WO2007131310A1

WO2007131310A1 - Anti-inflammatory and analgesic pharmaceutical composition containing safrole n-acylhydrazone derivatives, use, and method of preparation

Info

Publication number: WO2007131310A1
Application number: PCT/BR2007/000114
Authority: WO
Inventors: Carlos Alberto Manssour Fraga; Eiiezer Jesus De Lacerda Barreiro; Carolina De Mattos Duarte; Jorge Luiz Mendonça TRIBUTINO; Ana Luisa Palhares Miranda
Original assignee: Universidade Federal do Rio de Janeiro UFRJ
Current assignee: Universidade Federal do Rio de Janeiro UFRJ
Priority date: 2006-05-15
Filing date: 2007-05-14
Publication date: 2007-11-22
Anticipated expiration: 2008-11-15
Also published as: WO2007131310B1; BRPI0601885A

Abstract

New derivatives of the 6-nitro-N'-arylmethylidene-1,3-benzodioxol-5-carbohydrazide nucleus are described, having anti-inflammatory and analgesic properties, acting on the central nervous system, and are therefore useful in the treatment of acute and chronic inflammatory diseases and central hyperalgesia in mammals; pharmaceutical compositions containing said compounds and methods of preparation thereof are also described.

Description

Specification

ANTI-INFLAMMATORY AND ANALGESIC PHARMACEUTICAL COMPOSITION CONTAINING SAFROLE N-ACYLHYDRAZONE DERIVATIVES, USE, AND METHOD OF PREPARATION

Field of the Invention

The present invention is related to safrole Λ/-acylhydrazone derivatives. More specifically, the present invention is related to 6-nitro-ΛP-arylmethyldene- 1 ,3-benzodioxol-5-carbohydrazide derivatives, substituted or not, and its isosterics, having anti-inflammatory and/or analgesic properties acting on the central nervous system, also having antioxidant properties, particularly useful in the treatment of acute and chronic inflammatory diseases and/or central hyperalgesia. It also refers to a preparation process and pharmaceutical compositions containing the same.

Background of the Invention

Inflammation is a reaction of the vascularized tissue to injury, characterized by the extravasation of liquid and cells from the blood to the interstitial space. This constitutes an important defense mechanism, the ultimate purpose of which is to protect the organism from the initial causal agent of the cellular injury (micro-organisms, toxins) and from the consequences of the injury (cells and necrotic tissue).

Inflammation may be classified as acute or chronic. Acute inflammation lasts a relatively short time, from minutes to hours or for some days, and its main characteristics are: edema, leading to the exudation of plasmatic proteins, and the migration of leukocytes (mainly neutrophils) to the injured tissue.

Chronic inflammation is the sum of the organism's reactions as a result of the persistence of the aggressive agent, differing from acute inflammation by its long duration and by the absence or scarce evidence of the classic signs of inflammation.

In the inflammatory process, the migration of leukocytes to the site of inflammation is of vital importance, bearing in mind their role in the combat against the aggressive agents. They have the capacity to phagocyte and destroy aggressive agents, such as bacteria and antigens, besides degrading the tissue.

Leukocytic margination is the process by which leukocytes migrate from the blood column in the veins to occupy the periphery. Next, they adhere to the endothelium, moving to the endothelial surface (rolling phenomenon) and finally migrate through the venule wall, traversing the endothelial cells.

The whole process of leukocyte migration depends on the release of chemotactic substances, which guide the movement of these cells by forming a concentration gradient. In most cases of acute inflammation, neutrophils predominate at the injury site, being substituted by monocytes within 24 to 48 hours. The chemotactic agents (arachidonic acid (AA)) metabolites and cytokines), as components of the complementary system, are responsible for attracting these cells to the inflamed site, and are also responsible for increasing the inflammatory stimulus. The macrophage is crucial in chronic inflammation due to the large number of substances produced once it is activated.

The presence of the aggressive agent causes the release of chemical mediators of inflammation, the main role of which is the modulation of the entire inflammatory process. These mediators are of plasma origin or are produced by the ceils involved in the inflammatory process. The main mediators include vasoactive amines (histamine and serotonin), the hypotensor cynins (bradykinin), the complementary system, the coagulation system, the cytokines (IL-1 and TNF), the chemokynes, nitric oxide and eicosanoids. Pain can be defined as an unpleasant sensorial and emotional experience associated with potential or already occurred tissue damage, or described in terms of this pathology. It is one of the body's warning mechanisms designed to produce reactions that allow the individual to eradicate the aggressive agent, thus avoiding consequential tissue damage. Pain can be classified into three types: acute or physiological pain, chronic or inflammatory pain and neuropathic pain.

In acute or physiological pain, the high intensity stimuli capable of producing pain stimulate the nociceptors, present in the periphery of the skin, muscles, articulations and viscera, transmitting painful impulses via non- myelinic C fibers and myelinized Aδ fibers to the dorsal horn of the spinal chord. The information of pain is then conveyed along the central nervous system by the spinal chord, traversing the spinothalamic tract until it reaches the thalamus. Neurons of the second order perform synapsis with the neurons of the third order in the ventral portion of the thalamus, transmitting the impulse to the cerebral cortex, when the sensation of pain is then perceived. Descending tracts of the central nervous system are able to modulate the transmission of pain impulses in the dorsal horn, acting by monoaminergic mechanisms, involving the activation of ct₂-adrenergic and serotoninergic receptors. Around ten to fifteen neuromodulators or neurotransmitters have been said to be involved in the nociceptive tracts, acting both on the backbone and in the ascending and descending tracts in the processing of pain.

A tissue injury caused by stimuli of a chemical, physical or biological nature promote the release of pro-algesic chemical mediators at the site, such as serotonin, bradykinin, histamine, ATP, various monoamines, cytokines, chemokines, growth factors, excitory aminoacids and prostaglandins. These mediators are able to activate and sensibilize the nociceptores, increasing the excitability of these neurons, causing an exaggerated response to the pain stimuli, in a phenomenon called primary hyperalgesia. The stimulation of the nociceptores by the action of the chemical mediators induces a process of amplification of the response in the neurons of the dorsal horn, producing a sensibilization of the central nervous system in a phenomenon called wind-up. The vasodilatation and the action of the chemical mediators at the site of the issue injury induce a stimulation not only of the local nocicepetors, but also of those present in the areas surrounding the injury, thus promoting the onset of the phenomenon of secondary hyperalgesia or referred pain, where the sensation of pain is felt in places other than those where the tissue injury really occurred. Neuropathic pain is the result of an injury to the nervous tissue associated to trauma, surgery, amputation and radiation or with certain types of diseases such as herpes zoster, multiple sclerosis, arthritis and diabetes. The post-herpetic neuralgia and diabetic neuropathy are two very common neuropathic syndromes. This injury to the nervous fibers promotes an atypical (ectopic) stimulation of these fibers, inducing the processes of hyperexcitability and central sensibilization of the neurons of the dorsal horn, resulting in the phenomena of hyperalgesia and allodynia. Acetylsalicylic acid (AAS), just like indomethacin, ketoprofen, piroxicam and diclophenaco are part of the class of substances called non-steroidal antiinflammatory drugs (NSAIDs), and has been used clinically for many years.

