WO2016099835A1

WO2016099835A1 - Dermatological composition of papain in gel for the atraumatic removal of dermal necrosis and method of preparing same

Info

Publication number: WO2016099835A1
Application number: PCT/US2015/062727
Authority: WO
Inventors: Mauricio DOBBOLETTA
Original assignee: Brix Usa, Llc
Priority date: 2014-12-19
Filing date: 2015-11-25
Publication date: 2016-06-23
Also published as: AR098852A1

Abstract

A papain dermatological gel composition is disclosed for the atraumatic treatment of dermal necrosis which comprises papain with a final activity of at least 6,000 U/mg, wherein the papain is bio-encapsulated in a mixture of pH=7 buffer / C3-6 polyol / pectin / C2-6 alkanolamine / nonionic emulsifier, together with pharmaceutically acceptable dyeing agents, preservatives and solvents.

Description

DERMATOLOGICAL COMPOSITION OF PAPAIN IN GEL FOR THE ATRAUMATIC REMOVAL OF DERMAL NECROSIS AND METHOD OF PREPARING SAME DESCRIPTION

Field of the invention

This invention is related to the field of compositions for the treatment of the dermis, preferably those compositions related to the removal of necrotic tissue from the skin, more preferably gel compositions for the atraumatic and painless removal of necrotic dermal tissue.

Prior Art Background

Cutaneous or dermal necrosis consists in the cellular death of a portion of cutaneous tegument caused by insufficient blood supply. The most common cause of dermal necrosis is the vascular occlusion although it is also caused by exogenous agents and infections.

Several pathologies predominantly exhibit dermal necrosis; however, multiple dermatoses, sometime during their evolution, exhibit necrosis: malignant atrophic papulosis, acrosclerosis, certain stings, infant dermatomyositis, anthrax, zoster varicella virus, etc.

From a clinical point of view, the necrosis is shown as violet or blackish areas which generally turn to be covered by some eschar. Therefore, the clinical appearance of the dermal necrosis per se is not specific of any pathology.

So, in order to reach a diagnostic, thinking about a concrete disease for each case is based on the clinical history of the patient, the related diseases, other concomitant systemic or cutaneous symptoms, and the laboratory data.

Dermal necrosis is a serious process since once the necrosis is produced and developed, it is irreversible and the physician should rapidly determine the cause and administer the treatment whenever possible.

Necrosis can be defined as the pathologic cellular death able to be recognized by the morphological signs of necrophanerosis. These signs are in the cell cytoplasm: hypereosinophylia and loss of the normal structure; in the cell nucleous: pyknosis, karyolysis, or karyorrhexis. Pyknosis is the retraction of the nucleous with condensation of chromatin; karyolysis is the destruction of the nucleous; karyorrhexis is the fragmentation of the nucleous in pieces of condensed chromatin. The cytoplasm and nucleous alterations are concurrent. Pyknosis, karyolysis, and karyorrhexis are not part of the nuclear alteration; apparently they are different forms of reaction.

On the one hand, the pathological side of necrosis can be highlighted as the most serious manifestation of disease at a cellular level, and on the other hand, as the morphological base given by the signs of necrophanerosis in detecting the necrosis.

The first idea rules out from necrosis all the cellular death which is not the manifestation of disease, i.e., the apoptosis occurring during the normal cellular death in labile tissues, as those normally exposed to cell turnover such as the epidermic cells. The cellular death occurring during the remodeling process of organs under development, the cellular death of a dead organism, as well as the cellular death of cells separated from the organism and as a result of the action of fixative liquids are also ruled out, since they are not manifestations of disease.

The second idea rules out from necrosis other ways of pathological cellular death or apoptosis related to pathological conditions which are not manifested among the signs of the necrophanerosis, such as, the cellular death by which numeric atrophy may occur.

From the moment the noxa acts, morphological and functional cell alterations occur, which are reversible for a while if the noxious action stops. These lesions, called irreversible, are flocculations of the mitochondrial matrix. The cell is considered dead when the loss of vital functions is irreversible even though the noxious action is over. Signs of the cell vital functions are: cellular movements, granular staining of the cytoplasm with vital dyes without staining the nucleous, cellular respiration and the existence of membrane potential.

Necrobiosis is the cellular process occurring from the moment the cell dies to the moment the necrophanerosis occurs.

Necrophanerosis is related to structural changes which allow inferring the cell is dead. Necrolysis is the process of disintegration and destruction of the necrotic cells, which under certain conditions occurs together with polynuclear cell infiltration and removal of cellular detritus by the action of macrophages.

Necrosis includes the cellular death and the processes afterwards, i.e., necrobiosis, necrophanerosis, and necrolysis.

The nature of factors causing necrosis is as varied as those factors related to the general etiology.

Normally, two principal forms of necrosis are distinguished: coagulation necrosis and colliquative necrosis. The signs to distinguish these forms are mainly macroscopic and, therefore, they can be seen when the necrosis reaches certain dimension.

For a surgeon or a physician, the correct and suitable debridement of a cutaneous ulcer is quite significant even though there is no acute infection or significant necrosis. Considering new concepts such as the presence of metalloproteases, senscent cells or fibroblasts, bacterial biofilm, that is, all the factors that contribute to create an inflammatory, pathologic, chronic, local environment is of vital importance.

Knowing how to debride a wound is a quite necessary art that a physician should learn and develop so as to obtain the best favorable results for the patient; however, controlling the risk factors such as smoking, control of the diabetes, treatment of related venous or arterial vasculopathy, and the nutritional status are guiding elements that physicians should follow and evaluate when treating a patient with a wound of difficult cicatrization or a chronic ulcer.

So, the procedure by which all the materials incompatible with a wound undergoing healing process should be removed is known as debridement.

As per the Royal Academy Spanish Language Dictionary, to debride is "to divide, by using a cutting instrument, fibrous tissues originated from local strangulation or gangrene lesions" or "to separate the bridle or filaments that intertwine a sore producing pus".

This is the concept most widely accepted by the Latin America professionals.

To approach the debridement of chronic ulcers, several related aspects should be considered among which the following can be mentioned: inflammatory and infectious processes, how to treat an acute or chronic wound, knowledge of surgical and non-surgical debridement techniques, and identification of ischemic wounds.

It can be defined that all infected wounds are inflammatory wounds, but not all inflamed wounds are infected. All wounds should undergo a bacterial colonization phase, but not necessarily undergo a bacterial infection expression. Sixty % of chronic wounds have a bacterial biofilm, for which an infected wound exhibits erythema, edema, induration, and bad smell.

