[0042] Indeed, and contrary to the in vitro results demonstrating the biocidal effect of Zn-TMPyP, the inventors have advantageously demonstrated that Zn-TMPyP and TMPyP can be used as a selective herbicide when they are applied by foliar spraying. The inventors have in fact proved that Zn-TMPyP sprayed in a greenhouse causes the death of weeds and preserves the plants of agronomic interest. Plants of interest such as barley and tomato have been shown to grow perfectly whether under the condition "control" (spray of H ₂ 0) or following their treatment (foliar sprays of 50 mM Zn-TMPyP). Conversely, poppy and millet died completely following foliar treatment with Zn-TMPyP. The first symptoms appear 24 hours after the first treatment. After 48 hours, the poppy is completely destroyed. These results are unexpected with regard to the in vitro effects: whereas the growth of barley and tomato was drastically altered in vitro by 3.5 mM of Zn-TMPyP in root application, the application by foliar spray of 50 mM of Zn-TMPyP had no effect on the growth of plants of agronomic interest. On the other hand, the same dose of Zn-TMPyP leads to the death of weeds, testifying to the selective herbicidal effect when Zn-TMPyP is applied by foliar spraying.

Thus, Zn-TMPyP and TMPyP are advantageously selective foliar herbicides, preferentially when they are applied in a greenhouse.

[0045] A foliar herbicide is applied by spraying on the leaves and requires an adjuvant to help the active principle to penetrate inside the cells (basipetal translocation). Conversely, in the case of a root herbicide, the active principle is absorbed by the roots and carried by the sap (acropetic translocation).

[0046] Thus, and unexpectedly, while Zn-TMPyP has been shown to be a root herbicide in vitro, it is advantageously selective when applied by foliar spray. A selective herbicide is a herbicide which aims to eliminate weeds without damaging the growing space such as a greenhouse, a field, a meadow, a yard, an alley, a garden. It is therefore a question of fighting against weeds or “weeds” which are undesirable in crops. Advantageously, the cationic porphyrins according to the invention will destroy the weeds without damaging the plants of agronomic interest. The term “weed” is understood to mean any herbaceous or ligneous plant which is found in an agroecosystem without having been intentionally established there. For example, it will be an undesirable plant species present in a space where another plant species is grown.

According to one embodiment, the plants of agronomic interest will be chosen from plants of the genus Nicotiana, of the genus Solanum, of the genus Vitis, of the genus Hordeaum, of the genus Triticum, of the genus Zeya, of the genus Helianthus, of the genus Allium, genus Spinacia, genus Frugaria, genus Brassica.

The plants of agronomic interest will be chosen from tobacco (Nicotiana tabacum), potato (Solanum tuberosum), vine (Vitis vinifera), barley (Hordeum vulgare), tomato (Solanum lycopersicum), wheat (Triticum sativum), maize (Zeya mays), sunflower (Helianthus annuus), onion (Allium cepa), spinach (Spinacia oleracea), strawberry (Frugaria sp.) and turnip (Brassica rapa).

Thus, according to one embodiment, Zn-TMPyP and TMPyP are used for the treatment of crops chosen from tobacco crops (Nicotiana tabacum), potato (Solanum tuberosum), vine (Vitis vinifera), barley (Hordeum vulgare), tomato (Solanum lycopersicum), wheat (Triticum sativum), corn (Zeya mays), sunflower (Helianthus annuus), onion (Allium cepa), spinach (Spinacia oleracea ), strawberry ( Frugaria sp.) and turnip ( Brassica rapa).

According to a preferred embodiment, Zn-TMPyP and TMPyP, preferentially -Zn-TMPyP are used for the treatment of crops chosen from wheat (Triticum sativum), barley (Hordeum vulgare), maize ( Zeya mays), sunflower (Helianthus annuus), turnip (Brassica rapa), tomato (Solanum lycopersicum), onion (Allium cepa), spinach (Spinacia oleracea).

Zn-TMPyP and TMPyP will be used against weeds chosen from weeds of the genus Urtica, the genus Echinochloa, the genus Papaver, the genus Lolium, the genus Datura, the genus Stellaria, the genus Chenopodium, the genus Panicum, genus Taraxacum, genus Rumex, genus Solanum.