The analgesic and anti-inflammatory properties of these drugs are related to the inhibition of the prostaglandin endoperoxide synthase enzyme (PGHS) or cyclooxygenase (COX), reducing the production of the prostaglandins that, as seen earlier, are intimately related to the maintenance of the inflammatory processes, in the genesis of pain and platelet homeostasis.

With the discovery of a second isoform of this enzyme (PGHS-2), the hypothesis arose that the same was related to the production of prostaglandins in inflammatory processes, whereas the PGHS-1 would produce eicosanoids related to the maintenance of the normal physiological processes of most tissues, for instance, the production of PGE₂ having cytoprotective activity of the gastrointestinal tract. Based on this principle, it was postulated that the seriously adverse effects on the gastrointestinal tract associated to the chronic use of NSAIDs in inflammatory processes, such as rheumatoid arthritis, would be the result of the COX-1 inhibition [CASHMAN, J. N. (1996) Drugs 52, 13-23; VANE, J. et al. (1998) Ann. Rev. Pharmacol. Toxicol. 38, 97-120; DANNHARDT, G., KIEFER, W. (2001) Eur. J. Med. Chem. 36, 109-126; RIENDEAU, D. et al. (2001) J. Pharmacol. Exp. Ther. 296, 558-566], Said NSAIDs, referred to as classic, are competitive inhibitors in the cyclooxigenase site of the PGHS, without modulating the peroxidase function. Since then, the search for new selective NSAIDs for COX-2 has become an important therapeutic strategy. The synthesis of various compounds having selective inhibitory activity for COX-2 has been described, and these new molecules have anti-inflammatory activity, but without causing gastrotoxic effects in various preclinical and clinical trials. Currently three selective COX-2 drugs are on the market: celecoxib, etoricoxib and lumiracoxib. Rofecoxib and valdecoxib were recently withdrawn from the market because they increased the risk of myocardial infarction

With the advent of COX-3, the effect of selective COX-2 drugs on this isoform are not yet known Some authors have suggested a possible existence 5 of a third isoform expressed in the central nervous system, mainly by virtue of the fact that analgesic and antipyretic drugs such as paracetamol, have an unknown action mechanism, and do not act by inhibiting COX-1 or COX-2 Chandrasekaran and collaborators [CHANDRASEKARAN, N V et al (2002) Proc Natl. Acad. Sci. USA 99, 13926-13931] cloned and characterized COX-3,

I O a variant of COX-1 , having the same sequence of aminoacids in the catalytic site COX-3 in inhibited by various classic NSAIDs and also by paracetamol, which suggests that the analgesic and antipyretic mechanism of these drugs is by inhibition of this isoform Warner & Mitchell [WARNER, T D , MITCHEL, J A (2002) Proc. Natl. Acad. Sci. USA 99, 13371-13373] suggest the possible

1 5 existence of multiple COX-1 and COX-2 isoforms, and that certain NSAIDs could have a greater specificity for one or another isoform This could explain why different patients benefit from different types of NSAIDs Since the discovery of COX-3 is very recent, it still remains for the role of this isoform to be characterized in the inflammatory process

20 AAS is the most widely used clinical anti-platelet drug It is capable of irreversibly acetylating the hydroxyl residues of serine 529 in the COX-1 expressed on the platelet, preventing the link of AA to the active site of the enzyme Other NSAIDs reversibly compete with the AA for the link to the COX site As the platelets have no nucleus and have low protein synthesis capacity,

25 they cannot replace the constitutive COX-1 that underwent AAS acetylation After suspending administration of AAS, the COX-1 platelet activity only returns to normal if new platelets are added to the circulation AAS has been used clinically for primary and secondary prevention in patients with atherosclerotic vascular diseases

30 Bearing in mind the role of the leukotrienes in asthma and chronic inflammatory diseases such as ulcerative colitis and rheumatoid arthritis, the development of 5-LO inhibitors is a significant strategy Zileuton, a selective 5- LO inhibitor, is available on the market and has been used in anti-asthmatic therapy. However, clinical studies have shown that it is ineffective as monotherapy in patients suffering from rheumatoid arthritis and ulcerative colitis. COX inhibitors induce side effects in patients with asthma by increasing the production of leukotrienes caused by a greater availability of arachidonic acid for metabolization via the 5-LO pathway. A promising alternative to avoid these kinds of adverse effects of the NSAIDs would be the development of dual COX/5-LO inhibitors, leading to compounds having increased effectiveness and fewer adverse effects compared to the NSAIDs. Further, the dual inhibitors are therapeutically promising due to their anti-inflammatory effect with a greater scope of action than those of the classic NSAIDS, inhibiting both the production of inflammatory mediators deriving from 5-LO and the COX/5-LO. Various chemically distinct compounds have been described as dual COX/5-LO inhibitors, including BW-755c, CBS1108, ML3000 and tepoxalin.

Other approaches have been recorded in recent literature for the treatment of chronic inflammatory diseases, resulting in the reduction of tumoral necrosis factor α levels (TNF-α). The use of anti-TNF-α antibodies, such as etanercept, infliximab and adalimumab, has proven to be effective. Yet besides the discomfort of parenteral administration, clinical tests have associated the activation of latent tuberculosis and the increase in lymphomas in this therapy. These facts increment the advantages of developing new micro-molecules that can be orally administered for anti-TNF-α therapy.

In this context, thalidomide was approved for the treatment of lepromas, multiple myeloma and rheumatoid arthritis, due to its capacity to modulate the biosynthesis of cytokines such as TNF-α. Clinical tests show various other candidates as cytokine modulating drugs having different action mechanisms, but which ultimately reduce the biosynthesis of cytokines. There are phosphodiesterase-4 (PDE-4) inhibitors, such as rolipram, nitraquazone, theophiline and ariflo; p38 mitogen-activated (p38 MAPK) protein inhibitors, such as SB203580, BIRB796 and VX-745; TNF-α (TACE) converting enzyme inhibitors, such as marimastat; and KB (NF-KB) nuclear factor modulators, such as acanthoic acid, parthelenolide and ergolide. Other approaches such as the modulation of peripheral canabinoid receptors (CB2) have also been documented in recent literature, with a view to suppressing the biosynthesis of cytokines.