An uninfected wound classified as chronic may exhibit erythema without local signs of infection. The peri-ulcer erythema identifies the inflammation in response to an open wound. A chronic wound must be turned into an acute wound, since the bacterial biofilm is present in 60% of this type of ulceration.

Sometimes, erythema is seen around the ulcer which makes one think about the infection and cellulitis; in this case, the limb should be lifted so as to confirm their existence. If the erythema disappears or decreases, it is rubor belonging to an ischemic condition.

Chronic wounds must be turned into acute ones not only because it is an expression of the wound progress physiopathology but also because in an acute wound the phases of the healing process can be seen: inflammation, proliferation, and maturation and this pattern cannot be seen in chronic wounds which will never reach cicatrization.

In all chronic wounds, metalloproteases, which are part of the necrotic tissue, materials foreign to the ulcerous bed, and the presence of bacteria inside the biofilm should be removed, as they avoid release of cicatrization elements such as chemotactant, growth and mitogenic factors. It is of vital importance to change the surroundings of the lesion so as to generate new local sources for cure, such as in situ oxygen and nutrition.

Other two important factors to be removed from the chronic ulcers are the bacterial biofilm and the senescent cells. The senescent fibroblasts are responsible for not allowing the cicatrization phases of a wound to occur. Sixty % of chronic wounds contain bacterial biofilm, while it is present in only 6% of the acute wounds. It was demonstrated that the biofilm bacteria create some way of defense to the aggression caused by pomades, creams, or other therapeutic elements and produce the so called "bacterial (separation) detachment", which invades the site and is able to infect the chronic ulcer or wound. A wound may be colonized by aerobe bacteria and two species of anaerobe bacteria, which are present in 60% of the chronic wounds and on many occasions they are accompanied by an exudate able to decrease the content of water, electrolytes, nutrients, and growth factors.

Two ways of removing necrotic tissue are known: one way is surgical and the other by the action of proteolytic enzymes.

The surgical process entails the following difficulties: painful, traumatic, the necrotic tissue is removed and healthy tissue is affected as well, and never limited and precise enough to remove the necrotic tissue. Apart from being a time demanding process, it requires administering local anesthesia, using disposable material, etc., which ultimately turns to be uncomfortable for the patient and more expensive.

When using a bistoury or a pair of scissors as the physical means to debride, caution should be exerted so as to keep most of the healthy tissue viable by avoiding unnecessary aggression. It should be kept in mind that in the debridement areas neither electrocautery nor sutures should be used, only compression means can be used so as to produce hemostasia without trying to cauterize small vessels or to create foreign bodies in the area with suture knots.

The indispensable surgical material for a good debridement process will include: bistoury blades; curettes to remove proteinaceous clots which get accumulated and do not allow the granulation tissue to grow; "rongeur" for nipping away bone, metatarsal heads for bone biopsies. These materials can be extended including electric sagittal blades for nipping away bone. Other necessary instruments are straight or curved scissors and hemostatic or dissection forceps.

It is important to pay attention to the wounds and ulcers color during debridement. All black and grey tissue (fibrous and necrotic tissue) should be resected and leave as viable tissue only the one that appears reddish (granulation), white (tendines, fascia) or yellow (subcutaneous fat). Other detail to have in mind during debridement is that when an en-block resection is performed, an ulcerous site, only 2 mm around the ulcer should be taken as peri ulcerous tissue to be resected. This is because the senscent cells are located in this area, around the periphery. In the case of an erythematous edge greater than 5 mm in width, it should be resected because is an infected area.

In an ischemic wound, due to its complexity, well instrumented surgical treatment should be implemented. An ulcer or ischemic gangrene with clear signs of infection must be suitably and generally rapidly debrided. This is not the case for treating a dry, focused gangrene, able to justify this type of lesion, where the peripheral vascular condition should be carefully studied. In these cases, a direct arterial surgery could be recommended and debride the lesion one week after diagnosing the gangrenous condition.

The access to the injectable growth factors has allowed not accelerating the debridement process.

In order to avoid surgical procedures, mainly when treating biofilms unable to be dragged and which reappear after a few days, a hydrosurgical debridement piece was created which uses water or physiological saline solution at a pressure of about 0.1 MPa (15.0 psi) so as to debride wounds without causing damage by reducing the bacterial field and biofilm.

In those wounds for which immediate surgical or local area for debridement is not necessary, the dressings wet to dry can be used. These dressings are capable of removing dead tissue but may cause pain as a disadvantage.

There exist other techniques, such as the enzymatic debridement and the use of collagenases, which are considered as first line therapeutic within a nonsurgical debridement procedure.

For example, the maggot technique uses larvaes of Lucilia sericata fly to perform local debridement, since they are able to release a digestive proteolytic secretion which dissolves the necrotic tissue. It is also believed that this fly is able to secrete cytokines and growth factors, which increases tissue oxygenation. It has been broadly used for debridement although it is not effective in cases of venous ulcers.

Lately, low frequency ultrasound therapy has played a key role for debridement of this type of ulcer. In this procedure, a sort of cavitation producing energy able to alter cell membrane and activity is created, during which the necrotic tissue is removed, bacteria are destroyed, and the biofilm is broken. Some advantageous properties are attributed to this technology such as better oxygenation and the release of growth factors.

Surgical or non-surgical debridement on a wound which wants to be changed from chronic to acute is not the only issue to have in mind. Controlling the risk factors such as smoking, control of diabetes, treatment of related venous or arterial vasculopathies, and patient's nutritional status are elements to be followed and evaluated in a patient suffering from a hard cicatrization or chronic ulcer. Debridement on wounds even when they are not infected keeps on being an indispensable approach for treating these diseases.

At present, the use of atraumatic techniques for debridement on wounds is being imposed since most of the local healthy tissue is trying to be preserved. This healthy tissue is able to create the granulation and epithelization areas as from the release of more growth factors, oxygen, and nutrients.

Known enzymatic debriding agents up to present in the world market mainly consist of enzymes with topical antibiotics or only antibiotics. Known compositions are identified under the following commercial names: Iruxol, Clorfibrase, and Platsul.

Iruxol is a composition consisting of collagenase plus chloramphenicol. The disadvantages of this product are that not less than 25 days are needed to see granulation tissue, it is not practical because the surrounding area has to be protected with vaseline or any other healthy skin protecting product, and several healing procedures have to be performed until obtaining granulation tissue which leads to lesion cicatrization. It should be considered that each healing procedure takes at least 30 minutes.

Besides from the use of the topical antibiotic which can potentially produce bacterial resistance after a while, chloramphenicol causes bone marrow aplasia.

Reports from the Nation Ministry of Health in Argentina indicate that due to the aforementioned reason, Iruxol will be used until other alternative with less contraindications is developed.

