BR102017022753A2 - Anesthetic and Antiseptic Composition Based on Acmelella Oleracea for Integral Skin - Google Patents

Abstract

Anesthetic and antiseptic composition based on acmella oleracea for intact skin. The present patent application is an acmella oleracea based anesthetic and antiseptic composition for intact skin. The composition has the function of reducing the sensitivity of the skin, relieving the pain during the venipuncture procedure and the use of needle and / or syringe, which can be a valuable instrument to be used in the medical industry. The present invention is a product made from 90% acmella oleracea extract with 10% transcutol® which can be used as an aid in invasive skin needle procedures and may be used as a topical and antiseptic analgesic. . the aqueous extract of a. oleracea when manipulated with transcutol® has been shown to be effective in reducing pain level and reducing skin microbiota during venipuncture.

Description

“ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN”

Field of application [001] The present patent application is an anesthetic and antiseptic composition based on Acmella oleracea for healthy skin. The composition has the function of decreasing the sensitivity of the skin, relieving pain during the venipuncture procedure and when using a needle and / or syringe, which can be a valuable instrument to be used in the medical industry.

[002] The present invention is a product made from the extract of Acmella oleracea 90% with Transcutol® 10% that can be an aid in invasive procedures performed with a needle in the skin, and may be used as an analgesic topical and antiseptic. The aqueous extract of A. oleracea when manipulated with Transcutol ^® has been shown to be effective in reducing the level of pain and reducing the skin microbiota during venipuncture.

State of the art [003] Since the dawn of mankind, medicinal plants are resources for the treatment of various diseases. The World Health Organization (WHO) indicates that 80% of the world population has already used medicinal plants.

[004] Brazil has a rich history in the use of medicinal plants in the treatment of the population's health problems. At the national level, plants from Brazilian biomes have been used in traditional medicine by the population to treat a variety of diseases. It is believed that the care performed through the use of plants is favorable to human health as long as the user is aware of its purpose, risks and benefits. From the knowledge of medicinal plants, herbal medicines emerged. Phytotherapy, phyton (plant) and therapia (treatment) is described as an old practice, which has been growing among preventive and curative programs, stimulating research that evaluates plant extracts in the search for new products with greater therapeutic activity, low cost, biocompatibility and less toxicity. Herbal medicines are regulated drugs in Brazil that must meet similar quality and safety criteria to conventional ones.

[005] Most invasive skin procedures involve pain and discomfort, adequate topical analgesia is essential in the treatment of pain, especially with regard to

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2/10 regarding the performance of invasive procedures, such as venopuncture.

Venipuncture is a procedure that consists of accessing the bloodstream through appropriate devices, together with a careful selection of the puncture site and an efficient vein penetration technique.

[006] Transcutol® or diethylene glycol monoethyl ether is a substance that has a long history of use in cosmetics and topically applied products and is used in formulations as a co-surfactant and penetration enhancer.

[007] Since puncture is an invasive procedure, antisepsis measures are necessary to prevent bacterial contamination of the skin in the place where this procedure will be performed. Topical antiseptics should have rapid antimicrobial action, persistent residual effect and should not be toxic, allergenic or irritating.

[008] Currently, 70% alcohol is used in antisepsis of venipuncture in the vast majority of health services. The antiseptic activity of alcohol occurs by denaturing proteins and removing lipids, including the envelopes of some viruses. To achieve maximum germicidal activity, alcohol must be diluted with water, which allows protein denaturation. The recommended concentration to achieve greater microbicidal speed is 70% (KAMPF G, KRAMER A. Epidemiologic background of hand hygiene and evalution of the most important agents for scrubs and rubs. Clin Microbiol Rev. 2004; 17 (4): 863-893 ).

[009] Acmella oleracea (Spilanthes oleracea; Spilanthes acmella var.oleracea popularly known in Brazil as jambu is a vegetable distributed in tropical and subtropical regions, with important chemical properties.

[010] Some biological activities have been described for A. oleracea, such as anesthetic, anti-inflammatory, analgesic and antipyretic, anti-obesity, diuretic, antioxidant, antimutagenic, insecticide and bacteriostatic.