Summary of the Invention

The purpose of the present invention is to provide alternative therapeutic strategies to those resulting from the use of the anti-inflammatory and/or analgesic drugs currently available. In one aspect of the invention, the difficulties found with the anti-inflammatory and/or analgesic molecules available to-date are solved, since the molecules of the present invention inhibit cellular migration and edema, induce central analgesia and have antioxidant properties, without causing gastro-irritant effects. One of the purposes of the present invention is, therefore, to provide anti-inflammatory and/or analgesic molecules acting on the central nervous system with antioxidant properties, which are useful in the reversal of acute and/or chronic inflammation in mammals, preferably humans.

Another purpose of the present invention is to provide anti-inflammatory and/or analgesic pharmaceutical compositions acting on the central nervous system with antioxidant properties, which are useful in reversing acute and chronic inflammation and/or for the treatment of central hyperalgesia in mammals, preferably humans.

Given the advantages of molecule selectivity of the present invention, the pharmaceutical compositions of the present invention can be administered in a wider variety of presentation forms, resulting in benefits for the user and greater production flexibility. Accordingly, another purpose of the present invention is to provide alternatives for the limitations in administration of pharmaceutical compositions for the treatment of acute and chronic inflammatory diseases and/or central hyperalgesia. The molecules of the present invention have different synthesis pathways from other currently used to treat acute and chronic inflammatory diseases and/or central hyperalgesia, and it is easier to prepare drugs under these conditions. Therefore, another purpose of this present invention, in preparing a drug for the treatment of inflammatory disturbances and/or hyperalgesia in mammals, is to provide the use of 6-nitro-/V-arylmethyldene-1 ,3-benzodioxol-5- carbohydrazides derivatives, substituted or not, its isosterics and/or pharmaceutically acceptable salts, solvates or hydrates.

5 The synthetic stages involved in producing the compounds of the present invention are less complicated and less costly that those used previously, resulting in advantages from an industrial point of view. Hence, an additional purpose of the present invention is to provide methods to product the antiinflammatory and/or analgesic molecules listed herein. One of the purposes of I O the present invention is to disclose means of preparing n-acylhydrazone antiinflammatory and analgesic derivatives having antioxidant properties.

These and other purposes of the present invention will be better understood and appreciated after reading the detailed description of the invention and corresponding claims. 15

Description of the Drawings

Figure 1 - Shows the structures of the classic NSAIDS (Acetylsalicylic acid, Indomethacin, Ketoprofen, Paracetamol, Piroxicam and Diclophenaco).

Figure 2 - Shows the structures of the selective PGHS-2 inhibitors 20 (Celecoxib, Lumiracoxib, Rofecoxib, Valdecoxib and Etoricoxib).

Figure 3 - Shows the structures of the 5-LO and dual COX/5-LO inhibitors (Zileuton, BW-755c, CBS1108, ML3000 and Tepoxalin).

Figure 4 - Shows the structures of the PDE-4 inhibitors (Thalidomide, Rolipram, Nitraquazone, Theophiline and Ariflo).

25 Figure 5 - Shows the structures of the p38MAPK inhibitors (SB203580,

VX745 and BIRB796).

Figure 6 - Shows the structures of the TACE and NF-κB modulating inhibitors (Marimastat, Acanthoic acid, Parthenolide and Ergolide).

30 Detailed Description of the Invention

Having briefly referred to the purposes of the present invention, we shall now set forth a detailed description, whenever suitable using preferred embodiments of the invention. One of the most innovative characteristics of the present invention is the synthesis of new functionalized N-acylhydrazone derivatives of formula (I) safrole derivatives, rationally planned as new analgesic and anti-inflammatory agents. The main structural characteristic of these new derivatives is the Λ/-acylhydrazone standard, this being the pharmacophoric group involved in recognizing the pharmacological target of action; besides the other aromatic subunits, acting as secondary pharmacophoric groups, having biophoric characteristics (electronic and hydrophoic), needed for recognition by the pharmacological action target of action, by way of van der Waals interactions between the aromatic ring and similar sites. The use of this structural standard for analog anti-inflammatory agents having a centrally-acting analgesic and antioxidant action has not been described previously, and, therefore, the compounds described in this invention and its synthetic methodology constitute an innovation among centrally-acting anti-inflammatory and analgesic agents, even though other N-acylhydrazone derivatives have been documented in recent literature as enzyme inhibitors of the arachidonic acid cascade, having peripheral anti-inflammatory and analgesic action. It is therefore important to highlight that the purpose of this invention refers to new N-acylhydrazone compounds having molecular action mechanism different from those described previously. The new compounds described in this invention belong to the class of derivatives of the 6-nitro-/V-arylmethyldene-1,3-benzodioxol-5-carbohydrazide nucleus and its isosterics, of general structure (I):

wherein:

R is hydrogen, alkyl, cycloalkyl, phenyl-W, furyl, thiophenyl, pyridyl, pyrimidinyl, pyrrolyl, thiazolyl, quinazolyl or isoquinolyl; W is hydrogen, ortho-a\ky\, ortho-cyc\oa\ky\, orf/70-alkoxyl, orf/70-cycloalkoxyl, ortr/o-thioxyl, orf/70-aryoxyl, ortr/o-sulphones, orf/70-sulphates, ortho- sulphoxides, or#?o-sulphonates, orf/70-sulphonamides, ortho-ammo, ortho- amide, orf/70-halates, orf/70-carboalkoxyl, orf/70-carbothioaikoxyl, ortho- trihaloalkane, ortho-cyan, ortr/o-nitro, mefa-alkyl, mefø-cycloalkyl, mefa-aikoxyl, mefa-cycloalkoxyl, mefa-thioxyl, metø-aryoxyl, metø-sulphones, meta- sulphates, metø-sulphoxides, mefa-sulphonates, metø-sulphonamides, meta- amino, metø-amide, metø-halates, mefa-carboalkoxyl, mefa-carbothioalkoxyl, metø-trihaloalkane, mefa-cyan, mefa-nitro, para-alkyl, para-cycloalkyl, para- alkoxyl, para-cycloalkoxyl, para-thioxyl, para-aryoxyl, para-sulphones, para- sulphates, para-sulphoxides, para-sulphonates, para-sulphonamides, para- amino, para-amide, para-halates, para-carboalkoxyl, para-carbothioalkoxyl, para-trihaloalkane, para-cyan or para-nitro.