The pharmaceutical product Clorfibrase is a composition consisting of chloramphenicol, deoxyribonuclease, and fibrinolysin. The disadvantages of this product are the following: it takes not less than 30 days to produce granulation tissue, it is not practical to use since each healing procedure takes not less than 20 minutes depending on the lesion extension, and contains chloramphenicol, which produces bone marrow aplasia and bacterial resistance after prolonged use.

The product named Platsul in the way of transparent gel contains silver sulfadiazine. Besides its indication for treating necrosis, it is an antimicrobial agent useful for cicatrization but it does not help removing necrotic tissue.

The ideal skin necrosis treatment is the one that not only removes necrotic tissue, without damaging the healthy tissue, but is also painless, does not require much instrumentation, decreases the inflammation, and improves the oxygenation. Up to the present day, there is not any known product with all these characteristics.

Summary of the Invention

The present invention refers to a gel dermatological chemical composition for manual application, the objective of which is the atraumatic removal of dermal necrosis without implementing either surgical procedures or using any type of surgical instrument.

Therefore, the object of the present invention is a dermatological composition of papain in the form of a gel for the atraumatic treatment of dermal necrosis consisting of papain with a final activity of at least 6,000 U/mg, whereby the papain is bioencapsulated in a mixture of pH=7 buffer / C₂-6-poliol / pectin / C_2- 6 alkanolamine / non-ionic emulsifier together with pharmaceutically acceptable dyes, preservatives, and solvents.

Preferably, the buffer of pH=7 is selected from disodium or dipotassium acid phosphate (Na₂HP0 , K₂HP0 )/citric acid (C₆H₈0₇), sodium chloride (NaCI)/sodium citrate (C₆H₅0₇Na3.2H₂0)/sodium hydroxide (NaOH), sodium or potassium diacid phosphate (NaH₂P0 , KH₂P0 )/disodium or dipotassium acid phosphate (Na₂HP0₄, K₂HP0₄), tris-(hydroxymethyl)-aminometane (TRIS)/hydrochloric acid (HCI).

More preferably, the pH=7 buffer is KH₂P0₄/Na₂HP0₄.

In a preferred way, pectin is the pectin from apple bagasse (cores and skin) or citrus pectin. More preferably, the pectin used is the citrus pectin.

Also preferably, the C₃-6 polyol is selected from 1 ,2,3-propanetriol (glycerine), 1 ,2-propanediol (propylene glycol), 1 ,3-butanediol, 1 ,4-butanediol, 1 ,3-butenediol, 2,3-butenediol, 2,2-dimethyl-1 ,3-propanediol (neopentyl glycol), erythritol, sorbitol, mannitol, and the mixtures thereof.

More preferably, the C₃-6 polyol is 1 ,2-propanedyol (propylene glycol).

In a preferred way also, the C₂-6 alkanolamine can be monoethanolamine, diethanolamine, triethanolamine, and the mixtures thereof.

More preferably, the alkanolamine is triethanolamine.

Preferably, the non-ionic emulsifier is selected from the group consisting of: polyoxyethylene stearate 8 (E-430); polyoxyethylene stearate 40 (E-431 ); polyoxyethylene monolaurate 20 (20 OE or ethoxylated with 20 oxyethylenes) sorbitan, also known as polysorbate 20 or Tween 20 (E-432); polyoxyethylene monooleate 20 sorbitan, polysorbate 80 or Tween 80 (E-433); polyoxyethylene monopalmitate 20 sorbitan, polysorbate 40 or Tween 40 (E-434); polyoxyethylene monostearate 20 sorbitan, polysorbate 60 or Tween 60 (E-435); polyoxyethylene tristearate 20 sorbitan, polysorbate 65 or Tween 65 (E-436); and the mixtures thereof.

More preferably, the non-ionic emulsifier is polyoxyethylene monolaurate 20 sorbitan (polysorbate 20 or Tween 20).

Additionally, the composition further comprises a dye pharmaceutically acceptable selected from methylene blue, toluidine blue, methyl green, methylene green, gentian violet, malachite green, methylene yellow, and the mixtures thereof.

In a preferred way, the pharmaceutically accepted dye is toluidine blue.

Preferably, the preservative is selected from methylparaben, ethyparaben, propylparaben, and the mixtures thereof.

In a more preferred way, the composition comprises papain (30.000 U/mg) 20,00 % weight in weight (w/w), citrus pectin 7.00 % w/w, propylene glycol 15.00 % w/w, triethanolamine (99 %) 0.72 % w/w, TW20 (polyoxyethylene sorbitan monolaurate with 20 OE) 1 .00 % w/w, KH₂P0₄ 0.0378 % w/w, Na₂HP0₄ 0.1283 % w/w, toluidine blue (aqueous solution (aq.sol.) 1 % w/w) 0.20 % w/w, methylparaben 0.10 % w/w, propylparaben 0.05 % w/w, and enough quantity of distilled water for (q.s.) 100 % w/w.

Additionally, a method for preparing the papain composition for the atraumatic removal of dermal necrosis described is proposed and comprises the following:

a) adding the components of a pH=7 buffer to distilled water under constant agitation at about 1 ,500 rpm, one or more pharmaceutically acceptable preservatives and lastly papain 30,000 U/mg until complete dispersion at a neutral final pH (pH=7);

b) moisten while stirring pectin in a C3-6 polyol, adding later a non-ionic emulsifier and finally a C2-6 alkanolamine until neutralization of the pectin acidity forming a neutral encapsulating emulsion (pH=7); and

c) adding the neutral encapsulating emulsion from step b) to the dispersion obtained in step a) under stirring at low speed so as to obtain a buffer encapsulating emulsion (BEE), in the way of a stable gel of the papain dispersion.

Preferably, the method described comprises:

a) adding potassium monobasic phosphate, sodium diacid phosphate, methylparaben, propylparaben, and 30,000 U/mg of papain to distilled water under stirring at 1 ,500 rpm until complete dispersion at a neutral final pH (pH=7), obtaining a buffer dispersion;

b) moisten citrus pectin in propylene glycol and add sorbitan monooleate and finally triethanolamine under stirring until neutralization of the citrus pectin acidity so as to form an encapsulating neutral emulsion (pH=7); and

c) add the encapsulating neutral emulsion from step b) to the dispersion from step a) under stirring at low speed so as to obtain a buffer encapsulated emulsion (BEE), in a way of a stable gel of the papain dispersion.

Preferably, the method comprises the step of adding, under stirring, a dye dissolved in a pharmaceutically acceptable solvent to the papain dispersion.