According to one embodiment, Zn-TMPyP and TMPyP will be used against weeds chosen from nettle (Urtica dioi ^' ca), millet (Echinochloa frumentacea), poppy (Papaver rhoeas), ray Italian grass (Lolium multiforum), datura (Datura stramonium), white pimpernel (Stellaria media), lamb's-quarters (Chenopodium album), false millet (Panicum miliaceum), dandelion (Taraxacum officinale), rumex ( Rumex crispus), black moth (Solanum nigrum).

According to one embodiment, Zn-TMPyP and TMPyP, and preferably Zn-TMPyP, will be used against weeds chosen from black nightshade (Solanum nigrum), dandelion (Taraxacum officinale), lamb's-quarters ( Chenopodium album), dock ( Rumex crispus), Italian ryegrass ( Lolium multiforum), false millet ( Panicum miliaceum), white pimpernel ( Stellaria media), millet ( Echinochloa frumentacea), poppy (Papaver rhoeas ). [0056] Selective herbicidal composition

The present invention also relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least one surfactant. According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being at least 40mg/L.

According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being approximately 44mg/L.

According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being at least 60mg/L.

According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being approximately 66mg/L.

According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being at least 80mg/L.

According to one embodiment, the invention relates to a selective herbicidal composition comprising cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture and at least a surfactant the content of cationic porphyrins being approximately 88mg/L.

[0064] A person skilled in the art is able to determine the concentration to be applied as well as the frequency depending on the nature of the crops, the surface of the field, the weather conditions, etc.

[0065] Typically, the surfactant will be chosen from neutral and nonionic surfactants such as polyoxyethylene nonylphenylether, anionic surfactants such as products marketed under the name clasofl®, texapon®, cationic surfactants such as benzalkonium chloride (BAC) or Benzethonium chloride (BZT), neutral zwitterionic surfactants,

Preferably, the surfactant is polyoxyethylene nonylphenylether (Igepal CO-630, Sigma Aldrich).

According to one embodiment, the selective herbicidal composition comprises zinc-tetra (N-methylpyridyl) porphyrin tetrachloride and polyoxyethylene nonylphenylether.

[0068] Weeding process

The present invention also relates to a selective weed control process comprising at least one step of applying cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture in a growing space, said porphyrins being applied by foliar spraying.

[0070] Preferably, the growing space is chosen from among a greenhouse, a field, a meadow, a courtyard, an alley, a garden, a vegetable patch, etc.

The term “foliar spraying” means the application of the porphyrins according to the invention to the aerial parts of weeds and plants of agronomic interest in cultivation, and preferentially the application to the leaves of weeds and plants of agronomic interest in cultivation

According to one embodiment of the weed control method, TMPyP and Zn-TMPyP are sprayed at the cotyledon stage and/or at the first leaf stage such as the 2-leaf and/or 4-leaf stage.

The term “cotyledonary stage” or “cotyledonary stage” means the stage of emergence of the embryonic leaves.

[0074] The term "first leaf stage" means the stage of development of the true leaves.

[0075] Selective herbicidal method

The invention also relates to a method for selective weed control, the method comprising the application to a culture space of cationic porphyrins chosen from among zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or mixtures thereof, said porphyrins being applied by foliar spraying. [0077] Preferably, the growing space is chosen from among a greenhouse, a field, a meadow, a courtyard, an alley, a garden, a vegetable patch, etc.

According to one embodiment of the weed control method, TMPyP and Zn-TMPyP are sprayed at the cotyledon stage and/or at the 2-leaf and/or 4-leaf stage. Examples

[0079] Materials and Methods

In the examples which follow, the materials and methods described below were used.

[0081] Cationic porphyrins [0082] The cationic porphyrin metallated at the level of the tetrapyrrole nucleus used is the

Zinc-tetra (N-methylpyridyl) porphyrin tetrachloride named Zn-TMPyP (marketed by Frontier Scientific (Carnforth, UK)). The stock solution, prepared at 1 mM in water, is stored in the dark at room temperature.

[0083] Formulation for foliar spraying 50 mM Zn-TMPyP dissolved in water + 0.1% v/v of a neutral and non-ionic surfactant

(Polyoxyethylene nonylphenylether) (Igepal 00-630, Sigma Aldrich).