The new compounds of formula (I) were obtained in good to excellent chemical yields, using the synthetic methodology described herein, which is characterized by having few stages, providing high yields, based on commercially available compounds, qualifying this synthetic methodology for industrial use.

The compounds of the present invention were planned by way of converging syntheses, using classic reactions such as:

• aromatic nitration;

• hydrazinolysis;

• condensation of aldehydes with amines;

More specifically, the compounds of formula (I) of the present invention can be prepared using a method comprising the stages of:

• isomerization of the safrole, followed by ozonolysis for aldehyde synthesis (piperonal);

• nitration of piperonal in a concentrated acid means, regioselectively;

• oxidation of the 6-nitropiperonal to the 6-nitro-1 ,3-benzodioxol-5- methyl carboxylate ester; • obtaining the corresponding hydrazide by hydrazinolysis;

• condensation with the respective aldehydes to obtain the N- acylhydrazone ligands.

5 For purposes of illustration, this report will comprise a description of the synthesis of the following compounds:

• 6-nitro-1 ,3-benzodioxol-δ-carbaldehyde

• 6-nitro-1 ,3-benzodioxol-5-methyl carboxylate I O • 6-nitro-1,3-benzodioxol-5-carbohydrazide

• Λ/'-(2-furylmethyldene)-6-nitro-1 ,3-benzodioxol-5-carbohydrazide

• 6-nitro-Λ/'-(2-thienylmethyldene)-1 ,3-benzodioxol-5-carbohydrazide

• Λ/'-(3,5-ditercbutyl-4-hydroxyphenylmethyldene)-6-nitro-1 ,3- benzodioxol-5-carbohydrazide

15 • Λ/'-phenylmethyldene-6-nitro-1 ,3-benzodioxol-5-carbohydrazide

• 6-nitro-Λ/'-(2-pyridnylmethyldene)-1 ,3-benzodioxol-5-carbohydrazide

• 6-nitro-Λ/'-(4-pyridnylmethyldene)-1 ,3-benzodioxol-5-carbohydrazide

• 6-nitro-Λ/'-.(1 /-/-pyrrole-2-ylmethyldene)-1 ,3-benzodioxol-5- carbohydrazide 0

Next is a detailed description of the synthetic methods of this invention for some of the compounds claimed, including relevant spectroscopic data for the characterization thereof.

Persons skilled in the art will appreciate the invention disclosed herein,

25 which clearly provides a pharmaceutical composition for the treatment of inflammatory disturbances and/or hyperalgesia in mammals characterized by having a pharmaceutically acceptable carrier and/or dilutant, and at least one derivative of the 6-nitro-/V-arylmethyldene-1 ,3-benzodioxol-5-carbohydrazide nucleus, described in detail in this invention.

30 Persons skilled in the art will also appreciate another aspect of the present invention, which clearly provides the use of derivatives of the 6-nitro-Λ/'- arylmethyldene-1 ,3-benzodioxol-5-carbohydrazide nucleus, described in detail in this invention, characterized by the fact that it is for the preparatiop of a curative or prophylactic drug for inflammatory disturbances and/or hyperalgesia in mammals including man.

Persons skilled in the art will also appreciate another aspect of the present invention, which clearly provides a method of preparing the anti- inflammatory and analgesic n-acylhydrazone derivatives, explained below in detail. The following examples illustrate, but do not limit the present invention.

Example 1

Preparation of the 6-nitro-1 ,3-benzodioxol-5-carbaldehyde derivative

In a flask containing 1g (6.7mmol) of pulverized piperonal (Aldrich), in bain marie at room temperature, with a septum attached to an empty ball (pressure relief closed system), 3.3mL of concentrated HNO₃ was added. An immediate change of color from colorless to yellow was noted. After around 5 minutes of stirring, the formation of an orange gas (NO₂) and darkening of the reaction mixture (chestnut color) was noted, besides a sudden heating, which was controlled by adding ice to the bain marie. The start material dissolved completely. After around 10 minutes, the mixture was cooled in an ice bath, whereupon an intense precipitation of a light yellow solid was noted, and which was vacuum filtered, washed with around 20OmL of cold distilled water and recrystallized in ethanol/water.

Light yellow solid; 1.241g (95%); m.p. 91-92°C.

RMN ¹H (200MHz, DMSO-Cf₆, TMS) δ (ppm): 6,34 (s, 2H, H-2); 7,32 (s, 1 H, H- 4); 7,74 (s, 1 H, H-7); 10,09 (s, 1H, -CHO).

RMN ¹³C (50MHz, DMSO-Cf₆, TMS) δ (ppm): 104,85 (C-2); 105,43 (C-7); 107,48 (C-4); 127,92 (C-5); 146,13 (C-6); 151 ,76 (C-3); 152,33 (C-1); 188,58 (C=O). IV (υ_maχ, KBr) υ (cm^'1): 1682, 1518, 1368, 1336, 1126, 1119.

Example 2

Preparation of the 6-nitro-1,3-benzodioxol-5-methyl carboxylate derivative In a 25OmL flask containing 1.5g (7.7mmol) of 6-nitro-1 ,3-benzodioxol- 5-carbaldehyde, suspended in 1OmL of methanol, 1.3g (23.1 mmol) of potassium hydroxide solubilized in 2OmL of methanol was added, then 2.54g (lOmmol) of iodine was added, washing the beaker with a further 3OmL of 5 methanol, making a total of 6OmL of methanol. The reaction mixture continued to be stirred for 1.5 hours, whereupon it was noted that the start aldehyde had been totally consumed. Next 1OmL of solution saturated with sodium bisulphite was added, and it was noted that the chestnut color disappeared. The mixture was vacuum filtered and the solid was washed with 3OmL of methanol. The I O filtrate was evaporated under reduced pressure and recrystallized in ethanol/water.

Light yellow solid; 1.525g (88%); m.p. 102-103⁰C.

RMN ¹H (200MHz, CDCI₃, TMS) δ (ppm): 3,89 (s, 3H, OCH₃); 6,18 (s, 2H, H-7); 15 7,03 (s, 1 H, H-4); 7,38 (s, 1 H, H-2).

RMN ¹³C (50MHz, CDCI₃, TMS) δ (ppm): 53,45 (OCH₃); 103,77 (C-2); 105,08

(C-7); 108,60 (C-4); 123,98 (C-5); 143,21 (C-6); 149,78 (C-3); 151 ,49 (C-1);

165,89 (C=O).