Preferably, the dye is 1 % w/w toluidine blue dissolved in distilled water. Preferably, the stirring from step a) is performed at 1 ,500 rpm.

Also preferably, the stirring from step c) is performed from 20 to 150 rpm. In a preferred way, the citrus pectin/propylene glycol/triethanolamine concentration ratio ranges from 4/4/0.36 to 10/20/2 expressed in % w/w, respectively.

In a selected way, the citrus pectin/propylene glycol/triethanolamine concentration ratio is 7/15/0.72 expressed in % w/w.

Additionally, the method comprises the step of packing the so obtained gel in tubes.

Brief Description of the Figures

Figures 1 a and 1 b correspond to photographs representative of the Example of Application 1 .

Figures 2a and 2b correspond to photographs representative of the Example of Application 2.

Figures 3a and 3b correspond to the photographs representative of the I Example of Application 3.

Figures 4a and 4b correspond to the photographs representative of the

Example of Application 4.

Figures 5a, 5b, and 5c correspond to the representative photographs of the Example of Application 5.

Figures 6a and 6b correspond to representative photographs of the Example of Application 6.

Figures 7a and 7b correspond to representative photographs of the Example of Application 7.

Figures 8a and 8b correspond to representative photographs of the Example of Application 8.

Figures 9a and 9b correspond to representative photographs of the

Comparative Example 1 .

Figures 10a, 10b, and 10c correspond to representative photographs of the Comparative Example 2.

Figures 1 1 a and 1 1 b correspond to representative photographs of the Example of Application 9.

Figures 12a and 12b correspond to representative photographs of the Example of Application 10. Figures 13a and 13b correspond to representative photographs of the Example of Application 1 1 .

Figures 14a, 14b, and 14c correspond to representative photographs of the Comparative Example 3.

Figures 15a and 15b correspond to representative photographs of the

Example of Application 12.

Detailed Description of the Invention

In order to solve the prior art issues, a papain topical gel composition is proposed for dermatological use, which is recommended for the removal of dermal necrosis.

The gel composition as per the present invention comprises a concentrated proteolytic enzyme and a series of advantages, in comparison with the products known so far, were found, some of these advantages are: a granulation tissue is obtained within a time period of 12 to 48 hours; the necrosis is removed on an absolutely pain-free basis; since the gel is inhibited in contact with healthy tissue, it is not systemically absorbed and only acts on necrotic tissue; it is quite practical to use since the patient himself or a family member can apply it; the necrotic tissue, the infected fibrin, and the secretion are removed just by washing the area after treatment; it is bactericidal, anti-inflammatory, and vasodilator, all of which leads to a granulation tissue of unbeatable quality where a rapid cicatrization will be obtained later; it is accessible to be used by all the population.

Without limiting the product use, this product results to be mainly useful for treating: all etiology skin necrosis; venous ulcers; arterial ulcers; sacral, coxis eschars due to decubitus, etc.; necrosis secondary to trauma; second and third degree burns; abrasions secondary to trauma; necrosis due to vasculitic processes; skin necrosis secondary to insects stings; and necrosis in post-surgical scars.

In this way, the gel presented herein contains a single component based on papain which is different from the other known products in that the active ingredient, the papain enzyme, is forming a dispersion at values of at least 6,000 U/mg and is occluded in a buffer encapsulating emulsion (BEE), as an encapsulating, moisturizing, and stabilizing agent, which immobilizes and stabilizes it and exponentially increases the enzymatic activity of the final product within the storage time.

Therefore, a greater proteolytic effectivity to remove collagen fibers in necrotic tissue is reached without the need to isolate the surrounding healthy zone since the gel does not act on healthy tissue, is dermatologically safe and harmless, even for the ocular mucosa, is more resistant to storage, even under unfavorable conditions because no refrigeration is required, and has stronger antibacterial and antifungal potency with an increase in its antiseptic power at tissue level. Likewise, the proposed gel does not contain chloramphenicol in its formula, which causes bone marrow aplasia and bacterial resistance after prolonged use, for which the gel is of maximum safety.

The active ingredient is papain, a vegetal protease useful to eliminate necrotic tissue from cutaneous lesions in the presence of any encapsulating agent which provides stability such as a biopolymer.

Pectin is a polysaccharide, a high molecular weight macromolecule and one of the main constituents of the cellular wall in superior plants, responsible of the characteristic texture of vegetables. Pectins are broadly used in the food, pharmaceutical, and cosmetic industries mainly as thickening and stabilizing agents.

As per the present invention, compositions have been obtained with values of at least about 6,000 U/mg, which significantly outdo the accomplishments in the prior art.

In order to achieve this unexpected result, work proceeded on a papain bioencapsulation system by dispersing it in a pH=7 buffer / C_3-6 polyol / pectin- C₂-6 alkanolamine /non-ionic emulsifier.

A preferred embodiment of present invention is a gel papain dermatological composition for the atraumatic removal of dermal necrosis comprising papain with a final activity of at least 6,000 U/mg, where the papain is bioencapsulated in a mixture of pH=7 buffer / C3-6 polyol / pectin / C2-6 alkanolamine /non-ionic emulsifier, together with pharmaceutically acceptable dyes, preservatives, and solvents. As per the present invention, the pectin used to bioencapsulate the papain is from the apple bagasse (cores and skin) or citrus pectin. More preferably, for the gel composition of present invention citrus pectin is used.

Preferably, the pH=7 buffer is selected from disodium or dipotassium acid phosphate (Na₂HP0 , K₂HP0 )/cithc acid (C₆H₈0₇), sodium chloride (NaCI)/sodium citrate (C₆H₅0₇Na₃.2H₂0)/sodium hydroxide (NaOH), sodium or potassium diacid phosphate (NaH₂P0 , KH₂P0 )/disodium or dipotassium acid phosphate (Na₂HP0₄, K₂HP0₄), and tris-(hydroxymethyl)-aminometane (TRIS)/hydrochloric acid (HCI).

In a preferred embodiment, the pH=7 buffer used in the composition of the invention is the potassium diacid phosphate (KH₂P0₄)/disodium acid phosphate (Na₂HP0₄).

Likewise, also preferably, the C₃-6 polyol used in the dermatological composition of the invention is 1 ,2,3-propanetriol (glycerine), 1 ,2-propanediol (propylene glycol), 1 ,3-butanediol, 1 ,4-butanediol, 1 ,3-butenediol, 2,3-butenediol, 2,2-dimethyl-1 ,3-propanediol (neopentilglycol), erythritol, sorbitol, mannitol.

More preferably, the C₃-6 polyol is 1 ,2-propanediol (propyleneglycol).