[0084] Photostability / photodegradation test

The photostability/photodegradation of Zn-TMPyP was tested in water under a photoperiod of 16 h and in the dark with or without surfactant. Zn-TMPyP mixed with 0.1% v/v of surfactant is tested at 50 mM under photoperiod and in the dark by monitoring the absorption of the Soret band by UV-visible spectrometry. Monitoring the intensity of the Soret band as a function of irradiation time makes it possible to calculate the percentage of degraded porphyrins.

[0086] Plant models [0087] Nine plant species belonging to several large families were tested according to the conditions described below:

[0088] [Table 1]

[0089] Sterilization of spiders/carvopses

The seeds are sterilized according to the following protocol. Surface sterilization is first performed with 70% (v/v) ethanol for 2 minutes at room temperature. The ethanol is removed and the seeds are immersed in 20% (v/v) sodium hypochlorite for 15-20 minutes then rinsed thoroughly with sterile water. The barley caryopsis are left for at least one hour in water before culturing.

[0091] Seed germination

The germination medium for all the seeds is Gamborg's synthetic medium also called B5 (Duchefa Biochemie, Haarlem, Netherlands) supplemented with 2% (m/v) sucrose and solidified with 0.8% agar. (Difco, Dallas, USA). The pH is adjusted to 5.8 with 1 M sodium hydroxide before autoclaving. After autoclaving (120° C., 20 minutes, 1 bar) and cooling of the medium in Petri dishes and/or jars, porphyrin is added at the desired concentration. After sterilization, the seeds are deposited on the solidified B5 medium, which may or may not contain the porphyrin, under the laminar flow hood. The Petri dishes like the jars are placed in a culture chamber (photoperiod 16 h, 22°C, photon flux density -100 pmol.m-2.s-1 generated by cool daylight lamps (OSRAM Lumilux 24W)) . The jars are used for the transfer of the seedlings and allow them a more appropriate growth.

[0093] In Vitro Multiplication of the Clones [0094] The explants are multiplied in a sterile manner from seedlings 1 month old for the potato and two months old for the vines. Potato explants are multiplied on a synthetic Murashige and Skoog medium supplemented with 2% sucrose and myo-inositol (100 mg/L) pH 5.8. The vine explants are grown on ½ Chée and Pool medium supplemented with 2% sucrose, pH 5.9. The media are solidified with 0.8% Sobigel. Both media are supplied by Duchefa Biochemie, Haarlem, Netherlands. The media are autoclaved as previously described for the B5 medium. The vine explants are placed in a culture chamber at 25° C and those of potatoes at 23° C, same photoperiod and light intensity as for seed germination.

[0095] Cultivation in a greenhouse

[0096] The various seeds are cultured in a greenhouse in commercial soil (Universal soil, Fertiligene). The seeds were sown in the greenhouse under controlled temperature and humidity conditions (The temperature varies between 15°C and 30°C depending on the season, and the relative humidity varies from 50 to 70%.).

[0097] Two spot treatments (approximately 10 sprays each) spaced 48 hours apart were carried out one week after germination. The first treatment was carried out when the seedlings were at the cotyledon stage, the second when the seedlings were at the first leaf stage.

[0098] After two months, the seedlings were harvested. The roots and aerial parts were separated and the roots were washed with distilled water and dried on absorbent paper. These materials were then stored at -20°C until they were used for the various biochemical analyses.

[0099] Biochemical tests

[0100] Determination of total thiols

The seedlings (roots and aerial apparatus separated before freezing) are ground in liquid nitrogen. The powder is transferred to an Eppendorf tube and weighed (between 200 and 100 mg per extraction). The extract is acidified by 0.2 N HCL, vortexed and centrifuged

(13000g 10min 4°C). 500 pL of the supernatant is neutralized with 400 pL NaOH (0.2 N) and 50 pL Na ₂ HP0 ₄ . Then, 700 μL Na ₂ HP0 ₄ 0.12 M, 6 mM EDTA and 6 mM DTNB are added to 200 μL of the neutralized solution. After establishing a glutathione range, the total thiols contained in the samples are determined by UV-visible spectrometry at 412 nm and related to the fresh mass.