IV (υ_maχ, KBr) υ (cm^"1): 1717, 1509, 1362, 1272, 1108. 0

Example 3

Preparation of the 6-nitro-1,3-benzodioxol-5-carbohydrazide derivative

In a 5OmL flask containing 0.5g (2.22mmol) of 6-nitro-1 ,3-benzodioxol-

25 5-methyl carboxylate suspended in 2mL of ethanol, 6.7mL of 80% hydrazine hydrate was added. The reaction mixture continued to be stirred at room temperature for 2 hours, and a darkening of the reaction means and total consumption of the start ester was noted. The basic pH was neutralized with concentrated HCI in an ice bath and the means was concentrated under

30 reduced pressure, noting the precipitation of a yellow solid, which was intensified by an ice bath. The solid was filtered to give the desired product after recrystallization in ethanol/water. Yellow solid; 0.350 g (70%); m.p. 184-185°C.

RMN ¹H (200MHz, DMSO-Cf₆, TMS) δ (ppm): 4,42 (s, 2H, NH₂); 6,26 (s, 2H, H- 2); 7,03 (s, 1 H, H-4); 7,64 (s, 1H, H-7); 9,59 (s, 1 H, NH).

RMN ¹³C (50MHz₁ DMSO-Cf₆, TMS) δ (ppm): 104,24 (C-2); 105,32 (C-7); 108,54 5 (C-4); 128,27 (C-5); 141 ,93 (C-6); 148,79 (C-3); 151 ,71 (C-1); 165,17 (C=O). IV (υ_max, KBr) υ (cm^"1): 3336, 3199, 3120, 1670, 1519, 1502, 1475, 1328, 1257, 1030, 880.

Example 4

I O

Preparation of the 6-nitro-W-(W-arylmethyldene)-1,3-benzodioxol-5- carbohydrazides derivatives

In a flask containing 0.225g (1mmol) of 6-nitro-1 ,3-benzodioxol-5- carbohydrazide in 1OmL of absolute ethanol, 1.1mmol of the respective

15 aldehyde respective was added. The mixture was stirred for 1 hour, and it was noted that the start material had been totally consumed. After concentration of the means under reduced pressure, 15ml_ of cold water was added and the mixture was put in an ice bath. Intense precipitation of a solid was noted, which after recrystallization in ethanol/water, furnished the respective desired

20 products.

Λf'-(2-furylmethyldene)-6-nitro-1,3-benzodioxol-5-carbohydrazide

Yellow solid; 0,267g (88%); m.p. 226-228°C.

RMN ¹H (200MHz, DMSO-d₆, TMS) δ (ppm): 6,30 (s, 4H, H-2E/Z); 6,53 (d, 1 H, 25 J = 1 ,8 Hz, H-4" Z); 6,55 (d, 1H, J = 1 ,7 Hz, H-4"£); 6,72 (d, 1 H, J = 3,4 Hz, H-

3'¹Z); 6,95 (d, 1 H, J = 3,4 Hz, H-TE); 7,15 (s, 1 H, H-4'2); 7,30 (s, 1 H, H-4'E);

7,72 (s, 2H, H-AEIZ); 7,75 (s, 1 H, H-5"Z); 7,87 (s, 2H, H-IEIZ); 8,10 (s, 1H, H-

5"E); 1 1 ,87 (s, 1 H, H-2'Z); 11 ,90 (s, 1 H, H-2'E).

RMN ¹³C (50MHz, DMSO-Cf₆, TMS) δ (ppm): 104,40 (C-2); 104,68 (C-7); 105,45 30 (C-4); 108,52 (C-3"); 108,86 (C-3"); 112,49 (C-4"); 112,72 (C-4"); 128,57 (C-5);

134,88 (C-4'); 138,08 (C-4¹); 141 ,24 (C-6); 145,54 (C-5"); 145,91 (C-5"); 148,79

(C-1 a); 149,61 (C-3a); 152,23 (C-2"); 152,64 (C-2"); 161 ,86 (C-f); 167,89 (C-

1 '). IV (υ_max, KBr) υ (cm^"1): 3197, 3123, 3061, 2918, 2855, 1644, 1623, 1533, 1504,

1486, 1381, 1293, 1281, 1261, 1164, 1126, 1033, 1017, 939, 877, 763.

β-nitro-Λ/'-(2-thienylmethyldene)-1,3-benzodioxol-5-carbohvdrazide

5 Yellow solid; 0.287g (90%); m.p. 240-242⁰C.

RMN ¹H (200MHz, DMSO-Cf₆, TMS) δ (ppm): 6,30 (s, 4H, H-2E/Z); 7,03 (t, 1 H, J

= 4,3 Hz, H-4"Z); 7,15 (m, 2H, H-4"£,3"2); 7,31 (m, 2H, H-3"E,4'Z); 7,49 (m,

2H, H-4'E,5"Z); 7,71 (m, 3H, H-4E/Z,5"E); 8,13 (s, 2H, H-7E/Z); 11 ,88 (s, 1 H,

H-2'Z); 1 1 ,92 (s, 1 H, H-2'E). I O RMN ¹³C (50MHz, DMSO-cfe TMS) δ (ppm):104,37 (C-2); 104,46 (C-7); 105,42

(C-4); 108,60 (C-3"); 108,89 (C-3"); 128,14 (C-5); 128,21 (C-4"); 128,35 (C-4");

139,04 (C-4¹); 139,20 (C-4¹); 139,63 (C-5"); 141 ,52 (C-6); 143,41 (C-5"); 148,70

(C-1 a); 149,1 1 (C-3a); 152,22 (C-2"); 152,58 (C-2"); 161 ,74 (C-1¹); 167,75 (C-

1 '). 15 IV (Umax, KBr) υ (cm^"1): 3191 , 3073, 3060, 3032, 2992, 2915, 2844, 1639, 1608,

1591 , 1560, 1531 , 1503, 1485, 1380, 1290, 1264, 1222, 1162, 1125, 1033,

1015, 928, 878, 713.

Λ/'-(3,5-difercbutyl-4-hvdroxyphenvimethyldene)-6-nitro-1,3-benzodloxol-5- 20 carbohydrazide

Yellow solid; 0.362g (82%); m.p. 280-282⁰C.

RMN ¹H (200MHz, DMSO-d₆, TMS) δ (ppm): 1 ,32 (s, 36H, -CH₃E/Z); 5,72 (s,

1 H, -OH); 6,29 (s, 4H, H-2E/Z); 7,15 (s, 4H, H-2",6"E/Z); 7,35 (s, 2H, H-4E/Z);

7,67 (m, 1 H, H-4'Z); 7,73 (s, 1 H, H-4'E); 7,83 (s, 2H, H-IEIZ); 11 ,88 (s, 1 H, H- 25 2¹Z); 11 ,90 (s, 1 H, H-2'E).