Propylene glycol, or propane-1 ,2-diol, is an organic compound, an alcohol, or more precisely a colorless, tasteless, and odorless diol. It is an oily, clear, hygroscopic substance miscible in water, acetone, and chlorophorm. It is obtained by the hydration of propylene oxide.

The U.S. Food and Drug Administration has determined that the propyleneglycol is "generally considered safe" (GRAS, stands for Generally Recognized as Safe) for use in nutrition, cosmetics, and medicines. Propylene glycol is naturally metabolized in exercised muscles.

In this way, using this compound, the gel remains stable, even when it is uncapped, without either getting dry or forming a plug able to block the mouth of the container. On the other hand, the C_2-6 alkanolamine used in the composition as per the present invention to neutralize the acid pH can be monoethanolamine, diethanolamine, or triethanolamine. Preferably, the C_2-6 alkanolamine used is triethanolamine.

Triethanolamine is an organic chemical compound mainly formed by a tertiary amine and three hydroxile groups, of general chemical formula C6H15NO3. As with other amines, the triethanolamine acts as weak chemical base due to the electron pair available on the nitrogen atom.

It appears as a viscous liquid, although it may be solid when impure, depending on the temperature; it is a pale yellow or colorless limpid liquid, slightly hygroscopic and volatile, completely soluble in water and miscible with most oxygen-containing organic solvents. It has a slight ammonia-like odor.

This chemical is used for pH adjustment in cosmetics, hygiene, and cleaning preparations. Among the cosmetic and hygiene products, we may include skin lotions, eye gels, moisturizers, shampoos, shaving foams, etc.

A non-ionic emulsifier is also used in a preferred embodiment of the composition of the invention, which is selected from the group consisting of: polyoxyethylene stearate 8 (E-430); polyoxyethylene stearate 40 (E-431 ); polyoxyethylene monolaurate 20 (20 EO or ethoxylated with 20 oxyethylene groups) sorbitan (also known as polysorbate 20, Tween 20 or E-432); polyoxyethylene sorbitan monooleate 20 (polysorbate 80, Tween 80 or E-433); polyoxyethylene sorbitan monopalmitate 20 (polysorbate 40, Tween 40 or E-434); polyoxyethylene sorbitan monostearate 20 (polysorbate 60, Tween 60 or E-435); polyoxyethylene sorbitan tristearate 20 (polysorbate 65, Tween 65 or E-436); and mixtures thereof.

Particularly, according to a preferred embodiment of the composition of the invention, polyoxyethylene sorbitan monolaurate 20 (Tween 20) is used as the non-ionic emulsifier.

It is an additive used in the food industry which acts as a non-ionic, lipophilic detergent emulsifying and dissolving fat. It has been approved by the European Community for use in Food and it is identified as Emulsifier 433 (E-

433).

The composition according to the invention further comprises a pharmaceutically acceptable coloring agent.

Said pharmaceutically acceptable coloring agent is selected from methyl blue, toluidine blue, methyl green, methylene green, gentian violet, malachite green, methylene yellow, and mixtures thereof.

Particularly, the pharmaceutically acceptable coloring agent used in the composition of the invention is toluidine blue. Toluidine blue is a coloring agent which also acts as a powerful antimicrobial agent by fixing itself to the bacterial wall, being a non-toxic photosensitizing agent because most bacteria do not absorb visible light.

Finally, the preservative used in the gel composition is selected from the group consisting of methylparaben, ethylparaben, propylparaben and mixtures thereof. Preferably, in a preferred embodiment of the composition, a combination of methylparaben and propylparaben is used.

A preferred embodiment of the composition comprises papain (30,000 U/mg) 20.00% w/w, citrus pectin 7.00% w/w, propyleneglycol 15.00% w/w, triethanolamine (99%) 0.72% w/w, polyoxyethylene sorbitan monolaurate 20 EO (Tween 20) 1 .00% w/w, KH₂P0₄ 0.0378% w/w, Na₂HP0₄ 0.1283%, toluidine blue (1 % w/w aq. sol.) 0.20% w/w, methylparaben 0.10% w/w, propylparaben 0.05% w/w, and distilled water q.s. 100% w/w.

The secondary ingredients used are also used in the food industry, which grants toxicology safety.

It is another object of the present invention to provide a method of preparing the papain composition for the atraumatic removal of dermal necrosis as described above, which comprises:

a) adding to distilled water while stirring at approximately 1 ,500 rpm the components of a pH=7 buffer, one or more pharmaceutically acceptable preservatives, and finally 30,000 U/mg papain until complete dispersion to a final neutral pH (pH=7);

b) moisten pectin while stirring in a C3-6 polyol, then adding the non-ionic emulsifier, and finally a C_2-6 alkanolamine until neutralization of the pectin acidity thus forming a neutral encapsulating emulsion (pH=7); and

c) adding the neutral encapsulating emulsion from step b) to the dispersion obtained in step a) under low speed stirring to obtain a buffer encapsulating emulsion (BEE), in the form of a stable gel of the papain dispersion.

The described method preferably comprises:

a) adding to distilled water while stirring at approximately 1 ,500 rpm potassium monobasic phosphate, sodium diacid phosphate, methylparaben, propylparaben, and finally 30,000 U/mg papain until complete dispersion to a final neutral pH (pH=7); b) moisten while stirring citrus pectin in propylene glycol, adding sorbitan monooleate, and finally triethanolamine until neutralization of the citrus pectin acidity thus forming a neutral encapsulating emulsion (pH=7); and

The method further comprises the step of adding, while stirring, to the stable gel of the papain dispersion a pharmaceutically acceptable coloring agent dissolved in a pharmaceutically acceptable solvent.

Preferably, the coloring is 1 % w/w toluidine blue dissolved in distilled water.

Preferably, the stirring in step a) is performed at approximately 1 ,500 rpm, and the stirring in step c) is performed between approximately 20 and approximately 150 rpm.

More preferably, the concentration ratio of citrus pectin:propylene glycoktriethanolamine employed in the method ranges from 4:4:0.36 to 10: 10:2 expressed in % w/w, respectively.

Still more preferably, the concentration ratio of citrus pectin:propylene glycoktriethanolamine is 7:15:0.72 expressed in % w/w. In this system, the papain shows greater activity.

Once the desired dermatological composition is obtained, it is analyzed to verify it is in proper condition to be fractionated. For this purpose, the method comprises the step of packing the obtained gel into tubes made of pharmaceutically acceptable material.

In this way, the gel is packed into plastic tubes of 15, 30, and 60 g of capacity. The secondary packing is performed in cardboard boxes, each containing a printed tube and a package insert. These corner boxes are sealed so as to render them tamper-proof.