[011] The physicochemical composition of fresh leaves of A. oleracea contains calcium, phosphorus, iron, water, proteins, lipids, carbohydrates, fiber, ash, vitamin B1, B2, niacin, vitamin C and other chemical elements. The pharmacological effect is due to its chemical substances, among which trans-karyophylene, germacrene D, Ldodecene, spatulenol and spilantol. The presence of N-alkylamides suggests that the therapeutic effect of Acmella oleracea is related to its greater anesthetic activity. A. oleracea is a plant commonly used in cases of toothache and stomatitis, in

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3/10 form of gargle. In folk medicine, the plant is often used as a local anesthetic to combat toothache, due to the presence of spilantol, also used in toothpaste and chewing gum. Several patents related to the use of spilantol or A. oleracea extract in toothpastes and other oral hygiene formulations were filed in the 80s and 90s, especially in Japan (CAVALCANTI, VMS. Spilanthes acmella var. Oleracea spilantol extraction with dioxide of superocytic carbon. [Thesis], Campinas: State University of Campinas, Faculty of Chemical Engineering; 2008).

[012] The antifungal and bacteriostatic activity of spilantol has also been studied (BARBOSA AF, CARVALHO MG, SMITH RE, SABBA-SRUR AUO. Sphilantol: occurrence, extraction, chemistry and biological activities. Brazilian Journal of Pharmacognosy [Internet] 2015). Studies have also demonstrated its antibacterial and anti-inflammatory action (WU L, FAN N, LIN M, CHU L, HUANG S, HU C, HAN S. Anti-inflammatory effect so spilanthol from Spilanthes acmella on murine macrophage by down-regulation LPS- indiced inflammatory mediators. Journal of Agricultural and Food Chemistry. 2008; 53: 2341-2349); (ALCANTARA BN, KOBAYASHI YT, BARROSO KF, SILVA IDR, ALMEIDA MB, BARBOSA WLR. Pharmacognostic analyzes and evaluation of the in vitro antimicrobial activity of Acmella oleracea (L.) RK Jansen (Jambu) floral extract and fractions. J of Med Plants Res. 2014 Feb; 9 (4): 91-96).

[013] In a study on the anti-inflammatory and analgesic activity of Acmella oleracea, no adverse effect or mortality was observed in rats, administering up to 3g / kg orally, in the form of aqueous extract. The observation that spilantol inhibits several cytochromes, warns of the risk of adverse interactions between this substance and other drugs. The spilantol alkaloid present in the leaves, branches and flowers of A. oleracea (jambu) is described in patents as suitable for anesthetic, antiseptic, anti-wrinkle, toothpaste, gynecological, anti-inflammatory, with several products on the market, sold as medicine and cosmetic. This is the reason for the existence of five patents using A. oleracea registered with the United States Patent and Trademark Office (Uspto) in the period from 2000 to 2006 (one American, one French and three Japanese), seven with the World Intellectual Property Organization ( Wipo) (Japanese, American, English, Danish, Swiss, Brazilian and Australian) in the period from 2006 to

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2010 and one at the National Institute of Intellectual Property in 2005. A. oleracea is also used in the composition of anti-wrinkle creams.

[014] Nascimento (2012), in his master's dissertation, made an interesting survey of several patents containing the extract of A. oleracea, relating the presence of spilantol. These patents were applied for by Japanese industries for use as an additive to food and beverages; cleaning agent for body and hair preparations; use, together with a mixture of other plants, as an insecticidal product, for the control of insects and microorganisms in plants. In China, a patent has been applied for an oriental remedy for use in the treatment of common colds and flu composed of several plants, including A. oleracea (NASCIMENTO, AM. Polysaccharides and secondary metabolites of Spilanthes Oleracea L. (Jambu) [Dissertation] Curitiba: Federal University of Paraná, Sciences-Biochemistry; 2012).