[0102] Enzymatic activities

The seedlings (roots and aerial apparatus separated before freezing) are ground in liquid nitrogen. The powder is transferred into an Eppendorf tube in which 50 mg of PolyVynil PolyPyrrolidone (PVPP) (Sigma) have been weighed beforehand. The PVPP eliminates the polyphenols present in the organs that could interfere with biochemical assays. The homogenate is taken in extraction buffer (25 mM phosphate buffer pH 7.8, 10% (v/v) glycerol, and 1 mM EDTA). The tubes are then centrifuged for 20 minutes at 14,000 rpm at 4°C and the supernatants are stored at 4°C for the protein assay or stored at -20°C. For the protein assay, we use the Bradford assay method (Bradford, 1976) and BSA for the standard range.

[0104] Determination of catalase activity (CAT)

Catalase is an enzyme of the oxidoreductase family. Catalase activity can be quantified by measuring the disappearance of GH ₂ 0 ₂ by dismutation into H ₂ 0 and 0 ₂ . To determine this activity, we use the protocol carried out in the publication of Issawi. et al. (2018). The kinetics of H ₂ 0 ₂ disappearance is measured for 200 seconds every 20 seconds at 240 nm. The activity is calculated using the molar extinction coefficient of H ₂ 0 ₂ which is 43.6 rnM Lcnr ¹ [0106] Determination of the activity of guaiacol peroxidase (GPOX)

Guaiacol peroxidase is an enzyme present only in plants. It detoxifies the plant of oxygen peroxide molecules produced by the fungus. Its activity is measured by the oxidation of guaiacol in the presence of H ₂ 0 ₂ . To determine this activity, we use the protocol described in the publication by Issawi. et al. (2018). The kinetics of appearance of oxidized guaiacol is measured for 120 seconds every 10 seconds at 436 nm. The activity is calculated using the molar extinction coefficient of H ₂ 0 ₂ which is 25.5 mM Lcnr ¹ .

[0108] The biochemical tests were carried out on 3 independent experiments and statistically validated using the PAST software

Example 1: Demonstration of the selective foliar herbicidal effect of Zn-TMPyP vs. the in vitro biocidal effect of Zn-TMPvP

[0110] A. Biocidal effect in vitro: case of plants of agronomic interest [0111] The results are presented in Figure 1.

To avoid mechanical stresses, the results presented below concerning barley and tomato are results after 17 and 22 days respectively. Indeed, the seedlings will grow rapidly on the control media (without Zn-TMPyP) and touch the lid of the jars compared to the other cultures.

[0113] Barley

It clearly appears that the growth of barley is altered after one month of culture on a medium rich in Zn-TMPyP at 3.5 mM. In addition, the leaves show yellowing and subsequently wilting. On the other hand, in the control medium, the barley grows normally and shows rapid and perfect growth after 17 days. [0115] Tomato

In accordance with the results of the state of the art (Guillaumot et al.), the tomato (Marmande variety) shows significantly inhibited development after 22 days on the medium rich in Zn-TMPyP at 3.5 mM compared to the condition " witness ". [0117] Potato

The potato (Coquine variety) shows a slight delay in growth at the level of the stem and the roots on the treated medium compared to the seedlings of the control condition.

[0129] Tobacco and vine [0120] The tobacco and vine seedlings show considerable inhibition of their development after respectively one and a half and two months of culture on a medium rich in Zn-TMPyP at 3.5 mM ( Figure 1).

[0121] Conclusion: in vitro biocidal effect of Zn-TMPyP on plants of agronomic interest [0122] After treatment with Zn-TMPyP, plants of agronomic interest exhibit at best a delay in growth. In the majority of cases, the growth of these plants is inhibited by Zn-TMPyP, compared to the control medium. It thus appears very clearly that Zn-TMPyP has a biocidal effect in vitro on plants of agronomic interest.

[0123] B. Biocidal effect in vitro: case of weeds [0124] The results are presented in Figure 2.

[0125] Nettle and Arabidopsis

The nettle and arabidopsis show a very significant inhibition of growth on a medium treated with Zn-TMPyP (3.5 mM) after 18 days and 1 month respectively.

[0127] Poppy [0128] For the poppy, the seedlings show different responses which testify to a remarkable alteration and inhibition after 6 weeks.

[0129] Conclusion: in vitro biocidal effect of Zn-TMPyP on weeds [0130] In accordance with the results obtained on plants of agronomic interest, the growth of weeds is inhibited by Zn-TMPyP, testifying to its biocidal effect in vitro.

The in vitro results testify to a root herbicidal effect of Zn-TMPyP.