EXAMPLES

Example 1 : Manufacturing of the composition of the invention

In a 316 stainless steel reactor equipped with a lower dispersing disc and a

W-shaped blade for gel stirring, which are independently operated by means of a double shaft and an engine to stir the formula of this encapsulating system. The total amount of distilled water is added into the reactor with the dispersing system spinning at 1 .500 rpm; then the potassium monobasic phosphate and the sodium diacid phosphate are added; then, methyl paraben and propyl paraben; and finally 30,000 U/mg papain is added until complete dispersion.

Unlike other compositions, a papain dispersion in a pH=7 buffer solution is obtained instead of a solution in distilled water without any pH control over time, which is essential for enzymatic stability; a non-aqueous papain solution in polyethylene glycol is not obtained either, which is a product that would react with strong oxidizing agents. If the dispersing agent is stopped, most of the enzyme would precipitate, this is why the stirring continues and the following emulsion, separately prepared in a 316 stainless steel container, is added. The citrus pectin is moistened with propylene glycol, polyoxyethylene sorbitan monolaurate 20 EO (Tween 20) is added, an excellent moisturizing and tensioactive agent to significantly improve the power of the gel to moisturize and penetrate onto the wound, which increases the efficacy of the final product application. Then, triethanolamine is added to neutralize the acidity of the citrus pectin thus forming a neutral emulsion (pH=7), therefore the pH will not be modified when it is added to the previously described papain dispersion.

When adding the neutral encapsulating emulsion to the papain dispersion the W-shaped blade is turned on at approximately 150 rpm thus obtaining a stable gel from the papain dispersion. Once it is homogenous, the gel is ready to be fractionated.

Example 2: Selection of the papain concentration in citrus pectin: propylene glycol: triethanolamine.

1 g of citrus pectin is moistened and the formed emulsion is easily solubilized in distilled water. It is concluded that 1 g of citrus pectin is well moistened from 1 g of propylene glycol. Then 0.09 g of triethanolamine is added to achieve pH=7. Therefore, the mass ratio for citrus pectin:propylene glycoktriethanolamine is 1 :2:0.09 in % w/w, respectively. Different solutions were prepared by varying the concentration of citrus pectin:propylene glycoktriethanolamine while keeping the above developed ratio, namely: 4:8:0.36; 6:12:0.54; 7:15:0.72; 10:20:0.90 in % w/w, respectively, using distilled water as the solvent. A final 20% w/v concentration of papain enzyme was added to the solutions. In order to assess the effect of the different concentrations of citrus pectin:propylene glycoktriethanolamine on the enzyme, the enzymatic activity expressed by the papain was estimated through the Kunitz spectrophotometric method (1946). Said method employs a reaction mixture composed of 2 ml of 0.1 M buffer phosphate, pH=6; 1 mL of 1 % casein (substrate) and 1 ml of enzymatic extract sample (papain in citrus pectin:propylene glycoktriethanolamine); the mixture is incubated for 30 minutes at 37 °C, when this period is over the reaction is quenched with 20% trichloroacetic acid and it is allowed to stand for 20 minutes. To separate the enzymatic hydrolysis products, the mixture was centrifuged at 3,500 rpm for 30 minutes in an AWEL MF 20 Nuaire instrument.

Finally, the supernatant was separated and the peptide absorbance equivalent to 1 μg of tyrosine released per minute was measured at 280 nm in a T60-UV PG Instruments UV-VIS Spectrophotometer (190-1 100 nm) having a 2 nm bandwidth. A 0.005 ml μg^"1 cm^"1 molar extinction coefficient was used.

The results are shown in Figure 1 , where it can be seen that the papain enzyme immovilized in the neutral encapsulating emulsion increased its activity by 30% with respect to the free enzyme and, without intending to be tied to any theory, this result seems to indicate that the support somehow worked on the enzyme active conformation.

By using the concentration ratio 7:15:0.72 expressed in % w/w for the neutral encapsulating emulsion, an activity increase greater than 59% was obtained as compared to the enzyme in solution; this was the greatest value observed, and for this reason, that ratio was selected for the formulation of the composition according to the present invention as it showed the greatest activity. Higher concentrations showed decreased activity, even close to the free enzyme activity and, again, without intending to be tied to any theory, this may be due to steric hindrance of the active site of the enzyme and low diffusion rate of the substrate. 1 % polyoxythelene sorbitan monolaurate 20 EO is added to the selected encapsulating emulsion before neutralization with triethanolamine in order to increase its moisturizing power, then the papain dispersion prepared according to Example 1 and the coloring agent are also added, thus obtaining a preferred embodiment of the gel composition according to the present invention:

Example 3: Verification of the enzymatic stability and activity of the gel dermatological composition in relation to temperature.

Samples of a composition manufactured according to the method described in the present invention were taken with the objective of analyzing the residual enzymatic activity at different temperatures: room temperature and a higher temperature selected as the maximum temperature which the composition might be exposed to during storage at room temperature in summer.

One Sample (25 °C) was analyzed at a room temperature of 25 °C and the other Sample (36 °C) was analyzed at 36 °C after a storage period of 48 hours at that temperature; the results for both samples are shown in the above Table.

The enzymatic activity, far from decreasing, increased approximately by 5.12%. Therefore, it can be concluded that the composition of the present invention is acceptably stable as to be continuously marketed and used without the need to be kept under cooling conditions.

APPLICATION EXAMPLES

The present Application Examples were performed by means of an atraumatic restoring technique using the gel composition as per the present invention. The recommended procedure is based on applying the gel uniformly onto the necrotic skin zone, dressing the area, and repeat the procedure from 4 to 6 times a day for 1 to 2 days at the most until the necrotic dermal tissue is removed and granulation tissue is obtained. The results exhibited were obtained with patients of different ages and the gel composition was applied by physicians with minimal technical training so as to prove the efficacy of the composition of the present invention.

Each application lasts about 4 to 6 hours during which the gel is in contact with the necrotic tissue, the wound is covered with clean or aseptic cotton dressing, and there is no need to use either a bistoury or local anesthetics.

Four to six hours after each application the zone was washed with physiological solution and if any necrotic tissue still remained, this procedure was repeated until obtaining granulation tissue.

Whiting this recommended approach, the treatment total duration as well as the application frequency will depend on the wound seriousness and the patient progress.

The gel composition in the present invention has no contraindications and has no drug-drug interactions of any kind as well.

Example of Application 1

Twenty year old patient with a wound in his forearm close to the elbow after falling off a motorcycle 3 days ago and before starting the debridement treatment.