RMN ¹³C (50MHz, DMSO-Cf₆, TMS) δ (ppm): 30,56 (-CH₃); 34,98 (C-Me₃);

104,12 (C-7); 104,28 (C-2); 108,76 (C-4); 124,00 (C-2",6"); 124,47 (C-2",6");

125,69 (C-5); 128,61 (C-3",5"); 139,48 (C-1"); 141,99 (C-6); 144,99 (C-4');

148,59 (C-1a); 149,01 (C-4¹); 152,16 (C-3a); 156,28 (C-4"); 156,74 (C-4"); 30 167,69 (C-1').

IV (υ_maχ, KBr) υ (cm^"1): 3593, 3172, 3065, 2962, 2872, 1662, 1612, 1527, 1509,

1487, 1433, 1401, 1372, 1342, 1259, 1236, 1212, 1139, 1117, 1032, 927, 875, 766. /V'-phenylmethyldene-β-nitro-I.S-benzodioxol-δ-carbohvdrazide

Yellow solid; 0.297g (95%); m.p. 216-218°C.

RMN ¹H (200MHz, DMSO-Cf₆, TMS) δ (ppm): 6,31 (S₁ 4H, H-2E/Z); 7,40 (m, 12H₁ H-4E/2,2",3",4",5",6"£/Z); 7,75 (s, 2H, H-7E/Z); 7,99 (s, 1 H, H-4'Z); 8,22

(s, 1 H, H-4'E); 11 ,94 (s, 1 H₁ H-2'Z); 11,97 (s, 1H, H-2'E).

RMN ¹³C (50MHz, DMSO-d₆, TMS) δ (ppm): 104,40 (C-2); 104,54 (C-7); 105,42

(C-4); 108,58 (C-2"); 108,92 (C-6"); 128,24 (C-5); 129,236 (C-4"); 134,36 (C-

1"); 144,75 (C-4¹); 148,34 (C-6); 148,79 (C-4^P); 149,13 (C-1a); 152,43 (C-3a); 161 ,97 (C-1¹); 167,98 (C-1¹).

IV (υ_max, KBr) υ (cm^"1): 3222, 3114, 3059, 2916, 2851, 1646, 1607, 1533, 1503,

1485, 1381 , 1279, 1161 , 1123, 1033, 928, 878, 768, 693.

6-nitro-/V'-(2-pyridnylmethyldene)-1,3-benzodioxol-5-carbohydrazide Yellow solid; 0.264g (84%); m.p. 184-185°C.

RMN ¹H (200MHz, DMSO-d₆, TMS) δ (ppm): 6,32 (s, 4H, H-2E/2); 7,34 (m, 6H,

H-4£/Z,4Ε/Z,4Ε/Z); 7,99 (m, 6H, H-7£/Z,5",6"£/Z); 8,58 (d, 2H, J = 15,3 Hz, H-

3"BZ); 12,15 (s, 1 H, H-2'Z); 12,19 (s, 1 H, H-2'E).

RMN ¹³C (50MHz, DMSO-Cf₆, TMS) δ (ppm): 107,64 (C-2); 108,66 (C-7); 11 1 ,79 (C-5); 122,69 (C-6"); 123,68 (C-6"); 127,97 (C-4"); 128,25 (C-4"); 131 ,23 (C-6);

140,46 (C-5"); 140,61 (C-5"); 144,78 (C-1a); 148,42 (C-4¹); 151,81 (C-4¹);

152,09 (C-3"); 152,43 (C-3"), 155,50 (C-3a); 155,91 (C-1"); 156,19 (C-1");

165,35 (C-1 '); 171 ,34 (C-f).

IV (iw, KBr) υ (cm^"1): 3427, 3198, 3122, 3063, 2983, 2925, 2854, 1663, 1609, 1523, 1505, 1485, 1432, 1368, 1336, 1288, 1262, 1163, 1126, 1032, 921, 874,

777, 762, 640, 583.

6-nitro-Λ/'-(4-pyridnvimethyldene)-1,3-benzodioxol-5-carbohydrazide

Yellow solid; 0.279g (89%); m.p. 202-203⁰C. RMN ¹H (200MHz, DMSO-d₆, TMS) δ (ppm): 6,32 (s, 4H, H-2E/Z); 7,21 (s, 2H, H-4E/Z); 7,73 (d, 4H, J = 14 Hz, H-T, 6" BZ); 7,99 (s, 1 H, H-4'Z); 8,24 (s, 1 H, H- 4¹E); 8,63 (d, 4H, J = 14 Hz, H-3",5"E/Z); 12,27 (s, 1 H, H-2'Z); 12,30 (s, 1 H, H- 2' E). RMN ¹³C (50MHz, DMSO-Cf₆, TMS) δ (ppm): 107,64 (C-2); 107,79 (C-7); 108,61 (C-7); 111 ,74 (C-4); 112,04 (C-4); 124,33 (C-2",6"); 124,89 (C-2",6"); 131 ,04 (C-5); 131 , 16 (C-5); 144,74 (C-1"); 144,98 (C-1 "); 145,39 (C-4¹); 148,96 (C-4'); 152,13 (C-3",5^M); 152,46 (C-3",5"); 153,39 (C-6); 153,45 (C-6), 155,52 (C-1a); 155,94 (C-3a); 165,53 (C-1'); 171 ,54 (C-f).

IV (υ_maχ, KBr) υ (crrV¹): 3450, 3198, 3119, 3050, 2996, 2920, 2857, 1670, 1581 , 1530, 1508, 1489, 1421 , 1370, 1337, 1298, 1268, 1162, 1125, 1030, 923, 881 , 818, 744, 693, 537.

6-nitro-Λ/'-(1H-pyrrole-2-ylmethyldene)-1,3-benzodioxol-5-carbohydrazide

Yellow solid; 0.269g (89%); m.p. 202-203.

RMN ¹H (200MHz, DMSO-cfe, TMS) δ (ppm): 6,06 (d, 2H, J = 2,4 Hz, H-4"£/Z);

6,14 (d, 2H, J = 2,4 Hz, H-3"E/Z); 6,29 (s, 4H₁ H-2E/Z); 6,50 (s, 1 H, H-5"Z);

6,93 (s, 1 H, H-5"E); 7,11 (s, 1 H, H-4'Z); 7,27 (s, 1 H, H-4'£); 7,70 (s, 1 H, H-4Z); 7,72 (s, 1H, H-4E); 7,85 (s, 1H, H-7Z); 8,07 (s, 1 H, H-7E); 10,83 (s, 1 H, H-2'Z);

11 ,54 (s, 1 H₁ H-2Ε); 11 ,60 (s, 1 H, H-2'Z); 11 ,62 (s, 1 H, H-2'E).