[0132] C. Selective foliar herbicide effect: greenhouse trials [0133] The results are shown in Figures 3 to 6.

[0134] Tests on two cropping systems associated with their weeds were carried out:

- barley (grass) with poppy (dicot)

- tomato (dicot) with millet (grass) with a spray of 50 mM of Zn-TMPyP in two stages spaced 48 hours apart one week after germination. Seedlings were monitored every 48 h. The control medium was sprayed with water at the same frequency.

The results after one month and two months show that the plants of interest grow perfectly whether in the "control" condition or following their treatment (approximately 10 foliar sprays) with 50 mM of Zn-TMPyP . Conversely, poppy and millet died completely following their treatment. The first symptoms appear 24 hours after the first treatment and the poppy is destroyed after 48 hours after the first treatment (first 10 sprays) Whereas the growth of barley and tomato was drastically altered in vitro by 3.5mM Zn-TMPyP, foliar spray application of 50 mM Zn-TMPyP had no effect on the growth of plants of agronomic interest. On the other hand, the same dose of Zn-TMPyP leads to the death of weeds testifying to the selective herbicidal effect when Zn-TMPyP is applied by foliar spraying. These results testify to the selective foliar herbicidal effect of Zn-TMPyP.

These tests also confirm that the results obtained in vitro are not necessarily and automatically transposable to the greenhouse scale. Indeed, barley and tomato show significantly inhibited development in vitro. On the other hand, no visible effect is noticed on these greenhouse crops. On the other hand, the poppy is completely destroyed following foliar spraying in the greenhouse, while we observe different responses from the seedlings cultivated in vitro: some seedlings present a significant inhibition of growth and others show a delay in growth. growth.

D. Mechanism of Action

The total thiols, as well as the enzymatic activity (catalase activity and guaicol peroxidase activity) were determined on seedlings cultured in vitro on the medium rich in Zn-TMPyP 3.5 mM (FIG. 9) and on seedlings subjected to 50 mM Zn-TMPyP per foliar application (Figure 10).

The antioxidant machinery is significantly inhibited in the seedlings cultivated on the medium rich in Zn-TMPyP 3.5 mM. On the other hand, the molecules and the antioxidant enzymes remain constant in the seedlings treated with 50 mM of Zn-TMPyP under greenhouse except for an increase in thiols in the roots (FIG. 11).

[0140] These tests confirm that the ability of Zn-TMPyP to be a selective foliar herbicide was unexpected, based on in vitro tests.

Finally, it should be noted that the effect following foliar spraying of Zn-TMPyP is not predictable depending on the nature of the group of the plant: the development of barley (grass) and tomato ( dicot) is not affected following foliar spraying of Zn-TMPyP. In on the other hand, the opposite effect is obtained on the plants of the same group: the poppy (dicot) and the millet (grass) are completely dead after foliar spraying of Zn-TMPyP.

[0142] Example 2: Determination of the non-toxicity of Zn-TMPyP for peach worms (maggots) [0143] The maggots were placed in Petri dishes under photoperiod (16 h) or under dark conditions in a phytotron (24°C) for 24 hours. 30 maggots per petri dish were subjected to 10 sprays of H ₂ 0 or 50 mM of Zn-TMPyP. Percent viability was determined after 24 h.

5 to 10% of the maggots die under normal conditions (H ₂ O spray) and after treatment with Zn-TMPyP, establishing the safety of Zn-TMPyP on peach worms (FIG. 7).

Zn-TMPyP is advantageously more respectful of the environment by having a reduced impact on animal biodiversity.

[0146] Example 3: Photostability/photodegradation of Zn-TMPyP [0147] In the dark, Zn-TMPyP remains perfectly stable whether with or without surfactant for 29 days of follow-up (FIG. 8).

Under photoperiod (16 h light/8 h dark), the stability of Zn-TMPyP (with or without surfactant) drops sharply from the 10th day. This is completely photodegraded between the 17th and 19th day. As for a solution without surfactant, about 93% of Zn-TMPyP is photodegraded after 19 days under photoperiod, then this solution continues to photodegrade slowly to disappear after 29 days of photoperiod.

This follow-up was carried out by carrying out 3 independent experiments.