The wound was treated with the gel composition as per the present invention and as described before. Figure 1 a shows the wound before starting treatment and Figure 1 b shows the same wound after 2 hours of the gel application.

The process was pain-free and the wound was washed with physiological solution.

Example of Application 2

Twenty-two-year old patient with abrasion in his lower jaw bone due to fall off the motorcycle. The debridement treatment was performed with the gel composition of present invention after 48 hours of progression.

Figure 2a shows the wound before performing the treatment and Figure 2b shows the same wound after two hours of the gel composition application. Treatment was pain-free and the treated wound was washed only with physiological solution.

Example of Application 3

Twenty-seven-year old patient with significant abrasion on the right knee which was produced 48 which hours before (see Figure 3a).

The gel composition of the present invention was applied for two hours and, after washing it with physiological solution, the wound appeared as shown in Figure 3b. Treatment was pain-free.

Example of Application 4

Seventy-eight-year old patient with an ulcer on the right ankle of three years of progression (see Figure 4a).

After 12 hours of treatment with the composition of the present invention, the wound was cleaned with physiological solution and appeared as shown in Figure 4b.

The granulation tissue is visible without the presence of fibrin or secretion.

This was a pain-free accomplishment due to the application of the gel composition and later washing only with physiological solution.

Example of Application 5

Thirty- two-year old patient with a gasoline burn on the right shank of 4 days of progression (see Figure 5a).

The composition of the present invention was applied as shown in Figure

5b.

After twelve hours of treatment, by only washing the wound with physiological solution, and without pain, the lesion appeared as shown in Figure 5c. It can be seen that the inflamed area was significantly reduced.

Example of Application 6

Twenty-eight-year-old patient with abrasion on the left wrist of 72 hours of progression due to an accident in the street (see Figure 6a).

After 24 hours of treatment with the composition of the present invention, the wound progressed as shown in Fig. 6b.

Notice that the other smaller abrasions, where the gel composition of the present invention was not applied, remain intact as in the first photograph.

Example of Application 7 Figure 7 a shows the left knee of an eighteen-year-old patient with abrasion due to a fall of 24 hours of progression.

Figure 7b shows the wound after one hour of treatment with the composition of the present invention.

The wound was washed with physiological solution and the treatment was completely free of pain.

Example of Application 8

Figure 8a shows the left shoulder of a 25-year-old patient with abrasion caused by a fall while practicing sports.

Figure 8b shows the same wound after two hours of treatment with the gel composition of the present invention.

The treatment was pain-free and the wound was just washed with physiological solution after treatment.

Example of Application 9

Figure 1 1 a shows the right knee of a 24-year-old patient with a knee trauma due to a fall in the street after 24 hours of progression.

After a 2-hour treatment with the gel composition of the present invention, the wound progressed as shown in Figure 1 1 b.

Pain-free granulation tissue formation is seen.

Example of Application 10

Figure 12a shows the lower right limb of a 64-year old patient with post- trauma skin necrosis due to a fall after 7 days of progression.

Figure 12b shows the wound after twelve hours of treatment with the gel composition of the present invention.

It can be seen that the applied composition does not affect the healthy tissue which exhibits defined margins. The treatment was pain-free and only physiological solution was used to wash the wound.

Example of Application 11

Figure 13a shows the gluteal zone of an 82-year-old patient with an ulcer due to decubitus of 8 months of progression.

The gel composition of the present invention was applied to the wound for twelve hours in a way that the wound progressed as shown in Figure 13b. The wound shows granulation without either secretion or fibrin. Example of Application 12

Figure 15a shows a cutting wound with central necrosis plus peripheral infection in the left calf of a 40-year old patient after 3 days of progression.

Figure 15b shows the wound progress after 12 hours of treatment with the gel composition of the present invention.

Notice that, apart from having cleaned all the necrosis and fibrin, the peripheral cellulitis was reduced by 80%.

COMPARATIVE EXAMPLES

A series of Comparative Examples was performed to compare the effectivity between compositions of the prior the art versus the composition of the present invention.

In all the cases detailed below, patients had been unsuccessfully treated with some of the compositions of the prior art. Later, patients were treated with the gel composition of the present invention and positive results were obtained much sooner.

Comparative Example 1

Figure 9a shows a post-surgical wound after 6 months of progression in a 75-year-old patient. Throughout these six months, the wound was treated with silver sulfadiazine.

Figure 9b shows the same wound after treatment was switched to applying the composition of the present invention for twelve hours, where granulation tissue is seen.

Treatment was pain-free and the area was washed with physiological solution only.

Comparative Example 2

Figure 10a shows a vascular ulcer of 3 years of progression in an 82-year- old patient, which was previously treated with collagenase plus chloramphenicol. It is important to observe the fibrin, secretion, and central necrosis as a result of the treatment.

The ulcer treatment was modified by using the composition of the present invention, and after 12 hours the appearance of the ulcer was as shown in Figure 10b, i.e., 80 % of granulation tissue can be seen. Treatment continued by applying one more time the gel composition for 24 hours and the wound progressed as shown in Figure 10c. One-hundred % of granulation tissue was obtained in 48 hours of pain-free treatment.

Comparative Example 3

Figure 14a shows a vascular ulcer on the left lower limb of a 65 year-old patient with one year of progression during which time the lesion was treated with Iruxol and Platsul.

The wound treatment was switched to the gel composition of the present invention (see Figure 14b).

Twelve hours passed with the gel composition applied on the wound, which was later washed with physiological solution, and a favorable progression unreached with the previous treatment is shown in Figure 14c.

Claims

1 . A papain dermatological composition in the form of gel for the atraumatic treatment of dermal necrosis comprising papain with a final activity of at least 6.000 U/mg, wherein the papain is bioencapsulated in a mixture of pH=7 buffer / C3-6 polyol / pectin / C2-6 alkanolamine / non-ionic emulsifier, together with pharmaceutically acceptable dyes, preservatives, and solvents.

2. The composition according to claim 1 , wherein the pH=7 buffer is selected from the group consisting of: dissodium or dipotassium acid phosphate (Na₂HP0₄, K₂HP0₄)/cithc acid (C₆H₈0₇), sodium chloride (NaCI)/sodium citrate (C₆H₅0₇Na₃.2H₂0)/sodium hydroxide (NaOH), sodium or potassium diacid phosphate (NaH₂P0 , KH₂P0 )/disodium or dipotassium acid phosphate (Na₂HP0₄, K₂HP0₄), tris-(hydroxymethyl)-aminometane (TRIS)/hydrochloric acid (HCI).