RMN ¹³C (50MHz, DMSO-d₆, TMS) δ (ppm):107,56 (C-2); 107,98 (C-7); 108,57

(C-4); 11 1 ,67 (C-5"); 112,07 (C-5"); 112,95 (C-5); 115,49 (C-4"); 117,27 (C-4");

125,59 (C-3"); 126,40 (C-3"); 130,30 (C-1"); 130,41 (C-1"); 131 ,65 (C-6); 140,98 (C-4¹); 144,54 (C-4¹); 151 ,89 (C-1a); 15,49 (C-3a); 164,56 (C-1'); 170,56

(C-V).

IV (υ_max, KBr) υ (crrT¹): 3478, 3412, 3190, 3113, 3054, 2914, 2855, 1638, 1604,

1536, 1502, 1484, 1427, 1384, 1333, 1298, 1263, 1164, 1127, 1035, 928, 874,

744, 551.

The randomly chosen compounds were analyzed by RMN ¹H and ¹³C, in addition to IV-FT.

The new compounds of formula (I) was spectroscopically characterized and pharmalogically evaluated in in vivo and in vitro studies, with a view to determining the anti-inflammatory, analgesic and antioxidant profiles of the compounds. The animal models used in in vivo studies were carrageenan- induced mouse paw edema to investigate the anti-inflammatory profile; formalin-induced hyperalgesia in mice, and the hot plate test on mice to investigate the analgesic profile. The in vitro studies comprised a DPPH radical scavenging assay in order to investigate the antioxidant profile. The results obtained showed high anti-inflammatory, analgesic and antioxidant activities.

Particularly for the /V'-(3,5-ditercbutyl-4-hydroxyphenylmethyldene)-6~ 5 nitro-1 ,3-benzodioxol-5-carbohydrazide derivate (called "LASSBio-881") the results obtained had significant anti-inflammatory activity, with 31.7% inhibition of carrageenan-induced mouse paw edema (300 μmol/kg via oral; n=8 animals; ^*p<0,001 ANOVA ONE WAY followed by the Dunnet test). It was thus possible to identify that LASSBio-881 is anti-edematogenic. Moreover, the LASSBio-881

I O derivative was pharmacologically evaluated in terms of its analgesic properties, in the formalin-induced hyperalgesia model, showing significant activity in the neurogenic phase of the algesic process, with 44.5% inhibition (100 μmol/kg via oral; n=8 animals; *p<0,001 ANOVA ONE WAY followed by the Dunnet test). However, LASSBio-881 was not able to inhibit the inflammatory phase

15 significantly (13.4% ns) of the formalin-induced hyperalgesia test, indicating an analgesic central mechanism of analgesic action.

The pharmacological results provided by LASSBio-881 in the hot plate test in mice corroborate its central mechanism of analgesic action, and LASSBio-881 is capable of increasing the animal's latency time under thermal

20 stimulus by 51.2% (100 μmol/kg via oral; n=8 animals; ^*p<0,05 Student / test).

The radical scavenging effect was noted for the radical diphenylpycrylidrazine (DPPH). A decreased concentration was noted in the latter by UV-visible absorbance reading. Particularly the compound LASSBio- 881 (100 μM) was capable of stabilizing the radical by 35%.

25 From the results set forth above, we identified LASSBio-881 as a candidate for an anti-inflammatory drug, having central analgesic activity and antioxidant properties, proving to be useful in the treatment of acute and chronic inflammatory diseases in mammals, preferably humans. Likewise, LASSBio- 881 can be useful in treating central hyperalgesic processes associated to

30 tissue injury, inflammation or tumor growth in mammals, preferably humans.

Methodologies employed in the bio assays In vivo experiments

• Carrageenan-induced mouse paw edema

The compounds tested were administered orally, in a dose of 300 μmol/kg, using a 5% Arabic gum solution as vehicle. One hour after

5 administering, the edema was induced by subplantar injection of carrageenan 1% (1000 μg/paw). A solution of NaCI 0.9% (0.1 ml/paw) was administered in the opposite side paw. A reading was taken 3 hours after the subplantar administration of carrageenan by using a plethysmograph attached to a continual-flow peristaltic pump. The edema was expressed by the volume

K⁾ variation of the paws obtained from the difference between the paws with carrageenan and saline.

• Formalin-induced hyperalgesia in mice

The formalin assay consists of subplantar administration of formaldehyde

15 2,5% in albino mice of both sexes, weighing between 18-25g. The pain stimulus is then characterized for the time during which the animal continues to lick the paw that received the irritation stimulus. The formalin assay is characterized by having two distinct phases. The first phase (0-5 minutes after injecting formalin) is characterized by a so-called neurogenic phase, that is, having a particularly

20 relevant central component, incapable of being inhibited by centrally-acting analgesic drugs like morphine. The second phase (15-30 minutes after injecting formalin) is the so-called inflammatory phase in which mass production of inflammatory mediators such as prostaglandins occurs, being inhibited by classic NSAIDs such as acetylsalicylic acid and selective COX-2 inhibitors such ⁽ϋ>5 as nimesulid.

• Hot plate assay in mice

The central analgesic activity of the compounds was evaluated using the hot plate exposure test (55 ± 0.1 )°C, using Swiss mice of both sexes weighing 30 between 18-25g and kept without food for a period of eight hours.

The animals were place on a hot plate and their response to the thermal stimulation (withdrawal and licking of paw) were timed. A first reading was taken for adaptation of the animals and subsequently a control reading. The animals that did not respond to the hot plate stimulus for over 10 seconds were discarded. The derivatives were then administered orally in doses of 100, 300 and 500 μmol/kg. Four readings were taken at 30-minutes intervals after oral administration of the compounds. 5

In vitro assays

• DPPH radical scavenging assay

A 2ml ethanolic solution of the test compounds was added to 2ml of a

DPPH solution (1 x 10^'4 M) (Sigma), and the reaction mixture was stirred I O vigorously and kept at room temperature. The DPPH absorption was measured at 514nm. The average values were obtained from triplicate experiments.