[0150] Monitoring shows that the half-life of the weedkiller solution in water is two weeks when it is exposed to light (this is the case when the solution is sprayed on plants in nature ). The absolute stability of the solution in the dark will ensure the good storage of the product for long periods, in its opaque packaging, until its use.

Thus, the photodegradability of Zn-TMPyP makes it possible to limit its long-term effects and pleads in favor of its harmlessness for the environment and for humans.

Example 4 Selective Foliar Herbicidal Effect of Zn-TMPyP - Greenhouse Trial

[0153] Materials and Methods [0154] Additional greenhouse tests were carried out. All the plants underwent 5 treatments (foliar sprays) with 3 different C1, C2 or C3 doses of Zn-TMPyP, each treatment being spaced 48 hours apart. Treatments started at the cotyledon stage. Harvesting was carried out 6 weeks after germination (and 10 weeks for barley).

The doses of Zn-TMPyP are as follows:

C1=50mM; C2=75mM; 03=100mM

Zn-TMPyP was formulated with 0.1% v/v Polyoxyethylene nonylphenylether.

The species of plants used are shown in the table below: [0156] [Table 2]

[0157] Results

The results after 6 weeks (10 for barley) demonstrate that treatment with Zn-TMPyP drastically alters the growth of weeds, either in a dose-dependent manner or at a specific dose. Certain concentrations of Zn-TMPyP completely destroy weeds. (Figure 12).

On the contrary, and regardless of the concentrations applied, the plants of interest grow perfectly after application of Zn-TMPyP by foliar spraying. The growth of the plants of interest is in fact not altered. Only spinach growth is slightly affected. (Figure 13). These results confirm the selective foliar herbicidal effect of Zn-TMPyP.

Claims

[Claim 1] Use of cationic porphyrins selected from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride and tetra (N-methylpyridyl) porphyrin tetrachloride or mixtures thereof as a selective foliar herbicide.

[Claim 2] Use according to claim 1, characterized in that the porphyrins are zinc-tetra (N-methylpyridyl) porphyrin tetrachloride.

[Claim 3] Use according to claim 1 or 2, characterized in that said porphyrins are sprayed on a culture space chosen from a field, a greenhouse, a meadow, a courtyard, an alley, a garden, a vegetable patch.

[Claim 4] Use according to any one of Claims 1 to 3 for the treatment of cultures chosen from cultures of the genus Nicotiana, of the genus Solanum, of the genus Vitis, of the genus Hordeaum, of the genus Triticum, of the genus Zeya, of the genus Helianthus, genus Allium, genus Spinacia, genus Frugaria and genus (Brassica).

[Claim 5] Use according to any one of claims 1 to 4 for the treatment of crops chosen from wheat (Triticum sativum), barley (Hordeum vulgare), corn (Zeya mays), sunflower (Helianthus annuus), turnip (Brassica rapa), tomato (Solanum lycopersicum), onion (Allium cepa), spinach (Spinacia oleracea).

[Claim 6] Use according to any one of Claims 1 to 5, characterized in that the said cationic porphyrins have a herbicidal effect on weeds of the genus Urtica, of the genus Echinochloa, of the genus Papaver, of the genus Lolium, of the genus Datura, of the genus Stellaria, of the genus Chenopodium, of the genus Panicum, of the genus Taraxacum, of the genus Rumex, of the genus Solanum.

[Claim 7] Use according to any one of Claims 1 to 6, characterized in that the said cationic porphyrins have a herbicidal effect on weeds chosen from black nightshade (Solanum nigrum), dandelion (Taraxacum officinale), lamb's-quarters (Chenopodium album), dock (Rumex crispus), Italian rye (Lolium multiforum), false millet (Panicum miliaceum), white pimpernel (Stellaria media), millet (Echinochloa frumentacea), poppy (Papaver rhoeas). [Claim 8] Selective weed control process consisting of the application of cationic porphyrins chosen from zinc-tetra (N-methylpyridyl) porphyrin tetrachloride, tetra (N-methylpyridyl) porphyrin tetrachloride or their mixture, said porphyrins being applied by foliar spraying on a growing space such as a field, a greenhouse, a meadow, a courtyard, an alley, a garden, a vegetable patch. [Claim 9] Selective weed control method according to Claim 8 or 9, characterized in that the said porphyrins are applied at the cotyledon stage and/or at the first leaf stage such as the 2-leaf and/or 4-leaf stage.