3. The composition according to claim 2, wherein the pH=7 buffer is KH₂P0₄/Na₂HP0₄.

4. The composition according to claim 1 , wherein the C3-6 polyol is selected from the group consisting of: 1 ,2,3-propanetriol (glycerine), 1 ,2- propanediol (propylene glycol), 1 ,3-butanediol, 1 ,4-butanediol, 1 ,3-butenediol, 2,3- butenediol, 2,2-dimethyl-1 ,3-propanediol (neopentyl glycol), erythritol, sorbitol, mannitol, and their mixtures thereof.

5. The composition according to claim 4, wherein the C3-6 polyol is 1 ,2- propanediol (propylene glycol).

6. The composition according to claim 1 , wherein the pectin is pectin from apple or citrus bagasse.

7. The composition according to claim 6, wherein the pectin used is citrus pectin.

8. The composition according to claim 1 , wherein the C₂-6 alkanolamine is selected from the group consisting of: monoethanolamine, diethanolamine, triethanolamine, and the mixtures thereof.

9. The composition according to claim 8, wherein the C₂-6 alkanolamine is triethanolamine.

10. The composition according to claim 1 , wherein the non-ionic emulsifier is selected from the group consisting of: polyoxyethylene stearate 8 (E- 430); polyoxyethylene stearate 40 (E-431 ); polyoxyethylene sorbitan monolaurate 20 (20 EO or etoxylated with 20 oxyethylene groups) (also known as polysorbate 20, Tween 20 or E-432); polyoxyethylene sorbitan monooleate 20 (polysorbate 80, Tween 80 or E-433); polyoxyethylene sorbitan monopalmitate 20 (polysorbate 40, Tween 40 or E-434); polyoxyethylene sorbitan monostearate 20 (polysorbate 60, Tween 60 or E-435); polyoxyethylene sorbitan tristearate 20 (polysorbate 65, Tween 65 or E-436); and the mixtures thereof.

1 1 . The composition according to claim 10, wherein the non-ionic emulsifier is polyoxyethylene sorbitan monolaurate 20 (Tween 20).

12. The composition according to any of claims 1 , wherein the composition also comprises a pharmaceutically accepted dye selected from the group consisting of: methylene blue, toluidine blue, methyl green, methylene green, gentian violet, malachite green, methylene yellow, and the mixtures thereof.

13. The composition according to claim 12, wherein the pharmaceutically acceptable dye is toluidine blue.

14. The composition according to claim 1 , wherein the preservative is selected from the group consisting of: methylparaben, ethylparaben, propylparaben, and the mixtures thereof.

15. The composition according to claim 14, wherein the preservative is the mixture of methylparaben and propylparaben.

16. A papain dermatological composition in the form of gel for the atraumatic treatment of dermal necrosis, wherein the composition comprises 20.00 % w/w papain (30.000 U/mg), 7.00 % w/w citrus pectin, 15.00 % w/w propylene glycol, (99 %) 0.72 % w/w triethanolamine, 1 .00 % w/w polyoxyethylene sorbitan monolaurate 20 EO (TW 20), 0.0378 % w/w KH₂P0₄, 0.1283 % w/w Na₂HP0₄, 0.09% w/w toluidine blue (1 % w/w aq. sol.), 0.10 % w/w methylparaben, 0.05 % w/w propylparaben, and 100 % w/w distilled water q.s.

17. A method of preparing a papain dermatological composition in the form of gel for the atraumatic treatment of dermal necrosis, the method comprising:

a) adding to distilled water while stirring at approximately 1 ,500 rpm the components of a pH=7 buffer, one or more pharmaceutically acceptable preservatives, and finally 30,000 U/mg papain until complete dispersion into a final neutral pH (pH=7);

b) moisten pectin while stirring in a C3-6 polyol, then adding the non- ionic emulsifier, and finally a C_2-6 alkanolamine until neutralization of the pectin acidity thus forming a neutral encapsulating emulsion (pH=7); and

18. The method according to claim 17, comprising the step of adding under stirring to the stable gel of the papain dispersion a dyeing agent dissolved in a pharmaceutically acceptable solvent.

19. The method according to claim 18, wherein the dyeing agent is 1 % w/w toluidine blue dissolved in distilled water.

20. The method according to claim 17, wherein stirring in step a) is performed at 1 ,500 rpm.

21 . The method according to claim 17, wherein stirring in step c) is performed from 20 to 150 rpm.

22. The method according to any of claims 17, wherein the concentration ratio of citrus pectin/propylene glycol/triethanolamine employed ranges from 4/4/0.5 to 10/20/2 expressed in % w/w, respectively.

23. The method according to claim 22, wherein the concentration ratio of citrus pectin/propylene glycol/triethanolamine is 7/15/0.72 expressed in % w/w.

24. The method according to any of claims 17 to 23, wherein the method further comprises the step of packing the gel into tubes.

25. A method of preparing a papain dermatological composition in the form of gel for the atraumatic treatment of dermal necrosis, the method comprising:

a) adding to distilled water while stirring at approximately 1 ,500 rpm potassium monobasic phosphate, sodium diacid phosphate, methylparaben, propylparaben, and finally 30,000 U/mg papain until complete dispersion into a final neutral pH (pH=7);

b) moisten while stirring citrus pectin in propylene glycol, adding polyoxyethylene sorbitan monolaurate 20 (Tween 20), and finally triethanolamine until neutralization of the citrus pectin acidity thus forming a neutral encapsulating emulsion (pH=7); and

26. The method according to claim 25, wherein the method further comprises the step of packing the gel into tubes.

27. Use of papain bioencapsulated in a mixture of pH=7 buffer / C₃-6 polyol / pectin / C₂-6 alkanolamine / non-ionic emulsifier, wherein the papain has a final activity of at least 6.000 U/mg, for manufacturing a dermatological composition in the form of gel for the atraumatic treatment of dermal necrosis in a patient in need thereof.

28. The use according to claim 27, wherein the pH=7 buffer is KH₂P0₄/Na₂HP0₄.

29. The use according to claim 27, wherein the C3-6 polyol is 1 ,2- propanediol (propylene glycol).

30. The use according to claim 27, wherein the pectin is citrus pectin.

31 . The use according to claim 27, wherein the C_2-6 alkanolamine is triethanolamine.

32. The use according to claim 27, wherein the non-ionic emulsifier is polyoxyethylene sorbitan monolaurate 20 (Tween 20).

33. The use according to claim 27, wherein the pharmaceutically acceptable dye is toluidine blue.

34. The use according to claim 27, wherein the preservative is the mixture of methylparaben and propylparaben.

35. The use according to any of claims 27 to 34, wherein the composition is useful for the atraumatic removal of dermal necrosis without implementing either surgical procedures or using any type of surgical instrument.