[015] Nascimento (2012) also reports that the company Ballal BB has applied for a patent for a phytotherapic product against HIV infection and AIDS treatment, obtained from A. oleracea root powder. The Fortune Star company has applied for a patent for a herbal composition, including A. oleracea with fungicidal activity. A composition containing a mixture of several plants, with antibacterial and antimicrobial activity, was patented by the company Euro Natural Products. The company Dermane & Passaro describes a product for the cosmetic anti-wrinkle treatment based on pure spilantol or A. oleracea extract containing spilantol, capable of inhibiting the contraction of subcutaneous muscles, mainly of the face. The company Ikeda Bussa Co Ltd. has applied for a patent for cosmetic preparations for the body and face from A. oleracea root extract, to reduce wrinkles and anti-aging the skin. US2008 / 0069912 deals with the use of an A. oleracea extract for botox effect in a cosmetic anti-wrinkle composition. Natura Cosméticos S.A. claims a process for preparing chlorophyll-free A. oleracea extract and its use in cosmetic compositions. In patent number PI0802053-3, Natura, in partnership with UFBA and UNICAMP, registered a process of extraction, using supercritical fluid technology, of active compounds of plant material, with the main objective of obtaining spilantol and other alcamides from the A. oleracea plant.

[016] We are not aware of the use of A. oleracea manipulated with Transcutol ^® for topical analgesia of the skin in venipuncture.

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State of the art problems:

[017] In Brazil, there is a chronic lack of blood donors that can be explained by several reasons, including the fear of needle stick.

[018] EMLA® (Eutectic Mixture of Local Anesthetics) is a mixture of two local anesthetics (2.5% lidocaine and 2.5% prilocaine) with direct action on epidermis and dermis nociceptors and nerve endings, causing local analgesia . Sensitivity to pressure and touch can be maintained (FERREIRA, MARIANA SANTIAGO. Nursing care for children in pain. Diss. [Sn], 2014: 207-252).

[019] ELA-Max cream, marketed in Brazil under the name Dermomax (4% lidocaine), is also a topical formulation widely used in the temporary relief of pain associated with small cuts that affect only the epidermis, minor burns (1 ° degree), irritation and insect bites and is also used before procedures such as venipuncture, intradermal, subcutaneous or intramuscular injections or laser skin treatment.

[020] One of the biggest limitations to the use of topical anesthesia is the need to wait forty minutes for the topical anesthetic to contact the skin for absorption to occur.

Advantages of the invention:

[021] The main advantage of this product based on the extract of Acmella oleracea at 90% with Transcutol® at 10% is to significantly reduce pain and be effective in the antimicrobial action in venipuncture procedures within three minutes after application.

[022] In Watanabe's study (WATANABE KM, JAY J, ALICTO C, YAMAMOTO L. Improvement in Likelihood to Donate Blood After Being Offered a Topical Anesthesic. Hawaii Med J. 2011 feb; 70 (2): 28-29) adults completed a one-page survey on blood donation. Participants were asked about the likelihood of donating blood in the near future. And they were told about the possibility of using a topical anesthetic cream before the donation. Subsequently, the likelihood of donating blood was reassessed. The findings of this study suggest that the supply of a topical anesthetic had a positive effect on the study participants' likelihood of donating blood. This effect is even more positive among those who have never donated blood.

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Brief description of the Figures [023] Figure 1 - Cultivation of the Acmella oleracea plant in the botany laboratory of Univás.

[024] Figure 2: Obtaining the aqueous extract of Acmella oleracea.

[025] Figure 3: Dispersion of pain intensity points using the EVN scale.

Detailed description of the invention [026] The plant seedlings were obtained at an open market in the city of Belém-PA and grown in a greenhouse at the botany laboratory of Univás from May to November 2014, in Pouso Alegre-MG. The plant cultivation period was observed and the flowers were collected approximately 40 days after planting (FIGURE 1).

[027] The extraction of A. oleracea was performed using a method called "aqueous extraction by infusion" (EAI) (BARBOSA FS, LEITE GLD, PAULINO MAO, GUILHERME DO, MAIA JTLS, FERNANDES RC. Toxicity of extracts of Cyperus rotundus on Diabrotica speciosa Acta Sci., Agron. 2011 dec; 33 (4) in September 2015. The choice of extraction type was based on the use of an agent that would not interfere with the plant's antimicrobial activity, in this case, water distilled.