Claims

ClaimsANTI-INFLAMMATORY AND ANALGESIC PHARMACEUTICAL COMPOSITION CONTAINING SAFROLE N-ACYLHYDRAZONE DERIVATIVES, USE, AND METHOD OF PREPARATION

1. Pharmaceutical composition for the treatment of inflammatory disturbances and/or hyperalgesia in mammals characterized by having a pharmaceutically acceptable carrier and/or dilutant and at least one derivative from the 6-nitro-/V- arylmethyldene-1 ,3-benzodioxol-5-carbohydrazide nucleus of formula (I):

wherein: R is hydrogen, alkyl, cycloalkyl, phenyl-W, furyl, tiophenyl, pyridyl, pyrimidinyl, pyrrolyl, thiazolyl, quinazolyl or isoquinolyl;

W is hydrogen, orfΛo-alkyl, ortfjo-cycloalkyl, o/f/jo-alkoxyl, o/tfro-cycloalkoxyl, o/t/70-tioxyl, OAf/7o-aryoxyl, ortøo-sulphones, o/fr>o-sulphates, ortøo-sulphoxides, o/t/70-sulphonates, orfr/o-sulphonamides, or#?o-amino, ortho-am\de, ortho- halates, ortfro-carboalkoxyl, ortΛo-carbotioalkoxyl, ort/7o-trihaloalkane, ortho- cyan, o/t/70-nitro, mefa-alkyl, metø-cycloalkyl, mefa-alkoxyl, metø-cycloalkoxyl, mefa-tioxyl, mefa-aryoxyl, meta-su\ phones, mefa-sulphates, mefa-sulphoxides, mefa-sulphonates, mefa-sulphonamides, mefa-amino, mefa-amide, meta- halates, mefø-carboalkoxyl, metø-carbotioalkoxyl, mefa-trihaloalkane, meta- cyan, mefa-nitro, para-alkyl, para-cycloalkyl, para-alkoxyl, para-cycloalkoxyl, para-tioxyl, para-aryoxyl, para-sulphones, para-sulphates, para-sulphoxides, para-sulphonates, para-sulphonamides, para-amino, para-amide, para-halates, para-carboalkoxyl, para-carbotioalkoxyl, para-trihaloalkane, para-cyan or para- nitro; or its pharmaceutically acceptable salts, solvates and/or hydrates.

2. Pharmaceutical composition, according to claim 1 , characterized by the fact that said derivatives act on the central nervous system.

3. Pharmaceutical composition, according to claim 1 or 2, characterized by the fact that said derivatives have antioxidant properties.

4. Pharmaceutical composition, according to claims 1-3, characterized by the fact that said derivatives are useful in the treatment of symptoms associated to tissue injury, inflammation and/or tumor growth.

5. Pharmaceutical composition, according to claims 1-4, characterized by the fact that it is intended for the treatment of central hyperalgesia and/or acute and/or chronic inflammatory diseases in mammals, including humans.

6. Use of a derivative of the 6-nitro-/V-arylmethyldene-1 ,3-benzodioxol-5- carbohydrazide nucleus of formula (I):

wherein: R is hydrogen, alkyl, cycloalkyl, phenyl-W, furyl, tiophenyl, pyridyl, pyrimidinyl, pyrrolyl, thiazolyl, quinazolyl or isoquinolyl; W is hydrogen, orføo-alkyl, orf/?o-cycloalkyl, orffto-alkoxyl, orffto-cycloalkoxyl, ortøo-tioxyl, OA#?o-aryoxyl, o/fr/o-sulphones, o/fr/o-sulphates, o/t/?o-sulphoxides, orf/70-sulphonates, orføo-sulphonamides, ortho-ammo, ortho-amide, ortho- halates, orfΛo-carboalkoxyl, orfr/o-carbotioalkoxyl, o/f/7O-trihaloalkane, ortho- cyan, ortho-n\tro, mefø-alkyl, mefø-cycloalkyl, mefa-alkoxyl, mefø-cycloalkoxyl, mefa-tioxyl, mefa-aryoxyl, mefa-sulphones, mefø-sulphates, mefa-sulphoxides, mefa-sulphonates, mefø-sulphonamides, mefø-amino, mefa-amide, meta- halates, mefø-carboalkoxyl, metø-carbotioalkoxyl, mefa-trihaloalkane, meta- cyan, mefa-nitro, para-alkyl, para-cycloalkyl, para-alkoxyl, para-cycloalkoxyl, para-tioxyl, para-aryoxyl, para-sulphones, para-sulphates, para-sulphoxides, para-sulphonates, para-sulphonamides, para-amino, para-amide, para-halates, para-carboalkoxyl, para-carbotioalkoxyl, para-trihaloalkane, para-cyan or para- nitro; or its pharmaceutically acceptable salts, solvates and/or hydrates, characterized by the fact that it is for the preparation of a curative or prophylactic drug for inflammatory disturbances and/or hyperalgesia in mammals including man.

7. Method of preparing the n-acylhydrazone anti-inflammatory and analgesic derivatives of formula (I):

wherein:

R is hydrogen, alkyl, cycloalkyl, phenyl-W, furyl, tiophenyl, pyridyl, pyrimidinyl, pyrrolyl, thiazolyl, quinazolyl or isoquinolyl;

W e hydrogen, orf/jo-alkyl, ortfjo-cycloalkyl, orfΛo-alkoxyl, orfr/o-cycloalkoxyl, orf/7o-tioxyl, ortfjo-aryoxyl, orfr>o-sulphones, o/t/?o-sulphates, orfr/o-sulphoxides, ort/70-sulphonates, o/tr/o-sulphonamides, orf/70-amino, orf/70-amide, ortho- halates, o/t/70-carboalkoxyl, o/tr/o-carbotioalkoxyl, orfr/o-trihaloalkane, ortho- cyan, o/f/70-nitro, mefa-alkyl, mefa-cycloalkyl, mefa-alkoxyl, mefa-cycloalkoxyl, mefa-tioxyl, mefa-aryoxyl, mefa-sulphones, mefa-sulphates, mefa-sulphoxides, metø-sulphonates, metø-sulphonamides, mefa-amino, mefa-amide, meta- halates, mefa-carboalkoxyl, mefa-carbotioalkoxyl, metø-trihaloalkane, meta- cyan, mefa-nitro, para-alkyl, para-cycloalkyl, para-alkoxyl, para-cycloalkoxyl, para-tioxyl, para-aryoxyl, para-sulphones, para-sulphates, para-sulphoxides, para-sulphonates, para-sulphonamides, para-amino, para-amide, para-halates, para-carboalkoxyl, para-carbotioalkoxyl, para-trihaloalkane, para-cyan or para- nitro; or its pharmaceutically acceptable salts, solvates and/or hydrates, characterized by comprising the stages of: a) isomerization of the safrole, followed by ozonolysis for synthesis of the aldehyde (piperonal); b) nitration of piperonal in a concentrated acid means, regioselectively; c) oxidation of the 6-nitropiperonal to the 6-nitro-1 ,3-benzodioxol-5-methyl carboxylate ester; d) obtaining the corresponding hydrazide by hydrazinolysis; e) condensation with the respective aldehydes to obtain the N- acylhydrazone ligands.