[028] For the extraction, 106g of fresh flowers from the plant were harvested and transferred to a shredder plus 200 mL of distilled water. The mixture was ground for approximately 10 minutes until a pasty substance was obtained. This substance was transferred to a 500 mL Erlenmayer and incubated in a 60 ° C water bath for 60 minutes. After this period, the pasty substance was vacuum filtered to obtain the extract (FIGURE 2).

[029] The extract was sent to a compounding pharmacy to add Transcutol® in a concentration of 10% representing the maximum concentration of the extract capable of causing an effect with the minimum of Transcutol® that could be incorporated to be absorbed by the whole skin. The product was packed in a plastic dropper bottle.

Description of experiments:

[030] Study design: A randomized, controlled clinical trial with healthy volunteers was carried out.

[031] Location: Venous punctures were performed at Ambulatory I of Hospital das Clínicas Samuel Libânio in Pouso Alegre, MG. Microbiological tests were

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7/10 conducted at the Basic Research Laboratory of the Faculty of Health Sciences, Central Unit of the University of Vale do Sapucaí, in the city of Pouso Alegre, MG.

[032] Ethical aspects: In conducting this study, the determinations of Resolution 466/12, of the National Health Council, which provides for guidelines and standards that regulate research involving human beings, were observed and followed. The research was approved by the Research Ethics Committee of the University of Vale do Sapucaí, under the number 959.061.

[033] Sampling and Sample calculation: As no study was found in the literature to evaluate the analgesic activity of A. oleracea on intact skin applied topically, which would allow using data to calculate the sample size, a pilot study was carried out with 15 patients in each group and these results were used for sample calculation.

[034] For the calculation, the primary study outcome (pain) was used using the student t test (one-tailed hypothesis). Using a standard deviation of 2.6, a clinically relevant difference of 2 points on the visual numerical scale (EVN), with a test power of 90% and considering a significance level of 5%, the calculated number of individuals, per group, was 29.

[035] Eligibility criteria: Inclusion criteria: healthy volunteers, aged 18 to 65 years old of both genders without restriction on ethnicity, education and social class, who agreed to participate in the research by signing the Informed Consent Form (IC). Non-inclusion criteria: pregnant women, cancer patients, chronic kidney patients and refusal to participate in the research.

[036] Selection: The random sequence was generated by the Randomization Plan software. The individuals were selected in a systematic random order.

[037] Volunteers were randomly allocated to groups:

a) 70% alcohol group (n = 30): 70% alcohol was used in skin antisepsis.

b) A. oleracea group (n = 30): A. oleracea 90% extract was used in skin antisepsis.

[038] Procedures: The volunteers were recruited on a first-come, first-served basis at Ambulatory I at Hospital das Clínicas Samuel Libânio in Pouso Alegre, MG. Upon arriving at the clinic for blood collection, the volunteer was informed about the study and

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8/10 invited to participate. Accepting, by signing the Informed Consent Form, he was then submitted to the procedures determined by the study. Sample collections were carried out at the outpatient clinic in the morning, between 7:30 am and 10:00 am. All punctures were performed by the same phlebotomist who had no knowledge about the product used in the volunteers.

[039] Antimicrobial evaluation of A. oleracea: The first swab soaked in sterile saline was collected from the antecubital fossa in a region marked by a sterile qualitative filter paper field with a 5cm2 fenestra, from the Nalgo® brand. The swab was placed in a sterile glass tube with saline solution containing eight glass beads. Then the product selected was applied according to the antisepsis technique recommended by the HCSL clinical analysis laboratory. Three minutes were waited, which was the estimated time for the product to act.

[040] The second swab was collected in the same way as the first. The patient was then referred to the laboratory phlebotomist for venipuncture. At the end of the collections, the swabs were placed in a thermal plastic box and sent to the basic research laboratory of Univás for analysis.

[041] For the analysis, each swab was homogenized on an automatic shaker for one minute with eight sterile glass beads that favor the removal of microorganisms from the swab into the solution. After homogenizing the tube, 50 pL of the solution were seeded into plates containing Plate Count Agar Standart Methods Agar (PCA) culture medium and spread over the entire surface with the aid of a Drigalski loop.

[042] After sowing the samples, the plates were placed in the greenhouse at a temperature of 35 ° C for a period of 48 hours. After this period, the colony-forming units (UFC) were counted.

[043] Pain assessment: The volunteers did the pain assessment after the venipuncture using a one-dimensional scale, the visual numerical scale (EVN) (figure

3). EVN is an 11-point scale consisting of integers from 0 to 10; where 0 represents' 'no pain' 'and 10 represents' worst pain imaginable' '. The volunteers were instructed to select a single number that best represented the intensity of the pain.

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9/10 [044] Results: Among the 64 volunteers included in the study, four were excluded, with 30 volunteers participating in the group (a) 70% alcohol and 30 in the group (b) A. oleracea. In the study carried out, there was no significant difference in relation to the mean age between the groups analyzed using the Fisher Test (p = 0.906) (Table 1). Table 1: Age distribution of volunteers

Age Median Average Standard deviation P value General 19-65 51 48.73 12.47 Alcohol Group 70% 19-65 51 49.16 12.71 0.906 Group A oleracea 21-65 52 48.30 12.44

[045] For the analysis of gender, the Two Proportion test was used with statistical significance of p <0.05. There was no statistically significant difference between men (p = 0.806) and between women (p = 0.785) in the groups analyzed (Table 2) ·

Table 2: Gender of the volunteers

Genre Male Feminine General 55.0% 45.0% Alcohol Group 70% 56.67% 43.30% A. oieracea Group 53.33% 46.67% P value 0.806 0.785

[046] The intensity of pain analyzed by the EVN scale in group A. oleracea was statistically lower compared with group (a) alcohol (p = 0.0206 by the Man Whitney Test; Table 3).

[047] It can be seen through FIGURE 1 that the dispersion between the points for the (b) oleracea group was less than for the 70% alcohol group, demonstrating greater precision in the responses.

Table 3: Pain intensity according to the EVN scale.

Median Mean EVN pain Standard deviation P value Alcohol Group 70% 2.5 2.90 2.44 0.0206 Group A. oferacea 1.0 1.60 1.57

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10/10 [048] The results of the UFC count by the antisepsis technique are shown in Table 4. The groups were analyzed using the Chi-square Kruskal-Wallis test with statistical significance of p <0.05.

Table 4: UFC count before and after performing the antisepsis technique.

Group Before antisepsis After antisepsis Before / After p value

Median Average Median Average B 70% alcohol 41.5 109.20 1.0 25.27 0.0000 A. o / eraçea 11.5 03.23 1.0 46.23 0.0125

[049] The reduction in the CFU count of the skin microbiota (before / after) the use of alcohol was statistically significant with p = 0.0000. A similar significance was also observed (before / after) the use of 4. oleracea with p = 0.0125 (Table 4).

[050] Considering the wide dispersion of data from the microbiological analysis in the study, the comparative calculations between 70% alcohol before / after antisepsis and Acmella oleracea before / after antisepsis showed that there was no statistically significant difference between the medians of the two products (p = 0 , 0643 by the Wilcoxon Mann-Whitney Test).

Claims (5)

1) “ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN”, characterized by being composed of the base of the extract of 90% Acmella oleracea with 10% Transcutol®. 2) "ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN", according to claim 1, characterized by the fact that the composition is in the pharmaceutical form of solution intended for topical application. 3) “ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN”, according to claims 1 and 2, characterized by the fact that the solution is in the pharmaceutical form for use on intact skin. 4) "ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN", according to claims 1 to 3 characterized by the fact that the solution is in pharmaceutical form to reduce pain in three minutes after application and be effective in action antimicrobial in venipuncture procedures. 5) “ANESTHETIC AND ANTISEPTIC COMPOSITION BASED ON ACMELLA OLERACEA FOR INTEGRATED SKIN”, according to claims 1 to 4 is in the pharmaceutical form to be administered to humans and animals in venipuncture procedures and in the use of needle and / or syringe .